RESUMO
Bacteria classified in Mycoplasma (M. bovis and M. bovigenitalium) and Ureaplasma (U. diversum) genera are associated with granular vulvovaginitis that affect heifers and cows at reproductive age. The traditional means for detection and speciation of mollicutes from clinical samples have been culture and serology. However, challenges experienced with these laboratory methods have hampered assessment of their impact in pathogenesis and epidemiology in cattle worldwide. The aim of this study was to develop a PCR strategy to detect and primarily discriminate between the main species of mollicutes associated with reproductive disorders of cattle in uncultured clinical samples. In order to amplify the 16S-23S rRNA internal transcribed spacer region of the genome, a consensual and species-specific nested-PCR assay was developed to identify and discriminate between main species of mollicutes. In addition, 31 vaginal swab samples from dairy and beef affected cows were investigated. This nested-PCR strategy was successfully employed in the diagnosis of single and mixed mollicute infections of diseased cows from cattle herds from Brazil. The developed system enabled the rapid and unambiguous identification of the main mollicute species known to be associated with this cattle reproductive disorder through differential amplification of partial fragments of the ITS region of mollicute genomes. The development of rapid and sensitive tools for mollicute detection and discrimination without the need for previous cultures or sequencing of PCR products is a high priority for accurate diagnosis in animal health. Therefore, the PCR strategy described herein may be helpful for diagnosis of this class of bacteria in genital swabs submitted to veterinary diagnostic laboratories, not demanding expertise in mycoplasma culture and identification.
Assuntos
Doenças dos Bovinos/diagnóstico , Infecções por Bactérias Gram-Negativas/veterinária , Tenericutes/isolamento & purificação , Vulvovaginite/veterinária , Animais , Bovinos , Doenças dos Bovinos/microbiologia , Feminino , Infecções por Bactérias Gram-Negativas/diagnóstico , Infecções por Bactérias Gram-Negativas/microbiologia , Reação em Cadeia da Polimerase , RNA Bacteriano/genética , RNA Ribossômico 16S/genética , RNA Ribossômico 23S/genética , Especificidade da Espécie , Tenericutes/genética , Vagina/microbiologia , Vulvovaginite/diagnóstico , Vulvovaginite/microbiologiaRESUMO
Norovirus (NoV) is a member of the Caliciviridae family and is considered an emerging human enteric pathogen. NoVs are detected in farm animals such as cattle, sheep and pigs. Porcine NoV (PoNoV) is widespread worldwide, but frequency of infection is often low. This study aimed to investigate the natural PoNoV infection from adult animals of an important Brazilian pig-production region. Faecal samples (n = 112) of asymptomatic pigs aged 9 to 24 weeks old were collected from 16 grower-to-finish herds located in Paraná state, Brazilian Southern region, and evaluated for PoNoV presence. A reverse transcription-polymerase chain reaction (RT-PCR) assay was performed using specific primers that target a conserved region of the virus capsid gene (VP1). PoNoV was detected in 58 (51.8%) of the 112 faecal samples and in 14 (87.5%) of the 16 herds evaluated. Six of the obtained amplicons were submitted to phylogenetic genotyping analysis. The higher nucleotide (86.5-97.4%) and amino acid (100%) similarities of the sequences in this study were with the representative strains of the porcine NoV genogroup II genotype 11 (PoNoV GII-11). These results reveal that PoNoV infection is endemic in one of the most important pork production areas of Brazil and that the PoNoV GII-11 is prevalent in this region.
Assuntos
Infecções por Caliciviridae/veterinária , Norovirus/classificação , Sus scrofa/virologia , Doenças dos Suínos/epidemiologia , Animais , Brasil/epidemiologia , Infecções por Caliciviridae/epidemiologia , Infecções por Caliciviridae/virologia , Proteínas do Capsídeo/genética , Primers do DNA , Genótipo , Carne , Epidemiologia Molecular , Filogenia , RNA Viral/genética , Estações do Ano , Suínos , Doenças dos Suínos/virologiaRESUMO
Porcine teschovirus (PTV), Porcine sapelovirus (PSV) and Enterovirus G (EV-G) have been associated with enteric, respiratory, reproductive and neurological disorders. Although Brazil is the world's fourth largest producer and exporter of pork, no information on the occurrence of PTV, PSV and EV-G infections is available for Brazilian pig herds. This study aimed to investigate the occurrence of Porcine enteric picornavirus infections in pig farms located in three distinct geographical regions of Brazil. Forty randomly selected diarrhoeic and normal consistency faeces of suckling (n = 22) and nursery (n = 18) pigs from farms located in 21 distinct cities of the Southern, Southeast, and Midwest regions of Brazil were evaluated by nested-RT-PCR assays. Suckling piglets presented the expected amplicon size for PTV (158 bp) and EV-G (313 bp) in single and mixed infections in 40.9 % (9/22) of the faecal samples. PSV amplicon (212 bp) was not detected in this age group. For nursery pigs, Porcine enteric picornaviruses amplicons were present in 77.8 % (14/18) of the faecal samples. PTV and EV-G were detected in single and mixed infections, while PSV was detected only in two samples in co-infection with PTV and EV-G in this age group. The Brazilian regions evaluated presented at least two of the tested viruses. Sequencing analysis revealed high similarities to the related viruses (95.3 to 99.2 % for PTV, 94.2 to 98.5 % for PSV and 86 to 100 % for EV-G). For the first time PTV, PSV and EV-G have been molecularly detected and characterised in pig faecal samples in Brazil.
Assuntos
Infecções por Picornaviridae/veterinária , Picornaviridae/classificação , Doenças dos Suínos/virologia , Animais , Brasil/epidemiologia , Coinfecção , Fezes/virologia , Picornaviridae/genética , Infecções por Picornaviridae/epidemiologia , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Suínos , Doenças dos Suínos/epidemiologiaRESUMO
The aim of this study was to evaluate the frequency of Aichivirus C infection in the three major industrial pig-producing regions of Brazil. This retrospective study evaluated 63 faecal samples that were collected between 2004 and 2011 from suckling piglets (1 to 3 weeks old) belonging to 46 pig herds located in the South, Southeast and Central-west regions of Brazil. The presence of Aichivirus C in the samples was evaluated by reverse transcription-polymerase chain reaction assay. A 216-bp fragment of the aichivirus RNA-dependent RNA polymerase gene was amplified from 48 out of 63 (76.2 %) faecal samples. Aichivirus C was identified in all the evaluated regions, and variations in the frequency of virus detection among the different Brazilian regions were observed. Aichivirus C was detected at significantly higher rates in 2-week-old (18/20; 90.0 %) and 3-week-old (19/22; 86.4 %) piglets than in 1-week-old piglets (11/21; 52.4 %) (P < 0.05). The nucleotide sequences of three amplicons clustered together with other sequences previously isolated in Brazil, revealing that there were no phylogenetic differences in the strains of Aichivirus C by region or age. This study demonstrated that Aichivirus C has been present in Brazilian pig herds for years and has spread to the major pig-producing regions of the country.
Assuntos
Kobuvirus/isolamento & purificação , Infecções por Picornaviridae/veterinária , Doenças dos Suínos/epidemiologia , Doenças dos Suínos/virologia , Fatores Etários , Animais , Animais Lactentes , Brasil/epidemiologia , Distribuição de Qui-Quadrado , Fezes/virologia , Kobuvirus/genética , Filogenia , Infecções por Picornaviridae/epidemiologia , Infecções por Picornaviridae/virologia , Prevalência , RNA Viral/química , RNA Viral/genética , DNA Polimerase Dirigida por RNA/química , DNA Polimerase Dirigida por RNA/genética , Estudos Retrospectivos , Reação em Cadeia da Polimerase Via Transcriptase Reversa/veterinária , SuínosRESUMO
The aims of this study were to evaluate torque teno sus virus (TTSuV) infection in healthy slaughter-age pigs and to compare the similarities of the untranslated region (UTR) nucleotide (nt) sequences obtained from different biological samples of the same animals. Fifty-eight pigs were evaluated by PCR assay for the presence of TTSuV in paired samples of liver and serum (Group 1, n = 27) and lung and serum (Group 2, n = 31). All the pigs were positive for TTSuV infection in the organs sampled and 94.8 % (n = 55) presented with viraemia. The nt sequence similarities between the Groups 1 and 2 varied from 91.7 % to 96.6 % (TTSuV1) and 91 % to 95 % (TTSuV2). In Group 1, the nt sequence similarities were 93 % (TTSuV1) and 95.4 % (TTSuV2). In Group 2, the nt sequence similarities were 95 % (TTSuV1) and 91 % (TTSuV2). These results revealed the simultaneous infection with distinct strains of TTSuV1 and 2 in healthy pigs at slaughter age.
Assuntos
Fígado/virologia , Pulmão/virologia , Suínos/virologia , Torque teno virus/isolamento & purificação , Animais , Sequência de Bases , DNA Viral/química , DNA Viral/genética , Dados de Sequência Molecular , Reação em Cadeia da Polimerase/veterinária , Análise de Sequência de DNA , Suínos/sangue , Torque teno virus/genética , Regiões não TraduzidasRESUMO
O coronavírus bovino (BCoV) é o segundo mais importante agente etiológico viral envolvido em diarreias neonatais em bezerros de todo o mundo. Os relatos sobre a frequência da infecção por BcoV em rebanhos bovinos de corte criados extensivamente são incomuns no Brasil. Este estudo analisou 93 amostras fecais diarreicas de bezerros de corte com até 60 dias de idade, provenientes de 13 rebanhos comerciais distribuídos nos estados de Mato Grosso, Mato Grosso do Sul, Minas Gerais, Paraná e Rondônia. As amostras fecais foram colhidas no período de 2009-2012 e haviam sido previamente analisadas quanto à presença de rotavírus bovino grupo A (BoRVA), com resultados negativos. A presença do BCoV nas amostras fecais foi avaliada por meio da amplificação parcial do gene N pela técnica da semi-nested PCR. Em 33,3% (31/93) das amostras analisadas foi possível a amplificação de produtos com tamanho de 251 pb esperados para o BCoV. Os resultados deste estudo demonstraram que a coronavirose tem importante participação no complexo diarreia neonatal em rebanhos bovinos de corte criados extensivamente em várias regiões brasileiras.
Bovine coronavirus (BCoV) is the second most important viral agent involved in neonatal diarrhea in calves worldwide. The reports on the frequency of BCoV infection in beef cattle herds under extensive management are uncommon in Brazil. The present study analyzed 93 diarrheic fecal samples of calves up to 60 days of age from 13 commercial beef cattle herds located in the states of Mato Grosso, Mato Grosso do Sul, Minas Gerais, Paraná, and Rondônia. The fecal samples were collected during 2009-2012 and were previously analyzed for the presence of bovine rotavirus group A (BoRVA), with negative results. The presence of BCoV in the fecal samples was evaluated by the partial amplification of the N gene by using the semi-nested PCR technique. The expected products of 251 bp length were amplified 33.3% (31/93) of the analyzed diarrheic fecal samples. The results revealed that coronaviruses has important participation in the neonatal diarrhea complex of beef cattle herds reared extensively from the different geographical regions of Brazil.
RESUMO
O coronavírus bovino (BCoV) é o segundo mais importante agente etiológico viral envolvido em diarreias neonatais em bezerros de todo o mundo. Os relatos sobre a frequência da infecção por BcoV em rebanhos bovinos de corte criados extensivamente são incomuns no Brasil. Este estudo analisou 93 amostras fecais diarreicas de bezerros de corte com até 60 dias de idade, provenientes de 13 rebanhos comerciais distribuídos nos estados de Mato Grosso, Mato Grosso do Sul, Minas Gerais, Paraná e Rondônia. As amostras fecais foram colhidas no período de 2009-2012 e haviam sido previamente analisadas quanto à presença de rotavírus bovino grupo A (BoRVA), com resultados negativos. A presença do BCoV nas amostras fecais foi avaliada por meio da amplificação parcial do gene N pela técnica da semi-nested PCR. Em 33,3% (31/93) das amostras analisadas foi possível a amplificação de produtos com tamanho de 251 pb esperados para o BCoV. Os resultados deste estudo demonstraram que a coronavirose tem importante participação no complexo diarreia neonatal em rebanhos bovinos de corte criados extensivamente em várias regiões brasileiras.
Bovine coronavirus (BCoV) is the second most important viral agent involved in neonatal diarrhea in calves worldwide. The reports on the frequency of BCoV infection in beef cattle herds under extensive management are uncommon in Brazil. The present study analyzed 93 diarrheic fecal samples of calves up to 60 days of age from 13 commercial beef cattle herds located in the states of Mato Grosso, Mato Grosso do Sul, Minas Gerais, Paraná, and Rondônia. The fecal samples were collected during 2009-2012 and were previously analyzed for the presence of bovine rotavirus group A (BoRVA), with negative results. The presence of BCoV in the fecal samples was evaluated by the partial amplification of the N gene by using the semi-nested PCR technique. The expected products of 251 bp length were amplified 33.3% (31/93) of the analyzed diarrheic fecal samples. The results revealed that coronaviruses has important participation in the neonatal diarrhea complex of beef cattle herds reared extensively from the different geographical regions of Brazil.
RESUMO
Torque teno sus virus (TTSuV) is responsible for the infection of pig herds around the world. The aim of this study was to analyse the presence of natural infection by both species of TTSuV in suckling piglets from major pig-producing regions of Brazil. Faecal samples (n = 135) from 1 to 3-week-old suckling piglets from the Southern, Southeast and Midwest regions of Brazil were analysed by PCR assay to detect TTSuV1 and 2. TTSuV1 and 2 DNA was identified in 65 (48.1 %) and 23 (17 %) of piglet faecal samples, respectively. Co-infection by both species of TTSuV was detected in 17 (12.6 %) samples. Detection of TTSuV1 was significantly higher than that of TTSuV2 in the three Brazilian regions together (p < 0.05). Based on age of animals, TTSuV1 infection was statistically higher than TTSuV2 in each age group (p < 0.05). For all of the age groups together, no statistical difference was detected in the number of TTSuV1 and 2 positive results (p > 0.05). These findings revealed that TTSuV infection has disseminated in pig herds from different geographic Brazilian regions, and the presence of TTSuV in suckling piglet faecal samples suggested the early infection by the virus and the potential of these animals in spreading the virus.