TTC在乳及乳制品菌落总数检测中的应用

检测乳及乳制品菌落总数,为达到菌落在培养基中清晰可辨,在平板计数琼脂培养基中添加2,3,5-三苯基氯化四氮唑(TTC)。以灭菌乳、发酵乳、冰淇淋为样品基质,分别加入适当菌含量的大肠埃希氏菌、金黄色葡萄球菌、粪肠球菌、枯草芽胞杆菌工作菌悬液,制备适当浓度的TTC平板计数琼脂,同时以不添加TTC的平板计数琼脂作为对照,对同一样品进行菌落计数并同国标方法结果进行对比。TTC平板计数琼脂终浓度在0.001%~0.002%时能够使菌落显色,且与不添加TTC的对照组对比结果一致。在检测成分复杂的乳及乳制品样品时添加适宜浓度的TTC,有助于菌落的辨认,能够提高检验人员菌落计数的准确性,可用于检测乳及乳制品中菌落总数。     

采用ATP生物发光法分析6株常见细菌ATP含量差异

以ATP生物发光法为基础,辅以国标平板计数法,分析和测定了大肠埃希氏菌、枯草芽孢杆菌、蜡状芽孢杆菌、乳酸链球菌、乳酸片球菌等6株常见细菌中ATP含量的差异。结果显示:当单位体积菌悬液的ATP荧光值相同时,6株细菌菌落总数无明显差异;当菌悬液的菌落总数相同时,6株细菌的单位体积菌悬液ATP含量无明显差异。研究结果同时说明ATP生物发光法能够定量测定常见区域中的细菌总数。     

肉品中菌落总数和大肠菌群快速检测试纸片的研究

对肉品中菌落总数和大肠菌群快速检测试纸片进行了研究.选择中速定性滤纸为材料,分别浸入经不同特殊处理(不同浓度的TTC、琼脂、NaCl)的培养基中,40℃下烘干制得试纸片;将所制得的菌悬液涂布于成份不同的试纸片上,于32℃,培养20h(试纸片法),或涂布于平板计数琼脂上,于32℃,培养48h(国标法),然后计数,将二者得到的数据进行比较.结果表明:试纸片法与国标法具有很高的相似性.在100ml培养基中,TTC的浓度为0.02‰时试纸片的显色情况最好,生成的红色菌落数目最多.菌落生长受琼脂浓度的影响,当琼脂的浓度高于或低于0.15%时,细菌生长速度逐渐减慢.NaCl的浓度在0.4%时菌落生长呈现最好势态.     

两种不同检测方法分析不同类型天然和人工染菌食品中菌落总数的比较研究

目的：验证分析测试片检测不同类型食品菌落总数计数的适用性。 方法：与传统国标平板计数方法平行比较,应用测试片检测肉制品、面食、冷饮、奶制品、豆制品、果蔬和坚果7大类70份天然食品,以及以最常见的两种食源性微生物金黄色葡萄球菌和大肠埃希菌作为标准菌株,制备22的份不同食品基质的人工染菌样品。所有检测结果使用两种不同数据方法进行处理分析,以此探讨不同方法检测结果的相关性。 结果：依据ISO 16140-2 2016推荐的数据处理分析方法发现测试片和国标平板计数方法检测不同类型天然和人工染菌食品样品的菌落总数结果对数值差值的平均值均小于0.5;同时、单因素方差分析结果也证实两种方法检测结果无显著性差异（P>0.05）,相关系数高达0.938,不同方法检测结果之间呈正相关。以此发现本研究随机选择食品中菌落总数最高的分别为豆沙春卷、京酱肉丝和鲜猪皮。 结论：本研究证实测试片对不同类型食品菌落总数检测结果与国标平板计数方法结果一致性好,并且具有简单、快捷、易于标准化的优点,适用于不同层级食品承检机构。     

测试片法在食品菌落总数检测中的应用研究

目的探究菌落总数测试片法检测食品中菌落总数的可行性。方法用菌落总数测试片法和国家标准方法同时对制备的菌悬液、自然污染食品样品和人工污染食品样品的菌落总数进行检测,采用t-检验分析2种方法检测结果的差异,采用Pearson相关分析确定2种检测方法之间的相关性。结果菌悬液和自然污染食品样品菌落总数测试片法的变异系数为0.16%~3.76%,人工污染食品样品的变异系数为0.78%~8.61%;菌落总数测试片与国家标准方法在不同类型食品样品检测结果之间没有显著性差异(P<0.05),2种方法的检测结果呈正相关(r2>0.976,P<0.001)。结论菌落总数测试片法的检测效果与国家标准方法相当,可作为替代方法,用于检测食品中的菌落总数。     

应用干制培养基检测食品中金黄色葡萄球菌

目的建立金黄色葡萄球菌X干制培养基(Staphylococcus aureus X medium, XSA)检测食品中金黄色葡萄球菌的分析方法。方法依据GB 4789.10—2016《食品安全国家标准食品微生物学检验金黄色葡萄球菌检验》中前处理的方法制备样品,将样品添加到7.5%NaCl肉汤增菌后接种至干制培养基XSA上进行培养,通过鉴定实验进行定性检测和菌落平板计数进行定量检测,并用国标法同时进行定性及定量检测。结果在定性检测方面,该方法和国标方法假阴性率和假阳性率均为0%,但是检测时间由92 h缩短为48 h,在定量检测方面,该方法与国标法的对数偏差值的绝对值为0.3979<0.45,并且在6次检测同一个样品时的结果相对标准偏差较小,从而说明该方法和国标法的准确度相当,检测时长由78h缩短至24h,大大提高了检验效率。结论该方法操作简便快捷,结果稳定,适用于食品中金黄色葡萄球菌的检测。     

3种不同培养基和1种测试片检测水溶性化妆品中菌落总数的结果分析

目的使用t检验法分析3种不同培养基和1种测试片检测水溶性化妆品中菌落总数结果。方法参照《化妆品安全技术规范》(2015年版),使用卵磷脂吐温80营养琼脂、TTC营养琼脂、平板计数琼脂方法进行样品菌落总数检测,然后使用Petrifilm TM菌落测试片检测样品。运用配对t检验法评价4种不同方法对水溶性化妆品中菌落总数计数结果一致性的影响。结果在10~2、10~3 CFU/m L 2种不同浓度,平板计数琼脂法结果波动性最大,无法抑制菌落蔓延,而TTC营养琼脂和菌落测试片相对卵磷脂吐温80营养琼脂,具有抑制菌落蔓延生长、提高计数结果准确性的作用。结论在一定浓度下,可根据实际情况来选择相应培养基检测水溶性化妆品中菌落总数以提高检测效率和质量。     

手机拍照图像快速实现菌落计数

目的建立利用手机拍照, ImageJ软件自动实现菌落计数的方法。方法以大肠杆菌为测试菌,按照GB4789.2—2016《食品安全国家标准食品微生物学检验菌落总数测定》方法,获得菌落平板,手机拍照后,用Image J软件计数。并以井水、酒店预制菜、麻辣鱼仔熟食为样本比较了人工计数与Image J软件计数的差异性。结果 Image软件能对拍照后的菌落平板快速计数;人工计数和Image J软件计数结果无显著差异。结论本方法可重复性高,耗时短,克服了人工计数的缺点,保证检测溯源可查,有希望成为未来食品安全大数据体系数据采集端口。     

护手霜中菌落总数测定用培养基的探讨

检测化妆品菌落总数时,为使菌落在培养基中清晰可辨,允许在卵磷脂、吐温80一营养琼脂培养基中添加2,3,5-三苯基氯化四氮唑(以下简称TTC)。文章以护手霜为样品基质,分别加入适当菌含量的大肠埃希氏菌、金黄色葡萄球菌、蜡样芽孢杆菌、酵母工作菌悬液,制备5mg/100mL浓度的TTC卵磷脂、吐温80-营养琼脂,同时以不添加TTC卵磷脂、吐温80一营养琼脂平板进行验证,以营养琼脂培养基作为对照,对菌落计数结果进行对比。结论:添加了5mg/100mLTTC的培养基用于化妆品菌落总数检测时对G+有明显的抑制作用,建议在日常检测过程中需慎重选择。     

基于测试片法与平板计数法对不同类型食品中菌落总数结果的比较分析

目的比较菌落测试片法与平板计数法在不同类型食品中菌落总数测定结果的差异。方法在市面上随机抽取生肉制品、鲜奶制品、豆制品、粮食制品、冷冻饮品、坚果籽实、即食果蔬制品共112批,同时按菌落测试片法和平板计数法进行菌落总数测定,并对计数结果进行统计学分析。结果菌落测试片法与平板计数法的测定结果对数值差值的平均值|d log|= 0.06(<0.5),配对t检验P=0.860(>0.05)。结论菌落测试片法与平板计数法对上述7种类型的食品菌落总数测定结果无显著性差异,菌落测试片法可用于上述食品的菌落总数测定。     

Comparison of the compact dry TC and 3M petrifilm ACP dry sheet media methods with the spiral plate method for the examination of randomly selected foods for obtaining aerobic colony counts

Two hundred thirty-six randomly selected food and milk samples were examined to obtain aerobic colony counts by two dry sheet media methods and a standard Public Health Laboratory Service spiral plate method. Results for 40 samples were outside the limits of detection for one or more of the tested methods and were not considered. (The limits of detection for the spiral plate method were 200 to 1 x 10(8) CFU/ml for the spiral plate method and 100 to 3 x 10(6) CFU/ml for the dry sheet media methods.) The remaining 196 sets of results were analyzed further. When the results from the three methods were compared, correlation coefficients were all >0.80 and slopes and intercepts were close to 1.0 and 0.0, respectively. Mean log values and standard deviations were very similar for all three methods. The results were evaluated according to published UK guidelines for ready-to-eat foods sampled at the point of sale, which include a quality acceptability assessment that is based on aerobic colony counts. Eighty-six percent of the comparable results gave the same verdict with regard to acceptability according to the aerobic colony count guidelines. Both dry sheet media methods were comparable to the spiral plate method and can be recommended for the examination of food.     

Evaluation of the incubated plate count test for pasteurized milk

Factors were examined which may influence bacterial psychrotroph counts obtained by the European Community's directed preincubation test for pasteurized milk. Studies to determine the best protocol for sampling milk indicated that the least counting error was obtained by taking at least five consecutive samples of milk. Increasing the number of replicate plates or making subsamples did not decrease counting error. When milk samples from five commercial dairies were examined over an 11 month period there was a significant difference (p < 0·001) in counts obtained over the period, but this did not seem to be a seasonal effect. The counts between dairies were not significantly different nor was the position in the pasteurization run from which the samples were taken significant. The proportion of samples which failed the test standard was 9.7% over the period. The bacterial flora isolated from plate count agar were identified; Gram­negative rods were the most common isolates. The spiral plater system was compared with the pour plate method in the preincubation test and results were found to be in close agreement, suggesting that the spiral plater could be used to improve the current official methods. Varying the time and temperature of sample preincubation was found to give significantly different results in plate counts. This suggested that preincubation conditons were critical and that variation in conditions even within the specified ranges of the test could significantly alter the bacterial psychrotroph counts obtained. Because of these results, the validity of the statutory test was questioned.     

食品中金黄色葡萄球菌平板计数不确定度评定

为更好地评估食品中金黄色葡萄球菌指标。方法 按GB 4789.10—2010方法检验食品中金黄色葡萄球菌,按JJG 196—2006对菌落平板计数结果进行不确定度评定。结果 测量结果的扩展不确定度为U=0.072×lgT(T-食品中金黄色葡萄球菌菌落数),k=2。结论 测量不确定度评定可更为准确地评价食品质量,确保出厂食品的安全。     

Heterotrophic plate count methodology in the United States

In the United States (US), the history of bacterial plate counting (BPC) methods used for water can be traced largely through Standard Methods for the Examination of Water and Wastewater (Standard Methods). The bacterial count method has evolved from the original Standard Methods (1st edition, 1905) plate count which used nutrient gelatin and incubation at 20 degrees C for 48 h, to the HPC method options in the latest edition of Standard Methods that provide greater flexibility of application, depending on the data needs of the water analyst. The use of agar-agar as a gelling agent, replacing gelatin, allowed the use of higher incubation temperatures and resulted in the "body temperature count" (37 degrees C) found in the 3rd through the 8th edition of Standard Methods. The change from 37 degrees C incubation to 35+/-0.5 degrees C accommodated laboratories that did both milk and water analyses. By using a single temperature, fewer incubators were needed. The term "standard plate count" (SPC) first appeared in 1960 (11th edition) along with plate count agar. Incubation at 20 degrees C for the plate count was dropped from the 13th to 15th editions and few changes were made in the SPC method from the 11th edition through the 13th editions. Plate count analysis of bottled waters was included in the 14th edition (1975), calling for incubation at 35+/-0.5 degrees C for 72+/-4 h. Perhaps the most significant changes in plate count methods occurred with the 16th edition (1985). The term heterotrophic plate count replaced the standard plate count, and the spread plate (SP) and membrane filter (MF) methods were added along with new media for pour and spread plates (R2A agar and NWRI agar, both low nutrient) and for the membrane filter method (mHPC medium). The use of low nutrient media, lower incubation temperature, and longer incubation times, results in higher plate count results for most water samples. The options currently available, including low and high nutrient media, incubation temperatures (20 degrees C, 28 degrees C or 35 degrees C), plating methods (pour plate (PP), spread plate and membrane filter) and range of incubation times (24, 48, 72 h and 5-7 days) provide great flexibility in the application of the HPC analysis to drinking water.     

mini-MPN-STE-qPCR法快速定量检测食品中沙门氏菌

目的 建立一种快速、简易、可定量检测食源性沙门氏菌定量PCR(quantitative PCR, qPCR)方法。方法 依据沙门菌属invA基因序列设计染料法qPCR引物, 建立基于嵌合荧光法的沙门氏菌qPCR检测方法, 并通过短时间增菌(short time enrichment, STE)的5管微型多管发酵计数法（mini-most probable number, mini-MPN）进行定量检测, 构建mini-MPN-STE-qPCR法。使用人工污染的鸡肉混合液进行定量检测, 检测结果与传统MPN计数法和平板计数法进行比较分析。结果 建立的 qPCR法特异性良好, 方法灵敏度为50 CFU/mL, 结合48孔板min-MPN和4h 短时间增菌建立的mini-MPN-STE-qPCR法可将整个检测流程缩短至7 h。人工添加沙门氏菌的鸡肉混合液检测结果表明方法检出限为-0.347 log MPN/mL, Bland-Altman分析结果显示, 此法与传统MPN计数法相关系数R2=0.994, 与平板计数法相关系数R2=0.992。结论 该方法准确、简单易行、成本低、灵敏度高, 可用于食品中沙门菌的快速定量检测。     

A new medium for determining the total plate count in food

SimPlate for Total Plate Count-Color Indicator (TPC-CI, IDEXX Laboratories, Inc., Westbrook, Me.) is a new medium that incorporates the redox dye resazurin to detect and quantify bacteria in food. Enumeration is achieved by the most probable number method using a SimPlate device. Viable bacteria are detected in each well of the SimPlate device by the biochemical reduction of resazurin, which is blue, to the pink resorufin or the clear dihydroresorufin indicators. Results after 24 h of incubation for TPC-CI are highly correlated with standard plate count agar after 48 h of incubation. Correlation coefficients from studies conducted at five laboratories ranged from 0.94 to 0.98 in side-by-side comparisons against standard plate count agar. Four additional test sites, using alternative methods for determining the aerobic plate count in food, reported similar results in comparison studies (r = 0.91 to 0.97). The slopes from linear regression analysis at all sites ranged from 0.91 to 0.98, with y intercepts ranging from 0.11 to 0.84. Samples used for the validation of TPC-CI included raw food products (i.e., liver and grains), which may contain natural enzymes that interfere with enzyme-based detection methods. No interference was seen from the foods tested. These results suggest that TPC-CI is a suitable alternative to existing plate count methods and has reduced incubation time.     

Evaluation of various selective media for the detection of Pseudomonas species in pasteurized milk

Pseudomonas spp. are common gram-negative, post-pasteurization contaminants that contribute to spoilage of pasteurized dairy products. This study evaluated 5 common selective media for detecting Pseudomonas spp. in pasteurized milk. The performance of each selective medium for recovering 12 different Pseudomonas isolates (selected to represent a diversity of pasteurized milk isolates) was compared with that of standard plate count agar pour plates. Pseudomonas isolates showed varying abilities to produce colonies on different selective media. For 2 of 12 isolates, a 48-h incubation time was required for colony formation on any of the media tested. Violet red bile agar and coliform Petrifilm (3M, St. Paul, MN) were less effective than standard plate count agar pour plates at recovering Pseudomonas, regardless of incubation time, and MacConkey agar showed poor detection efficiency compared with SPCP after a 48-h incubation (R(2) = 0.26). Therefore, the use of violet red bile agar, MacConkey agar, or coliform Petrifilm may not be sufficient for detecting common Pseudomonas spp. in milk. The methods showing the highest detection efficiencies were crystal violet tetrazolium agar (CVTA) pour plates (R(2) = 0.95) and CVTA plates inoculated by spiral plating (R(2) = 0.89) incubated at 32 °C for 48 h. Overall, plating milk samples on CVTA followed by a 48-h incubation at 32 °C was the most effective selective method for recovering a diversity of Pseudomonas spp. from milk.     

PERFORMANCE AND MICROBIOLOGICAL GROWTH IN GROUND MEAT ASSOCIATED WITH THE USE OF THAWING TRAYS

The study goal was to determine whether trays marketed to hasten thawing of food would perform safely. Three brands of trays (Miracle Thaw, Super Defrost, and Defrost Master 2000) and aluminum skillets were purchased for experiments. Four frozen burgers with thermocouples inserted were placed onto a thawing device to thaw to 2.78C internal temperature. Experiments were replicated. Time and temperature (ambient, surface, internal) were recorded. Prethawing and thawed samples were analyzed for aerobic plate counts. Regression models were developed to identify sources of variation in time to thaw burgers, total bacterial count, and change in count pre/post thawing. All trays and aluminum skillets thawed burgers equally well (P>.05). Meat thawed within two hours (mean 83.6 min, maximum 119 min). Aerobic plate counts did not indicate spoilage of meat (mean 2372 cfu/g, maximum 17600 cfu/g). Emphasis on teaching home food preparers to refrigerate thawed foods within a two‐hour period is an educational priority.     

大肠菌群测量审核结果与分析

目的通过测量审核提升食品中大肠菌群检测能力和实验室质量管理水平。方法以测量审核作业指导书、GB 4789.3-2016《食品安全国家标准食品微生物学检验大肠菌群计数》第二法平板计数法为依据,采用平板计数法、PetrifilmTM大肠菌群测试片法同时检测样品。运用配对t检验法评价2种不同方法对乳粉中大肠菌群计数结果一致性的影响。结果平板计数法与PetrifilmTM大肠菌群测试片法检测结果无明显差异,样品|Z|2,结果满意。结论可根据实际情况来选择相应方法检测食品中大肠菌群以提高检测效率和质量。