Rotaviruses A and C in dairy cattle in the state of Rio de Janeiro, Brazil.

Stool samples were collected from calves from nine family-based small dairy farms in the state of Rio de Janeiro, for detection and characterization of rotavirus (RV) species A, B, and C (RVA, RVB, and RVC, respectively) by reverse transcription polymerase chain reaction. Twenty-six samples (27.7%) were positive for at least one of the species: 22 (23.4%) samples were positive only for RVA, 3 (3.2%) were positive for RVC, and one sample (1.1%) had co-infection of RVA and RVC. RVB was not detected. Seven (21.9%; n = 32) animals with diarrhea and 19 (30.1% n = 62) asymptomatic animals were positive, with no significant difference in positivity (p = 0.3677). RV was detected in all properties studied, at rates between 14.3 and 80%, demonstrating the widespread circulation of RV in four of the seven geographic regions of the state of Rio de Janeiro. Infection was more prevalent among animals ≤ 6 months of age. Sequence analysis of a portion of the RVA VP6-encoding gene identified the I2 genotype. RVC was also detected; to our knowledge, this is the first description of this agent in cattle in Brazil. The data presented here should add knowledge regarding the importance and prevalence of RV in our national territory, and may facilitate the planning and implementation of control and prevention measures for bovine rotavirus infections in Brazil.

Induction of Cell Death in the Human Acute Lymphoblastic Leukemia Cell Line Reh by Infection with Rotavirus Isolate Wt1-5.

Cancer is a major health problem that poses a great challenge to health care systems worldwide. Tools for cancer treatment have rapidly advanced in recent years, resulting in therapeutic strategies which are alternative and complementary to conventional treatment. To identify the cell surface receptors used by a tumor cell-adapted rotavirus and the cell death markers induced by its infection, we use Wt1-5, a rotavirus isolate recently adapted to tumor cells, to infect the human acute lymphoblastic leukemia cell line, Reh. The expression of cell surface receptors used by Wt1-5 was determined using flow cytometry and an antibody blocking assay to test for their implication in virus infection. Viral antigens and cell death markers induced by rotavirus infection were followed by flow cytometric analysis. The present study showed that rotavirus Wt1-5 was able to use cell surface proteins such as heat shock proteins (HSPs) 90, 70, 60 and 40, Hsc70, PDI and integrin ß3. Rotavirus Wt1-5 induced cytotoxic effects including changes in cell membrane permeability, alteration of mitochondrial membrane potential, DNA fragmentation and activation of cell death signaling. Wt1-5 deserves to be further studied as a candidate oncolytic agent due to its ability to induce apoptosis in lymphoblastic leukemia-derived cells.

Characterization of rotavirus possessing a DS-1-like VP3 gene from pigs in Brazil: Evidence for zooanthroponotic transmission.

Porcine group A rotavirus (RVA) strains SUI15A and SUI24A are suggested to have VP3 genes of human origin possessing DS-1-like backbone. The aim of the present study was to analyse the genome of two strains (SUI15A and SUI24A) and understand the evolution of a rare human-like M2 genotype in pigs. On partial genomic analysis, strains SUI24A (G3-P[13]-I5-R1-C1-M2-A8-N1-T7-E1-H1) and SUI15A (G3-P[x]-Ix-R1-C1-M2-Ax-Nx-T7-E1-H1) were found to have VP3 gene RVA different from those of typical porcine RVA strains described in Brazil and worldwide. This genotypic constellation was a novel constellation that has not been reported previously in both humans and pigs. Furthermore, on phylogenetic analysis, VP3 gene of strains appeared to be of human origin. Therefore, suggested to have evidence for human-to-porcine zooanthroponotic transmission.

Cynomolgus Monkeys (Macaca fascicularis) as an Experimental Infection Model for Human Group A Rotavirus.

Group A rotaviruses (RVA) are one of the most common causes of severe acute gastroenteritis in infants worldwide. Rotaviruses spread from person to person, mainly by faecal⁻oral transmission. Almost all unvaccinated children may become infected with RVA in the first two years of life. The establishment of an experimental monkey model with RVA is important to evaluate new therapeutic approaches. In this study, we demonstrated viral shedding and viraemia in juvenile⁻adult Macaca fascicularis orally inoculated with Wa RVA prototype. Nine monkeys were inoculated orally: seven animals with human RVA and two control animals with saline solution. During the study, the monkeys were clinically monitored, and faeces and blood samples were tested for RVA infection. In general, the inoculated animals developed an oligosymptomatic infection pattern. The main clinical symptoms observed were diarrhoea in two monkeys for three days, associated with a reduction in plasmatic potassium content. Viral RNA was detected in seven faecal and five sera samples from inoculated animals, suggesting virus replication. Cynomolgus monkeys are susceptible hosts for human Wa RVA infection. When inoculated orally, they presented self-limited diarrhoea associated with presence of RVA infectious particles in faeces. Thus, cynomolgus monkeys may be useful as animal models to evaluate the efficacy of new antiviral approaches.

Inflammatory and oxidative stress in rotavirus infection.

Rotaviruses are the single leading cause of life-threatening diarrhea affecting children under 5 years of age. Rotavirus entry into the host cell seems to occur by sequential interactions between virion proteins and various cell surface molecules. The entry mechanisms seem to involve the contribution of cellular molecules having binding, chaperoning and oxido-reducing activities. It appears to be that the receptor usage and tropism of rotaviruses is determined by the species, cell line and rotavirus strain. Rotaviruses have evolved functions which can antagonize the host innate immune response, whereas are able to induce endoplasmic reticulum (ER) stress, oxidative stress and inflammatory signaling. A networking between ER stress, inflammation and oxidative stress is suggested, in which release of calcium from the ER increases the generation of mitochondrial reactive oxygen species (ROS) leading to toxic accumulation of ROS within ER and mitochondria. Sustained ER stress potentially stimulates inflammatory response through unfolded protein response pathways. However, the detailed characterization of the molecular mechanisms underpinning these rotavirus-induced stressful conditions is still lacking. The signaling events triggered by host recognition of virus-associated molecular patterns offers an opportunity for the development of novel therapeutic strategies aimed at interfering with rotavirus infection. The use of N-acetylcysteine, non-steroidal anti-inflammatory drugs and PPARγ agonists to inhibit rotavirus infection opens a new way for treating the rotavirus-induced diarrhea and complementing vaccines.

Rotavirus vaccine stability in the aquatic environment.

AIMS: To evaluate the thermal and length of stability of the Rotaviruses (RV) vaccine (RotaTeq) in the aquatic environment. METHODS AND RESULTS: Surface freshwater, brackish and drinking water were spiked with RV vaccine strain and stored at 22 and 4°C. The virus infectivity and genome persistence were evaluated by plaque assay and RT-qPCR, respectively, up to 180 days. Infectious RV vaccine particles showed to be less stable in the brackish water matrix than in surface and drinking water either at 22 or 4°C. The estimated T90 values obtained by the linear regression model were 18, 55 and 59 days, respectively for brackish, surface and drinking water stored at 22°C and 68, 154 and 240 days at 4°C. As expected, the genome persistence showed to be less affected by length and temperature of storage in all the matrices evaluated. CONCLUSIONS: The evidence of high stability of the RV vaccine in water matrices reinforces the importance for surveillance of RV vaccines strains in the environment regarding the potential occurrence of unexpected infections and virus genomic reassortments. SIGNIFICANCE AND IMPACT OF THE STUDY: The presence of reassortants and the shedding of the live attenuated vaccine strains after vaccination can compromise the vaccine safety by introducing new viral variants in the environment.

Whole-genomic analysis of 12 porcine group A rotaviruses isolated from symptomatic piglets in Brazil during the years of 2012-2013.

Group A rotaviruses (RVAs) are leading causes of viral diarrhea in children and in the young of many animal species, particularly swine. In the current study, porcine RVAs were found in fecal specimens from symptomatic piglets on 4 farms in Brazil during the years of 2012-2013. Using RT-PCR, Sanger nucleotide sequencing, and phylogenetic analyses, the whole genomes of 12 Brazilian porcine RVA strains were analyzed. Specifically, the full-length open reading frame (ORF) sequences were determined for the NSP2-, NSP3-, and VP6-coding genes, and partial ORF sequences were determined for the VP1-, VP2-, VP3-, VP4-, VP7-, NSP1-, NSP4-, and NSP5/6-coding genes. The results indicate that all 12 strains had an overall porcine-RVA-like backbone with most segments being designated as genotype 1, with the exception of the VP6- and NSP1-coding genes, which were genotypes I5 and A8, respectively. These results add to our growing understanding of porcine RVA genetic diversity and will provide a platform for monitoring the role of animals as genetic reservoirs of emerging human RVAs strains.

Prevalence and genomic characterization of G2P[4] group A rotavirus strains during monovalent vaccine introduction in Brazil.

This study aims to: estimate the prevalence of G2P[4] rotaviruses in Brazil between 2001-2011 from patients with acute gastroenteritis; perform phylogenetic analyses of G2P[4] Brazilian strains (from vaccinated and non-vaccinated children) based on VP7 and VP8(∗) encoding genes and analyze the antigenic regions of these proteins comparing with RV1; and assess the full genetic background of eleven selected Brazilian strains. The G2P[4] detection rate among RVA positive samples was 0/157 in 2001, 3/226 (1.3%) in 2002, 0/514 in 2003, 0/651 in 2004, 31/344 (9%)/2005, 112/227 (49%)/2006, 139/211 (66%)/2007, 240/284 (85%)/2008, 66/176 (37.5%)/2009, 367/422 (87%)/2010 and 75/149 (50%)/2011. For the VP7 and VP8(∗) encoding genes, 52 sequences were analyzed and shared up to 99% nucleotide identity with other contemporary G2P[4] strains detected worldwide, grouping into different clusters. Most differences inside antigenic epitopes of VP7 and VP8(∗) have been maintained in the G2P[4] Brazilian strains along the years, and all were present before RV1 introduction. Eleven G2P[4] strains (4-vaccinated/7-non-vaccinated) were completely characterized and possessed the typical DS-1-like genotype constellation (G2-P[4]-I2-R2-C2-M2-A2-N2-T2-E2-H2) sharing up to 99% of nucleotide identity with contemporary worldwide strains. Reassortments between Brazilian G2P[4] human strains were observed. In conclusion, the data obtained in the current study suggests that implementation of RV1 vaccination might not influence the genetic diversity observed in G2P[4] analyzed strains. Several factors might have contributed to the increased prevalence of this genotype in Brazil since 2005: the introduction of RV1 into the Brazilian National Immunization Program has resulted in a decrease in the relative prevalence of predominant Wa-like RVA strains facilitating the increase of the heterotypic (DS-1-like) RVA strain G2P[4] in the Brazilian population; the genetic diversity found in different geographical regions throughout the years before, and after the introduction of RV1; the long period of low or no circulation of this genotype in Brazil previous to RV1 introduction could have created favorable conditions for the accumulation of immunological susceptible individuals.

Severe diarrhea outbreak in beef calves (Bos indicus) caused by G6P[11], an emergent genotype of bovine rotavirus group A / Surto de diarreia neonatal em bezerros de corte (Bos indicus) ocasionado por um genotipo emergente G6P[11] de rotavírus bovino grupo A

The episodes of diarrhea caused by neonatal bovine rotavirus group A (BoRVA) constitute one of the major health problems in the calf rearing worldwide. The main G (VP7) and P (VP4) genotypes of BoRVA strains involved in the etiology of diarrhea in calves are G6P[1], G10P[11], G6P[5], and G8P[1]. However, less frequently, other G and P genotypes have been described in BoRVA strains identified in diarrheic fecal samples of calves. This study describes the identification and molecular characterization of an emerging genotype (G6P[11]) in BoRVA strains involved in the etiology of a diarrhea outbreak in beef calves in a cattle herd of high production in extensive management system. The diarrhea outbreak, which showed high morbidity (60%) and lethality (7%) rates, occurred in calves (n= 384) Nelore (Bos indicus) up to 30-day-old from the State of Mato Grosso do Sul, Brazil. BoRVA was identified in 80% (16/20) of the fecal samples analyzed by polyacrylamide gel electrophoresis (PAGE) technique. In all PAGE-positive fecal samples were amplified products with 1,062-bp and 876-bp in the RT-PCR assays for VP7 (G type) and VP4 (VP8*) (P type) of BoRVA, respectively. The nucleotide sequence analysis of VP7 and VP4 genes of four wild-type BoRVA strains showed G6-III P[11]-III genotype/lineage. The G6P[11] genotype has been described in RVA strains of human and animal hosts, however, in calves this genotype was only identified in some cross-sectional studies and not as a single cause of diarrhea outbreaks in calves with high morbidity and lethality rates as described in this study...

Os episódios de diarreia neonatal ocasionados pelo rotavírus bovino grupo A (BoRVA) constituem-se em um dos principais problemas sanitários na criação de bezerros em todo o mundo. Os principais genotipos G (VP7) e P (VP4) de cepas de BoRVA envolvidos na etiologia da diarreia em bezerros são G6P[1], G10P[11], G6P[5] e G8P[1]. No entanto, com menor frequência, outros genotipos G e P têm sido descritos em cepas de BoRVA identificadas em amostras de fezes diarreicas de bezerros. Este estudo descreve a identificação e caracterização molecular de um genotipo emergente (G6P[11]) em cepas de BoRVA envolvidas na etiologia de um surto de diarreia em bezerros de um rebanho bovino de corte de alta produção em sistema de manejo extensivo. O surto, que apresentou altas taxas de morbidade (60%) e de letalidade (7%), ocorreu em bezerros (n=384) da raça Nelore (Bos indicus) com até 30 dias de idade, provenientes do estado do Mato Grosso do Sul, Brasil. O BoRVA foi identificado em 80% (16/20) das amostras fecais analisadas pela técnica de eletroforese em gel de poliacrilamida (PAGE). Em todas as amostras fecais PAGE-positivas foi possível a amplificação por RT-PCR de produtos com 1.062 pb e 876 pb referentes aos genes VP7 (G tipo) e VP4 (VP8*) (P tipo), respectivamente, de BoRVA. A análise da sequência de nucleotídeos dos genes VP7 e VP4 de quatro cepas de BoRVA demonstrou a presença do genotipo/linhagem G6-III P[11]-III. O genotipo G6P[11] tem sido descrito em cepas de RVA de hospedeiros humanos e animais. Contudo, em bezerros, este genotipo foi apenas identificado em alguns estudos transversais e não como a única causa de surtos de diarreia em bezerros com altas taxas de morbidade e...

Severe diarrhea outbreak in beef calves (Bos indicus) caused by G6P[11], an emergent genotype of bovine rotavirus group A / Surto de diarreia neonatal em bezerros de corte (Bos indicus) ocasionado por um genotipo emergente G6P[11] de rotavírus bovino grupo A

The episodes of diarrhea caused by neonatal bovine rotavirus group A (BoRVA) constitute one of the major health problems in the calf rearing worldwide. The main G (VP7) and P (VP4) genotypes of BoRVA strains involved in the etiology of diarrhea in calves are G6P[1], G10P[11], G6P[5], and G8P[1]. However, less frequently, other G and P genotypes have been described in BoRVA strains identified in diarrheic fecal samples of calves. This study describes the identification and molecular characterization of an emerging genotype (G6P[11]) in BoRVA strains involved in the etiology of a diarrhea outbreak in beef calves in a cattle herd of high production in extensive management system. The diarrhea outbreak, which showed high morbidity (60%) and lethality (7%) rates, occurred in calves (n= 384) Nelore (Bos indicus) up to 30-day-old from the State of Mato Grosso do Sul, Brazil. BoRVA was identified in 80% (16/20) of the fecal samples analyzed by polyacrylamide gel electrophoresis (PAGE) technique. In all PAGE-positive fecal samples were amplified products with 1,062-bp and 876-bp in the RT-PCR assays for VP7 (G type) and VP4 (VP8*) (P type) of BoRVA, respectively. The nucleotide sequence analysis of VP7 and VP4 genes of four wild-type BoRVA strains showed G6-III P[11]-III genotype/lineage. The G6P[11] genotype has been described in RVA strains of human and animal hosts, however, in calves this genotype was only identified in some cross-sectional studies and not as a single cause of diarrhea outbreaks in calves with high morbidity and lethality rates as described in this study. The monitoring of the G and P genotypes of BoRVA strains involved in diarrhea outbreaks in calves is important for both animal and public health by allowing the identification of the most frequent genotypes, the characterization of novel genotypes and to identify reassortments with genotypes described in animal and human hosts. The results of this study show the importance of the monitoring of the genotypes of BoRVA strains involved in episodes of bovine neonatal diarrhea as for characterization of frequency of occurrence and pathogenic potential of uncommon genotypes as for monitoring of the emergency of different BoRVA genotypes not included in commercial vaccines.(AU)

Os episódios de diarreia neonatal ocasionados pelo rotavírus bovino grupo A (BoRVA) constituem-se em um dos principais problemas sanitários na criação de bezerros em todo o mundo. Os principais genotipos G (VP7) e P (VP4) de cepas de BoRVA envolvidos na etiologia da diarreia em bezerros são G6P[1], G10P[11], G6P[5] e G8P[1]. No entanto, com menor frequência, outros genotipos G e P têm sido descritos em cepas de BoRVA identificadas em amostras de fezes diarreicas de bezerros. Este estudo descreve a identificação e caracterização molecular de um genotipo emergente (G6P[11]) em cepas de BoRVA envolvidas na etiologia de um surto de diarreia em bezerros de um rebanho bovino de corte de alta produção em sistema de manejo extensivo. O surto, que apresentou altas taxas de morbidade (60%) e de letalidade (7%), ocorreu em bezerros (n=384) da raça Nelore (Bos indicus) com até 30 dias de idade, provenientes do estado do Mato Grosso do Sul, Brasil. O BoRVA foi identificado em 80% (16/20) das amostras fecais analisadas pela técnica de eletroforese em gel de poliacrilamida (PAGE). Em todas as amostras fecais PAGE-positivas foi possível a amplificação por RT-PCR de produtos com 1.062 pb e 876 pb referentes aos genes VP7 (G tipo) e VP4 (VP8*) (P tipo), respectivamente, de BoRVA. A análise da sequência de nucleotídeos dos genes VP7 e VP4 de quatro cepas de BoRVA demonstrou a presença do genotipo/linhagem G6-III P[11]-III. O genotipo G6P[11] tem sido descrito em cepas de RVA de hospedeiros humanos e animais. Contudo, em bezerros, este genotipo foi apenas identificado em alguns estudos transversais e não como a única causa de surtos de diarreia em bezerros com altas taxas de morbidade e letalidade como descrito neste estudo. O monitoramento dos genotipos G e P de cepas de BoRVA envolvidos em surtos de diarreia em bezerros é relevante tanto para a saúde animal quanto para a saúde pública por possibilitar a identificação dos genotipos mais frequentes, a caracterização de novos genotipos e por identificar reassortments com genotipos descritos em hospedeiros humanos e animais. Os resultados deste estudo demonstram a importância do monitoramento dos genotipos de cepas de BoRVA envolvidas em surtos de diarreia neonatal bovina tanto para a caracterização da frequência de ocorrência e potencial patogênico de genotipos incomuns quanto para o monitoriamento da emergência de genotipos distintos daqueles incluídos em vacinas comerciais.(AU)

A decade of G3P[8] and G9P[8] rotaviruses in Brazil: epidemiology and evolutionary analyses.

This study aims to estimate the frequency of group A rotaviruses (RVA) infection with genotypes G3P[8] and G9P[8] in children that suffered from diarrheal disease (DD) between 2001 and 2011 in different Brazilian regions. In addition, the genetic diversity of G3P[8] and G9P[8] RVA strains recovered from vaccinated and non-vaccinated children was assessed. Laboratory-based RVA surveillance included 15,115 cases of DD, and RVA was detected by enzyme immune-assay and/or polyacrylamide gel electrophoresis in 3357 (22%) samples. RVA was genotyped by the semi-nested RT-PCR and among RVA-positive samples, 100 (2.9%) were G3 (63 G3P[8], 32 G3P not typed [NT], and 5 G3P[6]) and 378 (16.2%) were G9 (318 G9P[8], 59 G9P[NT], and 1 G9P[6]). From the G3 and G9 positive samples, 16 and 12, respectively, were obtained from children aged 4-48months vaccinated with the monovalent vaccine (Rotarix®, RV1). Phylogenetic analyses of the VP7 and VP8(∗) encoding genes were performed for 26 G3P[8] and 48 G9P[8] strains. VP8(∗) phylogenetic analysis revealed that all strains analyzed belonged to P[8] lineage III, whereas RV1 belongs to P[8]-I lineage. VP7 analysis revealed that all G3 and G9 strains belonged to G3-lineage III and G9-lineage III. The comparison of the VP7 and VP8(∗) antigenic epitopes regions of Brazilian strains with RV1 strain revealed several amino acid changes. However, no particular differences among Brazilian strains detected before and after vaccine introduction were observed, or among strains detected from vaccinated and non-vaccinated children. Complete genome characterization of four G3P[8] and seven G9P[8] strains revealed a typical conserved human Wa-like genomic constellation. Changes in the genetic diversity of G3P[8] and G9P[8] RVA detected from 2001 to 2011 in Brazil seemed not be related to RV1 introduction in Brazil.

Diarrhea outbreaks in suckling piglets due to rotavirus group C single and mixed (rotavirus groups A and B) infections / Surtos de diarreia em leitões lactentes por rotavírus grupo C em infecções singulares e mistas (rotavirus grupos A e B)

Porcine group A rotavirus (PoRVA) is a major cause of neonatal diarrhea in suckling and recently weaned piglets worldwide. The involvement of non-group A rotavirus in cases of neonatal diarrhea in piglets are sporadic. In Brazil there are no reports of the porcine rotavirus group C (PoRVC) as etiologic agent of the diarrhea outbreaks in piglets. The aim of this study was to describe the identification of rotavirus group C in single and in mixed infection with rotavirus groups A and B in three neonatal diarrhea outbreaks in suckling (<21-day-old) piglets, with 70 percent to 80 percent and 20 percent to 25 percent of morbidity and lethality rates, respectively, in three pig herds located in the state of Santa Catarina, Brazil. The diagnosis of PoRV in the diarrheic fecal samples was performed using polyacrylamide gel electrophoresis (PAGE) to identify the presence of porcine rotavirus groups A, B (PoRVB), and C, and by RT-PCR (PoRVA and PoRVC) and semi-nested (SN)-PCR (PoRVB) to partially amplify the VP4 (VP8*)-VP7, NSP2, and VP6 genes of PoRVA, PoRVB, and PoRVC, respectively. […] The PoRVB strains (first and second outbreaks) and the PoRVC strains (first, second, and third outbreaks) showed higher nt identity and clustered in the phylogenetic tree with PoRVB and PoRVC strains that belong to the N4 and I1 genotypes, respectively. This is the first description in Brazil of the involvement of PoRVC in the etiology of diarrhea outbreaks in suckling piglets. The results of this study demonstrated that PoRVC, in both single and mixed infections, is an important enteropathogen involved in neonatal diarrhea outbreaks in piglets and that the use of more sensitive diagnostic techniques allows the identification of mixed infections involving two or even three groups of PoRV, which may be more common than previously reported.

O rotavírus suíno grupo A (PoRVA) é uma das principais causas de diarreia neonatal em leitões lactentes e recém-desmamados em todo o mundo. As descrições do envolvimento de rotavírus não-grupo A em quadros de diarreia neonatal em leitões são esporádicas. No Brasil não há relatos do envolvimento do rotavírus suíno grupo C (PoRVC) na etiologia dos surtos de diarreia em leitões. O objetivo deste estudo foi descrever a identificação de rotavírus grupo C em infecções singulares e mistas com os rotavírus grupos A e B em três surtos de diarreia neonatal em leitões lactentes (<21 dias de idade), com taxas de morbidade de 70 por cento a 80 por cento e de letalidade de 20 por cento a 25 por cento, em três rebanhos suínos localizados no estado de Santa Catarina, Brasil. O diagnóstico de PoRV nas amostras de fezes diarreicas foi realizado por eletroforese em gel de poliacrilamida (PAGE) para identificar a presença dos grupos A, B (PoRVB), e C de rotavírus suíno e por RT-PCR (PoRVA e PoRVC) e semi-nested (SN)-PCR (PoRVB) com a amplificação parcial dos genes VP4 (VP8*)-VP7, NSP2 e VP6 de PoRVA, PoRVB e PoRVC, respectivamente. [...] As cepas de PoRVB (primeiro e segundo surtos) e as cepas de PoRVC (primeiro, segundo e terceiro surtos) mostraram maior identidade de nt com cepas de PoRVB e PoRVC que pertencem aos genotipos N4 e I1, respectivamente. Esta é a primeira descrição realizada no Brasil do envolvimento de PoRVC na etiologia de surtos de diarreia em leitões lactentes. Os resultados deste estudo demonstram que o PoRVC, tanto em infecções singulares quanto em infecções mistas, é um importante enteropatógeno envolvido em surtos de diarreia neonatal em leitões e que o uso de técnicas de diagnóstico mais sensíveis permite caracterizar que infecções mistas, com dois ou até mesmo com três grupos de PoRV, podem ser mais comuns do que anteriormente relatado.

Diarrhea outbreaks in suckling piglets due to rotavirus group C single and mixed (rotavirus groups A and B) infections / Surtos de diarreia em leitões lactentes por rotavírus grupo C em infecções singulares e mistas (rotavirus grupos A e B)

Porcine group A rotavirus (PoRVA) is a major cause of neonatal diarrhea in suckling and recently weaned piglets worldwide. The involvement of non-group A rotavirus in cases of neonatal diarrhea in piglets are sporadic. In Brazil there are no reports of the porcine rotavirus group C (PoRVC) as etiologic agent of the diarrhea outbreaks in piglets. The aim of this study was to describe the identification of rotavirus group C in single and in mixed infection with rotavirus groups A and B in three neonatal diarrhea outbreaks in suckling (<21-day-old) piglets, with 70 percent to 80 percent and 20 percent to 25 percent of morbidity and lethality rates, respectively, in three pig herds located in the state of Santa Catarina, Brazil. The diagnosis of PoRV in the diarrheic fecal samples was performed using polyacrylamide gel electrophoresis (PAGE) to identify the presence of porcine rotavirus groups A, B (PoRVB), and C, and by RT-PCR (PoRVA and PoRVC) and semi-nested (SN)-PCR (PoRVB) to partially amplify the VP4 (VP8*)-VP7, NSP2, and VP6 genes of PoRVA, PoRVB, and PoRVC, respectively. […] The PoRVB strains (first and second outbreaks) and the PoRVC strains (first, second, and third outbreaks) showed higher nt identity and clustered in the phylogenetic tree with PoRVB and PoRVC strains that belong to the N4 and I1 genotypes, respectively. This is the first description in Brazil of the involvement of PoRVC in the etiology of diarrhea outbreaks in suckling piglets. The results of this study demonstrated that PoRVC, in both single and mixed infections, is an important enteropathogen involved in neonatal diarrhea outbreaks in piglets and that the use of more sensitive diagnostic techniques allows the identification of mixed infections involving two or even three groups of PoRV, which may be more common than previously reported.(AU)

O rotavírus suíno grupo A (PoRVA) é uma das principais causas de diarreia neonatal em leitões lactentes e recém-desmamados em todo o mundo. As descrições do envolvimento de rotavírus não-grupo A em quadros de diarreia neonatal em leitões são esporádicas. No Brasil não há relatos do envolvimento do rotavírus suíno grupo C (PoRVC) na etiologia dos surtos de diarreia em leitões. O objetivo deste estudo foi descrever a identificação de rotavírus grupo C em infecções singulares e mistas com os rotavírus grupos A e B em três surtos de diarreia neonatal em leitões lactentes (<21 dias de idade), com taxas de morbidade de 70 por cento a 80 por cento e de letalidade de 20 por cento a 25 por cento, em três rebanhos suínos localizados no estado de Santa Catarina, Brasil. O diagnóstico de PoRV nas amostras de fezes diarreicas foi realizado por eletroforese em gel de poliacrilamida (PAGE) para identificar a presença dos grupos A, B (PoRVB), e C de rotavírus suíno e por RT-PCR (PoRVA e PoRVC) e semi-nested (SN)-PCR (PoRVB) com a amplificação parcial dos genes VP4 (VP8*)-VP7, NSP2 e VP6 de PoRVA, PoRVB e PoRVC, respectivamente. [...] As cepas de PoRVB (primeiro e segundo surtos) e as cepas de PoRVC (primeiro, segundo e terceiro surtos) mostraram maior identidade de nt com cepas de PoRVB e PoRVC que pertencem aos genotipos N4 e I1, respectivamente. Esta é a primeira descrição realizada no Brasil do envolvimento de PoRVC na etiologia de surtos de diarreia em leitões lactentes. Os resultados deste estudo demonstram que o PoRVC, tanto em infecções singulares quanto em infecções mistas, é um importante enteropatógeno envolvido em surtos de diarreia neonatal em leitões e que o uso de técnicas de diagnóstico mais sensíveis permite caracterizar que infecções mistas, com dois ou até mesmo com três grupos de PoRV, podem ser mais comuns do que anteriormente relatado.(AU)

Desenvolvimento de oligonucleotídeos iniciadores para sequenciamento dos genes codificadores das proteínas NSP1, NSP3 e VP6 de rotavírus suíno do grupo A / Development of primers for sequencing the NSP1, NSP3, and VP6 genes of the group A porcine rotavirus

Rotavírus é responsável pela ocorrência de diarreia em humanos e em várias espécies animais. Oito pares de oligonucleotídeos iniciadores (primers) foram desenvolvidos e utilizados como estratégia para o sequenciamento do tipo Sanger da região codificadora dos genes NSP1, NSP3 e VP6 com base nas áreas conservadas do genoma dos rotavírus do grupo A de suínos. Um total de 3 amostras previamente triadas como positivas para rotavírus do grupo A, tiveram os respectivos genes amplificados e seqüenciados por estes oligonucleotídeos iniciadores, possibilitando a caracterização das amostras circulantes, cujo conhecimento é essencial para a compreensão da epidemiologia da doença.(AU)

Rotavirus is the causative pathogen of diarrhea in humans and in several animal species. Eight pairs of primers were developed and used for Sanger sequencing of the coding region of the NSP1, NSP3, and VP6 genes based on the conserved regions of the genome of the group A porcine rotavirus. Three samples previously screened as positive for group A rotaviruses were subjected to gene amplification and sequencing to characterize the pathogen. The information generated from this study is crucial for the understanding of the epidemiology of the disease.(AU)

Desenvolvimento de oligonucleotídeos iniciadores para sequenciamento dos genes codificadores das proteínas NSP1, NSP3 e VP6 de rotavírus suíno do grupo A / Development of primers for sequencing the NSP1, NSP3, and VP6 genes of the group A porcine rotavirus

Rotavírus é responsável pela ocorrência de diarreia em humanos e em várias espécies animais. Oito pares de oligonucleotídeos iniciadores (primers) foram desenvolvidos e utilizados como estratégia para o sequenciamento do tipo Sanger da região codificadora dos genes NSP1, NSP3 e VP6 com base nas áreas conservadas do genoma dos rotavírus do grupo A de suínos. Um total de 3 amostras previamente triadas como positivas para rotavírus do grupo A, tiveram os respectivos genes amplificados e seqüenciados por estes oligonucleotídeos iniciadores, possibilitando a caracterização das amostras circulantes, cujo conhecimento é essencial para a compreensão da epidemiologia da doença.

Rotavirus is the causative pathogen of diarrhea in humans and in several animal species. Eight pairs of primers were developed and used for Sanger sequencing of the coding region of the NSP1, NSP3, and VP6 genes based on the conserved regions of the genome of the group A porcine rotavirus. Three samples previously screened as positive for group A rotaviruses were subjected to gene amplification and sequencing to characterize the pathogen. The information generated from this study is crucial for the understanding of the epidemiology of the disease.

Hospitalization due to norovirus and genotypes of rotavirus in pediatric patients, state of Espírito Santo

Viruses are the leading cause for hospitalization due to gastroenteritis worldwide. Group A rotaviruses (RV) are the most prevalent and are assorted in glycoproteins (G) and protease sensitive (P) dual genotypes based on polymorphic genes that encode the external VP7 and VP4 capsid proteins, respectively. Noroviruses (NoV) have increasingly answered by sporadic gastroenteritis. This study aimed to determine the prevalence of NoV and RV in 68 hospitalized children, between July 2004 and November 2006, at a pediatric hospital in Vitória city, state of Espírito Santo, Southeastern Brazil. Nucleic acid was extracted from fecal suspension following the guanidine-silica procedure. Reverse transcriptase-polymerase chain reaction (RT-PCR) and polyacrylamide gel electrophoresis were employed for NoV and RV detection, respectively. RV genotyping was accomplished using RT-PCR followed by heminested multiplex PCR with specific primers for the most prevalent types of G and P. Fecal samples were positive for NoV and RV in 39.7 percent (27/68) and 20.5 percent (14/68), respectively and together were responsible for 60 percent (41/68) of the cases. RV genotypes were: 50 percent G9P[8], 28.7 percent G2P[4], 7.1 percent G1P[8], G2P[8] and G?P[8]. Vomit was a prominent manifestation observed in 92 percent and 85 percent of the NoV and RV cases, respectively. The median hospitalization was 5 and 5.5 days for the patients infected with NoV and RV, respectively. The data showed that NoV prevailed over RV and it also corroborated the emergence of RV G9 genotype followed by G2P[4], reinforcing the need for RV genotype surveillance.

Rotavirus in south-americans indians communities / Rotavírus em comunidades indígenas sul-americanas

Rotaviruses are the most important etiologic agents of acute gastroenteritis in children under five years old. These viruses are responsible for several diarrhea outbreaks in both developed and developing countries. Acute diarrhea diseases are the foremost health problem in Indians communities. Serological survey among Brazilian Indians demonstrated seropositivity for rotavirus raged from 17% to 74%. Outbreaks investigations among these populations are extremely rare, especially in Brazilian southwestern region, standing out the importance in identifying the source of ongoing outbreaks, and for preventing the hereafter and/or most severe cases. The poor sanitation conditions and hygiene practices, as well as the possibility of native contact with wild virus reservoirs, account for enteropathogens spreading within Indian communities. Special precautions should currently be taken with respect to diarrhea monitoring among Amerindians, especially in the South American populations.

Rotavírus são os agentes etiológicos mais importantes das gastroenterites em crianças menores de cinco anos, responsáveis por vários surtos de diarréia em países desenvolvidos e em desenvolvimento. As doenças diarréicas agudas são os principais problemas de saúde dentro das comunidades indígenas. Vigilâncias sorológicas em índios brasileiros demonstraram 17% a 74% de soropositividade para rotavírus. A investigação de surtos entre essas populações são raras, principalmente na região sudeste do Brasil. A identificação da fonte original de tais surtos é importante para a prevenção de casos futuros, além de contribuir para o estabelecimento de um perfil epidemiológico. As precárias condições sanitárias e de práticas de higiene, assim como a possibilidade de contato dos índios com reservatórios selvagens, levam a dispersão de enteropatógenos dentro das comunidades indígenas. Atenções especiais devem ser adotadas em relação ao monitoramento de diarréia entre Ameríndios, especialmente os sul-americanos.