Medicine
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This repository contains the published and unpublished research of the Faculty of Medicine by the staff members of the faculty
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Item Diagnosing human cutaneous leishmaniasis using fluorescence in situ hybridization(Taylor & Francis Publishing, 2021) Kaluarachchi, T.J.; Wickremasinghe, R.; Weerasekera, M.; Yasawardene, S.; McBain, A.J.; Yapa, B.; de Silva, H.; Menike, C.; Jayathilake, S.; Munasinghe, A.; Wickremasinghe, R.; Ranasinghe, S.ABSTRACT: Cutaneous leishmaniasis (CL) is endemic in Sri Lanka. Giemsa-stained slit-skin-smears (SSS-Giemsa) and histology are routinely used in diagnosis with a sensitivity of 40-70%. PCR currently has limited accessibility. Therefore, we assessed the sensitivity and specificity of a previously described fluorescence in situ hybridization assay, on skin smears and biopsy samples to overcome the limitations encountered with routine diagnostic methods.Samples from a total of 123 suspected CL patients were collected and subjected to SSS-Giemsa, fluorescence in situ hybridization (FISH) on slit skin smears (SSS-FISH), formalin-fixed-paraffin-embedded-tissues stained with Hematoxylin & Eosin staining (FFPE-H&E) and FISH on formalin-fixed-paraffin-embedded-tissues (FFPE-FISH). Negative controls of 61 patient samples were collected from a CL non-endemic area and subjected to the same procedures. The gold standard PCR was used as a comparator. For FISH, two previously described cyanine 3 tagged Leihsmania genus-specific probes were used.Compared to PCR, SSS-Giemsa, SSS-FISH, FFPE-H&E, and FFPE-FISH had sensitivities of 76.5%, 79.1%, 50.4% and 80.9%, respectively. Routine diagnostic tests (SSS-Giemsa and FFPE-H&E) had a specificity of 100%. SSS-FISH and FFPE-FISH had specificities of 96.7% and 93.4%, respectively. FFPE-FISH had a statistically significant higher diagnostic performance than FFPE-H&E (p < 0.001). The relative performance of SSS-Giemsa, SSS-FISH and FFPE-FISH was similar (p > 0.05 for all comparisons).We conclude that FFPE-FISH is a more accurate diagnostic tool than FFPE-H&E. SSS-FISH did not have an additional advantage over SSS-Giemsa in diagnosis. However, SSS-FISH could be recommended as a minimally invasive method in studies assessing wound healing where immunological probes are used. KEYWORDS: Cutaneous leishmaniasis; Sri Lanka; fluorescence in situ hybridization.Item Pathogenesis of Leishmania donovani induced cutaneous leishmaniasis: role of Tumor Necrosis Factor α(Faculty of Medicine, University of Kelaniya, Sri Lanka, 2016) Manamperi, N.H.; Oghumu, S.; Pathirana, K.P.N.; Munidasa, U.A.D.D.; Somaratne, K.K.V.N.; Rathnayake, R.M.D.I.; Thilakaratne, S.M.; de Silva, M.V.C.; Pathmeswaran, A.; Abeyewickreme, W.; Satoskar, A.R.; Karunaweera, N.D.BACKGROUND: Cutaneous leishmaniasis (CL) in Sri Lanka is caused by the usually visceralizing Leishmania donovani. Host immune response plays a key role in the clinical presentation of leishmaniasis. Role of cytokines in pathogenesis of local lesions has not been studied. OBJECTIVE: To describe tissue cytokine expression with lesion progression with time in CL due to Leishmania donovani. METHODS: Skin biopsies from fifty eight patients with parasitologically or histopathologically confirmed CL and 30 healthy controls were analyzed for local tissue expression of Interleukin (IL)-12A, IL-4, IL-10, Interferon-gamma (IFNg) and Tumor Necrosis Factor-alpha (TNF-α). Cytokine mRNA was quantified by real-time RT- PCR using SYBR green. Relative copy numbers were calculated for each gene by 2-ΔΔCt method using β-actin as the reference gene and healthy controls as the calibrator. Spearman correlation was used to determine the correlation between cytokines and duration of active skin lesions. RESULTS: The study group consisted of 37 males (63.8%) and 21 females (36.2%) with a mean age of 35 (SD=12.05) years which ranged between 18-66 years. Mean duration of lesions was 6.8 (SD=9.10) months with a range of 1-48 months. The Spearman correlation coefficient for relative copy numbers and lesion duration was 0.220, 0.077, 0.073, 0.235 and 0.295 for IL-12A, IL-4, IL-10, IFNg and TNF-α respectively. There was a significant positive correlation between expression of TNF-α and lesion duration (p= 0.024). CONCLUSIONS: Localized expression of TNF-α increases with time in CL due to L. donovani in Sri Lanka, which indicates development of a pro-inflammatory immune response at the site of infection as the disease progresses.