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B-cell-specific peptides of leptospira interrogans LigA for diagnosis of patients with acute leptospirosis.

ABSTRACT: Leptospirosis is a reemerging infectious disease that is underdiagnosed and under-recognized due to low-sensitivity and cumbersome serological tests. Rapid reliable alternative tests are needed for early diagnosis of the disease. Considering the importance of the pathogenesis-associated leptospiral LigA protein expressed in vivo, we have evaluated its application in the diagnosis of the acute form of leptospirosis. The C-terminal coding sequence of ligA (ligA-C) was cloned into pET15b and expressed in Escherichia coli. Furthermore, the B-cell-specific epitopes were predicted and were synthesized as peptides for evaluation along with recombinant LigA-C. Epitope 1 (VVIENTPGK), with a VaxiJen score of 1.3782, and epitope 2 (TALSVGSSK), with a score of 1.2767, were utilized. A total of 140 serum samples collected from leptospirosis cases during the acute stage of the disease and 138 serum samples collected from normal healthy controls were utilized for evaluation. The sensitivity, specificity, positive predictive value, and negative predictive value were calculated for the recombinant LigA-C-specific IgM enzyme-linked immunosorbent assay (ELISA) and were found to be 92.1%, 97.7%, 92.8%, and 97.5%, respectively. Epitopes 1 and 2 used in the study showed 5.1 to 5.8% increased sensitivity over recombinant LigA-C in single and combination assays for IgM antibody detection. These findings suggest that these peptides may be potential candidates for the early diagnosis of leptospirosis.

SUBMITTER: Kanagavel M

PROVIDER: S-EPMC3957670 | biostudies-literature | 2014 Mar

REPOSITORIES: biostudies-literature

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B-cell-specific peptides of leptospira interrogans LigA for diagnosis of patients with acute leptospirosis.

Kanagavel Murugesan M Shanmughapriya Santhanam S Anbarasu Kumarasamy K Natarajaseenivasan Kalimuthusamy K

Clinical and vaccine immunology : CVI 20140108 3

Leptospirosis is a reemerging infectious disease that is underdiagnosed and under-recognized due to low-sensitivity and cumbersome serological tests. Rapid reliable alternative tests are needed for early diagnosis of the disease. Considering the importance of the pathogenesis-associated leptospiral LigA protein expressed in vivo, we have evaluated its application in the diagnosis of the acute form of leptospirosis. The C-terminal coding sequence of ligA (ligA-C) was cloned into pET15b and expres ...[more]

PMID: 24403522

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Project description:BackgroundLeptospirosis is caused by pathogenic spirochetes of the genus Leptospira spp. This zoonotic disease is distributed globally and affects domestic animals, including cattle. Leptospira interrogans serogroup Sejroe serovar Hardjo and Leptospira borgpetersenii serogroup Sejroe serovar Hardjo remain important species associated with this reproductive disease in livestock production. Previous studies on Brazilian livestock have reported that L. interrogans serovar Hardjo is the most prevalent leptospiral agent in this country and is related to clinical signs of leptospirosis, which lead to economic losses in production. Here, we described the isolation of three clinical strains (Norma, Lagoa and Bolivia) obtained from leptospirosis outbreaks that occurred in Minas Gerais state in 1994 and 2008.ResultsSerological and molecular typing using housekeeping (secY and 16SrRNA) and rfb locus (ORF22 and ORF36) genes were applied for the identification and comparative analysis of Leptospira spp. Our results identified the three isolates as L. interrogans serogroup Sejroe serovar Hardjo and confirmed the occurrence of this bacterial strain in Brazilian livestock. Genetic analysis using ORF22 and ORF36 grouped the Leptospira into serogroup Sejroe and subtype Hardjoprajitno. Genetic approaches were also applied to compare distinct serovars of L. interrogans strains by verifying the copy numbers of the IS1500 and IS1533 insertion sequences (ISs). The IS1500 copy number varied among the analyzed L. interrogans strains.ConclusionThis study provides evidence that L. interrogans serogroup Sejroe serovar Hardjo subtype Hardjoprajitno causes bovine leptospirosis in Brazilian production. The molecular results suggested that rfb locus (ORF22 and ORF36) could improve epidemiological studies by allowing the identification of Leptospira spp. at the serogroup level. Additionally, the IS1500 and IS1533 IS copy number analysis suggested distinct genomic features among closely related leptospiral strains.

Project description:Leptospirosis is a globally distributed bacterial infectious disease caused by pathogenic members of the genus Leptospira. Infection can lead to illness ranging from mild and non-specific to severe, with jaundice, kidney and liver dysfunction, and widespread endothelial damage. The adhesion of pathogenic Leptospira species (spp.), the causative agent of leptospirosis, to host tissue components is necessary for infection and pathogenesis. While it is well-established that extracellular matrix (ECM) components play a role in the interaction of the pathogen with host molecules, we have shown that pathogenic Leptospira interrogans binds to host cells more efficiently than to ECM components. Using in vitro phage display to select for phage clones that bind to EA.hy926 endothelial cells, we identified the putative lipoproteins LIC10508 and LIC13411, and the conserved hypothetical proteins LIC12341 and LIC11574, as candidate L. interrogans sv. Copenhageni st. Fiocruz L1-130 adhesins. Recombinant LIC11574, but not its L. biflexa homologue LBF1629, exhibited dose-dependent binding to both endothelial and epithelial cells. In addition, LIC11574 and LIC13411 bind to VE-cadherin, an endothelial cell receptor for L. interrogans. Extraction of bacteria with the non-ionic detergent Triton X-114 resulted in partitioning of the candidate adhesins to the detergent fraction, a likely indication that these proteins are outer membrane localized. All candidate adhesins were recognized by sera obtained from leptospirosis patients but not by sera from healthy individuals as assessed by western blot. This work has identified bacterial adhesins that are potentially involved in L. interrogans infection of the mammalian host, and through cadherin binding, may contribute to dissemination and vascular damage. Our findings may be of value in leptospirosis control and prevention, with the bacterial adhesins potentially serving as targets for development of diagnostics, therapeutics, and vaccines.

Dataset Information

B-cell-specific peptides of leptospira interrogans LigA for diagnosis of patients with acute leptospirosis.

Publications

B-cell-specific peptides of leptospira interrogans LigA for diagnosis of patients with acute leptospirosis.

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