Significance of the Gut Microbiome for Viral Diarrheal and Extra-Intestinal Diseases.

The composition of the mammalian gut microbiome is very important for the health and disease of the host. Significant correlations of particular gut microbiota with host immune responsiveness and various infectious and noninfectious host conditions, such as chronic enteric infections, type 2 diabetes, obesity, asthma, and neurological diseases, have been uncovered. Recently, research has moved on to exploring the causalities of such relationships. The metabolites of gut microbiota and those of the host are considered in a 'holobiontic' way. It turns out that the host's diet is a major determinant of the composition of the gut microbiome and its metabolites. Animal models of bacterial and viral intestinal infections have been developed to explore the interrelationships of diet, gut microbiome, and health/disease phenotypes of the host. Dietary fibers can act as prebiotics, and certain bacterial species support the host's wellbeing as probiotics. In cases of Clostridioides difficile-associated antibiotic-resistant chronic diarrhea, transplantation of fecal microbiomes has sometimes cured the disease. Future research will concentrate on the definition of microbial/host/diet interrelationships which will inform rationales for improving host conditions, in particular in relation to optimization of immune responses to childhood vaccines

Viroplasms: Assembly and Functions of Rotavirus Replication Factories.

Viroplasms are cytoplasmic, membraneless structures assembled in rotavirus (RV)-infected cells, which are intricately involved in viral replication. Two virus-encoded, non-structural proteins, NSP2 and NSP5, are the main drivers of viroplasm formation. The structures (as far as is known) and functions of these proteins are described. Recent studies using plasmid-only-based reverse genetics have significantly contributed to elucidation of the crucial roles of these proteins in RV replication. Thus, it has been recognized that viroplasms resemble liquid-like protein-RNA condensates that may be formed via liquid-liquid phase separation (LLPS) of NSP2 and NSP5 at the early stages of infection. Interactions between the RNA chaperone NSP2 and the multivalent, intrinsically disordered protein NSP5 result in their condensation (protein droplet formation), which plays a central role in viroplasm assembly. These droplets may provide a unique molecular environment for the establishment of inter-molecular contacts between the RV (+)ssRNA transcripts, followed by their assortment and equimolar packaging. Future efforts to improve our understanding of RV replication and genome assortment in viroplasms should focus on their complex molecular composition, which changes dynamically throughout the RV replication cycle, to support distinct stages of virion assembly

Whole genome analysis of selected human and animal rotaviruses identified in Uganda from 2012 to 2014 reveals complex genome reassortment events between human, bovine, caprine and porcine strains

Rotaviruses of species A (RVA) are a common cause of diarrhoea in children and the young of various other mammals and birds worldwide. To investigate possible interspecies transmission of RVAs, whole genomes of 18 human and 6 domestic animal RVA strains identified in Uganda between 2012 and 2014 were sequenced using the Illumina HiSeq platform. The backbone of the human RVA strains had either a Wa- or a DS-1-like genetic constellation. One human strain was a Wa-like mono-reassortant containing a DS-1-like VP2 gene of possible animal origin. All eleven genes of one bovine RVA strain were closely related to those of human RVAs. One caprine strain had a mixed genotype backbone, suggesting that it emerged from multiple reassortment events involving different host species. The porcine RVA strains had mixed genotype backbones with possible multiple reassortant events with strains of human and bovine origin.Overall, whole genome characterisation of rotaviruses found in domestic animals in Uganda strongly suggested the presence of human-to animal RVA transmission, with concomitant circulation of multi-reassortant strains potentially derived from complex interspecies transmission events. However, whole genome data from the human RVA strains causing moderate and severe diarrhoea in under-fives in Uganda indicated that they were primarily transmitted from person-to-person

Intracellular neutralisation of rotavirus by VP6-specific IgG

Funder: Wellcome Trust; funder-id: http://dx.doi.org/10.13039/100004440Rotavirus is a major cause of gastroenteritis in children, with infection typically inducing high levels of protective antibodies. Antibodies targeting the middle capsid protein VP6 are particularly abundant, and as VP6 is only exposed inside cells, neutralisation must be post-entry. However, while a system of poly immune globulin receptor (pIgR) transcytosis has been proposed for anti-VP6 IgAs, the mechanism by which VP6-specific IgG mediates protection remains less clear. We have developed an intracellular neutralisation assay to examine how antibodies neutralise rotavirus inside cells, enabling comparison between IgG and IgA isotypes. Unexpectedly we found that neutralisation by VP6-specific IgG was much more efficient than by VP6-specific IgA. This observation was highly dependent on the activity of the cytosolic antibody receptor TRIM21 and was confirmed using an in vivo model of murine rotavirus infection. Furthermore, mice deficient in only IgG and not other antibody isotypes had a serious deficit in intracellular antibody-mediated protection. The finding that VP6-specific IgG protect mice against rotavirus infection has important implications for rotavirus vaccination. Current assays determine protection in humans predominantly by measuring rotavirus-specific IgA titres. Measurements of VP6-specific IgG may add to existing mechanistic correlates of protection

Co-Surveillance of Rotaviruses in Humans and Domestic Animals in Central Uganda Reveals Circulation of Wide Genotype Diversity in the Animals

Rotavirus genotypes are species specific. However, interspecies transmission is reported to result in the emergence of new genotypes. A cross-sectional study of 242 households with 281 cattle, 418 goats, 438 pigs, and 258 humans in Uganda was undertaken between 2013 and 2014. The study aimed to determine the prevalence and genotypes of rotaviruses across co-habiting host species, as well as potential cross-species transmission. Rotavirus infection in humans and animals was determined using NSP3 targeted RT-PCR and ProSpecT Rotavirus ELISA tests, respectively. Genotyping of rotavirus-positive samples was by G- and P-genotype specific primers in nested RT-PCR assays while genotyping of VP4 and VP7 proteins for the non-typeable human positive sample was done by Sanger sequencing. Mixed effect logistic regression was used to determine the factors associated with rotavirus infection in animals. The prevalence of rotavirus was 4.1% (95% CI: 3.0-5.5%) among the domestic animals and 0.8% (95% CI: 0.4-1.5%) in humans. The genotypes in human samples were G9P[8] and P[4]. In animals, six G-genotypes, G3(2.5%), G8(10%), G9(10%), G11(26.8%), G10(35%), and G12(42.5%), and nine P-genotypes, P[1](2.4%), P[4](4.9%), P[5](7.3%), P[6](14.6%), P[7](7.3%), P[8](9.8%), P[9](9.8%), P[10](12.2%), and P[11](17.1%), were identified. Animals aged 2 to 18 months were less likely to have rotavirus infection in comparison with animals below 2 months of age. No inter-host species transmission was identified

Physicochemical analysis of rotavirus segment 11 supports a 'modified panhandle' structure and not the predicted alternative tRNA-like structure (TRLS)

.Rotaviruses are a major cause of acute gastroenteritis, which is often fatal in infants. The viral genome consists of 11 double-stranded RNA segments, but little is known about their cis-acting sequences and structural elements. Covariation studies and phylogenetic analysis exploring the potential structure of RNA11 of rotaviruses suggested that, besides the previously predicted "modified panhandle" structure, the 5' and 3' termini of one of the isoforms of the bovine rotavirus UKtc strain may interact to form a tRNA-like structure (TRLS). Such TRLSs have been identified in RNAs of plant viruses, where they are important for enhancing replication and packaging. However, using tRNA mimicry assays (in vitro aminoacylation and 3'- adenylation), we found no biochemical evidence for tRNA-like functions of RNA11. Capping, synthetic 3' adenylation and manipulation of divalent cation concentrations did not change this finding. NMR studies on a 5'- and 3'-deletion construct of RNA11 containing the putative intra-strand complementary sequences supported a predominant panhandle structure and did not conform to a cloverleaf fold despite the strong evidence for a predicted structure in this conserved region of the viral RNA. Additional viral or cellular factors may be needed to stabilise it into a form with tRNA-like properties

Lipidome analysis of rotavirus-infected cells confirms the close interaction of lipid droplets with viroplasms

10.1099/vir.0.049635-0Journal of General Virology94PART71576-158

Rotavirus infection

Q1Q1Artículo original1-16Rotavirus infections are a leading cause of severe, dehydrating gastroenteritis in children 200,000 deaths annually, mostly in low-income countries. Rotavirus primarily infects enterocytes and induces diarrhoea through the destruction of absorptive enterocytes (leading to malabsorption), intestinal secretion stimulated by rotavirus non-structural protein 4 and activation of the enteric nervous system. In addition, rotavirus infections can lead to antigenaemia (which is associated with more severe manifestations of acute gastroenteritis) and viraemia, and rotavirus can replicate in systemic sites, although this is limited. Reinfections with rotavirus are common throughout life, although the disease severity is reduced with repeat infections. The immune correlates of protection against rotavirus reinfection and recovery from infection are poorly understood, although rotavirus-specific immunoglobulin A has a role in both aspects. The management of rotavirus infection focuses on the prevention and treatment of dehydration, although the use of antiviral and anti-emetic drugs can be indicated in some cases

Uniformity of rotavirus strain nomenclature proposed by the Rotavirus Classification Working Group (RCWG)

In April 2008, a nucleotide-sequence-based, complete genome classification system was developed for group A rotaviruses (RVs). This system assigns a specific genotype to each of the 11 genome segments of a particular RV strain according to established nucleotide percent cutoff values. Using this approach, the genome of individual RV strains are given the complete descriptor of Gx-P[x]-Ix-Rx-Cx-Mx-Ax-Nx-Tx-Ex-Hx. The Rotavirus Classification Working Group (RCWG) was formed by scientists in the field to maintain, evaluate and develop the RV genotype classification system, in particular to aid in the designation of new genotypes. Since its conception, the group has ratified 51 new genotypes: as of April 2011, new genotypes for VP7 (G20-G27), VP4 (P[28]-P[35]), VP6 (I12-I16), VP1 (R5-R9), VP2 (C6-C9), VP3 (M7-M8), NSP1 (A15-A16), NSP2 (N6-N9), NSP3 (T8-T12), NSP4 (E12-E14) and NSP5/6 (H7-H11) have been defined for RV strains recovered from humans, cows, pigs, horses, mice, South American camelids (guanaco), chickens, turkeys, pheasants, bats and a sugar glider. With increasing numbers of complete RV genome sequences becoming available, a standardized RV strain nomenclature system is needed, and the RCWG proposes that individual RV strains are named as follows: RV group/species of origin/country of identification/common name/year of identification/G- and P-type. In collaboration with the National Center for Biotechnology Information (NCBI), the RCWG is also working on developing a RV-specific resource for the deposition of nucleotide sequences. This resource will provide useful information regarding RV strains, including, but not limited to, the individual gene genotypes and epidemiological and clinical information. Together, the proposed nomenclature system and the NCBI RV resource will offer highly useful tools for investigators to search for, retrieve, and analyze the ever-growing volume of RV genomic data.Fil: Matthijnssens, Jelle. Katholikie Universiteit Leuven; BélgicaFil: Ciarlet, Max. Novartis Vaccines & Diagnostics; Estados UnidosFil: McDonald, Sarah M.. National Institute Of Allegry & Infectious Diseases (niaid) ; National Institutes Of Health;Fil: Attoui, Houssam. Animal Health Trust.; Reino UnidoFil: Bányai, Krisztián. Hungarian Academy of Sciences; HungríaFil: Brister, J. Rodney. National Library Of Medicine; Estados UnidosFil: Buesa, Javier. Universidad de Valencia; EspañaFil: Esona, Mathew D.. Centers for Disease Control and Prevention; Estados UnidosFil: Estes, Mary K.. Baylor College of Medicine; Estados UnidosFil: Gentsch, Jon R.. Centers for Disease Control and Prevention; Estados UnidosFil: Iturriza Gómara, Miren. Health Protection Agency; Reino UnidoFil: Johne, Reimar. Federal Institute for Risk Assessment; AlemaniaFil: Kirkwood, Carl D.. Royal Children's Hospital; AustraliaFil: Martella, Vito. Università degli Studi di Bari; ItaliaFil: Mertens, Peter P. C.. Animal Health Trust.; Reino UnidoFil: Nakagomi, Osamu. Nagasaki University; JapónFil: Parreño, Gladys Viviana. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigación en Ciencias Veterinarias y Agronómicas. Instituto de Virología; ArgentinaFil: Rahman, Mustafizur. International Centre For Diarrhoeal Disease Research; BangladeshFil: Ruggeri, Franco M.. Istituto Superiore Di Sanita; ItaliaFil: Saif, Linda J.. Ohio State University; Estados UnidosFil: Santos, Norma. Universidade Federal do Rio de Janeiro; BrasilFil: Steyer, Andrej. University of Ljubljan; EsloveniaFil: Taniguchi, Koki. Fujita Health University School of Medicine; JapónFil: Patton, John T.. National Institute Of Allegry & Infectious Diseases (niaid) ; National Institutes Of Health;Fil: Desselberger, Ulrich. University of Cambridge; Estados UnidosFil: van Ranst, Marc. Katholikie Universiteit Leuven; Bélgic

Genomic analysis of codon, sequence and structural conservation with selective biochemical-structure mapping reveals highly conserved and dynamic structures in rotavirus RNAs with potential cis-acting functions

Rotaviruses are a major cause of acute, often fatal, gastroenteritis in infants and young children world-wide. Virions contain an 11 segment double-stranded RNA genome. Little is known about the cis-acting sequences and structural elements of the viral RNAs. Using a database of 1621 full-length sequences of mammalian group A rotavirus RNA segments, we evaluated the codon, sequence and RNA structural conservation of the complete genome. Codon conservation regions were found in eight ORFs, suggesting the presence of functional RNA elements. Using ConStruct and RNAz programmes, we identified conserved secondary structures in the positive-sense RNAs including long-range interactions (LRIs) at the 5′ and 3′ terminal regions of all segments. In RNA9, two mutually exclusive structures were observed suggesting a switch mechanism between a conserved terminal LRI and an independent 3′ stem–loop structure. In RNA6, a conserved stem–loop was found in a region previously reported to have translation enhancement activity. Biochemical structural analysis of RNA11 confirmed the presence of terminal LRIs and two internal helices with high codon and sequence conservation. These extensive in silico and in vitro analyses provide evidence of the conservation, complexity, multi-functionality and dynamics of rotavirus RNA structures which likely influence RNA replication, translation and genome packaging