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Prevalence of four enteropathogens in the faeces of young diarrhoeic dairy calves in Switzerland
F. Lanz Uhde, T. Kaufmann, H. Sager, S. Albini, R. Zanoni, E. Schelling, M. Meylan
The prevalences of Cryptosporidium parvum, rotavirus, bovine coronavirus (BCV), and enterotoxigenic Escherichia coli (E coli K99) were determined in diarrhoeic dairy calves aged one to 21 days on 71 dairy farms in western Switzerland during the winter of 2005 to 2006. Faecal samples from 147 untreated calves suffering from acute diarrhoea were analysed by standardised diagnostic methods, and the immunoglobulin status of each calf was evaluated. The prevalences of C parvum, rotavirus, BCV and E coli K99 were 550 per cent, 587 per cent, 78 per cent and 55 per cent, respectively. The proportions of herds positive for the respective pathogens among the herds with diarrhoeic calves were 417 per cent, 521 per cent, 21 per cent and 21 per cent. The immunoglobulin concentration in the serum of 905 per cent of the diarrhoeic calves was below 8 g/l.

DIARRHOEA is one of the most serious diseases of neonatal calves. It causes major economic losses due to mortality, poor growth and the costs of treatment. It is a complex, multifactorial disease that is affected by the intrinsic characteristics of the calf, its nutritional and immunological status, the management of the herd, the environment, and various infectious agents (Bendali and others 1999a, Scott and others 2004). Calves are at the greatest risk of developing diarrhoea in the first month of life and the incidence of diarrhoea decreases with age (Frank and Kaneene 1993, Bendali and others 1999b). The four major enteropathogens associated with neonatal calf diarrhoea worldwide are Cryptosporidium parvum, rotavirus, bovine coronavirus (BCV) and enterotoxigenic Escherichia coli (E coli K99) (Garcia and others 2000). Salmonella species can also cause diarrhoea and septicaemia, particularly when calves are brought together from different farms and raised intensively on milk replacer diets (Naylor 2002). Other agents, such as Campylobacter jejuni and Campylobacter sputorum biovar faecalis, Clostridium species, parvovirus, calicivirus, bredavirus and torovirus are also pathogenic in calves, but their importance in the field is uncertain (Mansfeld and others 2007). There has recently been interest in Clostridium difficile (Songer 2004, Arroyo and others 2005, Songer and Anderson 2006, Hammitt and others 2008) as a potential cause of enteritis in calves, but further studies are needed to evaluate its role. The prevalence of the different aetiological agents varies with the calf s age, and multiple infections are common. The clinical signs induced by the different microorganisms are similar, and an aetiological diagnosis can only be reached in the laboratory. The identification of the infectious agent(s) involved is important for planning preventive measures and controlling the disease. However, after an agent has been identified, it is difficult to determine whether it is responsible for the diarrhoea in the individual or in the herd, because most of the agents are also found in healthy calves (Saif and Smith 1985, Janke 1989, Fagan and others 1995). The main aim of the present study was to determine the prevalence of C parvum, rotavirus, BCV and E coli K99 in dairy calves aged one to 21 days suffering from acute diarrhoea in dairy herds in western Switzerland at the individual and at the herd level. The belief that C parvum is the most common organism identified in cases of neonatal diarrhoea in the region was examined. In addition, the results of standardised laboratory methods were compared with the results obtained with rapid tests, and the immunological status (the level of serum immunoglobulins) of the diarrhoeic calves was investigated.

MATERIALS AND METHODS Farms, animals and collection of samples A list of 420 rural veterinary practices in Switzerland was established, and from this list 70 practices located within a radius of approximately 80 km from the Clinic for Ruminants of the University of Berne were selected randomly. Veterinarians were asked by mail to participate in the study. The sample size for herds with diarrhoeic calves was calculated for an estimated prevalence of C parvum of 50 per cent among calves with diarrhoea, allowing for clustering within herds, a standard error less than 5 per cent, and an intraclass coefficient rho of 03 (Bennett and others 1991); a sample size of 210 diarrhoeic calves in 70 herds was derived. The participating veterinarians were asked to consider the following inclusion criteria: dairy farms with more than 15 milking cows, and samples to be collected from three calves aged one to 21 days with acute diarrhoea of less than 48 hours duration from each farm. The samples had to be collected between October 1, 2005 and April 30, 2006. Calves that had been treated with either antibiotics or halofuginone, or that suffered from other diseases such as pneumonia, omphalitis or polyarthritis, and calves from suckler herds were excluded from the study. A herd was classified as positive for one of the four pathogens if at least two calves excreted the pathogen. Samples of faeces and serum were collected by the veterinarians from calves fulfilling the inclusion criteria before they were treated, and sent to the Clinic for Ruminants within a day. The name of each calf s owner, and its identification number, age, breed, sex and clinical condition were recorded on a standardised form. The clinical data included the calf s body temperature, heart rate, respiratory rate, general condition, appetite, suckling reflex, degree of dehydration, and the consistency of its faeces. Laboratory analyses The faecal samples were analysed for the presence of C parvum, rotavirus, BCV and E coli K99 by standard diagnostic methods. Oocysts of C parvum were detected by a modified Ziehl-Neelsen staining method (Kaufmann 1996). The samples were tested for rotavirus by antigen-capture ELISA (Pathfinder Rotavirus EIA; Bio-Rad Laboratories) according to the manufacturers instructions. The excretion of BCV was diagnosed by a non-commercial biphasic antigen-capture ELISA assay in 96-well microELISA plates (Immulon M 129 A; Dynatech). For the diagnosis of E coli K99, each sample was cultured on minimal casein agar (MINCA agar medium;
The Veterinary Record, September 20, 2008

Veterinary Record (2008) 163, 362-366 F. Lanz Uhde, DrMedVet, T. Kaufmann, DrMedVet, M. Meylan, DrMedVet, FVH, PhD, DrHabil, Clinic for Ruminants, H. Sager, DrMedVet, FVH, PhD, Institute of Veterinary Parasitology, S. Albini, DrMedVet, FVH, Institute of Veterinary Bacteriology, R. Zanoni, DrMedVet, DrHabil, Institute of Veterinary Virology, Vetsuisse Faculty, University of Berne, Bremgartenstrasse 109a, 3012 Berne, Switzerland E. Schelling, DrMedVet, PhD, Swiss Tropical Institute, Basel, Switzerland Correspondence to Dr Meylan

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TABLE 1: Percentages of the 147 diarrhoeic calves showing different clinical signs and the scores assigned to calculate an overall clinical score Clinical observation General condition Normal Slightly affected Moderately affected Severely affected Appetite Good Reduced Anorectic Suckling reflex Good Moderate Absent Degree of dehydration Not dehydrated Slightly dehydrated Moderately dehydrated Severely dehydrated Consistency of faeces Mushy Watery Scoring 0 1 2 4 0 2 4 0 2 4 0 1 2 4 0 4 Percentage 388 537 41 34 490 435 75 566 340 95 531 374 81 14 422 578

ence of the four pathogens, either individually or in combination, was evaluated with a logistic regression model. Similarly, the immunoglobulin levels were evaluated for their association with the occurrence of the four pathogens. Possible associations between the immunoglobulin levels and the age of the calves (in days), and between the immunoglobulin levels and the clinical scores were investigated by a linear regression model. To compare the results of the rapid tests with the results of the standard diagnostic methods, the test results were recorded as either positive or negative for each pathogen. The sensitivity and specificity of the rapid tests were compared with those of the standard diagnostic tests (Thrusfield 2005). The data were analysed by using Intercooled STATA 8.2 (StataCorp). RESULTS Samples of faeces and serum from 174 diarrhoeic calves in 79 dairy herds were submitted by 32 veterinary practitioners between October 2005 and April 2006; 147 of the calves from 71 herds fulfilled the inclusion criteria and only these samples were used for further analyses. The targeted sample size of three calves from each herd was not achieved; samples from four calves were available from two herds, from three calves from 24 herds, from two calves from 22 herds, and only one sample was sent from the remaining 23 herds. The mean age of the calves was 83 days (95 per cent confidence interval [CI] 77 to 89, range one to 20 days). The distribution of the breeds was as follows: 327 per cent red Holstein, 231 per cent fleckvieh (red Holstein x simmental), 156 per cent Holstein Friesian, 41 per cent brown Swiss and 245 per cent mixed breeds. There were 483 per cent males and 517 per cent females. Nearly all the calves (925 per cent) showed few or no clinical signs of deterioration in condition, and the other 75 per cent showed moderate to severe signs of lethargy and weakness. The suckling reflex was present in 905 per cent of the calves (Table 1). Enteric pathogens were found in 912 per cent of the faecal samples. The calculated individual prevalences were 55 per cent for C parvum, 587 per cent for rotavirus, 78 per cent for BCV and 55 per cent for E coli K99 (Table 2). There were no significant associations between the pathogens diagnosed and clinical scores of the calves. Infections with only one pathogen were found in 586 per cent of the samples. The most frequent combination was rotavirus and C parvum (19 per cent). The proportions of single and combined infections are given in Table 3. The herd prevalences were 417 per cent for C parvum, 521 per cent for rotavirus, 21 per cent for BCV and 21 per cent for E coli K99. When the herd prevalences were calculated on the basis of the results from only the first two calves sampled in each herd they were slightly lower for rotavirus and C parvum than when they were calculated on the basis of all results from herds from which at least two samples were available without consideration of the number of samples (two or three) per herd (Table 4). The analyses of the faecal samples with the four immunochromatographic rapid tests were positive for C parvum in 449 per cent and for rotavirus in 469 per cent of the samples. None of the samples were positive for BCV and only one sample was positive for E coli K99. The sensitivity and specificity of the rapid tests were therefore calculated only for the FASTest CRYPTO Strip (sensitivity 835 per cent, specificity 100 per cent) and the FASTest ROTA Strip (sensitivity 841 per cent, specificity 100 per cent). The serum immunoglobulin concentration of 905 per cent of the calves was less than 8 g/l (Fig 1). There were no significant associations between the serum immunoglobulin

Axcell) mixed with PolyViteX (PVX-M; BioMrieux) and incubated aerobically at 37C for 24 hours. Colonies of E coli were then identified phenotypically. The presence of F5 (K99) was examined by slide agglutination with monovalent F5 (K99)-rabbit-antiserum (Statens Serum Institute). In parallel, all the faecal samples were analysed by four rapid tests: FASTest CRYPTO Strip, FASTest ROTA Strip, FASTest BCV Strip and FASTest E coli K99 Strip (MegaCor Diagnostics). They were based on immunochromatographic techniques and used as described by the manufacturer. The results were compared with those obtained by the established laboratory methods. The serum immunoglobulin levels were determined with the Paragon Serum Protein Electrophoresis Kit (Beckman Coulter) according to the manufacturers instructions. Statistical analyses For the individual calves with diarrhoea, logistic regression models with random effect on the herd level (xtlogit procedure) were used to calculate estimates of the prevalence of each of the four enteropathogens and for the 12 possible combinations. Clustering within herds (random effect in the model) was taken into consideration because the calves within the same herd would be less independent than calves from different herds (Bendali and others 1999b). At the herd level, the proportions of positive herds among the herds with diarrhoeic calves (herd prevalence) was calculated on the basis of the classification of herds as positive for a given pathogen when at least two calves excreted it; all other herds where at least two samples were available were considered negative for this infectious agent. Herds from which only one sample had been submitted were not included in the calculation of herd prevalences. In addition, the herd prevalences were calculated by using only the results from the first two calves examined in each herd, according to the same classification criteria. A clinical score for the calves was generated on the basis of the clinical data to test for possible associations between the clinical signs and infection with specific pathogens (Naciri and others 1999). The sum of the scores for the following five variables were used: general condition, appetite, suckling reflex, degree of dehydration and consistency of faeces (Table 1). The association of the clinical scores with the presThe Veterinary Record, September 20, 2008

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TABLE 2: Percentages of the 147 calves with diarrhoea that were positive for Cryptosporidium parvum, rotavirus, bovine coronavirus (BCV), and Escherichia coli K99 by standard laboratory methods and their mean (95 per cent confidence interval [CI]) calculated prevalences Infectious agent C parvum Rotavirus
BCV

TABLE 3: Percentages of the 147 calves with diarrhoea that were positive for Cryptosporidium parvum, rotavirus, bovine coronavirus (BCV), Escherichia coli K99 or combinations of these agents Infectious agent C parvum Rotavirus
BCV

Positive (%) 286 272 14 14 190 27 0 07 54 14 20 07 0 0 07 88

Percentage positive 537 558 88 95

Calculated prevalence (%) (95 per cent CI) 550 (405-688) 587 (429-728) 78 (27-209) 55 (17-167)

E coli K99

levels and the clinical scores or age of the calves or the pathogens diagnosed. DISCUSSION Diarrhoea in neonatal calves is one of the major problems in Swiss dairy herds (Sommer 1994, Strk and others 1997), but there have been few studies of the pathogens causing the problem (Luginbhl and Pfister 1996, Strk and others 1997, Luginbhl and others 2005). There is little information available for dairy calves owing to the small numbers of animals studied and the lack of detailed information on individual animals, but there have been larger scale investigations of suckler cow operations (Busato and others 1997a, b, 1998, Steiner and others 1997, Lentze and others 1999). The aim of this study was therefore to determine the prevalence of C parvum, rotavirus, BCV and E coli K99 in one to 21-dayold calves with acute diarrhoea in 71 dairy herds in western Switzerland with neonatal calf diarrhoea as a herd problem. The immunoglobulin levels in the serum of the calves with diarrhoea were also evaluated because a failure of passive transfer is a well-known risk factor for neonatal disease in calves (Barrington and others 2002). The incidence of diarrhoea in calves is highest during the winter months and the greatest risk for a calf occurs during its first month of life (Frank and Kaneene 1993, Bendali and others 1999a, Gutzwiller 2002, Scott and others 2004, Mansfeld and others 2007). For these reasons, samples were collected between October and April and only calves less than three weeks of age were included. The best diagnostic results are obtained early in the course of the disease (Scott and others 2004, Mansfeld and others 2007) and previous treatment with antimicrobials or halofuginone can lead to false-negative test results. The calves that had suffered from diarrhoea for more than 48 hours when examined by the veterinarian or had already been treated by the owner were therefore excluded from the study. After a pathogen had been diagnosed, the farmers wanted to take preventive measures (particularly treatment with halofuginone in the case of cryptosporidiosis) as soon as possible, and no further sampling was therefore possible from untreated calves after the initial diagnosis. When they were examined, the majority of the calves were in good general condition and systemic signs of disease were mostly mild. This was probably due to the fact that only calves that had had acute diarrhoea for less than 48 hours were sampled. As a result, no association was found between the pathogens detected and the clinical scores of the calves. The calves were sampled only once, and single sampling may give falsenegative results because rotavirus, BCV and cryptosporidia are excreted in a cyclical pattern. Furthermore, intermittent shedding of pathogens has been reported in both healthy and diseased animals (Trotz-Williams and others 2005, Singh and others 2006), and it is known that healthy control animals can

E coli K99 C parvum, rotavirus C parvum, BCV C parvum, E coli K99 Rotavirus, BCV Rotavirus, E coli K99 BCV, E coli K99 C parvum, rotavirus, BCV C parvum, rotavirus, E coli K99 C parvum, BCV, E coli K99 Rotavirus, BCV, E coli K99 C parvum, rotavirus, BCV, E coli K99 No pathogen

become diarrhoeic shortly after being sampled; the method of selecting healthy controls in herds with diarrhoea may therefore lead to bias (Busato and others 1998, Luginbhl and others 2005). Finally, it is often not possible to sample non-diarrhoeic calves on farms with a high morbidity. For these reasons a healthy control group within the herds under study was not included. Herds with three samples available were more likely to have two calves positive for one of the pathogens, and the herd prevalences were accordingly slightly higher in this analysis than when only the first two calves sampled were used in the analysis, a method that gives a more conservative estimate of the prevalences of the pathogens. The distribution and the relative prevalences of the different pathogens were the same in both analyses. The decision to restrict herd positivity to the herds in which pathogens had been found in at least two calves was made because the four pathogens have been reported to be excreted by healthy animals in a contaminated environment (Scott and other 2004). However, finding the same pathogen in two calves with acute diarrhoea suggests that it is likely to be implicated in the pathogenesis of the disease in the herd. Infection with more than one pathogen was observed in more than 40 per cent of the calves. In agreement with previous surveys worldwide, the most common mixed infection was rotavirus and C parvum (Snodgrass and others 1986, de la Fuente and others 1999, Garcia and others 2000, Joachim and others 2003, Hfle 2005). There is good evidence that rotavirus is a primary pathogen causing acute diarrhoea in neonatal calves (Torres-Medina and others 1985, Reynolds and others 1986, Snodgrass and others 1986, Busato and others 1998, Garcia and others 2000), and that it can be detected more frequently in the faeces of diarrhoeic than of non-diarrhoeic calves (De Rycke and others 1986, Snodgrass and others 1986, Busato and others 1998, Haschek and others 2006). In this study rotavirus was the infectious agent excreted most commonly by the calves with diarrhoea, not only alone but also in mixed infections with other pathogens, indicating that rotavirus has a predominant role in the pathogenesis of neonatal calf diarrhoea in Switzerland. C parvum was first reported in association with calf diarrhoea by Panciera and others (1971). It also has the potential to cause disease in immunocompromised human beings (Fayer and others 2000). Today, it is found worldwide with a prevalence in young calves of between 14 per cent and over 80 per cent, depending on the age, health and housing of the
The Veterinary Record, September 20, 2008

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Calves with diarrhoea (%)

TABLE 4: Mean (95 per cent confidence interval [CI]) herd prevalences of the four infectious agents calculated for 48 herds from which at least two samples were available Infectious agents Cryptosporidium parvum Rotavirus Bovine coronavirus Escherichia coli K99 Herd prevalence (%) calculated on the basis of results from Two or three calves in each herd First two calves only in each herd 417 (272-561) 521 (374-667) 21 (0-63) 21 (0-63) 354 (214-495) 375 (233-517) 0 21 (0-63)

50 40 30 20 10 0 0-2 >2-4 >4-6 >6-8 >8-10 Immunoglobulin concentration (g/l) >10

animals (Garber and others 1994, Lentze and others 1999, Naciri and others 1999, De la Fuente and others 1999, Garcia and others 2000, Lefay and others 2000, Joachim and others 2003, Trotz-Williams and others 2005, Haschek and others 2006, Singh and others 2006). A strong and highly significant association between infection with C parvum and the occurrence of diarrhoea has been observed in studies by Naciri and others (1999), Joachim and others (2003), TrotzWilliams and others (2005), Singh and others (2006) and Haschek and others (2006). Calves shedding oocysts have a three times higher risk of being diarrhoeic than uninfected calves. A highly significant association between the numbers of oocysts being shed and the presence of diarrhoea has also been suggested (Trotz-Williams and others 2005). The prevalence of C parvum in calves up to four weeks of age has probably been underestimated. Not all laboratories have included this parasite in their routine test panel and some researchers have considered cryptosporidiosis to be of minor importance. Although cryptosporidia are now acknowledged as primary pathogens, not all veterinarians seem to be aware of their importance (Joachim and others 2003). In the present study, 286 per cent of the diarrhoeic calves were shedding the organism alone, but it was not the most prevalent infectious agent associated with diarrhoea in the calves because rotavirus was detected more frequently. However, the estimates of prevalence had broadly overlapping confidence intervals, indicating that rotavirus and C parvum were of similar significance. The relatively low prevalence of BCV agrees with the results of other studies in Switzerland (Busato and others 1998, Luginbhl and others 2005), and similar observations have been made in Sweden (Bjrkman and others 2003). No classical cases of neonatal diarrhoea caused by E coli K99 were observed in either of these studies. In contrast, BCV appeared to be of major importance in Austria (Haschek and others 2006). The results of this study do not support an important role for BCV in causing diarrhoea in these calves. However, BCV is known to infect calves up to three months of age (Scott and others 2004), whereas the mean age of the calves was only about eight days. It is possible that BCV would have been more prevalent in older calves. The results of the study should only be extrapolated to older calves with caution. The relationship between the serum immunoglobulin concentration of neonatal calves and their risk of developing diarrhoea is controversial. Some authors have observed a clear correlation between diarrhoea and low levels of serum immunoglobulin (Barrington and others 2002, Lipp 2005), whereas others have observed no correlation (Donovan and others 1998, Gutzwiller 2002). The results of a survey by Gutzwiller (2002) indicated that high blood levels of maternal antibodies were of minor importance for the protection of calves against diarrhoea caused by viruses and cryptosporidia; it was hypothesised that immunoglobulins have to be present in the gut rather than in the blood in order to protect calves from viral enteritis. Donovan and others (1998) found that low serum immunoglobulin levels in newborn calves were a significant risk factor for septicaemia and pneumonia, but
The Veterinary Record, September 20, 2008

FIG 1: Percentages of the 147 calves with diarrhoea that had different ranges of immunoglobulin concentration in their serum

not for diarrhoea. However, Par and others (1993) suggested that high immunoglobulin levels decrease the duration of episodes of diarrhoea. The results of this study show that over 90 per cent of the calves with diarrhoea had less than 8 g/l of immunoglobulins in their serum (a level which is generally considered to be too low to provide adequate passive immunity), and that more than 50 per cent of them had less than 4 g/l. More than 10 g/l is considered to be necessary to provide ideal passive immunity (Besser and others 1991). These results indicate that it is important to ensure that neonatal dairy calves in Switzerland receive adequate quantities of colostrum. The results suggest that the rapid commercial tests for field use are satisfactory for the detection of C parvum and rotavirus in faecal samples. There were too few cases of BCV and E coli K99 for the results of the other two tests to be interpreted. The use of rapid tests solely for the two most prevalent pathogens cannot be recommended, because it is important to investigate all the possible causes in herds with neonatal calf diarrhoea; tests not only for rotavirus and C parvum but also for BCV and E coli should be included in investigations of diarrhoea in young calves. The results of this study show that rotavirus and C parvum are both widely prevalent in Swiss dairy herds in neonatal calves with diarrhoea. It is important to identify the infectious agents involved so that a suitable approach to the control of the disease can be taken, and appropriate advice on colostrum feeding, calf nutrition, hygiene and therapeutic regimens can be given (Scott and others 2004). The low levels of serum immunoglobulins in most of the calves indicate that they had received insufficient colostrum. ACKNOWLEDGEMENTS The authors thank the veterinary practitioners for their help with sample collection and the technical staff for the laboratory work. The rapid tests used in this study were in part provided by Veterinaria.

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simplified general method for cluster-sample surveys of health in developing countries. Rapport Trimestriel de Statistiques Sanitaires Mondiales 44, 98-106 BESSER, T. E., GAY, C. C. & PRITCHETT, L. (1991) Comparison of three methods of feeding colostrum to dairy calves. Journal of the American Veterinary Medical Association 198, 419-422 BJRKMAN, C., SVENSSON, C., CHRISTENSSON, B. & DE VERDIER, K. (2003) Cryptosporidium parvum and Giardia intestinalis in calf diarrhoea in Sweden. Acta Veterinaria Scandinavica 44, 145-152 BUSATO, A., LENTZE, T., HOFER, D., BURNENS, A., HENTRICH, B. & GAILLARD, C. (1998) A case control study of potential enteric pathogens for calves raised in cow-calf herds. Journal of Veterinary Medicine 45, 519-528 BUSATO, A., STEINER, L., MARTIN, S. W., SHOUKRI, M. M. & GAILLARD, C. (1997a) Calf health in cow-calf herds in Switzerland. Preventive Veterinary Medicine 30, 9-22 BUSATO, A., STEINER, L., TONTIS, A. & GAILLARD, C. (1997b) Frequency and etiology of calf losses and calf diseases in cow-calf farms. I. Methods of data collection, calf mortality, and calf morbidity. Deutsche Tierrztliche Wochenschrift 104, 131-135 DE LA FUENTE, R., LUZON, M., RUIZ-SANTA-QUITERIA, J. A., GARCIA, A., CID, D., ORDEN, J. A., GARCIA, S., SANZ, R. & GOMEZ-BAUTISTA, M. (1999) Cryptosporidium and concurrent infections with other major enteropathogens in 1 to 30-day old diarrheic dairy calves in central Spain. Veterinary Parasitology 80, 179-185 DE RYCKE, J., BERNARD, S., LAPORTE, J., NACIRI, M., POPOFF, M. R. & RODOLAKIS, A. (1986) Prevalence of various enteropathogens in the faeces of diarrhoeic and healthy calves. Annales de Recherches Vtrinaires 17, 159-168 DONOVAN, G. A., DOHOO, I. R., MONTGOMERY, D. M. & BENNET, F. L. (1998) Associations between passive immunity and morbidity and mortality in dairy heifers in Florida, USA. Preventive Veterinary Medicine 34, 31-46 FAGAN, J. G., DWYER, P. J. & QUINLAN, J. G. (1995) Factors that may affect the occurrence of enteropathogens in the feces of diarrhoeic calves in Ireland. Irish Veterinary Journal 48, 17-21 FAYER, R., MORGAN, U. & UPTON, S. J. (2000) Epidemiology of Cryptosporidium: transmission, detection and identification. International Journal for Parasitology 30, 1305-1322 FRANK, N. A. & KANEENE, J. B. (1993) Management risk factors associated with calf diarrhea in Michigan dairy herds. Journal of Dairy Science 76, 1313-1323 GARBER, L. P., SALAMAN, M. D., HURD, H. S., KEEFE, T. & SCHLATER, J. L. (1994) Potential risk factors for Cryptosporidium infection in dairy calves. Journal of the American Veterinary Medical Association 205, 86-91 GARCIA, A., RUIZ-SANTA-QUITERIA, J. A., ORDEN, J. A., CID, D., SANZ, R., GOMEZ-BAUTISTA, M. & DE LA FUENTE, R. (2000) Rotavirus and concurrent infections with other enteropathogens in neonatal diarrheic calves in Spain. Comparative Immunology, Microbiology and Infectious Diseases 23, 175-183 GUTZWILLER, A. (2002) Effect of colostrum intake on diarrhoea incidence in newborn calves. Schweizer Archiv fr Tierheilkunde 144, 59-64 HAMMITT, M. C., BUESCHEL, D. M., KEEL, M. K., GLOCK, R. D., CUNEO, P., DEYOUNG, D. W., REGGIARDO, C., TRINH, H. T. & SONGER, J. G. (2008) A possible role of Clostridium difficile in the etiology of calf enteritis. Veterinary Microbiology 127, 343-352 HASCHEK, B., KLEIN, D., BENETKA, V., HERRERA, C., SOMMERFELDSTUR, I., VILCEK, S., MOESTL, K. & BAUMGARTNER, W. (2006) Detection of bovine torovirus in neonatal calf diarrhoea in Lower Austria and Styria (Austria). Journal of Veterinary Medicine B 53, 160-165 HFLE, A. (2005) Querschnittsstudie zum Vorkommen von Campylobacter spp, Yersinia enterocolitica, Salmonellen, Rotaviren und Kryptosporidium parvum in Milchviehbetrieben mit Klberdurchfallproblemen des nrdlichen Baden-Wrtemberg, Inaugural dissertation, Berlin, Freie Universitt Berlin JANKE, B. H. (1989) Protecting calves from diarrhea. Veterinary Medicine 84, 803-811 JOACHIM, A., KRULL, T., SCHWARZKOPF, J. & DAUGSCHIES, A. (2003) Prevalence and control of bovine cryptosporidiosis in German dairy herds. Veterinary Parasitology 112, 277-288 KAUFMANN, J. (1996) Staining Cryptosporidia spp in faecal smears. In Parasitic Infections of Domestic Animals. Basel, Birkhuser. p 9 LEFAY, D., NACIRI, M., POIRIER, P. & CHERMETTE, R. (2000) Prevalence

of Cryptosporidium infection in calves in France. Veterinary Parasitology 89, 1-9 LENTZE, T., HOFER, D., GOTTSTEIN, B., GAILLARD, C. & BUSATO, A. (1999) Prevalence and importance of endoparasites in calves raised in Swiss cow-calf farms. Deutsche Tierrztliche Wochenschrift 106, 275-281 LIPP, K. (2005) Feldstudie zur kolostralen Immunglobulin-Versorgung neugeborener Klber in Abhngigkeit von der Verweildauer beim Muttertier. Inaugural dissertation. Munich, Ludwig-Maximilian-Universitt Mnchen LUGINBHL, A. & PFISTER, K. (1996) Die Kryptosporidiose des Kalbes als schwerwiegendes Bestandesproblem. Schweizer Archiv fr Tierheilkunde 138, 195-200 LUGINBHL, A., REITT, K., METZLER, A., KOLLBRUNNER, M., CORBOZ, L. & DEPLAZES, P. (2005) Feldstudie zu Prvalenz und Diagnostik von Durchfallerregern beim neonatalen Kalb im Einzugsgebiet einer schweizerischen Nutztierpraxis. Schweizer Archiv fr Tierheilkunde 147, 245-252 MANSFELD, R., HOEDEMAKER, M., MARTIN, R. & DE KRUIF, A. (2007) Diarrhoe whrend des ersten Lebensmonats. In Tierrztliche Bestandesbetreuung beim Milchrind. 2nd edn. Eds A. de Kruif, R. Mansfeld, M. Hoedemaker. Stuttgart, Enke. pp 152-156 NACIRI, M., LEFAY, M. P., MANCASSOLA, R., POIRIER, P. & CHERMETTE, R. (1999) Role of Cryptosporidium parvum as a pathogen in neonatal diarrhoea complex in suckling and dairy calves in France. Veterinary Parasitology 85, 245-257 NAYLOR, M. N. (2002) Neonatal ruminant diarrhea. In Large Animal Internal Medicine. 3rd edn. Ed B. P. Smith. St Louis, Mosby. pp 352-366 PANCIERA, R. J., THOMASSEN, R. W. & GARNER, F. M. (1971) Cryptosporidial infection in a calf. Veterinary Pathology 8, 479-484 PAR, J., THURMOND, M. C., GARDNER, I. A. & PICANSO, J. P. (1993) Effect of birth weight, total protein, serum IgG and packed cell volume on risk of neonatal diarrhoea in calves on two Californian dairies. Canadian Journal of Veterinary Research 57, 241-246 REYNOLDS, D. J., MORGAN, J. H., CHANTER, N., JONES, P. W., BRIDGER, J. C., DEBNEY, T. G. & BUNCH, K. J. (1986) Microbiology of calf diarrhoea in southern Britain. Veterinary Record 119, 34-39 SCOTT, P. R., HALL, G. A., JONES, P. W. & MORGAN, J. H. (2004) Calf diarrhoea. In Bovine Medicine. Diseases and Husbandry of Cattle. 2nd edn. Ed A. H. Andrews. Oxford, Blackwell Publishing. pp 185-214 SAIF, L. J. & SMITH, K. J. (1985) Enteric viral infections of calves and passive immunity. Journal of Dairy Science 68, 206-228 SINGH, B. B., SHARMA, R., KUMAR, H., BANGA, H. S., AULAKH, R. S., GILL, J. P. S. & SHARMA, J. K. (2006) Prevalence of Cryptosporidium parvum infection in Punjab (India) and its association with diarrhea in neonatal dairy calves. Veterinary Parasitology 140, 162-165 SNODGRASS, D. R., TERZOLO, H. R., SHERWOOD, D., CAMPBELL, I., MENZIES, J. D. & SYNGE, B. A. (1986) Aetiology of diarrhoea in young calves. Veterinary Record 119, 31-34 SOMMER, T. (1994) Ein Umfrageprojekt zur Klberhaltung in der Schweiz Ergebnisse zu Haltung, Ftterungstechnik, Krankheiten und Geburt. Schweizer Archiv fr Tierheilkunde 136, 382-389 SONGER, J. G. (2004) The emergence of Clostridium difficile as a pathogen of food animals. Animal Health Research Reviews 5, 321-326 SONGER, J. G. & ANDERSON, M. A. (2006) Clostridium difficile: an important pathogen of food animals. Anaerobe 12, 1-4 STRK, K. D., FREI-STHELI, C., FREI, P. P., PFEIFFER, D. U., DANUSER, J., AUDIGE, L., NICOLET, J., STRASSER, M., GOTTSTEIN, B. & KIHM, U. (1997) Frequency and cost of health problems in Swiss dairy cows and their calves (1993-1994). Schweizer Archiv fr Tierheilkunde 139, 343-353 STEINER, L., BUSATO, A., BURNENS, A. & GAILLARD, C. (1997) Frequency and etiology of calf losses and calf diseases before weaning in cow-calf farms. II. Microbiological and parasitological diagnoses in diarrheic calves. Deutsche Tierrztliche Wochenschrift 104, 169-173 THRUSFIELD, M. V. (2005) Diagnostic testing. In Veterinary Epidemiology. 3rd edn. Oxford, Blackwell Publishing. pp 305-330 TORRES-MEDINA, A., SCHLAFER, D. H. & MEBUS, C. A. (1985) Rotaviral and coronaviral diarrhoea. Veterinary Clinics of North America: Food Animal Practice 1, 471-493 TROTZ-WILLIAMS, L. A., JARVIE, B. D., MARTIN, S. W., LESLIE, K. E. & PEREGRINE, A. S. (2005) Prevalence of Cryptosporidium parvum infection in southwestern Ontario and its association with diarrhea in neonatal dairy calves. Canadian Veterinary Journal 46, 349-351

The Veterinary Record, September 20, 2008