Multidrug-resistant Salmonella enterica serovar Muenchen from pigs and humans and potential interserovar transfer of antimicrobial resistance

Abstract

Salmonella serovars are important reservoirs of antimicrobial resistance. Recently, we reported on multidrug-resistant (MDR) Salmonella enterica serovar Typhimurium strains among pigs with resistance to ampicillin, kanamycin, streptomycin, sulfamethoxazole, and tetracycline (resistance [R] type AKSSuT) and resistance to amoxicillin-clavulanic acid, ampicillin, chloramphenicol, streptomycin, sulfamethoxazole, and tetracycline (R type AxACSSuT). In the present study, 67 isolates (39 from humans and 28 from pigs) of clinically important Salmonella serovar Muenchen were characterized. Among the porcine isolates, 75% showed resistance to seven antimicrobials: ampicillin, chloramphenicol, streptomycin, sulfamethoxazole, tetracycline, amoxicillin-clavulanic acid, and kanamycin (R type ACSSuTAxK). One isolate from humans showed resistance to 10 of the 12 antimicrobials: ampicillin, chloramphenicol, streptomycin, sulfamethoxazole, tetracycline, amoxicillin-clavulanic acid, kanamycin, gentamicin, cephalothin, and ceftriaxone (R type ACSSuTAxKGCfCro). Pulsed-field gel electrophoresis revealed no clonality between the porcine and the human strains. The porcine and the human MDR strains carried class 1 integrons of 2.0 and 1.0 kb, respectively. Genes specific to the porcine strain included aadA2, aphA1-Iab, and tetA(B). DNA sequencing revealed that the porcine isolates carried bla(OXA-30) on a class 1 integron. Genes specific to the human strain included bla(TEM), strA, strB, cmlA, tetA(A), and aadA2. No bla(CMY-2) gene was detected. Serovar Muenchen strains of porcine and human origin were able to transfer resistance genes to laboratory strain Escherichia coli MG1655 by conjugation. Plasmid restriction with four restriction enzymes, EcoRI, BamHI, HindIII, and PstI, showed that the conjugative plasmids from porcine Salmonella serovar Muenchen and Typhimurium R-type MDR strains isolated from the same farms at the same time were similar on the basis of the sizes and the numbers of bands and Southern hybridization. The plasmid profiles among the Salmonella serovar Muenchen isolates from the two host species were different. This is the first report to show a high frequency of MDR Salmonella serovar Muenchen strains from pigs and a human strain that is similar to the MDR isolates with the AmpC enzyme previously reported among Salmonella serovars Newport and Typhimurium strains. The MDR strains from the two host species independently represent public health concerns, as Salmonella serovar Muenchen is among the top 10 causes of salmonellosis in humans.

FIG. 1.

PFGE fingerprints of 21 S. enterica serovar Muenchen isolates from swine and humans. ^a, two major clusters are shown. The vertical dotted line at 80% genetic similarity was considered an arbitrary threshold for grouping of the clusters into subgroups. Clusters 1 (with four subgroups, subgroups 1-A, 1-B, 1-C, and 1-D) and 2 (with one subgroup, subgroup 2-A) are shown. ^b, origin shows the different regions of North Carolina where the samples originated (NE, northeastern; SE, southeastern; NW, northwestern; SW, southwestern). All the swine isolates originated from the southeastern region of the state, the part of the state where pig production predominates.

FIG. 2.

PCR amplification of antimicrobial resistance genes and class 1 integron variable region among MDR Salmonella serovar Muenchen isolates from swine and humans. (A) Lanes: 1, sul1; 2, tetA(B); 3 and 4, qacE; 5 and 6, aphA1-Iab; and 7, molecular size marker (0.019 to 1.11 kb). (B) Lanes: 1, tetA(G); 2 and 3, tetA(A) (strains NCPH3633 and NCPH110); 4, strA; 5, qacE; 6, sul1; 7,bla_TEM; 8, aadA2; and 9, molecular size marker. (C) Lanes: 1, molecular size marker; 2, strain NCPH3633 (a human Salmonella serovar Muenchen isolate of R type ACSSuTAxKGCfCro), 3, isolate UCE1; 4, isolate UCE3; 5, isolate UCM6 (the isolates in lanes 3, 4, and 5 are swine Salmonella serovar Muenchen isolates of R type ACSSuTAxK); 6, isolate UT30 (Salmonella serovar Typhimurium phage type DT193, swine isolate of R type AKSSuTG); and 7, isolate UW05 (a Salmonella serovar Typhimurium phage type DT104 swine isolate of R type AxACSSuT).

FIG. 3.

Plasmid restriction analysis with four restriction enzymes: PstI, HindIII, EcoRI, and BamHI. Lanes: 1, Salmonella serovar Muenchen (human origin, p3633, R type ACSSuTAxKGCfCro); 2, Salmonella serovar Typhimurium (porcine origin, pUCE15, R type AKSSuT); 3, Salmonella serovar Muenchen (porcine origin, pUCE6, R type ACSSuTAxK); and M, molecular size marker. Relatively more similar plasmid restriction patterns were detected between Salmonella serovar Muenchen and Typhimurium isolates of porcine origin than Salmonella serovar Muenchen isolates of swine and human origin.

FIG. 4.

Southern hybridization of three strains: lane 1, Salmonella serovar Muenchen (human origin, p3633, R type ACSSuTAxKGCfCro); lane 2, Salmonella serovar Typhimurium (porcine origin, pUCE15, R type AKSSuT); lane 3, Salmonella serovar Muenchen (porcine origin, pUCE6, R type ACSSuTAxK), and lane M, molecular size marker. A DIG-labeled acetylphosphotransferase gene (aphA1-Iab) was used. Both MDR Salmonella serovar Typhimurium and MDR Salmonella serovar Muenchen isolates of porcine origin had the gene on the same restriction fragment, which was 9.5 kb.