流感病毒亚单位疫苗中间体中沙门菌快速检测方法的建立

目的：建立一种能够快速、准确地检测流感病毒亚单位疫苗中间体样品中沙门菌污染的方法。方法：首先利用选择性增菌培养基对样品进行增菌培养,然后提取样品中的细菌基因组DNA,通过沙门茵特异性引物对基因组DNA进行PCR扩增,以琼脂糖凝胶电泳实现对目的片段的检测。结果：该PCR反应体系的扩增检测灵敏度可达1pg的沙门菌DNA,利用选择性增菌培养配合该PCR体系可在最快24h内实现对沙门菌的准确检测,测定结果与传统方法相符。结论：此方法应用于流感病毒亚单位疫苗中间体的沙门菌检查,较之传统的培养法结合生化鉴定的方法,大大缩短了检测周期,降低了结果判读的难度,在实际生产中有很好的应用前景。     

鸡胴体中沙门菌的定量检测及风险评估

目的通过鸡胴体中沙门菌的定量检测,了解长春市市售鸡胴体中沙门菌的污染状况,为沙门菌的定量检测评估提供依据。方法前增菌:鸡胴体缓冲蛋白胨水淋洗后培养;增菌:TT肉汤和RV肉汤;分离:接种XLT4培养基;鉴定:API20E和沙门多价血清。结果沙门菌全年都有检出,阳性率15%~100%不等。7月份阳性率最高为100%;其次8月份为90%;6月份为80%。冷藏储存条件下鸡胴体样品中沙门菌的检出率及其含菌量均高于冷冻条件下。农贸市场鸡胴体样品中沙门菌的检出率及其含菌量均高于大型超市。要加强市售鸡胴体中各环节中沙门菌污染的监管。结论长春市市售鸡胴体中沙门菌的污染率为64.6%(155/240)。冷藏储存条件下鸡胴体样品中沙门菌的检出率及其含菌量均高于冷冻条件下。农贸市场鸡胴体样品中沙门菌的检出率及其含菌量均高于大型超市。要加强市售鸡胴体中各环节中沙门菌污染的监管。     

2005-2013年肠道致病菌中志贺菌和沙门菌的分布及检测方法经验总结

目的分析杭州市第三人民医院2005-2013年肠道致病菌中志贺菌和沙门菌的分布并对检测方法进行经验总结。方法采用定位显色培养基和营养琼脂初筛疑似志贺菌和沙门菌,并参考该院历年菌种分离情况进行血清凝集。采用EXCEL统计处理数据,χ2检验比较采用定位显色培养基和营养琼脂平板之前和之后,粪便培养志贺菌和沙门菌的分离率之间差异的显著性。结果采用定位显色培养基和营养琼脂平板初筛疑似菌株后,志贺菌和沙门菌的分离率差异存在统计学意义。结论采用定位显色培养基和营养琼脂初筛疑似菌株,并对本院历年菌种分离情况进行总结,对改善志贺菌和沙门菌的分离率,提升检验科微生物室的工作效率有较大帮助。     

不同霉菌培养基对Rhinocladiella sp.的影响及其生长水平预估的比较性研究

本实验使用孟加拉红琼脂、沙氏琼脂培养基和马铃薯葡萄糖琼脂3种市售霉菌培养基,以及改良孟加拉红和改良马铃薯葡萄糖琼脂2种自制改良培养基,按照GB 4789.15 2016 《食品安全国家标准食品微生物学检验霉菌和酵母计数》中的培养条件,对从变性淀粉样品中分离纯化得到的Rhinocladiella sp.137/143/155x214wat目标菌株进行了比较性研究。研究结果表明,使用沙氏琼脂培养基对纯化目标菌进行培养时,菌落数量能在培养第3 d得到显著(p=0.04)增长。在使用全部5种培养基对接种目标菌的无菌变性淀粉样品进行生长研究后,发现市售马铃薯葡萄糖琼脂在培养第3 d的增殖效果较其他4种培养基(p=0.01)有显著差距,而沙氏琼脂培养基则继续在缩短目标菌早期观测时间、预估菌落总体生长水平方面具有一定优势。     

反向线性探针杂交在检测实验动物沙门菌中的研究

目的建立简便、快速、准确、灵敏、特异的沙门菌检测方法。方法根据沙门菌argT基因序列设计通用引物和3'、5'均加有polyC的特异性探针。上游引物5'标记生物素,将探针线性固定在硝酸纤维素膜上,使沙门菌PCR扩增产物与探针进行杂交,通过优化杂交条件,建立反向线性探针杂交检测方法。利用该方法对重庆地区74只实验动物进行检测,同时与传统分离培养方法比较。结果反向线性探针杂交方法灵敏度高,对沙门菌PCR扩增产物在3ng/μL以上可有效检测。从细菌分离培养及DNA提取到PCR扩增及反向杂交结束仅需27h。该检测方法特异性高,对6种非沙门菌的检测中,其特异性为100%。应用传统分离培养方法和反向线性探针杂交方法分别检测42只KM小鼠和32只SD大鼠,两种方法检测结果一致性为100%。结论反向线性探针杂交检测方法,具有快速、可靠、敏感和特异的特点,可用于沙门菌感染时的检测,适合应用于实验动物沙门菌的监测。     

实验动物中沙门菌的实验室检测能力验证的结果与分析

目的通过实验动物中沙门菌检测能力验证计划,了解实验动物检测机构对沙门菌的检验能力,提高实验动物质量检测水平。方法按照CNAS批准的能力验证方案,通过冷冻干燥法制备含有沙门菌及干扰菌的实验动物粪便样品,经过稳定性和均匀性检验合格,作为能力验证样品。采用随机编号,经冷链运输发放给参加单位,并附作业指导书。在规定时限提交检验报告和原始记录复印件,其结果与样品预检结果一致的判为满意结果,不一致或未能提交结果的判为不满意结果。结果全国共有20个省市的30个实验室参加沙门菌能力验证项目,其中28个实验室获满意结果,占总参加机构的93.3%,不满意的2个实验室,占6.7%。采用分离培养方法的有29个实验室,采用PCR方法的有2个实验室。结论实验动物质量检测机构沙门菌检测能力较高,实施能力验证计划能够反映实验室的检测水平。     

市售鲫鱼肠道中沙门菌的调查研究

目的 为摸清市场中出售鲫鱼沙门菌的污染情况.方法 以徐州市某市场上出售的鲫鱼为材料,主要采用SS琼脂平板对120份肠道样品中的沙门菌进行初步鉴定和培养计数.结果 在120份鲫鱼肠道中共检出沙门菌32份,阳性率为26.7％ (32/120),菌群数量分布在(0.32 ～9.12)×103 CFU/g.结论 本研究对徐州地区市售鱼类沙门菌检疫及疾病防治等提供重要参考.     

鼠伤寒沙门菌pStSR100质粒对生物膜形成的影响

目的:鼠伤寒沙门菌在多种表面形成的生物膜对其致病性和引起食物中毒等方面起着重要作用,本研究探讨鼠伤寒沙门菌pStSR100质粒对细菌在不同材质表面生物膜形成的影响。方法:用LB(Luria-Bertani,LB)培养基和TSB(Tryptose Soya Broth,TSB)培养基分别将携带pStSR100质粒的野生株在96孔板与放置无菌小圆玻片的24孔板中静态培养48 h,用结晶紫半定量法确定生物膜形成的适宜培养基。将野生株与消除质粒的突变株,用结晶紫半定量法和激光共聚焦显微镜(Confocal Laser scanning microscopy,CLSM)观察其在聚苯乙烯培养板和小圆玻片表面形成生物膜的差异。结果:用LB培养时细菌生物膜的形成能力高于用TSB培养,LB培养基更适宜生物膜形成;结晶紫半定量法结果表明野生株比突变株在小圆玻片表面形成生物膜的能力明显增强,而在聚苯乙烯培养板表面两者则无明显差异;CLSM观察发现,野生株在小圆玻片表面形成融合成片的大克隆,突变株仅形成较小克隆。结论:鼠伤寒沙门菌pStSR100质粒能促进该菌在亲水性材质表面生物膜的形成,但其对该菌在疏水性材质表面生物膜的形成未见明显影响,这一新发现为进一步研究鼠伤寒沙门菌生物膜形成的调控机制,研制抗感染材料提供了理论和实验依据。     

不同方法检测鸡胴体中沙门菌结果的比较研究

对鸡胴体淋洗液样品进行沙门菌检测,样品经过前增菌和选择性增菌后,分别采用4种不同的方法进行检测,即普通PCR方法、实时荧光PCR方法、免疫学方法(VIDAS)和传统的微生物检验方法。共检测了56份样品,普通PCR检出阳性样品34份,实时荧光PCR阳性样品36份,VIDAS阳性样品28份;PCR和实时荧光定量PCR均无假阳性和假阴性结果。结果显示该3种检测方法均可以用于鸡胴体中沙门菌的快速检测。     

鼠伤寒沙门菌pStSR100质粒对生物膜形成的影响

目的：鼠伤寒沙门菌在多种表面形成的生物膜对其致病性和引起食物中毒等方面起着重要作用,本研究探讨鼠伤寒沙门菌pStSR100质粒对细菌在不同材质表面生物膜形成的影响。方法：用LB（Lufia—Bertani,LB）培养基和TSB（TryptoseSoyaBroth,TSB）培养基分别将携带pStSR100质粒的野生株在96孔板与放置无菌小圆玻片的24孔板中静态培养48h,用结晶紫半定量法确定生物膜形成的适宜培养基。将野生株与消除质粒的突变株,用结晶紫半定量法和激光共聚焦显微镜（ConfocalLaserscanningmicroscopy,CLSM）观察其在聚苯乙烯培养板和小圆玻片表面形成生物膜的差异。结果：用LB培养时细菌生物膜的形成能力高于用TSB培养,LB培养基更适宜生物膜形成;结晶紫半定量法结果表明野生株比突变株在小圆玻片表面形成生物膜的能力明显增强,而在聚苯乙烯培养板表面两者则无明显差异;CLSM观察发现,野生株在小圆玻片表面形成融合成片的大克隆,突变株仅形成较小克隆。结论：鼠伤寒沙门菌pStSR100质粒能促进该茵在亲水性材质表面生物膜的形成,但其对该菌在疏水性材质表面生物膜的形成未见明显影响,这一新发现为进一步研究鼠伤寒沙门菌生物膜形成的调控机制,研制抗感染材料提供了理论和实验依据。     

Salmonellae in Fish Meal Plants: Relative Amounts of Contamination at Various Stages of Processing and a Method of Control

Previous studies have shown that Menhaden fish meal, a common ingredient of animal feeds, is frequently contaminated with salmonellae. Animals that eat contaminated feed may become infected. If they, in turn, are eaten by humans, they may be a means by which salmonellae are introduced into the human population. Epidemiological studies of the fish-meal industry were carried out to determine the sources of salmonellae in fish meal and the factors affecting the persistence and survival of salmonellae during the processing of fish meal. Examination of 190 fish immediately after they came from the Gulf of Mexico revealed no salmonellae, but salmonellae were frequently isolated from samples of fish taken from the boats when they arrived at the plants. Salmonellae were also frequently isolated from dockside water at each of the plants. Approximately 50% of the samples taken in the raw fish processing areas were contaminated with salmonellae. The percentage of samples yielding salmonellae decreased progressively through the various sequences of processing, but more than 15% of the samples taken from the finished products were also positive. Salmonellae were isolated from the raw area of the plant most frequently while the plant was operating and less frequently when the plant was idle, whereas in the processing area of the plant the reverse was true. Salmonellae appeared to survive and multiply in the processing area of the plant while the plant was idle, which resulted in contamination of the first portion of each day's production. Salmonellae in the processed fish meal were reduced to nondetectable levels by reprocessing the first 45 min of each day's production.     

Investigation of Salmonella infection in goats fed corn silage grown on land fertilized with sewage sludge

A total of 36 goats were fed for 17 months with corn silage grown on land fertilized with human sewage sludge. These animals were investigated for salmonella infections. Salmonellae were not detected in cultures of fecal or silage samples. No significant agglutination titers were detected in goat sera examined. Salmonella newport C2 was isolated from the sludge used as fertilizer on the cornfields. The public health aspects of the findings are discussed as they relate to the increasing use of sewage sludge for agricultural fertilizers, as well as to the resultant effects on human food and livestock feed.     

Investigation of Salmonella infection in goats fed corn silage grown on land fertilized with sewage sludge.

A total of 36 goats were fed for 17 months with corn silage grown on land fertilized with human sewage sludge. These animals were investigated for salmonella infections. Salmonellae were not detected in cultures of fecal or silage samples. No significant agglutination titers were detected in goat sera examined. Salmonella newport C2 was isolated from the sludge used as fertilizer on the cornfields. The public health aspects of the findings are discussed as they relate to the increasing use of sewage sludge for agricultural fertilizers, as well as to the resultant effects on human food and livestock feed.     

Introduction of Salmonellae into a centralized laboratory animal facility by infected day old chicks

Fifty deliveries of day old chicks were examined for Salmonellae on arrival as part of the Animal Laboratories' routine microbiological monitoring programme. Paper floor inserts and faeces from the transport boxes were immersed in peptone water and then cultured in two different enrichment media. Salmonellae were isolated in six of the 50 samples; one isolate was identified as Salmonella muenchen and the other five as Salmonella cerro var. siegburg. The relevance of these findings to multispecies animal facilities is discussed.     

The Fluorescent Antibody Technique for the Detection of Salmonella in Routine Use

S ummary : The direct and indirect fluorescent antibody technique (FAT) were compared with cultural methods for detecting salmonellae in meat products, animal feedingstuffs, poultry carcase swabs, giblets and poultry plant and equipment swabs. Salmonellae were not isolated from meat products and fluorescent cells were not seen on slides prepared by either FAT. The indirect and direct FAT recorded 13% and 9% respectively, false positive results, with samples of animal feedingstuffs, but the direct FAT recorded a single false negative result. Salmonellae were not isolated from poultry carcase swabs but 3% and 4·5% respectively, of false positive results were obtained with the indirect and direct FAT. Salmonellae were isolated from both giblet samples and poultry plant swabs and both gave rise to false negative FAT results. Preliminary studies of the efficacy of the FAT for screening animal faecal material for salmonellae indicated that no single combination of enrichment broth and FAT gives unequivocal results, but the staining of smears from tetrathionate broth by either FAT gives rise to a high percentage of false negative results.     

Radiation Treatment of Foods: II. Public Health Significance of Irradiation-Recycled Salmonella1

Salmonellae resistant to gamma irradiation were developed by repeated irradiation and subculturing in a nutrient broth-yeast extract medium. Few differences were noted in the biochemical characteristics of parent and resistant cultures; however, microculture studies revealed variations in morphology and in cell division patterns. A considerable decrease in pathogenicity for day-old chicks was apparent with resistant cultures, but their phenol-water extracts were as toxic as parent material for 10-day chick embryos. Five serial chick passages did not reverse the reduced pathogenicity or aberrant morphology of a resistant Salmonella typhimurium culture. Results of phage typing of both parent and serially irradiated S. typhimurium were inconclusive, whereas the O-1 genus-specific phage lysed all parent serotypes tested but only one of the serially irradiated cultures. Agglutination of parent S. typhimurium cells with their homologous rabbit antiserum was unaffected by prior absorption with resistant strains. The results indicate that radiation recycling altered Salmonella into strains of lesser public health significance.     

High-resolution analysis of salmonellae from turtles within a headwater spring ecosystem

Sediments and water from the pristine headwaters of the San Marcos River, Texas, USA, as well as swabs from biofilms on the carapace and from the cloacae of 17 musk turtles (Sternotherus odoratus) and one snapping turtle (Chelydra serpentina serpentina) caught at the same site, were analysed for salmonellae by culture and molecular techniques. Whereas enrichment cultures from sediment and water samples were negative for salmonellae in PCR- and in situ hybridization-based analyses, both techniques detected salmonellae after enrichments from both carapace and cloacae of nine (i.e. of 53%) musk turtles. Further characterization of 10 isolates obtained from the enrichment cultures of four selected individuals and confirmed as salmonellae by PCR analysis was achieved by fingerprinting techniques (rep-PCR). The results show differences between individuals and, in one case, variation among isolates from a single individual. All isolates from two individuals displayed identical profiles. These profiles were different from those obtained from the isolates of the third individual, which were, themselves, also identical for all isolates. Salmonellae were much more diverse in samples from the carapace of the last individual with five different rep-PCR profiles retrieved. Serotyping of seven isolates representative for each rep-PCR profile identified all isolates as representing Salmonella enterica subspecies enterica serotype Rubislaw, which demonstrates the presence of different strains of potentially human pathogenic salmonellae naturally occurring on turtles even within pristine environments. The frequent detection of these organisms in biofilms on the carapace opens the door for speculations on the role of this habitat as a reservoir for salmonellae, and on potential implications for turtles acting as a dispersal vector.     

Screening of Feed Components for Salmonella with Polyvalent H. Agglutination

The application of polyvalent H serology for screening certain feed components for Salmonella was evaluated. In a comparative study of 1,894 suspicious or known positive samples, Salmonella organisms were detected in 1,141 samples with the conventional method and in 1,134 samples with the polyvalent H method. A statistical analysis of the results obtained by both methods indicated that the polyvalent H method is as reliable as the conventional method. Salmonellae can be detected by this method within 60 hr, whereas conventional methods require at least 4 days. The speed and reliability of the polyvalent H method are desirable for routine quality assurance.     

Interaction of Salt, pH, and Temperature on the Growth and Survival of Salmonellae in Ground Pork

The interaction of temperature, pH, and NaCl concentration on the growth and survival of several strains of salmonellae has been determined in broth and ground pork. Growth of 23 strains occurred in broth at 30 C over a wide range of pH-NaCl combinations; at 10 C, growth was limited to only a few combinations. Cultures which would not grow at 10 C because of the pH-NaCl effect survived for long periods, however. In contrast, cultures which would not grow at 30 C remained viable for only a short time. Results in fresh ground pork were in close agreement with the broth studies. Salmonellae would not grow in ground pork stored at 4 C but would grow in pork containing 3.5% salt stored at 10 C. Salmonellae grew competitively with the natural background flora at 10 C even when the salmonellae constituted less than 5% of the initial flora, and the background flora would grow at a lower temperature than the salmonellae. The data show that, whereas decreasing temperatures increase the inhibitory effects of pH and NaCl, they decrease the lethal effects.