Molecular detection and epidemiological correlates of Leishmania tropica and Leishmania major in clinical samples by using Nested PCR in Wasit, Iraq

Objective: Cutaneous leishmaniasis (CL) is an endemic parasitic disease in Iraq and across the Middle East, posing a persistent epidemiological and public health challenge. This study aimed to (i) evaluate the diagnostic performance of nested polymerase chain reaction (Nested PCR) in detecting Leishmania tropica and Leishmania major, (ii) compare its accuracy with conventional microscopy, and (iii) assess potential associations with demographic and clinical variables. Methods: A total of 69 clinical samples obtained from skin lesions were analyzed. DNA was extracted using a commercial kit and amplified by Nested PCR targeting kinetoplast DNA (kDNA). Gel electrophoresis confirmed amplification, and statistical analyses (T-test and Chi-square test) were applied to examine correlations between species distribution and demographic or clinical factors. Results: Nested PCR identified Leishmania DNA in 30 samples (43.5%), including 17 L. tropica and 13 L. major. Compared with PCR as the gold standard, microscopy demonstrated 100% sensitivity but lower specificity (84.6%). The highest infection rate occurred among children aged 1–10 years, with slightly higher prevalence among females, O+ blood group carriers, unemployed individuals, and those with lower education levels. Most patients presented with single, nodular lesions primarily affecting the extremities. However, statistical analyses revealed no significant associations between parasite species and demographic or clinical variables (p > 0.05). Conclusion: Nested PCR demonstrated superior sensitivity in diagnosing CL and differentiating between species, highlighting its advantage over conventional methods. These findings underscore the importance of expanding molecular surveillance and implementing region-specific control programs to reduce the burden of CL in endemic regions.