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To determine the excretion dynamics and genotypic characteristics of rotavirus A (RVA), a longitudinal observational study was performed in 10 pigs from 3 litters at a farrow-to-finish farm. A total of 400 fecal samples were directly collected from the rectums of individual pigs (aged 7 to 217 days) at 3- to 14-day intervals. Seventy-one samples (17.5%) were positive for RVA by reverse transcription-PCR designed to detect the VP7 and VP4 genes. At least 13 combinations of 5 G (G2, G4, G5, G9, and G11) and 6 P (P[6], P[7], P[13], P[23], P[27], and P[34]) genotypes were identified by direct sequencing of the PCR products. We were able to detect RVA VP7 sequences from each pig 4 to 6 times with intervals of 7 to 52 days (from 7 to 119 days of age). Each pig harbored RVAs with at least 3 to 6 different combinations of G and P genotypes, while repeated excretions of RVAs carrying the same combinations of G and P genotypes were also observed. Virus shedding and changes in G and P genotypes appeared to be associated with movement of the pigs into weaning, growing, and finishing barns. These results indicated that, over their lifetimes, pigs raised for meat frequently and intermittently excrete genetically diverse RVAs.

Rotavirus (RV) is an important cause of gastrointestinal disease in animals and humans. In this study, we developed an RT-PCR to detect RV group B (RVB) and characterized the VP7 (G) gene segment detected in porcine samples. One hundred seventy three samples were tested for RV group A (RVA), RVB, and C (RVC) by RT-PCR and examined for RV-like lesion using histopathology. A majority (86.4%) of the samples had mixed RV infections and co-infections of RVA/RVB/RVC were detected at a higher rate (24.3%) than previously reported. RVB was identified in 46.8% of the 173 samples. An adapted VP7 classification was developed using previously published (n=57) and newly sequenced (n=68) RVB strains, resulting in 20 G genotypes based on an 80% nucleotide identity cutoff value. Our results revealed a broad genetic diversity of porcine RVB strains, suggesting RVB has been the cause of common/pre-existing, yet undiagnosed, disease in pigs., Highlights ► Porcine RBV strains found in 46.8% of samples tested in the United States in 2009. ► Majority of RVB-positive samples (86.4%) contained mixed RV infections. ► Unexpected higher prevalence of RVC (50.0%) than RVA (27.1%) in

Using a total of 2018 fecal samples collected between 2003 and 2008 from foals with diarrhea, the molecular epidemiology of group A equine rotaviruses circulating in Japan was investigated by the reverse transcription-polymerase chain reaction (RT-PCR) typing and sequence analysis of the VP4 (P type) and VP7 (G type) genes. A total of 1149 samples showed positive reactions with RT-PCR, of which 462 samples (40.2%) were positive for G3 type, 502 samples (43.7%) were positive for G14 type, and 185 samples (16.1%) were positive for both G3 and G14 types. To examine P types, 59 G3 and 56 G14 positive samples were used. The majority of the samples (96.5%) were characterized as P[12] type. In a phylogenetic analysis, the VP4 gene of the P[12] type in Japan was found to be conserved for a long time. The VP7 sequences of the G3 type were found to be clustered in the same group as the HO-5 strain, which is a G3 strain that was isolated in 1982 in Japan. In contrast, the VP7 sequences of the G14 type, which were in circulation between 2003 and 2008, were clustered differently from those of the G14 type strains isolated in Japan in the late 1990 s. These results suggest that the VP7 gene of the G3 type has been conserved over 25 years, while the VP7 gene of the G14 type circulating between 2003 and 2008 appears to have re-emerged in or invaded Japan around 2000.

Enzyme-linked immunosorbent assays (ELISAs) were developed to detect group (gp) C rotavirus antigens and antibodies. Both assays were confirmed to be specific for gp C rotavirus by using serogroup A, B, and C rotaviruses; hyperimmune antisera to these serogroups of rotaviruses; and paired serum specimens from animals infected with gp C rotaviruses. The ELISA for antigen detection reacted not only with porcine gp C rotaviruses but also with human and bovine gp C rotaviruses. Following experimental challenge of gnotobiotic pigs with porcine gp C rotavirus, the virus was found by ELISA in all diarrheic feces. A high prevalence of antibodies to gp C rotaviruses was detected in sera from adult pigs (93 to 97%) and cattle (47 to 56%) in the United States and Japan. However, no antibody to gp C rotavirus was detected in the sera (n = 20) of adult horses in the United States. In human sera from Hokkaido, Japan, 3% of children and 13% of adults possessed antibody to gp C rotaviruses. These results suggest that the ELISA that we developed may be useful for surveying gp C rotavirus infections in animals and humans. On the basis of serology, gp C rotavirus infections are common in pigs and cattle in the United States and Japan, but they occur at lower levels in humans from the Hokkaido area of Japan.

A biotin-streptavidin-enhanced enzyme-linked immunosorbent assay (ELISA) which uses monoclonal antibodies (MAbs) for the detection of group C rotaviruses was developed. An assay in which plates were coated with three pooled MAbs and biotinylated polyclonal immunoglobulin G (IgG) (polyclonal antibody [PAb]) was used as the detector (MAb capture-PAb detector) was found to be the most sensitive and specific of the assays when it was compared with assays in which plates were coated with polyclonal antiserum and detection was done with either biotinylated polyclonal antiserum (PAb capture-PAb detector) or biotinylated pooled MAbs (PAb capture-MAb detector). The MAb capture-PAb detector ELISA detected 83% of samples confirmed to be positive for group C rotaviruses, whereas the PAb capture-PAb detector assay detected 63% of positive samples and the PAb capture-MAb detector assay detected 65% of positive samples. All three procedures detected both of the bovine and the two human group C rotaviruses, but none of the three procedures detected fecal samples containing group A and B rotaviruses or fecal samples negative for group C rotaviruses used in this study. The sensitivity of the MAb capture-PAb detector ELISA was determined by serially diluting fecal group C rotaviruses; antigens were detected in maximal positive dilution ranges of 1:1,000 to 1:3,000 for the samples tested. On the basis of the cell culture immunofluorescence assay infectivity titer of semipurified cell culture-passaged Cowden group C rotavirus, the sensitivity of the MAb capture-PAb detection ELISA for detection of homologous group C rotavirus was 53 fluorescent focus units per ml. Epitope mapping by use of the biotinylated MAbs in competition assay suggested that our MAbs may bind to three different but overlapping epitopes. These results suggest that the MAb capture-PAb detector ELISA can be used to study the epidemiology of group C rotaviruses in humans and animals.

The correlation between maternal serum antibodies in beef calves at 2 days old and protection against diarrhea induced by natural bovine rotavirus (BRV) infection was examined. Virus neutralizing (VN) antibody titers against BRV in sera from calves that developed diarrhea by BRV infection within 14 days of age (BRV-diarrheal calves) were significantly lower than those from calves that had no diarrhea. In the BRV-diarrheal calves, a positive correlation was found between the VN antibody titers and age of the onset of diarrhea. There were negative correlations between the VN antibody titers and duration of the diarrhea, VN antibody titers and cumulative diarrhea scores, and the VN antibody titers and duration of virus shedding. These results suggest that the VN antibody titers against BRV in newborn calf serum could be an indicator of protection against BRV-induced diarrhea.

To address the regulation and evolution of precursor protein cleavability in caliciviruses, we examined constraints on diversity of upstream regions of calicivirus precursor cleavage sites. We performed alanine scanning and supplementary mutagenesis of amino acids at P1, P2, P3, and P4 sites using four viruses representing the four major genera of the family Caliciviridae. This study complements previous mutagenesis studies and shows strong restrictions in mutations at the P1 and P4 sites for effective cleavage reactions. By contrast, such restrictions were less frequently observed at the P2 and P3 sites. Shannon entropy analysis of the reported sequences showed that the P2, P3, and P4 sites allow variations in amino acid size within a calicivirus genus whereas the P1 sites do not. Notably, the human sapovirus precursor protein exceptionally retains a basic rather than aromatic amino acid at the P4 site of the NS4/NS5 cleavage site in reported strains, and a substitution from basic to aromatic amino acid significantly enhanced cleavability at this site. Taken together, these data suggest the existence of (i) structural constraints on the P1 site that restrict size changes within each calicivirus genus, (ii) plastic substrate surfaces that accommodate size variation at the P2, P3, and P4 sites and modulate their own cleavabilities, and (iii) biological constraints on the P4 site that maintain the lower cleavability of the NS4/NS5 site in sapovirus.

A virus (designated the Shintoku strain) which was morphologically indistinguishable from group A rotaviruses was detected in the feces of adult cows with diarrhea in Japan. The virus contained 11 segments of double-stranded RNA and had an electrophoretic migration pattern in polyacrylamide gels similar to that of other group C rotaviruses (4-3-2-2). Feces containing the bovine virus reacted with antiserum to porcine group C rotavirus (Cowden strain) but not group A or B rotaviruses in immunoelectron microscopy. The virus was adapted to serial propagation in roller tube cultures of a rhesus monkey kidney cell line (MA104) by using high concentrations of trypsin. Evidence for viral replication in MA104 cell cultures was demonstrated by immunoelectron microscopy and indirect immunofluorescence by using antiserum to porcine group C rotavirus and by electrophoretic analysis of extracted viral double-stranded RNA. A significant antibody response against the isolate was detected in convalescent-phase sera of cows which excreted the virus: no increased antibody response to bovine group A rotavirus was observed. To our knowledge, this is the first isolation of a group C rotavirus from cattle.

The genetic diversity of gene 8 (encoding the outer capsid glycoprotein VP7) among group (Gp) C rotaviruses was examined by Northern and dot blot hybridization. A cDNA clone of the porcine Gp C Cowden strain gene 8 was labeled with 32P by nick translation and used as a probe. The gene 8 probe hybridized with the corresponding gene of one human (88-196) and four porcine (Cowden, NB, WH, and Wi) strains of Gp C rotaviruses under both moderate (50% formamide, 5X SSC, and 42 degrees) and high (50% formamide, 5X SSC, and 52 degrees) stringency conditions. However, under high stringency conditions little or no hybridization was detected with the corresponding gene of one bovine (Shintoku) and three other porcine (Ah, HF, and KH) strains of Gp C rotaviruses. In control experiments, the Cowden gene 8 probe did not hybridize with Gp A (Gottfried strain) or Gp B (Ohio strain) rotaviruses. These data demonstrate that the Cowden gene 8 probe is Gp C rotavirus-specific and that genetic diversity exists among Gp C rotaviruses in the gene encoding the outer capsid glycoprotein VP7. Our gene 8 probe may be useful in hybridization assays for serotyping Gp C rotaviruses, analogous to the use of VP7 probes for serotyping Gp A rotaviruses. However, final confirmation of our genetic approach to serotype Gp C rotaviruses awaits the serologic analysis of these viruses.

Fecal and nasal samples were collected from 180 calves with diarrhea and 36 clinically normal co-habitants, and tested for virus using HRT-18 cell cultures derived from human rectal adenocarcinoma. A cytopathic virus was isolated from 5 fecal and 56 nasal samples obtained from diarrheic calves. All calves in which the virus was isolated from diarrheic feces were positive for virus isolation from nasal swabs. The virus was also isolated from the nasal swabs of 10 clinically normal calves that were co-habitants with diarrheic calves. Because they were morphologically similar to coronavirus, agglutinated mouse erythrocytes and serologically identical with the Nebraska calf diarrhea coronavirus, new isolates were identified as bovine coronavirus. The demonstration of viral antigens in nasal epithelial cells by a direct immunofluorescence was in close agreement with the virus isolation in HRT-18 cell cultures. This is the first report on the isolation of bovine coronavirus from newborn calves with diarrhea in Japan. The evidence that the virus was frequently isolated from nasal swabs is of great interest for understanding the pathogenesis of bovine coronavirus infection.

A cytopathogenic virus (designated the Aichi/2004 strain) was isolated in a human rectal adenocarcinoma cell line (HRT-18) from the ileum contents of a calf with diarrhea. Oval and elongated particles, approximately 100 to 170 nm in diameter, with club-shaped projections were seen in the infected culture supernatant, and torovirus-like (tubular and torus nucleocapsid) structures were seen in the infected cells by electron microscopy. An antiserum against bovine torovirus (BToV) reacted with the infected cells by immunofluorescence and neutralized the isolate. However, antisera against bovine coronavirus (BCV) failed to react with the infected cells by immunofluorescence or did not neutralize the isolate. Further, the isolate was positive for BToV by reverse transcription-PCR (RT-PCR) targeting fragments of the nucleocapsid (N), membrane (M), and spike (S) genes. Comparison of the nucleotide sequences of the PCR products with those of the published N, M, and S genes (476 to 497, 672, and 687 to 690 nucleotides, respectively) of toroviruses showed high sequence identities (up to 99.4%, 98.7%, and 94.9% for the N, M, and S genes, respectively) between the isolate and BToVs. In contrast, the isolate was negative for BCV by RT-PCR. In a serological survey of serum samples from 355 calves at 33 farms, 92% of calves were positive for neutralizing antibodies to the isolate. These results indicate that the isolate in this study was BToV and that BToV infection might be common in cattle in Japan. To our knowledge, this is the first isolation of BToV in tissue culture.

Fecal samples from suckling ( n = 153) and weaned ( n = 116) piglets with diarrhea in Japan were examined for shedding of viral, bacterial, and parasitic pathogens using culture, microscopic, and polymerase chain reaction methods. In suckling piglets, diarrhea was attributed to infection with a single etiologic agent in 60.8% of cases and with combinations of agents in 22.2%. In weaned piglets, diarrhea was attributed to a single etiologic agent in 43.1% and to combinations of agents in 47.4% of cases. Rotavirus was the most prevalent agent in suckling (67.3%) and weaned (65.5%) piglets. The detection of other pathogens was associated with age of the animals examined. Coccidia were predominantly isolated from suckling piglets, whereas Escherichia coli was found predominantly in weaned piglets. Although a relationship was not observed between detection rate of rotavirus and age of piglets, a single group of rotavirus was detected in 87.5% of suckling piglets whereas multiple groups were detected in 51.6% of weaned piglets. The results of this study confirm that diarrhea in piglets can, to a variable degree, be causally associated with multiple agents. Additionally, these results suggest reasons why this syndrome can be difficult to control.

Type G9 of group A rotavirus (GAR) was shown to be predominant in a survey of VP7 (G) and VP4 (P) genotypes among porcine GARs associated with outbreaks of diarrhea in young pigs in Japan between 2000 and 2002. Comparison of the G9 VP7 gene sequences showed that the porcine G9 strains were more closely related to human G9 strains reemerging globally since the mid-1990s than to those from the mid-1980s. The VP7 gene sequences of porcine G9 strains from different farms were divergent (6.1 to 7.2% diference in nucleotides), suggesting that these G9 VP7 genes were not the result of recent introduction into the porcine population. Regarding the P genotype specificities of porcine G9 strains, while the majority of strains were close to unusual porcine P types (P[13] and P[23]), two strains were of the P[6] type, which has closer sequence identity with the human AU19 strain than with the porcine Gottfried strain. These unexpected results suggest that G9 GARs in the porcine population have spread more widely than previously thought and that the VP7 genes of porcine G9 strains and those of some human G9 strains detected recently may have a common progenitor., Facultad de Ciencias Veterinarias

Only two strains (Shintoku and porcine-like WD534tc) of group C rotavirus (GCR) from cattle have been reported to date. A GCR designated the Yamagata strain was the only pathogen detected in an outbreak of adult cow diarrhea accompanied by a decrease in milk production. The nucleotide sequences of the VP6 and VP7 genes from strain Yamagata were determined. Comparative sequence analysis showed that the sequence identities between strains Yamagata and Shintoku were markedly high in both VP6 gene (98.1%) and VP7 gene (93.5%), and that these strains belonged to the same clusters which were distinguished from GCRs from different host species in phylogenetic trees of these genes. These results suggested strongly that cattle species is one of the natural hosts of GCR infection, and that GCRs are a cause of adult cow diarrhea.

Serotype G14 was once considered to be uncommon among equine rotaviruses. While it sporadically emerged in some parts of the world, serotype G14 became the dominant G serotype among rotaviruses detected in foals with diarrhea in Japan in the late 1990s. However, it is not known how such recently emerging G14 rotaviruses are related in their overall genomic RNA constellation to prototype G14 strain identified earlier in the United States of America or how they were generated and why they have dominated over G3 equine rotaviruses. Genogrouping by RNA–RNA hybridization revealed that recently emerging serotype G14 equine rotavirus strains had an overall genomic RNA constellation that was highly conserved not only with contemporary and earlier G3 strains in Japan but also with prototype G3 and G14 strains previously identified in the United States of America. Japanese G14 rotavirus strains are likely to have originated form a VP7 gene substitution reassortant that had been formed earlier in the United States of America on the background of the then dominant G3 equine rotavirus.

Lactoferrin is a multifunctional, iron-binding glycoprotein found in physiological fluids of mammals. In the present study, a gene encoding the N-terminal half (N-lobe) of bovine lactoferrin was cloned and expressed in cultured insect cells using a baculovirus expression system. One mutation was found in the lactoferrin N-lobe gene, but it resulted in no amino acid substitution. The recombinant lactoferrin N-lobe was secreted into the culture medium and partially purified by means of an immobilized heparin column. The recombinant lactoferrin N-lobe secreted was not glycosylated, but it possessed antimicrobial activity toward Escherichia coli O111. The recombinant product synthesized and accumulated in the host cells exhibited greater electrophoretic mobility on SDS-PAGE than the secreted product and showed no potency to inhibit the growth of bacteria. It is thought that the product accumulated intracellularly lacks antimicrobial ability due to its degradation in the host cells or due to disruption of the active conformation.

A wasting disease characterized by progressive weight loss and dyspnea has been observed in weaning pigs on a farm in Yamagata Prefecture in 1998. Histopathologic findings in an affected pig were bronchointerstitial pneumonia and intracytoplasmic clusters of basophilic inclusions in macrophages of lymph nodes, which were similar to those in pigs with postweaning multisystemic wasting syndrome (PMWS) recently reported in North America and Europe. Porcine circovirus (PCV)-like particles were observed in bronchial lymph node of the pig by electron microscopy, and PCV antigens were detected in the lesions by immunohistochemical staining. PCV DNA was also detected in the lung and tonsil by PCR, and restriction fragment length polymorphism analysis of the PCR products with HinfI showed the same type of the PCV associated with PMWS (pmws PCV). Homology of nucleotide sequences between the PCR product and corresponding regions of published pmws PCV genomes was very high. These results indicated that virus detected in this study was pmws PCV. To our knowledge, this is the first report on the presence of pmws PCV in Japan.

豚繁殖・呼吸器障害症候群（以下PRRS）は，雌豚の繁殖障害と子豚の呼吸器症状を伴う疾病である。PRRSは全国の養豚場で大きな問題となっているが，その経済的な損失について調べた報告は少ない。そこで，関東地域の養豚場3戸を対象に，PRRSの発生に伴う経済的な損失について検討を行った。

The effects of a maternal vaccine against newborn calf diarrhea associated with group A bovine rotavirus (BRV), bovine coronavirus (BCV), bovine parvovirus and K99 Escherichia coli (E. coli) were examined on a beef cow-calf herd. After vaccination, serum or colostrum antibody titers to BRV, BCV and E. coli K99 in the vaccinated cows were significantly higher than those in unvaccinated control cows. Serum antibody titers to BRV, BCV and E. coli K99 in calves from the vaccinated cows were also significantly higher than those in calves from the control cows for 3-4 weeks after birth. These results suggested that the immunization of cows with the maternal vaccine enhanced the passive immunity levels in calves against BRV, BCV and K99 E. coli.

productive and respiratory syndrome. In: Diseases of swine, ed. Leman AD, Straw BE, Mengeling WL, et al., 7th ed., pp. 756762. Iowa State University Press, Ames, IA. 3. Benfield DA, Nelson EA, Collins JE, et al.: 1992, Characterization of swine infertility and respiratory syndrome (SIRS) virus (Isolate ATCC-VR 2332). J Vet Diagn Invest 4:127-133. 4. Christianson WT, Joo HS: 1994, Porcine reproductive and respiratory syndrome: a review. Swine Health Prod 2(2): 10-28. 5. Cohen SM, Ducharme CP, Carpenter CA, et al.: 1968, Rubella antibody in IgG and IgM immunoglobulins detected by immunofluorescence. J Lab Clin Med 72:760-766. 6. Goyal SM: 1993, Porcine reproductive and respiratory syndrome. J Vet Diagn Invest 5:656-664. 7. Hasche G, Stecher J, Gmelin K, et al.: 1984, Use of hepatitis B core antigen produced in E. coli to detect immunoglobulin M specific antibodies in an enzyme-linked immunosorbent assay. Eur J Clin Microbiol 3:30-34. 8. Kangro HO, Booth JC, Bakir TMF, et al.: 1984, Detection of IgM antibodies against cytomegalovirus: comparison of two RIA, ELISA and IFA test. J Med Virol 14:73-80. 9. Kim HS, Kwang J, Yoon IJ, et al.: 1993, Enhanced replication of porcine reproductive and respiratory syndrome (PRRS) virus in a homogeneous subpopulation of MA-104 cell line. Arch Virol 133:477-483. 10. Kimmel N, Friedman MC, Sarv I: 1982, Enzyme-linked immunosorbent assay (ELISA) for detection of herpes simplex virus-specific IgM antibodies. J Virol Methods 4:219-227. 11. McCaw MB, Molitor TW, Joo HS: 1992, Characterization of pseudorabies virus antibody response in young swine after infection and vaccination using an IgM antibody capture ELISA. J Clin Microbiol 30:346-350. 12. Wensvoort G, Terpstra C, Pol JMA, et al.: 1991, Mystery swine disease in the Netherlands: the isolation of Lelystad virus. Vet Q 13:121-130. 13. Yoon IJ, Joo HS, Christianson WT, et al.: 1992, Isolation of a cytopathic virus from weak pigs on farms with a history of swine infertility and respiratory syndrome. J Vet Diagn Invest 4:139-143. 14. Yoon IJ, Joo HS, Christianson WT, et al.: 1992, An indirect fluorescent antibody test for the detection of antibody to swine infertility and respiratory syndrome virus in swine sera. J Vet Diagn Invest 4:144-147. 15. Yoon IJ, Joo HS, Goyal SM, Molitor TW: 1994, A modified serum neutralization test for the detection of antibody to porcine reproductive and respiratory syndrome virus in swine sera. J Vet Diagn Invest 6:289-292.

Rotaviruses are members of the family Reoviridae and are a widespread cause of diarrhea in the young of many mammalian species, including humans. The rotavirus particles are nonenveloped and possess an icosahedral symmetry. The genome consists of 11 discrete segments of linear doublestranded RNA (dsRNA) and is enclosed within a doublelayered capsid. The dsRNA fragments are numbered 1 to 11 on the basis of their order of migration during polyacrylamide gel electrophoresis (PAGE). Rotaviruses are classified into serotypes based on the specificity of the outer capsid proteins VP7 (G types) and VP4 (P types) The more common method of serotyping is on the basis of G types (13), and currently 14 G types have been identified among group A rotaviruses. Among group A bovine rotaviruses (BRVs), at least 4 G types (G1, G6, G8, G10) have been reported based on the results of enzymelinked immunosorbent assay (ELISA), virus neutralization assays, nucleic acid hybridization assays, or sequence analysis. Rotaviruses are also typed using similar procedures on the basis of VP4 specificity, and currently there are at least 4 P types of BRV (P1, P5, P11, and P12). Upon analysis by PAGE, the 11 dsRNA segments of rotaviruses produce characteristic patterns referred to as genome electropherotypes. 35 Such electrophoretic analysis of BRV isolates is useful for obtaining epidemiologic information regarding the origin of the isolates, because each isolate produces a unique pattern. There have been numerous reports of short genome electropherotypes among human rotaviruses. Most of these short electropherotypes are restricted to subgroup I and share serotype G2 specificity. All BRV isolates tested so far, regardless of genome electropherotype, belong to subgroup I, and most possess long electropherotypes.38 Recently, there have been reports of short electropherotypes among BRV isolates, all restricted to serotype G6 1,22,37 Because there have been no reports of non-G6 BRV with a short genome pattern, we wanted to investigate the possibility that BRV short pattern isolates are restricted to serotype G6, similar to the situation observed for human G2 rotaviruses. In addition, we wanted to correlate the hybridization data with serology and cross-protection studies. In this report, we describe the serologic and molecular characterization (G and P specificity) of a bovine rotavirus strain, 2292B, with a short genome pattern. The reference viruses were grown in rhesus monkey kidney (MA104) cells in roller bottles and titrated by a plaque assay as previously described. The sources and serotypes (P and

The Cowden strain of porcine group C rotavirus and the Shintoku strain of bovine group C rotavirus were classified as different serotypes by two-way cross-neutralization tests. Two neutralization patterns against the Cowden and Shintoku strains were observed when hyperimmune or convalescent-phase antisera to three noncultivatable porcine group C rotaviruses and a human group C rotavirus were used in one-way cross-neutralization tests. Antisera to two porcine group C rotaviruses and the human group C rotavirus neutralized the Cowden strain at high titers but did not neutralize the Shintoku strain, suggesting that these three strains are serotypically related to the Cowden strain. The remaining antisera to a porcine group C rotavirus (HF strain) reacted with the Cowden and Shintoku group C rotaviruses in cell culture immunofluorescence tests but did not neutralize either virus in one-way cross-neutralization, suggesting that the HF strain belongs to a third serotype. However, confirmation of these findings requires additional analysis by two-way cross-neutralization. Our findings support the existence of at least two distinct serotypes of group C rotaviruses, and possibly a third, among animals and humans. The serotypic similarity observed between the Cowden strain and a human group C rotavirus suggests that the cultivatable Cowden strain and antiserum to this virus may provide important reagents for the diagnosis of group C rotaviruses in humans.

To further study the molecular characteristics of group (gp) C rotaviruses, we produced, cloned, and sequenced cDNA to gene 5 of the Shintoku strain of bovine gp C rotavirus. The resulting clone was specific for gene 5 and was genetically related to the human and porcine gp C rotaviruses, as demonstrated by Northern blot hybridization analysis. The Shintoku gene 5 is 1352 nucleotides in length and has one open reading frame encoding a polypeptide of 395 amino acids with a predicted molecular mass of 44.5 kDa. Comparative sequence analysis indicated that: (i) the Shintoku gene 5 protein shared 88.4 to 90.6% homology with the VP6 of the human (Bristol and 88–220) and porcine (Cowden) strains of gp C rotaviruses, but only low homology with the VP6 of bovine gp A (RF) and human gp B (ADRV) rotaviruses (41.3 and 16.3%, respectively); (ii) the predicted secondary structure was highly conserved among the gene 5 proteins of the bovine, porcine, and human gp C rotaviruses; and (iii) seven highly conserved regions were identified for the first time in the deduced primary amino acid sequences of gene 5 of gp C and gene 6 of gp A rotaviruses. However, only three of these highly conserved areas were present in the regions of VP6, where the secondary structure was predicted to be similar for the rotavirus strains examined. These three regions may contribute to common epitopes between the two groups of rotaviruses. Our results, in comparison with data for other rotaviruses, indicate that gene 5 of the bovine gp C rotavirus codes for the major inner capsid protein (VP6).

Two partial cDNA clones of genes 5 (encoding the major inner capsid protein VP 6) and 6 (encoding a nonstructural protein) of the porcine group (Gp) C rotavirus (Cowden strain) were radiolabeled with 32P and used individually as probes in Northern and dot blot hybridization assays. The specificity of each probe was tested against genomic dsRNA from: (1) porcine Gp A, B, and C rotaviruses; (2) Gp C rotaviruses from different species; and (3) porcine Gp C rotavirus field strains with varying electropherotype patterns. Neither probe hybridized with ds RNA from the porcine Gp A and B strains under the stringency conditions employed in the study. However, the gene 5 probe hybridized with the corresponding gene from the homologous porcine and the heterologous human and bovine Gp C rotaviruses tested. The gene 6 probe hybridized with the corresponding gene from the homologous Cowden strain, but hybridized weakly with gene 6 from the human and bovine Gp C rotaviruses. Both probes recognized all six different porcine Gp C field strains, although with varying intensities. Our results demonstrate that the gene 5 and 6 probes used in this study are specific for Gp C rotaviruses. However, evidence for greater genetic variation in the gene 6 among porcine, bovine and human Gp C strains suggested that the gene 5 probe may prove more broadly reactive among Gp C strains from different species. cDNA probes used in our study should prove useful for the detection of Gp C rotaviruses in feces and facilitate epidemiologic studies.

Three pregnant cows were inoculated intramuscularly with inactivated vaccine to bovine rotavirus (BRV) serotype 1 (BRV-1) and serotype 2 (BRV-2). Serum neutralizing antibody (NA) titers against both serotypes increased significantly after immunization. NA titers of colostrum obtained from immunized cows against BRV-1 and BRV-2 were 29286 and 38109, respectively, which were significantly higher than those from non-immunized control cows. Nine and 6 colostrum deprived calves were orally challenged with BRV-1 and BRV-2, respectively, and monitored for clinical manifestation and viral shedding. Five calves of them, 3 with BRV-1 and 2 with BRV-2, received 2 l of milk replacer supplemented with 10% immune colostrum 2 hr before challenge and twice daily for the first 5 days after challenge. Other 10 calves, 6 with BRV-1 and 4 with BRV-2, were fed only milk replacer as controls. All control calves developed severe diarrhea and shed a large amount of BRV in feces, beginning from 24 to 48 hr after challenge inoculation. On the contrary, all calves but one fed colostrum supplement remained clinically healthy after challenge, and BRV was not detected in their feces during feeding immune colostrum. The possibility that continuous feeding of immune colostrum is capable of preventing newborn calves from diarrhea associated with BRV and viral shedding was suggested.

Rotavirus B (RVB) has been identified as a causative agent of diarrhea in rats, humans, cattle, lambs, and swine. Recently, 20 RVB VP7 genotypes were determined based on an 80% nucleotide percent cut-off value. In this study, we sequenced the RVB VP6 gene segment from 80 RVB positive swine samples from the United States and Japan. Phylogenetic analyses, using the 30 available RVB VP6 sequences from GenBank and our 80 novel RVB VP6 sequences, revealed a large genetic diversity of RVB strains, mainly in pigs. For classification purposes, pairwise identity frequency analyses suggested an 81% nucleotide percent cut-off value, resulting in 13 RVB VP6 (I) genotypes. In addition, an intragenic recombinant RVB VP6 segment was identified from Japan. Furthermore, the data indicates frequent reassortment events occurred between the porcine RVB VP7 and VP6 gene segments. publisher: Elsevier articletitle: VP6 genetic diversity, reassortment, intragenic recombination and classification of rotavirus B in American and Japanese pigs journaltitle: Veterinary Microbiology articlelink: http://dx.doi.org/10.1016/j.vetmic.2014.05.015 content_type: article copyright: Copyright © 2014 The Authors. Published by Elsevier B.V. ispartof: Veterinary Microbiology vol:172 issue:3 pages:359-366 ispartof: location:Netherlands status: published

Background Both the G3P[12] and the G14P[12] type of equine group A rotavirus (RVA) have recently become predominant in many countries, including Japan. G3 types are classified further into G3A and G3B. The G3A viruses have been circulating in Europe, Australia, and Argentina, and the G3B viruses have been circulating in Japan. However, only an inactivated vaccine containing a single G3BP[12] strain is commercially available in Japan. To assess the efficacy of the current vaccine against recently circulating equine RVA strains, we examined antibody responses in pregnant mares to recent G3BP[12] and G14P[12] strains by virus neutralization test. Findings After vaccination in five pregnant mares, the geometric mean serum titers of virus-neutralizing antibody to recent G3BP[12] strains increased 5.3- to 7.0-fold and were similar to that against homologous vaccine strain. Moreover, antibody titers to recent G14P[12] strains were also increased 3.0- to 3.5-fold. Conclusions These results suggest that inoculation of mares with the current vaccine should provide foals with virus-neutralizing antibodies against not only the G3BP[12] but also the G14P[12] RVA strain via the colostrum.