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1.
Infect Drug Resist ; 12: 2627-2635, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31692544

RESUMO

Background: The concurrence of mcr and carbapenemase genes among Enterobacteriaceae has been a great clinical concern. In our study, we aimed to investigate the prevalence of mcr-positive carbapenem-resistant Enterobacteriaceae (CRE) in fresh vegetables and shed light on the possibility of transmission of mcr-positive CRE via fresh vegetables. Methods: In this study, 712 fresh vegetable samples from 10 provinces in China were collected between May 2017 and Dec 2018 and were screened for mcr and carbapenemase genes. Antibiotic susceptibilities for isolates co-harboring carbapenemase genes and mcr were determined by an agar dilution or a broth microdilution method. Pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST) analysis were also performed. Transferability of the carbapenemase/mcr-bearing plasmids was determined by conjugation, replicon typing and S1-PFGE-Southern blotting. The sequences of these plasmids were analyzed by using whole-genome sequencing with Illumina Hiseq platform. Results: Two E. coli isolates concomitantly carrying mcr-1 and bla NDM-5/9 from leaf rape and spinach, respectively, were found and both isolates showed multidrug resistance. Notably, mcr-1-positive 690 harboring bla NDM-5 and 701 carrying bla NDM-9 belonged to ST156 and ST2847, respectively, similar to the prevalent MLST types of E. coli co-carrying mcr-1 and bla NDM from avian in our previous study. mcr-1 was on ~33-kb IncX4 plasmid or ~60-kb IncI2 plasmid, while bla NDM-5/9 was on ~46-kb IncX3 plasmid or ~120-kb untypable plasmid. The plasmids were highly similar to those from animals and clinical patients reported in various countries.Conclusion: E. coli isolates concomitantly carrying mcr-1 and bla NDM-5/9 in fresh vegetables may serve as a direct source of pathogens in humans, and such discovery in fresh vegetables emphasizes the importance of prompt surveillance and intervention in limiting the spread of E. coli co-carrying bla NDM and mcr-1. To our knowledge, this is the first report of Enterobacteriaceae co-carrying bla NDM and mcr-1 in fresh vegetables.

2.
J Med Microbiol ; 68(6): 866-873, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31107201

RESUMO

PURPOSE: The emergence and spread of carbapenem-resistant hypervirulent Klebsiella pneumoniae (CR-hvKP) is causing worldwide concern, whereas NDM-producing hvKP is still rare. Here we report the complete genome sequence characteristics of an NDM-1-producing ST23 type clinical hvKP in PR China. METHODOLOGY: Capsular polysaccharide serotyping was performed by PCR. The complete genome sequence of isolate 3214 was obtained using both the Illumina Hiseq platform and Pacbio RS platform. Multilocus sequence type was identified by submitting the genome sequence to mlst 2.0 and the antimicrobial resistance genes and plasmid replicons were identified using ResFinder and PlasmidFinder, respectively. Transferability of the blaNDM-1-bearing plasmid was determined by conjugation experiment, S1 pulsed-field gel electrophoresis and Southern hybridization. RESULTS: Isolate 3214 was classified to ST23 and belonged to the K1 capsular serotype. The isolate's total genome size was 6 171 644 bp with a G+C content of 56.39 %, consisting of a 5 448 209 bp chromosome and seven plasmids. The resistome included 18 types of antibiotic resistance genes. Fourteen resistance genes including blaNDM-1 and blaCTX-M-14 were located on plasmids and five also including blaCTX-M-14 were in the chromosome. Plasmid pNDM_3214 carrying blaNDM-1 harboured six types of resistance genes surrounded by insertion sequences and was conjugative. The worldwide pLVPK-like virulence plasmid harbouring rmpA2 and rmpA was also found in this isolate. CONCLUSION: This study provides basic information of phenotypic and genomic features of ST23 CR-hvKP isolate 3214. Our data highlights the potential risk of spread of NDM-1-producing ST23 hvKP.


Assuntos
Antibacterianos/farmacologia , Farmacorresistência Bacteriana , Infecções por Klebsiella/microbiologia , Klebsiella pneumoniae/enzimologia , beta-Lactamases/metabolismo , China/epidemiologia , Eletroforese em Gel de Campo Pulsado , Humanos , Infecções por Klebsiella/tratamento farmacológico , Infecções por Klebsiella/epidemiologia , Klebsiella pneumoniae/genética , Klebsiella pneumoniae/imunologia , Klebsiella pneumoniae/patogenicidade , Tipagem de Sequências Multilocus , Plasmídeos/genética , Polissacarídeos Bacterianos/genética , Sorogrupo , Sorotipagem , Virulência , Sequenciamento Completo do Genoma , beta-Lactamases/genética
3.
Int J Antimicrob Agents ; 54(1): 89-94, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31034936

RESUMO

The presence of mobilized colistin resistance (mcr) genes is a global concern. However, data concerning mcr in fresh vegetables, a reservoir for antibiotic resistance genes, are still rare. In this study, mcr genes were analysed in 528 vegetable samples from 53 supermarkets or farmer's markets in 23 cities of 9 provinces in China, and the mcr-positive Enterobacteriaceae were characterized. Nineteen (3.6%) samples carried one or more mcr-positive isolates, and the highest three detection rates were found in carrot, pak choi and green pepper. Twenty-four mcr-1-positive isolates (23 Escherichia coli and one Enterobacter cloacae) were obtained, and E. coli isolates showed high genetic diversity. Different multilocus sequence type (MLST) isolates were also observed within the same sample. All 24 isolates showed multidrug resistance, and 14 carried blaCTX-M genes. Most isolates harbored similarly conjugative IncX4-type (∼33 kb) or IncI2-type (∼60 kb) mcr-1-bearing plasmids. The sequenced prevalent IncX4 plasmid and IncI2 plasmid from tomato were similar to the relevant plasmids from animals and clinical isolates in various countries. mcr-1-bearing IncHI2/ST3 plasmid highly similar to that carrying 14 resistance genes from E. coli of chicken was also observed. In conclusion, a high prevalence of mcr-1 in fresh vegetables was found in China, and the dissemination of mcr-1 was mediated by similar IncX4 or IncI2 plasmids. The plasmids from vegetables showed high similarity to plasmids from clinical isolates, indicating MCR-1-producers in ready-to-eat vegetables may pose a huge threat to public health and measures need to be taken to ensure food safety.


Assuntos
Antibacterianos/farmacologia , Colistina/farmacologia , Farmacorresistência Bacteriana , Enterobacter cloacae/isolamento & purificação , Escherichia coli/isolamento & purificação , Etanolaminofosfotransferase/genética , Verduras/microbiologia , China , Cidades , Enterobacter cloacae/classificação , Enterobacter cloacae/efeitos dos fármacos , Enterobacter cloacae/genética , Escherichia coli/classificação , Escherichia coli/efeitos dos fármacos , Escherichia coli/genética , Plasmídeos/análise
4.
Artigo em Inglês | MEDLINE | ID: mdl-30476621

RESUMO

INTRODUCTION: Haemophilus parasuis, one of the major swine pathogens, has at least fifteen different types, all of which have significant economic effects on the global swine industry. The aim of this study was to establish an experimental intraperitoneal infection model for H. parasuis in neutropenic guinea pigs. METHODS: Intraperitoneal administration of cyclophosphamide and Haemophilus parasuis was conducted in guinea pigs. Clinical signs, gross pathology, and histopathology were observed in neutropenic guinea pigs infected with H. parasuis. RESULTS: Intraperitoneal administration of 100 mg/kg cyclophosphamide led to immunosuppression with white blood cells, lymphocytes, and neutrophils all <1000 mm3, while no histological tissue damage was observed. Intraperitoneal administration of 109 colony-forming units (CFU) of H. parasuis led to typical respiratory symptoms, 90% morbidity, and 20% mortality in a 72 h-period. Bacteriological screening revealed that multiple organs, including the heart, liver, spleen, lungs, kidneys, and blood, were infected with H. parasuis. The threshold loads of bacteria in blood and the lungs were (7.04 ±â€¯0.53)log10 CFU/mL and (6.24 ±â€¯0.62)log10 CFU/g, respectively, at 3 d after infection. Gross pathology examination showed celiac effusion, intestinal mucosal hemorrhage, and liver, spleen, or lung swelling, necrosis, and hemorrhage. Congestion, mild interstitial pneumonia, inflammatory exudation, and endothelial cell proliferation were observed in the histological examination. DISCUSSION: All the results suggest that we have established an experimental intraperitoneal infection model for H. parasuis in neutropenic guinea pigs. It is especially useful as a tool for pharmacokinetics, pharmacodynamics, or a pharmacokinetics/pharmacodynamics (PK/PD) model of antimicrobial agents against respiratory disease.


Assuntos
Modelos Animais de Doenças , Infecções por Haemophilus/imunologia , Haemophilus parasuis/imunologia , Imunossupressores/administração & dosagem , Modelos Animais , Neutropenia/induzido quimicamente , Animais , Ciclofosfamida/administração & dosagem , Feminino , Cobaias , Infecções por Haemophilus/microbiologia , Infecções por Haemophilus/veterinária , Haemophilus parasuis/patogenicidade , Injeções Intraperitoneais , Pulmão , Masculino , Camundongos , Neutropenia/imunologia , Suínos
5.
Microb Drug Resist ; 25(1): 108-119, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30179517

RESUMO

The emergence and spread of multidrug resistance (MDR) plasmids carrying the colistin resistance gene mcr-1 has become a major public health concern. However, there is a paucity of data regarding the prevalence of mcr-1 plasmids concomitantly carrying blaCTX-M and oqxAB, an efflux pump that confers resistance to multiple agents. In this study, we determined the prevalence and characteristics of plasmids coharboring mcr-1, oqxAB, and blaCTX-M as well as those harboring oqxAB and blaCTX-M in Escherichia coli from food-producing animals. We isolated 493 E. coli strains, and mcr-1, blaNDM, and blaCTX-M were present in 140 (28.4%), 51 (10.3%), and 195 (39.6%) of the isolates, respectively. The two most prevalent plasmid-mediated quinolone resistance genes were oqxAB (34.5%) and qnrS (29.4%). Nine IncHI2/ST3 plasmids co-carrying mcr-1, oqxAB, and blaCTX-M were found, and similar IncHI2/ST3 plasmids mediated dissemination of these resistance genes. Two sequenced MDR IncHI2/ST3 plasmids coharboring mcr-1, oqxAB, and blaCTX-M showed high similarity to reference plasmid pHNSHP45-2, although they were from different regions in China. Colocalization of oqxAB and blaCTX-M on the same plasmid was found in 28 isolates, including the nine plasmids harboring mcr-1. The co-dissemination of oqxAB and blaCTX-M was mediated by diverse F33:A-:B- plasmids and similar IncHI2/ST3 plasmids. Pulsed-field gel electrophoresis and multilocus sequence typing analysis of donor isolates revealed heterogeneous patterns indicating that clonal dissemination was unlikely. The high incidence of similar IncHI2/ST3 plasmids simultaneously possessing mcr-1, oqxAB, and blaCTX-M poses a great threat to public health.


Assuntos
Farmacorresistência Bacteriana Múltipla/genética , Proteínas de Escherichia coli/genética , Escherichia coli/genética , Carne/microbiologia , Plasmídeos/genética , beta-Lactamases/genética , Animais , Antibacterianos/farmacologia , China , Colistina/farmacologia , Escherichia coli/efeitos dos fármacos , Alimentos , Testes de Sensibilidade Microbiana/métodos , Prevalência , Quinolonas/farmacologia
6.
Front Microbiol ; 9: 1147, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29910786

RESUMO

Vegetables harboring bacteria resistant to antibiotics are a growing food safety issue. However, data concerning carbapenem-resistant Enterobacteriaceae (CRE) in ready-to-eat fresh vegetables is still rare. In this study, 411 vegetable samples from 36 supermarkets or farmer's markets in 18 cities in China, were analyzed for CRE. Carbapenemase-encoding genes and other resistance genes were analyzed among the CRE isolates. Plasmids carrying carbapenemase genes were studied by conjugation, replicon typing, S1-PFGE southern blot, restriction fragment length polymorphism (RFLP), and sequencing. CRE isolates were also analyzed by pulsed-field gel electrophoresis (PFGE). Ten vegetable samples yielded one or more CRE isolates. The highest detection rate of CRE (14.3%, 4/28) was found in curly endive. Twelve CRE isolates were obtained and all showed multidrug resistance: Escherichia coli, 5; Citrobacter freundii, 5; and Klebsiella pneumoniae, 2. All E. coli and C. freundii carried blaNDM, while K. pneumoniae harbored blaKPC-2. Notably, E. coli with blaNDM and ST23 hypervirulent Klebsiella pneumoniae (hvKP) carrying blaKPC-2 were found in the same cucumber sample and clonal spread of E. coli, C. freundii, and K. pneumoniae isolates were all observed between vegetable types and/or cities. IncX3 plasmids carrying blaNDM from E. coli and C. freundii showed identical or highly similar RFLP patterns, and the sequenced IncX3 plasmid from cucumber was also identical or highly similar (99%) to the IncX3 plasmids from clinical patients reported in other countries, while blaKPC-2 in K. pneumoniae was mediated by similar F35:A-:B1 plasmids. Our results suggest that both clonal expansion and horizontal transmission of IncX3- or F35:A-:B1-type plasmids may mediate the spread of CRE in ready-to-eat vegetables in China. The presence of CRE in ready-to-eat vegetables is alarming and constitutes a food safety issue. To our knowledge, this is the first report of either the C. freundii carrying blaNDM, or K. pneumoniae harboring blaKPC-2 in vegetables. This is also the first report of ST23 carbapenem-resistant hvKP strain in vegetables.

7.
Artigo em Inglês | MEDLINE | ID: mdl-28069644

RESUMO

This study investigated the characteristics of Escherichia coli isolates carrying mcr-1-blaNDM from a chicken farm in China. Of the 78 E. coli isolates, 21 clonally unrelated isolates carried mcr-1-blaNDM Diverse IncI2 plasmids disseminated mcr-1, while the dissemination of blaNDM was mediated by diverse IncB/O plasmids. More striking was the colocalization of resistance genes mcr-1 and blaNDM-4 in an IncHI2/ST3 plasmid, which might pose a great challenge for public health.


Assuntos
Farmacorresistência Bacteriana Múltipla/genética , Infecções por Escherichia coli/veterinária , Proteínas de Escherichia coli/genética , Regulação Bacteriana da Expressão Gênica , Plasmídeos/metabolismo , Doenças das Aves Domésticas/epidemiologia , beta-Lactamases/genética , Criação de Animais Domésticos , Animais , Antibacterianos/farmacologia , Carbapenêmicos/farmacologia , Galinhas , China/epidemiologia , Células Clonais , Colistina/farmacologia , Escherichia coli/efeitos dos fármacos , Escherichia coli/genética , Escherichia coli/crescimento & desenvolvimento , Infecções por Escherichia coli/tratamento farmacológico , Infecções por Escherichia coli/epidemiologia , Infecções por Escherichia coli/transmissão , Proteínas de Escherichia coli/metabolismo , Transferência Genética Horizontal , Testes de Sensibilidade Microbiana , Plasmídeos/química , Doenças das Aves Domésticas/tratamento farmacológico , Doenças das Aves Domésticas/microbiologia , Doenças das Aves Domésticas/transmissão , beta-Lactamases/metabolismo
8.
Artigo em Inglês | MEDLINE | ID: mdl-27855074

RESUMO

We report the presence of mcr-1 in Escherichia coli and carbapenem-resistant Cronobacter sakazakii from the same diseased chicken. The mcr-1 gene linked with ISApl1 was located on two different IncI2 plasmids, including one multidrug plasmid in E. coli, whereas fosA3-blaNDM-9 was on an IncB/O plasmid in C. sakazakii The development of the fosA3-blaNDM-9 resistance region was mediated by IS26 The colocation of mcr-1 or blaNDM-9 with other resistance genes will accelerate the dissemination of the two genes.


Assuntos
Colistina/farmacologia , Cronobacter sakazakii/efeitos dos fármacos , Farmacorresistência Bacteriana/genética , Proteínas de Escherichia coli/genética , Escherichia coli/genética , Animais , Antibacterianos/farmacologia , Galinhas/microbiologia , China , Cronobacter sakazakii/genética , Cronobacter sakazakii/isolamento & purificação , Farmacorresistência Bacteriana/efeitos dos fármacos , Infecções por Enterobacteriaceae/microbiologia , Infecções por Enterobacteriaceae/veterinária , Escherichia coli/efeitos dos fármacos , Infecções por Escherichia coli/microbiologia , Infecções por Escherichia coli/veterinária , Testes de Sensibilidade Microbiana , Plasmídeos/efeitos dos fármacos , Plasmídeos/genética , beta-Lactamases/genética
9.
Antimicrob Agents Chemother ; 60(7): 4336-8, 2016 07.
Artigo em Inglês | MEDLINE | ID: mdl-27067314

RESUMO

We sequenced a novel conjugative multidrug resistance IncF plasmid, p42-2, isolated from Escherichia coli strain 42-2, previously identified in China. p42-2 is 106,886 bp long, composed of a typical IncFII-type backbone (∼54 kb) and one distinct acquired DNA region spanning ∼53 kb, harboring 12 antibiotic resistance genes [blaCTX-M-55, oqxA, oqxB, fosA3, floR, tetA(A), tetA(R), strA, strB, sul2, aph(3')-II, and ΔblaTEM-1]. The spread of these multidrug resistance determinants on the same plasmid is of great concern and, because of coresistance to antibiotics from different classes, is therapeutically challenging.


Assuntos
Antibacterianos/farmacologia , Escherichia coli/efeitos dos fármacos , Escherichia coli/genética , beta-Lactamases/metabolismo , China , Farmacorresistência Bacteriana Múltipla/genética , Plasmídeos/genética , beta-Lactamases/genética
10.
Front Microbiol ; 6: 964, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-26441898

RESUMO

The purpose of this study was to characterize a collection of 103 multidrug resistance IncF plasmids recovered from Escherichia coli of food producing and companion animals between 2003 and 2012. A total of 103 incF plasmids were characterized using an established PCR-based IncF replicon sequence typing (RST) system to identify FII, FIA, and FIB (FAB) groups. Plasmids were also analyzed using-restriction fragment length polymorphism (RFLP). Antibiotic Resistance determinants bla CTX-M , plasmid-mediated quinolone resistance (PMQR) genes and rmtB and plasmid addiction systems (PAS) were identified by PCR screening. A total of 20 different RSTs from 103 IncF plasmids were identified. The groups F2 and F33 with the RST formulae A-: B- were the most frequently encountered types (63.1%). The antibiotic resistance genes (ARGs) bla CTX-M , rmtB, and oqxB were carried by 82, 37, and 34 IncF plasmids, respectively. Most of these plasmids carried more than one resistance gene (59.2%, 61/103). The IncF plasmids also had a high frequency of addiction systems (mean 2.54) and two antisense RNA-regulated systems (hok-sok and srnBC) and a protein antitoxin-regulated system (pemKI) were the most prevalent. Not surprisingly, RFLP profiles among the IncF plasmids were diverse even though some shared identical IncF-RSTs. This is the first extensive study of IncF plasmid-positive E. coli isolates from animals in China. Our results demonstrate that IncF is the most prevalent plasmid family in E. coli plasmids and they commonly carry multiple resistance determinants that render them resistant to different antibiotic classes simultaneously. IncF plasmids also harbor addiction systems, promoting their stability and maintenance in the bacterial host, under changing environmental conditions.

11.
Mol Genet Genomics ; 290(4): 1543-9, 2015 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-25724693

RESUMO

Stalk rots are destructive diseases in maize around the world, and are most often caused by the pathogen Pythium, Fusarium and other fungi. The most efficient management for controlling stalk rots is to breed resistant cultivars. Pythium stalk rot can cause serious yield loss on maize, and to find the resistance genes from the existing germplasm is the basis to develop Pythium-resistance hybrid lines. In this study, we investigated the genetic resistance to Pythium stalk rot in inbred line Qi319 using F2 and F2:3 population, and found that the resistance to Pythium inflatum in Qi319 was conferred by two independently inherited dominant genes, RpiQI319-1 and RpiQI319-2. Linkage analysis uncovered that the RpiQI319-1 co-segregated with markers bnlg1203, and bnlg2057 on chromosome 1, and that the RpiQI319-2 locus co-segregated with markers umc2069 and bnlg1716 on chromosome 10. The RpiQI319-1 locus was further mapped into a ~500-kb interval flanked by markers SSRZ33 and SSRZ47. These results will facilitate marker-assisted selection of Pythium stalk rot-resistant cultivars in maize breeding. To our knowledge, this is the first report on the resistance to P. inflatum in the inbred line Qi319, and is also the first description of two independently inherited dominant genes conferring the resistance of Pythium stalk rot in maize.


Assuntos
Resistência à Doença/genética , Genes de Plantas/genética , Doenças das Plantas/genética , Zea mays/genética , Mapeamento Cromossômico , Cromossomos de Plantas/genética , DNA de Plantas/genética , Genes Dominantes/genética , Ligação Genética , Marcadores Genéticos/genética , Interações Hospedeiro-Patógeno , Endogamia , Doenças das Plantas/microbiologia , Reação em Cadeia da Polimerase , Pythium/fisiologia , Zea mays/microbiologia
12.
Int J Antimicrob Agents ; 44(3): 263-8, 2014 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-25108878

RESUMO

In this study, 130 non-Typhi Salmonella enterica isolates from chickens were analysed for the prevalence of plasmid-mediated quinolone resistance (PMQR) determinants as well as the co-existence of oqxAB and extended-spectrum ß-lactamase (ESBL) genes. The genes oqxAB, aac(6')-Ib-cr, blaCTX-M-9G and blaTEM were present alone or in combination in 40 (30.8%), 40 (30.8%), 55 (42.3%) and 2 (1.5%) isolates, respectively. Most of the oqxAB-blaCTX-M-9G-positive isolates (17/28) carried transferable ST2-IncHI2 plasmids containing an oqxAB cassette and blaCTX-M-14 flanked by insertion sequences IS10 or ISEcp1 upstream and IS903 downstream. The oqxAB-blaCTX-M-9G-positive isolates from a local area showed similar pulsed-field gel electrophoresis (PFGE) patterns, whilst the isolates from different areas were genetically divergent, suggesting that both clonal expansion in local areas and horizontal transmission contributed to the spread of ST2-IncHI2 plasmids containing oqxAB and blaCTX-M-14. This is the first report on the prevalence of ST2-IncHI2 plasmids concomitantly carrying oqxAB and blaCTX-M-14 in Salmonella and also the first description of the genetic environment of oqxAB-blaCTX-M. The genetic linkage of oqxAB-blaCTX-M-9G in non-Typhi Salmonella likely facilitates the spread of antibiotic-resistant Salmonella and poses a threat for clinical treatment of salmonellosis.


Assuntos
Farmacorresistência Bacteriana Múltipla , Proteínas de Membrana Transportadoras/genética , Plasmídeos/análise , Doenças das Aves Domésticas/microbiologia , Salmonelose Animal/microbiologia , Salmonella enterica/genética , beta-Lactamases/genética , Animais , Galinhas , DNA Bacteriano/química , DNA Bacteriano/genética , Proteínas de Membrana Transportadoras/metabolismo , Dados de Sequência Molecular , Salmonella enterica/enzimologia , Análise de Sequência de DNA , beta-Lactamases/metabolismo
13.
Microb Drug Resist ; 20(6): 641-50, 2014 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-24927154

RESUMO

To study the characteristics of plasmids harboring oqxAB among bla(CTX-M)-negative Escherichia coli isolates and search for oqxAB-harboring plasmids similar to plasmids carrying oqxAB-bla(CTX-M) reported previously, conjugation experiment was performed for 115 randomly selected oqxAB-positive but bla(CTX-M)-negative E. coli isolates from diseased animals in Guangdong, China. S1 nuclease pulsed-field gel electrophoresis (PFGE) and southern blotting experiments were performed to investigate the location of oqxAB and other resistance genes. The EcoRI digestion profiles of the plasmids with oqxAB were also analyzed. The clonal relatedness of donor isolates was investigated by PFGE. In this study, 32 oqxAB transconjugants were successfully obtained and most transconjugants showed multidrug resistances. Eleven replicon combination types were found in these transconjugants. floR and oqxAB were found on the same plasmids in all nine transconjugants resistant to florfenicol. The sequences between floR and oqxAB were identical in most transconjugants and the two genes were both linked with tnp in insertion sequences. Nine F18:A-:B1 plasmids with only oqxAB shared identical EcoRI digestion profiles and the profiles were also identical with that of a plasmid carrying oqxAB-bla(CTX-M) found previously. Co-transfer of plasmids carrying oqxAB and fosA3, respectively, was also observed in one isolate. This study demonstrates the dissemination of oqxAB among bla(CTX-M)-negative E. coli isolates was mainly mediated by identical F18:A-:B1 plasmids. A novel arrangement of regions between floR and oqxAB might play an important role in the dissemination of floR-oqxAB. This is the first description of the genetic environment of the relationship between oqxAB and floR in E. coli.


Assuntos
Proteínas de Escherichia coli/genética , Escherichia coli/genética , Plasmídeos/genética , Animais , Antibacterianos/farmacologia , China , Conjugação Genética/genética , Farmacorresistência Bacteriana Múltipla/genética , Eletroforese em Gel de Campo Pulsado/métodos , Escherichia coli/efeitos dos fármacos , Infecções por Escherichia coli/tratamento farmacológico , Microbiologia de Alimentos/métodos , Replicon/genética , Tianfenicol/análogos & derivados , Tianfenicol/farmacologia , beta-Lactamases/genética
14.
J Food Prot ; 76(12): 2018-23, 2013 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-24290675

RESUMO

A total of 247 Escherichia coli isolates (148 from diseased or dead poultry and 99 from diseased pets in the People's Republic of China) were screened for extended-spectrum ß-lactamase (ESBL) determinants by PCR and sequencing. Then, 16S rRNA methylase genes were detected among ESBL-producing isolates. Clonal relatedness of the E. coli isolates was examined by pulsed-field gel electrophoresis. Conjugation experiments were performed to investigate the association of 16S rRNA methylases and ESBLs, and plasmid contents were also characterized. Among 247 E. coli isolates, 74 (29.96%) isolates were positive for blaCTX-M genes, 42 from pets (12 from cats and 30 from dogs) and 32 from poultry (12 from chickens and 20 from ducks). The most common CTX-M type in isolates from pets was blaCTX-M-14, whereas blaCTX-M-27 was the most common for poultry. rmtB was dectected in 39 of the 74 blaCTX-M-positive isolates, 18 from pets and 21 from poultry. One strain from a pet was found to harbor blaCTX-M-14, blaCTX-M-15, and rmtB. blaCTX-M and rmtB were found to be colocated on the same transferable plasmid in 16 isolates. These genes were on the same or similar plasmids (eight F2:A-:B- and two IncN) in isolates from ducks, whereas they were colocated on the similar F2:A-:B- or similar F33:A-:B- plasmids in isolates of pets origin. In conclusion, similar F2:A-:B- plasmids and similar F33:A-:B- plasmids are responsible for the dissemination of both rmtB and blaCTX-Mgenes in E. coli isolates from poultry and pets, respectively.


Assuntos
Infecções por Escherichia coli/microbiologia , Proteínas de Escherichia coli/genética , Escherichia coli/enzimologia , beta-Lactamases/genética , Animais , Gatos , China , Conjugação Genética , Cães , Eletroforese em Gel de Campo Pulsado , Escherichia coli/isolamento & purificação , Infecções por Escherichia coli/genética , Infecções por Escherichia coli/veterinária , Metiltransferases/genética , Plasmídeos , Reação em Cadeia da Polimerase , Aves Domésticas , RNA Ribossômico 16S/genética
15.
PLoS One ; 8(9): e73947, 2013.
Artigo em Inglês | MEDLINE | ID: mdl-24040123

RESUMO

BACKGROUND: The association of PMQR and ESBLs in negative-bacteria isolates has been of great concern. The present study was performed to investigate the prevalence of co-transferability of oqxAB and bla CTX-M genes among the 696 Escherichia coli (E. coli) isolates from food-producing animals in South China, and to characterize these plasmids. METHODS: The ESBL-encoding genes (bla(CTX-M), bla(TEM) and bla(SHV)), and PMQR (qnrA, qnrB, qnrS, qnrC, qnrD, aac(6')-Ib-cr, qepA, and oqxAB) of these 696 isolates were determined by PCR and sequenced directionally. Conjugation, S1 nuclease pulsed-field gel electrophoresis (PFGE) and Southern blotting experiments were performed to investigate the co-transferability and location of oqxAB and bla(CTX-M). The EcoRI digestion profiles of the plasmids with oqxAB-bla(CTX-M) were also analyzed. The clonal relatedness was investigated by PFGE. RESULTS: Of the 696 isolates, 429 harbored at least one PMQR gene, with oqxAB (328) being the most common type; 191 carried bla(CTX-M), with bla(CTX-M-14) the most common. We observed a significant higher prevalence of bla(CTX-M) among the oqxAB-positive isolates (38.7%) than that (17.4%) in the oqxAB-negative isolates. Co-transferability of oqxAB and bla(CTX-M) was found in 18 of the 127 isolates carrying oqxAB-bla(CTX-M). These two genes were located on the same plasmid in all the 18 isolates, with floR being on these plasmids in 13 isolates. The co-dissemination of these genes was mainly mediated by F33:A-: B- and HI2 plasmids with highly similar EcoRI digestion profiles. Diverse PFGE patterns indicated the high prevalence of oqxAB was not caused by clonal dissemination. CONCLUSION: bla(CTX-M) was highly prevalent among the oqxAB-positive isolates. The co-dissemination of oqxAB-bla(CTX-M) genes in E. coli isolates from food-producing animals is mediated mainly by similar F33:A-: B- and HI2 plasmids. This is the first report of the co-existence of oqxAB, bla(CTX-M), and floR on the same plasmids in E. coli.


Assuntos
Escherichia coli/classificação , Escherichia coli/genética , Plasmídeos/genética , beta-Lactamases/genética , Animais , Antibacterianos/farmacologia , Galinhas , Conjugação Genética , Farmacorresistência Bacteriana/genética , Patos , Escherichia coli/efeitos dos fármacos , Escherichia coli/isolamento & purificação , Gansos , Testes de Sensibilidade Microbiana , Tipagem Molecular , Quinolonas/farmacologia , Suínos
16.
Microb Drug Resist ; 19(4): 331-5, 2013 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-23557071

RESUMO

qnrD, unlike other qnr genes, is mainly located on small nonconjugative plasmids. We investigated the presence of qnrD among 1,373 Enterobacteriaceae isolates in China. Twelve qnrD-positive strains were detected, and all were nonsusceptible to fluoroquinolones. The complete sequence of plasmids showed that the qnrD determinants were located on two plasmids with a respective size of ~4.2 and 2.7 k-bp. Interestingly, the identification of qnrD in this study revealed the highest prevalence of Proteeae among Enterobacteriaceae identified.


Assuntos
Animais Domésticos/microbiologia , Proteínas de Bactérias/genética , Infecções por Enterobacteriaceae/veterinária , Enterobacteriaceae/genética , Proteínas de Escherichia coli/genética , Produtos da Carne/microbiologia , Plasmídeos , Animais , Antibacterianos/farmacologia , Proteínas de Bactérias/classificação , Técnicas de Tipagem Bacteriana , China/epidemiologia , Conjugação Genética , Farmacorresistência Bacteriana/efeitos dos fármacos , Farmacorresistência Bacteriana/genética , Enterobacteriaceae/classificação , Enterobacteriaceae/isolamento & purificação , Infecções por Enterobacteriaceae/tratamento farmacológico , Infecções por Enterobacteriaceae/epidemiologia , Infecções por Enterobacteriaceae/microbiologia , Proteínas de Escherichia coli/classificação , Fluoroquinolonas/farmacologia , Humanos , Testes de Sensibilidade Microbiana , Dados de Sequência Molecular
18.
Foodborne Pathog Dis ; 10(1): 28-34, 2013 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-23320420

RESUMO

Development of antibiotic resistance may alter the virulence properties of bacterial organisms. In this study, nine clinical ceftriaxone-susceptible Salmonella enterica serotype Typhimurium strains were subjected to stepwise selection with increasing concentrations of ceftriaxone in culture media. Mutations in virulence-associated genes and antibiotic efflux genes were analyzed by polymerase chain reaction (PCR) and DNA sequencing. The expression levels of virulence genes invA and stn as well as efflux pump genes tolC, arcA, and arcB before and after the selection were measured by real-time quantitative reverse transcription-polymerase chain reaction (qRT-PCR). The stepwise selection resulted in the development of Salmonella strains that were highly resistant to ceftriaxone. Sequence analysis did not reveal any mutations or deletions in the examined virulence genes and regulatory gene, but a silent mutation (T423C) in acrR (encoding a repressor for the efflux pump) was detected in most of the ceftriaxone-resistant strains. The qRT-PCR revealed increased expression of the AcrAB-TolC efflux pump and decreased expression of invA and stn in the ceftriaxone-resistant strains. Moreover, decreased invasion into cultured epithelial cells and reduced growth rates were observed with the resistant strains. These results suggest that acquisition of ceftriaxone resistance is associated with the overexpression of the AcrAB-TolC efflux pump and leads to reduced virulence in Salmonella Typhimurium.


Assuntos
Antibacterianos/farmacologia , Ceftriaxona/farmacologia , Resistência às Cefalosporinas/genética , Regulação Bacteriana da Expressão Gênica , Salmonella typhimurium/efeitos dos fármacos , Salmonella typhimurium/patogenicidade , Animais , Proteínas da Membrana Bacteriana Externa/genética , Proteínas de Bactérias/genética , Sequência de Bases , Regulação para Baixo , Enterotoxinas/genética , Proteínas de Escherichia coli/genética , Células HeLa , Humanos , Testes de Sensibilidade Microbiana , Dados de Sequência Molecular , Mutação Puntual , Reação em Cadeia da Polimerase em Tempo Real , Proteínas Repressoras/genética , Infecções por Salmonella/microbiologia , Salmonella typhimurium/genética , Salmonella typhimurium/fisiologia , Análise de Sequência de DNA , Regulação para Cima , Virulência/genética
19.
Microb Drug Resist ; 19(3): 237-45, 2013 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-23289437

RESUMO

Plasmid-mediated quinolone-resistance (PMQR) genes were determined by polymerase chain reactions (PCRs) in 250 Escherichia coli isolates from food-producing animals in Guangdong, China, in 2009-2010. Then, the prevalence of plasmid-mediated ß-lactamase and 16S rRNA methylase genes was determined by PCRs among the PMQR-positive isolates. One hundred fifty-seven (62.8%) isolates were found to harbor at least one PMQR gene, and qnrS (84) and oqxAB (97) were the most two prevalent PMQR genes. ß-lactamase (ESBL and/or AmpC type) genes were detected in 106 of the 157 PMQR-positive strains. The bla(TEM-1) (78) was the most prevalent ß-lactamase gene in the 157 PMQR-positive isolates, followed by bla(CMY-2) (28), bla(CTX-M) (25), bla(SHV-1) (3), and bla(DHA-1) (3). Twenty-nine were detected to produce more than one type of ß-lactamase. The rmtB was the most prevalent 16S rRNA methylase gene detected (11.5%, 18/157), and armA was detected in only two (1.27%, 2/157) isolates, with one isolate coharboring rmtB and armA. Sixteen isolates were found to coharbor the three types of resistance genes detected in this study. Only 1 transconjugant JGDA2 harboring oqxAB, aac(6')-Ib-cr, bla(DHA-1), and rmtB was obtained from the 16 isolates harboring the three types of resistance genes, by conjugation experiment. The results of Southern blot hybridization revealed that oqxAB, bla(DHA-1), and rmtB were colocated on the same plasmid of ∼54 kb in the JGDA2. To our knowledge, this is the first description of the coexistence of the oqxAB, rmtB, and bla(DHA-1) resistance genes on the same plasmid in one E. coli strain.


Assuntos
Antibacterianos/farmacologia , Farmacorresistência Bacteriana/genética , Escherichia coli/efeitos dos fármacos , Quinolonas/farmacologia , Animais , Southern Blotting , China , Escherichia coli/genética , Escherichia coli/isolamento & purificação , Infecções por Escherichia coli/microbiologia , Infecções por Escherichia coli/veterinária , Plasmídeos , Reação em Cadeia da Polimerase , RNA Ribossômico 16S/genética , beta-Lactamases/genética , tRNA Metiltransferases/genética
20.
J Med Microbiol ; 61(Pt 11): 1591-9, 2012 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-22878251

RESUMO

In this study, the prevalence of plasmid-mediated quinolone resistance (PMQR) was investigated in 495 Escherichia coli isolates from diseased food-producing animals in Guangdong province, China. The quinolone resistance-determining regions (QRDRs) of the gyrA and parC genes were analysed for mutations in 55 isolates harbouring only oqxAB and all isolates harbouring other PMQR genes. Overall, 282 (57.0 %) E. coli isolates had at least one PMQR gene. oqxAB was detected in 215 isolates and predominated the PMQR genes, followed by qnrS (63 isolates), aac(6')-Ib-cr (56 isolates), qnrB (39 isolates) and qepA (18 isolates). qnrA, qnrC and qnrD were not found in any of the isolates. The rates of resistance to ciprofloxacin, enrofloxacin, levofloxacin and nalidixic acid were 75.2, 81.0, 70.5 and 97.4 %, respectively, among the 495 isolates. Eight types of mutation in gyrA were detected in 154 PMQR-positive isolates, and 147 isolates were found to have mutations in parC. PFGE analysis indicated that the PMQR-positive E. coli isolates were genetically diverse. This study demonstrated that the number of mutations in QRDRs of gyrA and/or parC was significantly associated with the MICs of quinolones (P<0.01). The rates of resistance to ciprofloxacin, enrofloxacin and nalidixic acid in PMQR-positive isolates were significantly higher than those in PMQR-negative isolates (P<0.05). In addition, the prevalence of oqxAB had significant Spearman correlation coefficients in relation to the MICs of all four tested quinolones (P<0.01).


Assuntos
DNA Girase/genética , DNA Topoisomerase IV/genética , Farmacorresistência Bacteriana Múltipla , Escherichia coli/efeitos dos fármacos , Quinolonas/farmacologia , Fatores R/genética , Animais , Antibacterianos/farmacologia , Galinhas , China/epidemiologia , DNA Girase/metabolismo , DNA Topoisomerase IV/metabolismo , Farmacorresistência Bacteriana Múltipla/genética , Patos , Escherichia coli/genética , Escherichia coli/metabolismo , Infecções por Escherichia coli/epidemiologia , Infecções por Escherichia coli/microbiologia , Infecções por Escherichia coli/veterinária , Proteínas de Escherichia coli/genética , Proteínas de Escherichia coli/metabolismo , Microbiologia de Alimentos , Regulação Bacteriana da Expressão Gênica/fisiologia , Mutação , Doenças das Aves Domésticas/epidemiologia , Doenças das Aves Domésticas/microbiologia , Suínos , Doenças dos Suínos/epidemiologia , Doenças dos Suínos/microbiologia
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