Leptospirosis is a zoonosis of worldwide distribution, which is cause of economic losses in farm animals with public health implications. Objetive. was to determine the presence of L. interrogans sensu lato in urine of cattle and water sources in rural areas from Córdoba (Colombia). Materials and methods. 78 urine samples were collected from cows and 39 samples of water for bacteriological culture and determination of the 16S rRNA gene from bacterial L. interrogans sensu lato. Although a definitive isolation was not achieved, L. interrogans sensu lato was detected by PCR in three urine samples and in one sample of waste water. Results. Circulation of L. interrogans sensu lato was confirmed by means PCR. Conclusions. These molecular evidences suggested the possibility of circulation of L. interrogans sensu lato in cows and water sources from rural areas of Ciénaga de Oro.