Possible clinical implications and future directions of managing bacterial biofilms in cutaneous leishmaniasis wounds.

Cutaneous leishmaniasis (CL) lesions are chronic and result in disfiguring scars. The microbiological aspects of these wounds have not been systematically investigated. We have recently reported that 61.5% of CL wounds in a Sri Lankan cohort harboured bacterial biofilms, mainly composed of bacilli, Enterobacteriaceae, and Pseudomonas, which could delay wound healing. We have additionally reported that biofilms were significantly associated patients over 40 years of age, discharge, pain and/or itching of the wound, and high pus cell counts. Using this as background knowledge and other relevant literature, we highlight the importance of investigating the role of biofilms in CL wound healing, clinical indicators, cost-effective laboratory tests involving less invasive sampling techniques for diagnosing biofilms and potential therapeutic options for biofilm-containing CL wounds, such as adjunctive application of wound debridement and antimicrobial treatment along with anti-parasitic drugs.

Distinct microbiome profiles and biofilms in Leishmania donovani-driven cutaneous leishmaniasis wounds.

The endemic strain of Leishmania donovani in Sri Lanka causes cutaneous leishmaniasis (CL) rather than more common visceral form. We have visualized biofilms and profiled the microbiome of lesions and unaffected skin in thirty-nine CL patients. Twenty-four lesions (61.5%) were biofilm-positive according to fluorescence in situ hybridization. Biopsies of biofilm-positive lesions were dominated by Pseudomonas, class Bacilli and Enterobacteriaceae and distinguished by significantly lower community evenness. Higher relative abundance of a class Bacilli OTU was detected in wound swabs versus contralateral skin. Wound swabs and biopsies had significantly distinct microbiome profiles and lower diversity compared to unaffected skin. Greater abundances of potentially pathogenic organisms were observed in wet ulcers, lesions with high parasite loads and large wounds. In summary, more than half of L. donovani associated CL wounds harboured biofilms and the wounds exhibited a distinct, less diverse, microbiome than unaffected skin.

Diagnosing human cutaneous leishmaniasis using fluorescence in situ hybridization.

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.

Therapeutic Response to Thermotherapy in Cutaneous Leishmaniasis Treatment Failures for Sodium Stibogluconate: A Randomized Controlled Proof of Principle Clinical Trial.

Treatment failure to intralesional sodium stibogluconate (IL-SSG) is a health challenge for cutaneous leishmaniasis (CL) in Sri Lanka. A randomized controlled proof of principle clinical trial, with two arms (viz., radio frequency-induced heat therapy [RFHT] by a ThermoMed™ device (Model 1.8, Thermosurgery Technologies, Inc., Phoenix, AZ) and thermotherapy by a handheld exothermic crystallization thermotherapy for CL [HECT-CL] device) was conducted on 40 CL treatment failures to IL-SSG, from three hospitals in Tangalle, Hambantota, and Anuradhapura, from January 2017 to January 2018, followed up for 180 days post-thermotherapy with a final follow-up in February 2020. Intention-to-treat cure rates were calculated at day 90 (initial cure rate) and at day 180 (final cure rate) posttreatment. Radio frequency-induced heat therapy group: the initial cure rate was 100% (20/20) and the final cure rate was 95% (19/20), with one patient relapsing. The HECT-CL group: both the initial and final cure rates were 80% (16/20), with no relapses and one excluded from the trial. In February 2020 (1.6-3 years posttreatment), 27 traceable patients (RFHT = 16, HECT-CL = 11) remained healed. Second-degree burns were observed with RFHT in 65% (13/20), with HECT-CL in 15% (3/20), which completely resolved subsequently. The cure rates between the two treatment groups were comparable (P = 0.15). Radio frequency-induced heat therapy consumed less time and required only a single hospital visit. Handheld exothermic crystallization thermotherapy for CL is potentially usable at community settings with both being less costly than IL-SSG. This study is the first proof that thermotherapy is an efficacious and safe treatment for CL patients in Sri Lanka, complicated by treatment failure to IL-SSG.

Vaginal stenosis after toxic epidermal necrolysis.

A 17-year-old adolescent girl developed severe toxic epidermal necrolysis after inhalation of methamphetamine. Recovery was complicated by vaginal synechiae and occlusion necessitating surgery.