Intra-genotypic Recombination Research Articles

Full genome sequence analysis of the predominant and uncommon G9P[4] rotavirus strains circulating in Tehran, Iran, 2021–2022: Evidence for inter and intra-genotype recombination

Group A rotaviruses (RVAs) are a major cause of acute gastroenteritis in children under 5 years of age worldwide. Herein, the genetic sequences of 11 RNA segments from three uncommon G9P[4] RVA strains found in the stool samples of children under 5 years of age in Iran were analyzed using next-generation sequencing (NGS) technology. The genomic constellations of these three uncommon G9P[4] strains indicated the presence of the double and quadruple reassortants of two G9P[4] strains, containing the VP7/NSP2 and VP7/VP2/NSP2/NSP4 genes on a DS-1-like genetic background, respectively. The genome of one strain indicated a Wa-like genetic backbone in a single-reassortant with the VP4 of the DS1-like human strains. With the exception of VP1, VP2, VP7, NSP2, NSP3, and NSP4 genes, which clustered with RVA of human origins belonging to cognate gene sequences of genogroup 1/2 genotypes/lineages, the remaining five genes (VP8/VP4, VP3, VP6, NSP1, NSP5) displayed direct evidence of recombination. It is presumed that the presence of uncommon G9P[4] strains in Iran is not linked to vaccination pressure, but rather to the high prevalence of RVA co-infection or the direct import of these uncommon RVA reassortants strains from other countries (especially those that have implemented RV vaccination).

Genetic characterization of duck hepatitis B viruses from Anhui Province, China.

Duck hepatitis B virus (DHBV) infection model was frequently used as the experimental model for human hepatitis B virus (HBV) research. In order to decipher the genetic characteristics of DHBVs from Anhui province of China, 120 duck liver tissue samples were collected and subjected to PCR screening, and 28 samples were detected as DHBV positive. Subsequently, five DHBV-positive samples were selected for genome-wide amplification and a comprehensive analysis. Comparative analysis of complete genome sequences using the MegAlign program showed that five strains of DHBVs shared 94.5-96.3% with each other and 93.2-98.7% with other reference strains in GenBank. The phylogenetic analysis showed that all five DHBV strains belonged to the evolutionary branch of "Chinese DHBV" isolates or DHBV-2. Importantly, three potential intra-genotypic recombination events, between strains AAU-6 and Guilin, strains AAU-1 and GD3, and strains AAU-6 and AAU-1, were respectively found using the RDP and SimPlot softwares and considered the first report in avihepadnaviruses. These results not only improve our understanding for molecular prevalence status of DHBV among ducks, but also provide a reference for recombination mechanism of HBV.

Diversity of Astrovirus in Goats in Southwest China and Identification of Two Novel Caprine Astroviruses.

ABSTRACTA total of 232 goat fecal samples (124 diarrheic and 108 nondiarrheic) collected from 12 farms in Southwest China were tested for astrovirus using RT-PCR. A total of 16.9% (21/124) of diarrheic and 20.4% (22/108) of nondiarrheic samples were astrovirus-positive, and no statistical difference was found in the detection rate between healthy and sick goats. Furthermore, 28 obtained complete ORF2 sequences could be classified into six genotypes according to the species classification criteria of the International Committee on Taxonomy of Viruses (ICTV). It is worth noting that, in addition to four known caprine astrovirus genotypes (MAstV–33, MAstV–34, Caprine Astrovirus G5.1, and Caprine Astrovirus G3.1), MAstV–13 and MAstV–24 genotypes were identified in goats. Interestingly, five of 19 ORF2 sequences in the Caprine Astrovirus G3.1 genotype showed possible intragenotypic recombination events. Furthermore, nearly complete caprine astrovirus genomes of MAstV–13 and MAstV–24 genotypes were obtained. The genome of the SWUN/ECJK3/2021 strain shared the highest similarity (62.0% to 73.9%) with astrovirus in MAstV–13, and clustered in the so-called human-mink-ovine (HMO) clade, which contained the majority of the neurotropic astrovirus strains. Moreover, the SWUN/LJK2-2/2020 strain showed the highest similarity (69.7% to 78.6%) and the closest genetic relationship to the known porcine and bovine astroviruses in MAstV–24. In conclusion, this study confirmed six genotypes of astrovirus circulating among goats in Southwest China, including MAstV–13 and MAstV–24 genotypes. These findings enhance our knowledge of the prevalence and diversity of astroviruses.IMPORTANCE Caprine astrovirus is a newly emerging virus, and information regarding its prevalence and molecular characteristics remains limited. In this study, six genotypes of astrovirus, including MAstV–13 and MAstV–24, were identified in goats, adding two novel caprine astrovirus genotypes to the four previously known genotypes, thereby enriching the diversity of the caprine astrovirus. Moreover, genomes of MAstV–13 SWUN/ECJK3/2021 and MAstV–24 SWUN/LJK2-2/2020 strains were obtained from goats, which aids in the understanding of the infection spectrum and host range of the two genotypes. This study is the first to demonstrate the presence of neurotropic-like astrovirus (MAstV–13) in goats, which has significant implications for the diagnosis of neurological diseases in goats.

Treatment Response and Drug Resistance Profiling of Genotype 6 of Hepatitis C Virus in HCV/HIV Co-Infected Patients: A Pilot Study from INDIA.

Hepatitis C Virus (HCV) genotype (GT) 6 demonstrates maximum genomic diversity out of all the known genotypes of HCV, attributable to its inherent intra-genotype and inter-genotype recombination property. This is the most common genotype seen in HCV/HIV co-infected cases. HIV/HCV co-infection is linked with increased genetic diversity in HCV structural genes. The detailed information on the distribution of HCV GT6, its subtypes, and resistance to currently available antiviral drugs is limited in the Indian subcontinent. Therefore, in this single-center retrospective cross-sectional study, we aimed to map the occurrence of HCV GT6, its subtypes and resistance-associated substitution (RAS), and its correlation with antiviral treatment response in HCV-infected patients. From a cohort of 2052 HCV-infected patients, the overall prevalence of GT6 was 2.5% (n = 53), with a maximum of 81.1% (n = 43) seen in HCV/HIV co-infected patients. Nine different subtypes, 6a, 6b, 6f, 6i, 6n, 6u, 6v, 6w, and 6xa, were detected in the Indian population for the first time, with a predominance of 6xa (41.5%), a rare subtype, followed by 6n (39.6%). The phylogenetic analysis by the neighbor-joining method revealed three prominent viral clades, 6v, 6n, and 6xa–6u. The baseline (before treatment initiation) plasma samples of all GT6-infected patients were retrieved from −80 °C and a part of the NS5a and NS5b region of the viral genome was analyzed for the presence of RAS. No RASs were seen in the NS5b region, while in two patients (3.7%) RASs were seen at baseline in the NS5a region of the virus. Sustained viral response (SVR) was attained in 81% (n = 43) of patients. No difference in GT6 subtype distribution or occurrence of RAS was seen between mono-infected HCV and HIV/HCV co-infected cases. Our study revealed that RAS at baseline did not influence the attainment of SVR and the currently available antiviral therapy is effective against GT6 mono-infected and HIV/HCV co-infected patients.

Genomic Epidemiology and Evolution of Duck Hepatitis A Virus.

Duck hepatitis A virus (DHAV), an avian picornavirus, causes high-mortality acute disease in ducklings. Among the three serotypes, DHAV-1 is globally distributed, whereas DHAV-2 and DHAV-3 serotypes are chiefly restricted to Southeast Asia. In this study, we analyzed the genomic evolution of DHAV-1 strains using extant GenBank records and genomic sequences of 10 DHAV-1 strains originating from a large disease outbreak in 2004–2005, in Hungary. Recombination analysis revealed intragenotype recombination within DHAV-1 as well as intergenotype recombination events involving DHAV-1 and DHAV-3 strains. The intergenotype recombination occurred in the VP0 region. Diversifying selection seems to act at sites of certain genomic regions. Calculations estimated slightly lower rates of evolution of DHAV-1 (mean rates for individual protein coding regions, 5.6286 × 10−4 to 1.1147 × 10−3 substitutions per site per year) compared to other picornaviruses. The observed evolutionary mechanisms indicate that whole-genome-based analysis of DHAV strains is needed to better understand the emergence of novel strains and their geographical dispersal.

Phylogenetic analysis and evolution of feline bocavirus in Anhui Province, eastern China

To understand the epidemic status of feline bocavirus (FBoV) in Anhui Province, eastern China, FBoV was successfully extracted from fecal samples of domestic cats, and five complete genomes were amplified in this study. Phylogenetic analysis showed that these five strains belong to three different FBoV genotypes. Recombination analysis showed that inter- and intra-genotype recombination events occurred. Selection pressure and codon usage bias analyses indicated that FBoV-1 and FBoV-3 continuously evolve toward adaptation, and selection pressure is the main factor for codon usage bias during evolution. This study provides the first molecular evidence of FBoV prevalence in eastern China, further enriching the available information on its genetics and evolutionary characteristics and providing a basis for further research on its evolution.

The emergence, evolution and spread of infectious bronchitis virus genotype GI-23.

Avian infectious bronchitis is a contagious viral disease, caused by avian infectious bronchitis virus (IBV), that leads to severe losses in the poultry industry all over the world. Since the 1950s, IBV has circulated in the Middle East and North Africa, and no tangible evidence has shown any effects of measures taken to control its spread or evolution. Furthermore, new IBV variants are continually discovered. Although several genetic studies on IBV have been conducted, many IBV strains from this region have either been misclassified or remain unclassified. The genotype 23 (GI-23) variant emerged and has prevailed in the Middle East by continuously evolving through inter- and/or intra-genotypic recombination. The GI-23 genotype is currently enzootic throughout Europe and Asia. Although many studies of protection against the circulating strains have been conducted, they have not been standardized according to regulatory requirements. In this review, we provide an overview of the evolution and genetic diversity of IBV genotypes and a genetic classification of IBV strains, with a focus on the GI-23 genotype. The high prevalence of IBV GI-23 strains necessitates the adoption of vaccination schemes using GI-23-based vaccines.Supplementary InformationThe online version contains supplementary material available at 10.1007/s00705-020-04920-z.

Novel genotype definition and genome characteristics of duck circovirus in central and Eastern China.

To explore genetic variations in duck circovirus (DuCV) and the molecular epidemiology of its infection, tissue samples were collected from 219 dead ducks from 20 farms in the central and eastern regions of China. All farms tested positive for DuCV, with duck-origin goose parvovirus, reovirus and Tembusu virus having co-infection rates of 100%, 0% and 0%, respectively. A total of 20 strains from the DuCV-positive flock were sequenced. The total sequence length was 1987-1996nt, and the sequences shared 82% (JX499186, DuCV2 from Sichuan province, China) to 99.7% (KY328304, DuCV1 from Shandong Province, China) sequence identity with DuCV sequences available in GenBank. Hyper-variable regions were mainly located in open reading frame (ORF)2, ORF3 and intergenic regions. The tertiary structure of ORF2 from four provinces (Henan, Anhui, Zhejiang and Fujian) in China showed a canonical viral jelly roll and the antigenic epitope of ORF2 located in the bulge of the protein surface. Overall, 15 of the 20 DuCV strains are possibly derived through inter-genotypic and intragenotypic recombination. Based on sequence and phylogenetic analyses, six strains from Fujian Province clustered into a novel genotype-DuCV-1d. These findings may enrich our understanding of DuCV evolution and circulation and lay the foundation for vaccine strain selection.

Genomic Analyses of Human Sapoviruses Detected over a 40-Year Period Reveal Disparate Patterns of Evolution among Genotypes and Genome Regions.

Human sapovirus is a causative agent of acute gastroenteritis in all age groups. The use of full-length viral genomes has proven beneficial to investigate evolutionary dynamics and transmission chains. In this study, we developed a full-length genome sequencing platform for human sapovirus and sequenced the oldest available strains (collected in the 1970s) to analyse diversification of sapoviruses. Sequence analyses from five major genotypes (GI.1, GI.2, GII.1, GII.3, and GIV.1) showed limited intra-genotypic diversification for over 20–40 years. The accumulation of amino acid mutations in VP1 was detected for GI.2 and GIV.1 viruses, while having a similar rate of nucleotide evolution to the other genotypes. Differences in the phylogenetic clustering were detected between RdRp and VP1 sequences of our archival strains as well as other reported putative recombinants. However, the lack of the parental strains and differences in diversification among genomic regions suggest that discrepancies in the phylogenetic clustering of sapoviruses could be explained, not only by recombination, but also by disparate nucleotide substitution patterns between RdRp and VP1 sequences. Together, this study shows that, contrary to noroviruses, sapoviruses present limited diversification by means of intra-genotype variation and recombination.

Emergence and genetic analysis of variant pathogenic 4/91 (serotype 793/B) infectious bronchitis virus in Egypt during 2019.

Infectious bronchitis virus (IBV) affects both vaccinated and unvaccinated flocks worldwide, with a significant impact on the poultry industry. The aim of the present study is to characterize an emerging variant pathogenic IBV originating from field outbreaks in vaccinated Egyptian layer flock. Samples were collected from disease-suspected flock with a history of administration of live and inactivated IBV vaccines (Ma5 type). Virus propagation in embryonated chicken eggs (ECEs), after three successive passages, revealed typical IBV lesions such as curling and dwarfism. The reported isolate was identified by a real-time reverse transcriptase PCR assay targeting nucleocapsid (N) gene and, further characterized by full-length spike (S1) gene sequencing. Phylogenetic analysis revealed clustering of the isolated virus within 4/91 genotype of GI-13 lineage. Deduced amino acid sequences identity revealed 75–76% and 88–90% similarity with the currently used classic (H120, Ma5, and M41) and variant vaccine strains (4/91 and CR88) in Egypt, respectively. Recombination analysis gave an evidence for distinct patterns of origin for the studied isolate providing another example of intra-genotypic recombination among IBVs and the first example of recombination within the GI-13 lineage in the Egyptian field. The studied isolate (IBV/CK/EG/Fadllah-10/2019) emerged as a result of recombination between the variant group (Egy/var I genotype, GI-23 lineage) as a major parent and the CR88 variant vaccine strain (4/91 genotype, GI-13 lineage) as minor parent. Our data suggest that both mutation and recombination may be contributing to the emergence of IBV variants which ascertain the importance of disease monitoring in vaccinated flocks as well as re-appropriation for the current vaccine strategies.Electronic supplementary materialThe online version of this article (10.1007/s11262-019-01693-9) contains supplementary material, which is available to authorized users.

Norovirus recombinants: recurrent in the field, recalcitrant in the lab - a scoping review of recombination and recombinant types of noroviruses.

Noroviruses are recognized as the major global cause of sporadic and epidemic non-bacterial gastroenteritis in humans. Molecular mechanisms driving norovirus evolution are the accumulation of point mutations and recombination. Intragenotypic recombination has long been postulated to be a driving force of GII.4 noroviruses, the predominant genotype circulating in humans for over two decades. Increasingly, emergence and re-emergence of different intragenotype recombinants have been reported. The number and types of norovirus recombinants remained undefined until the 2007 Journal of General Virology research article 'Norovirus recombination' reported an assembly of 20 hitherto unclassified intergenotypic norovirus recombinant types. In the intervening decade, a host of novel recombinants has been analysed. New recombination breakpoints have been described, in vitro and in vivo studies supplement in silico analyses, and advances have been made in analysing factors driving norovirus recombination. This work presents a timely overview of these data and focuses on important aspects of norovirus recombination and its role in norovirus molecular evolution. An overview of intergenogroup, intergenotype, intragenotype and 'obligatory' norovirus recombinants as detected via in silico methods in the field is provided, enlarging the scope of intergenotypic recombinant types to 80 in total, and notably including three intergenogroup recombinants. A recap of advances made studying norovirus recombination in the laboratory is given. Putative drivers and constraints of norovirus recombination are discussed and the potential link between recombination and norovirus zoonosis risk is examined.

Whole genome analysis of a novel neurotropic bovine astrovirus detected in a Japanese black steer with non-suppurative encephalomyelitis in Japan.

While neurotropic bovine astroviruses (BoAstVs) have been identified in North America and Europe, their presence has never been reported in Asia. In this study, we detected BoAstV in the brain of a steer showing neurological signs. Phylogenetic analysis revealed that the identified virus belongs to the Virginia/Human-Mink-Ovine clade, which contains most of the neurotropic astroviruses including the neurotropic BoAstVs. Similarity plot analysis showed that the virus was closely related to the American BoAstV NeuroS1 strain with respect to the ORF regions and to the European BoAstV CH13 strain in the 3' untranslated region, suggesting the occurrence of intra-genotypic recombination events.

Hepatitis E virus infection of slaughtered healthy pigs in Brazil.

Hepatitis E virus (HEV) is an emerging pathogen that can be transmitted through contaminated raw or undercooked meat derived from domestic pigs. HEV infections have been documented among pig herds, pig products and environmental samples raising concern about the spread of the virus. HEV genotypes 3 and 4 are considered zoonotic and have been linked to human cases. HEV was detected in 51 of 335 bile samples (15.2%) from healthy pigs in Minas Gerais, Brazil. Phylogenetic analysis of partial sequences from ORF1 and ORF2 regions yielded discordant results, assigning isolates to subtypes 3c and 3i, respectively, suggesting intragenotypic HEV recombination.

Genetic diversity and recombination of enterovirus G strains in Japanese pigs: High prevalence of strains carrying a papain-like cysteine protease sequence in the enterovirus G population.

To study the genetic diversity of enterovirus G (EV-G) among Japanese pigs, metagenomics sequencing was performed on fecal samples from pigs with or without diarrhea, collected between 2014 and 2016. Fifty-nine EV-G sequences, which were >5,000 nucleotides long, were obtained. By complete VP1 sequence analysis, Japanese EV-G isolates were classified into G1 (17 strains), G2 (four strains), G3 (22 strains), G4 (two strains), G6 (two strains), G9 (six strains), G10 (five strains), and a new genotype (one strain). Remarkably, 16 G1 and one G2 strain identified in diarrheic (23.5%; four strains) or normal (76.5%; 13 strains) fecal samples possessed a papain-like cysteine protease (PL-CP) sequence, which was recently found in the USA and Belgium in the EV-G genome, at the 2C–3A junction site. This paper presents the first report of the high prevalence of viruses carrying PL-CP in the EV-G population. Furthermore, possible inter- and intragenotype recombination events were found among EV-G strains, including G1-PL-CP strains. Our findings may advance the understanding of the molecular epidemiology and genetic evolution of EV-Gs.

A novel subgenotype 3a hepatitis E virus isolated from pigs in China.

Hepatitis E virus (HEV) infection is an emerging disease with zoonotic transmission that represents a serious public health concern, especially in developing countries. Here we characterize a novel HEV strain CCST-517, which possesses a complete genome sequence of 7284bp with typical HEV genome organization including 5' and 3' non-coding regions and three open reading frames. The sequence identities of CCST-517 with known HEV genotype 1, 2, 3, and 4 were 73.4-73.7, 73.2, 80.4-90.4, and 75.1-75.7%, respectively. Phylogenetic analysis clustered CCST-517 to the clade of HEV genotype 3a, together with the Japanese human HEV isolate (HE-JA10) and United States human HEV isolate (HEV-US2). Similarity plot analysis indicated that the fragment extending from 4500 to 5500nt included evidence of one intra-genotype recombination event in the genome sequence of the CCST-517 strain. To our knowledge, this is the first report of HEV genotype 3a with its complete genome sequence revealed in China. Our findings revealed a close phylogenetic relationship of CCST-517 to human HEV-US2 and HE-JA10, implying cross-species transmission of HEV between pigs and humans.

Whole genome analysis of porcine astroviruses detected in Japanese pigs reveals genetic diversity and possible intra-genotypic recombination

Porcine astroviruses (PoAstVs) are ubiquitous enteric virus of pigs that are distributed in several countries throughout the world. Since PoAstVs are detected in apparent healthy pigs, the clinical significance of infection is unknown. However, AstVs have recently been associated with a severe neurological disorder in animals, including humans, and zoonotic potential has been suggested. To date, little is known about the epidemiology of PoAstVs among the pig population in Japan. In this report, we present an analysis of nearly complete genomes of 36 PoAstVs detected by a metagenomics approach in the feces of Japanese pigs. Based on a phylogenetic analysis and pairwise sequence comparison, 10, 5, 15, and 6 sequences were classified as PoAstV2, PoAstV3, PoAstV4, and PoAstV5, respectively. Co-infection with two or three strains was found in individual fecal samples from eight pigs. The phylogenetic trees of ORF1a, ORF1b, and ORF2 of PoAstV2 and PoAstV4 showed differences in their topologies. The PoAstV3 and PoAstV5 strains shared high sequence identities within each genotype in all ORFs; however, one PoAstV3 strain and one PoAstV5 strain showed considerable sequence divergence from the other PoAstV3 and PoAstV5 strains, respectively, in ORF2. Recombination analysis using whole genomes revealed evidence of multiple possible intra-genotype recombination events in PoAstV2 and PoAstV4, suggesting that recombination might have contributed to the genetic diversity and played an important role in the evolution of Japanese PoAstVs.

The sequence of the full spike S1 glycoprotein of infectious bronchitis virus circulating in Egypt reveals evidence of intra-genotypic recombination.

Infectious bronchitis virus (IBV) continues to circulate worldwide, with a significant impact on the poultry industry and affecting both vaccinated and unvaccinated flocks. Several studies have focused on the hypervariable regions (HVRs) of the spike gene (S1); however, genetic and bioinformatics studies of the whole S1 gene are limited. In this study, the whole S1 gene of five Egyptian IBVs was genetically analyzed. Phylogenetic analysis revealed that the Egyptian IBVs are clustered within two distinct groups: the classic group resembling the GI-1 genotype (vaccine strains) and the variant group (field strains) of the GI-23 genotype. The variant genotype was divided into two distinct subgroups (Egy/var I and Egy/var II) resembling the Israeli variants IS/1494 and IS885 strain, respectively. Significant amino acid sequence differences between the two subgroups, especially in the epitope sites, were identified. A deletion at position 63 and an I69A/S substitution mutation associated with virus tropism were detected in the receptor-binding sites. The deduced amino acid sequence of HVRs of the variant subgroups indicated different genetic features in comparison to the classic vaccine group (H120 lineage). The Egyptian variant IBVs also contained additional N-glycosylation sites compared to the classical viruses. Recombination analysis gave evidence for distinct patterns of origin by recombination throughout the S1 gene, suggesting that the recent virus IBV-EG/1586CV-2015 emerged as a recombinant of two viruses from the variant groups Egy/var I and Egy/var II, providing another example of intra-genotypic recombination among IBVs and the first example of recombination within the GI-23 genotype. Our data suggest that both mutation and recombination may be contributing to the emergence of IBV variants. Moreover, we found that the commercially used vaccines are genotypically distant from the circulating field strains. Hence, continuous follow-up of the current vaccine strategy is highly recommended for better control and prevention of infectious bronchitis virus in the poultry sector in Egypt.

Prevalence and Phylogenetic Analysis of Human Bocaviruses 1-4 in Pediatric Patients with Various Infectious Diseases.

ObjectivesViral infections caused by human bocaviruses 1–4 (HBoV1-4) are more complicated than previously believed. A retrospective, large-scale study was undertaken to explore the prevalence of HBoV1-4 in pediatric patients with various infectious diseases and delineate their phylogenetic characteristics.MethodsClinical samples from four specimen types, including 4,941 respiratory, 2,239 cerebrospinal fluid (CSF), 2,619 serum, and 1,121 fecal specimens, collected from pediatric patients with various infectious diseases were screened for HBoV1-4. A 690-nt fragment in each specimen was then amplified and sequenced for phylogenetic analysis. Clinical characteristics of HBoV-positive patients with different specimen types available were evaluated.ResultsApproximately 1.2% of patients were confirmed as HBoV-positive, with the highest positive rate in patients with gastrointestinal infection (2.2%), followed by respiratory (1.65%), central nervous system (0.8%), and hematological infections (0.2%). A single genetic lineage of HBoV1 circulated among children over the 8-year period, while a new cluster of HBoV2, via intra-genotype recombination between HBoV2A and HBoV2B, was prevalent. Some patients had HBoV1-positive respiratory and serum specimens or fecal specimens. Several cases became HBoV1-positive following the appearance of respiratory infection, while several cases were positive for HBoV2 only in CSF and serum specimens, rather than respiratory specimens.ConclusionsA single genetic lineage of HBoV1 is speculated as a viral pathogen of respiratory infection and causes both comorbid infection and acute gastroenteritis. Additionally, a new cluster of HBoV2 is prevalent in China, which may infect the host through sites other than the respiratory tract.

Characterization of novel intragenotype recombination events among norovirus pandemic GII.4 variants

Recently, there has been an increase in the number of children hospitalized due to norovirus infection in Brazil. This is due both to the occurrence of more severe norovirus-related gastroenteritis cases after the introduction of the rotavirus vaccine and an increase in the tools for the detection of the disease. This pathogen is transmitted by the fecal-oral route, and the illness is characterized by diarrhea, vomiting, nausea and abdominal cramps. The genome of the virus is organized into three open reading frames showing strong mutation rates. Additionally, homologous recombination events, which can increase the virulence of the virus and lead to genotyping mistakes in molecular epidemiological studies, frequently occur. The purpose of this study was to describe two recombination events among different GII.4 variants that infected children who were hospitalized for severe acute gastroenteritis during distinct periods of time in Belém, Brazil. The recombination among the variants US95_96/Kaiso_2003 and Den Haag_2006b/Yerseke_2006a were observed in May 2003 and February 2009, respectively. In both cases, the association between the dominant variant at that point in time and another that was circulating at a low frequency in the population of Belém was demonstrated. Interestingly, the position of the breakpoint of the recombination event in the genome was the polymerase gene and was located at the nucleotide positions 4.834 and 5.002, which is an unusual location for the occurrence of recombination as other studies have previously reported the junction region as a breakpoint. In this study, both recombinant variant strains were related to severe cases of diarrhea that lead to hospitalization, demonstrating the viral evolution of GII.4 in response to selective pressures, which ultimately lead to the emergence of novel viral types in the pediatric population. The cases discussed here reinforce the need for continuous norovirus surveillance. To our knowledge, these two GII.4 variant recombinations have not yet been previously described.

Prevalence analysis of different human bocavirus genotypes in pediatric patients revealed intra-genotype recombination

BackgroundHuman bocavirus (HBoV) genotypes 1–4 have been detected worldwide in respiratory samples and stool samples, and are increasingly associated with respiratory and intestinal infections of previously unknown etiology in young children. Several studies revealed evidence of extensive recombination among HBoV genotypes at the NP1 and VP1 gene boundary region. This study explored the prevalence of HBoV genotypes in pediatric patients in Beijing, and studied their phylogeny. MethodsA total of 4941 respiratory specimens and 1121 fecal specimens were collected from pediatric patients with respiratory infections from January 2006 to December 2013, or with acute diarrhea from October 2010 to December 2012. Conventional PCR was used to detect HBoV1–4 within these samples. Gene fragments at the NP1 and VP1 gene boundary were amplified from HBoV-positive specimens, sequenced, and their phylogenetic inferences constructed using MEGA 6.0 software. Recombination events were identified with SimPlot software. ResultsHuman bocavirus 1, 2, and 3 were detected in 9 (0.80%), 15 (1.33%), and 1 (0.08%) of 1121 stool samples, respectively. However, only HBoV1 (82, 1.65%) was detected in respiratory specimens. Phylogenetic analysis of gene fragments at the HBoV NP1 and VP1 gene boundary indicated that HBoV1 sequences obtained from fecal or respiratory specimens across 8years were highly conserved (99–100%), while 15 HBoV2 sequences collected across 2years in Beijing were more diverse with up to 4.40% variation. Of the 15 HBoV2 sequences, 14 clustered into a new lineage divergent from other HBoV2 sequences in GenBank. Five HBoV2 genomic sequences were analyzed for recombination, revealing intra-genotype recombination between HBoV2A and HBoV2B. ConclusionsMore HBoV1 were detected in children with respiratory tract diseases, and HBoV2 in patients with acute diarrhea. Phylogenetic analysis revealed a new cluster of HBoV2 was prevalent in China, which may be the result of intra-genotype recombination between HBoV2A and HBoV2B.