Distribution of oxytetracycline resistance plasmids between aeromonads in hospital and aquaculture environments: implication of Tn1721 in dissemination of the tetracycline resistance determinant tet A - PubMed (original) (raw)

Distribution of oxytetracycline resistance plasmids between aeromonads in hospital and aquaculture environments: implication of Tn1721 in dissemination of the tetracycline resistance determinant tet A

G Rhodes et al. Appl Environ Microbiol. 2000 Sep.

Abstract

Oxytetracycline-resistant (OT(r)) mesophilic aeromonads were recovered from untreated hospital effluent (72 isolates) and from fish farm hatchery tanks (91 isolates) at sites within the English Lake District, Cumbria, England. The transfer of OT(r) plasmids from these isolates was investigated. Using Escherichia coli J53-1 as a recipient, 11 isolates from the hospital site and 6 isolates from the fish farm site transferred OT(r) plasmids (designated pFBAOT1 to 17). Original isolates were identified using fatty acid methyl ester and fluorescent amplified fragment length polymorphism comparisons as either Aeromonas hydrophila HG3 (eight isolates), A. veronii b.v. sobria HG8 (six isolates), and A. caviae HGB5 (one isolate). One isolate remained unidentified, and one could not be assigned a taxonomic designation beyond the genus level. Plasmids pFBAOT1 to -17 were screened for the presence of the tetracycline resistance determinants Tet A to E and Tet G. Only determinant Tet A (10 plasmids) was detected in these plasmids, with 7 tet gene determinants remaining unclassified. In all cases, Tet A was located on a 5.5-kb EcoRI restriction fragment. Hybridization with inc-rep probes N, P, Q, W, and U showed pFBAOT3, -4, -5, -6, -7, -9, and -11, from the hospital environment, to be IncU plasmids. Further, restriction fragment length polymorphism (RFLP) analyses and DNA probing demonstrated that pFBAOT plasmids were closely related to IncU OT(r) plasmids pASOT, pASOT2, pASOT3, pRAS1 (originally isolated from A. salmonicida strains from fish farms in Scotland and Norway, respectively), and pIE420 (isolated from a German hospital E. coli strain). In addition, DNA analyses demonstrated that plasmids pRAS1 and pIE420 had identical RFLP profiles and that all fragments hybridized to each other. The presence of tetracycline resistance transposon Tn1721 in its entirety or in a truncated form in these plasmids was demonstrated. These results provided direct evidence that related tetracycline resistance-encoding plasmids have disseminated between different Aeromonas species and E. coli and between the human and aquaculture environments in distinct geographical locations. Collectively, these findings provide evidence to support the hypothesis that the aquaculture and human compartments of the environment behave as a single interactive compartment.

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Figures

FIG. 1

FIG. 1

(A) RFLP profiles (generated by _Eco_RI digestion) of IncU OTr plasmids from aeromonads and corresponding Southern hybridizations with determinant Tet A and pASOT plasmid. Lanes: 1 and 11, λ DNA markers digested with _Hin_dIII and a 1-kb ladder, respectively; 2, pIE420; 3, pASOT; 4, pFBAOT3; 5, pFBAOT4; 6, pFBAOT5; 7, pFBAOT6; 8, pFBAOT7; 9, pFBAOT9; 10, pFBAOT11. (B) Hybridization of the Tet A determinant probe to plasmids. All plasmids harbored Tet A on a 5.5-kb fragment. (C) Hybridization of plasmid pASOT to IncU plasmids.

FIG. 2

FIG. 2

Comparison of plasmids pRAS1 and pIE420 by hybridization of radiolabeled plasmid pRAS1 to both plasmids. Lanes: 1, pRAS1; 2, pIE420.

FIG. 3

FIG. 3

Physical and genetic structure of transposon Tn_1721_ (length, 11,139 bp). The 38-bp inverted repeats IRL, IRR-I, and IRR-II are highlighted. The designation, position, and direction of transcription of genes are shown by the arrowed boxes. The region that comprises the minor transposon Tn_1722_ is highlighted, along with the location of _Eco_RI restriction sites. This diagram was adapted from Schnabel and Jones (39) and Tsuda et al. (45).

FIG. 4

FIG. 4

(A) _Eco_RI-generated RFLP patterns for IncU OTr plasmids. The gel was electrophoresed for an extended period in order to separate the fragments in the 5- to 7-kb range. Lanes: 1 and 12, λ DNA markers digested with _Hin_dIII and a 1-kb ladder, respectively; 2, pMT1286; 3, pASOT; 4, pIE420; 5, pFBAOT3; 6, pFBAOT4; 7, pFBAOT5; 8, pFBAOT6; 9, pFBAOT7; 10, pFBAOT9; 11, pFBAOT11; 12, 1-kb marker DNA. (B) Southern hybridization of the MCP probe to the DNA in panel A.

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