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/instrumentaçãoRESUMO
Avian metapneumoviruses (aMPV subtypes A-D) are respiratory and reproductive pathogens of poultry. Since aMPV-A was initially reported in Mexico in 2014, there have been no additional reports of its detection in the country. Using nontargeted next-generation sequencing (NGS) of FTA card-spotted respiratory samples from commercial chickens in Mexico, seven full genome sequences of aMPV-A (lengths of 13,288-13,381 nucleotides) were de novo assembled. Additionally, complete coding sequences of genes N (n = 2), P and M (n = 7 each), F and L (n = 1 each), M2 (n = 6), SH (n = 5) and G (n = 2) were reference-based assembled from another seven samples. The Mexican isolates phylogenetically group with, but in a distinct clade separate from, other aMPV-A strains. The genome and G-gene nt sequences of the Mexican aMPVs are closest to strain UK/8544/06 (97.22-97.47% and 95.07-95.83%, respectively). Various amino acid variations distinguish the Mexican isolates from each other, and other aMPV-A strains, most of which are in the G (n = 38), F (n = 12), and L (n = 19) proteins. Using our sequence data and publicly available aMPV-A data, we revised a previously published rRT-PCR test, which resulted in different cycling and amplification conditions for aMPV-A to make it more compatible with other commonly used rRT-PCR diagnostic cycling conditions. This is the first comprehensive sequence analysis of aMPVs in Mexico and demonstrates the value of nontargeted NGS to identify pathogens where targeted virus surveillance is likely not routinely performed.