Julia Monassa Fioretti

Surveillance of norovirus infections in the state of Rio De Janeiro, Brazil 2005-2008.

A 4‐year (2005–2008) norovirus (NoV) surveillance study was conducted in the state of Rio Janeiro, Brazil, to demonstrate the role of these viruses in outbreaks and sporadic cases of acute gastroenteritis. A cohort of 1,687 fecal samples was obtained from patients with gastroenteritis; 324 were rotavirus‐positive. Of the remainder 1,363 rotavirus‐negative samples, 1,087 samples were tested for NoV RNA in this study. The study enrolled 267 outpatients from Municipal Public Health Centers and 820 inpatients, whose samples were obtained by active surveillance in Public Hospitals. Fecal samples were tested by reverse transcription (RT) followed by polymerase chain reaction (PCR) using the MON 431–434 set of degenerate primers for NoV GI and GII detection, and there were 35.1% (381/1,087) positive samples for NoV, consisting of 30.2% (248/820) and 49.8% (133/267) from inpatient and outpatient, respectively. Children infected by NoV had significantly more frequent mucus in feces, vomiting and fever. No seasonal pattern in NoV infections was observed in patients admitted to hospital; however, two peaks of NoV infections were observed from ambulatory cases, suggesting that there was an occurrence of outbreaks in those time periods. Molecular characterization revealed GII to be the most prevalent genogroup, totaling 96.3% (104/108) of all sequences analyzed, and GII.4 was the genotype detected most frequently (80.7%), followed by GII.6, 3, 14, 7, and 8. Two GI strains, GI.2 and GI.3, were also observed. The number of outbreaks and sporadic cases described in this study highlights the need to implement diagnosis of NoV in surveillance laboratories. J. Med. Virol. 82:1442–1448, 2010.

Genetic diversity of noroviruses in Brazil

Norovirus (NoV) infections are a major cause of acute gastroenteritis outbreaks around the world. In Brazil, the surveillance system for acute diarrhoea does not include the diagnosis of NoV, precluding the ability to assess its impact on public health. The present study assessed the circulation of NoV genotypes in different Brazilian states by partial nucleotide sequencing analysis of the genomic region coding for the major capsid viral protein. NoV genogroup II genotype 4 (GII.4) was the prevalent (78%) followed by GII.6, GII.7, GII.12, GII.16 and GII.17, demonstrating the great diversity of NoV genotypes circulating in Brazil. Thus, this paper highlights the importance of a virological surveillance system to detect and characterize emerging strains of NoV and their spreading potential.

Temporal dynamics of norovirus GII.4 variants in Brazil between 2004 and 2012.

Noroviruses (NoVs) are the major cause of acute gastroenteritis outbreaks, and, despite a wide genetic diversity, genotype II.4 is the most prevalent strain worldwide. Mutations and homologous recombination have been proposed as mechanisms driving the epochal evolution of the GII.4, with the emergence of new variants in 1–3-year intervals causing global epidemics. There are no data reporting the dynamics of GII.4 variants along a specific period in Brazil. Therefore, to improve the understanding of the comportment of these variants in the country, the aim of this study was to evaluate the circulation of NoV GII.4 variants during a 9-year period in 3 out of 5 Brazilian regions. A total of 147 samples were sequenced, and a phylogenetic analysis of subdomain P2 demonstrated the circulation of six GII.4 variants, Asia_2003, Hunter_2004, Den Haag_2006b, Yerseke_2006a, New Orleans_2009, and Sydney_2012, during this period. The most prevalent variant was Den Haag_2006b, circulating in different Brazilian regions from 2006 to 2011. A Bayesian coalescent analysis was used to calculate the mean evolutionary rate of subdomain P2 as 7.3×10−3 (5.85×10−3–8.82×10−3) subst./site/year. These analyses also demonstrated that clade Den Haag_2006b experienced a rapid expansion in 2005 and another in 2008 after a period of decay. The evaluation of the temporal dynamics of NoV GII.4 in Brazil revealed a similar pattern, with few exceptions, to the worldwide observation. These data highlight the importance of surveillance for monitoring the emergence of new strains of NoV GII.4 and its impact on cases of acute gastroenteritis.

Noroviruses associated with outbreaks of acute gastroenteritis in the State of Rio Grande do Sul, Brazil, 2004-2011.

BACKGROUND Acute gastroenteritis norovirus (NoV) in a country of continental dimensions like Brazil has resulted in under-reporting of the number of outbreaks, as well as the genotypes associated. OBJECTIVES To demonstrate the role of NoV in outbreaks occurring in the State of Rio Grande do Sul, Southern Brazil, we determined its prevalence, as well as the genotypes associated, and evaluated clinical and epidemiological aspects. STUDY DESIGN NoV investigation was carried out in rotavirus group A negative stool samples from 2265 patients from 741 outbreaks that occurred in the State of Rio Grande do Sul, Brazil, during a period of eight years (2004-2011). NoV detection and nucleotide sequencing for genotype characterization was carried by using sets of primers targeting a conservative Rd-Rp polymerase genome region and the viral capsid gene, respectively. RESULTS NoVs were detected in 817 stool samples (36.1%) and associated with 327 outbreaks (44.1%). NoV GII.2, GII.3, GII.4, GII.6, GII.12, GII.13, GII.14, GII.15, GII.17, GII.21; and GI.1 and GI.3 were characterized. GII.4 was the most frequently detected (72.3%), with five variants identified (Asia_2003, Hunter_2004, Yerseke_2006a, Den_Haag_2006b, New Orleans_2009). This study describes the first detection of GI.1 and GII.13 and GII.15 in Brazil and demonstrates NoV winter-spring seasonality in this region of the country. CONCLUSIONS NoVs were responsible for almost 50% of outbreaks, with about 70% of them resulting from genotype GII.4 and its variants. The seasonality observed could help health authorities to establish a system of active surveillance in order to reduce NoV impact especially in congregate settings.

Bayesian coalescent inference reveals high evolutionary rates and expansion of Norovirus populations

Noroviruses (NoV) are a leading cause of outbreaks of nonbacterial acute gastroenteritis in humans worldwide and have become an important cause of hospitalization of children in South America. NoV belong to the family Caliciviridae and are non-enveloped single stranded, positive sense, RNA viruses. NoV of genotype GII/4 have emerged worldwide, causing four epidemic seasons of viral gastroenteritis during which four novel variants emerged. Despite the importance of NoV outbreaks, little is known about the evolutionary rates, viral spread and population dynamics of NoV populations. In order to gain insight into these matters, a Bayesian Markov chain Monte Carlo (MCMC) approach was used to analyze region D or full-length VP1 gene sequences of GII/4 NoV populations isolated in Brazil or Japan, respectively. The results of these studies revealed that the expansion population growth model was the best to fit the data in both datasets. The dates of the most common recent ancestors revealed that these viruses can quickly emerge in a geographical location. A mean evolutionary rate of 1.21 x 10(-2) nucleotide substitution/site/year (s/s/y) was obtained for the VP1 gene using full-length sequences. This rate is higher than the rates reported for other rapidly evolving RNA. Roughly similar rates (1.44 x 10(-2)s/s/y) were found using region D sequences, revealing the suitability of this region for evolutionary studies, in agreement with previous reports. High evolutionary rates and fast population growth may have contributed to the vigorous initial transmission dynamics of the GII/4 NoV populations studied.

Occurrence of human sapoviruses in wastewater and stool samples in Rio De Janeiro, Brazil.

To determine the prevalence and molecular epidemiology of human sapovirus (SaV) in both wastewater and stool samples in a 3‐year (2012–2014) surveillance study performed in Rio de Janeiro State, Brazil.

Surveillance of Noroviruses in Rio De Janeiro, Brazil: Occurrence of New GIV Genotype in Clinical and Wastewater Samples

Genogroup (G) IV norovirus (NoV) has been described in the literature as infectious agents in humans, although there are few reports regarding the frequency and spread of this virus, resulting in insufficient epidemiological data. The aim of this study was to investigate the occurrence of GIV norovirus in the State of Rio de Janeiro, Brazil in order to evaluate frequency, concentration, and genetic diversity using clinical and environmental approaches. For this purpose, 316 stool samples were collected from acute gastroenteritis cases reported over a period of three years. Wastewater samples were also obtained from the main wastewater treatment plant (WWTP) located in Rio de Janeiro throughout one year, totalizing 156 samples. All samples were submitted to quantitative analysis by TaqMan™ real-time PCR for GIV norovirus. Three out of 316 clinical samples were positive (0.9%) for GIV, with viral load ranging from 104 to 106 genome copies (CG) per gram. Regarding wastewater samples, GIV were detected in 52% of raw sewage, with viral load ranging from 104 to 106 CG per liter. Phylogenetic analysis revealed the circulation of a new GIV genotype in both clinical and environmental samples. To our knowledge, this is the first description of GIV norovirus in clinical samples in Brazil. These results demonstrate the importance of performing laboratory surveillance of clinical and environmental samples, assisting the comprehension of the epidemiology pattern of viruses with neglected diagnosis and indefinite impact in the population.

Molecular characterization of feline calicivirus variants from multicat household and public animal shelter in Rio de Janeiro, Brazil

The aim of this study was to perform the molecular characterization of conserved and variable regions of feline calicivirus capsid genome in order to investigate the molecular diversity of variants in Brazilian cat population. Twenty-six conjunctival samples from cats living in five public short-term animal shelters and three multicat life-long households were analyzed. Fifteen cats had conjunctivitis, three had oral ulceration, eight had respiratory signs (cough, sneeze and nasal discharge) and nine were asymptomatic. Feline calicivirus were isolated in CRFK cells and characterized by reverse transcription PCR target to both conserved and variable regions of open reading frame 2. The amplicons obtained were sequenced. A phylogenetic analysis along with most of the prototypes available in GenBank database and an amino acid analysis were performed. Phylogenetic analysis based on both conserved and variable region revealed two clusters with an aLTR value of 1.00 and 0.98 respectively and the variants from this study belong to feline calicivirus genogroup I. No association between geographical distribution and/or clinical signs and clustering in phylogenetic tree was observed. The variants circulating in public short-term animal shelter demonstrated a high variability because of the relatively rapid turnover of carrier cats constantly introduced of multiple viruses into this location over time.