Detection of a novel species A, DS-1-like, G4P[6] rotavirus strain from a Brazilian child with gastroenteritis.

Rotaviruses belonging to species A (RVA) remain among the most common causes of severe gastroenteritis in children aged <5 years, leading to substantial morbidity and mortality worldwide. Genome reassortment events between two human strains or human and animal strains represent one of the mechanisms which appear to generate the broad genetic variability of circulating. According to a nucleotide, sequence-based classification system, RVA strains are currently classified into three genotype constellations including Wa-like (genogroup I), DS-1-like (genogroup II), and AU-like (genogroup III). The present study reports the detection of an unusual RVA G4P[6] strain (coded as strain HSE005), which might have originated from a natural reassortment event between human and animal RVA strains. Molecular characterization of this isolate showed that it belonged to genogroup II, genotype G4P[6]. In addition, two genes (VP3 and NSP4) of this strain denoted evidence of reassortment events involving strains of distinct zoonotic evolutionary origins. Therefore, we propose that a new G4P[6] strain was identified, highlighting a possible first zoonotic transmission including a reassortment event that involved the VP3 gene.

The burden of norovirus disease in children: a multi-country study in Chile, Brazil, Thailand and the Philippines.

BACKGROUND: Noroviruses (NoVs) cause acute gastroenteritis (AGE) worldwide, affecting children in particular. We aimed to estimate the burden of disease due to NoV among children aged <6 years in Brazil, Chile, Philippines and Thailand. METHODS: This was a prospective, hospital-based, observational study. Children were recruited over one year between 2014 and 2017. Four cohorts were analysed: community-acquired AGE outpatients and inpatients, nosocomial AGE inpatients, and asymptomatic outpatients. We collected demographic and clinical data, and a stool sample that was tested for NoV. Positive samples were tested for Rotavirus (RV) and NoV-genotyped. Disease severity was assessed by the Vesikari and modified Vesikari scores. Prevalence and incidence of NoV-AGE were estimated by cohort and country. RESULTS: 1637 participants yielded valid laboratory results. The proportion of NoV-positive cases was 23.8% (95% CI 20.8-27.2) in the outpatient cohort, 17.9% (15.0-21.3) in the hospital cohort, 21.4% (12.7-33.8) in the nosocomial cohort and 9.6% (6.9-13.2) in the asymptomatic cohort. Genotype GII.4 was predominant (58%). Less than 4% samples had RV coinfection. In general, NoV-positive subjects had more severe presentations than NoV-negative subjects. CONCLUSIONS: NoV caused AGE with substantial burden throughout the studied settings, with higher relative frequency in Brazil where RV vaccination coverage is high.

Rotavirus antigenemia as a common event among children hospitalised for severe, acute gastroenteritis in Belém, northern Brazil.

BACKGROUND: Rotavirus antigenemia and RNAemia (the presence of rotavirus RNA in serum) have been commonly identified among paediatric patients with acute gastroenteritis. In this study we examined the association between rotavirus antigenemia and clinical features, and sought to determine the genotypes of rotaviruses detected in paired stool and serum samples. METHODS: Paired stool and serum samples were obtained from children hospitalised for acute gastroenteritis in Belém, Brazil, between June 2012 and June 2015. The 20-point Vesikari scoring system was used to assess the disease severity upon a retrospective medical record review. Stool and serum samples were primarily screened for the presence of rotavirus antigen using a commercial ELISA assay. The rotavirus isolates from stool and serum samples were genotyped by using the classical reverse-transcriptase polymerase chain reaction (RT-PCR) and/or through nucleotide sequencing of VP4 and VP7 genes. Viral load was estimated using real-time RT-PCR. RESULTS: In total rotavirus antigen was detected in 109 (24.2%) stool samples from 451 children, whereas antigenemia occurred in 38.5% (42/109) of these patients. We demonstrated that patients positive for rotavirus RNA in paired stool and serum samples were more likely to have a higher frequency of vomiting episodes in a 24-h period (p = 0.0035). Our findings also suggested that children not vaccinated against rotavirus are more likely to develop antigenemia, as compared to those given at least one vaccine dose (p = 0.0151). G12P [8] and G2P [4] genotypes were predominant throughout the study period, accounting for 52.3% (57/109) and 27.5% (30/109) of the typed isolates, respectively. Ten stool-serum pairs could be typed for VP4 and VP7 genes. Seven of these pairs showed concordant results with G2P [4] genotype being detected in stool and serum samples, whereas discrepancies between genotypes (G2P [4]/G2P[NT] and G12P [8]/G2P[NT]) were seen in three pairs. CONCLUSIONS: Rotavirus antigenemia and RNAemia occur in a significant number of children hospitalised for acute gastroenteritis in Belém, Brazil, and may contribute to a greater disease severity, particularly translated into a greater number of vomiting episodes. This study documented a high concordance of genotypes detected in a subgroup of paired stool and serum samples.

Emergence of G12P[6] rotavirus strains among hospitalised children with acute gastroenteritis in Belém, Northern Brazil, following introduction of a rotavirus vaccine.

Species A rotavirus still remains a major cause of acute gastroenteritis in infants and young children. Globally, six genotypes (G1P[8], G2P[4], G3P[8], G4P[8], G9P[8] and G12P[8]) account for >90% of circulating strains; however, genotype G12 in combination with P[6] or P[9] has been detected at increasing rates. We sought to broaden our knowledge about the rotavirus strains circulating during the early post-vaccine-introduction period. Stool samples were obtained from children hospitalised for acute gastroenteritis in Belém, Northern Brazil, from May 2008 to May 2011 and examined by reverse transcription polymerase chain reaction and nucleotide sequencing. A total of 122 out of the original 1076 rotavirus strains were judged to be non-typeable in the first analysis and were therefore re-examined. G2P[4] was the most prevalent genotype (58.0%), followed by G1P[8] (16.9%), and G12P[6] (7.5%). G12P[6] strains were identified at similar rates during the first (2.5%) and second (3.9%) years, and the rate jumped to 15.6% in the third year. Analysis of VP7 sequences of the G12P[6] strains showed that they belonged to lineage III. In addition, co-circulating G12P[6] strains displaying long and short RNA patterns were found to belong to the Wa-like and DS-1-like constellation, respectively. Additional unusual circulating strains G12P[9] and G3P[9] were also identified. This hospital-based study showed a high prevalence of G12P[6] strains in the third year of surveillance. Our results highlight the need for continuous longitudinal monitoring of circulating rotavirus strains after introduction of rotavirus vaccines in Brazil and elsewhere.

Detection and genotyping of enteric viruses in hospitalized children with acute gastroenteritis in Belém, Brazil: Occurrence of adenovirus viremia by species F, types 40/41.

Enteric adenovirus (AdV), sapovirus (SaV), and human astrovirus (HAstV) are important pathogens involved in the gastroenteritis etiology. In this study, a total of 219 fecal samples and sera were collected from children hospitalized for acute gastroenteritis (AGE) in two large pediatric hospitals in Belém, from March 2012 to April 2015. The samples were analyzed by polymerase chain reaction (PCR) for AdV and HAstV (astrovirus) detection, and Nested-PCR and qPCR for SaV detection. AdV was detected in 50.2% (110/219) of the cases, with 42.7% (47/110) being sequenced and classified as: species F (63.9% - 30/47), A (4.2% - 2/47), B (6.4% - 3/47), C (17.1% - 8/47), D (4.2% - 2/47), and E (4.2% - 2/47). Of the 110 AdV-positive feces samples, 80 paired sera presented sufficient amounts and were also tested for this virus, of which 51 (63.7%) showed positive results and 26 (70.3%) pairs (feces plus sera) presented concordant results after sequencing being classified as: species F (21/26; 80.8%), A (1/26; 3.8%), B (1/26; 3.8%), and C (3/26; 11.5%). Overall, HAstV rate in the feces samples was 1.8% (4/219), including both HAstV-1a (2/4; 50%) and HAstV-2c (2/4; 50%). SaV was detected in 4.6% (10/219) of the fecal samples, out of which 50% (5/10) of the positive samples were characterized into the genogroups GI.1 (1), GI.2 (2), and GII.4 (2). These findings highlighted the important contributions of AdV, HAstV, and SaV in the enteric virus spectrum in our region and showed the high genetic diversity of AdV. In addition, it demonstrated for the first time in Brazil, the circulation of AdV in the serum of hospitalized children with AGE.

Potential role of dengue virus, chikungunya virus and Zika virus in neurological diseases.

This study showed that laboratory markers of recent infection by dengue, Zika or chikungunya arboviruses were detected in the biological samples of approximately one-third of patients with encephalitis, myelitis, encephalomyelitis or Guillain-Barré syndrome, in a surveillance programme in Piauí state, Brazil, between 2015-2016. Fever and myalgia had been associated with these cases. Since in non-tropical countries most infections or parainfectious diseases associated with the nervous system are attributed to herpesviruses, enteroviruses, and Campylobacter jejuni, the present findings indicate that in tropical countries, arboviruses may now play a more important role and reinforce the need for their surveillance and systematic investigation in the tropics.

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

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.

Low genetic diversity of the Human T-cell Lymphotropic Virus (HTLV-1) in an endemic area of the Brazilian Amazon basin.

The Human T-cell Lymphotropic Virus (HTLV-1) is a Deltaretrovírus that was first isolated in the 1970s, and associated with Adult T-cell Leucemia-Lymphoma (ATLL), and subsequently to Tropical Spastic Paraparesis-Myelopathy (TSP/HAM). The genetic diversity of the virus varies among geographic regions, although its mutation rate is very low (approximately 1% per thousand years) in comparison with other viruses. The present study determined the genetic diversity of HTLV-1 in the metropolitan region of Belém, in northern Brazil. Blood samples were obtained from patients at the UFPA Tropical Medicine Nucleus between January 2010 and December 2013. The DNA was extracted and the PX region of the HTLV was amplified using nested PCR. The positive samples were then digested using the Taq1 enzyme for the identification and differentiation of the HTLV-1 and HTLV-2. The 5'LTR region of the positive HTLV-1 samples were amplified by nested PCR, and then sequenced genetically. The phylogenetic analysis of the samples was based on the maximum likelihood method and the evolutionary profile was analyzed by the Bayesian approach. Overall, 78 samples tested positive for HTLV-1, and 44 were analyzed here. The aA (cosmopolitan-transcontinental) subtype was recorded in all the samples. The following evolutionary rates were recorded for the different subtypes-a: 2.10-3, b: 2.69. 10-2, c: 6.23. 10-2, d: 3.08. 10-2, e: 6. 10-2, f: 1.78. 10-3, g: 2.2. 10-2 mutations per site per year. The positive HTLV-1 samples tested in the present study were characterized by their low genetic diversity and high degree of stability.

Potential role of dengue virus, chikungunya virus and Zika virus in neurological diseases

This study showed that laboratory markers of recent infection by dengue, Zika or chikungunya arboviruses were detected in the biological samples of approximately one-third of patients with encephalitis, myelitis, encephalomyelitis or Guillain-Barré syndrome, in a surveillance programme in Piauí state, Brazil, between 2015-2016. Fever and myalgia had been associated with these cases. Since in non-tropical countries most infections or parainfectious diseases associated with the nervous system are attributed to herpesviruses, enteroviruses, and Campylobacter jejuni, the present findings indicate that in tropical countries, arboviruses may now play a more important role and reinforce the need for their surveillance and systematic investigation in the tropics.

Detection and genotyping of human adenovirus and sapovirus in children with acute gastroenteritis in Belém, Pará, between 1990 and 1992: first detection of GI.7 and GV.2 sapoviruses in Brazil.

INTRODUCTION: Acute gastroenteritis (AGE) is one of the most common causes of morbidity and mortality, especially among children from developing countries. Human adenovirus (HAdV) and sapovirus (SaV) are among the agents that cause AGE. The present study aimed to detect and genotype HAdV and SaV in 172 fecal samples from children with AGE, collected during a surveillance study carried out in a low-income community in Belém, Pará, between 1990 and 1992. METHODS: HAdV was detected by nested PCR, using primers Hex1deg/Hex2deg and NeHex3deg/NeHex4deg. SaV was assayed by reverse transcription PCR (RT-PCR), nested PCR, and quantitative PCR. The nucleotide sequence was determined by direct cycle sequencing. RESULTS: Overall, 43% (74/172) of samples were positive for HAdV, of which 70.3% (52/74) were sequenced and classified as belonging to five different species, mostly A and F. For SaV, positivity was 5.2% (9/172) and genotypes GI.1, GI.7, GII.1, and GV.2 were detected. CONCLUSIONS: The present results reinforce the need for further studies to obtain epidemiological data about the circulation of these viruses in Brazil, especially in the Amazon Region, where data from the early 1990's are scarce. Furthermore, the study describes for the first time the detection of SaV genotypes GI.7 and GV.2 in Brazil, showing that these types circulated in the region more than 25 years ago.

Detection and genotyping of human adenovirus and sapovirus in children with acute gastroenteritis in Belém, Pará, between 1990 and 1992: first detection of GI. 7 and GV. 2 sapoviruses in Brazil

Abstract INTRODUCTION: Acute gastroenteritis (AGE) is one of the most common causes of morbidity and mortality, especially among children from developing countries. Human adenovirus (HAdV) and sapovirus (SaV) are among the agents that cause AGE. The present study aimed to detect and genotype HAdV and SaV in 172 fecal samples from children with AGE, collected during a surveillance study carried out in a low-income community in Belém, Pará, between 1990 and 1992. METHODS: HAdV was detected by nested PCR, using primers Hex1deg/Hex2deg and NeHex3deg/NeHex4deg. SaV was assayed by reverse transcription PCR (RT-PCR), nested PCR, and quantitative PCR. The nucleotide sequence was determined by direct cycle sequencing. RESULTS: Overall, 43% (74/172) of samples were positive for HAdV, of which 70.3% (52/74) were sequenced and classified as belonging to five different species, mostly A and F. For SaV, positivity was 5.2% (9/172) and genotypes GI.1, GI.7, GII.1, and GV.2 were detected. CONCLUSIONS: The present results reinforce the need for further studies to obtain epidemiological data about the circulation of these viruses in Brazil, especially in the Amazon Region, where data from the early 1990's are scarce. Furthermore, the study describes for the first time the detection of SaV genotypes GI.7 and GV.2 in Brazil, showing that these types circulated in the region more than 25 years ago.

Molecular analysis of norovirus in specimens from children enrolled in a 1982-1986 study in Belém, Brazil: A community-based longitudinal study.

Fecal specimens were collected during a longitudinal, community-based study in the city of Belém, North Brazil, that was conducted over 3 years (October 1982 to March 1986), in which 20 children were included from birth to 3 years of age. A total of 229 fecal samples were screened by real time RT-PCR targeting the junction region (ORF 1/2) of the norovirus (NoV) genome. NoV-positive samples were subjected to PCR and sequencing of the viral polymerase (ORF1) and viral protein 1 (VP1) genes (ORF2). The junction region was also sequenced to assess for recombination when ORF1 and ORF2 genotyping results were dissimilar. Samples classified as GII.P4/GII.4 were further characterized by sequencing the P2 subdomain of the viral capsid to determine possible alterations. An overall positivity of 16.1% (37/229) was observed, including GI (16.2%-6/37) and GII (83.8%-31/37) genogroups. Cases of NoV reinfection in at least 2-month intervals were observed, and 12 children developed at least one case of asymptomatic NoV infection. In total, 48.6% (18/37) NoV-positive samples were subjected to nucleotide sequencing analysis targeting the following polymerase genes: GI.P3 (n = 1), GII.Pa (n = 1), GII.Pc (n = 1), GII.P4 (n = 5), GII.P6 (n = 5), GII.P7 (n = 3), GII.P12 (n = 1), and GII.P22 (n = 1). For the VP1 gene, characterization was performed in 14 (77.8%) samples: GI.3 (n = 1), GII.2 (n = 1), GII.4 (n = 4), GII.6 (n = 4), GII.7 (n = 1), GII.12 (n = 1), GII.14 (n = 1), and GII.23 (n = 1). Recombination events were confirmed in three cases (GII.P12/GII.2, GII.P7/GII.14, and GII.Pa/GII.12), and four samples genotyped as GII.P4/GII.4 were analyzed to identify variants. None had contemporary counterparts. Three children developed consecutive NoV infections by different genotypes. The present report documents the importance of NoV as a cause of childhood infection during a longitudinal study conducted more than 30 years ago.

Molecular analysis of norovirus in specimens from children enrolled in a 1982-1986 study in Belém, Brazil: A community-based longitudinal study.

Fecal specimens were collected during a longitudinal, community-based study in the city of Belém, North Brazil, that was conducted over 3 years (October 1982 to March 1986), in which 20 children were included from birth to 3 years of age. A total of 229 fecal samples were screened by real time RT-PCR targeting the junction region (ORF 1/2) of the norovirus (NoV) genome. NoV-positive samples were subjected to PCR and sequencing of the viral polymerase (ORF1) and viral protein 1 (VP1) genes (ORF2). The junction region was also sequenced to assess for recombination when ORF1 and ORF2 genotyping results were dissimilar. Samples classified as GII.P4/GII.4 were further characterized by sequencing the P2 subdomain of the viral capsid to determine possible alterations. An overall positivity of 16.1% (37/229) was observed, including GI (16.2%-6/37) and GII (83.8%-31/37) genogroups. Cases of NoV reinfection in at least 2-month intervals were observed, and 12 children developed at least one case of asymptomatic NoV infection. In total, 48.6% (18/37) NoV-positive samples were subjected to nucleotide sequencing analysis targeting the following polymerase genes: GI.P3 (n = 1), GII.Pa (n = 1), GII.Pc (n = 1), GII.P4 (n = 5), GII.P6 (n = 5), GII.P7 (n = 3), GII.P12 (n = 1), and GII.P22 (n = 1). For the VP1 gene, characterization was performed in 14 (77.8%) samples: GI.3 (n = 1), GII.2 (n = 1), GII.4 (n = 4), GII.6 (n = 4), GII.7 (n = 1), GII.12 (n = 1), GII.14 (n = 1), and GII.23 (n = 1). Recombination events were confirmed in three cases (GII.P12/GII.2, GII.P7/GII.14, and GII.Pa/GII.12), and four samples genotyped as GII.P4/GII.4 were analyzed to identify variants. None had contemporary counterparts. Three children developed consecutive NoV infections by different genotypes. The present report documents the importance of NoV as a cause of childhood infection during a longitudinal study conducted more than 30 years ago.

Efficacy, safety and effectiveness of licensed rotavirus vaccines: a systematic review and meta-analysis for Latin America and the Caribbean.

BACKGROUND: RotaTeq™ (RV5; Merck & Co. Inc., USA) and Rotarix™ (RV1, GlaxoSmithKline, Belgium) vaccines, developed to prevent rotavirus diarrhea in children under five years old, were both introduced into national immunization programs in 2006. As many countries in Latin America and the Caribbean have included either RV5 or RV1 in their routine childhood vaccination programs, we conducted a systematic review and meta-analysis to analyze efficacy, safety and effectiveness data from the region. METHODS: We conducted a systematic search in PubMed, EMBASE, Scielo, Lilacs and the Cochrane Central Register, for controlled efficacy, safety and effectiveness studies published between January 2000 until December 2011, on RV5 and RV1 across Latin America (where both vaccines are available since 2006). The primary outcome measures were: rotavirus-related gastroenteritis of any severity; rotavirus emergency department visits and hospitalization; and severe adverse events. RESULTS: The results of the meta-analysis for efficacy show that RV1 reduced the risk of any-severity rotavirus-related gastroenteritis by 65% (relative risk (RR) 0.35, 95% confidence interval (CI) 0.25; 0.50), and of severe gastroenteritis by 82% (RR 0.18, 95%CI 0.12; 0.26) versus placebo. In trials, both vaccines significantly reduced the risk of hospitalization and emergency visits by 85% (RR 0.15, 95%CI 0.09; 0.25) for RV1 and by 90% (RR 0.099, 95%CI 0.012; 0.77) for RV5. Vaccination with RV5 or RV1 did not increase the risk of death, intussusception, or other severe adverse events which were previously associated with the first licensed rotavirus vaccine. Real-world effectiveness studies showed that both vaccines reduced rotavirus hospitalization in the region by around 45-50% for RV5 (for 1 to 3 doses, respectively), and, by around 50-80% for RV1 (for 1 to 2 doses, respectively). For RV1, effectiveness against hospitalization was highest (around 80-96%) for children vaccinated before 12 months of age, compared with 5-60% effectiveness in older children. Both vaccines were most effective in preventing more severe gastroenteritis (70% for RV5 and 80-90% for RV1) and severe gastroenteritis (50% for RV5 and 70-80% for RV1). CONCLUSION: This systematic literature review confirms rotavirus vaccination has been proven effective and well tolerated in protecting children in Latin America and the Caribbean.

An Analysis of the Associations among Cognitive Impulsiveness, Reasoning Process, and Rational Decision Making.

Impulsivity may lead to several unfortunate consequences and maladaptive behaviors for both clinical and nonclinical people. It has a key role in many forms of psychopathology. Although literature has discussed the negative impact of impulsivity, few have emphasized the relationship between cognitive impulsiveness and decision making. The aim of this study is to investigate the effects of cognitive impulsiveness on decision making and explore the strategies used by participants to solve problems. For this purpose, we apply two measures of impulsivity: the self-report Barratt Impulsiveness Scale (BIS-11) and the performance based Cognitive Reflection Test (CRT). Moreover, we evaluate participants' reasoning processes employed to answer CRT questions based on the calculation expressions, data organization, and erasures they made while answering the CRT (note that we utilized the instruments using pen and paper). These reasoning processes are related to the role of executive functions in decision making, and its relationship with impulsiveness. The sample consists of 191 adults, who were either professionals or undergraduate students from the fields of business, management, or accounting. The results show that cognitive impulsiveness may negatively affect decision making, and that those who presented the calculation to answer the CRT questions made better decisions. Moreover, there was no difference in the strategies used by impulsive vs. nonimpulsive participants during decision making. Finally, people who inhibited their immediate answers to CRT questions performed better during decision making.

Detection and genetic characterization of the emergent GII.17_2014 norovirus genotype among children with gastroenteritis from Northern Brazil.

Norovirus is the most important cause of viral gastroenteritis outbreaks worldwide. Recently, a novel GII.17 norovirus variant emerged and caused epidemics in Asian countries, replacing the GII.4 Sydney 2012 strain in hospitalized cases. In this study we describe the emergence of this novel NoV GII.17_2014 strain in Brazil.

Analysis of a genotype G3P[9] rotavirus a strain that shows evidence of multiple reassortment events between animal and human rotaviruses.

The species A rotaviruses (RVA) are important gastroenteric pathogens that infect humans and animals. RVA genotype G3P[9] has been described in human-animal reassortment events, and the complexity of its hosts motivates the genetic investigation of this strain. Therefore, the aim of this study is to analyse a G3P[9] sample that was detected in a child with acute gastroenteritis. The 1A3739 sample featured the constellation G3P[9]-I18-R3-C3-Mx-A19-N3-T3-E3-H6. The sequence for VP3 gene was not obtained. The phylogeny showed a closer relationship among genes VP7, VP1, NSP3, NSP4, and NSP5 with genes of animal origin, such as chiropter, alpaca, equine, and simian. In addition, the genes VP6 and NSP1 belong to the new genotypes I18 and A19, respectively. The emergence of strains such as these can interfere with the effectiveness of the RVA vaccine, and continuous monitoring is therefore important. Additional studies are needed to determine the evolutionary source and to identify a possible reservoir of RVA in nature.

Detection of a novel equine-like G3 rotavirus associated with acute gastroenteritis in Brazil.

Genotype G3P[8] of rotavirus A (RVA) is detected worldwide, usually associated with Wa-like constellation and exhibiting a long RNA migration pattern. More recently, a novel inter-genogroup, G3P[8] reassortant variant with a short electropherotype, has emerged in Asia, Oceania and Europe, denoting an overall potential of unusual rotavirus strains. During a RVA surveillance in Brazil, G3P[8] strains were found displaying a short electropherotype pattern, which had not been detected before in this region. This study aims to characterize the complete genome of 10 G3P[8] strains detected in the northern region of Brazil. All G3P[8] samples were subjected to partial sequencing, and the whole-genome phylogenetic analysis demonstrated that all strains possessed I2-R2-C2-M2-A2-N1-T2-E2-H2 genotype background, representing reassortants with an equine-like G3 VP7 and amino acid changes in VP4 and VP7 antigenic regions as compared to vaccine strains. Phylogenetic analysis demonstrated high nucleotide identity in almost all RNA segments of G3P[8] DS-1 samples detected in Asia, Oceania and Europe as well as G3P[4] strains in Japan. This study reports a novel, equine-like G3P[8] strain circulating in Brazil and isolated from children hospitalized for severe gastroenteritis, and highlights the complex dynamics of RVA molecular epidemiology. Our findings point to a novel RVA strain emerging in this region, and studies should be done to detect whether this may represent a challenge to current vaccine strategies.

Caliciviruses in hospitalized children, São Luís, Maranhão, 1997-1999: detection of norovirus GII. 12

ABSTRACT Gastroenteritis is one of the most common diseases during childhood, with norovirus (NoV) and sapovirus (SaV) being two of its main causes. This study reports for the first time the incidence of these viruses in hospitalized children with and without gastroenteritis in São Luís, Maranhão. A total of 136 fecal samples were tested by enzyme immunoassays (EIA) for the detection of NoV and by reverse transcription-polymerase chain reaction (RT-PCR) for detection of both NoV and SaV. Positive samples for both agents were subjected to sequencing. The overall frequency of NoV as detected by EIA and RT-PCR was 17.6% (24/136) and 32.6% (15/46), respectively in diarrheic patients and 10.0% (9/90) in non-diarrheic patients (p < 0.01). Of the diarrheic patients, 17% had fever, vomiting and anorexia, and 13% developed fever, vomiting and abdominal pain. Of the 24 NoV-positive samples, 50% (12/24) were sequenced and classified as genotypes GII.3 (n = 1), GII.4 (6), GII.5 (1), GII.7 (2), GII.12 (1) and GII.16 (1). SaV frequency was 9.8% (11/112), with 22.6% (7/31) in diarrheic patients and 4.9% (4/81) in nondiarrheic (p = 0.04) ones. In diarrheic cases, 27.3% had fever, vomiting and anorexia, whereas 18.2% had fever, anorexia and abdominal pain. One SaV-positive sample was sequenced and classified as GII.1. These results show a high genetic diversity of NoV and higher prevalence of NoV compared to SaV. Our data highlight the importance of NoV and SaV as enteropathogens in São Luís, Maranhão.

Characterization of novel intragenotype recombination events among norovirus pandemic GII.4 variants.

Recently, there has been an increase in the number of children hospitalized due to norovirus infection in Brazil. This is due both to the occurrence of more severe norovirus-related gastroenteritis cases after the introduction of the rotavirus vaccine and an increase in the tools for the detection of the disease. This pathogen is transmitted by the fecal-oral route, and the illness is characterized by diarrhea, vomiting, nausea and abdominal cramps. The genome of the virus is organized into three open reading frames showing strong mutation rates. Additionally, homologous recombination events, which can increase the virulence of the virus and lead to genotyping mistakes in molecular epidemiological studies, frequently occur. The purpose of this study was to describe two recombination events among different GII.4 variants that infected children who were hospitalized for severe acute gastroenteritis during distinct periods of time in Belém, Brazil. The recombination among the variants US95_96/Kaiso_2003 and Den Haag_2006b/Yerseke_2006a were observed in May 2003 and February 2009, respectively. In both cases, the association between the dominant variant at that point in time and another that was circulating at a low frequency in the population of Belém was demonstrated. Interestingly, the position of the breakpoint of the recombination event in the genome was the polymerase gene and was located at the nucleotide positions 4.834 and 5.002, which is an unusual location for the occurrence of recombination as other studies have previously reported the junction region as a breakpoint. In this study, both recombinant variant strains were related to severe cases of diarrhea that lead to hospitalization, demonstrating the viral evolution of GII.4 in response to selective pressures, which ultimately lead to the emergence of novel viral types in the pediatric population. The cases discussed here reinforce the need for continuous norovirus surveillance. To our knowledge, these two GII.4 variant recombinations have not yet been previously described.