Caracterização de cães com leptospirose, por meio de testes de diagnóstico sorológico, molecular e alterações clínicos-patológicas, e investigação do efeito antimicrobiano da nanopartícula ZnO:9Ag em cepas padrão de Leptospira spp

Abstract

Leptospirosis is a zoonosis of worldwide importance that causes deaths in both humans and animals. Infection in dogs and other species occurs through contact with urine or water contaminated by pathogenic bacteria of the genus Leptospira. The objective was to evaluate two diagnostic tests in the characterization of dogs with leptospirosis, to associate them with the main clinical-pathological alterations of suspected dogs, in addition to investigating the antimicrobial effect of the silver-doped Zinc Oxide nanoparticle (ZnO:9Ag) in standard strains Leptospira. spp. Five pathogenic species of Leptospira (L. interrogans, L. santarosai, L. borgpetersenii, L. kirschneri, L. noguchii) were selected, totaling ten standard strains. Minimum inhibitory concentration (MIC), cell viability by colorimetric assay (MTT), cell permeability test and release of intracellular content were used to verify the in vitro effect of streptomycin, penicillin G, doxycycline, tetracycline and ZnO:9Ag in the species of Leptospira. The safety and toxicity of ZnO:9Ag were analyzed in an alternative in vivo model, chicken embryo (EG). Of the 24 dogs evaluated, 8 (33.3%) were positive for leptospirosis, being 6 (25%) positive in the MAT, 2 (8.33%) positive in the qPCR and 1 (4.17%) positive in both tests. The predominant reactive serogroups were Icterohaemorrhagiae (33.33%) and Djasiman (33.33%), followed by Ballum (16.60%). The sensitivity and specificity of qPCR in relation to the gold standard test (MAT) was 16.7% and 88.9%, respectively. There were changes in the serum levels of creatinine, urea and FA in the positive group. All treatments exhibited anti Leptospira effects. Penicillin G had a lower MIC (<0.048), although ZnO:9Ag had a higher MIC (6.25 to 50 μg/mL), it also had an inhibitory effect, making it a good therapeutic option. Fifty percent (50%) of the strains showed lower viability with ZnO:9Ag (100 μg/mL), three in the species L. interrogans and two in the species L. kirschneri and L. noguchii. ZnO:9Ag was the only one that showed a significant change (p<0.05) in permeability in all species, and it was the treatment that most caused extravasation of intracellular content. The in vivo toxicity assay of ZnO:9Ag in chicken embryos revealed a difference with the negative control (p<0.05) only in the GGT enzyme. It is concluded for the diagnosis of acute canine leptospirosis, associating the two diagnostic tests MAT and qPCR and the clinical-pathological findings seems to be the best way, however the results suggest that the nanoparticle (ZnO:9Ag) can be potentially used as an agent alternative antibacterial against Leptospira spp., capable of inhibiting its growth, besides being safe and not very toxic to the organism.