Recombinant norovirus implicated in gastroenteritis outbreaks in Hiroshima Prefecture, Japan

Norovirus (NoV) is a major etiological agent of acute gastroenteritis outbreaks worldwide. A total of 314 fecal specimens collected from patients of 39 NoV gastroenteritis outbreaks in Hiroshima Prefecture, Japan, between December 2001 and April 2006 were tested for the occurrence of recombinant NoVs. Sixteen genotypes (GI/1, GI/2, GI/4, GI/7, GI/8, GI/11, GI/14, GII/2, GII/3, GII/4, GII/5, GII/6, GII/8, GII/12, GII/14, and GII/untypeable) were detected in the 39 outbreaks based on capsid sequences and GII/4 was predominant recently. Twelve strains detected in 11 (28.2%) of the 39 outbreaks were suspected to be recombinants by using Simplot and Recco analyses and five recombinant genotypes, GII/4-GII/12 (five strains), GIIb-GII/3 (four strains), GII/4-GII/2 (one strain), GII/4-GII/14 (one strain), and GI/2-GI/8 (one strain), were identified based on RNA-dependent RNA polymerase and capsid sequences. None of the strains genotyped as GII/4 based on the capsid sequence was identified as a recombinant. The putative recombination points in the recombinant strains were placed either upstream or downstream of the open reading frame (ORF) 1 and ORF2 overlap. The present study indicates the following: (a) recombination among ORFs is common in nature, (b) the involvement of recombinant NoVs in gastroenteritis outbreaks is extensive even in a local area such as Hiroshima Prefecture, Japan, and (c) the conserved region (ORF1 and ORF2 overlap) has a meaningful function against the recombination event.     

Existence of multiple genotypes associated with acute gastroenteritis during 6-year survey of norovirus infection in Japan

Norovirus (NoV) is recognized as one of the most common causative agent of diarrheal disease in young children worldwide. The current study was undertaken to determine the distribution of NoV genotypes in Japan. A total of 3,864 fecal specimens from children with acute gastroenteritis in five regions (Tokyo, Maizuru, Saga, Sapporo, and Osaka) of Japan from July 1995 to June 2001 were collected and then tested for the presence of NoV by RT-PCR. Three hundred sixty four were found to be positive for NoV, accounting for 11%. The highest prevalence of NoV infection was in November, December, and January as the early winter months in Japan. NoV was subjected to be further characterized to sequencing analysis. All NoVs belonged to two different genogroups I and II and these represented 3% and 97%, respectively. This finding indicated that NoV genogroup II was the dominant group causing acute gastroenteritis in Japan. Interestingly, NoV strains were classified into 16 distinct genotypes including genogroup II genotype 9 that was firstly identified in Japan. Of these, NoV genogroup II genotypes 3 and 4 dominated over other genotypes and became the leading strains in Japanese pediatric population. In conclusion, diarrhea due to NoV infection is still a health burden in Japan. This report also stresses the great genetic diversity as well as the importance of NoV causing the diarrhea in Japan.     

Prevalence of Norwalk-like virus infections in cases of viral gastroenteritis among children in Osaka City,Japan

Surveillance of Norwalk-like virus (NLV) infections in cases of pediatric gastroenteritis between April 1996 and March 2000 showed that NLVs were an important causative agent in viral gastroenteritis cases among children between November and January in those years. The predominant type of NLV was closely related to Lordsdale virus in genogroup 2. During the 1999-2000 season, Arg320-like strains, which may be genetic recombinants, suddenly appeared and spread.     

Fatal case of acute gastroenteritis with multiple viral coinfections

We report a fatal case of acute gastroenteritis in a child with autism spectrum disorder. Multiple viral coinfections were detected by PCR in the patient’s stool and digestive biopsy specimens. As viral detection is not necessarily associated with symptomatic disease, a semi-quantitative approach using cycle treshold values was proposed for the clinical interpretation of PCR. We discuss whether concomitant viral infections could be a risk factor for severe outcome in gastroenteritis cases. Individual risk factors are also addressed.     

Clostridium difficile is not associated with outbreaks of viral gastroenteritis in the elderly in the Netherlands

The coincidental increase in norovirus outbreaks and Clostridium difficile infection (CDI) raised the question of whether these events could be related, e.g. by enhancing spread by diarrhoeal disease outbreaks. Therefore, we studied the prevalence of C. difficile in outbreaks of viral gastroenteritis in nursing homes for the elderly and characterised enzyme immunoassay (EIA)-positive stool samples. Stool samples from nursing home residents (n = 752) in 137 outbreaks of viral aetiology were investigated by EIA for the presence of C. difficile toxins. Positive samples were further tested by a cell neutralisation cytotoxicity test, a second EIA and culture. Cultured isolates were tested for the presence of toxin genes, the production of toxins and characterised by 16S rRNA polymerase chain reaction (PCR) and sequencing. Twenty-four samples (3.2%) tested positive in the EIA. Of these 24 positive samples, only two were positive by cytotoxicity and three by a second EIA. Bacterial culture of 21 available stool samples yielded a toxinogenic C. difficile PCR ribotype 001 in one patient sample only. In conclusion, we found no evidence in this retrospective study for an association between viral gastroenteritis outbreaks and C. difficile. The high rate of false-positive EIA samples emphasises the need for second confirmation tests to diagnose CDI.     

Outbreak of sapovirus infection among infants and children with acute gastroenteritis in Osaka City, Japan during 2004-2005

One hundred and twenty five fecal specimens were collected from sporadic cases of acute gastroenteritis in a pediatric clinic in Osaka City, Japan from July 2004 to June 2005 and tested for the presence of rotavirus, norovirus, sapovirus, astrovirus, and adenovirus by RT-multiplex PCR. Among diarrheal viruses detected, norovirus was the most prevalent (19.2%, 24 of 125), followed by group A rotavirus (18.4%, 23 of 125), astrovirus (1.6%, 2 of 125), and adenovirus (0.8%, 1 of 125), respectively. Interestingly, sapovirus infection was identified with high incidence of 17.6% (22 of 125). Sapovirus was subjected to molecular genetic analysis by sequencing. It was found that sapovirus detected in this study was classified into four genotypes (GI/1, GI/4, GI/6, and GI/8), and sapovirus GI/6 was predominant, followed by GI/8 and accounted for 81.8% (18 of 22) and 9.1% (2 of 22), respectively. It was noteworthy that sapovirus GI/6 infection was apparently confined within the period of 5 months (August 2004 through December 2004). This pattern of infection implied the outbreak of sapovirus GI/6 in these patients, which was the first outbreak of acute gastroenteritis attributed to sapovirus in Osaka City. Another interesting feature of the study was the appearance of two novel, naturally occurring recombinant sapoviruses circulating in the Japanese population studied. This report confirmed the presence as well as the importance of sapovirus causing acute gastroenteritis among Japanese infants and children.     

Pulsed-field gel electrophoresis analysis of nasopharyngeal flora in children attending a day care center

To investigate how bacterial pathogens spread from child to child in a day care center, we monitored six children, two boys and four girls, born between August 1995 and November 1997, attending a day care center and analyzed nasopharyngeal samples from them using pulsed-field gel electrophoresis (PFGE). We obtained nasopharyngeal cultures from all of the affected children and almost all of the unaffected children between September 1998 and March 1999 after some children presented simultaneously with purulent rhinorrhea. Moreover, when a child was found to have acute otitis media, nasopharyngeal secretions from the child were independently cultured during treatment. During this period, 28 isolates of Moraxella catarrhalis, 13 of Streptococcus pneumoniae, and 4 of Haemophilus influenzae were recovered. PFGE gave 8 patterns for M. catarrhalis, 10 for S. pneumoniae, and 1 for H. influenzae. PFGE patterns demonstrated spread of M. catarrhalis between children. However, each occurrence of clusters of infection with M. catarrhalis lasted 2 to 6 weeks, with a change in PFGE pattern between occurrences of clusters. The M. catarrhalis strain infecting each child also changed. Similarly, the S. pneumoniae strain in each child also changed. In contrast, infection with H. influenzae persisted for about 3 months in an affected child.     

Immunogenicity and efficacy of a killed hepatitis A vaccine in day care center children

The objective of this study was to characterize the immune response of children after the use of two different vaccine doses and to evaluate whether vaccination benefits children attending day care centers in areas with high anti-HAV seroprevalence. The study was conducted in a day care center with a stable population in São Paulo, Brazil. Two groups of 20 children, all seronegative for hepatitis A antibodies, were assigned randomly to receive three times 0.5 and 1.0 ml of the vaccine, the second and third dose 1 and 6 months after the first dose, respectively. There were 27 children in the control group. All children in both vaccinated groups had protective levels of antibodies in the serum after two inoculations, and serious adverse reactions were not observed. In the eighth month of follow-up, a hepatitis A outbreak occurred in the day care center. Five children in the control group had high titers of IgM class anti-HAV, four with clinical manifestations of acute hepatitis. None of the vaccinated children developed symptoms or signs of hepatitis (P = 0.0125), and the estimate of vaccine efficacy was 100%. Two nonstudy children from the center also had clinical and serological evidence of acute hepatitis A. It is concluded that vaccination represents an important method for prevention of hepatitis A transmission in day care centers. The results of this pilot study justify further testing in larger groups. © 1996 Wiley-Liss, Inc.     

Role of Human Bocavirus infections in outbreaks of gastroenteritis

BACKGROUND: Human Bocavirus (HBoV) is considered to be responsible for lower respiratory tract infections in small children. Recent publications also reported the detection of HBoV in stool samples of gastroenteritis patients. Therefore HBoV might be associated with gastroenteritis cases. OBJECTIVES: To investigate the prevalence and the causative role of HBoV in gastroenteritis outbreaks in day care facilities for children. STUDY DESIGN: We examined 307 stool samples using a real time PCR protocol for HBoV load. Samples were collected from 48 independent outbreaks of gastroenteritis in day care facilities. RESULTS: HBoV was detected in 14/307 stool samples (4.6%). HBoV load in the samples was low and the 14 HBoV positive samples were distributed among 13 different outbreaks. Coinfections with Norovirus in single samples were frequent (57.1%), but no gastroenteritis outbreak could be associated with HBoV infection or coinfection. CONCLUSIONS: This study indicates that HBoV is not a causative agent for gastroenteritis outbreaks.     

Outbreak of acute gastroenteritis associated with group A rotavirus and genogroup I sapovirus among adults in a mental health care facility in Japan

An outbreak of acute gastroenteritis consisting of 57 cases occurred in a mental health care facility in Takasaki city, Japan during 6th February and 27th March 2002. A total of 18 fecal specimens collected from 17 residents and one member of the medical staff during this outbreak were tested for the presence of viral enteropathogens by RT-PCR and latex agglutination. Group A rotavirus and sapovirus were detected in 5 out of 18 fecal specimens (55.6%). To our knowledge, this is the first finding of an outbreak of gastroenteritis associated with co-circulation of different kinds of viruses such as group A rotavirus and sapovirus. All of group A rotaviruses were typed further as P[4]G2 strains. Both rotavirus and sapovirus were subjected to molecular analysis by sequencing. It was noteworthy that all rotaviruses and sapoviruses had high homologies, respectively, to each other and sapoviruses presented a potential novel sapovirus genogroup I (GI) genotype, which was obviously different from any GI genotypes (GI-a, b, c, and d). The outbreak associated with these viruses spread gradually from dormitory to dormitory, suggesting a spread by person-to-person contact, although investigation on the route of transmission of the outbreak is lacking. The findings confirm the presence of group A rotavirus and sapovirus are important in acute gastroenteritis among adults in Japan.     

Two successive outbreaks of SRSV-associated gastroenteritis in South Africa

Two successive outbreaks of gastroenteritis in South Africa were investigated to identify the aetiological agents. Some patients were involved in both outbreaks. Enteropathogenic bacteria or parasites were not evident in either outbreak. Small round structured viruses (SRSVs) were demonstrated in both outbreaks by direct electron microscopy. SRSV UK3/Hawaii virus was identified by immune electron microscopy as the causative agent in the first outbreak. Using new recombinant Norwalk virus (rNV) immunoassays for antibodies and antigen, Norwalk virus was implicated in the second outbreak. Preexisting antibodies to Norwalk virus were not protective and there was no cross protection between Hawaii and Norwalk viruses. There was no anamnestic response to Norwalk virus following the SRSV UK3/Hawaii outbreak although those affected had preexisting antibodies to Norwalk virus. To our knowledge, this is the first definitive diagnosis of SRSV-associated gastroenteritis in South Africa.     

广东省暴发性胃肠炎中诺如病毒的分子流行病学特点分析

目的研究广东省暴发性胃肠炎中诺如病毒的分子流行病学特点。方法对2003年至2004年的13起急性胃肠炎患者的粪便和肛拭子标本,采用针对诺如病毒的特异引物和逆转录-聚合酶链反应(RT-PCR)技术进行检测,再经核苷酸序列测定分析病毒的基因型,同时收集患者相关流行病学资料。结果13起暴发性胃肠炎中有8起是由诺如病毒引起的,时间主要集中在每年的10月至12月,基因分析显示这些病毒都属基因Ⅱ组,但分属3个亚型,6株病毒属GⅡ-4型。结论诺如病毒是广东省暴发性非细菌性胃肠炎的重要病原体之一,具有分布广、流行时间集中在秋冬季的特点。引起暴发的病毒主要为基因Ⅱ组,在现阶段存在优势毒株。     

GII.17 norovirus infections in outbreaks of acute nonbacterial gastroenteritis in Osaka City,Japan during two decades

Norovirus (NoV) is a major cause of viral gastroenteritis, and GII.4 has been the predominant genotype worldwide since the mid-1990s. During the 2014 to 2015 winter, a rare genotype, NoV GII.17, emerged and became prevalent mainly in East Asia. Over the past two decades, NoV molecular surveillance in Osaka City, Japan, has revealed that NoV GII.17 was detected for the first time in February 2001 and that NoV GII.17-associated outbreaks remarkably increased during the 2014 to 2015 season, with higher incidence recorded in January to March 2015. Genetic analysis indicated that 28 GII.17 outbreak strains were closely related to the novel GII.P17-GII.17 variants represented by the Kawasaki308/2015/JP strain, similar to that in other regions. Statistical analysis showed that NoV GII.17 infections were more common in adults than GII.3 and GII.4 infections, suggesting that the affected adults most likely did not have antibodies against NoV GII.17 and the novel GII.17 variant had recently appeared. Regarding transmission, food was one of the most important factors involved in the spread of NoV GII.17 among adults; 61% of GII.17 outbreaks were foodborne, with oysters being the most common vehicle. Interplay between pathogens, hosts, and environmental factors was considered to be important in the 2014 to 2015 NoV GII.17 epidemic.     

Transition of genotypes associated with norovirus gastroenteritis outbreaks in a limited area of Japan, Hiroshima Prefecture, during eight epidemic seasons

The transition of genotypes implicated in 102 NoV gastroenteritis outbreaks in Hiroshima Prefecture, Japan, during eight epidemic seasons was investigated. Eighteen genotypes were implicated in the outbreaks, with the chronological characteristics as in GII.3, GII.4, GII.5 and GII.12. In GII.4 variants, amino acid changes and positively selected sites were of note and significantly concentrated in the surface-exposed P2 subdomain of the VP1 protein. Notably, variant-specific epitopes at which positively selected sites are located may be significant for distinguishing a new GII.4 variant. The interaction of these genetic changes with developing immunity seems to influence NoV epidemics.     

Astrovirus as a cause of gastroenteritis in Japan.

We used an enzyme immunoassay (EIA) to screen for astrovirus in stool specimens from outbreaks and sporadic cases of gastroenteritis collected between 1982 and 1992 in six prefectural public health institutes in Japan. Three outbreaks of gastroenteritis involving schoolchildren and adults were confirmed to be attributable to astrovirus. Astrovirus was detected in 6 to 10% of the specimens from patients with sporadic gastroenteritis from whom no other bacterial or viral agent had been identified. Among the sporadic cases, astrovirus was most frequently detected in infants less than 1 year of age, and the incidence peaked in March and April. Using specimens from recent outbreaks, we found that the EIA was more sensitive than electron microscopy (EM) for the detection of astrovirus, and many EM-negative specimens were positive by EIA. However, some stool specimens previously found to have astrovirus-like particles by EM were negative by EIA, perhaps because of inadequate storage conditions, such as long-term storage and repeated freezings and thawings. Our results indicate that astrovirus is more commonly associated with childhood gastroenteritis than has been previously appreciated and suggest that further studies to examine the epidemiology and disease burden of this virus are needed.