Culture-Based Isolation of Leptospira Species from Human Urine at a Tertiary Care Hospital in Bangladesh.

Leptospirosis is a significant zoonotic disease and previous studies have confirmed its presence in Bangladesh. Despite its potential to cause severe illness and death, the disease is frequently under-recognized because of limited diagnostic facilities and insufficient clinical awareness. This cross-sectional descriptive type of observational study was aimed to isolate Leptospira spp. from human urine by culture in a resource-limited setting at Mymensingh Medical College, Mymensingh, Bangladesh from October 2024 to March 2025. A total of 20 urine samples from clinically suspected cases of leptospirosis were collected and cultured in modified Ellinghausen-McCullough-Johnson-Harris (EMJH) medium. The isolates were confirmed by nested PCR and Sanger sequencing targeting the 16S rRNA gene. Four isolates (20.0%) were recovered and identified as Leptospiraspp. by dark field microscopy and nested PCR. Culture positivity was highest (25.0%) in patients presenting within 5-7 days of illness and declined with longer fever duration, with no isolates detected after 15-20 days. A lower positivity rate was observed among patients with recent antibiotic use (1 case) compared to those without antibiotic exposure (3 cases). Phylogenetic analysis identified all isolates as Leptospira wolffii, closely related to previously reported strains from the Mymensingh area. These findings provide microbiological evidence of the ongoing circulation of Leptospira wolffii and demonstrate the utility of urine culture, alongside molecular methods, to enhance diagnostic capacity and epidemiological understanding in endemic settings.