2021年广西腹泻病人来源的鼠伤寒沙门菌及其单相变种比较分析

目的 通过研究比较2021年广西壮族自治区腹泻病人来源的鼠伤寒沙门菌及其单相变体1，4，［5］，12∶i∶-（S.1，4，［5］，12∶i∶-）流行特征与耐药情况，更好地了解鼠伤寒沙门菌及其单相变体多重耐药克隆的流行病学及其在传播的潜力。方法 对来自病例监测分离的276株鼠伤寒沙门菌用血清凝集方法进行第一次分型，当第二项鞭毛抗原诱导三次后依然不凝集，再用多重PCR的方法进行第二次分型，用微量肉汤稀释法进行药敏试验。结果 经多重PCR确认为鼠伤寒沙门菌单相变体的有201株（72.8%），鼠伤寒沙门菌75株。鼠伤寒沙门菌对磺胺类药物、氯霉素的耐药率显著高于鼠伤寒沙门菌单相变体（P<0.05）。鼠伤寒沙门菌单相变体对氨苄西林、头孢噻肟、萘啶酸、庆大霉素、四环素5种抗生素的耐药率显著高于鼠伤寒沙门菌（P<0.05）。两种沙门菌对3类及以上抗生素耐药率达到79%以上，共同多重优势耐药谱为耐氨苄西林、氯霉素、甲氧苄啶/磺胺甲唑和四环素。结论 鼠伤寒沙门菌单相变体已经超过鼠伤寒沙门菌成为广西壮族自治区腹泻病人中最优势的血清型。两种沙门菌耐药情况不容乐观，特别是多重耐药菌株的快速增加。需要有针对性地加强对食源性鼠伤寒沙门菌和鼠伤寒沙门菌单相变体进行耐药监测，揭示其耐药性的潜在决定因素，并展开有效的干预措施。     

沙拉沙星体外诱导鼠伤寒沙门菌耐药实验

目的为获得体外鼠伤寒沙门菌(Salmonella typhimurium)对盐酸沙拉沙星(sarafloxacin hydrochloride)的活性耐药菌株,研究诱导其产生耐药的盐酸沙拉沙星最低浓度,为评价食品中沙拉沙星残留导致的微生物耐药提供依据。方法设置盐酸沙拉沙星0.001、0.002 5、0.005、0.025、0.05、0.1μg/ml实验组和空白对照组、NaOH溶剂对照组,采用NCCLS法测试最小抑菌浓度,对鼠伤寒沙门菌进行耐药诱导,耐药判断为≥8×MIC(0.25μg/ml)。对产生耐药的鼠伤寒沙门菌的gyrA基因片段进行PCR扩增,焦磷酸凝胶测序法鉴定gyrA基因常见5个位点观察突变。结果盐酸沙拉沙星在0.005μg/ml水平传到第10代时,MIC增大32倍,抑菌浓度增加到1μg/ml,耐药菌增殖停止;传到第25代时,鼠伤寒沙门菌gyrA基因Ser83位点发生突变,获得多株稳定、有生物活性的耐药菌株。沙拉沙星浓度≥0.025μg/ml时,细菌不生长;沙拉沙星浓度≤0.002 5μg/ml时细菌生长,不产生耐药。结论沙拉沙星浓度为0.005μg/ml时可体外诱导鼠伤寒沙门菌产生耐药,经传代后获得具生物活性的耐药菌株。     

2014—2019年成都市新都区鼠伤寒沙门菌耐药特性和分子分型研究

目的 了解新都区2014—2019年分离的54株鼠伤寒沙门菌耐药情况和分子分型特征.方法 对从病例监测、食物中毒和食品监测分离的54株鼠伤寒沙门菌用微量肉汤稀释法进行药敏试验,用脉冲场凝胶电泳(pulsed field gel electrophoresis,PFGE)进行分子分型.结果 54株鼠伤寒沙门菌耐药率为98...     

鼠伤寒沙门菌多重耐药性机制研究进展

AcrAB外流泵系统的过度表达和拓扑异构酶基因突变是导致鼠伤寒沙门菌产生多重耐药性 (MAR)的重要原因。AcrAB外流泵系统的合成有多重耐药调节子 (marRAB)调控 ,AcrAB与多种抗生素的耐药有关。拓扑异构酶 (topoisomerase)的突变主要与氟喹诺酮类药物的耐药性有关 ,突变主要发生在gyrA ,parC基因上     

四川省鸭和猪源鼠伤寒沙门菌脉冲场凝胶电泳分型与耐药比较分析

目的比较研究四川省鸭和猪源鼠伤寒沙门菌脉冲场凝胶电泳(pulsed field gel electrophoresis,PFGE)分型与耐药特征,为分析鼠伤寒沙门菌食品来源提供支持。方法将鸭和猪源监测中分离的47株鼠伤寒沙门菌进行PFGE分型,并采用最小抑菌浓度法测定9种药物敏感性。结果鸭和猪源中分离的鼠伤寒沙门菌在PFGE分型和耐药上存在差异。鸭源菌株聚集于簇I,而猪源菌株聚集于簇II。簇II菌株对氨苄西林、环丙沙星、氯霉素、庆大霉素、四环素、复方新诺明6种药物的耐药率高于簇I菌株。不同的PFGE分型鼠伤寒沙门菌在监测地区和耐药谱存在差异。SCSTm-10型菌株均分离于绵阳地区,SCSTm-11型主要分离于资阳地区。SCSTm-10型菌株对复方新诺明和环丙沙星的耐药率高于SCSTm-11型菌株。结论 PFGE分型可以获取四川省鼠伤寒沙门菌分离来源、分离地点和耐药的聚类信息,为鼠伤寒沙门菌食物中毒提供流行病学调查支持。     

广西鼠伤寒沙门菌1,4,[5],12∶i∶-耐药性及分子分型特征研究

目的 通过研究2018年广西壮族自治区食源性鼠伤寒沙门菌单相变体1,4,[5],12∶i∶-(S.1,4,[5],12∶i∶-)耐药情况和分子分型特征,建立广西壮族自治区食源性S.1,4,[5],12∶i∶-数据库。 方法 对来自病例监测和食品监测分离的124株S.1,4,[5],12∶i∶-用微量肉汤稀释法进行药敏试验,用脉冲电场凝胶电泳(PFGE)进行分子分型。结果 124株S.1,4,[5],12∶i∶-均对亚胺培南敏感,对另外13种抗生素产生不同程度的耐药,对四环素耐药率最高(96.8%,120/124),其次为氨苄西林(95.2%,118/124),三重及以上耐药率为26.6%(33/124)。47.1% S.1,4,[5],12∶i∶-分离株具有氨苄西林、氯霉素、磺胺类、四环素(ACSuT)多重耐药。PFGE结果分为104种带型,相似度为72.9%～100%,优势带型为JPXX01.GX0114(5株)和JPXX01.GX0294(4株),均来源于食源性疾病监测。结论 广西壮族自治区S.1,4,[5],12∶i∶-耐药情况不容乐观,特别对青霉素类和四环素类耐药严重。S.1,4,[5],12∶i∶-分子型别呈高度散发状态,及时对S.1,4,[5],12∶i∶-进行分子溯源,可以提高广西壮族自治区在散发病例中识别潜在的食源性S.1,4,[5],12∶i∶-暴发事件的能力。     

鼠伤寒沙门菌婴幼儿分离株耐药基因及毒力基因研究

目的基于全基因组测序结果,分析鼠伤寒沙门菌耐药表型和耐药基因的关系,研究其耐药发生机制和毒力因子。方法利用美国临床和实验室标准委员会(CLSI)推荐的微量肉汤稀释法对5岁以下腹泻患儿粪便中分离的321株鼠伤寒沙门菌开展药物敏感性试验,从中选择不同耐药表型的3株鼠伤寒沙门菌(S1:敏感;S2:TET耐药;S3:ESBLs阳性,10重耐药)进行全基因组测序,通过与抗生素抗性基因数据库(ARDB)和病原毒力因子数据库(VFDB)比对以及美国国立生物技术信息中心(NCBI)核酸数据库人工检索,对耐药基因和毒力基因进行注释。结果 321株鼠伤寒沙门菌对IPM和MEM均敏感,对TET和AMP的耐药率最高,分别为84.4%(271/321)和83.8%(269/321),206株菌株显示为耐3类以上抗生素(64.2%,206/321),11株CIP-CTX耐药株均为ESBLs阳性(3.4%,11/321);全基因组测序结果显示,S1、S2和S3的全基因组大小依次为4 876 427、4 970 690、5 133 380 bp,预测编码基因数分别为4 825、4 936和5 082个,GC含量依次为52.18%、52.14%、51.87%;对S1、S2和S3基因组中18、20和32个耐药基因进行注释,除共有基因外,S2主要携带tet A和sul2基因,S3主要携带sul1/2/3、tetB、AAC(3)-IV、AAC(6')-I、ANT(2″)-I、ANT(3″)-I、aph A1、bla_(CTX-M-14)、bla_(OXA-1)、catB3、cml_e1和cml_e3基因。与阳性鼠伤寒沙门菌株LT2的gyr A基因进行序列比对发现,S3 gyr A基因第87位密码子由GAC突变为AAC(Asp87→Asn);分别注释S1、S2和S3基因组的130、129和120个毒力基因,发现主要以三型分泌系统和粘附因子为主。结论 5岁以下腹泻患儿粪便样品中鼠伤寒沙门菌耐药现象严重;全基因组测序分析发现耐药基因与耐药表型一致,且存在多种毒力因子;为后续开展耐药基因传播机制和菌株致病性研究,评估其对人群的健康风险,防控鼠伤寒沙门菌感染提供技术支持。     

北京市食源性非伤寒沙门菌的分子分型和耐药性研究

目的了解北京市食源性非伤寒沙门菌的分子特征及耐药情况。方法对2004—2010年北京市食源性致病菌监测网收集的100株沙门菌进行脉冲场凝胶电泳(PFGE)分型和抗生素敏感性检测。结果 100株非伤寒沙门菌通过PFGE分型分为62个不同的带型,每个带型包含1~11株菌。抗生素敏感性结果显示,100株菌中有55株菌表现为对至少1种抗生素耐药,其中多重耐药菌株15株。菌株对各抗生素的耐药率为萘啶酸40%、四环素30%、氯霉素15%、庆大霉素10%、甲氧苄啶/磺胺甲恶唑10%、环丙沙星9%、头孢西丁1%、头孢噻肟0%。结论沙门菌PFGE带型和耐药谱均与血清型存在很高的一致性。提示北京市食源性非伤寒沙门菌的耐药情况比较严重,开展对该菌分子分型与耐药特征分析的联合监测意义重大。     

中国食源性鼠伤寒沙门菌株耐药谱及PFGE分型研究

目的了解掌握中国食品中鼠伤寒沙门菌的耐药状况，并对2002—2005年国家食源性疾病监测网分离的23株鼠伤寒沙门菌进行耐药性监测及PFGE分型研究。方法利用血清学方法对2002—2005年食源性疾病监测网分离的沙门菌进行分型，并运用CLSI（Clinical and Laboratory Standards Institute）推荐的纸片法对鼠伤寒沙门菌株进行耐药性检测。采用脉冲场凝胶电泳法（PFGE）进行PFGE分型。结果发现15株多重耐药鼠伤寒沙门菌，其中耐4～5种抗生素的6株（40％），耐6～9种抗生素的5株（33．3％），耐10种抗生素的4株（26．7％）；可分为16个PFGE型。其中5个PFGE型的菌株数超过1株。结论我国食源性鼠伤寒沙门菌分离株的多重耐药性严重，PFGE分型方法对鼠伤寒沙门菌的分型能力较好，同一PFGE型菌株的耐药谱非常接近。     

市售活鸡和腹泻患者中非伤寒沙门菌分子特征和耐药性研究

分析市售活鸡及腹泻患者中非伤寒沙门菌的分子特征的相似性及耐药性,预防沙门菌的感染流行。方法 收集2012年1月—2013年10月从1 054份腹泻患者粪便样品及440份农贸市场的活鸡肛拭子中分离出的非伤寒沙门菌株93株,进行血清分型,对常见的非伤寒沙门菌采用脉冲场凝胶电泳(PFGE)进行分子分型、Bionumerisc v4.0进行聚类分析、纸片扩散法(K-B法)进行药物敏感性试验。结果 1 054份腹泻样品中共检出非伤寒沙门菌88株,分离率为8.35%,分为25种血清型,肠炎沙门菌居多。其中0～2岁的婴幼儿检出率较高,占52.27%(46/88)。PFGE结果:16株肠炎沙门菌分成14个PFGE型、12株斯坦利沙门菌分成11个PFGE型、14株鼠伤寒沙门菌分成14个PFGE型、6株德尔卑沙门菌分成5个PFGE型;对菌株间的相似性进行比较发现两个100%同源的肠炎沙门菌PFGE型,其他菌株的分子特征的相似性不大,鸡源菌株与人源菌株的分子特征相似性不大;对12种抗菌药物的耐药率在50%以下,不同的血清型耐药率各不相同,耐药较严重的为德尔卑沙门菌。440份活体鸡肛拭子分离出的非伤寒沙门菌5株,分离率为1.14%,共分得4种血清型,其中姆班达卡沙门菌病人中未检出。结论 该地区由非伤寒沙门菌引起腹泻的感染率高,尤以婴幼儿多见,患者与鸡未发现有同一克隆株,对常用抗生素有很好的敏感性。     

Antimicrobial resistance of Salmonella enterica serovar Typhimurium isolated from cattle in Japan

The antimicrobial susceptibility of 144 Salmonella enterica serovar Typhimurium isolates collected from all over Japan between 1973 and 1998 were investigated. All the isolates exhibited resistance to four or more antimicrobials and 22 resistance patterns were observed. Isolates showing resistance patterns to ampicillin (A), chloramphenicol (C), streptomycin (S), sulfonamides (Su) and tetracycline (T), which are typical resistance patterns for S. Typhimurium DT104 (DT104), were predominant. Thirty-six of the 68 isolates that exhibited resistance to five or more antimicrobials (ACSSuT+) were identified as DT104 by phage typing. Another 103 S. Typhimurium strains gathered from cattle between 1977 and 1999 in a limited area of Japan were analyzed for molecular epidemiological studies. Results using fluorescent amplified-fragment length polymorphism and pulsed-field gel electrophoresis suggest that clonal exchange of S. Typhimurium among cattle in Japan has occurred since 1992, and that contemporary strains show a remarkable degree of homogeneity with DT104 at a molecular level. The clonal replacement by DT104 affected the antimicrobial resistance pattern of S. Typhimurium from cattle in Japan.     

Interaction of Salmonella enterica subspecies enterica serovar Typhimurium and mung bean (Phaseolus aureus) plants

The effect of Salmonella enterica subspecies enterica serovar Typhimurium, a zoonotic serovar, on mung bean (Phaseolus aureus) cultivar Pant Mung-3 plants was studied. Inoculation of mung bean seeds with Salmonella Typhimurium (7.2 x 10(5) CFU/ml) reduced sprouting rate (P < 0.07). This effect was more pronounced at higher levels of contamination. In the soil inoculated with Salmonella Typhimurium (7.2 x 10(6) CFU/g), germination was retarded and the number of defective sprouts was also significantly higher (P < 0.002). Salmonella Typhimurium grew inside germinating seeds and plant tissues and persisted in seedlings, adult plants, and harvested seedlings dried and stored at room temperature (30 degrees C) up to 45 days. Phaseolus aureus plants grown in sterile soil was resistant to Salmonella Typhimurium infection at 15 days of age and cleared Salmonella from all the aerial parts within 3 h of infection. However, Salmonella Typhimurium could be reisolated from the basal area of the stem and from soil even after 45 days of exposure to the pathogen.     

Quantification of low and high levels of Salmonella enterica serovar Typhimurium on leaves

Precise and rapid quantification of low levels of pathogens associated with fresh produce may be particularly challenging and yet evermore necessary to guarantee microbial safety of the produce and to carry out research on the subject of persistence of the pathogens. Here, microbiological and molecular based methods were examined for their ability to precisely quantify different amounts of Salmonella enterica serovar Typhimurium artificially inoculated on parsley leaves. Recovery of S. Typhimurium from parsley by mechanical detachment using stomacher, mortar and pestle, vortex, sonicator or homogenizer followed by plating resulted in underestimation with less than 1% recovery when leaves were inoculated with 3.5–6.5 log CFU/g. Lower levels were undetectable by most assayed methods, and only recovery with mortar and pestle or adding of enrichment step resulted in partial detection of 300 CFU/g. Implementation of PCR based methods with/without pre extraction of the DNA from the contaminated leaves resulted in more accurate values of the pathogen (about 20% of the initial inocula) and as low as 300 CFU/g were detected even without an enrichment step. These methods can be applied to study transfer of Salmonlla from contaminated water or soil to plants using low and more reasonable levels of contamination.     

Survival of Campylobacter jejuni and Salmonella enterica Typhimurium in vacuum-packed, moisture-enhanced pork

The abilities of Campylobacter jejuni and Salmonella enterica Typhimurium to survive in vacuum-packaged, moisture-enhanced pork stored at 4 or 10°C were examined. Pork loins were surface inoculated with either C. jejuni or Salmonella Typhimurium and then moisture enhanced to a target of 10 or 20%. The enhanced pork loins were sliced 1 cm thick and vacuum packaged. A pork loin without moisture enhancement was sliced and vacuum packaged as a control. Samples were collected, plated, and the numbers of surviving organisms were determined periodically during storage at 4 and 10°C. The numbers of C. jejuni or Salmonella Typhimurium in samples with different moisture enhancement levels were similar (P > 0.05). No significant differences (P > 0.05) in C. jejuni counts were observed between samples at 10°C and those at 4°C. In contrast, the numbers of Salmonella Typhimurium in samples at 10°C had significantly (P < 0.05) increased (0.41 log CFU/g) from those at the refrigerated temperature of 4°C. Vacuum storage at 4 and 10°C for 28 days did not result in dramatic reductions in the mean numbers of C. jejuni and Salmonella Typhimurium. Our findings indicate that vacuum packaging under chilled conditions will not add substantially to safety for moisture-enhanced pork. Strict hygienic practices or the implementation of decontamination technologies is recommended.     

Characterization of antimicrobial resistance in Salmonella enterica serovar Typhimurium isolates from food animals in the U.S.

Salmonella enterica subspecies enteric serovar Typhimurium is the second most common serovar implicated in human diseases in the United States. In this study, 120 S. Typhimurium isolates from animal sources were characterized by antimicrobial susceptibility testing, antimicrobial resistance gene detection and plasmid analysis. Overall, 94 (78%) of the isolates were resistant to one or more antimicrobials and 63 (53%) were resistant to at least five antimicrobials. Resistance was most commonly observed to streptomycin (62%), sulfisoxazole (62%), or tetracycline (61%). When resistance was detected, a corresponding resistance gene was detected in 89% of cases. Class 1 integrons were detected in 51 isolates, all which contained the aadA2 gene. A plasmid Inc group was detected in 68 (57%) isolates. Thirty nine (57%) of these isolates were resistant to 5 or more antimicrobials.     

Fermented whey dairy beverage offers protection against Salmonella enterica ssp. enterica serovar Typhimurium infection in mice

Fermented whey dairy beverages are dairy products obtained by fermentation from a mixture of milk and whey. These beverages have important health benefits, which could be improved with the addition of probiotic cultures. This study assessed the protective effect of the cosupplementation of a probiotic culture (Lactobacillus casei 01) with a fermented whey dairy beverage against infection by Salmonella enterica ssp. enterica serovar Typhimurium in a murine model. Two fermented whey dairy beverages were prepared: conventional (FWB; starter culture) and probiotic (PFWB; starter and probiotic cultures). In the first set of experiments, Balb/C female mice were treated with FWB or PFWB, challenged with Salmonella Typhimurium, and analyzed for clinical signs, weight loss, and mortality for 20 d postinfection. In the second set of experiments, mice were treated with FWB or PFWB, challenged with Salmonella Typhimurium, and killed on d 10 postinfection. The liver, colon, and ileum were used for myeloperoxidase, eosinophil peroxidase, and histological analysis and translocation to the liver. The contents from the small intestine were used for secretory IgA determination. The FWB treatment showed a better effect on animal survival (70%), translocation of the pathogen to the liver (2 out of 10), histopathology (fewer lesions), and inflammation than PFWB, which presented 50% animal survival, translocation in 5 out of 10 animals, and higher lesions. The control group presented 40% animal survival, translocation in 6 out of 10 animals, and severe lesions. Therefore, FWB was deemed to have a greater protective effect against Salmonella Typhimurium infection in the murine model compared with PFWB.     

Development and Characterization of Monoclonal and Polyclonal Antibodies Against Salmonella enterica Typhimurium for Biosensor Detection

ABSTRACT: For the construction of a biosensor to specifically and effectively detect Salmonella enterica Typhimurium, a monoclonal antibody (mAb) 1B4 and a polyclonal antibody (pAb) S48 were developed against purified outer membrane proteins (OMPs) of Salmonella enterica Typhimurium. Gel electrophoresis data indicated that the apparent molecular weight of the main OMP was 55 kD. The mAb 1B4 strongly reacted to Salmonella enterica Typhimurium and Salmonella enterica Paratyphi and had no cross-reaction to other gram-negative and gram-positive bacteria that were tested. The pAb S48 also showed high reactivity to Salmonella enterica Typhimurium, but had cross-reactivity with other tested bacteria. Because of the high specificity and sensitivity of 1B4 against S. enterica Typhimurium, it was immobilized onto a sensor platform to capture bacteria. The captured bacteria were visualized byalight microscope fortheir detection. The detection limit of the biosensorwas 1 × 10² colony forming units/mL in bacterial culture and in a milk sample, and no other bacteria interfered with the binding of targeted cells. Therefore, a biosensor was developed that would rapidly detect Salmonella in the laboratory and in a food processing plant.     

Identification of Salmonella enterica serovar Typhimurium using specific PCR primers obtained by comparative genomics in Salmonella serovars

Salmonella enterica serovar Typhimurium is a major foodborne pathogen throughout the world. Until now, the specific target genes for the detection and identification of serovar Typhimurium have not been developed. To determine the specific probes for serovar Typhimurium, the genes of serovar Typhimurium LT2 that were expected to be unique were selected with the BLAST (Basic Local Alignment Search Tool) program within GenBank. The selected genes were compared with 11 genomic sequences of various Salmonella serovars by BLAST. Of these selected genes, 10 were expected to be specific to serovar Typhimurium and were not related to virulence factor genes of Salmonella pathogenicity island or to genes of the O and H antigens of Salmonella. Primers for the 10 selected genes were constructed, and PCRs were evaluated with various genomic DNAs of Salmonella and non-Salmonella strains for the specific identification of Salmonella serovar Typhimurium. Among all the primer sets for the 10 genes, STM4497 showed the highest degree of specificity to serovar Typhimurium. In this study, a specific primer set for Salmonella serovar Typhimurium was developed on the basis of the comparison of genomic sequences between Salmonella serovars and was validated with PCR. This method of comparative genomics to select target genes or sequences can be applied to the specific detection of microorganisms.     

Heat tolerance of Salmonella enterica serovars Agona, Enteritidis, and Typhimurium in peanut butter

Recent large foodborne outbreaks caused by Salmonella enterica serovars have been associated with consumption of foods with high fat content and reduced water activity, even though their ingredients usually undergo pasteurization. The present study was focused on the heat tolerance of Salmonella enterica serovars Agona, Enteritidis, and Typhimurium in peanut butter. The Salmonella serovars in the peanut butter were resistant to heat, and even at a temperature as high as 90 degrees C only 3.2-log reduction in CFU was observed. The obtained thermal inactivation curves were upwardly concave, indicating rapid death at the beginning (10 min) followed by lower death rates and an asymptotic tail. The curves fitted the nonlinear Weibull model with beta parameters < 1, indicating that the remaining cells have a lower probability of dying. beta at 70 degrees C (0.40 +/- 0.04) was significantly lower than beta at 80 degrees C (0.73 +/- 0.19) and 90 degrees C (0.69 +/- 0.17). Very little decrease in the viable population (less than 2-log decrease) was noted in cultures that were exposed to a second thermal treatment. Peanut butter is a highly concentrated colloidal suspension of lipid and water in a peanut meal phase. We hypothesized that differences in the local environments of the bacteria, with respect to fat content or water activity, explained the observed distribution and high portion of surviving cells (0.1%, independent of the initial cell number). These results demonstrate that thermal treatments are inadequate to consistently destroy Salmonella in highly contaminated peanut butter and that the pasteurization process cannot be improved significantly by longer treatment or higher temperatures.