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Construction and Characterization of sao Gene Knock-out Mutant of Streptococcus suis Serotype 2 Chinese Highly Virulent Strain 05ZYH33
2型猪链球菌中国强毒株05ZYH33 sao基因敲除突变株的构建及其生物学功能

LIU Wen-Jing,PAN Xiu-Zhen,LI Xian-Fu,WANG Chang-Jun,TANG Jia-Qi,
刘文静
,潘秀珍,李先富,王长军,唐家琪

微生物学通报 , 2010,
Abstract: 构建2型猪链球菌(Streptococcus suis serotype 2, S. suis 2)中国强毒株05ZYH33的sao基因敲除突变株。构建中间为壮观霉素抗性基因, 两侧为sao编码基因上下游同源序列的基因敲除载体, 同源重组筛选sao基因敲除突变株。PCR、RT-PCR、Western Blot对疑似突变株进行验证, 实验结果均证实sao基因完全被spc抗性基因替代, 成功构建了突变株05ZYH33Δsao。对野生株和突变株进行菌落溶血活性、生长特性、小鼠致病性比较, 结果表明sao基因的敲除并未使野生型菌株在以上三方面产生明显的变化。筛选获得的05ZYH33 sao基因突变株为进一步研究sao基因在05ZYH33致病过程中的作用奠定了基础。
2型猪链球菌溶血素多克隆和单克隆抗体制备及鉴定
Preparation and identification of polyclonal and monoclonal antibody of Suilysin in highly virulent strains of Streptococcus suis serotype 2
 [PDF]

孙雯,郑峰
- , 2017,
Abstract: 目的 制备并鉴定2型猪链球菌(Streptococcus suis serotype 2,S.suis 2)四川分离菌株 05ZYH33 的溶血素(Suilysin,SLY)多克隆和单克隆抗体。方法 利用重组 SLY 蛋白制备 SLY 兔多抗,优化间接 ELISA 体系检测多抗效价,Western blot 分析 SLY 多抗特异性,检测 SLY 兔多抗对人外周血杀菌作用的影响。利用杂交瘤细胞融合技术制备 SLY 单抗,筛选杂交瘤细胞株并制备小鼠腹水,间接 ELISA 和 Western blot 鉴定 SLY 单抗的效价和特异性。结果 间接 ELISA 显示 SLY 多抗效价高达1∶204 800,Western blot 显示 SLY 多抗有较高的特异性和免疫反应性,人外周血杀菌试验显示,SLY 多抗能加强人外周血对 05ZYH33 的杀伤作用。间接 ELISA 检测杂交瘤细胞 2B6 和 5F7 培养上清效价分别是1∶800和1∶3 200,单抗效价都为1∶204 800。间接 ELISA 和 Western blot 鉴定显示,单抗 2B6 和 5F7 具有很强的特异性和免疫反应性。结论 成功制备了效价高、特异性强的 S.suis 2 05ZYH33 SLY 多克隆和单克隆抗体。
Comparative Genomic Hybridization Identifies Virulence Differences in Streptococcus suis  [PDF]
Han Zheng, Ruiting Lan, Xiao Zheng, Zhigang Cui, Zhijie Liu, Xuemei Bai, Shaobo Ji, Marcelo Gottschalk, Jianguo Xu
PLOS ONE , 2014, DOI: 10.1371/journal.pone.0087866
Abstract: Streptococcus suis is an important zoonotic pathogen. However, identification of virulent S. suis strains is complicated because of the high diversity of the species. Here we evaluated the genetic difference among S. suis strains using comparative genomic hybridization (CGH) and virulence variation in vivo and in vitro. We showed that different clades differed in their ability to activate TLR2/6 in vitro and their capacity to induce cytokine production in vivo as well as their resistance to phagocytosis and survival in vivo. Our data showed the S. suis strains tested can be classified into three groups having differing levels of virulence: epidemic and highly virulent strains were clustered into clade Ia (epidemic and highly virulent group, E/HV group), virulent strains were clustered into clade Ib (virulent group, V group), and intermediately or weakly virulent strains were clustered into other clades (intermediately or weakly virulent group, I/WV group). Our study provided further insight into the genomic and virulence variation of S. suis.
Cloning,Expression and Animal Experiment of the Immunopotential Fragment of mrp Gene from Streptococcus suis type 2
猪链球菌2型mrp基因免疫功能片段的克隆、表达及动物试验

FAN Hong Jie LU Cheng Ping,TANG Jia Qi,
范红结
,陆承平,唐家琪

微生物学报 , 2004,
Abstract: 根据猪链球菌2型(Streptococcus suis type 2)国外分离株的溶菌酶释放蛋白(Muramidasereleased protein, MRP)的基因序列,设计并合成一对引物,利用PCR技术扩增了江苏分离株的开放阅读框298~827bp间529bp的基因片段,并定向克隆至pET32a(+)表达载体中。重组质粒经限制性酶切鉴定和测序,转化至大肠杆菌BL21,经IPTG诱导,可表达分子量约42kD的蛋白。经过镍亲和层析柱层析,获得纯化的重组蛋白。以重组蛋白免疫Balb/c小鼠,以5LD50猪链球菌强毒株攻击后小鼠的相对存活率达625%。证实所表达的MRP片段为重要的保护性抗原。
Characterization of porcine dendritic cell response to Streptococcus suis
Marie-Pier Lecours, Mariela Segura, Claude Lachance, Tufaria Mussa, Charles Surprenant, Maria Montoya, Marcelo Gottschalk
Veterinary Research , 2011, DOI: 10.1186/1297-9716-42-72
Abstract: Streptococcus suis is a major swine pathogen associated mainly with meningitis, although other pathologies have also been described such as septicemia with sudden death, endocarditis, arthritis, and pneumonia [1]. Among 35 serotypes described, serotype 2 is considered the most virulent and the most frequently isolated from both diseased pigs and humans. Consequently, most studies on virulence factors and the pathogenesis of infection have been carried out with this serotype [2]. Until recently, S. suis disease in humans has been considered as rare and only affecting people working with pigs or pork by-products. However, with a rising incidence in humans over the last years, S. suis is now considered as an important emerging zoonotic agent, especially in Asian countries, where S. suis has recently been identified as the leading cause of adult meningitis in Vietnam, the second in Thailand, and the third in Hong Kong. In 2005, an important outbreak occurred in China and resulted in 200 human cases with a fatality rate near 20% [1]. In humans, S. suis is mainly responsible for meningitis, septicemia and streptococcal toxic shock-like syndrome [1,3,4].Despite the increasing number of studies, the pathogenesis of the S. suis infection is still not completely understood and, to date, attempts to control the infection are hampered by the lack of an effective vaccine. The mechanisms involved in the host innate and adaptive immune responses toward S. suis as well as those used by S. suis to subvert these responses are unknown. Several virulence factors have been proposed to be involved in the pathogenesis of S. suis infection [5]. Among them, the capsular polysaccharide, which confers to the bacteria antiphagocytic properties, has been demonstrated as a critical virulence factor [2,6,7] and its structure was recently described [8]. In fact, non-encapsulated mutants were shown to be avirulent in mice and pig models of infection [2]. Among several proteins and enzymes, a hemoly
Genetic diversity of Streptococcus suis isolates as determined by comparative genome hybridization
Astrid de Greeff, Henk J Wisselink, Freddy M de Bree, Constance Schultsz, Christoph G Baums, Hoa Thi, Norbert Stockhofe-Zurwieden, Hilde E Smith
BMC Microbiology , 2011, DOI: 10.1186/1471-2180-11-161
Abstract: In this study, the correlation between gene content, serotype, phenotype and virulence among 55 S. suis strains was studied using Comparative Genome Hybridization (CGH). Clustering of CGH data divided S. suis isolates into two clusters, A and B. Cluster A isolates could be discriminated from cluster B isolates based on the protein expression of extracellular factor (EF). Cluster A contained serotype 1 and 2 isolates that were correlated with virulence. Cluster B mainly contained serotype 7 and 9 isolates. Genetic similarity was observed between serotype 7 and serotype 2 isolates that do not express muramidase released protein (MRP) and EF (MRP-EF-), suggesting these isolates originated from a common founder. Profiles of 25 putative virulence-associated genes of S. suis were determined among the 55 isolates. Presence of all 25 genes was shown for cluster A isolates, whereas cluster B isolates lacked one or more putative virulence genes. Divergence of S. suis isolates was further studied based on the presence of 39 regions of difference. Conservation of genes was evaluated by the definition of a core genome that contained 78% of all ORFs in P1/7.In conclusion, we show that CGH is a valuable method to study distribution of genes or gene clusters among isolates in detail, yielding information on genetic similarity, and virulence traits of S. suis isolates.Streptococcus suis forms a problem in the swine industry. Clinically healthy sows carry S. suis in their nasal cavities and on their tonsils, and transmit the bacteria to their piglets [1], that develop a variety of infections, such as septicaemia, meningitis, polyarthritis, and endocarditis, and often do not survive [2]. S. suis occasionally causes meningitis, arthritis or endocarditis in humans. However, recently several large human outbreaks of S. suis have been described in China [3,4], and Thailand [5], whilst S. suis meningitis has become endemic in Vietnam [6,7], suggesting that isolates that are more virulent t
Effects of live Brucella abortus strain 19 vaccine on reindeer later challenge exposed with Brucella suis type 4
Robert A. Dieterich,Jamie K. Morton
Rangifer , 1987,
Abstract: Twelve reindeer (Rangifer tarandus) were vaccinated with Brucella abortus strain 19 vaccine and challenge exposed with B. suis type 4 two and one-half months later during mid-gestation. An additional 10 reindeer served as non-vaccinated controls. A sharp serologic titer response was observed in both vaccinates and controls. Brucella suis type 4 was isolated from tissues and blood from most controls (8 of 10, and 7 of 10 respectively). Seven of 11 vaccinated cows aborted, gave birth to weak fawns that died, or were not pregnant at the completion of the experiment. Brucella suis type 4 was isolated from the tissue of 4 of 12 vaccinates at necropsy. It was concluded that, under the conditions of this experiment, B. abortus strain 19 vaccine in reindeer did not provide adequate protection against challenge exposure with virulent B. suis type 4 organisms.
Mutations in the Gene Encoding the Ancillary Pilin Subunit of the Streptococcus suis srtF Cluster Result in Pili Formed by the Major Subunit Only  [PDF]
Nahuel Fittipaldi,Daisuke Takamatsu,María de la Cruz Domínguez-Punaro,Marie-Pier Lecours,Diane Montpetit,Makoto Osaki,Tsutomu Sekizaki,Marcelo Gottschalk
PLOS ONE , 2012, DOI: 10.1371/journal.pone.0008426
Abstract: Pili have been shown to contribute to the virulence of different Gram-positive pathogenic species. Among other critical steps of bacterial pathogenesis, these structures participate in adherence to host cells, colonization and systemic virulence. Recently, the presence of at least four discrete gene clusters encoding putative pili has been revealed in the major swine pathogen and emerging zoonotic agent Streptococcus suis. However, pili production by this species has not yet been demonstrated. In this study, we investigated the functionality of one of these pili clusters, known as the srtF pilus cluster, by the construction of mutant strains for each of the four genes of the cluster as well as by the generation of antibodies against the putative pilin subunits. Results revealed that the S. suis serotype 2 strain P1/7, as well as several other highly virulent invasive S. suis serotype 2 isolates express pili from this cluster. However, in most cases tested, and as a result of nonsense mutations at the 5′ end of the gene encoding the minor pilin subunit (a putative adhesin), pili were formed by the major pilin subunit only. We then evaluated the role these pili play in S. suis virulence. Abolishment of the expression of srtF cluster-encoded pili did not result in impaired interactions of S. suis with porcine brain microvascular endothelial cells. Furthermore, non-piliated mutants were as virulent as the wild type strain when evaluated in a murine model of S. suis sepsis. Our results show that srtF cluster-encoded, S. suis pili are atypical compared to other Gram-positive pili. In addition, since the highly virulent strains under investigation are unlikely to produce other pili, our results suggest that pili might be dispensable for critical steps of the S. suis pathogenesis of infection.
Acute meningitis by Streptococcus suis
Maria-Jesus Corrales-Arroyo,Maria Angeles Del Real-Francia,Amalia Hernandez-Gonzalez,Jose Manuel Morales Puebla
Journal of Microbiology and Infectious Diseases , 2012,
Abstract: Streptococcus suis is a coccus Gram positive, anaerobic optional. Human infection by this microorganism is a zoonoticdisease that usually presents as purulent meningitis. Mortality is low but is common sequelae. A case of meningitis byS. suis secondary to contact with pigs is presented here. A 35-year-old male patient was admitted to the hospital complainingof high fever, malaise, vomiting and headache. A physical examination revealed decreased level of consciousness,with adequate response to painful stimulus and his eyes with deconjugated gaze. S. suis was isolated in bloodculture. He was treated with cefotaxime, vancomycin and acyclovir in the intensive care unit. He experienced progressiveimprovement. He was discharged with severe deafness and a minimally unstable gait as sequellae. J Microbiol Infect Dis2012; 2(4): 160-162Key words: Streptococcus suis, meningitis, deafness.
Meningitis por Streptococcus suis
Geffner Sclarsky,D. E.; Moreno Mu?oz,R.; Campillo Alpera,Ma.S.; Pardo Serrano,F.J.; Gómez Gómez,A.; Martínez-Lozano,Ma.D.;
Anales de Medicina Interna , 2001, DOI: 10.4321/S0212-71992001000600007
Abstract: human infection by streptococcus suis (s. suis) is a zoonosis, with a known occupational risk and clinical presentation mainly as a purulent meningitis with low mortality and frequent hearing loss and ataxia sequela. less than 150 human cases have been reported since original one thirty years ago. there is a geographical distribution most patients living in northen europe and south asia. s. suis disease in human has been reported in two patients in spain the last years. we present two patients with s. suis meningitis, both were men with occupation related by pork meet, and good outcome. they come at our hospital in a lapse of one month. both had neurosensorial hearing loss and walking ataxia. one patient had peripheral facial paralysis and diplopia because of paresia of contralateral sixth nerve, with complete resolution at three months.the rare presentation of s. suis meningitis in our country must not forget us to record the working risk at anamnesis.
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