纺织品中铜绿假单胞菌快速检测方法研究
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摘要
探讨纺织品中铜绿假单胞菌的快速检测方法。将免疫磁珠富集技术和环介导等温扩增技术结合起来,利用免疫磁珠富集技术富集样品中铜绿假单胞菌,富集液提取DNA,进行环介导等温扩增技术特异扩增。以49株铜绿假单胞菌菌株和21株非铜绿假单胞菌菌株测定特异性,以49株铜绿假单胞菌阳性菌液稀释成不同梯度测试灵敏度。结果表明:该方法特异性、灵敏度符合要求,检测限可达4.5CFU/mL,且简化了测试流程,使检测时间显著缩短。认为:所建立的纺织品中铜绿假单胞菌快速检测方法鉴别准确、快速,尤其适用于基层检验、检疫机构。
The rapid detecting method for pseudomonas aeruginosa in textiles was discussed.Immunomagnetic enrichment method and loop-mediated isothermal amplification were combined.The immunomagnetic enrichment method was adopted to enrich pseudomonas aeruginosa in samples.DNA was extracted from enriched liquid.Specific amplification was performed by loop-mediated isothermal amplification.The specificity was tested by forty-nine pseudomonas aeruginosa bacterial strain and twenty-one non-pseudomonas aeruginosa bacterial strain.Positive bacteria solution of forty-nine pseudomonas aeruginosa was diluted into different gradients to test sensitivity.The test results show that the specificity and sensitivity of the test method can meet the requirement.The detecting limitation can reach 4.5 CFU/mL.Moreover,the testing flow is simplified and the detecting time is obviously reduced.It is considered that the established detecting method for pseudomonas aeruginosa in textiles is accurate and rapid.It is especially suitable for basic inspection and quarantine organization.
The rapid detecting method for pseudomonas aeruginosa in textiles was discussed.Immunomagnetic enrichment method and loop-mediated isothermal amplification were combined.The immunomagnetic enrichment method was adopted to enrich pseudomonas aeruginosa in samples.DNA was extracted from enriched liquid.Specific amplification was performed by loop-mediated isothermal amplification.The specificity was tested by forty-nine pseudomonas aeruginosa bacterial strain and twenty-one non-pseudomonas aeruginosa bacterial strain.Positive bacteria solution of forty-nine pseudomonas aeruginosa was diluted into different gradients to test sensitivity.The test results show that the specificity and sensitivity of the test method can meet the requirement.The detecting limitation can reach 4.5 CFU/mL.Moreover,the testing flow is simplified and the detecting time is obviously reduced.It is considered that the established detecting method for pseudomonas aeruginosa in textiles is accurate and rapid.It is especially suitable for basic inspection and quarantine organization.
引文
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[2]MUHAMMAD SALMAN,AAMIR ALI,ABDULHAQUE.A Novel Multiplex PCR for Detection of Pseudomonas Aeruginosa:A Major Cause of Wound Infections[J].Pakistan Journal of Medical Sciences,2013,29(4):957-961.
[3]BRADBURY R S,RODDAM L F,MERRITT A,et al.Virulence Gene Distribution in Clinical,Nosocomial and Environmental Isolates of Pseudomonas Aeruginosa[J].Journal of Medical Microbiology,2010,59(8):881-890.
[4]NIKBIN V S,ASLANI MM,SHARAFI Z,et al.Molecular Identification and Detection of Virulence Genes Among Pseudomonas Aeruginosa Isolated from Different Infectious Origins[J].Iranian Journal of Microbiology,2012,4(3):118-123.
[5]邸秀珍,王睿.耐碳青霉烯类铜绿假单胞菌耐药机制的研究现状[J].中国临床药理学杂志,2015,26(8):669-672.
[6]何小曼,曾繁启.免疫磁珠分离技术在微生物学检测中的应用[J].中国生物制品学杂志,2008,21(11):1030-1032.
[7]覃昱.应用免疫磁珠分离及LAMP技术快速检测配方奶粉中克罗诺杆菌[D].广州:南方医科大学,2014.
[8]CURTIS K A,RUDOLPH D L,OWEN S M.Rapid Detection of HIV-1 by Reverse-Transcription,Loop-mediated Isothermal Amplification(RTLAMP)[J].Journal of Virological Methods,2008,151(2):264-270.
[9]UEDA Shigeko,KUWABARA YOSHIHIRO.The Rapid Detection of Salmonella from Food Samples by Loop-mediated Isothermal Amplification(LAMP)[J].Biocontrol Science,2009,14(2):73-76.
[10]BARBARA H I,DAVID K.NAD-Dependent Inhibition of Protein Synthesis by Seudomonas Aeruginosa Toxin[J].Proceedings of the National Academy of Sciences of the United States of America,1975,72(6):2284-2288.
[11]刘伟,侯丽萍,赵良娟,等.环介导等温扩增技术检测食品中铜绿假单胞菌[J].食品研究与开发,2012(5):140-142.
[12]王欢,黄庆,史大川,等.基于颜色判定的环介导恒温扩增技术快速检测铜绿假单胞菌的研究[J].第三军医大学学报,2012(11):2264-2268.
[13]姜莉莉,樊兆斌,高文玉,等.水貂绿脓杆菌环介导等温扩增快速检测方法的建立[J].中国农学通报,2012,28(29):79-82.