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Acesso aberto (Open Access)
Caracterização das infecções por norovírus nas hospitalizações pediátricas por gastrenterite na cidade de Belém, Pará
(Universidade Federal do Pará, 2012-04-20) SIQUEIRA, Jones Anderson Monteiro; GABBAY, Yvone Benchimol; http://lattes.cnpq.br/1579859438466504
The Norovirus (NoVs), Caliciviridae family, is related with acute gastroenteritis (AGE) in people of all ages groups. Its importance as cause of outbreaks have been confirmed, which occur mainly indoors. Its transmission is mainly by the fecal-oral route through contaminated water and food, and person to person contact. The disease usually is characterized by diarrhea, vomiting, nausea and abdominal cramps. The aim of this study was to demonstrate the importance of NoVs as pathogen associated with hospital admissions of children with AGE in Belém, Pará. The collection of fecal specimens occurred from May 2008 to April 2011, being only tested the samples with negative results for rotavirus. The enzyme immunoassay (EIA) and the reverse transcription-polymerase chain reaction (RT-PCR) were used for NoVs detection. The samples with positive results by EIA and negative by RT-PCR, were submitted to the semi-nested RT-PCR, and the ones with remain negative, were tested by the real-time PCR. A total of 483 samples were analyzed with a positivity of 35.4% (171/483). Adopting the RT-PCR as the reference method, the EIA had a sensitivity of 85.9% and a specificity of 93.4% with excellent reproducibility between them (kappa = 0.8, p <0.0001). The 22 samples positive only by EIA were tested first by semi-nested RT-PCR and after by real-time PCR, with a positivity of 63.6% (14/22) and 75% (6/8), respectively. The partial nucleotide sequencing of ORF1 region demonstrated the presence of GII.4d (80.8%-42/52), GII.7 (7.7%-4/52) and GII.b (11.5%-6/52) genotypes. Sequencing was performed in 64.3% (9/14) of the samples positive only by semi-nested RT-PCR, also corresponding to ORF1, which 55.6% (5/9) were classified as GII.4d and 44.4% (4/9) as GII.b. Of the six samples classified as GII.b, five were characterized as GII.3 when sequenced with primers specifics for the capsid region, suggesting the possibility of recombinants samples. A higher infection rate was observed in children under 2 years of age (90.1%-154/171) and the main symptoms were vomiting (95.8%-137/143) and dehydration (94.4%-118/25), considering the diarrhea was an inclusion criterion. Most infected children had more than 9 days of diarrhea (41.2%), 4 evacuations per day (43.9%) and more than 5 episodes of vomiting (90%) during hospitalization. Regarding seasonality, three peaks of positivity were observed in September and October 2008 (63.6%), and in February 2010 (62.1%). Any correlation with the climatic parameters of rainfall, humidity and temperature was demonstrated. This study confirmed the importance of NoVs as viral enteropathogen associated with AGE among hospitalized children in Belém, imposing the necessity of an active surveillance, in order to avoid possible morbidity caused by this virus in childhood.
Acesso aberto (Open Access)
Detecção e genotipagem de norovírus em diferentes amostras de água e esgoto não tratado na cidade de Belém, Pará, Brasil, 2008 a 2010
(Universidade Federal do Pará, 2014) TEIXEIRA, Dielle Monteiro; GABBAY, Yvone Benchimol; http://lattes.cnpq.br/1579859438466504
Enteric viruses excreted in feces from infected individuals dispersed in aquatic environments by sewage discharge. Among these viruses, the norovirus (NoV) is actually considered the main cause of gastroenteritis outbreaks worldwide, resulting from the ingestion of contaminated food and water as well as is also associated with hospitalizations. This research aimed to detect and partially characterize the human NoV (GI/GII) in different water matrices and in untreated sewage from Metropolitan Region of Belem. The study involved superficial waters from bay (Ver-o-Peso), river (Acai’s Port), stream (Tucunduba) and two lakes (Bolonha and Agua Preta), as well as treated water (WTP-Bolonha) and untreated sewage (SLP-UNA), monthly collected over two years . The water and sewage (2 liters) were initially concentrated on filtering membranes to obtain a final volume of 2 mL. The nucleic acid was extracted by silica method and submitted to semi nested RT-PCR (reverse transcription Polymerase chain reaction) using NoV GI and GII specific primers. The cDNA obtained after reverse transcription was also used to investigate the GI/GII by TaqMan® real time PCR. The positive samples for both molecular methods were analyzed for 5’end ORF2 by nested (for GI) and semi nested (for GII) in order to obtain amplicon for identification of circulating strains, being further purified using a commercial kit and submitted to molecular characterization in the automated sequencer. The obtained sequences were edited, aligned and compared to others available in gene bank (NCBI) and in the site NoV genotyping tool. In the period of November 2008 to October 2010, 168 water and sewage samples were collected and analyzed for NoV presence, obtaining a positivity of 33.9% (57/168) of which 21.1% (12/57) were positive only by TaqMan® real time PCR, 19.3% (11/57) only by semi nested and 59.6% (34/57) for both. Considering the two methodologies used, in the positive cases GI (82.5% - 47/57) was most frequent than GII (79.0% - 45/57). However, in most samples there was coexistence of the two genogroups (61.4% - 35/57), mainly in the Tucunduba and SLP-UNA samples, considered the most NoV contaminated sites. On the other hand, in WTP-Bolonha this agent was not found. Of 57 positive samples by TaqMan® real time PCR and/or semi nested RT-PCR, 53 were retested for 5’end ORF2, since four samples showed insufficient quantity of material which allowed a new analyze, so, in 47.2% (25/53) the NoV genome was detected, of these 12% (3/25) belonging to GI, 24% (6/25) to GII and 64% (16/25) for both. The most frequent GI and GII genotypes were GI.8 (n=8) and GII.4 (n=12), respectively, but others genotypes were also observed with lower incidence as GII.6 (n=3), GII.9 (n=2), GII.12 (n=1), GII.14 (n=1), GI.1 (n=1) and GI.4 (n=2). Due to low quality of sequences obtained, eight samples could not be genotyped for GI and three for GII. Of 96 samples with concentration of thermotolerant coliforms above the recommended, 34 (35.4%) were also NoV positive. Increase on conductivity and total dissolved solids was observed in materials from Ver-o-Peso and Tucunduba, as well as the turbidity was notably higher in these places and the Acai’s Port. In the less rainy period (July to November) there was a trend in positivity increasing for NoV, and in the highest rainfall (December to June) a decrease in the incidence of this agent was noted. The results obtained in the present study indicate the circulation of NoV GI and GII in aquatic environments in Belem, revealing the degradation that these water bodies have suffered, as a result of poverty or lack of sanitation in our city, allowing the permanence of pathogens in these ecosystems, along with its effluents.