Human GAS Virulence Genes in Bovine GCS | CDC EID

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Volume 16, Number 1–January 2010
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Human Group A Streptococci Virulence Genes in Bovine Group C Streptococci
Márcia G. Rato, Ricardo Bexiga, Sandro F. Nunes, Cristina L. Vilela, and Ilda Santos-Sanches
Author affiliations: Universidade Nova de Lisboa, Caparica, Portugal (M.G. Rato, I. Santos-Sanches); Universidade Técnica de Lisboa, Lisbon, Portugal (R. Bexiga, S.F. Nunes, C.L. Vilela); and Cambridge University, Cambridge, UK (S.F. Nunes)

Suggested citation for this article

Abstract
Phage-encoded virulence genes of group A streptococci were detected in 10 (55.6%) of 18 isolates of group C streptococci that had caused bovine mastitis. Bovine isolates carried other genetic determinants, such as composite transposon Tn1207.3/F10394.4 (100%) and antimicrobial drug resistance genes erm(B)/erm(A) (22.2%), linB (16.6%), and tet(M)/tet(O) (66.7%), located on mobile elements.

Strains of Streptococcus dysgalactiae subsp. dysgalactiae are described as α-hemolytic or nonhemolytic (Lancefield group C) and associated only with animal infections (bovine mastitis), a disease with major economic consequences for the dairy industry (1). Group A streptococci (GAS)–specific phage-associated virulence determinants encoding pyrogenic exotoxins or superantigens (speM, ssa), which are strongly associated with severe diseases such as scarlet fever, streptococcal toxic shock syndrome, and rheumatic fever, have been described among human group C streptococci (GCS) or group G streptococci (GGS) (S. dysgalactiae subsp. equisimilis) (2) but not among α-hemolytic GCS (S. dysgalactiae subsp. dysgalactiae) of bovine origin. In contrast, M protein or M-like proteins were found in human GGS/GCS (S. dysgalactiae subsp. equisimilis) and in animal GCS (S. dysgalactiae subsp. dysgalactiae) but only in β-hemolytic strains (3).

Composite transposons and other genetic determinants also considered to be located in specific mobile elements such as macrolide (either encoding methylases [erm genes] or efflux pumps [mef genes]) and tetracycline resistance determinants (tet genes) have been found among streptococcal species of human origin. We studied a collection of field isolates of bovine GCS S. dysgalactiae subsp. dysgalactiae to search for genetic determinants, particularly those carried by mobile elements known to be transferred among human GAS and GGS/GCS.

The Study
We studied 18 α-hemolytic S. dysgalactiae subsp. dysgalactiae field isolates of Lancefield group C that had caused bovine subclinical mastitis. Isolates were obtained from 304 milk samples of 248 cows from 8 farms in Portugal that were included in the study. Detailed information regarding isolation methods and identification of field isolates by biochemical methods was described in a study of the subclinical mastitis–associated pathogen S. uberis (4). To confirm identification of S. dysgalactiae subsp. dysgalactiae, the 16S rRNA gene was amplified by PCR and sequenced (5). SmaI/cfr9I-digested DNA banding patterns were obtained by pulsed-field gel electrophoresis for clone identification as described (4).

All genes analyzed by PCR are shown in the Appendix Table. The emm gene subtyping was performed as described (www.cdc.gov/ncidod/biotech/strep/M-ProteinGene_typing.htm). Primers used and conditions for PCR were essentially as described elsewhere (Appendix Table).

Samples without DNA and strains lacking (negative) or carrying (positive) specific genes were used as controls in the PCR. Results were consistent in 2 or 3 PCRs that included these controls. Sequencing of all virulence gene amplicons was performed with the same primers used for amplification (STAB-Vida, Lisbon, Portugal). All sequences were compared with sequences in GenBank by using the BLAST alignment tool (www.ncbi.nlm.nih.gov/BLAST).

Antimicrobial drug resistance against macrolides (erythromycin), lincosamides (pirlimycin), and tetracycline was determined as described (10). Macrolide resistance phenotypes identified were M (resistance to macrolides) and MLSB (resistance to macrolides, lincosamides and streptogramins B).

We detected bacteriophage-associated virulence genes speM, speK, speC, spd1, and speL. Overall, speM was found in 10 (55.6%) of 18 bovine GCS isolates, speK in 9 (50%), speC and spd1 in 6 (33%), and speL in 4 (22.2%). All but 1 of the PCR products showed expected sizes (Appendix Table). Tn1207.3/Φ10394.4 composite transposon left junction amplicon showed a size of 380 bp instead of 453–6,807 bp as described for GAS (9). No amplification was observed for the right junction of this genetic element.

Suggested Citation for this Article
Rato MG, Bexiga R, Nunes SF, Vilela CL, Santos-Sanches I. Human group A streptococci virulence genes in bovine group C streptococci. Emerg Infect Dis [serial on the Internet]. 2010 Jan [date cited]. Available from http://www.cdc.gov/EID/content/16/1/116.htm
DOI: 10.3201/eid1601.090632

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