Concurrent colonization of rodent kidneys with multiple species and serogroups of pathogenic Leptospira

Authors: HAMOND, CAMILA - US Department Of Agriculture (USDA); LECOUNT, KAREN - US Department Of Agriculture (USDA); BROWNE, A. SPRINGER - US Department Of Agriculture (USDA); ANDERSON, TAMMY - US Department Of Agriculture (USDA); STUBER, TODD - US Department Of Agriculture (USDA); HICKS, JESSICA - US Department Of Agriculture (USDA); CAMP, PATRICK - US Department Of Agriculture (USDA); FERNANDES, LUIS - US Department Of Agriculture (USDA); VAN DER LINDEN, HANS - Amsterdam University College; GORIS, MARGA - Amsterdam University College; Bayles, Darrell; SCHLATER, LINDA - US Department Of Agriculture (USDA); Nally, Jarlath

Submitted to: Applied and Environmental Microbiology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 8/25/2023
Publication Date: 10/11/2023
Citation: Hamond, C., LeCount, K., Browne, A., Anderson, T., Stuber, T., Hicks, J., Camp, P., Fernandes, L.G., Van Der Linden, H., Goris, M.G., Bayles, D.O., Schlater, L.K., Nally, J.E. 2023. Concurrent colonization of rodent kidneys with multiple species and serogroups of pathogenic Leptospira. Applied and Environmental Microbiology. 89(10). https://doi.org/10.1128/aem.01204-23.
DOI: https://doi.org/10.1128/aem.01204-23

Interpretive Summary: Pathogenic Leptospira, the causative agent of leptospirosis, can infect humans, and domestic and wildlife animals. The disease is typically transmitted via the urine of chronically infected, asymptomatic animals. More than 38 species of pathogenic Leptospira have been identified to date comprising hundreds of serovars. In order to design effective animal vaccines and diagnostics, it is essential to be able to know the species and serovars of Leptospira circulating in domestic and wildlife animals. This is accomplished using a range of assays, including molecular tests and culture of Leptospira. Culture is definitive and provides the added benefit of having isolates that can be completely characterized for research purposes. However, culture is inherently difficult and not routinely performed. Our previous work with rats trapped on the U.S. Virgin Islands indicated that several were infected with more than one species and serovar of pathogenic Leptospira and were thus shedding multiple species and serovars at the same time. To prove this definitively, novel culture strategies were developed to isolate individual colonies from infected rat samples. Characterization of these individual isolates confirm that rodents can be colonized with more than one species/serogroup of pathogenic Leptospira concurrently. The identification and characterization of multiple species/serogroups of Leptospira from individual reservoir hosts of infection is essential to understand the epidemiology of leptospirosis and highlights their role in the transmission of disease to both human and domestic animal populations.

Technical Abstract: Rodents are important reservoir hosts of pathogenic leptospires in the U.S. Virgin Islands. Our previous work using PCR, culture, and whole genome sequencing (WGS) determined that the kidneys of trapped rodents were colonized with L. borgpetersenii serogroup Ballum (n=48) and/or L. kirschneri serogroup Icterohaemorrhagiae (n=3). In addition, kidneys from nine rodents appeared to be colonized with a mixed population comprising more than one species/serogroup of pathogenic leptospires. The aim of this study was to validate colonization with multiple species/serogroups by characterizing clonal isolates derived from cultures of pathogenic leptospires containing mixed species. Cultures of presumptive mixed species (designated LR1, LR5, LR37, LR57, LR60, LR61, LR68, LR70 and LR72) were propagated in different media including HAN liquid media incubated at both 29' and 37', and T80/40/LH incubated at 29'. Polyclonal reference antisera specific for serogroup Ballum and Icterohaemorrhagiae was used to enrich for different serogroups followed by subculture on agar plates. Individual colonies were then selected and propagated in liquid media for genotyping and serotyping. From nine cultures of mixed species/serogroups, at least one clonal isolate was separated from five of the mixed cultures: L. borgpetersenii serogroup Ballum in LR1, LR5 and LR37, and L. kirschneri serogroup Icterohaemorrhagiae in LR60 and LR72. In four of the cultures with mixed species (LR57, LR61, LR68 and LR70), clonal isolates of both L. borgpetersenii serogroup Ballum and L. kirschneri serogroup Icterohaemorrhagiae were recovered. Our results definitively establish that rodents can be colonized with more than one species/serogroup of pathogenic Leptospira concurrently. The identification and characterization of multiple species/serogroups of Leptospira from individual reservoir hosts of infection is essential to understand the epidemiology of leptospirosis and highlights their role in the transmission of disease to both human and domestic animal populations.