The wide variety of pathogenic Leptospira serovars and the weak protection offered by the available vaccines encourage the search for protective immunogens against leptospirosis. We found that the secretin GspD of the type II secretion system (T2S) of Leptospira interrogans serovar Canicola was highly conserved amongst pathogenic serovars and was expressed in vivo during infection, as shown by immunohistochemistry. Convalescent sera of hamsters, dogs, and cows showed the presence of IgG antibodies, recognizing a recombinant version of this protein expressed in Escherichia coli (rGspDLC) in Western blot assays. In a pilot vaccination study, a group of eight hamsters was immunized on days zero and 14 with 50 µg of rGspDLC mixed with Freund’s incomplete adjuvant (FIA). On day 28 of the study, 1,000 LD50 (Lethal Dose 50%) of a virulent strain of Leptospira interrogans serovar Canicola (LOCaS46) were inoculated by an intraoral submucosal route (IOSM). Seventy-five percent protection against disease (p = 0.017573, Fisher’s exact test) and 50% protection against infection were observed in this group of vaccinated hamsters. In contrast, 85% of non-vaccinated hamsters died six to nine days after the challenge. These results suggest the potential usefulness of the T2S secretin GspD of Leptospira as a protective recombinant vaccine against leptospirosis.

The bacterial Quorum Sensing system refers to the recognition of signaling molecules called autoinducers produced by bacteria when a certain cell density is reached in the environment. Those cell-density-dependent autoinducers regulate and coordinate diverse functional processes, such as bioluminescence, biofilm production, sporulation, and even the expression of some virulence factors, among others. There is a wide variety of autoinducers, and for Gram-negative bacteria, the canonical autoinducers are the N-acyl-homoserine lactones (AI-1). Presently, the production of autoinducers in Leptospira has not been described; therefore, the objective of this study was to detect and identify autoinducers in this bacterial genus. We report here the expression of AI-1 in cultures ≥2.4 × 10⁸ of Leptospira meyeri. Ethyl acetate extracts of Leptospira culture supernatants were capable of activating the β-galactosidase system in the biosensor Agrobacterium tumefaciens strain NTL4. Partial identification of the leptospiral supernatant extracts was done by thin-layer chromatography (TLC), showing a similar retention factor to the synthetic standard N-Octanoyl-DL-homoserine lactone (C8-AHL) in the Leptospira supernatant extracts. In addition, infrared spectroscopy (IR) analysis showed peaks corresponding to the lactone and amide groups in both the C8-AHL standard and the Leptospira meyeri culture extracts. Moreover, High-Performance Liquid Chromatography–Mass Spectrometry (HPLC-MS/MS) confirmed the same retention time (10.7 ± 0.1 min) in both the Leptospira meyeri supernatant extracts and the C8-AHL standard. These results show that Leptospira meyeri synthesizes N-acyl homoserine lactone family autoinducers, particularly the N-Octanoyl-DL-homoserine lactone, and lay the groundwork for future research on Quorum Sensing systems in Leptospira.