Trimethoprim Resistance Plasmids Research Articles

IS257-mediated cointegration in the evolution of a family of staphylococcal trimethoprim resistance plasmids

Analyses of the Staphylococcus epidermidis multiresistance plasmids pSK697 and pSK818 have revealed them to be closely related to the trimethoprim resistance plasmid pSK639, also isolated from S. epidermidis. pSK697 and pSK818 were found to contain a cointegrated copy of a second plasmid related to the S. epidermidis multidrug antiseptic and disinfectant resistance plasmid pSK108 and the S. aureus tetracycline resistance plasmid pT181, respectively. In contrast to pSK639, both plasmids were found to contain a third copy of IS257, such that the integrated plasmids in both cases are flanked by a copy of this element. This organization and the presence of duplicated sequences at the extremities of the integrated plasmids implicate IS257 in the formation of these cointegrate plasmids. Sequence analysis of the IS257 elements from these plasmids has provided insights into the probable mechanism of cointegration, viz., nonresolved replicative transposition of IS257.

Trimethoprim resistance in urinary pathogens in northern Scotland: epidemic spread of a resistance plasmid encoding the type Ib trimethoprim-resistant dihydrofolate reductase

The prevalence of trimethoprim resistance in enterobacterial urinary pathogens from hospitalised patients in the Angus district of northern Scotland (22.8%) was twice that found in similar isolates from patients attending general practitioners (11.2%). Thirty-three of the 143 trimethoprim-resistant strains were shown to harbour transferable plasmids conferring high-level trimethoprim resistance. In total, 17 different plasmid types were distinguished. Two plasmids, pUK1184 and pUK1185, accounted for 36% of the trimethoprim resistance plasmids and were shown by restriction endonuclease digestion fingerprints to be closely related to plasmid pUK28, previously demonstrated to be endemic in urinary pathogens in the Edinburgh area. Only 21% of the plasmids were shown to encode the type Ia trimethoprim-resistant dihydrofolate reductase, whereas 70% of the trimethoprim resistance plasmids were found to encode the type Ib dihydrofolate reductase. Hybridisation of the trimethoprim resistance plasmids identified in this study with gene probes specific for the integrase genes of transposons Tn7 and Tn21 indicates that the dhfrIa is rarely present within Tn7 or related transposons in these plasmids and may be more prevalent within Tn21-like transposons. In contrast, with the exception of the two endemic plasmids that harboured the dhfrIb gene within a Tn7-like transposon, the majority of dhfrIb genes were not found to be associated with either Tn7- or Tn21-like structures.

Distribution and transferability of plasmids encoding trimethoprim resistance in urinary pathogens from Greece

Of 505 strains of Enterobacteriaceae responsible for significant bacteriuria and isolated from hospital patients in two Greek cities in 1989, 151 strains (30%) were resistant to trimethoprim (MIC greater than or equal to 4 mg/L) and 220 (44%) were resistant to sulphamethoxazole (MIC greater than or equal to 64 mg/L); 127 (84%) of the trimethoprim-resistant strains exhibited high-level resistance (MIC greater than 1024 mg/L) and 121 (80%) were additionally resistant to four or more other antibiotics. Plasmids were detected in 141 (93%) of the trimethoprim-resistant strains. Trimethoprim resistance was encoded on self-transmissible plasmids in 79 (52%) of the resistant strains, and in a further seven strains (5%), plasmids coding for trimethoprim resistance could be mobilised by X+ factor. Co-transfer of various other antimicrobial resistances with trimethoprim resistance was observed, tetracycline resistance being the most common. The low degree of linkage observed between trimethoprim resistance and resistance to streptomycin and spectinomycin suggests that Tn7 is relatively uncommon in Greece. Classification of trimethoprim-resistance plasmids on the basis of their antimicrobial-resistance patterns and molecular mass revealed 39 different profiles. Overall, these findings differ from those from other European countries where the prevalence of transferable high-level trimethoprim resistance is low and where chromosomal Tn7-encoded trimethoprim resistance is common.

Trimethoprim resistance in urinary bacteria isolated from patients attending general practitioners

The incidence of trimethoprim resistance amongst enterobacterial strains responsible for significant bacteriuria in patients attending general practitioners in Edinburgh was 11.4%. Two-thirds of these resistant strains were highly resistant to trimethoprim. Trimethoprim resistance was more prevalent in bacteria isolated from men and from elderly patients. Less than half of the highly resistant strains possessed trimethoprim resistance plasmids; however, amongst those that did, one plasmid predominated. This plasmid was identical with the most successful trimethoprim resistance plasmid in hospital enterobacterial isolates at that time.

Restriction enzyme fingerprinting of trimethoprim resistance plasmids.

Restriction enzyme fingerprinting was applied to 72 transferable trimethoprim resistance plasmids to examine aspects of their epidemiology and molecular relatedness. These plasmids had previously been divided into 25 groups according to differences in mol. wts and in antimicrobial resistance determinants. Restriction enzyme fingerprinting allowed the plasmids to be further divided into 44 different groups. The groups based on molecular weight and resistance patterns often, but not invariably, corresponded with those based on restriction enzyme fingerprints. Some plasmids with the same mol. wt and resistance pattern had different digest fingerprints and conversely, although more rarely, plasmids which differed in molecular weight by as much as 10 MDa or in resistance pattern by one resistance marker, had indistinguishable fingerprints. The plasmids were initially divided into three broad categories according to which restriction enzymes gave fingerprints of 6-20 fragments. These categories differed in the molecular weights of the plasmids contained, the numbers of resistance markers, and the proportions of the plasmids which carried transposon Tn7. Some plasmids were more widespread and persistent than others with the same mol. wt and resistance pattern but with a different restriction enzyme fingerprint. Thus, application of this technique has shown the trimethoprim resistance plasmids studied to be more diverse than was indicated by determination of mol. wt and resistance pattern, and has indicated changes in the plasmid pool over the 3 years during which they were collected.

Trimethoprim resistance in commensal bacteria isolated from farm animals.

Trimethoprim resistance was examined in faecal bacteria obtained from chickens, sheep, cattle and pigs. The incidence of trimethoprim resistance in porcine strains was 17% (157/922) and, whereas 15.8% (146/922) of these bacteria were highly resistant, only 4% (37/922) of the isolates possessed trimethoprim resistance plasmids. Highly resistant porcine strains were obtained from 44% of the pig farms (41/93) but transferable trimethoprim resistance was found in isolates from 11% (10/93) of the farms. There was an association between the carriage of trimethoprim resistance plasmids and certain farms. Most of the resistance plasmids were not identical with those found in human clinical bacteria but one porcine plasmid was the same as the most ubiquitous trimethoprim resistance plasmid in Edinburgh.

Detection of New Low-Level Trimethoprim Resistance Plasmids

Detection of New Low-Level Trimethoprim Resistance Plasmids

Trimethoprim resistance plasmids in Escherichia coli isolated from cases of diarrhoea in cattle, pigs and sheep.

A total of 1572 isolates of Escherichia coli obtained from the faeces of young farm animals with diarrhoea over the period 1980-1983 were screened for resistance to trimethoprim (Tp). Resistance to Tp was detected in 263/954 (28%) of bovine isolates, 59/441 (13%) of porcine isolates and 15/177 (9%) of ovine isolates. Seventy-five resistant isolates from separate outbreaks of infection on farms within a 25 mile radius of Nottingham were examined in detail. Sixty-eight (91%) of the 75 isolates were resistant to greater than 1024 mg Tp/l and 34 (50%) of these 'highly resistant' isolates (45% of total resistant isolates) transferred their Tp resistance to E. coli K12. A further 13 (17%) isolates were demonstrated to carry non-self-transferable plasmids which were capable of being mobilized to E. coli K12 by the broad host range plasmid RP4. Thirty-one self-transferable Tp R plasmids were divided between the following incompatibility groups: IncB (14 plasmids), IncFII (4 plasmids), IncH2 (1 plasmid), IncI alpha (10 plasmids), IncI delta (1 plasmid) and IncP (1 plasmid). In terms of antibiotic resistance patterns and incompatibility properties, many of these plasmids closely resembled those isolated from human patients in the same area, suggesting that there may be a common pool of Tp R plasmids.

A follow-up survey of transferable, plasmid-encoded trimethoprim resistance in a general hospital (1975-1983).

Urinary isolates of bacteria resistant to trimethoprim were collected at the Whittington Hospital, London, during five three-monthly periods during 1975, 1977, 1979, 1981, and 1983. Seventy-six transferable trimethorpim-resistant isolates so obtained were examined for their antibiotic resistance patterns, trimethoprim resistance (TpR) levels and plasmid molecular weights. Overall between 1975 and 1983 the frequency of transferable TpR rose from 10% to 40% of trimethoprim-resistant isolates, caused mainly by an increasing number of Escherichia coli isolates which cotransferred antibiotic resistance to trimethoprim, streptomycin and spectinomycin. From 1975-1979 the rise in the frequency of transferable TpR was due to a gradual increase amongst in-patient isolates but since 1979 transferable TpR rose rapidly due to a ten-fold increase in out-patient isolates.

Trimethoprim resistance in urinary coliforms from patients in the community: plasmids and R-transfer.

A study of urinary coliform isolates, from patients attending their general practitioner, between 1981 and 1983 showed that 10.2% were resistant to trimethoprim. Forty-four trimethoprim-resistant Escherichia coli were collected for further study and the results compared with those obtained from hospital isolates collected during 1979-80 and 1982. Thirty-two (73%) were highly resistant to trimethoprim (MIC greater than 1024 mg/l) a similar proportion to that in the hospital isolates. The highly resistant isolates bore a close resemblance to the hospital isolates in that 66% carried multiple plasmids, 97% were multiply drug-resistant and all were highly resistant to sulphamethoxazole. The frequency of antimicrobial resistance transfer (59%) was similar to that from the hospital isolates. More than half of the trimethoprim resistance plasmids characterized were indistinguishable in terms of mol.wt and resistance pattern from those found in isolates from the hospital collections. Our results suggest that, whereas trimethoprim-resistant E. coli are less commonly isolated from patients in the community than in hospitals, many of the community isolates may have originated from a hospital source.

Trimethoprim resistance plasmids in faecal bacteria

Summary Of 320 patients surveyed in a general hospital's medical wards during a 6-month period in 1981, 18 (5.6%) harboured enterobacteria which contained resistance plasmids conferring resistance to trimethoprim (Tp). At the beginning of the study period, Tp-containing therapy was not the first choice for treatment of infections caused by these bacteria and the incidence of plasmid-determined resistance was 10%. When Tp alone was used as the first choice of treatment of urinary tract infections and the use of this antimicrobial was correspondingly increased, the proportion of strains that contained Tp resistance plasmids (R-plasmids) decreased to 4%. Relatively more patients with Tp-resistant strains in the bowel had significant bacteriuria compared with those without Tp-resistant organisms. The continuing occurrence of Tp-R-plasmid containing strains in the gut was associated with continuing antimicrobial therapy; the strains generally disappeared after antimicrobial therapy was stopped.

Evidence for an epidemic trimethoprim-resistance plasmid in fecal isolates of Escherichia coli from citizens of the United States studying in Mexico.

A previous study revealed the emergence of high-level resistance to trimethoprim-sulfamethoxazole in strains of Escherichia coli isolated from 95% of students from the United States who were taking either trimethoprim alone or trimethoprim-sulfamethoxazole for prophylaxis of travelers' diarrhea while in Guadalajara, Mexico. Many of these strains were subsequently demonstrated to cotransfer resistance to trimethoprim along with that to streptomycin and ampicillin. The present study demonstrated that at least 12 (60%) of 20 transferable (tra+) trimethoprim-resistance plasmids studied possessed both an identical Hind III restriction pattern and type I dihydrofolate reductase genes. Donor strains were shown to be distinct; this finding suggested that a common tra+ trimethoprim-resistance plasmid was widely disseminated among E. coli strains in Guadalajara. These results may explain in part the surprising degree of resistance encountered in trimethoprim-consuming persons in that region.

Bactericidal activity of trimethoprim alone and in combination with sulfamethoxazole on susceptible and resistant Escherichia coli K-12

The combined effects of trimethoprim and sulfamethoxazole on the viability of Escherichia coli K-12 and resistant strains possessing resistance plasmids were examined in minimal medium. When methionine, glycine, and adenine were present, sulfamethoxazole could enhance trimethoprim activity against E. coli K-12 so that the combination was bactericidal. However, this enhancement occurred over a narrow range of trimethoprim concentrations (0.04 to 0.2 mg liter-1) and only when the sulfamethoxazole concentration was more than 10 times that of trimethoprim. Under certain conditions, sulfamethoxazole enhanced trimethoprim bactericidal activity against E. coli K-12 carrying plasmid R1 at concentrations of sulfamethoxazole far below those required to inhibit the organism, but there was no such enhancement with the same host containing the SSu plasmid. Similar differences were found with strains possessing trimethoprim resistance plasmids R483 and R751. Sulfamethoxazole can promote a bactericidal response with trimethoprim in E. coli K-12 and some of its resistant derivatives, but only under a narrow range of concentrations.

Occurrence of transposable trimethoprim resistance in clinical isolates of Escherichia coli devoid of self-transmissible resistance plasmids.

Fifty trimethoprim-resistant clinical isolates of Escherichia coli, devoid of self transmissible trimethoprim resistance plasmids, were examined for the presence of trimethoprim resistance transposons. Trimethoprim resistance was mobilized from 12 strains by transposition onto plasmid RP4. The trimethoprim resistance transposons isolated comprised two groups: those with and without linked streptomycin resistance.

Distribution of transferable trimethoprim and gentamicin resistance plasmids in Enterobacteriaceae

Following the employment of trimethoprim/sulfonamid and gentamicin in the general clinical praxis transferable plasmids with respective resistance function were identified at first in enteric bacteria from sewage before they could be detected in patient strains. Whereas the trimethoprim resistance plasmids represent different incompatibility groups (C, FII, I5, K, M, N, S, U) the gentamicin resistance plasmids are very similar IncM members of 62 MD in size. Therefore a clonal distribution of a particular gentamicin resistance plasmid has to be taken into consideration.

Verbreitung transferabler Trimethoprimresistenz‐ und Gentamicinresistenzplasmide in Enterobacteriaceae

AbstractFollowing the employment of trimethoprim/sulfonamid and gentamicin in the general clinical praxis transferable plasmids with respective resistance functions were identified at first in enteric bacteria from sewage before they could be detected in patient strains. Whereas the trimethoprim resistance plasmids represent different incompatibility groups (C, FII, I5, K, M, N, S, U) the gentamicin resistance plasmids are very similar IncM members of 62 MD in size. Therefore a clonal distribution of a particular gentamicin resistance plasmid has to be taken into consideration.

Characterization of a R plasmid-associated, trimethoprim-resistant dihydrofolate reductase and determination of the nucleotide sequence of the reductase gene.

The trimethoprim-resistant dihydrofolate reductase associated with the R plasmid R388 was isolated from strains that over-produce the enzyme. It was purified to apparent homogeneity by affinity chromatography and two consecutive gel filtration steps under native and denaturing conditions. The purified enzyme is composed of four identical subunits with molecular weights of 8300. A 1100 bp long DNA segment which confers resistance to trimethoprim was sequenced. The structural gene was identified on the plasmid DNA by comparing the amino acid composition of the deduced proteins with that of the purified enzyme. The gene is 234 bp long and codes for 78 amino acids. No homology can be found between the deduced amino acid sequence of the R388 dihydrofolate reductase and those of other prokaryotic or eukaryotic dihydrofolate reductases. However, it differs in only 17 positions from the enzyme associated with the trimethoprim-resistance plasmid R67.

Trimethoprim resistance plasmids of different origin encode different drug-resistant dihydrofolate reductases

Abstract Several plasmids mediating resistance to folic acid analogs were studied. The plasmids were in part newly isolated from clinical material and in part R factors studied earlier, such as R483, R721, R751, and R388. By gel chromatography, plasmid-carrying bacterial strains were all found to produce drug-resistant dihydrofolate reductases of a molecular weight distinctly larger than that of the chromosomal enzyme of the host. By gel electrophoresis and zymographic detection technique, analog inhibition characteristics, heat sensitivity, and pH optimum curves, the dihydrofolate reductases induced by R483, R751, and R388, respectively, could be clearly discerned as separate enzymes. Of the newly isolated plasmids all but one coded for a dihydrofolate reductase similar to that of R483. The aberrant one seemed to yield a new enzyme variant as judged from its drug inhibition characteristics and its pH optimum profile. Large differences in drug insensitivity were observed, thus the R751 and R388 enzymes were virtually insensitive to folic acid analogs, whereas the corresponding enzymes from the newly isolated plasmids, and from R483 showed a substantially higher sensitivity. On the other hand these latter enzymes were overproduced, in that the plasmid-carrying bacteria showed a 10- to 20-fold higher content of dihydrofolate reductase than the plasmid-free host strain. Among newly isolated trimethoprim-resistant strains, one was found which overproduced dihydrofolate reductase about 200-fold. In this case the enzyme was only slightly more resistant to folic acid analogs than the chromosomal Escherichia coli K-12 enzyme, and did not seem to be plasmid borne.

Isolation of a nontransmissible antibiotic resistance plasmid by transductional shortening of R factor RP1.

A plasmid segregant carrying tetracycline and carbenicillin resistance markers has been isolated from R factor RP1 by transductional shortening with phage P22. The new plasmid RP1-S2, which has a molecular weight of 23 times 10-6, has lost the transfer, phage sensitivity, and neomycin resistance functions of RP1. It combines readily with a W group plasmid, R388, to form a transmissible carbenicillin and trimethoprim resistance plasmid, RWP1.