Evaluation of different ways of presenting LipL32 to the immune system with the aim of developing a recombinant vaccine against leptospirosis

Centro de Biotecnologia, Universidade Federal de Pelotas, Pelotas, Brazil.
Canadian Journal of Microbiology (Impact Factor: 1.22). 05/2007; 53(4):472-9. DOI: 10.1139/w06-138
Source: PubMed


Leptospirosis, caused by bacteria of the genus Leptospira, is a direct zoonosis with wide geographical distribution. The implications in terms of public health and the economical losses caused by leptospirosis justify the use of a vaccine against Leptospira in human or animal populations at risk. In this study, we used the external membrane protein LipL32 as a model antigen, as it is highly immunogenic. The LipL32 coding sequence was cloned into several expression vectors: (i) pTarget, to create a DNA vaccine; (ii) pUS973, pUS974, and pUS977 for expression in BCG (rBCG); and (iii) pAE, to express the recombinant protein in Escherichia coli, for a subunit vaccine. Mice were immunized with the various constructs, and the immune response was evaluated. The highest humoral immune response was elicited by the subunit vaccine (rLipL32). However, with rBCG, the titer was still rising at the end of the experiment. The serum of vaccinated animals was able to recognize LipL32 on the membrane of the Leptospira, detected by indirect immunofluorescence. A monoclonal antibody anti-LipL32 was shown to inhibit the growth of Leptospira in vitro, indicating potential protection induced by the LipL32 antigen.

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Available from: Odir Dellagostin, Mar 07, 2014
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    • "Q72PR0 Putative glutamine synthetase protein Catalytic activity; ligase activity [22] Q72Q79 Fructose-bisphosphate aldolase Lyase activity [21] Q72R58 Flagellin protein Structural molecule activity [21] Q 7 2 R U 5 L i p L 4 5 U n k n o w n [21] Q72S54 Flagellin protein Structural molecule activity [21] Q72S55 Flagellin protein Structural molecule activity [21] Q 7 2 S G 6 ATP-dependent Clp protease, proteolytic subunit Peptidase activity; nucleotide binding; hydrolase activity [23] Q 7 2 S M 7 L i p L 3 2 U n k n o w n [21] Q72T03 Peroxiredoxin Antioxidant activity; oxidoreductase activity [22] Q 7 2 T 2 7 P u t a t i v e c i t r a t e l y a s e Catalytic activity; lyase activity; ion binding [21] Q72U13 Elongation factor Ts Nucleic acid binding [21] Q72V20 Hypothetical protein LIC10483 Unknown [21] Q 7 2 V D 7 Electron transport flavoprotein beta subunit Electron carrier activity [21] Q72WD5 DNA polymerase III beta subunit Nucleic acid binding; transferase activity; hydrolase activity Total conserved proteins 16 (Appendices 3–5 in the Supplementary Material), sixteen of which have been previously reported in the literature as immunogenic (Table 2) through immune studies in the hamster [20] and mice models [23] and immunoblotting with serum from infected humans [22] and mice [21]. Further work is required to determine if any of the identified conserved proteins could be used as viable targets for therapeutic drugs, antimicrobials, and/or vaccines; however this does suggest that proteomic comparison of serovars could also be used as an effective screening tool; further refinement of the conserved proteome presented herein is suggested as the proteomes of additional serovars are published. "
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    • "Presently, molecular techniques have been used for leptospirosis vaccine development. These vaccines were developed from component proteins of the leptospiral cell such as lipopolysaccharides (LPSs), lipoproteins, outer-membrane proteins (OMPs) and virulence factors (Gebriel et al., 2006; Haake et al., 2000; Ren et al., 2003; Seixas et al., 2007). Some vaccines were successful at the laboratory level, but have not been developed for clinical applications . "
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    • "In addition, the full-length LipL32 protein was also expressed as a secreted protein. Previously we reported the expression of recombinant LigANI in E. coli with a yield of 6-10 mg/L [13], while recombinant LipL32 was expressed at 40 mg/L [29]. In this study we report that large-scale expression in P. pastoris resulted in yields of over 250 mg/L for both rLigANI and rLipL32, without the need for subsequent solubilisation and/or re-folding steps. "
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