Genetic heterogeneity of Mycoplasma hyopneumoniae in pigs has been reported, however there has been limited reproducibility on the molecular methods employed so far. The aim of this study was to modify and standardize a high-resolution multiple locus variable number tandem repeat analysis (MLVA), to investigate the genetic variability of M. hyopneumoniae circulating in the United States of America (USA), Brazil, Mexico and Spain. The MLVA was standardized on the basis of the number of tandem repeats in two Mycoplasma adhesins, P97 and P146, which are proteins involved in the adherence of the pathogen to cilia. A total of 355 samples obtained from the four countries were analyzed. The Simpson's diversity index for the assay was D=. 0.976 when samples from all countries were combined. A large number of MLVA types (. n=. 139) were identified, suggesting that multiple M. hyopneumoniae variants are circulating in swine. The locus P97 had 17 different types with 2-18 repeats. The P146 locus showed higher heterogeneity, with 34 different types, ranging from 7 to 48 repeats. MLVA types that presented more than 30 repeats in P146 were found in Spain and Brazil, while shorter repeats were observed in the USA and Mexico. This simplified MLVA method proved to be an efficient tool for typing M. hyopneumoniae with a high degree of stability, repeatability, and discriminatory power. In conclusion, M. hyopneumoniae showed a high variable number tandem repeat heterogeneity and this assay can be applied in molecular epidemiology investigations within farms and productions systems.
Bibliographical noteFunding Information:
The authors thank Dr. Anna Perez de Rozas from CReSA for providing Spanish isolates, Jason Daniels for his help with sample processing, the UMN-VDL for providing USA and Mexican samples, and Microvet for its support in providing samples from Brazil. This project was funded by the Minnesota Pork Board and the Pork Checkoff , and the Swine Disease Eradication Center . L.F. Dos Santos was supported by Capes Foundation, Ministry of Education of Brazil (proc. No: BEX17617/12-0) and Fundação de Amparo à Pesquisa do estado de Minas Gerais FAPEMIG . M.A.S. Moreira was supported by CNPQ.
© 2014 Elsevier B.V.
- Enzootic pneumonia
- Molecular typing
- Multiple locus variable tandem analysis (MLVA)