Synthetic gene as target to assess the sensitivity of PCR to detect Trichinella spp. larvae in meat from a non-endemic region
| Autor: | Maria Isabel Botelho Vieira, Luiz Carlos Kreutz, Ana Paula Andreolla, Lucas de Figueiredo Soveral, Jalusa Deon Kich, Thiago de Souza Rosés, Rafael Frandoloso |
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| Rok vydání: | 2019 |
| Předmět: |
Meat
040301 veterinary sciences Swine Trichinella Argentina Biology Genome Polymerase Chain Reaction Sensitivity and Specificity law.invention 0403 veterinary science Plasmid Food Animals Synthetic gene law Zoonoses Genotype Genes Synthetic Animals Non endemic Polymerase chain reaction DNA Primers Swine Diseases Larva 0402 animal and dairy science Trichinellosis 04 agricultural and veterinary sciences biology.organism_classification 040201 dairy & animal science Virology Animal Science and Zoology Brazil |
| Zdroj: | Tropical animal health and production. 52(2) |
| ISSN: | 1573-7438 |
| Popis: | Trichinellosis is a zoonotic disease exotic in Brazil but commonly found worldwide including South American countries like Argentina. International trading of swine meat needs an official Trichinella-free diagnosis commonly carried out by pepsin-HCl digestion of diaphragm tissue fragments followed by microscopic examination for the presence or absence of Trichinella larvae. The easiness of this diagnostic method allows it to be performed at slaughtering plants but, in contrast, it lacks sensitivity and does not allow species differentiation, which is fundamental for determining geographical and species distribution of different genotypes. In our study, we aimed to evaluate a highly sensitive diagnostic method based on the polymerase chain reaction (PCR) that would allow us to detect and classify different species of Trichinella. Thus, we designed a synthetic gene and selected five sets of primers targeting specific regions of the Trichinella genome. The synthetic gene was cloned into a plasmid and then used to optimize PCR conditions. Using our PCR, we were able to detect 0.001 pg of the synthetic gene, which corresponded to 0.01 larvae. Then, we collected 175 samples of Suidae (domestic and wild boars) diaphragm fragments that were pooled into groups, digested with pepsin-HCl, and had the DNA extracted for analysis by PCR. The clinical samples evaluated were negative by PCR. Our results indicate that the PCR-based method might be a useful diagnostic method complementary to the pepsin-HCl digestion method currently in use, mostly in non-endemic areas. |
| Databáze: | OpenAIRE |
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