Dataset Information

Norovirus RNA in serum associated with increased fecal viral load in children: Detection, quantification and molecular analysis.

ABSTRACT: Worldwide, norovirus (NoV) is a major cause of acute gastroenteritis (AGE) responsible for pandemics every ~3 years, and over 200,000 deaths per year, with the majority in children from developing countries. We investigate the incidence of NoV in children hospitalized with AGE from Belém, Pará, Brazil, and also correlated viral RNA levels in their blood and stool with clinical severity. For this purpose, paired stool and serum samples were collected from 445 pediatric patients, ?9 years between March 2012 and June 2015. Enzyme-linked immunosorbent assay (EIA) was used to detect NoV in stool and reverse transcription quantitative PCR (RT-qPCR) used to quantify NoV RNA levels in sera (RNAemia) and in the positive stool. Positives samples were characterized by the partial ORF1/2 region sequence of viral genome. NoV antigen was detected in 24.3% (108/445) of stool samples, with RNAemia also present in 20.4% (22/108). RNAemia and a high stool viral load (>107 genome copies/gram of faeces) were associated with longer hospitalizations. The prevalent genotypes were GII.4 Sydney_2012 (71.6%-58/81) and New Orleans_2009 (6.2%-5/81) variants. Eight other genotypes belonging to GII were detected and four of them were recombinant strains. All sera were characterized as GII.4 and shared 100% similarity with their stool. The results suggest that the dissemination of NoV to the blood stream is not uncommon and may be related to increased faecal viral loads and disease severity.

SUBMITTER: Reymao TKA

PROVIDER: S-EPMC6028094 | biostudies-literature | 2018

REPOSITORIES: biostudies-literature

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Norovirus RNA in serum associated with increased fecal viral load in children: Detection, quantification and molecular analysis.

Reymão Tammy Kathlyn Amaral TKA Fumian Tulio Machado TM Justino Maria Cleonice Aguiar MCA Hernandez Juliana Merces JM Bandeira Renato Silva RS Lucena Maria Silvia Sousa MSS Teixeira Dielle Monteiro DM Farias Fredison Pinheiro FP Silva Luciana Damascena LD Linhares Alexandre Costa AC Gabbay Yvone Benchimol YB

PloS one 20180702 7

Worldwide, norovirus (NoV) is a major cause of acute gastroenteritis (AGE) responsible for pandemics every ~3 years, and over 200,000 deaths per year, with the majority in children from developing countries. We investigate the incidence of NoV in children hospitalized with AGE from Belém, Pará, Brazil, and also correlated viral RNA levels in their blood and stool with clinical severity. For this purpose, paired stool and serum samples were collected from 445 pediatric patients, ≤9 years between ...[more]

PMID: 29965979

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Project description:BACKGROUND: Reverse transcription-polymerase chain reaction (RT-PCR) is the main method for laboratory diagnosis of norovirus-associated infectious intestinal disease (IID). However, up to 16% of healthy individuals in the community, with no recent history of IID, may be RT-PCR positive; so it is unclear whether norovirus is actually the cause of illness in an IID case when they are RT-PCR positive. It is important to identify the pathogen causing illness in sporadic IID cases, for clinical management and for community based incidence studies. The aim of this study was to investigate how faecal viral load can be used to determine when norovirus is the most likely cause of illness in an IID case. METHODS: Real-time RT-PCR was used to determine the viral load in faecal specimens collected from 589 IID cases and 159 healthy controls, who were infected with genogroup II noroviruses. Cycle threshold (Ct) values from the real-time RT-PCR were used as a proxy measure of viral load. Receiver-operating characteristic (ROC) analysis was used to identify a cut-off in viral load for attributing illness to norovirus in IID cases. RESULTS: One hundred and sixty-nine IID cases and 159 controls met the inclusion criteria for the ROC analysis. The optimal Ct value cut-off for attributing IID to norovirus was 31. The same cut-off was selected when using healthy controls, or IID cases who were positive by culture for bacterial pathogens, as the reference negative group. This alternative reference negative group can be identified amongst specimens routinely received in clinical virology laboratories. CONCLUSION: We demonstrated that ROC analysis can be used to select a cut-off for a norovirus real time RT-PCR assay, to aid clinical interpretation and diagnose when norovirus is the cause of IID. Specimens routinely received for diagnosis in clinical virology laboratories can be used to select an appropriate cut-off. Individual laboratories can use this method to define in-house cut-offs for their assays, to provide the best possible diagnostic service to clinicians and public health workers. Other clinical and epidemiological information should also be considered for patients with Ct values close to the cut-off, for the most accurate diagnosis of IID aetiology.

Project description:The sooty mangabey (SM) (Cercocebus atys) is the natural host of a simian immunodeficiency virus, termed SIVsm, which gave rise to human immunodeficiency virus type 2. Data on the geographic distribution, prevalence, and genetic diversity of SIVsm in the wild remains limited. To address this issue, noninvasive strategies based on screening SM fecal and urine specimens for SIVsm-specific antibodies and virion RNA (vRNA) were developed, and the results were correlated with viral loads in plasma. Twenty-three SIVsm-infected and 27 uninfected SMs were evaluated. Time-matched urine, fecal and plasma samples were collected over a 2-month period from 16 captive naturally infected SMs. The remaining 7 infected and 27 uninfected SMs were sampled once. Each specimen was subjected to enhanced chemiluminescence-Western blot analysis and nested reverse transcriptase (RT) PCR. The results showed that urine was highly sensitive (96%) and specific (100%) for detection of SIVsm antibodies, while fecal detection was much less sensitive (16%). Conversely, vRNA detection was more sensitive in feces (50%) than in urine (2%) samples. Fecal-vRNA detection correlated with viral loads in plasma (P < 0.002). SMs with detectable fecal vRNA had a mean viral load in plasma of 458,006 copies/ml, while those with undetectable fecal vRNA had a mean viral load in plasma of 29,428 copies/ml. Moreover, for every log increase in the viral load in plasma, the odds of detecting virus in fecal samples increased 87-fold. Genetic diversity of SIVsm in the SM colony was characterized by sequencing partial gag (846 bp) and gp43 (439 bp) fragments. Surprisingly, four new SIVsm lineages were identified, two of which were initially detected by fecal RT-PCR. This study documents the suitability of noninvasive methods for the detection and molecular characterization of new SIV variants. These assays will be useful for studying the phylogeny and epidemiology of SIVsm infections in the wild, and they hold promise as tools for investigating natural SIV infections in endangered nonhuman primates.

Dataset Information

Norovirus RNA in serum associated with increased fecal viral load in children: Detection, quantification and molecular analysis.

Publications

Norovirus RNA in serum associated with increased fecal viral load in children: Detection, quantification and molecular analysis.

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