Location: Infectious Bacterial Diseases Research
2024 Annual Report
Objectives
Objective 1: Characterize leptospires circulating in dairy cows to advance the development of efficacious intervention strategies.
Sub-objective 1.1: Detect and classify leptospires circulating in dairy cows.
Sub-objective 1.2: Characterization of recent isolates of pathogenic leptospires.
Sub-objective 1.3: Enhanced bacterin intervention strategies.
Objective 2: Characterize the pathogenesis of DD for the development of more effective intervention strategies.
Sub-objective 2.1: Characterization of Treponema and other pathogens in DD.
Sub-objective 2.2: Development of host immune response to antigens from DD lesions.
Sub-objective 2.3: Development of effective intervention strategies for DD.
Approach
Leptospirosis and digital dermatitis (DD) are two different diseases caused by two separate groups of bacteria in the Phylum Spirochaetes that have substantial impact on livestock production. Multiple serovars of Leptospira interrogans are the leading cause of acute lethal leptospirosis in humans and domestic animals while serovar Hardjo of L. borgpetersenii is the leading cause of bovine disease, causing reproductive failure and persistent shedding via urine to maintain disease transmission. Current bovine bacterin vaccines are limited in efficacy. To improve on this, Objective 1 will characterize leptospires circulating in dairy cows to advance the development of efficacious intervention strategies. It is first necessary to identify those species and serovars of Leptospira currently circulating in animal populations, as described in subobjective 1.1. Given the importance of L. borgpetersenii to animal disease, subobjective 1.2. will include comparative genomics and proteomics of multiple serovars within Leptospira borgpetersenii to identify conserved pathogenic mechanisms of infection and candidate vaccinogens as potential recombinant subunit vaccines. We hypothesize that efficacy of bacterins can be further improved to provide heterologous protection using growth media that more closely emulates that encountered during host infection, as proposed in subobjective 1.3. Unlike leptospirosis, DD is an infectious polymicrobial skin infection in which Treponema species are found at the invading edge of the lesions. Causing painful ulcerative proliferative and necrotizing lesions on the skin at or near the hooves, DD is a significant cause of lameness in both dairy and feedlot cattle. Aside from lameness being a significant animal welfare issues, DD leads to decreased production, higher treatment costs and premature culling. Objective 2 will characterize the pathogenesis of DD for the development of more effective intervention strategies. Since the etiology of DD is not fully characterized, subobjective 2.1 will use bacterial 16S rRNA gene sequencing to characterize bacterial community of early DD lesions as induced in a sheep model, compare the sheep model to naturally infected bovine DD lesions in order to determine a core consortium of Treponema and other pathogens present. After obtaining isolates representing this core consortium, a defined mixture of Treponema species and other bacterial pathogens will be used to induce lesions in the sheep model. Subobjective 2.2. will then characterize the bovine innate immune responses to Treponema, and how that may affect lesion healing. Since current mitigation strategies use heavy metal, formalin-containing footbaths or topical antibiotics, subobjective 2.3. will evaluate novel alternative antimicrobial compounds for topical treatment of DD lesions. Understanding pathogenic mechanisms used by Spirochetes, specific species causing disease and host-pathogen interactions, are critical for the development of efficacious diagnostics, vaccines, and therapeutics for control of infection in domestic livestock.
Progress Report
To address Objective 1, studies were conducted to identify and characterize Leptospira circulating in livestock and the environment. Leptospira were cultured from Iowa water sources, dairy cows in California, bulls in Colorado, bulls in Texas and horses in Kentucky. Samples included urine and semen and viable leptospira were detected in frozen bovine semen. As vaccine protection is generally limited to that leptospira serovar, understanding circulating serotypes and strains is critical for successful disease intervention strategies because it is likely that current bacterin may not contain some key serotypes and strains. Novel approaches have also been developed to genetically manipulate pathogenic Leptospira to identify virulence factors that may contribute to development of a cross protective vaccine.
To address Objective 2, long-acting vaccine delivery platforms using recombinant proteins from Haemonchus contortus were evaluated in sheep. This work demonstrated the utility of single dose vaccines in inducing sustained immunity in livestock as a model for characterizing responses of sheep to subunit vaccines against other pathogens, like Treponema. The ability of novel biocides to eliminate bacterial pathogens was obtained, specifically demonstrating hydrazone containing biocides are bactericidal on mastitis pathogens and as surface treatments for eliminating Treponemes that cause digital dermatitis. These biocides offer producers environmentally safe alternatives to antibiotics. National distribution and prevalence of digital dermatitis and ovine foot-rot is being characterized using samples obtained in the 2024 NAHMS (National Animal Health Monitoring Service).
Accomplishments
1. New species of Leptospira in Iowa water sources. In a collaborative project with scientists at the National Veterinary Service Laboratory, ARS scientists in Ames, Iowa surveyed water sources in Iowa for the presence of Leptospira. Water samples were tested by culture and PCR for the presence of environmental and pathogenic leptospirosis species. Data demonstrated that 78% of samples were culture positive for Leptospira with 9.5% containing pathogenic Leptospira. Whole genome sequencing of isolates identified three novel nonpathogenic species and two novel pathogenic species. This study emphasizes the diversity of Leptospira circulating in Iowa water sources, facilitates development of improved diagnostics to detect Leptospira, and emphasizes the role of water in the transmission of leptospirosis to humans and livestock. This work will be of interest to livestock owners, regulatory and public health personnel, and researchers with interest in leptospirosis.
2. Horses are reservoir hosts of Leptospira. In collaborative studies with scientists at the University of Kentucky, ARS scientists in Ames, Iowa demonstrated that mares can be reservoir hosts of animal leptospirosis. Results demonstrated asymptomatic mares actively excrete different species and serovars of Leptospira. Isolates included Leptospira interrogans serovar Copenhageni and L. kirschneri serovar Rushan, an isolate not previously identified in the U.S. The lipopolysaccharide (LPS) of L. kirschneri serovar Rushan differs from the LPS of serovars included in commercial bacterin vaccines suggesting that current vaccines may not protect against this serovar. The unique structure of its LPS also suggests it may not be detected by current leptospirosis diagnostic assays used in the U.S. This work will be of interest to livestock owners, regulatory personnel, and researchers with interest in leptospirosis.
3. Proteomic profiles of L. borgpetersenii serovar Hardjo a primary cause of leptospirosis in cattle. Leptospirosis is a zoonotic disease that causes reproductive losses and infertility in cattle with L. borgpetersenii serovar Hardjo being the most prevalent isolate associated with infections in cattle. Pathogenic species of Leptospira are known to modify antigen expression when exposed to different environmental cues, including temperature, during host infection. ARS scientists in Ames, Iowa, characterized differences in expression of more than 3,000 proteins of two serovar Hardjo strains after incubation at two temperatures (29 and 37 C). Proteins expressed at high levels by serovar Hardjo at body temperature were identified.for consideration in developing improved bacterin vaccines. This work will be of interest to commercial vaccine producers, stakeholders, regulatory personnel, and researchers with interest in leptospirosis and leptospirosis vaccines.
Review Publications
Shiel, R.E., Nolan, C.M., Nally, J.E., Refsal, K.R., Mooney, C.T. 2021. Qualitative and semiquantitative assessment of thyroid hormone binding proteins in greyhounds and other dog breeds. Domestic Animal Endocrinology. 76:1-8. https://doi.org/10.1016/j.domaniend.2021.106623.
Putz, E.J., Fernandes, L.G., Bayles, D.O., Lippolis, J.D., Nally, J.E. 2022. Proteomic dataset comparing strains of Leptospira borgpetersenii serovar Hardjo cultured at different temperatures. Data in Brief. 45. https://doi.org/10.1016/j.dib.2022.108713.
Anderson, T., Hamond, C., Haluch, A., Toot, K., Nally, J.E., LeCount, K., Schlater, L.K. 2023. Animals exposed to Leptospira Serogroups not included in bacterins in the United States and Puerto Rico. Tropical Medicine and Infectious Disease. 8(3). https://doi.org/10.3390/tropicalmed8030183.
Putz, E.J., Fernandes, L.G., Sarlo Davila, K.M., Whitelegge, J., Lippolis, J.D., Nally, J.E. 2024. Proteomic profiles of Leptospira borgpetersenii serovar Hardjo strains JB197 and HB203 cultured at different temperatures. Journal of Proteomics. 295. https://doi.org/10.1016/j.jprot.2024.105106.
LeCount, K., Fox, K., Anderson, T., Bayles, D.O., Stuber, T., Hicks, J., Schlater, L.K., Nally, J.E. 2023. Isolation of Leptospira kirschneri serovar Grippotyphosa from a red panda (Ailurus fulgens) after antimicrobial therapy: case report. Frontiers in Veterinary Science. 9. https://doi.org/10.3389/fvets.2022.1064147.
Hamond, C., LeCount, K., Anderson, T., Putz, E.J., Stuber, T., Hicks, J., Camp, P., Van Der Linden, H., Bayles, D.O., Schlater, L.K., Nally, J.E. 2024. Isolation and characterization of saprophytic and pathogenic strains of Leptospira from water sources in the Midwestern United States. Frontiers in Water. 6. https://doi.org/10.3389/frwa.2024.1278088.
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.
Stone, N.E., McDonough, R.F., Hamond, C., LeCount, K., Busch, J.D., Dirsmith, K.L., Rivera-Garcia, S., Soltero, F., Arnold, L.M., Weiner, Z., Galloway, R.L., Schlater, L.K., Nally, J.E., Sahl, J.W., Wagner, D.M. 2023. DNA capture and enrichment: a culture-independent approach for characterizing the genomic diversity of pathogenic leptospira species. Microorganisms. 11(5). https://doi.org/10.3390/microorganisms11051282.
Hamond, C., Adam, E., Stone, N.E., LeCount, K., Anderson, T., Putz, E.J., Camp, P., Hicks, J., Stuber, T., Van Der Linden, H., Bayles, D.O., Sahl, J.W., Schlater, L.K., Wagner, D.M., Nally, J.E. 2024. Identification of equine mares as reservoir hosts for pathogenic species of Leptospira. Frontiers in Veterinary Science. 11. https://doi.org/10.3389/fvets.2024.1346713.
Hamond, C., Dirsmith, K.L., LeCount, K., Soltero, F.V., Rivera-Garcia, S., Camp, P., Anderson, T., Hicks, J.A., Galloway, R., Sutherland, G., Schafer, I.J., Goris, M.G., Van Der Linden, H., Stuber, T., Bayles, D.O., Schlater, L., Nally, J.E. 2022. Leptospira borgpetersenii serovar Hardjo and Leptospira santarosai serogroup Pyrogenes isolated from bovine dairy herds in Puerto Rico. Frontiers in Veterinary Science. 9. https://doi.org/10.3389/fvets.2022.1025282.
Canales, N., Bustamante, H., Wilson-Welder, J.H., Thomas, C., Ramirez, E., Salgado, M. 2022. First molecular confirmation of Treponema spp. in lesions consistent with digital dermatitis in Chilean dairy cattle. Pathogens. https://doi.org/10.3390/pathogens11050510.
Wilson-Welder, J.H., Han, S., Bayles, D., Alt, D.P., Kanipe, C.R., Garrison, K., Mansfield, K., Olsen, S.C. 2024. Correlation of lesion severity with bacterial changes in Treponeme-associated hoof disease from free-roaming wild elk (Cervus canadensis). Animal Microbiome. 6. Article 20. https://doi.org/10.1186/s42523-024-00304-9.
Lynn, L.E., Scholes, R.C., Kim, J., Wilson-Welder, J.H., Orts, W.J., Hart-Cooper, W.M. 2024. Antimicrobial, preservative, and hazard assessments from eight chemical classes. ACS Omega. 9(16):17869–17877. https://doi.org/10.1021/acsomega.3c08672.