我国市售婴儿配方乳粉中蜡样芽胞杆菌污染及其毒力基因调查

目的了解国内市售婴儿配方乳粉中蜡样芽胞杆菌污染及毒力基因分布情况。方法采用MPN法定量检测婴儿配方乳粉中的蜡样芽胞杆菌,在对分离菌株正确鉴定的基础上,开展腹泻型和呕吐型毒素产生相关毒力基因的分布研究。结果 135份婴儿配方乳粉中有57份检出蜡样芽胞杆菌,检出率为42.22%。平均污染水平为7.14 MPN/g。国产产品蜡样芽胞杆菌污染较进口产品重,网售产品较超市销售产品重,差异有统计学意义(P0.05)。共发现24种蜡样芽胞杆菌毒力基因携带模式,其中nhe基因携带率最高,达92.98%(53/57),其次为ent FM基因(71.93%),70.18%(40/57)的菌株同时携带nhe和ent FM基因。亚型分型结果显示nhe A、nhe B和nhe C基因的携带率分别为88.72%、88.72%和49.12%。溶血素BL基因携带率分别为hbl A 24.56%、hbl C 22.81%和hbl D 17.54%,cyt K基因携带率为22.81%。有8株菌既携带nhe的3个基因又携带hbl的3个基因。结论我国市售婴儿配方乳粉蜡样芽胞杆菌污染较重,分离到的蜡样芽胞杆菌菌株普遍携带毒力基因,建议加强对婴儿配方乳粉中蜡样芽胞杆菌污染的监管,并开展膳食暴露该菌对婴儿健康影响的风险评估,为制定婴儿配方乳粉中蜡样芽胞杆菌的限量标准提供依据。     

Growth and toxin profiles of Bacillus cereus isolated from different food sources

Eleven strains of Bacillus cereus isolated from milk and meat products have been used to study growth and sporulation profiles in detail. Polymerase chain reaction (PCR) using primers detecting cold shock protein A gene signatures (cspA), showed that none of the strains were the newly suggested species in the B. cereus group, B. weihenstephanensis, comprising psychrotolerant cereus strains, although one of the strains grew at 4 degrees C, two at 6 degrees C and seven grew at 7 degrees C. One of the two strains that grew at 6 degrees C had a maximum growth temperature of 42 degrees C, while the remaining 10 strains all grew at temperature of 43 degrees C or higher. Only three strains grew at 48 degrees C. At 42 degrees C, the generation time varied between 11 and 34 min. Spore germination was much faster for the two strains that grew at 6 degrees C than for the other nine strains in milk at 7 degrees C and 10 degrees C. All strains were cytotoxic and contained the non-haemolytic enterotoxin gene (nhe), 10 strains contained the enterotoxin T gene (bceT), and only six had the gene (hbl) encoding haemolytic enterotoxin. Two strains showed some microheterogeneity in the nhe operon. but contained all three genes. We can conclude that true B. cereus strains can have growth profiles as expected for B. weihenstephanensis, and that nhe and bceT were not correlated with growth profiles. However, the two psychrotolerant strains with minimal growth temperature of 4 degrees C and 6 degrees C did not contain hbl, as judged from our PCR results.     

糙米中蜡样芽孢杆菌的分离鉴定及其毒力基因与药敏性检测

本文从一份糙米样品中分离纯化出5株细菌,分别命名为BR1、BR2、BR3、BR4和BR5,对其进行形态学观察、生理生化鉴定、16S rRNA及gyrB基因测序鉴定,并对其毒力基因及药物敏感性等生物学特征进行研究。结果表明,5株分离菌株生化特性符合蜡样芽孢杆菌（Bacillus cereus）的特性,结合16S rRNA及gyrB基因测序结果,菌株BR1和BR4鉴定为蜡样芽孢杆菌。毒力基因的PCR检测结果表明,菌株BR1和BR4携带肠毒素基因（hbl、nhe、entFM和bceT）和细胞毒素K基因（cytK）,未检测到呕吐型毒力基因（cer和ces）。菌株BR1和BR4对青霉素、氨苄西林和头孢噻肟等药物耐药,对阿莫西林中度敏感,对诺氟沙星、环丙沙星和庆大霉素等药物敏感。本研究通过分离鉴定糙米源蜡样芽孢杆菌,发现分离菌株携带多种毒力基因,并且对青霉素类、头孢菌素类及磺胺类等药物耐药,表明糙米中的蜡样芽孢杆菌有引起食源性疾病的风险。     

即食米面制品中蜡样芽胞杆菌分离鉴定及毒力基因研究

为探究即食米面制品中蜡样芽胞杆菌的污染状况及食源性蜡样芽胞杆菌中毒力基因的分布规律,以餐饮服务场所采集245份即食米面制品为蜡样芽胞杆菌的污染调查材料,通过国标法及持家基因分离鉴定得到49株蜡样芽胞杆菌阳性株,并对其进行13种毒力基因的PCR检测。结果表明:蜡样芽胞杆菌的平均检出率为10.61%(26/245),阳性检出样品主要为盒饭及米饭,其蜡样芽胞杆菌平均检出水平为3032 CFU/g,其中以盒饭的检出程度最高。49株分离菌株均检出两种或两种以上的毒力基因,非溶血性肠毒素基因nhe和entFM检出率最高,分别达到100%及91.84%,是分离菌株的主要毒力基因;溶血素基因hblA、hblC、hblD检出率分别为20.41%、38.78%和40.82%。同时携带nhe三个基因又携带hblA、hblC和hblD基因的强毒株有7株,占14.29%,且这7株菌均含有entFM基因。呕吐型毒力基因ces、cer、EM1仅2株检出,检出率为4.08%。本研究对即食米面制品中蜡样芽胞杆菌的监控、预警及爆发引起食物中毒后追踪其感染源和传播途径及构建基因指纹图谱库和分子溯源平台具有指导意义。     

Toxin genes profiles and toxin production ability of Bacillus cereus isolated from clinical and food samples

Bacillus cereus can cause diarrheal and emetic type of food poisoning but little study has been done on the main toxins of food poisoning caused by B. cereus in Korea. The objective of this study is to characterize the toxin gene profiles and toxin-producing ability of 120 B. cereus isolates from clinical and food samples in Korea. The detection rate of nheABC, hblCDA, entFM, and cytK enterotoxin gene among all B. cereus strains was 94.2, 90.0, 65.8, and 52.5%, respectively. The ces gene encoding emetic toxin was not detected in all strains. Bacillus cereus strains carried at least 1 of the 8 enterotoxin genes were classified into 12 groups according to the presence or absence of 8 virulence genes. The 3 major patterns, I (nheABC, hblCDA, entFM, and cytK gene), II (nheABC, hblCDA and entFM gene), and VI (nheABC and hblCDA gene), accounted for 79.2% of all strains (95 out of 120 B. cereus isolates). Non-hemolytic enterotoxin (NHE) and hemolysin BL (HBL) enterotoxins were produced by 107 and 100 strains, respectively. Our finding revealed that NHE and HBL enterotoxins encoded by nhe and hbl genes were the major toxins among B. cereus tested in this study and enterotoxic type of B. cereus was predominant in Korea.     

Broad distribution of enterotoxin genes (hblCDA, nheABC, cytK, and entFM) among Bacillus thuringiensis and Bacillus cereus as shown by novel primers

Eight new pairs of PCR primers were designed and efficiently detect eight toxin genes (hblC, hblD, hblA, nheA, nheB, nheC, cytK, and entFM) in 411 B. cereus strains (121 food- and 290 soil isolates) and 205 B. thuringiensis strains (43 serovars, 10 food- and 152 soil isolates). According to the presence of these eight toxin genes, they were divided into four groups among the total 616 isolates. In Group I, all eight genes occurred simultaneously in 403 (65.42%) isolates, while Group II (134 isolates or 21.75%) and Group III (46 isolates or 7.47%) were devoid of hblCDA and cytK, respectively. In Group IV, there were thirty-three isolates which lacked both hblCDA and cytK. The presence of hblCDA in B. thuringiensis strains (86.80%) was significantly higher (P<0.05) than in B. cereus strains (66.18%) whereas no significant difference in nheABC, cytK and entFM occurrence was detected between both bacterial groups. Both nheABC and entFM genes were found in all B. cereus and B. thuringiensis strains (616 strains in total), while the cytK gene could be detected in 365 (88.80%) of the B. cereus and 172 (83.90%) of the B. thuringiensis strains. None of the 616 tested strains showed the presence of only a single or two genes in either the hbl or nhe operons. The eight primer pairs designed for this multiplex PCR allowed rapid detection of eight toxin genes from boiled cells with high sensitivity, gave 100% reproducibility, and did not cross-react to 32 other bacterial strains.     

Phenotypic and genotypic comparisons reveal a broad distribution and heterogeneity of hemolysin BL genes among Bacillus cereus isolates

The presence of hemolysin BL (HBL; components L(2), L(1), and B)-encoding genes (hblC, hblD, and hblA) from 339 Bacillus cereus strains isolated in Thailand was determined. PCR analysis showed that all three hbl genes were detected in 222 strains (65.5%). Two, one or no hbl genes were detected in 3 (0.9%), 6 (1.8%), and 108 (31.8%) strains, respectively. Among the 222 strains in which all three hbl genes were detected, 210 (61.9%) displayed discontinuous hemolysis (DH) characteristic of HBL producers, while 12 (3.5%) showed continuous hemolysis (CH) on sheep blood agar. Among strains in which none of the hbl genes was detected, 97 (28.6%) displayed CH while 11 (3.2%) did not show hemolytic activity. Three strains in which two hbl genes were detected showed CH. hblC was present in five of six strains where only one hbl gene was detected, and all of them (designated SS-00-15, TG-00-06, TG-00-14, F-00-25, and NR-01-49) showed DH. The HpaII restriction profiles of PCR fragments amplified from the hblC-A region in these five strains using hblC forward (FHC) and hblA reverse (RHA(2)) primers displayed heterogeneous patterns, which indicated sequence variation. Western blot analysis using polyclonal antibodies (Pab) raised against HBL components purified from strain F837/76 showed that three of the five strains produced all three components, whereas strain TG-00-06 did not give a signal for any component, and strain TG-00-14 produced B and L(1) but not L(2). The production of L(2) by these five strains was further analyzed using the Oxoid RPLA test. Three strains gave high titers (>64) whereas strains TG-00-06 and TG-00-14 showed lower titers of 16 and 32, respectively. The data show that HBL-encoding genes are widely distributed among B. cereus isolated in Thailand and there is a high degree of heterogeneity in both the genes and proteins. This is the first report of a B. cereus strain showing DH in which all three hbl genes and their proteins were not detected by both PCR primers and antibodies derived from prototype and type strains. The data also suggest that the L(2) component from strains TG-00-06 and TG-00-14 may be antigenically very different from that of most B. cereus isolates.     

Kefiran antagonizes cytopathic effects of Bacillus cereus extracellular factors

Kefiran, the polysaccharide produced by microorganisms present in kefir grains, is a water-soluble branched glucogalactan containing equal amounts of D-glucose and D-galactose. In this study, the effect of kefiran on the biological activity of Bacillus cereus strain B10502 extracellular factors was assessed by using cultured human enterocytes (Caco-2 cells) and human erythrocytes. In the presence of kefiran concentrations ranging from 300 to 1000 mg/L, the ability of B. cereus B10502 spent culture supernatants to detach and damage cultured human enterocytes was significantly abrogated. In addition, mitochondrial dehydrogenase activity was higher when kefiran was present during the cell toxicity assays. Protection was also demonstrated in hemolysis and apoptosis/necrosis assays. Scanning electron microscopy showed the protective effect of kefiran against structural cell damages produced by factors synthesized by B. cereus strain B10502. Protective effect of kefiran depended on strain of B. cereus. Our findings demonstrate the ability of kefiran to antagonize key events of B. cereus B10502 virulence. This property, although strain-specific, gives new perspectives for the role of bacterial exopolysaccharides in functional foods.     

红河州食源性蜡样芽胞杆菌毒力基因研究

目的了解红河州食源性蜡样芽胞杆菌毒力基因的携带情况,为食源性食物中毒的防治提供参考依据。方法采用PCR方法对2011年1月至2014年9月红河州收集到的31株食源性蜡样芽胞杆菌10种毒力基因进行检测。结果分离菌株普遍携带肠毒素毒力基因并且所有菌株均至少携带一种肠毒素基因,而呕吐型毒力基因只有一株菌携带。结论红河州食源性蜡样芽胞杆菌基因携带以腹泻型为主,且携带率较高,对食品安全和公共健康存在潜在威胁。     

石家庄市131株食源性蜡样芽胞杆菌毒力基因的分布

目的研究石家庄市食源性蜡样芽胞杆菌毒力基因的分布及毒力活性,了解蜡样芽胞杆菌的潜在威胁。方法采用PCR方法,对食品风险监测中分离到的131株蜡样芽胞杆菌进行肠毒素、呕吐毒素9种毒力基因扩增检测,用血平板检测的方法分析蜡样芽胞杆菌的毒力。结果毒力基因携带率较高,至少携带一个毒力基因的菌株达到检出菌总数的99.2%(130/131),溶血素BL基因(hbl ACD)和肠毒素FM基因(ent FM)是石家庄市食源性蜡样芽胞杆菌的主要毒力基因;检出的蜡样芽胞杆菌均产生溶血素BL,检出率为100%。结论腹泻型肠毒素在食品中的分布比较广泛,检出的蜡样芽胞杆菌均具有溶血素,对进食者存在潜在的危险性,今后应加强监控蜡样芽胞杆菌的污染,预防和控制蜡样芽胞杆菌食源性疾病的发生。     

食品中蜡样芽孢杆菌的分离及携带毒力基因的检测

为了确定成都市食品中蜡样芽孢杆菌所携带的毒力基因情况,本实验对市售食品共采样130份,通过菌落在MYP培养基上形态特征对蜡样芽孢杆菌进行初步分离,再利用16Sr RNA测序结果进行比对分析,并检测分离菌株携带的管家基因gyr B、rpo B、Vrr A、gro EL进一步确认,最后检测分离菌株携带毒力基因情况。结果表明130份样品中共检出23株蜡样芽孢杆菌,检出率为17.7%;23株分离菌株中,蜡样芽孢杆菌的4个管家基因gyr B、rpo B、Vrr A、gro EL检出率为100%,毒力基因的检测结果表明,nhe B和ent FM在16株分离菌株中检出,检出率为69.6%;nhe A和nhe C在14株分离菌株中检出,检出率为60.9%;hbl D在11株分离菌株中检出,检出率为47.8%;cyt K在10株分离菌株中检出,检出率为43.5%;bce T在9株分离菌株中检出,检出率为39.1%;hbl A和hbl C在8株分离菌株中检出,检出率为34.8%;cer和ces在2株分离菌株中检出,检出率为8.7%;未发现分离菌株携带hbl B和Hly基因。研究结果表明市售食品中分离的蜡样芽孢杆菌毒力基因携带率较高,对食品安全具有潜在威胁,应当引起有关部门注意。     

Effect of Bacillus cereus exocellular factors on human intestinal epithelial cells

To gain insight on the biological effects of the exocellular factors produced by Bacillus cereus, culture filtrate supernatants of different strains were coincubated with differentiated Caco-2 cells. Exocellular factors were able to detach enterocyte-like cells from the substratum after 1 h of incubation. In addition, microvilli effacing and dramatic changes on the cellular surface of enterocytes were found after incubation periods as short as 20 min. Since cell detachment was not inhibited by fetal calf serum, thiol activated cholesterol-binding cytolysin, cereolysin O, does not seem to be involved. Also, translocation of phosphatidylserine from the inner to the outer leaflets of the plasma membrane was demonstrated by using fluorescein isothiocyanate (FITC)-Annexin V. In contrast to the high capability of detaching Caco-2 cells shown by all the strains under study, the mitochondrial dehydrogenase activity was lowered by culture filtrate supernatants in a strain-dependent manner. For strain M2, the decrease in dehydrogenase activity was already evident after 30 min of incubation. Production of biologically active factors depends on the growth phase, and maximal activity was found in late exponential-early stationary phases. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) of concentrated exocellular factors showed a very complex scenery supporting the multifactorial character of the biological activity of B. cereus.     

海口市5类食品中蜡样芽胞杆菌药敏和毒力基因检测及分子分型研究

目的 了解海口市5类食品中蜡样芽胞杆菌污染状况、毒力基因携带类型、药物敏感特点和分子分型特征。方法 参照GB 4789.14—2014《食品安全国家标准 食品微生物学检验 蜡样芽胞杆菌检验》对海南粉、海南粉配料、奶粉类、快餐类和糕点类5类食品进行分离鉴定蜡样芽胞杆菌,并应用普通聚合酶链式反应(PCR)方法进行菌群特异性基因groEL和10种毒力基因检测;采用微量肉汤稀释(MIC)法检测菌株对抗生素的敏感性;应用脉冲场凝胶电泳(PFGE)技术对菌株进行分子分型。结果 626份不同食品样品中有197份检出蜡样芽胞杆菌,检出率为31.5%,其中海南粉检出率最高,为63.1%(140/222)。所有菌株均至少携带1种毒力基因,entFM基因携带率最高,为99.0%(195/197),ces基因携带率最低,为2.5%(5/197);同时携带nheA、nheB和nheC基因的菌株占88.8%(175/197),同时携带hblA、hblC和hblD基因的菌株占13.7%(27/197)。所有菌株对庆大霉素和氯霉素的敏感率均为100.0%(197/197),对万古霉素和环丙沙星敏感率分别为99.5%(196/197)和92.9%(183/197),对青霉素和复方新诺明的耐药率分别为100.0%(197/197)和90.9%(179/197)。PFGE分型结果显示,所有菌株可分为30个簇,117种带型。结论 海口市5类食品均存在蜡样芽胞杆菌污染,其中海南粉污染较重,对食品安全构成潜在的威胁;可依据菌株的毒力基因、分子分型和抗生素敏感性特点进行针对性防控和重点监管。     

我国食源性蜡样芽孢杆菌毒力基因和药物敏感性研究

目的 了解我国不同地区食源性蜡样芽孢杆菌的毒力基因携带特点及其对抗生素的敏感性,为食源性食物中毒的防治提供参考依据.方法 采用PCR方法对2011年我国不同地区收集的238株食源性蜡样芽孢杆菌10种毒力基因进行检测;采用微量肉汤稀释法测定其抗生素敏感性.结果 溶血素BL基因、肠毒素T基因和细孢毒素K基因是我国食源性蜡样芽孢杆菌的主要毒力基因,至少携带一个毒力基因的菌株达到检出菌总数的87.4％;蜡样芽孢杆菌对庆大霉素、万古霉素、环丙沙星、复方新诺明的敏感率为100％,对红霉素、四环素、氯霉素、克林霉素的敏感率分别为88.8％、90.2％、99.6％、87.1％,对氨苄西林和头孢噻肟的敏感率仅为0.4％和5.4％.结论 我国食源性蜡样芽孢杆菌毒力基因携带率较高,对食品安全和公共健康构成潜在的威胁;蜡样芽孢杆菌对氨苄西林、头孢噻肟的敏感性差,不应作为经验用药和预防用药.     

Bacillus cereus group strains, their hemolysin BL activity, and their detection in foods using a 16S RNA and hemolysin BL gene-targeted multiplex polymerase chain reaction system

Hemolysin BL (HBL) is a major virulence factor for Bacillus cereus group strains. It is also a target enterotoxin for the most commonly used B. cereus detection kit, i.e., the B. cereus enterotoxin (diarrheal type) reversed passive latex agglutination (BCET-RPLA) test kit. A survey of the HBL activities and the cytotoxicities to the Chinese hamster ovary (CHO) cells for the B. cereus group strains, however, showed that although only part of the B. cereus group strains are HBL active, all strains show cytotoxicity to the CHO cells. Thus, methods that allow the detection of not only the HBL but also of the B. cereus group strains are important. In this study, by comparison of the gene sequences of the 16S rRNA for B. cereus group and other bacteria strains, we designed primers B16S1 and B16S2 specific to all the B. cereus group strains. In addition, because HBL is a major enterotoxin, we also designed HBL gene-specific polymerase chain reaction (PCR) primers, i.e., Hm1 and Hm2, that generated the same results as those of the hemolysis and BCET-RPLA assays. Primers B16S1/B16S2 and Hm1/Hm2 could be combined into a multiplex PCR system for the simultaneous detection of B. cereus group cells and the possible presence of their HBL enterotoxins. Also, all these PCR systems allowed the detection of n x 10(0) CFU B. cereus cells per g of food sample if an 8-h enrichment step was performed prior to the PCR.     

生鲜食品中蜡样芽孢杆菌毒力基因及耐药性研究

目的:研究生鲜食品中蜡样芽孢杆菌的毒力基因及耐药性。方法:在成都市周边农贸市场和路边小摊采集各种生鲜食品共100份,采用MYP选择性培养基初步分离蜡样芽孢杆菌菌株,采用16S rRNA序列比对分析鉴定得到蜡样芽孢杆菌分离菌株,采用PCR检测分离鉴定菌株中蜡样芽孢杆菌13个毒力基因的携带情况,进一步采用纸片扩散法检测菌株对18种抗生素的耐药性。结果:从100份生鲜食品样本中,检出蜡样芽孢杆菌24株,检出率为24%,其中,路边小摊的检出率(70%)高于农贸市场(18.9%)。呕吐型基因ces和cer的检出率较低,仅在1株中发现;腹泻型基因nheC有24株检出;cytK有13株检出;bceT有12株检出;hblA有11株检出;nheB有10株检出;hblC、hblD、nheA各有9株检出;hly-Ⅱ有7株检出;hblB、entFM检出率为0;蜡样芽孢杆菌的4个看家基因rpoB、gyrB、groEL、vrrA在24株分离菌株中检出率为100%。同时发现,生鲜食品来源的蜡样芽孢杆菌分离菌株对杆菌肽B、磺胺甲亚唑、苯唑西林、青霉素表现出较高耐药性,耐药率大于96%;而对阿米卡星、氯霉素、庆大霉素、亚胺培南耐药率小于7%。结论:本研究通过分离鉴定生鲜食品源蜡样芽孢杆菌,研究其毒力基因携带及耐药性,为进一步评价生鲜食品中蜡样芽孢杆菌的安全风险提供参考数据。     

Effects of chitosan and a low-molecular-weight chitosan on Bacillus cereus and application in the preservation of cooked rice

Shrimp chitosan with 95% deacetylation and low-molecular-weight chitosan (LMWC) isolated from chitosan hydrolysate were investigated for their effects on the growth of Bacillus cereus and for use in the preservation of cooked rice. Four strains of Bacillus cereus were used: standard strain BCRC 10603 and three isolates (nos. 1 through 3) from cooked rice. The antibacterial activity of chitosan against B. cereus was greatly decreased when the reaction pH was changed from 6.0 to 7.0, but LMWC activity was less affected by this pH change. The susceptibility of B. cereus cells to chitosan decreased with increasing of cell age, in accordance with the relative electronegativity of the cell surface. B. cereus spores were more sensitive to LMWC and chitosan than were vegetative cells. Addition of 80 ppm LMWC and chitosan in sterile saline (pH 7.0) greatly reduced the D-value for the tested four strains at 90 degrees C from 30.77 to 46.51 min to 7.47 to 10.17 min and 4.68 to 7.91 min, respectively, and at 100 degrees C from 1.95 to 2.56 min to 0.89 to 0.93 min and 0.72 to 0.80 min, respectively. Addition of 2,000 ppm LMWC to raw rice water before steam cooking effectively inhibited increases in total aerobic bacteria and B. cereus in cooked rice stored at 37 and 18 degrees C.     

Establishment of a novel multiplex PCR assay and detection of toxigenic strains of the species in the Bacillus cereus group

Five different enterotoxins and one emetic toxin of Bacillus cereus have been characterized. To amplify all of the enterotoxin and emetic-specific sequences of the species in the B. cereus group, a multiplex PCR with 12 primer pairs was established. In developing the assay method, a common terminal sequence at the 3' ends of all primers was chosen and a hot start Taq polymerase was used to overcome primer dimer formation. The assay was successfully applied to analyze the toxigenic potential of 162 food-poisoning and food-related strains. Results showed that there were 10 toxigenic patterns for all the test strains. All of the B. cereus strains carried at least one toxin gene. More than 70% of Bacillus mycoides strains carried no known toxin genes. The toxin profiles and toxin genes of B. mycoides strains were significantly different from B. cereus strains (P < 0.05), although the two species were closely related. The results suggest that many B. mycoides strains might be less prone to cause food poisoning. They also indicate the importance of detecting the toxin genes together with the detection of the species in the B. cereus group.     

基于全基因组测序的蜡样芽孢杆菌食品分离株分子特征及耐药性研究

目的 研究上海市食品中分离的蜡样芽孢杆菌(Bacillus cereus)的基因组特征及其耐药性。方法 本研究对上海市食品中分离的蜡样芽孢杆菌进行鉴定、药物敏感性实验和全基因组测序,利用BioNumerics软件对测序数据进行拼接组装,利用拼接数据进行多位点序列分型(MLST)、毒力基因和耐药基因分析,并与PubMLST数据库中的ST型进行比较;对耐药基因和耐药表型进行比较分析。结果 本研究中37株蜡样芽孢杆菌均可分型,其中7株为ST新型。37株蜡样芽孢杆菌分为34个ST型,其中3株菌为ST26型,其余ST型各占1株,呈高度多样性。37株蜡样芽孢杆菌均携带nheA、nheB、nheC基因,hblACD基因簇的携带率为37.8%,而完整ces呕吐基因簇的携带率只有16.2%,其中2株为分离自即食食品的ST26型。药敏实验显示37株蜡样芽孢杆菌对氨苄西林、青霉素和苯唑西林耐药性较高,与其携带耐药基因种类并不完全一致。结论 上海市食品中分离的蜡样芽孢杆菌菌株分子分型呈现高度多样性,应加强食品中蜡样芽孢杆菌分子生物学监测,为预防控制由其引起的食源性疾病提供科学依据。     

温州地区蜡样芽胞杆菌分型特征研究

目的了解温州地区不同来源蜡样芽胞杆菌毒力基因分布、生化分型和多位点序列分型(MLST)特征。方法参照GB 4789.14—2014对127株蜡样芽胞杆菌进行鉴定和生化分型,同时采用PCR方法检测10种毒力基因,并进行MLST基因分型。结果 127株蜡样芽胞杆菌同时携带Nhe A、Nhe B、Nhe C基因的菌株占94.5%(120/127),而同时携带hbl A、hbl C、hbl D基因的占9.4%(12/127),携带ces基因的占7.9%(10/127);127株菌株除10株不能进行生化分型外,其余117株分为2型(0.8%,1/127)、5型(3.9%,5/127)、8型(1.6%,2/127)、9型(63.8%,81/127)和10型(22.0%,28/127);通过MLST分析,72株菌株共分为28个ST序列型,ST26(18.1%,13/72)、ST144(15.3%,11/72)、ST92(6.9%,5/72)和ST164(6.9%,5/72)为常见ST型别。28个ST型聚类分析显示温州地区蜡样芽胞杆菌分为4个序列群和13个单态群。结论温州地区蜡样芽胞杆菌毒力基因携带率较高,遗传关系具有多样性。