Rotavirus VP7 Research Articles

In Silico screening of T-cell and B-cell Epitopes of Rotavirus VP7 and VP4 proteins for Effective Vaccine Design

Rotavirus is one of the deadliest causative agents of childhood diarrhea which causes half a million child death across the globe, mostly in developing countries. However, effective vaccine strategies against rotavirus are yet to be established to prevent these unwanted premature deaths. In this regard, in silico vaccine design for rotavirus could be a promising alternative for developing countries due to its efficiency in shortening valuable time and cost. The present study described an epitope-based peptide vaccine design against rotavirus, using a combination of T-cell and B-cell epitope predictions and molecular docking approach. To perform this, sequences of rotavirus VP7 and VP4 proteins were retrieved from the NCBI database and subjected to different bioinformatics tools to predict most immunogenic T-cell and B-cell epitopes. From the identified epitopes, the sequence VMSKRSRSL of VP7 and TQFTDFVSL of VP4 was identified as the most potential epitopes based on their antigenicity, conservancy and interaction with major histocompatability complex I (MHC-I) alleles. Moreover, the peptide VMSKRSRSL interacted with human leukocyte antigen, HLA-B*08:01 and TQFTDFVSL interacted with HLA-A*02:06 with considerable binding energy and affinity score. Combined population coverage for our identified epitopes was found 70.53% and 45.64% for world population and South Asian population respectively. All these results suggest that, the epitopes identified in this study could be a very good vaccine candidate for the strains of rotavirus circulating in Bangladesh. However, as this study is completely dependent on computational prediction algorithms, further in vivo screening is required to come up in a precise conclusion about these epitopes for effective rotavirus vaccination.
 Bangladesh J Microbiol, Volume 35 Number 1 June 2018, pp 45-55

Rotavirus VP6 as a potential vaccine candidate.

By the age of 5years, virtually all children have been infected by group A rotavirus (RVA), which is responsible for around half million mortality annually prior to vaccination. Relatively high rate of the morbidity and mortality highlights the necessity of applying preventive procedures particularly in developing countries. Two live attenuated RVA vaccines (Rotarix and RotaTeq) are licensed and now being used in many countries worldwide. Although these vaccines are shown to reduce the mortality up to 50%, several key questions yet remained to answer. Indeed, the licensed RV vaccines were found to be less effective in countries of sub-Saharan Africa and Southeast Asia. Therefore, developing next generation RVA vaccines is warranted. VP6 is highly abundant and conserved protein that forms the middle layer of RV particles and was shown to be both antigenic and immunogenic. Although it does not induce neutralizing antibodies, different VP6 preparations were found to induce homologous and cross-reactive immune responses with partial protection from RVA replication. Although the molecular mechanisms are not fully elucidated, VP6-based RVA vaccine candidates are worthy of further consideration. This review aims to focus on different aspects of VP6 protein and its potentiality for an alternative RV vaccine against RV disease.

The C Terminus of Rotavirus VP4 Protein Contains an Actin Binding Domain Which Requires Cooperation with the Coiled-Coil Domain for Actin Remodeling.

The interactions between viruses and actin cytoskeleton have been widely studied. We showed that rotaviruses remodel microfilaments in intestinal cells and demonstrated that this was due to the VP4 spike protein. Microfilaments mainly occur in the apical domain of infected polarized enterocytes and favor the polarized apical exit of viral progeny. The present work aims at the identification of molecular determinants of actin-VP4 interactions. We used various deletion mutants of VP4 that were transfected into Cos-7 cells and analyzed interactions by immunofluorescence confocal microscopy. It has been established that the C-terminal part of VP4 is embedded within viral particles when rotavirus assembles. The use of specific monoclonal antibodies demonstrated that VP4 is expressed in different forms in infected cells: classically as spike on the outer layer of virus particles, but also as free soluble protein in the cytosol. The C terminus of free VP4 was identified as interacting with actin microfilaments. The VP4 actin binding domain is unable to promote microfilament remodeling by itself; the coiled-coil domain is also required in this process. This actin-binding domain was shown to dominate a previously identified peroxisomal targeting signal, located in the three last amino acids of VP4. The newly identified actin-binding domain is highly conserved in rotavirus strains from species A, B, and C, suggesting that actin binding and remodeling is a general strategy for rotavirus exit. This provides a novel mechanism of protein-protein interactions, not involving cell signaling pathways, to facilitate rotavirus exit.IMPORTANCE Rotaviruses are causal agents of acute infantile viral diarrhea. In intestinal cells, in vitro as well as in vivo, virus assembly and exit do not imply cell lysis but rely on an active process in which the cytoskeleton plays a major role. We describe here a novel molecular mechanism by which the rotavirus spike protein VP4 drives actin remodeling. This relies on the fact that VP4 occurs in different forms. Besides its structural function within the virion, a large proportion of VP4 is expressed as free protein. Here, we show that free VP4 possesses a functional actin-binding domain. This domain, in coordination with a coiled-coil domain, promotes actin cytoskeleton remodeling, thereby providing the capacity to destabilize the cell membrane and allow efficient rotavirus exit.

Rotavirus VP3 targets MAVS for degradation to inhibit type III interferon expression in intestinal epithelial cells.

Rotaviruses (RVs), a leading cause of severe diarrhea in young children and many mammalian species, have evolved multiple strategies to counteract the host innate immunity, specifically interferon (IFN) signaling through RV non-structural protein 1 (NSP1). However, whether RV structural components also subvert antiviral response remains under-studied. Here, we found that MAVS, critical for the host RNA sensing pathway upstream of IFN induction, is degraded by the RV RNA methyl- and guanylyl-transferase (VP3) in a host-range-restricted manner. Mechanistically, VP3 localizes to the mitochondria and mediates the phosphorylation of a previously unidentified SPLTSS motif within the MAVS proline-rich region, leading to its proteasomal degradation and blockade of IFN-λ production in RV-infected intestinal epithelial cells. Importantly, VP3 inhibition of MAVS activity contributes to enhanced RV replication and to viral pathogenesis in vivo. Collectively, our findings establish RV VP3 as a viral antagonist of MAVS function in mammals and uncover a novel pathogen-mediated inhibitory mechanism of MAVS signaling.

Isolation and characterization of a new candidate human inactivated rotavirus vaccine strain from hospitalized children in Yunnan, China: 2010-2013.

AIMTo determine the distribution of rotavirus VP7 gene in hospitalized children in Yunnan, China.METHODSA total of 366 stool specimens were collected from hospitalized children in hospitals in Yunnan Province from September 2010 to December 2013. The genomic RNA electropherotypes and the G genotypes of the rotaviruses were determined. A phylogenetic analysis of the VP7 gene was performed. Rotavirus isolation was performed, and characterized by plaque, minimum essential medium, and all genes sequence analysis. Quantification of antibodies for inactivated vaccine prepared with ZTR-68 was examined by enzyme-linked immunosorbent assay and microneutralization assay.RESULTSGroup A human rotavirus was detected in 177 of 366 (48.4%) stool samples using a colloidal gold device assay. The temporal distribution of rotavirus cases showed significant correlation with the mean air temperature. Rotaviruses were isolated from 13% of the rotavirus-positive samples. The predominant genotype was G1 (43.5%), followed by G3 (21.7%), G9 (17.4%), G2 (4.3%), G4 (8.7%), and mixed (4.3%) among a total of 23 rotavirus isolates. A rotavirus strain was isolated from a rotavirus-positive stool sample of a 4-month-old child in The First People’s Hospital of Zhaotong (2010) for use as a candidate human inactivated rotavirus vaccine strain and for further research, and was designated ZTR-68. The genotype of 11 gene segments of strain ZTR-68 (RVA/Human-wt/CHN/ZTR-68/2010/G1P[8]) was characterized. The genotype constellation of strain ZTR-68 was identified as G1-P[8]-I1-R1-C1-M1-A1-N1-T1-E1-H1. The VP7 and VP4 genotypes of strain ZTR-68 were similar to Wa-like strains.CONCLUSIONSA high prevalence of the G1, G2, and G3 genotypes was detected from 2010 to 2012. However, a dominant prevalence of the G9 genotype was identified as the cause of gastroenteritis in children in Yunnan, China, in 2013. A candidate human inactivated rotavirus vaccine strain, designated ZTR-68 was isolated, characterized, and showed immunogenicity. Our data will be useful for the future formulation and development of a vaccine in China.

Bioengineered Norovirus S60 Nanoparticles as a Multifunctional Vaccine Platform.

Homotypic interactions of viral capsid proteins are common, driving viral capsid self-formation. By taking advantage of such interactions of the norovirus shell (S) domain that naturally builds the interior shells of norovirus capsids, we have developed a technology to produce 60-valent, icosahedral S60 nanoparticles through the E. coli system. This has been achieved by several modifications to the S domain, including an R69A mutation to destruct an exposed proteinase cleavage site and triple cysteine mutations (V57C/Q58C/S136C) to establish inter-S domain disulfide bonds for enhanced inter-S domain interactions. The polyvalent S60 nanoparticle with 60 exposed S domain C-termini offers an ideal platform for antigen presentation, leading to enhanced immunogenicity to the surface-displayed antigens for vaccine development. This was proven by constructing a chimeric S60 nanoparticle displaying 60 rotavirus (RV) VP8* proteins, the major RV-neutralizing antigen. These S60-VP8* particles are easily produced and elicited high IgG response in mice toward the displayed VP8* antigens. The mouse antisera after immunization with the S60-VP8* particles exhibited high blockades against RV VP8* binding to its glycan ligands and high neutralizing activities against RV infection in culture cells. The three-dimensional structures of the S60 and S60-VP8* particles were studied. Furthermore, the S60 nanoparticle can display other antigens, supporting the notion that the S60 nanoparticle is a multifunctional vaccine platform. Finally, the intermolecular disulfide bond approach may be used to stabilize other viral-like particles to display foreign antigens for vaccine development.

Rotavirus VP6 protein mucosally delivered by cell wall-derived particles from Lactococcus lactis induces protection against infection in a murine model.

Rotaviruses are the primary cause of acute gastroenteritis in children worldwide. Although the implementation of live attenuated vaccines has reduced the number of rotavirus-associated deaths, variance in their effectiveness has been reported in different countries. This fact, among other concerns, leads to continuous efforts for the development of new generation of vaccines against rotavirus.In this work, we describe the obtention of cell wall-derived particles from a recombinant Lactococcus lactis expressing a cell wall-anchored version of the rotavirus VP6 protein. After confirming by SDS-PAGE, Western blot, flow cytometry and electronic immunomicroscopy that these particles were carrying the VP6 protein, their immunogenic potential was evaluated in adult BALB/c mice. For that, mucosal immunizations (oral or intranasal), with or without the dmLT [(double mutant Escherichia coli heat labile toxin LT(R192G/L211A)] adjuvant were performed. The results showed that these cell wall-derived particles were able to generate anti-rotavirus IgG and IgA antibodies only when administered intranasally, whether the adjuvant was present or not. However, the presence of dmLT was necessary to confer protection against rotavirus infection, which was evidenced by a 79.5 percent viral shedding reduction.In summary, this work describes the production of cell wall-derived particles which were able to induce a protective immune response after intranasal immunization. Further studies are needed to characterize the immune response elicited by these particles as well as to determine their potential as an alternative to the use of live L. lactis for mucosal antigen delivery.

Emergence of G8P[6] rotavirus strains in Korean neonates

BackgroundRotaviruses are the major causes of pediatric gastroenteritis worldwide. The genotypic distribution of rotavirus strains shows temporal and geographical fluctuations, and knowledge of the molecular epidemiology of rotaviruses is important for the development of vaccines and diagnostic reagents. We investigated VP4 and VP7 capsid genotypes of rotaviruses isolated from 211 stool specimens collected from Korean neonates in a neonatal intensive care unit from September 2017 to March 2018.ResultsOf 211 stool specimens, 15 specimens (7.1%) were rotavirus-positive. Eleven specimens (73.3%) were G8P[6] type and 4 (26.7%) were G4P[6] type. Sequence analysis revealed that all G8 sequences in this study showed the highest nucleotide identity to G8 sequences of G8P[8] rotavirus strains isolated in Vietnam in 2014, and P[6] gene sequences showed the highest nucleotide identity to P[6] sequences of G4P[6] strains detected in Korea in 2012. Only one amino acid difference in VP7 was found in 3 of the 11 G8P[6] strains in this study, but multiple amino acid substitutions in VP7 were detected between these G8P[6] strains and the commonly used vaccine strains.ConclusionsThis study showed that rotavirus G8P[6] strains were firstly detected at high frequency in Korean neonates from September 2017 to March 2018. These new rotavirus G8P[6] strains were estimated to be derived from reassortment events between the G8 of G8P[8] strains in Asian region and the P[6] of G4[6] in Korea. Whether the emergence of this unusual G8P[6] strain reflects continuous prevalence or transient occurrence will require continuous monitoring of rotavirus epidemiology.

Glycan Binding Specificity and Mechanism of Human and Porcine P[6]/P[19] Rotavirus VP8*s.

Rotaviruses (RVs), which cause severe gastroenteritis in infants and children, recognize glycan ligands in a genotype-dependent manner via the distal VP8* head of the spike protein VP4. However, the glycan binding mechanisms remain elusive for the P[II] genogroup RVs, including the widely prevalent human RVs (P[8], P[4], and P[6]) and a rare P[19] RV. In this study, we characterized the glycan binding specificities of human and porcine P[6]/P[19] RV VP8*s and found that the P[II] genogroup RV VP8*s could commonly interact with mucin core 2, which may play an important role in RV evolution and cross-species transmission. We determined the first P[6] VP8* structure, as well as the complex structures of human P[19] VP8*, with core 2 and lacto-N-tetraose (LNT). A glycan binding site was identified in human P[19] VP8*. Structural superimposition and sequence alignment revealed the conservation of the glycan binding site in the P[II] genogroup RV VP8*s. Our data provide significant insight into the glycan binding specificity and glycan binding mechanism of the P[II] genogroup RV VP8*s, which could help in understanding RV evolution, transmission, and epidemiology and in vaccine development.IMPORTANCE Rotaviruses (RVs), belonging to the family Reoviridae, are double-stranded RNA viruses that cause acute gastroenteritis in children and animals worldwide. Depending on the phylogeny of the VP8* sequences, P[6] and P[19] RVs are grouped into genogroup II, together with P[4] and P[8], which are widely prevalent in humans. In this study, we characterized the glycan binding specificities of human and porcine P[6]/P[19] RV VP8*s, determined the crystal structure of P[6] VP8*, and uncovered the glycan binding pattern in P[19] VP8*, revealing a conserved glycan binding site in the VP8*s of P[II] genogroup RVs by structural superimposition and sequence alignment. Our data suggested that mucin core 2 may play an important role in P[II] RV evolution and cross-species transmission. These data provide insight into the cell attachment, infection, epidemiology, and evolution of P[II] genogroup RVs, which could help in developing control and prevention strategies against RVs.

Intradermal and intranasal immunizations with oligomeric middle layer rotavirus VP6 induce Th1, Th2 and Th17 T cell subsets and CD4+ T lymphocytes with cytotoxic potential

Rotavirus (RV) inner capsid VP6 protein is a potential non-live vaccine candidate due to high degree of conservation and immunogenicity, and ability to self-assemble into oligomeric structures, including nanotubes. These VP6 structures induce strong humoral and T cell immunity and protect mice against RV challenge. It has been suggested that intracellular neutralization by IgA antibody and VP6-specific CD4+ T cells mediate protection. We investigated generation of diverse CD4+ T cell subsets by intradermal and intranasal delivery of recombinant VP6 (rVP6) nanotubes in BALB/c mice. Production of antiviral cytokine interferon-γ (IFN-γ), interleukin-4 (IL-4) and pro-inflammatory cytokine IL-17 was analyzed following in vitro stimulation of immune cells. Cell surface CD107a expression was measured to determine VP6-specific cytotoxic T cells. Both parenteral and mucosal immunization with oligomeric rVP6 induced VP6-specific Th1, Th2 and Th17 cells. For the first time, cytotoxicity-related degranulation (CD107a surface expression) indicated that RV VP6-specific CD4+ T cells had cytotoxic T lymphocyte (CTL) phenotype. These findings demonstrate an ability of rVP6 nanostructures to induce heterogeneous CD4+ T cells with different effector functions, including CTLs with potential to lyse RV-infected cells, suggesting an additional mechanism of RV VP6-induced protection.

Design and testing of a highly conserved human rotavirus VP8* immunogenic peptide with potential for vaccine development

Rotavirus infection of young children particularly below five years of age resulting in severe diarhoea, is the cause of a large number of infant deaths all over the world, more so in developing countries like India. Vaccines developed against this infection in the last two decades have shown mixed results with some of them leading to complications. Oral vaccines have not been very effective in India. Significant diversity has been found in circulating virus strains in India. Development of a vaccine against diverse genetic variants of the different strains would go a long way in reducing the incidence of infection in developing countries.Success of such a vaccine would depend to a large extent on the antigenic peptide to be used in antibody production. The non-glycosylated protein VP4 on the surface capsid of the virus is important in rota viral immunogenicity and the major antigenic site(s) responsible for neutralization of the virus via VP4 is in the VP8* subunit of VP4. It is necessary that the peptide should be very specific and a peptide sequence which would stimulate both the T and B immunogenic cells would provide maximum protection against the virus. Advanced computational techniques and existing databases of sequences of the VP4 protein of rotavirus help in identification of such specific sequences. Using an in silico approach we have identified a highly conserved VP8* subunit of the VP4 surface protein of rotavirus which shows both T and B cell processivity and is also non-allergenic. This sub-unit could be used in in vivo models for induction of antibodies.

Genotypic characterization of rotavirus in children under 5 years circulating in C\xf4te D\u2019Ivoire from 2010 to 2013

BackgroundRotavirus infection is the most common cause of severe gastroenteritis in children under five years of age in both developed and developing countries. The World Health Organisation (WHO) recommends the surveillance of rotavirus strains prior to vaccine introduction in all applicable countries. The objective of this study was to describe the epidemiological characteristics as well as to determine the circulating genotypes of rotaviruses in Côte d’Ivoire prior to vaccine introduction.MethodsThe study included children under five years of age who met the inclusion criteria after informed consent had been sort from their parents or guardians. Rotavirus VP6 antigens were detected for each stool sample using Enzyme Immunoassay (EIA). Genotyping of positive EIA samples was performed by reverse-transcriptase-PCR (RT-PCR) assays.ResultsA total of 684 children were recruited. Children aged between 6 and 11 months were the most represented with 34%. Rotavirus VP6 antigens were found in 27.1% (186/684) of samples tested. Commonly detected G genotypes included G12 (46.6% (82/176) and G1 (13.1% (23/176) whilst P[8] (49.8% (91/183) was the most predominant P genotype. Rotavirus G12P[8] was the most predominant strain circulating in Côte d’Ivoire within the period of study and constituted 26.6% of all strains detected.ConclusionThe monitoring of circulating strains will help guide decision-makers in the choice of vaccine. Genotypic variability of circulating rotavirus strains over the years implies there is a need for continuous rotavirus strain surveillance even after vaccine introduction.

Specificity and affinity of neuraminic acid exhibited by canine rotavirus strain K9 carbohydrate-binding domain (VP8*).

The outer capsid spike protein VP4 of rotaviruses is a major determinant of infectivity and serotype specificity. Proteolytic cleavage of VP4 into 2 domains, VP8* and VP5*, enhances rotaviral infectivity. Interactions between the VP4 carbohydrate-binding domain (VP8*) and cell surface glycoconjugates facilitate initial virus-cell attachment and subsequent cell entry. Our saturation transfer difference nuclear magnetic resonance (STD NMR) and isothermal titration calorimetry (ITC) studies demonstrated that VP8*64-224 of canine rotavirus strain K9 interacts with N-acetylneuraminic and N-glycolylneuraminic acid derivatives, exhibiting comparable binding epitopes to VP8* from other neuraminidase-sensitive animal rotaviruses from pigs (CRW-8), cattle (bovine Nebraska calf diarrhoea virus, NCDV), and Rhesus monkeys (Simian rhesus rotavirus, RRV). Importantly, evidence was obtained for a preference by K9 rotavirus for the N-glycolyl- over the N-acetylneuraminic acid derivative. This indicates that a VP4 serotype 5A rotavirus (such as K9) can exhibit a neuraminic acid receptor preference that differs from that of a serotype 5B rotavirus (such as RRV) and the receptor preference of rotaviruses can vary within a particular VP4 genotype.

Human Group C Rotavirus VP8*s Recognize Type A Histo-Blood Group Antigens as Ligands.

Group/species C rotaviruses (RVCs) have been identified as important pathogens of acute gastroenteritis (AGE) in children, family-based outbreaks, as well as animal infections. However, little is known regarding their host-specific interaction, infection, and pathogenesis. In this study, we performed serial studies to characterize the function and structural features of a human G4P[2] RVC VP8* that is responsible for the host receptor interaction. Glycan microarrays demonstrated that the human RVC VP8* recognizes type A histo-blood group antigens (HBGAs), which was confirmed by synthetic glycan-/saliva-based binding assays and hemagglutination of red blood cells, establishing a paradigm of RVC VP8*-glycan interactions. Furthermore, the high-resolution crystal structure of the human RVC VP8* was solved, showing a typical galectin-like structure consisting of two β-sheets but with significant differences from cogent proteins of group A rotaviruses (RVAs). The VP8* in complex with a type A trisaccharide displays a novel ligand binding site that consists of a particular set of amino acid residues of the C-D, G-H, and K-L loops. RVC VP8* interacts with type A HBGAs through a unique mechanism compared with that used by RVAs. Our findings shed light on the host-virus interaction and the coevolution of RVCs and will facilitate the development of specific antivirals and vaccines.IMPORTANCE Group/species C rotaviruses (RVCs), members of Reoviridae family, infect both humans and animals, but our knowledge about the host factors that control host susceptibility and specificity is rudimentary. In this work, we characterized the glycan binding specificity and structural basis of a human RVC that recognizes type A HBGAs. We found that human RVC VP8*, the rotavirus host ligand binding domain that shares only ∼15% homology with the VP8* domains of RVAs, recognizes type A HBGA at an as-yet-unknown glycan binding site through a mechanism distinct from that used by RVAs. Our new advancements provide insights into RVC-cell attachment, the critical step of virus infection, which will in turn help the development of control and prevention strategies against RVs.

Expression and characterization of a novel truncated rotavirus VP4 for the development of a recombinant rotavirus vaccine

The outer capsid protein VP4 is an important target for the development of a recombinant rotavirus vaccine because it mediates the attachment and penetration of rotavirus. Due to the poor solubility of full-length VP4, VP8 was explored as candidate rotavirus vaccines in the past years. In previous studies, it has been found that the N-terminal truncated VP8 protein, VP8-1 (aa26-231), could be expressed in soluble form with improved immunogenicity compared to the core of VP8 (aa65-223). However, this protein stimulated only a weak immune response when aluminum hydroxide was used as an adjuvant. In addition, it should be noted that the protective efficacy of VP4 was higher than that of VP8 and VP5. In this study, it was found that when the N-terminal 25 amino acids were deleted, the truncated VP4∗ (aa26-476) containing VP8 and the stalk domain of VP5 could be expressed in soluble form in E. coli and purified to homogeneous trimers. Furthermore, the truncated VP4 could induce high titers of neutralizing antibodies when aluminum adjuvant was used and conferred high protective efficacy in reducing the severity of diarrhea and rotavirus shedding in stools in animal models. The immunogenicity of the truncated VP4 was significantly higher than that of VP8∗ and VP5∗ alone. Taken together, the truncated VP4∗ (aa26-476), with enhanced immunogenicity and immunoprotectivity, could be considered as a viable candidate for further development and has the potential to become a parenterally administered rotavirus vaccine.

Evaluation of the immunogenicity of a recombinant HSV-1 vector expressing human group C rotavirus VP6 protein

Group C Rotavirus (RVC) has been associated globally with sporadic outbreaks of gastroenteritis in children and adults. RVC also infects animals, and interspecies transmission has been reported as well as its zoonotic potential. Considering its genetic diversity and the absence of effective vaccines, it is important and necessary to develop new generation vaccines against RVC for both humans and animals. The aim of the present study was to develop and characterize an HSV-1-based amplicon vector expressing a human RVC-VP6 protein and evaluate the humoral immune response induced after immunizing BALB/c mice. Local fecal samples positive for RVC were used for isolation and sequencing of the vp6 gene, which phylogenetically belongs to the I2 genotype. We show here that cells infected with the HSV[VP6C] amplicon vector efficiently express the VP6 protein, and induced specific anti-RVC antibodies in mice immunized with HSV[VP6C], in a prime-boost schedule. This work highlights that amplicon vectors are an attractive platform for the generation of safe genetic immunogens against RVC, without the addition of external adjuvants.

Rotavirus strain distribution in Ghana pre- and post- rotavirus vaccine introduction

BackgroundGhana introduced the monovalent rotavirus vaccine (Rotarix) into its national paediatric vaccination programme in May2012. Vaccine introduction was initiated nationwide and achieved >85% coverage within a few months. Rotavirus strain distribution pre- and post-RV vaccine introduction is reported. MethodsStool samples were collected from diarrhoeic children <5 years of age hospitalized between 2009 and 2016 at sentinel sites across Ghana and analyzed for the presence of group A rotavirus by enzyme immunoassay. Rotavirus strains were characterized by RT-PCR and sequencing. ResultsA total of 1363 rotavirus EIA-positive samples were subjected to molecular characterization. These were made up of 823 (60.4%) and 540 (39.6%) samples from the pre- and post-vaccine periods respectively. Rotavirus VP7 genotypes G1, G2 and G3, and VP4 genotypes P[6] and P[8] constituted more than 65% of circulating G and P types in the pre–vaccine period. The common strains detected were G1P[8] (20%), G3P[6] (9.2%) and G2P[6] (4.9%).During the post-vaccine period, G12, G1 and G10 genotypes, constituted more than 65% of the VP7 genotypes whilst P[6] and P[8] made up more than 75% of the VP4 genotypes. The predominant circulating strains were G12P[8] (26%), G10P[6] (10%) G3P[6] (8.1%) and G1P[8] (8.0%). We also observed the emergence of the unusual rotavirus strain G9P[4] during this period. ConclusionRotavirus G1P[8], the major strain in circulation during the pre-vaccination era, was replaced by G12P[8] as the most predominant strain after vaccine introduction. This strain replacement could be temporary and unrelated to vaccine introduction since an increase in G12 was observed in countries yet to introduce the rotavirus vaccine in West Africa. A continuous surveillance programme in the post-vaccine era is necessary for the monitoring of circulating rotavirus strains and the detection of unusual/emerging genotypes.

Viral metagenomics analysis of poultry faeces in live poultry market, Haikou, China

Objective: To conduct in-depth study of the distribution and diversity of viruses in poultry is of great importance in monitoring the emergence of interspecies transmission of novel viruses that may cause epidemics with public health significance. Poultry is an economically important source of meat, eggs and feathers which plays an important role as natural reservoirs of many pathogenic viruses. Compared with wild animals, poultry have more frequent interactions and therefore opportunities to transmit their viruses to human. Methods: To study the viromes of different types of poultry in Hainan, China, we used metagenomics for deep viral nucleic acid sequencing of the faecal samples collected from chickens, ducks and pigeons from a live poultry market in Haikou. Result: The poultry viromes were identified by sequence similarity comparisons of viral reads (BLASTxE score, <5) against viral reference database. A total of 15 309 viral reads were obtained, approximately 13 063, 1 370 and 876 viral reads were generated from the chicken, duck, pigeon faecal samples, respectively. The majority of the sequences were homologous to the animal virus of Adenoviridae, Herpeaviridae, Picobirnaviridae, Reoviridae, Retroviridae, Circoviridae, Paramyxoviridae, Astroviridae, Caliciviridae, Coronaviridae, Picornaviridae, and Orthomyxoviridae. The VP4 and VP7 segments of a pigeon rotavirus, similar to fox rotavirus in group A, were sequenced and phylogenetically analyzed. The near full genome of a pigeon circovirus was also analyzed. Conclusion: The major types of poultry in a Haikou harbor many different families of viruses in their feces which may have the potential for interspecies transmissions. Further studies should be conducted to identify the most prevalent and important viruses among a larger number of poultry in Haikou and other areas in Hainan.

Use of the Synthetic Gene Encoding the Truncated Human Rotavirus VP8* Protein in Escherichia coli for Production of Vaccine Candidates or Development of Diagnostic Antibodies

Use of the Synthetic Gene Encoding the Truncated Human Rotavirus VP8* Protein in Escherichia coli for Production of Vaccine Candidates or Development of Diagnostic Antibodies

Phylogenetic comparison of the VP7, VP4, VP6, and NSP4 genes of rotaviruses isolated from children in Nizhny Novgorod, Russia, 2015-2016, with cogent genes of the Rotarix and RotaTeq vaccine strains.

Group A rotaviruses (RVA) are one of the leading causes of gastroenteritis in young children worldwide. The introduction of universal mass vaccination around the world has contributed to a reduction in hospitalizations and outpatient visits associated with rotavirus infection. Continued surveillance of RVA strains is needed to determine long-term effects of vaccine introduction. In the present work, we carried out the analysis of the genotypic diversity of RVA strains isolated in Nizhny Novgorod (Russia) during the 2015-2016 epidemic season. Also we conducted a comparative analysis of the amino acid sequences of T-cell epitopes of wild-type and vaccine (RotaTeq and Rotarix) strains. In total, 1461 samples were examined. RVAs were detected in 30.4% of cases. Rotaviruses with genotype G9P[8] (40.5%) dominated in the 2015-16 epidemic season. Additionally, RVAs with the following genotypes were detected: G4P[8] (25.4%), G1P[8] (13%), G2P[4] (3.2%). Rotaviruses with genotypes G3P[9], G6P[9], and G1P[9] totaled 3%. The number of partially typed and untyped RVA samples was 66 (14.9%). The findings of a RVA of G6P[9] genotype in Russia were an original observation. Our analysis of VP6 and NSP4 T-cell epitopes showed highly conserved amino acid sequences. The found differences seem not to be caused by the immune pressure but were rather related to the genotypic affiliations of the proteins. Vaccination against rotavirus infection is not included in the national vaccination schedule in Russia. Monitoring of the genotypic and antigenic diversity of contemporary RVA will allow providing a comparative analysis of wild-type strains in areas with and without vaccine campaign.