A serological survey of canine leptospirosis in the city of Belgrade, Serbia

Canine leptospirosis is a zoonosis caused by bacteria belonging to the genus Leptospira. Dogs are one of the animal species involved in the cycle of preservation and transmission of leptospirosis in urban areas. Serological testing for the presence of specific antibodies against Leptospira spp. in dogs was continuously performed between 2010 and 2020 in the city of Belgrade. At the request of the owners themselves, other veterinary laboratories or laboratory clinics, 179 blood sera from 179 dogs were examined in the Laboratory for Immunology, Scientific Institute of Veterinary Medicine of Serbia. Blood sera samples from dogs were examined using the standard microscopic agglutination test (MAT) for the presence of specific antibodies against seven different serovars of Leptospira: Pomona, Icterohaemorrhagiae, Grippotyphosa, Sejroe, Canicola, Bataviae, and Australis. The number of seropositive dogs was 17/179 (9.5%). Among all examined sera, the highest titre of seropositive samples was to serovar Icterohaemorrhagiae (10/17, 58.8%), followed by Pomona (4/17, 23.5%), and serovar Canicola (3/17, 17.6%). Specific antibodies for serovars Grippotyphosa, Sejroe, Bataviae and Australis were not detected in any of the dog sera. Cross-reaction (the presence of two or three titres with different values where one of them was higher than others) between different serovars was diagnosed in a low number of sera (n=4), with the following serovars: Icterohaemorrhagiae and Pomona (n=3) and Pomona and Canicola (n=1). The confirmed specific antibody titres for Leptospira spp. were between 1:100 to 1:3000 (5 sera had titres of 1:100, 7 had titres of 1:300, 4 had titres of 1:1000, and 1 serum had a titre 1:3000). Monitoring canine leptospirosis is a useful tool in preventing leptospirosis in Belgrade.

Brote de 4 casos leptospirosis de un grupo familiar en zona rural tropical de Colombia: un reporte de caso

La leptospirosis es una zoonosis con manifestaciones clínicas causadas por espiroquetas patógenas del género Leptospira spp. Su curso puede ser desde enfermedad leve hasta un síndrome ictero-hemorrágico severo denominado enfermedad de Weil. Se estudió un brote epidemiológico constituido por una serie de cuatro casos de leptospirosis de severidad moderada a severa, ocurridos en una zona rural de clima tropical en los llanos orientales de Colombia, a 450 metros sobre el nivel del mar en una familia visitante en Puerto Lleras, Meta, Colombia. Probablemente secundario al consumo de aguas contaminadas por orina de roedores. Las pacientes fueron ingresadas bajo la sospecha de un síndrome ictérico de origen infeccioso con un falso positivo para antígeno de superficie de hepatitis B. Dos pacientes desarrollaron síndrome de Weil asociado a microangiopatía trombótica por lo que requirieron manejo en unidad de cuidados intensivos (UCI) sin embargo, una de ellas fallece y las otras dos pacientes desarrollan signos y síntomas moderados evidenciando un curso variable de la enfermedad. En nuestro país existe una sobre notificación de síndromes febriles, ictéricos e icterohemorragicos de diferentes etiologías y difícil diagnóstico por lo que la leptospirosis tiende a ser confundida o ignorada como diferencial en muchos casos.

Sensitivity and Specificity of Serology, Histopathology, and Molecular Tests in the Detection of Leptospirosis from Slaughtered Cattle in Indonesia

Leptospira spp. is a pathogenic bacteria that causes leptospirosis in humans and cattle. The World Health Organization (WHO) recommends the microscopic agglutination test (MAT) as the laboratory gold standard in the detection of leptospirosis. However, the limitation of MAT triggers the laboratory technicians to develop alternative laboratory tests against leptospirosis. The current study aimed to compare the sensitivity and specificity of histopathology special stain using modified Gram staining (MGS) and molecular test using reverse transcriptase-polymerase chain reaction (RT-PCR), compared to the MAT for Leptospira spp. detection in cattle. This study used a total of 38 serum and 38 kidney samples from the cattle slaughtered in the Sidoarjo slaughterhouse, Indonesia. The collected serum samples were tested against MAT and RT-PCR. The kidneys were processed for histopathology using MGS. The result indicated that 16 (42.10%) of the tested samples were positive against MAT, 6 (15.78%) were positive against MGS, and 18 (47.36%) were positive against RT-PCR. The RT-PCR indicated better sensitivity and lower specificity, compared to MAT and MGS. The findings revealed that the RT-PCR is an appropriate laboratory test for detecting cattle leptospirosis with better sensitivity and specificity. Therefore, this method can be suggested to substitute MAT and overcome its limitations.

A Unique Case of Fatal Coinfection Caused by Leptospira spp. and Hepatozoon canis in a Red Fox Cub (Vulpes vulpes)

Red foxes are the most abundant wild carnivore species in Europe commonly exposed to pathogenic Leptospira and Hepatozoon canis. Despite high seroprevalence, the clinical disease caused by these pathogens in red foxes has never been reported. Herein, we report the first-ever case of a fatal Leptospira spp. and H. canis coinfection in a two-month-old red fox cub with acute haemolytic anaemia, mild bronchopneumonia, intraalveolar haemorrhage, and tubulonephrosis. The presence of pathogenic Leptospira spp. DNA was detected in the kidney and lung tissues of the infected animal. In contrast to our previous knowledge, we believe that such fatal cases due to concomitant infection by Leptospira spp. and H. canis, especially in young animals, may commonly occur in nature. However, further studies are required to identify other factors that possibly contribute to the severity and the pathogenic effect of Leptospira spp. and H. canis infections in red foxes.

First Isolation and Molecular Typing of Pathogenic and Intermediate Leptospira Species from Urine of Symptomatic Dogs

Aim of this study was to evaluate, the presence and diversity of Leptospira spp. in blood and urine samples collected from 175 owned-dogs from Sardinia, Italy. After determination of leptospiral infection by microscopic agglutination test (MAT), urine from MAT-positive dogs were examined by real-time polymerase chain reaction (lipL32 rt-PCR) and then isolated by culture. In order to characterize obtained serovars, positive cultures were then subjected to 16S rRNA and secY sequencing, phylogenetic analysis and Multilocus Sequence Typing (MLST). Results showed that seven dogs (4%; 95% CI: 0–55) had Leptospira DNAs in their urine and five strains were isolated from urine cultures. The three different sequence types (ST17, ST198 and ST24) belonging to Leptospira interrogans genomospecies identified by MLST analyses in this study, confirmed that the leptospiral infection was widespread in Sardinian dogs. We also reported the first characterization of a new Leptospira spp. isolated from urine of one dog living in the study area. Whole genome sequencing and phylogenetic analysis, confirmed that this genospecies was closely related to Leptospira hovindhougenii, an intermediate Leptospira spp. with unknown pathogenicity previously isolated from a rat in Denmark. Further studies are required to clarify whether healthy dogs that shed leptospires in their urine could represent a zoonotic risk for humans in this region.

ESTUDO RETROSPECTIVO DA LEPTOSPIROSE CANINA EM HOSPITAL VETERINÁRIO ESCOLA NO MUNICÍPIO DE LONDRINA, PR

O objetivo do trabalho foi relatar a ocorrência de casos de cães com Leptospira spp. no Hospital Veterinário de Londrina. Foram avaliados 5.454 prontuários, durante os meses de janeiro a dezembro de 2016, e selecionados os que apresentaram solicitação de exame para Lepstopira por meio da técnica de campo escuro (CE). Nos positivos pela técnica CE dados sobre o sexo, sinais clínicos, endereço, data do atendimento e resultado na SAM foram coletados. A análise dos dados foi realizada pelo programa EpiInfo (7.2.3.1), dos 5.454 prontuários nos quais foi solicitado exame CE em 7,06% (385/5.454), e nestes, 13,76% (53/385) foi identificada a presença da espiroqueta. Dos animais positivos na técnica CE, 20,75% (11/53) foram reagentes na SAM. O sorovar Canicola foi o mais frequente em 90,91% (10/11) dos animais, apresentando títulos variados de 100 a 12.800. Para o sorovar Pomona 9,09% (1/11) dos animais apresentaram títulação de 100. Os sinais clínicos mais observados fora apatia em 58,49% (31/53), azotemia em 49,06% (26/53) e êmese em 49,06% (26/53) dos casos. Quanto ao sexo, os machos constituíram 71,70% dos casos e as fêmeas 28,30% (p=0,0015). Os resultados demonstraram a presença de Leptospira spp. entre os animais atendidos no hospital veterinário de Universidade Estadual de Londrina e reforçam a importância do sorovar Canicola entre os cães, visto que este foi o mais frequente entre os sorovares e que apresentou os maiores títulos.

Detección de Leptospira spp. (Spirochaetales: Leptospiraceae) en muestras ambientales de regiones habitadas por poblaciones vulnerables del norte argentino

LLa leptospirosis continúa siendo hoy en día un problema para la salud pública, principalmente en poblaciones de bajos recursos socioeconómicos. En este trabajo se presenta la detección de leptospiras patógenas en muestras ambientales (aguas y barros) provenientes de regiones del norte argentino (provincias de Formosa, Salta, Santiago del Estero, Misiones y Chaco) con variadas características climatológicas habitadas por poblaciones vulnerables. De las 89 muestras analizadas, en el 24,7% fue posible detectar molecularmente la presencia de leptospiras patógenas. La prevalencia por tipo de muestra fue de 27,8% para las aguas y 11,8% para los barros. Todas las localidades muestreadas presentaron al menos una muestra positiva a alguna de las pruebas realizadas, por lo que el presente trabajo refleja la necesidad de profundizar los estudios de la leptospirosis en distintas regiones de la Argentina.

Zingiber zerumbet (L.) Smith Hexane Crude Extract Caused DNA Damage on Leptospira spp.

Numerous attempts have been made to control leptospirosis by using chemoprophylaxis, but with limited success. The present study was done to investigate the antileptospiral potential of hexane, ethyl acetate and methanol extracts of Zingiber zerumbet rhizomes. The extracts were assayed for antileptospiral activity using broth microdilution method against Leptospira interrogans (serovar Batavie, Canicola, Australis) and Leptospira biflexa serovar Patoc. The Z. zerumbet hexane extract exhibited antileptospiral activity, with IC50 values of 248 μg/mL against L. interrogans serovar Canicola, IC50 of 125 μg/mL against L. interrogans serovar Australis, IC50 of 15.63 μg/mL against L. interrogans serovar Batavie and IC50 of 109 μg/mL against L. biflexa serovar Patoc. However, both ethyl acetate and methanol extracts did not show any distinct antileptospiral activity. Since the hexane extract of Z. zerumbet showed antileptospiral activity, the DNA-damaging properties of this extract were tested according to their IC50 and IC25 values that were specific to each serovars. The DNA-damaging properties were determined by treating the selected Leptospira spp. with the hexane extract and subjecting its DNA to electrophoresis and analysis on agarose gels. The results demonstrated that the hexane extract had DNA-damaging properties towards L. biflexa serovar Patoc and L. interrogans serovar Australis, as proven by the appearance of fragmented DNA on the gels. We conclude that the Z. zerumbet hexane extract could inhibit the growth of Leptospira spp. serovar Patoc and Australis through DNA-damaging activity and thus, could be a potential antileptospiral agent. Further studies are needed to investigate the potential of this hexane extract as an antileptospiral agent using in vivo rat models of leptospirosis.