Leptospirosis Vaccine: Search for Subunit Candidates (original) (raw)
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PLOS ONE, 2023
Leptospirosis is a public health concern with lethality around 15% of the total cases. The current vaccines against Leptospira infection based on bacterins have several limitations, which require urgent development of new ones. In this context, groundbreaking approaches such as peptide-vaccines could be used to come around with promising results. Our goal was to identify conserved and immunogenic epitopes from the lipoprotein LruC that could interact with Major Histocompatibility Complex (MHC) I and II. LruC is a conserved lipoprotein expressed during leptospirosis that is considered among vaccine candidates and can be used as source for development of peptide-based vaccines. We searched for peptides that would be recognized by antibodies from either serum of hamsters previously immunized with low-LPS bacterin vaccines or from serum of patients diagnosed with leptospirosis. Immuno properties of seven peptides from LruC protein were evaluated in silico and by Dot Blot assay, and validate by ELISA. Preliminary results pointed one promising peptide that was recognized by the sera. In conclusion, the immunoinformatic approach helps the search and screening of peptides, while the Dot Blot assay, a simple and effective tool, helps to test and validate them. Thus, these prospective techniques together were validated to identify and validate potential peptides for further investigation as peptide-based vaccines or diagnostic methods.
Journal of Medical Microbiology, 2010
Leptospirosis is a worldwide zoonosis caused by pathogenic Leptospira. The whole-genome sequence of Leptospira interrogans serovar Copenhageni together with bioinformatic tools allow us to search for novel antigen candidates suitable for improved vaccines against leptospirosis. This study focused on three genes encoding conserved hypothetical proteins predicted to be exported to the outer membrane. The genes were amplified by PCR from six predominant pathogenic serovars in Brazil. The genes were cloned and expressed in Escherichia coli strain BL21-SI using the expression vector pDEST17. The recombinant proteins tagged with N-terminal 6¾His were purified by metal-charged chromatography. The proteins were recognized by antibodies present in sera from hamsters that were experimentally infected. Immunization of hamsters followed by challenge with a lethal dose of a virulent strain of Leptospira showed that the recombinant protein rLIC12730 afforded statistically significant protection to animals (44 %), followed by rLIC10494 (40 %) and rLIC12922 (30 %). Immunization with these proteins produced an increase in antibody titres during subsequent boosters, suggesting the involvement of a T-helper 2 response. Although more studies are needed, these data suggest that rLIC12730 and rLIC10494 are promising candidates for a multivalent vaccine for the prevention of leptospirosis.
Leptospirosis is a worldwide zoonosis caused by pathogenic Leptospira. The whole-genome sequence of L. in-terrogans serovar Copenhageni together with bioinformatics tools represent a great opportunity to search for novel anti-gen candidates that could be used as subunit vaccine against leptospirosis. We focused on six genes encoding for con-served hypothetical proteins predicted to be exported to the outer membrane. The genes were amplified by PCR from Lep-tospira interrogans genomic DNA and were cloned and expressed in Escherichia coli. The recombinant proteins tagged with N-terminal hexahistidine were purified by metal-charged chromatography. The immunization of hamsters followed by challenge with lethal dose of virulent strain of Leptospira showed that the recombinant proteins Lsa21, Lsa66 and rLIC11030 elicited partial protection to animals. These proteins could be used combined or in a mixture with novel adju-vants in order to improve their effectiveness.
Clinical and Vaccine Immunology, 2013
Toward developing an effective vaccine capable of conferring heterologous protection, the putative lipoprotein LemA, which presents an M3 epitope similar to that of Listeria, was evaluated as a vaccine candidate in the hamster model of leptospirosis. LemA is conserved (>70% pairwise identity) among the pathogenic Leptospira spp., indicating its potential in stimulating a cross-protective immune response. Using different vaccination strategies, including prime-boost, DNA vaccine, and a subunit preparation, recombinant LemA conferred different levels of protection in hamsters. Significant protection against mortality was observed for the prime-boost and the DNA vaccine strategies, which showed 87.5% (P < 0.01) and 62.5% (P < 0.05) efficacy, respectively. Although the subunit vaccine preparation protected 50.0% of immunized hamsters, the level of protection was not significant. None of the hamsters in the control groups survived challenge with a virulent strain of Leptospira interrogans serogroup Icterohaemorrhagiae. Characterization of the immune response found that the strongest antibody response was stimulated by the subunit vaccine preparation, followed by the prime-boost strategy. The DNA vaccine failed to elicit an antibody response in immunized hamsters.
Subunit Approach to Evaluation of the Immune Protective Potential of Leptospiral Antigens
Clinical and Vaccine Immunology, 2011
Leptospirosis is the most widespread zoonosis in the world. Current vaccines are based on whole-cell preparations that cause severe side effects and do not induce satisfactory immunity. In light of the leptospiral genome sequences recently made available, several studies aimed at identification of protective recombinant immunogens have been performed; however, few such immunogens have been identified. The aim of this study was to evaluate 27 recombinant antigens to determine their potential to induce an immune response protective against leptospirosis in the hamster model. Experiments were conducted with groups of female hamsters immunized with individual antigen preparations. Hamsters were then challenged with a lethal dose of Leptospira interrogans. Thirteen antigens induced protective immune responses; however, only recombinant proteins LIC10325 and LIC13059 induced significant protection against mortality. These results have important implications for the development of an efficacious recombinant subunit vaccine against leptospirosis.
Journal of Vaccines & Vaccination, 2011
Leptospirosis is a re-emergent zoonosis characterized by an acute febrile and systemic illness in humans caused by pathogenic spirochetes belonging to the genus Lepstospira. This disease has global distribution, and it is more frequent in tropical and subtropical areas. The complete genomic sequence of Leptospira species offered the possibility to identify potential vaccine candidates for leptospirosis, since environmental control measures are difficult to implement and there is not an ideal vaccine available for human use. Secreted and surface exposed molecules are potential targets for inducing protective immune response in the host. Although we selected six predicted sequences coding for putative outer membrane proteins with unknown function to be analyzed as vaccinal candidates against leptospirosis and for biological characterization, only the lic13435 gene was expressed and purified. The lic13435 gene is specific for pathogenic leptospires suggesting a possible virulence and/or pathogenicity associated function. The recombinant protein was purified and tested as vaccine candidate against leptospirosis. The immunization with the recombinant protein was able to produce a significant immune response in hamsters. Nevertheless, the animals were not protected against leptospirosis.
Clinical and Vaccine Immunology, 2013
The leptospiral immunoglobulin-like (Lig) proteins LigA and LigB possess immunoglobulin-like domains with 90-amino-acid repeats and are adhesion molecules involved in pathogenicity. They are conserved in pathogenic Leptospira spp. and thus are of interest for use as serodiagnostic antigens and in recombinant vaccine formulations. The N-terminal amino acid sequences of the LigA and LigB proteins are identical, but the C-terminal sequences vary. In this study, we evaluated the protective potential of five truncated forms of LigA and LigB proteins from Leptospira interrogans serovar Canicola as DNA vaccines using the pTARGET mammalian expression vector. Hamsters immunized with the DNA vaccines were subjected to a heterologous challenge with L. interrogans serovar Copenhageni strain Spool via the intraperitoneal route. Immunization with a DNA vaccine encoding LigBrep resulted in the survival of 5/8 (62.5%) hamsters against lethal infection (P < 0.05). None of the control hamsters or animals immunized with the other vaccine preparations survived. The vaccine induced an IgG antibody response and, additionally, conferred sterilizing immunity in 80% of the surviving animals. Our results indicate that the LigBrep DNA vaccine is a promising candidate for inclusion in a protective leptospiral vaccine.
Vaccine, 2010
Leptospira is an important infectious Gram-negative bacterium causing Leptospirosis in mammals. Outer membrane proteins (OMPs) are key molecules in the interface between the cell and its environment. A group of putative leptospiral outer membrane proteins with an OmpA-like domain, comprising Lp0056, Lp0222, Lp3615, Lp3685, Lp4337 and Lbp328, were identified by bioinformatic methods and expressed as GST-tag fusion proteins. All these recombinant proteins were screened for immune-protective potential in hamsters challenged with Leptospira interrogans serovar Pomona. Out of these six proteins, three fusion proteins including Lp4337, Lp3685 and Lp0222 were found to be protective on the basis of survival. The immune response generated against these three recombinant proteins was evaluated on the basis of antibody production, lymphocyte proliferation and cytokine profiles in response to recall antigens whereas protective efficacy was assessed on the basis of survival and histopathological lesions in the vital organs (kidney, liver, and lung). Our results show that all three recombinant proteins generated strong immune responses, enhanced survival and reduced the severity of histopathological lesions. Of the proteins studied, Lp4337 generated the strongest immune response and was able to impart maximum protection (75%), followed by Lp3685 (58%), whereas Lp0222 generated lowest immune response correlating to protection (42%) against lethal infection of leptospira in the immunized animals. In contrast, control animals injected with PBS demonstrated low survival and had significant lesions. All these results clearly indicate that these three OmpA-like proteins may serve as novel vaccine candidates for leptospirosis.
Frontiers in Cellular and Infection Microbiology
Leptospirosis is a neglected disease of man and animals that affects nearly half a million people annually and causes considerable economic losses. Current human vaccines are inactivated whole-cell preparations (bacterins) of Leptospira spp. that provide strong homologous protection yet fail to induce a cross-protective immune response. Yearly boosters are required, and serious side-effects are frequently reported so the vaccine is licensed for use in humans in only a handful of countries. Novel universal vaccines require identification of conserved surface-exposed epitopes of leptospiral antigens. Outer membrane β-barrel proteins (βb-OMPs) meet these requirements and have been successfully used as vaccines for other diseases. We report the evaluation of 22 constructs containing protein fragments from 33 leptospiral βb-OMPs, previously identified by reverse and structural vaccinology and cell-surface immunoprecipitation. Three-dimensional structures for each leptospiral βb-OMP were ...
Antigenic potential of a recombinant polyvalent DNA vaccine against pathogenic leptospiral infection
Antigenic potential of a recombinant polyvalent DNA vaccine against pathogenic leptospiral infection, 2018
Leptospirosis is a serious epidemic disease caused by pathogenic Leptospira species. The disease is endemic in most tropical and sub-tropical regions of the world. Currently, there is no effective polyvalent vaccine for prevention against most of the circulating serovars. Moreover, development of an efficient leptospiral vaccine capable of stimulating cross-protective immune responses against a wide range of serovars remains a daunting challenge. This, in part, is associated with the extensive diversity and variation of leptospiral serovars from region to region. In this study, a multi-epitope DNA vaccine encoding highly immunogenic epitopes from LipL32 and LipL41 was designed using in-silico approach. The DNA encoding antigenic epitopes was constructed from conserved pathogenic Leptospira genes (LipL32 and LipL41). Immunization of golden Syrian hamsters with the multi-epitope chimeric DNA vaccine resulted in the production of both agglutinating and neutralizing antibodies as evidence by MAT and in-vitro growth inhibition tests respectively. The antibodies produced reacted against eight different serovars and significantly reduced renal colonization following in vivo challenge. The vaccine was also able to significantly reduce renal colonization which is a very important factor responsible for persistence of leptospires among susceptible and reservoir animal hosts. In conclusion, the leptospiral multi-epitope chimeric DNA vaccine can serve as a potentially effective and safe vaccine against infection with different pathogenic leptospiral serovars