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.

Attenuated nephritis in inducible nitric oxide synthase knockout C57BL/6 mice and pulmonary hemorrhage in CB17 SCID and recombination activating gene 1 knockout C57BL/6 mice infected with Leptospira interrogans.

The aims of this study were to investigate the frequency of pulmonary hemorrhage (PH) in mice unable to produce functional B and T lymphocytes and to explore the effect of an inducible nitric oxide synthase gene (Inos) knockout (KO) on the frequency/severity of interstitial nephritis in vivo. We studied the outcome of infection by the virulent Leptospira interrogans serovar Copenhageni strain Cop. The animals used were Inos KO mice, recombination activating gene 1 (Rag1) KO mice, CB17 severe combined immunodeficiency (SCID) mice, and the respective wild-type (WT) C57BL/6 and BALB/c controls. The Inos KO and WT mice survived with no clinical symptoms of leptospirosis. The frequency and severity of nephritis was significantly lower in the Inos KO mice. All of the Rag1 KO and SCID animals died of acute leptospirosis, whereas all of the WT mice survived. PH was observed in 57 and 94% of Rag1 KO mice and in 83 and 100% of SCID mice, using inoculum doses of 10(7) and 10(6) leptospires, respectively. There was no evidence of PH in the WT controls. In conclusion, the loss of the Inos gene had a negligible effect on the outcome of leptospiral infection, although we observed a reduced susceptibility for interstitial nephritis in this group. Of note, the absence of functional B- and T-cell lymphocytes did not preclude the occurrence of PH. These data provide evidence that PH in leptospirosis may not be related only to autoimmune mechanisms.

Detection and quantification of Leptospira interrogans in hamster and rat kidney samples: immunofluorescent imprints versus real-time PCR.

A major limitation in the clinical management and experimental research of leptospirosis is the poor performance of the available methods for the direct detection of leptospires. In this study, we compared real-time PCR (qPCR), targeting the lipL32 gene, with the immunofluorescent imprint method (IM) for the detection and quantification of leptospires in kidney samples from the rat and hamster experimental models of leptospirosis. Using a virulent strain of Leptospira interrogans serovar Copenhageni, a chronic infection was established in the rat model, which were euthanized 28 days post-infection, while the hamster model simulated an acute infection and the hamsters were euthanized eight days after inoculation. Leptospires in the kidney samples were detected using culture isolation, qPCR and the IM, and quantified using qPCR and the IM. In both the acute and chronic infection models, the correlation between quantification by qPCR and the IM was found to be positive and statistically significant (P<0.05). Therefore, this study demonstrates that the IM is a viable alternative for not only the detection but also the quantification of leptospires, particularly when the use of qPCR is not feasible.

Different outcomes of experimental leptospiral infection in mouse strains with distinct genotypes.

The mouse disease model has the advantage of a broad array of immunological and genetic tools available for basic research. Some studies on transgenic and/or mutant mouse strains as models for experimental leptospirosis have been reported; however, the wider use of such models is hampered by a poor understanding of the outcome of experimental leptospiral infection among the different mouse strains available. Here, the outcome of infection by a virulent strain of Leptospira interrogans serogroup Icterohaemorrhagiae strain Cop was studied in four commonly used wild-type mouse strains: A, CBA, BALB/c and C57BL/6. The end points evaluated in this study were survival, presence of kidney lesions, leptospiral load in kidney samples, microscopic agglutination test titre and anti-leptospiral IgG antibody levels. As expected, none of the mouse strains were susceptible to lethal leptospirosis. However, these strains developed specific pathologies associated with sublethal leptospirosis. The A and C57BL/6 strains exhibited a high leptospiral load in kidney samples and the CBA and C57BL/6 strains developed severe inflammatory lesions, whilst the BALB/c strain proved to be the most resistant to subclinical leptospirosis.

An imprint method for detecting leptospires in the hamster model of vaccine-mediated immunity for leptospirosis.

In determining the efficacy of new vaccine candidates for leptospirosis, the primary end point is death and an important secondary end point is sterilizing immunity. However, evaluation of this end point is often hampered by the time-consuming demands and complexity of methods such as culture isolation (CI). In this study, we evaluated the use of an imprint (or touch preparation) method (IM) in detecting the presence of leptospires in tissues of hamsters infected with Leptospira interrogans serovar Copenhageni. In a dissemination study, compared to CI, the IM led to equal or improved detection of leptospires in kidney, liver, lung and blood samples collected post-infection and overall concordance was good (kappa=0.61). Furthermore, in an evaluation of hamsters immunized with a recombinant leptospiral protein-based vaccine candidate and subsequently challenged, the agreement between the CI and IM was very good (kappa=0.84). These findings indicate that the IM is a rapid method for the direct observation of Leptospira spp. that can be readily applied to evaluating infection in experimental animals and determining sterilizing immunity when screening potential vaccine candidates.