Determination of bovine rotavirus G serotype by polymerase chain reaction

A total of 113 diarrheic samples comprising of 68 buffalo calves and 45 cow calves were screened by RNA-PAGE for the detection of presence of rotavirus. RNA-PAGE analysis of these samples revealed 11 (9.73%) was found positive for rotavirus. Out of 68 faecal samples of buffalo calves tested for viral gastroenteritis, 8 (11.76%) were found positive for rotavirus. Similarly, out of 45 faecal samples of cattle calves tested for viral gastroenteritis, 3 (6.66%) was found positive for rotavirus. Rotavirus-positive samples represented long electropherotype. All RNA-PAGE-positive faecal samples for rotavirus subjected to RT-PCR for VP7 gene, ten samples yielded a specific product of 1,013 bp of VP7 gene. All the PCR-positive samples of the present study were subjected to genotyping with primers for G6, G8 and G10 genotype. All positive samples showed G10 genotype. This indicates that G10 may be predominant genotype among bovine calves in Mumbai region in India.     

Molecular characterization of bovine rotavirus circulating in beef and dairy herds in Argentina during a 10-year period (1994-2003)

Group A bovine rotavirus (BRV) is one of the main causes of neonatal calf diarrhea. The present study reports the incidence of rotavirus diarrhea and the genotypes of BRV strains circulating in beef and dairy herds from Argentina, during a 10-year period (1994-2003). Group A BRV was detected in 62.5% (250/400) of the total studied cases of diarrhea. Positive cases were analyzed by heminested multiplex RT-PCR for P and G genotypes identification. Sixty percent of them were typed as P[5]G6, 4.4% P[11]G10, 4.4% P[11]G6 and 2.4% P[5]G10. Additionally, 9.2% of the cases were initially typed as G8 combined with P[5] or P[11], but sequence analysis revealed they belonged to genotype G6, lineage Hun4-like. Partial typing was assessed in 12.0% of the cases. One of the partially typed samples was closely related to genotype G15. BRV was detected in 71% and 58% of the outbreaks registered in beef and dairy farms, respectively. A clear differential distribution of G/P types was found according to the herd type. P[5]G6 was the prevalent strain in beef herds, while P[11] was the prevalent P-type in dairy herds (71%), associated in similar proportions with G6 and G10, These findings indicate that BRV genotypes included in the current commercially available rotavirus vaccines (G6, G10, P[5] and P[11]) should protect calves from most Argentinean field strains. Nevertheless, continuous surveillance is necessary to detect the emergence of new variants.     

Genetic diversity and novel combinations of G4P[19] and G9P[19] porcine rotavirus strains in Thailand

Several epidemiological studies reported the detection of rotavirus strains bearing unusual combinations of genetic background of human and porcine rotaviruses. This observation supports the hypothesis of interspecies transmission of rotaviruses in humans and pigs. The aims of this study were to investigate the genotypes and molecular characteristics of rotaviruses in piglets with diarrhea in several farms from two provinces in Thailand. A total of 207 fecal specimens collected from diarrheic piglets were screened for the presence of groups A, B, and C rotaviruses. Group A rotaviruses were detected in 41 out of 207 (19.8%) fecal specimens tested. A wide variety of G-P combination rotavirus strains were detected in this study. The G4P[6] was identified as the most prevalent genotype (39.0%), followed by G4P[23] (12.2%), G3P[23] (7.3%), G4P[19] (7.3%), G3P[6] (4.9%), G3P[13] (4.9%), G3P[19] (4.9%), G9P[13] (4.9%), G9P[19] (4.9%), G5P[6], and G5P[13] each of 2.4%. Furthermore, G5 and G9 in combinations with P-nontypeable strains were also found at each consisting of 2.4% (n = 1) of the collection. It was interesting to note that among diversified porcine rotavirus strains, novel combinations of G4P[19] and G9P[19] strains were detected for the first time in this study. Nucleotide sequences of VP4 and VP7 of these strains were closely related to human rotaviruses reported previously. The data implies that these porcine rotaviruses were probably generated in nature from the reassortment between the viruses of human and porcine origin. This study provides valuable epidemiological information and molecular characteristics of porcine rotaviruses circulating in piglets with diarrhea in northern Thailand.     

Detection and genetic characterization of porcine group A rotaviruses in asymptomatic pigs in smallholder farms in East Africa: Predominance of P[8] genotype resembling human strains

Viral enteritis is a serious problem accounting for deaths in neonatal animals and humans worldwide. The absence of surveillance programs and diagnostic laboratory facilities have resulted in a lack of data on rotavirus associated diarrheas in pigs in East Africa. Here we describe the incidence of group A rotavirus (RVA) infections in asymptomatic young pigs in East Africa. Of the 446 samples examined, 26.2% (117/446) were positive for RVA. More nursing piglets (78.7%) shed RVA than weaned (32.9%) and grower (5.8%) pigs. RVA incidence was higher in pigs that were either housed_free-range (77.8%) or tethered_free-range (29.0%) than those that were free-range or housed or housed-tethered pigs. The farms with larger herd size (>10 pigs) had higher RVA prevalence (56.5%) than farms with smaller herd size (24.1–29.7%). This study revealed that age, management system and pig density significantly (p < 0.01) influenced the incidence of RVA infections, with housed_free-range management system and larger herd size showing higher risks for RVA infection. Partial (811–1604 nt region) sequence of the VP4 gene of selected positive samples revealed that different genotypes (P[6], P[8] and P[13]) are circulating in the study area with P[8] being predominant. The P[6] strain shared nucleotide (nt) and amino acid (aa) sequence identity of 84.4–91.3% and 95.1–96.9%, respectively, with known porcine and human P[6] strains. The P[8] strains shared high nt and aa sequence identity with known human P[8] strains ranging from 95.6–100% to 92–100%, respectively. The P[13] strains shared nt and aa sequence identity of 83.6–91.7% and 89.3–96.4%, respectively, only with known porcine P[13] strains. No P[8] strains yielded RNA of sufficient quality/quantity for full genome sequencing. However analysis of the full genome constellation of the P[6], two P[13] and one untypeable strains revealed that the P[6] strain (Ke-003-5) genome constellation was G26-P[6]-I5-R1-C1-M1-A8-N1-T1-E1-H1, P[13] strains (Ug-049 and Ug-453) had G5-P[13]-I5-R1-C1-M1-A8-N1-T7-E1-H1 while the untypeable strain (Ug-218) had G5-P[?]-I5-R1-C1-M1-A8-N1-T1-E1-H? In conclusion, P[6] and P[8] genotypes detected were genetically closely related to human strains suggesting the possibility of interspecies transmission. Further studies are required to determine the role of RVA in swine enteric disease burden and to determine the genetic/antigenic heterogeneity of the circulating strains for development of accurate diagnostic tools and to implement appropriate prophylaxis programs.     

Longitudinal study of bovine rotavirus group A in newborn calves from vaccinated and unvaccinated dairy herds

Reports of rotavirus excretion in calves usually result from cross-sectional studies, and in face of the conflicting results regarding protection of calves born to vaccinated dams against diarrhea, the aim of the present study was to evaluate rotavirus excretion in dairy calves born to vaccinated or unvaccinated dams, to identify the genotypes of bovine rotavirus group A (RVA) strains isolated from these animals as well as to investigate characteristics of the disease in naturally occurring circumstances throughout the first month of life. Five hundred fifty-two fecal samples were taken from 56 calves, 28 from each farm and, in the vaccinated herd, 11/281 samples (3.91%) taken from six different calves tested positive for RVA while in the unvaccinated herd, 3/271 samples (1.11%) taken from 3 different calves tested positive. The genotyping of the VP7 genes showed 91.2% nucleotide sequence identity to G6 genotype (NCDV strain), and for the VP4 gene, strains from the vaccinated herd were 96.6% related to B223 strain, while strains from the unvaccinated herd were 88% related to P[5] genotype (UK strain). Genotypes found in this study were G6P[11] in the vaccinated herd and G6P[5] in the unvaccinated herd. All calves infected with rotavirus presented an episode of diarrhea in the first month of life, and the discrepancy between the genotypes found in the commercial vaccine (G6P[1] and G10P[11]) and the rotavirus strains circulating in both vaccinated and unvaccinated herds show the importance of keeping constant surveillance in order to avoid potential causes of vaccination failure.     

Prevalence of serotypes G6 and G10 group A rotaviruses in dairy calves in Quebec.

Fecal samples from diarrheic and nondiarrheic dairy calves (1 to 3 weeks old) from 12 regions of Quebec, collected between 1992 and 1994, were screened for group A bovine rotavirus (BRV) using a combination of 2 VP6-specific monoclonal antibodies (MAbs) in an enzyme-linked immunosorbent assay (ELISA). The overall prevalence of BRV infection was 26.4% (107/405). In diarrheic calves, BRV infection reached 74.3% (55/74), but only 15.7% (52/331) in nondiarrheic calves. BRV-positive samples were serotyped by enzyme-linked immunosorbent assay (ELISA) using G6 and G10 specific MAbs. The analysis of 107 field samples revealed that, in diarrheic calves, 34.5% (19/55) were G6, 27.2% (15/55) were G10, 9% (5/55) were G6 and G10 positive, and 29.9% (16/55) were G6 and G10 negative. In nondiarrheic calves, 19.2% (10/52) were G6, 19.2% (10/52) were G10, 7.6% (4/52) were G6 and G10 positive, and 53.6% (28/52) were G6 and G10 negative. Rotavirus dsRNA was extracted from BRV-positive samples and examined by polyacrilamide gel electrophoresis (PAGE). Of 107 samples tested, 74 (69.1%) were positive, and all the samples demonstrated a typical group A rotavirus migration pattern.     

G and P genotype profiles of rotavirus A field strains circulating in beef and dairy cattle herds in Brazil, 2006–2015

The aim of this retrospective study was to use RT-PCR and nucleotide sequencing analysis to determine the G (VP7 gene) and P (VP4 gene) genotypes of 155 Brazilian bovine rotavirus A (RVA) wild-type strains detected in diarrheic calves from all Brazilian geographical regions from 2006 to 2015. The RVA strains evaluated belonged to the G6, G10, P[5], and P[11] genotypes. The G6P[5] genotype was prevalent (65.5%; P < 0.05) in beef, and the G10P[11] (38.4%) and G6P[11] (30.8%) genotypes were more prevalent in dairy cattle herds. The Midwest was the region with the highest number of genotyped RVA strains, where the genotypes G6, P[5], and P[11] were identified. Genotype combination G6-IV/P[5]-IX, prevalent in beef herds, and G6-III/P[11]-III or G10-IV/P[11]-III, prevalent in dairy herds, were detected. In addition, for the first time in Brazil, we detected the P[5] and P[11] genotype RVA strains that belong to lineage II and VII, respectively.     

Isolation and molecular characterisation of equine rotaviruses from Germany

A total of 26 rotavirus positive faecal samples of diarrhoeal foals, and 8 equine rotavirus isolates were examined. Viral RNA patterns were generated, G typing was performed by PCR, and a P[12]-specific DNA probe was developed for P typing. Furthermore, five equine rotavirus isolates were sequenced in the genomic regions coding for VP7 and part of VP4. Rotaviruses of genotype G3 P[12] were found in 22 faecal samples and G14 P[12] type could be found in 4 faecal samples. These findings confirm that in Germany G3 P[12] is the predominating type of equine rotaviruses.     

Giardia and Cryptosporidium infections in neonatal reindeer calves: Relation to the acute phase response

This longitudinal observational study was conducted to investigate the spontaneous effect of Giardia and Cryptosporidium infections on acute phase response (APR) in reindeer calves (Rangifer tarandus tarandus) in Finnish Lapland.Serum (n = 609) and faecal samples (n = 366) were collected from 54 reindeer calves aged zero to 33 days. The samples were analysed for Giardia, Cryptosporidium, acute phase proteins (APP) and γ-globulins.Linear regression models were used to investigate associations of early Giardia infection (before 12 days of life) with the response of APPs and acquiring of passive immunity.Giardia was detected in 100% and Cryptosporidium in 23% of calves. There was a negative association between early Giardia infection and γ-globulin concentrations (p = 0.032) and a positive association with serum amyloid A (SAA) concentrations (p = 0.042). The results suggest a protective effect of colostrum against Giardia infection and that early infection may induce activation of APR.     

Detection and characterisation of bovine rotavirus in Ireland from 2006–2008

Background

Worldwide, Group A bovine rotavirus (RVA boRV) is one of the main causes of neonatal calf diarrhoea. Currently, limited epidemiological and sequence data exists on the RVA disease in bovines in Southern Ireland only. The aim of the study was to generate epidemiological and sequence data of RVA boRV distributed over a wide geographical area in Ireland.

Findings

272 stool samples were obtained from symptomatic calves and analysed to identify the prevalent G and P genotypes. Viral type combinations including G6P[5], G6P[11] and G10P[11] genotype were the most frequently identified. The G6P[5] combination was predominant throughtout the study, accounting for 70% (n = 191). Sequence analysis of the VP7 gene revealed that Irish G6 strains fell within Lineage IV, similiar to previous reports in Ireland.

Conclusion

The detection of unusual G and P combinations may have an impact on rotavirus control programmes and current vaccines may need to incorporate new strains, as the current vaccine available may not offer protection against all of these circulating types.     

Effects of Pre-Weaning Vitamin E,Selenium, and Copper Supplementation on the Performance,Acute Phase Protein Concentration,and Immune Function of Stressed Beef Calves

Two experiments were conducted to determine the effects of pre-weaning vitamin E, Se, and Cu supplementation on performance and immune response in stressed calves. In Exp. 1, 71 Hereford x Angus calves were individually creep fed: 1) control supplement (CON), 2) control plus 500 IU vitamin E + 0.3 mg Se/kg DM (E), 3) control plus 10 mg Cu/kg DM (CU), or 4) a combination of E and CU treatments (ECU). In Exp. 2, 80 Hereford (Angus calves were individually creep fed: 1) control supplement (CON), 2) control plus 0.3 mg Se/kg DM (SE), 3) control plus 500 IU vitamin E + 0.3 mg Se/kg DM (LOWE), 4) control plus 1000 IU vitamin E + 0.3 mg Se/kg DM (MEDE), or 5) control plus 1500 IU vitamin E + 0.3 mg Se/kg DM (HIE). Treatments continued for 49 (Exp. 1) or 53 d (Exp. 2) prior to weaning. At weaning all calves were transported to feedlot facilities. In Exp. 1, vitamin E tended (P<0.09) to improve post-weaning ADG and reduce (P<0.06) plasma haptoglobin (Hp), but had no effect on plasma α-tocopherol. Dietary Cu tended to increase (P<0.01) liver Cu stores, and antibody titers to bovine viral diarrhea (BVD) were greater (P<0.04) at weaning in CU and E calves. In Exp. 2, vitamin E tended to increase serum α-tocopherol (P<0.06) and cortisol (P<0.08). Vitamin E and Se supplementation may improve post-weaning performance and decrease plasma Hp concentrations in stressed calves.     

Diversity and zoonotic potential of rotaviruses in swine and cattle across Europe

Group A rotaviruses can infect both humans and animals. Individual rotavirus strains can occasionally cross species barriers and might hereby contribute to the emergence of new genotypes in heterologous hosts. The incidence and impact of zoonotic rotavirus are not well defined, and one reason for this is a lack of data about strains circulating in suspected reservoir animal hosts. In this study we report the incidence, genetic diversity, and molecular epidemiology of rotaviruses detected in domestic cattle and swine in 6 European countries. From 2003 to 2007, 1101 and more than 2000 faecal specimens were collected from swine and cattle, both healthy and diarrhoeic, and tested for rotaviruses. Viruses from positive stools were genotyped and a subset of strains was characterized by nucleotide sequencing and phylogenetic analysis of the VP7 (G) and VP4 (P) genes. Rotaviruses were detected in 43% of bovine samples and in 14% of porcine samples. In cattle, 10 different combinations of G and P types were identified and the most common strains were G6P[11] and G6P[5]. In swine, the number of identified G-P combinations was higher (n=21), however, no single combination was predominant across Europe. Newly described genotype specificities, P[27] and P[32], were identified in swine. When compared at the nucleotide sequence level, the identified porcine rotavirus strains and contemporary human strains grouped together phylogenetically, whereas bovine rotavirus strains formed separate clades. These data demonstrate large genetic diversity of porcine and bovine rotavirus strains across Europe, and suggest that livestock herds may serve as potential reservoirs for human infections.     

Detection of group a rotavirus strains circulating in calves in Tunisia

Faecal samples were collected from 89 dairy calves to determine the prevalence of rotavirus infection in Tunisia and the genomic diversity of bovine rotavirus strains. After screening of all faecal samples by enzyme-linked immunosorbent assay, rotavirus strains were analysed by RNA polyacrylamide gel electrophoresis and characterized antigenically by monoclonal antibodies to the VP6 subgroup. The VP7 genotype was determined by nested RT-PCR. Of the 89 calves tested, 27 (30%) were positive for rotavirus antigen. Four different long electrophoretypes were identified. All VP6 typeable strains carried the subgroup I specificity. G8 genotype was the most prevalent, but G6 and mixed strains G(6 + 8) were also detected.     

Lack of evidence for fecal shedding of Mycobacterium avium subsp. paratuberculosis in calves born to fecal culture positive dams

The objective was to detect presence of calves excreting Mycobacterium avium subsp. paratuberculosis (MAP) in their feces as a consequence of being born to MAP fecal culture positive (vs. negative) dams. For each cow that was about to calf, approximately 10 g of feces was collected manually by the herdsmen from the rectum using a disposable plastic examination sleeve within 48–72 h prior to actual calving. Between 1 and 3 d of birth, herd personnel collected approximately 10 g of fecal samples followed by monthly visits to the farm at which time 10 g of fecal samples were again collected by study investigators from each calf at approximately 30, 60 and 90 d of age. Mycobacterium avium subsp. paratuberculosis was recovered from 8% (5/60) of the cows that gave birth to calves. However, MAP was not recovered from any of the fecal samples (0/240) collected from study calves. Findings of the present study suggest lack of evidence for fecal excretion of MAP in calves born to fecal culture positive (vs. negative) dams in a heavily infected herd.     

Molecular characterization of bovine rotavirus strains circulating in northern Italy, 2003-2005

A total of 232 stools collected from calves with rotavirus infection in herds located in northern Italy from 2003 to 2005 was investigated. Determination of the rotavirus G and P types was carried out using nested RT-PCR. G6 was the most prevalent genotype, accounting for 78.5% of samples, G10 accounted for 9.9% of samples and viruses of G8 type were found in 4.7% of samples. In 3% of samples, viruses were not classified due to concomitant infection with more G type strains, whereas viruses in 3.9% of samples could not be characterized with any of the G-specific primers used in this study. Most common P types were P[11] and P[5], accounting for 65.1% and 25%, respectively. In 2.6% of cases, samples reacted with multiple P-specific primers; no P[1] serotype was identified. The G6P[11] combination was predominant throughout the study period, i.e. 52.5% in 2003, 50% in 2004 and 40% in 2005. The incidence of G6P[5] increased from 13.1% in 2003 to 27% in 2004 and 25.5% in 2005. The G10P[11] combination decreased markedly from 18% in 2003 to 2.6% in 2004, rising again to 7.3% in 2005. G8P[11] viruses were similarly present in 2003 (5%) and 2004 (4.3%), declining slightly in 2005 (1.8%).