Whole genome sequencing of a rare rotavirus from archived stool sample demonstrates independent zoonotic origin of human G8P[14] strains in Hungary.

Genotype P[14] rotaviruses in humans are thought to be zoonotic strains originating from bovine or ovine host species. Over the past 30 years only few genotype P[14] strains were identified in Hungary totaling<0.1% of all human rotaviruses whose genotype had been determined. In this study we report the genome sequence and phylogenetic analysis of a human genotype G8P[14] strain, RVA/Human-wt/HUN/182-02/2001/G8P[14]. The whole genome constellation (G8-P[14]-I2-R2-C2-M2-A11-N2-T6-E2-H3) of this strain was shared with another Hungarian zoonotic G8P[14] strain, RVA/Human-wt/HUN/BP1062/2004/G8P[14], although phylogenetic analyses revealed the two rotaviruses likely had different progenitors. Overall, our findings indicate that human G8P[14] rotavirus detected in Hungary in the past originated from independent zoonotic events. Further studies are needed to assess the public health risk associated with infections by various animal rotavirus strains.

Equine-like G3 rotavirus in Hungary, 2015 - Is it a novel intergenogroup reassortant pandemic strain?

Novel, intergenogroup reassortant G3 rotavirus strains are spreading in at least three continents: Asia, Australia, and Europe. The present study provides evidence that a closely related G3P[8] strain circulated in Hungary during 2015. Whole genome sequencing and phylogenetic analysis showed that the identified strain continues to evolve by reassortment. This observation demonstrates the genomic plasticity of the novel strain, which is thought to be a prerequisite of the success of emerging rotavirus genotypes.

Complete genome analysis of contemporary G12P[8] rotaviruses reveals heterogeneity within Wa-like genomic constellation.

G12 rotaviruses are globally emergent rotaviruses causing severe childhood gastroenteritis. Little is known about the evolution and diversity of G12P[8] rotaviruses and the possible role that widespread vaccine use, globally, has had on their emergence. In Sicily, Italy, surveillance activity for rotaviruses has been conducted uninterruptedly since 1985, thus representing a unique observatory for the study of human rotaviruses in the pre- and post-vaccine era. G12 rotaviruses were first detected only in 2012 and between 2012 and 2014 they accounted for 8.7% of all rotavirus-associated infections among children, with peaks of 27.8% in 2012/2013 and 21% in 2014. We determined and analyzed the full-genome of 22 G12P[8] rotaviruses collected during the 2012-2014. Although all G12P[8] rotaviruses exhibited a typical Wa-like genotype constellation (G12P[8]-I1-R1-C1-M1-A1-N1-T1-E1-H1), phylogenetic analysis allowed distinguishing either two or three (sub)lineages in each genome segment. On the basis of the segregation patterns into lineages/sublineages, 20 G12P[8] rotaviruses could be grouped into three stable major genomic sub-constellations, whilst two strains displayed unique genome architectures, likely due to ressortment with co-circulating strains. Altogether, these findings indicate that the onset and prolonged circulation of G12 rotaviruses was due to repeated introductions of different G12 rotaviruses circulating globally. Importantly, as regional rotavirus vaccination was initiated in 2012 reaching a 45% coverage in newborns in 2014, a correlation between the appearance and spread of G12 rotaviruses and the enacted vaccination program could not be drawn. Constant epidemiologic surveillance remains important to monitor the epidemiological dynamics of human rotaviruses.

Zoonotic transmission of rotavirus: surveillance and control.

Group A rotavirus (Rotavirus A, RVA) is the main cause of acute dehydrating diarrhea in humans and numerous animal species. RVA shows vast diversity and a variety of human strains share genetic and antigenic features with animal origin RVA strains. This finding suggests that interspecies transmission is an important mechanism of rotavirus evolution and contributes to the diversity of human RVA strains. RVA is responsible for half a million deaths and several million hospitalizations worldwide. Globally, two rotavirus vaccines are available for routine use in infants. These vaccines show a great efficacy profile and induce protective immunity against various rotavirus strains. However, little is known about the long-term evolution and epidemiology of RVA strains under selective pressure related to vaccine use. Continuous strain surveillance in the post-vaccine licensure era is needed to help better understand mechanisms that may affect vaccine effectiveness.

Canine rotavirus C strain detected in Hungary shows marked genotype diversity.

Species C rotaviruses (RVC) have been identified in humans and animals, including pigs, cows and ferrets. In dogs, RVC strains have been reported anecdotally on the basis of visualization of rotavirus-like virions by electron microscopy combined with specific electrophoretic migration patterns of the genomic RNA segments. However, no further molecular characterization of these viruses was performed. Here, we report the detection of a canine RVC in the stool of a dog with enteritis. Analysis of the complete viral genome uncovered distinctive genetic features of the identified RVC strain. The genes encoding VP7, VP4 and VP6 were distantly related to those of other RVC strains and were putatively classified as G10, P8 and I8, respectively. The new strain was named RVC/Dog-wt/HUN/KE174/2012/G10P[8]. Phylogenetic analyses revealed that canine RVC was most closely related to bovine RVC strains with the exception of the NSP4 gene, which clustered together with porcine RVC strains. These findings provide further evidence for the genetic diversity of RVC strains.

Identification of a multi-reassortant G12P[9] rotavirus with novel VP1, VP2, VP3 and NSP2 genotypes in a child with acute gastroenteritis.

The G12 rotavirus genotype is globally emerging to cause severe gastroenteritis in children. Common G12 rotaviruses have either a Wa-like or DS-1-like genome constellation, while some G12 strains may have unusual genome composition. In this study, we determined the full-genome sequence of a G12P[9] strain (ME848/12) detected in a child hospitalized with acute gastroenteritis in Italy in 2012. Strain ME848/12 showed a complex genetic constellation (G12-P[9]-I17-R12-C12-M11-A12-N12-T7-E6-H2), likely derived from multiple reassortment events, with the VP1, VP2, VP3 and NSP2 genes being established as novel genotypes R12, C12, M11 and N12, respectively. Gathering sequence data on human and animal rotaviruses is important to trace the complex evolutionary history of atypical RVAs.

Emergence of multireassortant bluetongue virus serotype 4 in Hungary.

The genome sequence and the phylogenetic relationships of a serotype 4 bluetongue virus (BTV-4) emerged during 2014 in Hungary are described in this study. Genome segment 2 encoding the major neutralization antigen, VP2, shared moderate sequence similarity (nt, ⩽ 94.3%) with the corresponding gene of contemporary and historic homotypic bluetongue viruses, whereas genome segments S1, S4, S5, S7-S10 were typically more closely related to the cognate genes of heterotypic isolates. Importantly, in many gene phylogenies the Hungarian BTV-4 strain showed genetic relationship to BTV strains identified in outbreaks in the western Mediterranean basin. Our results indicate the identified Hungarian bluetongue virus strain evolved through reassortment involving multiple genome segments from various heterotypic bluetongue viruses.

Full genome characterization of human Rotavirus A strains isolated in Cameroon, 2010-2011: diverse combinations of the G and P genes and lack of reassortment of the backbone genes.

Over the past few years whole genome sequencing of rotaviruses has become a routine laboratory method in many strain surveillance studies. To study the molecular evolutionary pattern of representative Cameroonian Rotavirus A (RVA) strains, the semiconductor sequencing approach was used following random amplification of genomic RNA. In total, 31 RVA strains collected during 2010-2011 in three Cameroonian study sites located 120 to 1240 km from each other were sequenced and analyzed. Sequence analysis of the randomly selected representative strains showed that 18 RVAs were Wa-like, expressing G1P[6], G12P[6], or G12P[8] neutralization antigens on the genotype 1 genomic constellation (I1-R1-C1-M1-A1-N1-T1-E1-H1), whereas 13 other strains were DS-1-like, expressing G2P[4], G2P[6], G3P[6], and G6P[6] on the genotype 2 genomic constellation (I2-R2-C2-M2-A2-N2-T2-E2-H2). No inter-genogroup reassortment in the backbone genes was observed. Phylogenetic analysis of the Cameroonian G6P[6] strains indicated the separation of the strains identified in the Far North region (Maroua) and the Northwest region (Bamenda and Esu) into two branches that is consistent with multiple introductions of G6P[6] strains into this country. The present whole genome based molecular characterization study indicates that the emerging G6P[6] strain is fully heterotypic to Rotarix, the vaccine introduced during 2014 in childhood immunization program in Cameroon. Continuous strain monitoring is therefore needed in this area and elsewhere to see if G6s, besides genotype G1 to G4, G8, G9 and G12, may become a new, regionally important genotype in the post vaccine licensure era in Africa.

Large-scale whole genome sequencing identifies country-wide spread of an emerging G9P[8] rotavirus strain in Hungary, 2012.

With the availability of rotavirus vaccines routine strain surveillance has been launched or continued in many countries worldwide. In this study relevant information is provided from Hungary in order to extend knowledge about circulating rotavirus strains. Direct sequencing of the RT-PCR products obtained by VP7 and VP4 genes specific primer sets was utilized as routine laboratory method. In addition we explored the advantage of random primed RT-PCR and semiconductor sequencing of the whole genome of selected strains. During the study year, 2012, we identified an increase in the prevalence of G9P[8] strains across the country. This genotype combination predominated in seven out of nine study sites (detection rates, 45-83%). In addition to G9P[8]s, epidemiologically major strains included genotypes G1P[8] (34.2%), G2P[4] (13.5%), and G4P[8] (7.4%), whereas unusual and rare strains were G3P[8] (1%), G2P[8] (0.5%), G1P[4] (0.2%), G3P[4] (0.2%), and G3P[9] (0.2%). Whole genome analysis of 125 Hungarian human rotaviruses identified nine major genotype constellations and uncovered both intra- and intergenogroup reassortment events in circulating strains. Intergenogroup reassortment resulted in several unusual genotype constellations, including mono-reassortant G1P[8] and G9P[8] strains whose genotype 1 (Wa-like) backbone gene constellations contained DS1-like NSP2 and VP3 genes, respectively, as well as, a putative bovine-feline G3P[9] reassortant strain. The conserved genomic constellations of epidemiologically major genotypes suggested the clonal spread of the re-emerging G9P[8] genotype and several co-circulating strains (e.g., G1P[8] and G2P[4]) in many study sites during 2012. Of interest, medically important G2P[4] strains carried bovine-like VP1 and VP6 genes in their genotype constellation. No evidence for vaccine associated selection, or, interaction between wild-type and vaccine strains was obtained. In conclusion, this study reports the reemergence of G9P[8] strains across the country and indicates the robustness of whole genome sequencing in routine rotavirus strain surveillance.

Review of global rotavirus strain prevalence data from six years post vaccine licensure surveillance: is there evidence of strain selection from vaccine pressure?

Comprehensive reviews of pre licensure rotavirus strain prevalence data indicated the global importance of six rotavirus genotypes, G1P[8], G2P[4], G3P[8], G4P[8], G9P[8] and G12P[8]. Since 2006, two vaccines, the monovalent Rotarix (RV1) and the pentavalent RotaTeq (RV5) have been available in over 100 countries worldwide. Of these, 60 countries have already introduced either RV1 or RV5 in their national immunization programs. Post licensure vaccine effectiveness is closely monitored worldwide. This review aimed at describing the global changes in rotavirus strain prevalence over time. The genotype distribution of the nearly 47,000 strains that were characterized during 2007-2012 showed similar picture to that seen in the preceding period. An intriguing finding was the transient predominance of heterotypic strains, mainly in countries using RV1. Unusual and novel antigen combinations continue to emerge, including some causing local outbreaks, even in vaccinated populations. In addition, vaccine strains have been found in both vaccinated infants and their contacts and there is evidence for genetic interaction between vaccine and wild-type strains. In conclusion, the post-vaccine introduction strain prevalence data do not show any consistent pattern indicative of selection pressure resulting from vaccine use, although the increased detection rate of heterotypic G2P[4] strains in some countries following RV1 vaccination is unusual and this issue requires further monitoring.