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ARS Home » Midwest Area » Ames, Iowa » National Animal Disease Center » Infectious Bacterial Diseases Research » Research » Publications at this Location » Publication #420023

Research Project: Intervention Strategies for Spirochete Diseases

Location: Infectious Bacterial Diseases Research

Title: Investigations into the growth and formation of biofilm by Leptospira biflexa at temperatures encountered during infection

Author
item DOS SANTOS RIBEIRO, PRISCYLA - University Medical Center Amsterdam
item Stasko, Judith
item Shircliff, Adrienne
item FERNANDES, LUIS - Oak Ridge Institute For Science And Education (ORISE)
item Putz, Ellie
item ANDREASEN, CLAIRE - Iowa State University
item AZEVEDO, VASCO - Federal University Of Bahia (UFBA)
item RISTOW, PAULA - University Medical Center Amsterdam
item Nally, Jarlath

Submitted to: Biofilms
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 12/9/2024
Publication Date: 6/20/2025
Citation: Dos Santos Ribeiro, P., Stasko, J.A., Shircliff, A.L., Fernandes, L.G., Putz, E.J., Andreasen, C., Azevedo, V., Ristow, P., Nally, J.E. 2025. Investigations into the growth and formation of biofilm by Leptospira biflexa at temperatures encountered during infection. Biofilms. https://doi.org/10.1016/j.bioflm.2024.100243.
DOI: https://doi.org/10.1016/j.bioflm.2024.100243

Interpretive Summary: Leptospirosis is a neglected zoonotic disease of global significance cause by atypical and unusual bacteria that belong to the genus Leptospira. The transmission of leptospirosis involves a complex epidemiological cycle whereby leptospires are shed from persistently infected reservoir hosts into the environment where they can survive and persist to maintain transmission to new hosts. Despite the sensitivity of Leptospira spp. to harsh conditions encountered in changes to salinity, osmolarity, and pH, these bacteria are widespread in bodies of water, sewage, and soil, and endure adverse environmental circumstances. One mechanism for bacteria to survive unfavorable environments is to form biofilms since they can provide resistance to antibiotics, disinfectants, and the host immune system. Leptospires form biofilms in vitro, in situ in rice fields and unsanitary urban areas, and in vivo while colonizing rodent kidneys. In this study, the growth and formation of biofilms by a model leptospire was investigated not just at 29°C, but at 37°C/5% CO2, a temperature similar to that encountered during host infection. Planktonic free-living leptospires grow in HAN media at both 29°C and 37°C/5% CO2, but cells grown at 37°C/5% CO2 are longer (18.52 µm +/- 3.39) compared to those at 29°C (13.93 µm +/- 2.84). Biofilms formed at 37°C/5% CO2 had more biomass compared to 29°C, protein content within the biofilm matrix was more prominent than double-stranded DNA, and biofilms featured a distinct protein layer. Additionally, the model enabled effective protein extraction for proteomic comparison across different biofilm phenotypes. Results highlight an important role for proteins in biofilm matrix structure by leptospires and the identification of their specific protein components holds promise for strategies to mitigate biofilm formation.

Technical Abstract: The genus Leptospira comprises unique atypical spirochete bacteria that includes the etiological agent of leptospirosis, a globally important zoonosis. Biofilms are microecosystems composed of microorganisms embedded in a self-produced matrix that offers protection against hostile factors. Leptospires form biofilms in vitro, in situ in rice fields and unsanitary urban areas, and in vivo while colonizing rodent kidneys. The complex three-dimensional biofilm matrix includes secreted polymeric substances such as proteins, extracellular DNA (eDNA), and saccharides. The genus Leptospira comprises pathogenic and saprophytic species with the saprophytic L. biflexa being commonly used as a model organism for the genus. In this study, the growth and formation of biofilms by L. biflexa was investigated not just at 29°C, but at 37°C/5% CO2, a temperature similar to that encountered during host infection. Planktonic free-living L. biflexa grow in HAN media at both 29°C and 37°C/5% CO2, but cells grown at 37°C/5% CO2 are longer (18.52 µm +/- 3.39) compared to those at 29°C (13.93 µm +/- 2.84). Biofilms formed at 37°C/5% CO2 had more biomass compared to 29°C, as determined by crystal violet staining. Confocal microscopy determined that the protein content within the biofilm matrix was more prominent than double-stranded DNA, and featured a distinct layer attached to the solid substrate. Additionally, the model enabled effective protein extraction for proteomic comparison across different biofilm phenotypes. Results highlight an important role for proteins in biofilm matrix structure by leptospires and the identification of their specific protein components holds promise for strategies to mitigate biofilm formation.