To investigate the prevalence of bovine papillomavirus (BPV) in bovine papilloma and healthy skin, DNA extracted from teat papillomas and healthy teat skin swabs was analysed by PCR using the primer pairs FAP59/FAP64 and MY09/MY11. Papillomavirus (PV) DNA was detected in all 15 papilloma specimens using FAP59/FAP64 and in 8 of the 15 papilloma specimens using MY09/MY11. In swab samples, 21 and 8 of the 122 samples were PV DNA positive using FAP59/FAP64 and MY09/MY11, respectively. Four BPV types (BPV-1, -3, -5 and -6), two previously identified putative BPV types (BAA1 and -5) and 11 putative new PV types (designated BAPV1 to -10 and BAPV11MY) were found in the 39 PV DNA-positive samples. Amino acid sequence alignments of the putative new PV types with reported BPVs and phylogenetic analyses of the putative new PV types with human and animal PV types showed that BAPV1 to -10 and BAPV11MY are putative new BPV types. These results also showed the genomic diversity and extent of subclinical infection of BPV.

Human norovirus (NoV) strains cause a considerable number of outbreaks of gastroenteritis worldwide. Based on their capsid gene (VP1) sequence, human NoV strains can be grouped into two genogroups (GI and GII) and at least 14 GI and 17 GII genotypes (GI/1-14 and GII/1-17). Human NoV strains cannot be propagated in cell-culture systems, but expression of recombinant VP1 in insect cells results in the formation of virus-like particles (VLPs). In order to understand NoV antigenic relationships better, cross-reactivity among 26 different NoV VLPs was analysed. Phylogenetic analyses grouped these NoV strains into six GI and 12 GII genotypes. An antibody ELISA using polyclonal antisera raised against these VLPs was used to determine cross-reactivity. Antisera reacted strongly with homologous VLPs; however, a number of novel cross-reactivities among different genotypes was observed. For example, GI/11 antiserum showed a broad-range cross-reactivity, detecting two GI and 10 GII genotypes. Likewise, GII/1, GII/10 and GII/12 antisera showed a broad-range cross-reactivity, detecting several other distinct GII genotypes. Alignment of VP1 amino acid sequences suggested that these broad-range cross-reactivities were due to conserved amino acid residues located within the shell and/or P1-1 domains. However, unusual cross-reactivities among different GII/3 antisera were found, with the results indicating that both conserved amino acid residues and VP1 secondary structures influence antigenicity.

Infectious acute gastroenteritis is an important public health problem worldwide. A total of 639 stool specimens were tested for the presence of diarrhea pathogens. The specimens were from outpatients with acute gastroenteritis who consulted the pediatric clinic in Kumamoto Prefecture, Japan, from June 2002 to December 2007. Of these, 421 (65.9%) were positive for diarrhea pathogens. Among them were norovirus (NoV) in 260 (61.8%), sapovirus (SaV) in 81 (19.2%), rotavirus in 49 (11.6%), adenovirus in 19 (4.5%), enterovirus in 13 (3.1%), astrovirus in 9 (2.1%), kobuvirus in 1 (0.2%), and bacterial pathogens in 11 (2.6%). Mixed infection (co-infection of viruses) was found in 22 (5.2%) of the 421 pathogen-positive stool samples. NoV was the most prevalent pathogen throughout the study period; however, the SaV detection rate was unexpectedly high and was found to be the secondary pathogen from 2005 to 2007. Genetic analysis of SaV with 81 strains demonstrated that SaV strains belonging to genogroup IV emerged in 2007, and dynamic genogroup changes occurred in a restricted geographic area. This study showed that SaV infection is not as rare as thought previously.

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Sapporo-like viruses (SLV) are a causative agent of gastroenteritis in humans. SLV-specific primers were newly designed in capsid protein-coding region and 529 fecal samples collected from gastroenteritis patients were tested. Thirty-five samples (6.6%) were found to be positive by reverse transcriptase-polymerase chain reaction (RT-PCR). Out of 35 positive samples, 25 were classified into a genogroup SG-I, typified by the Sapporo virus, and 9 were classified as a genogroup SG-II, typified by the London virus. Interestingly, one sample could not be classified into any genogroup, which suggests that it may be part of a new SLV genogroup. The RT-PCR used in this study appeared to be capable of widely detecting SLV genogroups, and it was seen to be powerful enough for molecular epidemiological as well as phylogenetic studies on SLV.

Sapovirus (SV) causes gastroenteritis in humans and comprises genetically divergent viruses. A nested reverse transcription-polymerase chain reaction (RT-PCR) targeting the capsid-protein-coding region was developed using universal and genogroup-specific primer sets. The universal primers were capable of detecting human SV genogroups I, II, IV and V. Genetic analysis of the amplified products enabled us to phylogenetically determine the genotypes of the viruses. In addition, genogroup-specific primers that amplified different lengths of the amplicon depending on the genogroup were developed. These genogroup-specific primers were also used as inner primers for the nested PCR. These two simple RT-PCR methods are powerful tools for both detection and epidemiological studies of human SV.

Noroviruses (NVs) are common pathogens that consist of genetically divergent viruses that induce gastroenteritis in humans and animals. Between September 1999 and June 2004, 1,898 samples obtained from patients showing sporadic or outbreak gastroenteritis in Chiba Prefecture, Japan, were tested for NVs by reverse transcription-PCR. NVs were detected in 603 samples. Approximately 80% were positive for genogroup GII, 13% were positive for genogroup GI, and the remaining 7% were positive for both genogroups. Phylogenetic analysis showed that the GI and GII genogroups could be further divided into 13 and 16 genotypes (including new genotypes), respectively. The GII-4 genotype, which included five small genetic clusters (subtypes), was the most common in this study and was detected in approximately 40% of positive samples. The P2 regions of 10 strains belonging to each of the five GII-4 subtypes showed 5 to 18% amino acid diversity. The amino acid substitutions accumulated in the protruding (P) region during the 5-year study period. Our data suggest that highly variable NV strains are circulating in Chiba Prefecture, with a high rate of genetic change observed during the 5-year study period.The genus Norovirus is a member of the family Caliciviridae. Caliciviruses contain a positive-sense single-stranded RNA genome and include a further three genera, Vesivirus, Lagovirus, and Sapovirus (2,3,8). Noroviruses (NVs) have three major open reading frames (ORFs) that encode nonstructural, capsid, and minor structural proteins, respectively (8). They are one of the most common causes of gastroenteritis and have been detected in fecal samples from both humans (12,15,28) and animals (20,30,37). Human-associated NV outbreaks resulting from ingestion of contaminated water or food, such as oysters (4,5,18,23), and outbreaks in public places, particularly hospitals, schools, and cruise ships (9,11,22,36), pose an important public health problem.Reverse transcription-PCR (RT-PCR) and sequencing of the partial viral genome are the most popular and useful procedures for obtaining epidemiological and genetic information on NVs. Human NVs can be divided into two genogroups, genogroups GI and GII, by genetic analysis of the RNA polymerase and capsid regions (1, 15), with several genotype classifications having been reported independently (1,16,33). Recently, based on the genotype classification of Katayama et al. (16), Kageyama et al. (15) reported on a detailed scheme for the genotyping of NVs based on distribution analysis by using the pairwise distance of the capsid N-terminal/shell domain. They classified the GI and GII genogroups into 14 and 17 genotypes, respectively.During the winter of 2002-2003, an increase in NV outbreaks was reported in Europe and the United States (6, 21). Moreover, worldwide, the GII-4 genotype (Bristol virus-like genotype) has been shown to be the predominant strain of NV associated with gastroenteritis (13,21,(34)(35)(36). Changes in the phylogenetic and genetic characteristics of GII-4 genotype strains...

Two G12 human rotavirus strains, CP727 and CP1030, were isolated from the respective diarrheic stools of an infant and an adult in Japan. VP7 gene sequences of strains CP727 and CP1030 showed high identity with that of the G12 prototype strain L26, and with those of G12 strains reported recently from Thailand, the United States, and India. VP4 gene sequences of strains CP727 and CP1030 showed the highest identity with those of P[9] rotaviruses. In Northern blot hybridization, strains CP727 and CP1030 were found to be closely related to strain AU-1 (G3P[9]); nine RNA segments hybridized to each other. Moreover, all segments each of the two Japanese G12 strains hybridized to those of the Thai G12 strain T152. These results suggest that Japanese G12 strains detected in this study are reassortants between a L26-like strain and a strain in the AU-1 genogroup. A similar reassortant was found in the Thai G12 strain T152.