Gastroenteric Viruses Detection in a Drinking Water Distribution-to-Consumption System in a Low-Income Community in Rio de Janeiro.

The availability of drinking water is one of the main determinants of quality of life, disease prevention and the promotion of health. Viruses are important agents of waterborne diseases and have been described as important markers of human faecal contamination. This study aimed to investigate viruses' presence as an indicator of drinking water quality in low-income communities in the Manguinhos area, Rio de Janeiro, Brazil. Three hundred and four drinking water samples (2L/each) were collected along the drinking water distribution-to-consumption pathway in households, as well as healthcare and school units. Water samples were collected both directly from the water supply prior to distribution and after storage in tanks and filtration units. Using qPCR, viruses were detected 50 times in 45 water samples (15%), 19 of these being human adenovirus, 17 rotavirus A and 14 norovirus GII. Viral loads recovered ranged from 5E+10 to 8.7E+106 genome copies/Liter. Co-detection was observed in five household water samples and there was no difference regarding virus detection across sampling sites. Precarious and inadequate environmental conditions characterized by the lack of local infrastructure regarding basic sanitation and waste collection in the territory, as well as negligent hygiene habits, could explain viral detection in drinking water in regions with a water supply system.

Detection of salivirus in raw sewage samples in Rio de Janeiro, Brazil

Gastroenteric viruses are important pathogens related to cases of acute gastroenteritis, affecting millions of people worldwide with a major impact on children under five in developing countries. The introduction of metagenomic approach techniques in the 2000s has allowed the description of new viruses, among them Salivirus, which has been associated worldwide with cases of diarrhea. This study aimed to detect salivirus in raw sewage samples from a wastewater treatment plant (WWTP) collected between June 2013 and May 2014 in Rio de Janeiro, Brazil. Fifty-two samples collected weekly were tested by using a real-time quantitative PCR (qPCR). Salivirus genome was detected in 71.1% (37/52) of the samples, with viral concentration ranging from 7.56 x 104 to 7.20 x 106 genomic copies per liter. Higher viral loads were detected in the summer and fall of 2014, although these data were not sufficient to infer seasonality for this virus. The high prevalence of salivirus in sewage samples highlights the importance of viral research in wastewater to generate data on salivirus circulation, increasing understanding regarding its dissemination in the population.

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.

Enteric viruses in HIV-1 seropositive and HIV-1 seronegative children with diarrheal diseases in Brazil.

Diarrheal diseases (DD) have distinct etiological profiles in immune-deficient and immune-competent patients. This study compares detection rates, genotype distribution and viral loads of different enteric viral agents in HIV-1 seropositive (n = 200) and HIV-1 seronegative (n = 125) children hospitalized with DD in Rio de Janeiro, Brazil. Except for group A rotavirus (RVA), which were detected through enzyme immunoassay, the other enteric viruses (norovirus [NoV], astrovirus [HAstV], adenovirus [HAdV] and bocavirus [HBoV]) were detected through PCR or RT-PCR. A quantitative PCR was performed for RVA, NoV, HAstV, HAdV and HBoV. Infections with NoV (19% vs. 9.6%; p<0.001), HBoV (14% vs. 7.2%; p = 0.042) and HAdV (30.5% vs. 14.4%; p<0.001) were significantly more frequent among HIV-1 seropositive children. RVA was significantly less frequent among HIV-1 seropositive patients (6.5% vs. 20%; p<0.001). Similarly, frequency of infection with HAstV was lower among HIV-1 seropositive children (5.5% vs. 12.8%; p = 0.018). Among HIV-1 seropositive children 33 (16.5%) had co-infections, including three enteric viruses, such as NoV, HBoV and HAdV (n = 2) and NoV, HAstV and HAdV (n = 2). The frequency of infection with more than one virus was 17 (13.6%) in the HIV-1 negative group, triple infection (NoV + HAstV + HBoV) being observed in only one patient. The median viral load of HAstV in feces was significantly higher among HIV-1 positive children compared to HIV-1 negative children. Concerning children infected with RVA, NoV, HBoV and HAdV, no statistically significant differences were observed in the medians of viral loads in feces, comparing HIV-1 seropositive and HIV-1 seronegative children. Similar detection rates were observed for RVA, HAstV and HAdV, whilst NoV and HBoV were significantly more prevalent among children with CD4+ T lymphocyte count below 200 cells/mm3. Enteric viruses should be considered an important cause of DD in HIV-1 seropositive children, along with pathogens more classically associated with intestinal infections in immunocompromised hosts.

The Impact of the Extreme Amazonian Flood Season on the Incidence of Viral Gastroenteritis Cases.

During the Amazonian flood season in 2012, the Negro River reached its highest level in 110 years, submerging residential and commercial areas which appeared associated with an elevation in the observed gastroenteritis cases in the city of Manaus. The aim of this study was to evaluate the microbiological water quality of the Negro River basin during this extreme flood to investigate this apparent association between the illness cases and the population exposed to the contaminated waters. Forty water samples were collected and analysed for classic and emerging enteric viruses. Human adenoviruses, group A rotaviruses and genogroup II noroviruses were detected in 100, 77.5 and 27.5% of the samples, respectively, in concentrations of 103-106 GC/L. All samples were compliant with local bacteriological standards. HAdV2 and 41 and RVA G2, P[6], and P[8] were characterised. Astroviruses, sapoviruses, genogroup IV noroviruses, klasseviruses, bocaviruses and aichiviruses were not detected. Statistical analyses showed correlations between river stage level and reported gastroenteritis cases and, also, significant differences between virus concentrations during this extreme event when compared with normal dry seasons and previous flood seasons of the Negro River. These findings suggest an association between the extreme flood experienced and gastrointestinal cases in the affected areas providing circumstantial evidence of causality between the elevations in enteric viruses in surface waters and reported illness.

Detection and Molecular Characterization of Gemycircularvirus from Environmental Samples in Brazil.

Gemycircularvirus (GemyCV) is a group of viruses which has been recently proposed as a new viral genus detected in fecal and environmental samples around the world. GemyCVs have been detected in human blood, brain tissue, cerebrospinal fluid, and stool sample. In the present study, we demonstrate for the first time, through molecular detection and characterization, the presence of GemyCVs in environmental samples from Brazil. Our results show a percentage of positivity ranging from 69 (25/36) to 97 % (35/36) in river water samples collected in Manaus, Amazon region, and wastewater from a wastewater treatment plant located in Rio de Janeiro, respectively, revealing GemyCVs as an important environmental contaminant.

A non-enteric adenovirus A12 gastroenteritis outbreak in Rio de Janeiro, Brazil

A gastroenteritis outbreak that occurred in 2013 in a low-income community in Rio de Janeiro was investigated for the presence of enteric viruses, including species A rotavirus (RVA), norovirus (NoV), astrovirus (HAstV), bocavirus (HBoV), aichivirus (AiV), and adenovirus (HAdV). Five of nine stool samples (83%) from patients were positive for HAdV, and no other enteric viruses were detected. Polymerase chain reaction products were sequenced and subjected to phylogenetic analysis, which revealed four strains and one strain of non-enteric HAdV-A12 and HAdV-F41, respectively. The HAdV-A12 nucleotide sequences shared 100% nucleotide similarity. Viral load was assessed using a TaqMan real-time PCR assay. Stool samples that were positive for HAdV-A12 had high viral loads (mean 1.9 X 107 DNA copies/g stool). All four patients with HAdV-A12 were < 25 months of age and had symptoms of fever and diarrhoea. Evaluation of enteric virus outbreaks allows the characterisation of novel or unique diarrhoea-associated viruses in regions where RVA vaccination is routinely performed.

A non-enteric adenovirus A12 gastroenteritis outbreak in Rio de Janeiro, Brazil.

A gastroenteritis outbreak that occurred in 2013 in a low-income community in Rio de Janeiro was investigated for the presence of enteric viruses, including species A rotavirus (RVA), norovirus (NoV), astrovirus (HAstV), bocavirus (HBoV), aichivirus (AiV), and adenovirus (HAdV). Five of nine stool samples (83%) from patients were positive for HAdV, and no other enteric viruses were detected. Polymerase chain reaction products were sequenced and subjected to phylogenetic analysis, which revealed four strains and one strain of non-enteric HAdV-A12 and HAdV-F41, respectively. The HAdV-A12 nucleotide sequences shared 100% nucleotide similarity. Viral load was assessed using a TaqMan real-time PCR assay. Stool samples that were positive for HAdV-A12 had high viral loads (mean 1.9 X 107 DNA copies/g stool). All four patients with HAdV-A12 were < 25 months of age and had symptoms of fever and diarrhoea. Evaluation of enteric virus outbreaks allows the characterisation of novel or unique diarrhoea-associated viruses in regions where RVA vaccination is routinely performed.

Surveillance of Human Astrovirus Infection in Brazil: The First Report of MLB1 Astrovirus.

Human astrovirus (HAstV) represents the third most common virus associated with acute diarrhea (AD). This study aimed to estimate the prevalence of HAstV infection in Brazilian children under 5 years of age with AD, investigate the presence of recently described HAstV strains, through extensive laboratory-based surveillance of enteric viral agents in three Brazilian coastal regions between 2005 and 2011. Using reverse transcription-polymerase chain reaction (RT-PCR), the overall HAstV detection rate reached 7.1% (207/2.913) with percentage varying according to the geographic region: 3.9% (36/921) in the northeast, 7.9% in the south (71/903) and 9.2% in the southeast (100/1.089) (p < 0.001). HAstV were detected in cases of all age groups. Detection rates were slightly higher during the spring. Nucleotide sequence analysis of a 320-bp ORF2 fragment revealed that HAstV-1 was the predominant genotype throughout the seven years of the study. The novel AstV-MLB1 was detected in two children with AD from a subset of 200 samples tested, demonstrating the circulation of this virus both the in northeastern and southeastern regions of Brazil. These results provide additional epidemiological and molecular data on HAstV circulation in three Brazilian coastal regions, highlighting its potential to cause infantile AD.

Aichi Virus Positivity in HIV-1 Seropositive Children Hospitalized with Diarrheal Disease.

Aichi viruses (AiV) have been detected in patients with diarrheal diseases (DD). The aim of this study was to assess AiV infection rates in hospitalized children with DD, including 123 HIV-1 seropositive and 125 HIV-1 seronegative patients, in two public pediatric hospitals in Rio de Janeiro, Brazil. AiV was investigated by nested RT-PCR. The AiV-positive samples were also tested for specie A rotavirus, norovirus, astrovirus, enteric adenovirus and bocavirus in order to assess co-infections. AiV parcial genome sequencing and phylogenetic analyses were performed. AiV were detected in 9/123 (7.32%) of the HIV-1 seropositive subjects and 1/125 (0.8%) of the HIV seronegative patients with DD (p = 0.019). The phylogenetic analysis of positive samples disclosed that: i) 13 samples were characterized as genotype A, with one of them being from the HIV-1 seronegative patient; ii) one sample from a HIV-1 seropositive patient was characterized as genotype B. AiV genotype A was grouped into 3 genetic clusters. Data suggest that AiV may be an opportunistic pathogen infecting children with AIDS and DD.

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.

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.