Production of virulence-related proteins by Canadian strains of Streptococcus suis capsular type 2.

The production of muramidase-released protein (MRP), extracellular protein factor (EF) and hemolysin (suilysin) by 101 Canadian field strains of Streptococcus suis capsular type 2 is described. Most strains (72%) isolated from diseased pigs were MRP-EF- and only 1 strain was MRP+EF+. This strain was also the only 1 to produce the hemolysin. Thirteen strains (15%) were MRP+ EF- and only 3 strains were MRP* EF-. All the strains isolated from clinically healthy pigs as well as a bovine and 2 human isolates had a MRP-EF- phenotype. In addition, 7 strains (8%) had a MRPS phenotype, which had so far been described for S. suis capsular type 1. In conclusion, most Canadian field isolates of S. suis capsular type 2 tested in this study do not produce the virulence-related proteins described so far for this bacterial pathogen.     

Experimental airborne transmission of Streptococcus suis capsular type 2 in pigs.

Experimental airborne transmission of Streptococcus suis type 2 was studied in specific pathogen free piglets. Forty piglets were allotted to five groups of eight 7-week-old animals and housed in three separated units. Negative control pigs (group 1) were housed in unit A and infected batches were housed in units B (group 2) and C (groups 4). In units B and C, non-inoculated groups (groups 3 and 5, respectively), 40 cm distant from the respective inoculated group and without any physical contact between them, also took place. Six animals of groups 2 and 4 were inoculated intravenously with 2 x 10(8) colony forming units (cfu) of a mild and a high virulent S. suis strains, respectively. The remaining animals in these groups and pigs from groups 1, 3, 5 received broth medium in the same way. Differences among virulence of S. suis capsular type 2 were observed in inoculated pigs of groups 2 and 4. Pigs from group 2 became carriers, showing only mild symptoms. By contrast, animals from group 4 presented an acute form of the disease. All the indirect contact pigs in groups 3 and 5 had S. suis in palatine tonsils from day 6 after the infection and they presented clinical manifestations similar to those observed in experimentally infected pigs. Two direct contact animals were also contaminated in the upper respiratory tract but surprisingly they did not show any symptoms. Airborne transmission of S. suis in experimentally pigs was demonstrated in the present study. Indirect infections, as described in this study, are a more realistic way to infect pigs than other experimental procedures and may be used to further study the pathogenesis of the infection caused by this important pathogen.     

Adherence of Streptococcus suis capsular type 2 to porcine lung sections.

The present study was undertaken to evaluate the ability of 33 Streptococcus suis capsular type 2 isolates to adhere to frozen sections of porcine lung. Twenty isolates originated from diseased pigs and 13 from the nasal cavities of clinically healthy pigs. All isolates from diseased animals adhered to lung sections; isolates from pneumonia adhered, in general, in greater numbers than isolates from meningitis. Only four isolates from clinically healthy animals showed a weak adherence to lung sections. Hydrophobic surface properties were also evaluated. All isolates tested appeared to possess a hydrophilic cell surface. The thickness of the capsular material correlated well with the degree of adherence. However, when the adherence capacity of a noncapsulated mutant was compared with that of the parent strain, it was found that the mutant strain had at least the same adherence capacity as the capsulated parent strain. The data suggest that S. suis capsular type 2 isolates involved in pathological conditions can adhere to porcine lung tissue. The adherence activity does not seem to involve hydrophobic interactions. The amount of capsular material seems to influence the adherence activity, but is probably not the only mechanism involved.     

Production of capsular material by Streptococcus suis serotype 2 under different growth conditions.

The procedure currently used for the production of Streptococcus suis antigen is very long and includes several subcultures. The aim of the present work was to study the in vitro production of capsular material by S. suis serotype 2 after each of these subcultures. The amount of capsular material produced was evaluated by electron microscopy using bacterial cells grown on blood-agar plates and in Todd-Hewitt broth (THB) or THB supplemented with serum. In addition, the production of antibodies in rabbits with antigens produced using different growth conditions was compared. Antigens produced after only three subcultures possessed as much capsular material as cells obtained after the complete procedure and induced a similar antibody response. The use of serum as a supplement to the broth did not assure a higher production of capsule; in addition, antibody titers obtained with antigens produced in THB were as high as those obtained with antigens produced in THB supplemented with serum. We recommend the use of three subcultures in nonsupplemented broth for the production of immunogens. This revised protocol offers two main advantages: it is less time-consuming because of the limited number of subcultures and is also less expensive since nonsupplemented broths are used.     

Isolation and characterization of Streptococcus suis capsular types 9-22

The incidence and biochemical patterns of Streptococcus suis capsular types 9-22 are presented. Of 148 untypeable (with types 1-8 antisera) isolates of S. suis recovered from diseased pigs, 10% were not capsulated. Of the remaining 134 isolates, only 53% belonged to capsular types 9-22; capsular types 22 and 9 were the most prevalent, representing 19% and 13%, respectively. Capsular type 15 (de Moor's group T Streptococcus) is reported here for the first time in North America since it was described in 1963 in Europe. Of 188 untypeable isolates recovered from clinically healthy pigs, 25% were noncapsulated. Of the remaining 141 isolates, 90% belonged to the new capsular types, and 87% were identified as 1 of 4 types: 17, 18, 19, and 21. Capsular types 12 and 20 were not detected among the Canadian isolates. Almost half of strains were arginine dihydrolase-negative, and 45% fermented mannitol, which is seldom a positive test with capsular types 1-8. Although some strains were negative with salicin or trehalose, none were negative for both sugars. Only 54% of isolates tested with 1 rapid multitest system were correctly identified as S. suis. A tentative biochemical profile that might be used with a microplate identification system is also presented. Biochemical identification using the conventional system instead of the rapid multitest system is preferable.     

Experimental infection of specific pathogen free piglets with French strains of Streptococcus suis capsular type 2.

A standardized model of Streptococcus suis type 2 infection in specific-pathogen-free piglets, housed in high-security barns, was used to compare the virulence of 3 French field strains of S. suis serotype 2 isolated from tonsils of a healthy pig (strain 65) or from diseased pigs (meningitis, strain 166', or septicemia, strain 24). In one of the 2 trials, 7-week-old pigs, in 3 groups of 8, were inoculated intravenously with 2 x 10(8) colony-forming units of S. suis type 2. In each group, 1 uninfected animal was a sentinel. Eight animals were also used as negative control group. The experiment was repeated under similar conditions with strains 65 and 166'. Virulence differed markedly among these S. suis strains when clinical signs, zootechnical performances, lesions, and bacteriological data were analyzed. Strain 65 did not induce clinical signs in inoculated pigs. In contrast, pigs infected with the other 2 strains exhibited clinical signs and typical lesions of S. suis type 2 infections. Differences in virulence were also observed between the 2 virulent strains. Sentinel animals exhibited the same manifestations as those recorded in inoculated piglets. Results were similar in the second trial, indicating that under the present experimental conditions, results were reproducible. The standardized conditions described in this study could be a useful tool to further study about the S. suis infection.     

Antibody response to an autogenous vaccine and serologic profile for Streptococcus suis capsular type 1/2

An autogenous vaccine was developed, using sonicated bacteria, with a strain of Streptococcus suis capsular type 1/2. The objectives of this study were to evaluate the antibody response following vaccination and to assess the changes in antibody levels in pigs from a herd showing clinical signs of S. suis capsular type 1/2 infection in 6- to 8-week-old pigs. An enzyme-linked immunosorbent assay using the vaccine antigen was standardized. Results from a preliminary study involving 2 control and 4 vaccinated 4-week-old pigs indicated that all vaccinated pigs produced antibodies against 2 proteins of 34 and 43 kDa, respectively, and, in 3 out of 4 vaccinated pigs, against the 117-kDa muramidase-released protein. For the serologic profile, groups of 30 pigs from the infected herd were blood sampled at 2, 4, 6, 8, and 10 weeks of age. The lowest antibody level was observed between weeks 6 and 8, presumably corresponding to a decrease in maternal immunity. A marked increase was seen at 10 weeks of age, shortly after the onset of clinical signs in the herd. For the vaccination field trial, newly weaned, one-week-old piglets were divided into 2 groups of 200 piglets each (control and vaccinated); blood samples were collected from 36 piglets in each group at 2-week intervals for 12 weeks. A significant increase (P 0.05) in antibody response was observed 4 weeks following vaccination and the level of antibodies stayed high until the end of the experiment. In the control group, the increase was only observed at 13 weeks of age, probably in response to a natural infection. The response to the vaccine varied considerably among pigs and was attributed, in part, to the levels of maternal antibodies at the time of vaccination. No outbreak of S. suis was observed in the control or vaccinated groups, so the protection conferred by the vaccine could not be evaluated.     

Prevalence, capsular type and antimicrobial susceptibility of Streptococcus suis isolated from slaughter pigs in Korea.

This study was undertaken to determine the prevalence, capsular serotype, and antimicrobial susceptibility of Streptococcus suis isolated from slaughter pigs. Capsular serotype and antimicrobial susceptibility were determined by coagglutination test and agar dilution minimum inhibitory concentration, respectively. Streptococcus suis was isolated from 55 of the 406 palatine tonsillar samples tested (13.8%) and 14 of the 29 sampled herds (48.3%). Of the 55 isolates recovered from slaughter pigs, 26 (47.3%) were untypeable. Of the remaining 29 isolates, capsular serotypes 9 (9 isolates) and 16 (4 isolates) were the most common, followed by capsular serotypes 4 (3 isolates) and 7 (3 isolates). Every capsulated isolate was typeable and no palatine tonsillar sample yielded more than one serotype. Most of isolates were susceptible to low concentrations (MIC90) of amoxicillin (2 microg/mL), ceftiofur (1 microg/mL), and penicillin (1 microg/mL). No correlation was found between antimicrobial susceptibility and capsular serotype.     

猪链球菌通用及2型TaqMan荧光PCR检测技术建立与应用

在对设计合成的引物和6-carboxy-fluorescein(6-FAM)荧光素标记的TaqMan水解探针进行筛选的基础上,通过反应体系优化,建立了检测猪链球菌通用和2型的2种荧光PCR检测技术,实现了对猪链球菌的快速检测和定型。敏感性、特异性、重复性、感染组织模拟试验及临床样品的检测结果均表明建立的方法快速、特异、灵敏,整个检测过程(包括样品的前处理时间)可在1.5 h内完成,检测培养物和模拟组织样品的灵敏度可达10 CFU～100 CFU。     

猪链球菌9型荚膜多糖cps9G基因的原核表达

本研究旨在克隆表达猪链球菌9型荚膜多糖部分抗原表位。设计1对引物,采用PCR法以猪链球菌9型河南分离株PY-2的基因组DNA为模板扩增出cps9G全基因序列。用限制性核酸内切酶BamHⅠ、XhoⅠ进行双酶切后,将其定向克隆到pET32a（＋）中,构建重组表达质粒pET32a—cps9G,并将其转化至大肠杆菌BL21（DE3）感受态细胞中进行诱导表达,并优化表达条件。结果表明,经1．2mmol／L IPTG诱导4h后SDS-PAGE分析表明,重组菌株表达出了约50700的融合蛋白条带,与预期一致。Western-blotting分析表明,该融合蛋白具有特异的生物学活性。这为以后建立快速、简便、特异的免疫学检测方法及制备单克隆抗体提供了较好的抗原,同时为研制猪链球菌亚单位疫苗和诊断试剂盒奠定了基础。     

猪链球菌2型毒力相关蛋白

猪链球菌 (Streptococcus suis)是重要的猪病原菌 ,并能感染人 ,可致人和猪死亡。猪链球菌病多发于 3～ 12周龄断奶后的仔猪 ,几乎世界各国都有发生 ,对养猪业危害严重 ,同时事关公共卫生。目前已鉴定了 35种血清型的猪链球菌 ,其中最重要、最常见的是 2型 [1 ] 。我国于 1991年在广东省首次证实此菌的存在 ,1998年在江苏的人群和猪群中暴发由 2型所致的链球菌病 [2 ] ,其毒力因子的研究 ,尤其值得关注。1　毒力相关蛋白的发现　　基于传统观念 ,最初有人提出荚膜多糖决定猪链球菌 2型的侵袭力 [3 ,4] ,但后来的研究不能证实荚膜多糖是该菌…