2003至2012年石家庄地区细菌性食物中毒病原菌特征研究

探讨石家庄地区细菌性食物中毒的特征和分布规律,为食物中毒事件的判定、预防和防控提供科学依据。收集2003至2012年石家庄地区细菌性食物中毒检验报告进行汇总分析。10a间石家庄地区共报告242起细菌性食物中毒,其中111起检出致病菌,检出率为45．87％（111／242）;单一致病菌和混合致病菌所致食物中毒分别占80．18％（89／111）和19．82％（22／111）。共检出7种病原菌,以变形杆菌和金黄色葡萄球菌占首位,其他还有沙门菌、蜡样芽胞杆菌、致泻大肠埃希菌、副溶血性弧菌、产气荚膜梭菌;食物中毒样品中以食品检出率最高,其次为患者粪便、剩余食品、呕吐物、涂抹物;食物中毒全年皆可发生,主要集中在4至9月份;食物中毒总体呈下降趋势。石家庄地区细菌性食物中毒病原菌呈多样性,以变形杆菌和金黄色葡萄球菌为主;食物中毒发生具有明显的季节性;近2a,沙门菌食物中毒有增多的迹象,应根据以上特点加强食品卫生监督管理,防控细菌性食物中毒的发生。     

食源性金黄色葡萄球菌脉冲场凝胶电泳分型分析

对石家庄地区食源性金黄色葡萄球菌进行脉冲场凝胶电泳(PFGE)分型,建立石家庄市金黄色葡萄球菌PFGE分型的数据库,探讨引起食物中毒金黄色葡萄球菌菌株和其他食源性金黄色葡萄球菌的分子特征。利用PFGE分型技术对2010至2012年石家庄市的75株食源性金黄色葡萄球菌进行全基因组分析,DNA酶切图谱用BioNumerics软件进行聚类分析。75株食源性金黄色葡萄球菌的PFGE图谱的相似性系数在54%～100%之间,经聚类分析得到45个PFGE型别,6起食物中毒的金黄色葡萄球菌分别由PFGE01型、PFGE02型、PFGE11型、PFGE12型、PFGE15型和PFGE45型引起。建立的金黄色葡萄球菌PFGE分型数据库分子型别较多,且存在差异明显的多个克隆系,对石家庄地区金黄色葡萄球菌引起食物中毒的防控和溯源工作有重要意义。     

PCR检测食品中金黄色葡萄球菌肠毒素基因

目的：检测食品中金黄色葡萄球菌肠毒素基因谱携带情况,探讨与食物中毒发病相关的金黄色葡萄球菌基因类型。方法：取食品中检出的金黄色葡萄球菌进行分离培养鉴定,通过PCR技术检测样本18种不同型的肠毒素基因的携带情况,并进行统计学分析。结果：食品中检出的金黄色葡萄球菌肠毒素基因有较高的阳性检出率,主要检出sea、seb、sei、seg、sel、sem、sen等基因,其中sea基因检出率最高,达到77.27%;sei、seg、sem、sen基因具有相关性,检出率为9.09%;seb、sel基因的检出率为4.55%。结论：在食品中主要检出金黄色葡萄球菌肠毒素sea、sei、seg、sem、sen、seb、sel等基因,其中,sei、seg、sem、sen基因的相关性还有待进一步深入研究。     

PCR技术检测食品中金黄色葡萄球菌肠毒素B基因

目的:金黄色葡萄球菌B型肠毒素是污染食品引起食物中毒的主要原因之一,针对进出口食品卫生监测的需要,研究一种简便、快速、准确的实验方法.方法:利用聚合酶链反应技术(PCR)采用特异的模板探针引物进行杂交,最后通过电泳技术与阳性对照进行比对,来判断阴阳性结果.结果:本方法检出率高,每克样品中有4个金黄色葡萄球菌即可检出,24小时即可报告结果.结论:PCR方法检测食品中金黄色葡萄球菌肠毒素B基因快速、准确,检测周期短,既可提高检出率,又可节省检测时间.     

探讨一起金葡菌引起食物中毒的检验结果分析

目的:探讨一起食物中事件中的致病因素以及与金葡菌的关系。方法:采集15份与食物中毒有关联的样本。根据GB4789.10-2010【食品安全国家标准食品微生物学检验金黄色葡萄球检验】对目的菌进行分离葡萄球菌及生化反应。结果:在二人呕吐物、三份豆沙、一份大米中检测出了金黄色葡萄球菌。结论:根据病人的临床反映、流行病学调查、以及微生物学检测的结果来分析,已经可以证实,这是一起由金葡菌感染而引起的食物中毒事件。     

金黄色葡萄球菌引起食物中毒的作用机制与其耐药性的研究进展

葡萄球菌广泛分布于自然界中,如空气、土壤、水以及物体的表面,在人和动物的皮肤表面部、鼻咽、肠道也常可发现葡萄球菌。大部分葡萄球菌是非致病菌,少数可引起人或动物致病,金黄色葡萄球菌（Staphylococcus aureus,金葡菌）即为最主要的致病性葡萄球菌。金葡菌是一种革兰氏阳性球菌,是医院感染常见的病原体之一,同时也是引起食品污染和细菌性食物中毒的一种重要细菌,其产生的毒素可使人中毒,带来非常严重的公共卫生负担。本文拟对金葡菌的病原与病理学特性,金葡菌与食物中毒,抗生素滥用与金葡菌耐药性等方面做简要综述。     

金黄色葡萄球菌引起食物中毒的作用机制与其耐药性的研究进展

葡萄球菌广泛分布于自然界中,如空气、土壤、水以及物体的表面,在人和动物的皮肤表面部、鼻咽、肠道也常可发现葡萄球菌.大部分葡萄球菌是非致病菌,少数可引起人或动物致病,金黄色葡萄球菌(Staphylococcus aureus,金葡菌)即为最主要的致病性葡萄球菌.金葡菌是一种革兰氏阳性球菌,是医院感染常见的病原体之一,同时也是引起食品污染和细菌性食物中毒的一种重要细菌,其产生的毒素可使人中毒,带来非常严重的公共卫生负担.本文拟对金葡菌的病原与病理学特性,金葡菌与食物中毒,抗生素滥用与金葡菌耐药性等方面做简要综述.     

沿海地区食物中毒病原分析

目的:探讨大连沿海地区引起食物中毒的病原菌。方法:选取近5年来大连沿海地区10起细菌性食物中毒病人急性中毒期的呕吐物、血液、粪便、可疑食物、使用过的器皿等标本进行病原学检测。结果:引起沿海地区食物中毒的主要致病菌为副溶血性弧菌、沙鱼弧菌、金黄色葡萄球菌肠毒素、类志贺邻单胞菌等。结论:为防止细菌性食物中毒的发生,应提倡居民及游客食用新鲜食物,煮熟后方可食用,相关食物加工场所应及时清洗和消毒,保持清洁,避免交叉感染。     

噬菌体JD007在即食牛奶中对金黄色葡萄球菌抑菌作用

目的评估噬菌体JD007作为食品添加剂在即食牛奶中防控金黄色葡萄球菌的效果,并与食品添加剂nisin抑菌效果作比较,以评估两者是否有协同抑菌作用。方法应用双层琼脂平板法对噬菌体滴度测定和扩增,应用超高速密度梯度离心对其进行纯化;使用稀释平板涂布法对细菌进行计数进而评估杀菌效果。结果噬菌体JD007对10株(共11株)分离自食品的金黄色葡萄球菌有杀菌作用,该噬菌体在铺有金黄色葡萄球菌的双层琼脂平板上可形成透明抑菌圈。噬菌体感染细菌的感染复数MOI=10、100时,无论是25℃还是4℃,在牛乳中都能显著抑制金黄色葡萄球菌生长,抑菌效果比nisin显著;而且噬菌体JD007和nisin在4℃有协同抑菌作用。结论噬菌体JD007对食品来源金黄色葡萄球菌有较广的杀菌谱,在室温和4℃时,能显著抑制牛乳中金黄色葡萄球菌的生长,具有作为食品防腐剂的潜能。     

金黄色葡萄球菌的检测方法优化及其在舟山市水产品加工厂中的污染状况调查

金黄色葡萄球菌(Staphylococcus aureus)是引起食物中毒的重要致病菌。为简化金黄色葡萄球菌复杂的检测方法,并了解舟山市水产品中该致病菌的污染状况,通过比较已公开的3套针对耐热核酸酶编码基因的PCR鉴定体系,根据特异性和灵敏性实验筛选了1套最优的针对金黄色葡萄球菌的PCR鉴定体系。基于这一体系和国家规定的标准方法,对舟山市水产品加工厂各个环节采集的120份样品中金黄色葡萄球菌的污染状况进行了调查。结果显示,120份样品中金黄色葡萄球菌的检出率为8.3%,PCR检测方法与国标法检出阳性率比较无显著差异,而且集中在原料和环境这两个环节检出金黄色葡萄球菌,在半成品和成品中却未检测到。由此,可以认为舟山市水产品加工厂的原料和环境中存在着一定的安全隐患,而成品相对较为安全。     

Structural elucidation of the extracellular and cell-wall teichoic acids of Staphylococcus aureus MN8m, a biofilm forming strain

Extracellular teichoic acid, an essential constituent of the biofilm produced by Staphylococcus epidermidis strain RP62A, is also an important constituent of the extracellular matrix of another biofilm producing strain, Staphylococcus aureus MN8m. The structure of the extracellular and cell wall teichoic acids of the latter strain was studied by NMR spectroscopy and capillary electrophoresis-mass spectrometry. Both teichoic acids were found to be a mixture of two polymers, a (1-->5)-linked poly(ribitol phosphate), substituted at the 4-position of ribitol residues with beta-GlcNAc, and a (1-->3)-linked poly(glycerol phosphate), partially substituted with the D-Ala at 2-position of glycerol residue. Such mixture is unusual for S. aureus.     

Structural evidence of α-aminoacylated lipoproteins of Staphylococcus aureus

Bacterial lipoproteins are known to be diacylated or triacylated and activate mammalian immune cells via Toll-like receptor 2/6 or 2/1 heterodimer. Because the genomes of low G+C content gram-positive bacteria, such as Staphylococcus aureus, do not contain Escherichia coli-type apolipoprotein N-acyltransferase, an enzyme converting diacylated lipoproteins into triacylated forms, it has been widely believed that native lipoproteins of S. aureus are diacylated. However, we recently demonstrated that one lipoprotein SitC purified from S. aureus RN4220 strain was triacylated. Almost simultaneously, another group reported that another lipoprotein SA2202 purified from S. aureus SA113 strain was diacylated. The determination of exact lipidated structures of S. aureus lipoproteins is thus crucial for elucidating the molecular basis of host-microorganism interactions. Toward this purpose, we intensively used MS-based analyses. Here, we demonstrate that SitC lipoprotein of S. aureus RN4220 strain has two lipoprotein lipase-labile O-esterified fatty acids and one lipoprotein lipase-resistant fatty acid. Further MS/MS analysis of the lipoprotein lipase digest revealed that the lipoprotein lipase-resistant fatty acid was acylated to α-amino group of the N-terminal cysteine residue of SitC. Triacylated forms of SitC with various length fatty acids were also confirmed in cell lysate of the RN4220 and Triton X-114 phase in three other S. aureus strains, including SA113 strain and one Staphylococcus epidermidis strain. Moreover, four other major lipoproteins including SA2202 in S. aureus strains were identified as N-acylated. These results strongly suggest that lipoproteins of S. aureus are mainly in the N-acylated triacyl form.     

Induced thermotolerance under nonisothermal treatments of a heat sensitive and a resistant strain of Staphylococcus aureus in media of different pH

AIMS: The aim was to assess the induced thermotolerance under nonisothermal treatments of two strains of Staphylococcus aureus in media of different pH. METHODS AND RESULTS: Staphylococcus aureus ATCC 25923 was more heat resistant than S. aureus ATCC 13565 at any pH investigated under isothermal conditions. At pH 7.4, the D58 value of the resistant strain was approx. 30 times greater. Both strains showed a higher heat resistance at pH 4.0 than at pH 7.4. In contrast, under nonisothermal treatments (0.5-2 degrees C min(-1)), both strains were more heat resistant when treated at pH 7.4 than at pH 4.0 due to heat adaptation at the higher pH. At the slowest heating up rate tested at pH 7.4, the initially heat-sensitive strain nearly reached the thermotolerance of the heat-resistant strain. CONCLUSIONS: The induced thermotolerance under nonisothermal treatments depended on the treatment medium pH and the microbial strain tested. The induced thermotolerance in a sensitive strain can be greater than in a heat-resistant strain, showing similar resistance under nonisothermal conditions. SIGNIFICANCE AND IMPACT OF THE STUDY: This work shows data of interest about mechanisms of microbial resistance and adaptation to heat. Moreover, it contributes to the development of more adequate combined processes for food preservation.     

Effect of combined physico-chemical preservatives on enterocin AS-48 activity against the enterotoxigenic Staphylococcus aureus CECT 976 strain

AIMS: Control of the enterotoxigenic Staphylococcus aureus CECT 976 strain by enterocin AS-48 in laboratory cultures, and behaviour of the AS-48 activity in the presence of food preservatives. METHODS AND RESULTS: Enterocin AS-48 shows inhibitory activity on the majority of the Staphylococcus species tested. This enterocin has a bactericidal and bacteriolytic mode of action on S. aureus CECT 976, a strain selected for this study by its enterotoxigenic character (SEA production). The inhibitory effect of AS-48 was pH and temperature dependent, and enterocin activity was higher at pH 5. The minimum bactericidal concentration (MBC) of AS-48, decreased from 15 microg ml(-1) at 37 degrees C to 10 microg ml(-1) at 15 degrees C. Sublethally injured cells showed an increased sensitivity with a MBC of 5 microg ml(-1). In this way, the highest effectiveness of Ent AS-48 against S. aureus CECT 976 was obtained at 4 degrees C in combination with high concentrations of NaCl (6 and 7%). Interestingly, enterotoxin SEA production by strain CECT 976 was markedly inhibited by subinhibitory concentrations of Ent AS-48. These low concentrations also provoked a delay of bacterial growth. CONCLUSION: The results presented indicated that Ent AS-48 has a potential for application as a protective agent against S. aureus in foods. SIGNIFICANCE AND IMPACT OF THE STUDY: In this study, we have established the conditions for an efficient inhibition of growth and enterotoxin production by S. aureus CECT 976 in culture media by a combination of environmental factors and Ent AS-48.     

Assessing and analysing contamination of a dairy products processing plant by Staphylococcus aureus using antibiotic resistance and PFGE

A dairy product processing plant was studied for 2.5 years to examine contamination with Staphylococcus aureus and try to correlate the source of contamination. Cultures were submitted to an antibiotic susceptibility test (AST) and characterised by Pulsed-field Gel Electrophoresis (PFGE) analysis. Results showed that 35.2% (19/51) of food handlers were asymptomatic carriers of S. aureus, and that 90.4% (19/21) of raw milk sampled was contaminated. Staphylococcus aureus was isolated from only 10 samples among more than 3200 investigated dairy products. No S. aureus contamination was found on machinery. The AST analysis demonstrated sensitivity of tested S. aureus to oxacillin, cephalothin, vancomycin, gentamicin, and sulfamethoxazole/trimethoprim. AST analysis generated eight different phenotypic profiles, but did not allow us to identify the source of contamination in seven of ten final products. PFGE analysis proved to be a sensitive method as it generated 42 different DNA banding profiles among the 48 S. aureus investigated, demonstrating a lack of predominance of endemic strains in the plant, contrary to suggestions raised by antibiotic resistance typing. Based on PFGE genotyping, S. aureus strains isolated from four contaminated final products were similar to four S. aureus isolated from raw milk. Five final products contained S. aureus different from all other strains collected, and one showed similarity to a strain isolated from a food handler. These results suggest contamination by raw milk as the main source of contamination of the final dairy products.     

Variation in resistance of natural isolates of Staphylococcus aureus to heat, pulsed electric field and ultrasound under pressure

AIMS: To study and compare the resistance of 15 Staphylococcus aureus isolates to heat, pulsed electric field (PEF) and ultrasound (UW) under pressure (manosonication, MS). METHODS AND RESULTS: Survival curves to heat (58 degrees C), to PEF (22 kV cm(-1), 2 micros square wave pulses) and to UW under pressure (117 microm, 20 kHz, 200 kPa) were obtained and inactivation parameters (decimal reduction times for heat and UW under pressure, and b-values for PEF) were calculated. A wide resistance variation to heat treatment, but not to PEF and MS, was observed amongst the 15 strains. CONCLUSIONS: There was no relationship between the resistances to the three physical agents studied. Staphylococcus aureus was relatively resistant to MS but sensitive to PEF. Heat resistance varied with strain and was positively correlated to carotenoid pigment content. SIGNIFICANCE AND IMPACT OF THE STUDY: Results would help in defining safe food preservation processes. Care should be taken to choose the most adequate strain of S. aureus to model food preservation processing.     

安徽不同地区奶站牛奶中金黄色葡萄球菌及其肠毒素污染状况评估

目的评定安徽地区各奶站牛奶中金黄色葡萄球菌以及肠毒素的污染情况。方法通过从安徽省不同地区30所奶站采集乳样,进行乳源性金黄色葡萄球菌的分离与生化鉴定,并采用PCR技术对分离出的菌株进行金黄色葡萄球菌肠毒素血清型鉴定。结果安徽省30个奶站中有4个地区奶站的牛奶中污染金黄色葡萄球菌;从污染牛奶中共分离出5株金黄色葡萄球菌,检出率为16.7%。经鉴定,所分离出的金黄色葡萄球菌中2株为肠毒素A型,1株为肠毒素C型,2株为同时产肠毒素A和肠毒素C。结论安徽省不同地区奶站中的牛奶污染的金黄色葡萄球菌产肠毒素类型以肠毒素A为主。     

Whole-genome sequence of Staphylococcus aureus strain LCT-SA112

Staphylococcus aureus is a facultative anaerobic Gram-positive coccal bacterium. S. aureus is the most common species of Staphylococcus to cause staphylococcal infections, which are very common in clinical medicine. Here we report the genome sequence of S. aureus strain LCT-SA112, which was isolated from S. aureus subsp. aureus CGMCC 1.230.     

Cefotaxime-hydrolysing activity of the β-lactamase of Klebsiella oxytoca D488 could be related to a threonine residue at position 140

The chromosomally encoded beta-lactamase of Klebsiella oxytoca D483 strain, active against all third-generation cephalosporins but ceftazidime, was purified to homogeneity. The pure protein was digested by trypsin, Staphylococcus aureus V8 protease or proteinase Asp-N. Amino acid sequences of the HPLC-separated proteolytic peptides were determined by manual Edman degradation. Overlapping fragments gave the alignment of the 263 residues of the beta-lactamase which presented 90% homology with the beta-lactamase of the K. oxytoca E23004 strain and about 40% homology with the other enzymes of the structural class A. The cefotaximase activity might result from interaction of a threonine residue at position 140 (position 165 in the numbering of Ambler) with the oxyimino group of the antibiotic.