金黄色葡萄球菌肠毒素与多位点序列分型分子分型研究进展

金黄色葡萄球菌是引起临床疾病、社区感染和食物中毒的常见革兰氏阳性菌;肠毒素是参与金黄色葡萄球菌引起食物中毒、猩红热、全身感染等疾病的重要毒力因子;目前已发现27种金黄色葡萄球菌肠毒素/类肠毒素,不同来源金黄色葡萄球菌携带肠毒素/类肠毒素存在差异;多位点序列分型(MLST) 是一门新兴的基于细菌多个管家基因位点差异性分析、可长期监测细菌遗传进化趋势的分子分型技术;截至2021年初MLST已建立128种细菌的分型数据库,金黄色葡萄球菌数据库中已有6583种分型,金黄色葡萄球菌ST分型存在明显的地区流行性,各地已形成独特优势克隆。目前研究表明金黄色葡萄球菌肠毒素/类肠毒素携带与MLST分型间存在一定的相关性,本文将围绕金黄色葡萄球菌肠毒素/类肠毒素、MLST分型及相关性进行综述。     

噬菌体编码的金黄色葡萄球菌肠毒素A的 研究进展

金黄色葡萄球菌(Staphylococcus aureus)是一种人畜共患的食源性致病菌,并且能够产生不同种类的毒素,其中金黄色葡萄球菌肠毒素(staphylococcal enterotoxin,SE)是引起食物中毒的一类主要毒素。金黄色葡萄球菌肠毒素A(staphylococcal enterotoxin A,SEA)在食物中毒事件中的检出率最高,毒性最强,成为目前研究的热点。编码金黄色葡萄球菌肠毒素A的基因sea是由噬菌体携带并在菌体之间传播的。本研究主要综述了编码SEA的噬菌体与SEA之间的关系、SEA的分类、在不同的食品基质中环境因子对SEA产生量、sea基因相对表达量的影响及SEA新型的检测方法等,并介绍了目前金黄色葡萄球菌肠毒素研究的难点及研究方向的分析。     

金黄色葡萄球菌肠毒素在食源性微生物中的研究进展

由细菌污染引起的食源性疾病依然是影响人类公共健康和食品安全的最大问题之一。其中,金黄色葡萄球菌是人类的一种重要病原菌,引起许多严重感染。金黄色葡萄球菌属于革兰氏阳性球菌,广泛分布于自然界,是引起化脓性疾病的重要病原菌,也是引起食品污染和细菌性食物中毒的一种重要细菌。食品受金黄色葡萄球菌污染后,不仅会腐败变质,而且部分菌株产生金黄色葡萄球菌肠毒素（Staphylococcal enterotoxins,SEs）而引起食物中毒,由金黄色葡萄球菌肠毒素引起的食物中毒占整个细菌性食物中毒的首位。所以,对SEs的研究以及快速、精确的检测和筛查,成为关键环节。本文拟对金黄色葡萄球菌肠毒素生物学性状、致病．洼、检验方法等方面的研究进展进行综述。当然,对金黄色葡萄球菌及其主要致病因子肠毒素的研究有待进一步深入与发展。     

2009—2020年北京市朝阳区113株食源性金黄色葡萄球菌耐甲氧西林和肠毒素/类肠毒素基因携带与分子分型分析

目的 了解北京市朝阳区2009~2020年113株食源性金黄色葡萄球菌甲氧西林耐药、肠毒素/类肠毒素基因携带和分子分型情况。 方法 K-B法检测头孢西丁药物敏感性,PCR法扩增mecA基因;实时荧光定量PCR检测28种肠毒素/类肠毒素基因;脉冲场凝胶电泳(Pulsed field gel electrophoresis,PFGE)和多位点序列分型(Multilocus sequence typing,MLST)进行分子分型。 结果 检出7株MRSA(6.19%,7/113),6株为ST59,1株为ST9;所有菌株至少携带1种肠毒素/类肠毒素基因,最多携带14种,共检出24种;经典sea-see五型肠毒素携带率为45.13%。MLST共分24型,ST1、ST398、ST7、ST59为优势型;发现ST6656、ST6688、ST6709、ST6745、ST6754等5个新型别。PFGE共分25型,H型(11/113)、I型(9/113)、J型(29/113)是优势分型,占比43.36%。 结论 本地区食源性金黄色葡萄球菌MRSA菌株分离率低于临床株;PFGE结果显示本地区食源性金黄色葡萄球菌优势菌株集中;优势ST型别主要与医院、社区感染和动物相关,新发现五个ST型别;肠毒素/类肠毒素携带与ST型别存在一定关联性;ST1-MSSA- sea-seh-sek-seq-selw-selx是优势株;仅检测5种经典肠毒素不能代表金黄色葡萄球菌引起食物中毒的致病因子,应扩大肠毒素检测范围。     

食源性金黄色葡萄球菌肠毒素基因型分布研究

目的 采用PCR法扩增食源性金黄色葡萄球菌中肠毒素基因以了解该菌肠毒素基因携带情况,比较食物中毒和食品监测来源菌株中肠毒素基因检出率差异.方法 合成sea、seb、sec、sed和see五种肠毒素基因特异性引物,用常规PCR方法扩增食物中毒和食品监测来源菌株中各自肠毒素基因,同时采用mini-VIDAS检测食物中毒来源菌株中肠毒素.结果 110株菌株中有30株检出肠毒素基因,检出率为27.3％,肠毒素基因阳性菌株均只检出1种肠毒素基因.其中来自2起食物中毒的14株菌株均检出seb型肠毒素和相关基因,检出率为100％.来源于食品监测样本的96株菌株中有16株检出肠毒素基因,检出率为16.7％,包括sea型4株、seb型2株、sec型4株、sed型6株.结论 在宁波市食品监测中所分离的金黄色葡萄球菌所携带的肠毒素基因主要有sea、seb、sec和sed四型,而seb型肠毒素是引起金黄色葡萄球菌肠毒素所致食物中毒的主要因素.     

2003年～2015年金黄色葡萄球菌食物中毒事件特征分析

为了解2003年～2015年金黄色葡萄球菌引起的食物中毒事件发生规律,对中国知网收录的2003年～2015年金黄色葡萄球菌食物中毒暴发资料进行统计与分析。结果为共收集到发生在2003年～2015年间的金黄色葡萄球菌食物中毒事件86起,累计发病2 431人,死亡0人。肠毒素A（enterotoxin,sea）所引起的食物中毒事件最多,其次为肠毒素C（enterotoxin C,sec）。金黄色葡萄球菌所引起的食物中毒事件大多发生在夏、秋两季,然而食物中毒人数以春季最多,其次为夏季和秋季,冬季中毒人数最少。发生在集体食堂的食物中毒事件最多。肉及其制品引起的食物中毒事件数最多。因此,应针对重点人群、重点场所开展主动监测,指导合理加工或保藏食物,防止食物交叉污染或腐败变质,从而防止食物中毒事件的发生。     

金黄色葡萄球菌肠毒素及移动基因元件研究进展

金黄色葡萄球菌肠毒素是一种热源性的超抗原。食用被肠毒素污染的食物能够引起食物中毒,导致恶心、呕吐、腹痛有时伴随腹泻。本文综述了肠毒素的分类命名、理化性质、以及编码不同肠毒素的基因在移动基因元件中的存在情况。这些移动基因元件在金黄色葡萄球菌毒力传播和进化过程中起着至关重要的作用,了解它们对于金黄色葡萄球菌流行病学溯源以及理解毒力机制具有一定的指导意义。     

一起食物中毒事件中产独特肠毒素表型的金黄色葡萄球菌病原学分析

目的对一起疑似为金黄色葡萄球菌所导致的食物中毒事件进行葡萄球菌肠毒素检测,结合金黄色葡萄球菌病原学分析,为明确食物中毒诊断提供依据。方法根据流行病学调查,采用ELISA方法对可疑食物进行葡萄球菌肠毒素检测,同时对可疑食物和患者呕吐物进行金黄色葡萄球菌分离,运用Vitek2 Compact全自动细菌鉴定仪和血浆凝固酶试验鉴定为金黄色葡萄球菌,采用脉冲场凝胶电泳(PFGE)对病原菌进行同源性分析,以ELISA方法对检出的金黄色葡萄球菌菌株进行肠毒素检测,用PCR方法对肠毒素基因进行分型。结果食物和患者样品中分别分离出2和11株金黄色葡萄球菌,PCR方法及ELISA方法对肠毒素分型结果显示,其中12株同时存在SEA、SEB、SED、SEE 4种肠毒素及相关基因,PFGE聚类分析显示,其中12株产肠毒素金黄色葡萄球菌具有高度同源性。结论本起食物中毒事件为具有独特肠毒素表型的金黄色葡萄球菌导致,在金黄色葡萄球菌中毒实验室调查过程中,肠毒素检测结合病原菌溯源分析可以为相关公共卫生事件提供科学依据。     

Mini VIDAS与国标法检测金黄色葡萄球菌的比较及其在食物中毒中的应用

笔者使用Mini VIDAS方法和GB4789.10-2010第一法检测金黄色葡萄球菌,并对二者进行比较。GB4789.10-2010第一法(国标法),只检验金黄色葡萄球菌,对可疑食物中毒样品或产生葡萄球菌肠毒素的金黄色葡萄球菌菌株的鉴定,应检验其肠毒素。利用Mini VIDAS检测金黄色葡萄球菌肠毒素与检测金黄色葡萄球菌致病菌相比较,从检测时间、检出率、中毒判断等方面来说明检测肠毒素的重要性。Mini VIDAS检测金黄色葡萄球菌肠毒素检测时间一般为25.5小时,最快只需要90分钟,而GB4789.10-2010第一法检测金黄色葡萄球菌平均检测时间为78小时,而且检出率较前者低。可见,Mini VIDAS能在较短时间内直接检出金黄色葡萄球菌肠毒素,并克服了食物中毒时致病菌已死,无法检出,造成判断失误的弊端。     

蘑菇罐头生产中金黄色葡萄球菌肠毒素产生原因的研究

金黄色葡萄球菌肠毒素(简称肠毒素)引起的食物中毒,占整个细菌性食物中毒的50%左右.食用含肠毒素的食物后、发病快、一般     

Staphylococcal enterotoxins and enterotoxin-like toxins with special reference to dairy products: An overview

Staphylococcal Enterotoxins (SEs) have been raising health concerns for food safety due to their association with staphylococcal foodborne poisoning (SFP). As superantigens, they also cause the life threatening toxic shock syndrome (TSS), the transmission of which via food cannot be ruled out despite the lack of epidemiological evidence. To date, at least 23 of these exotoxins are known and separated into SEs and Staphylococcal Enterotoxin-like (SEl) depending on whether or not they invoke emesis. This work presents an up-to-date overview on the presently known SEs/SEls from the perspective of their classification, pathogenesis, and genetic organisation. The incidence of these toxins in dairy products, the risk this poses to the public health, and possible control means are also reviewed.     

Distribution of newly described enterotoxin-like genes in Staphylococcus aureus from food

Extensive analysis of the Staphylococcus aureus genome has allowed the identification of new genes encoding enterotoxin-like superantigens (SEls). Some of these are thought to be involved in staphylococcal food poisoning, while others do not elicit any emetic effect. The potential impact of these members of the enterotoxin-like family on the human organism seems to rely mainly on their superantigenic activity. In this paper the distribution of the genes coding for enterotoxin-like superantigens in S. aureus isolated from food was studied. Fifty isolates of S. aureus were examined and 27 were shown to be enterotoxigenic. Only 9 of the 27 strains carried genes encoding enterotoxins SEA-SEE. In 18 SEA-SEE-negative strains the presence of newly described enterotoxin genes was detected. All SEA-SEE-positive strains simultaneously carried genes of new SEls. We show that the gene encoding SElH (staphylococcal enterotoxin-like enterotoxin H) was the most frequently detected (n=14), while genes encoding SElI together with SElG accompanied by the other genes of the egc locus were detected in three strains. We also detected the presence of three less investigated genes: sep, sel, and sek. These genes were present in eight, two, and one isolate, respectively. In one strain, sep was accompanied by genes of other SEls, while in the remaining seven it was the only enterotoxin-like gene detected. The high prevalence of newly discovered enterotoxin genes, including the genes encoding emetic toxins, was demonstrated in food-derived strains. This supports the need for additional work on its role in food poisoning and, alternatively, to monitor its presence in S. aureus isolated from food. Our results suggest that yet unknown genetic elements encoding enterotoxin genes may exist.     

Dose-response Modelling of Staphylococcal Enterotoxins Using Outbreak Data

Staphylococcal food poisoning (SFP) is one of the most common food-borne diseases and results from the ingestion of staphylococcal enterotoxins (SEs). Yet, small amount of data are available for establishing a dose response. The objective of this work was to build a dose response relation based on the systematic investigations carried out during recent years in France. Over the period 2010-2014, more than 60 SFP outbreaks involving SEs, mainly from France, were microbiologically investigated. The enterotoxins were characterized as well as quantified. Attack rates, appearance times and natures of symptoms collected during epidemiological investigations were related to microbiological data. The outbreaks collected focused on enterotoxins SEA, SEB, SEC, and SED. Distribution of appearance times of symptoms and their natures were not influenced by the type of enterotoxins. The US EPA benchmark dose (BMD) methodology was then used to establish dose response. Attack rates of SFP outbreaks were modelled as a function of ingested doses and a BMD have been estimated for SEA.     

Incidence of Staphylococcus aureus and associated risk factors in Nham,a Thai fermented pork product

Staphylococcus aureus is one of the most prevalent bacterial pathogens causing food-borne disease worldwide. Staphylococcal food poisoning is caused by ingestion of staphylococcal enterotoxins (SEs) pre-formed in the implicated food. In this study, the incidences of S. aureus and classical SEs (SEA-SEE) contamination in ‘Nham’, a traditional Thai fermented pork product, were determined. Among 155 Nham samples tested, as high as 39.35% of the samples were positive for S. aureus (2–3500 MPN/g), but none were positive for the SEs. The risk factors for S. aureus contamination were highly correlated with the manufacturer and the pH of the product. A predictive model determined the probability of the presence of S. aureus to be ≤0.24 at the pH ≤ 4.6. During the fermentation process, the number of S. aureus slightly increased in the first day and decreased afterward. S. aureus counts continued to decrease when Nham was stored refrigerated. The negative result for enterotoxins and low counts of S. aureus in Nham surveyed in this study, and reduction of the pathogen counts during fermentation and storage suggested that there is very low risk of staphylococcal food poisoning from consuming properly fermented Nham.     

金黄色葡萄球菌肠毒素检测方法及前处理技术研究进展

金黄色葡萄球菌是常见的食源性致病菌之一,肠毒素是导致金黄色葡萄球菌食物中毒的最主要原因。现有肠毒素检测方法包括免疫学方法、分子生物学方法、生物传感器方法和质谱法等。各方法均存在优劣,酶联免疫学作为现有国家、行业标准推荐方法,虽然灵敏度高,但存在交叉反应,且价格昂贵。而质谱分析技术法具有高通量、专属性强等优点,已成为近年肠毒素检测技术研究热点。由于食品基质含有脂质,糖类等多种复杂成分,对肠毒素的分离与检测具有较强的干扰,因此选择恰当的样品前处理方法是实现肠毒素的准确检测的关键。本文分别从肠毒素性质、肠毒素检测方法及原理、样品前处理方法等方面对现有研究进展进行阐述。通过对现有技术方法的比较分析,基于高效液相分离技术结合质谱检测技术开发肠毒素检测方法,不仅能够从食品中实现多种肠毒素的高效分离富集,还具有灵敏度高、专属性强等优点,是未来肠毒素检测方法的主要研究方向。     

食物中毒事件中致病菌肠毒素检测和分子分型研究

目的 对一起食物中毒事件样本进行检测,查明食物中毒原因,并对分离菌株进行相关性分析。方法 参照国家标准方法GB4789.10-2016,对采集样本进行细菌学检验。应用酶联免疫吸附法(ELISA)检测葡萄球菌肠毒素。通过脉冲场凝胶电泳(PFGE)进行分子分型,分析菌株之间的相关性。结果 食物中毒事件中5名患者洗胃液、食物、分离菌株,均检测出E型葡萄球菌肠毒素。3株分离菌株PFGE分子分型提示来源不同克隆株,除了产生E型葡萄球菌肠毒素外,还有B、C、D型。结论 该起食物中毒由不同PFGE型别的产毒金黄色葡萄球菌污染食物引起,菌株产葡萄球菌肠毒素的型别不完全相同。金黄色葡萄球菌引起的食物中毒应关注多污染源及菌株肠毒素基因表达情况。     

The detection of enterotoxins and toxic shock syndrome toxin genes in Staphylococcus aureus by polymerase chain reaction

A simple polymerase chain reaction (PCR)-based procedure was developed for the detection of fragments of staphylococcal enterotoxins (SEs) SEA, SEB, SEC, SED, SEE, SEG, SEH, and SEI together with the toxic shock syndrome toxin (TSST-1) genes of Staphylococcus aureus. One hundred and twenty-nine cultures of S. aureus were selected, 39 of which were recovered from 38 suspected staphylococcal food-poisoning incidents. The method was reproducible, and 32 different toxin genotypes were recognized. The presence of SE genes was associated with S. aureus strains reacting with phages in group III, and the TSST-1 gene with phages in group I. There was a 96% agreement between the PCR results for detection of SEA-D and TSST-1 as compared with a commercial reverse passive latex agglutination assay for the detection of SEs from cultures grown in vitro. Enterotoxin gene fragments were detected in S. aureus cultures recovered from 32 of the 38 suspected staphylococcal food poisoning incidents, and of these, 17 were associated with SEE, SEG, SEH, and SEI in the absence of SEA-D. Simple PCR procedures were also developed for the detection of SE directly in spiked food samples, and this was most successfully achieved in mushroom soup and ham. Detection was less successful in three types of cheese and in cream. SEA or SEB were detected by enzyme-linked immunosorbent assay in three food samples (two of which were associated with food poisoning incidents) naturally heavily contaminated with S. aureus: the appropriate SEA or SEB gene fragments were detected directly in these three foods by PCR.