AP631炭疽噬菌体裂解蜡样芽胞杆菌的发现和鉴定

目的 AP631炭疽噬菌体裂解蜡样芽胞杆菌的发现和鉴定。方法通过革兰氏染色、菌落形态特征、噬菌体裂解试验、青霉素敏感试验、溶血试验、生化试验和蛋白质毒素结晶试验对炭疽疫点现场分离的5株菌进行蜡样芽胞杆菌的鉴定,通过PCR扩增、小白鼠毒力测定试验和MLST基因检测等方法对该5株菌和本实验室保存的3株被AP631炭疽噬菌体裂解的蜡样芽胞杆菌进行毒力和基因测定。结果确定了8株可被AP631炭疽噬菌体不完全裂解的蜡样芽胞杆菌。从疫点采集分离的5株菌青霉素敏感试验阴性,炭疽毒力基因PCR扩增阴性,排除了炭疽芽胞杆菌;蛋白质毒素结晶试验染色试验为阴性,排除了苏云金芽胞杆菌;根据溶血试验和生化反应等一系列鉴别实验鉴定为蜡样芽胞杆菌。8株菌的MLST基因检测发现5株菌有新的等位基因位点或ST型。结论本研究首次证明我国使用的炭疽芽胞杆菌诊断AP631噬菌体可以不完全裂解部分蜡样芽胞杆菌,存在非特异性裂解现象,提示在今后使用噬菌体鉴定炭疽杆菌时,对于外环境分离到的可疑芽胞杆菌,特别要注意关于噬菌体不完全裂解的问题,这将为炭疽诊断标准的正确制订提供新的认识并具有重要的指导意义。     

TaqMan-LNA探针多重实时荧光定量PCR检测炭疽芽胞杆菌的研究

目的设计检测多个靶基因的炭疽多重荧光定量PCR反应系统,建立快速准确的基因诊断方法,解决蜡样芽胞菌群基因诊断交叉反应问题。方法选择炭疽杆菌PX01毒素质粒基因pagA,PX02毒素质粒基因capB,以及染色体上的前噬菌体λBa03基因PL3作为靶基因,利用在线荧光定量PCR引物设计软件设计、优化多重PCR引物以及相应的TaqMan探针序列,对探针进行LNA标记并优化荧光PCR反应体系,构建多重实时荧光定量PCR反应系统。以3对引物PCR扩增并克隆靶基因的质粒标准品,倍比稀释后进行敏感度试验,以炭疽、蜡样芽胞杆菌、苏云金杆菌、蕈状杆菌以及常见肠道致病菌进行特异度试验,利用炭疽弱毒株污染土壤及奶粉进行模拟检测。结果 TaqMan-LNA多重实时荧光定量PCR可迅速检出3个炭疽杆菌靶基因,常用的探针法荧光定量PCR反应液均适用;pagA、CapB、PL3基因克隆标准品序列正确;该方法对pagA、CapB、PL3基因的检出限分别为63、398、114copies/μl,且仅检出炭疽杆菌基因,其余4种蜡样芽胞菌扩增均阴性。土壤及奶粉样品检测均阳性,其中pagA近检出限为6.6×104/ml,PL3基因检出限为6.6×105/ml。结论 TaqMan-LNA多重实时荧光定量PCR方法具有适用性广,检测迅速,灵敏度及特异度高等特点,能够将炭疽杆菌与蜡样芽胞菌群中其他细菌相鉴别,可试用于炭疽杆菌的感染检测。     

市售婴幼儿食品蜡样芽胞杆菌污染状况调查

目的 了解甘肃省市售婴幼儿食品蜡样芽胞杆菌污染状况.方法 2018-2020年随机抽取甘肃省14个地(州、市)的百货商场、便利店/零售店、超市和网店等场所在售婴幼儿食品640份,应用平板计数法检测蜡样芽胞杆菌,以荧光PCR方法检测呕吐型基因.结果 640份样本中,蜡样芽胞杆菌总阳性率为15.00％(96/640),其中婴儿配方食品17.98％(41/228)、较大婴儿和幼儿配方食品18.64％(22/118)、婴幼儿谷类辅助食品11.22％(33/294);网店阳性率最高26.04％(25/96),呕吐型蜡样芽胞杆菌阳性率为6.25％(6/96);定量结果大于100 cfu/g的阳性样本占16.67％(16/96).结论 甘肃省市售婴幼儿食品存在蜡样芽胞杆菌污染,有潜在食品安全隐患,建议相关监管部门严格监督检查.     

蜡样芽胞杆菌溶血性肠毒素的提取及生物学活性分析

蜡样芽胞杆菌是国内引起细菌性食物中毒的常见细菌之一, 提取其溶血性肠毒素有助于阐明它的发病机理,有可能制备诊断试剂, 从而达到预防和控制蜡样芽胞杆菌食物中毒的目的。从国内引起食物中毒的１００ 株蜡样芽胞杆菌中,选出试验用菌株。从培养后的上清液中, 经过盐析、离子交换柱层析、制备性凝胶电泳提取蜡样芽胞杆菌溶血素。溶血素由 Ｂ、 Ｌ１ 和 Ｌ２ ３ 种成份组成, 分子量分别为３５ 、３６ 和４５ｋ Ｄａ ３ 种成份的协同作用在血琼脂上引起靶状溶血, 使兔皮肤血管通透性增加, 兔小肠肠段结扎产生积液反应, Ｖｅｒｏ 细胞形态学改变。提示蜡样芽胞杆菌溶血素具有肠毒素的性质, 试验也证实所提取的溶血素有免疫原性和免疫反应性     

贵州省2006-2011年炭疽芽胞杆菌检出情况与致病性研究

目的了解贵州省2006—2011年炭疽芽胞杆菌感染和分布情况,为贵州省炭疽疫情的预防控制和炭疽病09分子流行病学研究提供科学依据。方法对来自贵州省2006—2011年不同地区09830份疑似炭疽标本（外环境标本667份、患者标本151份和牲畜标本12份）,采用传统的革兰染色镜检、普通营养琼脂平板和血琼脂平板培养基分离培养炭疽芽胞杆菌,运用青霉素抑制试验和噬菌体裂解试验对可疑炭疽芽胞杆菌菌落进行鉴定。采用Real—time PCR技术检测88株经传统细菌学方法鉴定为炭疽芽胞杆菌菌株pX01质粒上的PA基因和pX02质粒上的CAP基因。结果从830份标本中分离出88株炭疽芽胞杆菌,总检出率为10．60％。2007年分离出炭疽杆菌的阳性标本最多,占36．36％;其次为2009年,占29．55％。阳性标本主要分布在黔西南州,其次为毕节地区和黔南州;册亨县、织金县和望谟县为炭疽杆菌检出数较多的监测县。88株炭疽芽胞杆菌CAP基因均为阳性。除2007年2株炭疽菌株PA基因为阴性外,其余86株PA基因均为阳性。结论贵州省炭疽疫源地面积广大,污染严重;检出的88株炭疽芽胞杆菌中97．73％的菌株同时具有两种毒力质粒,具有强致病性;该研究结果对贵州省炭疽疫情的预防控制及分子流行病学研究具有重要意义。     

小切口非超声乳化白内障摘除联合人工晶体植入术后感染性眼内炎的临床分析

目的探讨基层医院小切口非超声乳化白内障摘除联合人工晶体植入术后感染性眼内炎的危险因素、治疗和预后。方法回顾性分析2005年5月～2010年7月本院收治的8例(8只眼)感染性眼内炎患者的临床资料,分析感染性眼内炎的危险因素、病程、治疗及预后。结果 8例感染性眼内炎患者主要临床表现为眼红痛和视力障碍。8例患者中3只眼接受抗生素等药物综合治疗,4只眼行玻璃体切除、晶状体后囊膜切除及人工晶体取出术联合综合药物治疗,1例行眼球摘除术。6例患者保存了视力,1例眼球萎缩。细菌培养1例为表皮葡萄球菌感染,1例为肺炎链球菌感染。结论糖尿病患者白内障术后感染性眼内炎的发生几率较大,眼外伤、恶性肿瘤也可能是白内障术后眼内炎发生的危险因素,玻璃体手术和规范用药是有效的治疗手段。     

山东省一起皮肤炭疽疫情来源炭疽芽胞杆菌的分子分型及基因溯源分析

目的 对一起皮肤炭疽暴发疫情来源炭疽芽胞杆菌开展分子分型及基因溯源分析，为炭疽的有效防控提供重要的基础性数据。方法 本研究对2021年山东省曹县一起炭疽疫情样本中分离的8株炭疽芽胞杆菌进行药物敏感性实验、MLVA-15和canSNP分型、全基因组测序(WGS)和SNPs分析。结果 药敏结果显示，所有分离株耐药性一致。8株菌MLVA-15分型为同一基因型，该基因型在国内首次发现，命名为MLVA15-CHN77型；canSNP分型均为A.Br.001/002型。WGS结果分析显示，8株菌具有相同的耐药基因和毒力基因谱。SNPs分析显示，8株暴发菌株聚成一簇。结论 基于全基因组序列的分型和溯源策略，是炭疽芽胞杆菌传统分型方法的必要补充。     

洋快餐引起的蜡样芽孢杆菌食物中毒

本文报道一起由蜡样芽孢杆菌污染食物引起2人食物中毒的事件。根据流行病学调查,临床症状,实验室检验结果,确认该起食物中毒是由外带洋快餐受蜡样芽胞杆菌污染所致。     

甘肃省居民米面及节令食品中蜡样芽孢杆菌污染现状调查分析

目的了解甘肃省居民米面及节令食品中蜡样芽孢杆菌污染状况,为食品安全性风险监测提供依据。方法按《全国食源性致病菌监测网工作手册》中蜡样芽孢杆菌检验标准操作程序操作。结果 2009~2011年甘肃省地方特色米面食品和节令食品中蜡样芽孢杆菌检出率分别为3.33%、15.65%、7.68%,总检出率为8.26%;地方特色米面食品蜡样芽孢杆菌检出率为13.03%,显著高于节令食品的2.20%;第一、四季度蜡样芽孢杆菌检出率显著高于第二、三季度。结论甘肃省地方特色米面食品蜡样芽孢杆菌污染较为严重,提示甘肃省食品监管部门对此类食品应加强监督管理;今后应加强对蜡样芽胞杆菌在食物中的含菌量及产毒状况的监测研究。     

辽宁省炭疽芽胞杆菌MLVA分型研究

目的 了解辽宁地区炭疽芽胞杆菌的流行特征及菌型基因特征。方法 通过多位点可变数目串联重复序列(Multiple locus variable numbers of tandem repeats analysis,MLVA)分型实验,对2001—2011年辽宁省分离到的6株炭疽芽胞杆菌分离株DNA进行检测,DNA指纹图谱使用BioNumerics 4.0 软件进行统计分析,得出聚类分析结果。结果 聚类分析发现,6株炭疽芽胞杆菌株可分为2个基因型。对于炭疽暴发而言,其可变数目串联重复序列遗传标记具有高度相似性。结论 炭疽芽胞杆菌基因组中的串联重复序列可作为炭疽芽胞杆菌基因分型的指标,在炭疽暴发事件中的病原体溯源上具有重要的意义。     

蜡样芽孢杆菌食物中毒的研究进展

蜡样芽孢杆菌是芽孢杆菌属的一种,会引起食物中毒,因此与人类关系密切。本文概述了蜡样芽孢杆菌的病原学特征、致病性及其引起食物中毒的流行病学特征,以提出科学的防控措施来有效控制该菌在食品中的污染,保护公众健康。     

The incidence of Bacillus cereus in foods in Central Thailand

Thirty of 53 different raw and cooked or processed foods obtained in Central Thailand were demonstrated to contain Bacillus cereus. The level of contamination was found to depend on the type of food sampled; the mean was 3.2 X 10(6) organisms/gm on raw and cooked or processed foods ready for consumption, and 2.8 X 10(8) organisms/gm on cooked or processed foods kept overnight at kitchen temperature. B. cereus was commonly found on rice (both uncooked and boiled or fried), dried chilli pepper, shrimp paste, and on certain kinds of cooked foods seasoned with these products. Most of the total of 275 B. cereus isolates were found to possess biochemical reactions typical for B. cereus. Variations were also examined among several isolates. Nineteen per cent of the isolates did not produce acid from salicin. Certain types of the food tested were also demonstrated by volunteer experiments to carry enterotoxigenic B. cereus.     

ACE和α-adducin基因联合作用和左心室肥厚

目的探讨ACE I/D和-αadducin G/T基因联合作用和左心室肥厚的关系。方法聚合酶链反应(PCR)-聚丙烯酰胺凝胶电泳检测基因型,左心室重量指数(LVMI)衡量左心室肥厚程度。结果ACE基因III、D、DD三种基因型的左心室重量指数差异无统计学意义,-αadducin基因GG、GT、TT三种基因型的左心室重量指数差异无统计学意义,ACE和-αadducin基因联合基因型的左心室重量指数差异无统计学意义,但通过多元线性回归发现携带D等位基因的GG、GT、TT三种基因型左心室重量指数与基因型有关,TT型左心室重量指数最大,GG型最小。结论携带D等位基因的TT型高血压患者可能易发生靶器官损害(左心室肥厚)。     

Successful non-surgical treatment of brain abscess and necrotizing fasciitis caused by Bacillus cereus

Musculoskeletal and central nervous system infections caused by Bacillus cereus are very rare. Only a few cases have been reported, whose clinical courses strongly suggested that surgical procedures combined with appropriate antimicrobial therapy are necessary to cure these infections. A 60-year-old man with severe neutropenia due to myelodysplastic syndrome, developing necrotizing fasciitis and brain abscess caused by Bacillus cereus is reported. Without performing any surgical procedures, the patient was successfully treated with systemic antimicrobial therapy combined with granulocyte colony stimulating factor, which contributed to the increase in the neutrophil count.     

Association of H pylori cagA and vacA genotypes and IL-8 gene polymorphisms with clinical outcome of infection in Iranian patients with gastrointestinal diseases

AIM: To find out if a functional promoter polymorphism in the IL-8 gene along with cagA status and polymorphisms in vacA gene influence the type of diseases in Iranian patients infected by H pylori. METHODS: IL-8 -251 A/T polymorphism was genotyped by oligonucleotide allele specific PCR (ASO-PCR) in a sample of 233 patients with H pylori infection undergoing upper gastrointestinal endoscopy. The presence of cagA gene and polymorphisms in vacA gene was also determined by PCR. Association of these genetic polymorphisms with the development of gastritis, peptic ulcers as well as gastric cancer was tested. RESULTS: When the patients with different clinical manifestations were compared according to the presence of cagA gene or various vacA genotypes, only the vacA genotypes were significantly different among gastritis, peptic ulcer and gastric cancer patients (x2= 17.8; P=0.001). Furthermore, there was a significant difference in the frequency of IL-8 -251 A/T genotypes between patients with gastric cancer and benign diseases (x2= 10.47; P = 0.005). CONCLUSION: The IL-8 -251 A/T polymorphism and the polymorphisms in H pylori vacA gene are involved in limiting the infection outcome to gastritis and peptic ulcer or in favoring cancer onset in Iranian patients.     

2006-2016年贵州省炭疽芽胞杆菌SNR分型分析

目的 对贵州省2006-2016年分离的炭疽芽胞杆菌进行单核苷酸重复序列分型分析(SNR),了解菌株SNR型别特征,为炭疽疫情监测、调查与溯源提供技术手段和科学依据.方法 提取贵州省2006-2016年间不同地区的炭疽芽胞杆菌菌株基因组DNA,应用SNR各位点引物进行PCR扩增,将扩增产物进行毛细管电泳分析获得各位点序...     

基因探针和PCR方法在菌痢流行病学研究中的应用

目的 探讨基因探针和PCR方法在F2a志贺氏菌痢暴发的流行病学研究中的意义。方法 对43株自患者粪便和食物中分离到的F2a志贺氏菌进行16srRNA、ipaH基因Southern杂交，随机PCR和set1／set2毒力基因PCR分析。结果 随机PCR、ipaH基因Southern杂交将43株F2a志贺氏菌分为两个不同的基因型，set1／set2毒力基因PCR分为三个不同的基因型，16srRNA基因Southern杂交均为同一基因型。结论 基因分型方法能更准确、深入地揭示菌痢暴发过程中各分离株之间的流行病学联系。其中set1／set2毒力基因PCR方法具有较高的分辨率，在F2a志贺氏菌的分子流行病学研究中具有重要的作用。     

Bacillus cereus endocarditis: report of a case and review.

Bacillus cereus is a ubiquitous organism that infrequently causes serious infections. We report a patient with B. cereus endocarditis involving a mechanical aortic valve. Data for 10 cases of B. cereus endocarditis reported in the literature are summarized. B. cereus is resistant to many commonly used antibiotics, a finding that has clinical significance for empirical antibiotic selection in patients with suspected endocarditis. Infection in patients with valvular heart disease in the few cases reported is associated with significant mortality and morbidity.     

Comparative study of PCR-based Cryptosporidium discriminating techniques with a review of the literature

We identified the species or genotypes of the six Cryptosporidium isolates from patients and C. parvum strain HNJ-1 using the seven previously described species-differentiation and genotyping PCR protocols for detection of Cryptosporidium parasites. In addition, we also discussed about the usefulness of these PCR-based protocols on the basis of the reports previously published. Cryptosporidium diagnostic fragment was amplified by PCR with each primer pair, targeting the 18S ribosomal RNA (18SrRNA), Cryptosporidium oocyst wall protein (COWP). Heat shock protein 70 (HSP70), Polythreonine (Poly-T), Thrombospondin related adhesive protein of Cryptosporidium-1 (TRAP-C1), and unknown gene locus, in all isolates from patients and the strain HNJ-1. The RFLP profiles of 18SrRNA, COWP, HSP70, Poly-T, and TRAP-C1 PCR products in all isolates from patients were found to be the same among isolates, and were correspondent to those of C. parvum human genotype. While the RFLP profiles of HNJ-1 were strictly different from those of isolates from patients, and were correspondent to C. parvum cattle genotype. In addition, nucleotide sequences in 18 SrRNA gene of all isolates from patients and HNJ-1 were found to be identical to that of C. parvum, human or cattle genotype, respectively. Therefore, the isolates from patients and HNJ-1 were identified as C. parvum human and cattle genotype, respectively. According to the reports related to the PCR-based protocols applied in the present study, RFLP profiles targeting the HSP70, Poly-T, TRAP-C1 genes had been revealed in only a few species or genotypes, but those of 18SrRNA and COWP genes were in all species and genotypes. However, we supposed that it was difficult to distinguish between human or cattle genotype and other species or genotypes by RFLP profiles of 18SrRNA or COWP because the RFLP profiles of human or cattle genotype were identical or similar to those of other species or genotypes. On the other hand, it has been known that the nucleotide sequences in 18SrRNA or COWP gene are different among Cryptosporidium species and/or genotypes. Therefore, the direct sequencing method targeting the variable regions which can be used to distinguish among Cryptosporidium species, as well as the genotypes within C. parvum in either 18SrRNA or COWP gene is the most useful tool for accurate identification of Cryptosporidium isolates.