Serological and molecular findings in diagnosis of leptospirosis serovar hardjo in a dairy bovine herd

Abstract

The cattle are considered hosts of the Hardjo serovar, causing economic damages due to the reproductive failures like abortions and infertility. The serovar Hardjo usually remains in the reproductive tract and also in the renal tubules where it is eliminated intermittently in the urine for months. Placental remnants, the aborted fetus and contaminated urine promote the permanence of this bacterium within the herd for years. Thus, the objective of this study was to monitor for prolonged period, cows naturally infected with Leptospira ssp. through microbiological culture, serological examination and DNA detection of the pathogen in the urine. The dairy herd was composed of 50 breeding cows with a history of abortion and infertility, without leptospirosis vaccine and located in the northern region of Paraná. Blood and urine samples were collected and laboratorial diagnosis were performed five times at intervals of four months. Blood samples were collected from the all 50 animals and the serum was submitted to the microscopic agglutination test (MAT) for the detection of anti-leptospira antibodies. Of the total cows, 20 showed antibody titres ≥ 1: 100 in MAT and urine samples were collected from only those animals with higher titers to perform nested-PCR (n-PCR) and bacterial isolation per culture. In addition, two urine samples from five animals with antibody titers < 1: 100 were collected in MAT for n-PCR. Serovar Hardjo was considered the most frequent during the serological monitoring of the animals evaluated. The n-PCR technique was able to detect leptospiral DNA in the urine of animals with MAT ≥ 1: 100 antibody titers and urine from animals whose titers were < 1: 100. Sequencing of the leptospiral amplicons shared 100% nucleotide sequence identity with the Leptospira interrogans species. Positive n-PCR results from animals with titers of < 1: 100 suggest that the cut-off of MAT is could be not sufficient to detect renal carriers, so it is also important to use n-PCR as an additional diagnostic tool for identify infected animals with Hardjo serovar and whose serology was negative. reprodutivo e também nos túbulos renais onde é eliminado de forma intermitente na urina por meses. Restos placentários, o feto abortado e a urina contaminada favorecem a permanência dessa bactéria dentro do rebanho por anos. Assim, o objetivo deste estudo foi monitorar por período prolongado, vacas naturalmente infectadas por Leptospira ssp. através de cultura microbiológica, exame sorológico e detecção de DNA do patógeno na urina. O rebanho leiteiro estudado era constituído por 50 vacas reprodutoras com histórico de abortos e infertilidade, sem uso de vacina contra a leptospirose e localizada na região Norte do Paraná. As amostras de sangue e urina foram coletadas e submetidas a análises laboratoriais em cinco vezes em intervalos espaçados de quatro meses. As amostras de sangue foram coletadas dos 50 animais e o soro foi submetido ao teste de soroaglutinação microscópica (SAM) para detecção de anticorpos anti-leptospira. Do ...

Culture Strategies for Isolation of Fastidious Leptospira Serovar Hardjo and Molecular Differentiation of Genotypes Hardjobovis and Hardjoprajitno

“…EMJH culture media (liquid or semi-solid) is widely used to isolate Leptospira (Rodríguez et al, 2002 ; González et al, 2006 ; Zacarias et al, 2008 ; Miraglia et al, 2009 ; Chakraborty et al, 2011 ). For many years, our leptospirosis research group has unsuccessfully attempted to isolate serovar Hardjo from bovine urine using unmodified EMJH, even after obtaining positive serology for this serovar (Hashimoto et al, 2017 ).…”

Serological diagnosis and molecular characterization of Leptospira spp. in the blood and urine of bovine females from refrigerated slaughterhouses

“…Therefore, the results of the present study show that it is important to combine serological and molecular diagnosis techniques (HASHIMOTO et al, 2017), because they complement one another. Thereby, even when no antibody was detected in the serum sample of one animal, there may be DNA in the urine sample of this animal.…”

Bovine genital leptospirosis: Findings in bulls maintained in Caatinga biome conditions

“…There was variation in reactive serogroups (Sejroe, Tarassovi, Canicola e Grippotyphosa), which suggests different sources of infection, although there is a possibility that cattle without signs and symptoms of infection may carry and disseminate other strains within the species in an adaptive process (Pinto et al 2017). As in other regions of Brazil (Hashimoto et al 2017, Miashiro et al 2018, Pinna et al 2018, Guedes et al 2019a, 2019b, Pimenta et al 2020) and the world (Loureiro et al 2016, Sejroe was the most prevalent serogroup in Caatinga biome cattle.…”