Intergenogroup Recombinant Sapovirus in Japan, 2007�C2008

We investigated the incidence of sapovirus (SaV)–associated gastroenteritis in infants and children in Japan during 2007–2008 and characterized the diversity of SaV-positive strains. SaV was detected in 19 (4%) of 477 fecal specimens. The leading genogroup (79%, 15 cases) comprised intergenogroup recombinant SaVs (GII/GIV).     

Repeated circulation over 6 years of intergenogroup mono-reassortant G2P[4] rotavirus strains with genotype N1 of the NSP2 gene

Human rotavirus A, a major cause of acute diarrhoea in infants and young children worldwide, contains the genome of 11 segments of double-stranded RNA, which is grouped into three major genotype constellations: the Wa genogroup (G1-P[8]-I1-R1-C1-M1-A1-N1-T1-E1-H1), the DS-1 genogroup (G2-P[4]-I2-R2-C2-M2-A2-N2-T2-E2-H2), and the AU-1 genogroup (G3-P[9]-I3-R3-C3-M3-A3-N3-T3-E3-H3). Recently, a G2P[4] strain detected in Kenya in 1982 was reported to be a nonstructural protein 2 (NSP2) gene mono-reassortant between the Wa and DS-1 genogroups. While NSP2 induces heterotypic antibody responses in children, thereby being the likely target of immune selective pressure, it was not known how frequently these NSP2 mono-reassortants circulated among children. In a previous 10 year epidemiological study, out of 100 G2P[4] rotavirus specimens that were typed into 22 distinct electropherotypes (i.e., strains), RNA-RNA hybridization identified that 12 strains were mono-reassortants involving either the NSP2 or the NSP3 gene. The aim of this study was therefore to determine the nucleotide sequences of all 11 genes of the representative mono-reassortant strain (AU605) and the sequences of the reassorted genes of the other mono-reassortant strains. The genome constellation of AU605 was G2-P[4]-I2-R2-C2-M2-A2-N1-T2-E2-H2, indicating that it was a mono-reassortant strain carrying a Wa-like NSP2 genotype on the DS-1 genogroup background. The reassorted gene of the other 11 strains also had the genotype N1 in the NSP2 gene. Given that electrophoretically-identical rotaviruses represent a single strain, the 12 NSP2 mono-reassortants detected in six rotavirus seasons accounted for 36% (36 of 100) of G2P[4] rotavirus specimens in the 10 year collection. The circulation of NSP2 mono-reassortant strains was observed in epidemic seasons when the G2P[4] genotype was not dominant. Taken together with their repeated occurrence at higher-than-expected frequencies, the identification of NSP2 mono-reassortant G2P[4] strains suggests that such genome constellation may also be viable in nature, and that they circulated less rarely than generally believed.     

Characteristics of the mosaic genome of a human parechovirus type 1 strain isolated from an infant with pneumonia in China

Human parechoviruses (HPeVs) belong to the Parechovirus genus of the large and growing family of Picornaviridae with a non-enveloped, single-stranded and positive-sense RNA. An HPeV strain was isolated from the nasopharyngeal aspirate specimen of a 2 months old infant hospitalized with pneumonia in Beijing, China and nominated as BJ-37359 followed the code of the specimen. Strain BJ-37359 was identified as HPeV1 by whole genome sequencing. The full genome of strain BJ-37359 consisted of 7336 nucleotides (nt), excluding a poly (A) tail and contained an ORF of 6537 nt flanked by 5′UTR of 709 nt and 3′UTR of 90 nt. Phylogenetic analyses revealed that strain BJ-37359 were clustered together with HPeV1 strains in the structural capsid protein region, while uncoupling in the non-structural gene regions. Analyses with Simplot and Bootscan indicated that multiple recombination events occurred in the non-structural region and VP0 region of strain BJ-37359 with other HPeV1, and other types might have contributed to the recombination, especially HPeV6 and HPeV7 strains. Recombination analyses indicated that strain BJ-37359 may have a mosaic genome with new genomic recombination breakpoints.     

Detection and molecular characterization of sapoviruses in dogs

Caliciviruses are important human and animal pathogens. Novel caliciviruses have been identified recently in dogs, raising questions about their pathogenic role and concerns regarding their zoonotic potential. By screening stool samples of young or juvenile dogs using RT-PCR assays, sapoviruses (SaVs) were found in 7/320 (2.2%) samples of animals with acute gastroenteritis while they were not detected in healthy animals (0/119). The sequence of a nearly 3 kb portion at the 3′ end of the genome, encompassing the RNA-dependent RNA polymerase (RdRp), the capsid region (ORF1) and the ORF2 were determined for three strains. A distinctive genetic feature in canine SaVs was a 4-nucleotide (nt) interval between the ORF1 and ORF2. Two strains (Bari/4076/07/ITA and Bari/253/07/ITA) were very closely related in the RdRp and capsid regions to the strain AN210D/09/USA (90.4–93.9% nt), while strain Bari/5020/07/ITA displayed only 71.0–72.0% nt identity to this group of canine SaVs and 76.0% to strain AN196/09/USA. Overall, these findings indicate that the canine SaVs detected in Italy may represent distinct capsid types, although all currently known SaVs segregate into the novel proposed genogroup, tentatively named as GXIII.     

Genome characteristics and molecular evolution of the human sapovirus variant GII.8

Human sapovirus is regarded as an important viral agent for acute diarrhea worldwide. GII.8, a recently reported genotype, has been detected in a few countries and regions. In this study, we obtained the first genome sequence of a sapovirus GII.8 strain isolated in mainland China, and comprehensively analyzed the genetic diversity and evolutionary process of this genotype. The viral genome of the new GII.8 Guangzhou strain GZ2014-L231 comprised 7433 nucleotides, including two ORFs. Pairwise alignments of the new genome with representative sequences of different genotypes showed inconsistent homology between different protein-encoding regions, of which NS1 and VP2 were found as the variable proteins, and NS3, NS5, and NS6/7 were found as the conserved ones. Compared with other reported GII.8 genomes, the Guangzhou strain introduced 34 new nucleotide changes and one new amino acid change. Phylogenetic analysis based on full-length VP1 sequences demonstrated that 11 GII.8 strains could be divided into 4 clusters A-D, with 88 SNP and 10 SAP spots occurred during their evolutionary process. The Guangzhou strain has higher homology with seven GII.8 strain detected after 2014, especially the US and Peruvian strains of 2015/2016, which have the identical VP1 amino acid sequences. Using a Bayesian coalescent method based on VP1 sequences, GII.8 was predicted to emerge in 2001 with the evolution rate of 1.45 × 10⁻³ nucleotide substitutions/site/year (strict clock). In summary, the data in this study not only provided reference data from mainland China for sapovirus researches in future, but also firstly described the evolutionary process of the GII.8 genotype.     

Emergence of intragenotype recombinant sapovirus in Japan.

Sapovirus is an important causative agent of sporadic cases as well as of outbreaks of acute gastroenteritis in humans worldwide. A total of 603 fecal specimens collected from July 2005 to June 2006 from children with acute gastroenteritis in five localities in Japan (Maizuru, Tokyo, Sapporo, Saga, and Osaka) were screened for sapovirus by RT-PCR. It was found that 17 specimens were positive for sapovirus and it represented 2.8%. Interestingly, intragenotype recombinant sapovirus GI/1 emerged with 76.4% (13 of 17) and rapidly became the leading cause of acute gastroenteritis in Japan for the first time. The lower frequency of sapovirus GI/2 and GI/4 (each of 11.8%), which were the second prevailing genotypes, was also detected. A novel nomenclature of sapovirus was proposed, in which worldwide sapovirus strains were classified into seven genogroups. Of these, novel sapovirus genogroups VI and VII demonstrated the very low homologies, only 32.8-41.6% at the amino acid level and 43.6-49.9% at the nucleotide level, to those of sapovirus genogroups I-V. Of note, two distinct clusters of sapovirus were co-circulating in porcine. Interestingly, the worldwide sapovirus strains shared the 25 nucleotide-conserved region, covering the polymerase-capsid junction which differed according to each species due to multiple nucleotide substitutions. The finding suggests that the sapovirus recombination between human and animal hardly takes place in nature. This is also the first, to our best knowledge, demonstrating the emergence of the intragenotype recombinant sapovirus with its causing diarrheal illness in Japan.     

Phylogenetic analyses of Norovirus strains detected in Uruguay reveal the circulation of the novel GII.P7/GII.6 recombinant variant

Noroviruses (NoV) are one of the major etiological agent of acute gastroenteritis (AGE) outbreaks worldwide. Distinct NoV genotypes have been associated with different transmission patterns and disease severity in humans. Therefore, it is important to identify genetically different NoV genotypes circulating in a particular region. However, genotyping has become a challenge due to recombination events occurring mainly nearby ORF1/ORF2 junction of NoV genome, leading to distinct genotypes with polymerase and capsid regions derived from parenteral strains. Taking this into account, ORF1/ORF2 sequences were obtained from NoV strains collected from patients with AGE in Uruguay. This study reveals in silico evidences of recombination events taking place in four out of six strains analyzed for which its polymerase gene and its capsid region correspond to GII.P7 and to GII.6 genotype, respectively. These results also reveal the circulation of a GII.P7/GII.6 recombinant variant in the natural populations of NoV strains in the northwestern region of Uruguay. As far as we know this is the first report about the circulation of a NoV GII.P7/GII.6 recombinant variant in the Americas.     

The origin of two rare human P[10] rotavirus strains

The Group A rotavirus (RVA) P[10] is a rare genotype of the RVA VP4 gene. To date, the whole genome sequence of only a single P[10] RVA strain, RVA/Human-tc/IDN/69M/1980/G8P4[10], has been determined, revealing a DS-1-like genotype constellation. Whole genomic analyses of P[10] RVA strains with other VP7 genotypes are essential to obtain conclusive data on the origin and genetic diversity of the P10] RVAs. In the present study, the whole genome of a human G4P[10] RVA strain, RVA/Human-tc/IDN/57M/1980/G4P[10], was analyzed. Strain 57M exhibited an unusual G4-P[10]-I1-R1-C1-M1-A1-N1-T2-E1-H2 genotype constellation, and was found to originate from intergenogroup reassortment events involving acquisition of RVA strain 69M-like VP4, NSP3 and NSP5 genes by a co-circulating Wa-like human G4 RVA strain. Although the reference P[10] strain, 69M, exhibits a DS-1-like genotype constellation, the exact origin of this RVA remains to be elucidated. By detailed phylogenetic analyses, we found that the VP1-VP3, VP6, NSP2 and NSP4 genes of 69M originated from artiodactyl and/or artiodactyl-like human P[14] strains, whilst its NSP1, NSP3 and NSP5 genes were more related to those of typical human DS-1-like strains than those of other RVAs. On the other hand, the origin of the VP4 gene of 69M could not be established. Nevertheless, these observations clearly indicated that strain 69M might have originated from reassortment events involving at least the artiodactyl or artiodactyl-like human RVAs and the typical human DS-1-like strains. The present study provided rare evidence for intergenogroup reassortment events involving co-circulating typical human Wa-like RVAs and unusual RVAs of the DS-1-like genogroup, and revealed the presence of artiodactyl-like genes in a human P[10] strain, highlighting the complex evolutionary patterns of the P[10] RVAs.     

Novel recombinant sapovirus

We determined the complete genome sequences of two sapovirus strains isolated in Thailand and Japan. One of these strains represented a novel, naturally occurring recombinant sapovirus. Evidence suggested the recombination site was at the polymerase-capsid junction within open reading frame one.     

A novel feline norovirus in diarrheic cats

By screening a collection of fecal samples from young cats housed in three different shelters in South Italy, noroviruses (NoVs) were found in 3/48 (6.2%) specimens of animals with enteritis signs while they were not detected in samples collected from healthy cats (0/57). Upon sequence analysis of the short RNA-dependent RNA polymerase (RdRp) region, the three strains displayed the highest nucleotide (nt) and amino acid (aa) identities to the prototype GIV.2 strain lion/Pistoia/387/06/ITA (91.0–93.0% nt and 97.0–98.0% aa). The sequence of ~ 3.4-kb portion at the 3′ end of the genome of a NoV strain, TE/77-13/ITA, was determined. In the full-length ORF2, encoding the VP1 capsid protein, the virus was genetically closest to the canine GVI.2 NoV strains C33/Viseu/2007/PRT and FD53/2007/ITA (81.0–84.0% nt and 93.0–94.0% aa identities), suggesting a recombination nature, with the cross-over site being mapped to the ORF1-ORF2 junction. Based on the full-length VP1 amino acid sequence, we classified the novel feline NoV, together with the canine strains Viseu and FD53, as a genotype 2, within the genogroup GVI. These findings indicate that, as observed for GIV NoV, GVI strains may infect both the canine and feline host. Unrestricted circulation of NoV strains in small carnivores may provide the basis for quick genetic diversification of these viruses by recombination. Interspecies circulation of NoVs in pets must also be considered when facing outbreaks of enteric diseases in these animals.     

Complete nucleotide sequence analysis of the norovirus GII.17: A newly emerging and dominant variant in China, 2015

Norovirus is an important pathogen which accounts for majority of the viral related acute gastroenteritis. Recently, a variant of genotype GII.17 was reported to be predominant over GII.4 and accounted for several acute gastroenteritis outbreaks in Asia. In the current study, the full genome of a norovirus strain ZHITHC-12 isolated during this outbreak period in China was identified and characterized. The viral genome was 7557 nucleotides in length and a phylogenetic analysis based on full length genome sequences indicated that ZHITHC-12 belonged to GII.17 genotype. A further phylogenetic analysis based on all available polymerase and capsid sequences showed that ZHITHC-12 was in Cluster III on both phylogenetic trees and grouped with other strains also isolated during 2013 to 2015. Moreover, homology modeling analysis based on GII norovirus capsid 5BSX template revealed that substitutions, mutations, and more importantly, deletions and insertions, occurred at or near the putative epitopes and histo-blood group antigen (HBGA) binding sites in its protruding P2 domain, which might confer new antigenic or biological properties for this novel variant. In summary, the first full genome and capsid protein structure of a novel norovirus GII.17 variant isolated in China was extensively characterized. The data would be helpful not only for the epidemiology study, but also for the diagnostic tool development and effective vaccine design in the future.     

Novel intergenotype human norovirus recombinant GII.16/GII.3 in Bangladesh

Noroviruses (NoVs) are one of the major etiological agents of acute gastroenteritis in all age groups. In this study, we identified an intergenotype NoV recombinant strain in the fecal specimens of two male infants with acute diarrhea in Bangladesh. Phylogenetic analysis showed that the identified strains were recombinant NoV strains with a GII.3 capsid and a GII.16 polymerase gene. The recombination breakpoint was located in the ORF1/ORF2 overlap region. To the best of our knowledge this is the first report of a NoV recombinant GII.16/GII.3 strain worldwide.     

Analysis of uncommon norovirus recombinants from Manaus,Amazon region,Brazil: GII.P22/GII.5, GII.P7/GII.6 and GII.Pg/GII.1

Norovirus (NoV) is responsible for outbreaks and sporadic cases of nonbacterial acute gastroenteritis in humans worldwide. The virus consists of small round particles containing a single-stranded RNA genome that is divided into three Open Reading Frames. NoV evolves via mechanisms of antigenic drift and recombination, which lead to the emergence of new strains that are capable of causing global epidemics. Recombination usually occurs in the ORF1/ORF2 overlapping region and generates strains with different genotypes in the polymerase and capsid region. The primary objective of this study was to analyze recombination in positive-NoV samples. Specimens were collected during 2011, 2012 and 2014, from children under two years of age presenting gastrointestinal symptoms such as vomiting and diarrhea. The partial polymerase (B region), capsid (D region) genes and the ORF1-ORF2 overlap regions were sequenced in each sample. The recombinant analyses were performed in the Simplot software v.3.5.1 and RDP4 Beta v. 4.6 program. These analyses showed that GII.Pg/GII.1, GII.P7/GII.6, and GII.P22/GII.5 were recombinant strains. To our knowledge, this is the first time that the GII.P22/GII.5 and GII.Pg/GII.1 strains were described in South America and the GII.P7/GII.6 was detected in Northern of Brazil.     

Sapovirus outbreaks in long-term care facilities, Oregon and Minnesota, USA, 2002-2009

We tested fecal samples from 93 norovirus-negative gastroenteritis outbreaks; 21 outbreaks were caused by sapovirus. Of these, 71% were caused by sapovirus genogroup IV and 66% occurred in long-term care facilities. Future investigation of gastroenteritis outbreaks should include multi-organism testing.     

Molecular analysis of echovirus 13 isolates and aseptic meningitis,Spain

Echovirus 13 (EV13), considered rare, was reported worldwide in 2000, mostly related to aseptic meningitis outbreaks. In Spain, 135 EV13 isolates were identified. The genetic relationships between 64 representative strains from Spain and other reported isolates from the United States, Germany, Italy, Japan, and Sweden were described by analyzing the partial sequence of the major capsid protein (VP1) gene. The strains from Spain were clearly identified as EV13 (79.5% similarity with the EV13 reference strain) and were grouped phylogenetically into two different clusters (by origination on either the Iberian Peninsula or Canary Islands). Isolates from Germany from 2000 clustered with the Canary Islands group. The isolates from other countries obtained before 2000 were genetically distant. Changes in EV13 coding sequence involved several differences in the C-terminal extreme of the VP1 protein. Part of the neutralizing antigenic site III has been described in this genome region in poliovirus and swine vesicular disease virus.     

Molecular evolution and epidemiology of echovirus 6 in Finland

Echovirus 6 (E-6) (family Picornaviridae, genus Enterovirus) is one of the most commonly detected enteroviruses worldwide. The aim of this study was to determine molecular evolutionary and epidemiologic patterns of E-6. A complete genome of one E-6 strain and the partial VP1 coding regions of 169 strains were sequenced and analyzed along with sequences retrieved from the GenBank. The complete genome sequence analysis suggested complex recombination history for the Finnish E-6 strain. In VP1 region, the phylogenetic analysis suggested three major clusters that were further divided to several subclusters. The evolution of VP1 coding region was dominated by negative selection suggesting that the phylogeny of E-6 VP1 gene is predominantly a result of synonymous substitutions (i.e. neutral genetic drift). The partial VP1 sequence analysis suggested wide geographical distribution for some E-6 lineages. In Finland, multiple different E-6 lineages have circulated at the same time.     

一起急性胃肠炎疫情中重组诺如病毒基因特征分析

目的 明确成都市2018年5月某建筑工地发生的一起急性胃肠炎疫情中的诺如病毒的基因型别,并对其遗传进化和分子流行病学特征进行分析研究。方法 对实时荧光定量PCR检测结果为阳性的标本采用常规RT - PCR扩增其RNA依赖的RNA聚合酶区和衣壳蛋白区并测序,与国内外参考株进行序列比对和构建进化树等基因特征分析。结果 序列比对和进化树分析结果显示:此次疫情中检出的诺如病毒核苷酸同源性为100%;其RNA依赖的RNA聚合酶区与北京株处于同一分支,同源性95.4%;而衣壳蛋白区与莫斯科株同源性最高,为96.4%。结合SimPlot对其进行分析,重组可能位点在213 bp处,定位在ORF1/2交叠区。结论 引起本次急性胃肠炎疫情的重组诺如病毒基因型为GI.P4 - GI.5,这是成都市首次检出诺如病毒GI群重组株。