Asymptomatic Enteric Virus Infections and Association with the Gut Microbiome in Rural Residents of Northern Laos.

Viral gastrointestinal infections are an important public health concern, and the occurrence of asymptomatic enteric virus infections makes it difficult to prevent and control their spread. This study aimed to determine the prevalence of and factors associated with asymptomatic enteric virus infection in adults in northern Laos. Fecal samples were collected from apparently healthy participants who did not report diarrhea or high fever at the time of the survey in northern Laos, and enteric viruses were detected using polymerase chain reaction (PCR) and reverse transcription (RT)-PCR. Individual characteristics, including the gut microbiome, were compared between asymptomatic carriers and noncarriers of each enteric virus. Of the participants (N = 255), 12 (4.7%) were positive for norovirus genogroup I (GI), 8 (3.1%) for human adenovirus, and 1 (0.4%) for norovirus GII; prevalence tended to be higher in less-modernized villages. Gut microbial diversity (evaluated by the number of operational taxonomic units) was higher in asymptomatic carriers of norovirus GI or human adenovirus than in their noncarriers. Gut microbiome compositions differed significantly between asymptomatic carriers and noncarriers of norovirus GI or human adenovirus (permutational analysis of variance, P <0.05). These findings imply an association between asymptomatic enteric virus infection and modernization and/or the gut microbiome in northern Laos.

Association between the dietary inflammatory index and disability in Japanese older people.

OBJECTIVES: To examine the demographic and lifestyle characteristics related to the dietary inflammatory index (DII™) score and to evaluate the association between DII score and disability among older people in Japan. DESIGN: Cross-sectional design. The DII score was calculated from nutrient intake information obtained from a FFQ. Disability was assessed using the Tokyo Metropolitan Institute of Gerontology Index of Competence questionnaire. Overall disability and disability in each component of everyday competence, that is, instrumental activities of daily living (IADL), intellectual activities and social participation, were assessed. Those with a deficit in one or more activities were defined as disabled. SETTING: Five non-urban areas in Japan. PARTICIPANTS: A total of 1642 Japanese older people aged 65 years or older. RESULTS: Women, residents of Oga-shi, and those with a higher education and greater frequency of shopping followed a more anti-inflammatory diet, while those living alone and residents of Minamiawaji-shi had higher dietary inflammation. A pro-inflammatory diet was associated with higher odds of overall disability and disability in each component of competence: overall disability, OR (95 % CI) = 1·26 (1·16, 1·36); IADL disability, OR (95 % CI) = 1·16 (1·07, 1·26); disability in intellectual activities, OR (95 % CI): 1·30 (1·20, 1·40); and disability in social participation, OR (95 % CI) = 1·20 (1·11, 1·29). CONCLUSIONS: Sex, living alone, education, frequency of shopping and area of residence were shown to be determinants of DII score in Japanese older people. DII score was positively associated with disability.

Role of rotavirus vaccination on G9P[8] rotavirus strain during a seasonal outbreak in Japan.

Although two live oral rotavirus (RV) vaccines, Rotarix and RotaTeq, play a critical role toward reducing disease severity, hospitalization, and death rate in RV infections, regular monitoring of vaccine effectiveness (VE) is yet necessary because the segmented genome structure and reassortment capability of RVs pose considerable threats toward waning VE. In this study, we examined the VE by a test-negative study design against G9P[8]I2 strain during a seasonal outbreak in February-May, 2018, in an outpatient clinic in Kyoto Prefecture, Japan. It remains important because G9P[8]I2 strain remains partially heterotypic to these vaccines and predominating in post-vaccination era. During year-long surveillance, RV infections were detected only from February to May. During this outbreak, 33 (42.3%) children out of 78 with acute gastroenteritis (AGE) remained RV-positive, of which 29 (87.8%) children were infected with G9P[8]I2. Two immunochromatographic (IC) assay kits exhibited 100% sensitivity and specificity to detect G9P[8]I2 strain. Only 23.2% children were found to be vaccinated. Yet, significant VE 69.7% (95% CI: 2.5%-90.6%) was recognized against all RV strains that increased with disease severity. Similar significant VE 71.8% (95% CI: 1%-92%) was determined against G9P[8]I2 strain. The severity score remained substantially low in vaccinated children. Our data reveal that vaccine-preventable G9P[8]I2 strain yet may cause outbreak where vaccination coverage remains low. Thus, this study emphasizes the necessity of global introduction of RV-vaccines in national immunization programs of every country.

The virucidal effect against murine norovirus and feline calicivirus as surrogates for human norovirus by ethanol-based sanitizers.

This study examined the virucidal effects of five types of alcohol-based sanitizers including malic acid and sodium malate, or monoethanolamin, in 58 vol % ethanol (pH 4.0, pH 7.1, pH 11.8), 65 vol % ethanol (pH 4.2), and 75 vol % ethanol (pH 4.4) against murine norovirus (MNV) and feline calicivirus (FCV). The virus titer of MNV was reduced in an ethanol dose-dependent manner under the same pH (about 4.0) condition. Virucidal effect against MNV was correlated with pH when the concentration of ethanol was constant (58 vol %). All the ethanol-based sanitizers provided sufficient virucidal effects against FCV. In conclusion, the virucidal effect of the ethanol-based sanitizer at low concentration of ethanol against norovirus (NoV) is increased when the pH is adjusted to a neutral state.

Flow cytometric assay using two fluorescent proteins for the function of the internal ribosome entry site of hepatitis C virus.

The initiation of translation in hepatitis C virus (HCV) occurs at the internal ribosome entry site (IRES) located at the 5'-end of its genomic RNA. To study the function of HCV IRES, we constructed a reporter plasmid that generates a bicistronic mRNA encoding two fluorescent proteins: cap-dependent DsRed2 and IRES-dependent Azami Green (AG). We introduced the plasmid into Huh7.5.1 and HEK293 cells and measured the relative IRES activity from the ratio of AG's signal to DsRed2's in individual cells using flow cytometry. To compare our method and a conventional biochemical method, we constructed a structurally similar reporter in which Renilla and Firefly luciferases replace DsRed2 and AG, respectively. With these systems, we found that the IRES A164G substitution decreased its activity, that interferon alpha affected the IRES activity in a cell type-specific manner, and that a synthetic micro-RNA targeting IRES was able to suppress the gene expression. In conclusion, the two methods were comparable in sensitivity in the studies of IRES mutations and host cell types. We discussed the significance of our findings and potential advantage of the cytometric assay: application to the molecular study of the HCV translation and to screening anti-IRES drugs.

Development of a simple system for screening anti-hepatitis C virus drugs utilizing mutants capable of vigorous replication.

Replication of infectious hepatitis C virus in Huh7 cells, a human hepatocyte cell line, has become possible due to the unique properties of the JFH1 isolate. Developing reporter virus systems for a simple titration has been attempted by integrating heterologous reporter genes into the JFH1 genome, resulting in a big infectivity reduction that limits the usefulness of such reporter systems. To overcome this problem, JFH1-infected Huh7 cells were cultured continuously for 2 years to obtain Huh7-adapted JFH1 variants capable of yielding up to 1000-fold higher titers. Sequence analysis of variant genome RNA suggested that this adapted population consisted mainly of two variants. By joining the 5'-half of the obtained representative viral complementary DNA (cDNA) fragments of the variants with the 3'-half of the wild-type's, two prototype clones, A/WT and B/WT, were constructed. Replication of A/WT and B/WT viruses in Huh7 cells showed up to 100-1000-fold higher titers than the wild-type. A Renilla luciferase cDNA was inserted into the Nonstructural Protein 5A region of the A/WT and B/WT cDNA to generate A/WT-Rluc and B/WT-Rluc, respectively. Transfection of Huh7 cells with in vitro-transcribed A/WT-Rluc and B/WT-Rluc RNA resulted in production of infectious viruses with approximately 15- and 25-fold higher titers, respectively, than the wild-type RNA. The replication of A/WT-Rluc and B/WT-Rluc viruses was more vigorous than the wild-type even with insertion of the luciferase cDNA showing a good correlation of luciferase activities with infectious titers. Furthermore, interferon-alpha inhibited the replication of A/WT-Rluc and B/WT-Rluc viruses in a dose-dependent manner as determined by a luciferase assay. These results imply that our system is potentially a tool useful for screening anti-hepatitis C virus drugs in a simple and time/cost-saving manner.

Throughput expression of multiple G-protein coupled receptors for HIV infection in choriocarcinoma cells, trophoblasts, and breast milk cells.

The chemokine receptors, which belong to G-protein coupled receptors (GPCRs) and become co-receptors when HIV enters the cell, have been mentioned in recent research. Numerous studies have reported that the cellular mechanism of HIV crossing the placental barrier is still not totally understood. This study was conducted to investigate whether the mRNAs of nineteen typs of GPCRs and CD4 were expressed in choriocarcinoma cell lines, trophoblasts, and breast milk cells by using RT-PCR. It was found that the expression of GPCRs varied in different cell lines. Of note is that CD4 could not be expressed in either choriocarcinoma cells or trophoblasts. It was noteworthy that mRNAs of multiple GPCRs were identified in choriocarcinoma cells, trophoblasts, and breast milk cells for the first time. The expression amounts of these mRNAs were further measured by quantitative RT-PCR. Interestingly, mRNAs of CCR9/CCR10 were strongly expressed in trophoblasts. This study provided further insights to the cellular mechanism of mother-to-child transmission of HIV.

Development of a rapid immunochromatographic test for noroviruses genogroups I and II.

Norovirus (NoV) is known to cause acute gastroenteritis in children worldwide. Although reverse transcription-PCR (RT-PCR) method is considered to be the "gold standard" for diagnosis of this viral infection, it requires skillful personnel and well-equipped laboratory. In this study, a rapid and easily performable diagnostic kit was developed using immunochromatographic method with rabbit polyclonal antibodies raised against recombinant virus-like particles (rVLPs) of most prevalent genotypes, genogroup II genotypes 3 and 4. This kit was evaluated for reactivity to rVLPs and detection of natural viruses in stool samples collected from children with diarrhea in comparison to the results obtained by RT-PCR. In the prospective assessment, the kit showed agreement rate of 84.1%, sensitivity of 69.8% and specificity of 93.7%. Genotyping of the RT-PCR positive samples by sequence analysis revealed that some heterogeneous genotypes were also detected while some in homogeneous genotypes occasionally showed false negative records resulting in lower sensitivity. No cross-reactivity with other common viral pathogens was observed. Taken together with the result of the detection limit of viral load as small as approximately 10(6-7)copies/g of stool, the current immunochromatography test is justified for screening for NoV infection with simple laboratory support.

Sequence analysis of Vietnamese P[6] rotavirus strains suggests evidence of interspecies transmission.

Nucleotide and amino acid sequences of the VP8* gene of five Vietnamese P[6] rotavirus strains detected from hospitalized patients with acute gastroenteritis were analyzed and compared with other human and porcine P[6] rotaviruses. It is of interest that these strains had greatest identity with two Italian porcine rotavirus strains, 134/04-10 and 134/04-11. To our knowledge, these five Vietnamese rotaviruses are the rare P[6] rotavirus strains belonging to lineage I that cluster into sublineage Ic with porcine rotaviruses, and not into sublineage Ia, as other human P[6] rotaviruses have done so far. Sequence analysis of the VP7 gene of these P[6] rotavirus strains was also performed. The results showed that the Vietnamese G9P[6] strain had high similarity with other human G9 rotaviruses, confirming a human-animal reassortant virus, whereas other three G4P[6] strains had best identity with porcine G4 rotavirus strains, suggesting interspecies transmission of rotavirus between porcine and humans. This result provides the important data on molecular characteristics of Vietnamese rotaviruses, and highlights interspecies transmission events of rotaviruses in Vietnam as well as in Asia.

Detection and genetic characterization of norovirus in oysters from China and Japan.

A total of 225 oysters from China and Japan were collected during October 2005 to September 2006 and were then tested for the presence of norovirus by RT-nested PCR. The detection rate of norovirus was different between China and Japan, accounting for 14.6% (19 of 130) and 25.3% (24 of 95), respectively. In China, norovirus in oyster was detected continuously from July to February with the highest prevalence in August, October and November (each of 21%, 4 of 19). On the other hand, norovirus in Japan was found year-round with highest prevalence in March and October (each of 20.8%, 5 of 24). Norovirus strains detected were subjected to further characterization by sequence analysis. It was found that the norovirus strains belonged to only two distinct genotypes, the GII/3 (known as the Mexico virus cluster) and the GII/4 (known as the Lordsdale virus cluster). In China, the norovirus GII/4 was the most predominant, accounting for 78.9% (15 of 19). In contrast, it was interesting that both the norovirus GII/4 and the norovirus GII/3 were co-predominant with a prevalence of 50% (12 of 24) in Japan. Another interesting feature of the study was that the norovirus GII/4 strains in oysters from both countries were grouped into two distinct variant clusters known as the Farmington Hills variant and the Hunter variant. More than 102 copies of norovirus were detected in 41 of 43 oysters. This study provided additional evidence of the presence of norovirus in oysters and is also the first report to demonstrate the existence of norovirus variants in oysters.

Photocatalytic inactivation of diarrheal viruses by visible-light-catalytic titanium dioxide.

Titanium dioxide (TiO2) that had been irradiated with visible light (VL) was demonstrated to inactivate rotavirus, astrovirus, and feline calicivirus (FCV). The virus titers were dramatically reduced after exposure for 24 hrs to the VL-catalytic TiO2. The addition of bovine serum albumin could protect the virus against inactivation by VL-catalytic TiO2 in a dose-dependent manner. This finding implied that the VL-catalytic TiO2 products might somehow interact initially with the viral proteins in the process of virus inactivation. Moreover, we showed partial degradation of the rotaviral dsRNA genome. This was more prominent when the virus was exposed to the VL-catalytic TiO2 treatment for at least 2 days. An attempt was made to elucidate the mechanism underlying the inactivation of the viruses. It was found that upon activation of TiO2 with VL by using a white fluorescent lamp, the reactive oxygen species such as superoxide anions (O2-) and hydroxyl radicals (*OH) were generated in a significant amount after stimulation for 8, 16, and 24 hrs. We therefore assume that virus inactivation by VL-catalytic TiO2 might occur through the generation of O2- and *OH followed by damage to the viral protein and genome. This is the first report, to the best of our knowledge, demonstrating the inactivation of rotavirus, astrovirus and FCV by the presence of TiO2 film under VL as well as describing its mechanism.

Molecular and epidemiological trend of norovirus associated gastroenteritis in Dhaka City, Bangladesh.

BACKGROUND: Diarrhea, over the years, has killed millions of people and continues to be a major threat in Bangladesh. OBJECTIVES: To determine the incidence of norovirus infection in infants and young children with acute gastroenteritis in Dhaka City, Bangladesh and to determine the genogroup and genotype in norovirus-positive stool specimens. STUDY DESIGN: Fecal specimens were collected from infants and children with acute gastroenteritis in Dhaka City, Bangladesh from October 2004 to September 2005, and examined for norovirus by reverse transcription-polymerase chain reaction. RESULTS: Noroviruses were detected in 41 of 917 fecal specimens. Molecular analysis of norovirus was carried out by sequencing methods. Only norovirus GII/4 strains were detected during this study. The dominant genotype throughout the study period was GII/4. Norovirus infections were most commonly observed in winter and rainy seasons in Dhaka City. The common clinical symptoms in norovirus-infected patients were diarrhea (90%), vomiting (75%) and abdominal pain (46%). CONCLUSIONS: This is the first epidemiological research of norovirus in Bangladesh. Norovirus is an important enteropathogen responsible for viral gastroenteritis among infants and children in Bangladesh.

Characterization of a broadly reactive monoclonal antibody against norovirus genogroups I and II: recognition of a novel conformational epitope.

Norovirus, which belongs to the family Caliciviridae, is one of the major causes of nonbacterial acute gastroenteritis in the world. The main human noroviruses are of genogroup I (GI) and genogroup II (GII), which were subdivided further into at least 15 and 18 genotypes (GI/1 to GI/15 and GII/1 to GII/18), respectively. The development of immunological diagnosis for norovirus had been hindered by the antigen specificity of the polyclonal antibody. Therefore, several laboratories have produced broadly reactive monoclonal antibodies, which recognize the linear GI and GII cross-reactive epitopes or the conformational GI-specific epitope. In this study, we characterized the novel monoclonal antibody 14-1 (MAb14-1) for further development of the rapid immunochromatography test. Our results demonstrated that MAb14-1 could recognize 15 recombinant virus-like particles (GI/1, 4, 8, and 11 and GII/1 to 7 and 12 to 15) and showed weak affinity to the virus-like particle of GI/3. This recognition range is the broadest of the existing monoclonal antibodies. The epitope for MAb14-1 was identified by fragment, sequence, structural, and mutational analyses. Both terminal antigenic regions (amino acid positions 418 to 426 and 526 to 534) on the C-terminal P1 domain formed the conformational epitope and were in the proximity of the insertion region (positions 427 to 525). These regions contained six amino acids responsible for antigenicity that were conserved among genogroup(s), genus, and Caliciviridae. This epitope mapping explained the broad reactivity and different titers among GI and GII. To our knowledge, we are the first group to identify the GI and GII cross-reactive monoclonal antibody, which recognizes the novel conformational epitope. From these data, MAb14-1 could be used further to develop immunochromatography.

Anti-norovirus polyclonal antibody and its potential for development of an antigen-ELISA.

Norovirus (NoV) capsid proteins were expressed as virus-like particles (VLPs) by using recombinant baculovirus in insect cells, which had 5 genotypes in genogroup I and 11 genotypes in genogroup II, and the VLPs were used as immunogens. Polyclonal antibody against the VLP of GII/3 genotype showed broad-range cross-reactivity, reacting not only with intra-genogroup strains, but also inter-genogroup strains, by antibody-ELISA using 16 kinds of VLPs. Furthermore, antigen-ELISA was conducted in sandwich enzyme-linked immunosorbent assay (ELISA) using the polyclonal antibody for capturing antigens, and three kinds of monoclonal antibodies against the VLP of GII/4 genotype for detecting antigens. This format successfully detected eight genotypes of NoV from clinical specimens and proved that polyclonal antibody, which has broad-range cross-reactivity, was capable of detecting various types of genotypes from clinical specimens.

Sequence analysis of the VP7 gene of human rotavirus G1 isolated in Japan, China, Thailand, and Vietnam in the context of changing distribution of rotavirus G-types.

Over the last decade, rotavirus G1 has represented the most common genotype worldwide. Since 2000, the prevalence of rotavirus G1 has decreased in some countries such as Japan and China. To monitor the trend of the VP7 encoding gene of rotavirus G1, we performed a sequence analysis of 74 G1 rotavirus strains isolated in Japan, China, Thailand, and Vietnam during the period from 2002 to 2005. The phylogenetic tree showed that all of the studied G1 strains from the four countries clustered into lineage III, the same as the majority of the G1 strains isolated in China and Japan in 1990 and 1991. Examination of the deduced amino acid sequences of the G1 strains from China and Japan revealed an amino acid substitution at position 91 (Asn instead of Thr) in antigenic region A when compared to the G1 strains isolated in China and Japan in 1990, 1991, and global reference strains. For the G1 strains from Thailand and Vietnam, there were three amino acid substitutions, not belonging to any antigenic regions. The study showed that there have been no considerable changes of human rotavirus G1 isolated in Japan, China, Thailand, and Vietnam. Further studies need to be carried out for a better understanding of why such changes in the prevalence of rotavirus G1 occur in these countries.

Risk factors for protein-energy malnutrition in children under 5 years: study from Luangprabang province, Laos.

BACKGROUND: Laos is one of the poorest countries in which chronic malnutrition is highest. The aim of the present study was to determine the prevalence of and to identify risk factors associated with protein-energy malnutrition (PEM) in children under 5 years of age in Luangprabang province, Laos. METHODS: This cross-sectional study was undertaken from March to May 2004. Anthropometric measurements of 798 children were done and data were transformed into height-for-age, weight-for-age and weight-for-height ratios. Mothers were also interviewed with a semi-structured questionnaire. Anthropometric data were entered into Nutstat in Epi-Info 2000 and transferred to SPSS for analysis. RESULTS: There was a high prevalence of stunting, underweight and wasting, that is, 54.6%, 35%, and 6% respectively. It was also noted that children aged 12-23 months and Khmu ethnic children had a higher prevalence of stunting (65% and 66%) and underweight (45% and 40%), respectively. However, it was also found that boys were more prone to be stunted and underweight. Furthermore, restricted intake of meats, vegetables during illness, and low maternal education were main risk factors for child malnutrition in the study area. CONCLUSION: Socioeconomic-demographic factors, low maternal education, poor nutrition knowledge for mother and feeding practices for sick children are affecting children's health regarding stunting and underweight. It is recommended that an improvement in societal infrastructure, better maternal education and nutrition are needed to address the child malnutrition issue.

Diversity of viruses associated with acute gastroenteritis in children hospitalized with diarrhea in Ho Chi Minh City, Vietnam.

A molecular epidemiological study on common diarrheal viruses was conducted in Ho Chi Minh City, Vietnam between October 2002 and September 2003. Fecal samples were collected from 1,010 hospitalized children with acute gastroenteritis. Those samples were screened for groups A, B, and C rotavirus, adenovirus, genogroups I and II norovirus (NoV), sapovirus (SaV), and human astrovirus (HAstV) by RT-multiplex PCR, and the positive specimens were characterized further by ELISA, nested PCR, or sequencing. Among the diarrheal viruses detected, group A rotavirus was the most common, with a proportion of 67.4%, whereas NoV GII, adenovirus, SaV, and HAstV were also found in 5.5, 3.2, 0.8, and 0.6%, respectively. It is noteworthy that the group C rotavirus was first reported in Vietnam, with a proportion of 0.5% in this study. Fifty-six of 1,010 (5.5%) samples were found positive with more than one viral agent, in which 25 samples contained both group A rotavirus and NoV GII. Group A rotavirus could be identified throughout year with the peaks in both the dry and rainy season, whereas other viruses prevailed mainly in the rainy season. G-typing for the group A rotavirus showed that genotype 1 was still the most prevailing (33.0%), but interestingly, serotype 9 was emergent and became the third most common rotavirus G-type in these samples (13.7%). The four most common G-P combinations globally, G1P[8], G2P[4], G3P[8], and G4P[8] were found in 46.8% of rotavirus-positive samples, and it is of interest that one unusual rotavirus G9P[19] strain was first detected in Vietnam. The majority of NoV strains belonged to GII/4, and SaV strains mainly clustered with the Manchester strain (GI/1). Twenty-seven out of 32 adenovirus strains were identified as serotype 41. All HAstVs belonged to genotype 1. The results indicated clearly the impact of viral agents causing gastroenteritis and the importance of vaccination against diarrhea in Vietnam.

Prevalence of sapovirus infection among infants and children with acute gastroenteritis in Dhaka City, Bangladesh during 2004-2005.

Sapovirus, a member of the family Caliciviridae is one of the major causative agents of viral gastroenteritis affecting all age group. Sapovirus was detected in 25 of 917 stool specimens from infants and children with acute gastroenteritis in a Children Hospital in Dhaka City, Bangladesh during 2004-2005. All fecal specimens were examined for sapovirus by reverse transcription-polymerase chain reaction. Molecular analysis of sapovirus was carried out by sequencing methods. Sapovirus detected in this study was clustered into only one distinct genogroup I. Sapovirus GI/1 was predominant, followed by GI/2 and accounted for 92% (23 of 25) and 8% (2 of 25), respectively. The results clearly indicated that sapovirus infections were observed most commonly in the autumn to winter seasons (September to January) in Dhaka City. The common clinical symptoms of sapovirus infected patients were dehydration (88%), vomiting (76%), and abdominal pain (60%). This is the first report of sapovirus in Bangladesh.

Emergence of new variant rotavirus G3 among infants and children with acute gastroenteritis in Japan during 2003-2004.

A total of 402 fecal specimens from infants and children with acute gastroenteritis in five places (Tokyo, Maizuru, Saga, Sapporo, and Osaka) in Japan from July 2003 to June 2004 were collected and then tested for the presence of rotavirus by RT-PCR. Of these, 83 were positive for rotavirus and this accounted for 20.6%. Rotavirus was further characterized to G-types (VP7 genotypes) and P-types (VP4 genotypes). Interestingly, an emergence of rotavirus G3 was identified with an exceptionally high prevalence (97.5%; 81 of 83), followed by rotavirus G2 (2.5%; 2 of 83). The P-types of 19 rotavirus strains, which could not be typed by RT-PCR, were determined as P[8] with multiple point mutations at the VP4 primer-binding site by sequencing analysis. The predominant genotype was G3P[8] (95.2%, 79 of 83), followed by a number of unusual combinations G3P[4] (2.4%, 2 of 83), and G2P[8] (2.4%, 2 of 83). Another interesting feature of the study was the demonstration of a great genetic diversity in new variant rotavirus G3 strains circulating in Japan. In comparison with rotavirus G3 strains circulating in 1990-1995 in Japan, a wide range of amino acid substitutions (up to 16) of new variant rotavirus G3 VP7 genes was identified. Of note, the changes at positions 96, 99, and 100 were revealed to be located in the antigenic region A, and 213 in the antigenic region C. To the best of our knowledge, this is the first reporting of an emergence of new variant rotavirus G3 together with a sudden disappearance of G1, G4, and G9 in infants and children with rotavirus infection-associated gastroenteritis in Japan.

Novel porcine rotavirus of genotype P[27] shares new phylogenetic lineage with G2 porcine rotavirus strain.

A novel and unusual strain of porcine rotavirus (PoRV) CMP034 was isolated from a 7-week-old piglet during the epidemiological survey of porcine rotavirus infection in Chiang Mai province, Thailand from June 2000 to July 2001. Molecular characterization of gene VP4 by sequence analysis showed a low level of amino acid sequence identity, ranging from 56.7% to 76.6%, while comparison of VP8* portion showed 41.8% to 69.9% identity, with the 26 P genotypes recognized to date. Phylogenetic analysis of the VP4 sequence revealed that CMP034 was only distantly related to the other 26 P genotypes and was located in a separate branch. Sequence analysis of gene VP7 showed the highest level of amino acid identity (94.7%) with the PoRV G2-like reference strain 34461-4 but a lower level of identity with those of human G2 rotaviruses, ranging from 87.7% to 88.0%. Phylogenetic analysis of gene VP7 revealed two major lineages among G2 rotavirus strains based on the host origin. PoRV strain CMP034 clustered exclusively with G2-like PoRV strain 34461-4 in a novel lineage that is distinct from the major G2 human lineage. Moreover, strain CMP034 displayed a porcine-like VP6 and NSP5/6 with subgroup I specificity, while bearing an NSP4 with some genetic group B human-like characteristics. These findings provide evidence that CMP034 should be considered as a novel VP4 genotype P[27].