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The patients with bigger skin lesion in CL and higher level of immunoglobulin or splenomegaly in VL, had the higher parasite load detected by qPCR. Two VL patients with parasite converted to microscopically negative after treatment were detected positive with qPCR. Total 20 skin lesion samples from current CL patients and 20 bone marrow and/or PBMC samples from current VL patients were all detected as positive with qPCR without cross-reaction with samples from patients with malaria, brucellosis and dengue or normal donors. This method was clinically applied to quantitatively detect skin lesion samples from 20 cutaneous leishmaniasis (CL) and bone marrow and/or PBMC samples from 30 current and cured visceral leishmaniasis (VL) patients, and blood samples from 11 patients with other infections and 5 normal donors as well. It also detected Pan- Leishmania protozoa including Leishmania donovani, Leishmania infantum and Leishmania major without cross-reaction with other pathogen DNAs available in our lab. The designed mkDNA-based qPCR was able to detect as low as one copy of Leishmania mkDNA or DNA from single parasite.
was developed to detect different species of Leishmania. Thus, a quantitative real-time PCR (qPCR) based on the conversed regions of kinetoplast minicircle DNA (mkDNA) of Leishmania spp. There is an urgent need for developing a sensitive and easily operated diagnostic method for the diagnosis and disease management of leishmaniasis. The low sensitivity and inconvenience of microscopic examination could cause misdiagnosis and relapse of leishmaniasis. At present, the clinical diagnosis and treatment monitoring still rely on parasitological culture and microscopy that needs experienced technicians. Leishmaniasis is still a serious neglected tropical disease that may cause death in infected individuals. 4Department of Pediatrics and National School of Tropical Medicine, Baylor College of Medicine, Houston, TX, United States.3Warwick Medical School, Warwick University, Coventry, United Kingdom.2Beijing Institute of Tropical Medicine, Beijing, China.1Emergency and Critical Care Center, Beijing Friendship Hospital, Capital Medical University, Beijing, China.Waterfield 3, Bin Zhan 4, Lei Wang 1,2 * and Guowei Yang 1,2 * Yun Wu 1,2 †, Xiaojun Tian 1,2 †, Nan Song 1,2, Minjun Huang 1,2, Zhaoyong Wu 1,2, Shaogang Li 1,2, Nicholas R.