猪链球菌的鉴定及其主要毒力基因的多重PCR检测

通过革兰氏染色镜检、生化试验及PCR鉴定,对分离的猪源链球菌进行初步研究.同时根据猪链球菌主要毒力因子基因Sly、MRP、EF的核苷酸序列,设计并合成了3对特异性引物,通过体系和条件优化,建立多重PCR检测方法,对实验菌株及阴性对照菌株进行检测分析.结果从不同地区分离到的30株猪源链球菌中,确认16株为猪链球菌.多重PCR检测结果显示,Sly检出率为5/16,MRP检出率为4/16,EF检出率为2/16,阴性对照菌株毒力因子检测结果均为阴性.分析结果表明,该多重PCR体系可用于猪链球菌毒力相关因子Sly、MRP、EF的基因检测,特异性和敏感性较好.     

猪链球菌9型国内分离株毒力基因的检测及小鼠免疫试验

通过检测7株猪链球菌9型（SS9）国内分离株的毒力基因,比对7株SS9荚膜合成基因cps9j的300bp碱基序列,以及进行7株SS9对小鼠的毒力试验和免疫保护试验,初步探讨了国内SS9的毒力特性。结果显示,7株SS9均缺失胞外因子（epf）和溶血素（sly）基因,溶茵酶释放蛋白基因（mrp）存在于GZ0665和GD0606株,off2仅存于GZ0665株;病猪脑脊液分离株GZ0665与从健康猪扁桃体分离的6株SS9的cps9j同源性低;在7株SS9分离株和SS2分离株HA9801中,只有GZ0665和GD0606株能致死小鼠;用灭活的GZ0665菌株免疫的小鼠对同源菌株GZ0665及GD0606株攻击的保护率均为100％（10／10）,用灭活的GD0606和未灭活的GD0627菌株免疫的小鼠对GD0606株攻击的保护率分别为80％（8／10）和50％（5／10）,而对GZ0665株攻击的保护率分别为60％（6／10）和30％（3／10）。证实,SS9病猪脑脊液分离株的毒力基因有别于SS9健康猪扁桃体分离株和SS2。表明,小鼠可以用来区分SS9病猪脑脊液分离株与健康猪扁桃体分离株的致病力。     

猪链球菌毒力因子的研究进展

猪链球菌病是一种重要的人畜共患病,给人类和养猪业的健康发展造成极大的危害。猪链球菌的毒力因子包括荚膜多糖(CPS)、溶菌酶释放蛋白(MRP)、胞外因子(EF)、溶血素(SLY)、毒力相关序列(ORF2)、纤连蛋白和血纤蛋白原结合蛋白(FBPS)、谷氨酸脱氢酶(GDH)、IgG结合蛋白等,它们与猪链球菌的致病性有着密切的关系。笔者通过总结毒力因子对猪链球菌致病机理的影响,以期为猪链球菌病疫苗的研制和治疗提供理论基础。     

Serotype- and virulence-associated gene profile of Streptococcus suis isolates from pig carcasses in Chiang Mai Province,Northern Thailand

In this present study, the serotype of 40 Streptococcus suis isolates from submaxillary glands of pig carcasses sold in wet markets in Chiang Mai Province, northern Thailand, was investigated. Eleven serotypes, including types 2, 3, 4, 5, 7, 8, 9, 17, 21, 22 and 31, were found in the isolates by a Multiplex PCR combined with serum agglutination. Of the eleven serotypes present, type 3 was the most prevalent, while types 2, 4, 5 and 21 were of primary interest due to their human isolate serotype. The mrp+/epf − /sly − genotype was found to be the most prevalent genotype. This study indicates the importance of effective control of human S. suis infection due to raw pork or pig carcass handling in northern Thailand.     

猪链球菌病研究进展

猪链球菌(Streptococcus suis)是一种重要的人兽共患病病原体,能够引起猪的疫病,人类感染该菌可导致脑膜炎、败血症,甚至死亡.该菌对养猪业造成严重经济影响,对公共卫生事业构成巨大威胁.由于猪链球菌血清型较多,抗原结构复杂,在临床上又可呈现不同的症状,如败血症、脑膜脑炎、心内膜炎、肺炎、化脓性淋巴结炎、关节炎等,还常常与其他疾病发生合并感染.近几年来,该病的发病率和死亡率有逐年上升趋势.链球菌的致病性已引起临床工作者和科研人员的广泛关注和重视,国内外许多学者对猪链球菌做了大量的研究工作,已发现的猪链球菌2型的致病因子包括荚膜多糖、溶菌酶释放蛋白、细胞外因子、蛋白质片段和IgG结合蛋白、溶血素等.     

Identification and characterization of Streptococcus suis

A total of 188 Streptococcus suis strains isolated from pigs with various lesions, mainly bronchopneumonia, were identified using classical biochemical methods and using the API 20 STREP system. The serological classification was performed by precipitation reactions after Lancefield extraction, slide-coagglutinations and capsular reaction tests. Fifty-nine strains belonged to Types 1, 1/2 or 2, 78 strains to Types 3-8 and 51 strains were non-typable. The results of the different tests and the biochemical differences between the different serotypes are discussed.     

猪链球菌2型的鉴定及其毒力因子检测

猪链球菌的感染不仅对养猪业造成巨大的损失,同时对公共卫生尤其是相关从业人员的生命造成威胁,严重感染引起死亡.根据参考文献,设计了1对猪链球菌2型特异性引物.对四川分离的7株链球菌、江苏分离的1株、北京分离的6株和菌种中心保存的5株C群马链球菌兽疫亚种、1株D群链球菌、4株R型链球菌、2株S群链球菌和E、F、G、K、L、M、N、O、P群链球菌各1株进行了检测.结果7株四川分离株均为2型;1株江苏分离株为2型;6株北京分离株有3株为2型;1株R群、2株s群链球菌为2型猪链球菌.对检测出的14株猪链球菌2型的毒力因子做进一步检测.MRP基因检出率为92.9%(13/14);EF基因检出率为100%(14/14);sly基因检出率为71.4%(10/14);Cps2A基因检出率为92.9%(13/14);gapdh基因检出率为64.3%(9/14);FBPS基因检出率为92.9%(13/14);orf2基因检出率为71.4%(10/14).毒力因子全为阳性的菌株有6株,MRP-EF+菌株有2株.本研究从259份健康猪扁桃体中分离到16株2型猪链球菌,健康猪带菌率为6.2%(16/259).16株猪链球菌2型中,MRP、EF、Cps2A全为阴性,只从1株2型猪链球菌中检测到了sly基因(1/16);gapdh基因检出率为56.25%(9/16);FBPS基因检出率为75.0%(12/16);Orf2基因检出率为37.5%(6/16).     

饲料霉菌复合污染的危害

霉菌污染越来越成为饲料工业和畜牧养殖业中的隐形杀手。论文主要论述了饲料霉菌复合污染给饲料工业和养殖业带来的危害及如何预防饲料霉变,减少霉菌复合污染给畜牧业带来的经济损失。     

猪链球菌对红霉素耐药性的研究

从发病猪体内分离、鉴定猪链球菌，采用肉汤稀释法和纸片琼脂扩散法筛选对红霉素耐药的猪链球菌，用双纸片法确定耐药株的耐药表型，通过聚合酶链反应检测对红霉素耐药的基因ermb/mefA。猪链球菌对红霉素的耐药表型为cMLS表型，即同时对克林霉素耐药。在3株红霉素耐药株中扩增到ermb基因，其余未能检测到ermb或mefA基因。     

猪致病性链球菌的分离鉴定

采集33例临床症状疑似猪链球菌感染猪的全血及内脏器官进行涂(触)片染色镜检,取有球菌感染的样品16份进行血琼脂平板分离纯化试验;根据血琼脂平板上菌落形态、溶血情况及对单个菌落涂片染色镜检结果,共筛选出12份可疑样品,进一步采用液体培养基对12份样品进行增菌及纯化培养;选取纯化培养菌进行链球菌生化试验,结果共鉴定出11株纯化的猪链球菌;11株猪链球菌均对小鼠表现出高致病力.     

Serological characterization and antimicrobial susceptibility of Streptococcus suis isolates from diagnostic samples in Denmark during 1995 and 1996

At the Danish Veterinary Laboratory Streptococcus suis infections in pigs were diagnosed in 114 cases in 1995 and in 151 cases in 1996. Isolates were serotyped using specific antisera against type 1 through 28 and a total of 67 cases from 1995 and 113 cases in 1996 were tested for resistance to 11 antimicrobial agents. The majority of cases were lung diseases (57%), followed by septicaemia (16%), meningitis (15%) and endocarditis (8%). Almost 96% of the isolates could be typed using the 28 antisera. The most common serotype was serotype 2 (29%), followed by serotype 7 (17%), and serotypes 3, 4 and 8 (9–10%). The remaining serotypes were observed in frequencies of less than 5%. Serotype 7 was more commonly isolated from septicaemia than the other serotypes. Serotype 2 was more commonly isolated from pigs older than 4 weeks compared to the other serotypes. Most isolates were susceptible to amoxycillin+clavulanate, ampicillin, ceftiofur, enrofloxacin, penicillin, spectinomycin, tiamulin and trimethoprim+sulphadiazine. A high frequency (>30%) of resistance to tetracycline was observed. Among isolates of serotype 2, 9.7% were resistant to lincomycin and 12.9% to spiramycin. Among other serotypes 56.8% were resistant to lincomycin and spiramycin. The differences in susceptibility between isolates of serotype 2 and the other serotypes were statistically significant. Compared to a previous Danish study the distribution of serotypes of S. suis causing infections among pigs in Denmark has changed during the last 15 years.     

猪链球菌鉴定方法的研究进展

猪链球菌是一种重要的人畜共患病病原,现已对我国部分省市的养殖业产生了巨大的影响。该菌血清型众多,生化特征变化较大,鉴定困难。本文就其中几种鉴定方法,如生化鉴定、血清学鉴定、分子水平鉴定等方面进行了综述。     

Mutagenesis of Streptococcus equi and Streptococcus suis by transposon Tn917

Genetic tools for studying streptococci are much less sophisticated than those that are available for many other bacterial genera. In this paper, we describe the development of a transposon mutagenesis system that we have used to mutate two important veterinary streptococci, Streptococcus equi and Streptococcus suis. The system uses a temperature-sensitive suicide vector to deliver Tn917 via electroporation, transposing Tn917 into the chromosomal DNA of the two streptococci. The transposon insertions can be rescued from the streptococcal chromosomes by plasmid rescue and selection in E. coli, with subsequent insertion site analysis by DNA sequencing. Transposition appeared to have occurred in an essentially random fashion when chromosomal DNA of S. suis and S. equi mutants was analysed by Southern blotting. However, when analysis of 60 S. equi mutants was carried out using the S. equi genome sequence database, 60% of transposon insertions had occurred within a 15 kb region of the genome whereas the other insertions appeared to have occurred essentially randomly. This finding suggests that Southern blot analysis for assessing the randomness of transposon libraries may need to be interpreted with caution. However, this observation notwithstanding, the Tn917 based system described in this paper will facilitate the study of S. suis and S. equi.     

猪链球菌病的诊断和防治

1　临床症状及剖检变化某猪场存栏猪 5 0 0 0余头 ,于 1 999年 1 0月发生该病 ,从保育猪舍开始 ,2～ 3天发病蔓延到整个猪场 ,先后发病 82 0头 ,经紧急治疗仍死亡 3 2 5头 ,病死率为 3 9.6 %。表现突然发病不食 ,体温 41～43℃ ,结膜潮红、流泪。有浆液性鼻液 ,呼吸困难 ,腹下、四肢、耳尖等处皮肤呈出血性紫斑 ,少部分育成病猪表现明显的神经症状 ,共济失调 ,倒地四肢做游泳状 ,磨牙或口吐白沫 ,大多于 1～ 3天内死亡 ,症状较轻及治愈者 ,偶留有跛行或生长不良。剖检变化差异较大 ,主要为全身淋巴结充血肿胀 ,脾脏肿大 1～ 3倍 ,边缘发生出…     

Streptococcus suis isolated from pigs in Finland

A total of 58 Streptococcus suis strains were isolated from deceased pigs submitted to the National Veterinary Institute, Regional Laboratory in Kuopio, Finland, over a 3 1/2 year period, most frequently from cases of pneumonia. The bacteria were isolated from cases of meningitis, sepsis, rhinitis, endocarditis and abortion. S. suis was also isolated from nasal cavity, lung and brain of some sick piglets without signs of inflammation. Further S. suis was detected in 12 out of 107 tonsils of healthy fatteners tested. S. suis strains were identified by biochemical methods followed by typing. The most common capsular types were 7, 3 and 2, respectively. Only one type 1 strain and no types 6 and 9 strains were found. All S. suis strains tested were sensitive to penicillin and ampicillin.S. suis is not uncommon in Finnish pig herds. S. suis may be regarded as a potentially pathogenic organism which under certain predisposing conditions may cause serious disease.     

湖南省猪链球菌的分离与鉴定

从病猪脏器病料中通过细菌培养、生化试验分离鉴定出了121株猪链球菌.经PCR技术鉴定马链球菌兽疫亚种28株,占23.1%,猪链球菌24株,占19.8% ,其中猪链球菌2型4株,占3.3%,猪链球菌9型2株,占1.7%,猪链球菌1型和7型都没检测到,证实省内除了猪链球菌2型外,也存在其他血清型的猪链球菌.     

Conjugation of antibiotic resistance in Streptococcus suis.

Forty-eight clinical isolates of Streptococcus suis were examined for antibiotic sensitivity and the presence of plasmid DNA. It was determined that isolates from this study showed a substantial increase in resistance to erythromycin (ery), clindamycin, and tetracycline (tet) compared to a similar study conducted five years earlier. Eleven of the 48 isolates contained plasmid DNA as revealed by DNA isolation and gel electrophoresis. Plasmid DNA from four strains resistant to the above three antibiotics was tested for the ability to transform an antibiotic sensitive recipient. No transformation of antibiotic resistance could be demonstrated. In other experiments, the above four strains, along with four plasmid-negative triply resistant strains were tested for the ability to transfer tet or ery resistance to tet and ery sensitive recipients by conjugation. In each mating, antibiotic resistance was transferred at frequencies averaging 2.4 x 10(-6) recombinants/recipient for ery and 3.4 x 10(-6) recombinants/recipient for tet resistance. DNA from each clinical specimen, as well as the recombinants mentioned above was probed with tn916. Autoradiographs revealed that several clinical isolates and recombinants bound the probe. It is concluded that conjugation of antibiotic resistance in these clinical strains is possibly mediated by a transposon similar to tn916.     

Isolation of Streptococcus suis from swine in Nebraska

Streptococcus suis was isolated from swine with localized and diffuse infections. The isolates (n = 121) were unable to grow in broth containing 6.5% NaCl. Their abilities to hydrolyze esculin were variable, as were their fermentative reactions in lactose, trehalose, raffinose, and inulin. Sorbitol was not attacked by any strain. Serologic reactions were obtained between the isolates and coagglutination reagents prepared from commercially available antisera to streptococcal groups D, R, or S.