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      Diagnosis of acute canine leptospirosis using multiple laboratory tests and characterization of the isolated strains

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          Abstract

          Background

          Dogs presenting with acute leptospirosis may present non-specific clinical and laboratory findings, and the definitive diagnosis may require additional confirmatory tests, including bacterial culture, for the direct or indirect identification of the pathogen. The present study describes the diagnosis of leptospirosis in suspected dogs based on the use of multiple diagnostic tests, including serological, molecular and bacteriological tests, along with the characterization of the recovered leptospiral strains.

          Results

          Urine, serum and blood samples were collected from 33 dogs with suspected clinical leptospirosis treated at the University of São Paulo Veterinary Hospital Service (Hovet FMVZ-USP) between 2013 and 2016. Only dogs with high blood urea nitrogen and creatinine levels in association with multiple clinical manifestations of the disease were included. Leptospiral culture, PCR and serology (Microscopic agglutination test - MAT) were performed in blood and urine samples taken from all suspected dogs at clinical presentation, and an additional prospective MAT titration was performed in seven dogs. Infection could be identified exclusively by PCR in 10 dogs (30.3%), exclusively by MAT in four dogs (12.1%) and by both tests in four dogs, totaling 18 dogs (54.5–95%CI: 37.6–71.5). Six out of eight MAT-confirmed cases presented with the highest titers against the Icterohaemorrhagiae serogroup. Leptospires were recovered from urine samples from two PCR-positive dogs, and both strains could be characterized by Multilocus Sequence Analysis and serogrouping as L. interrogans serogroup Icterohaemorrhagiae. Both isolates were shown to be pathogenic in the hamster model.

          Conclusions

          The simultaneous use of MAT and PCR was able to increase the diagnosis of leptospirosis in clinically suspected cases. Despite the increasing incidence of new serovars affecting dogs being reported in different locations, our results suggest that leptospiral strains belonging to the Icterohaemorrhagiae serogroup are still a major causative agent of canine leptospirosis in São Paulo, Brazil.

          Electronic supplementary material

          The online version of this article (10.1186/s12917-018-1547-4) contains supplementary material, which is available to authorized users.

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          Most cited references37

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          Emergence, control and re-emerging leptospirosis: dynamics of infection in the changing world.

          Globally, leptospirosis poses an increasing public health problem, as evidenced by markedly increasing incidence rates and multiple outbreaks in all continents. Yet, the disease is severely neglected and hence, its global burden is largely unknown. The estimated incidence of about half a million severe human cases annually is probably an underestimation while the burden for animal health is unknown. It is anticipated that current international initiatives will assess the global burden of leptospirosis, while mathematical modelling of transmission dynamics will allow the identification and testing of appropriate intervention and outbreak response measures within the coming years. © 2011 The Authors. Clinical Microbiology and Infection © 2011 European Society of Clinical Microbiology and Infectious Diseases.
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            Polymerase chain reaction for detection of Leptospira spp. in clinical samples.

            A sensitive assay for Leptospira spp., the causative agent of leptospirosis, was developed on the basis of the polymerase chain reaction (PCR). A 331-bp sequence from the Leptospira interrogans serovar canicola rrs (16S) gene was amplified, and the PCR products were analyzed by DNA-DNA hybridization by using a 289-bp fragment internal to the amplified DNA. Specific PCR products also were obtained with DNA from the closely related nonpathogenic Leptospira biflexa but not with DNA from other spirochetes, such as Borrelia burgdorferi, Borrelia hermsii, Treponema denticola, Treponema pallidum, Spirochaeta aurantia, or more distant organisms such as Escherichia coli, Staphylococcus aureus, Mycobacterium tuberculosis, and Proteus mirabilis. The assay was able to detect as few as 10 bacteria. Leptospira DNA was detected in urine from experimentally infected mice. In addition, the test was found to be suitable for diagnosing leptospirosis in humans. Cerebrospinal fluid and urine from patients with leptospirosis were positive, whereas samples from control uninfected patients were negative.
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              Urban epidemic of severe leptospirosis in Brazil

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                Author and article information

                Contributors
                brunomiotto@hotmail.com
                ba_tozzi@hotmail.com
                manoela.penteado@hotmail.com
                aline_gil@hotmail.com
                luzanolli@gmail.com
                (55 11) 3091-7927 , (55 11) 9 9616-1453 , marcosbryan@usp.br
                morenoam@usp.br
                wlilenbaum@id.uff.br
                mitika.hagiwara@gmail.com
                Journal
                BMC Vet Res
                BMC Vet. Res
                BMC Veterinary Research
                BioMed Central (London )
                1746-6148
                17 July 2018
                17 July 2018
                2018
                : 14
                : 222
                Affiliations
                [1 ]ISNI 0000 0004 1937 0722, GRID grid.11899.38, Departamento de Clínica Médica (Department of Veterinary Clinics), Faculdade de Medicina Veterinária e Zootecnia (School of Veterinary Medicine and Animal Science), , Universidade de São Paulo (University of São Paulo), ; São Paulo, SP 05508-270 Brazil
                [2 ]ISNI 0000 0004 1937 0722, GRID grid.11899.38, Departamento de Medicina Veterinária Preventiva e Saúde Animal (Department of Veterinary Preventive Medicine and Animal Health), Faculdade de Medicina Veterinária e Zootecnia (School of Veterinary Medicine and Animal Science), , Universidade de São Paulo (University of São Paulo), ; São Paulo, SP 05508-270 Brazil
                [3 ]ISNI 0000 0001 2184 6919, GRID grid.411173.1, Departamento de Microbiologia e Parasitologia (Department of Microbiology and Parasitology), , Universidade Federal Fluminense, ; Niterói, RJ 24210-130 Brazil
                Article
                1547
                10.1186/s12917-018-1547-4
                6050646
                30016949
                c4f62853-10ae-4cb6-9a5d-32db3d702346
                © The Author(s). 2018

                Open AccessThis article is distributed under the terms of the Creative Commons Attribution 4.0 International License ( http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver ( http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.

                History
                : 1 January 2018
                : 9 July 2018
                Funding
                Funded by: FundRef http://dx.doi.org/10.13039/501100001807, Fundação de Amparo à Pesquisa do Estado de São Paulo;
                Award ID: 2012/14681-7
                Award ID: 2012/13022-0
                Award ID: 2013/17136-2
                Funded by: FundRef http://dx.doi.org/10.13039/501100003593, Conselho Nacional de Desenvolvimento Científico e Tecnológico;
                Award ID: 164284/2014
                Categories
                Research Article
                Custom metadata
                © The Author(s) 2018

                Veterinary medicine
                leptospirosis,dogs,canine,pcr,mat,acute infection,culture,sequencing,mlst,icterohaemorrhagiae
                Veterinary medicine
                leptospirosis, dogs, canine, pcr, mat, acute infection, culture, sequencing, mlst, icterohaemorrhagiae

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