Identification of Antibiotic Resistance Cassettes in Class 1 Integrons in Aeromonas spp. Strains Isolated From Fresh Fish (Cyprinus carpio L.)

Forty-six Aeromonas spp. strains were isolated from fresh fish and investigated for their antimicrobial susceptibility, detection of Class 1 integrons by PCR, and arrangement of gene cassettes. Selected isolates were further characterized by enterobacterial repetitive intergenic consensus-PCR. Twenty isolates were found to carry Class 1 integrons. Amplification of the variable regions of the integrons revealed diverse bands ranging in size from 150 to 1,958 pb. Sequence analysis of the variable regions revealed the presence of several gene cassettes, such as adenylyl transferases (aadA2 and aadA5), dihydrofolate reductases (dfrA17 and dfrA1), chloramphenicol acetyl transferase (catB3), β-lactamase (oxa2), lincosamide nucleotidil transferase (linF), aminoglycoside-modifying enzyme (apha15), and oxacillinase (bla _OXA-10). Two open reading frames with an unknown function were identified as orfC and orfD. The aadA2 cassette was the most common integron found in this study. Interestingly, five integrons were detected in the plasmids that might be involved in the transfer of resistance genes to other bacteria. This is a first report of cassette encoding for lincosamides (linF) resistance in Aeromonas spp. Implications on the incidence of integrons in isolates of Aeromonas spp. from fresh fish for human consumption, and its possible consequences to human health are discussed.     

Identification and Characterization of Integron-Mediated Antibiotic Resistance in the Phytopathogen Xanthomonas oryzae pv. oryzae

Four streptomycin-resistant isolates of Xanthomonas oryzae pv. oryzae (YNA7-1, YNA10-2, YNA11-2, and YNA12-2) were examined via PCR amplification for the presence of class 1, class 2, and class 3 integrons and aadA1 and aadA2 genes, which confer resistance to streptomycin and spectinomycin. The class 1 integrase gene intI1 and the aminoglycoside adenylyltransferase gene aadA1 were identified in all four resistant isolates but not in 25 sensitive isolates. PCR amplifications showed that 7790-bp, 7162-bp, 7790-bp, and 7240-bp resistance integrons with transposition gene modules (tni module) in 3′ conserved segments existed in YNA7-1, YNA10-2, YNA11-2, and YNA12-2, respectively. Subsequent analysis of sequences indicated that the integrons of YNA7-1 and YNA11-2 carried three gene cassettes in the order |aacA3|arr3|aadA1|. The integron of YNA10-2 carried only |arr3|aadA1| gene cassettes. The integron of YNA12-2 lacked a 550-bp sequence including part of intI1 but it still carried |aacA3|arr3|aadA1| gene cassettes. The analysis of inactive mutants and complementation tests confirmed that the aacA3 gene conferred resistance to tobramycin, kanamycin, gentamicin and netilmicin; the arr3 gene conferred resistance to rifampicin; and the aadA1 gene conferred resistance to streptomycin and spectinomycin. The resistance phenotypes of the four isolates corresponded with their resistance gene cassettes, except that YNA7-1 and YNA12-2 did not show rifampicin resistance. Sequence comparison revealed that no gene cassette array in GenBank was in the same order as in the integrons of the four resistant isolates in this study and the aadA1, which was identical in the four resistant isolates, showed 99% identity with aadA1 sequences in GenBank. The result of a stability test showed that the resistance phenotype, the aadA1 gene, and the intI1 gene were completely stable in YNA7-1 and YNA12-2 but unstable in YNA10-2 and YNA11-2. To our knowledge, this is the first report of resistance integron in a phytopathogenic bacteria.     

Antibiotic Resistance in Food-Borne Bacterial Contaminants in Vietnam

This study was conducted to examine the rate of contamination and the molecular characteristics of enteric bacteria isolated from a selection of food sources in Vietnam. One hundred eighty raw food samples were tested; 60.8% of meat samples and 18.0% of shellfish samples were contaminated with Salmonella spp., and more than 90% of all food sources contained Escherichia coli. The isolates were screened for antibiotic resistance against 15 antibiotics, and 50.5% of Salmonella isolates and 83.8% of E. coli isolates were resistant to at least one antibiotic. Isolates were examined for the presence of mobile genetic elements conferring antibiotic resistance. Fifty-seven percent of E. coli and 13% of Salmonella isolates were found to contain integrons, and some isolates contained two integrons. Sequencing results revealed that the integrons harbored various gene cassettes, including aadA1, aadA2, and aadA5 (resistance to streptomycin and spectinomycin), aacA4 (resistance to aminoglycosides), the dihydrofolate reductase gene cassettes dhfrXII, dfrA1, and dhfrA17 (trimethoprim resistance), the beta-lactamase gene bla_PSE1 (ampicillin resistance), and catB3 (chloramphenicol resistance). Plasmids were also detected in all 23 antibiotic-resistant Salmonella isolates and in 33 E. coli isolates. Thirty-five percent of the Salmonella isolates and 76% of the E. coli isolates contained plasmids of more than 95 kb, and some of the isolates contained two large plasmids. Conjugation experiments showed the successful transfer of all or part of the antibiotic resistance phenotypes among the Salmonella and E. coli food isolates. Our results show that enteric bacteria in raw food samples from Vietnam contain a pool of mobile genetic elements and that the transfer of antibiotic resistance can readily occur between similar bacteria.     

Novel Integron Gene Cassette Arrays Identified in a Global Collection of Multi-Drug Resistant Non-Typhoidal Salmonella enterica

Investigation of integron carriage in a global collection of multi-drug resistant Salmonella enterica identified 3 unique class 1 integron gene cassette arrays not previously reported in this species. The present study used PCR and DNA sequence analysis to characterize the structure of these gene cassette arrays. A ~4.0 kb integron containing the gene cassette array arr2/cmlA5/bla _OXA10 /aadA1 was found in isolates belonging to serovars Isangi and Typhimurium from South Africa. A ~6.0 kb integron containing the gene cassettes aac(6′)IIc/ereA2/IS1247/aac/arr/ereA2 was found in isolates belonging to serovar Heidelberg from the Philippines. In this gene cassette array, the insertion sequence, IS1247, and two putative resistance genes, disrupt the erythromycin resistance gene cassette. Finally, a ~6.0 kb integron containing the gene cassette qacH/dfrA32/ereA1/aadA2/cmlA/aadA1 was found in serovar Stanley isolates from Taiwan. This integron, which has not been previously reported in any bacterial species, contains a new dihydrofolate reductase gene cassette sequence designated dfrA32, with only 90% sequence similarity to previously reported dfrA cassettes. The S. enterica integrons described in the present study represent novel collections of resistance genes which confer multi-drug resistance and have the potential to be widely disseminated among S. enterica as well as other bacterial species.     

Achromobacter xylosoxidans: An Emerging Pathogen Carrying Different Elements Involved in Horizontal Genetic Transfer

In the last few years, numerous cases of multidrug-resistant Achromobacter xylosoxidans infections have been documented in immunocompromised and cystic fibrosis patients. To gain insights into the molecular mechanisms and mobile elements related to multidrug resistance in this bacterium, we studied 24 non-epidemiological A. xylosoxidans clinical isolates from Argentina. Specific primers for plasmids, transposons, insertion sequences, bla _ampC, intI1, and intI2 genes were used in PCR reactions. The obtained results showed the presence of wide host range IncP plasmids in ten isolates and a high dispersion of class 1 integrons (n?=?10) and class 2 integrons (n?=?3). Four arrays in the variable region (vr) of class 1 integrons were identified carrying different gene cassettes as the aminoglycoside resistance aac(6′)-Ib and aadA1, the trimethoprim resistance dfrA1 and dfrA16, and the β-lactamase bla _OXA-2. In only one of the class 2 integrons, a vr was amplified that includes sat2-aadA1. The bla _ampC gene was found in all isolates, confirming its ubiquitous nature. Our results show that A. xylosoxidans clinical isolates contain a rich variety of genetic elements commonly associated with resistance genes and their dissemination. This supports the hypothesis that A. xylosoxidans is becoming a reservoir of horizontal genetic transfer elements commonly involved in spreading antibiotic resistance.     

Characterization of antimicrobial resistance of Pseudomonas aeruginosa isolated from canine infections

Aims: To determine the prevalence of Pseudomonas aeruginosa among dogs with suspected soft tissue infections and to characterize these isolates. Methods and Results: Swabs were taken from infected soft tissues of 402 dogs. Pseudomonas aeruginosa strains were confirmed phenotypically and tested for susceptibility to 11 antimicrobial agents and genotyped by SpeI pulsed‐field gel electrophoresis (PFGE). The genetic basis of fluoroquinolone (FQ) resistance and the presence of integrons were also characterized. A total of 27 (6·7%) dogs tested positive for Ps. aeruginosa. Fourteen different SpeI patterns were observed in 25 typeable strains. Among the β‐lactams, three isolates presented resistance to ticarcillin and carbenicillin, while only one isolate exhibited resistance to ceftazidime. Among the aminoglycosides (AGs), three strains showed resistance to amikacin, and four strains exhibited resistance to gentamicin and tobramycin. Four strains with mutations that led to the substitution of Thr at position 83 with Ile in GyrA and the exchange of Ser at position 87 with Leu in ParC displayed resistance to all tested FQs. These strains also carried class 1 integrons and showed resistance to between 6 and 10 antimicrobials. These integrons included four different gene cassettes (aacA4‐aadA1, bla_OXA‐31‐aadA2, aadA1‐arr‐3‐catB3 and cmlA5‐cmlA‐aadA1). Conclusions: A small proportion of infected dogs treated in two animal hospitals in Beijing, China carried Ps. aeruginosa isolates. Low levels of resistance to anti‐pseudomonal agents were observed in these strains. Significance and Impact of the Study: This study is the first report on the antimicrobial resistance profiles of Ps. aeruginosa isolated from infected canine origin in China. Additionally, this is the first report of the oxacillin resistance gene bla_OXA‐31 in a canine Ps. aeruginosa isolate.     

Prevalence and antimicrobial resistance in Salmonella enterica isolated from broiler chickens,pigs and meat products in Thailand–Cambodia border provinces

This study aimed to examine the prevalence and antimicrobial resistance (AMR) of Salmonella isolates from broiler chickens, pigs and their associated meat products in the Thailand–Cambodia border provinces. A total of 941 samples were collected from pigs and broiler chickens at slaughter houses and from carcasses at local fresh markets in Sa Kaeo, Thailand (n = 554) and Banteay Meanchey, Cambodia (n = 387) in 2014 and 2015. From these samples, 345 Salmonella isolates were collected from Sa Keao (n = 145; 23%) and Banteay Meanchey (n = 200; 47%) and assayed for antimicrobial susceptibility, class 1 integrons and extended‐spectrum β‐lactamase (ESBL) genes. Serovars Typhimurium (29%) and Rissen (29%) were the most common serotypes found in Thai and Cambodian isolates, respectively. Multidrug resistance was detected in 34% and 52% of isolates from Sa Keao and Banteay Meanchey, respectively. The majority of the Thai isolates were resistant to ampicillin (72.4%), whereas most Cambodian isolates were resistant to sulfamethoxazole (71%). Eleven isolates from Sa Keao and 44 from Banteay Meanchey carried class 1 integrons comprising resistance gene cassettes. The most common gene cassette array was dfrA12‐aadA2 (61.1%). Six isolates were ESBL producers. The β‐lactamase genes found included bla_TEM‐1, bla_CTX‐M‐55 and bla_CMY‐2. Some of these class 1 integrons and ESBL genes were located on conjugative plasmid. In conclusion, multidrug‐resistant Salmonella are common in pigs, chickens and their products in the Thailand–Cambodia border provinces. Our findings indicate that class 1 integrons play a role in spread of AMR in the strains in this study.     

Occurrence and molecular characteristics of antimicrobial resistance of Escherichia coli from broilers,pigs and meat products in Thailand and Cambodia provinces

Nine hundred and forty‐one samples were collected in Sa Keao, Thailand (n = 554) and Banteay Meanchey, Cambodia (n = 387) from July 2014 to January 2015. A total of 667 Escherichia coli isolates (381 isolates from Sa Keao and 286 isolates from Banteay Meanchey) were obtained and examined for antimicrobial susceptibility, class 1 integrons, ESBL genes and horizontal transfer of resistance determinants. Prevalence of E. coli in pig and broiler carcass samples from slaughterhouses and fresh markets was 36–85% in Sa Keao and 11–69% in Banteay Meanchey. The majority of these isolates were multidrug resistant (75.3%). Class 1 integrons were common in both Thai (47%) and Cambodian (62%) isolates, of which four resistance gene cassette arrays including aadA1, dfrA1‐aadA1, dfrA12‐aadA2 and aadA2‐linF were identified. Class 1 integrons in two broiler isolates from Sa Keao (dfrA12‐aadA2) and one broiler isolate from Banteay Meanchey (dfrA1‐aadA1) were horizontally transferable. Sixteen isolates were confirmed to be ESBL‐producing strains with ESBL gene bla_CTX‐M‐15, broad spectrum β‐lactamase gene bla_TEM‐1 and the AmpC gene bla_CMY‐2 being detected. The bla_TEM‐1 gene was most prevalent and located on a conjugative plasmid.     

Class 1 Integrons and Antibiotic Resistance of Clinical Acinetobacter calcoaceticus–baumannii Complex in Poznań, Poland

Sixty-three clinical isolates of Acinetobacter calcoaceticus–baumannii complex were analyzed for the presence of integrons and antimicrobial resistance. Class 1 integrons were detected in 40 (63.5 %) isolates. None of them had class 2 or class 3 integrons. The majority of the integrons contained aacC1–orfA–orfB–aadA1 gene cassette array. The presence of integrons was associated with the increased frequency of resistance to 12 of 15 antimicrobials tested, multi-drug resistance phenotype, and the overall resistance ranges of the strains.     

Occurrence and transmission of class 1 and 2 integrons among phenotypic highly ampicillin-resistant avian <Emphasis Type="Italic">Escherichia coli</Emphasis> isolates from Pakistan

Thirty four avian Escherichia coli isolates were collected from different cities of Punjab province, Pakistan during 2008–2009. Twenty five phenotypic highly ampicillin-resistant (MICs ≥ 256 μg/ml) avian E. coli strains were selected for the investigation of occurrence and transmission of class 1, 2 and 3 integrons and β-lactamase genes. Amoxicillin, sulfonamide, trimethoprim, enrofloxacin, pefloxacin and tetracycline were the most common phenotypic resistant among ampicillin-resistant avian E. coli strains. Integrons and β-lactamase were found 60 and 72% respectively. Class 1 and 2 integrons were found 52 and 8%, while class 3 integrons were not found in all strains. All class 1 positive strains had variable fragments associated with gene cassettes dfrA7, dfrA1-aadA1, aadA1, aadA22 and dfrA12-orfF-aadA2 respectively, which confer resistance to trimethoprim and streptomycin. Class 2-positive strains had similar gene cassettes array dfrA1-sat1-aadA1 conferring resistance to trimethoprim, streptothricin and spectinomicin/streptomycin. Integrons are frequently found in β-lactamase positive isolates and widely disseminate multidrug resistance genes but they do not play role in the spreading of β-lactamase genes. Class 1 integrons gene cassette aadA22 is reported for the first time in avian E. coli. Findings of this study may provide important and useful information reflecting specific antibiotic selective pressure in Punjab province, Pakistan.     

Class 1 integrons in Pseudomonas aeruginosa isolates from clinical settings in Amazon region, Brazil

A hundred and six Pseudomonas aeruginosa isolates from clinical cases were screened using PCR for the presence of integrons and associated resistance gene cassettes. Forty-four isolates harboured class 1 integrons (41.5%), of which 29 isolates (66%) also carried gene cassettes. The aacA gene was most frequently found within class 1 integrons (69%), followed by blaOXA family genes (52%). From class 1 integron-positive strains, we detected a total of 15 isolates (34%) carrying no gene cassettes. Restriction fragment-length polymorphism analysis of the integrons variable region revealed some identical structures, as well as distinct profiles indicating heterogeneity among these cassette regions. Multiresistance was observed in 71% of isolates, nevertheless no strong correlation was observed between integron presence and multiresistance. This is the first report showing class 1 integron prevalence and gene cassette content in P. aeruginosa isolates from clinical settings in the Brazilian Amazon.     

Diverse Gene Cassettes in Class 1 Integrons of Facultative Oligotrophic Bacteria of River Mahananda,West Bengal,India

Background

In this study a large random collection (n = 2188) of facultative oligotrophic bacteria, from 90 water samples gathered in three consecutive years (2007–2009) from three different sampling sites of River Mahananda in Siliguri, West Bengal, India, were investigated for the presence of class 1 integrons and sequences of the amplification products.

Methodology/Principal Findings

Replica plating method was employed for determining the antibiotic resistance profile of the randomly assorted facultative oligotrophic isolates. Genomic DNA from each isolate was analyzed by PCR for the presence of class 1 integron. Amplicons were cloned and sequenced. Numerical taxonomy and 16S rRNA gene sequence analyses were done to ascertain putative genera of the class 1 integron bearing isolates. Out of 2188 isolates, 1667 (76.19%) were antibiotic-resistant comprising of both single-antibiotic resistance (SAR) and multiple-antibiotic resistant (MAR), and 521 (23.81%) were sensitive to all twelve different antibiotics used in this study. Ninety out of 2188 isolates produced amplicon(s) of varying sizes from 0.15 to 3.45 KB. Chi-square (χ²) test revealed that the possession of class 1 integron in sensitive, SAR and MAR is not equally probable at the 1% level of significance. Diverse antibiotic-resistance gene cassettes, aadA1, aadA2, aadA4, aadA5, dfrA1, dfrA5, dfrA7, dfrA12, dfrA16, dfrA17, dfrA28, dfrA30, dfr-IIe, blaIMP-9, aacA4, Ac-6′-Ib, oxa1, oxa10 and arr2 were detected in 64 isolates. The novel cassettes encoding proteins unrelated to any known antibiotic resistance gene function were identified in 26 isolates. Antibiotic-sensitive isolates have a greater propensity to carry gene cassettes unrelated to known antibiotic-resistance genes. The integron-positive isolates under the class Betaproteobacteria comprised of only two genera, Comamonas and Acidovorax of family Comamonadaceae, while isolates under class Gammaproteobacteria fell under the families, Moraxellaceae, Pseudomonadaceae, Aeromonadaceae and Enterobacteriaceae.

Conclusions

Oligotrophic bacteria are good sources of novel genes as well as potential reservoirs of antibiotic resistance gene casettes.     

Class 1 and class 2 integrons and plasmid-mediated antibiotic resistance in coliforms isolated from ten rivers in northern Turkey

We aimed to determine the molecular mechanisms of antibiotic resistance in coliforms isolated from ten rivers in northern region of Turkey. A total of 183 isolates were tested for antimicrobial susceptibility by disk diffusion and agar dilution methods. Resistance to ampicillin, streptomycin, trimethoprim, tetracycline, and chloramphenicol was detected in 58%, 51.9%, 24%, 28.4%, and 12.5%, respectively. Twelve (6.5%) phylogenetically distant organisms were detected to harbor self-transmissible plasmids ranging 52 to >147 kb in sizes. Resistances to ampicillin, tetracycline, trimethoprim, streptomycin, and nalidixic acid were commonly transferable traits. Transferable nalidixic acid-resistant strains harbored qnrS gene, which was the first report of plasmid-mediated quinolone resistance in bacteria of environmental origin in Turkey. Fourteen and five coliforms harbored class 1 and class 2 integrons, respectively, and some of them were located on transferable plasmids. Sequence analyses of variable regions of the class 1 and 2 integrons harbored various gene cassettes, dfrA1, dfr2d, dfrA7, dfrA16, dfrA17, aadA1, aadA5, bla _oxA-30, and sat1. A gene cassette array, dfrA16 has been demonstrated for the first time in a Citrobacter koseri isolate. Class 1 and class 2-bearing strains were clustered in different groups by BOX-PCR fingerprinting. Rivers in the northern Turkey may act as receptacle for the multi-drug resistant enterobacteria and can serve as reservoirs of the antimicrobial resistance determinants in the environment. The actual risk to public health is the transfer of resistance genes from the environmental bacteria to human pathogens. This study was presented in part at the 2^nd World Conference on Magic Bullets, held October 3–5, 2008 in Nurnberg, Germany.     

Characterization and Chromosomal Mapping of Antimicrobial Resistance Genes in Salmonella enterica Serotype Typhimurium

Two hundred and twenty-six Salmonella enterica serotype Typhimurium isolates were examined for the presence of integron-associated gene cassettes. All but two of the non-DT104 isolates, together with DT104 isolates, contained gene cassettes. Amplicons of 1.5 kbp each were found in two non-DT104 isolates, encoding a dhfrI gene (trimethoprim resistance) linked to an aadA gene (streptomycin and spectinomycin resistance), by site-specific recombination. DT104 isolates of resistance (R) type ACSSuT possessed the recently described 1.0- and 1.2-kbp gene cassettes. Macrorestriction analysis with XbaI and DNA probing mapped ant(3")-1a, bla_PSE-1, and dhfrI genes to large multiresistant gene clusters in a DT170a isolate and a DT193 isolate. In contrast, all DT104 isolates (R-type ACSSuT) possessed a conserved 10-kbp Xba1 DNA fragment. Our study highlights the occurrence of integrons (and antimicrobial resistance determinants) among serotype Typhimurium isolates other than DT104. Larger and previously unrecognized multiresistance gene clusters were identified in these isolates by DNA probing.     

Genetic determinants of antibacterial resistance among nosocomial Escherichia coli, Klebsiella spp., and Enterobacter spp. isolates collected in Russia within 2003-2007

Nosocomial bacterial isolates collected within 2003-2004 (n=411) and 2005-2007 (n=422) were highly resistant to cephalosporins III-IV and antibacterials of other groups (aminoglycosides, fluoroquinolons, chloramphenicol, and co-trimoxazole). Genes encoding TEM, SHV, CTX-M, OXA-2, and AmpC types of beta-lactamases (BLs) in the E. coli, Klebsiella spp., and Enterobacter spp. isolates were detected using polymerase chain reaction (PCR). Prevalent CTX-M-type BLs were detected in 85% of the E. coli, 87% of the Klebsiella spp., and 38% of the Enterobacter spp. isolates of the first strain collection and in 94% of the E. coli, 91% of the Klebsiella spp., and 38% of the Enterobacter spp. isolates of the second one. Genes belonging to three subtypes of blacTx-M genes were identified: bla(CTX-M-1) (228 bla(CTX-M-15) and six bla(CTX-M-3) of the first strain collection; 275 bla(CTX-M-15), three bla(CTX-M-3), and one bla(CTX-M-22) of the second one), bla(CTX-M-2) (one bla(CTX-M-5) of the first strain collection and one bla(CTX-M-2) of the second one), bla(CTX-M-9) (17 bla(CTX-M-14) and one bla(CTX-M-9) of the first strain collection; seven bla(CTX-M-14) and one bla(CTX-M-9) of the second one). Three isolates of the first strain collection and one isolate of the second one carried two genes belonging to two different subtypes, i.e., bla(CTX-M-15) and bla(CTX-M-14) simultaneously. The bacterial isolates had high levels of associative resistance to ciprofloxacin, co-trimoxazole, gentamicin, amikacin, and chloramphenicol associated with the resistance gene cassettes aadA1, aadA2, aadA5, aadB, aacA4, aac(6')Ib; dfrA1, dfrA5, dfrA12, dfrA17, cmlA1, ereA2, and catB8 in the class 1 integrons and the resistance gene cassettes dfrA1, sat1, and aadA1 in the class 2 integrons.     

Molecular characterization of antimicrobial resistance in Salmonella isolated from animals in Japan

Aims:  To investigate the prevalence of integrons and antimicrobial resistance genes in Salmonella recovered from animals in Japan.
Methods and Results:  Forty-eight out of ninety-four (51·1%) Salmonella isolates showed multidrug resistance phenotypes and harboured at least one antimicrobial resistance gene. Twenty-two out of forty-seven (46·8%) Salmonella enterica serovar Typhimurium that were multidrug-resistant were of definitive phage type DT104. Class 1 integrons were identified in 34/94 isolates (36·2%): 21 isolates containing two gene cassettes, aadA2 and bla _PSE–1, and 13 containing one gene cassette, aadA1 , aadA2 or bla _PSE–1. Class 2 integrons containing estX - sat2 - aadA1 gene cassettes were only identified in Salmonella Enteritidis. The β-lactamase-encoding gene, bla _TEM, was only detected in S. Typhimurium. The plasmid-mediated quinolone resistance gene, qnrS1 , was identified in S. Typhimurium and Salmonella Thompson.
Conclusions:  Our results characterized integrons and antimicrobial resistance genes in Salmonella of animal origin. To the best of our knowledge, this is the first report of qnrS in Salmonella from Japan and also the first report of qnrS in S . Thompson.
Significance and Impact of the Study:  Little is known about the molecular basis of antimicrobial resistance in Salmonella isolated from animals. This study provides useful data on the incidence of integrons and resistance genes in Salmonella of animal origin.     

pIMP-PH114 Carrying bla IMP-4 in a Klebsiella pneumoniae Strain is Closely Related to Other Multidrug-Resistant IncA/C2 Plasmids

The IncA/C plasmids are broad host-range vehicles which have been associated with wide dissemination of CMY-2 among Enterobacteriaceae of human and animal origins. Acquired metallo-β-lactamases (MBLs) such as the IMP-type enzymes are increasingly reported in multidrug-resistant Gram-negative bacteria worldwide, particularly in Enterobacteriaceae. We described the complete sequence of the first IMP-4-encoding IncA/C₂ plasmid, pIMP-PH114 (151,885 bp), from a sequence type 1 Klebsiella pneumoniae strain that was recovered from a patient who was hospitalized in the Philippines. pIMP-PH114 consists of a backbone from the IncA/C₂ plasmids, with the insertion of a novel Tn21-like class 1 integron composite structure (containing the cassette array bla _IMP-4-qacG-aacA4-catB3, followed by a class C β-lactamase bla _DHA-1 and the mercury resistance operon, merRTPCADE) and a sul2-floR encoding region. Phylogenetic analysis of the IncA/C repA sequences showed that pIMP-PH114 formed a subgroup with other IncA/C plasmids involved in the international spread of CMY-2, TEM-24 and NDM-1. Identical bla _IMP-4 arrays have been described among different Enterobacteriaceae and Acinetobacter spp. in China, Singapore and Australia but the genetic context is different. The broad host range of IncA/C plasmids may have facilitated dissemination of the bla _IMP-4 arrays among different diverse groups of bacteria.     

Transmission and characterization of <Emphasis Type="Italic">bla</Emphasis><Subscript>NDM-1</Subscript> in <Emphasis Type="Italic">Enterobacter cloacae</Emphasis> at a teaching hospital in Yunnan,China

Background

In recent years, New Delhi metallo-beta-lactamases 1 (bla _NDM-1) has been reported with increasing frequency and become prevalent. The present study was undertaken to investigate the epidemiological dissemination of the bla _NDM-1 gene in Enterobacter cloacae isolates at a teaching hospital in Yunnan, China.

Methods

Antimicrobial susceptibility testing was performed using VITEK 2 system and E test gradient strips. The presence of integrons and insertion sequence common region 1 were examined by PCR and sequencing. Clonal relatedness was assessed by pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing. Conjugation experiments and Southern blot hybridization were performed to determine the transferability of plasmids.

Results

Ten E. cloacae isolates and their Escherichia coli transconjugants were exhibited similar resistant patterns to carbapenems, cephalosporins and penicillins. 8 (80%) of E. cloacae isolates carried class 1 integron and 1 (12.5%) carried class 2 integron. Integron variable regions harbored the genes which encoded resistance to aminoglycosides (aadA1, aadA2, aadA5, aadB, aac(6′)-Ib-cr), sulfamethoxazole/trimethoprim (dfrA17, dfrA12, dfrA15) and Streptozotocin (sat2). Six E. cloacae isolates belonged to ST74 and exhibited highly similar PFGE patterns. Each isolate shared an identical plasmid with ~33.3 kb size that carried the bla _NDM-1 gene, except T3 strain, of which the bla _NDM-1 gene was located on a ~50 kb plasmid.

Conclusions

Our findings suggested that plasmid was able to contribute to the dissemination of bla _NDM-1. Hence, more attention should be devoted to monitor the dissemination of the bla _NDM-1 gene due to its horizontal transfer via plasmid. In addition, nosocomial surveillance system should actively monitor the potential endemic clone of ST74 to prevent their further spread.