鲜牛奶中耐药性细菌的分离与鉴定  

Isolation and identif ication of antibiotics resistant bacteria from fresh milk

作　　者：张晓梅 杨洪江[1] 

机构地区：[1]天津科技大学工业发酵微生物教育部重点实验室,天津市工业微生物重点实验室,天津科技大学生物工程学院,天津300457

出　　处：《食品科技》2012年第6期84-88,共5页Food Science and Technology

基　　金：国家科学自然基金面上项目(30970114)

摘　　要：目的:分离鉴定鲜牛奶中耐药性细菌的分布。方法:利用含有四环素(16μg/mL)、环丙沙星(4μg/mL)或庆大霉素(16μg/mL)的Luria-Bertani平板,分离样品中的耐药性细菌;采用K-B纸片法对分离菌株的耐药性进行确认;利用血平板测定分离菌株的溶血性;利用16S rRNA方法对分离菌株进行鉴定。结果:对30份采自河北张家口地区新鲜无菌牛奶样品的平板筛选结果显示,具有耐受1种以上抗菌素细菌的样品为23份(76.67%),具有耐受2种以上抗菌素细菌的样品为7份(23.33%),具有耐受3种以上抗菌素细菌的样品为1份(3.33%);共分离得到耐受四环素菌株37株、耐受环丙沙星8株、耐受庆大霉素菌株7株;随机挑选6株(每种抗菌素耐受细菌选取2株)进行16S rRNA鉴定,结果为黏质沙雷氏菌(Serratia marcescens)1株、铜绿假单胞菌(Pseudomonas aeruginosa)2株、琼氏不动杆菌(Acinetobacter junii)1株、阪崎肠杆菌(Cronobacter sakazakii)1株和金黄色葡萄球菌(Staphylococcus aureus)1株;6株细菌中α溶血3株,β溶血1株,γ溶血2株;纸片法验证结果显示,只有1株分离细菌对庆大霉素敏感。结论:鲜牛奶样品中普遍存在多种耐药性细菌,抗菌素平板可以用于初步分析牛奶样品的耐药性细菌。Objective： Isolation and identification of antibiotics resistant bacteria in fresh milk. Methods： Luria-Bertani plates containing tetracycline （16 μg/mL）, ciprofloxacin （4 μg/mL） or gentamicin （16 μg/ mL） were used in isolating the antibiotics resistant bacteria from samples. K-B disk diffusion method was used to confirm the resistant phenotypes. Blood agar plates were used in hemolysis assay. 16S rRNA analysis method was used to identify the isolates. Results： 30 fresh milk samples collected from Zhangjiakou were screened for antibiotic resistant bacteria. The screening results showed that 23 （76.67%） samples had isolates resistant to more than one antibiotic, 7（23.33%） samples had isolates resistant tomore than two antibiotics, and 1 （3.33%） sample had isolates resistant to three antibiotics tested. Totally, 37 strains were isolated resistant to tetracycline, 8 strains were isolated resistant to ciprofloxacin, and 7 strains were isolated resistant to gentamicin. Six strains （two from each antibiotic resistant bacteria group） were randomLy selected for 16S rRNA analysis and they were identified as Serratia marcescens （1）, Pseudomonas aeruginosa （2）, Acinetobacter junii （1）, Cronobacter sakazakii （1） and Staphylococcus aureus （1）. Hemolysis test results showed that 3 strains with a-hemolytic phenotype, 1 strains with 13-hemolytic phenotype, and 2 strains with y-hemolytic phenotype. Furthermore, antibiotic resistant phenotypes of the six isolates were validated K-B disk diffusion method and only one isolate sensitive to gentamicin. Conclusion： Fresh milk is the reservoir of a variety of antibiotics resistant bacteria, antibiotic plates can be used for preliminary screening of antibiotics resistant bacteria from fresh milk samples.

关 键 词：牛奶 耐药细菌 分离鉴定 K-B纸片法 

分 类 号：TS252.1[轻工技术与工程—农产品加工及贮藏工程]