en Diagnostic/screening methods and protocols Diagnostic/screening methods and protocols Original article Original article <strong>Introduction</strong>: Leptospirosis is a significant public health problem in the Caspian littoral of Iran comprising Gilan, Mazandaran, and Golestan provinces.&nbsp; In Golestan province, serology assays indicated anti-<em>Leptospira</em> antibodies in animals and humans; however, no reliable record of infections in patients with signs and symptoms of the disease is available. <strong>Methods</strong>: We employed the indirect immunofluorescent antibody assay (IFA), and two PCR assays, a real-time PCR (qPCR) and a nested-PCR targeting 16S rDNA (<em>rrs</em>) sequence for diagnosis of leptospirosis in febrile patients in Golestan province. <strong>Results</strong>: Out of 52 febrile patients, 25 (48.07%) had antibody titers &ge;1/80 by IFA, and were defined as positive. In 7, 9, and 7 individuals, the antibody titers were 1/40, 1/20, and 1/10, respectively, and 4 had no antibodies. The qPCR and nested PCR detected leptospiral DNA in 55.75% and 67.3% of the patients&#39; sera, respectively. The two PCR assays had a Kappa agreement of 0.53 (<em>P</em>< 0.0001), suggesting a moderate agreement, and showed a significant reverse association with the IFA titers (<em>P</em><0.05). <strong>Conclusion</strong>: Our results suggest that higher antibody titers accompanied the spirochete&#39;s removal from blood by the patient&#39;s immune response. Hence, a reliable diagnosis of <em>Leptospira</em> infection necessitates deploying a DNA-based method alongside serology. Leptospirosis, IFA, PCR, Golestan, Iran, Diagnosis, Kappa statistics 98 103 http://jommid.pasteur.ac.ir/browse.php?a_code=A-10-26-2&slc_lang=en&sid=1 Saied Reza Naddaf saiedrezanaddaf@gmail.com 0000-0002-0751-8692 Yes Department of Parasitology, Pasteur Institute of Iran, Tehran, Iran Reyhaneh Mohabati 0000-0003-1524-7199 No Department of Virology, Pasteur Institute of Iran, Tehran, Iran Rouhollah Vahabpor 0000-0002-3102-553X No Department of Medical Lab Technology, School of Allied Medical Sciences, Shahid Beheshti University of Medical Sciences, Tehran, Iran Sabah Naeimi 0000-0002-1717-6411 No Department of Parasitology, Pasteur Institute of Iran, Tehran, Iran Sana Eybpoosh 0000-0002-2304-8352 No Department of Epidemiology and Biostatistics, Research Centre for Emerging and Reemerging Infectious Diseases, Pasteur Institute of Iran, Tehran, Iran