Tylosin Resistance in Arcanobacterium pyogenes Is Encoded by an Erm X Determinant

Abstract

Arcanobacterium pyogenes, a commensal on the mucous membranes of many economically important animal species, is also a pathogen, causing abscesses of the skin, joints, and visceral organs as well as mastitis and abortion. In food animals, A. pyogenes is exposed to antimicrobial agents used for growth promotion, prophylaxis, and therapy, notably tylosin, a macrolide antibiotic used extensively for the prevention of liver abscessation in feedlot cattle in the United States. Of 48 A. pyogenes isolates, 11 (22.9%) exhibited inducible or constitutive resistance to tylosin (MIC of >128 g/ml). These isolates also exhibited resistance to other macrolide and lincosamide antibiotics, suggesting a macrolide-lincosamide resistance phenotype. Of the 11 resistant isolates, genomic DNA from nine hybridized to an erm(X)-specific probe. Cloning and nucleotide sequencing of the A. pyogenes erm(X) gene indicated that it was >95% similar to erm(X) genes from Corynebacterium and Propionibacterium spp. Eight of the erm(X)-containing A. pyogenes isolates exhibited inducible tylosin resistance, which was consistent with the presence of a putative leader peptide upstream of the erm(X) open reading frame. For at least one A. pyogenes isolate, 98-4277-2, erm(X) was present on a plasmid, pAP2, and was associated with the insertion sequence IS6100. pAP2 also carried genes encoding the repressor-regulated tetracycline efflux system determinant Tet 33. The repA gene from pAP2 was nonfunctional in Escherichia coli and at least one A. pyogenes isolate, suggesting that there may be host-encoded factors required for replication of this plasmid.The use of antimicrobial agents as feed additives for disease prophylaxis and growth promotion in the beef cattle industry is a common practice in the United States. One of the major targets is liver abscessation, which is second only to respiratory disease in terms of economic losses to cattle feedlots. Control of liver abscesses has primarily depended on the use of feed additives, such as the macrolide tylosin, chlortetracycline, and oxytetracycline (10, 26), with tylosin the most effective and commonly used feed additive (11). A study involving almost 7,000 feedlot cattle demonstrated that the use of tylosin as a feed additive reduced the incidence of liver abscessation by 73% and increased weight gain and feed conversion by 2.3 and 2.6%, respectively (G. J. Vogel and S. B. Laudert, abstract, J. Anim. Sci. 72[Supp. 1]:293, 1994). Correspondingly, tylosin use is extensive, with 42.3% of feedlot cattle receiving tylosin as a feed additive, primarily for the prevention of liver abscesses (26).Fusobacterium necrophorum is the primary etiological agent of bovine hepatic abscessation (18), but Arcanobacterium pyogenes is a synergistic pathogen, being found in 10 (9) to 90% (12) of abscesses. Interestingly, in cattle fed tylosin, the incidence of hepatic abscesses containing A. pyogenes increased from 10 to 53% (9).The resistance of A. pyogenes to tylosin has only recently been documented. In a survey of A. pyo...

Facile Recovery of Individual High-Molecular-Weight, Low-Copy-Number Natural Plasmids for Genomic Sequencing

“…One large region resembles part of the 28-kb Corynebacterium glutamicum plasmid pTET3 (NC_003227) (31), including a tetracycline resistance determinant and a class 1 integron with a truncated integrase and an aminoglycoside adenyltransferase cassette, aadA9 (Table 3). An adjacent, smaller region resembles the 9-kb Arcanobacterium pyogenes plasmid pAP2 (NC_005206) (17), including a macrolide resistance determinant. In addition, BLASTX identified a region of pLEW279a with approximately 60% amino acid identity to RepA replication initiation proteins of C. glutamicum plasmids pTET3 and pCG4 (29 kb; NC_004945) and a region with approximately 50% amino acid identity to TraA transfer proteins of Corynebacterium plasmids pGA2 (19 kb; NC_004535) and pNG2 (15 kb; NC_005001).…”

Genomics ofActinobacteria: Tracing the Evolutionary History of an Ancient Phylum

“…The replicases of pEC2, pEC3, and pKW4 have a number of features resembling those of the C. diphtheriae plasmid pNG2, including the RepA replication protein, indicating that they replicate by the rolling-circle mechanism (423-426, 429, 430, 496). Plasmids of this family have been found only in corynebacteria, except for pAP2 from Arcanobacterium pyogenes (212), but may, like pNG2, have a broad host range (212,351,390). The pEC2 plasmid of C. efficiens YS-314 is apparently a natural hybrid of two pNG2-like plasmids, since it contains two typical repA genes (CEP005 and CEP013).…”

Development and Application of Real-Time PCR Assays for Quantification of erm Genes Conferring Resistance to Macrolides-Lincosamides-Streptogramin B in Livestock Manure and Manure Management Systems

“…The use of tylosin on animals significantly increased the resistance by gut commensal bacteria to MLS B (9,17). Resistance to tylosin in a food animal production environment was found to be encoded by erm genes (1,18,19,43). The erm genes encode 23S rRNA adenine-specific N 6 -methyltransferases, which methylate the 23S rRNA of bacteria (28).…”