Complete genome sequence of a highly divergent astrovirus isolated from a child with acute diarrhea

<p>Abstract</p> <p>Background</p> <p>Astroviruses infect a variety of mammals and birds and are causative agents of diarrhea in humans and other animal hosts. We have previously described the identification of several sequence fragments with limited sequence identity to known astroviruses in a stool specimen obtained from a child with acute diarrhea, suggesting that a novel virus was present.</p> <p>Results</p> <p>In this study, the complete genome of this novel virus isolate was sequenced and analyzed. The overall genome organization of this virus paralleled that of known astroviruses, with 3 open reading frames identified. Phylogenetic analysis of the ORFs indicated that this virus is highly divergent from all previously described animal and human astroviruses. Molecular features that are highly conserved in human serotypes 1–8, such as a 3'NTR stem-loop structure and conserved nucleotide motifs present in the 5'NTR and ORF1b/2 junction, were either absent or only partially conserved in this novel virus.</p> <p>Conclusion</p> <p>Based on the analyses described herein, we propose that this newly discovered virus represents a novel species in the family Astroviridae. It has tentatively been named Astrovirus MLB1.</p

Identification of a novel picornavirus related to cosaviruses in a child with acute diarrhea

Diarrhea, the third leading infectious cause of death worldwide, causes approximately 2 million deaths a year. Approximately 40% of these cases are of unknown etiology. We previously developed a metagenomic strategy for identification of novel viruses from diarrhea samples. By applying mass sequencing to a stool sample collected in Melbourne, Australia from a child with acute diarrhea, one 395 bp sequence read was identified that possessed only limited identity to known picornaviruses. This initial fragment shared only 55% amino acid identity to its top BLAST hit, the VP3 protein of Theiler's-like virus, suggesting that a novel picornavirus might be present in this sample. By using a combination of mass sequencing, RT-PCR, 5' RACE and 3' RACE, 6562 bp of the viral genome was sequenced, which includes the entire putative polyprotein. The overall genomic organization of this virus was similar to known picornaviruses. Phylogenetic analysis of the polyprotein demonstrated that the virus was divergent from previously described picornaviruses and appears to belong to the newly proposed picornavirus genus, Cosavirus. Based on the analysis discussed here, we propose that this virus represents a new species in the Cosavirus genus, and it has tentatively been named Human Cosavirus E1 (HCoSV-E1)

Rapid and Sensitive Detection of Rotavirus Molecular Signatures Using Surface Enhanced Raman Spectroscopy

Human enteric virus infections range from gastroenteritis to life threatening diseases such as myocarditis and aseptic meningitis. Rotavirus is one of the most common enteric agents and mortality associated with infection can be very significant in developing countries. Most enteric viruses produce diseases that are not distinct from other pathogens, and current diagnostics is limited in breadth and sensitivity required to advance virus detection schemes for disease intervention strategies. A spectroscopic assay based on surface enhanced Raman scattering (SERS) has been developed for rapid and sensitive detection of rotavirus. The SERS method relies on the fabrication of silver nanorod array substrates that are extremely SERS-active allowing for direct structural characterization of viruses. SERS spectra for eight rotavirus strains were analyzed to qualitatively identify rotaviruses and to classify each according to G and P genotype and strain with >96% accuracy, and a quantitative model based on partial least squares regression analysis was evaluated. This novel SERS-based virus detection method shows that SERS can be used to identify spectral fingerprints of human rotaviruses, and suggests that this detection method can be used for pathogen detection central to human health care

Klassevirus 1, a previously undescribed member of the family Picornaviridae, is globally widespread

Background Diarrhea is the third leading infectious cause of death worldwide and is estimated to be responsible for approximately 2 million deaths a year. While many infectious causes of diarrhea have been established, approximately 40% of all diarrhea cases are of unknown etiology. In an effort to identify novel viruses that may be causal agents of diarrhea, we used high throughput mass sequencing to analyze stool samples collected from patients with acute diarrhea. Results Sequences with limited similarity to known picornaviruses were detected in a stool sample collected in Australia from a child with acute diarrhea. Using a combination of mass sequencing, RT-PCR, 5' RACE and 3' RACE, a 6383 bp fragment of the viral genome was sequenced. Phylogenetic analysis demonstrated that this virus was highly divergent from, but most closely related to, members of the genus Kobuvirus. We have tentatively named this novel virus klassevirus 1. We also detected klassevirus 1 by RT-PCR in a diarrhea specimen collected from a patient in St. Louis, United States as well as in untreated sewage collected in Barcelona, Spain. Conclusion Klassevirus 1 is a previously undescribed picornavirus that is globally widespread and present on at least three continents. Further investigations to determine whether klassevirus 1 is a human pathogen are needed

The NICE-GUT trial protocol:A randomised, placebo controlled trial of oral nitazoxanide for the empiric treatment of acute gastroenteritis among Australian Aboriginal children

Diarrhoeal disease is the second leading cause of death in children under 5 years globally, killing 525 000 annually. Australian Aboriginal and Torres Strait Islander (hereafter Aboriginal) children suffer a high burden of disease. Randomised trials in other populations suggest nitazoxanide accelerates recovery for children with Giardia, amoebiasis, Cryptosporidium, Rotavirus and Norovirus gastroenteritis, as well as in cases where no enteropathogens are found. This double blind, 1:1 randomised, placebo controlled trial is investigating the impact of oral nitazoxanide on acute gastroenteritis in hospitalised Australian Aboriginal children aged 3 months to <5 years. Dosing is based on age-based dosing. The primary endpoint is the time to resolution of 'significant illness' defined as the time from randomisation to the time of clinical assessment as medically ready for discharge, or to the time of actual discharge from hospital, whichever occurs first. Secondary endpoints include duration of hospitalisation, symptom severity during the period of significant illness and following treatment, duration of rehydration and drug safety. Patients will be followed for medically significant events for 60 days. Analysis is based on Bayesian inference. Subgroup analysis will occur by pathogen type (bacteria, virus or parasite), rotavirus vaccination status, age and illness severity. Ethics approval has been granted by the Central Australian Human Research Ethics Committee (HREC-14-221) and the Human Research Ethics Committee of the Northern Territory Department of Health and Menzies School of Health Research (HREC2014-2172). Study investigators will ensure that the trial is conducted in accordance with the principles of the Declaration of Helsinki. Individual participant consent will be obtained. Results will be disseminated via peer-reviewed publication. ACTRN12614000381684

Metagenomic Analysis of Human Diarrhea: Viral Detection and Discovery

Worldwide, approximately 1.8 million children die from diarrhea annually, and millions more suffer multiple episodes of nonfatal diarrhea. On average, in up to 40% of cases, no etiologic agent can be identified. The advent of metagenomic sequencing has enabled systematic and unbiased characterization of microbial populations; thus, metagenomic approaches have the potential to define the spectrum of viruses, including novel viruses, present in stool during episodes of acute diarrhea. The detection of novel or unexpected viruses would then enable investigations to assess whether these agents play a causal role in human diarrhea. In this study, we characterized the eukaryotic viral communities present in diarrhea specimens from 12 children by employing a strategy of “micro-mass sequencing” that entails minimal starting sample quantity (<100 mg stool), minimal sample purification, and limited sequencing (384 reads per sample). Using this methodology we detected known enteric viruses as well as multiple sequences from putatively novel viruses with only limited sequence similarity to viruses in GenBank

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

Whole Genome In-Silico Analysis of South African G1P[8] Rotavirus Strains before and after Vaccine Introduction over a Period of 14 Years

Rotavirus G1P[8] strains account for more than half of the group A rotavirus (RVA) infections in children under five years of age, globally. A total of 103 stool samples previously characterized as G1P[8] and collected seven years before and seven years after introducing the Rotarix® vaccine in South Africa were processed for whole-genome sequencing. All the strains analyzed had a Wa-like constellation (G1-P[8]-I1-R1-C1-M1-A1-N1-T1-E1-H1). South African pre- and post-vaccine G1 strains were clustered in G1 lineage-I and II while the majority (84.2%) of the P[8] strains were grouped in P[8] lineage-III. Several amino acid sites across ten gene segments with the exception of VP7 were under positive selective pressure. Except for the N147D substitution in the antigenic site of eight post-vaccine G1 strains when compared to both Rotarix® and pre-vaccine strains, most of the amino acid substitutions in the antigenic regions of post-vaccine G1P[8] strains were already present during the pre-vaccine period. Therefore, Rotarix® did not appear to have an impact on the amino acid differences in the antigenic regions of South African post-vaccine G1P[8] strains. However, continued whole-genome surveillance of RVA strains to decipher genetic changes in the post-vaccine period remains imperative

Immunogenicity of a Third Scheduled Dose of Rotarix in Australian Indigenous Infants: A Phase IV, Double-blind, Randomized, Placebo-Controlled Clinical Trial

BackgroundRotarix (GlaxoSmithKline) oral rotavirus vaccine is licensed as 2 doses in the first 6 months of life. In settings with high child mortality rates, clinical protection conferred by 2 doses of Rotarix is reduced. We assessed vaccine immune response when an additional dose of Rotarix was given to Australian Aboriginal children 6 to MethodsORVAC is a 2-stage, double-blind, randomized, placebo-controlled trial. Australian Aboriginal children 6 to ResultsBetween March 2018 and August 2020, a total of 253 infants were enrolled. Of these, 178 infants (70%) had analyzable serological results after follow-up; 89 were randomized to receive Rotarix, and 89 to receive placebo. The proportion with seroresponse was 85% after Rotarix compared with 72% after placebo. There were no occurrences of intussusception or any serious adverse events.ConclusionsAn additional dose of Rotarix administered to Australian Aboriginal infants 6 to Clinical trials registrationNCT02941107

Comparative field performance and adherence to test results of four malaria rapid diagnostic tests among febrile patients more than five years of age in Blantyre, Malawi

<p>Abstract</p> <p>Background</p> <p>Malaria rapid diagnostics tests (RDTs) can increase availability of laboratory-based diagnosis and improve the overall management of febrile patients in malaria endemic areas. In preparation to scale-up RDTs in health facilities in Malawi, an evaluation of four RDTs to help guide national-level decision-making was conducted.</p> <p>Methods</p> <p>A cross sectional study of four histidine rich-protein-type-2- (HRP2) based RDTs at four health centres in Blantyre, Malawi, was undertaken to evaluate the sensitivity and specificity of RDTs, assess prescriber adherence to RDT test results and explore operational issues regarding RDT implementation. Three RDTs were evaluated in only one health centre each and one RDT was evaluated in two health centres. Light microscopy in a reference laboratory was used as the gold standard.</p> <p>Results</p> <p>A total of 2,576 patients were included in the analysis. All of the RDTs tested had relatively high sensitivity for detecting any parasitaemia [Bioline SD (97%), First response malaria (92%), Paracheck (91%), ICT diagnostics (90%)], but low specificity [Bioline SD (39%), First response malaria (42%), Paracheck (68%), ICT diagnostics (54%)]. Specificity was significantly lower in patients who self-treated with an anti-malarial in the previous two weeks (odds ratio (OR) 0.5; p-value < 0.001), patients 5-15 years old versus patients > 15 years old (OR 0.4, p-value < 0.001) and when the RDT was performed by a community health worker versus a laboratory technician (OR 0.4; p-value < 0.001). Health workers correctly prescribed anti-malarials for patients with positive RDT results, but ignored negative RDT results with 58% of patients with a negative RDT result treated with an anti-malarial.</p> <p>Conclusions</p> <p>The results of this evaluation, combined with other published data and global recommendations, have been used to select RDTs for national scale-up. In addition, the study identified some key issues that need to be further delineated: the low field specificity of RDTs, variable RDT performance by different cadres of health workers and the need for a robust quality assurance system. Close monitoring of RDT scale-up will be needed to ensure that RDTs truly improve malaria case management.</p