Plasmid-associated transfer of tetracycline resistance in Bacteroides ruminicola

Abstract

Tetracycline resistance was transferred at frequencies between l0-7 and 10-6 per recipient cell in anaerobic matings between two strains of the strictly anaerobic rumen bacterium Bacteroides ruminicola. The donor strain, 223/M2/7, was a multiple-plasmid-bearing tetracycline-resistant strain from the ovine rumen, and the recipient, F101, was a rifampin-resistant mutant of B14, a bovine strain belonging to B. ruminicola subsp. brevis. Resistance transfer could occur in the presence of DNase, but not in dummy mating mixtures in which filtrate from a donor culture replaced donor cells. Acquisition of tetracycline resistance by the recipient was accompanied by the appearance of a 19.5-kilobase pair plasmid (designated pRRI4) which was homologous with a plasmid of similar size and restriction pattern present in the donor strain. A transconjugant (F115) carrying pRRI4 was also able to act as a donor of tetracycline resistance and plasmid DNA in matings with another recipient. Derivatives of F115 that had spontaneously lost tetracycline resistance lacked detectable plasmid DNA. It is concluded that pRRI4 mediated the transfer of tetracycline resistance. Transfer of resistance was not detectably enhanced by pregrowth of the donor in medium containing tetracycline. Transfer of tetracycline resistance was not detected from 223/M2/7 to a strain, 23, belonging to B. ruminicola subsp. ruminicola.

Use of antibiotic resistance mutations to track strains of obligately anaerobic bacteria introduced into the rumen of sheep

“…For selective enumeration of strain F100, rifampicin and tetracycline were added to give final concentrations of 50 pg/ml and 10 pg/ml, respectively, to the molten medium (held at 60°C). Plates were poured immediately in an anaerobic atmosphere and incubated as described previously (Flint et al 1988a). Dilutions of homogenized rumen samples were prepared anaerobically in a solution containing (g/l): casitone, 1; NaHCO,, 5 ; cysteine, 1; 1 ml/l ti Tween 80 and 150 ml/l each of minerals solutions I and 11.…”

Evidence for recent intergeneric transfer of a new tetracycline resistance gene, tet(W), isolated from Butyrivibrio fibrisolvens, and the occurrence of tet(O) in ruminal bacteria

“…Strains of obligately anaerobic rumen bacteria that show resistance to tetracycline have been isolated from cattle and sheep, which have been fed diets free of antibiotics (Flint et al, 1988a). Prevotella strains are known to harbour tet (Q) genes; in one case, a plasmid-encoded tet (Q) gene was shown to be transmissible to rumen Prevotella and to human Bacteroides spp., while in other strains the tet (Q) marker was located chromosomally (Flint et al, 1988b;Shoemaker et al, 1992;Salyers et al, 1995a). The nature and location of tetracycline resistance genes in other predominant rumen genera such as Selenomonas and Butyrivibrio has remained unknown, but a recent investigation of Tc R in Butyrivibrio showed that two strains were able to transfer Tc R at high frequency in vitro via a chromosomal element.…”

Molecular genetics of obligate anaerobes from the rumen

“…Bacteriophage-like particles are abundant, and phage have been isolated that infect some of the major bacterial species such as Selenomonas ruminantium and P. ruminicola [21,22]. Extrachromosomal plasmid DNA appears to be abundant in several rumen species such as S. ruminantium, but rare in others such as P. ruminicola and F. succinogenes [2,23,24]. It may be argued that other conditions limit gene transfer; these include the high levels of nucleases in rumen fluid and the low growth rates of many organisms in anaerobic environments.…”

“…Coliform bacteria have been shown to exchange antibiotic resistance plasmids, but only under conditions where the host animal is starved and rumen volatile fatty acid levels are severely depressed [26]. Strains showing resistance to tetracycline are found belonging to some of the major obligately anaerobic bacterial species and tetracycline resistance in a P. ruminicola strain was found to be due to a plasmid-encoded tetQ gene that shows a high degree of sequence homology with chromosomally located tetQ genes from hind gut Bacteroides [24,27]. Under laboratory conditions this plasmid was capable of transmission to another strain of P. ruminicola, and to strains of human hind gut Bacteroides [24,27].…”

“…Strains showing resistance to tetracycline are found belonging to some of the major obligately anaerobic bacterial species and tetracycline resistance in a P. ruminicola strain was found to be due to a plasmid-encoded tetQ gene that shows a high degree of sequence homology with chromosomally located tetQ genes from hind gut Bacteroides [24,27]. Under laboratory conditions this plasmid was capable of transmission to another strain of P. ruminicola, and to strains of human hind gut Bacteroides [24,27]. The possibility clearly exists that genetic interchange involving obligate anaerobes can occur naturally between related organisms from different habitats [27].…”