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Influence of Diet on the Shedding of Candida glabrata by Experimentally Infected Preweaned Calves
The Veterinary Journal 2002, 164, 275±279 doi:10.1053/tvjl.2002.0731, available online at http://www.idealibrary.com on
Influence of Diet on the Shedding of Candida glabrata by Experimentally Infected Preweaned Calves D. ELAD, J. BRENNER, A. MARKOVITCS, S. SHLOMOVITZ, J. BASAN and N. SCHWARTZ The Kimron Veterinary Institute, P.O. Box 12, Bet Dagan, 50250 Israel
SUMMARY The influence of three diets, comprising dam's milk (DM) from the same farm, commercial milk replacer with (CMRL) or without 3.2% lactose (CMR), on the duration and intensity of Candida glabrata shedding in the faeces of preweaned calves following experimental oral infection was examined. Shedding of other potential enteric pathogens was also monitored. The duration and intensity of C. glabrata shedding in DM-fed calves were reduced significantly compared with the calves fed the two diets based on milk replacers. Consequently, feeding calves with DM might disrupt the infective cycle, resulting in the yeast's elimination from a farm. In the CMR and CMRL groups, the periods of intensive shedding of C. glabrata and rotavirus overlapped but no diarrhoea was associated with the shedding of either microorganism. There was no evidence that lactose diminished colonization in vivo. # 2002 Published by Elsevier Science Ltd.
KEYWORDS: Candida galbrata; calves; shedding; diet.
INTRODUCTION Faecal samples and intestinal contents of preweaned calves contain yeasts, among which Candida glabrata has been shown to be the most prevalent (Gedek, 1969; Elad et al., 1998) with the abomasum the target organ for C. glabrata colonization. It has been suggested that the presence of yeasts in the gastrointestinal tract (GIT) might be associated with neonatal calf diarrhoea (Elad et al., 1998) in naturally infected animals. Lactose has been reported to reduce the adhesion of C. glabrata to epithelial cell cultures (Brown et al., 1998). It is notable that the formulation of commercial milk replacers for newborn calves differs significantly from that of dam's milk (DM) and that one of the components missing in the former is lactose. The prevalence of yeasts in general, and of C. glabrata in particular, in the GIT of milk-fed calves is lower than in calves fed with milk replacers
(Gedek, 1969). The objective of our experiment was to assess the influence of feeding two milk replacers of different composition or DM, on the ability of C. glabrata to colonize the GIT of preweaned calves as expressed by the yeast's shedding. The transfer of immunoglobulins (Ig) by colostrum (lactogenic immunity) is of paramount importance in ruminants (Brenner, 1991). Low levels of circulatory immunoglobulins (<8 mg ml ÿ 1) (McGuire & Adams, 1982), resulting from intestinal malabsorption or colostral deficiencies, may result in the morbidity and mortality of young animals from various pathogens, including opportunists (Blom, 1982; Saif & Smith, 1985). To exclude the possible influence of a deficiency in lactogenic immunity on the outcome of the experiment, postcolostral levels of immunoglobulins were assessed. MATERIALS AND METHODS
Animals
Correspondence to: Daniel Elad, DVM, PhD, Head, Department of Clinical Bacteriology and Mycology, Kimron Veterinary Institute, P.O. Box 12, 50250 Bet Dagan, Israel. Tel.: 972-3-9681688; Fax: 972-3-9681753; E-mail: elad@ agri.huji.ac.il
Calves were housed in individual ground-level pens, and were attended by personnel experienced in the rearing of young cattle. During the first 4 days of their lives, the calves were fed 2 litres of first
1090-0233/02/$ ± see front matter # 2002 Published by Elsevier Science Ltd.
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THE VETERINARY JOURNAL, 164, 3
colostrum, pooled from cows on the farm, at 12 h intervals. Dams were immunized with Rotavac (Coopers), an inactivated anti-rotavirus and F5 enterotoxigenic Escherichia coli vaccine. From the fourth day onwards the calves were fed, twice daily, with 2 litres of either DM or 250 g Bullstart Beef (Denkavit) suspended in tepid water, with or without 3.2% (w/v) lactose. Dry food, composed of corn and cottonseed, concentrates, hay and milk powder, was available ad libitum in addition to fresh water. Calves were assigned to the experimental groups by order of birth. A faecal sample from each calf was examined before its inclusion in the experiment, as described below.
Yeast strain
The C. glabrata strain used for the experimental infection was isolated from the intestinal contents of a diarrhoeic calf (not from the farm of origin of the experiment animals), which had been submitted for routine examination to the Kimron Veterinary Institute. Identification was made with the ID 32C kit (BioMeÂrieux SA). The strain was lyophilized and kept in vials at ÿ20 C. Before the experiment, a colony forming unit count (CFU) was performed on two vials by culturing serial dilutions on Sabouraud Dextrose Agar. Based on this count, a vial was rehydrated and aliquots of 5 ml saline containing 108 CFU of C. glabrata were prepared. Calves were inoculated within 30 min of rehydration.
Experimental design
The experiment was conducted on 15 calves, divided into three groups of five animals: Group 1: calves fed with commercial milk replacer (CMR) Group 2: calves fed with commercial milk replacer with 3.2% lactose (CMRL) Group 3: calves fed with dam's milk (DM)
Inoculation
Each calf, aged 5±6 days, was given 108 CFU of C. glabrata, suspended in 5 ml sterile saline, per os. Thereafter faecal samples were taken daily from the rectum of each calf. Calves were considered negative if no yeasts were isolated on two consecutive days.
Microbiological examinations
Tests to detect yeasts, bacteria (F5 ETEC, Salmonella spp.), viruses (rotavirus, coronavirus) and protozoa (Cryptosporidium parvum) were performed on all faecal samples, including the ones taken on the day before inoculation, as previously described
(Elad et al., 1998). Growth of C. glabrata on CHROMagar Candida (CHROMagar) was scored as 1 (low) for 20 CFU or less, 2 (intermediate) for more than 20 CFU of non-confluent growth and 3 (high) for confluent growth.
Levels of lactogenic immunity
Levels of circulatory immunoglobulins in sera, sampled before inoculation with C. glabrata, were measured by a commercial agar-gel-precipitating test (VMRD) as previously described (Brenner et al., 1992).
Statistical analysis
For the statistical analysis, the sum of scores observed for each calf while shedding the yeast was computed. Due to the relatively low number of animals, the Mann±Whitney U-test was chosen to compare any differences between the experimental groups. A twotailed distribution of the data was assumed. RESULTS
Microbiological examinations
All the calves included in this experiment were free of detectable levels of C. glabrata at the onset of the study but started shedding the yeast 24 h after the inoculation and continued doing so for variable periods. Detailed results and shedding score sums are presented in Fig. 1 and Table I respectively. No statistically significant differences (P 0.55) were found between the C. glabrata shedding score sums of the two milk replacer-fed groups. All calves in these two groups shed intermediate (score 2) quantities of C. glabrata for varying periods. Two calves in the CMR group and three calves in the CMRL group shed high (score 3) quantities of the yeast. In the DM-fed calves, shedding was observed for two 2±12 days but only one calf was colonized by C. glabrata, as judged by the duration and quantity of excreted yeast, whereas three others excreted low (score 1) levels of the yeast for the 2±3 days that followed inoculation, perhaps indicating transition through the GIT with no colonization. One additional calf excreted low levels of C. glabrata for 6 days, possibly as a result of a limited, transitory colonization. The difference between the number of C. glabrata shedding score sums of the DM group and that of each of the other two groups was statistically significant (P 0.0159 in both cases). Rotavirus was found in ten of the calves for various periods during the experiment. In calves belonging to the CMR and CMRL groups, excretion of rotavirus frequently coincided with the most intensive
DIET AND SHEDDING OF CANDIDA GLABRATA BY CALVES
277
Fig. 1. Duration and degree of Candida glabrata shedding by calves in various dietary groups after oral inoculation of 108 CFU of the yeast.
connection between the diarrhoea and the intensity of C. glabrata and/or rotavirus shedding was found.
Table I Sum of Candida glabrata shedding scores Diet
Milk replacer without lactose Milk replacer with lactose Dam's milk
Serum immunoglobulins
Calf 1
2
3
4
5
11
15
33
23
38
30
12
20
23
14
14
6
3
3
2
C. glabrata shedding. Detailed results are presented in Fig. 1. This phenomenon was not observed in the DM-fed animals. No F5 ETEC, Salmonella spp., C. parvum or coronavirus were found in the faecal samples on any occasion in any calf.
Clinical relevance of colonization by C. glabrata and rotavirus
Intermittent mild diarrhoea was observed in all calves in the experiment, for various periods. No
No calf was hypogammaglobulinaemic (i.e. with serum Ig levels <8 mg mlÿ1).
DISCUSSION Our results indicate that the ability of lactose to reduce the adhesive capability of C. glabrata observed in vitro (Brown et al., 1998) was not demonstrable in vivo. The statistical analysis indicates that calves fed with DM shed C. glabrata for shorter periods and with less intensity than those fed with either CMR or CMRL. In fact, based on the shedding pattern, C. glabrata did not colonize the GIT of at least three, possibly four of the five calves fed with DM. Although the antimicrobial activity of milk, especially the lactoperoxidase and lactoferrin system, against various microorganisms, including C. albicans, C. tropicalis and Cryptococcus neoformans, has been investigated, primarily in vitro (Pruitt &
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THE VETERINARY JOURNAL, 164, 3
Kamau, 1994; Tomita, 1994; Vorland et al., 1998), information regarding the specific activity of milk on fungal pathogens in vivo is scant. Cow's milk increased the risk of diaper rash in infants compared with human milk (Lopez-Martinez & Ruiz-Maldonado, 1982), possibly indicating that milk's antimicrobial activity is species related. Bovine lactoferricin had a pronounced antibacterial and antimycotic activity, in fact more so than similar peptides of human, murine and caprine origin (Vorland et al., 1998). Other factors may, however, contribute to inhibiting the colonization of the GIT by C. glabrata, as this yeast is more resistant than several other yeasts to the activity of bovine lactoferrin in vitro (Xu et al., 1999). We have observed on several farms (unpublished data) that a change in diet from CMS to DM will stop calves shedding C. glabrata. The results of our experiment indicate that a `susceptibility window' for infection with C. glabrata exists in calves up to the age of about 3 weeks (Elad et al., 1998). Following that age, probably because of the development of a competitive microflora, C. glabrata is no longer shed. Consequently, the infective cycle in a herd is likely to be maintained by direct passage from one calf to another. A similar occurrence was seen in the present study, in calves not included in the experiment. The low level and brief period of C. glabrata shedding in DM-fed calves might render the continuity of the infective cycle impossible and consequently lead to the disappearance of the yeast from the farm. In CMR- and CMRL-fed calves, an increase in C. glabrata shedding occurred concurrently with an increase in rotavirus shedding (to detectable levels). This phenomenon was not observed in DM-fed calves. This might be the result of an interaction between these microorganisms or result from the activity of one or more additional factors influencing the shedding of both. The connection between C. glabrata and rotavirus shedding underlines the so far overlooked importance of the former as a component in the multifactorial nature of enteric diseases in newborn animals (Brenner et al., 1993). The clinical significance of colonization by C. glabrata in calves is not clear. The possibility of a connection between the colonization of the GIT of preweaned calves by C. glabrata and diarrhoea was suggested previously (Elad et al., 1998), but such an association was not clearly established in the present trial. Thus, C. glabrata does not differ in two characteristics from other putative enteric pathogens, such as salmonella, rotavirus and protozoa as, firstly, it is present in apparently healthy animals
and, secondly, experimental infection does not readily reproduce the clinical signs that the microorganisms cause under natural conditions. Consequently, further investigations of the host±parasite interaction between C. glabrata and calves are necessary to elucidate its clinical significance. REFERENCES BLOM, J. Y. (1982). The relationship between serum immunoglobulin values and the incidence of respiratory diseases and enteritis in calves. Nordisk Veterinñr Medicin 34, 276±88. BRENNER, J. (1991). Passive lactogenic immunity in calves: a review. Israel Journal of Veterinary Medicine 46, 1±12. BRENNER, J., NERIA, A., ASKENAZY, G., PAZ, R., MEIROM, R., UNGAR-VARON, H. & TRAININ, Z. (1992). A lactogenicimmune-deficiency-syndrome in cows: unexplained phenomenon. Veterinary Immunology and Immunopathology 32, 315±324. BRENNER, J., ELAD, D., MARKOVITCS, A., GRINBERG, A. & TRAININ, Z. (1993). Epidemiological study of calf diarrhoea in Israel ± a one year survey of faecal samples. Israel Journal of Veterinary Medicine 48, 113±16. BROWN, A. J. P., CORMACK, B. P., GOW, N. A. R., KVAAL, C., SOLL, D. R. & SRIKANTHA, T. (1998). Advances in the molecular genetics of Candida albicans and Candida glabrata. Journal of Medical Mycology 36 (Suppl. 1), 230±7. ELAD, D., BRENNER, J., MARKOVICS, A., SCHLOMOVITS, S. & BASAN, J. (1998). Possible involvement of Candida glabrata in neonatal calf diarrhoea. Mycopathologia 141, 7±14. GEDEK, B. (1969). Besiedlung des Verdauungstraktes vom Kalb mit Hefen. Zentralblatt fuÈr Bakteriologie, Parasitenkunde, Infektionskrankheiten und Hygiene Abteilung 1 Origineale 211, 94±101. LOPEZ-MARTINEZ, M. & RUIZ-MALDONADO, R. (1982). Candidiasis in children with diaper rash. Study of 140 cases. Medicina Cutanea Ibero-Latina-Americana 10, 225±230. MCGUIRE, T. C. & ADAMS, D. S. (1982). Failure of colostral immunoglobulin transfer to calves: prevalence and diagnosis. Compendium on Continuing Education 4, S35±S40. PRUITT, K. M. & KAMAU, D. N. (1994). Quantitative analysis of bovine lactoperoxidase system components and of the effect of the activated system on bacterial growth and survival. In: Proceedings of the International Dairy Federation Seminar: Indigenous Antimicrobial Agents of Milk, Recent Developments, Aug. 31±Sep. 1, 1993, pp. 73±88, Uppsala, Sweden: International Dairy Federation, Brussels. SAIF, L. J. & SMITH, K. L. (1985). Enteric infections of calves and passive immunity. Journal of Dairy Science 68, 206±28. TOMITA. M. (1994). Active peptides of lactoferrin. In: Proceedings of the International Dairy Federation Seminar: Indigenous Antimicrobial Agents of Milk, Recent Developments, Aug. 31±Sep. 1, 1993, pp. 7±13, Uppsala, Sweden: International Dairy Federation, Brussels.
DIET AND SHEDDING OF CANDIDA GLABRATA BY CALVES
VORLAND, L. H., ULVATNE, H., ANDERSEN, J., HAUKLAND, H. H., REKDAL, é., SVENDSEN, J. S. & GUTTENBERG, T. J. (1998). Lactoferricin of bovine origin is more active than lactoferricins of human, murine and caprine origin. Scandinavian Journal of Infectious Diseases 30, 513±517.
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XU, Y. Y., SAMARANAYAKE, Y. H., SAMARANAYAKE, L. P. & NIKAWA, H. (1999). In-vitro susceptibility of Candida species to lactoferrin. Medical Mycology 37, 35±41. (Accepted for publication 11 March 2002)
Book Review doi: 10.1053/tvjl.2001.0694, available online at http://www.idealibrary.com on
Digestive Physiology of Pigs
Eds. Lindberg, J.E. and Ogle, B. Wallingford, Oxon, CABI Publishing, 2001. 416 pp. £75 (hard) ISBN 0851995179
The 8th Symposium on digestive Physiology in Pigs was held at the Swedish University of Agricultural Science, Uppsala, on 20±22 June 2000. This book is the proceedings of that meeting and so is not a comprehensive textbook. Rather, it describes a diversity of individual research projects, 107 in all. Sadly, only four of these are single-author reports. The structure is provided by placing the contributions within six areas: Gut development and function (15); the gastrointestinal immune system (3); nutrient absorption and utilization by the gut (7); digestive processes (43); econutrition and health maintenance (20) and finally free communications (19). Each starts with a valuable review. The contributions that follow are mini-papers, two to four pages in length, with a standard format and fortified by tables and figures. At the end of the book is a useful 10-page subject index. Technically the book is excellent, well printed, well illustrated, easy to read and to find your way around, and barely a mistake could I find. And for those with a limited attention span there is much to be said for contributions only a couple of pages in length! Proceedings give more than a view of current research; they reveal some of the limitations of the research world too. It seems, at least in the prosperous Western world, that pig research is shaped by the nature of the pig industry. There, pigs are usually highly selected, intensively reared, prolific, productive and profitable, and almost all
the research was on such pigs. And since the Symposium took place in Europe, under the auspices of the European Association of Animal Production, it is not surprising that three-quarters of the contributions were from European laboratories, Denmark heading the field, with the remainder (bar two from Brazil) from the USA, Canada and Australia. The preface remarks that the Symposium attracted delegates from countries worldwide, but in fact no contribution came from anywhere in Asia or Africa. Many of the multitude of pigs in these continents, China especially, complement mankind as scavengers and foragers, rather than competing for arable crops, so it seems a pity this book ignores them. The physiology and metabolism of a foraging sow, a splendidly adapted herbivore with a really quite large fermentation vat in its caeco-colon, may be able to interest the curious physiologist quite as much as the grazing ruminant does. In his concluding remarks, J.P. Laplace, the wise old grandfather of much current pig research, suggests it is time for gut physiologists to move onward from their traditional concern with digestibility. They should now focus on cellular mechanisms in the gut and their genetic basis, and on the intricate interactions of nutrition, gut function and the immune and endocrine systems of the body. The 8th Symposium showed that some movement in these directions is underway. We must wait for the 9th Symposium, in Canada, to see how far it will carry physiological understanding.
R. N. B. KAY
Influence of Diet on the Shedding of Candida glabrata by Experimentally Infected Preweaned Calves D. ELAD, J. BRENNER, A. MARKOVITCS, S. SHLOMOVITZ, J. BASAN and N. SCHWARTZ The Kimron Veterinary Institute, P.O. Box 12, Bet Dagan, 50250 Israel
SUMMARY The influence of three diets, comprising dam's milk (DM) from the same farm, commercial milk replacer with (CMRL) or without 3.2% lactose (CMR), on the duration and intensity of Candida glabrata shedding in the faeces of preweaned calves following experimental oral infection was examined. Shedding of other potential enteric pathogens was also monitored. The duration and intensity of C. glabrata shedding in DM-fed calves were reduced significantly compared with the calves fed the two diets based on milk replacers. Consequently, feeding calves with DM might disrupt the infective cycle, resulting in the yeast's elimination from a farm. In the CMR and CMRL groups, the periods of intensive shedding of C. glabrata and rotavirus overlapped but no diarrhoea was associated with the shedding of either microorganism. There was no evidence that lactose diminished colonization in vivo. # 2002 Published by Elsevier Science Ltd.
KEYWORDS: Candida galbrata; calves; shedding; diet.
INTRODUCTION Faecal samples and intestinal contents of preweaned calves contain yeasts, among which Candida glabrata has been shown to be the most prevalent (Gedek, 1969; Elad et al., 1998) with the abomasum the target organ for C. glabrata colonization. It has been suggested that the presence of yeasts in the gastrointestinal tract (GIT) might be associated with neonatal calf diarrhoea (Elad et al., 1998) in naturally infected animals. Lactose has been reported to reduce the adhesion of C. glabrata to epithelial cell cultures (Brown et al., 1998). It is notable that the formulation of commercial milk replacers for newborn calves differs significantly from that of dam's milk (DM) and that one of the components missing in the former is lactose. The prevalence of yeasts in general, and of C. glabrata in particular, in the GIT of milk-fed calves is lower than in calves fed with milk replacers
(Gedek, 1969). The objective of our experiment was to assess the influence of feeding two milk replacers of different composition or DM, on the ability of C. glabrata to colonize the GIT of preweaned calves as expressed by the yeast's shedding. The transfer of immunoglobulins (Ig) by colostrum (lactogenic immunity) is of paramount importance in ruminants (Brenner, 1991). Low levels of circulatory immunoglobulins (<8 mg ml ÿ 1) (McGuire & Adams, 1982), resulting from intestinal malabsorption or colostral deficiencies, may result in the morbidity and mortality of young animals from various pathogens, including opportunists (Blom, 1982; Saif & Smith, 1985). To exclude the possible influence of a deficiency in lactogenic immunity on the outcome of the experiment, postcolostral levels of immunoglobulins were assessed. MATERIALS AND METHODS
Animals
Correspondence to: Daniel Elad, DVM, PhD, Head, Department of Clinical Bacteriology and Mycology, Kimron Veterinary Institute, P.O. Box 12, 50250 Bet Dagan, Israel. Tel.: 972-3-9681688; Fax: 972-3-9681753; E-mail: elad@ agri.huji.ac.il
Calves were housed in individual ground-level pens, and were attended by personnel experienced in the rearing of young cattle. During the first 4 days of their lives, the calves were fed 2 litres of first
1090-0233/02/$ ± see front matter # 2002 Published by Elsevier Science Ltd.
276
THE VETERINARY JOURNAL, 164, 3
colostrum, pooled from cows on the farm, at 12 h intervals. Dams were immunized with Rotavac (Coopers), an inactivated anti-rotavirus and F5 enterotoxigenic Escherichia coli vaccine. From the fourth day onwards the calves were fed, twice daily, with 2 litres of either DM or 250 g Bullstart Beef (Denkavit) suspended in tepid water, with or without 3.2% (w/v) lactose. Dry food, composed of corn and cottonseed, concentrates, hay and milk powder, was available ad libitum in addition to fresh water. Calves were assigned to the experimental groups by order of birth. A faecal sample from each calf was examined before its inclusion in the experiment, as described below.
Yeast strain
The C. glabrata strain used for the experimental infection was isolated from the intestinal contents of a diarrhoeic calf (not from the farm of origin of the experiment animals), which had been submitted for routine examination to the Kimron Veterinary Institute. Identification was made with the ID 32C kit (BioMeÂrieux SA). The strain was lyophilized and kept in vials at ÿ20 C. Before the experiment, a colony forming unit count (CFU) was performed on two vials by culturing serial dilutions on Sabouraud Dextrose Agar. Based on this count, a vial was rehydrated and aliquots of 5 ml saline containing 108 CFU of C. glabrata were prepared. Calves were inoculated within 30 min of rehydration.
Experimental design
The experiment was conducted on 15 calves, divided into three groups of five animals: Group 1: calves fed with commercial milk replacer (CMR) Group 2: calves fed with commercial milk replacer with 3.2% lactose (CMRL) Group 3: calves fed with dam's milk (DM)
Inoculation
Each calf, aged 5±6 days, was given 108 CFU of C. glabrata, suspended in 5 ml sterile saline, per os. Thereafter faecal samples were taken daily from the rectum of each calf. Calves were considered negative if no yeasts were isolated on two consecutive days.
Microbiological examinations
Tests to detect yeasts, bacteria (F5 ETEC, Salmonella spp.), viruses (rotavirus, coronavirus) and protozoa (Cryptosporidium parvum) were performed on all faecal samples, including the ones taken on the day before inoculation, as previously described
(Elad et al., 1998). Growth of C. glabrata on CHROMagar Candida (CHROMagar) was scored as 1 (low) for 20 CFU or less, 2 (intermediate) for more than 20 CFU of non-confluent growth and 3 (high) for confluent growth.
Levels of lactogenic immunity
Levels of circulatory immunoglobulins in sera, sampled before inoculation with C. glabrata, were measured by a commercial agar-gel-precipitating test (VMRD) as previously described (Brenner et al., 1992).
Statistical analysis
For the statistical analysis, the sum of scores observed for each calf while shedding the yeast was computed. Due to the relatively low number of animals, the Mann±Whitney U-test was chosen to compare any differences between the experimental groups. A twotailed distribution of the data was assumed. RESULTS
Microbiological examinations
All the calves included in this experiment were free of detectable levels of C. glabrata at the onset of the study but started shedding the yeast 24 h after the inoculation and continued doing so for variable periods. Detailed results and shedding score sums are presented in Fig. 1 and Table I respectively. No statistically significant differences (P 0.55) were found between the C. glabrata shedding score sums of the two milk replacer-fed groups. All calves in these two groups shed intermediate (score 2) quantities of C. glabrata for varying periods. Two calves in the CMR group and three calves in the CMRL group shed high (score 3) quantities of the yeast. In the DM-fed calves, shedding was observed for two 2±12 days but only one calf was colonized by C. glabrata, as judged by the duration and quantity of excreted yeast, whereas three others excreted low (score 1) levels of the yeast for the 2±3 days that followed inoculation, perhaps indicating transition through the GIT with no colonization. One additional calf excreted low levels of C. glabrata for 6 days, possibly as a result of a limited, transitory colonization. The difference between the number of C. glabrata shedding score sums of the DM group and that of each of the other two groups was statistically significant (P 0.0159 in both cases). Rotavirus was found in ten of the calves for various periods during the experiment. In calves belonging to the CMR and CMRL groups, excretion of rotavirus frequently coincided with the most intensive
DIET AND SHEDDING OF CANDIDA GLABRATA BY CALVES
277
Fig. 1. Duration and degree of Candida glabrata shedding by calves in various dietary groups after oral inoculation of 108 CFU of the yeast.
connection between the diarrhoea and the intensity of C. glabrata and/or rotavirus shedding was found.
Table I Sum of Candida glabrata shedding scores Diet
Milk replacer without lactose Milk replacer with lactose Dam's milk
Serum immunoglobulins
Calf 1
2
3
4
5
11
15
33
23
38
30
12
20
23
14
14
6
3
3
2
C. glabrata shedding. Detailed results are presented in Fig. 1. This phenomenon was not observed in the DM-fed animals. No F5 ETEC, Salmonella spp., C. parvum or coronavirus were found in the faecal samples on any occasion in any calf.
Clinical relevance of colonization by C. glabrata and rotavirus
Intermittent mild diarrhoea was observed in all calves in the experiment, for various periods. No
No calf was hypogammaglobulinaemic (i.e. with serum Ig levels <8 mg mlÿ1).
DISCUSSION Our results indicate that the ability of lactose to reduce the adhesive capability of C. glabrata observed in vitro (Brown et al., 1998) was not demonstrable in vivo. The statistical analysis indicates that calves fed with DM shed C. glabrata for shorter periods and with less intensity than those fed with either CMR or CMRL. In fact, based on the shedding pattern, C. glabrata did not colonize the GIT of at least three, possibly four of the five calves fed with DM. Although the antimicrobial activity of milk, especially the lactoperoxidase and lactoferrin system, against various microorganisms, including C. albicans, C. tropicalis and Cryptococcus neoformans, has been investigated, primarily in vitro (Pruitt &
278
THE VETERINARY JOURNAL, 164, 3
Kamau, 1994; Tomita, 1994; Vorland et al., 1998), information regarding the specific activity of milk on fungal pathogens in vivo is scant. Cow's milk increased the risk of diaper rash in infants compared with human milk (Lopez-Martinez & Ruiz-Maldonado, 1982), possibly indicating that milk's antimicrobial activity is species related. Bovine lactoferricin had a pronounced antibacterial and antimycotic activity, in fact more so than similar peptides of human, murine and caprine origin (Vorland et al., 1998). Other factors may, however, contribute to inhibiting the colonization of the GIT by C. glabrata, as this yeast is more resistant than several other yeasts to the activity of bovine lactoferrin in vitro (Xu et al., 1999). We have observed on several farms (unpublished data) that a change in diet from CMS to DM will stop calves shedding C. glabrata. The results of our experiment indicate that a `susceptibility window' for infection with C. glabrata exists in calves up to the age of about 3 weeks (Elad et al., 1998). Following that age, probably because of the development of a competitive microflora, C. glabrata is no longer shed. Consequently, the infective cycle in a herd is likely to be maintained by direct passage from one calf to another. A similar occurrence was seen in the present study, in calves not included in the experiment. The low level and brief period of C. glabrata shedding in DM-fed calves might render the continuity of the infective cycle impossible and consequently lead to the disappearance of the yeast from the farm. In CMR- and CMRL-fed calves, an increase in C. glabrata shedding occurred concurrently with an increase in rotavirus shedding (to detectable levels). This phenomenon was not observed in DM-fed calves. This might be the result of an interaction between these microorganisms or result from the activity of one or more additional factors influencing the shedding of both. The connection between C. glabrata and rotavirus shedding underlines the so far overlooked importance of the former as a component in the multifactorial nature of enteric diseases in newborn animals (Brenner et al., 1993). The clinical significance of colonization by C. glabrata in calves is not clear. The possibility of a connection between the colonization of the GIT of preweaned calves by C. glabrata and diarrhoea was suggested previously (Elad et al., 1998), but such an association was not clearly established in the present trial. Thus, C. glabrata does not differ in two characteristics from other putative enteric pathogens, such as salmonella, rotavirus and protozoa as, firstly, it is present in apparently healthy animals
and, secondly, experimental infection does not readily reproduce the clinical signs that the microorganisms cause under natural conditions. Consequently, further investigations of the host±parasite interaction between C. glabrata and calves are necessary to elucidate its clinical significance. REFERENCES BLOM, J. Y. (1982). The relationship between serum immunoglobulin values and the incidence of respiratory diseases and enteritis in calves. Nordisk Veterinñr Medicin 34, 276±88. BRENNER, J. (1991). Passive lactogenic immunity in calves: a review. Israel Journal of Veterinary Medicine 46, 1±12. BRENNER, J., NERIA, A., ASKENAZY, G., PAZ, R., MEIROM, R., UNGAR-VARON, H. & TRAININ, Z. (1992). A lactogenicimmune-deficiency-syndrome in cows: unexplained phenomenon. Veterinary Immunology and Immunopathology 32, 315±324. BRENNER, J., ELAD, D., MARKOVITCS, A., GRINBERG, A. & TRAININ, Z. (1993). Epidemiological study of calf diarrhoea in Israel ± a one year survey of faecal samples. Israel Journal of Veterinary Medicine 48, 113±16. BROWN, A. J. P., CORMACK, B. P., GOW, N. A. R., KVAAL, C., SOLL, D. R. & SRIKANTHA, T. (1998). Advances in the molecular genetics of Candida albicans and Candida glabrata. Journal of Medical Mycology 36 (Suppl. 1), 230±7. ELAD, D., BRENNER, J., MARKOVICS, A., SCHLOMOVITS, S. & BASAN, J. (1998). Possible involvement of Candida glabrata in neonatal calf diarrhoea. Mycopathologia 141, 7±14. GEDEK, B. (1969). Besiedlung des Verdauungstraktes vom Kalb mit Hefen. Zentralblatt fuÈr Bakteriologie, Parasitenkunde, Infektionskrankheiten und Hygiene Abteilung 1 Origineale 211, 94±101. LOPEZ-MARTINEZ, M. & RUIZ-MALDONADO, R. (1982). Candidiasis in children with diaper rash. Study of 140 cases. Medicina Cutanea Ibero-Latina-Americana 10, 225±230. MCGUIRE, T. C. & ADAMS, D. S. (1982). Failure of colostral immunoglobulin transfer to calves: prevalence and diagnosis. Compendium on Continuing Education 4, S35±S40. PRUITT, K. M. & KAMAU, D. N. (1994). Quantitative analysis of bovine lactoperoxidase system components and of the effect of the activated system on bacterial growth and survival. In: Proceedings of the International Dairy Federation Seminar: Indigenous Antimicrobial Agents of Milk, Recent Developments, Aug. 31±Sep. 1, 1993, pp. 73±88, Uppsala, Sweden: International Dairy Federation, Brussels. SAIF, L. J. & SMITH, K. L. (1985). Enteric infections of calves and passive immunity. Journal of Dairy Science 68, 206±28. TOMITA. M. (1994). Active peptides of lactoferrin. In: Proceedings of the International Dairy Federation Seminar: Indigenous Antimicrobial Agents of Milk, Recent Developments, Aug. 31±Sep. 1, 1993, pp. 7±13, Uppsala, Sweden: International Dairy Federation, Brussels.
DIET AND SHEDDING OF CANDIDA GLABRATA BY CALVES
VORLAND, L. H., ULVATNE, H., ANDERSEN, J., HAUKLAND, H. H., REKDAL, é., SVENDSEN, J. S. & GUTTENBERG, T. J. (1998). Lactoferricin of bovine origin is more active than lactoferricins of human, murine and caprine origin. Scandinavian Journal of Infectious Diseases 30, 513±517.
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XU, Y. Y., SAMARANAYAKE, Y. H., SAMARANAYAKE, L. P. & NIKAWA, H. (1999). In-vitro susceptibility of Candida species to lactoferrin. Medical Mycology 37, 35±41. (Accepted for publication 11 March 2002)
Book Review doi: 10.1053/tvjl.2001.0694, available online at http://www.idealibrary.com on
Digestive Physiology of Pigs
Eds. Lindberg, J.E. and Ogle, B. Wallingford, Oxon, CABI Publishing, 2001. 416 pp. £75 (hard) ISBN 0851995179
The 8th Symposium on digestive Physiology in Pigs was held at the Swedish University of Agricultural Science, Uppsala, on 20±22 June 2000. This book is the proceedings of that meeting and so is not a comprehensive textbook. Rather, it describes a diversity of individual research projects, 107 in all. Sadly, only four of these are single-author reports. The structure is provided by placing the contributions within six areas: Gut development and function (15); the gastrointestinal immune system (3); nutrient absorption and utilization by the gut (7); digestive processes (43); econutrition and health maintenance (20) and finally free communications (19). Each starts with a valuable review. The contributions that follow are mini-papers, two to four pages in length, with a standard format and fortified by tables and figures. At the end of the book is a useful 10-page subject index. Technically the book is excellent, well printed, well illustrated, easy to read and to find your way around, and barely a mistake could I find. And for those with a limited attention span there is much to be said for contributions only a couple of pages in length! Proceedings give more than a view of current research; they reveal some of the limitations of the research world too. It seems, at least in the prosperous Western world, that pig research is shaped by the nature of the pig industry. There, pigs are usually highly selected, intensively reared, prolific, productive and profitable, and almost all
the research was on such pigs. And since the Symposium took place in Europe, under the auspices of the European Association of Animal Production, it is not surprising that three-quarters of the contributions were from European laboratories, Denmark heading the field, with the remainder (bar two from Brazil) from the USA, Canada and Australia. The preface remarks that the Symposium attracted delegates from countries worldwide, but in fact no contribution came from anywhere in Asia or Africa. Many of the multitude of pigs in these continents, China especially, complement mankind as scavengers and foragers, rather than competing for arable crops, so it seems a pity this book ignores them. The physiology and metabolism of a foraging sow, a splendidly adapted herbivore with a really quite large fermentation vat in its caeco-colon, may be able to interest the curious physiologist quite as much as the grazing ruminant does. In his concluding remarks, J.P. Laplace, the wise old grandfather of much current pig research, suggests it is time for gut physiologists to move onward from their traditional concern with digestibility. They should now focus on cellular mechanisms in the gut and their genetic basis, and on the intricate interactions of nutrition, gut function and the immune and endocrine systems of the body. The 8th Symposium showed that some movement in these directions is underway. We must wait for the 9th Symposium, in Canada, to see how far it will carry physiological understanding.
R. N. B. KAY