Arrested Development of Nematodes and some Related Phenomena

Arrested DeveloDment of Nematodes and some Related Phenomena 1

. .

J F MICHEL

Central Veterinary Laboratory. Weybridge. Surrey. England I . Introduction .................................................................................... I1. Dictyocaulidae. Heligmosomatidae ...................................................... 111.

IV.

Dictyocaulus viviparus ..................................................................... Dictyocaulus filaria ......................................................................... Nipposirongylus brasiliensis............................................................... Trichostrongylidae ...................... Ostertagia circumcincia ............. Osiertagia osteriagi ...... .............................................................. Trichostrongylusspp . . . . .............................................................. Marshallagia marshalli... .............................................................. Cooperia spp.................................................................................. Nemaiodirus spp ............................................................................ Hyostrongylus rubidus ..................................................................... Graphidium strigosum ..................................................................... Obeliscoides cuniculi.... ............................................................. Haemonchusplacei ........................................................................ Haemonchus contortus .....................................................................

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V. VI . VII .

VIII

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IX .

X. XI .

Oesophagosiomum radiatum ... Oesophagostomumspp . of pigs ......................................................... The Spring Rise .............................................................................. Ancylostomatidae.............................................................................. Ancylostoma spp. Uncimria spp....................................................... Strongyloididae........................................... Strongyloides spp ...................................... Ascaridae ....................................................................................... Ascaridia galli .............................................................................. Ascaris lumbricoides, Ascaris mum ................................................... Toxocara canis .............................................................................. Toxocara cati ................................................................................. Neoascaris vitulorum ........................................................... Some other ascarids .............................................................. Heterakidae .................................................................................... Heterakis gallinae........................................................................... Spiruridae ....................................................................................... Habronema spp............................................................................ Discussion ....................................................................................... References .......................................................................................

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280 281 281 282 283 284 284 281 295 296 296 298 300 301 301 303 303 307 307 308 310 31 1 311 312 322 322 326 326 328 328 330 331 333 334 334 336 336 336 336 331 343

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I. INTRODUCTION The ability to interrupt its development is a necessary qualification of almost any parasite. Most parasitic nematodes have one or more clearly defined resting stages, development from which depends on the reception of some specific signal or stimulus. The question has been discussed in some detail by Rogers (1961). The phenomenon that is the subject of the present article is the temporary cessation of development of nematodes at a precise point in early parasitic development, where such an interruption contains a facultative element, occurring only in certain hosts, certain circumstances or at certain times of year and often affecting only a proportion of the worms. Where the growth and development of a proportion of the worms is arrested while the remainder proceed normally, bimodal size distributions tend to result and the occurrence of such distributions in nematode populations is almost diagnostic of the phenomenon and helps to distinguish it from a general slowing of growth and stunting of adult worms which are common effects of innate or acquired resistance (see for example Winfield, 1933; Mayhew el af., 1960; Bawden, 1969a; or Michel et al., 1972~). Dunsmore (1961) draws a useful distinction between arrested larvae, the development of which has temporarily ceased at an early stage, and retarded worms which have reached the final morphological stage but have attained a size less than normal. In much of the literature this distinction is not drawn; worms classified as “immature” may be arrested larvae, retarded or normally developing immature worms or stunted adults. Even where immature worms are classified in a more precise way confusion may arise, for where worms intermediate in stage or size between arrested larvae and adults are found in animals that have not been exposed to infection for some time, it is difficult to determine whether development has been retarded or whether the worms present were at one time arrested and have since resumed their development. It is the purpose of this paper to review some of the available information on arrested development as it illuminates the question of what factors cause development to be interrupted and the circumstances in which it is resumed. Arrested development has been reported to occur in a great number of hostparasite systems but there appear to be differences between them in points of detail. While there are also some striking similarities, it would not be justifiable to discuss the phenomenon as though it were identically the same in all systems. Accordingly the subject will be dealt with by parasite genera, an attempt being made to present as complete a picture as the literature permits. In a final section similarities and differences will be discussed. Although a very large number of papers deal, either directly or indirectly, with arrested development, their coverage is very uneven and it is evident, not only that the phenomenon will in time be observed in a great many more species, but also that many basic observations have yet to be made. Great differences in emphasis will be noticed in the accounts which follow. This is also a reflection of the literature which does not at present permit the construction of a balanced account of any but a very few species.

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11. DICTYOCAULIDAE, HELIGMOSOMATIDAE Dictyocaulus viviparus

Taylor and Michel(l952) found worms at the early fifth stage in the lungs of clinically affected cattle which had been withheld from the possibility of re-infection for periods of up to eight weeks. In experimentally infected calves early fifth stage worms down to 0.58 mm in length were present as much as 80 days after infection. Michel (1955) showed that such worms could persist without appreciable growth for several months. When resistant calves were given a large challenge infection and killed at different times from 24 to 21 5 days later all the worms recovered from their lungs were immature. In susceptible calves only a small proportion were arrested in their development. It was concluded that the inhibition of development was a consequence of the resistance of the host. It may be asked, however, to what extent these results were due to the greater loss of developing or mature worms from the resistant host. If the arrested worms persisted while developing worms were lost, then clearly, where a constant proportion of the worms that became initially established became arrested, and if many of the developing worms were subsequently lost, the proportion of immature worms in the population remaining would be greater than if developing worms were not lost. An interpretation of this kind might indeed explain the results of Michel (1955) and seems adequately to account for those of Weber (1958). It cannot, however, dispose of the results of Michel et al. (1965) which clearly indicate that in cattle immunized either with X-irradiated larvae or with untreated larvae, a larger proportion of the worms initially established is arrested than in susceptible controls. Similarly, the results of Parfitt and Sinclair (1967) who immunized calves against D . viviparus by giving them larvae of D.filaria indicate that a larger proportion of the worms initially established were arrested in the immunized than in the susceptible calves. The results of both Michel et al. (1965) and of Parfitt and Sinclair (1967) contain evidence of arrested development of a substantial proportion of the worms in susceptible animals. Of the worms recovered from susceptible calves by Parfitt and Sinclair 24 and 30 days after infection, 20 % were less than 3 mm long. On two out of the five occasions on which the cattle of Michel et al. were challenged, over 50 "/o of the worms recovered from susceptible controls after 30days were under 3mm in length; on the other three occasions the number of arrested worms was negligible. There was no obvious feature common to the two occasions and absent from the other three unless it be that the two occasions were in autumn and the other three in spring. That seasonal factors may be involved in the inhibition of development of D . viviparus is not improbable and is suggested by the observations of R. P. Gupta (personal communication, 1969). As early as 1948, Wetzel expressed the view that infections of D. viviparus were carried on from year to year by silent carriers and Michel (1955) suggested that arrested larvae might be of importance in this regard. A considerable volume of circumstantial evidence supports this. A survey conducted in Scottish knackeries by Jarrett et al. (1955) showed that the percentage of stirks i n which lungworms were found

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increased from seven in February to 31 in April. A similar survey by Cunningham et al. (1956) at two knackeries revealed a very small incidence in January but 33 %and 41 in March. The incidence in cows was much lower but showed the same trend. If it may be assumed that a similar sample of animals was examined in winter and in spring, then the number of worms actually increased at a time when for a number of reasons new infection was unlikely. But the methods employed in these surveys were not calculated to detect very small worms and it is likely that the apparent increase in the spring was due to the development of arrested worms. Swietlikowski (1959) in Poland, found that the number of heifers passing lungworm larvae in their faeces increased in the spring, which implies that heifers which had passed no larvae during the winter began to do so in the spring. According to Malczewski (1970b) Swietlikowski interpreted this as being due to a greatly extended prepatent period. Similar observations were made in Canada by Gupta and Gibbs (1970) who showed that the number of lungworm larvae in the faeces of yearling cattle fell to a minimum in February and increased again in May. In Austria, Supperer and Pfeiffer (1971) regularly examined the faeces of young cattle on a number of farms and found that 11 out of 64 passed lungworm larvae in the spring after being negative during the winter. Not only were these animals housed but a number had received anthelmintic treatment which might be expected to remove adult lungworms. Meanwhile,post rnortern examinations of cattle during the winter had frequently revealed the presence of early fifth stage larvae only, in the lungs. Supperer and Pfeiffer (1971) and Pfeiffer (1971) were in little doubt that D. viviparus persists through the winter at this stage and resumes its development in the spring. They point out that if the increased larval output in the spring were attributable to worms that had persisted as adults, the number of these would tend to decrease through the winter; but in the Scottish knackery surveys they increased. Supperer and Pfeiffer (1971) observed that it was yearling cattle rather than cows that acted as silent carriers, a fact noted also by Enigk and Duwel(l962) and by Grafner et al. (1965), and gained the impression that calves that acquired their first appreciable infection in the autumn were particularly likely to show patent infections in the following spring. Since calves acquire a resistance to the establishment of new lungworm infection very quickly (Michel, 1962), only calves first exposed (to more than very slight infection) in autumn could acquire appreciable numbers of worms at this time and this may suggest that larvae picked up in autumn are particularly likely to become arrested. All the evidence seems to suggest that worms arrested in their development persist in the host for much longer than those that have developed normally and the phenomenon may therefore be seen as a means of carrying on the infection from one year to the next. Certainly, the free-living stages do not survive on the pasture at all well (Rose, 1956). Dictyocaulusfilaria The finding of immature lungworms up to 100 days after experimental infection was reported by Taylor and Michel (1952) who suggested that it

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occurred chiefly in resistant sheep. SokoliC et al. (1963) stated that the development of D . jilaria in their artificially immunized sheep was inhibited but they did not make it clear whether they found some stunting or whether a proportion of the worms was at a very much earlier stage than the remainder. There are grounds for believing that the phenomenon of arrested develop ment plays an important part in the epidemiology of D.jilaria infection, for it appears that disease not uncommonly appears some months after the worms are acquired. Thus it is suggested that in some Middle Eastern countries infection is picked up on mountain grazings but outbreaks of lungworm disease do not occur until the sheep have been removed from sources of infection for some time and are weakened by deficient nutrition. Not dissimilar is the observation that while in England infection is chiefly acquired in late summer and autumn, outbreaks of disease commonly occur in January and February. Thesuggestion implicit in this, that arrested worms may be prompted to develop when host resistance declines, should be regarded with caution. Information is needed on seasonal changes in populations of adult and immature lungworms in sheep; at the present time there is no published information on the subject. Nippostrongylus brasiliensis While the relationship between N . brasiliensis and the rat has been more intensively studied than any other host-helminth system, there are few accounts of arrested development. Schwartz et al. (1 931) found that in rats killed 13-1 6 days after a first infection small numbers of third stage larvae were still present in the lungs. 13-16 days after a second infection the number of third stage larvae in the lungs was very much greater. Such work as touched on this subject during the next 25 years tended only to complicate and confuse the issue. Sarles and Taliaferro (1936) reported that in the previously infected rat, N . brasiliensis in the lungs were smaller than in the susceptible rat and took longer to migrate to the intestine. When such worms were transferred, after 63 days, to susceptible rats they grew normally and Sarles and Taliaferro therefore concluded that the worms were not permanently affected but “had merely been inhibited in their development”. Taliaferro and Sarles (1937, 1939) noted that in immunized rats larvae in the skin and the lungs were temporarily immobilized by a tissue reaction. If this reaction was intense, precipitates formed around the anterior end of the larvae which perished. This combination of observations and ideas led to the conclusion that inhibited development was necessarily the consequence of an acquired resistance. The work of Porter (1935), however, shows that innate resistance may have the same effect. Porter found that in an abnormal host, the deer mouse, N . brasiliensis migrates normally to the lungs but then remains there without further development. Although the phenomena described by these authors concerned third stage larvae in the skin or lungs, Chandler (1932) observed an apparently similar phenomenon affecting fourth stage larvae in the intestine. In a second infection “a few worms although they reached the intestine had failed to grow or develop at all after reaching the fourth larval stage. A small percentage of such are

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sometimes found even in first infections”. In an experiment in which rats received from one to six infections of 200 larvae at intervals of 7 days, the number of adult worms was much the same as in rats which had received the last infection only, but the number of inhibited fourth stage larvae tended to increase with each infection. Chandler deduced that the inhibition of development was an effect of an increasing host resistance but another interpretation of his results is possible. According to this a constant proportion of each dose of larvae became arrested and adult worms were lost after a short time. The adult worms present when the rats were killed were therefore those of the last infection only while the fourth stage larvae represented an accumulation from all the infections that the rat had received. Some doubt must remain whether Chandler was dealing with arrested development in the strict sense. Thus he found (Chandler, 1935) that worms that became established in previously infected rats showed great variation in their rate of growth, every stage from the early fourth to the mature adult being present. When Chandler (1936) transferred retarded or inhibited worms obtained in this way to susceptible rats they all grew to maturity but when they were transferred to resistant rats no further development occurred. Subsequent work with this host-parasite system has not been concerned specifically with arrested development and a great many questions remain to be answered. In particular it would be interesting to know whether the development of N . brasiliensis can be arrested in the intestine as well as in the lungs. Size distributions of worms in the intestine which might show part of the population to remain stationary while the rest grow normally, do not appear to have been published. Such data have, however, been presented by Twohy (1956) in the case of Nippostrongylus larvae in the lungs of rats. Twohy found that size distributions of worms from this site were bimodal and while the larger mode moved to the right with the passage of time as the worms grew, the smaller, representing worms between 600 and 700 pm long remained in the same place. The growth of all the worms was punctuated by periods during which increase in length was negligible and it is interesting that one of these periods occurred when the worms are between 600 and 700 pm long. This may suggest that some particular stimulus is needed to induce worms to develop beyond this stage. Resumed growth may depend on the satisfaction of some nutritional requirement, a suggestion that emerges from the work of Weinstein (1958) and of Weinstein and Jones (1959) who, in the course of experiments on the in vitro culture of Nippostrongylus,found that a proportion of worms reared in incomplete media failed to develop beyond exactly this point but persisted in good condition for a considerable time. When the medium was supplemented with vitamins and amino acids, growth was resumed. 111. TRICHOSTRONGYLIDAE Ostertagia circumcincta

From his studies of the parasitic development of Ostertagia circumcincta in sheep, Sommerville (1953) concluded that the larvae, having penetrated the

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mucosa on the third or fourth day after infection, could proceed in one of three ways: (1) They could leave the mucosa shortly after the third ecdysis; (2) they could grow within the mucosa to emerge from it at any stage; and (3) they could develop no further than the early fourth stage and remain in the gastric pits for up to 3 months or probably longer. In later work Sommerville (1963) examined the structure of experimentally established populations more critically and found size distributions to be bimodal. But while, among worms in the fundus, this situation continued at least until the 56th day, at the pyloric end of the stomach the large worms were lost and only the arrested larvae remained. While adult worms are lost more rapidly than the arrested larvae, the number of these also decreases with the passage of time (Sommerville, 1954; Armour et ul., 1966), presumably as they develop. Gordon (1953) also reported that the development of Ostertugiu spp. of sheep can be arrested and Horak and Clark (1964) in experimental infections of several sheep and a goat found up to 30% of the worms to be still in the fourth stage 4 weeks after infection. Arrested development is not, however, seen in all experimental infections of 0.circumcinctu (see for example, Threlkeld, 1934). By means of a very simple experiment, Dunsmore (1960) made a significant advance. He infected one small group of lambs with a single dose of 1000 larvae and a second group with 100 000 and killed all the lambs 14 days later. At the higher infection rate a very much greater proportion of the worms was arrested than at the lower rate. The size distributions are reproduced in Fig. 1. The peculiar significance of these results is that they cannot be explained in terms of a constant proportion of the worms initially established being arrested and more developing worms being lost from the larger infection. Even if it is assumed that as great a proportion of the large dose of larvae became established as of the small, the arrested worms represent 1.5 % of those at the low dose and 20.8 ”/, at the high dose. It is very difficult to avoid the conclusion that the size of the infection does affect the proportion of the worms that fail to develop beyond the early fourth stage.

Length (mm)

FIG.1 . Frequency distributionsof thelength of Ostertagia circumcinctain sheep which received loo0 larvae (broken line) or 1000oO larvae (solid line). Reproduced from Dunsmore (1960).

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Of course this does not mean that the loss of adult worms cannot also influence the proportion of arrested worms present. Connan (1 969) for example, who gave lambs 75 000 larvae on a single occasion and killed them 24 days later, found that there was an inverse relationship between the number of worms present and the percentage that were at the early fourth stage. Presumably this means that similar numbers were established initially in all the lambs but there was variation in the rate at which developing worms were lost. Dunsmore (1961) has shown that immune mechanisms of the host may be involved in the inhibition of development. He demonstrated that when sheep which had been subjected to whole body irradiation or which had received cortisone were given 100000 larvae, a very much smaller proportion was arrested than in control sheep. (Conversely James and Johnstone (1 967a) have suggested that the administration of cortisone to sheep causes the development of arrested worms to be resumed.) Having found that both arrested and developing 0. circumcincta could be removed with the anthelmintic thiabendazole, Dunsmore treated naturally infected sheep in this way and then infected these resistant but worm-free sheep with 60 000 larvae. When the sheep were slaughtered 23 days later most of the worms present were at the early fourth stage. Finally Dunsmore (1963) tried to show that the presence of adult worms also played a part in the inhibition of development and that when adult worms were removed, arrested larvae would be permitted to resume their development. In a first experiment the phenothiazine treatment used to remove the adult worms was only partly successful but there was some indication that the number of arrested worms decreased while the number of adults remained the same. In a second experiment anthelmintic treatment was more effective and a decrease in arrested worms and a corresponding increase in the number of developing worms were demonstrated. But because an essential control group had been omitted it was not possible to conclude with certainty that the development of arrested forms had indeed been prompted by the removal of adults. There is some evidence to suggest that the arrested development of 0. circumcincta is also subject to seasonal effects. Connan (1968a) found that Ostertagia spp. present in ewes were predominantly at the early fourth stage in the winter and mainly adult from April onwards. Reid and Armour (1 972) studied the larvae on a pasture grazed by ewes and lambs by putting susceptible lambs on it for short periods at different times during one year. They found that the development of some of the worms acquired by lambs grazing in late August was arrested and that the proportion failing to develop reached a peak in lambs grazing in December. Connan (1968a), in a similar investigation, had found some worms to be arrested in lambs grazing in September, considerably more in January and March and none in May or July. In the belief that some seasonal factor was probably acting on the larvae during their free-living existence, Connan(1969) investigated theeffect ofstoring larvae at 4°C and studied the parasitic development of larvae which had been stored at this temperature for 6 and 12 months respectively, but without demonstrating any difference in the number of worms arrested. Results discussedelsewherein this review suggest that even 6 months’ storage is too long

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and any changes in the larvae that might have occurred would already have been reversed. The question of whether environmental factors acting on the free-living stages causes parasitic development of 0. circumcincta to be arrested therefore remains open. Connan (1969) also reported a difference, in the extent to which their development was arrested, between different isolates of 0. circumcincta. A strain which had been maintained in the laboratory through more than 12 generations was apparently less prone to arrested development than one recently isolated. Even this strain did not become arrested if lambs were infected with freshly cultured larvae. Ostertagia ostertagi Arrested development has been more extensively studied in infections of 0. ostertagi in cattle than in any other host-parasite system but an understanding of the phenomenon is still far from complete. The first report appears to be that of Porter and Cauthen (1946) who found immature 0. ostertagi in a calf several months after the last of several experimental infections. Threlkeld and Johnson (1 948) encountered the phenomenon in calves used to study the survival of larvae on pasture. Some of these calves harboured immature worms, in one case in large numbers, 1 5 days after being housed. Vegors (1957) found large numbers of fourth stage larvae in cattle removed from pasture in late spring and then housed for 30 days in conditions calculated to prevent accidental infection. In the following year Vegors (1958) showed that the number of fourth stage larvae in cattle taken off pasture in early summer decreased only slightly during 28 days of housing. Martin et al. (1957) described a number of outbreaks in housed cattle in several of which most of the Ostertagia recovered were immature although circumstantial evidence indicated that the worms had been picked up some months earlier. There is also evidence of arrested development in the results of Herlich (1960) who found, in cattle that had been on pasture from February until April, that half the 0. ostertagi present were immature. Since there are no grounds for thinking that the population was being turned over at an uncommonly rapid rate or that new infection was being picked up at an increasing rate, it may be concluded that the development of at least some of the immature worms had been arrested. More detailed study of the phenomenon began about this time. Following on the work of Sommerville (1953) with 0. circumcincta, Threlkeld (1958) made similar observations on the development of 0. ostertagi and found that in this species also the time taken for development to be completed was very variable. The histotrophic phase might be terminated at any time between 96 h and 3 months or more after infection and development could be arrested. In preliminary experiments on the regulation of populations of 0. ostertagi in calves Michel(l963) observed that in calves which received constant numbers of infective larvae daily the number of early fourth stage worms increased steadily. It was evident that the development of these worms was arrested because, at the peak, the number present was equivalent to all the larvae that became established during 5 1 days. Moreover these worms were remarkably

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uniform in size and stage of development. There was also good evidence that the arrested larvae could and did resume their development. In two calves which were apparently incapable of a normal response to infection and which failed to show the usual manifestations of resistance, the number of early fourth stage larvae was very much smaller than in normally responsive calves killed at a corresponding stage in the experiment. For this and other possibly inadequate reasons, Michel concluded that the inhibition of development was an expression of host resistance. Evidence of the same tendency was that of Ross (1963a) who found that a larger proportion of a second infection was arrested than of a first infection. Ross and Dow (1964) were not, however, able to repeat this result. Michel (1969a) observed a linear increase in burdens of early fourth stage larvae, in calves which were infected daily, which indicated that a constant 1 1 % of the larvae that became established were arrested and that while adult worms were lost after a short life span, the arrested worms accumulated. This result did not suggest that host resistance was the cause of arrested development. Nor indeed did the work of Michel and Sinclair (1969) who found that equal numbers of worms were arrested in cortisone-treated and in untreated control calves which were infected daily. In cortisone-treated calves receiving 3000 larvae per day, arrested larvae accumulated twice as rapidly as in calves given 1500 larvae per day. Host resistance does, however, appear to play some part, though possibly a small one. Michel et al. (1973a) found that a larger proportion of a challenge infection was arrested in calves that had received larvae daily for some time than in previously uninfected calves. That the presence of adult worms may also be involved was suggested by the results of Michel (1963). Adult worms were regularly removed from calves infected daily by means of the anthelmintic Neguvon which is without effect on immature Ostertagia. In calves given this treatment, arrested worms did not accumulate as they did in untreated calves. When anthelmintic treatment was discontinued the number of arrested worms built up rapidly. It became obvious, however, that the chief cause of arrested development was not known. Attempts to establish large burdens of arrested worms experimentally were largely unsuccessful. Ritchie et al. (1966) could detect no arrested worms in calves which had received 100 000 larvae on a single occasion and Anderson et al. (1967) failed to build up significant burdens of arrested worms either by giving four doses of 100 OOO larvae at weekly intervals or by giving 20 doses of lo00 larvae over 4 weeks followed by a final dose of 400 000 or 800000. The essential clue to the causation of arrested development of 0. ostertagi was the observation by Anderson et al. (1966) that large burdens of arrested worms were associated with autumn grazing. In an experiment on infested experimental paddocks they showed (Anderson et al., 1965a) that a large proportion of larvae picked up in the autumn failed to develop and this was so both in calves that had grazed throughout the season and in so-called tracers, young susceptible calves which were put on the pasture for a short period and then housed long enough to allow normally developing worms to develop to the fifth stage. Clearly host resistance was not involved in these conditions and

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Anderson et al. suggested that experience by the free-living stages of autumn conditions induced a state resembling diapause. In further observations in which tracer calves were used on commercial farms, Armour et al. (1969a) confirmed the seasonal pattern, finding that no larvae were arrested in June as against 65 % in October, the same percentage being recorded whether the calves were killed 4 or 27 days after removal from the pasture. In somewhat similar observations in New Zealand, Brunsdon (1972) found that while the percentage of early fourth stage larvae in the worm burden of calves grazing continually on experimental paddocks rose from April to reach 84% in September, in tracer calves put onto the pasture for short periods at different stages in the experiment, the proportion of arrested worms was at no time greater than 8 %. Brunsdon rightly concluded that the arrested worms accumulated while the adult worms did not and that large burdens of arrested forms could be built up even if only a small proportion of the worms ingested became arrested. Armour et al. (1967) illuminated another formerly puzzling feature by their discovery that while the stock of 0. ostertagi used by many workers in Britain and other countries which was originally isolated at Weybridge in 1959 showed a very limited propensity for arrested development, a recent isolate from a farm in Ayrshire became arrested far more readily. Michel (1967) questioned whether the results described in this preliminary report could not be interpreted as showing an effect of worm numbers, but subsequent and fuller accounts by Armour et al. (1967b, 1969b) showed that there was no connection between the number of worms present and the proportion arrested. They considered that there was variation between strains in their ability to respond to seasonal environmental factors. The nature of these factors was not spelled out in detail but Armour (1970) refers to experiments in which larvae of both the “Weybridge strain” and of the “field strain” were exposed to “autumn conditions” in a climatic chamber for a period of 10 weeks, with the result that the development of a large proportion of larvae of the field strain but not of the Weybridge strain was arrested when they were fed to calves. Michel (1967) had pointed out that the changes produced in the larvae by exposure to seasonal factors must be reversible because the identical larvae which became arrested if picked up by calves in the autumn developed normally if picked up 4 or 5 months later. Because many of his larvae had died during their 10 weeks exposure in the climatic chamber, Armour (1970) suggested that a fixed and possibly small proportion of larvae of the field strain were susceptible to the factors inducing arrested development and that these persisted better on the pasture so that their relative abundance increased in the autumn. In spring their relative number decreased again as eggs that had overwintered on the pasture hatched and developed. An explanation of a similar kind had been proposed by Sollod (1967) who suggested that the “field strain” was really a mixture of strains (i.e. polymorphic) and that one strain (morph) overwintered in the host in a dormant state and did not survive well on the pasture while the other survived well on the pasture but was not subject to the inhibition of development. Sollod‘s theory does not explain the increase in the proportion of larvae

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that become arrested in the autumn and Armour’s fails to account for the decrease in spring without invoking the persistence of eggs through the winter and their hatching in the spring, a process which does not occur to any significant extent. Moreover, more recent work by Michel, Lancaster and Hong (unpublished observations, 1972) has shown that the changes are indeed reversible. They stored larvae at 10°C and at 6 week intervals determined the percentage that became arrested when fed to calves. There was a negligible mortality among the larvae during the period of storage and their infectivity remained the same. The results are shown in Fig. 2. As yet there is no evidence to suggest that larvae which are inherently more prone to arrested development survive less well on the pasture than larvae which are incapable of interrupting their parasitic development. That there is a genetic basis for variation in the propensity for arrested development emerges from an experiment described by Michel et al. (1973b) which indicates that the progeny of worms which have overwintered in the host have a greater aptitude for arrested development than the progeny of worms that overwintered on the pasture. By selection, considerable changes in the composition of populations may be rapidly achieved. It would be misleading, therefore, to think of distinct strains of 0. ostertagi native to different regions. The characteristics of a population will depend on the manner in which the cattle are managed and populations will vary from farm to farm. It is not hard to identify the selection pressures in consequence of which the inhibition of development in response to seasonal factors may have evolved.

Weeks

FIG.2. The percentage of Ostertugia ostertagi which became arrested in infections in calves with fresh larvae or larvae stored for various periods from 6 to 24 weeks, showing that the proportion that became arrested reached a peak after 12 weeks’ storage and then declined.

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The life of adult worms in the host is short (Michel, 1963, 1970a) and worm eggs which reach the pasture in late autumn or winter have a negligible chance of developing to the infective stage (Rose, 1961). Accordingly, worms that infect the host between autumn and mid-winter leave hardly any progeny. The arrested development of worms picked up at this time enables them to persist in the host until the spring, when conditions are again favourable for the development of the free-living stages. It is of interest in this connection that in parts of Australia it is Ostertagia larvae which are picked up in the spring that are inhibited in their development (Hotson, 1967; Anderson, 1972; Anon, 1973): Australian 0. ostertagi becomes arrested in the same calendar month as its British ancestors and while it is tempting to think of a biological clock keeping perfect time for 150 generations, it is more probable that the change was due to the operation of a different selection pressure, namely the loss of the progeny of any worms which were reproducing during the hot dry summer. The rapidity with which selection can produce measurable changes in a population explains why the stock of 0. ostertagi maintained at Weybridge since 1959 has almost entirely lost its ability to become arrested. The procedure has been to infect a calf, usually with larvae that have been stored for 3 or 4 months and to culture its faeces when the egg output was at its peak, i.e. early in the infection. By this procedure a large part of the worms capable of being arrested would fail to develop and the resulting culture would contain only the progeny of worms that had not responded to the effects of storage. Now that the issues involved are recognized another isolate is being subjected to selection with the opposite tendency. Calves are infected with larvae that have been stored, worms that develop directly are removed by anthelmintic treatment and cultures are made when arrested worms have developed to maturity. If arrested development depends, to so large a degree, on seasonal factors, it may be expected that the relative numbers of immature and adult worms should show a seasonal pattern. Such a pattern was demonstrated by Ross (1965). Although the sample of animals that he examined was small and heterogeneous, his results are of the greatest interest. For a period of 12 months Ross examined four abomasa per month from cattle between 6 and 24 months old from an Irish knackery. His results are reproduced in Fig. 3. Numbers of immature Ostertagia were high in the winter and low in the summer. Adult worms showed peaks in March, due presumably to yearling animals in which arrested worms were developing, and in September due to calves in their. first grazing season (see Michel, 1969b). Malczewski (1970b) who examined 12-18 month-old cattle in a slaughterhouse in Poland also found that numbers of immature Ostertugia rose from September to a peak in November and then declined while numbers of adults increased to their maximum in May. A similar pattern is seen in the results of Bessonov (1967) who studied the incidence, in cattle reaching a slaughter-house in Kazhakstan, of nodules containing Ostertugia larvae. He found that their numbers increased from September to January and decreased in the spring. In autumn and winter the nodules contained larvae between 1 and 2mm long; in late winter and early spring the larvae appeared to be growing. In New

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7

40-000

30,000 -

a c

:

-

D

a 20,000v)

?

a“

10,000 -

I

I

I

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FIG.3. Seasonal changes in burdens of immature (broken line) and mature (solid line) Osrertugia ostertugi in cattle, 6 2 4 months old. Reproduced from Ross (1965).

Zealand, Brunsdon (1971) found that numbers of early fourth stage 0. ostertagi in calves maintained on experimental paddocks rose in winter and fell in late spring. While there is no room for doubt that arrested larvae are capable of resuming their development, the question of what factors may stimulate this remains unresolved. The matter is of more than academic interest, for such resumed development can be the cause of severe disease. Martin et al. (1957) described ten outbreaks of disease which they attributed to this cause. What were evidently very similar outbreaks had been reported, though without a clear analysis of the origin of the worms, by Ackert and Muldoon (1920) in Kansas, by Wetzel (1950) in Germany and by Threlkeld and Bell (1952) in Virginia. More recent reports are those of Burger et al. (1966) in N.W. Germany and of Raynaud (1968) in France. The old term “winter ostertagiasis” has, been largely superseded by the alternative “type I1 ostertagiasis” proposed by Anderson et al. (1965b). The distinction between type I and type I1 consists only of the source of the worms which are recently acquired in the first case and have developed from arrested forms in the second. The term “pre-type I1 ostertagiasis” to denote the presence of large numbers of arrested larvae is unsatisfactory because it implies that where such burdens are present, clinical ostertagiasis must inevitably follow. This does not appear to be so; large burdens of arrested worms are extremely common, especially in out-wintered cattle, but outbreaks of winter ostertagiasis are relatively rare. Because populations of adult worms are turned over rapidly, it may be expected that arrested worms will give rise to disease only if large numbers

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develop over a short time. This could come about in a number of ways. The larvae could resume their development after a fixed interval in the host so that if large numbers were acquired over a short period, large numbers would develop over a short space of time. Alternatively the larvae could develop after a fixed time from receipt of the signal in response to which development had been halted. Both suggestions would, however, be hard to reconcile with the observations that although cattle which are housed in late autumn acquire the whole of their burden over a period of 6 weeks or less, outbreaks of winter ostertagiasis may occur in such animals from December to June. According to another view some specific stimulus is required to induce the development of arrested larvae in large numbers and a possible factor might be associated with parturition. A condition resembling winter ostertagiasis has been described as occurring in newly calved heifers by Hotson (1967) and, in personal communications, by White (1970), de Chaneet (1971) and Whitten (1972). Similar cases have also been described by Bailey and Herlich (1953) and by Smith and Jones (1962). Outbreaks such as these do not provide conclusive evidence, however, that some factor associated with parturition has stimulated development. Large burdens of adult worms could arise because the usual loss of adult worms failed to occur. Small numbers of arrested Ostertagia appear to resume their development constantly. Adult worms that are lost are as readily replaced from a reservoir of arrested larvae as by the acquisition of new infection. Even in cattle withheld from further infection, the population of late fourth and fifth stage worms is in a state of dynamic equilibrium as adult worms are lost and replaced by development from the early fourth stage. How this equilibrium is maintained is still far from clear. The view of Michel (1963) that a constant proportion of the burden of early fourth stage larvae resumes its development every day is certainly mistaken. Although the burdens of arrested larvae that may be found in any homogeneous group of cattle in winter are extremely variable, the numbers of developing and adult worms present tend to be very uniform (Michel, unpublished observations, 1966; Egerton et al., 1970). Arrested larvae of 0.ostertagi are highly resistant to the action of anthelmintics (Armour et al., 1967; Reid et al., 1968; Baker and Walters, 1971). Consequently treatment of an animal carrying a population of arrested worms produces only a very temporary effect on the number of adults, because the worms removed are promptly replaced by the development of a corresponding number of larvae. Michel(1970b) has shown that repeated anthelmintic treatment can ultimately deplete the burden of arrested worms. In the absence of anthelmintic treatment the number of adult worms depends on the average life span of adult worms, which does not vary greatly, and on the rate of recruitment. Where new infection is prevented, the number of arrested worms resuming development per unit time determines the rate of recruitment and hence the number of adults. But at the same time the number of adults must play an important part in regulating the rate at which arrested larvae resume their development. A more dramatic development of large numbers of larvae may destroy the delicate regulatory mechanism that this implies. In a recent series of observations

3

_ _ -- --

Early 4 t h stage larvae ------------ - - _ _ _ _ _ _ _ _ _-_ ____

-

- 2-

n

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I

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-

o . . . ;.............,....:...

FIG.4. Burdens of early fourth stage, late fourth stage an4 fifth stage Ostertagia ostertagiin a group of yearling cattle which grazed infected pastures until the end of December and were then housed.

Michel, Lancaster and Hong (unpublished observations, 1973) housed a group of yearling cattle in December, after they had acquired large burdens of arrested worms on an experimentally infested pasture, and then killed animals at intervals of time. Results of worm counts are shown in Fig. 4. The number of developing and adult worms remained at a moderate level until March when they greatly increased. In consequence the stock of early fourth stage larvae had largely been depleted by April. Meanwhile it was evident that the population of adult worms was turned over rapidly. This interesting observation could not identify what caused the great increase, in the number of arrested worms that developed per unit of time, and further observations are being made. Some evidence of a possible involvement of host nutrition on the decline of burdensof arrested larvae is contained in a number of papers from Experiment, Georgia. Young cattle in this part of the U.S.A. are customarily wintered on temporary pastures sown especially for the purpose. These pastures can hardly be a significant source of infection. They are clean when the cattle are put on in November and December and it seems improbable that an appreciable infestation can be built up before they are taken off again. The large burdens of immature Ostertugiu that cattle coming off such pastures in the spring are found to carry have almostcertainly been acquired before the cattle were put on to the winter pasture. Vegors et ul. (1955) showed that if the yearlings were given supplementary feed while on the winter pasture they carried fewer worms, both mature and .immature in the following May. In somewhat similar circumstances Vegors et ul. (1955) found more immature worms in cattle receiving a diet low in protein. Similarly Ciordia et al. (1971) noted that cattle grazing winter pastures at a high stocking rate made poor weight gains and carried more worms than comparable cattle at a lower stocking rate. Up to 90 % of the Ostertugia were at the early fourth stage. It seems likely that in these cases the rate of loss of adult worms was reduced in poorly nourished cattle and hence the burden of arrested worms was not depleted so rapidly. That other factors may

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also affect the rate of this depletion is suggested by the results of Ciordia et al. (1 972). Trichostrongylus spp. Records of arrested development of Trichostrongylusspp. are few. According to Michel (1952a), T. retortaeformis is arrested at the late third stage and a large proportion of the larvae given to previously infected rabbits may fail to develop beyond this stage. In susceptible rabbits a very much smaller proportion of the worms is arrested. Burdens of adult T. retortaeformis are regulated by the occurrence of an abrupt elimination of fifth stage worms whenever the biomass of worms exceeds a critical level (Michel, 1952b). In an animal carrying large numbers of arrested larvae and not exposed to further infection, the faecal egg counts show successive peaks separated by periods during which no eggs are passed. Meanwhile the number of late third stage larvae shows a logarithmic decrease. In rabbits infected daily the burden of late third stage larvae rises to a peak from which there is a logarithmic decrease. The number of fourth and fifth stage worms follow a similar though rather blunter curve displaced to the right by one prepatent period. It was demonstrated (Michel, 1953) that the number of late third stage larvae began to decrease when the rabbits became refractory to the establishment of new worms. When the administration of infective larvae to some of the larvae was stopped at this point the infection ran the same course as in rabbits which continued to receive infective larvae. From this it was concluded that in both groups all the fourth and fifth stage worms that were present late in the infection were derived from arrested worms that had resumed their development. Size distributions suggested that they were doing so in batches. There are obviously some points of similarity between these phenomena and those seen in some other host-parasite systems. More worms appear to be arrested in previously infected than in susceptible rabbits. A greater proportion of a large dose of larvae is arrested than of a small dose. That the arrested larvae resume their development in batches suggests a regulatory mechanism whereby the presence or absence of adult worms controls resumed development. The development of Trichostrongylusaxei can also be arrested. Herlich and Merkal (1963) found that in experimental infections 1 % of the worms were immature 49 days after the administration of infective larvae to susceptible calves. In calves that had been immunized in various ways between 2 % and 4%of the worms were immature. The number of animals was small but the difference appeared to be a real one and seemed not to be due to differences in the number of adult worms that had been lost. Nor could the immature worms have persisted from the immunizing infection, for some of the calves had been immunized by the implantation of mature worms. Development may also be arrested in hosts of an unsuitable species. Rohrbacher (1960) infected rabbits with T.axei of bovine origin and found that up to 100% of the worms were still immature 3-4 weeks later, although there was considerable variation between replicates. Supplementation of the standard

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ration with ascorbic acid increased the percentage of immature worms but this effect was not apparent in pregnant does. Unfortunately neither Herlich nor Rohrbacher indicated the stage of their immature worms and this must weaken the conclusion that arrested development of T. axei is associated with an innate or an acquired resistance of the host. It may be significant in this connection that Denham (1969) who infected immunized lambs with T. colubriformis found stunted adult worms but no arrested larvae. Hart (1964), however, presented what he felt to be circumstantial evidence of arrested development of T. axei in Nigerian Zebu cattle. Marshallagia marshalli

Vural et al. (1972) gave a single infection of M . marshalli to sheep which had had some previous experience of infection, and found that two-thirds of the worms present 30 days later were still in the early fourth stage. Cooperia spp.

In experimental infections of calves with mixed trichostrongylid worms Goldberg (1959) found that early fourth stage Cooperia larvae (the adults present were C . oncophora and C .puncrata) were present 48 days after infection although accidental infection was very unlikely. About the same time Sommerville (1960) was studying the development of Cooperia curticei in sheep (Fig. 5). He noted that the development of some worms was arrested just before the fourth ecdysis and that after the majority of the worms had reached this point the size distribution of the population

7 1

+

Age of population (days)

FIG.5. Growth curve of Cooperiu curticei in sheep. +Adult females; x adult males; 0

fourth stage females; 0 fourth stage males; Asexes not differentiated.Reproduced from Sommerville (1 960).

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tended to become bimodal. Normal growth of all the worms was briefly interrupted at the end of the fourth stage and it was precisely at this point that development of some was arrested for long periods. This is shown in Fig. 5 . Sommerville thought that this was not a coincidence but that worms close to an ecdysis were “more likely to be adversely affected by unfavourable features of the environment”. The development of Cooperia oncophora is arrested at the early fourth stage (Michel et al., 1970). Records regarding other species are imprecise and refer either to “immature worms” or to “fourth stage larvae”. An observation of a strange kind may be mentioned at this point. Stewart (1958) gave a vast number of larvae of C . punctata to steers newly taken off pasture and found up to 600 ensheathed third stage larvae in their abomasa as late as 9 days later. He suggested that exsheathement had been inhibited as the result of host resistance. Herlich (1965a) observed that calves which had been infected once only with 350 000 larvae or more of C.pectinata or C .punctata harboured immature worms 6 weeks later. The number of these immature worms was, however, very variable and ranged from 1 % to 70 % of the total. That the large numbers of larvae administered were probably not the cause of arrested development, emerges from the work of Herlich (1965b) who used much smaller infections. In studying cross resistance between C . oncophora and C . pectinata both in calves and in lambs he found variable numbers of immature worms 20 and 28 days after infection but it was evident that neither acquired resistance nor the inherent unsuitability of the host was a primary cause of arrested development. This was also the conclusion of Goldberg (1973) on the grounds that he had observed development to be arrested in primary infections of susceptible calves and that on another occasion he had seen no arrested larvae in resistant calves. On the other hand the results of Herlich (1967) with C. pectinata show that the administration of 30000 larvae divided into 10 daily doses results in the presence of more arrested worms, 33 days after the last dose, than are to be found if the same number of larvae is given on a single occasion. In the animals dosed daily, both the proportion and the absolute number of immature worms was greater. These results and those of a second similar experiment could not be explained otherwise than by postulating that a greater proportion of the worms initially established was arrested in the animals given larvae daily. There is evidence that arrested development of C . oncophora may be caused by seasonal factors. Anderson et al. (1965b) mentioned that this species tended to be arrested in calves grazing in the autumn and Michel et al. (1970) have shown that when successive pairs of young susceptible calves grazed an infested pasture for short periods and were then killed after a period of housing, the percentage of the worms they carried, which failed to develop beyond the early fourth stage, increased from September to a peak of over 90% in December and then fell again to reach a very low level in March (Fig. 6). Brunsdon (1972), working in New Zealand, confirmed these findings. Since cattle become refractory to C. oncophora infection fairly rapidly, observations on seasonal changes in burdens of mature and arrested Cooperia in slaughter-house or knackery material could well be misleading. In England, C.

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20

FIG.6. The proportion of Cooperia oncophora which became arrested in pairs of calves exposed to infection for short periods on pasture, each pair at a different time of year. Reproduced from Michel et al. (1970).

oncophora larvae tend to be absent from young cattle during the winter following their first grazing season because larvae, picked up during the critical period, no longer become established. Nonetheless, Ivanova (1969) who examined cattle coming into a slaughter-house in Western Siberia found that burdens of immature C. oncophora and C. punctata were small in summer and large in winter. Nematodirus spp.

Gibson (1959) infected lambs of various ages with large numbers of larvae of N . battus and N.Jilicollis on a single occasion. The youngest lamb, aged 8 weeks, died and many adult worms and very few immature worms were found in it. The other lambs, which were between 28 and 89 weeks old at the time of infection, survived and when they were slaughtered either 4 or 12 weeks after infection, most of the worms recovered from them were immature and in some animals it appeared that a loss of adult worms had occurred. Gibson also infected two lambs, 58 and 98 weeks old respectively, which had been infected previously and these, when slaughtered 4 weeks later, also had large burdens of immature Nematodirus and few adults. These results do not give any clear indication of the causes of arrested development in this system because the only animal in which the phenomenon did not occur differed from the others not only in being younger, but also in that it succumbed to the infection. It seemed not improbable, however, that some host response was involved. But in a subsequent experiment Gibson (1963) showed that about 50 % of immature worms were present both in previously infected 14 week-old lambs and in susceptible controls, when they were killed 8 weeks after receiving a large challenge infection of N . battus. Nonetheless, Donald et al. (1964) and Dineen et al. (1965a) saw the inhibition of development as an effect of host resistance. They derived this view from an experiment in which groups of lambs received 50 OOO or 130OOO larvae of N . spathiger on a single occasion or 50 000divided into 25 daily doses. Lambs were killed on the 21st, 40th and 74th days and, on the basis of the percentage of worms in the fourth stage on these days, it was concluded that development was

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arrested (at the late fourth stage) to a greater degree in the larger infection than in the smaller, and to an even greater extent in lambs infected daily. The results of a second experiment of the same kind (Dineen et af., 1965a) showed the same differences but the percentage of arrested larvae in all groups was lower, a circumstance which Dineen et al. attributed to the greater inherent susceptibility of the particular batch of lambs. It is obvious, however, that the proportion of the worms that are at the late fourth stage may be high, either because the number of worms arrested at this stage is great, or because the number of adult worms remaining is small. Many of the worms in the lambs of Donald and his colleagues were lost during the course of the experiment, the number of adults decreasing faster than the number of fourth stage larvae. When absolute numbers of fourth stage larvae are considered, it becomes difficult, indeed, to discern differences in the extent to which development was arrested in the three groups of lambs. Dineen et af. (1965a) consider that arrested development is a major expression of population control. They visualize that the inhibition of development, and its subsequent resumption, depend on a response by the host to antigenic stimulation about a threshold, a decline, through the loss or removal of worms, reducing the host’s resistance and permitting the development of some arrested worms. This, in turn, increases the level of antigenic stimulation with a consequent increase in resistance. Such a sensitive regulatory mechanism resembles another proposed by Dineen (1963) as controlling the loss of adult worms but it has been questioned by Michel(1969a) whether populations of adult worms are really regulated in this way. If satisfactory evidence of a specific effect of host resistance on arrested development of Nematodirus spp. is still lacking, the results of Mapes and Coop (1970) do at least suggest that the phenomenon may be induced by a change of conditions in the gut. They gave a moderate number of N. battus larvae to lambs a few days after these had received one million larvae of Haemonchus contortus, an extreme measure which had markedly raised the pH of the anterior small intestine. Not only were the N . battus, present after 15 days in such pretreated animals, markedly smaller than in control lambs but their size distribution was distinctly bimodal. The Nematodirus population could be divided into worms which had ceased to grow at a length of approximately 3 mm and worms which had developed normally. In a second very similar experiment, Mapes and Coop (1971) failed to demonstrate arrested development but in a third experiment they found a bimodal size distl-ibution in the fourth stage worms of a first infection and showed that, after the passage of time, only the first mode remained while the other disappeared as the worms developed to the fifth stage (Mapes and Coop, 1972). The apparent difficulty in repeating observations on arrested development suggested that some factor or factors not controlled by Mapes and Coop or by Dineen et al. (1965a), and of greater importance, was operating. That seasonal factors acting on the larvae might be involved is suggested by the work of Reid and Armour (1972) who turned out susceptible lambs for short periods at different times of year and killed them after a period of housing sufficient to allow developing worms to reach maturity. Although the numbers

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recovered were not large, it appeared that the development of N . Jilicollis larvae picked up from the pasture in September and October was arrested. Ayalew et al. (1973) studied the worm burdens of ewes in Canada in June and in December. Although they also found only moderate numbers of Nematodirus spp. it was evident that they were predominantly immature in December and mainly adult in June.

Hyostrongylus rubidus Kotlhn and Hirt (1928) noticed nodules in the stomachs of pigs and found that they contained immature H. rubidus. Since these nodules were rather rare while adult H. rubidus were very common Kotlhn and Hirt doubted whether a histotrophic phase was a normal part of the life-history of this species. On re-examining this material, and on the basis of further observations Kotlan (1949, 1954) stated that while a period of development in the gastric glands was normal, it was possible to distinguish between a regular histotrophic phase of short duration and an irregular histotrophic phase in which the larvae persisted in the glands for an extended period without developing further. In these cases, development was suspended about the third moult and the larvae showed no tendency to emerge from the mucosa. A rather larger proportion of the worms behaved in this way in rabbits than in pigs. Kotlhn (1952) thought that an irregular histotrophic phase always ended in the death of the larva but this view is no longer tenable. Burden et al. (1970) produced experimental evidence of arrested development of H. rubidus. The phenomenon tended to occur on a skghtly larger scale in old pigs than in young, but differences between different experiments were of a very much more striking order. This may suggest that uncontrolled factors acting possibly on the free-living stages may play a part. Connan (1971a), in the course of a survey of worm burdens in adult pigs, noted a seasonal trend in numbers of early fourth stage Hyostrongylus larvae which rose in autumn, reached a peak in December and declined to low levels in the spring. Over 5 % of the pigs examined had burdens of early fourth stage larvae in excess of 20000. The largest burdens of adult worms occurred in lactating sows and tended to persist in sows in poor condition. Hyostrongylosis is regarded primarily as a disease of adult pigs. Castle (1932) and Nicolson and Gordon (1959) described outbreaks in sows shortly after farrowing and Dodd (1960) stressed the economic importance of such outbreaks. Hyostrongylosis is much less common in young pigs although, according to Thoonen and Vercruysse (I 951), it can occur. Burden and Kendall (1973b) failed to demonstrate an effect of pregnancy, parturition or lactation on the course of experimental infections in gilts which they infected with 100 larvae daily for 40 weeks, the last dose being given shortly before farrowing. But the results of theirpost mortem worm counts are interesting because they led to a novel theory on the regulation of populations of arrested larvae. In earlier work Burden and Kendall(1973a) had found that in pigs infected every day, numbers both of adults and of larvae remained relatively constant. They deduced that worms were being lost after a short adult life and the population was in a state of dynamic equilibrium. When

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Burden and Kendall (1973b) stopped infecting their gilts after 40 weeks they saw little change in the number of fourth stage larvae although it was to be expected that these would rapidly decline. To explain this finding they suggested that the number of arrested larvae that can exist in a pig at one time is limited and that new worms can only establish themselves if a space has been left for them by the maturation of older larvae. On the basis of Burden and Kendall’s results, this limit would be less than 1000 larvae and it would therefore be difficult to account by means of this theory for the very much larger burdens of fourth stage larvae occasionally encountered, as for example by Connan (197 I a). Another interpretation of Burden and Kendall’s (1973b) results is however possible. The number of pigs that they used was inevitably small and variation in worm burdens was considerable but scrutiny of the figures shows that when infection was discontinued, the number of fourth stage larvae fell to approximately half its former level. This implies that at this point in time half the larvae present were developing to maintain a constant number of adults while the other half were arrested. Graphidium strigosum Martin et al. (1 957) refer briefly to the observation that when a very large number of infective larvae of G . strigosum was given to rabbits, the majority of the worms present 7 weeks later was immature. Obeliscoides cuniculi

0 . cuniculi, a parasite of the cottontail, can very successfully parasitize the domestic rabbit and it is therefore of interest for experimental purposes. As shown by Sollod (1968), development can be arrested at the early fourth stage, the larvae having doubled their length since beginning their parasitic development. Size distributions of worms from infections of different ages in susceptible rabbits are bimodal, the first mode in every case being centred on 2 mm (Chitranukroh, personal communication, 1972). Russell et al. (1 966) infected rabbits with infective larvae ranging in number from 2500 to 25 000 per kg liveweight. When the rabbits were killed 50 days later it was found that the number of adult worms present was the same in all groups but that the number of arrested fourth stage larvae was roughly proportional to the number of infective larvae administered. The results are summarized in Fig. 7. Russell et al. interpreted these findings in the following terms: “Excess larvae remain in a state of temporarily arrested development awaiting either a decline in host resistance allowing development of additional adults or until some or all of the adult population dies or is eliminated by the host.” They visualized arrested development as being due to host resistance and affecting all worms above a critical number. But at the lower rates of infection the worms recovered represented a greater proportion of the larvae administered than at the higher rates and a more plausible interpretation might be that a constant proportion of the worms initially established is arrested and that subsequently a loss of developing worms reduces the number that grow to maturity to a constant

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Lorwe ,,’

-0 4 -

n

x

gE

3~

2I I

5

1 I I 10 15 20 lnoculum (larvae per kg ,x 10’)

I

25

FIG.7. The relationship between the number of Obeliscoides cuniculi administered to rabbits and the number of larval and adult worms present after 50 days. Drawn from data presented by Russell el al. (1966).

level. On thebasis of the transitory weight loss observedby Russell et al. 5 days after infection it seems not unlikely that this loss of developing worms occurred quite early in the infection. Evidence in support of this second interpretation is furnished by the results of Hutchinson et al. (1972) who demonstrated that, with doses of infective larvae ranging from 5000 to 15000, there was an apparent effect of dose size on arrested larvae when these were expressed as a percentage of the number of worms recovered 14 days after infection, but not when they were expressed as a percentage of the number of larvae administered. “These results”, they concluded, “show that a fairly constant proportion of the inoculum remains as inhibited larvae.” This must imply that the smaller percentage established at higher infection rates is due to a loss of worms which occurs after the worms reach the early fourth stage and which is well under way before the 14th day. It means also that arrested worms are exempt from this loss. An important advance was the finding by Stockdale et al. (1970) that storage of the infective larvae at 4”C, before they were administered to rabbits, increased the proportion that became arrested. (It may be significant in this connection that the larvae used by Russell etal. (1966) had been stored at this temperature while a sufficient number for the experiment was collected together.) Fernancb et al. (1 97 1) maintained infective larvae at 4°C and showed that the proportion that subsequently became arrested in rabbits rose to a peak after approxii mately 8 weeks’ storage and then declined again. At a storage temperature of 17°C the proportion that became arrested rose to and declined from a rather earlier and considerably lower peak. By infecting rabbits with mixtures of stored and fresh larvae Fernando and her colleagues were able to show that the worms which developed most quickly were not exerting an inhibitory effect on the remainder.

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In later work, Hutchinson et al. (1972) showed that storing larvae at 15°C and then dropping the temperature to 5°C greatly increased the number that was arrested. On the basis of these experiments, they regard arrested development as a seasonal phenomenon due chiefly to the effect of decreasing temperature on the infective larvae and possibly associated with over-winter survival. It is perhaps significant that populations of 0. cuniculi in snowshoe hares were found by Erickson (1944) to increase in February and March. Very probably only adult worms were counted. Dorney (1963) found egg counts of cottontails due to 0. cuniculi to increase in the same 2 months. It seems very likely that this increase in the spring may be associated with the development of arrested worms at this time. Haemonchus placei

Bremner (1956) described the early parasitic development of H . placei in calves and noted that a small proportion of the larvae failed to develop beyond the early fourth stage. He suggested that either the worms developing most quickly produced some substance which inhibited the development of the slowest or else that a reaction on the part of the host was involved. About the same time Roberts (1957) also encountered the phenomenon of arrested development in experimentally infected calves. Calves which had received a single infection of 50000 larvae were found to harbour up to 1000 fourth stage Haemonchus 18 weeks later. When such animals were reinfected with up to half a million larvae, up to 17000 fourth stage larvae were present 10 weeks later but there was no obvious relationship between the number of larvae administered and the number of arrested worms recovered. Broadly similar results were obtained by Ross (1 963b). Roberts (personal communication, 1957) was of the opinion that large burdens of arrested H . placei could be built up in calves by infecting them repeatedly and that if adult worms were subsequently removed by anthelmintic treatment, all the arrested worms would develop and give rise to disease. A paper by Roberts and Keith (1959) is not infrequently quoted as containing evidence of this but it indicates, rather, that anthelmintic treatment, if repeated, retards the development of a resistance to the establishment of worms. When calves were infected either daily or at longer intervals, given anthelmintic treatment periodically and finally challenged with,5000 larvae, they were found to harbour more worms both mature and immature than calves which had not received anthelmintic treatment. Haemonchus contortus

The realization that the development of this much studied parasite could be arrested came surprisingly late. Field et al. (1 960) observed an increase in the Haemonchus egg count in the faeces of recently lambed ewes which had been housed, since 6 weeks before lambing, in conditions calculated to prevent accidental infection. They concluded that the increase must have been due to the resumed development of arrested larvae. In much of the literature, the arrested development of Haemonchus and a number of other species is linked

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with the increase in faecal egg count and in worm numbers seen in lactating animals. In the present review this subject is treated separately because some of the assumptions linking it with arrested development d o not appear to be warranted. The worms that take part in the so-called post-parturient rise are often arrested worms which have resumed their development, as Field et al. surmised, but it does not follow that it is necessarily events associated with parturition or lactation that trigger the resumption of development. Veglia (1915) and Stoll (1943) described the transitory and superficial histotrophic phase of H. contortus and showed that most of the worms returned to the surface after 40 h. Between 12 "/d and 16 72, however, penetrated deeper than the gastric pits. Malczewski (1970a) noted that 15 % of the larvae were still in the mucosa on the sixth day. But it is clear from the work of Blitz and Gibbs (1971) that arrested development is not inevitably associated with this abnormal histotrophic phase. Although larvae are arrested at the stage at which emergence from the mucosa normally occurs, the arrested larvae are not necessarily retained in the tissues. Blitz and Gibbs (1971a) found that arrested H. c o n f o r mwere at the early fourth stage and between 1.1 and 1.2 mm long. They were characterized by the presence in their gut cells of rod-shaped crystalline inclusions which did not disappear until the larvae resumed their development and completed the fourth stage. A possible connection between arrested development and host resistance, hinted at by Silverman and Patterson (1960), cannot be lightly dismissed. Christie and Brambell ( I 967) compared worms from a 7 day-old infection in susceptible lambs with worms of equal age from lambs which had been immunized with 10 doses, each of 20000 larvae, followed by anthelmintic treatment. The size distributions of worms from the two groups of lambs were entirely different. Worms from the immunized lambs showed little variation about a mode of 1.15 mm (cf. Blitz and Gibbs, 1971a) while those from susceptible lambs were considerably larger (see Fig. 8). Although in these very large infections the worms did not persist for long in the immunized animals,

Body length of worms (mrn)

FIG.8. Frequency distributions of the length of 7 day-old Huemonrhus contortus in immunized (protected) and susceptible (control) lambs. Reproduced from Christie and Brambell (1967).

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Christie and Brambell saw the failure of the worms to develop as essentially the same phenomenon as had been described by Field et al. (1960), by Gibbs (1964) and by Dineen et al. (1965b). Arrested development of H. contortus has also been demonstrated in lambs infected at regular intervals, either daily or weekly. Pradhan and Johnstone (1 972) found fourth stage larvae to accumulate in such lambs and argued that their development must be arrested because after a time their number exceeded the larvae that had been administered over the preceding 12 days. Development may also be arrested in very large infections. Silverman et al. (1970) noted a much greater variation in the rate of development of Haemonchus in large infections than in small ones. Christie (1970) compared a single infection of 2000 larvae with one of 1OOOO00 larvae and found that after 14 days all the worms of the large infection were arrested while the size distribution of worms from the small infection was bimodal, only a proportion of the worms having remained at the early fourth stage. It is of courvejust possible that by the fourteenth day all the developing worms had already been lost from the large infection. Malczewski (1970a), who infected lambs with 100000 larvae and found 17% of the worms to be arrested, noted that worm numbers rapidly declined, arrested larvae persisting rather longer than the developing worms. The connection between arrested development and either the size of the infection or the resistance of the host is a theme that recurs in the work of Dineen and his colleagues at the McMaster Institute. Dineen et al. (1965b) showed that when 3000 larvae were given to lambs as 30 daily doses of 100, a larger proportion became arrested than when they were all given on a single occasion. They discussed this difference in terms of whether the worms had passed a vulnerable stage in their development when an immune response developed. In a second experiment, Dineen and Wagland (1966) gave single infections of from 500 to 3000 larvae to groups of lambs and found that both the proportion of the worms still present after 56 days and the percentage that had not developed beyond the early fourth stage was the same in both groups. Animals which had been immunized in this way were now given anthelmintic treatment andchallenged with afurther 30001arvae.When they were slaughtered another 56 days later, rather more arrested larvae were present in the previously infected than in susceptible control lambs. After a second challenge the difference was even greater. This would be compelling evidence of an effect of previous infection on the inhibition of development, were it possible to exclude the possibility that arrested worms had persisted from a previous infection. The same difficulty arises also in the interpretation of the results of Wagland and Dineen (I 967) and of Donald et al. (1 969), although the general tendency of their results indicates that host resistance does play a part, though probably a small one, in causing development of H . contortus to be arrested. In experimental infections arrested development appears to be rather fickle in its appearance. Colglazier et af. (1969) found that a large proportion of Haemonchus in a mixed infection was still at the fourth stage after 35 days. When they attempted to repeat this observation in apparently identical circumstances, all the worms developed. There are many accounts of the occurrence of early fourth stage larvae of H .

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contortus in naturally infected sheep. Benz and Todd (1 969) examined groups of sheep taken off pasture in Wisconsin at different times of year and found that considerable numbers of fourth stage larvae were present between September and March. Turner and Wilson (1962) found all of a fairly large burden of H. contortus in a lamb slaughtered in Maryland in February to be immature and concluded, in view of the circumstances, that infection could not have occurred recently. A number of authors describe a seasonal pattern in burdens of arrested and adult Huemonchus in sheep. Viljoen (1964), Rossiter (1964) and Muller (1968) in South Africa, James and Johnstone (1967b) in Australia and Blitz and Gibbs (1972b), Ayalew and Gibbs (1973) and Ayalew et ul. (1973) in Canada have all found that in winter burdens consist predominantly of immature worms while in summer most of the worms are adults. Malczewski (1970b) in Poland and Hart (1964) in Nigeria have published similar findings concerning H . contortus in cattle. Using tracer lambs, Connan (1 97 1b) showed that larvae picked up from the pasture in the autumn tend to be arrested. Blitz and Gibbs (1972a) carried out a number of experiments to demonstrate that environmental factors acting on the free-living stages would cause subsequent parasitic development to be arrested. Artificial treatments proved rather ineffective, but exposing larvae on the pasture throughout September resulted in 96% of them being arrested when fed to lambs. It was concluded that exposure to decreasing temperature or photoperiod had to be of some duration to condition the larvae effectively. The decrease in the number of immature worms in sheep and the increase in the number of adults appears to be due to the resumed development of arrested worms. Ross and Gordon (1936) commented that lambing ewes and ewes with lambs at foot might be severely affected by Huemonchus infection and indeed Gibbs (1964) described an outbreak in which several deaths occurred in housed ewes which had just lambed or were just about to. The circumstances indicated that the worms had not been recently acquired and Gibbs formed the opinion that they had been picked up some months previously and had persisted in some quiescent form. Blitz and Gibbs (1 97 1 b) transplanted arrested H . contortus to worm-free pregnant ewes in January and deduced from the faecal egg counts that the worms did not develop until April after the ewes had lambed. Clearly the susceptibility of the new host was not a sufficient stimulus to induce the larvae to develop. This ensued either at a pre-determined time of year or in response to events associated with lambing. On the whole the evidence seems to favour the conclusion that development is linked to the time of year. Proctor and Gibbs (l968a) concluded that in ewes the timing and extent of that part of the so-called post-parturient rise in egg count that was attributable to H . contortus depended more on the time of year than on lambing. CvetcoviC et ul. (1971) have provided very striking evidence of this. Although Blitz and Gibbs (1972b) did not think it necessary to postulate that events associated with lambing triggered the development of arrested worms which probably occurred spontaneously, the possibility of such a connection cannot be entirely excluded. Connan ( I 968a) found that in a group of

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bred ewes all the worms had matured 1 month after lambing while in a comparable group of barren ewes killed at the same time, most of the worms were still immature. The evidence of O’Sullivan and Donald (1973) also suggests that arrested H . contortus persist longer in empty ewes. It may also be significant thatwhile Gibbs (1969) found that maturation of arrested worms took 1 month in pregnant and parturient ewes, Blitz and Gibbs (1972b) found the process to extend over 2 months in barren and virgin ewes. Gibbs (1969) has suggested that two steps are involved, first a stimulus depending on the time of year and mediated through the host’s endocrine system acts on the quiescent larvae, only few of which develop unless and until this stimulus is reinforced by humoral changes associated with parturition and lactation. It is the opinion of the present writer that lactation affects the loss of adult worms rather than the development of arrested larvae. If this is so, the results of Connan (1968a) and of Blitz and Gibbs (1972b) would imply that, even if the development of arrested larvae is prompted by stimuli associated with the time of year, the process can be delayed if adult worms are allowed to accumulate. The loss of adult worms from sheep appears to be very rapid in most circumstances. Donald (personal communication, 1970) has calculated a mean life span of the order of 25 days and Whitlock et al. (1972) have observed a decline in the burdens of sheep removed from pasture at a rate between 5 % and 20 per day. Clearly, if arrested larvae are not subject to this loss, and in spite of the observations of Malczewski (1970a) it appears that they are not, then where the free-living stages are incapable of surviving through the winter on the pasture, arrested development must represent an important means whereby the parasite can be carried on from one season to the next. Tetley (1959) in New Zealand pointed to the importance of worms that had over-wintered in the ewe as the source of pasture infestations in the summer and Connan (1971b) thinks that, in Eastern England at least, worms which have persisted through the winter as arrested forms in the ewes, are the source of the infective larvae that are the cause of disease in lambs and which, if picked up in the autumn, become arrested and so persist through the next winter. In temperate climates H. contortus appears to be monocyclic and Ayalew and Gibbs (1973) have commented on the very restricted season during which larvae can develop from egg to adult without interruption. Blitz and Gibbs (1972a) saw arrested development as a phenomenon akin to diapause and believed other causes to be incidental. They apparently visualize that the evolution of this mechanism has resulted in a point of potential discontinuity in development and that this happens to be susceptible to other factors. The idea is avaluable one but it is open to question whether their distinction between “specific inhibition”, denoting a response to seasonal factors, and “non-specific inhibition”, relating to all the rest, is particularly useful.

<:

IV. TRICHONEMATIDAE Trichonema spp. Trichonema spp. were the subject of an interesting and widely discussed observation by Gibson (1953). A number of naturally infected horses were

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housed and treated with the anthelmintic phenothiazine. Adult Trichonema were passed in the dung and the faecal egg count fell to zero. After some weeks egg counts rose to near their former level. The horses were given phenothiazine again, and again adult worms appeared in the dung and Trichonema eggs disappeared, only to reappear after several weeks. This sequence of events was repeated a number of times in exactly the same way except that the number of worms passed in the faeces after anthelmintic treatment and the faecal egg count both tended to decrease from one occasion to the next. Gibson considered three possible explanations. The results of separate tests seemed to rule out the possibility that the action of the drug on adult worms was incomplete. Accidental infection of the horses was not considered likely because very stringent measures were taken to avoid it and because this explanation would demand that the rate of accidental infection decreased steadily throughout the experiment. Gibson therefore suggested that worms inhibited in their development and insusceptible to the action of phenothiazine were the source of the adult worms which seemed to reappear after anthelmintic treatment. He proposed that the development of the arrested larvae was in some way prevented by the presence of adult worms and that the removal of these by anthelmintic treatment permitted another batch to grow to maturity. It is possible that this explanation is the correct one but an alternative is available which does not postulate a specific effect of adult worms in inhibiting development. If a population of Trichonema spp. were in a state of dynamic equilibrium, adult worms being lost constantly and replaced by the development of small numbers out of a large reservoir of arrested larvae, then periodic anthelmintic treatment would result in precisely the phenomena observed by Gibson (1953). Oesophagostomum columbianum

Curtice (1890) noticed large numbers of nodules in the intestinal wall of sheep and found each to contain a larval worm 3-4 mm long. He experienced considerable difficulty in relating these larvae to adult worms-a circumstance that suggests a marked discontinuity of size and stage-but finally linked them to a hitherto unidentified species which he called Oesophagostomum columbianum. The nodules and their formation were described by Theiler (1 921). Eosinophil leucocytesaccumulate around the larvae which have penetrated into the mucosa. The larvae may remain in the resulting green caseous nodules for some time but finally the nodules become calcified and at this stage live larvae can no longer be demonstrated in them. Veglia (1924) found that the larvae return to the intestinal lumen 6-8 days after infection and immediately after completing the third ecdysis but “external factors and age of the animals may delay exit of the larvae”. Veglia (1928) reported that in lambs a substantial proportion of the nodules still contained larvae 2 months after infection and that in adult sheep live larvae could be found in the nodules for up to 6 months. The larvae continued to emerge from the nodules but in resistant adult sheep they failed to persist in the gut lumen and were lost in the faeces.

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Fourie (1936) distinguished between a normal and an abnormal pattern of development. Between 24 and 96 h after infection the larvae penetrated the mucosa and encysted close to the muscularis mucosae. About the 5th day they performed what Fourie termed a second parasitic migration and returned to the lumen. A proportion behaved differently and penetrated the muscularis mucosae with the formation of a fibrous capsule. Either the larvae emerged from these capsules at a later time (the result, according to Fourie, of a fortunate accident), or they died. A few penetrated to the peritonea1 cavity and finally perished in other organs. Monnig (1938) was of the opinion that a tissue reaction and nodule formation occurred only in resistant sheep but this was almost certainly an over-simplification. This early work on Oe. columbianum had a marked influence on the development ofideas on the subject ofarrested development which came to be associated with a tissue reaction occurring in resistant but not in susceptible hosts and with abnormal migratory behaviour. The possibility that such a connection might be fortuitous was not entertained until recently. It is now evident, however, that nodule formation occurs in susceptible as well as in resistant hosts and that it is not necessarily associated with arrested development of the larvae. Shelton and Griffiths (1967) found that typically the larvae return to the lumen of the intestine on the 5th day after infection and that a nodule forms around the products of the third moult. Such nodules disappear by the 25th day. In a second or subsequent infection the number of fourth stage larvae which fail to return to the intestinal lumen is very much greater than in a first infection and the number of eosinophils that accumulate around such larvae is so great that a very long lasting nodule then results. Shelton and Griffiths (1968) provided further evidence to show that the number of larvae remaining in the intestinal wall was greater in sheep that had experienced infection previously. Sarles (1944) had come to a similar conclusion when he found that in 8month-old lambs infected daily the proportion that gave rise to nodules increased with the number of larvae administered. The relevant changes in the host may be local in character. Dobson (1966) showed that the area over which caseous nodules occurred increased with successive infections. The innate resistance of an unsuitable host may also prevent development. Herlich (1970) noticed that in the calf, Oe. columbianum did not proceed beyond the fourth stage but remained alive in nodules for several weeks. The sex of the host may also be involved. Dobson (1964) showed that a primary infection in male lambs yielded more worms than one in female lambs but that there were more nodules in female lambs than in males. This effect appears to be restricted to the development of the worms, for Bawden (1969b) demonstrated that Oe. columbianum is more prolific in female than in male lambs. If arrested Oe. columbianum could resume their development in appreciable numbers, the phenomenon of arrested development could play a significant part in the epidemiology of oesophagostomiasis. Gordon (1948) was in no doubt that they could and did do so. According to him, “the third stage larva 11

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enters the bowel wall and remains there for periods ranging from a few days to several months before returning to the lumen of the colon to resume its development”. Gordon (1949) derived evidence in support of this from an experiment in which he housed infected sheep to prevent further infection and treated them with phenothiazine after 5, 9 and 12 months. On each occasion Oe. columbianum of various sizes were expelled in the faeces and on each occasion the number was slightly smaller. Because phenothiazine at the dosage employed had been shown to be capable of expelling all adult Oe. columbianum, Gordon argued that worms must continue to emerge from the mucosa for at least a year. Gordon (1952) did not fail to see the significance of this. He saw the larvae in the bowel wall as “a kind of resting stage which may serve to carry over the parasite from one favourable season to another. The young Oesophagostomum may overwinter in the bowel wall and emerge in the spring when the weather is suitable for the development of the free-living stages”. Surprisingly little has been done in the ensuing 20 years to confirm and amplify this important suggestion. Even seasonal changes in populations of adult and arrested worms have scarcely been investigated. Sarles (1944) had noticed that considerable numbers of Oesophagostomum nodules were commonly encountered in the autumn in lambs in American grassland flocks but this does not necessarily mean that many arrested worms were present. Rossiter (1964) in the Eastern Province of South Africa found that fourth stage Oesophagostomumlarvae in a group of wethers were numerous from December to July and few in number during the rest of the year. The number of adult worms showed the opposite tendency. Meanwhile Viljoen (1964) in a similar series of observations in the Eastern Karoo found that the number of fourth stage larvae was high in winter and low in summer while the number of adult worms showed a similar fluctuation but just 2 months behind. No doubt, as further information becomes available, the phenomenon and its significance will be better understood. Oesophagostomum venulosum

It appears unlikely on general grounds that nodule formation is a necessary accompaniment to arrested development. It is a matter of some interest, therefore, whether arrested development occurs in infections of Oe. venulosum in which nodules do not seem to have been either observed or reported. Published evidence is, however, remarkably scanty. The observations of Goldberg (1951) indicate that the life history of Oe. venulosum is similar to that of other species of Oesophagostomum. The third stage larvae encyst in the mucosa of the small intestine on the 3rd day and reemerge on the 4th day, the third ecdysis being completed just after emergence. By the 5th day most of the larvae are already in the large intestine. Goldberg reports finding a small number of fourth stage larvae in the large intestine of a lamb 30 days after experimental infection but does not specify their precise stage of development. In a goat, Goldberg (1952) found a large proportion of the worms to be at the fourth stage 39 days after reinfection. There is, however, no report of an extended histotrophic phase in the small intestine.

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Oesophagostomum radiatum Oe. radiatum causes the formation of nodules in the ileum ofcalves and spends a variable time in them. Anantaraman (1942) was of the opinion that no development beyond the early fourth stage occurs unless the larva emerges from its nodule. Normally this happens on the 8th day and the larvae move to the caecum shortly after emergence but Anantaraman found some live larvae still in nodules in the ileum 2 months after experimental infection. Goldberg (1 959) found fourth stage larvae in the small intestine of calves 40 days after experimental infection and in the large intestine after 71 days. Similarly, Roberts et al. (1962) found fourth stage larvae in experimentally infected calves 11 weeks after the last infection. The largest number of arrested worms was found in calves which had received infective larvae daily before receiving a challenge infection. (The evidence suggests that the challenge infection failed to become established.) While this need not mean that the inhibition of development was caused by the resistance of the host, an observation by Douvres (1960) may suggest that it can be. Douvres found that the growth in vitro of Oe. radiatum is inhibited by the inclusion in the medium of serum or extracts of large intestine from resistant, but not from worm-free calves.

Oesophagostomum spp. of pigs

In describing the parasitic development of Oesophagostomum longicaudum, Spindler (1933) did not observe that the larvae could persist in the mucosa beyond the normal period. Kotlhn (1948), however, was led to the conclusion that what he called an irregular histotrophic phase did occur. Having penetrated into the mucosa, some larvae persisted there for a prolonged period without developing further. Kotlan believed that such larvae finally perished. It was also noticed that even those larvae which did develop might remain in the mucosa for a somewhat variable period. Shorb and Shalkop (1959) studied experimental infections of Oe. quadrispinulatum and found that, in addition to worms that developed normally, what they took to be third and fourth stage larvae were present in considerable numbers for at least 65 days. Like K o t l h , Shorb and Shalkop believed that these larvae would never emerge from the nodules and would perish. Two further observations by Kotlin (1948) are of interest: first that, in experimental infections of Oe. dentatum, a greater proportion of the larvae undergo an irregular histotrophic phase in heavy infections than in light; and second that in the rabbit, an unnatural host, the number arrested in their development is greater than in the pig. This second observation is given particular significance by the work of Jacbos et al. (1968,1971) who showed that in guinea pigs and rats the larvae of Oesophagostomum spp. from pigs would persist in the mucosa of the large intestine for considerable periods and that these dormant larvae resumed their development when fed to pigs. Thus, like some other nematodes Oesophugostomum spp. show a tendency to be arrested in abnormal hosts which can then act as paratenic hosts. The parallel between arrested development as defined

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in the introduction to this review and the ability of nematodes to suspend their development in transport hosts is discussed further in a later section. Evidence that larvae arrested in the pig are also capable of resuming their development is largely indirect and concerns the phenomenon known as the peri-parturient rise which is dealt with in the next section.

V. THESPRINGRISE

Worms that have been arrested in their development but which have subsequently grown to maturity are frequently involved in a phenomenon, or group of phenomena, variously known as the spring rise, the post parturient rise or, more recently, the lactation rise. This has come to be so firmly linked, in the literature, with arrested development, the assumption being made that events associated with parturition or lactation trigger the development of arrested larvae, that the matter must be examined here, even though it is the opinion of the writer that a rather different process underlies the rise. If the evidence is still incomplete and the position has been, and to some extent still is, confused, this is largely because unsuitable techniques have been employed. Most of the work has been based on faecal egg counts and the unwarranted deductions that are so often drawn from them. The story starts with a report by Zavadovskii and Zviagintsev (1933) on a seasonal fluctuation in the number of Nematodirus eggs in the faeces of camels, a fluctuation that was quite possibly due to changes in the number of infective larvae available to them. This report prompted Taylor (1935) to publish observations on what appeared to be a rather different phenomenon in ewes. Taylor had made monthly egg counts from May 1932 until August 1933 on composite samples of faeces from 12 ewes which lambed in February and March and were housed during these 2 months. The counts rose from March, to reach a peak in June, and then declined, most of the decrease having occurred by August. Because the conditions in which the ewes were kept in the early spring precluded accidental infection with strongylate nematodes, Taylor concluded that this very striking increase in egg count was not attributable to newly acquired worms but to an increase in the egg output of the worms which, he thought, might be occasioned by a loss of resistance associated with the strain of lambing and lactation. This interesting observation appears to h v e been overlooked for the next decade. Certainly, no effort was made to follow it up. In 1944 Hawkins et al. published observations on the egg counts of ewes which were housed from November until late May, and lambed in March. The counts were low during the winter; the count due to Ostertagia spp. rose in April and May while a peak due to Haemonchus occurred a little later. Even though the lambs did not become infected while in the barn, Hawkins et al. believed that the ewes had picked up larvae there. Seghetti and Marsh (1945) made somewhat similar observations on ewes which were not only kept in conditions in which re-infection was unlikely but which were also given anthelmintic treatment 6 weeks and 1 week before lambing. In spite of this, an increase in worm egg output was observed after

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lambing. Hawkins and de Freitas (1947) also observed a spring increase in the egg counts of housed ewes and suggested that it was due partly to new infection and partly to an increased rate of ovulation. Meanwhile, D. 0. Morgan and his collaborators had embarked on a study of seasonal changes in the worm burdens of Scottish hill sheep which, they felt, should present a simple situation uncomplicated by changes of pasture or by supplementary feeding. They observed a marked seasonal fluctuation in the faecal egg counts of ewes, with a peak in April and May and minimal counts in January and February. Yearling sheep also showed an increase in early spring. Although Harbour et al. (1946) had shown that infestations on the herbage of hill pastures remained at a fairly high level at least until Christmas, Morgan and Sloan (1947) thought that only few larvae could be available to sheep in the late winter. They believed that the spring increase in egg count might be due to one of three causes: (a) that a larger proportion than usual of the worms picked up succeeded in establishing themselves because of lowered host resistance; (b) that the fecundity of the worms was increased for the same reason; or (c) that there was a natural seasonal rhythm of egg-laying by the worms. Cushnie and White (1948) who had made similar observations during the hard winter of 1946-1947 also felt that few larvae could have been picked up before the spring, because the pastures lay deep in snow and the ewes had subsisted on silage. Morgan et al. (1950) continued the systematic examination of faeces samples from an increasing number of hefts, with basically similar results. Within any year, there was a marked similarity in the egg count pattern of different hefts, but there were some differences between years. Thus, a rather higher peak in the spring of 1947 might be associated, according to Morgan et al., with the severe preceding winter. This theme of stress, associated with poor nutrition and exposure to severe conditions, reducing host resistance features, is also to be found in Paver et al. (1955) who noticed unusually high peaks of egg count in 1947 and 1951 following severe winters. White and Cushnie (1952) were not, however, able to reduce the spring rise by supplementary feeding. Morgan et af. (1951) continued their investigations by purchasing an entire flock of sheep and slaughtering groups of 50 at the end of August, in January, a week before lambing began in April and at the beginning of June when egg output was at its peak. Their worm counts showed conclusively that the spring rise was associated with an increase in worm numbers which began in February and was on a scale which made it quite unnecessary to postulate an increase in the fecundity of the worms. Ostertagia spp., Trichostrongylus axei and, to a lesser extent, Cooperia curticeiwere involved. Because the development of arrested larvae was beginning to be discussed about this time as a possible cause of the spring rise, Morgan et al. (1951) slaughtered two ewes every fortnight during the first 6 months of the year and examined the mucosae with the aid of a compressorium. Because they failed to find significant numbers of immature worms during the winter, they came to the conclusion that the worms involved in the spring rise were picked up from the pasture during the late winter and early spring.

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Wilson et al. (1953) also slaughtered hill sheep periodically, and found that worm numbers increased in spring, in wethers as well as in ewes. Haemonchus contortus, Ostertagia spp. and Trichostrongylus spp. were involved. Wilson et al. also failed to find arrested worms during the winter and found the absence of any H . contortus particularly hard to understand since it was known that this species does not survive at all well on the pasture. According to Parnell (personal communication, 1971) the possibility cannot be excluded that the methods of detection employed by Morgan et al. (1951) and by Wilson et al. (1953) were less than fully effective. Morgan et al. (1951) had noticed a tendency for the spring rise to occur earliest in the first ewes to lamb and to be less marked in barren ewes. Parnell et al. (1954), working with lowland flocks, also noticed that the onset of the rise in individual sheep was closely correlated with the date of parturition. In the same year Crofton (1954) published the results of his study of faecal egg counts of ewes in lowland flocks, which clearly demonstrated a close correlation between the date of lambing and of what now came to be called the postparturient rise. Crofton claimed that the peak egg count of individual ewes was much higher than the peak of mean counts for the whole flock, and was of much shorter duration. In individual ewes the rise lasted for 2 weeks, while for the flock as a whole it lasted as long as the lambing period. The rise in individual ewes occurred very accurately between 6 and 8 weeks after lambing. It could occur from February to June in the same flock and whether late or early, its extent was about the same. Crofton considered that the rise could hardly be due to the uptake of new infection because it was so steep, and he did not think many infective larvae were available on the pasture in late winter and early spring. Hence, arrested worms must be the source of the increased worm burden and he proposed the theory that the post-parturient rise was due to a progressive loss of host resistance which he divided into three stages. While the ewe was in the resistant state the worms that it ingested were nearly all eliminated without development (i.e. failed to become established). Loss of resistance, due to a decreased challenge, led to the accumulative stage during which more of the worms ingested became established but their development was inhibited. During the final stage, the loss of resistance, such infective larvae as were ingested became established, and both they and the arrested worms which had accumulated were able to develop. If the increase in worm numbers was due to a loss of resistance, the termination of the post-parturient rise must be due to an increase in resistance. Crofton noticed that the fall in egg count coincided with a sharp decrease in the ewes’ milk yield.and suggested that the decline in resistance, to which he attributed the rise, was accelerated by the strain of milk production. In barren ewes and in wethers, there was also a rise in the spring but it was very much less marked than in ewes that lambed. In attributing the spring rise in barren ewes and wethers to a “basic rhythm of reproduction” Crofton hinted at a mechanism involving the host’s endocrine system but he questioned whether the rise seen in unbred animals could be identical with the postparturient rise. This doubt arose also from his observations on ewes lambing

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in the autumn (Crofton, 1958), for while wethers and barren ewes showed a small increase in egg count in spring only, ewes that lambed in the autumn manifested a post-parturient rise in autumn, and in ewes that lambed twice a year, two rises were seen. These findings were confirmed by Downey (1968). The view current in the 1950s and 1960s was that the post-parturient rise was explicable in terms of immune phenomena. Soulsby (1957a) studied levels of circulating antibody in a flock of ewes and claimed that the spring rise was preceded by a fall in titre (which in his published data is none too obvious to the untrained eye) and that the fall in egg output by which the phenomenon was terminated was accompanied by a rise in titre. On the basis of these observations he erected what could be regarded as the classical theory (Soulsby, 1957a, 1961a), namely, that limited exposure to infection during the winter, possibly in combination with deficient nutrition, leads to a loss of immunity resulting in reactivation of dormant larvae and/or susceptibility to the establishment of new worms. The termination of the spring rise, according to this theory, is in the nature of a flock “self-cure” of the kind described by Stoll(l929) and Stewart (1950) and, as such, precipitated by an increased uptake of worms and followed by a state of resistance to the establishment of new worms. In the years that followed, this theory became increasingly difficult to sustain. Soulsby (1966) showed that the egg output of ewes could not be increased by the administration of corticosteroids and while he reported that the akylating agent chlorambucil did produce this effect, Brunsdon (1966~)and Gibbs (1969) were unable to confirm this. As a legacy from the classical theory, the notion that the termination of the spring rise is identical with the self-cure mechanism and depends on a threshold of antigenic stimulation, dies very hard (see for example, Brunsdon, 1966b; Arundel and Ford, 1969; and Ayalew and Gibbs, 1973). That it is not necessary to invoke this mechanism to explain the observed phenomena will be argued below. The observations and conclusions of Crofton (1954, 1958) have been amplified and modified. Brunsdon (1964a) and Dunsmore (1965) failed to confirm that the rise in individual ewes was short-lived; they found it to be of much the same form and duration as that of the flock as a whole. While Condy (1961) and Brunsdon (1964a) agreed that the peak occurred 6-8 weeks after lambing, Spedding and Brown (1956) and Large et al. (1959) observed it to coincide with, or immediately follow, lambing. Herweijer (1965) saw the peak 2-3 weeks after lambing and Dunsmore (1965) and Wensvoort (1961) saw it 4 weeks after lambing. Brunsdon (1964a) and Dunsmore (1965) noted considerable individual variation in the timing of the peak which might even precede parturition. Large et al. (1959) found that egg counts began to increase well in advance of lambing and Brunsdon (1 966b) observed them to increase gradually from tupping time. Most authors, among themBrunsdon (1964a), Field et al. (1960)and Spedding and Brown (1956) agree that in wethers, unmated ewes or ewes that abort, there is a spring rise similar to that seen in parturient ewes but considerably smaller. Dunsmore (1969, Arundel and Ford (1969) and Brunsdon (1966b),

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however, observed no rise at all in such animals. But it appears that on some occasions no rise occurs even in parturient ewes (Lewis and Stauber, 1969) and that it may fail to appear in some individuals (Connan, 1967a). That the post-parturient rise can occur in ewes that have been withheld from infection for some weeks or even months before lambing, has been shown by Spedding and Brown (1956), Field et al. (1960), Brunsdon (1966) and, in yearling sheep, by Naerland (1952). But if the ewes had been withheld from infection since the previous summer the post-parturient rise was greatly reduced (Spedding and Brown, 1956).It has also been shown that anthelmintic treatment of the ewes, before lambing, does not prevent the rise (Large ~t al., 1959; Dunsmore, 1965; Field et al., 1960). In view of these observations it became the general consensus of opinion that the post-parturient rise was attributable to arrested worms which had been stimulated to resume their development by events associated with lambing. In a careful analysis of the evidence, Dunsmore (1965) argued convincingly that the stimulus must be endocrine in nature. Evidence soon began to accumulate, however, which indicated that the phenomenon was more complex than had been envisaged. First it became clear that lactation, rather than parturition, exerted a crucial influence on the rise in egg count. Ewes whose lambs are weaned at, or shortly after, birth show a negligible increase in egg output (Connan, 1968b; Jansen, 1968; Arundel and Ford, 1969; OSullivan and Donald, 1970; Brunsdon and Vlassoff, 1971a). The evidence is so striking that a close connection between lactation and the post-parturient rise cannot be doubted. In the words of Brunsdon and Vlassoff (1971a) "the fact that failure to suckle lambs prevents the occurrence of the post-parturient rise appears to preclude the effects of hormones of pregnancy and parturition and points to the primary importance. . . of changes that occur during lactation." This may oversimplify the matter slightly, for not only can the increase in egg count begin before lambing but, as shown by Brunsdon (1964, 1967, 1970) and others, the post-parturient rise is not infrequently terminated before the lambs are weaned. A number of attempts have been made to induce an increase in the faecal egg count of virgin ewes by the administration of prolactin, or by stimulating its release by means of stilboestrol or acetyl promazine, but without very striking effect, Salisbury and Arundel (1970), Gibbs (1969) and Blitz and Gibbs (1972b) achieving only a limited increase in the size of the spring rise and failing to alter its timing. Secondly, evidencebegan to accumulate that the worms in the post-parturient rise are not always arrested worms. Connan (1968a),who followed changes in populations of Ostertagia spp. in ewes, of which some were housed and some on pasture, concluded that in those on pasture there was a real increase in worm numbers during the post-parturient period. While in the housed ewes the arrested larvae had all developed by May, i.e. 2 months after lambing, in the pastured ewes immature worms continued to be present. Connan concluded that lactation not only prompted the development of arrested worms, but also reduced the host's resistance to the establishment of newly acquired larvae. He also deduced that there was an increase in the fecundity of the worms. While

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some ewes with significant burdens had negligible egg counts during the winter, the increase in egg count after parturition was proportionately greater than the increase in the number of mature worms. A somewhat similar study conducted in Australia where H . conrortus, Ostertagia spp. and Trichostrongylus colubriformis were the most common species, led O’Sullivan and Donald (1 970) to much the same conclusions. They gave anthelmintic treatment to ewes, 3 months before lambing, and verified that no worms, either mature or immature, remained. These ewes, together with undosed controls, now remained on the pasture and the same postparturient rise was seen in all. In the conditions of this trial the development of arrested larvae evidently made little or no contribution to the rise which must therefore be attributed to new worms. O’Sullivan and Donald suggested that while worms become established in lactating ewes, dry ewes are refractory, but it is not easy to derive this from the evidence that they presented because, as measured by tracer lambs, larvae ceased to be available on the pasture while the ewes were in milk. These publications lent support to the view that lactation, or changes associated with it, reduced the resistance of the host and that this permitted newly acquired worms to become established, arrested worms to resume their development and adult females to ovulate without restraint. There was a little discussion as to the specificity of this loss of resistance. While Connan (1968b) thought that the post-parturient rise involved whatever nematodes happened to be present, Brunsdon (1970) disagreed on the grounds that the rise attributable to different species was of rather different form and timing and that some species that were available were not involved (Brunsdon and Vlassoff, 1971b). It does not seem likely that this theory, envisaging a loss of resistance and of its several effects on the establishment, development and reproduction of worms, will survive. The results of Connan (1968a) and of O’Sullivan and Donald (1 970) do not unequivocally demonstrate these effects but they do strongly suggest that mature worms are lost less rapidly from lactating than from dry ewes. The results of Connan (1971~)who recovered more worms towards the end of lactation from ewes on a low plane of nutrition than from well fed ewes lend themselves to the same interpretation. It may be significant that Connan (1970) and Dineen and Kelley (1972), who studied the effect of lactation in rats on their response to infection with Nippostrongylus brasilensis, demonstrated an effect on the expulsion of worms but not on other manifestations of resistance (Connan, 1972). The post-parturient rise will almost certainly prove explicable in terms of an effect of lactation on the persistence and activity of adult worms; the precipitating cause of the resumed development of arrested forms will be found elsewhere. The very interesting and significant observation by Morgan Parnell and Rayski (1951) that an increase in worm numbers to a high peak in March occurred in yearling sheep as well as in ewes should have made it obvious that the development of arrested worms need not be connected with lambing at all. While this appears to have been overlooked, other, less direct, indications have not. Thus Jansen (1968) and Herweijer (1969) noticed a change, with time, in

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the species composition of the eggs passed during the post-parturient rise. Of more obvious significance is the finding that, especially where H . contortus is the dominant species, the magnitude of the rise is greatly affected by the date of lambing. Brunsdon (1967) showed that ewes which lambed 2 months later than the rest of the flock showed no post-parturient rise, and Salisbury and Arundel (1970) found that ewes lambing early had a smaller and more transitory rise. Cvetkovid et al. (1971) showed that there was an optimum time of lambing for the occurrence of a maximal post-parturient rise due to H. contortus. Moreover the spring rise in unmated ewes occurred at this precise time as did a slight rise in ewes that had not yet lambed. Proctor and Gibbs (1968b) and Ayalew and Gibbs (1973) in whose ewes H . contortus was also the dominant species observed that the post-parturient rise tended to occur at a certain time of year and was not related to the time of lambing. These results suggest that the development of different species may occur at slightly different times of year, or possibly in response to different stimuli, and that while arrested H . contortus develop during a short period, the development of Ostertagia circumcincta, the dominant species in Crofton’s (1954) ewes, is spread over a much longer period. Blitz and Gibbs (1972b) have expressed the view that the development of arrested H . contortus occurs at the same time in all sheep, possibly in response to some seasonal stimulus mediated by a neurosecretory mechanism of the host (Gibbs, 1969), and while in dry sheep they are subject to what Blitz and Gibbs term “some protracted form of self-cure”, this does not operate during lactation, so that the worms persist. An explanation of this kind will almost certainly prove to be the correct one but the part played by the self-cure mechanism in the post-parturient rise and its termination requires some comment. It is generally assumed that because worm numbers decrease at the end of lactation, this must necessarily be a new event, the active expulsion of worms by the mechanism visualized by Stewart (1950). This assumption is unjustified. Like Ostertagia ostertagi in cattle (Michel, 1963, 1969a, 1970a), Nematodirus spathiger’in sheep (Dineen et al., 196Sa) or Haemonchus contortus in sheep (Whitlock et al., 1972), it may be expected that populations of most other strongylate nematodes tend to be in a state of dynamic equilibrium and to turn over rapidly. There are grounds for believing that in old hosts the rate of turnover is particularly rapid. In the opinion of the writer, all the features of the post-parturient rise can be explained by postulating that the loss of worms, to which all populations are subject, is suspended in the lactating animal. Whatever their source, whether derived from arrested worms or from larvae newly acquired from the pasture, worms which reach maturity during lactation persist. Meanwhile the life span of adult worms in the non-lactating animal is not sufficiently long either for large burdens to occur or for the worms to reach an age at which they become prolific egg layers. At the end of lactation, worms are again lost at the same rateas before and the population inevitably declines. In so far as the post-parturient rise was due to arrested worms, this source of supply will have been largely exhausted and the

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worm burden will fall to the point at which the loss is balanced by recruitment in the form of larvae acquired from the pasture. It is not impossible that, during lactation, restraint on the egg output of worms is relaxed, but the observation of Connan (1968a) who measured an increase in egg output per female and of Brunsdon and Vlassoff (1971a) who observed a decrease in the number of mature females without eggs in their uteri, can be explained otherwise. If a rapid turnover of worms occurs, most worms will be too young to be ovulating freely, and if the loss of worms is suspended, the worms will be older and more prolific. Events surrounding the post-parturient rise may therefore be visualized as follows. During the autumn and early winter, a proportion of the infective larvae picked up by the ewes will become established and while some develop, the remainder will be arrested. The life of the worms that develop will be short so that their numbers will remain small, but the arrested worms will accumulate. In the spring, the arrested worms will resume their development, H . contortus over a fairly short period, Ostertagia spp. over a rather longer period. In barren ewes and wethers, worm numbers will increase to the point where this new supply of developing worms is balanced by an increased loss. This increase in worm numbers may be reflected in an increased faecal egg count. In the lactating ewe, the loss of worms ceases and adult worms therefore accumulate, being derived from arrested worms, new worms from the pasture, or both. The worms will persist to a greater age and will therefore be more prolific. Where H. contortus is the dominant species and the ewes lamb before the arrested worms have developed, the post-parturient rise will be small or it will be delayed; in ewes lambing after the arrested worms have developed and have been lost, the post-parturient rise will be small or absent. Where the dominant species persists well on the pasture through the winter so that new infection is available in the spring, or where arrested forms develop over a long period, there will be a close correlation between the date of lambing and the post-parturient rise. When the lambs are weaned, the normal loss is resumed (in poorly nourished animals or animals in poor condition this may be delayed) and worm numbers and faecal egg count fall to the level appropriate to the rate of recruitment. Accordingly, the decrease is characteristically of logarithmic form (see for example, Spedding, 1956). The post-parturient rise observed by Crofton (1958), in ewes lambing in autumn, was presumably attributable to words picked up from the pasture at that time and it is to be expected that if his ewes had been housed, no rise would have occurred. Similarly the observations of Southcott et al. (1972) on the effect of the date of lambing on the post-parturient rise suggest that in those of their ewes which lambed in summer, the rise was due to larvae picked up from the pasture, while in those lambing in winter the worms either largely or wholly derived from arrested forms. Mention must however be made of an observation by Gibbs (1969) who noticed a slight increase in the egg count of empty ewes at the same time as the post-parturient rise in autumn lambing ewes. It is likely that there was an increase in the number of larvae available on the pasture at just this time.

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The worm eggs passed during the course of the post-parturient rise play an important part in the epidemiology of parasitic gastroenteritis. As early as 1933 Schmid, who noticed seasonal changes in the egg output of ewes, recognized the part they played as a source of infection for the lambs. Hawkins et al. (1944) considered that H. contortus and Oe. columbianum were carried on from year to year by the breeding flock, rather than on the pasture. I t was clear to Morgan and Sloan (1947) that on account of the spring rise, contamination of the pasture was “greatest at the time of year when the lambs are very young and therefore most susceptible to worms”. Crofton (1958) also saw the post-parturient rise as a device for synchronizing the availability of infective stages with the occurrence of susceptible hosts but he thought that the ewes only contributed the initial infection which was then built up by the lambs through several worm generations. Thinking along a similar line, Brunsdon (1964) suggested that the magnitude of the post-parturient rise might determine the speed at which worm burdens were built up in the lambs. It was shown by Heath and Michel(l969) and by Boag and Thomas (1971) that where residual infestations on the pasture in spring were small, the postparturient rise was the source of virtually all the larvae picked up by the lambs to the end of August and therefore the cause of most outbreaks of parasitic gastroenteritis. A number of attempts to control nematode parasitism in lambs, by anthelmintic treatment of the ewes, have therefore been successful (Leiper, 1951 ; Nunns et al., 1965; Brunsdon, 1966a) but where an appreciable infestation has overwintered on the pasture this approach is much less effective (Arundel and Ford, 1969; Thomas and Boag, 1971; Donnelly et al., 1972). The occurrence of a spring rise or post-parturient rise is not restricted to sheep. Hansen and Shivnani (1956) showed that the number of eggs of Haemonchus, Cooperia, Ostertagia, Bunostomum, Nematodirus and Trichostrongylus in the faeces of yearling cattle rose in April to reach a peak a t the end of that month. In the light of evidence discussed in earlier sections it is probable that the increase was due to the development of arrested worms. Corticelli and Lai (1960) reported an increase in the faecal egg counts of cows in Sardinia either coincident with, or immediately following calving, and Michel et al. (1972a) observed a similar increase, though of smaller extent, in beef cows calving at all times of year. Unpublished observations by the same authors indicate that in dairy heifers a peak of worm egg output occurs a few days after calving. Observations by Schatzle (1964) are also of interest. He followed the faecal egg counts of ungulates of 20 different species, and maintained in a variety of conditions, in the Zoological Gardens of Munich, and found that almost without exception, they rose in March and declined in July. Since the mean ambient temperature did not exceed 4°C until April, it seemed improbable that these increased counts could be due to an increased uptake of infective larvae. Dunn (1965) noticed that in roe deer in Scotland, burdens of nematodes of a number of species were at their highest between March and June. In pigs, worm egg counts due to Oesophagostomum spp. and Hyostronglyus rubidus also show a marked increase during lactation. The first report of the phenomenon by Connan (under the name of Barnett, 1966) was followed by a similar one by Jacobs (1966). Both authors found that faecal egg counts (which

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were chiefly attributable to Oesophagostomum spp.) of sows were low in the first half of pregnancy, tended to rise during the second half, reached a maximum during lactation and fell suddenly, when the piglets were weaned, to the level characteristic of early pregnancy. This periparturient rise in egg output was associated with changes in population structure; there were more adult worms during lactation and they were larger and more prolific. Jacobs was reluctant to regard this phenomenon as analogous with the post-parturient rise in sheep on the possibly inadequate grounds that while trichostrongylid worms were dominant in the rise in sheep, Hyostrongylus played a minor role in sows. This impression may merely be due to the greater prevalence and greater prolificacy of Oesophagostomum spp. and indeed Connan (1967b) found that numbers of Hyostrongylusin pregnant sows increased at the same time as those of Oesophagostomum spp. He argued that since parasites as diverse as these were involved, the rise must be due, as in the ewe, to a non-specific change in the immune status of the host. Connan (1967b) found large burdens of early fourth stage Hyostrongylus larvae in sows and considered them to be arrested because of their uniformity and the manner in which the sows had been managed. He noted that a periparturient rise could occur in sows that had negligible access to new infection and considered that the worms involved were arrested larvae resuming their development. Like Jacobs (1966), Connan found that the periparturient rise could be prematurely terminated by weaning piglets and that if the piglets were separated from the sow immediately after birth, the rise was entirely suppressed. These points were confirmed by Barth (1968) and by Jacobs and Dunn (1968) but Thomas and Smith (1968) did not see an immediate decrease in the egg counts of sows when their piglets were weaned. In the course of a survey of Hyostrongylus infections in sows, Connan (1971a) found more adult worms in lactating sows than in sows at other stages of the reproductive cycle and there were indications that worms persisted longer in sows that were in poor condition. This point had also been made by Connan (1967b), and Barth (1968) reported that frequently sows with such persistent burdens failed to breed. There appears to be a seasonal fluctuation in burdens of arrested worms in sows. Connan(l97la)found that early fourth stage H.rubidusincrease to a peak about December and decrease in the spring. Poelvoorde (1973) has described an increase, in early spring,in egg counts due to Oesophagostomurn spp. of housed boars. The increase seen by Connan (1971a) in the number of adult H. rubidus during lactation was not associated with any very obvious decrease in the number of early fourth stage larvae and Connan came to the conclusion that the increase in adult worms was due not so much to any increase in the number of worms resuming their development, as to a “failure of the self-cure mechanism”. Presumably, he visualized that the usual turnover of the population was in abeyance during lactation. It appears likely that arrested H. rubidus are resuming their development over a long period in the spring, but work on this and on the extent of the periparturient rise at different times of year is clearly needed.

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Changes resembling the spring rise have also been observed in the worm burdens of rabbits. Dunsmore (1966b, c) has shown that at some sites in Australia, Graphidium strigosum and Passalurus ambiguus are more numerous in female rabbits than in males during the breeding season. This was true also of Trichostrongylus retortaeformis although here the difference was largely due to a decrease in male rabbits (Bull, 1959, 1964; Dunsmore, 1966a; Dunsmore and Dudzinski, 1968). It is argued that since parasites as different as G. strigosum and P . ambiguus show similar fluctuations, the susceptibility of the host rather than the availability of new infection must be the cause. Outside the breeding season male rabbits tend to carry more worms than female rabbits.

VI. ANCYLOSTOMATIDAE Ancylostoma spp., Uncinaria spp.

Arrested development of hookworms shows many of the features seen in the systems already discussed, but the phenomenon appears to play a somewhat different role in the transmission of infection and the story is complicated by a diversity of migratory behaviour. While it was originally believed that infection with Ancylostoma spp. was by the oral route (Looss, 1897), Looss (1901) noticed that percutaneous infection was possible and rapidly came to the conclusion that it was the more important route. Within a surprisingly short space of time and working in appalling conditions (in a room used for the examination of post mortem material from the victims of a cholera epidemic) he had shown that from the skin, the larvae migrate to the lungs and reach the alimentary tract via the trachea (Looss, 1905). He had also made the very interesting observation that in old hosts a greater or lesser number of larvae remain in the tissues without developing and formed the impression that they could remain there almost indefinitely. In the years that followed there was considerable discussion of the question whether the tracheal migration (described in detail by Fiilleborn, 1914) was obligatory or whether larvae, orally administered, could develop in the gut directly as Leuckart (1 876) had concluded from possibly inadequate evidence. The experimental infections of Looss (1901) had suggested that when orally administered, Ancylostoma larvae penetrate the wall of mouth and oesophagus, and Miyagawa (1913, 1916) was of the opinion that until they had passed through the lungs, the larvae were susceptible to digestion, so that any larvae which reached the gut, without having performed a tracheal migration, would perish. Yokogawa (1926), however, showed that the larvae were not susceptible to digestion and it soon became clear that direct development in the gut could and did occur (Yokogawa and Oiso, 1925a; Oiso and Kawanishi, 1927; Foster and Cross, 1934).This was true also of Uncinariastenocephala (Fulleborn, 1926a). The majority of A . caninum larvae that are ingested develop directly and only a small proportion follows the tracheal route (Yokogawa and Oiso,

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1925b; Shirai, 1926; Matsusaki, 1950; Enigk and Stoye, 1968). Whether larvae which reach the intestine complete their development there or penetrate the mucosa and migrate to the lungs, appears to depend on the nature and condition of the host. Thus, when rabbits, guinea pigs or mice are infected orally the larvae pass via the liver and portal circulation to the lungs and via the trachea back to the gut (Yokogawa, 1926; Yokogawa and Oiso, 1926). In young dogs, according to Scott (1928) and Rohde (1959), the larvae do not perform this migration but they do so in cats and in rats. This, however, may not be solely a question of the unsuitability of the host for when larvae of A . duodenale are fed to dogs, a host in which only few worms persist and none mature, no migration occurs (Yokogawa and Oiso, 192%). While larvae of A . caninum make some growth in the lungs of puppies, they do not do so in the lungs of abnormal hosts (Nakajima, 1931). Not all the larvae reaching the lungs in the pulmonary artery penetrate into the alveoli. According to Miyagawa (1913, 1916) some pass into the peripheral circulation and, in abnormal hosts, the majority do so. In this case they encyst in the muscles and other tissues (Matsusaki, 1950). This demands that the larvae must be capable of passing through the capillary bed of the lungs and in the case of Uncinaria stenocephala this has been shown to be possible (Fulleborn, 1926b). Herrick (1928) had noticed that few worms become established in the gut of old dogs and Miller (1965) suggests that as in unsuitable hosts, so in old dogs the larvae of A . caninum migrate and take the somatic route. Since adult bitches are more resistant to hookworm infection than adult dogs, Miller speculated that more arrested larvae are likely to be stored in the tissues of bitches than of dogs. The possible significance of this is discussed below. Dormancy of A . caninum larvae in the tissues of bitches is not as well studied or documented a phenomenon as its peculiar interest would justify. Scott (1928) showed that after oral infection, some of the worms did not develop beyond the third stage and persisted for a considerable period in the lung, liver, stomach and both large and small intestines. A larger proportion of the larvae was arrested in old dogs than in young and in cats than in dogs. In cats many worms were arrested but many more could no: be accounted for in the alimentary tract, liver and lungs and it is possible that these found their way to other organs and persisted there. Shirai (1926) had also observed that in what he termed “improper hosts” the larvae failed to reach the intestine and many remained in the lungs without developing. Even in dogs, larvae which remain in the lungs for more than the very short time that is normal, fail to grow. Schwartz and Alicata (1934) noticed that larvae still present in the lungs 96 h after either oral or percutaneous infection had not grown at all while those that had reached the alimentary tract were considerably more advanced in growth and development. This does not mean that the lungs are necessarily an unsuitable site for growth, but rather that those larvae that fail to develop are also more likely to remain longer in the lungs. When arrested worms from rats were fed to dogs and cats the proportion appropriate to the particular host developed and the rest remained in an arrested state (Scott, 1928).

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Sarles (1929) observed arrested development of A . caninurn in the alimentary tract of dogs infected by the percutaneous route. A proportion of the worms did not develop beyond the third stage and when some of the worms were lost, between the 5th and 7th day after infection, the arrested larvae were not affected. Sarles found that the development of a greater proportion of the worms was arrested in old dogs than in young but in view of the way in which experimentalanimals were procured at that time, it is not possible to determinewhether the difference was due to age or to prior experience of infection. Circumstantial evidence that A . duodenale can be arrested in the alimentary tract of humans is presented by Brumpt and H o (1953) who not only observed inordinately extended prepatent periods in a number of cases but also recovered immature worms after anthelmintic treatment from the faeces of patients who had been withheld from infection for a considerable period. Arrested development as described by Scott (1928) or by Sarles (1929) should not be confused, on the one hand, with the immobilization, walling off and destruction of some larvae seen by Kerr (1936) in mice nor, on the other hand, with the slower growth that occurs in resistant dogs (Kotliin, 1960). It is evident that A . caninurn shows a marked tendency to interrupt its development in the parasitic third stage and that according to their migratory behaviour and the circumstances, the arrested larvae may persist in a variety of sites. The tendency for the larvae to migrate and to follow a somatic pathway so that they reach, and remain in, the tissues is greater in abnormal hosts than in dogs, and greater in old dogs, particularly in bitches, than in puppies. In so far as worms arrested in abnormal hosts are capable of continuing their development when ingested by young dogs, A . caninurn can be said to make use of paratenic hosts. The persistence of arrested larvae in the tissues of bitches has a very similar significance, for the bitch too can act as an intermediate host of a special kind. Early records of prenatal infection with hookworms were based on the finding of eggs in the faeces of pups so young that, granting a prepatent period of normal length, they must have been infected before birth. Thus, Howard (1917a, b) detected hookworm eggs in the faeces of a 14 day-old infant whose mother had suffered from ground-itch during pregnancy. de Langen (1923) found eggs in the faeces of a 6 day-old infant. Adler and Clark (1922) found eggs of A . caninurn in the faeces of day-old pups and Ackert and Payne (1923) found mature Necator suillus in piglets 26 days old (the normal prepatent period being 6 weeks). Attempts to induce prenatal infection experimentally soon followed. Foster (1932, 1935) infected pregnant bitches and demonstrated eggs in the faeces of their pups I1 days after birth. It is probable that the infection in the pups did derive from the larvae administered to their dams but since the previous history of the bitches was obscure, an element of doubt remains. This doubt was rather less in the case of the results of Clapham (1962) who kept her bitches in conditions calculated to prevent accidental infection and obtained very similar results. According to Miller (1966) the infection of bitches at any time, before they are mature, before mating or during pregnancy, invariably leads to prenatal infection of the pups. This fact, together with the circumstance that the infection

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in the pups always becomes patent about the same time, suggests that larvae lying dormant in the bitch are involved and that their migration and resumed development must be accurately synchronized with parturition. The subject was given a great impetus by the astonishing discoveries of Olsen and Lyons (1962, 1965) on the life-history of Uncinaria lucasi, a parasite of the fur seal which they studied on the Pribiloff Islands of Alaska. They found no worms in the gut of adult seals; only the pups were infected. But the worms in the pups were all at the same stage of development and pups became infected even when infective larvae were demonstrably absent from the environment. Moreover, pups reared artificially did not become infected. Olsen and Lyons were able to show that late third stage larvae were present in the blubber of all classes of seals and that for a short time after parturition they were present in the milk. It appeared that gut infections derived only from larvae transmitted in the milk; infective larvae, which were present in the sand from August onwards, could not give rise to infections in the gut butwheningested, migrated to the blubber and persisted there. Commonly, large infestations of infective larvae in the sand survive from one season to the next and are still present when the seals return from the sea, but in some years few or none survive. The evolution of this life history should be seen against the background of this circumstance. Clearly, the transmission of partly developed worms in the colostrum could provide one explanation of the early patency of other hookworm infections in the recently born. Indeed, Stone and Girardeau (1966) recovered larvae of A . caninum from the colostrum of a bitch and Enigk (1970) claims to have made a similar observation even earlier. Enigk and Stoye (1968) infected bitches kept in worm-free surroundings at different stages of pregnancy and delivered the pups by Caesarian section. Pups reared artificially did not become infected, but those reared on their dams did. Meanwhile third stage larvae were recovered from the milk, peak numbers occurring on the 6th day and the last larva being seen on the 20th. Larvae of Uncinaria stenocephala have also been recovered from the milk of bitches (Enigk, 1970). Transmission via the colostrum appears to be a far more common route than via the placenta. Indeed, there does not seem to be any really conclusive evidence, such as the finding of hookworm larvae in foetuses, that prenatal infection with hookworms occurs at all. There is little evidence as to whether seasonal factors play any part in the arrested development of hookworms. Recently, however, Schad et al. (1973) have reported that in humans in India egg counts due to A . duodenale tend to rise some time before the beginning of the monsoon. If this were due to new infection, it must have been acquired in very arid conditions and at a time when hookworm larvae had been shown not to be available in the soil. Seasonal fluctuations in egg counts had also been observed by others. Schad et al. (1973) provided fairly good evidence that in an experimentally infected volunteer more than 22 weeks elapsed before the worms grew to maturity. They conclude that seasonal factors are responsible for the arrested development of the worms and the synchronization of their development with the advent of conditions that favour free-living development and transmission. There is

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every reason to believe that these observations and conclusions will be confirmed and amplified within the next few years. Beyond an oblique reference by Soulsby and Owen (1965) to arrested development of Bunostomum trigonocephafum,very little has been written on the subject. More detailed study of the development and migrations of the hookworms of ruminants should, however, prove rewarding. Percutaneous infection is, according to Stoye (1 965) and Westen (1967), more effective than oral infection in setting up worm burdens and Enigk (1970) reports that after oral infection many larvae become encapsulated in the liver. The fate and potentialities of such larvae is not revealed and awaits investigation.

Strongyloides spp.

VII. STRONGYLOIDIDAE

There are as yet no reports of the arrested development of Strongyfoides spp. either in the gut or at sites along the normal migratory pathway. But both prenatal and colostral transmission liave been demonstrated and the evidence suggests that arrested larvae are stored in the maternal tissues and that their reactivation is synchronized with a stage of pregnancy or with parturition. Enigk (1952) found eggs of S. ransomi in the faeces of 4 5 day-old piglets whose dam had been experimentally infected during the second half of pregnancy, and deduced that infection must have occurred before birth, even though no larvae could be detected in new-born litter mates. Very similar results were obtained by Frickers (1953) who found eggs in the faeces of even younger piglets. Stewart et af.(1963) did succeed in finding larvae of S. ransomi in the lungs and liver of one stillborn piglet and of another killed at birth. To explain the absence of worms from the gut, they suggested that migration of the larvae to the intestines was in some way inhibited until after birth. Stone (1964) also produced satisfactory evidence of prenatal infection, recovering 160 larvae from the new-born piglet of a naturally infected sow and 111 larvae from the muscles of another new-born piglet whose dam had received an experimental infection of 20 million larvae 2 months before farrowing. The relatively small numbers of larvae that could be recovered from new-born piglets puzzled a number of investigators. Supperer (1965) who found few or no larvae in the gut, lungs or other organs of new-born piglets whose litter mates showed patent infections 3 days after birth, speculated that the worms were chiefly in the blood at the time of birth. In an attempt to account for the fact that in young piglets the worms nearly always reached maturity at the same time after birth, Enigk (1952) suggested that only larvae which found their way to the foetus in the last 3 weeks of pregnancy could survive and that any that arrived sooner would perish. Pfeifferand Supperer (1966), however, established patent infections in 4-5 dayold piglets by infecting their dams 5 weeks before service. Moreover, they gave the sows anthelmintic treatment 15 and 18 days after infection. These puzzling findings were largely explained by the work of Moncol and Batte (1966) who showed that piglets which had sucked from their dams became infected, while those that were separated from the sow at birth and

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fed on her colostrum, after it had been filtered, remained free of worms. These findings were confirmed by Supperer and Pfeiffer( 1967)and by Stewart( 1969). There were, however, small differences between their findings in points of detail. Moncol and Batte found the greatest numbers of larvae in the colostrum before farrowing and a decrease already evident 12 h after parturition; Supperer and Pfeiffer found large numbers 44 h after farrowing and somewhat fewer after 68 h. According to Enigk (1970) the colostral route is far more important than the prenatal and indeed, Supperer and Pfeiffer report that in all theirresearches they found only one single larva in a new-born or foetal pig. Strongyloides transmitted via the colostrum can be the source of damaging infections. Stewart et al. (1968) for example, have described serious outbreaks in piglets so young that the worms could not have been acquired otherwise than by this route. Supperer (1965) noted that what at that time he took for prenatal infection was more marked in the litters of old sows than those of gilts and this suggests that in resistant pigs the larvae are less well able to develop and more likely to accumulate as arrested forms. Just where such larvae are stored until finally they migrate to the mammary gland is not known although it is to be presumed that they will have taken a somatic rather than a tracheal path. Miyagawa (1916) found S. stercoralis larvae in the kidneys of percutaneously infected dogs. Although the practical difficulties may prove considerable, the migrations of S. ransomi in sows offer an interesting field of study. Fulleborn and Shilling-Torgau (1911) and Fulleborn (1927) showed that unlike hookworms, S. stercoralis could not develop directly in the gut but that a migration through the tissues, preferably the lungs, was obligatory. In unsuitable hosts like the rabbit, development was not completed but the larvae would persist in the skin. When such larvae from rabbits were fed to dogs they developed directly without the need for a migration. This led Fulleborn (1927) to the conclusion that “unsuitable” hosts of S. stercoralis could be regarded as intermediate hosts. Supperer and Pfeiffer (1967) have pointed out that when S. ransomi is transmitted through the colostrum, the sow is acting as an intermediate host in which all but the gut phase of development is completed. It seems that S.papillosus can also be transmitted via the colostrum. Supperer and Pfeiffer (1962) have demonstrated that calves whose dams were experimentally infected began to pass eggs in their faeces 2 days earlier than calves given infective larvae immediately after birth. Bezubik (1969) has questioned whether prenatal infection with S. papillosus can occur at all because he could find no larvae in the progeny of experimentally infected rabbits. Because they could not recover S. papillosus larvae from the trachea of a percutaneously infected cow while larvae were plentiful in the tracheal mucus of a calf infected similarly, Supperer and Pfeiffer (1964) concluded that in the old (or resistant) host, the normal migration was prevented. They thought that this might increase the chances of prenatal infection, the larvae attempting to satisfy a frustrated urge to migrate. More probably, an abnormal migration leads to the accumulation in the somatic tissues of arrested larvae which may subsequently migrate to the mammary gland. It is possible that whenarrested Strongyloidesarestimulated by events associ-

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ated with parturition, their activity may not be confined to migration towards the mammary gland. Taylor (1955) has quoted an observation, at second hand, to the effect that eggs of Strongyloides westeri appear in the faeces of mares “about the commencement of activity of the mammary gland”. Ascaridia galli

VIII. ASCARIDAE

Variation in the migratory behaviour of Ascarids and the idea that there are “abnormal” as well as “normal” patterns, has engaged the attention of helminthologists for many years. Guberlet (1924) and Ackert (1924, 1931) found that while the great majority of A . galli in experimentally infected chickens develop in the small intestine, a few are recoverable from the liver, lungs and trachea, indicating that a tracheal migration can occur. It is interesting, in this connection, that Ascaridia columbae normally, and perhaps invariably, performs a tracheal migration @Iwang and Wehr, 1958). It soon became clear that, in addition to the abnormal tracheal migration of a small proportion of A . galli larvae, development in the intestine was also variable. While Ackert (1931) found that development occurs in the lumen of the posterior duodenum for the first 9 days after infection, and in the mucosa from the 10th to the 17th day, Tugwell and Ackert (1950) showed that a minority of individuals began the tissue phase as early as the 1st day and continued at least to the 26th day. Moreover, it appears, froni the work of a number of authors, that not all the larvae undergo a tissue phase at all (Tugwell and Ackert, 1952; Todd and Crowdus, 1952; Hansen el al., 1954; Horton-Smith and Long, 1956). Although a growth curve published by Ackert (1931) appears smooth, others by Tugwell and Ackert (1952) and reproduced in Fig. 9 show that neither larvae in the intestinal lumen nor those in the mucosa increase in length between the 10th and 14th days after infection. While larvae in the L”

24 22 -

- 20 -+E IS16 8

P f

g

J

1412 l08 -

6-

Age of A galli ( days )

FIG.9. Growth curves of Ascaridin galli in the mucosa (broken line) and in the lumen (solid line) of the intestineof chickens. Reproduced from Tugwell and Ackert (1952).

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lumen then resume their growth, those that remain in the mucosa do not. This must lead to a bimodal size distribution, and these were observed by Madsen (1962a) who pointed out (Madsen, 1962b)that such distributions were the only satisfactory criterion of arrested development. On this basis it is clear that arrested development commonly occurs in infections of A. galli but it is not easy to distinguish, in the older literature, between this phenomenon and retarded growth or stunting. Thus, the mean length of the worms is depressed by the age of the host or its previous experience of infection (Ackert and Jones, 1929), by its breed or the innate resistance of individuals (Ackert et al., 1935; Todd and Hansen, 1951) or by concurrent infection with Heterakis or Histomonus (Madsen, 1962a). Conversely the length of the worms is increased by dietary deficiency of the host (Ackert et al., 1931; Ackert and Beach, 1933) or by periodic bleeding (Ackert and Porter, 1932). Of a rather different order of magnitude is the effect demonstrated by Roberts (1936) who showed a negative regression of host age on both worm numbers and mean length which, in the youngest chickens, was six times as great as in the oldest. Graham et al. (1932), in attempting to separate the worms of a second infection from those of a first infection on the basis of their size, found not only that there was considerable variation in the rate of growth of worms of the second infection, but that some worms of the first infection appeared to develop very much more slowly than the remainder. Arrested development was clearly involved in the results of Ikeme (1970) who compared the worm burdens of chickens infected with 1000 eggs daily for 6 weeks with those of chickens receiving 10 eggs daily. At the low infection rate the first adult worms were seen in the 3rd week and no third stage larvae were present after the 7th week. At the high infection rate, no adult worms were seen until the 10th week and third stage larvae were present to the end of the experiment in the 19th week. That the parasitic development of A. galli might be influenced by seasonal factors, possibly acting on the free-living stages, was suggested by Itagaki (1927) who found that in spring and in autumn when climatic conditions favoured free-living development, parasitic development was direct. In the dry conditions of summer and to some extent in winter also, development in the host was delayed and the worms were enclosed in nodules in the intestinal wall. Ackert et al. (1947) noticed that in infections set up by the administration of old eggs of A. galli not only were fewer worms established than in infections with freshly cultured eggs, but the worms did not grow as rapidly. Hansen el al. (1953) who compared eggs cultured in air with eggs cultured in water found that the latter developed more slowly in the host, apparently because they underwent an extended tissue phase. Without doubt, there is a connection of sorts between arrested development and a histotrophic phase. While a tissue phase is not obligatory and while not all worms that enter the mucosa are arrested it does appear that all arrested worms are in the tissues. Evidently the larvae are not arrested because they are in the tissues but they remain in the tissues because they are arrested. Madsen (1962b), however, equated arrested development with abnormal migration. He saw the phenomenon, in all host-parasite systems, as depending on the

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resistance of the host and a lack of vigour on the part of the larva which could therefore be captured with resulting nodule formation. This seems an extreme view and one which perhaps does not distinguish clearly enough between an extended histotrophic phase and the encapsulation and destruction of occasional larvae described, in the case of A. galli, by Sadun (1950). While the literature cited above suggests that A . galli is arrested in the third stage, an element of doubt is introduced by a paper by Tongson and McCraw (1967) who studied the effect of the age of the host on the development of the worms and found that second stage larvae were present at least to the 48th day and that their relative number tended to increase with time, presumably as the larger worms were lost. The proportion of these larvae was greater in older hosts. It may be relevant that the development of many other ascarids is arrested at the second stage. Ascaris lumbricoides, Ascaris suum Stewart (1916a, b, c) described the tracheal migration of A. lumbricoides in experimentally infected rats which, he thought, represented an intermediate host. In the following year Ransom and Foster (1917) showed that a tracheal migration occurred in the pig also and that no intermediate host was needed. Ransom and Cram (1921) using mice, guinea pigs and rabbits, demonstrated that the larvae pass to the lungs not only via the portal circulation but that some travel via the lymphatics and the thoracic duct. Some larvae appeared also in the peripheral lymph nodes, indicating that they had passed through the capillaries of the lung and into the peripheral circulation. Cameron (1934) was of the opinion that this happened in pigs also. According to Asada (1926) an accessory route via the abdominal cavity is also used. Fiilleborn (I 921a) had also concluded that some larvae passed from the lungs into the general circulation and had recovered them from the brain and other tissues. Larvae which had taken the somatic route either died or became encapsulated in the tissues in which case, according to Ransom and Cram, they could survive for a considerable time. The significance of these encapsulated larvae is not known. The possibility that, as suspected by Stewart (1917), rodents can, after all, act as intermediate hosts should not perhaps be entirely ruled out although Ishii (1959) has shown that larvae of A . mum, which have developed in mice for more than 1 day, are no longer infective to pigs. It is not known whether a somatic migration can occur in old or resistant pigs. Presumably investigators are intimidated by the practical difficulty of hunting for a few minute larvae in several hundred pounds of sow. But a number of workers have satisfied themselves that prenatal infection (or colostral transmission) of A. suum does not occur (Shillinger, 1924; Martin, 1926; Alicata, 1926; Kelley and Olsen, 1961; Olsen and Gaafar, 1963). Soulsby (1957b, C, 1961a, b) has shown that in guinea pigs the larvae of A. suum undergo a period of slow growth in the late second stage from the 1st to the 3rd day after infection. In guinea pigs which had been immunized in various ways, larvae failed to grow beyond this stage. I n less effectively immunized guinea pigs the mean length of the worms increased slowly but it is not clear

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whether this was because all the worms grew slowly or because some remained at a length of 250 pm while the remainder grew normally. Crandall and Arean (1964) also found development to be arrested at this size and stage. They placed second stage larvae in millipore diffusion chambers in the body cavity of immune and susceptible mice and found that while the larvae survived equally well in either, they grew only in susceptible mice. In immune mice they remained a t precisely the same stage as in Soulsby’s experiments. On the basis of experiments such as these, Soulsby (1961a) saw arrested development as an effect of host resistance and visualized the stage at which development was halted as being specifically susceptible to the action of host antibodies. Having concluded, from experiments with other species of nematodes, that moulting is often accompanied by the release of antigens, he found particular significance in the circumstance that development of A . suum is arrested just before an ecdysis. He believed that the larvae must grow to some extent in the host before they can elicit an immune response.

Toxocara canis Fulleborn (1921b) noticed that larvae of T. canis can become encysted in the muscles and other tissues not only of dogs but also of mice and guinea pigs. These encysted larvae, which Fulleborn visualized as having strayed accidentally from their normal migratory path, could persist for several months. They neither grew nor developed, remaining at the same size as when hatched from the egg. When encysted larvae were fed to other guinea pigs, they migrated to the tissues and encysted again. Fiilleborn thought it likely that dogs might be infected by feeding them encysted larvae and that all manner of animals might serve as intermediate hosts including humans, in which symptoms of disease might be caused. Both Fiilleborn’s findings and his speculations have been amply confirmed. When infective eggs of A . mum are fed to mice, the larvae migrate to the somatic tissues including the brain and spinal cord and remain there, alive and active, for several months without measurable growth (Hoeppli et al., 1949; Beaver et af., 1952; Sprent, 1955a; Nichols, 1956; Ishii, 1959). When either infective eggs or encysted larvae recovered from mice are fed to young dogs, they develop to maturity after performing a tracheal migration (Schacher, 1957; Sprent, 1953a, 1957, 1958a; Webster, 1958). When infective eggs or encysted larvae from mice are fed to old dogs, on the other hand, they migrate to the somatic tissues and encyst (Sprent, 1953a, 1958a). Encysted larvae can survive in the tissues of dogs for at least a year and probably longer (Douglas and Baker, 1965). When encysted larvae from mice are fed to other mice they migrate to the tissues and encyst again and if these cysts are fed to yet other mice the process is repeated (Ishii, 1959). Not all encysted larvae which have been fed to other mice follow the same migratory path. A few perform a tracheal migration and reach the gut but it appears that these reinvade the tissues (Oshima, 1961a). While there is considerable variation in migratory behaviour it is clear that the larvae of T.canis tend to follow a somatic route in hosts that are unsuitable on account of age or species, and a tracheal migration in young dogs which

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may be seen as presenting more favourable conditions. Even in young dogs, however, the larvae may proceed to the lungs by three different routes, the portal circulation and liver, the lymphatics and thoracic duct and the body cavity and liver (Yokogawa, 1923; Matoff, 1968). Pregnancy and parturition also affect the migration of the larvae. Oshima (1961b) has shown that the proportion of larvae invading mice which encyst in the tissues decreases to a minimum at parturition. Apparently they migrate to the gut where, however, they do not persist. This finding illuminates the significance of the occurrence of a somatic migration in dogs. While small mammals such as rats and mice clearly act as paratenic hosts, mature bitches can fulfil a similar function because prenatal infection and colostral transmission are common. It is interesting, therefore, that Ehrenford (1957) found, during the course of a survey, that patent infections occurred much less frequently in bitches than in male dogs. According to Webster (1956, 1958) more larvae migrate to the tissues of bitches than of dogs. Fulleborn (192 1a) examined 5 day-old pups and found T. canis larvae in their intestines, of a size and stage of development indicating them to be 2-3 weeks old. There is, however, more direct evidence of intrauterine infection. Fulleborn (1921b) administered larvae, recovered from the tissues of a guinea pig, to a pregnant bitch by the intravenous route and found very many more larvae in the new-born pups than were normally present in pups out of bitches not experimentally infected. Shillinger (1923) obtained broadly similar results, as did Shillinger and Cram (1923) who gave anthelmintic treatment to their bitches before infecting them and who maintained them in very clean conditions thereafter. Further evidence of prenatal infection was provided by Augustine (1927) who demonstrated patent infections, 3 weeks after birth, in pups artificially reared on cow’s milk, and by Strasser (1964) who obtained similar results with specific pathogen-free pups. Petrov (1941) found migrating larvae of T. canis in still-born fox cubs. The results of Fulleborn (1921b), Shillinger (1923), Shillinger and Cram (1923) and Augustine (1927) constitute a series in which bitches were infected 8-33 days before parturition and in all cases there were no worms in the intestines of the pups at the time of birth; they were either in the liver or the lungs and all were at the same stage of development. The development of prenatally acquired T. canis larvae and its accurate synchronization has been studied in some detail by Sprent (1957, 1958a) and Sprent and English (1958). It was the opinion of Augustine (1 927) that development of the larvae was retarded in the foetus but prenatal infection can occur if the bitches are infected before service and withheld from infection throughout pregnancy. Yutuc (1949) gave repeated anthelmintic treatment to naturally infected bitches and maintained them, during pregnancy, in conditions which precluded accidental infection, but the pups were infected nonetheless. It is evident that larvae encysted in the bitch migrate to the uterus and it is probable that it is the timing of this migration which is related to the stage of pregnancy, although an additional and more precise control in the foetus is possible. No matter when the bitch is infected, eggs appear in the faeces 01 the pups 21 days after birth (English and Sprent, 1965; Enigk, 1970) or accord

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ing to Yutuc (1954) after 24.8 days. This compares with a prepatent period of 30 days in pups fed infective eggs shortly after birth (Noda, 1957). Douglas and Baker (1959, 1965) have shown that irrespective of when the bitch was infected, the foetus is invaded about the 42nd day of pregnancy provided that the bitch was infected more than 14 days earlier. Noda (1959), however, claims to have observed prenatal infection of pups born to bitches infected only 2$ and 5 days before parturition. Douglas and Baker infected bitches on a single occasion and were able to show that at least two successive litters became prenatally infected. (According to Enigk (1970) three successive litters can be infected by a single infection of the bitch.) Moreover, there was remarkable synchronization between the appearance of eggs of T.canis in the faeces of the pups and of the bitch. Kaemmerer and Stendel (1964) had suspected that the appearance of worm eggs of the faeces of nursing bitches might,be spurious because the bitches ingest the faeces of the pups, but Douglas and Baker (1965) found that the bitches continued to pass eggs after the pups were removed from them, A similar observation is that of Schlaaf (1959) who recorded the appearance of eggs of T.canis in the faeces of a jackal and of a dingo during lactation. It seems that when encysted larvae in the bitch are reactivated, presumably in response to some signal from the host’s endocrine system, they may migrate to the gut as well as to the uterus and develop at the same rate in either case (suggesting that the primary mechanism synchronizing the development of the larvae in the foetus with parturition is located in the bitch and not in the foetus). It appears further that the larvae can migrate to the mammary gland, for Enigk and Stoye (1968) have demonstrated their presence in the milk of bitches, so that presumably colostral transmission is possible.

Toxocara cati As visualized by Stewart (1918) T.cati uses paratenichosts,and manyspecies can serve in this capacity. The range extends from earthworms and cockroaches to man (Sprent, 1956). The larvae commonly occupy the brain and spinal cord (Beaver et al., 1952) and infection in man may therefore be attended with dangerous consequences. The part played by T. cati and T.canis as the cause of disease classified as “visceral larva migrans” has been copiously reviewed by Beaver (1956) and by Sprent (1969). According to the measurements of Sprent (1956) second stage larvae encysted in abnormal hosts are of exactly the same size as larvae newly hatched from the egg. When infectiveeggs of T.catiare fed to cats larvae are recoverable from liver and lungs and those which complete their migration to the alimentary tract develop. Some larvae can be recovered from the muscles and these show no growth (Sprent, 1955b). In mice that have been given infective eggs, larvae can be recovered from the liver, lungs and somatic tissues but do not reach the alimentary tract. In cats, therefore, a tracheal migration is normal, in mice a somatic migration. When infected mice are fed to cats the worms develop directly without the need for a migration and development is quicker in the mouseinfected cat than in the egg-infected one (Sprent and English, 1958).Although a

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somatic migration can occur in cats, albeit rarely, prenatal infection and colostral transmission have not been reported and are believed not to occur (Sprent and English, 1958; Egnik, 1970). Neoascaris vitulorum

Evidence of arrested development of N . vitulorum is at best indirect. Prenatal infection appears to be common and there is a good measure of uniformity in the age at which calves develop patent infections. Many workers have observed either eggs in the faeces of young calves or worms, at an advanced stage of development, in their intestines (MacFie, 1922; Boulenger, 1922; Griffiths, 1922; Molintas, 1938; Vaidyanathan, 1949) and it has been suggested that this is the only route by which infection can occur because Brumpt(1936), Herlich and Porter (1953, 1954), Refuerzo and Albiz-Jiminez (1954) and Cvetkovit and NeveniC: (1960) failed to produce patent infections in calves of various ages by the administration of infective eggs. When Herlich (1953) fed infective eggs to mice, his subsequent examination of various tissues led him to the conclusion that the larvae performed a tracheal migration. But for a time at least, larvae were recovered from the kidneys. Irfan and Sarwar (1954) reported that in guinea pigs the larvae did not proceed beyond the liver and that they appeared to remain there for an indefinite period. Refuerzo et al. (1952) showed that when infective eggs were orally administered to calves, they could subsequently be recovered from liver and lungs but did not reach the alimentary tract. This rather scanty collection of evidence may suggest that a somatic migration occurs both in laboratory animals and in cattle, a conclusion also reached by Jansen (1963). It appears probable that larvae in the tissues can resume activity after a considerable time. Infection of the dam at any time from 19 to 191 days before parturition (and presumably even earlier than this) has been shown to result in infection in the calves (Srivastava and Mehra, 1955; Cvetkovid and Nevenid, 1960) ;butthe first appearance of eggs in the faeces of the calves occurs during a relatively short period around 24 days after birth (Molintas, 1938; Vaidyanathan, 1949; Refuerzo and Albiz-Jiminez, 1954; Herlich and Porter, 1954; Ranatunga, 1960) but exceptionally it may be as early as the 10th or 12th day (Cvetkovid and Nevenid, 1960; Ranatunga, 1960). Warren (1969) has shown that infection can take place via the colostrum and believes this to be by far the most importarit route. He could find no larvae in foetal or new-born calves. Calves which sucked from their experimentally infected dams acquired infections which were patent after 33 days, while handreared calves did not. Meanwhile larvae resembling those of N . vitulorum but twice as big as those freshly hatched from the egg, were present in the milk from the 2nd to the 18th day. Some other ascarids The life histories of the Ascaridae have been very adequately reviewed by Sprent (1 954) and it would be inappropriate to d o more here than touch on some

parallels with phenomena discussed in other sections. Sprent (1952a) examined

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the migratory behaviour of a number of species in experimentally infected mice and found them to conform to two basic patterns. Ascaris suum and Parascaris equorum, parasites of herbivores, performed a tracheal migration but, having reached the alimentary tract of the mouse, did not survive there for long. Ascaris columnaris, A. mustelarum, A. devosi (Sprent, 1952b), Toxascaris leonina and T. transfuga, all parasites of carnivores, tended to migrate to the somatic tissues of mice where, according to Sprent (1952b), they could persist, alive and active, for at least 6 months. In a number of cases it has been shown that such dormant larvae, from mice or other rodents, are infective to the carnivorous host (Matoff and Wassilef, 1958; Sprent, 1952b; Tiner, 1949) and it is accepted that the ascarids of carnivores use intermediate hosts. Tiner (1949) has shown that, in the case of two species, the larvae do sufficient damage in the brain of the intermediate host to increase materially the chance that it will be taken by a predator. In some cases the use of an intermediate host appears to be obligatory, as for example A . devosi and A. columnaris (Sprent, 1952b), while in others, as for example Toxascaris leonina, the final host can as readily be infected with embryonated eggs as with mice carrying dormant larvae (Matoff and Wassilef, 1958; Sprent, 1958b). In the dog or cat infected with T. leonina larvae from mice, the worms develop directly without any tissue migration and are therefore more advanced than in infections established with infective eggs. This implies that one tissue phase is necessary before the worms will develop to maturity and that it may occur either in the intermediate host or in the intestinal wall of the definitive host. Most species show some variation in migratory behaviour. In a number, a proportion of the larvae infecting mice follow the tracheal route (Sprent, 1952a). Tiner (1953) has shown that 10% of A. columnaris larvae become encapsulated in the wall of the caecum while the remainder pass via the liver and lungs to the somatic tissues. Toxascaris transfuga also shows this diversity of behaviour. While some larvae are enclosed in nodules in rectum and caecum, others perform a tracheal migration and yet others become encapsulated in various somatic tissues (Sprent, 1951). While, in the egg-infected definitive host, T. leonina spends some time in the duodenal wall, before returning to the lumen, Wright (1935) has pointed out that, particularly in heavy infections, some penetrate to the abdominal cavity, liver, lungs and other organs, those reaching the lungs returning to the gut via the trachea. Sprent (1959) has suggested that after its development in the intestinal wall, T. leonina migrates towards the lumen in suitable hosts and towards the abdominal cavity in unsuitable hosts. There are considerable differences between species in the amount of growth which they make in the intermediate host (Sprent, 1958b). While Toxocara ranis and T. cat; neither grow nor moult in the mouse, Toxacaris leonina and A . devosidevelop to the third stage, the latter also doubling in length. As might be expected where the carnivorous host swallows its prey whole, the ascarids of snakes grow to a considerable size in mice. There must be some doubt whether ascarid larvae in the tissues of the intermediate host can be regarded as arrested at a precise point in their development. According to Tiner (1953),

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for example, there is great variation in the length of larvae of A . columnark, and of an ascarid of the racoon, in the brain of mice. Sprent (1955a) has considered the possibility that whether the larvae follow a tracheal or a somatic migration depends in part on whether they grow before reaching the lungs, small larvae being able to pass through the lung capillaries to reach the peripheral circulation, while large larvae are filtered out and break into the alveoli. This may be a fruitful line of thought, for in hosts which are unfavourable through an innate or acquired resistance, growth of the larvae may be inhibited and a somatic migration therefore more likely to ensue. This may occur also in very heavy infections. In time it may prove that, like Toxocara canis, a number of other species can encyst in the somatic tissues of the definitive host and that prenatal or colostral infection can occur. Heterakis gallinae

IX. HETERAKIDAE

According to Dorman (1 928) H . gallinae does not migrate beyond the alimentary tract of chickens, It does, however, spend a brief period between the 2nd and 5th day after infection among the glands of the caecal mucosa (Uribe, 1922; Roberts, 1937)andcompletesthesecondmoultduringthisperiod.Growth, during the tissue phase, is very slow (Lund, 1958a). Uribe (1922) thought that the occasional encystment of H. gallinae in the caecal wall indicated an abnormal host-parasite relationship and Lund ( 1 958b), finding fewer worms to be mature in previously infected chickens, after challenge, than in susceptible controls, suggested that this was due to a delayed emergence of worms from the caecal mucosa. Madsen (1962a) has found bimodal (and at times skewed) size distributions in populations of H. gallinae from chickens infected once only, the smaller mode corresponding with the size of worms i n the tissue phase, and it may be concluded that it is at this stage that development may be arrested. This is not to say, however, that this connection between arrested development and the tissue phase is not fortuitous. Habronema spp.

X. SPIRURIDAE

An isolated 40 year-old reference to arrested development of Habronema spp. in horses is of considerable interest. Schwartz et al. (1931) reported the finding, by A. McIntosh, of Habronema larvae in the stomach of horses near Washington, D.C. These larvae were no more advanced than those that occur in the intermediate host and they were noticed between the months of December and March when the flies which act as intermediate hosts were not present. This not only tends to demonstrate that the development of the larvae in the horses had been arrested but also suggests what part the phenomenon was playing in the life history of the parasite. That Hahronrnia spp. show an inclination to interrupt their development at the third stage, is also illustrated by the fact that in cutaneous and pulmonary habronemiasis the larvae do not develop beyond this point.

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XI. DISCUSSION In the introduction to this review, the phenomenon with which it was to deal was specified as the temporary cessation of development at an early parasitic stage, which did not occur invariably and which, when it did occur, tended to affect only a proportion of the worms. The evidence which has been reviewed suggests that this was a reasonable definition and that it is possible to discuss the phenomenon in general terms and as it occurs in a wide range of nematode species. But it is also evident that arrested development as defined above is closely related to the failure of a number of species to develop in their intermediate hosts. It would hardly be reasonable, for example, rigidly to separate arrested development of Toxocara canis or Oesophagostomum dentatum in the bitch or the sow from their failure to develop in rats and mice, on the grounds that in the one case the worms are affected only in certain circumstances while in the other all are always affected. It can be argued, rather, that a fundamental propensity for arrested development has evolved in a number of directions in different host-parasite systems, against the background of different selection pressures. Arrested development has been observed in a considerable number of nematode species and it is certain that the record is incomplete. With the passage of time the phenomenon will be reported from a great many more host-parasite systems. Admissible evidence for its occurrence may be of three kinds: (a) the finding in a worm population of a large proportion at precisely the same immature stage when there has been no sudden uptake of worms for some time; (b) the presence of worms at an immature stage in animals that have been withheld from infection for a period of time longer than the normal prepatent period; (c) the occurrence of bimodal size distributions in populations of worms from hosts not exposed to a corresponding pattern of infection. The presence of a large proportion of immature worms in animals that are currently exposed to infection, may mean merely that the population is being turned over rapidly, but in this case a continuous range of stages would be present. The stipulation that in cases of arrested development as strictly defined, the worms should have interrupted their development at one precise point, seems to be justified by the large number of cases in which this occurs. The developmental course of most nematodes appears to include a critical or particularly sensitive step at which development is readily halted by unfavourable factors, or development beyond which demands some special stimulus. Not infrequently, a momentary hesitation at this point is a feature of normal development and sometimes it immediately precedes moulting and corresponds with a lethargus. A number of authors have commented on this, from their own points of view. Thus, Soulsby (1960, 1966) who regarded arrested development purely as a consequence of host resistance, thought that the antigens emitted at the time of moulting were involved and that “the inhibition of, for example, a receptor mechanism for further development” might be affected. Rogers and Sommerville (1969) with a particular interest in the signals which cause resting stages to resume development, noted that many nematodes are arrested just

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before a moult and, since it is not uncommon for nematodes to stop growing when they moult, suggested that “requirements for growth to re-start may be critical so that nematodes would be more vulnerable to adverse environments when moulting than at any other time”. Such critical steps in development are not confined to early parasitic development. An example illustrating this is provided by the vulva1 flap of Ostertugiu ostertagi which tends to be incompletely formed in adverse conditions, its development being far more likely to be halted at one particular stage than at any other (Michel et al., 1972b). Where arrested development occurs in a number of different circumstances and different causative factors appear to be concerned, the worms are nonetheless arrested at precisely the same stage. There is evidently considerable variation, both between and within species of nematodes, in their aptitude for arrested development and also in the factors in response to which development is interrupted. These causative factors are of two kinds: (I) Seasonal factors acting on the free-living stages, such as changes in temperature and, probably, photoperiod. (2) Factors affecting the suitability of the host as an environment for worm development. Among these are the unsuitability of abnormal hosts, innate resistance due to age or sex of the host and individual or breed characteristics, and the effects of previous infection, the concurrent presence of mature worms or very large inocula. There is as yet no adequate evidence that interaction of factors of these two kinds is necessarily involved as envisaged by Madsen (1962b). His ideas did, however, play an important part in the development of the subject and are therefore of considerable interest. Madsen, who believed arrested development to be identical with an extended tissue phase and who was writing principally about Ascaridiagalli, expressed himself as follows: “On infection with a certain dosage of eggs we have a very comp!ex dynamic interplay of factors on both worms and the host (including immunizing phenomena) which influence in varying degrees the entry, or not, of the worms into the mucosa. If the worms enter the mucosa then the various factors determine to what degree, at what time and for how long the association with the mucosa will prevail. A strong association obviously strongly impairs the growth of the larvae. Depending on the course of the said interplay, various degrees of plurimodal distributions of lengths of worms occur in single dose experimental infections.” The chief importance of Madsen’s views was that it put a period to the assumption that arrested development had necessarily a single cause and that the factors causing development to be resumed were necessarily the converse of those which had induced it to be halted. In many cases, conclusions regarding the cause of arrested development prove on closer examination to be unfounded because they are based on the proportion of the worms, present in the animals at the end of an experiment, which are arrested. Since, characteristically, arrested worms persist in the host for longer than developing or adult worms, factors or circumstances which lead to a rapid loss of adults are mistakenly identified as a cause of arrested

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development. In the same way, where a constant proportion of the worms that become established is arrested, the percentage of arrested worms, in animals which have been exposed to infection for a long time, will be far greater than in comparable animals that have been exposed to the same infection for a short time only, for while the arrested worms steadily accumulate, the burden of adults is maintained at a level related to the rate at which new infection is acquired. The significance of arrested development may be visualized in two different ways. Some workers see the phenomenon as performing a regulatory function, tending to maintain a constant burden of adult worms. According to this view, when the rate of new infection is high, some worrhs are diverted to a store, where they persist in an inactive and innocuous state, and they are mobilized from this store when worm numbers and the rate of new infection are low. On this basis, arrested development is seen as a consequence of host resistance and effects of the weight of infection and of the presence of adult worms are regarded as central phenomena. The renewed development of arrested larvae is believed to be controlled by a sensitive feedback mechanism which allows larvae to develop in sufficient numbers to replace adult worms that are lost. A massive resumption of development is regarded as a consequence of a depressed immune status. There is evidently some difficulty in accommodating the sensitivity of the first mechanism and the crudity of the second within the framework of a single theory (Dunsmore, I963 ; Soulsby, 1957c, 1958, 1960, 1961a ; James and Johnstone, 1967a). Foregoing sections have tended to show that evidence for a direct effect of immunity or of infection size on arrested development is rather less convincing than at one time it appeared, but an irreducible core of evidence remains and there can be no doubt that, in some systems, host resistance does play a crucial part, while in others the resumed development of arrested larvae is controlled by a feedback mechanism of the type mentioned. The other view, towhich the present writer subscribes, sees arrested development as fulfilling the function of synchronizing the life history of the parasite either with that of its host or with seasonal changes in the outside environment. Punctuality (which may be defined as being in the right place at the right time) is the essence of successful parasitism and the most effective aid to punctuality is the ability to mark time. As a means of synchronization, arrested development implies a response to signals either to induce development to be halted or to cause it to be resumed, or both. NOWthat it is coming to be recognized that the interruption of development at particular seasons may be regarded as playing an essential part in many life histories (see, for example, Muller, 1968; Malczewski, 1970b; or Blitz and Gibbs, 1972b), it is probable that considerable progress will be made during the next decade in elucidating the nature d these signals in a great number of host-parasite systems. When means have been found of inducing development to be arrested at will, in experimental infections, it will be possible to work out whether resumed development depends on the receipt of signals via the host or whether it is spontaneous, and to discover the nature of such signals.

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Since these questions are susceptible to experimental investigation, it would be idle to speculate at this stage, but it is clear that an endoparasite has access to rather limited sources of information regarding the time of year or conditions in the outside world. One possible line of communication would certainly be through the neurosecretory system of the host as suggested by Gibbs ( I 969). Indeed the endocrine system of the host not uncommonly acts as a regulator of parasite activity, especially where, to facilitate the infection of a new generation of susceptible hosts, the activity of the parasite is synchronized with reproduction of the host. A brief consideration of some of the evidence regarding the effect of the sex of the host and of gonadic hormones on the parasite and of some examples of their role in synchronizing the life of host and parasite seems appropriate at this point. Commonly, male hosts are more susceptible to nematode infection than are females. In mice infected with Aspiculuris tetraptera twice as many worms are established in 2-6 week-old males than in females and the susceptibility of males is reduced by the administration of oestrogens while that of both sexes is reduced by gonadectomy (Roman, 1951 ; Mathies, 1954, 1959). According to Stahl (1961) the difference does not become apparent until the mice are 10 weeks old but there is a difference at an earlier age in the rate at which the worms are lost from the host. Pregnant and non-pregnant mice are, however, equally susceptible (Dunn and Brown, 1962). More Ascaridia galli develop in male chicks than in females, provided they are more than 9 weeks old when infected (Todd and Hollingsworth, 1952). The resistance of immature female chicks can be increased by the administration of diethyl stilboestrol (Ackert and Dewhirst, 1950). Male mice are more susceptible to Nematospiroides dubius than females-and this is equally true of rats, an abnormal host (Dobson, 1961a, b). While there is no difference in the numbers of Nipposfrongylus brasiliensis established in male or female rats, in hamsters (an abnormal host) 23 times as many were established in males as in females and again the difference increased with the age of the host(Haley, 1958).Ehrenford (1956) found patent infections of Toxocara canis to be three times as frequent in male dogs as in bitches, the difference increasing with age. In not every case are males more susceptible than females. Dobson (1964) for example, could not detect any effect of the sex ofthe host on burdens of Haemonchus contortus in lambs. Stewart et a/. ( I 969) found gilts to be more susceptible to Strongyloides spp. and Ascaris mum than male pigs of the same age and Scrivener (1 964) found female lambs to be more susceptible than male lambs to Ostrrtagia spp. The activity of the worms or their pathogenic effect may also be affected by the sex of the host. Whitlock (1937) observed that nearly all cases of severe syngamiasis i n partridges that came to his notice were in females. Clapham ( I 939) confirmed this but demonstrated that equal numbers of Syngamus trachea became established in experimentally infected partridges of either sex. All classes of parasites afford examples of the synchronization of the host and parasite, apparently mediated in most if not in all by the host’s endocrine system. The haemosporidian Leucocytozoon simondi disappears from the

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blood of ducks in the winter and reappears, with increased schizogony, at the beginning of the breeding season. Both can be advanced by artificially increasing the length of daylight to which the ducks are exposed (Huff, 1942; Chernin, 1952). Sexual reproduction of Opalina ranarum and consequent formation of cysts (which infect the tadpoles) coincides with reproductive activity of the frog and appears to be due to the frog’s gonadal hormones (Bieniarz, 1950; ElMoftyand Smyth, 1960). The hormones associated with moulting of Cryptocercus punctulutus, a wood-eating blattid, prompt gametogenesis in its protozoan parasites (Cleveland and Nutting, 1955; Nutting and Cleveland, 1958). The relative abundance of a number of helminth parasites in male and female frogs changes during the breeding season and numbers in males can be affected by the administration of oestradiol (Lees and Bass, 1960). The gonads of Polystoma stellai, a trematode of the frog, can be induced to mature by means of pituitary implants which are believed to act by the release of gonadotrophic hormone (Stunkard, 1959). A phenomenon that may be of a similar kind is the destrobilization of cestodes; this has been observed when the host hibernates (see for example, Ford, 1972). Perhaps the best known example of reproduction of a parasitic insect being synchronized with breeding of the host is provided by Spilopsyllus cuniculi, ovarian development of which begins 10 days before the doe gives birth, so that gravid fleas are found only when there are young rabbits in the nest (Allan, 1956; Mead-Briggs and Rudge, 1960; Rothschild, 1961). Apparently this is due to the direct action of cortisone on the flea (Rothschild and Ford, 1964a, b). Similarly, reproduction of the cattle louse Dermalinia bovis is greatly stimulated by the administration of corticosteroids to the host (Michel and Sinclair, unpublished observations, 1964). The helminth parasites of fish also provide examples of synchronization of their maturation and reproduction with the seasons and the availability of intermediate hosts, and in a number of cases the signals received by the parasite appears to be mediated by the host. Chubb (1963) showed that the pseudophyllean cestode Triaenophorus nodulosus did not begin to grow in the pike until the winter, so that eggs were not released until the spring. It was evident, however, that growth and development were prompted by something other than falling temperature. The acanthocephalan Neoechinorlzyncus rutili in the stickleback shows an annual cycle of maturation, the number of gravid adults increasing in the spring when conditions favour infection of the ostracod intermediate host (Walkey, 1967). Kennedy (1969) has shown that maturation of the cestode Caryoplzyllaeus laticeps coincides with spawning by the host. In all these cases parasites persist in a relatively inactive or immature form until called into activity by some signal which, in most cases, appears to be transmitted by the host’s endocrine system. In nematodes, arrested development can clearly fulfil a similar synchronizing function and two main trends are discernible. (1) On the one hand are devices which postpone the development of the worms to maturity until a favourable time of year. In many nematode infections the life of adult worms is short. In an infection of more than minimal size the worms are lost after an adult life extending, in some cases, to 3 weeks or even less. Since free-living development is almost invariably far more successful at some 12

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times of year than at others, it follows that a severe selection pressure is exerted against any worms that reach maturity at the wrong time. In such circumstances it is to be expected that strains of worms will kvolve, which become arrested in response to the receipt, by the free-living stages, of appropriate signals from the environment, such as changes in temperature or photoperiod. Development will be resumed shortly before conditions favouring free-living development return. Whether, in any particular system, the resumption of development depends on a signal transmitted by the host or whether the worms develop spontaneously after a fixed lapse of time, must await experimental proof but either arrangement seems eminently possible. However prompted, resumed development may coincide with the host’s breeding season so that infective stages are plentiful when a new generation of susceptible hosts becomes available. This was discussed at some length in the section devoted to the “spring rise” and the conclusion was reached that while the output of worm eggs from parturient or lactating hosts was indeed greatly increased, the mechanisms concerned acted chiefly on the longevity of adult worms while seasonal factors determined the development of arrested larvae: (2) The second trend is best illustrated by the nematodes of carnivores which make use of intermediate hosts. Here there is a tendency for development to be arrested also in the definitive host, if by virtue of age or previous infection it has become resistant. The dormancy of larvae in intermediate hosts may fall outside the strict (but arbitrary) definition of arrested development used in the present review, since it seems to lack a facultative element. But any arbitrarily assembled group has ill-defined edges and it is impossible to ignore the parallels between the interruption of development in the abnormal or intermediate host on the one hand, and the resistant definitive host on the other, because both must be regarded as hosts unsuitable for the completion of development, and in both cases the temporary cessation of development serves a similar function. The significance of prenatal and colostral infection is seen by Enigk (1970) in terms of the heavy contamination of the environment before the young can become resistant. This implies that the young will be reinfected from the environment and that free-living development is completed more rapidly than the development of host resistance. Of greater significance, probably, is the part played by very early infection in inducing a state of immunological tolerance. Kassai and Aitken (1967) have shown that if newborn rats are infected with Nippostrongylus brasiliensis, the worms persist for very much longer than they do in rats first inf6cted at a slightly greater age. To this extent prenatal and colostral infection may be regarded as another device for overcoming the limitations set by the normally short reproductive life of the worms. In this broad group of infections, factors associated with conditions in the host are likely to be dominant as causes of arrested development. The effects of an innate resistance, whether due to the species, breed, individual character, age or sex of the host are very similar to those of an acquired resistance (see Michel, 1968) and it is to be expected that larvae may react similarly to these different situations. The connection between encystment in the tissues and arrested development offers an interesting subject for study. The view of Kotlan (1952), Madsen (1962b) and others, that arrested development and an

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extended tissue phase are identical, is no longer tenable and it is legitimate to ask, where both occur together, whether the larvae are arrested because they are situated in the tissues or whether they come to be encysted in the tissues because their development is arrested. For example if larvae of a nematode which normally performs a tracheal migration fail to grow before they reach the lungs they may be more likely to reach the somatic tissues. Other factors are, however, likely to be involved and the possibility must be considered that aberrant migratory behaviour may not merely be the result of accident, as early workers tended to think, but that because of their inherent nature or environmental conditioning, aberrant larvae are “doing their own thing”. Two basic patterns are therefore discernible. One is directed to synchronization with the seasons, with arrested development depending on the receipt by the free-living stages of signals from the environment; the other is directed to synchronization with reproduction of the host and development tends to be arrested in abnormal or resistant hosts and resumed in response to signals related to pregnancy or parturition. But these patterns are in no sense distinct; there are many areas, and indeed points of detail, in which they overlap. REFERENCES

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Barth, D. (1968). Hyostrongylus rubidus, ein weitverbreiteter Schweineparasit in Deutschland. Tierdrztl. Umsch.23,115-122. Bawden, R. J. (1%9a). Some effects of the diet of mice on Nematospiroides dubius (Nematoda). Parasitology 59,203-213. Bawden, R. J. (1969b). The establishment and survival of Oesophagostomum columbianum in male and female sheep given high and low protein diets. Aust. J. agric. Res. 20, 1151-1159. Beaver, P. C. (1956). Larva migrans. ExplParasit. 5,587-621. Beaver, P. C., Snyder, C. H., Canera, G. M., Dent, J. H. and Lafferty, J. W. (1952). Chronic eosinophilia due to visceral larva migrans. Pediatrics 9, 7-19. Benz, G. W. and Todd, A. C. (1969). Observations on the organization of nematode developmental stages present in some natural and experimental infections. Trans. Am. microsc. SOC.88,89-94. Bessonov, A. S. (1967). The dynamics of nodular infection of the stomach of cattle in ostertagiasis, caused by Ostertagia ostertagi and the diagnostic significance of ostertagia nodules. Tematichieskiy sbornik rabot PO gel’mintologic sel’skokhozyaystvernikh zhivotaikh 13, 242-251. Bezubik, B. (1969). Negative attempts to obtain transuterine infections with Strongyloidespapillosus in the rabbit. Actaparasit.polon. 17, 11-16. Bieniarz, J. (1950). Influence of vertebrate gonadotropic hormones upon the reproductive cycle of certain protozoa in frogs. Nature, Lond. 165,650-651. Blitz, N. M. and Gibbs, H. C. (1971a). Morphological characterization of the stage of arrested development of Haemonchus contortus in sheep. Can. J. Zool. 49, 991-995. Blitz, N. M. and Gibbs, H. C. (1971b). An observation on the maturation of arrested Haemonchus contortus larvae in sheep. Can. J. comp. Med. 35, 178-180. Blitz, N. M. and Gibbs, H.C. (1 972a).Studies on the arrested development of Haemonchus contortus in sheep. 1. The induction of arrested development. Int. J. Parasit. 2, 5-12. Blitz, N. M. and Gibbs, H. C. (1972b). Studies on the arrested development of Haemonchus contortus in sheep. 11. Termination of arrested development and the spring rise phenomenon. Znt. J. Parasit. 2, 13-22. Boag, B. and Thomas, R. J. (1971). Epidemiological studies on gastro-intestinal nematode parasites of sheep. Infection patterns on clean and autumn-contaminated pasture. Res. vet. Sci. 12,132-1 39. Boulenger, C. L. (1922). On Ascaris vitulorum Goeze. Parasitology 14, 87-92. Bremner, K. C. (1956). The parasitic life-cycle of Haemonchusplacei (Place) Ransom (Nematoda: Trichostrongylidae). Aust. J. Zoo[. 4, 146-151. Brumpt, E. ( I 936). “Pr6cis de Parasitologie” (5th edition). Masson, Paris. p. 865. Brumpt, L. C. and Ho, T. S. (1953). Diapause des ankylostomes chez les grands anbmiques. C.r. SPanc. SOC.Biol. 167, 1064-1066. Brunsdon, R. V. (1964). The seasonal variations i n the nematode egg counts of sheep: A comparison of the spring rise phenomenon in breeding and unmated ewes. N.Z. vet. J. 12, 75-80. Brunsdon, R. V. (1966a). Importance of the ewe as a source of trichostrongyle infection for lambs: control of the spring-rise phenomenon by a single postlambing anthelmintic treatment. N.Z. vet. J. 14, 118-125. Brunsdon, R. V. (1966b3. A comparison of the spring-rise phenomenon in the faecal nematode-egg counts of housed sheep with that of sheep grazing infective pasture, N.Z. vet. J. 14, 145-151.

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