Biofilme de cepas autóctones de leptospira interrogans: investigação na infecção canina aguda e efeito disruptivo in vitro por antibióticos e ácido p-cumárico

Abstract

Leptospirosis is a zoonotic infectious disease caused by bacteria of the genus Leptospira. Dogs are highly susceptible, often presenting renal impairment with clinical and epidemiological significance. Leptospira can form biofilms in the environment, in chronic hosts and in vitro, however, this feature has not been investigated in acute infections. Biofilm formation represents a One Health threat, as it is associated with therapeutic failures and the persistence of pathogenic bacteria in the human-animal-environment interface. This study aimed to investigate biofilms in the renal parenchyma of dogs naturally infected with acute leptospirosis. Anti-lipL32 immunohistochemistry and Alcian Bleu and periodic acid-Schiff staining were applied to identify Leptospira and detect components of the exopolysaccharide matrix. Additionally, four strains (C20, C29, C51 and C82), isolated from these dogs, were subjected to antimicrobial susceptibility assays in both planktonic and biofilm (7 and 21 days) forms, using doxycycline, ciprofloxacin and p-coumaric acid (p-CA) at concentrations ranging from 0,02μg/mL to 1,600μg/mL. After antimicrobial exposure, the disruption of biofilm architecture was evaluated by scanning electron microscopy. No histological staining of the extracellular matrix was detected; however, incipient biofilms, in the form of L. interrogans aggregates, were observed. Biofilm maturity negatively affected antimicrobial efficacy, with lower susceptibility observed in biofilms as early as 7 days compared to planktonic forms. In both growth conditions, p-CA exhibited lower MICs than the tested antibiotics. Despite variations in biofilm architecture after disruption, intact bacterial cells were present in all treatments. These findings highlight the need to investigate the role of bacterial aggregates in in vivo biofilm progression, the specific mechanisms of p-CA action in L. interrogans biofilms, and more effective therapeutic combinations than conventional treatments.