Molecular characterization of Corynebacterium pseudotuberculosis isolates using ERIC-PCR Alessandro de Sa ´ Guimara ˜es a,b,d,1 , Elaine Maria Seles Dorneles a,1 , Giovanna Ivo Andrade a , Andrey Pereira Lage a,c , Anderson Miyoshi b , Vasco Azevedo b,c , Aurora Maria Guimara ˜es Gouveia a,c,1 , Marcos Bryan Heinemann a,c,1, * a Universidade Federal de Minas Gerais, UFMG, Escola de Veterina ´ria, Departamento de Medicina Veterina ´ria Preventiva, Laborato ´rio de Sanidade de Ovinos e Caprinos, Av. Antoˆnio Carlos 6627, Caixa Postal 567, Campus da UFMG, CEP 30123-970, Belo Horizonte, Minas Gerais, Brazil b Universidade Federal de Minas Gerais, UFMG, Instituto de Cieˆncias Biolo ´gicas, Departamento de Biologia Geral, Laborato ´rio de Gene ´tica Celular e Molecular, Av. Antoˆnio Carlos 6627, Campus da UFMG, CEP 30123-970, Belo Horizonte, Minas Gerais, Brazil c Grupo de Extensa˜o da Pesquisa em Ovinos e Caprinos – GEPOC, Brazil d Empresa Brasileira de Pesquisa Agropecua ´ria, EMBRAPA, R. Eugeˆnio do Nascimento 610, Dom Bosco, CEP 36038-330, Juiz de Fora, Minas Gerais, Brazil 1. Introduction Corynebacterium pseudotuberculosis is a relevant animal pathogen belonging to the family of Corynebacteriaceae classified into two biotypes based on hosts preferences and nitrate-reducing activity (Biberstein et al., 1971). In sheep, it is responsible for caseous lymphadenitis characterized by abscesses in the superficial and visceral lymph nodes and is widely disseminated among the sheep population in Veterinary Microbiology 153 (2011) 299–306 A R T I C L E I N F O Article history: Received 30 November 2010 Received in revised form 23 May 2011 Accepted 7 June 2011 Keywords: Corynebacterium pseudotuberculosis Caseous lymphadenitis RAPD ERIC-PCR A B S T R A C T Caseous lymphadenitis is an infectious sheep and goats disease caused by Corynebacterium pseudotuberculosis and characterized by abscesses in superficial and visceral lymph nodes. C. pseudotuberculosis strains isolated from these hosts have been shown to be very difficult to type by the existing methods. The aim of this study is evaluating the potential of the Enterobacterial Repetitive Intergenic Consensus (ERIC-PCR) as a tool for molecular typing of C. pseudotuberculosis strains isolated in sheep. One hundred and twenty seven isolates of C. pseudotuberculosis were isolated from lesions suspected to have had caseous lymphadenitis collected from sheep at the slaughterhouse. Animals were from 24 flocks in 13 municipalities of the Minas Gerais State, Brazil. Species identification of the isolates was performed by routine biochemical tests and mPCR. Fingerprint was performed by RAPD using ERIC-1R, ERIC-2 and ERIC-1R + ERIC-2 primers. Seventeen different genotypes were generated by ERIC 1-PCR, 21 genotypes by ERIC 2-PCR and 21 genotypes by ERIC 1 + 2-PCR. Hunter-Gaston Discrimination Index (HGDI) found for ERIC 1, ERIC 2, ERIC 1 + 2 PCR were 0.69, 0.87, and 0.84, respectively. For most herds evaluated observed at most three different genotypes among isolates from animals of these property, in all ERIC-PCR assays. However a few flocks observed between four and nine genotypes per flock. The W Kendall value found for correlation among the three techniques of ERIC-PCR was 0.91 (P < 5.01 Â 10 À6 ). The results show that ERIC-PCR has good discriminatory power and advantages over other DNA-based typing methods, making it a useful tool to discriminate C. pseudotuberculosis isolates. ß 2011 Elsevier B.V. All rights reserved. * Corresponding author. Tel.: +55 31 3409 2077; fax: +55 31 3409 2080. E-mail addresses: alessandrodesa@uol.com.br (A.d.S. Guimara ˜es), selesdorneles@yahoo.com.br (E.M.S. Dorneles), giovannaivo@yahoo.com.br (G.I. Andrade), alage@vet.ufmg.br (A.P. Lage), miyoshi@icb.ufmg.br (A. Miyoshi), vasco@icb.ufmg.br (V. Azevedo), auroragouveia@terra.com.br (A.M.G. Gouveia), mabryan@ufmg.br (M.B. Heinemann). 1 These authors contributed equally to this work. Contents lists available at ScienceDirect Veterinary Microbiology jo u rn al ho m epag e: ww w.els evier.c o m/lo cat e/vetmic 0378-1135/$ – see front matter ß 2011 Elsevier B.V. All rights reserved. doi:10.1016/j.vetmic.2011.06.002