化妆品中金黄色葡萄球菌检验能力验证方法及分析

文章依据《化妆品安全技术规范》2015年版和《NIFDC-PT-188化妆品金黄色葡萄球菌检验能力验证作业指导书》，分别对编号为TC1880001和TC18801010的两份样品中的金黄色葡萄球菌进行定性检测，并结合使用BAX?system Q7系统对增菌液进行初步筛查，同时使用VITEK 2COMPACT全自动细菌鉴定系统对分离出的菌株加以鉴定，试图通过外部能力验证，提高对化妆品中金黄色葡萄球菌的检验能力。最终结果表明编号为TC1880001和TC18801010的两个样品中金黄色葡萄球菌检验结果均为阴性。笔者对此次能力验证结果较为满意，并表示检测化妆品中金黄色葡萄球菌时具备常规法和实时荧光PCR法两种能力，能满足日常对化妆品金黄色葡萄球菌的检测监管需求。     

实验动物金黄色葡萄球菌的实验室检测能力验证结果评价

目的通过实施实验动物金黄色葡萄球菌检出能力验证计划,了解实验动物检测机构检验能力,提高实验动物质量检测水平。方法按照CNAS批准的能力验证方案,通过低温冻干制备样品,经过稳定性和均匀性检验合格,作为能力验证样品。采用随机编号,发样给参加单位,并附作业指导书。在规定时限提交检验报告和原始记录复印件,其结果与样品预检结果一致的判为满意结果,不一致或未能提交结果的判为不满意结果。结果共有28个实验室参加本次能力验证项目,其中获得满意验证结果的实验室为22个,占总参加机构的78.57%;未得到满意验证结果的实验室为6个,占21.43%。采用国标检测方法的有27个,采用PCR检测方法的有1个。结论实验动物质量检测机构对金黄色葡萄球菌的检测能力较高,实施能力验证计划能够反映实验室的检测水平。     

实验动物中沙门菌的实验室检测能力验证的结果与分析

目的通过实验动物中沙门菌检测能力验证计划,了解实验动物检测机构对沙门菌的检验能力,提高实验动物质量检测水平。方法按照CNAS批准的能力验证方案,通过冷冻干燥法制备含有沙门菌及干扰菌的实验动物粪便样品,经过稳定性和均匀性检验合格,作为能力验证样品。采用随机编号,经冷链运输发放给参加单位,并附作业指导书。在规定时限提交检验报告和原始记录复印件,其结果与样品预检结果一致的判为满意结果,不一致或未能提交结果的判为不满意结果。结果全国共有20个省市的30个实验室参加沙门菌能力验证项目,其中28个实验室获满意结果,占总参加机构的93.3%,不满意的2个实验室,占6.7%。采用分离培养方法的有29个实验室,采用PCR方法的有2个实验室。结论实验动物质量检测机构沙门菌检测能力较高,实施能力验证计划能够反映实验室的检测水平。     

兔出血症病毒抗体的实验室检测能力验证结果评价

目的通过兔出血症病毒抗体检测能力验证计划,了解实验动物检测机构检验能力,提高实验动物质量检测水平。方法按照CNAS批准的能力验证方案,通过筛选血清制备样品,经过稳定性和均匀性检验合格,作为能力验证样品。采用随机编号,发样给参加单位,并附作业指导书。在规定时限提交检验报告和原始记录复印件,其结果与样品预检结果一致的判为满意结果,不一致或未能提交结果的判为不满意结果。结果来自14个省市自治区的20个实验室报名参加本次比对实验,均在规定时间内反馈了检测结果,17个实验室检测结果为合格或优秀,占参加比对实验室的85%。在20个实验室中,14个实验室采用了酶联免疫吸附实验(ELISA)方法,6个实验室采用血凝抑制实验(HAI)方法。结论全国各实验动物检测机构兔出血症病毒抗体总体检测能力较高,实施能力验证计划能够反映实验室的检测水平。     

实验小鼠肾匀浆中酯酶-3的实验室测定能力验证结果评价

目的通过实验小鼠肾匀浆中酯酶-3项目的检测比对,了解全国实验动物质量检测实验室的水平,促进各实验室加强质控。方法按照CNAS批准的能力验证方案,制备样品并将稳定性和均匀性合格的样品作为能力验证样品,进行随机编号,随作业指导书一起发放给参加单位。在规定时限提交检验报告和原始记录复印件,其结果与样品预检结果完全一致的判为优秀结果;除杂合型判定以外的结果均一致的判定为满意结果,否则判为不满意结果。结果共10个实验室报名参加本次比对试验,其中优秀结果 0个实验室;满意结果的有9个实验室,占参加比对实验室的90.0%;不满意结果 1个实验室,占参加比对实验室的10.0%。结论本次能力验证项目反映了各参与实验室在实验小鼠肾匀浆中酯酶-3的总体检测能力较高,但检测细节及部分实验室技术水平还有待提高。     

实验小鼠肾匀浆中苹果酸酶-1和异柠檬酸脱氢酶-1实验室检测能力验证结果评价

目的通过实验小鼠肾匀浆中苹果酸酶-1和异柠檬酸脱氢酶-1检测能力验证计划,了解实验动物检测机构检验能力,提高实验动物质量检测水平。方法按照CNAS批准的能力验证方案,通过样品制备,经过稳定性和均匀性检验合格,作为能力验证样品。采用随机编号,发样给参加单位,并附作业指导书。在规定时限提交检验报告和原始记录复印件,其结果与样品预检结果一致的判为满意结果,不一致或未能提交结果的判为不满意结果。结果共有10个实验室参加本次能力验证项目,其中满意结果的实验室8个,占总参加机构的80%,不满意的实验室有2个,占20%。结论实验动物质量检测机构在实验小鼠肾匀浆中苹果酸酶-1和异柠檬酸脱氢酶-1的检测能力较高,实施能力验证计划能够反映实验室的检测水平。     

实验小鼠呼肠孤病毒Ⅲ型抗体的实验室检测能力验证结果评价

目的通过实验小鼠呼肠孤病毒Ⅲ型抗体检测能力验证计划,了解实验动物检测机构检验能力,提高实验动物质量检测水平。方法按照CNAS批准的能力验证方案,血清经过标定后,经过稳定性和均匀性检验合格,作为能力验证样品。采用随机编号,发样给参加单位,并附作业指导书。在规定时限提交检验报告和原始记录复印件,其结果与样品预检结果一致的判为满意结果,不一致或未能提交结果的判为不满意结果。结果共有17个省市的27个实验动物检测机构报名参加本次能力验证项目。27个检测机构均提交了满意结果,占总参加机构的100%。采用酶联免疫吸附实验(ELISA)方法的有26个检测机构,采用免疫荧光实验(IFA)方法的有1个检测机构。结论实验动物质量检测机构实验小鼠呼肠孤病毒Ⅲ型抗体检测能力较高,实施能力验证计划能够反映实验室的检测水平。     

欧盟转基因生物安全检测技术现状及启示北大核心CSCD

为了了解欧盟转基因生物安全检测技术发展现状,提高我国转基因安全监管水平。本研究根据对欧盟8家转基因检测相关机构现状的调查和分析结果,阐述了转基因检测过程中的关键技术要点及欧盟的实施情况,具体包括抽样与制样方法,转基因检测方法的循环验证与参数要求,样品的检测策略与实验室环境要求,以及检测结果的表述与不确定度评估。并结合我国转基因生物安全检测发展现状,提出了开展检测方法的实验室联合验证和加强转基因定量PCR检测技术的研发和应用的建议。     

阪崎肠杆菌能力验证样品均匀性和稳定性的研究

目的制备出以脱脂奶粉为基质的均匀性好、稳定性强的阪崎肠杆菌(Enterobacter sakazakii)能力验证样品。方法研究奶粉基质的粒度、奶粉和菌粉的混合比例,得到阪崎肠杆菌能力验证样品的最佳均匀性条件;研究包装形式及贮存温度对样品稳定性的影响,得到阪崎肠杆菌能力验证样品的最佳稳定性条件。结果要保证样品足够均匀,奶粉最佳粒径范围为120μmD180μm,1 g菌粉最多与300 g奶粉进行混合;贮存温度对样品稳定性有较大影响,高温明显降低样品稳定性;真空包装可以显著提高样品稳定性。结论以奶粉为基质的阪崎肠杆菌能力验证样品的制备能够更准确地考查乳品微生物检验人员的检测水平,为国内微生物能力验证水平的提高奠定了基础。     

欧盟转基因生物安全检测技术现状及启示

为了了解欧盟转基因生物安全检测技术发展现状,提高我国转基因安全监管水平。本研究根据对欧盟8家转基因检测相关机构现状的调查和分析结果,阐述了转基因检测过程中的关键技术要点及欧盟的实施情况,具体包括抽样与制样方法,转基因检测方法的循环验证与参数要求,样品的检测策略与实验室环境要求,以及检测结果的表述与不确定度评估。并结合我国转基因生物安全检测发展现状,提出了开展检测方法的实验室联合验证和加强转基因定量PCR检测技术的研发和应用的建议。     

多重PCR快速检测化妆品中三种致病菌的研究

利用多重PCR技术建立快速检测化妆品中三种致病菌的方法。根据已报道的大肠杆菌phoA基因、铜绿假单胞菌外膜蛋白基因oprL和金黄色葡萄球菌特异性序列SmaI选择特异性引物,对人工染菌化妆品进行多重PCR检测。结果显示,三种致病菌的基因组DNA均可与各自引物特异性结合,扩增产物大小分别为622 bp、504 bp和426 bp。该方法用于人工污染的化妆品中,大肠杆菌的检出限浓度为103 CFU/mL,铜绿假单胞菌和金黄色葡萄球菌的检出限浓度为105 CFU/mL。作者建立的多重PCR方法可同时快速、特异地对化妆品中三种致病菌进行检测,在化妆品行业具有较大的应用价值。     

Microbial quality of tropical inland waters and effects of rainfall events

Novel markers of fecal pollution in tropical waters are needed since conventional methods recommended for other geographical regions may not apply. To address this, the prevalence of thermotolerant coliforms, enterococci, coliphages, and enterophages was determined by culture methods across a watershed. Additionally, human-, chicken-, and cattle-specific PCR assays were used to identify potential fecal pollution sources in this watershed. An enterococcus quantitative PCR (qPCR) assay was tested and correlated with culture methods at three sites since water quality guidelines could incorporate this technique as a rapid detection method. Various rainfall events reported before sample collection at three sites were considered in the data analyses. Thermotolerant coliforms, enterococci, coliphages, and enterophages were detected across the watershed. Human-specific Bacteroides bacteria, unlike the cattle- and chicken-specific bacteria, were detected mostly at sites with the corresponding fecal impact. Enterococci were detected by qPCR as well, but positive correlations with the culture method were noted at two sites, suggesting that either technique could be used. However, no positive correlations were noted for an inland lake tested, suggesting that qPCR may not be suitable for all water bodies. Concentrations of thermotolerant coliforms and bacteriophages were consistently lower after rainfall events, pointing to a possible dilution effect. Rainfall positively correlated with enterococci detected by culturing and qPCR, but this was not the case for the inland lake. The toolbox of methods and correlations presented here could be potentially applied to assess the microbial quality of various water types.     

耐甲氧西林金黄色葡萄球菌及其肠毒素的检测和耐药性分析

目的评价乳胶结合试验检测耐甲氧西林金黄色葡萄球菌(MRSA)及其肠毒素(SE),并进行耐药性分析.方法收集130株金黄色葡萄球菌临床分离株,通过药敏试验将其分为耐甲氧西林金黄色葡萄球菌和甲氧西林敏感金黄色葡萄球菌(MSSA),用反向间接血凝试验(RPHA)检测金黄色葡萄球菌肠毒素.结果67株MR-SA产肠毒素,19株MSSA产肠毒素,MRSA产肠毒素率为100%,MSSA产肠毒素率为30%.结论实验室应重视金黄色葡萄球菌肠毒素的检测.     

Correlation of bacterial indicator organisms with Salmonella spp., Staphylococcus aureus and Candida albicans in sea water

The value of total coliforms, faecal coliforms and faecal streptococci in predicting the presence of Salmonella spp. and the numbers of Staphylococcus aureus and Candida albicans in sewage polluted coastal water were assessed. All indicators had strong positive association with Salmonella and moderate positive correlations with Staph. aureus and C. albicans. Total coliforms correlated better with salmonellas and Staph. aureus than did the two faecal groups. Regression analysis revealed that total coliforms have a better value as predictors of the presence of Salmonella and Staph. aureus , while faecal coliforms are better predictors of C. albicans , in moderately polluted areas. The conclusion reached is that enumeration of total coliforms is sufficient to predict the presence of Salmonella spp. or Staph. aureus in sea water moderately affected by sewage pollution, without the additional measurement of faecal coliforms and faecal streptococci.     

Evaluation of coliforms as indicators of water quality in India.

The total and thermotolerant coliform counts in rural drinking water derived from ground water, piped supplies and surface water are reported for a number of areas in India. To evaluate these counts as indicators of recent faecal contamination the total coliforms and thermotolerant coliforms isolated have been identified. Thermotolerant Escherichia coli formed 11.7% of the total coliforms and 75.1% of the thermotolerant coliforms. Citrobacter sp. (20.4%) and Klebsiella sp. (50.9%) were the other common total coliforms isolated and, among the thermotolerant coliforms, Klebsiella sp. (16.4%) was the only other organism frequently encountered. The total coliform counts were significantly correlated with water temperature. The applicability in tropical areas of standards developed for temperate climates is discussed.     

多重PCR检测奶牛乳腺炎金黄色葡萄球菌粘附素基因clfa A、clfa B、fnbp A和fnbp B

利用多重PCR检测金黄色葡萄球菌粘附素基因clfa A、clfa B、fnbp A和fnbp B的方法,对奶牛乳腺炎金黄色葡萄球菌临床分离株进行聚集因子主效基因的分析。通过设计合成的特异性引物对金黄色葡萄球菌模板进行PCR扩增,将目的基因回收并连接到T载体,鉴定后进行测序验证,然后对本实验室所分离鉴定的金葡菌临床分离株进行多重PCR检测。PCR产物经过电泳成像显示,clfa A和clfa B分别在292bp和205bp处出现特异性条带;fn-bp A和fnbp B分别在524bp和642bp处出现特异性条带。通过对29株金葡菌临床分离株多重PCR检测发现:能扩增出clfa A、clfa B、fnbp A和fnbp B的分别有26株、12株、28株和3株。建立的多重PCR检测金黄色葡萄球菌粘附素基因的方法具有良好的特异性和可靠性,并且发现clfa A和fnbp A基因存在于绝大部分的金黄色葡萄球菌中。     

Evaluation of coliforms as indicators of water quality in India

P.W. RAMTEKE, J.W. BHATTACHARJEE, S.P. PATHAK AND N. KALRA. 1992. The total and thermotolerant coliform counts in rural drinking water derived from ground water, piped supplies and surface water are reported for a number of areas in India. To evaluate these counts as indicators of recent faecal contamination the total coliforms and thermotolerant coliforms isolated have been identified. Thermotolerant Escherichia coli formed 11.7% of the total coliforms and 75.1% of the thermotolerant coliforms. Citrobacter sp. (20.4%) and Klebsiella sp. (50.9%) were the other common total coliforms isolated and, among the thermotolerant coliforms, Klebsiella sp. (16.4%) was the only other organism frequently encountered. The total coliform counts were significantly correlated with water temperature. The applicability in tropical areas of standards developed for temperate climates is discussed.     

Survival and transport of bacteria in egg washwater

An evaluation of methods for monitoring the quality of water used to wash eggs at grading stations was undertaken to improve maintenance of bacterial viability during overnight sample transport. Bacterial content of samples at analysis would then better reflect conditions at the time eggs were washed. The interactive effects of temperature and the highly alkaline water conditions upon viability were the subjects of this study. Nine transport methods were examined for their efficacy in recovering total and coliform bacteria from recycled water used to wash eggs, and these were compared with samples analyzed at two commercial egg grading stations. Samples were shipped under test to the laboratory for analysis the following day. The survival of Staphylococcus aureus and Escherichia coli was also examined, but in a synthetic washwater matrix under various combinations of temperature (6 to 32 degrees C) and pH (9.5 to 10.5) to determine whether there was likely to be a different response to variations in transport treatment among gram-positive and -negative bacteria. S. aureus was much more resistant to the lethal effects of high pH and moderate temperature than E. coli. These results indicated that samples of high pH should be held (transported) at less than or equal to 13 degrees C to optimize bacterial survival. Considering cost, ease of manipulation, and the ability to protect both coliforms and the bacterial population as a whole, the method of choice for transport of industrial samples was the direct addition of washwater to containers in which powdered KH2PO4 and Na2S2O3 had been placed to yield final concentrations, when dissolved, of 0.2 and 0.05% (wt/vol), respectively.     

Survival and transport of bacteria in egg washwater.

An evaluation of methods for monitoring the quality of water used to wash eggs at grading stations was undertaken to improve maintenance of bacterial viability during overnight sample transport. Bacterial content of samples at analysis would then better reflect conditions at the time eggs were washed. The interactive effects of temperature and the highly alkaline water conditions upon viability were the subjects of this study. Nine transport methods were examined for their efficacy in recovering total and coliform bacteria from recycled water used to wash eggs, and these were compared with samples analyzed at two commercial egg grading stations. Samples were shipped under test to the laboratory for analysis the following day. The survival of Staphylococcus aureus and Escherichia coli was also examined, but in a synthetic washwater matrix under various combinations of temperature (6 to 32 degrees C) and pH (9.5 to 10.5) to determine whether there was likely to be a different response to variations in transport treatment among gram-positive and -negative bacteria. S. aureus was much more resistant to the lethal effects of high pH and moderate temperature than E. coli. These results indicated that samples of high pH should be held (transported) at less than or equal to 13 degrees C to optimize bacterial survival. Considering cost, ease of manipulation, and the ability to protect both coliforms and the bacterial population as a whole, the method of choice for transport of industrial samples was the direct addition of washwater to containers in which powdered KH2PO4 and Na2S2O3 had been placed to yield final concentrations, when dissolved, of 0.2 and 0.05% (wt/vol), respectively.