腹泻仔猪肠道致病性大肠埃希氏菌和沙门氏菌的分离鉴定与耐药性研究

为探明仔猪细菌性腹泻肠道致病性大肠埃希氏菌和沙门氏菌流行的血清型、耐药表型及耐药基因型,本试验采集了贵州省5个地（州）市7个规模化养猪场的128份腹泻仔猪肠道样本,并对采集的样本进行了大肠埃希氏菌和沙门氏菌分离与鉴定,通过动物试验鉴定菌株的致病性,利用血清学方法鉴定其血清型,并通过药敏纸片法对主要致病菌进行耐药性研究,采用PCR技术检测各致病菌株耐药相关基因,分析细菌耐药表型和耐药基因型相关性。结果显示,本研究共分离鉴定到78株致病性大肠埃希氏菌与21株沙门氏菌,致病性大肠埃希氏菌血清型以O138、O87为主,沙门氏菌血清型以鼠伤寒沙门氏菌、肠炎沙门氏菌居多;药敏试验结果表明,本试验分离到的78株致病性大肠埃希氏菌对β-内酰胺类药物耐药率达80%以上,对其他种类的抗菌药耐药率均超过40%,分离鉴定的21株沙门氏菌对氨基糖苷类药物耐药率达50%以上,对其他种类的抗菌药耐药率均达20%以上;本试验分离鉴定的致病性大肠埃希氏菌共检出12种耐药相关基因,沙门氏菌共检出10种耐药相关基因,两种细菌耐药基因型与耐药表型符合率均达60%以上,且均为多重耐药。本研究为仔猪腹泻的综合防控提供了理论依据。     

腹泻兔病原菌的分离鉴定及耐药性检测

为了确定娄底地区兔腹泻的病原菌、致病性及耐药性,对采集的203份患腹泻的兔肛拭子、粪便进行病原菌分离鉴定,结果分离得到了30株大肠埃希氏菌、20株铜绿假单胞菌、17株产气荚膜梭菌、15株金黄色葡萄球菌;人工接种小鼠试验、KB药敏纸片法分别检测病原菌的致病性及耐药性,结果表明分离得到20株大肠埃希氏菌、15株铜绿假单胞菌、12株产气荚膜梭菌、10株金黄色葡萄球菌对小鼠具有致病性;分离的15株致病性铜绿假单胞菌对阿莫西林、氨曲南、庆大霉素等7种药物耐药率在46.7%以上,分离的12株致病性产气荚膜梭菌对环丙沙星、磺胺嘧啶、氟苯尼考等8种药物耐药率在41.7%以上,分离的10株致病性金黄色葡萄球菌对磺胺嘧啶、氟苯尼考、庆大霉素等7种药物耐药率在80.0%以上,分离的20株大肠埃希氏菌对磺胺嘧啶、多黏菌素、强力霉素等7种药物耐药率在45.0%以上,4种病原菌对其他药物具有不同的耐药性。     

致羔羊腹泻大肠杆菌血清型鉴定与耐药性分析

为了分析白城地区致羔羊腹泻大肠杆菌流行的血清型与耐药性,从采集的腹泻羔羊粪便样品127份中分离鉴定得到了56株为致病性大肠杆菌。采用玻板凝集法和K-B药敏纸片法分别检测56株致病性大肠杆菌分离株的血清型分布和耐药性情况。结果显示,56株致病性大肠杆菌分离株属于9个血清型,致病性大肠杆菌分离株以O119、O101、O24为流行的优势血清型;56株致病性大肠杆菌分离株且呈现多重耐药性,对头孢克肟、头孢曲松两种药物的耐药率最低,在12.4%~28.6%之间;对阿米卡星、新霉素、多西环素等7种药物的耐药率在50.0%~78.6%之间;对大观霉素、环丙沙星、氧氟沙星等5种药物的耐药率在82.1%~96.4%之间。     

温州地区鸡肠道大肠埃希菌耐药性分析及血清型鉴定

为研究温州地区鸡肠道大肠埃希菌的耐药性和血清型,从27家养鸡场采集泄殖腔拭子355份,分离并鉴定出346株鸡大肠埃希菌,分离率为97.46%。在346株分离株中,对3种以上抗菌药物耐药的菌株占84.39%(292/346),呈现多重耐药性。O血清型鉴定结果显示,分离菌株分别属于13个血清型,其中O138、O146、O136、O45占分离菌株的39.02%(135/346),为优势血清型。     

禽源大肠埃希氏菌分离株的病原特性鉴定

1998～2002年从上海地区72个养禽场分离出84株禽源大肠埃希氏菌,结合培养特性、生化特性及血清型等对其进行了鉴定。其形态、染色及生化特性等基本符合大肠埃希氏菌的特点。用138种大肠埃希氏菌"O"抗原定型血清检测84株分离株,其中O7811株,O210株,O978株,O356株,O1256株,O225株,O925株,O1235株,O964株,O263株,O23、O120各2株,O118、O128、O138、O83各1株。其余13株不在其检测范围。药敏试验结果表明,分离菌株已对多种抗生素产生了很强的耐药性。对其中29株菌株进行了小鼠致病性试验,致病率高达72 8%。     

川渝部分地区仔猪源大肠埃希菌的血清型鉴定及耐药性监测

本研究旨在调查四川和重庆地区仔猪源大肠埃希菌的耐药性及血清型流行状况。本试验采用玻片凝集法和纸片扩散法对采自两地不同区域的43个猪场共273株大肠埃希菌进行血清型鉴定和耐药性测定,结果表明,273株大肠埃希菌分属43个血清型,优势血清型由高及低依次为O138、O20、O8、O60、O141、O7、O38,合占检出菌株的46.83%(96/205);对14种药物存在不同程度的耐药性:对氨苄西林、磺胺嘧啶、四环素呈高度耐药,耐药率均在90%以上,较为敏感的丁胺卡那霉素、头孢他啶敏感率分别为53.11%、49.45%,大部分菌株呈多重耐药,共151种耐药谱型。由此可见,川渝地区部分规模化猪场大肠埃希菌的流行血清型多、耐药程度严重。故掌握优势血清型,筛选有效抗菌药物,采取对应措施防治大肠杆菌病已势在必行。     

仔猪腹泻大肠埃希菌的分离鉴定及药敏试验

为了了解甘肃省武威市凉州区部分地区引起仔猪黄白痢的致病性大肠埃希菌的主要血清型及其致病性,从凉州区4家规模化猪场采集了60份仔猪肛门拭子,用细菌分离培养方法分离出20株大肠埃希菌菌株,然后经形态观察、生化试验,确定了该次分离菌株为致病性大肠埃希菌。经血清型鉴定,表明20株分离菌株分布在8个血清型中,其中优势血清型为O8（9株）,占总数的45%。药物敏感性试验结果表明,分离菌株对临床常用药物具有抗药性。该研究能够对凉州区仔猪腹泻病的流行病学研究、诊断和免疫防制提供科学参考。     

奶牛粪样中致病性大肠埃希菌的分离与鉴定

大肠埃希菌血清型众多,一些具有特殊血清型的大肠埃希菌对人和动物有致病性。本试验从包头市侯家营奶牛场77头外表健康成年泌乳荷斯坦奶牛采集新鲜粪样,进行细菌分离培养,共分离到疑似大肠埃希菌69株,经过形态观察、生化鉴定、动物致病性试验和血清型鉴定,其中12株为致病性大肠埃希菌,均为强毒株,占分离菌数的17.4%,另57株为非致病性大肠埃希菌,占分离菌数的82.6%。同时对12株致病性大肠埃希菌进行了"O"血清型鉴定,分属7种血清型O78、O6、O9、O98、O1、O8、O149,分别占定型菌株的25.0%(3/12)、8.3%(1/12)、16.7%(2/12)、8.3%(1/12)、8.3%(1/12)、8.3%(1/12)、25.0%(3/12)。     

携带耶尔森强毒力岛的大肠埃希菌耐药性及其对雏鸭的致病性试验

为了解105株动物源携带耶尔森强毒力岛的大肠埃希菌耐药状况和致病性,分别对各分离菌进行28种抗菌药物的敏感性测定,并选择不同毒力基因型的10株大肠埃希菌进行雏鸭的致病性试验.结果表明,分离菌对四环素、多西环素、氨苄西林、阿莫西林、复方磺胺甲(噁)唑、链霉素耐药性较高,耐药率均在80%以上;敏感率在80%以上的药物有头孢...     

宁夏部分地区腹泻羔羊源大肠埃希菌的分离鉴定与药敏试验

为掌握宁夏地区羔羊腹泻源大肠埃希菌的耐药性,采集腹泻羔羊肛拭子90份,开展大肠埃希菌的分离鉴定及药敏试验。结果分离到78株大肠埃希菌,对头孢类、氨基糖苷类、喹诺酮类、氟苯尼考、青霉素类、四环素等抗菌药物有较高的敏感性,对万古霉素、复方新诺明、克林霉素耐药(耐药率78.21%~94.87%)。结果表明,宁夏地区羔羊腹泻源大肠埃希菌对抗菌药物保持较高的敏感率,耐药现象并不严重。     

仔猪腹泻致病性大肠杆菌分型鉴定及耐药性分析

为了解贵州省规模化养猪场腹泻仔猪致病性大肠杆菌流行情况及耐药性变化,本研究运用凝集试验、PCR和药敏纸片琼脂扩散法等方法对分离的78株致病性大肠杆菌进行血清型、毒力基因及耐药性分析。结果显示,78株致病性大肠杆菌以O138、O87血清型为主,占定型菌株的60.8%;其中62株致病性大肠杆菌检出毒力基因,检出率为79.5%,可分为8种毒力基因类型,分属肠致病性大肠杆菌（EPEC）、肠产毒性大肠杆菌（ETEC）和肠聚集性大肠杆菌（EAEC）,毒力基因eaeA、elt和escV检出率较高,分别为38.5%、28.2%和21.8%;分离到的致病性大肠杆菌对β-内酰胺类药物高度耐药,均为多重耐药株,耐药种类可达8种以上。结果表明,当前贵州省规模化养猪场腹泻仔猪致病性大肠杆菌的毒力基因检出率较高且基因型复杂,耐药性严重。本试验结果可为规模化养猪场防控仔猪腹泻提供基础资料及理论依据。     

规模化养鸭场鸭源大肠杆菌分离株的致病性及系统发育分析

为研究鸭源大肠杆菌(E.coli)的致病性,明确鸭病原性E.coli与人和其他动物E.coli的亲缘关系及O血清型与菌种间遗传关系的相关性,本研究从我国西南地区规模化养鸭场患典型E.coli败血症的雏鸭体内分离37个血清型共82个E.coli分离株(其中优势血清型32株,其他血清型50株),以每株0.2 mL (109 cfu/mL)腿部肌肉接种7日龄健康鸭进行致病性试验;并对其中14个鸭源E.coli代表株进行16S rRNA基因克隆、测序及系统发育分析.结果表明:所有分离株全部为致病性E.coli,其中高致病株、中等致病株和低致病株分别占受试菌株的80.5％(66/82)、17.1％ (14/82)和2.4％(2/82),4个优势血清型全部为高致病株和中等致病株,分别占受试菌株的84.4％(27/32)和15.6％(5/32);14个代表株鸭源E.coli的16S rRNA形成2个主要分支,3株新发现的血清型单独形成一个较远的分支,11株常见血清型与人及其他动物源E.coli形成另一个大的分支,其中有6株与人的O157亲缘关系较近,2株与出血性E.coli O157:H7 sakai聚为一个小分支;从遗传进化的关系分析,这些菌株有可能是人类的潜在病原菌;同时还发现不同血清型的分离株可以聚为一支,而同一血清型的分离株可以处于不同的分支,常规的O血清学分型不能体现菌种间遗传关系的远近.     

Isolation,Identification and Drug Resistance Analysis of Interstinal Pathogenic Escherichia coli and Salmonella Isolated from Diarrhea Piglets

In order to find out the serotype, resistant phenotype and genotype of Escherichia coli (E. coli) and Salmonella in piglets, this study collected 128 samples of diarrhea piglets from seven large-scale pig farms in five cities in Guizhou province, and the E. coli and Salmonella were isolated and identified. The pathogenicity of the strain was identified by animal test. The drug resistance of the main pathogen was tested by drug susceptibility paper. The resistance gene of each pathogen was detected by PCR. The drug resistance and genotype correlation of the bacterial were analyzed. The results showed that 78 strains of pathogenic E. coli and 21 strains of Salmonella were isolated and identified in this study. The serotypes of pathogenic E. coli were predominantly O138 and O87. Salmonella Typhimurium and Salmonella Enteritidis were predominant serotypes. The susceptibility test showed that the resistant strains of 78 strains of E. coli were more than 80% resistant to β-lactams and more than 40% for other antibacterials. The resistance rate of 21 strains of Salmonella to aminoglycosides was more than 50% and more than 20% to other types of antibacterials; 12 and 10 kinds of drug resistance-related genes of E. coli and Salmonella were detected, respectively; The coincidence rate of resistant genotype and phenotype of two kinds of bacteria were above 60%, and both were multiple drug resistance. This study provided a theoretical basis for comprehensive prevention and control of piglets diarrhea.     

Rapid identification of virulence genes in enterotoxigenic Escherichia coli isolates associated with diarrhoea in Queensland piggeries

OBJECTIVE: To identify virulence genes in enterotoxigenic E coli (ETEC) isolates associated with diarrhoea in neonatal, 1 to 3 week-old and weaned pigs in southeast Queensland. DESIGN: Multiplex PCR and serotyping were applied to E coli isolates obtained over a 5-year period (1998-2002) from cases diagnosed at Toowoomba Veterinary Laboratory. PROCEDURE: A total of 126 isolates from 25 different Queensland piggeries were tested for haemolytic activity on 5% sheep blood agar and by multiplex PCR for the presence of five commonly recognised fimbrial (F4, F5, F6, F41 and F18) and three enterotoxin genes (STa, STb, LT). A subset of 62 representative isolates were serotyped by slide agglutination. For comparative purposes, multiplex PCR was also performed on the DNA of 31 ETEC isolates from 9 serotypes originating from piggeries in southern New South Wales. RESULTS: A total of 113 (89.7%) of the isolates from Queensland possessed ETEC virulence genes, including 14 of 15 isolates from neonatal pigs (93.3%), 18 of 23 isolates from 1 to 3 week old pigs (78.3%) and 81 of 88 isolates from weaned pigs (92.1%). F4:STa:STb:LT (serotype O149) was the most prevalent pathotype in neonatal and 1-3 week old pigs and F4:STa:STb:LT (serotype O149) and F18:STa:STb:LT (serotype O141) were most prevalent in weaned pigs. In comparison, isolates obtained from neonatal pigs from New South Wales belonged to a more diverse range of pathotypes and serotypes. CONCLUSION: Multiplex PCR was a rapid and specific method for detecting the presence of ETEC virulence genes in porcine E coli isolates. For isolates obtained from cases of suspected colibacillosis in Queensland, growth of a heavy pure culture of haemolytic E coli was a sensitive prognostic indicator of the presence of ETEC virulence genes in the isolate. ETEC pathotypes and serotypes remained stable in Queensland piggeries over the five-year study period and appear to have changed little over the last three decades.     

共和县地区猪场仔猪致病性大肠杆菌血清型鉴定

采集青海省共和县不同地区4个猪场仔猪粪样316份,并采用玻板凝集法进行了O抗原血清型鉴定。结果发现:316份仔猪粪样经分离鉴定出致病性大肠杆菌176株,155株鉴定出抗O血清型。该地区流行优势血清型为O11(13株)、O60(12株)、O119(10株)、O137(9株)、O141(10株)、O144(12株)、O147(12株)和O149(8株),分别占8.38%、7.74%、6.45%、5.81%、6.45%、7.74%、7.74%和5.16%。8种优势血清型占总定型菌株的53.5%(83/155)。     

临床型乳房炎奶牛致病性大肠杆菌血清型研究

为有效防治乳房炎以及进一步研究致病性大肠杆菌的危害性,本研究对呼和浩特市某奶牛场16头患临床型乳房炎的奶牛进行采样分析了致病性大肠杆菌。结果表明,在16份粪样、乳样中致病性大肠杆菌的检出率分别为100%和68.75%,血样中未检出致病性大肠杆菌。经动物试验表明,所检出的致病菌均有强致病力。在血清型鉴定中,粪样中分离鉴定出致病性大肠杆菌为O51、O55、O127、O7、O91、O101、O112、O92、O142血清型较多;乳样中分离出的致病性大肠杆菌为O55、O101、O112、O134、O51、O53、O78、O91、O126、O128、O158血清型较多。且粪样中分离出的致病性大肠杆菌42种血清型,乳样中分离出的致病性大肠杆菌33种血清型中具有25种血清型相同,分别为:O8、O35、O44、O51、O55、O61、O62、O65、O69、O80、O91、O92、O95、O96、O98、O99、O100、O101、O112、O120、O127、O134、O142、O153、O158。因此可见,作为环境性病原菌,大肠杆菌侵入乳房而引发了奶牛乳房炎。     

猪胸膜肺炎放线杆菌四川株的分离鉴定及药物敏感性试验

从四川某猪场疑似胸膜肺炎放线杆菌感染的病料中分离到一株革兰氏阴性小球杆菌。经染色镜检、生化试验、PCR检测、血清型检测,致病性试验和药敏试验,鉴定该菌为猪胸膜肺炎放线杆菌1型,且具有较强致病性。药敏试验结果表明,该菌对四环素、甲氧苄啶,以及链霉素、庆大霉素、卡那霉素、丁胺卡那、新霉素等氨基糖苷类药物高度耐药,对氯霉素、氟苯尼考、氨苄西林、阿莫西林、羧苄西林和恩诺沙星、诺氟沙星等喹诺酮类受试药物敏感。     

In vitro and in vivo characterization of avian Escherichia coli. II. Factors associated with pathogenicity

The pathogenicity of 197 Escherichia coli isolates obtained from clinically affected commercially grown broiler chickens and normal hatchery chicks was assessed by inoculating day-old broilers intratracheally. The degree of pathogenicity (high, intermediate, low) was judged according to mortality and lesions occurring within 7 days following inoculation. Serotype, metabolic activity, motility, and in vitro antibiotic sensitivity of each isolate were evaluated and related to pathogenicity. Seventy-five of the isolates of high to intermediate pathogenicity belonged to serogroup O2, O78, or O35. In addition, 51 pathogenic E. coli isolates could not be serotyped, and several had multiple serotypes. Most isolates had similar metabolic activity, as determined by amino acid decarboxylation and carbohydrate fermentation, regardless of pathogenicity. An exception was the fermentation of adonitol, which occurred more frequently with the highly pathogenic strains. Motility and in vitro antibiotic sensitivity were not related to pathogenicity. An age-associated resistance to intratracheal E. coli administration occurred by 15 days of age in uncompromised birds. Relative susceptibility of birds older than 2 weeks to intratracheal and/or intravenous E. coli inoculation could be increased by prior exposure to pathogenic reovirus 1733, adenovirus 3167, or infectious bursal disease virus (IBDV). Birds infected with IBDV at 3 weeks failed to clear apathogenic and pathogenic E. coli from circulating blood.