Pathology of calves with diarrhoea in southern Britain

Twenty-one moribund calves with diarrhoea were purchased from 11 farms, their faeces examined for enteropathogens and samples of intestinal tissue removed under anaesthesia. Lesions and presence of enteropathogens on the mucosal surface were scored by histological examination of immunostained paraffin sections. Two or more enteropathogens were detected in 19 calves. Cryptosporidium appeared to be the principal cause of diarrhoea in six calves, rotavirus in four, Salmonella typhimurium in two, bacteria adherent to the surface of the large intestine in two, coronavirus in one and K99+ Escherichia coli in one calf. Diarrhoea in four calves was the consequence of mixed infections in which no one enteropathogen appeared to predominate. In one calf no enteropathogen was detected. Diarrhoea was associated with infections and lesions throughout the small and large intestines. The enteropathogens most frequently associated with lesions in the small intestines were rotavirus, coronavirus and cryptosporidium; in the large intestines they were coronavirus and bacteria apparently adherent to the mucosal surface.     

Aetiology of diarrhoea in young calves

Faeces samples were collected from 302 untreated calves on the day of onset of diarrhoea and from 49 healthy calves at 32 farms experiencing outbreaks of diarrhoea. At least four diarrhoeic calves were sampled on each farm, and samples were examined for rotavirus, coronavirus, cryptosporidium, enterotoxigenic Escherichia coli and Salmonella species. Although all these enteropathogens were excreted more frequently by the diarrhoeic than by the healthy calves, the difference was significant overall only for rotavirus. Rotavirus was excreted by 18 per cent of healthy calves, coronavirus by 4 per cent, cryptosporidium by 14 per cent, and no enterotoxigenic E coli or Salmonella species were detected. The most common enteropathogen in diarrhoeic calves was rotavirus, which was excreted by more than half the diarrhoeic calves on 18 farms. Coronavirus was excreted at a similar high prevalence on one farm, cryptosporidium on five farms and enterotoxigenic E coli on three farms. Concurrent infection with two or more microorganisms occurred in 15 per cent of diarrhoeic calves. There was no difference in the isolation rate of campylobacters between diarrhoeic and healthy calves.     

Evaluation of ELISA and electron microscopy for the detection of coronavirus and rotavirus in bovine faeces

Enzyme-linked immunosorbent assay (ELISA) was compared with electron microscopy in the examination of faeces from experimental calves and showed 100 per cent agreement in the detection of 19 bovine coronavirus and 15 bovine rotavirus electron microscope positive samples. In a limited field survey of calf diarrhoea 75 selected faeces were examined independently by ELISA and electron microscopy and the agreement between the two tests was 95 per cent for coronavirus and 84 per cent for rotavirus. A further comparison was made with 74 samples submitted for routine diagnosis and this yielded agreements of 82 per cent (coronavirus) and 89 per cent (rotavirus). Factors contributing to discrepant results were examined and the relative advantages and disadvantages of the two tests for routine detection of these enteric viruses are discussed.     

Longitudinal survey of rotavirus infection in calves

A longitudinal survey of rotavirus infection in heifer calves was carried out on a closed Friesian dairy herd over two successive calving seasons. Rotavirus was detected by electron microscopy in the faeces of 45 of 57 (79 per cent) calves examined. On average the virus was first detected at 6.1 days of age. Clinically the disease associated with rotavirus infection was of mild to moderate severity. Only one infected calf required intravenous fluid therapy. Diarrhoea or excretion of abnormal faeces was associated with rotavirus infection in 58 per cent of infected calves, while in the remaining 42 per cent infection was subclinical. The cycle of rotavirus infection was broken by thorough cleansing and disinfection of the calf house.     

Role of Cryptosporidium parvum as a pathogen in neonatal diarrhoea complex in suckling and dairy calves in France.

This study was carried out to find the importance of Cryptosporidium parvum in diarrhoea of neonatal calves in two types of breeding - suckling and dairy calves - in France. Different agents causing neonatal diarrhoea, E. coli, rotavirus, coronavirus, Salmonella and Cryptosporidium were systematically researched in faeces. 1. Suckling calves: In 40 livestock farms selected for diarrhoea, 311 calves 4 to 10 days old which had diarrhoea for less than 24h or no diarrhoea, were included in the study. A prophylaxis of neonatal diarrhoea had been carried out in 21 of the 40 livestock farms. On D0 (inclusion day), the mean age was 6 days, 82% presented a good initial general condition and 76.2% had a good appetite; 48.6% were diarrhoeic but 91.3% presented no sign of dehydration. Only 6.1% were infected by E. coli K99, 14.3% by rotavirus, 6.8% by coronavirus, 0.3% by Salmonella but 50% excreted C. parvum oocysts. This later percentage increases up to 84% and 86% by D3 and D7, respectively . We note that 16% of the 4-day-old calves on D0 are excreting oocysts and this percentage increases as a function of the age of the calf on D0 to reach 90% to 95% by the age of 8 days. 10 out of 12 dead calves excreted C. parvum oocysts. From D0 to D14 the other pathogen agents show a relative or a decreasing stability. 2. Dairy calves: 382 calves which had diarrhoea for less than 24 h or no diarrhoea, aged 8 to 15 days coming from six industrial livestock farms were included in the study. On D0, 99% of the calves presented a good initial general condition, 99.7% had a good appetite and no calf was dehydrated. At this date (D0), 16.8% of the calves excreted cryptosporidia. This percentage increases up to 23% and 51.8% on D3 and D8, respectively, then decreases to 31.9% on D14. The pressure of the other pathogenicagents remains relatively stable, excepted for rotavirus on D7 (from 9.9% on D0 to 27.2% on D7, then 12.6% on D14) which does not explain the concomitantpeak in diarrhoea because the infection by rotavirus on D7 is more frequent in non-diarrhoeic calves than in diarrhoeic calves. Our results show that Cryptosporidium prevalence is higher in suckling than in dairy calves and C. parvum constitutes actually in both cases the major aetiological agent of neonatal diarrhoea.     

Intestinal antibody response after vaccination and infection with rotavirus of calves fed colostrum with or without rotavirus antibody

The intestinal and systemic antibody response of calves vaccinated and/or challenged with rotavirus was studied employing isotype-specific ELISAs for the detection of IgG1, IgG2, IgM and IgA antibodies to rotavirus. Monoclonal antibodies to bovine immunoglobulin isotypes of proven specificity were used as conjugated or catching antibody. Five days after oral inoculation (dpi) of a 5-day-old gnotobiotic calf with rotavirus, IgM rotavirus antibodies were excreted in faeces, followed 5 days later by IgA rotavirus antibodies. The increase in IgM rotavirus antibody titre coincided with the inability to detect further rotavirus excretion. Faeces IgM and IgA rotavirus antibody titres fell to low levels within 3 weeks post infection. IgG1 and IgG2 rotavirus antibodies were not detected in faecal samples. In serum, antibodies to rotavirus of all four isotypes were detected, starting with IgM at 5 dpi. Two SPF-calves, which were fed colostrum free of rotavirus antibodies, were vaccinated with a modified live rotavirus vaccine and challenged with virulent rotavirus 6 days later. Upon vaccination, the calves showed an antibody response similar to the response of the infected gnotobiotic calf. Intestinal IgM rotavirus antibodies were excreted before or on the day of challenge and appeared to be associated with protection against challenge infection with virulent virus and rotavirus-induced diarrhoea. In 3 control calves, which were challenged only, the antibody patterns also resembled that of the gnotobiotic calf and again the appearance of IgM rotavirus antibodies coincided with the end of the rotavirus detection period. Two other groups of 3 SPF-calves were treated similarly, but the calves were fed colostrum with rotavirus antibodies during the first 48 h of life. These calves excreted passively acquired IgG1 and IgG2 rotavirus antibodies in their faeces from 2 to 6 days after birth. After vaccination, no IgM or IgA antibody activity in serum or faeces was detectable. Upon challenge, all calves developed diarrhoea and excreted rotavirus. Seven to 10 days after challenge low levels of IgM rotavirus antibody were detected for a short period. These data indicate that the intestinal antibody response of young calves to an enteric viral infection is associated with the excretion of IgM antibodies, immediately followed by IgA antibodies. This response is absent or diminished in calves with passively acquired specific antibodies which may explain the failure to induce a protective intestinal immune response by oral vaccination with modified live rotavirus of calves fed colostrum containing rotavirus antibodies.     

The studies on the aetiology of diarrhoea in neonatal calves and determination of virulence gene markers of Escherichia coli strains by multiplex PCR

The purpose of this study was to determine aetiological agents of diarrhoea in neonatal calves and to investigate virulence gene markers of Escherichia coli strains isolated from calves by multiplex polymerase chain reaction (PCR). Eighty-two diarrhoeic calves and 18 healthy calves were used as subjects. Faeces were taken from the rectums of all the calves and were subjected to bacterial culture. Antigen enzyme-linked immunosorbent assay (ELISA) was performed to detect rotavirus, coronavirus and E. coli K99 in faeces of all the calves. A multiplex PCR was used to characterize E. coli strains in all the calves. Escherichia coli was isolated from 37 faeces samples, Enterococcus ssp. was isolated from 22 faeces samples and Salmonella was isolated from one faeces sample in diarrhoeic calves. Furthermore, only E. coli was isolated from all 18 faeces samples of healthy calves. Of the 37 E. coli isolated from diarrhoeic calves, K99 (18.9%), F41 (18.9%), heat-stable enterotoxin a (STa) (18.9%), Shiga toxin 1 (Stx1; 13.5%) and Shiga toxin 2 (Stx2; 5.4%) and intimin (8.1%) genes were identified by multiplex PCR. Of the 18 E. coli isolated from healthy calves, K99 (16.6%) and intimin (55.5%) genes were identified by PCR. A total of 15 rotavirus, 11 coronavirus and 11 E. coli K99 were detected in diarrhoeic calves by the antigen ELISA. As a result, this study shows that rotavirus, coronavirus, E. coli and Enterococcus ssp. were determined to play a role in the aetiology of diarrhoea in the neonatal calves. K99, F41, STa, Stx1 and Stx2 were found as the most common virulence gene markers of E. coli strains isolated from calves with diarrhoea. Multiplex PCR may be useful for characterization of E. coli isolated from calves.     

Enteric infections in veal calves: A longitudinal study on four veal calf units

Summary

Forty‐five calves on four veal calf units were monitored during the first four weeks after their arrival. Faecal samples were collected on alternate days and screened for the presence of rotaviruses, bovine coronavirus, Cryptosporidium oocysts, K99 positive strains of E. coli and Salmonella spp. Rotaviruses and Cryptosporidium were the most commonly detected agents (78% and 60% respectively of the calves). Bovine coronavirus was detected in the faeces of 18% of the calves, whilst K99 positive E. coli was only found in 2 samples from one calf. Salmonella spp. were not isolated from any of the 646 faecal samples examined. Shedding of rotaviruses occurred in a bimodal pattern beginning in the first week of the survey. Cryptosporidium oocysts were detected most frequently in the interval between the two peaks of rotavirus shedding. The presence of rotaviruses or Cryptosporidium oocysts in faeces was not strongly associated with scour, nor were combined infections with these agents or the cases of bovine coronavirus infection. The condition of the calves throughout the survey was generally satisfactory.     

Agents associated with neonatal diarrhoea in Ethiopian dairy calves

Faeces samples collected from diarrhoeic dairy calves in the first 8 weeks of life were examined for the presence of 5 enteropathogens. The majority of the 108 diarrhoea cases occurred in the first 5 weeks of life and a commercial ELISA kit detected bovine enteric coronavirus (BEC) in 38.9%, serogroup A rotavirus (RV) in 16.7% and K99 (F5) fimbrial adhesin-positive Escherichia coli (K99 ETEC) in 11.1 per cent. Concurrent infections of these enteropathogens were detected in 14.8% of samples (30.8% of samples positive for these agents). No evidence of cryptosporidial infection was found using a differential staining method on faecal smears nor was salmonella excretion detected. On 2 of the 8 farms only BEC was present; the other 6 farms were positive for all 3 agents. It is concluded that BEC is the major infectious cause of neonatal calf diarrhoea in the Ethiopian dairy herds studied with RV and K99 ETEC also contributing to morbidity, either alone or as mixed infections.     

Microbiology of calf diarrhoea in southern Britain

Faeces samples from calves with diarrhoea in 45 outbreaks were examined for six enteropathogens. Rotavirus and coronavirus were detected by ELISA in 208 (42 per cent) and 69 (14 per cent) of 490 calves respectively; calici-like viruses were detected by electron microscopy in 14 of 132 calves (11 per cent). Cryptosporidium were detected in 106 of 465 (23 per cent), Salmonella species in 58 of 490 (12 per cent) and enterotoxigenic Escherichia coli bearing the K99 adhesin (K99+ E coli) in nine of 310 calves (3 per cent). In the faeces of 20 per cent of calves with diarrhoea more than one enteropathogen was detected; in 31 per cent no enteropathogen was found. Faces samples from 385 healthy calves in the same outbreaks were also examined. There was a significant statistical association of disease with the presence of rotavirus, coronavirus, Cryptosporidium and Salmonella species (P less than 0.001). Healthy calves were not examined for calici-like viruses and the association of K99+ E coli with disease was not analysed because there were too few positive samples. Rotavirus infections were more common in dairy herds and single suckler beef herds whereas Salmonella infections were more often found in calf rearing units. Cryptosporidium were more common in single and multiple suckler beef herds. K99+ E coli were found in one dairy herd and one multiple suckler beef herd both with unhygienic calving accommodation. Variations in coronavirus detection among different farm types were not statistically significant. In this survey rotavirus was the most commonly detected agent in calf diarrhoea and Cryptosporidium were found in approximately one quarter of affected calves. Infection with Salmonella species was widespread, but K99+ E coli infections were less common in the United Kingdom than in other countries.     

Evaluation of a combined rotavirus and enterotoxigenic Escherichia coli vaccine in cattle

A vaccine of rotavirus and K99 antigen from enterotoxigenic Escherichia coli was emulsified in oil adjuvant and administered intramuscularly to pregnant cows. Calves born to and reared on vaccinated dams were protected against experimental rotavirus infection at five days old when compared with calves from unvaccinated control cows. Field trials of the vaccine were carried out in 40 commercial herds, in which half the cows in each herd were selected at random for vaccination and half were left unvaccinated. In 31 herds (2641 cows) there was no significant diarrhoea problem (less than 10 per cent morbidity); these herds were excluded from further analysis. The nine remaining herds did experience a calf diarrhoea problem of greater than 10 per cent morbidity, but on four farms the disease was associated with cryptosporidiosis and on a fifth no enteropathogens were detected; these five farms (461 cows) were also excluded from further analysis. Of the remaining four herds, two beef suckler herds (105 cows) had concurrent rotavirus and cryptosporidial infections, and vaccination was associated with a decreased excretion of rotavirus but not with a decreased incidence of diarrhoea. In the other two dairy herds (68 cows) with prevaccination rotavirus problems, there was a significantly decreased incidence of diarrhoea in calves born to vaccinated cows. No natural field challenge of enterotoxigenic E coli was encountered on any of the trial farms.     

Enteric infections in veal calves: a longitudinal study on four veal calf units

Forty-five calves on four veal calf units were monitored during the first four weeks after their arrival. Faecal samples were collected on alternate days and screened for the presence of rotaviruses, bovine coronavirus, Cryptosporidium oocysts, K99 positive strains of E. coli and Salmonella spp. Rotaviruses and Cryptosporidium were the most commonly detected agents (78% and 60% respectively of the calves). Bovine coronavirus was detected in the faeces of 18% of the calves, whilst K99 positive E. coli was only found in 2 samples from one calf. Salmonella spp. were not isolated from any of the 646 faecal samples examined. Shedding of rotaviruses occurred in a bimodal pattern beginning in the first week of the survey. Cryptosporidium oocysts were detected most frequently in the interval between the two peaks of rotavirus shedding. The presence of rotaviruses or Cryptosporidium oocysts in faeces was not strongly associated with scour, nor were combined infections with these agents or the cases of bovine coronavirus infection. The condition of the calves throughout the survey was generally satisfactory.     

Rotavirus infection in newborn calves. I. Evaluation trials of certain enzymes in faeces

The purpose of this work was to investigate the relationship between rotavirus infection, diagnosed in faeces using the serological test, in calves with diarrhoea and the activity of lysozyme, alpha-amylase, lipase and trypsin in the same material. The faeces were taken from 28 sick and 1 healthy animals, aged from 1 to 5 days, for microbiological and enzymatic examination. No correlation was found between lysozyme activity in faeces and rotavirus infection. Similar results were obtained with the other enzymes.     

Rotavirus infections in calves in dairy herds

Neonatal calf diarrhoea was studied in 115 calves of one dairy herd from January 1976 to June 1977. Two syndromes could be distinguished: a mild and short lasting 'early diarrhoea' within the first three days of life and a usually more severe 'late diarrhoea' from the fourth to the 14th day of life. The latter type of diarrhoea occurred almost exclusively during the first half of the year. Thirty-four out of 45 calves with late diarrhoea excreted rotaviruses, whereas only one of 34 calves with early diarrhoea excreted virus. In addition, rotavirus excretion was found in 11 calves that either remained healthy or had recovered from diarrhoea before virus was first detected. Similar findings were obtained in the spring of 1978 but in addition bovine coronavirus was recovered from four calves with late diarrhoea and from three healthy calves. One faecal sample obtained from a calf with diarrhoea on day 2 yielded K99+ Escherichia coli. Nearly all cows excreted rotavirus-specific antibodies in their colostrum but no relationship was found between the initial colostral antibody titre against rotavirus and the development of rotavirus-associated diarrhoea in the calf.     

Group A rotavirus excretion patterns in naturally infected pigs

Cross-sectional and cohort studies were conducted in a piggery to investigate the excretion pattern of group A rotavirus in pigs. The cross-sectional survey revealed that 47 (9 per cent) of 521 pigs sampled were excreting rotavirus in the faeces. No rotavirus antigen was detected in the faeces of pigs either less than one week or over two months old. The prevalence of infection increased with age over the sucking period, and was greatest at five weeks old. Diarrhoea was observed in only eight (17 per cent) of the pigs excreting rotavirus. Sixteen piglets from four litters were selected and faecal samples collected daily from each animal from birth to two months old. All the piglets excreted group A rotavirus and the range of ages at which they first became infected was between 13 and 39 days. The average duration of excretion in individual piglets was 7.4 days. Ten of 13 sucking piglets which excreted rotavirus developed diarrhoea soon after it was first detected.     

Association of diarrhea in cattle with torovirus infections on farms.

An epidemiologic survey was performed to determine the incidence of torovirus infections in 2 disease entities of cattle: diarrhea of replacement calves up to 2 months old, and winter dysentery of adult cattle. Samples were obtained from 187 diarrheal and 115 healthy calves from 15 farms, as well as 149 diarrheal and 67 healthy cows from 27 farms with or without winter dysentery. Enzyme-linked immunosorbent assays for detection of torovirus, rotavirus, and coronavirus antigen in feces, and of torovirus and coronavirus antibodies in serum were used to monitor infections in these groups. Torovirus was detected in 9 of the 15 farms in the study, and in 6% of calves with diarrhea, which was significantly higher than in healthy calves (2%). Seroconversion to torovirus was found significantly more often after winter dysentery episodes than on farms without a disease history; coronavirus seroconversion was less common.     

Pathology of natural rotavirus infection in clinically normal calves

During a longitudinal study of the epidemiology of rotavirus infection in a calf rearing unit, excretion of virus in faeces was detected by enzyme-linked immunosorbent assay in 40 of 48 (83 per cent) unweaned calves aged between three days and five weeks. Fifty per cent of the infected calves had no clinical signs of disease. Enterocytes containing rotavirus antigen and intestinal lesions were found in all of 12 clinically normal calves selected for necropsy between days 1 and 4 of virus excretion. Stunting and fusion of villi, exfoliation, disarrangement and vacuolation of enterocytes and the presence of cuboidal enterocytes were observed in infected calves but not in rotavirus-free control calves. Lesions predominated in the upper small intestine, where rotavirus was most abundant, especially on the first two days of virus excretion. The numbers of enterocytes infected with rotavirus diminished before the lesions resolved.     

Studies on rotavirus infection and diarrhoea in young calves.

The occurrence of diarrhoea in calves was monitored during the first three weeks of life. Calves fed amounts of colostrum sufficient to produce serum Ig levels in excess of 30 mg per ml did not develop diarrhoea, whereas calves fed less colostrum did. Rotaviruses and mycoplasma-like particles were observed in the faeces of calves with and without diarrhoea. The epidemiology of rotavirus infection in calves is discussed.     

Diagnosis of bovine coronaviruses using mouse erythrocyte suspensions stabilized with formalin

Bovine coronavirus isolated from calf faeces diseased with gastroenteritis and passaged to colostrum-free calves agglutinated mouse and rat erythrocytes. The agglutination reaction depended on temperature and took place only at a temperature of 4 degrees C. At a temperature of 37 degrees C the agglutinate broke down within 15 minutes. The coronavirus could be detected by the haemagglutination test in the contents of the small and large intestines and in the faeces of experimentally and naturally infected calves. The agglutination capacity of mouse erythrocytes was not affected by careful fixation of these erythrocytes with formalin and subsequent lyophilization and remained unchanged for as long as 52 weeks of storage at a temperature of 4 degrees C. It was demonstrated by a comparative examination of 182 samples of the faeces of calves suffering from diarrhoea that haemagglutination test was as sensitive as electron microscopy.     

Epidemiological studies of parvovirus infections in calves on endemically infected properties

Bovine parvovirus serology and virus excretion were monitored in calves located on three endemically infected North Queensland properties. Maternally derived serum antibody to bovine parvovirus was found to have a half-life of 19 days. On all three properties, calves developed intestinal bovine parvovirus infection with seroconversion soon after weaning. This occurred more promptly where the environment was subject to heavier bovine parvovirus contamination due to management practices. The concurrent presence of moderate levels of residual serum antibody had only minor influence on the onset of the infection. On one beef cattle property, onset of intestinal bovine parvovirus infection was associated with an outbreak of post-weaning diarrhoea. Anthelmintic treatment trials indicated that this syndrome was unrelated to helminth burdens, though coccidiosis appeared responsible for occasional subsequent cases of dysentery. It was considered that bovine parvovirus may have significantly contributed to the development of the diarrhoea syndrome, in conjunction with substantial weaning stresses.