Identification of Collagenase as a Critical Virulence Factor for Invasiveness and Transmission of Pathogenic Leptospira Species

Background. Leptospirosis is a global zoonotic disease. Transmission of Leptospira from animals to humans occurs through contact with water contaminated with leptospire-containing urine of infected animals. However, the molecular basis for the invasiveness of Leptospira and transmission of leptospirosis remains unknown. Methods. Activity of Leptospira interrogans strain Lai colA gene product (ColA) to hydrolyze different collagenic substrates was determined by spectrophotometry. Expression and secretion of ColA during infection were detected by reverse-transcription quantitative polymerase chain reaction and Western blot assay. The colA gene-deleted (△colA) and colA gene-complemented (C△colA) mutants were generated to determine the roles of ColA in transcytosis in vitro and virulence in hamsters. Results. Recombinant or native ColA hydrolyzed all the tested substrates in which type III collagen was the favorite substrate with 2.16 mg/mL Km and 35.6 h⁻¹ Kcat values. Coincubation of the spirochete with HUVEC or HEK293 cells directly caused the significant elevation of ColA expression and secretion. Compared with wild-type strain, △colA mutant displayed much-attenuated transcytosis through HEK293 and HUVEC monolayers, and less leptospires in blood, lung, liver, kidney and urine and 25-fold-decreased 50% lethal dose and milder histopathological injury in hamsters. Conclusions. The product of colA gene is a collagenase as a crucial virulence factor in the invasiveness and transmission of L. interrogans.