Persistence of antibiotic resistance: evaluation of a probiotic approach using antibiotic-sensitive Megasphaera elsdenii strains to prevent colonization of swine by antibiotic-resistant strains

Abstract

Megasphaera elsdenii is a lactate-fermenting, obligately anaerobic bacterium commonly present in the gastrointestinal tracts of mammals, including humans. Swine M. elsdenii strains were previously shown to have high levels of tetracycline resistance (MIC=64 to >256 μg/ml) and to carry mosaic (recombinant) tetracycline resistance genes. Baby pigs inherit intestinal microbiota from the mother sow. In these investigations we addressed two questions. When do M. elsdenii strains from the sow colonize baby pigs? Can five antibiotic-sensitive M. elsdenii strains administered intragastrically to newborn pigs affect natural colonization of the piglets by antibiotic-resistant (AR) M. elsdenii strains from the mother? M. elsdenii natural colonization of newborn pigs was undetectable (<10(4) CFU/g [wet weight] of feces) prior to weaning (20 days after birth). After weaning, all pigs became colonized (4 × 10(5) to 2 × 10(8) CFU/g feces). In a separate study, 61% (76/125) of M. elsdenii isolates from a gravid sow never exposed to antibiotics were resistant to chlortetracycline, ampicillin, or tylosin. The inoculation of the sow's offspring with mixtures of M. elsdenii antibiotic-sensitive strains prevented colonization of the offspring by maternal AR strains until at least 11 days postweaning. At 25 and 53 days postweaning, however, AR strains predominated. Antibiotic susceptibility phenotypes and single nucleotide polymorphism (SNP)-based identities of M. elsdenii isolated from sow and offspring were unexpectedly diverse. These results suggest that dosing newborn piglets with M. elsdenii antibiotic-sensitive strains delays but does not prevent colonization by maternal resistant strains. M. elsdenii subspecies diversity offers an explanation for the persistence of resistant strains in the absence of antibiotic selection.

Fig. 1.

Timeline for Megasphaera elsdenii inoculation and fecal sample collection (experiment B). Numbers represent days relative to farrowing (birth) date: above timeline, when offspring pigs were inoculated intragastrically with a mix of five M. elsdenii strains, and below timeline, when fecal samples were taken for enumeration and selective isolation of M. elsdenii. Pigs were weaned at 20 days after farrowing. Three fecal samples were taken from the sow at times indicated and analyzed for M. elsdenii.

Fig. 2.

SNP differences in hgd-odt assembled sequences of M. elsdenii strain 14-14 and inoculum strains used in these studies. The hgd and odt genes of M. elsdenii strains were amplified and sequenced. Portions of the gene sequences were spliced together at the added GGGGG sequence in the figure. SNPs in the assembled sequences were used to fingerprint M. elsdenii inoculum strains recovered from pigs. Shaded letters represent consensus nucleotides and, when there are equal numbers of nucleotides at a base position, the strain 14-14 nucleotide at that position. An hgd amplicon for strain F103 was not detected. Complete hgd and odt sequences for M. elsdenii strain 14-14 have been deposited in GenBank.