RESUMO
Molecular analysis was conducted on 36 Mycoplasma gallisepticum DNA extracts from tracheal swab samples of commercial poultry in seven South African provinces between 2009 and 2012. Twelve unique M. gallisepticum genotypes were identified by polymerase chain reaction and sequence analysis of the 16S-23S rRNA intergenic spacer region (IGSR), M. gallisepticum cytadhesin 2 (mgc2), MGA_0319 and gapA genetic regions. The DNA sequences of these genotypes were distinct from those of M. gallisepticum isolates in a database composed of sequences from other countries, vaccine and reference strains. The most prevalent genotype (SA-WT#7) was detected in samples from commercial broilers, broiler breeders and layers in five provinces. South African M. gallisepticum sequences were more similar to those of the live vaccines commercially available in South Africa, but were distinct from that of F strain vaccine, which is not registered for use in South Africa. The IGSR, mgc2 or MGA_0319 sequences of three South African genotypes were identical to those of the ts-11 vaccine strain, necessitating a combination of mgc2 and IGSR targeted sequencing to differentiate South African wild-type genotypes from ts-11 vaccine. To identify and differentiate all 12 wild-types, mgc2, IGSR and MGA_0319 sequencing was required. Sequencing of gapA was least effective at strain differentiation. This research serves as a model for the development of an M. gallisepticum sequence database, and illustrates its application to characterize M. gallisepticum genotypes, select diagnostic tests and better understand the epidemiology of M. gallisepticum.
Assuntos
Galinhas/microbiologia , Bases de Dados de Ácidos Nucleicos , Infecções por Mycoplasma/veterinária , Mycoplasma gallisepticum/genética , Doenças das Aves Domésticas/microbiologia , Animais , Proteínas de Bactérias/genética , Sequência de Bases , Análise por Conglomerados , Primers do DNA/genética , DNA Bacteriano/química , DNA Bacteriano/genética , DNA Espaçador Ribossômico/química , DNA Espaçador Ribossômico/genética , Genótipo , Dados de Sequência Molecular , Infecções por Mycoplasma/microbiologia , Mycoplasma gallisepticum/classificação , Mycoplasma gallisepticum/isolamento & purificação , Reação em Cadeia da Polimerase/veterinária , Análise de Sequência de DNA/veterinária , África do Sul , Traqueia/microbiologiaRESUMO
Recent reports have shown an increased recovery of cells from flocked nylon swabs which may improve the specimen quality and the real sensitivity of diagnostic tests in a clinical setting. In this study, the detection of Mycoplasma gallisepticum (MG) and M. synoviae (MS), using dry swabs of different materials (nylon flocked, cotton, and polyester), was investigated using real-time TaqMan PCR protocols. Different types of samples, including dilutions of pure broth cultures of MG and MS as well as swabs from tracheas of experimentally infected chickens and field cases of infection, were analyzed. There were no statistical differences in real-time PCR results among the different swab types (P < 0.05), indicating that this is not likely to be a significant factor in MG and MS detection by this method.
Assuntos
Galinhas , Infecções por Mycoplasma/veterinária , Mycoplasma gallisepticum/isolamento & purificação , Mycoplasma synoviae/isolamento & purificação , Doenças das Aves Domésticas/diagnóstico , Manejo de Espécimes/métodos , Animais , Fibra de Algodão , DNA Bacteriano/análise , Infecções por Mycoplasma/diagnóstico , Infecções por Mycoplasma/microbiologia , Nylons , Poliésteres , Doenças das Aves Domésticas/microbiologia , Reação em Cadeia da Polimerase em Tempo Real/veterinária , Sensibilidade e Especificidade , Manejo de Espécimes/veterinária , Traqueia/microbiologiaRESUMO
Two groups of Mycoplasma gallisepticum (MG) isolates (n = 24) from Jordan were analyzed by molecular methods and compared with other Middle Eastern isolates, related international isolates, and reference strains. The first group (n = 19) was isolated from July 2004 to January 2005 (isolation period A), and the newer group (n = 5) from June 2007 to April 2008 (isolation period B). The groups of isolates are from chicken flocks from northern Jordan, but are not from the same farms. None of the flocks were vaccinated for MG. Random amplified polymorphic DNA analysis, targeted sequencing of the partial MG cytadhesin 2 (mgc2), and the MG 16S-23S rRNA intergenic spacer region (IGSR) divided the Jordanian isolates into two groups. All of the 19 isolates from time period A, in addition to two isolates from time period B, were indistinguishable from the F strain. Three of five isolates from time period B were characterized as wild types and were indistinguishable from each other. The wild-type field strain was readily distinguished from the F strain. It was 91% and 96.4% similar to the F strain based on Clustal-W alignments of sequences of mgc2 and IGSR, respectively. Sequence similarity of mgc2 gene of the Jordan wild-type strain to isolates from Israel and Egypt ranged from 96.5% to 100%, whereas for IGSR it was 99.4%-100%. We theorize that the F-strain live MG vaccine, commonly used in Jordan prior to 2007, was transmitted to nonvaccinated poultry in the region and was a predominant genotype during time period A.
