Genetic diversity of G9P[8] rotavirus strains circulating in Italy in 2007 and 2010 as determined by whole genome sequencing

This study reports the molecular characterization of G9P[8] rotavirus strains from children with acute diarrhea identified in different cities of Italy, in 2007 and 2010. Seventeen samples exhibited a G9P[8] genotype by RT-PCR and semi-nested PCR. Preliminary sequence analysis of the VP7 and VP8^* encoding genes revealed nucleotide identities ranging between 96% and 100%. Full genome sequencing of four G9P[8] strains selected in different cities or years showed that the investigated Italian strains possessed a complete Wa-like genotype constellation. However, phylogenetic analyses assigned strains to different clusters reflecting point mutations and possibly earlier reassortment between Wa-like RVA strains. Deduced amino acid sequence of the VP7 and VP4 genes for the G9P[8] strains revealed at least five substitutions in relevant antigenic sites of both proteins.     

Whole genome analyses of DS-1-like Rotavirus A strains detected in children with acute diarrhoea in southern Mozambique suggest several reassortment events

We report the first whole genome constellations of Mozambican rotavirus A strains detected between 2012 and 2013 in the Mavalane General Hospital in Maputo city and Manhiça District Hospital in the Manhiça district. Consensus sequences for ten DS-1-like strains (G2P[4] and G8P[4]) were identified with an Illumina Miseq platform using cDNA prepared from dsRNA extracted from stool samples, without genome amplification or prior adaptation to cell culture. Comparison of previously reported genotyping results and the consensus sequences described in this study, indicated that the genotype primers specific for G12 and P[4] might require revision. Phylogenetic analyses indicated diversity among the G2P[4] Mozambican strains and suggested reassortment between G2P[4] and G8P[4] Mozambican strains, as well as the intragenogroup reassortment of all the genome segments encoding VP1, 2, 3 and 6 for strain RVA/Human-wt/MOZ/0045/2012G8P[4]. These results highlight the necessity to determine whole genome constellations to confirm surveillance data in Africa and to monitor the growing diversity in DS-1-like strains.     

Multiple reassortment and interspecies transmission events contribute to the diversity of feline, canine and feline/canine-like human group A rotavirus strains

RNA–RNA hybridization assays and complete genome sequence analyses have shown that feline rotavirus (FRV) and canine rotavirus (CRV) strains display at least two distinct genotype constellations (genogroups), represented by the FRV strain RVA/Cat-tc/AUS/Cat97/1984/G3P[3] and the human rotavirus (HRV) strain RVA/Human-tc/JPN/AU-1/1982/G3P3[9], respectively. G3P[3] and G3P[9] strains have been detected sporadically in humans. The complete genomes of two CRV strains (RVA/Dog-tc/ITA/RV198-95/1995/G3P[3] and RVA/Dog-tc/ITA/RV52-96/1996/G3P[3]) and an unusual HRV strain (RVA/Human-tc/ITA/PA260-97/1997/G3P[3]) were determined to further elucidate the complex relationships among FRV, CRV and HRV strains. The CRV strains RV198-95 and RV52-96 were shown to possess a Cat97-like genotype constellation. However, 3 and 5 genes of RV198-95 and RV52-96, respectively, were found in distinct subclusters of the same genotypes, suggesting the occurrence of reassortment events among strains belonging to this FRV/CRV/HRV genogroup. Detailed phylogenetic analyses of the HRV strain PA260-97 showed that (i) 8 genome segments (VP3, VP4, VP6, VP7 and NSP2-5) clustered closely with RV198-95 and/or RV52-96; (ii) 2 genome segments (VP1 and VP2) were more closely related to HRV AU-1; and (iii) 1 genome segment (NSP1) was distantly related to any other established NSP1 genotypes and was ratified as a new NSP1 genotype, A15. These findings suggest that the human strain PA260-97 has a history of zoonotic transmission and is likely a reassortant among FRV/CRV strains from the Cat97 and AU-1-like genogroups. In addition, a potential third BA222-05-like genogroup of FRV and HRV strains should be recognized, consisting of rotavirus strains with a stable genetic genotype constellation of genes also partially related to bovine rotavirus (BRV) and bovine-like rotaviruses. The detailed phylogenetic analysis indicated that three major genotype constellations exist among FRV, CRV and feline/canine-like HRV strains, and that reassortment and interspecies transmission events contribute significantly to their wide genetic diversity.     

Whole genome analysis of multiple rotavirus strains from a single stool specimen using sequence-independent amplification and 454? pyrosequencing reveals evidence of intergenotype genome segment recombination

Infection of a single host cell with two or more different rotavirus strains creates conditions favourable for evolutionary mechanisms like reassortment and recombination that can generate novel strains. Despite numerous reports describing mixed rotavirus infections, whole genome characterisation of rotavirus strains in a mixed infection case has not been reported. Double-stranded RNA, exhibiting a long electropherotype pattern only, was extracted from a single human stool specimen (RVA/Human-wt/ZAF/2371WC/2008/G9P[8]). Both short and long electropherotype profiles were however detected in the sequence-independent amplified cDNA derived from the dsRNA, suggesting infection with more than one rotavirus strain. 454? pyrosequencing of the amplified cDNA revealed co-infection of at least four strains. Both genotype 1 (Wa-like) and genotype 2 (DS-1-like) were assigned to the consensus sequences obtained from the nine genome segments encoding NSP1-NSP5, VP1-VP3 and VP6. Genotypes assigned to the genome segments encoding VP4 were P[4] (DS-1-like), P[6] (ST3-like) and P[8] (Wa-like) genotypes. Since four distinct genotypes [G2 (DS-1-like), G8, G9 (Wa-like) and G12] were assigned to the four consensus nucleotide sequences obtained for genome segment 9 (VP7), it was concluded that at least four distinct rotaviruses were present in the stool. Intergenotype genome recombination events were observed in genome segments encoding NSP2, NSP4 and VP6. The close similarities of some of the genome segments encoding NSP2, VP6 and VP7 to artiodactyl rotaviruses suggest that some of the infecting strains shared common ancestry with animal strains, or that interspecies transmission occurred previously. The sequence-independent genome amplification technology coupled with 454? pyrosequencing used in this study enabled the characterisation of the whole genomes of multiple rotavirus strains in a single stool specimen that was previously assigned single genotypes, i.e. G9P[8], by sequence-dependent RT-PCR.     

Prevalence and genome characterization of porcine rotavirus A in southern Mozambique

Rotavirus A (RVA) is an important pathogen causing gastroenteritis in many species, including humans and pigs. The objective of this study was to determine the prevalence of RVA in pigs from smallholdings and commercial farms in southern Mozambique and characterize the complete genomes of selected strains. RVA was detected at a rate of 11.8% (n = 288), of which 7.6% was detected at commercial farms and 4.2% at smallholdings. The whole genomes of eight rotavirus strains were determined using an Illumina MiSeq platform. Seven displayed a G9P[13] and one a G4P[6] genotype combination, all with a typical porcine backbone (I1/5-R1-C1-M1-A1/8-N1-T1/7-E1-H1). Phylogenetic analysis indicated that the seven G9P[13] strains were in fact one strain that circulated on a commercial pig farm. The genome segments of this strain clustered with diverse segments of human and porcine RVA strains from various Asian countries. Analysis of the G4P[6] strain revealed four distinct genome segments (VP2, VP4, VP6 and VP7) and five genome segments closely related to South African porcine rotavirus strains (NSP1, NSP3, NSP4, NSP5 and VP1). These results suggest that both the G4P[6] and the G9P[13] strains possibly emerged through multiple reassortment events. The presence of these strains on the commercial farms and smallholdings calls for a more in-depth surveillance of rotavirus in Mozambique.     

Evidence of discordant genetic linkage in the VP4, VP6, VP7 and NSP4 encoding genes of rotavirus strains from adolescent and adult patients with acute gastroenteritis

The vast diversity within rotavirus strains circulating in the developing countries continues to be a major challenge for the efficacy of currently used preset rotavirus vaccines. The sequence analysis and phylogeny of multiple genes of rotavirus strains enable identification of reassortant strains and their human or animal origin. The objective of this study was to monitor the genetic linkage between the rotavirus VP4(P), VP6(I), VP7(G) and NSP4(E) encoding genes. The G, P, I and E genotypes of a total of 80 rotavirus strains isolated from adolescent and adult cases of acute gastroenteritis at the two time points [1993–1996 (n = 67) and 2004–2007 (n = 13)] were determined by nucleotide sequencing and phylogenetic analysis. The rotavirus strains from the 1990s and 2000s revealed common combinations of genotypes (G1–P[8]–I1–E1, G2–P[4]–I2–E2, G3–P[8]–I1–E1 and G4–P[8]–I1–E1) in 47.8% and 30.8%, unusual combinations of the same genotypes (G2–P[8]–I2–E2, G9–P[6]–I1–E1, G9–P[6]–I1–E2, G9–P[6]–I2–E1 and G4–P[4]–I1–E2, G1–P[4]–I2–E1, G9–P[4]–I1–E1) in 7.5% and 23% and mixed infections of different G and P genotypes in 31.3% and 46.2%, respectively. Discordance in the association of I with E, G with I and E and P with I and E genotypes was found to be contributed respectively by 23.8–38.5%, 40.3–69.8% and 49.3–61.5% of the rotavirus strains at the two time points.The data suggest relatively high occurrence of intergenogroup reassortment in circulating rotavirus strains emphasizing the need for continuous surveillance and whole genome sequence based characterization of rotavirus strains for better understanding of their evolution and ecology.     

Genetic characterization of group A rotavirus strains circulating among children with acute gastroenteritis in Japan in 2004-2005.

A total of 752 fecal specimens collected from July 2004 to June 2005 from children with acute gastroenteritis in four localities in Japan (Maizuru, Tokyo, Sapporo, and Osaka) were screened for group A rotavirus by RT-PCR. It was found that 82 (10.9%) specimens were positive for group A rotavirus. The G-(VP7 genotypes) and P-(VP4 genotypes) types were further investigated. The P-types of 18 rotavirus strains, which could not be typed by RT-PCR, were determined by sequencing analysis. Of these, 94% (17/18) were P[8] with multiple point mutations at the VP4 primer-binding site. Another sample turned out to be a rare genotype P[9], which was closely related to feline rotavirus. The predominant genotype was G1P[8] (46.4%), followed by G3P[8] (32.9%) and G2P[4] (12.2%). A number of unusual combinations including, G1P[4] (1.2%), G2P[8] (1.2%), G3P[9] (1.2%), G1G3P[8] (1.2%), and G2G3P[8] (3.7%), were also detected. A new nomenclature of P[8] was proposed, in which worldwide rotavirus P[8] strains were classified into four sub-lineages, namely IA, IB, IIA, and IIB. A wide range of amino acid substitutions (up to 22) specific for P[8] lineages and sub-lineages were also identified. Interestingly, only short amino acid motifs located at positions 32-35, 121-135, and 195-236 of VP4 correctly defined the phylogenetic P[8] lineages and sub-lineages. Of note, at least two distinct clusters of rotavirus P[8] were co-circulating in the Japanese pediatric population studied.     

Full genomic characterization of a novel genotype combination,G4P[14], of a human rotavirus strain from Barbados

Since 2004, the Pan American Health Organization (PAHO) has carried out rotavirus surveillance in Latin America and the Caribbean. Here we report the characterization of human rotavirus with the novel G–P combination of G4P[14], detected through PAHO surveillance in Barbados. Full genome sequencing of strain RVA/Human-wt/BRB/CDC1133/2012/G4P[14] revealed that its genotype is G4-P[14]-I1-R1-C1-M1-A8-N1-T1-E1-H1. The possession of a Genogroup 1 (Wa-like) backbone distinguishes this strain from other P[14] rotavirus strains. Phylogenetic analyses suggested that this strain was likely generated by genetic reassortment between human, porcine and possibly other animal rotavirus strains and identified 7 lineages within the P[14] genotype. The results of this study reinforce the potential role of interspecies transmission in generating human rotavirus diversity through reassortment. Continued surveillance is important to determine if rotavirus vaccines will protect against strains that express the P[14] rotavirus genotype.     

Complete genetic characterization of human G2P[6] and G3P[6] rotavirus strains

During the 2008–2009 rotavirus season, 10 G3P[6] rotavirus strains were isolated for the first time in Belgium, while an outbreak of G2P[6] strains occurred in the USA in 2005–2006. Partial sequencing of the 11 genome segments of the 10 Belgian G3P[6] strains revealed a clonal origin. Two of these strains, and a G2P[6] strain representative of the American outbreak, were selected and sequenced completely to analyze their evolutionary relationships. Genetic analysis revealed that all strains possessed a DS-1-like genotype constellation. The 2 Belgian G3P[6] strains showed >99% sequence identity at the nucleotide level and the American G2P[6] strain was phylogenetically closely related to the Belgian P[6] strains. These data suggest that reassortment(s) involving VP7 occurred recently, and that the prevalence of DS-1-like P[6] rotavirus strains need to be closely monitored because the currently licensed RVA vaccines contain neither the P[6] genotype nor strains with a complete human DS-1 genotype constellation.     

Molecular characterization of rotavirus strains from pre- and post-vaccination periods in a country with low vaccination coverage: The case of Slovenia

Rotavirus vaccination started in Slovenia in 2007 on a voluntarily basis. The vaccination rate is relatively low (up to 27%) and no increasing trend is observed. We present rotavirus genotype distribution among children hospitalized for rotavirus gastroenteritis in Slovenia. Eight consecutive rotavirus seasons were followed, from 2005/06 to 2012/13, and 113 strains of the most common rotavirus genotypes were randomly selected for molecular characterization of rotavirus VP7 and VP4 (VP8¹) genome segments. During the vaccine introduction period, from 2007 to 2013, rotavirus genotype prevalences changed, with G1P[8] decreasing from 74.1% to 8.7% between 2007/08 and 2010/11 seasons, replaced by G4P[8] and G2P[4], with up to 52.0% prevalence. Comparable analysis of VP7 and VP8¹ genome fragments within G1P[8] genotype lineages revealed considerable differences for rotavirus strains circulating before and during the vaccination period. The G1P[8] rotavirus strains from the pre-vaccination period clustered in a phylogenetic tree within Rotarix®-like VP7 and VP8¹ lineages. However, since 2007, the majority of G1P[8] strains have shifted to distant genetic lineages with lower nucleotide (88.1–94.0% for VP7 and 86.6–91.1% for VP8¹) and amino acid (93.8–95.2% for VP7 and 85.3–94.6% for VP8¹) identities to the vaccine Rotarix® strain. This change also resulted in a different deduced amino acid profile at the major VP7 and VP8¹ antigenic epitopes.     

Genotypic linkages of VP4, VP6, VP7, NSP4, NSP5 genes of rotaviruses circulating among children with acute gastroenteritis in Thailand

Rotavirus is the main cause of acute viral gastroenteritis in infants and young children worldwide. Surveillance of group A rotavirus has been conducted in Chiang Mai, Thailand since 1987 up to 2004 and those studies revealed that group A rotavirus was responsible for about 20-61% of diarrheal diseases in hospitalized cases. In this study, we reported the continuing surveillance of group A rotavirus in 2005 and found that group A rotavirus was detected in 43 out of 147 (29.3%) stool samples. Five different G and P genotype combinations were detected, G1P[8] (27 strains), G2P[4] (12 strains), G9P[8] (2 strains), G3P[8] (1 strain), and G3P[10] (1 strain). In addition, analysis of their genotypic linkages of G (VP7), P (VP4), I (VP6), E (NSP4), and H (NSP5) genotypes demonstrated that the rotaviruses circulating in Chiang Mai, Thailand carried 3 unique linkage patterns. The G1P[8], G3P[8], and G9P[8] strains carried their VP6, NSP4, NSP5 genotypes of I1, E1, H1, respectively. The G2P[4] strains were linked with I2, E2, H2 genotypes, while an uncommon G3P[10] genotype carried unique genotypes of I8, E3 and H6. These findings provide the overall picture of genotypic linkage data of rotavirus strains circulating in Chiang Mai, Thailand.     

Full genomic analyses of human rotavirus strains possessing the rare P[8]b VP4 subtype

Rotaviruses with the P[8] VP4 genotype are a major cause of acute infantile diarrhea. The P[8] genotype is classified into two genetically distinct subtypes, P[8]a and P[8]b. Most of the P[8] strains belong to subtype P[8]a, whilst P[8]b strains are rare. To date, the whole genomes of a few P[8]a strains have been analyzed, whilst there are no reports on full genomic analysis of the P[8]b strains. To determine the genetic makeup of the rare P[8]b strains and their overall genetic relatedness to the P[8]a strains, the present study analyzed the full genomes of a human G9P[8]b strain, MMC38, and a G1P[8]b strain, MMC71, detected in Bangladesh in 2005. By nucleotide sequence identities and phylogenetic analyses, strains MMC38 and MMC71 exhibited a human rotavirus Wa-like genotype constellation. Except for the VP4 gene, all the genes of strains MMC38 and MMC71 were closely related to cognate genes of the contemporary and more recent human Wa-like G1P[8]a, G9P[8]a, G11P[8]a, G11P[25], G12P[6] and G12P[8]a strains, including those from Bangladesh. Therefore, strains MMC38 and MMC71 possessed the genetically distinct P[8]b VP4 gene on a common human Wa-like genetic backbone, pointing towards their possible origin from reassortment events between common human Wa-like strains and unidentified rotavirus strains possessing the rare P[8]b-like VP4 gene. Since strains with this stable Wa-like genetic backbone can spread rapidly, and it is not certain as to whether the current rotavirus vaccines will be equally efficacious against the P[8]b strains as the P[8]a strains, proper detection of P[8]b strains and their whole genomic analyses might be of public health significance. To our knowledge, the present study is the first report on full genomic analysis of the rare P[8]b rotavirus strains.     

Whole-genomic analysis of G3P[23], G9P[23] and G3P[13] rotavirus strains isolated from piglets with diarrhea in Thailand, 2006–2008

Group A rotavirus (RVA) is the most common cause of severe acute viral gastroenteritis in humans and animals worldwide. This study characterized the whole genome sequences of porcine RVAs, 2 G3P[23] strains (CMP40/08 and CMP48/08), 1 G9P[23] strain (CMP45/08), and 1 G3P[13] strain (CMP29/08). These strains were collected from diarrheic piglets less than 7 weeks of age in 4 pig farms in Chiang Mai, Thailand, in 2008. The VP7-VP4-VP6-VP1-VP2-VP3-NSP1-NSP2-NSP3-NSP4-NSP5 genes of CMP40/08 and CMP48/08 strains were assigned as G3-P[23]-I5-R1-C1-M1-A8-N1-T1-E1-H1 genotypes based on their nucleotide sequences and phylogenetic analyses. The CMP29/08 strain was different from the CMP40/08 and CMP48/08 strains only in the VP4 gene, since it was assigned as P[13] genotype. Furthermore, the VP7 gene of the CMP45/08 strain was classified as genotype G9, and the NSP3 gene as T7 genotype. The finding of this study supports the porcine-origin of T7 genotype, although the NSP3 gene of this strain was similar to the bovine UK strain at the highest nucleotide sequence identity of 92.6%. Whole genome sequence analysis of the porcine RVAs indicated that multiple inter-genotypic and intra-genotypic reassortment events had occurred among the porcine RVAs circulating in this studied area. Interestingly, the VP7 gene of the CMP45/08 strain, and the VP1, NSP2, and NSP4 genes of all four porcine RVAs strains described in this study revealed much similarity to those of two porcine-like human RVA strains (RVA/Human-tc/THA/Mc323/1989/G9P[19] and RVA/Human-tc/THA/Mc345/1989/G9P[19]) detected in Thailand in 1989. The present study provided important information on the evolution of porcine RVA.     

Reassortment of Human Rotavirus Gene Segments into G11 Rotavirus Strains

G11 rotaviruses are believed to be of porcine origin. However, a limited number of G11 rotaviruses have been recently isolated from humans in combination with P[25], P[8], P[6], and P[4]. To investigate the evolutionary relationships of these strains, we analyzed the complete genomes of 2 human G11P[25] strains, 2 human G11P[8] strains, and 3 porcine reference strains. Most of the 11 gene segments of these 7 strains belonged to genotype 1 (Wa-like). However, phylogenetic clustering patterns suggested that an unknown G11P[25] strain with a new I12 VP6 genotype was transmitted to the human population, in which it acquired human genotype 1 gene segments through reassortment, resulting in a human G11P[8] rotavirus strain with an entire human Wa-genogroup backbone. This Wa-like backbone is believed to have caused the worldwide spread of human G9 and G12 rotaviruses. G11 human rotavirus strains should be monitored because they may also become major human pathogens.     

Molecular characterization of two rare human G8P[14] rotavirus strains,detected in Italy in 2012

Since 2007, the Italian Rotavirus Surveillance Program (RotaNet-Italy) has monitored the diversity and distribution of genotypes identified in children hospitalized with rotavirus acute gastroenteritis.We report the genomic characterization of two rare human G8P[14] rotavirus strains, identified in two children hospitalized with acute gastroenteritis in the southern Italian region of Apulia during rotavirus strain surveillance in 2012.Both strains showed a G8-P[14]-I2-R2-C2-M2-A11-N2-T6-E2-H3 genomic constellation (DS-1-like genomic background). Phylogenetic analysis of each genome segment revealed a mixed configuration of genes of animal and zoonotic human origin, indicating that genetic reassortment events generated these unusual human strains. Eight out of 11 genes (VP1, VP2, VP3, VP6, VP7, NSP3, NSP4 and NSP5) of the Italian G8P[14] strains exhibited close identity with a Spanish sheep strain, whereas the remaining genes (VP4, NSP1 and NSP2) were more closely related to human strains. The amino acid sequences of the antigenic regions of outer capsid proteins VP4 and VP7 were compared with vaccine and field strains, showing high conservation between the amino acid sequences of Apulia G8P[14] strains and human and animal strains bearing G8 and/or P[14] proteins, and revealing many substitutions with respect to the RotaTeq™ and Rotarix™ vaccine strains. Conversely, the amino acid analysis of the four antigenic sites of VP6 revealed a high degree of conservation between the two Apulia strains and the human and animal strains analyzed.These results reinforce the potential role of interspecies transmission and reassortment in generating novel rotavirus strains that might not be fully contrasted by current vaccines.     

Analysis of complete genome sequences of G9P[19] rotavirus strains from human and piglet with diarrhea provides evidence for whole-genome interspecies transmission of nonreassorted porcine rotavirus

Whole genomes of G9P[19] human (RVA/Human-wt/THA/CMH-S070-13/2013/G9P[19]) and porcine (RVA/Pig-wt/THA/CMP-015-12/2012/G9P[19]) rotaviruses concurrently detected in the same geographical area in northern Thailand were sequenced and analyzed for their genetic relationships using bioinformatic tools. The complete genome sequence of human rotavirus RVA/Human-wt/THA/CMH-S070-13/2013/G9P[19] was most closely related to those of porcine rotavirus RVA/Pig-wt/THA/CMP-015-12/2012/G9P[19] and to those of porcine-like human and porcine rotaviruses reference strains than to those of human rotavirus reference strains. The genotype constellation of G9P[19] detected in human and piglet were identical and displayed as the G9-P[19]-I5-R1-C1-M1-A8-N1-T1-E1-H1 genotypes with the nucleotide sequence identities of VP7, VP4, VP6, VP1, VP2, VP3, NSP1, NSP2, NSP3, NSP4, and NSP5 at 99.0%, 99.5%, 93.2%, 97.7%, 97.7%, 85.6%, 89.5%, 93.2%, 92.9%, 94.0%, and 98.1%, respectively. The findings indicate that human rotavirus strain RVA/Human-wt/THA/CMH-S070-13/2013/G9P[19] containing the genome segments of porcine genetic backbone is most likely a human rotavirus of porcine origin. Our data provide an evidence of interspecies transmission and whole-genome transmission of nonreassorted G9P[19] porcine RVA to human occurring in nature in northern Thailand.     

Detection of the first G6P[14] human rotavirus strain from a child with diarrhea in Egypt

We report the first detection of a G6P[14] rotavirus strain in Egypt from the stool of a child participating in a hospital-based diarrhea surveillance study conducted throughout the year 2004. Rotavirus infection was initially detected using a rotavirus group A VP6 enzyme immunoassay; the P (VP4) and G (VP7) genotypes of the strain were identified by RT-PCR. We sequenced the VP7 gene and the VP8* portion of the VP4 gene and the strain displayed the strongest identity to the VP7 [>94% nucleotides (nt), >97% amino acids (aa)] and VP4 (>93% nt, >98% aa) sequences of PA169, a novel G6P[14] strain first isolated from a child in Italy during the winter of 1987. Additional sequencing and analysis of the other remaining structural (VP1–VP3, VP6) and non-structural (NSP1–NSP5) proteins support this animal-to-human reassortment theory. According to the full genome classification system, the G6P[14] strain (EGY3399) was assigned to G6-P[14]-I2-R2-C2-M2-A11-N2-T6-E2-H3 genotypes. The greatest similarity of EGY3399 NSP4 and NSP5 gene sequences were to those of ovine and simian origin, respectively. Coupled with other observations, our results suggest G6P[14] isolates rarely cause severe diarrhea in Egyptian children, and support other studies that indicate animal rotavirus contribute to the genetic diversity of rotavirus detected from humans through interspecies transmission and single or multiple segments reassortment.     

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.     

Characterization of human rotaviruses circulating in Iraq in 2008: Atypical G8 and high prevalence of P[6] strains

Fecal samples from 976 children with gastroenteritis were collected and analyzed for group A rotavirus (RVA), in three different cities in Iraq between January 2008 and December 2008. RVA antigen was detected in 394 (40%) of the samples, and 98 samples were available for further genotype analyses using multiplex RT-PCR and sequence analyses for untypeable strains. The G/P-genotype combination was determined for 69 samples, and 19, 2 and 8 samples remained P-untypeable, G-untypeable and G/P-untypeable (UT), respectively. The most prevalent genotype was G2 (40%, 39/98) most often associated with P[6]. G1 was the second most common genotype (16%, 16/98) mainly associated with P[8] and P[UT]. G3, G4 and G9 were detected at a lower prevalence (3%, 11%, 3%, respectively), mainly associated with P[6]. Surprisingly, five G8P[6], and seven G12 RVA strains in combination with P[6] and P[8] were also detected for the first time in Iraq. Overall, a striking high prevalence of 47% of the analyzed samples possessed the P[6] genotype (65% of the P-typed RVA strains). Atypical genotype combinations such as G1P[4], G1P[6], G2P[8] or strains with mixed G-types were detected sporadically. The detection of unusual G8P[6] RVA strains prompted us to further analyze the NSP2, NSP3, NSP4 and NSP5 gene segments of three selected G8P[6] strains, resulting in their designation to the N2, T2, E2 and H2 genotypes, respectively. The VP7, VP4, NSP2, NSP3 and NSP5 gene segments clustered closely with common human RVA strains, whereas the NSP4 gene sequences were found to cluster with animal derived RVA strains, suggesting a potential reassortment event. The high prevalence of RVA strains with the G8, G12 and P[6] genotypes in combination with a DS-1-like genotype constellation in Iraq, needs to be monitored closely as these RVA strains might challenge the effectiveness of current RVA vaccines.