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

Manoela de Souza Penteado,Walter Lilenbaum,Aline Gil Alves Guilloux,Andrea Micke Moreno,Luisa Zanolli Moreno,Bruno Alonso Miotto,Marcos Bryan Heinemann,Mitika Kuribayashi Hagiwara,Barbara Furlan Tozzi

doi:10.1186/s12917-018-1547-4

Manoela de Souza Penteado, Walter Lilenbaum + Show 7 more

Open Access

https://doi.org/10.1186/s12917-018-1547-4

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Abstract

BackgroundDogs 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.ResultsUrine, 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.ConclusionsThe 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.

Highlights

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
Acute leptospirosis was diagnosed in eight dogs (24.2%) by Microscopic agglutination test (MAT): six dogs without a history of recent vaccination had titers ≥800 in a single serum sample; prospective evaluation performed in seven dogs revealed seroconversion in two other dogs, which showed fourfold increase in MAT titers against Icterohaemorrhagiae serogroup
Most dogs diagnosed by Polymerase Chain Reaction (PCR) and DNA sequencing were identified using urine samples, and even though the detection of leptospiral DNA in urine specimens taken from dogs with multiple signs of leptospirosis is highly suggestive of acute leptospiral infection, it may not distinguish dogs with acute infection from those with chronic renal carriage of leptospires associated with other underlying diseases causing clinical manifestations similar to leptospirosis

Summary

Introduction

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. Clinical and laboratory findings are usually non-specific, and a definitive diagnosis requires additional confirmatory tests for the direct or indirect identification of the pathogen, such as dark-field microscopy, Polymerase Chain Reaction (PCR), bacterial culture and Microscopic agglutination test (MAT) [1]. Since the exact time of infection at clinical presentation is typically unknown, and given that the early and accurate identification of infected dogs is crucial to alter the course of the disease with appropriate drug therapy [5], the use of multiple simultaneous tests may improve the chance for a correct diagnosis and consequent therapeutic success [6]. MAT has a poor ability to predict the infecting serovar and may not distinguish between infection and vaccine-induced titers [10]

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