常规培养法和酶联荧光免疫分析法在测定空肠弯曲菌中的对比

目的运用常规培养法和酶联荧光免疫分析法对相同食品进行空肠弯曲菌检测,对所得结果进行对比分析。方法对市面上销售的150瓶装酸奶同时采用常规培养法和酶联荧光免疫分析法进行检测。结果采用常规培养法检测出空肠弯曲菌3瓶,其阳性率为2.0%;采用酶联荧光免疫分析法检测出空肠弯曲菌7瓶,其阳性率为4.7%。结论酶联荧光免疫分析法的检出率较高,且方法操作简单、省时,建议有条件的检测机构尽量采用酶联荧光免疫分析法对空肠弯曲菌进行检测。     

滤膜驱动分离培养法在一起空肠弯曲菌腹泻疫情中的应用

目的应用滤膜驱动分离培养法对一起腹泻疫情的系列样本进行空肠弯曲菌的检测和培养,并对该方法及其增菌效果进行评价。方法收集一起腹泻疫情的18份粪便标本,采用实时荧光PCR方法为疫情定性为空肠弯曲菌感染,应用滤膜驱动分离培养法对粪便标本进行空肠弯曲菌增菌和分离培养,并对增菌效果进行比较分析。结果采用荧光PCR方法检测到18份标本中有11份空肠弯曲菌核酸阳性,阳性率为61.11%(11/18);应用滤膜驱动分离培养法,对18份标本进行空肠弯曲菌的分离和鉴定,共分到8株菌,分离率为44.44%(8/18);其中3份核酸阳性标本,由于菌液含量低,无法用培养方法检出。11份核酸阳性粪便标本经增菌培养后,Ct平均值由原来的27.44变为23.50,增菌效果显著(P0.05)。结论采用滤膜驱动分离培养法对粪便标本尤其是冻融粪便进行空肠弯曲菌的增菌和分离培养,能有效提高菌株的分离率,避免资源浪费,可做为常规监测或大规模样本筛检的可行性检测方法。     

口服红霉素要注意十大事项

<正>红霉素一种大环内酯类抗生素,对葡萄球菌属、各组链球菌、革兰阳性杆菌、支原体、衣原体、螺旋体等均有较强的抗菌活性。用于治疗敏感细菌所致的呼吸道感染、金葡菌性皮肤感染、沙眼衣原体引起的结膜炎、空肠弯曲菌性肠炎等。对感染军团菌、肺炎支原体、空肠弯曲肠菌等,红霉素为首选用药。为安全有效使用红霉素,应了解使用红霉素的注意事项。     

尿培养标本直接细菌鉴定和药敏试验的价值

目的:探究直接细菌鉴定和药敏试验在尿培养标本检测中的应用价值。方法:选择我院自2017年1月至2017年9月收集的75份疑似尿路感染住院患者的中段尿标本作为研究对象,将尿标本平均分成三份,分别采用直接方法(美华MA120系统)、常规培养法以及经典手工方法进行细菌鉴定和药敏试验,并且以经典手工方法检测结果作为金标准,对比另外两种检测方法的细菌鉴定和药敏实验正确率、错误率。结果:常规培养法对细菌鉴定的诊断的准确率(86.7%)与直接检测法诊断的准确率(85.3%)对比无显著差异,常规培养法对株革兰阴性球菌药敏分析诊断的准确率(95.3%)与直接检测法诊断的准确率(93.0%)对比无显著差异,无统计学意义(P0.05);常规培养法对株革兰阳性球菌药敏分析诊断的准确率(93.8%)明显高于直接检测法诊断的准确率(87.5%),两组对比差异显著(P0.05)。结论:对尿培养标本采用直接细菌鉴定和药敏试验进行检测,其对细菌鉴定和阴性球菌药敏分析诊断的准确率均较高,对阳性球菌药敏分析诊断的准确率略低于常规培养方法,但其操作更简便和用时更少,因此其检测结果可作为临床尿路感染患者合理使用抗菌药物的有效参考依据。     

PCR技术快速检测空肠弯曲菌方法的建立

目的建立空肠弯曲菌的快速检测方法。方法用PCR法扩增标本菌株中弯曲菌属16SrRNA片段、空肠弯曲菌M apA片段和结肠弯曲菌CeuE片段,通过电泳后对产物进行分析。结果11份标本中有7份检出弯曲菌属16SrRNA片段,其中3份样品中检测到空肠弯曲菌M apA片段,4份样品中检测到结肠弯曲菌CeuE片段。结论PCR技术可用于空肠弯曲菌的快速检测。     

抽气加烛缸法培养空肠弯曲菌的初步探讨

作者根据原有烛缸法的基础上再抽去部分的气体,并用化学方法增加适量的CO_2来检测弯曲菌空肠亚种,获得满意的结果。实验方法:取腹泻患者粪便标本,分离接种于弯曲菌空肠亚种的选择性平板培养基中。将分离好的平皿置于干燥器中,称取碳酸氢钠5.5g、酒石酸4g与一只     

环介导等温扩增法检测空肠弯曲菌

目的 空肠弯曲菌是重要的食源性疾病病例标本的病原菌检测项目,用环介导等温扩增(Loop-mediated isothermal amplification,LAMP)的方法,对食源性致病菌空肠弯曲菌进行快速检测。方法 以空肠弯曲菌 mapA 基因序列为检测靶基因,建立了空肠弯曲菌的LAMP 快速检测方法。空肠弯曲菌的LAMP反应由DNA扩增试剂盒提供的反应体系加入引物和模板进行。做了空肠弯曲菌DNA模板灵敏度的检测、菌液灵敏度的检测和特异性检测。结果 我们方法对空肠弯曲菌DNA 的检测灵敏度为84.5fg/反应管;对空肠弯曲菌菌液检测灵敏度为0.8 CFU/反应管;特异性100%。结论 对食源性疾病腹泻样本的空肠弯曲菌检测项目,可以采用mapA-LAMP方法进行等温扩增初筛,初筛阳性的样品再有针对性地进行后续的传统培养。     

空肠弯曲菌感染与Guillain-Barré综合征

目的 :探讨空肠弯曲菌感染与Guillain -Barr啨综合征 (GBS)发病的关系以及空肠弯曲菌感染和抗 -GM1抗体与疾病严重程度的关系。方法 :用ELISA方法检测 85例GBS患者血清中的空肠弯曲菌IgG、IgM、IgA抗体及抗 -GM1抗体 ,同时从患者的大便中培养空肠弯曲菌 ;根据Hughes报道的方法对病情轻重进行评分。结果 :GBS患者空肠弯曲菌感染率为 5 1 8% ,抗 -GM1抗体检出率为 42 4% ;近期空肠弯曲菌感染者抗 -GM 1抗体检出率(6 3 6 % )显著高于未感染者 (19 5 % ) (P <0 .0 0 1) ;空肠弯曲菌感染组与非感染组以及抗 -GM1抗体阳性组与阴性组之间病情轻重均无显著性差异 (P均 >0 .0 5 )。结论 :空肠弯曲菌感染是GBS的重要诱发因素之一 ;血清中抗-GM 1抗体的存在与近期空肠弯曲菌感染有关 ,但空肠弯曲菌感染及抗 -GM 1抗体的存在与疾病严重程度无关。     

人源空肠弯曲菌抗生素敏感性检测

为进一步掌握空肠弯曲菌的药敏规律,指导治 疗与预防,我们将1997年分离到的人源空肠弯曲菌 进行了14种抗菌药物敏感性测定,结果报告如下。 l 材料与方法 1.l 菌株来源 16株空肠弯曲菌为1997年8月 于河北省满城县儿童粪便标本中分离所得。 1.2 培养基 Mueller-Hinton(MH)培养基和微氧 袋均为英国Oxoid公司产品。 1.3 药敏纸片中国北京药品生物制品检定所产 品,有效期内使用。     

空肠弯曲菌的一种简易保存法

<正> 空肠弯曲菌是引起人类腹泻的常见病原菌之一。但因本菌分离后死亡较快,难于保存,影响了有关研究工作的开展。采用半固体酵母浸膏培养基保存于37℃,该菌可存活半年以上,获得较满意的效果。菌种来源:20株空肠弯曲菌自腹泻病人粪便中分离,6株自鸡粪便中分离,全部菌株均经生化试验鉴定符合空肠弯曲菌特征。菌种保存用培养基——半固体酵母浸膏培养     

应用PCR方法对生鸡肉中分离到的空肠弯曲菌进行鉴定

目的：对北京市市场销售的肉制品的空肠弯曲菌污染的状况进行调查,并建立快速的弯曲菌PCR鉴定的方法。方法：采用中国疾病预防控制中心下发的监测方法对北京市超市和集贸市场销售的鸡肉采集100件并进行检测,生化鉴定采用API Campy系统进行鉴定,同时采用针对空肠弯曲菌h ipO设计的引物对分离的弯曲菌进行鉴定。结果：共有4件检出空肠弯曲菌,采用PCR方法进行鉴定的结果与生化方法鉴定的结果一致。结论：在北京销售的鸡肉中存在空肠弯曲菌的污染,带菌率为4%,针对h ipO设计的引物可以对空肠弯曲菌进行快速鉴定。     

常见食源性致病菌基因芯片鉴定技术

目的：建立鉴定食源性致病菌的基因芯片技术。方法：采用复合PCR方法扩增致病菌特异性DNA片段，结合基因芯片杂交技术鉴定不同的食源性致病菌。结果：本研究完成了采用3对引物的复合PCR扩增；不同致病菌特异性核苷酸片段同时布阵在一张芯片上；标记物质采用了非放射性的生物素，经不同标准菌种、实际检验样品和水平测试样品的考核验证，该鉴定系统灵敏度达620cfu／g，特异性高，基因芯片质量稳定，该鉴定系统基本含盖了目前食源性致病菌鉴定的需要。结论：常见食源性致病菌基因芯片鉴定技术，可为常规细菌检验方法的最终鉴定提供进一步佐证，尤其在一些培养条件苛刻的致病菌（产单核李斯特菌、弯曲菌等）的鉴定，以及在VITEK仪无法鉴定和手工鉴定判断误差大的情况下，基因芯片方法将发挥其独特的技术性优势，大大提高检验鉴定结果的准确性。     

Rapid detection of point mutations in domain V of the 23S rRNA gene in erythromycin-resistant Campylobacter isolates by pyrosequencing

Campylobacter is one of the most common foodborne pathogens causing acute gastroenteritis in humans. Erythromycin, a macrolide antibiotic, is the first-choice treatment for Campylobacter infections, and failure to eradicate Campylobacter is usually due to macrolide resistance. The most important mechanism responsible for macrolide resistance in Campylobacter is mediated by point mutations at position 2074 or 2075 in the peptidyl-transferase region of domain V of the 23S rRNA gene. In this study, the minimum inhibitory concentrations of 58 Campylobacter isolates (C. jejuni: n?=?37; C. coli: n?=?21) obtained from chickens were measured by agar dilution. Isolates were subjected to both pyrosequencing and Sanger sequencing methods to detect the 2074 and 2075 point mutations and evaluate the efficacy of the pyrosequencing method. The A2075G mutation was found to be the predominant mutation associated with erythromycin resistance. Compared with traditional methods, pyrosequencing is a novel, rapid, low-cost, and quantitative technology for detecting erythromycin resistance in Campylobacter.     

Detection of Campylobacter jejuni from the skin of broiler chickens, ducks, squab, quail, and guinea fowl carcasses

Campylobacter jejuni is often found on broiler carcasses and can cause gastroenteritis in humans. Both carcass rinses and swabs of the skin have been utilized to ascertain the prevalence of C. jejuni in the processing plant. Not all poultry commodities are equally capable of carrying C. jejuni on the carcass skin. Our objective was to measure the probability of C. jejuni detection (sensitivity) for the skin swabbing method followed by enrichment in semisolid media, and to ascertain the sensitivity of this method for commercial broiler, duck, squab, quail, and guinea fowl. The probability of detecting skin contaminated with C. jejuni was significantly higher for broiler chicken compared to retail duck, squab, quail, or guinea fowl for 10 or 100 colony-forming units (CFU)/in2 of skin (1 in2 = 1 square inch = 2.5 x 2.5 cm). Thirty-three percent (10 CFU/in2) and 100% (100 CFU/in2) of skin samples from broilers were positive for C. jejuni at the levels inoculated while 7-20% and 47-80% of skin samples were detected as contaminated with C. jejuni at 10 or 100 CFU/in2 for retail duck, squab, quail, and guinea fowl, respectively. Our method of using skin swabs and enrichment with semisolid media generated a sensitivity of almost 100% for detecting C. jejuni at 1000 or 10,000 CFU/in2 skin regardless of poultry species. The level of contamination that our method could detect with 50% and 90% reliability (DT50 and DT90) was 14 and 79 (broilers); 67 and 406 (squab); 39 and 226 (quail); 69 and 400 (guinea fowl); 69 and 400 (duck) CFU/in2 of skin, respectively.     

应用多重PCR快速检测空肠弯曲菌

目的:建立能快速、特异检测空肠弯曲菌的多重PCR技术。方法:选用针对空肠弯曲菌外膜蛋白A(mapA)基因和马尿酸酶(hipO)基因的2对引物,在同一扩增体系中进行PCR,优化反应体系,测定特异性和灵敏度,并进行了鸡肉模拟样品检测。结果:该方法扩增目的基因片段分别为589 bp和323 bp,特异性和灵敏度均高。细菌纯培养物的检测灵敏度为105 cfu/ml,鸡肉模拟样品42℃预增菌36 h后检测灵敏度能达到101 cfu/ml。结论:初步建立能快速、灵敏、特异地测定空肠弯曲菌的多重PCR检测技术。     

Campylobacter jejuni in acute diarrhoea in infancy and its relation to faecal leucocytic count.

A prospective study was carried out to identify the relative risk of Campylobacter jejuni infection in 50 infants with acute diarrhoea, 24 infants with acute resistant diarrhoea and 25 healthy normal infants as a matching control group. Faecal samples were collected from the three groups and were cultured on both selective media for Campylobacter and other media for isolation of other organisms. Direct stool smears, stained with methylene blue, were examined for detection of faecal leucocyte in all samples. Campylobacter jejuni were isolated from 4 cases (8.0%) of the acute diarrhoeal group and 4 cases (16.6%) of acute resistant diarrhoeal group. The other bacterial pathogens isolated from our cases were Salmonella, Shigella, E. Coli, Proteus mirabilis, Vibrio Parahaemolytious Klebsiella, Streptococcus faecalis and Candida albicans. All cases from whom Campylobacter was isolated were bottle fed and their ages were below 6 months. Smears for faecal leucocytes were positive in 100% of Campylobacter isolated cases, 60% of Salmonella, 50% of Shigella, 14% of E. Coli and 100% were negative in all other cases. Thus it can be recommended that any case presenting with acute diarrhoea should be initially screened by faecal leucocytic counting, positive cases should be cultured for Campylobacter jejuni detection in addition to cultures for other organisms detection.     

Specific detection of Campylobacter jejuni from faeces using single nucleotide polymorphisms

Specimens of human faeces were tested by a rapid strategy for detection of Campylobacter jejuni lineages by the presence of specific single nucleotide polymorphisms (SNPs) based on the C. jejuni multi locus sequence typing (MLST) scheme. This strategy was derived from analysis of the MLST databases to identify clonal complex specific SNPs followed by the design of real-time PCR assays to enable identification of six major C. jejuni clonal complexes associated with cases of human infection. The objective was to use the MLST SNP-based assays for the direct detection of C. jejuni by clonal complex from specimens of human faeces, and then confirm the accuracy of the clonal complex designation from the SNP-based assays by performing MLST on the cultured faecal material, this targeted at determining the validity of direct molecular specimen identification. Results showed it was possible to identify 38% of the isolates to one of the six major MLST clonal complexes using a rapid DNA extraction method directly from faeces in under 3 h. This method provides a novel strategy for the use of real-time PCR for detection and characterization beyond species level, supplying real-time epidemiological data, which is comparable with MLST results.     

Detection of Campylobacter jejuni in dairy farm environmental samples using SYBR Green real-time polymerase chain reaction

The aim of this study was to evaluate a SYBR Green based real-time PCR assay using well-characterized primers to detect Campylobacter jejuni in naturally contaminated dairy farm environmental samples. Specificity of the assay was determined with 62 C. jejuni strains and 120 non-C. jejuni strains. Peak melting temperature obtained with melting curves specific for C. jejuni was 77.5 degrees C. Standard curves were constructed using mean threshold cycle (C(T)) and various concentrations of C. jejuni ranging from 10(0) to 10(8) colony forming units (CFU)/mL, which resulted in a linear relationship between C(T) and log input DNA. Correlation coefficients of standard curves based on pure culture of C. jejuni in broth and spiked cells in lagoon water were R(2) = 0.995 (slope = 3.21) and R(2) = 0.988 (slope = 3.22), respectively, and sensitivity limits were <10 and >10(3) CFU/mL, respectively. After 24-h enrichment, total C. jejuni counts of all samples spiked with 10(0) CFU/mL reached >10(5) CFU/mL, and the detection limit was improved from >10(3) CFU/mL to <10 CFU/mL of inoculum in broth. Eighty-two dairy farm environmental samples, including fecal slurry, feed/silage, lagoon water, drinking water, bulk tank milk, farm soil, and bedding material, were analyzed. The real-time PCR assay detected C. jejuni in 25 (30.4%) of 82 samples, with 17 (68%) of these samples being culture positive for C. jejuni. All samples that were positive by standard culture methods were also positive by the real-time PCR method. Mean C( T ) values of 48-h enriched cultures for 17 PCR-positive/culture-positive samples and eight PCR-positive/culture-negative samples were 21.4 +/- 3.6, and 34.6 +/- 1.5 (p < 0.0001), respectively. C( T ) values for negative samples were >38.0. These results indicate that the SYBR Green real-time PCR assay provides a specific, reproducible, and simple method for detecting C. jejuni in dairy farm environmental samples.