市售生鲜鸡肉5 种主要食源性致病菌污染检测及分布概率评估

采用聚合酶链式反应技术对上海市52 个市场的208 份鸡腿、鸡爪、鸡翅、鸡胗、鸡胸肉中出血性大肠杆菌O157∶H7、沙门氏菌、志贺氏菌、单增李斯特菌、阪崎肠杆菌5 种食源性致病菌进行快速检测,以了解不同鸡肉食用部位各种致病菌的分布规律,并根据β概率分布确立最具代表性的部位作为评估靶点。结果显示：在208 份样品中,大肠杆菌O157∶H7和沙门氏菌未检出;阪崎肠杆菌在鸡翅中检出1 株;志贺氏菌检出2 株,分布于鸡翅、鸡胗;单增李斯特菌检出11 株,检出率达5.29%,且在鸡翅中检出率最高（4 株）,鸡腿（3 株）、鸡爪（3 株）部位检出率相当,鸡胗检出率最低（1 株）。     

多重荧光定量聚合酶链反应法检测肉制品中的3种食源性致病菌

摘要：目的 建立多重荧光定量聚合酶链反应法（quantitative polymerase chain reaction,qPCR）快速检测肉制品中沙门氏菌、单增李斯特菌、大肠杆菌O157:H7 3种食源性致病菌的方法。方法 根据沙门氏菌的invA基因、单增李斯特菌的hemolysin基因、大肠杆菌O157:H7的rfbE基因序列分别设计特异性引物及探针,通过优化反应体系,测定其灵敏度、特异性和重复性,建立了可同时检测上述3种食源性致病菌多重qPCR方法。结果 该方法只扩增3种靶细菌,对其他供试菌不扩增;沙门氏菌、单增李斯特菌、大肠杆菌O157:H7的检出限分别为101、102、102拷贝数/μL,并且拥有良好的重复性和特异性。人工污染的猪肉样品经SLE（Salmonella、Listeria monocytogenes and Escherichia coli 3种菌株的共增菌培养基）培养基富集8 h后,分别可以检测到初始菌液浓度为1.56×102 CFU/25 g的沙门氏菌,2.13×102 CFU/25 g的单增李斯特菌,2.32×102 CFU/25 g的大肠杆菌O157:H7。结论 所建立的多重qPCR灵敏度高、特异性强、重复性好,可同时检测肉制品中沙门氏菌、单增李斯特菌和大肠杆菌O157:H7 3种食源性致病菌,在提高食品安全和保护人类健康方面有重要意义。     

徐州市2007-2011年食品中食源性致病菌监测结果分析

目的 了解徐州市食品中食源性致病菌污染状况,为食源性疾病监测提供科学依据.方法 依据《全国食源性致病菌监测工作手册》对食源性致病菌进行监测.结果 2007-2011年间共监测630份食品,总检出率为15.08％:检出沙门菌、大肠杆菌O157:H7、单增李斯特菌、副溶血性弧菌、金黄色葡萄球菌;阪崎杆菌、志贺菌、创伤弧菌未检出.生肉类和水产品污染较严重,生肉以单增李斯特菌和沙门菌污染程度高,检出率为分别为18.28％和10.75％;水产品以副溶血性弧菌检出率最高,为23.08％.结论 徐州地区食品中存在食源性致病菌污染,卫生监督部门应加强食品安全管理.     

2012-2014年安康市食品风险监测概况

调查分析安康市食品污染状况,预防食物中毒暴发流行。按照《全国食源性疾病监测工作手册》的要求采样、检测、保存菌株、网络报告。共计监测672份样品,检出致病菌50株,总检出率7.44%。检出沙门、单增李斯特菌、金黄色葡萄球菌、蜡样芽孢杆菌、大肠杆菌O157∶H7,检出率分别为0.17%、3.21%、2.87%、6.76%、0.83%;阪崎肠杆菌、致泻大肠杆菌、大肠杆菌O104∶H4、志贺氏菌均未检出。安康市食品致病菌污染状况复杂,单增李斯特菌、金黄色葡萄球菌、蜡样芽孢杆菌检出率较高,尤其应注意单增李斯特菌引起的食物中毒。     

2005-2007年宝鸡市食源性致病菌污染监测与分析

目的了解宝鸡市食源性致病菌污染状况。方法按照GB/T 4789—2003食品微生物检验标准方法检测。结果2005-2007年共对302份食品进行食源性致病菌污染监测,检出致病菌86株,检出率为28.5%,其中单增李斯特菌52株,检出率为17.2%,副溶血性弧菌24株,检出率为38.1%,沙门菌8株,检出率为2.6%,出血性大肠埃希菌O157∶H72株,检出率为0.7%。5类食品中鲜冻水产、生畜禽肉、速冻食品致病菌检出率较高,分别为55.6%、30.7%、25.0%%,熟肉制品、生食蔬菜中也有检出,分别为7.8%、7.3%。结论鲜冻水产、生畜禽肉、速冻食品是食源性致病菌污染的主要食品,存在食物中毒隐患,应引起足够重视。     

天津市2015~2019年生鲜果蔬病原微生物污染排查分析

为排查天津市即食生鲜果蔬病原微生物污染情况,2015~2019年在生产环节和流通环节抽取生菜、番茄、黄瓜、苦菊、桃和梨等6种即食果蔬品种共计654批次样品,采用国家标准方法分析样品中菌落总数、大肠菌群、沙门氏菌、金黄色葡萄球菌、单增李斯特氏菌、蜡样芽孢杆菌和大肠杆菌O157:H7污染状况进行风险分析。结果显示,654批次样品中检出食源性致病菌135批次,检出率为20.6%,其中包括沙门氏菌1份,金黄色葡萄球菌6份,蜡样芽孢杆菌128份,其他致病菌均未检出。食源性致病菌检出率较高的样品多出现在超市和农贸市场,表明即食果蔬致病菌的污染易发生在流通环节。即食生鲜果蔬中食源性致病菌污染对消费者健康存在潜在的安全隐患,需加强采后流通环节污染防控,保障即食果蔬食品安全,防止食源性疾病暴发。     

汕头市2005—2007年食源性致病菌监测

目的了解汕头市2005—2007年食品中沙门菌、单核细胞增生性李斯特菌、肠出血性大肠杆菌（O157：H7）、副溶血性弧菌的污染情况。方法按“《广东省食源性致病菌监测计划》检测技术要求”的检验方法进行。结果在采集的237份食品样品中,共检出5份沙门菌、8份单核细胞增生性李斯特菌和13份副溶血性弧菌,检出率分别为2．50％、3．38％和32．50％。未检出肠出血性大肠杆菌（O157：H7）。以水产品和生肉食品食源性致病菌污染最为严重。结论应加强水产品和生肉食品食源性致病菌污染的监测。     

2009年嘉兴市食源性病原菌监测分析

目的监测2009年嘉兴市食品中致病菌污染状况。方法共采集275份生熟食品样品,分离沙门菌、单核细胞增生李斯特菌、大肠杆菌O157:H7、金黄色葡萄球菌和副溶血性弧菌。结果 75份生肉、水产品食源性病原菌总污染率45.3%,其中检出沙门菌6株,单核细胞增生李斯特菌12株,金黄色葡萄球菌4株,副溶血性弧菌18株。未检出大肠杆菌O157:H7。200份即食食品总污染率3.0%,以金黄色葡萄球菌污染为主。结论 2009年嘉兴市主要污染食品品种是冷藏冷冻生肉,污染的食源性致病菌以单核细胞增生李斯特菌和副溶血性弧菌为主。     

2004-2007年肇庆市食品中食源性致病菌监测与分析

目的了解肇庆市食品中食源性致病菌的污染状况和污染水平,初步确定高危食品的种类,为食源性疾病的监测提供科学的依据。方法按全国食品污染物监测网的工作手册,2004-2007年采集4个监测点的5大类食品(生肉、熟肉、水产品、生吃水产品和生吃蔬菜)共计325份,对沙门菌、单核细胞增生李斯特菌、大肠杆菌O157∶H7和副溶血性弧菌4种食源性致病菌进行监测分析。结果325份食品样品中,检出致病菌27株(8.31%)。其中沙门菌16株(4.92%),单核细胞增生李斯特菌10株(3.08%),大肠杆菌O157∶H7未检出,80份水产品中检出副溶血性弧菌1株(1.25%)。生肉的污染情况最为严重,检出率为11.35%,其次是熟肉,检出率为9.68%;水产品的检出率为5.45%,生食水产品的检出率为4.00%,生食蔬菜的检出率为2.56%。结论沙门菌和单核细胞增生李斯特菌对肇庆市食品的污染普遍存在。生肉、熟肉的污染尤为严重,是重要的高危食品。     

多重PCR法检测鲜切哈密瓜中3种食源性致病菌

建立可同时检测鲜切哈密瓜中的单增李斯特菌、鼠伤寒沙门氏菌和大肠杆菌O157:H7的多重聚合酶链式反应(polymerase chain reaction,PCR)检测方法。根据单增李斯特菌inl A基因、鼠伤寒沙门氏菌inv A基因、大肠杆菌O157:H7的wzy基因设计3对特异性引物。对多重PCR反应体系中的引物浓度、退火温度、Mg2+浓度、d NTP浓度进行了优化,并确定适宜的多重PCR反应体系及反应条件。结果表明:25μL反应体系,10×PCR buffer为2.5μL,Mg Cl2(25 mmol/L)为3.5μL,d NTPs(2.5 mmol/L)为2μL,inl A基因上下游引物(5μmol/L)为1μL,inv A基因上下游引物(5μmol/L)为1μL,wzy基因上下游引物(5μmol/L)为2μL,单增李斯特菌DNA模板为1μL,鼠伤寒沙门氏菌DNA模板为1μL,大肠杆菌O157:H7 DNA模板为1μL,ex Taq DNA聚合酶(5 U/L)为0.3μL,加dd H2O补足25μL。反应条件为95℃预变性3 min;94℃变性30 s,53.9℃退火30 s,72℃延伸30 s,32个循环;72℃延伸10 min。鲜切哈密瓜中人工接种的目标菌的灵敏度为单增李斯特菌2.7×104 CFU/g,大肠杆菌O157:H7 3.3×104 CFU/g以及鼠伤寒沙门氏菌3.8×104 CFU/g。该技术可为快速检测鲜切哈密瓜中病原菌污染度及其控制提供参考依据。     

OCCURRENCE OF ESCHERICHIA COLIO157:H7/O157, LISTERIA MONOCYTOGENES AND SALMONELLA SPP. IN BEEF SLAUGHTERHOUSE ENVIRONMENTS, EQUIPMENT AND WORKERS

Escherichia coli O157/O157:H7, Listeria monocytogenes and Salmonella spp. are major foodborne pathogens and the emergence of these pathogens has been reported in many countries. The aim of this study was to investigate contamination of the beef slaughterhouse environment, equipment and workers with E. coli O157/O157:H7, L. monocytogenes and Salmonella spp. For this study, 500 swab samples were taken from 19 different points in five privately owned slaughterhouses, their periphery, slaughterhouse equipment and slaughterhouse employees. The presence of E. coli O157:H7/O157, L. monocytogenes and Salmonella spp. was determined with the application of the immunomagnetic separation method. Our study showed that the swabs taken from the five slaughterhouses contained E. coli O157:H7 in the environment, equipment, abattoir workers and water with a frequency 0.31, 1, 1.42 and 0%, respectively; while E. coli O157 was evident in the environment, equipment, abattoir workers and water with a ratio of 15, 10, 10 and 0%, respectively; L. monocytogenes was detected in the environment, equipment, abattoir workers and water at a ratio of 4.37, 15, 5.71 and 0%, respectively; and Salmonella spp. occurrence in the environment, equipment, abattoir workers and water at a ratio of 3.43, 16, 11.42, and 0%, respectively. Implementing hazard analysis critical control point principles in work procedures would definitely reduce the gross contamination occurring in abattoirs.

PRACTICAL APPLICATIONS

This study has revealed the effect of personnel and equipment on the contamination routes of E. coli O157:H7, Listeria monocytogenes and Salmonella spp. in meat in slaughterhouses and showed that especially L. monocytogenes and Salmonella spp. may pose a higher risk than E. coli O157:H7 in slaughterhouses.     

Multiplex PCR for simultaneous detection of Salmonella spp., Listeria monocytogenes, and Escherichia coli O157:H7 in meat samples

A multiplex PCR method was developed for simultaneous detection of Salmonella spp., Listeria monocytogenes, and Escherichia coli O157:H7 in meat samples. DNA detection sensitivity for this method was 10(3) CFU/ml for each pathogen. When this protocol was used for the detection of each of the above pathogenic bacteria in spiked pork samples, 1 cell per 25 g of inoculated sample could be detected within 30 h. In the samples of naturally contaminated meat, Salmonella spp., L. monocytogenes, and E. coli O157:H7 were detected over the same time period. Excellent agreement was obtained for the results of multiplex PCR and the conventional culture method, which suggests that the multiplex PCR is a reliable and useful method for rapid screening of meat products for Salmonella spp., L. monocytogenes, and E. coli O157:H7 contamination.     

冰鲜鸡肉中3 种主要致病菌的共修复- 增菌条件研究

研究冰鲜鸡肉中鼠伤寒沙门氏菌(S. typhimurium)、单增李斯特菌(L.monocytogenes)和大肠杆菌(E. coli) O157:H7的共修复-增菌条件。通过传统技术和PCR技术相结合的方法,研究肉鸡屠宰浸烫脱毛工艺对3种目标菌的热激损伤及共修复-增菌条件,并利用共修复-增菌结果,对生产线和超市的冰鲜鸡肉样品中3种目标菌的污染状况进行调查。结果表明：肉鸡屠宰浸烫脱毛工艺条件能使部分S. typhimurium、L.monocytogenes和E. coli O157:H7处于亚致死状态;TSB-YE培养基对热损伤S. typhimurium、L.monocytogenes和E. coli O157:H7的修复效果最好,且对前两种菌的修复时间为2h,对后一种菌的修复时间为4h;TSB-YE培养基在16h对3种致病菌的共增菌效果最好;运用共修复-增菌条件,对80份实际样品中的3种致病菌检出率分别为：S. typhimurium 22.5% (18/80)、L.monocytogenes 11.3% (9/80)和E. coli O157:H7 18.8% (15/80)。     

猪肉加工、流通过程中主要食源性病原细菌的监测

猪肉安全是食品安全的一个重要组成部分,食源性病原细菌是猪肉安全的重点监测对象。选择猪肉加工、流通过程的主要环节,应用PCR技术对沙门氏菌、产单核细胞李斯特菌、空肠弯曲菌、结肠弯曲菌和大肠杆菌O157等主要食源性病原细菌进行跟踪监测。结果表明:在所监测的6个环节中,均存在不同程度的病原菌污染,其中取内脏环节污染率最高,达49%;在监测的病原菌中,结肠弯曲菌的污染率最高,平均污染率为8.4%。取内脏、冷藏和销售环节是猪肉发生病原菌交叉污染的关键点。     

A note on the incidences of Salmonella spp., Listeria spp. and Escherichia coli O157:H7 serotypes in Turkish sausage (Soudjouck)

The incidence of Salmonella spp., Listeria monocytogenes and Escherichia coli O157:H7 was determined in 100 Turkish sausage (soudjouck) samples collected from shops and markets in the Afyon province, Turkey. Salmonella spp. were detected in 7% of the samples. All of the isolates were S. enterica Paratyphi B. In addition, Listeria spp. were detected in 9% of the samples. Its distribution was 7% L. monocytogenes and 1% each of L. ivanovii and L. innocua. Serological study of the seven L. monocytogenes isolates showed that three of these were 1/2 ab, three were 5/6 ab and one was 1 ab. E. coli O157:H7 was not detected in any of the samples. The pH values of the samples ranged from 4.8 to 6.5. In conclusion, increasing number of listeriosis and salmonellosis cases in Turkey and the contamination levels found indicate that risk assessment and improved preventive measures are required for these sausages.     

鲜切果蔬中4 种病原微生物多重PCR检测技术

研发可同时检测鲜切果蔬中的单核细胞性李斯特菌、鼠伤寒沙门菌、金黄色葡萄球菌和大肠杆菌O157:H7的多重聚合酶链式反应（polymerase chain reaction,PCR）检测方法。根据单核细胞性李斯特菌inlA基因、鼠伤寒沙门菌invA基因、大肠杆菌O157:H7 wzy基因、金黄色葡萄球菌nuc基因设计及筛选出4?对引物。对多重PCR体系及条件进行优化。该方法对单核细胞性李斯特菌、鼠伤寒沙门菌、金黄色葡萄球菌和大肠杆菌O157:H7的检出限分别为3.5×106、1.6×105、2.4×105、4.8×105?CFU/mL。将优化的多重PCR方法对不同接种量富集后验证,结果表明,经过9?h富集后,该方法检出限为1?CFU/mL。该方法在鲜切莴苣、鲜切黄瓜、鲜切木瓜、鲜切哈密瓜中应用同样可扩增出4?条目标菌。因此,利用所建立的多重PCR方法对鲜切果蔬中侵染的病原菌检出限可达到1?CFU/g。该方法相较于传统的培养检测方法具有节约大量的劳力、试剂、时间等优点,检测时间也由原来的5～7?d缩短至9～11?h,对于企业或分析检验中心大批量样品的监测具有指导意义。     

Thermal inactivation of Escherichia coli O157:H7, Salmonella, and Listeria monocytogenes in breaded pork patties

ABSTRACT:  Decimal reduction times ( D -values) and thermal resistance constants ( z -values) for 3 foodborne pathogenic bacteria in formulated ready-to-eat breaded pork patties were determined with thermal inactivation studies. Meat samples, inoculated with Escherichia coli O157:H7, Salmonella , and Listeria monocytogenes cultures or uninoculated controls, were packaged in sterile bags, immersed in circulated water bath, and held at 55, 57.5, 60, 62.5, 65, 67.5, and 70 °C for different durations of time. The D - and z -values were determined by using a linear regression model. Average calculated D -values for E. coli O157:H7, Salmonella , and L . monocytogenes at a temperature range of 55 to 70 °C were 32.11 to 0.08 min, 69.48 to 0.29 min, and 150.46 to 0.43 min, respectively. Calculated z -values for E. coli O157:H7, Salmonella , and L. monocytogenes were 5.4, 6.2, and 5.9 °C, respectively. The results of this study will be useful to food processors to validate thermal lethality of the studied foodborne pathogens in ready-to-eat breaded pork patties.     

HYGIENIC PARAMETERS, TOXINS AND PATHOGEN OCCURRENCE IN RAW MILK CHEESES

In total, 71 samples of retail raw milk cheeses produced or imported in Belgium and samples of Belgian farmhouse cheeses were examined for cotiforms, β-glucuronidase positive Escherichia coli, Escherichia coli O157 , Staphylococcus aureus, Salmonella spp. , Listeria spp. and Listeria monocytogenes. The presence of staphylococcal enterotoxins was investigated on samples with S. aureus counts higher than 10³ cfu/g. The incidence of coliforms, β-glucuronidase positive E. coli and S. aureus was higher in soft than in blue veined, semi-hard, hard and fresh cheeses. Four mold-ripened soft cheeses were positive for E. coli O157. One of the 4 cheeses was positive for verotoxin VT2. Staphylococcal enterotoxins were detected in 1 soft redsmear cheese, which was positive for L. monocytogenes. L. monocytogenes was also detected in one fresh cheese . Salmonella was not detected in any of the 71 raw milk cheeses.     

Survey of retail alfalfa sprouts and mushrooms for the presence of Escherichia coil O157:H7, Salmonella,and Listeria with BAX,and evaluation of this polymerase chain reaction-based system with experimentally contaminated samples

BAX, a polymerase chain reaction (PCR)-based pathogen detection system, was used to survey retail sprouts and mushrooms for contamination with Escherichia coli O157:H7, Salmonella, Listeria spp., and Listeria monocytogenes. No Salmonella or E. coli O157:H7 was detected in the 202 mushroom and 206 alfalfa sprout samples screened. L. monocytogenes was detected in one sprout sample, and seven additional sprout samples tested positive for the genus Listeria. BAX also detected Listeria species in 17 of the mushroom samples. Only 6 of 850 PCR assays (0.7%) failed to amplify control DNA, and therefore reagent failures and the inhibition of PCR by plant compounds were rare. The sensitivity of the detection system was evaluated by assaying samples inoculated with 10 CFU of each of the pathogens. One hundred seventy-two alfalfa sprout samples were inoculated with E. coli O157:H7, and two sets of 130 samples were experimentally contaminated with Salmonella Enteritidis and L. monocytogenes. The frequency of detection depended on the protocols used for inoculation and culturing. Inoculation of samples with approximately 10 CFU from frozen stocks yielded detection rates of 87.5 and 94.5% for L. monocylogenes and Salmonella Enteritidis, respectively, in mushrooms. The corresponding rates for alfalfa sprouts were 94.5 and 76.3%. The E. coli O157:H7 detection rate was 100% for mushrooms but only 48.6% for sprouts when standard BAX culture protocols were used. The substitution of an overnight incubation in modified E. coli medium for the 3-h brain heart infusion incubation increased the rate of E. coli O157:H7 detection to 75% for experimentally contaminated sprouts. The detection rate was 100% when E. coli O157:H7 cells from a fresh overnight culture were used for the inoculation. Test sensitivity is therefore influenced by the type of produce involved and is probably related to the growth of pathogens in the resuscitation and enrichment media.     

2007年温州地区食源性致病菌污染调查分析

目的了解2007年温州地区食品中沙门菌、单核细胞增生李斯特菌、大肠杆菌O157∶H7、空肠弯曲菌、金黄色葡萄球菌、副溶血性弧菌、香港海鸥菌的污染状况,提高温州地区食源性疾病检测、预警和控制能力,有效地预防、预测食源性疾病的暴发。方法依据国家食源性疾病监测网2007年度工作手册进行。结果检测生鸡肉、生猪肉、生牛肉、生羊肉、散装熟肉制品、海水鱼、淡水鱼、冷菜等八大类,共检出各类病原菌74株。其中以金黄色葡萄球菌的总检出率为最高,检出20株,检出率达15.04%;其次为副溶血性弧菌,检出39株,检出率为9.75%;单核细胞增生李斯特菌和沙门菌分别检出9株和6株,检出率分别为3.77%和2.51%,未检出大肠杆菌O157∶H7、空肠弯曲菌、香港海鸥菌。结论温州地区各类食品存在一定的食源性致病菌污染,其中冷菜、海水鱼、散装熟肉制品是主要污染品种。