Comb bulding by the oriental hornet (Vespa orientalis)

Anim. Behav.,1976, 24, 72--83

COMB BUILDING BY THE ORIENTAL HORNET

(VESPA ORIENTALIS)

BY JACOB ISHAY

Department of Physiology and Pharmacology, Sackler School of Medicine, Tel-Aviv University, Tel-Aviv Abstract. Oriental hornet young queens and workers have been induced to build combs in observation

boxes. Both workers and queens displayed building activity over a period of 2 to 6 weeks, which is approximately the time range necessary for the development of one generation (from egg to pupa or maximally from egg to imago). Building activity and the comb architecture have been found to be influenced by such factors as: size of group, age of hornets, amputation of wings, tarsi and tips of antennae and light, and by feeding with mannose and barbiturates. Cell size is influenced by various drugs. The earliest report concerning the annual cycle and nest structure of wasps is that Reaumur (1742) on Paravespula vulgaris, the prevalent wasp species in Europe. Other investigators have since studied the numerous aspects of wasp and hornet nest-building, such as the way in which the hexagonal cells in the comb are formed (Waterhouse 1864), the nest-building technique (Eberhard 1969), the stages involved in the construction of the individual comb cells (Deleurance 1947, 1955, 1957), the digging of the space to house the nest (Janet 1903), the manner in which wasps overcome the difficulties involved in widening and expanding the nest (Kemper 1961), the weight of the soil particles carried off in the course of nest-digging (Ishay, BytinskiSalz & Shulov 1967), the repairing of experimentally-induced damage to the nest (Weyrauch 1935, 1936, 1937), the influence of geotaxis on the direction of cell-building (Montagner 1964) and changes in the orientation of the pedicle fastening the comb to the roof of the nest (Wafa & Sharkawi 1972; Ishay 1973). Most investigators have concerned themselves with the nests built by complete colonies. Edgeworth (1864), however, has observed nestbuilding by groups of workers only, but in these instances rather than build cells, the workers primarily built envelope-like structures which protected the nest against adverse climatic conditions. Janet (1903), in contrast, did observe cell-building by worker groups only. In Vespinae the colony is founded by a solitary queen, who builds a small nest to house the initial brood. The first cells to be constructed are worker cells. Larger cells are built in the autumn by the workers and these serve to rear queens (Blackith 1958). In Australia, wasp colonies survive through two seasons, and in the winter following the first season, the young queens persist building in the original nest,

but build only queen cells. The extensive data on the annual cycle and nest-construction of numerous Vespinae species have been summarized by Wilson (1971) and Spradbery (1973). During years of observations we noticed that while going into hibernation hornet queens turned their wings along the side of the body in such a way that their distal edges touched and rested on the substratum in a characteristic manner (Plate I, Fig. 1A). In that position, they rested for long periods of time, almost without movement. On the other hand, queens bearing amputated wing edges were unable to go into hibernation and remained restless during the whole winter (Ishay, unpublished observations). These observations induced us to assume that the position of the wings on the hornet dorsum and the presence of the distal wing edges are important for their resting and sensorial perception. The present paper describes experiments and observations made on groups of (or individual) workers and queens of Vespa orientalis, in an attempt to elucidate the various aspects of nest-building by this hornet species; that is to say: (1) To attempt to induce building by young queens in the autumn, or during the hibernation period. (2) To gain information on what brings about the building by workers during the active season. (3) To attempt to influence the building through anatomical (amputation of wings, tips of antenna or tarsi), physiological (cooling for 4 to 6 days at 4 to 8~ influence of age, queen pheromone), pharmacological (feeding on atropine, morphine, pentobarbital ( = nembutal), mammalian oestrogens and testosterone), dietetic (feeding on mono- and oligosaccharides and animal proteins), sociological (isolation or 72

ISHAY: COMB BUILDING BY THE HORNET grouping of animals), and physical (influence of daylight, total darkness, direction of light) means.

Methods The observations and experiments related here were carried out between 1970 and 1974, during the active season of V. orientalis (May to December) and occasionally also throughout the winter. Experiments were done in the vesparium of Tel-Aviv University. Queens and workers, individually or in groups and even in colonies, were maintained in breeding boxes as described elsewhere (Ishay 1962, 1964). The hornets were offered honeybee or wasp pupae and 30 per cent sucrose solution as food. and clumps of soil and bits of paper as building materials. The indoor temperature was maintained at 27 to 28~ which is close to optimal for this species of hornet (Ishay et al. 1967). Each hornet used in this study was marked by a number on the back of the thorax within 24 hr of eclosion. Experimental Procedure A. The following experiments were carried out in daylight on single queens and on groups of queens: (1) Physiological effects on the building were tested by observing the building activity of queens following exposure for 4 to 6 days to temperatures between 6 and 8~ (2) The effect of the oestrogen di-ethinyl. estradiol was tested by feeding the hormones to the queens. (3) Anatomical effects on the building were tested by: (a) Amputation of one pair of wings; (b) Amputation of both pairs of wings. (4) Physical effects on building were tested by exposing the queens for 2 weeks to stroboscopic illumination at the rate of five flashes per second. B. The following experiments were carried out in daylight (except where otherwise specified) on workers: (1) Effect of social stimulation (or crowding effect) on the building activity. This was tested by placing the workers from the moment of eclosion: (a) Singly in the breeding box; (b) in groups of two to four idividuals; (c) in groups of five to ten individuals; (d) in groups of 20 to 200 individuals. C. The following experiments were all carried out on groups of five or more workers maintained in the breeding box only:

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(1) The correlation between induced anatomical changes and symmetry of the building architecture was tested by: (a) Partial or total amputation of the wings (fore wings and/or hind) wings; (b) ligation of the wings (at their bases, their middle or at the tips; each pair separately or both pairs together, the latter procedure resulting in crossing of the wings; (c) gluing of the wings at rest as they are either spread or folded; (d) attaching (by glue) a prosthetic wing or two above the natural wings so that the former project some 5 to 10 mm beyond the latter; (e) amputation of the terminal segments of one or both antennae, and (f) amputation of the tip of the tarsus in one foreleg. Further details on the experiments are given in the results section. (2) The effect of light was tested in the following manner: (a) By maintenance of the newly ecloded workers for 3 weeks in total darkness and their subsequent transfer to daylight; (b) by the reverse procedure, i.e. initial maintenance in daylight and subsequent transfer to total darkness; (c) by constant exposure to white, ultra-violet or stroboscopic (white) light. (3) The effect of the direction of illumination was tested by illuminating the breeding box (a) from the side (white light) (b) from the bottom and (c) from the top. (4) The effect of diet on the building was tested by feeding the hornets on: (a) Sucrose solution with or without animal proteins; (b) various mono-, di- and tri-saccharides other than sucrose; (c) sucrose solution together with the mentioned sugars; (d) animal proteins only. (5) The effects of the following hormones were tested: (a) C 16 lactone, the queen pheromone of V. orientalis. This was tested directly on workers that had been given the hormone in the food as well as indirectly on workers that had been maintained for several days together with a fertilized queen and thus received the hormone from her; (b) the androgen testosterone, which is the principal testicular hormone in man, (c) the oestrogen di-ethinyl-estradiol, which is a semi-synthetic oestrogen of high potency. (6) Physiological effects on the building were tested by: (a) observing the building activities of workers following exposure for several days to temperatures between 6 and 8~ (b) observing the building activities of workers after initial amputation of the wings and subsequent exposure to temperatures of 6 to 8~ for

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several days; (c) studying the influence of age on correctness of building. (7) The effect of prevention of building was studied by: (a) withholding of building materials; (b) maintenance of the hornets i n breeding boxes in which an excess of vacant cells from a previous construction were available; (c) maintenance of the hornets in breeding boxes whose geometric structure did not enable normal building. These consisted of breeding boxes whose roofs were only 1.5 cm high, or narrow and tall breeding boxes which did not enable the building of more than two cells across, or breeding boxes whose roof was made of a repellent material such as glass wool. (8) The effect of the following pharmacological substances was tested: (a) atropine, which is a competitive antagonist of acetylcholine (muscarinic effects); (b) morphine, which is the most important narcotic analgesic; (c) pentobarbital ( = nembutal), which is a short-acting barbiturate and general anaesthetic. Except where otherwise specified, all results are summed from at least ten separate trials. The majority of hornets used in the present study were workers and queens that had ecloded in the laboratory from combs collected in the field in the environs of Tel-Aviv, as described previously (Ishay 1964). A few of the experimental hornets were obtained from purely vesparium reared colonies. The combs of a single colony yield, within 2 weeks in the middle of the season, up to 2000 workers, and at the end of the active season (September to November) about 1000 to 3000 queens. The number of colonies discovered from time to time in the field is well beyond the experimental needs. Our vesparium can hold eighty breeding boxes, within which various experiments can be run concurrently with no inconvenience. Where necessary specific experimental procedures are described in greater detail under the Results section.

Results Young Queens during the Months of October to December Effects of cooling. Young queens (before or ~fter fecundation) maintained individually or in groups of up to 200 individuals in the breeding boxes displayed no building activity in the vesparium. However, if groups of queens of ten or more individuals are maintained for 4 to 6 days at 6 to 8~ and then are returned to normal laboratory temperature (27 to 28~ this

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treatment induces some of the young queens to construct a natural comb into which they oviposit. The comb construction and ovlposition are similar to those of queens under natural conditions in the spring (Plate I, Fig. 1), but with two salient differences as follows: (1) A number of young queens may construct a common comb into which they oviposit jointly without any show of animosity. In nature, in contrast, a queen that founds a nest in the spring is markedly territorial, does not permit any rival queens nearby and any such encounter, whether accidental or experimentally programmed, will result in the founding queen attempting to sting the trespassing queen to death. (2) The young queens construct queen cells, as do worker hornets in the natural nest in the autumn, but unlike wild queens in the spring (which construct worker cells). Under stroboscopic light however, they built combs of worker cells. Building by young queens with amputated wings. If the breeding box already contains combs with queen cells, amputated queens oviposit in the queen cells and attend the ecloding brood; if the combs contain worker cells, the young queens correct some of the worker cells (by deposition of building material at their outlets till the elongated cells acquire the volume of queen cells) and then oviposit in them. They will oviposit in queen cells of combs which are placed horizontally, i.e. at 90 ~ to the normally down--facing comb, but not in combs which are turned upside down, with the cells outlets facing up. Young queens that have had one pair of wings amputated, commence spontaneous building activity, if the breeding box does not contain ready-made combs (from a previous construction). The amputated queens spontaneously engage in two types of construction as follows: (1) Firstly they build a conglomerate of cells in a single layer which partly overlap each other like the tiles of a roof and which are fastened at one end to the side of the breeding box. The volume of these cells is equal to that of queen cells. Construction of the cells commences from the bottom up. At first a vertical partition is constructed at about the centre of one side of the box. On this partitition, two cone-shaped cells are constructed, one on each side of the vertical partition. Next, another two cells are constructed, starting from about half-way up the initial cells and so on, working upwards.

ISHAY: COMB BUILDING BY THE HORNET It should be noted that all of these cells are fastened directly to the wall of the box for at least half their length and that there is no pedicle or compact comb shape, with cells at a uniform height. The queens oviposit in these cells and for about 2 weeks nurse the ecloding brood jointly. (2) Subsequently, the amputated queens abandon the original comb (and brood), and adjacent to it but somewhat higher up, they construct one to three queen-celled combs which are attached to one side of the breeding box through a horizontal pedicle. In this case, too, construction commences with the building of a thin partition (of the same thickness as the cell walls) but whereas in the earlier construction the partition served as a common wall for two adjacent cells, in the new combs the partition serves merely as a pedicle which separates the comb from the wall of the breeding box, much the same as the pedicle of a naturally-constructed comb suspends and thus separates the comb from the roof of the nest. At the distal end of this partition, the queens jointly construct a queencelled comb into which they jointly oviposit. Effects of feeding drugs. Young queens maintained in groups whose diet is made to include di-ethinyl-oestradiol (which is incidentally the human anticonceptional pill) dissolved in sugar solution (1 mg per 100 ml) engage in building activities during many hours of the day. They do not construct new combs but rather collect building materials and either deposit them in irregular layers at the outlets of impaired cells or use them to extend the length of existing cells and pedicles. In no instance does this construction serve any apparent effective purpose, inasmuch as no new cells or envelopes are built but rather existing structures are merely modified. These modifications are noticeably irregular and are independent of which way up the comb is i.e. the cells are extended in any direction, even if the comb is situated at right-angles or inversely to the natural position. Moreover, the queens never oviposit into the cells that they have altered. Young queens maintained individually do not display any building activity under any of the treatments described above. Attempts to induce groups of young queens to build between January and March have not succeeded.

Workers Effects of group size. (1) Workers who upon eclosion are kept together in groups of five

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to fifty individuals, will within a few days commence building combs with worker cells, much the same as the wild queen does in the spring. The number and size of the combs are dependent of the number of workers which normally make up the comb-construction team. Thus, when a large number of workers are present in the breeding box, several combs are constructed simultaneously which ultimately merge into a single comb extending the entire length of the roof of the breeding box. All cells whose bottom part is already completed are promptly oviposited into by workers participating in their building. (2) In breeding boxes housing two to four workers only, construction is delayed till the second week of the workers' time together. In such cases, the pedicle joining the comb to the wall of the breeding box is not suspended from the roof of the box but at approximately 45 ~ from one of its vertical sides. (3) Workers kept singly in breeding boxes do not engage in any building activity. Effects of wing amputation. Wasps and hornets eclode with the fore-wings fully spread, and only on the second day of life do they fold their wings longitudinally when at rest (Schremmer 1962; Kemper & D~hring 1967). This~is the source of the old name Diploptera given to Vespidae that fold their wings (Imms 1960). If the bind wings are amputated prior to folding of the fore-wings, then the latter remain spread for life. If only one hind~wing is snipped off, then only the corresponding fore-wing remains unfolded. A hornet that has had one or both hind wings amputated is incapable of flying. If the hind wings are amputated 1 or 2 days after the folding of the fore-wings, then the fore-wings remain folded during rest, and are spread during flying attempts. The following treatments of their wings cause the hornets to build laterally-attached, i.e. incorrectly built combs: (1) The amputation of at least one hind wing (Fig. 2:9 bottom) or half of one fore-wing (Fig. 2:3). (2) The gluing on of one or two extra wings to the fore-wings (so that they project 5 to 10 mm past them). (3) Tying together one pair of wings at their bases, leaving the contralateral pair free. The wings that are bound together, although intact, cannot be spread or moved (Fig. 2:6 bottom), freely and usually hang useless along the side of the body.

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Fig. 2. Schematic description of various experiments on wings: 1. Unfolded pair of wings. 2. Folded wings as in resting position. 3. Amputation of the fore wings up to the marker lines. 4, Snipping the outer margins. 5. Snipping the inner margins. 6. Trimming of both outer and inner margins or ligating the bases of each pair of wings separately. 7. Snipping off a triangle-shaped portion from the tip of each fore-wing. 8 and 9. Boring three to four circular holes (t .5 mm in diameter) in one or both fore-wings. 10. Making three longitudinal incisions in each fore-wing. 11. Fixation of the wings by gluing them or by ligating them at their bases, or at their tips (according to the experiment). 12. Amputation of the hind-wing and 13. Snipping the inner margin of the fore-wing to prevent attachment of the hind-wing. An example of two incorrect combs is given in Plate II, Fig. 3. In the majority of such combs, the planes along the roofs of the cells and particularly along the cell entrances are horizontally unbalanced and there is also marked irregularity in the size of the cells. When the two pairs of wings are bound at their bases and crossed over, the hornets cease building altogether (Fig. 2:11). When only the tips of the fore-wings are bound together and crossed over, the hornets construct only envelopes but not the stem or the comb proper. Fig. 2 schematically describes the various experiments on hornet wings. Similar construction is also displayed by workers whose wings are left intact but the tarsus of one foreleg or the first segment of one antenna is covered with cement or, alternatively, its tip is damaged (by heat) or removed. The

hornets were anaesthetized with ether and the legs or antennae were wrapped in dental cement (Harvard cement, Dichter & Hofmann Dental Geselschaft, Berlin) which hardens rapidly, and cannot be removed by the hornets. Control groups anaesthetized as well did not show any change in their building activity. The same results were obtained with hornets anaesthetized several times consecutively. Covering with cement or damaging the tip of the distal segments of both antennae causes a worker to cease normal building altogether. Such workers cluster in groups of eight to fifteen on the roof of the breeding box, where they may deposit building material in irregular fashion, but they evidently cannot construct a comb pedicle or cells. Incidentally, the flight of such workers is apparently unimpaired, for they fly in natural and co-ordinated fashion both in the breeding

ISI-IAY: COMB B U I L D I N G BY THE H O R N E T PLATE

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Fig. 1. Young queens of V. orientalis that had been cooled for several days. Some of them remained inactive upon return to room temperature, but others started constructing a normal comb in one of the cells, an egg laid by one of the workers may be seen. Notice the folding of the wings along the body sides and folding of the edges of the distal part of the wings, while resting upon the substrate. (A).

Ishay, Anim. Behav., 24, 1

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Fig. 3. Incorrect combs built by workers with amputated wings. Fig. 4. Building in total darkness: The comb consist on single cells attached one to another and on one side of the box. Ishay, Attire. Behav., 24, 1

ISI-IAY: COMB BUILDING BY THE HORNET box and ill the courtyard of the vcsparium in the same way as their undamaged nest mates. The following treatments of the wings do not result in changes in the building pattern: (1) Snipping off the outer margins of the fore wings (Fig. 2:4). (2) Snipping off the inner margins of the fore wings (up to the hind wings) (Fig. 2:5). (3) Concurrent trimming of both the inner and outer margins of the fore wings (Fig. 2:6 top end). (4) Boring three circular holes (1.5 mm in diameter) in one or both fore wings (Fig. 2:8; 9 top end). (5) Snipping off a triangle-shaped portion from the tip of each fore-wing (Fig. 2:7). (6) Making three longitudinal incisions in each fore-wing (Fig. 2:10fore wings). (7) Ligating t h e bases of each pair of wings separately, so that the bound wing pairs are arranged symetrically on both sides of the body (Fig. 2:6 bottom). (8) Gluing of each wing separately in the following position: (a) spread (Fig. 2:1); (b) folded (Fig. 2:2). Fixation of the wings causes the hornet to build a long narrow ledge rather than a stem but the comb itself is constructed properly. In hornets aged 3 to 4 days or more, incorrect comb construction occurs only if the hind-wings are amputated, and there is no building whatsoever if their wings.are crossed by ligating all four wings together at their bases. Effect of light. The vesparium is dimly lit throughout the day so that no photography is possible without additional illumination. In such dim light, there is usually correct, spontaneous comb building by the hornets. Correct building occurs also when the breeding boxes are illuminated from the side or bottom by an electrical light (40-W incandescent bulb). However, in breeding boxes with translucent ceilings, illumination through the top results in incorrect comb building. Incorrect building occurs also if the box is illuminated from the side by ultraviolet light (252 gm). Hornets aged 3 to 4 days or more build incorrectly only if their breeding boxes are illuminated from the top. Building in total darkness. Hornets maintained, from eclosion, in the dark (by wrapping their breeding boxes in numerous black sheets), construct single cells in one layer on one side of the box. These discrete cells do not conjoin to form a comb and each is attached to the wall of the box by its side and top, with the cell entrance facing down. Such cells may be constructed in vertical or horizontal rows, but in repeat experiments no definite building pattern was discernible, the cells fastened irregularly

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in most parts of the wall (Plate lI, Fig. 4). Hornets that had spent several days in light, when placed in total darkness, construct normally as in light, i.e. the stem is attached to the roof of the breeding box and the comb is symetrical and comprised of regularly shaped cells. Hornets which i~litially built scattered cells in the dark, when subjected to illumination resort to perfect comb construction. Effects of feeding on various sugars and drugs. Hornets maintained on water and meat adlibitum in the absence of sucrose solution, survive for only a few days and cease construction altogether. When fed on sucrose solution only, the hornets survive for up to 2 weeks but do not build. Numerous sugars, such as glucose, fructose, maltose, lactose, trehalose, melibiose, raffinose of melezitose, when served along or together with the standard sucrose solution, fail to alter the building habits or the development & t h e eggs oviposited in the cells. Feeding on mannose or galactose only, causes death of the hornets within a few days, probably due to starvation resulting from failure of the hornets to feed on these sugar solutions. If these two sugars are given while dissolved in sucrose solution (100 mg per ml sucrose solution) the hornets do not die but construction is delayed tilt the third week of life. (Hornets fed on the standard diet of sucrose solution and meat start building on or about the fourth day of life.) The addition of mannose or pentobarbital (1 mg per 1 ml solution) to the diet results in incorrect comb building. The following drugs and hormones when added to the sucrose solution, do not alter the direction of comb building or the orientation of the comb: atropine (1 mg per 1 ml), morphine (100 mg per 1 ml), di-ethinyl-estradiol (1 mg per I ml) and testosterone (1 mg per i ml). With the exception of morphine, all the substances listed above induce the building of larger cells than normal. Especially large are the cells built under the influence of pentobarbital, which attain the size of queen cells even during the season when only worker cells are usually constructed. State of ovaries during construction. Hornets that had commenced building were dissected and were found to contain developed ovaries, each containing usually two to three eggs of large size similar to that of oviposited eggs. In each worker group there was more than one worker engaged in oviposition at the same time,

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so that no dominance of a single worker with respect to oviposition occurred. Dissection of workers that had failed to build over a prolonged period showed that none of them had developed ovaries. Effects of cooling and of queen pheromones. Workers lacking one or both pairs of wings, if maintained in a refrigerator (4 to 8 ~ for 4 to 6 days and then restored to the breeding box build correctly (see Ishay 1973). If workers from queen-right colonies that had spent several days in the nest and had come into close contact with correctly built cells as well as with pheromones released by the queen now had their wings cut off, they mostly retained correct building habits, i.e. they built combs whose pedMe is fastened to the roof of the breeding box. Whenever a comb is constructed (whether by workers or by the queen), the outlets of the cells always face downwards. If, however, the breeding box is gradually rotated or tilted, the workers correspondingly alter the direction of the cell outlets, thus suggesting that the cell outlets are always associated with a clear-cut, positive geotropism. Prohibition of building. In experimental colonies, if numerous combs are intentionally planted to present a total number of cells considerably greater than customary, none of the workers engages in building activity. If for some reason the workers fail to build during the initial phase of life, whether because of a surplus of cells or a paucity of building materials or inadequate places for building in the breeding box, they will not do so at a later stage. If a group of workers that had completed building a correct comb with a number of cells is transferred into a new breeding box, the workers once more build a similar, i.e. correct, comb. However, upon a second or third transfer, the workers build an incorrect comb, similar to that by wingamputated hornets; in such cases, the comb is built on a clod of soil on the floor of the breeding box, a phenomenon not observed in any of the other experiments. This transfer experiment was run on these groups of hornets who were 24 hr old at its start. Building by mixed groups (with intact and amputated wings, or previously exposed or not to a fertilized queen). Ordinarily, if the breeding box contains five to seven workers, these all form a single work team which starts construction at one point,

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building first a pedicle and then a small comb, like the young wild queens do in the spring. On the other hand, if the nest contains fifteen or more workers, the workers commence simultaneous construction at two or more different points, which usually results in two or more combs some distance apart (in a nest containing a fertilized queen, the construction is always from one central point). If the hornets in groups without a queen are winged, all their combs will be suspended from the roof of the breeding box, as occurs in nature, but if the wings of the workers are amputated, the combs will be fastened one above the other and vertical to one side of the box. If the larger construction teams contain workers that had undergone various treatments (e.g. amputation or non-amputation of the wings, exposure or non-exposure to a queen for short intervals), then the construction is mixed, i.e. some of them are correct, some are incorrect, but the combs are still built some distance apart. As construction proceeds, most of the combs usually become enveloped in one or more envelopes, the workers enfolding together both correct and incorrect combs. However, whereas the comb-building workers form into construction teams made up of five to seven individuals, with each team squatting over its own comb, ovipositing into its own cells and protecting its own brood, the envelopes, in contrast, are constructed as a joint effort, with members from the various construction teams forming into extra-territorial, ad hoc work groups. While there is a certain measure of aggressiveness in the relations between the territorial, comb-building teams, the intra-team relationships are markedly tolerant. The activities of each member within a team are undoubtedly intermittent. Thus any member of the team may alternately forage for food, share its food with the others, feed the larvae, ventilate or guard the comb and engage in construction. However, the appearance of an alien worker (from another group) on the comb elicits aggressive behaviour in the 'native' population. Individuals of the local team approach the intruder in a threatening manner and this is usually sufficient to scare it away. The occasional unintimidated intruder is nipped mandibularly, wrestled with and ultimately pushed off the comb. Only rarely is there recourse to fatal stinging of the intruder. Effect of age. We deemed it of interest to study the building habits of individual workers within

ISHAY: COMB BUILDING BY THE HORNET a construction team made up of seven workers only. We studied such individual building in parallel in thirty separate breeding boxes, each with only seven workers inside. All hornets used in this experiment were from the combs of a single nest collected in the field in the last week of July. Each construction team was comprised of workers that had ecloded on the same day and all the teams were made up of the hatch of six consecutive days. Each worker was marked by a number on the back of the thorax and was put on record if it carried out at least one building activity per day, like accretion to one of the cells or to the comb envelope. From a practical standpoint, it is almost impossible to follow the activities of each worker over a 24-hr period in such a large number of breeding boxes. However, workers engaged in building do so over a relatively long stretch of time (20 to 30 min) and this, frequently, several times a day. So it is possible to record whether a worker has done any building at all, though more difficult to quantify the amount of building because quite frequently a worker demolishes an earlier construction and builds another. Moreover, at times the building is intermittent, i.e. a worker may build one day but not the following day. Consequently, the duration of building of any one worker was considered to be the time elapsed between the first and the last day of building. By observing the behaviour of the workers, one can easily determine which of them is not constructing at the moment or even predict which will not build anymore. Workers engaged in building are most of the time seen hanging by their legs from the lip of the comb cells, with their backs facing down. The cells usually contain brood at various stages. The workers nurse the brood during most hours of the day and rest at night in the afore-mentioned position, i.e. suspended under the cell outlets. Occasionally they descend the comb to forage for food or building materials, but promptly return, inspect the cells and suspend themselves underneath them. Cessation of building. This occurs in two stages: (1) The worker spends increasing periods of time on the upper surface of the comb rather than suspended underneath the cell outlets. It guards over the comb and occasionally brings food t o the brood or to other team-mates but does not forage for building materials.

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(2) The worker descends from the comb and spends most hours of the day on the floor of the nest or in the vicinity of the comb, but not on the comb proper or above it. If the breeding box is opened, such workers fly out and may spend long hours outside, frequently returning only with the dark of evening, or may never return. This is in marked contrast to the behaviour of workers who are building, who adhere to the comb evenupon a slight touch of the breeding box and if allowed outside the box, they fly a short distance and return to the comb ahnost immediately. Figure 5 presents in graph form the span of building (in days) of 210 workers. About 7 per cent of the workers never engage in any building, a phenomenon observed also in experiments with larger worker groups. Of the total workers, 86 per cent built for time intervals ranging from 3 days to 3 weeks, while the remaining 7 per cent may build for longer periods of up to 43 days. None of the workers was seen to build on the first day of life, less than 10 per cent on the second day and almost half the workers on the fifth day, but maximal participation in the building was observed only on the fifteenth day of life. The effective building span is thus from the second day of life till about 2 to 3 weeks later, when most workers cease building. Discussion It seems generally true that almost any hornet, be it queen or worker, engages at one time or another in building activities, but whereas workers build spontaneously during the first days of life, queens do so, under natural conditions, only after the workers have ceased building, i.e. after the few winter months. The experiments described demonstrate that experimentally at least, this difference is not essential, because the young, as yet unfertilized queens, can be induced to build cells and oviposit soon after their eclosion, by experimental manipulation: either by amputating their wings or by placing them in the cool for several days. In the latter procedure, the cooling possibly interrupts the diapause of the queen and stimulates development of its ovaries, a process which in nature occurs in the spring. It is more difficult, however, to explain the induction of building by amputation of the wings. Studies on other social insects, like those of ant colonies, have shown that after the queen is fecundated during the nuptial flight, she

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DAYS Fig. 5. Graphic presentation of the duration of building by the workers. The graph is based on a study of 210 workers from the same colony who

were maintainedin groups of sevenindividualsin thirtyseparate but identical breeding boxes.

alights on the ground and before entering the future nest, she sheds her wings. A similar observation has been made for the queens of termites (Imms 1960). The observation on the position of the wings of hibernating hornet queens have suggested that the distal edges of the fore-wings, at least, play some role in their sensorial perception. Various experiments described above have confirmed that assumption. It is possible therefore, that in hornet queens the amputation of the wings stimulates the development of the ovaries, and this will set off a sequertce of instinctive activities which commences with the gathering of building materials, the building of cells and the process of oviposition. On the matter of geotropism, workers and queens construct cells whose outlet faces down, but young queens that receive oestrogens in

their diet build in irregular fashion, with no regard to geotropic considerations. We may speculate that in the young queen, the building and oviposition are strongly depressed, which is ecologically reasonable when one considers that were the queen to expend energy and found a nest in the autumn, both she and the nest would face extinction due to the adverse conditions in the winter. Apparently di-ethinylestradiol partially counteracts this depression, enabling the queen to 'dabble' in construction; however, the genes responsible for programming the sequence of building activities in the desired geotropic and geometric directions are apparently still depressed, so that the queen's building is disorganized and haphazard. Amputation of the wings apparently counteracts the depression even further, while cooling of the queen for several days probably extinguishes the

ISHAY: COMB BUILDING BY THE HORNET depression altogether, enabling proper building, the development of the ovaries and oviposition, and stimulating all the instincts associated with nursing of the brood. All young queens that build in the autumn construct queen cells only. This has been reported also by Spradbery (1973) for young queens of the Australian Paravespula germaniea. It would seem that in V. orientalis too, the building of queen cells by young queens is due to climatic factors. On the other hand constant stroboscopic illumination probably nullifies or modifies the solar spectrum effects which find their expression in the spectral make-up of solar radiation during autumn, or in the length of the day, and consequently the young queens construct combs with worker cells even during the autumn. As to the problem of interrelations between queens: the attitude of one queen towards the other is apparently quite different during the autumn and winter than during the active period of the spring and summer. Thus several queens may exist peacefully in the winter following fertilization and up to the spring when each founds a new colony. At this point however, mutual antagonism sets in, so that any encounter between two rival queens will lead to violent combat to the death. We were able to induce young queens to build only in the autumn, at a time when even in nature they co-exist peacefully both inside and outside the nest. Why it was impossible to experimentally induce queens to build between January and March remains unclear. Neither is it clear why queens in the spring, following the hibernating period, become aggressive towards one another, individualistic and fanatically territorial with regards to the new nest founded by them. According to Nixon (1934) the functions of the wings are flight, ventilation and possibly communication of an alarm signal. In young hornets who have had their hind wings amputated, the typical spread of the fore-wings apparently points to a compensatory effect. The wings figure importantly in the building, at least in young hornets: they apparently enable the initiation of spatial perception (Ishay 1975), probably via mechanoreceptors such as the chordotonal sensillae which, in insects, are usually located on the bases of the wings (Dethier 1963). Amputation of the wings or induced changes in their symmetry result in changes in comb construction. On the other hand, if part of the contour of the wing is re-

81

tained intact, the peripheral perception is apparently unimpaired and construction proceeds in normal fashion. In more mature hornets the asymmetry produced by amputation or 'augmentation' of the wings is less important, but crossing of the wings does inhibit the building activities (and the ripening of the ovaries). All changes following wing amputation probably stem from erroneous and asymmetric perception of the Gestalt (Wertheimer 1912) and points to a variable degree of adaptation level to the new situation (Helson 1969). On the other hand, pentobarbital probably damages the sensory-motor system of the insect, even in doses that do not cause anaesthesia, and leads to such phenomena as lateral comb building and larger-sized comb cells. Witt (1956) noted that spiders fed on pentobarbital built denser webs than normal, which means that in this case the changes in building take the opposite form to that in the hornets, It seems that the queen pheromone exerts organizational influence on the construction by the workers. In the presence of a queen which secretes the pheromone, the building activities are centralized, i.e. the combs and cells are built close together and only in the primary nest (Ishay 1973). In the absence of a queen, but provided the synthetic pheromone is given with the food, even the amputated hornets build combs correctly. Such effects were not observed when they are fed on mammalian oestrogens or androgens. Mannose is a toxic sugar for bees (Sols Cadenas & Alvarado 1960; Arnold, Seitz & Lohr 1974) as well as hornets, while galactose is toxic for hornets (Fischl, Ishay & Talmor 1975). Both sugars cause delayed building and mannose also induces incorrect building. All the other sugars tested did not induce visual changes in building. Damaging of the tarsi or the tip of the antenna apparently also causes asymmetry in the spatial perception but, surprisingly enough, hornets with only a single intact antenna still construct combs with cells of proper shape and size. Workers which are not fed animal proteins (besides carbohydrates) do not build, probably because the stimulus for the production of oocytes in the ovaries is lacking and because ripening of the ovaries is apparently the primary drive towards building in workers deprived of a queen. In the absence of edible sugars (e.g. sucrose) in the food the workers fail to build and die within a few days because they cannot

82

ANIMAL

BEHAVIOUR,

obtain the energy necessary for daily activities from proteins, i.e. unlike the larvae, they are incapable of gluconeogenesis (Ishay et al. 1967). Substances which delay the onset of building or which induce incorrect building (e.g. improper attachment Of the combs, and the construction of cells which are not uniform in size, shape or levelling) are probably toxic or exert a nonspecific effect on various parameters associated with the building. The cell outlets invariably face down throughout all the described manipulations and this is probably a clear-cut case of geotaxis. Light is required for the construction of a compact comb, but the building itself does not necessitate light inasmuch as in nature the oriental hornet usually constructs its nest underneath the soil surface. Correct combs are built in the dark by hornets that had spent several days in the light. It is possible that some exposure to light is necessary for the proper orientation of the hornet prior to the start of construction and that failure to do so renders them incapable of constructing compact combs. The building of a comb on a clod of soil on the floor as in the transfer experiments, is unlike anything we have previously observed in the comb building of hornets. Ordinarily the hornets build the comb underneath the roof of the breeding box, which is the highest point in the box, and this occurs also in nature. We may conjecture that construction of a comb without the usual pedMe and directly onto a clod of soil results from the urgent need of the workers to oviposit the eggs that had ripened in their ovaries, and hence the 'short-cut'. In other words, rather than expend time in studying the architecture of the breeding box, hauling the soil from the floor to the roof of the box and engaging in the construction of a pedicle and symmetrical cells around it, the workers simply process the building materials in situ and oviposit immediately. Once they have started ovipositing in a particular site, they do not leave it but rather continue to expand the comb already started, in this case, close to the floor of the breeding box. It is also possible, however, that upon repeated transfer from box to box, the workers lose part of their spatial orientation, possibly something to do with their perception of height. The change in the building by yotmg amputated queens from ahnost single wall-attached cells to a comb structure probably points to

24,

1

preferences for stimuli of progressively increasing complexity with increased exposure in the choice situation. Building near, or on the ground by consecutively transferred groups of workers represents most probably a satiation effect due to the continuous period of exposure (Walker 1970). It seems to us that the manner of construction by hornets maintained in the dark from the moment of eclosion or by hornets maintained in groups of two to four individuals, and the failure of single hornets to construct, are all manifestations of sensory deprivation. Additionally, however, they point to a b~tsic difference between queens and workers: the former build correctly spontaneously (in the spring) only when they are solitary, whereas the latter require some social interaction with their team-mates for correct building. Our general impression is that both queens and workers engage in building and nursing the brood for a period of time sufficient to rear one generation of hornets at least from egg to pupa, a process requiring 16 to 21 days (Ishay 1962; W a f a & Sharkawi 1972) and at most, from egg to imago (29 to 42 days). The development time of the brood of other species of Vespinae is not much different than that for the brood of V. orientalis (Janet 1895; Spradbery 1973; Kemper & D6hring 1967). It would seem that the building of the initial comb and the nursing of the contained brood by the queen lasts as long as is necessary to raise a first generation of progeny, and that each of the ecloding workers in the natural nest is capable of advancing t h e construction and c a r e of the brood through one more generation. However, whereas workers grown in queenless colonies build combs and then oviposit and raise one generation (of males only), the workers of a queen-right colony build the combs and nurse one generation of brood but do not oviposit. It follows then that in a natural nest, the queen ensures the constancy of oviposition while the young workers ensure, each in its own time, the continuity of construction in the nest. In the autumn, despite the presence of numerous workers in the nest, construction ceases because none of the workers is young enough. At that time, there is eclosion of young queens which, however, do not engage i n building until they have undergone hibernation. Thus w e have a situation where the old queen does not continue to oviposit, the young queens are not yet building, while the workers are all old and do

ISHAY: COMB B U I L D I N G BY THE H O R N E T n o t b u i l d o r n u r s e t h e b r o o d . All these ditions, t a k e n t o g e t h e r , are w h a t r e n d e r the of hornets discontinuons and explain c o l o n i e s o f V. orientalis are a n n u a l a n d perennial.

connest why not

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