空肠弯曲菌生物学特性上的一些歧异

对195例不同年龄腹泻患者的粪便、690只健康和腹泻畜禽的直肠和泄殖腔拭子及108份腹泻死亡畜禽的脏器材料进行了空肠弯曲菌培养,共分离鉴定458株弯曲菌(其中空肠弯曲菌445株、结肠弯曲菌13株),就其一些生物学特性进行了观察,采用Lior生物学分型法进行了354株弯曲菌(空肠弯曲菌352株、结肠弯曲菌2株)的分型。虽然表明绝大多数生物学性状符合已有文献描述,但也发现在形态、培养和生理生化特性及抗菌药抗性上存有一些歧异,其中最为主要的是483%(215/445)的空肠弯曲菌和231%(3/13)的结肠弯曲菌有萘啶酮酸抗性,11%(5/445)的空肠弯曲菌和76%(1/13)的结肠弯曲菌有噻孢霉素抗性,抗菌药抗性与菌株来源有关(P<0005)。352株空肠弯曲菌生物学分型结果表明在这些动物体内生物型Ⅰ(409%)和Ⅱ(582%)占优势,同一动物体内可有该菌的2个生物型分布。     

中国小型猪空肠弯曲菌的首次分离及其鉴定

目的对中国小型猪空肠弯曲菌进行分离鉴定。方法采集发病小型猪标本,采用细菌学分离培养、生化鉴定、药敏试验、血清学试验、PCR检测等方法进行鉴定,并对细菌的分子生物学特征进行分析。结果分离到1株细菌,经鉴定为空肠弯曲菌（CJp0812）。用空肠弯曲菌flaA基因特异引物对CJp0812细菌的PCR扩增为阳性,经核苷酸序列测定分析证实上述序列与空肠弯曲菌NCTC11168基因序列（GenBank登录号：NC002163）同源性高达99%。结论首次从中国小型猪中分离到了空肠弯曲菌,为进一步研究该菌及开展流行病学调查提供了科学依据。     

基于全基因组序列的弯曲菌特征分析

空肠弯曲菌(Campylobacter jejuni)和结肠弯曲菌(Campylobacter coli)是引起人类腹泻的主要致病菌。传统生化方法在鉴定弯曲菌时存在步骤多、耗时长、通量低等问题。本研究通过利用生物信息学方法对弯曲菌全基因组进行序列、基因注释、耐药基因、多位点序列分型以及CRISPR-Cas系统等分析,挖掘能够有效区分空肠弯曲菌和结肠弯曲菌的高分辨力特征。实验结果表明,空肠弯曲菌和结肠弯曲菌在基因组序列长度、GC含量、基因数量、多位点序列分型以及CRISPR-Cas系统等方面存在显著差异。同时,研究还发现了一段在空肠弯曲菌基因组中广泛存在的高分辨力CRISPR重复序列。这些特征可用于构建能够准确鉴别空肠弯曲菌和结肠弯曲菌的生物信息学方法。     

139株空肠弯曲菌耐药性分析

空肠弯曲菌(Campylobacter jejuni)是最常见的食源性病原菌之一。本研究采用微量肉汤稀释法对分离得到的139株空肠弯曲菌(117株为禽源样本分离株,22株为人源样本分离株)进行耐药性检测。通过对最小抑菌浓度(MIC)的判定结果得出:120株(86. 33%)空肠弯曲菌分离株对6类9组临床常用的抗生素表现出不同程度的耐药,其中禽源空肠弯曲菌耐药率为83. 76%,22株人源空肠弯曲菌均表现出耐药性。对喹诺酮类抗生素表现出高度耐药(环丙沙星80. 58%,萘啶酸77. 70%);对四环素类表现为中等耐药(四环素53. 24%);对部分大环内酯类、氨基糖苷类、林可酰胺类表现为低耐药(庆大霉素7. 19%,阿奇霉素5. 76%,克林霉素6. 47%);对酰胺醇类、部分大环内酯类表现为敏感(氟苯尼考0%,红霉素0%、泰利霉素0%)。139株空肠弯曲菌共产生14种耐药谱型,以TET-CIP-NAL谱型最多,占比38. 13%,耐三重及以上抗生素的多重耐药菌株占比53. 24%。禽源菌株中多重耐药占比46. 15%,人源菌株中多重耐药占比90. 91%。研究结果显示空肠弯曲菌耐药现状不容乐观,尤其对喹诺酮类与四环素类抗生素耐药性较为突出,且过半数菌株为多重耐药。本研究为食源性空肠弯曲菌的防控及临床用药提供参考。     

华南四省食品中空肠弯曲菌分离株的毒力相关基因分析和ERIC-PCR分型

【目的】了解空肠弯曲菌(Campylobacter jejuni)在华南四省食品中的污染状况及其遗传多样性,为食源性病原菌的风险监测和控制提供基础数据。【方法】采用传统国标和MPN组合的方法对华南四省市售生鲜禽畜肉及其制品、生食蔬菜、熟食、水产品、速冻食品、奶制品、食用菌7大类食品进行C.jejuni污染调查,对分离到的C.jejuni菌株进行12种毒力相关基因检测分析和ERIC-PCR分型。【结果】2011-2012年共检测样品558份,检出阳性样品14份,检出率2.51%,其中肉与肉制品的检出率达13.2%,污染量达8.77 MPN/g,获得C.jejuni14株。毒力相关基因分析显示9种毒力相关基因的携带率超过50%,而pldA和wlaN的携带率均为14.3%,virB11的携带率为0。ERIC-PCR分析结果显示15株C.jejuni可分为3簇共10种基因型,其中簇A中有4株的指纹图谱基本相同。【结论】华南四省中的生鲜禽肉类样品,存在不同程度的空肠弯曲菌污染,尤其是福州的鸭肉样品,其MPN值大于110 MPN/g,应加强生禽肉类的食品安全防控。     

汕头市区空肠弯曲菌的电镜观察

自从1972年Dekeyser等首次证实空肠弯曲菌(Campylobacter jejuni)是腹泻较常见的病原菌,我国1981年上海首次分离该菌以来,国内许多地区已先后开展有关分离鉴定工作,初步表明空肠弯曲菌在我国并不少见,本菌的重要性正日益引起重视。但有关该菌的电镜观察,国内外只见零星报道。本文就其电镜形态及其与致病的关系作初步探讨。     

从腹泻患儿粪便中检出空肠弯曲菌

近年来,空肠弯曲菌在世界各国已成为引起腹泻的重要病原菌之一。1982年在我国上海首次报道后,各地已相继有报道。为了解空肠弯曲菌在我市儿童中发病情况和探索其病原特点,我院肠道门诊于1986年9月～1987年8月从549例腹泻患儿粪便中分离出12株空肠弯曲菌,其阳性     

空肠弯曲菌Ⅵ型分泌系统研究进展

Ⅵ型分泌系统(Type Ⅵ Secretion System,T6SS)是一种倒置于细胞膜上的类噬菌体样结构,能够输送效应蛋白并在定殖和生态位建立中发挥作用.近年来,在空肠弯曲菌(Campylobacter jejuni)中发现了T6SS同源基因且能够表达组装成结构完整的T6SS,但T6SS对空肠弯曲菌的毒力影响尚不清...     

MALDI-TOF-MS方法快速鉴定食品中空肠弯曲菌的技术研究

运用基质辅助激光解吸电离飞行时间质谱(matrix-assisted laser desorption/ionization time of-flight mass spectrometry,MALDI-TOF-MS)技术快速鉴定食品中空肠弯曲菌.通过对该方法的样品前处理的选择、稳定性、特异性等方面进行研究,确定了方法...     

空肠弯曲菌增菌方法的研究与血清学分型

本文对空肠弯曲菌增菌培养基作了改进,以增菌法与直接分离法(直接法)对比进行实验。从379份腹泻成人粪便中分离出29株空肠弯曲菌,其中增菌法分离出29株(100%),直接法15株(51.72%)。增菌法提高检出率48.28%,P<0.001,两法差异非常显著,符合率为96.3%,说明增菌法优于直接法。另外,用烛缸培养法从鸡和猪粪便中分离出空肠弯曲菌分别为50和32株。对人、鸡和猪粪便中检出的110株菌作了血清学和生物学分型。从分出的15个血清型谱中发现不同来源菌株有共同性血清型,多集中于45型     

Prevalence of Campylobacter spp. in turkey meat from a slaughterhouse and in turkey meat retail products

One hundred and forty-four samples of chilled turkey meat from six flocks, taken directly from the slaughterhouse, and 100 samples of turkey meat retail products were examined. Over one-quarter (29.2%) of the tested samples from the slaughterhouse were Campylobacter positive, showing high variability in the flocks. The lowest percentage of Campylobacter-positive samples was found in flocks I and III (8.3%), whereas, in flock VI, 91.7% of the samples were Campylobacter positive. Turkey meat retail products showed a prevalence of 34% for Campylobacter. Heat-treated meat was negative for Campylobacter. Quantitative studies of the samples taken at the slaughterhouse revealed a mean log range of 1.9-2.5 CFU g(-1)Campylobacter spp. Results from the quantification of retail products gave a mean log value of 2.1 CFU g(-1).     

PCR-ELISAs for the detection of Campylobacter jejuni and Campylobacter coli in poultry samples

Campylobacter species, primarily Campylobacter jejuni and Campylobacter coli, are regarded as a major cause of human gastrointestinal disease, commonly acquired by eating undercooked chicken. We describe a PCR-ELISA for the detection of Campylobacter species and the discrimination of C. jejuni and C. coli in poultry samples. The PCR assay targets the 16S/23S ribosomal RNA intergenic spacer region of Campylobacter species with DNA oligonucleotide probes designed for the specific detection of C. jejuni, C. coli, and Campylobacter species immobilized on Nucleo-Link wells and hybridized to PCR products modified with a 5' biotin moiety. The limit of detection of the PCR-ELISA was 100-300 fg (40-120 bacterial cells) for C. jejuni and C. coli with their respective species-specific oligonucleotide probes and 10 fg (4 bacterial cells) with the Campylobacter genus-specific probe. Testing of poultry samples, which were presumptive positive for Campylobacter following culture on the Malthus V analyzer, with the PCR-ELISA determined Campylobacter to be present in 100% of samples (n = 40) with mixed cultures of C. jejuni/C. coli in 55%. The PCR-ELISA when combined with culture pre-enrichment is able to detect the presence of Campylobacter and definitively identify C. jejuni and C. coli in culture-enriched poultry meat samples.     

Isolation and phylogenetic analysis of arcobacter spp. in ground chicken meat and environmental water in Japan and Thailand

Prevalence of Arcobacter spp. in chicken meat samples and environmental water samples in Japan and Thailand was investigated. Arcobacter was isolated from 48% of chicken meat samples (20/41) and 23% of river water samples (4/17) from Japan, and 100% of chicken meat samples (10/10) and 100% of canal water samples (7/7) from Thailand. A. butzleri was among the species isolated from all positive samples. About 10% genetic diversity was seen in the rpoB-rpoC in Arcobacters, and phylogenetic trees were divided into two clusters. In both countries, the results suggested that chicken and environmental water were highly contaminated with a genetically diverse population of Arcobacter.     

Antimicrobial resistance of Campylobacter isolates from retail meat in the United States between 2002 and 2007

The emergence of antimicrobial resistance in Campylobacter spp. has been a growing public health concern globally. The objectives of this study were to determine the prevalence, antimicrobial susceptibility, and genetic relatedness of Campylobacter spp. recovered by the National Antimicrobial Resistance Monitoring System (NARMS) retail meat program. Retail meat samples (n = 24,566) from 10 U.S. states collected between 2002 and 2007, consisting of 6,138 chicken breast, 6,109 ground turkey, 6,171 ground beef, and 6,148 pork chop samples, were analyzed. A total of 2,258 Campylobacter jejuni, 925 Campylobacter coli, and 7 Campylobacter lari isolates were identified. Chicken breast samples showed the highest contamination rate (49.9%), followed by ground turkey (1.6%), whereas both pork chops and ground beef had <0.5% contamination. The most common resistance was to doxycycline/tetracycline (46.6%), followed by nalidixic acid (18.5%), ciprofloxacin (17.4%), azithromycin and erythromycin (2.8%), telithromycin (2.4%), clindamycin (2.2%), and gentamicin (<0.1%). In a subset of isolates tested, no resistance to meropenem and florfenicol was seen. C. coli isolates showed higher resistance rates to antimicrobials, with the exception of doxycycline/tetracycline, than those seen for C. jejuni. Pulsed-field gel electrophoresis (PFGE) fingerprinting resulted in 1,226 PFGE profiles among the 2,318 isolates, with many clones being widely dispersed throughout the 6-year sampling period.     

Prevalence of Campylobacter spp., Escherichia coli, and Salmonella serovars in retail chicken, turkey, pork, and beef from the Greater Washington, D.C., area.

A total of 825 samples of retail raw meats (chicken, turkey, pork, and beef) were examined for the presence of Escherichia coli and Salmonella serovars, and 719 of these samples were also tested for Campylobacter spp. The samples were randomly obtained from 59 stores of four supermarket chains during 107 sampling visits in the Greater Washington, D.C., area from June 1999 to July 2000. The majority (70.7%) of chicken samples (n = 184) were contaminated with Campylobacter, and a large percentage of the stores visited (91%) had Campylobacter-contaminated chickens. Approximately 14% of the 172 turkey samples yielded Campylobacter, whereas fewer pork (1.7%) and beef (0.5%) samples were positive for this pathogen. A total of 722 Campylobacter isolates were obtained from 159 meat samples; 53.6% of these isolates were Campylobacter jejuni, 41.3% were Campylobacter coli, and 5.1% were other species. Of the 212 chicken samples, 82 (38.7%) yielded E. coli, while 19.0% of the beef samples, 16.3% of the pork samples, and 11.9% of the turkey samples were positive for E. coli. However, only 25 (3.0%) of the retail meat samples tested were positive for Salmonella. Significant differences in the bacterial contamination rates were observed for the four supermarket chains. This study revealed that retail raw meats are often contaminated with food-borne pathogens; however, there are marked differences in the prevalence of such pathogens in different meats. Raw retail meats are potential vehicles for transmitting food-borne diseases, and our findings stress the need for increased implementation of hazard analysis of critical control point (HACCP) and consumer food safety education efforts.     

Quantification of thermophilic <Emphasis Type="Italic">Campylobacter</Emphasis> spp. in broilers during meat processing

Campylobacter spp. is a common cause of gastrointestinal illness. Since animal products, especially poultry meat, are an important source of human outbreaks of campylobacteriosis, tracing back to processing and initial production is of great interest. Samples were collected at a German poultry slaughterhouse for the estimation of the prevalence of Campylobacter at different processing steps. Quantification of Campylobacter in each of the samples was also performed. Out of 99 samples examined, 51 (51.5%) were positive for Campylobacter, with bacterial counts ranging from log(10) 6.5 cfu sample(-1) for carcasses to log 3.6 cfu ml(-1) for scalding water. The Campylobacter isolates (n = 51) were subtyped by pulsed-field gel electrophoresis using SmaI and KpnI restriction enzymes. Molecular typing showed a multitude of strains with different molecular patterns. Strains found in cloacal swabs before processing could also be isolated from carcasses at different processing steps.     

Development of a combined PCR-culture technique for the rapid detection of Arcobacter spp. in chicken meat

A combined PCR-culture technique was developed to detect Arcobacter spp. in fresh chicken meat. Following a short selective enrichment of chicken samples, bacterial DNA was extracted and amplified using primers targeted at the genes encoding 16S rRNA of Arcobacter spp. The selected primers amplify a 181-bp fragment from all Arcobacter spp., whereas no PCR product is generated for other bacteria, including the closely related Campylobacter and Helicobacter species. The assay was used to screen 96 retail-purchased chicken samples for the presence of Arcobacter spp. Fifty-three percent of the samples analysed were positive for this micro-organism. The assay is simple and sensitive and reduces the amount of time required to positively detect Arcobacter spp. in poultry meat.     

Evaluation of RapidChek Select for the screening of Salmonella in meat and meat products

The efficacy of the RapidChek Select, an alternative rapid method based on lateral flow technology, for the screening of Salmonella in meat and meat products, was compared with the current ISO reference culture-based method. Of the 265 routine samples examined, 61 were found to be positive for Salmonella by both methods. The percentage of agreement between the results of two methods was determined as 98%. All presumptive positive results obtained by the RapidCheck Select were confirmed to be positive by ISO method. For five samples ISO method gave positive result, while RapidChek Select gave negative result. The limit of detection (LOD(50)) of RapidChek Select and ISO methods for minced beef meat samples were 1.00 cfu/25 g and 0.63 cfu/25 g, respectively. For sausage samples, LOD(50) of both methods were 2.00 cfu/25 g. As a result, the high agreement between two methods and the comparable detection limits of two methods showed that the RapidChek Select is an efficient alternative method for the screening of Salmonella in meat and meat products.     

A novel method and simple apparatus for the detection of thermophilic Campylobacter spp. in chicken meat products

Conventional procedures for isolation of thermophilic Campylobacter spp. from chicken are complex, labor intensive, and time-consuming. The objective of this study was to create a novel Campylobacter culturing apparatus. A main concept of the device was based on the ability of Campylobacter to pass through a 0.45 microm pore size filter in viscous media. Preliminary study demonstrated that only viable Campylobacter moved through the membrane filter and could multiply in the enrichment culture. C. jejuni, C. coli, C. lari, and C. upsaliensis in the chicken samples were detected at cell concentrations as low as 10 cfu/g, after 24 h incubation at 42 degrees C. In total, 84 retail chicken samples were comparatively studied using both conventional method and apparatus. Sixteen samples (19.05%) were positive by the apparatus method; 14 (16.66%) of these positive samples contained C. coli and 2 (2.38%) contained C. jejuni. With the conventional method, 7 (8.33%) samples were positive 7 (8.33%) with C. coli. In conclusion, the apparatus detected more positive samples than did the conventional culture method.     

Sensitivity of Campylobacter spp. to irradiation in poultry meat

The sensitivity of Campylobacter jejuni (three strains), Camp. coli (three strains), Camp. fetus (one strain) and Camp. lari (one strain) to irradiation in poultry meat was investigated. There was no significant difference in the counts obtained on Blood or Skirrows agar. Preston agar gave a significantly lower recovery of the pathogens after irradiation so these results were not included in calculations of D ₁₀ values. The D ₁₀ values ranged from 0.12 to 0.25 kGy and there was a significant difference in the radiation sensitivity between different Campylobacter spp. and within strains of the same species. These values indicate that Campylobacter spp. are more radiation-sensitive than Salmonella and Listeria monocytogenes irradiated under similar conditions. Therefore irradiation treatments suggested to eliminate the latter from poultry carcasses would also be sufficient to remove Campylobacter.