Tapered optical fiber DNA biosensor for detecting Leptospira DNA

Objective: To establish a DNA detection platform based on a tapered optical fiber to detect Leptospira DNA by targeting the leptospiral secY gene.Methods: The biosensor works on the principle of light propagating in the special geometry of the optical fiber tapered from a waist diameter of 125 to 12 μm. The fiber surface was functionalized through a cascade of chemical treatments and the immobilization of a DNA capture probe targeting the secY gene. The presence of the target DNA was determined from the wavelength shift in the optical transmission spectrum.Results: The biosensor demonstrated good sensitivity, detecting Leptospira DNA at 0.001 ng/μL, and was selective for Leptospira DNA without cross-reactivity with non-leptospiral microorganisms. The biosensor specifically detected DNA that was specifically amplified through the loop-mediated isothermal amplification approach.Conclusions: These findings warrant the potential of this platform to be developed as a novel alternative approach to diagnose leptospirosis.

Severe community-acquired pneumonia caused by Leptospira interrogans:A case report and review of literature

BACKGROUND Leptospira is an uncommon pathogen for adult severe community-acquired pneumonia and its nonspecific manifestations and limited diagnostic tests make it difficult to identify.Although conventional penicillin remains efficacious to treat leptospirosis,failure in early diagnosis and treatment can lead to progression into a deadly syndrome with multiple organ dysfunction.Next generation sequencing is of great value to understand cases with infection of unknown cause,which could help in the diagnosis of uncertain Leptospira infection.CASE SUMMARY We recently managed a patient with fever,cough and dyspnea on admission that progressed into persistent adult respiratory distress syndrome,hemoptysis and hematuria after admission.In this case,the rare Leptospira infection was clouded by the positive influenza tests at admission,delaying early Leptospira-targeted antibiotics administration.Next generation sequencing,a novel molecular diagnostic tool,provided a key hint to uncover the crucial pathogen,Leptospira interrogans,further supported by the possible occupational exposure history.Subsequent conventional penicillin and mechanical respiratory support were administrated to cure the patient successfully without any sequela.CONCLUSION Clinicians must pay attention to possible exposure history and keep uncommon Leptospira in mind when managing pneumonia with unknown causes.

Spatial Analysis of <i>Leptospira</i>in Rats, Water and Soil in Bantul District Yogyakarta Indonesia

Leptospirosis is a potential threat to public health. An increasing number of people infected with Leptospira were reported in Bantul District, Yogyakarta special region with a case fatality rate (CFR) of 7.8%. Infected areas in the district have increased from 2 to 15 sub districts. Leptospirosis is caused by Leptospira bacteria and spread by direct contact with infected rodents and indirect contact through contaminated water or soil. Leptospira in rats, water and soil were detected using real-time quantitative polymerase chain reaction (qPCR). The sites of sampled materials were geocoded using Global Positioning System (GPS). Spatial analysis was used to predict the spread of Spira. This study aims to perform the mapping, clustering, and predicting the spread of Leptospira in Bantul Yogyakarta Indonesia. Data were collected from three sub-districts: Sedayu, Sewon and Bantul. The result showed that 38.04% from 368 samples were Spira positive. There were four significant clusters of infection spread source. Spira is predicted to spread in, and out from, Bantul District.

Comparative analysis of current diagnostic PCR assays in detecting pathogenic Leptospira isolates from environmental samples

Objective:To compare the efficiency of routine diagnostic PCR assays in detecting pathogenic Leptospira isolated from water and soils.Methods:Seven routine assays targeting six genes(lip L32,fla B,gyr B,lfb1,sec Y and lig B)were evaluated and compared on the cultures of two groups of pathogenic Leptospira from different sources.One group included 19 described reference strains recovered from infected human or animals,and another group included 22 environmental isolates from recreational and residential sites in Malaysia.The latter have been confirmed for presence of pathogenic Leptospira DNA.PCR positivity or detection sensitivity of each assay was determined and compared between the two groups.Results:Validation on reference strains showed 100.0%PCR sensitivity for all assays except lig B-PCR(95.0%)that failed to amplify Leptospira interrogans serovar Pomona.In marked contrast,there was a notable decline in sensitivity in the environmental isolates(lip L32-PCR,95.5%;fla B-PCR,90.9%;gyr B-PCR,77.3%;lfb1-PCR,59.1%;sec Y-PCRs,40.9%G1/G2-PCR,36.4%;lig B-PCR,13.6%),implying a large genetic distance between the two groups,as well as nucleotide polymorphism among environmental isolates.Conclusions:High proportion of false-negative PCR results suggests a need of prudent selection of primers in detecting environmental pathogenic Leptospira.These findings offer valuable insights on the extensive biodiversity of genus Leptospira and its impact on the efficacy and development of molecular detection tool.

First histopathological study in kidneys of rodents naturally infected with Leptospira pathogenic species from Yucatan, Mexico

Objective: To report the renal histological lesions in synanthropic rodents, Mus musculus and Rattus rattus, naturally infected with Leptospira spp., captured in a rural community in Yucatan, Mexico. Methods: Kidney samples of synanthropic rodents were collected from a rural community in Yucatan, Mexico. Polymerase chain reaction was used to detect Leptospira spp. infection. Tissue kidney was fixed in 10% buffered formalin, processed according to the usual techniques for paraffin inclusion, cut and stained with hematoxylin and eosin, and examined using a conventional electronic microscope. Results: A total of 187 rodents were captured. Nine individuals(4.8%) were positive for Leptospira spp. in the molecular analysis. All renal lesions observed in the histopathological study had been reported previously for Leptospira spp. infection. Conclusions: The histopathological lesions are present in the kidneys, plus the results of the polymerase chain reaction confirm that these rodents are true carriers of Leptospira spp.

Genetic variation of Leptospira isolated from rats catched in Yogyakarta Indonesia

Objective: To detect genetic variations among pathogenic Leptospira isolated from rats using 16 S r RNA gen as chronometer. Methods: This is an observational study with cross sectional design. Rats saples were taken in Yogyakarta Special Region of Indonesia. Leptospira in the rats was detected by two methods ie. real time PCR(q PCR) by using primers correspond to16 S r RNA gene of Leptospira, and standard PCR by using dif erent set of primer correspond to the 16 S r RNA gene of Leptospira. The standard PCR amplicon then subjected for DNA sequencing. Analysis genetic variation was performed using MEGA 6.2. Software. Results:There were 99 DNA samples from rats included in this study. Detection of Leptospira by using q PCR revealed 25 samples positive for pathogenic Leptospira, while only 6 samples were able to be detected using standard PCR. The new primer set correspond to 16 S r RNA gene was able to detect specii cally pathogenic Leptospira in the rats. Sequencing analysis of 6 PCR amplicons showed that the Leptospira which infect the rats catched in Yogyakarta genetically close related with pathogenic Leptospira which were isolated from human, animal, rodents, and environment. Conclusions: It can be considered that rats are the most important vector and reservoir of Leptospira.

LipL21 mRNA expression in lungs of hamsters infected with pathogenic Leptospira

Objective:Pulmonary haemorrhage is an increasing cause of death in leptospirosis patients.However,molecular mechanism underlying pathologies in this organ is not clearly understood.It has been shown that sodium transport was disturbed following Leptospira infection.LipL21 is the second abundant outer membrane protein found only in pathogenic Leptospira.Its expression in vivo has been shown which suggests that this protein may be involved in survival in hosts or pathogenesis.However,the expression of this protein in host organs and its role in lung pathology has not been demonstrated.In this study we demonstrated the expression of LipL21 in lungs of hamsters infected with pathogenic Leptospira.Methods:Lung tissues were collected from Golden Syrian hamsters injected with Leptospira interrogans serovar Pyrogenes at days 3,5 and 7 post-infection.Four hamsters were used for each time point.Lungs from non-infected hamsters were collected as a control group.LipL21 mRNA expression in lung tissues was investigated by reverse transcription and nested PCR.Results:LipL21 mRNA expression was detected in all lung tissues from hamsters infected with pathogenic Leptospira.No PCR product was detected when tissues from non-infected hamsters were investigated.Conclusion:Our data demonstrated that LipL21 is expressed in lungs of hamsters infected with pathogenic Leptospira.Additional experiments such as quantitation and localization of LipL21 expression in lungs will provide further information whether this protein is involved in pathogenesis.

问号钩体(Leptospira interrogans)Ⅲ型分泌系统相关基因分析

根据鼠疫耶尔森菌Ⅲ型分泌系统和大肠杆菌鞭毛系统中已知蛋白,以NCB I/B last软件搜索问号钩体全基因组中与之高度同源的蛋白;用TMHMM-2.0对所得蛋白跨膜区进行分析;用In terpro对所得蛋白结构域进行分析,初步整理出问号钩体Ⅲ型分泌系统组成,发现10个被注释为鞭毛系统的相关基因和Ⅲ型分泌系统有关,并和鼠疫耶尔森菌Ⅲ型分泌系统相关蛋白有较高同源性。6个为跨内膜蛋白,4个为胞内蛋白,并共同组成Ⅲ型分泌系统的内膜孔道,但没能发现组成外膜孔道的蛋白组分。问号钩体Ⅲ型分泌系统与鞭毛组装系统共用相同的部分组分,但钩体Ⅲ型分泌系统和经典Ⅲ型分泌系统有一定程度的差别:它缺乏外膜孔道结构,且分泌产物先进入胞周间隙。

High-coverage proteome analysis reveals the first insight of protein modification systems in the pathogenic spirochete Leptospira interrogans

细螺旋体病是感染人和大量动物的类螺旋体的病原的螺旋菌引起的普遍 zoonotic 疾病。由联合计算预言和高精确性的双人脚踏车团系列，我们修订了螺旋体 interrogans serovar Lai，为细螺旋体病负责的免费生活的病原的螺旋菌，为新奇基因的提供的实质的肽证据和新基因边界的染色体注解。随后，我们介绍了蛋白质表示和多重 posttranslational 修正(PTM ) 的高范围的 proteome 分析。约 64.3% 预言的 L。interrogans 蛋白质被检测 2 编目 540 蛋白质。同时，多重 PTM 的侧面并发地被建立，在总数 32 phosphorylated， 46 acetylated 和 155 methylated 蛋白质包含。在 serovar Lai 的 PTM 系统显示出唯一的特征。L 的唯一的象真核细胞一样特征。interrogans 蛋白质修正在 phosphorylation 和精氨酸 methylation 被表明。这系统的分析在建议的初核质而且一个看法提供高范围的蛋白质表示和多重修正的全面信息不仅 evolutionarily 原始的 L。interrogans 与优核质在蛋白质修正系统分享重要类似。

THE CONSTRUCTION AND EXPRESSION OF RECOMBINANT SHUTTLE PLASMID WITH OMPL1 GENE FROM LEPTOSPIRA INTERROGANS SEROVAR LAI STRAIN 017 IN BACILLE CALMETTE-GUERIN

Objective.To construct recombinant BCG again st leptospirosis.Methods.We amplified the entire open readin g frame of the OmpL1gene from the genome of the leptospire serovar Lai strain 017.Two recombin ant plasmids pBQ1and pBQ2were constructed by oriented ligation based on the E.coli-BCG shuttle plasmids pMV261and pMV361respectively.The recombinant plasmids were transformed into BCG by electroporation.The rBCGs bearing pBQ1and pBQ2were induced by high temperature of 45℃.Results.The expressed product,a 35kD prote in was detected by SDS-PAGE.The resu lt indicates that pBQ1and pBQ2can express OmpL1in rBCG.Conclusion.The technical methods in this study may help detect the immunogenicity a nd immunoprotection of OmpL1and develop more safe,highl y effective rBCG bearing leptospira l antigen with long-lasting protection.

Determination of the genus-specific antigens in outer membrane proteins from the strains of Leptospira interrogans and Leptospira biflexa with different virulence

Objective:To determine the existence of genus-specific antigens in outer membrane proteins (OMPs) of leptospira with different virulence. Methods: Microscope agglutination test (MAT) was applied to detect the agglutination between commercial rabbit antiserum against leptospiral genus-specific TR/Patoc I antigen and 17 strains of Leptospira interrongans belonging to 15 serogroups and 2 strains of Leptospira biflexa belonging to 2 serogroups.The outer envelopes (OEs) of L.interrogans serogroup Icterohaemorrhagiae serovar lai strain lai (56601) with strong virulence and serogroup Pomona serovar pomona strain Luo (56608) with low virulence,and L.biflexa serogroup Semaranga serovar patoc strain Patoc I without virulence were prepared by using the method reported in Auran et al.(1972).OMPs in the OEs were obtained by treatment with sodium deoxycholate. SDS-PAGE and western blot were used for analyzing the features of the OMPs on electrophoretic pattern and the immunoreactivity to the antiserum against TR/Patoc I antigen, respectively. Results:All the tested strains belonging to different leptospiral serogroups agglutinated to the antiserum against leptospiral genus-specific TR/Patoc I antigen with agglutination titers ranging from 1:256-1:512. A similar SDS-PAGE pattern of the OMPs from the three strains of leptospira with different virulence was shown and the molecular weight of a major protein fragment in the OMPs was found to be approximately 60 KDa.A positive protein fragment with approximately 32 KDa confirmed by Western blot,was able to react with the antiserum against leptospiral genus-specific TR/Patoc I antigen, and was found in each the OMPs of the three stains of leptospira.Conclusion: There are genus-specific antigens on the surface of L.interrogans and L.biflexa. The OMP with molecular weight of 32 KDa may be one of the genus-specific protein antigens of leptospira.

Production of reactive oxygen species and expression of inducible nitric oxide synthase in rat isolated Kupffer cells stimulated by Leptospira interrogans and Borrelia burgdorfen

瞄准：为了评估反应的氧种类(ROS ) 和可诱导的氮的氧化物的表示的生产，在老鼠的 synthase (i NOS ) 孤立肾脏钩端螺旋体和包柔氏螺旋体属 burgdorferi 刺激的 Kupffer 房间(KC ) 。方法：老鼠 Kupffer 房间被肝的灌注用 0.05% 胶原酶分开，并且由 Percoll 坡度净化了。净化的 Kupffer 房间是有活着的 L 的测试在试管内。interogans 和 B。burgdorferi 准备。ROS 的生产被化合光决定，而 i NOS 蛋白质表达式被西方的污点试金用 anti-iNOS 抗体计算。结果：B。burgdorferi 并且到一更少的程度 L。interrogans 与一座山峰导致了 ROS 生产在感染以后的 35 min。化合光信号日益增多地减少了并且由孵化的 180 min 是无法发现的。Leptospirae 和 borreliae 在在 6 个小时达到顶点并且仍然是明显的 Kupffer 房间导致了增加的 i NOS 表情在感染以后的 22 h。结论：螺旋菌的两个类在老鼠 Kupffer 细胞导致了 ROS 和 i NOS 生产。自从在 leptospiral 以及在 borrelial 感染的肝损坏的原因，仍然是未知的，我们建议那螺旋体和肝的包柔氏螺旋体属损坏能被氧激进分子开始调停，并且然后被氮的氧化物部分地至少维持。

A molecular dynamics simulation study of peptide deformylase from Leptospira interrogans complex:Exploring the closing mechanism of the substrate pocket

探索底层衣袋的关门机制，我们从螺旋体 interrogans 在 ligand 免费、 actinonin 固定的肽 deformylase 上独立执行 16,000 ps 分子动态学模拟。我们 CD 环，吸水的禁止者和恐水病的簇为半开的符合构造的形成是必要的结果表演，和 Tyr71 在调停起一个重要作用 CD 环的运动。actinonin 固定的 LiPDF 建筑群的平均 MD 结构来临到晶体结构。这些与实验一致很好。

Molecular cloning of genes flhA and flhB_2 for flagellar biosynthesis of Leptospira interrogans and functional prediction of the prokaryotic expressing products

To determine the pathogenic potential of the leptospiral flagella-associated proteins, the genes flhA and flhB2 encoding the biosynthesis of flagella of leptospira interrogans serogroup icterohaemorrhagiae serovar lai stain 56601 were cloned and their prokaryotic expression systems were constructed. It was demonstrated that the cloned flhA and flhB2 genes had 2118 bp in length and showed 100% and 99.9% of homologies in their nucleotide sequences and 100% and 98.8% of homologies in their putative amino acid sequences respectively, in comparison with those of previously reported. The prokaryotic expression systems under the induction with IPTG could efficiently express the target proteins rFlhA and rFlhB2 with the outputs of approximate 10% of the total bacterial proteins. Based on the sequences of the cloned genes flhA and flhB2, the structural features in associated with pathogenesis and the functions of the target proteins were analyzed with bioinformatics softwares, in which the FlhA was found to have 7 major transmembrane helices, while the FlhB2 had 5 ones. The conserved domains in the FlhA showed high similarity to those of the FHIPEP of the other bacterial FlhA and EscV families, but the conserved domains in the FlhB2 were similar to those of bac-export-2 and EscU families, EscV and EscU families being the protein products of the type Ⅲ secretion system in association with pathogenesis. The FlhA and FlhB2 also contained protein kinase C （PKC） and protein tyrosine kinase （PTK） phosphorylation sites, indicating that PKC and PTK of host cells were involved in the internalization and intracellular proliferation in the pathogenesis of microorganisms. All these data leads to a conclusion that the flhA and flhB2 genes of L.interrogans are relatively conserved and their gene products have great potential in the pathogenesis of this organism.

Genotypic Analysis, Construction of the Expression System and Immunological Identification of the Recombinant Proteins of the LipL32 Gene in the Dominant Serogroups of Leptospira interrogans in China

To investigate the existence of the major outer membrane protein (MOMP) gene LipL32 in 15 dominant Chinese strains of 15 serogroups of Leptospira interrogans and 2 international strains of 2 serogroups of Leptospira biflexa, and to clone and construct the expression system as well as to identify the recombinant proteins, genomic DNAs from strains of leptospira were prepared by routine phenol-chloroform method, and the fragments of the LipL32 gene with the whole length from the strains were amplified with high fidelity PCR. The target amplification products were sequenced after T-A cloning, and the expression system for the genes were thereby constructed. Expression of the recombinant proteins was identified by using SDS-PAGE after induction with IPTG at different dosages. Western blot assays with rabbit antiserum against the whole cell of TR/PatocⅠ of Leptospira and immunized serum with rMOMPs were used to determine the immunoreactivity and immunogenicity of the recombinant proteins. Microscopic agglutination test was used to determine the cross- agglutination titres in rabbit sera immunized with rMOMPs, and the cell adherence model of Leptospira was used to examine the blocking effects of rabbit antisera against these rMOMPs. It was found that the LipL32 gene could be found in all the 17 strains of Leptospira mentioned above with two different genotypes, i.e. LipL32/1 and LipL32/2. Amounts of expressions of rMOMP1 and rMOMP2 after IPTG accounted for 40% and 10% of the total bacterial proteins respectively. Both rMOMP1 and rMOMP2 could combine with the rabbit antiserum against leptospiral TR/PatocⅠ, and could induce the production of agglutination antibodies to these 17 strains of Leptospira with 1∶2 to 1∶64 MAT titres. The rabbit anti-rMOMP1 and anti-MOMP2 antibodies at 1∶2 to 1∶16 dilutions could efficiently block adherence of Leptospira. It concludes that all the Leptospira tested in the present study possess LipL32/1 or LipL32/2 genes, and the constructed expression system can express the rMOMP1 and rMOMP2. These recombinant proteins are showed to have good immunogenicity and satisfactory immunoreactivity.

2020-2022年湖南省鼠类钩端螺旋体病监测与菌株分型

目的初步了解湖南省钩端螺旋体病主要宿主动物鼠类的流行病学概况及分离菌株血清学和MLST基因型特征。方法2020-2022年在湖南省湘潭、涟源、双峰及浏阳4个钩体病监测点,开展鼠种构成调查、钩体分离培养、菌株鉴定工作。采用暗视野显微凝集试验(Microscopic agglutination test,MAT)对分离菌株开展血清群鉴定。利用MLST(Multilocus sequence typing)方法开展基因分型分析,分别对7个位点(glmU、pntA、sucA、tpiA、pfkB、mreA和caiB)进行普通PCR扩增、测序、序列分析。结果4个湖南钩体病监测点共捕啮齿类和食虫目动物461只,平均鼠密度为2.52%,共分离菌株56株,来源于黑线姬鼠、黄胸鼠和鼩鼱3个种类,钩体分离率是12.15%。湖南省4个监测点以黑线姬鼠为优势鼠种,占88.72%。经MAT鉴定显示:56株湖南钩体分离株隶属于黄疸出血群和澳洲群两个血清群,黄疸出血群为当地主要流行血清群,占94.64%。MLST分型结果显示:56株湖南省分离株ST型别包括ST1、ST128和ST1053个基因型,其中ST1(57.14%)和ST128(37.50%)为湖南主要流行ST型。利用BioNumerics软件进行UPGMA聚类分析显示:56株菌株被分为3个不同Clades,分别对应3个不同ST型,ST型分布在不同动物种类和地域上呈现明显差异。结论黄疸出血群、ST1和ST128为湖南省主要流行型别,初步了解湖南省钩端螺旋体病鼠类动物流行病学概况及流行菌株分子分型特征,对湖南省钩体病的防控和疫苗制备提供科学依据。

2021—2023年青岛市小兽携带鼠传病原体监测分析

目的调查青岛市小兽携带病原体基本情况,为鼠传疾病的防控工作提供理论依据。方法2021—2023年在青岛市西海岸新区、即墨区、胶州市、平度市和莱西市采集小兽肝、脾、肾和肺组织的核酸样本,采用实时荧光定量PCR(Quantitative Real-time PCR,qPCR)检测致病性钩端螺旋体和巴尔通体,反转录实时荧光定量PCR(Reverse transcription-qPCR,RT-qPCR)方法检测汉坦病毒和新型布尼亚病毒。对种群构成比、病原体感染率进行统计学分析。结果本次共捕获小兽624只,其中褐家鼠最多,占比为4311%,小家鼠占比为3686%、黑线姬鼠占比为1394%、鼩鼱占比为609%。病原学检测结果显示,鼠传病原体中汉坦病毒感染率为337%,致病性钩端螺旋体感染率为096%,巴尔通体感染率为112%;未检出新型布尼亚病毒。褐家鼠和小家鼠均检出3种病原体,以褐家鼠携带病原体感染率最高;监测生境中农田耕地生境鼠病原体检出感染率最高为820%。结论青岛市存在汉坦病毒、巴尔通体、致病性钩端螺旋体的鼠传病原感染情况,应加强农村地区的鼠传病原体监测和防控工作,降低人群鼠传病原体的感染风险。

钩端螺旋体OmpR家族相关TCS 鉴定及功能初探

目的建立钩端螺旋体(钩体)OmpR家族双组分系统(TCS)的鉴定方法,初步探讨OmpR家族TCS在钩体感染人巨噬细胞过程中对钩体主要外膜蛋白(OMPs)表达水平的调控作用。方法采用生物信息学技术预测致病性钩体赖株TCS的组氨酸激酶(HK)和应答调节蛋白(RR)OmpR功能结构域。采用激酶活性检测试剂盒和细菌双杂交技术进一步验证各HK激酶活性以及各HK与RR的相互作用。采用qRT-PCR法检测HK抗血清封闭前后钩体主要OMPs基因mRNA水平的变化。结果钩体56601株中LA2828、LA1710含有HATPase_c激酶催化结构域,LA2827、LA1709含有Response_reg与OmpR功能结构域,提示其分别可构成HK/OmpR-TCS。体外重组HK-2828和HK-1710蛋白均具有激酶活性,且分别与OmpR-2827和OmpR-1709存在直接的相互作用。HK-2828抗体封闭处理可使感染过程中显著下降的外膜蛋白编码基因lipL32、lipL41和ompL1的mRNA水平明显回升。结论成功构建钩体OmpR家族-TCS鉴定及功能探究体系,为阐明感染过程中问号钩体OMPs表达改变调控机制,以及制定基因工程疫苗OMPs抗原筛选新策略提供依据。

钩端螺旋体病流行过程及其影响因素概述

钩端螺旋体病是由致病性钩端螺旋体所引起的一种人兽共患传染病,呈全球性分布,可引发靶器官衰竭,甚至死亡。随着全球气候变暖、旅游开发、动物栖息地破坏、生态系统失衡等问题的出现,钩端螺旋体病人兽共患的风险显著加大。本文对钩端螺旋体病的病原学、流行病学及其疾病影响因素进行综述,旨在全面认识和有效防控该疾病,并为今后深入研究奠定基础,对保护人群生命健康具有重要意义。

Identification of Potential Leptospira Phosphoheptose Isomerase Inhibitors Through Virtual High-Throughput Screening

The life-threatening infections caused by Leptospira serovars demand the need for designing anti-leptospirosis drugs. The present study encompasses exploring inhibitors against phosphoheptose isomerase （GmhA） of Leptospira, which is vital for lipopolysaccharide （LPS） biosynthesis and is identified as a common drug target through the subtractive genomic approach. GmhA model was built in Modeller 9v7. Structural refinement and energy minimization of the predicted model was carried out using Maestro 9.0. The refined model reliability was assessed through Procheck, ProSA, ProQ and Profile 3D. The substrate-based virtual high-throughput screening （VHTS） in Ligand.Info Meta-Database tool generated an in-house library of 354 substrate structural analogs. Furthermore, structure-based VHTS from the in-house library with different conformations of each ligand provided 14 novel competitive inhibitors. The model together with insight gained from the VHTS would be a promising starting point for developing anti-leptospirosis competitive inhibitors targeting LPS biosynthesis pathway.