七日热群钩端螺旋体的一个新血清型

A23株钩端螺旋体系1962年8月3日分离富云南省勐腊县钩端螺旋体病患者血液。经交叉凝集试验、交又凝集素吸收试验和园子血清凝集试验证明是七日热群保林卡那亚群钩端螺旋体的一个新血清型。建议命名为致病性钩端螺旋体曼I芏血清型(Leptospira interrogansserovar manzhucng)。此型感染在勐腊已发现6例。     

北京地区首次发现赖型钩端螺旋体

采用分群和分型因子血清经显微镜凝集试验（ＭＡＴ）对由北京市顺义县某农场稻田周围黑线姬鼠肾组织分离获得的４株钩端螺旋体（钩体）进行了血清学分类检定。均属黄疸出血群赖型钩体。这在北京地区尚属首次分离发现。     

利用患者血清鉴定钩端螺旋体感染血清型的研究

1996年7月与1997年8月,湖北仙桃、枝江和浙江龙游地区先后爆发钧体病流行。为了解当地的主要流行菌型,并进～步验证采用患者血清鉴定感染型别的可能性,我们选择了从湖北和浙江上述地区采集的47例确诊钩体患者血清进行了血清学分型。现将结果报告如下。1材料与方法1．三血清样品从湖北仙桃和枝江地区采集的30例确诊钩体病患者血清由湖北省卫生防疫站提供;从浙江龙游地区采集的问例确诊钩体病患者血清由浙江省卫生防疫站提供。1．2参考菌株15群群代表参考菌株及相应各群各型参考菌株均由检定所提供。豆．3检测方法以培养5～7d生长良好的15…     

中国钩端螺旋体rRNA基因多态性分析

以ＤｉｇｄＵＴＰ标记的１６ＳｒＲＮＡ及２３ＳｒＲＮＡ基因为探针,分析了八个血清群５４个血清型６４株国内外致病性钩端螺旋体参考株和２７株野生株染色体经限制性内切酶ＥｃｏＲⅠ消化后的ｒＲＮＡ基因限制性图谱。结果发现,９１株菌中共有５６个核糖核酸型（Ｒｉｂｏｔｙｐｅ,简称ＲＴ）,除部分血清群中少数不同的血清型有相同的ＲＴ型外,大部分血清型都有独特的ＲＴ型,同一血清群往往拥有共同的核心片段;除黄疸出血群的黄疸出血型外,同一血清型的国内和国际参考株的ＲＴ型不相同;大多数野生株的ＲＴ和相应血清型国内参考株相同,差异也只表现为谱形上个别带型的缺少和增加,所研究的波摩那型野生株的ＲＴ型和国际参考株相同而和国内参考株不同     

秋季群钩端螺旋体的一个新血清型

1962年7月1日自云南省西双版纳州勐腊县钩端螺旋体病患者的血培养物中分离出钩端螺旋体A6株。经交叉显凝试验和交叉凝集索吸收试验,证明它是秋季群钩端螺旋体的一个新血清型。建议命名为致病性钩端螺旋体南腊血清型(Leptospina inteirogans serovar nanla),A6株为参考株。     

七日热群钩端螺旋体的两个新血清型

本文报告七日热群钩端螺旋体的两个新血清型。A10株钩端螺旋体系I 962年自勐腊县钩端螺旋体病患者分离,命名为云南型钩端螺旋体(Leptospira interrogans serovar yunnan)H27株钩端螺旋体系1964年自河口县钩端螺旋体病患者分离,命名为河口型钩端螺旋体(Le-ptospira interrogans serovar hekou)。     

致病性钩端螺旋体的三个新血清型

自云南省分离出3株(M48、A31、A82)钩端螺旋体,经鉴定它们分别属于3个新血清型。M48系1960年从耿马县猪肾分离,A31与A82分别于1962、1970年自勐腊县病人血培养物中分离。拟分别命名为耿马型(M48)、版纳型(A31)和勐捧型(A82)钩端螺旋体[Leptospirainterrogans serovars gengma (M48),banna (A3I)和mengpeng (A82)]。其中耿马型应属于塔拉索夫血清群;版纳型和勐捧型的血清学性质界于塔拉索夫和歇尔曼二群之间,其血清学归属尚难肯定。     

钩端螺旋体四个新血清型的鉴定

本文报告从云南省的病人和动物分离出的钩端螺旋体地方株中{个新血清型的鉴定结果。检定L82株及S590株为爪哇群,分别命名镇康型(Zhenkang)、勐马型(Mongma);S621株为致热群盂连型(Manglian);L100株为塔拉索夫群云县型(Yunxian)钩端螺旋体。     

我国钩端螺旋体型因子血清研究概况

我国的钩端螺旋体抗原分析和因子血清研究殆与Kmety同时于20世纪50年代末起步。但研究对象和目的大不相同,体会有所不同。现将个人体会结合国内外研究情况作梗概介绍。供同仁参考。     

临床标本钩端螺旋体L型的分离与形态学鉴定

应用钩体Ｌ型培养基成功地从钩体病神经系统后发症患者的血液和／或脑脊液标本中分离到钩体Ｌ型。将Ｌ型阳性培养物转种于钩体Ｌ型固体平板培养,发现基落形态呈丝状（Ｆ）型。对所分离到的Ｌ型进行形态学鉴定表明：Ｌ型呈球状体、螺旋状的丝状体等多形性改变,电镜下见Ｌ型推动部分或全部细胞壁,Ｌ型经免疫标记技术鉴定证实为钩体的Ｌ型。     

Immunodominant antigens recognized by the human immune response to infection by organisms of the species Leptospira interrogans serogroup Australis

Abstract Serum samples from patients infected by organisms of Leptospira interrogans serogroup Australis were tested by Western blot to determine the nature of major antigens that are involved in the immune response. Although there was some patient-to-patient variability, immunodominant genus-specific antigens were found to be proteins of apparent molecular ratio 68, 46 and 35-kDa, and lipopolysaccharide (LPS) sub-units in the 35-14-kDa region. Serogroup epitopes specific for Australis were exclusively saccharides of about 32 and 24 kDa: a serovar-specific antigen for serovar lora was of 38–40 kDa and behaved like a protein. Antibodies to the LPS serogroup-specific antigens and to the 38–40 kDa protein were long-lasting and consequently suggest that these immunodominant epitopes are important in resistance to re-infection.     

问号钩体粘附侵袭相关基因特征分析

使用NCBI ,Swissprot/TrEMBL ,ProDom ,Pfam ,Tmpred ,SignalP ,ClustW等网络资源和软件 ,根据问号钩体黄疸出血型赖株粘附侵袭相关基因诠释结果 ,对mce ,invA ,mviN和atsE四个粘附侵袭相关基因编码蛋白的结构域、跨膜区域和信号肽等进行了详细分析 ,并使用Bioedit,Mega2软件进行氨基酸多重序列比较并绘制系统发生树。结果显示 ,mce和mviN为穿膜蛋白 ,invA和atsE为菌体内蛋白质 ;许多对哺乳动物和对植物致病的微生物具有mce ,invA ,mviN和atsE四个粘附侵袭相关基因 ,其表达的蛋白质在感染宿主过程中起重要作用 ,钩体的粘附侵袭相关蛋白与它们在一级结构上有较高相似性。据生物信息学结果推测 ,问号钩体黄疸出血型赖株粘附侵袭相关基因和钩体致病性间有密切关系 ,其编码蛋白在致病过程中可能起重要作用     

Molecular and phenotypic characterization of Leptospira johnsonii sp. nov., Leptospira ellinghausenii sp. nov. and Leptospira ryugenii sp. nov. isolated from soil and water in Japan

In a previous study, 50 of 132 soil samples collected throughout Japan were found to be Leptospira‐positive. In the present study, three strains identified in the collected specimens, three, E8, E18 and YH101, were found to be divergent from previously described Leptospira species according to 16S ribosomal RNA gene sequence analysis. These three strains have a helical shape similar to that of typical Leptospira and were not re‐isolated from experimental mice inoculated with the cultured strains. Upon 16S ribosomal RNA gene sequence analysis, E8 was found to belong to the intermediate Leptospira species clade and E18 and YH101 to belong to the saprophytic Leptospira species clade. Based on analyses of genome‐to‐genome distances and average nucleotide identity in silico using whole genome sequences and DNA–DNA hybridization in vitro, these isolates were found to be distinct from previously described Leptospira species. Therefore, these three isolates represent novel species of the genus Leptospira for which the names Leptospira johnsonii sp. nov., (type strain E8 ^T, = JCM 32515 ^T = CIP111620 ^T), Leptospira ellinghausenii sp. nov., (type strain E18 ^T, = JCM 32516 ^T = CIP111618 ^T) and Leptospira ryugenii sp. nov., (type strain YH101 ^T, = JCM 32518 ^T = CIP111617 ^T) are proposed.     

Re-characterization of an extrachromosomal circular plasmid in the pathogenic Leptospira interrogans serovar Lai strain 56601

In China, Leptospira interrogans serovar Lai strain 56601 （str.56601） is one of main pathogenic strains that cause severe leptospirosis in both human and animals. The genome of this organism was completely sequenced in 2003. However, in 2011, we identified and corrected some assembly errors in the str.56601 genome due to the repeat sequences widely distributed in the Leptospira genome. In this study, we re-analyzed the previously reported mobile, phage-related genomic island in the chromosome and rectified detailed sequence information in both the plasmid and chromosome using various experimental methods. The presence of a separate circular extrachromosomal plasmid was also confirmed, and its location in the genomi~ region was determined relative to the genomic island reported in L. interrogans serovar Lai by a combination of pulsed-field gel electrophoresis -based and plasmid extraction-based Southern blot analysis. This report confirmed that the sep arate extrachromosomal circular plasmid is not integrated into the chromosome of. interrogans str.56601 and markedly improved our understanding of the genomic organization, evolution, and pathogenesis of L. interrogans. In particular, characterization of this extrachromosomal cir cular plasmid will contribute to the development of genetic manipulation systems in pathogenic Leptospira species.     

Genetic differences among the LPS biosynthetic loci of serovars of Leptospira interrogans and Leptospira borgpetersenii

The gene organization in the lipopolysaccharide biosynthetic (rfb) locus was analyzed in seven Leptospira interrogans serovars within serogroup Icterohemorrhagiae, seven non-Icterohemorrhagiae serovars and one Leptospira borgpetersenii serovar. Two groups of loci were delineated based on DNA hybridization and sequence analysis. Group 1 contained the two Hardjo subtypes, Hardjoprajitno and Hardjobovis. Group 2 (containing Copenhageni, Pomona, Naam, Mwogolo, Smithi, Lai, Canicola, Autumnalis, Pyrogenes, Australis and Icterohemorrhagiae) differed from Group 1 in its organization upstream of orf11, where five ORFs (32, 33, 34, 35, 37) were identified that were not contained in the Group 1 loci. These ORFs encoded a putative epimerase (orf32), a glycosyltransferase (orf33), two integral membrane proteins (orfs 34 and 35), and a galactosyltransferase (orf37). Serovars Australis, Pomona and Autumnalis did not contain orf37. Serovar Bataviae was excluded from the grouping because of its unique genetic organization upstream of orf13. In the Group 2 loci, comparison of the genetic layout at the 5' end revealed differences which included mutations disrupting reading frames in either or both orf34 and orf35 and apparent allelic differences between orf33 homologs that may be sufficient to account for the genetic basis of serovar identity.     

Identification of three extra-chromosomal replicons in Leptospira pathogenic strain and development of new shuttle vectors

Background

The genome of pathogenic Leptospira interrogans contains two chromosomes. Plasmids and prophages are known to play specific roles in gene transfer in bacteria and can potentially serve as efficient genetic tools in these organisms. Although plasmids and prophage remnants have recently been reported in Leptospira species, their characteristics and potential applications in leptospiral genetic transformation systems have not been fully evaluated.

Results

Three extrachromosomal replicons designated lcp1 (65,732 bp), lcp2 (56,757 bp), and lcp3 (54,986 bp) in the L. interrogans serovar Linhai strain 56609 were identified through whole genome sequencing. All three replicons were stable outside of the bacterial chromosomes. Phage particles were observed in the culture supernatant of 56609 after mitomycin C induction, and lcp3, which contained phage-related genes, was considered to be an inducible prophage. L. interrogans–Escherichia coli shuttle vectors, constructed with the predicted replication elements of single rep or rep combined with parAB loci from the three plasmids were shown to successfully transform into both saprophytic and pathogenic Leptospira species, suggesting an essential function for rep genes in supporting auto-replication of the plasmids. Additionally, a wide distribution of homologs of the three rep genes was identified in L. interrogans isolates, and correlation tests showed that the transformability of the shuttle vectors in L. interrogans isolates depended, to certain extent, on genetic compatibility between the rep sequences of both plasmid and host.

Conclusions

Three extrachromosomal replicons co-exist in L. interrogans, one of which we consider to be an inducible prophage. The vectors constructed with the rep genes of the three replicons successfully transformed into saprophytic and pathogenic Leptospira species alike, but this was partly dependent on genetic compatibility between the rep sequences of both plasmid and host.

Electronic supplementary material

The online version of this article (doi:10.1186/s12864-015-1321-y) contains supplementary material, which is available to authorized users.     

Serological and molecular characterization of leptospira serovar Kenya from captive African giant pouched rats (Cricetomys gambianus) from Morogoro Tanzania

Two identical leptospiral isolates coded Sh9 and Sh25 obtained from the urine of captive African giant pouched rats (Cricetomys gambianus), destined for use as biodetector of antipersonnel landmines were typed as serovar Kenya using cross-agglutination absorption test and DNA fingerprinting with the insertion element sequences IS1533 and IS1500 derived primers. The two isolates were previously characterized using cultural and serological-microagglutination test as pathogenic leptospires of the serogroup Ballum, closely related to serovars Kenya and Peru. To our knowledge, this is the first reported in-depth characterization of leptospira isolates from Tanzania.     

问号钩端螺旋体基因组DNA芯片的研制

目的 研制并初步评估问号钩端螺旋体(简称钩体)赖型赖株的基因组DNA芯片。方法 利用Primegens引物设计软件筛选出问号钩体赖型赖株全基因组中的特异性基因进行引物设计。对成功设计出相应引物的3 290个基因用聚合酶链反应方法进行扩增,以纯化后的产物点样制备芯片。并用双色荧光杂交策略对芯片质量进行了初步平估。结果 共获得3 290个基因产物用于点样。参考株自身杂交实验结果表明:该芯片有较高的点一致性、信噪比和较低的假阳性率。结论 成功制备了包含问号钩体赖型赖株3 290个目的基因的基因组DNA芯片,并可用于基于该芯片的问号钩体比较基因组学的研究。     

Characterization of cheW genes of Leptospira interrogans and their effects in Escherichia coli

The motility and chemotaxis systems are critical for the virulence of leptospires. In this study, the phylogenetic profiles method was used to predict the interaction of chemotaxis proteins. It was shown that CheW1 links to CheA1, CheY, CheB and CheW2, CheW3 links to CheA2, MCP （LA2426）, CheB3 and CheD1; and CheW2 links only to CheW1. The similarity analysis demonstrated that CheW2 of Leptospira interrogans strain Lai had poor homology with Chew of Escherichia coli in the region of residues 30-50. In order to verify the function of these proteins, the putative cheW genes were cloned into pQE31 vector and expressed in wild-type E. coli strain RP437 or chew defective strain RP4606. The swarming results indicated that CheW1 and CheW3 could restore swarming of RP4606 while CheW2 could not. Overexpression of CheW1 and CheW3 in RP437 inhibited the swarming of RP437, whereas the inhibitory effect of CheW2 was much lower. Therefore, we presumed that CheW1 and CheW3 might have the function of CheW while CheW2 does not. The existence of multiple copies of chemotaxis homologue genes suggested that L. interrogans strain Lai might have a more complex chemosensory pathway.     

Effect of osmolarity and viscosity on the motility of pathogenic and saprophytic Leptospira

The motility of bacteria is an important factor in their infectivity. In this study, the motility of Leptospira, a member of the spirochete family that causes a zoonotic disease known as leptospirosis, was analyzed in different viscous or osmotic conditions. Motility assays revealed that both pathogenic and saprophytic strains increase their swimming speeds with increasing viscosity. However, only pathogenic Leptospira interrogans maintained vigorous motility near physiological osmotic conditions. This suggests that active motility in physiological conditions is advantageous when Leptospira enters hosts and when it migrates toward target tissues.