Changing distribution of norovirus genotypes and genetic analysis of recombinant GIIb among infants and children with diarrhea in Japan

A total of 402 fecal specimens collected during July 2003-June 2004 from infants and children with acute gastroenteritis, encompassing five localities (Maizuru, Tokyo, Sapporo, Saga, and Osaka) of Japan, were tested for the presence of norovirus by RT-PCR. It was found that 58 (14.4%) fecal specimens were positive for norovirus. Norovirus infection was detected throughout the year with the highest prevalence in December. Norovirus GII was the most predominant genogroup (98.3%; 57 of 58). The genotypes detected in this study were GI/4, GII/2, GII/3, GII/4, and GII/6. Of these, NoV GII/3 (known as the Arg320 virus cluster) was the most predominant genotype (43.9%), followed by NoV GII/4 (the Lordsdale virus cluster; 35.1%) and others. Two norovirus strains clustered with a "new variant designated GIIb" and a "new variant of GII/4" were found circulating in Japan for the first time. It was interesting to note that NoV GIIb and NoV GII/3 appeared to be the recombinant strains and the recombination site was demonstrated at the overlap of ORF1 and ORF2. The majority (96%) of the dominant norovirus strains were identified as the recombination of GII/3 capsid and GII/12 polymerase. The recombination in the NoV GIIb capsid gene at the breakpoint located at P1 domain was also identified. Obviously, NoV GIIb isolate in Japan had double recombination. This is the first report demonstrating the existence of different "new variants" co-circulating in Japanese infants and children with acute gastroenteritis.     

Molecular epidemiology of enteric viruses in children with sporadic gastroenteritis in Valencia, Venezuela

The epidemiology and clinical symptoms in infants and young children with acute sporadic viral gastroenteritis due to viral etiologies other than rotaviruses have not been studied thoroughly in developing countries. Fecal specimens from 480 children <5 years of age who were admitted to a large children's hospital in the city of Valencia, Venezuela, with acute diarrhea during January to December 2003 were collected and screened by ELISA and RT-PCR for rotavirus, adenovirus, norovirus, sapovirus, and astrovirus. Viral isolates were partially characterized by phylogenetic analysis. Norovirus viral load was determined by qRT-PCR. Viruses were identified in 205 (43%) of the 480 stool samples collected. Rotavirus was the virus detected most frequently (21%), followed by norovirus (13%), adenovirus (5%), sapovirus (3%), and astrovirus (2%). Viral infection rates were highest in the 6- to 11-month-old group (49%) and lowest in children >24 months old. Norovirus GII was more prevalent (90%) than GI (10%). Enteric adenovirus (serotypes 40/41) was present in 43% of the adenovirus-positive samples. Rotavirus infection caused more severe clinical symptoms than the other viruses detected, with more vomiting (84%) and dehydration (11%) that led to hospital admission of 20% of the children with acute gastroenteritis. Rotavirus and norovirus showed marked and opposite seasonal patterns. No association was observed between disease severity and viral load in children infected with norovirus. These results not only confirm the impact of rotavirus infection in Venezuela but also indicate that other enteric viruses, especially noroviruses, contribute significantly to sporadic acute gastroenteritis and to the burden of disease.     

A single-tube multiplex PCR for rapid detection in feces of 10 viruses causing diarrhea

A novel multiplex polymerase chain reaction assay was developed to identify 10 viruses in a single tube. The assay was targeted to detect group A and C rotaviruses, adenovirus, norovirus GI, norovirus GII, sapovirus, astrovirus, Aichi virus, parechovirus, and enterovirus. A total of 235 stool samples were collected from infants and children with acute gastroenteritis in Kyoto, Japan, from 2008 to 2009, then tested by this novel multiplex PCR and compared with a multiplex PCR described previously, which used 3 primer sets. The novel multiplex PCR could detect the targeted viruses in 111 of the 235 (47.2%) stool samples. Of these, 9 out of 10 types of viruses were identified, including group A rotavirus, norovirus GII, enterovirus, sapovirus, adenovirus, parechovirus, group C rotavirus, astrovirus, and norovirus GI. In contrast, the multiplex PCR that used 3 sets of primers could detect the targeted viruses in 109 of the 235 (46.4%) stool samples. Among these, 8 types of viruses were identified, including group A rotavirus, norovirus GII, enterovirus, adenovirus, parechovirus, group C rotavirus, sapovirus, and astrovirus. The results suggested that the new multiplex PCR is useful as a rapid and cost effective diagnostic tool for the detection of major pathogenic viruses causing diarrhea.     

Detection and molecular characterisation of rotavirus and norovirus infections in Jordanian children with acute gastroenteritis

Rotavirus and norovirus are globally important causes of paediatric gastroenteritis, but no studies of viral genotypes have been reported from Jordan. We undertook a molecular epidemiological study in children hospitalised with acute gastroenteritis in Jordan between January 2006 and December 2007. Among 368 children, rotavirus and norovirus infections were detected in 49.5% and 11.4% of children, respectively. Rotavirus genotypes P[8],G1 (56%), P[4],G2 (14%) and P[8],G9 (13%) were most commonly identified, consistent with results of global rotavirus surveillance studies. Norovirus GII.3 was the most commonly detected genotype, followed by GII.4, contrasting with most studies in which GII.4 has predominated.     

Association between Norovirus and Rotavirus Infection and Histo-Blood Group Antigen Types in Vietnamese Children

Norovirus and rotavirus are the two most important causes of acute gastroenteritis in children worldwide. Both norovirus and rotavirus recognize human histo-blood group antigens (HBGAs), and multiple binding patterns for HBGAs have been reported. To explore the role of HBGAs in host susceptibility to norovirus and rotavirus, we conducted a cross-sectional study in children hospitalized with diarrhea in northern Vietnam from September 2010 through September 2012. Of 260 children with paired stool and saliva samples, 158 (61%) were classified as HBGA secretors (Le^a−b+), 31 (12%) were nonsecretors (Le^a+b−), and 71 (27%) were partial secretors (Le^a+b+). Norovirus was detected in 50 patients (19%), with viral genotypes GII.3 (n = 28) and GII.4 (n = 22) being the most common. All children infected with norovirus strains of genotype GII.4 were either HBGA secretors or partial secretors. Of the 28 GII.3 cases, 12 involved HBGA secretors, 11 partial secretors, and 5 nonsecretors. A total of 85 children tested positive for rotavirus, 74 of whom were infected with genotype P[8], 5 with P[4], and 6 with P[6]; all were HBGA secretors or partial secretors. This is the first epidemiological study demonstrating in a population that HBGA phenotype is a key susceptibility factor for both norovirus and rotavirus infections in children.     

Molecular epidemiology of noroviruses associated with acute sporadic gastroenteritis in children: Global distribution of genogroups,genotypes and GII.4 variants

Noroviruses are a leading cause of epidemic and sporadic acute gastroenteritis worldwide. The development of sensitive molecular diagnostic techniques has revolutionized our understanding of norovirus epidemiology over the past two decades, but norovirus strain types associated with sporadic gastroenteritis remain poorly described. Therefore, we conducted a systematic review of studies performed after 2000 to clarify the genotypic distribution of noroviruses in children (≤18 years of age) with sporadic acute gastroenteritis. Genogroup GII norovirus was the most prevalent, accounting for 96% of all sporadic infections. GII.4 was the most prevalent genotype, accounting for 70% of the capsid genotypes and 60% of the polymerase genotypes, followed by the capsid genotype GII.3 (16%) and the polymerase genotype GII.b (14%). The most common ORF1/ORF2 inter-genotype recombinants were GII.b, GII.12, and GII.4 polymerase genotypes combined with the capsid genotype GII.3, accounting for 19% of all genotyped strains. The distribution of GII.4 variants over the last decade was dominated by successive circulation of GII.4/2002, GII.4/2004, GII.4/2006b, and GII.4/2008 with GII.4/2006b continuing to date. Genotypes GII.4 and GII.3 have predominated in children during the past decade; this is most notable in the global emergence of GII.4 variant noroviruses. As the burden of rotavirus disease decreases following the introduction of childhood immunization programs, the relative importance of norovirus in the etiology of acute childhood gastroenteritis will likely increase. In order for a successful norovirus vaccine to be developed, it should provide immunity against strains with capsid genotypes GII.4 and GII.3.     

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.     

Molecular and epidemiological features of GIIb norovirus outbreaks in Victoria,Australia, 2002–2005

The epidemiology of GIIb norovirus outbreaks and the characteristics of GIIb open reading frame (ORF) 2 recombinant forms are poorly understood and this study examined these questions using norovirus‐associated gastroenteritis outbreaks in Victoria, Australia, during 2002–2005. Twenty‐one GIIb outbreaks were detected and were the second most common ORF 1 norovirus outbreak genotype (5%) after GII.4 (90%). Both GIIb and GII.4 outbreaks peaked in warmer months of the year but their periodicity was different. ORF 2 sequencing analysis was carried out in the two regions previously designated C and D. RT‐PCR region D primers were less sensitive than region C primers. No evidence of recombination between regions C and D was found. ORF 2 genotypes for the 21 GIIb outbreaks were: GII.1 (10 outbreaks), GII.3 (10 outbreaks) and, apparently for the first time, GII.13 (1 outbreak). GIIb outbreaks could occur in a broad range of settings and there was no correlation between ORF 2 genotype and setting except that all 5 outbreaks involving mainly young children were associated with GIIb/GII.3. J. Med. Virol. 81:1652–1660, 2009. © 2009 Wiley‐Liss, Inc.     

Molecular epidemiology of norovirus infection among children with acute gastroenteritis in Shanghai,China, 2001–2005

Norovirus is one of the major causes of outbreaks and sporadic cases of acute gastroenteritis in young children worldwide. Obtaining local baseline information regarding this virus is important for developing and evaluating prevention strategies of norovirus transmission in children. The age, seasonal distribution and circulating genotypes of norovirus in Shanghai, China, between 2001 and 2005 were determined. Of 5411 stool specimens collected from children under 5 years of age who were hospitalized with acute gastroenteritis 3,975 were rotavirus‐negative, indicating the presence of another causative agent. From these specimens, 484 were selected at random for genotyping, and 45 were norovirus‐positive. Norovirus infection was detected in all age groups, but infants less than 6 months old showed the lowest prevalence (5.4%). Norovirus infections peaked from August to November. Among the 37 identified norovirus strains, 2 were GII‐3, 2 were GII‐7, and 33 were GII‐4 genotypes. This study demonstrated the impact of norovirus infection causing acute gastroenteritis in hospitalized children and the importance of vaccination against norovirus diarrhea in Shanghai, China. J. Med. Virol. 81:1826–1830, 2009. © 2009 Wiley‐Liss, Inc.     

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.     

Genetic relatedness of noroviruses identified in sporadic gastroenteritis in children and gastroenteritis outbreaks in northern Alberta

We compared the proportion and genotype distribution of norovirus (NoV) identified in sporadic acute gastroenteritis in children younger than 7 years old with the NoV strains found in outbreaks from January 2003 through April 2004 in northern Alberta, Canada. Eight genogroup I (GI) and 133 GII NoV cases were detected in 1,166 cases of acute sporadic childhood gastroenteritis with a monthly detection rates varying from 6.0% to 20.4% and no sporadic gastroenteritis case in October 2003. Seventy-eight outbreaks (65%) tested positive for NoV during the study period with an obvious winter predominance and no NoV outbreaks in August, September, and October 2003. Three GI and 51 GII strains from the sporadic childhood gastroenteritis cases and seven GI and 37 GII strains from gastroenteritis outbreaks were sequenced and analyzed. Strains belonging to the GII.4 cluster predominated in outbreaks (68%) while the strains in sporadic childhood gastroenteritis demonstrated significant heterogeneity with the majority belonging to the GII.3 cluster (36%). Further analysis of NoV strains from 34 sporadic childhood gastroenteritis cases and 38 gastroenteritis outbreaks in chronologically and geographically related groups failed to demonstrate clear link between strains circulating in the setting of sporadic childhood gastroenteritis and those found in outbreaks.     

Norovirus genotypes causing gastroenteritis outbreaks in Finland 1998-2002.

BACKGROUND: Outbreak investigation methods for enteric viruses were improved in 1990s when gene amplification techniques were established in viral laboratories. OBJECTIVES: The objective of the study was to determine the causative agents for Finnish viral gastroenteritis outbreaks. Our aim was also to further characterise the norovirus strains, reveal the temporal occurrence of norovirus (NV) genotypes and to study some epidemiological aspects concerning the outbreaks. STUDY DESIGN: A total of 416 Finnish viral gastroenteritis outbreaks that occurred during 5 years (1998-2002), excluding those among hospitalised children, were investigated for enteric viruses. Stool samples were screened by electron microscopy as well as analyzed by specific noro- and astrovirus RT-PCR tests. Amplicon sequence analysis was used to find out norovirus genotypes. RESULTS: Noroviruses caused 252 (60.6%) of the outbreaks; other viruses, astro- or rotavirus, caused four epidemics. Norovirus epidemics occurred in all kinds of settings, most often in hospitals (30.6%) and in restaurants and canteens (14.3%). Both NV genogroups were found every year, but NV GGII outbreaks always outnumbered those of GGI. All but one outbreak at hospitals and nursing homes were of genotype GII. Polymerase sequence analysis revealed a variety of NV genotypes; six GI and at least eight GII genotypes. The GI.3 Birmingham-like and GII.4 Bristol-like genotype appeared every year, whereas the other types were circulating for shorter periods or sporadically. During the study period the genotypes GII.4 (Bristol), GII.1 (Hawaii), an emerging genotype GIIb, and a new variant of GII.4 predominated in that order. Indication for rapid genetic changes in the genotype GII.4 was also noticed. CONCLUSIONS: Noroviruses were the most prevalent causative agents in the outbreaks. Many NV genotypes were circulating, and a shift in the predominant genotypes was evident between epidemic seasons.     

Molecular epidemiology of norovirus gastroenteritis outbreaks in New Zealand from 2002-2009

Noroviruses are the most common cause of acute non-bacterial gastroenteritis outbreaks worldwide, including New Zealand. New Zealand has a population of 4.4 million, which allows for centralized outbreak surveillance and a Norovirus Reference Laboratory, which facilitates efficient diagnosis, surveillance, and tracking of norovirus outbreaks. Norovirus outbreak strains are identified, sequenced, and compared with international reference strains. Between January 2002 and December 2009, 1,206 laboratory-confirmed norovirus outbreaks were recorded. The predominant outbreak settings were healthcare institutions for the elderly and acute care patients. Other outbreak settings included catering establishments, cruise ships, homes, community events, school camps, child-related settings, and consumption of contaminated shellfish. Of the 1,206 outbreaks, 105 (8.7%) were caused by norovirus genogroup I (GI) strains, 1,085 (89.9%) were caused by genogroup II (GII) strains, and both GI and GII strains were detected in 9 (0.8%) outbreaks. The genogroup was not identified in 7 (0.6%) outbreaks. A range of norovirus genotypes, including GI genotypes 1-6, GII genotypes 2-8, and GII.12, were associated with these outbreaks. The predominant genotype was GII.4, which was identified in 825 (68.4%) outbreaks. Norovirus GII.4 variant strains, including 2002 (Farmington Hills), 2004 (Hunter), 2006a (Laurens, Yerseke), 2006b (Minerva), and 2010 (New Orleans) implicated in overseas outbreaks also occurred in New Zealand, providing evidence of global spread.     

Molecular epidemiology of norovirus infections in sporadic cases of viral gastroenteritis among children in Northern Italy

Surveillance of norovirus infections in sporadic cases of pediatric gastroenteritis admitted to a main hospital in Northern Italy during a full-year period (2002) showed that noroviruses (10.4%) were the second most common causative viral agent, following rotaviruses (21.1%), and noroviruses (81%) were mostly implicated in mixed infections. The epidemic period of norovirus was September-December, with September and November as months of major prevalence (33.3 and 38.5%, respectively). Six distinct norovirus genotypes were detected (GI.7, GII.1, GII.2, GII.4, GII.7, GII, not assigned named GIIb), and the predominant genotype was GII.4. A "new GII.4 2002 variant" accounted for 82.9% of total strains. Since the severity of norovirus symptoms does not usually require admission to hospital, the burden of norovirus disease in the general children population may be much higher than that suggested by the present hospital-based investigation.