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1.
Mol Biol Rep ; 51(1): 777, 2024 Jun 21.
Artigo em Inglês | MEDLINE | ID: mdl-38904698

RESUMO

BACKGROUND: Senecavirus A (SV-A) is an RNA virus that belongs to the genus Senecavirus within the family Picornaviridae. This study aimed to analyze factors that can influence the molecular diagnosis of Senecavirus A, such as oligonucleotides, RNA extraction methods, and RT-qPCR kits. METHODS: Samples from suspected cases of vesicular disease in Brazilian pigs were analyzed for foot-and-mouth disease, swine vesicular disease, and vesicular stomatitis. All tested negative for these diseases but positive for SV-A. RT-qPCR tests were used, comparing different reagent kits and RNA extraction methods. Sensitivity and repeatability were evaluated, demonstrating efficacy in detecting SV-A in clinical samples. RESULTS: In RNA extraction, significant reduction in Cq values was observed with initial dilutions, particularly with larger supernatant volumes. Trizol and Maxwell showed greater sensitivity in automated equipment protocols, though results varied in tissue tests. RT-qPCR kit comparison revealed differences in amplification using viral RNA but minimal differences with plasmid DNA. Sensitivity among methods was comparable, with slight variations in non-amplified samples. Repeatability tests showed consistent results among RT-qPCRs, demonstrating similarity between methods despite minor discrepancies in Cq values. CONCLUSIONS: Trizol, silica columns, and semi-automated extraction were compared, as well as different RT-qPCR kits. The study found significant variations that could impact the final diagnosis.


Assuntos
Infecções por Picornaviridae , Picornaviridae , RNA Viral , Doenças dos Suínos , Animais , Picornaviridae/genética , Picornaviridae/isolamento & purificação , Suínos , Infecções por Picornaviridae/diagnóstico , Infecções por Picornaviridae/veterinária , Infecções por Picornaviridae/virologia , RNA Viral/genética , Doenças dos Suínos/virologia , Doenças dos Suínos/diagnóstico , Sensibilidade e Especificidade , Reação em Cadeia da Polimerase em Tempo Real/métodos , Doença Vesicular Suína/diagnóstico , Doença Vesicular Suína/virologia , Febre Aftosa/diagnóstico , Febre Aftosa/virologia , Brasil , Reprodutibilidade dos Testes
2.
Braz J Microbiol ; 55(2): 1961-1966, 2024 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-38589741

RESUMO

The FTA card has emerged as a promising alternative for nucleic acid extraction. The FTA card is a filter paper impregnated with chemicals that preserve and stabilize the genetic material present in the sample, allowing for its storage and transport at room temperature. The aim of this study was to test the card for the detection of RNA and DNA nucleic acids. Two RNA viruses (Senecavirus A and classical swine fever virus) and two DNA viruses (African swine fever virus and suid alphaherpesvirus 1) were tested, and in all cases, there was a decrease in sensitivity. The methods exhibited good repeatability and demonstrated a rapid and practical use for sample transport and nucleic acid extraction.


Assuntos
Vírus da Febre Suína Africana , Animais , Suínos , Vírus da Febre Suína Africana/isolamento & purificação , Vírus da Febre Suína Africana/genética , Vírus da Febre Suína Clássica/genética , Vírus da Febre Suína Clássica/isolamento & purificação , Herpesvirus Suídeo 1/isolamento & purificação , Herpesvirus Suídeo 1/genética , RNA Viral/genética , RNA Viral/isolamento & purificação , Medicina Veterinária/métodos , Doenças dos Suínos/virologia , Doenças dos Suínos/diagnóstico , Vírus de DNA/genética , Vírus de DNA/isolamento & purificação , Picornaviridae/genética , Picornaviridae/isolamento & purificação , Picornaviridae/classificação , Sensibilidade e Especificidade , DNA Viral/genética , Vírus de RNA/genética , Vírus de RNA/isolamento & purificação , Vírus de RNA/classificação , Infecções por Vírus de DNA/veterinária , Infecções por Vírus de DNA/diagnóstico , Infecções por Vírus de DNA/virologia , Manejo de Espécimes/métodos , Manejo de Espécimes/instrumentação
3.
Mol Biol Rep ; 50(4): 3439-3450, 2023 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-36757549

RESUMO

BACKGROUND: Megalocytiviruses (MCV) are double-stranded DNA viruses that infect fish. Two species within the genus are epidemiologically important for fish farming: red sea bream iridovirus (RSIV) and infectious spleen and kidney necrosis virus (ISKNV). The objective of this work was to study regions that allow the differentiation and correct diagnosis of RSIV and ISKNV. METHODS: The regions ORF450L, ORF342L, ORF077, and the intergenic region between ORF37 and ORF42R were sequenced and compared with samples from the database. RESULTS: The tree constructed using the sequencing of the PCR product Megalocytivirus. ORF077 separated the three major clades of MCV. RISV genotypes were well divided, but not ISKNV. All qPCRs tests showed acceptable repeatability values, that is, less than 5%. CONCLUSION: Two qPCRs for ISKNV detection and two for RSIV were considered suitable for use in the diagnosis and typing of MCV. The results of this study demonstrate the importance of an accurate evaluation of methodologies for the differentiation of MCV.


Assuntos
Infecções por Vírus de DNA , Doenças dos Peixes , Iridoviridae , Iridovirus , Animais , Iridoviridae/genética , Reação em Cadeia da Polimerase em Tempo Real , Infecções por Vírus de DNA/genética , Infecções por Vírus de DNA/veterinária , Filogenia
4.
Braz J Microbiol ; 53(3): 1691-1699, 2022 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-35553417

RESUMO

The vesicular stomatitis virus belongs to the Rhabdoviridae family, genus Vesiculovirus. Four species (New Jersey, Indiana, Cocal, and Alagoas) are responsible for disease outbreaks in Western Hemisphere countries. In Brazil, the Alagoas virus is responsible for the main outbreaks of the disease, mainly in the states of the Northeast, Midwest, and Southeast regions of the country. The present study aimed to perform the genetic characterization of 41 vesicular stomatitis virus samples. RNA was extracted using Trizol and used to amplify part of gene P. Amplicons were sequenced using the Sanger method. The phylogenetic trees generated showed that Alagoas vesiculoviruses were positioned into three groups: group A formed by the first virus isolate; group B by isolates from states in the Northeast region; and group C by isolates from the states of Bahia, Goiás, and Tocantins. Their divergence to date has generated the formation of two genotypes evolving independently in regions that until the present study had little geographic overlap.


Assuntos
Estomatite Vesicular , Animais , Brasil/epidemiologia , Filogenia , Vírus da Estomatite Vesicular Indiana/genética , Vesiculovirus/genética
5.
Braz J Microbiol ; 53(2): 1065-1075, 2022 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-35394611

RESUMO

The classical swine fever virus is the etiologic agent of one of the diseases with the greatest impact on swine farming worldwide. An extensive area of Brazil is considered free of the disease, but some states in Northeast Brazil have registered outbreaks since 2001. The objective of this study was to analyze the genetic variations of the virus and its spread over time and space. Partial sequences of the viral E2 protein obtained from samples collected during the Brazilian outbreaks were compared with sequences from the GenBank database (NCBI). The results demonstrated the continuous presence of the virus in the state of Ceará, with diffusion to at least two other states. The Brazilian Northeast virus presents specific polymorphisms that separate it from viruses isolated in other countries.


Assuntos
Vírus da Febre Suína Clássica , Peste Suína Clássica , Vírus , Animais , Brasil/epidemiologia , Peste Suína Clássica/epidemiologia , Peste Suína Clássica/genética , Vírus da Febre Suína Clássica/genética , Surtos de Doenças , Filogenia , Suínos , Proteínas Virais/genética
6.
Intervirology ; 59(1): 20-9, 2016.
Artigo em Inglês | MEDLINE | ID: mdl-27362774

RESUMO

OBJECTIVE: The aim of this study was to use partial Ul44 sequences (glycoprotein C) of Suid herpesvirus 1 to examine the evolution and dynamics of the virus in different periods and hosts. METHODS: Phylogenetic trees were constructed using the software MrBayes after analysis in the software jModelTest to evaluate the best phylogenetic models. The software SplitsTree 4.0 was used to create phylogenetic networks, and the BEAST program was used to generate data on phylogeography. Replication kinetics and serum neutralization tests were applied to tree strains from different phylogenetic groups. RESULTS: Ul44 sequences derived from domestic swine and wild swine clustered in different clades and had different selective pressures depending on the host. We found no differences in replication kinetics and serum neutralization tests in the strains tested. Data show that the evolution of herpesviruses is complex, and different genetic groups may be evolving at different rates. Ul44 is an important marker for molecular evolution and epidemiology studies, but it is not useful for biological information.


Assuntos
Evolução Molecular , Herpesvirus Suídeo 1/genética , Pseudorraiva/virologia , Doenças dos Suínos/virologia , Animais , Animais Selvagens/virologia , Herpesvirus Suídeo 1/crescimento & desenvolvimento , Herpesvirus Suídeo 1/fisiologia , Interações Hospedeiro-Patógeno , Testes de Neutralização , Filogenia , Filogeografia , Pseudorraiva/sangue , Pseudorraiva/epidemiologia , Alinhamento de Sequência , Software , Sus scrofa/virologia , Suínos/virologia , Replicação Viral
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