四川泡豇豆原料表皮附生乳酸菌抗生素抗性及抗性基因污染分析
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摘要
以鲜豇豆为研究对象,利用生理生化特征、16S r RNA、抗生素抗性和抗性基因检测分析其表皮附生乳酸菌对氯霉素(CHL)、四环素(TET)和氨苄青霉素(AMP)的抗性与抗性基因。结果表明:从鲜豇豆表皮共分离得到182株乳酸菌,分别属于Weissella confuse(50.0%)、Weissella cibaria(15.4%)、Enterococcus sulfurous(7.1%)和Lactococcus lactis subsp.lactis(27.5%)。32株(17.6%)乳酸菌对CHL和TET有抗性,其中,6株W.confuse(3.3%)和4株L.lactis subsp.Lactis(2.1%)对CHL单一抗性,4株W.confuse(2.1%)和5株L.lactis subsp.Lactis(2.7%)对TET单一抗性,6株W.confuse(3.2%)、2株W.cibaria(1.1%)、1株E.sulfurous(0.5%)和4株L.lactis subsp.Lactis(2.1%)对CHL和TET二重抗性;编码外排泵基因efr A、efr B、nor B、nor C、nor E和sug E中,efr B的检出率最高,达31.3%,efr B和sug E的检出率最低,为3.1%。TET被检抗性基因tet(A)、tet(B)、tet(C)、tet(D)、tet(H)和tet(L)中,tet(C)的检出率最高,达90.9%,tet(B)的检出率最低,为4.5%。
The epibiotic lactic acid bacteria(LABs) on the fresh cowpea,which resist to chloramphenicol(CHL)tetracycline(TET) and ampicillin(AMP),were investigated by the physiological and biochemical characteristics,16 S r RNA,antibiotic resistance and resistant genes. 182 isolates in current study were assigned to Weissella confuse(50.0%),Weissella cibaria(15.4%),Enterococcus sulfurous(7.1%) and Lactococcus lactis subsp. lactis( 27. 5 %), respectively. And 32 isolates( 17. 6 %) were found to be against CHL and TET with one or two resistance,including 6 W. confuse(3.3%) and 4 L. lactis subsp. lactis(2.1%) with resistance to CHL,and 4W. confuse(2.1%) and 5 L. lactis subsp. lactis(2.7%) with resistance to TET. 6 W. confuse(3.2%),2 W. cibaria(1.1%),1 E. sulfurous(0.5%) and 4 L. lactis subsp. lactis(2.1%) with resistances to CHL and TET. In the effux pump genes efr A,efr B,nor B,nor C,nor E and sug E,efr B had the highest detection rate with 31.3%,but efr B and sug E with only 3.1%. Among the TET resistant genes tet(A),tet(B),tet(C),tet(D),tet(H) and tet(L),tet(C)had the highest detection rate,90.9%,and tet(B) was found with 4.5% lowest detection rate.
The epibiotic lactic acid bacteria(LABs) on the fresh cowpea,which resist to chloramphenicol(CHL)tetracycline(TET) and ampicillin(AMP),were investigated by the physiological and biochemical characteristics,16 S r RNA,antibiotic resistance and resistant genes. 182 isolates in current study were assigned to Weissella confuse(50.0%),Weissella cibaria(15.4%),Enterococcus sulfurous(7.1%) and Lactococcus lactis subsp. lactis( 27. 5 %), respectively. And 32 isolates( 17. 6 %) were found to be against CHL and TET with one or two resistance,including 6 W. confuse(3.3%) and 4 L. lactis subsp. lactis(2.1%) with resistance to CHL,and 4W. confuse(2.1%) and 5 L. lactis subsp. lactis(2.7%) with resistance to TET. 6 W. confuse(3.2%),2 W. cibaria(1.1%),1 E. sulfurous(0.5%) and 4 L. lactis subsp. lactis(2.1%) with resistances to CHL and TET. In the effux pump genes efr A,efr B,nor B,nor C,nor E and sug E,efr B had the highest detection rate with 31.3%,but efr B and sug E with only 3.1%. Among the TET resistant genes tet(A),tet(B),tet(C),tet(D),tet(H) and tet(L),tet(C)had the highest detection rate,90.9%,and tet(B) was found with 4.5% lowest detection rate.
引文
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[3]Sharma P,Tomar S K,Goswami P,et al.Antibiotic resistance among commercially available probiotics[J].Food Research International,2014,57:176-195.
[4]Campos J,Mourao J,Pestana N,et al.Microbiological quality of ready-to-eat salads:an underestimated vehicle of bacteria and clinically relevant antibiotic resistance genes[J].Int J Food Microbiol,2013,166(3):464-470.
[5]Fernandez-Fuentes M A,Abriouel H,Morente E O,et al.Genetic determinants of antimicrobial resistance in Gram posistiver bacteria from organic foods[J].Int J Food Microbiol,2014,172:49-56.
[6]Zhang X X,Zhang T,Fang H H.Antibiotic resistance genes in water environment[J].Appl Microbiol Biot,2009,82(3):397-414.
[7]Jiang L,Hu X L,Xu T,et al.Prevalence of antibiotic resistance genes and their relationship with antibiotics in the Huangpu River and the drinking water sources[J].Sci Total Environ,2013,458-460:267-272.
[8]凌代文.乳酸细菌分类鉴定及实验方法[M].北京:中国轻工业出版社,1999.
[9]Xiang W L,Liang H Z,Liu S,et al.Isolation and performance evaluation of halotolerant phosphate solubilizing bacteria from the rhizospheric soils of historic Dagong Brine Well in China[J].World Journal of Microbiology and Biotechnology,2011,27(11):2629-2637.
[10]Klare I,Konstabel C,Muller-Bertling S,et al.Evaluation of new broth media for microdilution antibiotic susceptibility testing of Lactobacilli,Pediococci,Lactococci,and Bifidobacteria[J].Applied and Environmental Microbiology,2005,71(12):8982-8986.
[11]Beatriz Rojo-Bezares,Yolanda S,Patricia P,et al.Assessment of antibiotic susceptibility within lactic acid bacteria strains isolated from wine[J].International Journal of Food Microbiology,2006,111:234-240.
[12]杨芳,陶然,杨扬,等.人工湿地中抗生素抗性大肠杆菌和抗性基因的去除与分布[J].环境工程学报,2013,7(6):2057-2062.
[13]Jia S Y,He X W,Bu Y Q,et al.Environmental fate of tetracycline resistance genes originating from swine feedlots in river water[J].Journal of Environmental Science and Health,2014,49(8):624-631.
[14]Aminov R I,Chee-Sanford J C,Garrigues N,et al.Development,validation,and application of PCR primers for detection of tetracycline efflux genes of gram-negative bacteria[J].Applied and Environmental Microbiology,2002,68(4):1786-1793.
[15]He G X,Zhang C,Crow R R,et al.Sug E,a new member of the SMR family of transporters,contributes to antimicrobial resistance in Enterobacter cloacae[J].Antimicrobial Agents and Chemotherapy,2011,55(8):3954-3957.
[16]Patel D,Kosmidis C,Seo,et al.Ethidium bromide MIC screening for enhanced efflux pump gene expression or efflux activity in Staphylococcus aureus[J].Antimicrobial Agents and Chemotherapy,2010,54(12):5070-5073.
[17]Swick M C,Morgan-Linnell S K,Carlson K M,et al.Expression of multidrug efflux pump genes acr AB-tol C,mdf A,and nor E in Escherichia coli clinical isolates as a function of fluoroquinolone and multidrug resistance[J].Antimicrobial Agents and Chemotherapy,2011,55(2):921-924.
[18]Lavilla L L,Benomar N,Sanchez V A,et al.Role of efr AB efflux pump in biocide tolerance and antibiotic resistance of Enterococcus faecalis and Enterococcus faecium isolated from traditional fermented foods and the effect of EDTA as efr AB inhibitor[J].Food Microbiology,2014(44):249-257.
[19]Cytryn E,Rijn J V,Schramm A,et al.Identification of bacteria potentially responsible for oxic and anoxic sulfide oxidation in biofilters of a recirculating mariculture system[J].Applied and Environmental Microbiology,2005,71(10):6134-6141.
[20]Nayak S,Prasanna R,Prasanna B M,et al.Analysing diversity among Indian isolates of Anabaena(Nostocales,Cyanophyta)using morphological,physiological and biochemical characters[J].World Journal of Microbiology Biotechnology,2007,23(11):1575-1584.
[21]Stackebrandt E,Goebel B M.Taxonomic note:a place for DNA-DNA reassociation and 16S r RNA sequence analysis in the present species definition in bacteriology[J].International Journal of Systematic Bacteriology,1994,44(4):846-849.