Immunization against Leishmania major Infection Using LACK- and IL-12-Expressing Lactococcus lactis Induces Delay in Footpad Swelling

Abstract

Background Leishmania is a mammalian parasite affecting over 12 million individuals worldwide. Current treatments are expensive, cause severe side effects, and emerging drug resistance has been reported. Vaccination is the most cost-effective means to control infectious disease but currently there is no vaccine available against Leishmaniasis. Lactococcus lactis is a non-pathogenic, non-colonizing Gram-positive lactic acid bacterium commonly used in the dairy industry. Recently, L. lactis was used to express biologically active molecules including vaccine antigens and cytokines.Methodology/Principal findingsWe report the generation of L. lactis strains expressing the protective Leishmania antigen, LACK, in the cytoplasm, secreted or anchored to the bacterial cell wall. L. lactis was also engineered to secrete biologically active single chain mouse IL-12. Subcutaneous immunization with live L. lactis expressing LACK anchored to the cell wall and L. lactis secreting IL-12 significantly delayed footpad swelling in Leishmania major infected BALB/c mice. The delay in footpad swelling correlated with a significant reduction of parasite burden in immunized animals compared to control groups. Immunization with these two L. lactis strains induced antigen-specific multifunctional TH1 CD4+ and CD8+ T cells and a systemic LACK-specific TH1 immune response. Further, protection in immunized animals correlated with a Leishmania-specific TH1 immune response post-challenge. L. lactis secreting mouse IL-12 was essential for directing immune responses to LACK towards a protective TH1 response.Conclusions/SignificanceThis report demonstrates the use of L. lactis as a live vaccine against L. major infection in BALB/c mice. The strains generated in this study provide the basis for the development of an inexpensive and safe vaccine against the human parasite Leishmania.

Display of recombinant proteins at the surface of lactic acid bacteria: strategies and applications

“…After oral administration, only the strain secreting the antigen and co-expressing IL-12 induced protection against the parasite [105]. In another study these same authors showed that subcutaneous co-administration of L. lactis secreting IL-12 with either a strain secreting LACK or a strain with a cell wall anchored (CWA)-LACK led to induction of protective immunity [106]. The lack of effectiveness of the CWA-LACK strain in oral immunization could be due to the fact that the expression strain was an alr mutant, which could lead to impaired cell-wall strength and, possibly, antigen anchoring.…”

Mucosal targeting of therapeutic molecules using genetically modified lactic acid bacteria: an update

“…Subcutaneous immunization with L. lactis expressing the cell wall–anchored form of A2 induced the higher levels of antigen‐specific serum antibodies, while mice immunized with L. lactis producing the cytoplasmic form of A2 demonstrated the highest reduction in liver parasitemia after visceral L. donovani challenge. Later, the same group reported the construction of different lactococci strains expressing one of the best‐studied Leishmania major antigens, the Leishmania homologue of activated C kinase (LACK), in the cytoplasmic, secreted, or cell wall–anchored forms, and a strain secreting biologically active mouse IL‐12 (Hugentobler et al ., , b). Subcutaneous co‐immunization with live L. lactis strains expressing the cell wall–anchored form of LACK and secreting IL‐12 significantly delayed footpad swelling in L. major ‐infected mice.…”

“…Furthermore, immunization with these two strains induced antigen‐specific multifunctional TH1 CD4 + and CD8 + T cells and a systemic LACK‐specific TH1 immune response. This same group then evaluated the effect against L. major infection in mice after oral immunization with recombinant L. lactis strains deficient in alanine racemase (alr‐), an enzyme that participates in cell wall synthesis (Grangette et al ., ), expressing LACK antigen in the cytoplasmic, secreted, or cell wall–anchored forms alone or in combination with a L. lactis strain secreting mouse IL‐12 (Hugentobler et al ., , b). They showed that oral immunization using live lactococci secreting both LACK and IL‐12 was the only treatment that partially protected the mice against subsequent L. major challenge.…”

Cutaneous Leishmaniasis: Update on Vaccine Development

“…85,86 Consistent with this concept, inclusion of IL-12 as part of a DNA vaccine cocktail improved protection against L major challenge. [87][88][89] However, administration of IL-12 in humans is toxic; thus, this strategy is not suitable for human vaccination. 90,91 Several studies have demonstrated that complete clearance of the parasite by a T H 1 immune response, which is desirable for the safety of a patient, is, however, not sufficient in mediating long-term immunity.…”