Norovirus Recombinant Strains Isolated from Gastroenteritis Outbreaks in Southern Brazil, 2004–2011

Submitted by Sandra Infurna ([email protected]) on 2016-12-26T14:14:01Z No. of bitstreams: 1 juliana_andrade_etal_IOC_2016.pdf: 1969643 bytes, checksum: df814dc238ff8e34e22c2f7e11107106 (MD5)Approved for entry into archive by Sandra Infurna ([email protected]) on 2016-12-26T14:22:45Z (GMT) No. of bitstreams: 1 juliana_andrade_etal_IOC_2016.pdf: 1969643 bytes, checksum: df814dc238ff8e34e22c2f7e11107106 (MD5)Made available in DSpace on 2016-12-26T14:22:45Z (GMT). No. of bitstreams: 1 juliana_andrade_etal_IOC_2016.pdf: 1969643 bytes, checksum: df814dc238ff8e34e22c2f7e11107106 (MD5) Previous issue date: 2016Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Biologia Comparada e Ambiental. Rio de Janeiro, RJ, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Biologia Comparada e Ambiental. Rio de Janeiro, RJ, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Biologia Comparada e Ambiental. Rio de Janeiro, RJ, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Biologia Comparada e Ambiental. Rio de Janeiro, RJ, Brasil.Noroviruses are recognized as one of the leading causes of viral acute gastroenteritis, responsible for almost 50% of acute gastroenteritis outbreaks worldwide. The positive single-strand RNA genome of noroviruses presents a high mutation rate and these viruses are constantly evolving by nucleotide mutation and genome recombination. Norovirus recombinant strains have been detected as causing acute gastroenteritis outbreaks in several countries. However, in Brazil, only one report of a norovirus recombinant strain (GII.P7/GII.20) has been described in the northern region so far. For this study, 38 norovirus strains representative of outbreaks, 11 GII.4 and 27 non-GII.4, were randomly selected and amplified at the ORF1/ORF2 junction. Genetic recombination was identified by constructing phylogenetic trees of the polymerase and capsid genes, and further SimPlot and Bootscan analysis of the ORF1/ORF2 overlap. Sequence analysis revealed that 23 out of 27 (85%) non-GII.4 noroviruses were recombinant strains, characterized as: GII.P7/GII.6 (n = 9); GIIP.g/GII.12 (n = 4); GII.P16/GII.3 (n = 4); GII.Pe/GII.17 (n = 2); GII.P7/GII.14 (n = 1); GII.P13/GII.17 (n = 1); GII.P21/GII.3 (n = 1); and GII.P21/GII.13 (n = 1). On the other hand, among the GII.4 variants analyzed (Den Haag_2006b and New Orleans_2009) no recombination was observed. These data revealed the great diversity of norovirus recombinant strains associated with outbreaks, and describe for the first time these recombinant types circulating in Brazil. Our results obtained in southern Brazil corroborate the previous report for the northern region, demonstrating that norovirus recombinant strains are circulating more frequently than we expected. In addition, these results emphasize the relevance of including ORF1/ORF2-based analysis in surveillance studies as well as the importance of characterizing strains from other Brazilian regions to obtain epidemiological data for norovirus recombinant strains circulating in the country

Human Bocavirus in Brazil: Molecular Epidemiology, Viral Load and Co-Infections

Human bocavirus (HBoV) is an emerging virus and has been detected worldwide, especially in pediatric patients with respiratory and gastrointestinal infection. In this study, we describe HBoV prevalence, genotypes circulation and DNA shedding, in stool samples from children up to two years of age in Brazil. During 2016 and 2017, 886 acute gastroenteritis (AGE) stool samples from ten Brazilian states were analyzed by TaqMan®-based qPCR, to detect and quantify HBoV. Positive samples were genotyped by sequencing the VP1/2 overlap region, followed by phylogenetic analysis and co-infections were accessed by screening other gastroenteric viruses. HBoV was detected in 12.4% (n = 110) of samples, with viral load ranging from 1.6 × 102 to 1.2 × 109 genome copies per gram of stool. From these, co-infections were found in 79.1%, and a statistically lower HBoV viral load was found compared to viral loads of rotavirus, norovirus and adenovirus in double infected patients (p < 0.05). No significant differences were found between HBoV viral load in single or co-infections, age groups or genotypes. Phylogenetic analysis identified the circulation of HBoV-1 in 38%, HBoV-2 in 40% and HBoV-3 in 22%. Continuous HBoV monitoring is needed to clarify its role in diarrhea disease, especially in the absence of classic gastroenteric viruses

Nosocomial acute gastroenteritis outbreak caused by an equine-like G3P[8] DS-1-like rotavirus and GII.4 Sydney[P16] norovirus at a pediatric hospital in Rio de Janeiro, Brazil, 2019

Worldwide, rotavirus (RVA) and norovirus are considered major etiological agents of acute gastroenteritis (AGE) in pediatric population admitted to hospitals. This study describes the investigation of nosocomial infections caused by emergent RVA and norovirus strains reported at a pediatric hospital in southern Brazil in May 2019. This outbreak affected 30 people among children and adults. Nine stool samples (eight children and one nurse) were obtained and analyzed by RT-qPCR to detect and quantify RVA and norovirus. Positive samples were genotyped by sequencing and subjected to phylogenetic analysis. We detected RVA in 44.4% (4/9) and norovirus in 55.5% (5/9) at high viral loads, ranging from 3.5 × 107 to 6.1 × 107 and 3.2 × 102 to 3.2 × 109 genome copies/g of stool, respectively. Co-infections were not observed. RVA VP4 and VP7 gene sequencing in combination with polyacrylamide gel electrophoresis identified the circulation of equine-like G3P[8] DS-1-like, and the partial sequencing of the other nine genes revealed that strains possessed I2-R2-C2-M2-A2-N1-T2-E2-H2 genotype background. The emergent recombinant norovirus variant, GII.4 Sydney[P16], was identified by ORF1-2 sequencing. Active surveillance and effective prevention measures should be constantly reinforced to avoid the spread of nosocomial viral infections into hospitals, which could severely affect pediatric patients admitted with underlying health conditions

Rotavirus A during the COVID-19 Pandemic in Brazil, 2020–2022: Emergence of G6P[8] Genotype

Rotavirus A (RVA) remains a leading cause of acute gastroenteritis (AGE) hospitalizations in children worldwide. During the COVID-19 pandemic, a reduction in vaccination coverage in Brazil and elsewhere was observed, and some reports have demonstrated a reduction in AGE notifications during the pandemic. This study aims to investigate the diversity and prevalence of RVA genotypes in children and adults presenting with AGE symptoms in Brazil during the COVID-19 pandemic between 2020 and 2022. RVA was screened using RT-qPCR; then, G and P genotypes were characterized using one-step multiplex RT-PCR. A total of 2173 samples were investigated over the three-year period, and we detected RVA in 7.7% of samples (n = 167), being 15.5% in 2020, 0.5% in 2021, and 13.8% in 2022. Higher RVA prevalence was observed in the Northeastern region (19.3%) compared to the Southeastern (6.1%) and Southern regions (5.5%). The most affected age group was children aged between 0 and 6 months old; however, this was not statistically significant. Genotyping and phylogenetic analysis identified the emergence of G6P[8] during the period; moreover, it was detected in 10.6% of samples in 2020 and in 83.5% in 2022. In contrast, the prevalence of G3P[8], the previous dominant genotype, decreased from 72.3% in 2020 to 11.3% in 2022. We also identified unusual strains, such as G3P[9] and G9P[4], being sporadically detected during the period. This is the first report on the molecular epidemiology and surveillance of RVA during the COVID-19 pandemic period in Brazil. Our study provides evidence for the importance of maintaining high and sustainable levels of vaccine coverage to protect against RVA disease. Furthermore, it highlights the need to maintain nationwide surveillance in order to monitor future trends and changes in the epidemiology of RVA in Brazil

Detection of norovirus epidemic genotypes in raw sewage using next generation sequencing

Submitted by Sandra Infurna ([email protected]) on 2019-05-02T12:45:46Z No. of bitstreams: 1 TulioFumian_JuliaFioretti_etal_IOC_2019.pdf: 6296886 bytes, checksum: a655648de44ac1c27815339298dbc2da (MD5)Approved for entry into archive by Sandra Infurna ([email protected]) on 2019-05-02T12:54:52Z (GMT) No. of bitstreams: 1 TulioFumian_JuliaFioretti_etal_IOC_2019.pdf: 6296886 bytes, checksum: a655648de44ac1c27815339298dbc2da (MD5)Made available in DSpace on 2019-05-02T12:54:52Z (GMT). No. of bitstreams: 1 TulioFumian_JuliaFioretti_etal_IOC_2019.pdf: 6296886 bytes, checksum: a655648de44ac1c27815339298dbc2da (MD5) Previous issue date: 2019University of New South Wales. School of Biotechnology and Biomolecular Sciences.Faculty of Science. Sydney, Australia / Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Virologia Comparada e Ambiental. Rio de Janeiro, RJ, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Virologia Comparada e Ambiental. Rio de Janeiro, RJ, Brasil.University of New South Wales. School of Biotechnology and Biomolecular Sciences.Faculty of Science. Sydney, Australia.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Virologia Comparada e Ambiental. Rio de Janeiro, RJ, Brasil.University of New South Wales. School of Biotechnology and Biomolecular Sciences.Faculty of Science. Sydney, Australia.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Virologia Comparada e Ambiental. Rio de Janeiro, RJ, Brasil.Noroviruses are a leading cause of epidemic and pandemic acute gastroenteritis (AGE) worldwide, and contaminated food and water are important routes for its transmission. Raw sewage has been used for viral surveillance to monitor the emergence of new norovirus strains with the potential to cause epidemics. In this study, we investigated norovirus occurrence and norovirus RNA levels in 156 samples collected from May 2013 to May 2014, across three different stages (52 samples each) of a wastewater treatment plant (WWTP) in Rio de Janeiro, Brazil. We also explored norovirus GII diversity in raw sewage samples by next-sequencing generation (NGS). In addition, we examined norovirus prevalence and molecular epidemiology from acute gastroenteritis cases. Using RT-qPCR, norovirus GI and GII was detected in 38.5% and 96.1% of raw sewage samples, 40.4% and 96.1% of primary effluent samples and 1.9% and 5.8% of final effluent samples, respectively. Norovirus RNA levels varied from 4 to 6.2 log10 genome copies per litre (gc L-1) for GI and from 4.4 to 7.3 log10 gc L-1 for GII. Using MiSeq NGS, we identified 13 norovirus genotypes over the one-year period, with six dominant capsid genotypes, including GII.4, GII.17, GII.5, GII.2, GII.3 and GII.1. GII.4 noroviruses were the most prevalent in wastewater samples (68.5%), and a similar trend was observed in AGE cases (71%). The emergent GII.17 was the second most prevalent genotype (14.3%) identified in the raw sewage samples, however, it was not detected in clinical cases. Due to the high burden of norovirus outbreaks and the lack of vaccine and antiviral drugs, it is essential to understand the genotypic diversity of norovirus at the population level. Complementary data obtained from both clinical and environmental (sewage) samples proved to be an effective strategy to monitor the circulation and emergence of norovirus epidemic genotypes

Phylogenetic analyses of NoV GII sequences based on the polymerase and capsid regions.

<p>(a) Phylogenetic tree of 231 bp within the polymerase region (3’-ORF1). (b) Phylogenetic tree of 277 bp within the capsid region (5’-ORF2). References strains of NoV genotypes are named according to GenBank with their respectively accession numbers. Brazilian GII.4 strains are marked with a filled diamond. The scale bar at the bottom of the tree indicates distance. Bootstrap values (2000 replicates) are shown at the branch nodes and values lower than 60% are not shown.</p

Simplot and Bootscan analyses of the NoV recombinant GII.P7/GII.6 detected in four different years.

<p>(a) LVCA7842, (b) LVCA14441, (c) LVCA15109, and (d) LVCA17819. For similarity plot, the y-axis gives the percentage of identity within a sliding window of 200 bp wide, with a step size between plots of 20 bp. The site where the two NoV parental strains of genotypes GII.6 (JX989075) and GII.7 (AF414409) have equal identity to the recombinant (crossed by the vertical blue lines) is the predicted site of recombination. For Bootscan, the y-axis gives the percentage of bootstrap support values of permutated trees using a sliding window of 200 bp wide with a step size between plots of 20 bp. GII.17 strain (AY502009) was used as an outlier sequence.</p

Similarity plots of the NoV recombinant types detected in Southern Brazil during the 2004–2011 period.

<p>SimPlot was constructed using Simplot version 3.5.1 with a slide window width of 200 bp and a step size of 20 bp. The vertical axis indicates nucleotide identities (%) between the query sequences (Brazilian samples) and the reference strains. NoV parental strains are indicated by the accession numbers and their respectively ORF1-based genotypes (indicated by the letter P) or ORF2-based genotypes.</p

Phylogenetic analyses of NoV GII.4 sequences based on the polymerase region (ORF1) and capsid region (ORF2).

<p>(a) Phylogenetic tree of 231 bp within the polymerase region (3’ ORF1). (b) Phylogenetic tree of 277 bp within the capsid region (5’ ORF2). References strains of NoV GII.4 variants are named according to GenBank with their respectively accession numbers. Brazilian GII.4 strains are marked with a filled diamond. The scale bar at the bottom of the tree indicates distance. Bootstrap values (2,000 replicates) are shown at the branch nodes and values lower than 60% are not shown.</p