Campylobacter spp. and their antimicrobial resistance patterns in poultry: an epidemiological survey study in Turkey

The current study aimed at determining the prevalence and the antimicrobial resistance profiles of thermophilic Campylobacter spp. infecting broiler chickens. A total of 240 caecal samples from six slaughterhouses were examined for the presence of Campylobacter spp. C. jejuni was detected in 40.4% (97/240) of the samples and C. coli in 12.1% (29/240). The agar disc diffusion method and the E-test were used for testing the antimicrobial susceptibility of C. jejuni and C. coli isolates. C. jejuni isolates were most resistant to nalidixic acid (79.4%) followed by tetracycline (76.3%), ciprofloxacin (74.2%) and enrofloxacin (15.5%). Among the C. coli isolates, the frequency of resistance to nalidixic acid and ciprofloxacin was the same at 65.5%. The predominant profiles of multidrug resistance to three or more antimicrobials in C. jejuni and C. coli were determined as tetracycline/nalidixic acid/ciprofloxacin resistance (48.5%) and tetracycline/nalidixic acid/ciprofloxacin/enrofloxacin resistance (51.7%), respectively. To prevent the transmission of antimicrobial-resistant bacteria of animal origin to humans, it should be noted that high proportions of multidrug resistance were found in both species.     

应用PCR技术检测空肠弯曲菌及结肠弯曲菌的研究

本研究根据空肠弯曲菌及结肠弯曲菌的16s rRNA基因相同保守序列来设计引物,建立了一种PCR检测方法,该方法具有较强的特异性及灵敏性,其对空肠弯曲菌及结肠弯曲菌的最低检测限度为5 CFU/mL,它无需增菌培养过程可以直接对样品进行检验,在对鸡肉样品空肠弯曲菌及结肠弯曲菌的检测中取得了较好的效果.     

Sensitivity of Direct Culture,Enrichment and PCR for Detection of Campylobacter jejuni and C. coli in Broiler Flocks at Slaughter

Broiler chicken flocks are a significant source of Campylobacter jejuni and Campylobacter coli that result in the major public health problem of campylobacteriosis. Accurate estimates of the prevalence of both C. coli and C. jejuni in flocks would enhance epidemiological understanding, risk assessment and control options. This study combined results from a panel of 10 detection tests (direct culture, enrichment and PCR) on caecal samples from flocks at slaughter. A parallel interpretation approach was used to determine the presence of Campylobacter spp. and for C. jejuni and C. coli individually. The sample was considered positive if at least one method detected the target and this interpretation was taken to represent a ‘proxy gold standard’ for detection in the absence of a gold standard reference test. The sensitivity of each individual method to detect Campylobacter spp., C. jejuni and C. coli was then estimated relative to the proxy gold standard. Enrichment in adapted Exeter broth (deficient in polymyxin B) with a resuscitation step was 100% sensitive, whilst direct culture on modified charcoal cefoperazone deoxycholate agar (mCCDA) was highly sensitive (97.9%). Enrichment methods using Preston broth and Bolton broth were significantly less sensitive. Enrichment in Exeter broth promoted the recovery of C. jejuni, whilst enrichment in Bolton broth favoured C. coli. A RT‐PCR detection test could identify 80% of flocks that were co‐colonised with both species. This study found that 76.3% (n = 127) of flocks were colonised with Campylobacter spp. The majority (95.9%) of Campylobacter‐positive flocks were colonised with C. jejuni; however, approximately one‐third of positive flocks were simultaneously colonised with both C. jejuni and C. coli. The findings highlight the impact of different detection methodologies on the accuracy of the estimated incidence of both C. jejuni and C. coli entering the abattoir within broiler flocks and the associated public health risks.     

空肠弯曲菌保存方法的研究

本文研究了１５株不同菌株和同一菌株在不同保存温度和培养基中的保存效果，结果表明：用同一菌株作该菌保存方法研究缺乏整体代表性；在所试的两种冰冻温度中，以－２５℃极显著地优于－１０℃保存（Ｐ＜０．０１）。在－２５℃下冻融１次保存该菌是最佳保存条件，于Ｐ－基、ＦＭＰＧ和ＭＳＧ培养基中，菌株１００％存活的期限分别为１２个月以上、６～９个月间（ＦＭＰＧ和ＭＳＧ），菌株５０％存活的期限三者均在１２个月以上。     

空肠弯曲杆菌耐药性及其主要相关基因初步分析

从307份鸡盲肠内容物样品中分离到47株空肠弯曲杆菌,并测定这些空肠弯曲杆菌耐药性,结果显示对喹诺酮类药(环丙沙星)耐药性为31.9%;对氨苄青霉素,克林霉素和红霉素耐药性较高,分别达到85.1%、83.0%和83.0%;而对痢特灵和庆大霉素则较为敏感,敏感度分别为85.1%和74.5%。有89.4%的菌株显示多重耐药性。利用MAMAPCR技术,对获得的47株空肠弯曲杆菌进行检测,结果显示对环丙沙星耐药的15株空肠弯曲杆菌均检测出其在gyrA基因257位发生点突变,30株对喹诺酮药物敏感的菌株均未检出。另外2株对环丙沙星耐药的菌株,有1株检测出点突变。     

空肠弯曲杆菌生物学特性及分型技术研究新进展

Campylobacter jejuni (C.jejuni) is a major pathogen of human campylobacteriosis, as well as one of the major food-borne pathogen. It can not only cause human diarrhea, but also cause many systemic diseases such as endocarditis, arthritis, septisis. The most serious complication it can cause is the Guillain Barre syndrome. In developed countries, C.jejuni even has a trend to excess Salmonella and be reported as next Salmonella, which has caused a worldwide concern. Analysis of biological characteristics for C.jejuni can help us to distinguish it with other pathogens. Typing methods play important roles in the identification, monitoring and prevention of C.jejuni. It can also help us to understand its virulence and resistance mechanisms to C.jejuni. This paper reviews advances of C.jejuni in biological characteristics, serotyping and genotyping methods so as to help us further understand its new progress in these areas.     

空肠弯曲菌耐药机制研究进展

空肠弯曲菌是一种全球关注的人兽共患病病原菌,由于滥用抗生素而造成其耐药性不断增强,成为公共卫生日益关注的问题.各类抗生素作用位点的基因突变是诱发空肠弯曲菌对各类临床常用的抗生素产生耐药的主要原因,同时细菌对药物的外排机制也在其对抗生素耐药过程中发挥了一定的作用.文章对空肠弯曲菌对各类临床常用抗生素的耐药机制做一综述.     

动物源性空肠弯曲杆菌耐药性地域差异性分析

从上海家禽批发市场采集鸡盲肠内容物样品,分离到47株空肠弯曲杆菌,从上海屠宰场采集猪盲肠内容物样品,分离到99株空肠弯曲杆菌,分别测定分析了这些空肠弯曲杆菌耐药性; 通过对不同产地鸡源空肠弯曲杆菌耐药性状况比较结果显示,江苏南通、江苏海门和浙江金华分离到的鸡源空肠弯曲杆菌对9种抗生素(见表1)都有不同程度的耐药性,对万古霉素、红霉素和氨苄青霉素的耐药性达到100%。江苏海门、上海崇明和浙江温岭、金华等地的菌株分别对克林霉素和氨苄青霉素显示了高耐药性,分别达到90%、100%;通过对不同产地猪源空肠弯曲杆菌耐药性状况比较结果显示,山东莘县分离到的猪源空肠弯曲杆菌对9种抗生素都有不同程度的耐药性,对万古霉素、红霉素、克林霉素和氨苄青霉素的耐药性达到80%以上。河南孟津和安徽淮北分离到的猪源空肠弯曲杆菌对除头孢拉定之外的8种抗生素都有不同程度的耐药性,对氨苄青霉素的耐药性达到100%。结果显示动物源性空肠弯曲杆菌耐药性存在明显的地域差异性。     

空肠弯曲菌致病机理研究进展

空肠弯曲菌是一种人畜共患病病原菌 ,可以引起人和动物发生多种疾病 ,并且是一种食物源性病原菌 ,被认为是引起全世界人类细菌性腹泻的主要原因 ,对空肠弯曲菌的致病机理的研究越来越多。其致病因素包括粘附、侵袭、产生毒素和分子模拟机制等四个方面 ,通过分子分子模拟机制可以引起最严重的并发症 -格林 -巴利综合征。空肠弯曲菌可以通过产生细胞紧张性肠毒素、细胞毒素和细胞致死性膨胀毒素而致病。就空肠弯曲杆菌致病机理研究现状作了概述。     

空肠弯曲杆菌鞭毛的研究进展

空肠弯曲杆菌是一种人畜共患的食物源性病原菌,可引起人和动物细菌性腹泻,且该菌的感染率逐年递增。鞭毛是细菌菌体的一种特殊结构,与菌体的运动性密切相关,有助于其躲避有害环境,同时鞭毛在细菌的致病性等方面也起着重要作用。研究发现,空肠弯曲杆菌的发病机制与鞭毛在宿主上皮细胞的定植力、黏附、侵袭力及毒素的产生密切相关。文章概述了空肠弯曲杆菌鞭毛结构、功能、调控机制及相关基因等方面的研究进展,通过归纳总结已知基因缺失突变对鞭毛的影响,从分子水平了解鞭毛的调控机制,从而探讨空肠弯曲杆菌的致病机理,以期为降低其感染率提供理论依据。     

Prevalence and antimicrobial susceptibility of Campylobacter in broiler flocks in Japan

Campylobacter was isolated from 67 (47.2%) of 142 broiler flocks between September 2009 and February 2010. The prevalence of Campylobacter in broiler flocks was significantly lower during January and February than it was from September to December. Campylobacter colonization was more common in flocks that were not provided with a disinfected water supply, which was consistent with the findings of a previous study. The prevalence of antimicrobial drug‐resistant Campylobacter spp. was investigated, and the minimum inhibitory concentrations of eight antimicrobial agents were determined for 122 Campylobacter jejuni isolates and 46 Campylobacter coli isolates from broiler flocks between 2007 and 2010. In this study, 29.5% (36/122) of C. jejuni isolates and 41.3% (19/46) of C. coli isolates were resistant to enrofloxacin (ERFX), whereas all isolates were susceptible to erythromycin. Furthermore, the ERFX‐resistant isolates were tested for susceptibility to other classes of antimicrobial agents, and 55.6% (20/36) of ERFX‐resistant C. jejuni isolates and 47.4% (9/19) of ERFX‐resistant C. coli isolates were resistant to at least one of aminobenzyl penicillin, dihydrostreptomycin and oxytetracycline. To avoid an impact of antimicrobial drug‐resistant Campylobacter spp. on the efficacy of antimicrobial treatment for human campylobacteriosis, prudent use of antimicrobial agents is a requisite. The use of antimicrobial agents should be accompanied by various approaches such as prevention of Campylobacter colonization in broiler flocks with the aim of lowering the occurrence of Campylobacter infection in humans.     

鸡空肠弯曲菌灭活苗的制备及其免疫保护试验研究

用分离到的空肠弯曲菌制备鸡空肠弯曲菌甲醛灭活疫苗 ,并进行了该病的免疫保护实验。制备 2× 10 9个 / m L浓度的细菌悬液 ,4m L / L浓度的甲醛溶液作用 12 h灭活后制成空肠弯曲菌灭活苗 ,免疫 2周龄的 SPF雏鸡 ,14 d后 ,进行攻击实验 ,观察其临床症状 ;并分别于感染后 7d和 14 d捕杀 ,观察病理学变化及进行细菌分离实验以揭示其保护率。结果表明 ,鸡空肠弯曲菌灭活苗的保护率可达 10 0 % ,说明该苗经口免疫后安全可靠 ,无毒副作用     

Low levels of antibacterial drug resistance expressed by Gram-negative bacteria isolated from poultry carcasses in New Zealand

Abstract

AIM: To provide baseline data on the levels and patterns of antibacterial drug resistance expressed by Gram-negative bacteria isolated from poultry carcasses in New Zealand.

METHODS: Between July and December 2006, isolates of Escherichia coli (n=407) and Salmonella spp. (n=3) originating from carcass-rinse samples were submitted by testing laboratories affiliated to five major poultry processing plants. Isolates of Campylobacter jejuni (n=193) originating from retail poultry carcasses in 2005–2006 were retrieved from the Massey University archives. All isolates underwent disc diffusion susceptibility testing against panels of 12 (Enterobacteriaceae) and six (Campylobacter spp.) antibacterial drugs. Cephalothin-resistance in isolates of E. coli was confirmed using ETest strips, and confirmation of the resistance phenotypes for a subset of C. jejuni isolates used microbroth dilution assays. Patterns within the resistance phenotypes of the isolates were investigated using hierarchical clustering, and logistic regression modelling.

RESULTS: The majority of isolates (71.5% E. coli, 99% C. jejuni, and all three Salmonella spp. isolates) were fully susceptible to the drugs that were tested. Four (1%) E. coli isolates showed resistance to three or more drugs. The proportions of susceptible E. coli differed between the five processing plants. Resistances were detected in E. coli isolates, using disc diffusion to cephalothin (18.2%), ampicillin (4.4%), tetracycline (4.4%) and gentamicin (1.5%). There was an association between cephalothin-resistant isolates of E. coli and decreased susceptibility to gentamicin. Using ETests to ascertain the minimum inhibitory concentrations (MIC) of E. coli for cephalothin gave inconsistent results. One of 193 C. jejuni isolates was resistant to erythromycin, and microbroth dilution assays confirmed that this panel of C. jejuni was generally susceptible to antibacterial drugs.

CONCLUSIONS: The levels of resistance shown by Gram-negative bacteria isolated from chicken carcasses in New Zealand are among the lowest reported around the world. No resistance to extended-spectrum cephalosporin drugs was detected in E. coli, suggesting that CTX-M and AmpC beta-lactamases are rare or absent. Salmonella spp. are rarely isolated from poultry carcasses during routine testing in New Zealand, and the isolates identified during this study were fully susceptible to the drugs tested. A panel of C. jejuni isolates originating from retail poultry carcasses were susceptible to first-line and second-line antibacterial drugs. The use of cephalothin as a marker of resistance to first-generation cephalosporins may not be appropriate for non-type-specific E. coli of animal origin.     

The pathology of natural and experimentally induced Campylobacter jejuni abortion in sheep

We describe here the gross and microscopic lesions in 18 experimentally induced and 120 natural Campylobacter abortions. In natural Campylobacter abortions, gross lesions were reported infrequently; placentitis was recorded in 6% and hepatic lesions in 4% of our field cases. Placentitis was the microscopic lesion identified most consistently in natural abortions (93%) and was often observed in association with abundant bacterial colonies in chorionic villi (54%) and less often with placental vasculitis (13%). In natural abortions, suppurative fetal pneumonia (48%), necrosuppurative hepatitis (16%), and purulent meningitis (7%) were also observed. The better-preserved specimens from experimentally induced abortions were utilized to define placental changes more precisely. Placentitis was identified in all 18 experimentally induced abortions and was observed most consistently in the chorionic villus stroma (100%), often accompanied by suppurative surface exudate (89%). An inflammatory infiltrate was less commonly identified in the cotyledonary hilus (39%) and intercotyledonary placenta (22%). Bacteria were visualized in H&E-stained sections in 89% of placentas from experimentally infected ewes, primarily as well-demarcated bacterial colonies within subtrophoblastic, sinusoidal capillaries (89%), in the cotyledonary villus stroma (89%), and within the cytoplasm of trophoblasts (22%). Transmission electron microscopy and immunohistochemistry confirmed that the vast majority of the well-demarcated bacterial colonies characteristic of Campylobacter abortion were within subtrophoblastic sinusoidal capillaries. The most characteristic microscopic lesions identified in cases of Campylobacter abortion in sheep were placentitis with placental bacterial colonies, placental vasculitis, and fetal pneumonia.     

空肠弯曲菌肠毒素产生最佳培养条件的研究

通过对C.jejuni产生CE的最佳培养基及多粘菌素B、Fe     

Characterization of Campylobacter Jejuni/Coli Isolated from Different Sources

Characteristics of 50 strains of G. jejuni/coli isolated from rectal swabs, caecal contents or bile of cow, sheep, goat, swine, broiler chicken and man were investigated. All the strains grew at 30 °C, 35°C 42°C, but not at 25 °C. The strains were nalidixic acid sensitive. Differences were noticed in salt-tolerance, in the growth with 2,3,5 tri-phenyltetracolium chloride and in the hippurate hydrolysis. Growth of the strains with certain bile salts was also investigated and the results are discussed.     

Combined Campylobacter jejuni and Campylobacter coli Rapid Testing and Molecular Epidemiology in Conventional Broiler Flocks

Campylobacter spp. are important causes of bacterial zoonosis, most often transmitted by contaminated poultry meat. From an epidemiological and risk assessment perspective, further knowledge should be obtained on Campylobacter prevalence and genotype distribution in primary production. Consequently, 15 Austrian broiler flocks were surveyed in summer for their thermophilic Campylobacter spp. contamination status. Chicken droppings, dust and drinking water samples were collected from each flock at three separate sampling periods. Isolates were confirmed by PCR and subtyped. We also compared three alternative methods (culture‐based enrichment in Bolton broth, culture‐independent real‐time PCR and a lateral‐flow test) for their applicability in chicken droppings. Twelve flocks were found to be positive for thermophilic Campylobacter spp. during the entire sampling period. Seven flocks (46.6%) were contaminated with both, C. jejuni and C. coli, five flocks harboured solely one species. We observed to a majority flock‐specific C. jejuni and C. coli genotypes, which dominated the respective flock. Flocks within a distance <2 km shared the same C. jejuni genotypes indicating a cross‐contamination event via the environment or personnel vectors. Multilocus sequence typing (MLST) of C. jejuni revealed that the majority of isolates were assigned to globally distributed clonal complexes or had a strong link to the human interface (CC ST‐446 and ST4373). The combination of techniques poses an advantage over risk assessment studies based on cultures alone, as, in the case of Campylobacter, occurrence of a high variety of genotypes might be present among a broiler flock. We suggest applying the lateral‐flow test under field conditions to identify ‘high‐shedding’ broiler flocks at the farm level. Consequently, poultry farmers and veterinarians could improve hygiene measurements and direct sanitation activities, especially during the thinning period. Ultimately, real‐time PCR could be applied to quantify Campylobacter spp. directly from chicken droppings and avoid non‐interpretable results achieved by culture‐dependent methods.     

河南省鸡源结肠弯曲杆菌的分离鉴定及耐药性分析

[目的]掌握河南省鸡源结肠弯曲杆菌的分布和对抗菌药物的耐药情况。[方法]从养殖场抽取鸡泄殖腔拭子460份进行结肠弯曲杆菌分离培养,采用PCR、基质辅助激光解吸电离飞行时间质谱进行鉴定,并通过微量肉汤稀释法对分离菌株进行药物敏感性分析。[结果]共分离结肠弯曲杆菌42株,分离率为9.13%;分离的结肠弯曲杆菌对环丙沙星、萘啶酸、阿奇霉素耐药严重,耐药率分别为95.24%、90.48%、71.43%。[结论]河南省鸡源结肠弯曲杆菌耐药情况较为严重,应规范使用抗菌药物。     

Genetic diversity among Campylobacter jejuni isolates from healthy livestock and their links to human isolates in Spain

This study was aimed at determining the genetic diversity of Campylobacter jejuni from healthy ruminants and poultry, and study by multilocus sequence typing (MLST) their links to human isolates in Spain. MLST analysis of 160 animal isolates generated 45 sequence types (STs, nine of them new to this study), that clustered into 18 clonal complexes (CC) and nine singletons. The 71 isolates from humans generated 28 STs (13 CC plus four singletons). Only 11 STs and nine CCs were shared by humans and animals (particularly from dairy cattle and sheep), mainly corresponding to sporadic cases rather than outbreaks, probably as an adaptation of the general human population to the types commonly circulating in livestock. PCR analysis of the distribution of four virulence-associated genes detected the cdtABC gene cluster in all 160 isolates but with a 700-bp deletion in four of them, and amplified the virB11, cgtB and wlaN genes in 4.7%, 21.3% and 21.9% respectively. A subset of 87 C. jejuni animal isolates analysed using flaA PCR-RFLP, MLST and pulsed-field gel electrophoresis generated 31, 38 and 55 types respectively. The combined typing approach used provided reliable inter-strain relationships, confirming the co-existence of several strains in some farms, but also identifying identical genotypes sampled over a wide temporal span in different environments and hosts. Typing results confirmed a high genetic diversity of C. jejuni in our region and suggested that ruminants are also important sources for human infection. MLST data provided will help to obtain a more comprehensive image of the population structure of C. jejuni and establish reliable source attribution schemes.