Equine Leptospirosis Research Articles

Placental abnormalities associated with Leptospira interrogans infection in naturally infected mares

The reproductive features of equine leptospirosis are often neglected. Equine genital leptospirosis is characterized as a silent chronic syndrome, and besides abortions, leads to placental abnormalities, stillbirths, and birth of weak foals. This study aimed to study the occurrence of placental abnormalities associated with Leptospira interrogans infection in naturally infected mares under field conditions. The studied herd had a high occurrence of placentitis and abortions. Ten pregnant mares, eight with placental abnormalities on ultrasonography and were selected. Serum and cervicovaginal mucus (CVM) samples were collected for serology and PCR, respectively. Positive samples in lipL32-PCR were submitted to the sequencing of the secY gene. In lipL32-PCR of CVM, five out of 10 (50%) mares were positive and all were characterized as Leptospira interrogans. Our results highlight the presence of placental abnormalities in the reproductive subclinical leptospirosis syndrome. We encourage field veterinarians to include leptospirosis testing in their reproductive management.

Identification of equine mares as reservoir hosts for pathogenic species of Leptospira.

Equine leptospirosis can result in abortion, stillbirth, neonatal death, placentitis, and uveitis. Horses can also act as subclinical reservoir hosts of infection, which are characterized as asymptomatic carriers that persistently excrete leptospires and transmit disease. In this study, PCR and culture were used to assess urinary shedding of pathogenic Leptospira from 37 asymptomatic mares. Three asymptomatic mares, designated as H2, H8, and H9, were PCR-positive for lipL32, a gene specific for pathogenic species of Leptospira. One asymptomatic mare, H9, was culture-positive, and the recovered isolate was classified as L. kirschneri serogroup Australis serovar Rushan. DNA capture and enrichment of Leptospira genomic DNA from PCR-positive, culture-negative samples determined that asymptomatic mare H8 was also shedding L. kirschneri serogroup Australis, whereas asymptomatic mare H2 was shedding L. interrogans serogroup Icterohaemorrhagiae. Sera from all asymptomatic mares were tested by the microscopic agglutination test (MAT) and 35 of 37 (94.6%) were seropositive with titers ranging from 1:100 to 1:3200. In contrast to asymptomatic mares, mare H44 presented with acute spontaneous abortion and a serum MAT titer of 1:102,400 to L. interrogans serogroup Pomona serovar Pomona. Comparison of L. kirschneri serogroup Australis strain H9 with that of L. interrogans serogroup Pomona strain H44 in the hamster model of leptospirosis corroborated differences in virulence of strains. Since lipopolysaccharide (LPS) is a protective antigen in bacterin vaccines, the LPS of strain H9 (associated with subclinical carriage) was compared with strain H44 (associated with spontaneous abortion). This revealed different LPS profiles and immunoreactivity with reference antisera. It is essential to know what species and serovars of Leptospira are circulating in equine populations to design efficacious vaccines and diagnostic tests. Our results demonstrate that horses in the US can act as reservoir hosts of leptospirosis and shed diverse pathogenic Leptospira species via urine. This report also details the detection of L. kirschneri serogroup Australis serovar Rushan, a species and serotype of Leptospira, not previously reported in the US.

Leptospirosis in horses: Sentinels for a neglected zoonosis? A systematic review.

Leptospirosis is considered a neglected tropical zoonosis in low-income countries due to surveillance system limitations and non-specificity of symptoms. Humans become infected through direct contact with carrier animals or indirectly through Leptospira-contaminated environments. Conventionally, equines have been considered an uncommon source of leptospirosis, but recent publications in Latin America suggest that their role in the maintenance and dispersion of the bacteria could be more relevant than expected, as horses are susceptible to a wide variety of zoonotic Leptospira spp. from domestic and wild animals with which they share the environment. A systematic review of the published literature was conducted to compile the available information on Leptospira spp. in Ecuador, with a special focus on equine leptospirosis, to better understand the epidemiology of the bacterium and identify possible knowledge gaps. A systematic review of the published literature was conducted in PubMed, SciELO and Web of Science databases to compile the available information on Leptospira spp. in Ecuador, with a special focus on equine leptospirosis, to better understand the epidemiology of the bacterium. We used a combination of the terms (Leptospira OR Leptospirosis) AND Ecuador, without restrictions on language or publication date. Our literature review reveals that published scientific information is very scarce. Eighteen full-text original scientific articles related to Leptospira or leptospirosis cases in Ecuador were included in the systematic review. Most of the studies reported data obtained from one of the four regions (Coast), and specifically from only one of the 24 Provinces of Ecuador (Manabí), which evidence a large information bias at the geographical level. Furthermore, only the studies focused on humans included clinical signs of leptospirosis and there is only one study that analyzes the presence of Leptospira spp. in water or soil as a risk factor for pathogen transmission. Finally, only one study investigated Leptospira in horses. Since sentinel species can provide useful data on infectious diseases when epidemiologic al information is lacking, and horses could be considered excellent sentinel species to reveal circulating serovars, we propose developing a nationwide surveillance system using horses. This cost-effective epidemiological survey method provides a baseline for implementing specific prevention and control programs in Ecuador and neighboring developing countries.

Seroprevalence of Equine Leptospirosis in the State of Goiás, Brazil.

Leptospirosis, caused by Leptospira, is a zoonotic disease that, in horses, is linked to abortions, uveitis, and sporadic occurrences of liver and kidney disease, often resulting in significant economic losses for farmers. Research on the prevalence of leptospirosis in horses in the central-west region of Brazil has been relatively scarce. Thus, the present study aimed to determine the prevalence of leptospirosis in equine herds in the state of Goiás (Central Brazil). Blood samples were collected from 894 equids at 294 randomly selected farms divided into three different strata according to their herd characteristics. The microscopic agglutination test for the detection of anti-Leptospira agglutinins was carried out and the results showed that among the 294 sampled farms, 213 (72.9%; CI 95% 71.7-78.9) had one or more animals positive for leptospirosis, and of the 894 horses sampled, 513 (61.6%; CI 95% 54.3-69.0) were seropositive for leptospirosis. Djasiman, Icterohaemorrhagiae, and Australis were the most prevalent serogroups. The results showed a high prevalence of seropositive animals and a widespread distribution of positive farms in the state of Goiás. Thus, environmental sanitation measures and health education to prevent and control equine leptospirosis in the state are required.

Serological Surveillance of Equine Leptospirosis in Croatia in the Period From 2012 to 2022: A Key Insight Into the Changing Epizootiology.

Leptospirosis is re-emerging zoonotic bacterial disease of global importance that affects domestic and wild animals and humans. Due to the public health importance, control of disease in Croatia is being implemented by monitoring the seroprevalence of equine leptospirosis and it is regulated by the law. In the period from 2012 to 2022, a total of 61,724 serum samples from apparently healthy horses were admitted to the Laboratory for leptospires, Faculty of Veterinary Medicine University of Zagreb. Serum samples were tested for Leptospira spp. antibodies using the microscopic agglutination test (MAT). Samples were considered seropositive with a cut-off titre 1:200 for Bratislava and 1:400 for other Leptospira spp. serovars. Out of 61,724 serum samples tested, 6,665 (10.80%) were found seropositive for at least one Leptospira serovar. In the studied period, seroprevalence varied between 5.00% and 15.94%. The highest seroprevalence was found for serovar Pomona (41.98%) and serovar Grippotyphosa (31.34%), followed by Sejroe (8.03%), Icterohaemorrhagiae (7.05%) and Bratislava (6.47%). Results indicated that horses in Croatia are particularly exposed to Leptospira spp. infections. The most prevalent presumed infective serovar was Pomona increasing each year. Investigated horses were randomly selected and healthy and most seropositive horses have anamnestic titre due to previous infection. This is the first study in Europe reporting such high seropositivity for the serovar Pomona in apparently healthy horses. According to the results of the present study, the question arises of the possible evolutionary adaptation of the pathogenic serovar Pomona as dominant for horses.

Equine leptospirosis: results of laboratory control in the Russian Federation

An analysis of the results of equine leptospirosis diagnostics by methods used in practice in state veterinary laboratories of the Russian Federation (bacteriological, serological (microagglutination reaction and enzyme-linked immunosorbent assay (ELISA)), molecular genetic (polymerase chain reaction method) for the period 2018-2020 is presented in the article. The veterinary laboratories of the Russian Federation received 225,582 horses blood serum samples for research on leptospirosis, for which research was carried out in the microagglutination reaction – 1,516,087, in ELISA – 1,039, 12,770 samples reacted positively. The analysis of the reported data for the last three years in the 4-vet form showed that the epizootic situation in equine leptospirosis in the Russian Federation remains difficult. The level of infection of horses with Leptospira averaged 5.7%. In 2020, the largest number of positive responders in microagglutination reaction were detected in 9 regions; among the studied animals, specific leptospirotic antibodies were found from 1.6% to 65.0% of cases. The dominant serogroups of leptospira in horses in the Russian Federation are Icterohaemorrhagiae and Grippotyphosa, up to 34% and 19.7%, respectively. Over the past three years, 1,907 urine samples from horses have been examined by dark-field microscopy; live leptospira have been found in only 1 case. To improve the epizootic situation in horses leptospirosis in Russia, it is necessary to increase the number of bacteriological and molecular biological studies for the timely detection of the causative agent of the disease – L. interrogans. In disadvantaged regions, it is necessary to use antileptospirosis vaccines in accordance with the specific etiological structure of animal leptospirosis, which is relevant for a specific territory.

Frecuencia de leptospirosis en equinos: revisión de literatura

Equine leptospirosis has been associated with abortions, renal failure and uveitis, which constitutes a health and economic problem. However, it is generally asymptomatic, which makes this disease of little interest in equines in comparison with other domestic and wild species. At present, the real epidemiological situation of leptospirosis in equines is unknown; just as the disease develops in these animals. A search of scientific literature was carried out; with the objective of knowing which are the serovars of genus leptospira reported in equines from 2010 to 2020. In the literature review, 21 articles were selected, in which it was identified that Leptospira Bratislava, is the most frequently reported (11.43%, IC95%: 6-19.1%) is in contact with equines, followed by Leptospira icterohaemorrhagiae (9.52%, IC95%: 4.6-16.8%).

Evaluation of new leptospiral antigens for the diagnosis of equine leptospirosis: An approach using pan-genomic analysis, reverse vaccinology and antigenic selection.

The current gold standard diagnostic test for leptospirosis is the microscopic agglutination test (MAT), which has many drawbacks; therefore, the development of a better and easier serological test for leptospirosis is needed. To apply reverse vaccinology (RV) and antigenic selection on the assortment of leptospiral targets and evaluate their potential for use as reagents for the diagnosis of equine leptospirosis. Cross-sectional study. The antigenic selection parameters were: proteins with antigenicity score ≥0.5 (VaxiJen), at least one B cell epitope and size between 10 and 275KDa. New leptospiral proteins were cloned, expressed and serologically screened against equine sera (n=128) on a single analysis and comparative combinations. Sensitivity (Se) and specificity (Sp), accuracy, positive predictive value (PPV) and negative predictive value (NPV) were calculated. A BLAST with nucleotide and protein sequences was used to identify the serovar or species specificity. This cross-sectional analysis had three main limitations: (a) The equine sera used in these tests were limited to sera submitted to the Animal Health Diagnosis Center and were only tested against seven serovars; (b) MAT results were considered being 'perfect', and the highest titre presented was considered being the infecting serovar, which may not hold true; (c) The strains used to represent the serovars and the limited number of different serovars and species included in the genetic analysis, which leads to the possibility that these proteins might be present in different species or serovars that perhaps would be seroprevalent in another geographic region. The new leptospiral antigens described in this research could increase the sensitivity and specificity of ELISA for detection of Leptospira exposure and the detection of leptospirosis in horses along with support from other clinical signs. Some of these new antigens might be used to improve the detection of infecting serovar.

Seroprevalence and risk factors associated with equineleptospirosis in the metropolitan region of Salvador and Recôncavo Baiano region, Bahia state (NE Brazil).

Equine leptospirosis, although usually asymptomatic, has been associated with recurrent uveitis, abortion, and other systemic signs, constituting a major economic loss in the equine agribusiness sector. The occurrence of anti-Leptospira spp. agglutinins were investigated in 1200 serum samples of horses from 27 municipalities of the Recôncavo Baiano region, Bahia state (NE Brazil), besides the risk factors related to animals and their cattle farms. The microscopic agglutination test (MAT) was performed using 13 serogroups of Leptospira spp. as antigens. From 1200 analyzed equines, 97 (8%) were considered as positive, which obtained titer equal to or higher than 200. In 22 (78.6%) from the 28 properties, at least one animal was detected as seropositive. The most prevalent serogroup in animals, raised in stables, was the Australis (serovar Bratislava) 67%; in mounted police animals was the Sejroe (serovar Wolffi and Hardjobovis) 50%; and equestrian animals was the Australis (serovar Bratislava) 25% and Icterohaemorrhagiae (serovar Copenhageni) 25%. Equine leptospirosis is present in most cattle farms of the region studied. The risk factors of major importance regarded the semi-extensive livestock farming, the land topography of the cattle farm, the contact with animals in neighboring properties, and cohabitation with goats.

Застосування полівалентної інактивованої вакцини (варіант Equi) проти лептоспірозу коней

Застосування полівалентної інактивованої вакцини (варіант Equi) проти лептоспірозу коней

Lipidomic analysis of immune activation in equine leptospirosis and Leptospira-vaccinated horses.

Currently available diagnostic assays for leptospirosis cannot differentiate vaccine from infection serum antibody. Several leptospiral proteins that are upregulated during infection have been described, but their utility as a diagnostic marker is still unclear. In this study, we undertook a lipidomics approach to determine if there are any differences in the serum lipid profiles of horses naturally infected with pathogenic Leptospira spp. and horses vaccinated against a commercially available bacterin. Utilizing a high-resolution mass spectrometry serum lipidomics analytical platform, we demonstrate that cyclic phosphatidic acids, diacylglycerols, and hydroperoxide oxidation products of choline plasmalogens are elevated in the serum of naturally infected as well as vaccinated horses. Other lipids of interest were triacylglycerols that were only elevated in the serum of infected horses and sphingomyelins that were increased only in the serum of vaccinated horses. This is the first report looking at the equine serum lipidome during leptospiral infection and vaccination.

Evidence of Clinicopathological Changes during Equine Leptospirosis

Evidence of Clinicopathological Changes during Equine Leptospirosis

Molecular characterisation of Leptospira strains in Pakistan

Abstract Introduction: Leptospirosis affects a wide range of mammals, humans, and even a few poikilothermic animal species. In Pakistan, serological studies of equine leptospirosis have reported a prevalence of over 40%, but no study has ever been conducted towards molecular detection of Leptospira in horses. Material and Methods: Blood samples from 128 horses were screened using ELISA and 41 positive samples were examined for the presence of leptospiral DNA using specific primers for 16S rRNA gene. Results: Out of 41 tested samples, 20 samples were found to be PCR-positive, revealing a fragment of 306 bp after gel electrophoresis. Sequencing and phylogenetic analysis of positive samples revealed circulation of pathogenic Leptospira spp. in Pakistani horses. No evidence of circulation of intermediate species was found in this study. Conclusion: This study reports the first molecular evidence of equine leptospirosis in Pakistan and lays ground for further research in this area. It also confirms the efficiency of 16S rRNA for the diagnosis of equine leptospirosis.

Presence of leptospires on genital tract of mares with reproductive problems

Leptospirosis is a zoonotic disease of global importance, and has a worldwide distribution. Equine leptospirosis is commonly manifested by recurrent uveitis, reproductive disorders, as abortions, embryonic absorption, stillbirth and the birth of weak foals. The aim of this study was to verify the presence of Leptospira sp or its DNA in genital tract of mares with reproductive problems. A total of 38 mares with reproductive problems were studied. All the mares were sampled for blood (for serology), urine (for culturing and qPCR), vaginal fluid-VF and endometrial biopsy-EB (for culturing, qPCR and indirect immunofluorescence). PCRs products were sequenced for secY gene. Seventeen (44.7%) serum samples were reactive, predominantly against serogroups Australis (76.4%) and Pomona (23.6%). No positive culture was obtained, but DNA was detected by qPCR on urine samples (26.3%), VF (44.7%) and EB (18.4%) collected 2 months or longer following diagnosis of early fetal death and endometritis. Leptospira cell aggregations were visible by indirect immunofluorescence on 57.1% (4/7) EBs and 17.6% (3/17) VFs. A total of 18 amplicons showed interpretable sequences. Out of those 18 amplicons, 15 presented 100% of identity with the species L. interrogans (sv Bratislava and Pomona), while three were L. borgpertersenii. This study suggests the presence of leptospires in the uterus of mares with reproductive problems. Moreover, serology was shown not to be indicated for the diagnosis of presumptive Leptospira infection in early gestation. The most common agent of the genital infection in those mares was L. interrogans, most probably sg Australis.

Recombinant antigens rLipL21, rLoa22, rLipL32 and rLigACon4-8 for serological diagnosis of leptospirosis by enzyme-linked immunosorbent assays in dogs.

Animal leptospirosis is one of the most common zoonotic diseases in the United States and around the world. In a previous study, we applied four recombinant antigens, rLipL21, rLoa22, rLipL32 and rLigACon4-8 of Leptospira interrogans (L. interrogans) for the serological diagnosis of equine leptospirosis (Ye et al, Serodiagnosis of equine leptospirosis by ELISA using four recombinant protein markers, Clin. Vaccine. Immunol. 21:478–483). In this study, the same four recombinant antigens were evaluated for their potential to diagnose canine leptospirosis by ELISA. A total of 305 canine sera that were Leptospira microscopic agglutination test (MAT)-negative (n = 102) and MAT-positive (n = 203) to 5 serovars (Pomona, Grippotyphosa, Icterohaemorrhagiae, Canicola and Hardjo) were tested. When individual recombinant antigens were used, the sensitivity and specificity of ELISA were 97.5% and 84.3% for rLigACon4-8; 89.7% and 81.4% for rLoa22; 92.6% and 84.3% for rLipL32 and 99.5% and 84.3% for rLipL21, respectively compared to the MAT. The sensitivity and specificity of ELISA were, 92.6% and 91.2% for rLigACon4-8 and rLipL32, 97.5% and 84.3% for rLigACon4-8 and rLipL21, 89.7% and 87.3% for rLigACon4-8 and rLoa22, 89.7% and 87.3% to rLipL21 and rLoa22, 92.6% and 91.2% for rLipL21 and rLipL32 and 89.2% and 94.1% for rLoa22 and rLipL32 when one of the two antigens was test positive. The use of all four antigens in the ELISA assay was found to be sensitive and specific, easy to perform, and agreed with the results of the standard Leptospira Microscopic Agglutination test (MAT) for the diagnosis of canine leptospirosis.

Serodiagnosis of Equine Leptospirosis by Enzyme-Linked Immunosorbent Assay Using Four Recombinant Protein Markers

Leptospirosis, caused by Leptospira spp., is one of the most common zoonotic diseases in the world. We tested four recombinant proteins of Leptospira interrogans, namely, rLipL21, rLoa22, rLipL32, and rLigACon4-8, to evaluate their potential for use as antigens for the diagnosis of equine leptospirosis. We employed equine sera (n = 130) that were microscopic agglutination test (MAT) negative and sera (n = 176) that were MAT positive for the 5 serovars that most commonly cause equine leptospirosis. The sensitivity and specificity of ELISA compared to MAT were 82.39% and 86.15%, respectively, for LigACon4-8, 77.84% and 92.31%, respectively, for Loa22, 77.84% and 86.15%, respectively, for LipL32, and 84.66% and 83.85%, respectively, for LipL21. When one of the two antigens was test positive, the sensitivity and specificity of ELISA were 93.75% and 78.46%, respectively, for rLigACon4-8 and LipL32, 93.18% and 76.15%, respectively, for rLigACon4-8 and LipL21, 89.77% and 80.77%, respectively, for rLigACon4-8 and Loa22, 91.48% and 78.46%, respectively, for LipL21 and Loa22, 93.75% and 76.92%, respectively, for LipL21 and LipL32, and 90.34% and 80.77%, respectively, for Loa22 and LipL32. In conclusion, we have developed an indirect ELISA utilizing rLigACon4-8, rLoa22, rLipL32, and rLipL21 as diagnostic antigens for equine leptospirosis. The use of four antigens in the ELISA was found to be sensitive and specific, the assay was easy to perform, and the results concurred with the results of the standard Leptospira MAT.

Development of an Enzyme-Linked Immunosorbent Assay Using a Recombinant LigA Fragment Comprising Repeat Domains 4 to 7.5 as an Antigen for Diagnosis of Equine Leptospirosis

Leptospira immunoglobulin (Ig)-like (Lig) proteins are a novel family of surface-associated proteins in which the N-terminal 630 amino acids are conserved. In this study, we truncated the LigA conserved region into 7 fragments comprising the 1st to 3rd (LigACon1-3), 4th to 7.5th (LigACon4-7.5), 4th (LigACon4), 4.5th to 5.5th (LigACon4.5-5.5), 5.5th to 6.5th (LigACon5.5-6.5), 4th to 5th (LigACon4-5), and 6th to 7.5th (LigACon6-7.5) repeat domains. All 7 recombinant Lig proteins were screened using a slot-shaped dot blot assay for the diagnosis of equine leptospirosis. Our results showed that LigACon4-7.5 is the best candidate diagnostic antigen in a slot-shaped dot blot assay. LigACon4-7.5 was further evaluated as an indirect enzyme-linked immunosorbent assay (ELISA) antigen for the detection of Leptospira antibodies in equine sera. This assay was evaluated with equine sera (n = 60) that were microscopic agglutination test (MAT) negative and sera (n = 220) that were MAT positive to the 5 serovars that most commonly cause equine leptospirosis. The indirect ELISA results showed that at a single serum dilution of 1:250, the sensitivity and specificity of ELISA were 80.0% and 87.2%, respectively, compared to those of MAT. In conclusion, an indirect ELISA was developed utilizing a recombinant LigA fragment comprising the 4th to 7.5th repeat domain (LigACon4-7.5) as a diagnostic antigen for equine leptospirosis. This ELISA was found to be sensitive and specific, and it yielded results that concurred with those of the standard MAT.

Equine leptospirosis in tropical Northern Queensland

Determine leptospiral serodiversity, serodominant serovars and prevalence in the horse population of Northern Queensland (NQ), Australia, with special focus on the Atherton Tableland and Townsville-Burdekin regions. Cross-sectional survey. Serum samples from 429 horses originating from 172 horse-owning properties in NQ were collected at regional horse shows and veterinary clinics. Samples were analysed using a microscopic agglutination test. Owners were interviewed on potential risk factors associated with leptospirosis. Of the 172 properties that submitted samples, horses from 60 properties (35%, 95% confidence interval (CI) 31-39%) in NQ showed evidence of exposure to one or more serovars of Leptospira at a serum dilution of 1 : 100. The proportion of seropositive properties in the Atherton Tableland and Townsville-Burdekin regions was 25% (95% CI 17-33%) and 36% (95% CI 30-43%), respectively. In total, 81 horses had titres to a single serovar and the remained had titres to two or more serovars. Although 20 serovars representing 17 serogroups were detected, serovar Arborea was serodominant and detected in horses from 29 properties. Geldings were more at risk (odds ratio 3.3) of being seropositive to serovar Arborea. The detection of serovar Arborea-seropositive horses was strongly associated (P < 0.05) with the average annual rainfall each property received and the presence of pigs and sugarcane fields in the vicinity. Serological evidence of leptospirosis in horses in NQ indicates that prevalence is high and horses have been exposed to a wide range of serovars.

Leptospirosis in horses

Leptospirosis in horses has been considered a relatively uncommon infection. However, recent data suggest that the infection is widespread, with the incidence and infecting serovars varying considerably in different geographical regions. The majority of infections remain asymptomatic. Clinical signs in equine leptospirosis resemble those seen in other animal species. However, leptospirosis as a cause of acute respiratory distress is becoming more frequently recognised. A particular feature of equine leptospirosis is post infection recurrent uveitis (moon blindness or periodic ophthalmia), which appears to be mediated by autoimmune mechanisms involving cross reactivity between ocular tissues and leptospiral membrane proteins. There are no leptospiral vaccines licensed for use in horses, with no prospect for any becoming available in the foreseeable future. Accordingly, prevention of equine leptospirosis must rely on good hygiene practices, minimisation of rodent contact, and vaccination of other species of production and companion animals.

Seroprevalence study of leptospirosis in horses in northern Poland

Serological surveys in many laboratories around the world have indicated that leptospirosis occurs frequently in horses, and can be caused by a wide variety of Leptospira serovars. The infections are...