Submitted to: American Society for Microbiology
Publication Type: Abstract Only
Publication Acceptance Date: 5/22/2003
Publication Date: 5/22/2003
Citation: STANTON, T.B., MCDOWALL, J.S., RASMUSSEN, M.A. SELECTIVE ISOLATION OF SWINE MEGASPHAERA ELSDENII STRAINS WITH DIVERSE TETRACYCLINE RESISTANCE GENOTYPES, INCLUDING GENE MOSAICS OF TET(O) AND TET(W). AMERICAN SOCIETY FOR MICROBIOLOGY. 2003. ABSTRACT P. 684.
Technical Abstract: In that tetracycline has been commonly added to swine feed for disease prevention and growth promotion purposes, the microbial ecosystem of the swine intestinal tract seems a good choice for investigating tet gene ecology. As a basis for such investigations, we are analyzing tetracycline-resistant (Tc**r) intestinal anaerobes and their resistance mechanisms. Previously we isolated eight Tc**r Megasphaera elsdenii strains by plating swine cecal samples on CCA agar medium containing 64-256 ug chlortetracycline/ml (Abstract Z-42, ASM 2002 Annual Mtg). M. elsdenii is a lactate-fermenting, anaerobic coccus and common resident of mammalian GI tracts. Two novel tet genes (designated 'tetOWO', for the sake of convenience) were identified in the Tc**r strains. The genes are mosaic, i.e., formed by double crossover recombination between tet(O) and tet(W) genes. Considering that mosaic genes have not been described for other bacteria, we hypothesized that tet(O) and tet(W) recombination within M. elsdenii may produce tetOWO genes. If so, additional M. elsdenii strains may have alternative tet genotypes. Culture medium Me109M was formulated to include lactate (1%, w/v) and monensin (50 ug/ml) for selectively isolating M. elsdenii from swine feces. Under anaerobic conditions, fecal bacterial counts were 6 x 10**10 cfu/gm on CCA agar and 7 x 10**8 cfu/gm on Me109M. About 1/3 of the colonies on Me109M were M. elsdenii strains. Thirty-one M. elsdenii strains were analyzed by PCR assays developed to differentiate among tet(O), tet(W), and tetOWO genes. Nine strains contained no detectable tet genes. Twelve had tetOWO genes. Eight strains displayed a new mosaic genotype, tetOW, produced from a single crossover recombination between tet(O) and tet(W). Two strains contained tet(W). A bovine rumen M. elsdenii strain, isolated separately, contained a tet(O) gene. We are currently examining a possible correlation between tet genotype and resistance phenotype (tetracycline MIC values). These results indicate M. elsdenii strains have diverse tet genotypes, and are preliminary, indirect evidence that tet genes can evolve through recombination in this species.