Phenotypic and genotypic characterization of antimicrobial resistance in Escherichia coli O111 isolates

OBJECTIVES: The aim of this study was to generate baseline data on the prevalence and molecular basis of antimicrobial resistance in Escherichia coli O111 isolates. METHODS: A total of 105 epidemiologically unrelated E. coli O111 isolates from humans and cattle (isolated between 1983 and 2003) were tested for susceptibility to 17 antimicrobial agents by broth microdilution. Resistant isolates were screened by molecular methods for resistance genes, class 1 and 2 integrons and mutations in the quinolone-resistance determining regions. RESULTS: Resistance was found in 76% of the isolates, with a prevalence of 72% for multiresistance. The most prevalent resistances were to streptomycin, sulfamethoxazole and tetracycline (72-68%), followed by spectinomycin, ampicillin and kanamycin/neomycin (39-25%). For each antimicrobial agent, the predominant resistance genes were ampicillin, bla(TEM) (94%); chloramphenicol, catA1 (100%); gentamicin, aac(3)-IV and aac(3)-II (50% each); kanamycin, aphA1 (100%); streptomycin, aadA1- like (66%); sulfamethoxazole, sul1 (59%); tetracycline, tet(A) (86%); and trimethoprim, dfrA1-like (83%). Class 1 integrons were found in 41% of the isolates. They carried aadA1, dfrA1-aadA1 and dfrA15-aadA1. A class 2 integron (dfrA1-sat1-aadA1) was found in one isolate. Only three isolates (3%) were resistant to nalidixic acid (reduced susceptibility to ciprofloxacin), with a single mutation in the gyrA gene. CONCLUSIONS: E. coli O111 strains exhibit a wide repertoire of genetic elements to sustain antimicrobial pressure. Two specific antimicrobial resistance pheno/genotypes, [STR-SPT]-SUL-TET/aadA1-sul1-tet(A) and STR-SUL-TET-AMP-[KAN-NEO]/strA/B-sul2-tet(A)-bla(TEM)-aphA1, are predominant.     

Occurrence of integrons and resistance genes among sulphonamide-resistant Shigella spp. from Brazil

OBJECTIVES: To determine the occurrence of class 1 and 2 integrons and antimicrobial resistance genes among sulphonamide-resistant Shigella strains isolated in Brazil during 1999-2003. METHODS: Sixty-two Shigella (Shigella flexneri, n = 47 and Shigella sonnei, n = 15) were tested against 21 antimicrobial agents. The presence of integrons classes 1 and 2 and antimicrobial resistance genes was investigated by PCR using specific primers. RESULTS: A total of eight antimicrobial resistance profiles were identified, with the profile of resistance to sulfamethoxazole, trimethoprim, spectinomycin, streptomycin and tetracycline being the most common among S. sonnei, and additionally to ampicillin and chloramphenicol among S. flexneri. Class 1 integrons were found in only two strains, whereas class 2 integrons were found in 56 (90.3%) of the strains. All class 2-positive strains had a similar fragment of 2214 bp harbouring a gene cassette array conferring resistance to trimethoprim, streptothricin and spectinomycin/streptomycin. The genes coding for resistance to chloramphenicol (catA1), tetracycline [tet(A) and tet(B)] and ampicillin (bla(OXA) and bla(TEM)), were detected in resistant strains. CONCLUSIONS: The detection of class 1 and 2 integrons and additional antimicrobial resistance genes allowed us to identify the most frequent antimicrobial resistance patterns of Shigella spp. isolated in Brazil.     

Trends in antimicrobial resistance, phage types and integrons among Salmonella serotypes from pigs, 1997-2000

OBJECTIVES: The objectives of this study were to determine antimicrobial resistance and to identify phage types and class 1 integrons among non-typhoidal Salmonella isolates from 24 pig farms in North Carolina collected between 1997 and 2000. METHODS: A total of 1314 isolates of 30 serotypes from pig faecal samples were collected and analysed over a 3 year period. The isolates were characterized using antimicrobial susceptibility testing, phage typing, PCR and DNA sequencing for class 1 integrons. RESULTS: A high frequency of resistance to antimicrobial agents including tetracycline (85%), ampicillin (47%), co-amoxiclav (23%) and chloramphenicol (21%) was detected. Two multidrug resistance patterns were common in Typhimurium (including variant Copenhagen): isolates with co-amoxiclav, ampicillin, chloramphenicol, streptomycin, sulfamethoxazole and tetracycline (R-type AxACSSuT) [36%] and isolates with ampicillin, kanamycin, streptomycin, sulfamethoxazole and tetracycline (R-type AKSSuT) [45%] resistance patterns. Definitive Type 104 (DT104) was the most common (34%) among eight phage types identified. AKSSuT was found among non-DT104 phage types, particularly DT21 and DT193. Class 1 integrons were detected among various serotypes including Typhimurium, Derby, Muenchen, Worthington, Bere and Muenster. aadA was the most common resistance gene insert, and the oxa30 beta-lactamase resistance gene was also identified among serovar Muenchen. CONCLUSIONS: In this study, two most important multidrug resistance patterns (AxACSSuT and AKSSuT) and phage types of public health significance (DT104 and DT193) constituted two-thirds of the serotype Typhimurium isolates. The findings imply that pigs raised in the commercial production system may pose a risk in serving as reservoirs of resistant Salmonella.     

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

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.     

Molecular mechanisms of resistance in multidrug-resistant serovars of Salmonella enterica isolated from foods in Germany

OBJECTIVES: The objectives of this study were to determine antimicrobial susceptibility and to characterize the molecular mechanisms of multidrug resistance among German food-borne Salmonella isolates of different serovars. METHODS: A total of 319 epidemiologically independent multidrug-resistant isolates from German foodstuffs comprising 25 different serovars were tested for their antimicrobial susceptibility by broth microdilution. The presence of antimicrobial resistance genes, integrons of classes 1 and 2 and their integrated resistance gene cassettes as well as the Salmonella genomic island 1 (SGI1) was investigated by PCR and DNA sequencing. Localization of integrons and relevant resistance genes was done by Southern hybridization. Sequence analysis revealed mutations in the quinolone resistance-determining region of the gyrA gene. RESULTS: The most prevalent resistances found in the multidrug-resistant serovars of Salmonella enterica from foods were to streptomycin (94%), sulfamethoxazole (92%), tetracycline (81%), ampicillin (73%), spectinomycin (72%), chloramphenicol (48%) and trimethoprim (27%). Twenty-four resistance genes covering six antimicrobial families (beta-lactams, aminoglycosides, phenicols, sulphonamides, tetracycline, and trimethoprim) were identified in the food isolates, many of them integrated as gene cassettes in class 1 and class 2 integrons. Class 1 integrons were detected in 65% of the multidrug-resistant Salmonella isolates comprising 16 different serovars, while class 2 integrons were found in 10% of the isolates belonging to two serovars only. The results demonstrate a clear predominance of both SGI1-borne resistance genes and class 1 integrons in Salmonella serovar Typhimurium DT104 and of class 2 integrons in Salmonella serovar Paratyphi B (d-tartrate positive). Nalidixic acid resistance found in 15% of the isolates was associated with single mutations in the gyrA gene. CONCLUSIONS: This study confirms the role of foods of animal and other origin as a reservoir of multidrug-resistant Salmonella and underlines the need for continuing surveillance of food-borne zoonotic bacterial pathogens along the food chain.     

Molecular characterization of integrons in non-typhoid Salmonella serovars isolated in Japan: description of an unusual class 2 integron

OBJECTIVES: To characterize class 1 and class 2 integrons among non-typhoid Salmonella enterica serovars in Japan, and also to monitor the spread of the multidrug-resistant S. enterica serovar Typhimurium DT104. METHODS: A total of 105 Salmonella isolates were included in this study. The broth microdilution method was used to determine the MIC values of a range of antibiotics for these isolates. PCR and DNA sequencing were used for screening and characterization of class 1 and class 2 integrons. RESULTS: PCR sequencing analysis revealed the presence of seven profiles of class 1 integrons in addition to a new type of class 2 integron. The identified gene cassettes within class 1 integrons were as follows; aadA1, aadA2 and aadA5, which confer resistance to streptomycin and spectinomycin; aadB, which confers resistance to gentamicin, kanamycin and tobramycin; dfrA1 and dfrA17, which confer resistance to trimethoprim; bla(PSE-1), which confers resistance to ampicillin; catB3, which confers resistance to chloramphenicol; and sat1, which confers resistance to streptothricin. Two strains of the multidrug-resistant S. Typhimurium DT104 were characterized in this study. DNA sequencing of class 2 integrons identified one with an unusual array of gene cassettes, sat, sat1 and aadA1. CONCLUSIONS: In this study, we characterized the antibiotic resistance gene cassettes within class 1 integrons in different isolates of non-typhoid Salmonella serovars, and we also identified a new type of class 2 integron.     

Phenotypic and genotypic characterization of antimicrobial resistance in German Escherichia coli isolates from cattle,swine and poultry

OBJECTIVE: Phenotypic and genotypic characterization of the antimicrobial resistance of German Escherichia coli strains isolated during 1999-2001 from cattle, swine and poultry. MATERIALS AND METHODS: Three hundred and seventeen isolates were tested for their resistance to 17 antimicrobial agents by broth microdilution. Resistant strains were screened by molecular methods for resistance genes, integrons and mutations in quinolone-resistance determining regions. RESULTS: Resistance was found in 40% and multiresistance in 32% of the strains. The resistance was significantly higher in isolates from poultry (61%) and swine (60%) than from cattle (25%) (P < 0.01). The most prevalent resistances were to sulfamethoxazole, tetracycline, streptomycin, ampicillin and spectinomycin (30-15%). For each antibiotic, the predominant resistance genes were: ampicillin, blaTEM1-like (92%); chloramphenicol, catA (68%) and cmlA1-like (36%); gentamicin, aac(3)-IV (60%); kanamycin, aphA1 (100%); streptomycin, aadA1-like (61%) and strA/B (59%); sulfamethoxazole, sul2 (66%), sul1 (42%) and sul3 (14%); tetracycline, tet(A) (66%) and tet(B) (42%); and trimethoprim, dfrA1-like (77%), dfrA17 (13%) and dfrA12 (7%). Class 1 integrons were found in 30% of the strains. They carried dfrA1-aadA1a (40%), aadA1a (29%), sat1-aadA1a (16%), dfrA17-aadA5 (11%), oxa1-aadA1a (5%) and dfrA12-aadA2 (3%). Eleven percent of the strains were resistant to nalidixic acid. Of these, 61% presented a reduced susceptibility to ciprofloxacin (MIC = 0.12-2 mg/L) and single mutations in gyrA or gyrA and parC genes, and 39%, full resistance to ciprofloxacin (MIC > or = 4 mg/L) and double and single mutations in gyrA and parC, respectively. CONCLUSION: The study gives baseline information on the magnitude of the resistance problem and its genetic background in contemporary German E. coli from food-producing animals.     

Class 1 integron-associated gene cassettes in Salmonella enterica subsp. enterica serovar Agona isolated from pig carcasses in Brazil

OBJECTIVES: Two multiresistant Salmonella enterica subsp. enterica serovar Agona isolates from pig carcasses were investigated for antimicrobial resistance genes and their location with particular reference to the detection of class 1 integrons. METHODS: The two S. Agona isolates were investigated for their in vitro susceptibility to antimicrobial agents and their plasmid content. The resistance genes and class 1 amplicons were identified by PCR assays. Amplicons of class 1 integrons were cloned and sequenced. Transferability of resistance plasmids was confirmed by conjugation. RESULTS: Both S. Agona isolates carried conjugative plasmids of approximately 150 kb which harboured all resistance genes detected in the respective isolates. S. Agona 231 was resistant to chloramphenicol by catA1, to tetracycline and minocycline by tet(B), and to sulphonamides by sul1. In addition, it harboured a streptomycin resistance gene strA and a class 1 integron with a new aadA variant designated aadA23, which mediates resistance to streptomycin and spectinomycin. S. Agona 242 also carried the genes catA1, tet(B), and sul1. Moreover, it harboured a second sulphonamide resistance gene, sul2, and a class 1 integron with intact gene cassettes carrying new variants of the trimethoprim resistance gene dfrA15b or the chloramphenicol resistance gene cmlA4. The third gene cassette consisted of a truncated aadA2 gene. CONCLUSIONS: The results of this study show that large conjugative multiresistance plasmids are present in S. Agona from pigs. Analysis of the class 1 integrons revealed the presence of new variants of resistance genes so far not detected in Salmonella isolates.     

Characterization of antimicrobial resistance and class 1 integrons found in Escherichia coli isolates from humans and animals in Korea

OBJECTIVES: Antimicrobial resistance and class 1 integrons found in Escherichia coli isolates from humans and animals in Korea were characterized. METHODS: E. coli isolates were examined for susceptibility to antimicrobial agents. Integrase genes were amplified. Gene cassette regions for classes 1 and 2 integrons were amplified and sequenced. Conjugal transfer and Southern hybridization were performed to determine the genetic localization of class 1 integrons. The clonal relationship of E. coli isolates carrying an identical cassette array was analysed by PFGE. RESULTS: Commensal E. coli isolates from animals were highly resistant to commonly used antimicrobial agents such as tetracycline, sulfamethoxazole, streptomycin, ampicillin and carbenicillin. Integrons were most prevalent in commensal E. coli isolates from poultry (44%), followed by clinical isolates from humans (33%), commensal isolates from swine (23%) and humans (13%). dfrA17-aadA5, dfrA12-orfF-aadA2 and aadA1 were found most frequently in E. coli isolates from humans, poultry and swine, respectively. Class 1 integrons were mostly located in conjugative plasmids. E. coli isolates carrying an identical cassette array were phylogenetically unrelated. CONCLUSIONS: The use of antibiotics is strongly associated with antimicrobial resistance. E. coli isolates from different sources may select a specific gene cassette by antibiotic selective pressure, which results in differences in class 1 integrons. The horizontal transfer of class 1 integrons through conjugative plasmids seems to be responsible for wide dissemination of a particular type of class 1 integron.     

A European survey of antimicrobial susceptibility among zoonotic and commensal bacteria isolated from food-producing animals

OBJECTIVE: To study antimicrobial resistance in zoonotic bacteria isolated from food animals in different countries using uniform methodology. METHODS: Samples were taken at slaughter from chickens, pigs and cattle in four EU countries per host. Escherichia coli (indicator organism; n = 2118), Salmonella spp. (n = 271) and Campylobacter spp. (n = 1325) were isolated in national laboratories and MICs tested in a central laboratory against, where appropriate, ampicillin, cefepime, cefotaxime, ciprofloxacin, chloramphenicol, erythromycin, gentamicin, nalidixic acid, streptomycin, tetracycline and trimethoprim/sulfamethoxazole. RESULTS: Isolation rates were high for E. coli, low for Salmonella and intermediate for Campylobacter. MIC results showed resistance prevalence varied among compounds, hosts and countries. For E. coli and Salmonella, resistance to newer compounds (cefepime, cefotaxime, ciprofloxacin) was absent or low, but to older compounds (except gentamicin), resistance was variable and higher. E. coli isolates from Sweden showed low resistance, whereas among isolates from Spain (pigs), resistance to ampicillin, chloramphenicol, streptomycin, tetracycline and trimethoprim/sulfamethoxazole was higher; the UK, France, the Netherlands, Germany, Italy and Denmark were intermediate. For Campylobacter spp. isolates from chickens, nalidixic acid and ciprofloxacin resistance was >30% in France and the Netherlands, >6% in the UK and zero in Sweden. Nalidixic acid resistance was high in cattle (20%-64%), whereas ciprofloxacin resistance was markedly lower in cattle, variable in pigs (3%-21%) and highest in Sweden. Generally, Campylobacter coli was more resistant than Campylobacter jejuni. CONCLUSION: Antimicrobial resistance among enteric organisms in food animals varied among countries, particularly for older antimicrobials, but resistance to newer compounds used to treat disease in humans was generally low.     

Occurrence of integrons and antimicrobial resistance genes among Salmonella enterica from Brazil

OBJECTIVES: To determine the occurrence of antimicrobial resistance genes and role of integrons among 135 antimicrobial-resistant Salmonella enterica from Brazil. METHODS: The presence of antimicrobial resistance genes, class 1 and 2 integrons and gene cassettes was analysed by PCR and sequencing. The genetic location of class 1 integrons was determined in 25 isolates by hybridization and plasmid transfer experiments. RESULTS: Fifty-five of the isolates were positive for class 1 integrons. Integron-positive isolates represented 17 different serovars and were mainly from human (n=28) and animal (n=13) sources. The gene cassette arrangements could be determined in 51 of the positive isolates, which harboured one [dfrA22, aadA1 or orf3 (putative trimethoprim resistance)], two [aadA1-dfrA1, aac(6')-Ib-orf1 (unknown function) or aacA4-aadA1], three [dfrA15b-cmlA4-aadA2, orf2 (unknown function)-dfrA5-orfD] or four [orf4-aacA4-blaOXA-30 (interrupted by an IS1 element)-aadA1] cassettes in their variable region. Only one isolate harboured a class 2 integron with the gene cassette array dfrA1-sat-aadA1. Several integron unrelated resistance genes were also detected in the isolates. Sulphonamide resistance was primarily mediated by sul2 and sul3, tetracycline resistance by tet(B) and tet(A), chloramphenicol resistance by catA1, streptomycin resistance by strA and ampicillin resistance by blaTEM. blaCTX and blaCMY-2 were found in cephalosporin-resistant isolates. Mating and hybridization experiments demonstrated that a high-molecular-weight plasmid mediated the gene transfer of integrons and additional resistance determinants. CONCLUSIONS: The present study revealed that integron-mediated resistance genes contributed to the multiresistance phenotype observed in the isolates, but most resistance genes were located outside the integron structure, as independent genes. However, they might be located on the same conjugative plasmid.     

Characterization of antimicrobial resistance and class 1 and 2 integrons in Salmonella enterica isolates from different sources in Portugal

OBJECTIVES: The antimicrobial resistance profiles of 1183 Salmonella isolates collected during 2002-2003 from several sources (human, food products and environment) were evaluated. The occurrence, distribution and cassette content of class 1 and 2 integrons among the sulphonamide-resistant population, as well as the role of particular clones to the spread of these genetic elements, were investigated. METHODS: The isolates were examined for susceptibility to antimicrobial agents. The characterization of class 1 and 2 integrons was investigated using PCR, PCR-RFLP (restriction fragment length polymorphism) and sequencing in the sulphonamide-resistant isolates. Conjugation assays and clonality analysis by PFGE were performed. RESULTS: The most common resistance phenotypes were to nalidixic acid, tetracycline, streptomycin, sulfamethoxazole and ampicillin (ranging from 31% to 17%). Resistance to sulphonamides (n=200) was associated with resistance to other antimicrobial agents, with 75% of the isolates carrying one or two class 1 integrons while only 3% simultaneously carried class 1 and 2 integrons. Integrons were observed among at least 11 serotypes (mainly Typhimurium) and in a reduced number of PFGE clones (20). Eight class 1 integron types were found, with the aadA genes (aadA1, aadA2 and aadA5) alone or downstream of a trimethoprim (dfrA1, dfrA12 and dfrA17) or a beta-lactamase resistance gene (blaoxa-30) and the blaPSE-1 gene alone. Most of the class 1 integron types were shared by several clones from the same or different serotypes obtained either from humans or food products of animal origin, especially pork products. However, some Typhimurium-specific integrons were found: aadA2 plus blaPSE-1 and blaoxa-30-aadA1. CONCLUSIONS: Apart from the hypothetical contribution of the conjugative transfer of integrons, the incidence of Salmonella carrying these genetic units seems to rely on the ability of certain clones to spread or persist in particular animal niches. Our data suggest that food-producing animals might be simultaneously considered as a reservoir of clones and integrons carrying antibiotic resistance genes, thus making the food chain, especially pork products, a possible source of multidrug-resistant isolates in humans.     

New cluster of plasmid-located class 1 integrons in Vibrio cholerae O1 and a dfrA15 cassette-containing integron in Vibrio parahaemolyticus isolated in Angola

The resistance profile and its correlation with mobile genetic elements were investigated in 11 Vibrio cholerae O1 and 2 Vibrio parahaemolyticus clinical isolates, as well as in 1 V. cholerae O1 and 1 V. cholerae non-O1 environmental isolate, isolated between 1991 and 1996 in different provinces of Angola. All clinical isolates of V. cholerae O1 were resistant to ampicillin, chloramphenicol, trimethoprim, sulfamethoxazole, and tetracycline. They also contained a large conjugative plasmid (p3iANG) with a set of three class 1 integrons harboring dfrA15, blaP1, and qacH-aadA8 cassettes, which code for resistance to trimethoprim, beta-lactams, quaternary ammonium compounds, and aminoglycosides, clustered in a 19-kb region. Chloramphenicol (cat1), kanamycin (aph), sulfonamide (sul2), and tetracycline (tetG) resistance genes were also carried on the plasmid within the same 19-kb region. A chromosomal integron containing the dfrA15 cassette was also revealed in V. parahaemolyticus strains. SXT integrase genes were present in six V. cholerae isolates but apparently were not associated with known SXT-associated resistance genes. This study indicates that plasmids and integrons contributed mainly to the circulation of multiple-drug resistance determinants in Vibrio strains from Angola.     

Antimicrobial susceptibility and genetic relatedness of Salmonella serovars isolated from animal-derived dog treats in the USA

OBJECTIVES: The objectives of this study were to determine the potential risk of dog treats in transmitting Salmonella to humans in the USA, and to characterize genetic relatedness and antimicrobial resistance among the isolates. METHODS: A total of 158 dog treats derived from pig ears and other animal parts were randomly collected nationwide and assayed for the presence of Salmonella. The Salmonella isolates were characterized using serotyping, pulsed-field gel electrophoresis (PFGE) and antimicrobial susceptibility testing. RESULTS: Forty-one percent (65/158) of samples were positive for Salmonella. Eighty-four Salmonella isolates, comprising 24 serotypes, were recovered from the 65 positive samples. Fourteen samples were contaminated with more than one Salmonella serotype. PFGE analysis of 78 Salmonella isolates yielded 64 patterns. S. Infantis with PFGE patterns indistinguishable from those of strains identified in Canadian outbreaks in 1999 were recovered in several dog treat products. The majority of Salmonella isolates were susceptible to the antimicrobials tested; however, resistance was observed to tetracycline (26%), streptomycin (23%), sulfamethoxazole (19%), chloramphenicol (8%) and ampicillin (8%). Twenty-eight (36%) Salmonella isolates were resistant to at least one antimicrobial and 10 (13%) isolates displayed resistance to four or more antimicrobials. Two isolates were identified as S. Typhimurium DT104 with the characteristic penta-resistance phenotype (ampicillin, chloramphenicol, streptomycin, sulfamethoxazole and tetracycline). One S. Brandenburg isolate was resistant to eight antimicrobials. Seven Salmonella isolates also contained class I integrons encoding resistance genes to aminoglycosides, beta-lactam and streptothricin antimicrobials. CONCLUSIONS: The study indicates that animal-derived dog treats in the USA could be a potential source of animal and human infections with Salmonella, including multidrug-resistant Salmonella strains.     

Genetic relatedness of a rarely isolated Salmonella: Salmonella enterica serotype Niakhar from NARMS animal isolates

BACKGROUND: In the United States, Salmonella enterica serotype Niakhar is infrequently isolated. Between 1997 and 2000, the animal arm of the National Antimicrobial Resistance Monitoring System-Enteric Bacteria (NARMS) assayed a total of 22,383 Salmonella isolates from various animal sources (swine, cattle, chickens, turkeys, cats, horses, exotics and dogs) for antimicrobial susceptibility. Isolates originated from diagnostic and non-diagnostic submissions. OBJECTIVES: To study the phenotypic and genotypic characteristics of Salmonella Niakhar. METHODS AND RESULTS: Only five (0.02%) of the 22,383 isolates were identified as Salmonella Niakhar. Antimicrobial resistance testing indicated that three isolates were pan-susceptible, one isolate was resistant to ampicillin and one isolate was resistant to ampicillin, chloramphenicol, ciprofloxacin, kanamycin, nalidixic acid, streptomycin, sulfamethoxazole, tetracycline and trimethoprim/sulfamethoxazole. RAPD-PCR analysis, PFGE and ribotyping indicated that two pan-susceptible isolates were genetically similar, whereas the three remaining isolates were genetically different. The one Salmonella Niakhar isolate that was multiresistant harboured a class I integron, intI1 and two large plasmids. CONCLUSIONS: This study represents the first report of a ciprofloxacin-resistant Salmonella isolate from the animal arm of NARMS.     

Molecular basis of resistance to trimethoprim, chloramphenicol and sulphonamides in Bordetella bronchiseptica

OBJECTIVES: To date, little is known about the molecular basis of antimicrobial resistance in Bordetella bronchiseptica, an important respiratory tract pathogen in pigs, dogs and cats. The aim of this study was to identify genes coding for trimethoprim resistance present in porcine B. bronchiseptica and to determine their localization, transferability and association with other resistance genes. METHODS: Six B. bronchiseptica isolates with elevated MICs of trimethoprim were investigated by PCR for the presence of trimethoprim resistance genes and their association with class 1 integrons. The amplicons obtained were cloned and sequenced. Plasmid localization of these integrons was confirmed by transformation and conjugation. Isolates carrying the same integron were compared for their genetic relatedness by XbaI and SpeI pulsed-field gel electrophoresis (PFGE). RESULTS: Five B. bronchiseptica isolates carried a class 1 integron with two gene cassettes, one carrying the trimethoprim resistance gene dfrA1 and the other the chloramphenicol resistance gene catB3. This integron was present on a common conjugative plasmid in four of the five isolates and on the chromosome in the remaining isolate. All five B. bronchiseptica isolates proved to be related on the basis of their PFGE patterns. Another isolate had a class 1 integron with a dfrB1 and a catB2 cassette on a structurally different conjugative plasmid. The sulphonamide resistance gene sul1 was detected in the 3'-conserved segment of both types of integrons. CONCLUSIONS: This is the first report of trimethoprim, chloramphenicol and sulphonamide resistance genes and class 1 integrons in B. bronchiseptica isolates.     

Comparative in vitro activities of selected antimicrobial agents against Edwardsiella tarda.

MICs of 14 antimicrobial agents for 29 strains of Edwardsiella tarda were determined by an agar dilution method. Of the agents tested, ciprofloxacin, enoxacin, and norfloxacin were the most active on a weight basis. All strains were also susceptible to clinically achievable concentrations of ampicillin, chloramphenicol, tetracycline, trimethoprim, sulfamethoxazole, trimethoprim plus sulfamethoxazole, cefotaxime, and gentamicin. Ninety percent of the strains demonstrated high-level resistance to polymyxin B and colistin.     

儿童肠球菌多重耐药与Ⅰ类整合子的检测

目的了解儿童临床分离肠球菌的耐药特征及其多重耐药与Ⅰ类整合子的相互关系。方法采用琼脂稀释法测定常用抗菌药物对152株肠球菌的最低抑菌浓度（MIC）,用聚合酶链反应（PCR）检测肠球菌Ⅰ类整合子和Ⅰ类整合酶基因。结果屎肠球菌对氨苄西林、阿莫西林-克拉维酸、环丙沙星的耐药率分别为96．8％、95．2％和84．1％,粪肠球菌对上述3种抗菌药物的耐药率分别为23．6％、18．0％和49．4％,屎肠球菌的耐药率明显高于粪肠球菌（P〈0．001）;粪肠球菌中有2株对万古霉素的MIC为8μg/mL,粪肠球菌和屎肠球菌对替考拉宁均敏感。儿童多重耐药肠球菌发生率高达93．7％。屎肠球菌耐药模式以耐氨苄西林、红霉素、环丙沙星、利福平、四环素、高水平庆大霉素6种抗菌药物为主,占屎肠球菌的59％,粪肠球菌以耐四环素、红霉素、利福平、氯霉素、高水平庆大霉素5种抗菌药物为主,占粪肠球菌的26％。全部152株肠球菌未检测到Ⅰ类整合子,仅有5株检测到Ⅰ类整合酶基因。结论儿童肠球菌多重耐药十分严重,儿童肠球菌多重耐药与Ⅰ类整合子和Ⅰ类整合酶基因尚无明显关系。     

Molecular characteristics of class 1 and class 2 integrons and their relationships to antibiotic resistance in clinical isolates of Shigella sonnei and Shigella flexneri

OBJECTIVES: To analyse the gene cassettes and determine the roles of class 1 and class 2 integrons in antibiotic-resistant strains of Shigella sonnei (n=31) and Shigella flexneri (n=33). METHODS: Various molecular techniques, including PCR and Southern-blotting analysis, were used to analyse various markers of class 1 and class 2 integrons in these 64 S. sonnei and S. flexneri isolates collected in Hangzhou, China. The gene cassette arrays in integrons were identified by DNA sequencing and/or restriction fragment length polymorphism. Two genomic DNA fragments, one containing intI1 from a S. flexneri isolate that contains intI1 but lacks 3'-conserved region and another containing intI2 from a S. sonnei isolate, were cloned into pUC19 vectors and sequenced. The links between integron gene cassette arrays and antibiotic resistance were analysed. RESULTS: Class 2 integrons were present in 80.6% (25/31) of the S. sonnei isolates and 87.9% (29/33) of the S. flexneri isolates. All of these integron 2-positive isolates contained constant gene cassette arrays of dfrA1+sat1+aadA1 which confer resistance to trimethoprim and streptomycin. It was demonstrated that the class 2 integron was located in the Tn7 region inside the attTn7 locus downstream of glmS in Shigella. Class 1 integrons were found in 9.4% (6/64) of Shigella spp. isolates. An atypical class 1 integron without a 3'-conserved segment on the Shigella chromosome, termed Shigella atypical class 1 integron (SAI), was present in 84.9% (28/33) of S. flexneri isolates. The SAI contained two gene cassettes, bla(OXA30) and aadA1; however, the SAI conferred resistance to ampicillin, but not to streptomycin, in Escherichia coli host. The bla(OXA30) and aadA1 cassettes of the SAI seemed to be always coordinately excised or integrated. CONCLUSIONS: Multiple and complex mechanisms involving mobile genetic elements in class 1 and class 2 integrons and antibiotic resistance have been developed in the evolution of Shigella strains.     

Antibiotic resistance genes, integrons and multiple antibiotic resistance in thirty-five serotypes of Salmonella enterica isolated from humans and animals in the UK

OBJECTIVES: To examine 397 strains of Salmonella enterica of human and animal origin comprising 35 serotypes for the presence of aadB, aphAI-IAB, aadA1, aadA2, bla(Carb(2)) or pse1, bla(Tem), cat1, cat2, dhfr1, floR, strA, sul1, sul2, tetA(A), tetA(B) and tetA(G) genes, the presence of class 1 integrons and the relationship of resistance genes to integrons and antibiotic resistance. RESULTS: Some strains were resistant to ampicillin (91), chloramphenicol (85), gentamicin (2), kanamycin (14), spectinomycin (81), streptomycin (119), sulfadiazine (127), tetracycline (108) and trimethoprim (45); 219 strains were susceptible to all antibiotics. bla(Carb(2)), floR and tetA(G) genes were found in S. Typhimurium isolates and one strain of S. Emek only. Class 1 integrons were found in S. Emek, Haifa, Heidelberg, Mbandaka, Newport, Ohio, Stanley, Virchow and in Typhimurium, mainly phage types DT104 and U302. These strains were generally multi-resistant to up to seven antibiotics. Resistance to between three and six antibiotics was also associated with class 1 integron-negative strains of S. Binza, Dublin, Enteritidis, Hadar, Manhattan, Mbandaka, Montevideo, Newport, Typhimurium DT193 and Virchow. CONCLUSION: The results illustrate specificity of some resistance genes to S. Typhimurium or non- S. Typhimurium serotypes and the involvement of both class 1 integron and non-class 1 integron associated multi-resistance in several serotypes. These data also indicate that the bla(Carb(2)), floR and tetA(G) genes reported in the SG1 region of S. Typhimurium DT104, U302 and some other serotypes are still predominantly limited to S. Typhimurium strains.