PCR performance for the diagnosis of cutaneous leishmaniasis caused by Leishmania viannia complex using biopsy samples, compared with exudate samples from skin lesions on filter paper.

BACKGROUND: Cutaneous leishmaniasis (CL) is generally diagnosed by molecular methods, including PCR, using biopsy samples, skin scrapings and clinical exudates. In this study, we assessed the PCR performance for diagnosis of CL using skin of biopsy samples vs PCR of skin lesion exudate samples on filter paper and compared the diagnostic concordance of PCR using both sampling methods. METHODS: We assessed the PCR performance using 80 skin biopsy samples and 80 filter paper samples containing exudates from skin lesions obtained from 74 patients with clinical suspicion of CL in Cusco, Peru. RESULTS: : PCR using skin biopsy samples had superior diagnostic accuracy compared with filter paper PCR (62.5% [50/80] vs 38.7% [31/80], respectively; p˂0.005) and the diagnostic concordance between both sampling methods was 'moderate' (kappa coefficient=0.50, 95% CI 0.98 to 1.0). CONCLUSIONS: PCR using biopsy samples remains the standard for diagnosis of CL.

Estimation of Biological Parameters of Cutaneous Ulcers Caused by Leishmaniasis in an Animal Model Using Diffuse Reflectance Spectroscopy.

Cutaneous leishmaniasis (CL) is a neglected tropical disease that requires novel tools for its understanding, diagnosis, and treatment follow-up. In the cases of other cutaneous pathologies, such as cancer or cutaneous ulcers due to diabetes, optical diffuse reflectance-based tools and methods are widely used for the investigation of those illnesses. These types of tools and methods offer the possibility to develop portable diagnosis and treatment follow-up systems. In this article, we propose the use of a three-layer diffuse reflectance model for the study of the formation of cutaneous ulcers caused by CL. The proposed model together with an inverse-modeling procedure were used in the evaluation of diffuse-reflectance spectral signatures acquired from cutaneous ulcers formed in the dorsal area of 21 golden hamsters inoculated with Leishmanisis braziliensis. As result, the quantification of the model's variables related to the main biological parameters of skin were obtained, such as: diameter and volumetric fraction of keratinocytes, collagen; volumetric fraction of hemoglobin, and oxygen saturation. Those parameters show statistically significant differences among the different stages of the CL ulcer formation. We found that these differences are coherent with histopathological manifestations reported in the literature for the main phases of CL formation.

Role of Histopathology in the Diagnosis of Cutaneous Leishmaniasis: A Case-Control Study in Sri Lanka.

Cutaneous leishmaniasis (CL) displays a spectrum of manifestations clinically and histologically. Then, it becomes a diagnostic challenge and must discern from the other clinical and histological mimics, especially when the Leishman-Donovan bodies are inattentive. In this study, we compared the distinguishing histomorphological characteristics of CL against the other skin diseases with similar clinical and histological features. Skin biopsies of 181 patients, which suspect CL clinically, are evaluated histologically. Pertaining to the first case-control comparison, which performed between skin lesions of CL with or without discernible organisms and the other granulomatous dermatitis, highlighted that the ill-formed coalescent granulomata (OR = 14.83) and diffuse dense dermal plasma cell infiltrate (OR = 74.25) are significantly associated with the skin lesions of CL. The second case-control analysis was between CL without discernible organisms and the other granulomatous dermatitis, and identified a significant association in the presence of ill-formed coalescent granulomata (OR = 16.94) and diffuse dense (>50/HPF) dermal plasma cell infiltrate (OR = 74.5) in the skin lesions of CL. Pertaining to epidermal changes, acanthosis (OR = 2.38), spongiosis (OR = 9.13), and the presence of ulceration (OR = 20.26) are among the major concerns in CL. In conclusion, in the presence of clinical suspicion, dermal granulomata in ill-formed coalescent morphology with high plasma cell density in a diffuse arrangement are positive factors for the diagnosis of CL, especially when the discernible Leishmania amastigotes are absent. Resource utilization such as polymerase chain reaction and other ancillary techniques during the diagnosis of CL can be minimized by using a range of histopathological features and special attention should be focused on this in the future.

A Potential Role for Mononuclear Phagocytes in Cutaneous Ulcer Development in Human Immunodeficiency Virus-Leishmania braziliensis Coinfection.

Skin ulcer development in cutaneous leishmaniasis due to Leishmania braziliensis infection is associated with a mononuclear cell infiltrate and high levels of tumor necrosis factor (TNF). Herein, we show that despite the absence of Leishmania-driven TNF, a cutaneous leishmaniasis patient with acquired immunodeficiency syndrome developed a skin ulcer. The presence of mononuclear phagocytes and high levels of TNF, chemokine (C-C motif) ligand 2 (CCL2), and metalloproteinase-9 in tissue are identified as potential contributors to immunopathology observed in L. braziliensis-infected patients.

Quantification of Leishmania (Viannia) Kinetoplast DNA in Ulcers of Cutaneous Leishmaniasis Reveals Inter-site and Inter-sampling Variability in Parasite Load.

BACKGROUND: Cutaneous leishmaniasis (CL) is a skin disease caused by the protozoan parasite Leishmania. Few studies have assessed the influence of the sample collection site within the ulcer and the sampling method on the sensitivity of parasitological and molecular diagnostic techniques for CL. Sensitivity of the technique can be dependent upon the load and distribution of Leishmania amastigotes in the lesion. METHODOLOGY/PRINCIPAL FINDINGS: We applied a quantitative real-time PCR (qPCR) assay for Leishmania (Viannia) minicircle kinetoplast DNA (kDNA) detection and parasite load quantification in biopsy and scraping samples obtained from 3 sites within each ulcer (border, base, and center) as well as in cytology brush specimens taken from the ulcer base and center. A total of 248 lesion samples from 31 patients with laboratory confirmed CL of recent onset (≤3 months) were evaluated. The kDNA-qPCR detected Leishmania DNA in 97.6% (242/248) of the examined samples. Median parasite loads were significantly higher in the ulcer base and center than in the border in biopsies (P<0.0001) and scrapings (P = 0.0002). There was no significant difference in parasite load between the ulcer base and center (P = 0.80, 0.43, and 0.07 for biopsy, scraping, and cytology brush specimens, respectively). The parasite load varied significantly by sampling method: in the ulcer base and center, the descending order for the parasite load levels in samples was: cytology brushes, scrapings, and biopsies (P<0.0001); in the ulcer border, scrapings had higher parasite load than biopsies (P<0.0001). There was no difference in parasite load according to L. braziliensis and L. peruviana infections (P = 0.4). CONCLUSION/SIGNIFICANCE: Our results suggest an uneven distribution of Leishmania amastigotes in acute CL ulcers, with higher parasite loads in the ulcer base and center, which has implications for bedside collection of diagnostic specimens. The use of scrapings and cytology brushes is recommended instead of the more invasive biopsy.

Imported new world cutaneous leishmaniasis in a traveller from Slovakia.

We present a case of imported leishmaniasis in a 31-year-old woman from Slovakia, who visited the countries of South America for three months in 2011. On 29 and 31 August 2011, she was probably infected with Leishmania parasites in the jungles of Ecuador. Approximately one week after returning to Slovakia, a small papules appeared on patient's left leg. Another wound was found after two weeks. Both ulcers were enlarging. We proved amastigote forms of Leishmania spp. only in repeated dermal scrapings from the edge of the ulcer by Giemsa staining after negative results from examination of a wound scrape and biopsy specimen. We identified the species Leishmania (Viannia) panamensis as a causative agent by using the polymerase chain reaction (PCR) method and subsequent sequencing of the ITS region. Closure of wounds and scab formation were observed after 20 days of treatment with sodium stibogluconate. In the control microscopic examination after the end of the treatment, parasites were not present, and the PCR confirmed the negative result (Fig. 2, Ref. 31).

Ulcerative penile leishmaniasis in a child.

Penile ulcers may be caused by several different agents. Rarely, cutaneous leishmaniasis may also be accompanied by penile ulcers. We report a five-year-old boy with who had an ulcer on the glans penis. Smears from the ulcer demonstrated amastigotes, biopsy showed histopathological features of leishmaniasis and Leishmania was grown in culture. Treatment with meglumine antimoniate injections led to improvement.

Comparative analysis of the tissue inflammatory response in human cutaneous and disseminated leishmaniasis

Cutaneous leishmaniasis (CL) is the most frequent clinical form of tegumentary leishmaniasis and is characterised by a single or a few ulcerated skin lesions that may disseminate into multiple ulcers and papules, which characterise disseminated leishmaniasis (DL). In this study, cells were quantified using immunohistochemistry and haematoxylin and eosin staining (CD4+, CD68+, CD20+, plasma cells and neutrophils) and histopathology was used to determine the level of inflammation in biopsies from patients with early CL, late CL and DL (ulcers and papules). The histopathology showed differences in the epidermis between the papules and ulcers from DL. An analysis of the cells present in the tissues showed similarities between the ulcers from localised CL (LCL) and DL. The papules had fewer CD4+ T cells than the DL ulcers. Although both CD4+ cells and macrophages contribute to inflammation in early CL, macrophages are the primary cell type associated with inflammation intensity in late ulcers. The higher frequency of CD20+ cells and plasma cells in lesions demonstrates the importance of B cells in the pathogenesis of leishmaniasis. The number of neutrophils was the same in all of the analysed groups. A comparison between the ulcers from LCL and DL and the early ulcers and papules shows that few differences between these two clinical forms can be distinguished by observing only the tissue.

Comparative analysis of the tissue inflammatory response in human cutaneous and disseminated leishmaniasis.

Cutaneous leishmaniasis (CL) is the most frequent clinical form of tegumentary leishmaniasis and is characterised by a single or a few ulcerated skin lesions that may disseminate into multiple ulcers and papules, which characterise disseminated leishmaniasis (DL). In this study, cells were quantified using immunohistochemistry and haematoxylin and eosin staining (CD4+, CD68+, CD20+, plasma cells and neutrophils) and histopathology was used to determine the level of inflammation in biopsies from patients with early CL, late CL and DL (ulcers and papules). The histopathology showed differences in the epidermis between the papules and ulcers from DL. An analysis of the cells present in the tissues showed similarities between the ulcers from localised CL (LCL) and DL. The papules had fewer CD4+ T cells than the DL ulcers. Although both CD4+ cells and macrophages contribute to inflammation in early CL, macrophages are the primary cell type associated with inflammation intensity in late ulcers. The higher frequency of CD20+ cells and plasma cells in lesions demonstrates the importance of B cells in the pathogenesis of leishmaniasis. The number of neutrophils was the same in all of the analysed groups. A comparison between the ulcers from LCL and DL and the early ulcers and papules shows that few differences between these two clinical forms can be distinguished by observing only the tissue.

Dermoscopic features of cutaneous leishmaniasis.

BACKGROUND: Previous studies have demonstrated that dermoscopy improves accuracy in diagnosing pigmented and non-pigmented tumors. Recently, there has been increasing evidence that dermoscopy can also be useful in the diagnosis of some skin infections. OBJECTIVE: We sought to describe the dermoscopic features of cutaneous leishmaniasis. METHOD: Dermoscopic examination (using the Derm Lite FOTO, 3Gen) of 144 CL lesions in 82 patients was performed. This study was conducted in the Imam Reza and Ghaem hospitals (Mashhad, Iran) from October 2010 to September 2011. RESULTS: We observed the following dermoscopic features: generalized erythema (81.9%), white starbursts (60.4%), yellow hue (43.8%), yellow teardrop-like structures (41.7%), central ulcers (59%), hyperkeratosis (33.3%), and milia-like cysts (4.9%). We also observed vascular structures, including dotted vessels (61.1%), hairpin vessels (37.5%), linear irregular vessels (30.6%), comma-shaped vessels (29.9%), glomerular vessels (22.9%), arborizing telangiectasia (10.4%), and corkscrew vessels (4.2%). LIMITATIONS: Biopsy and pathologic evaluation were not performed due to ethical considerations. CONCLUSION: Important vascular patterns seen in melanocytic and non-melanocytic tumors were frequently observed in this infection. Dermoscopy may be a promising tool to predict the clinical course in cases of cutaneous leishmaniasis.

Cardiac echinococcosis associated with cerebrovascular occlusive disease and subcutaneous bullous eruptions and ulcers.

Cardiac involvement is an uncommon complication of echinococcosis. It is usually asymptomatic and may only be diagnosed incidentally. A 7-year-old boy was admitted with acute stroke and bullous and ulcerated skin lesions. He was diagnosed with cardiac echinococcosis complicated by systemic emboli to the central nervous system and superficial cutaneous arteries. In endemic areas, echinococcosis should be considered in the differential diagnosis of cardiac disease and unexplained cerebral embolism.

Reappraisal of the immunopathogenesis of disseminated leishmaniasis: in situ and systemic immune response.

Disseminated leishmaniasis (DL) is an emerging form of Leishmania braziliensis infection characterised by multiple cutaneous lesions on different parts of the body and a high rate of mucosal involvement. Systemic production of TNFα and IFNγ in DL patients is lower than in cutaneous leishmaniasis (CL) caused by L. braziliensis, which may account for parasite dissemination due to the decreased ability to control parasite growth. In this study, the systemic and in situ immune response of DL and CL patients was characterised through evaluation of chemokine and cytokine production. In situ evaluation showed similar production of IFNγ, TNFα, IL-10, transforming growth factor-beta (TGFß), chemokine (C-C motif) ligand 2 (CCL2), CCL3, CCL11 and chemokine (C-X-C motif) ligand 10 (CXCL10) in papular and ulcerative lesions from DL as well as in ulcerated lesions from CL. Serum levels of CXCL9, a chemokine that attracts T-cells, was higher in serum from DL than from CL. These data indicate that a decrease in the type 1 immune response in peripheral blood of DL patients is due to attraction of Leishmania antigen-activated T-cells to the multiple cutaneous lesions. This may account for the absence of or few parasites in the lesions and for the development of ulcers similar to those observed in CL.

A cluster of cutaneous leishmaniasis associated with human smuggling.

Cutaneous leishmaniasis (CL) is rarely seen in the United States, and the social and geographic context of the infection can be a key to its diagnosis and management. Four Somali and one Ethiopian, in U.S. Border Patrol custody, came to the United States by the same human trafficking route: Djibouti to Dubai to Moscow to Havana to Quito; and then by ground by Columbia/Panama to the United States-Mexico border where they were detained. Although traveling at different times, all five patients simultaneously presented to our institution with chronic ulcerative skin lesions at different sites and stages of evolution. Culture of biopsy specimens grew Leishmania panamensis. Soon thereafter, three individuals from East Africa traveling the identical route presented with L. panamensis CL to physicians in Tacoma, WA. We document here the association of a human trafficking route and new world CL. Clinicians and public health officials should be aware of this emerging infectious disease risk.

Inmunoterapia de la leishmaniasis cutánea: factores que influencian su efectividad / Immunotherapy of cutaneous leishmaniasis: factors that influence their effectiveness

En Venezuela se utiliza la inmunoterapia por su bondad terapéutica y operacional en el tratamiento de la leishmaniasis cutánea y se aprecian diferencias de su efectividad entre los servicios locales de salud que la emplean. En el estado Mérida, donde se tiene un bajo nivel de falla terapéutica, se desconocen los factores de riesgo ó de protección que la determinan. Por ello se planteó realizar un estudio epidemiológico de casos y controles para evaluar los factores individuales demográficos, clínicos, inmunológicos y adherencia terapéutica que influyen en dicha efectividad. El ajuste con regresión logística determinó según definición: a) Demográfica: como riesgo las edades extremas, tabaquismo y de oficio doméstico, con protección en el alfabeto educativamente, b) Clínica: como riesgo la forma intermedia, cinco o más lesiones, infección secundaria y ubicación en pié, con protección en la forma localizada y tamaño menor a 60 mm, c) Inmunológica: de riesgo las bajas respuestas a leishmánina y PPD, d) Adherencia terapéutica: son riesgo la aplicación tópica y aseo local inadecuados. Se concluye, hay factores de riesgo y protección que modelan la eficacia de la inmunoterapia, lo que en consecuencia demanda una dinámica vigilancia clínico-epidemiológica para potenciar dicha terapéutica.

Immunotherapy is used in Venezuela as treatment of cutaneous leishmaniasis and there are differences among the health services that use it. In Merida State there are a low proportion of failures, but the factors that are related to this failure are unknown. A case-control study was planned to evaluate the demographics, clinical, immunological factors and the treatment compliance that can be related to the effectiveness. Logistic regression showed that the factors related to failure were: a) demographics: lower and older ages, smoking, domestic labor and illiteracy; b) clinical: intermediate leishmaniasis, five or more lesions, aggregated infection, lesions in feet, and lesion size above 60 mm; c) Immunology: low reactivity to Montenegro and tuberculin tests; d) treatment compliance: use of incorrect topical substances and inadequate cleaning of ulcer. As a conclusion, there are several factors that influence treatment response, that require clinical and epidemiological surveillance to increase the effect of therapy.

A chronic mutilating rhinopathy with a delayed diagnosis of mucocutaneous leishmaniasis.

BACKGROUND: Mucocutaneous leishmaniasis is a granulomatous disease clinically characterized by ulcerated skin and mucosal lesions. Mucocutaneous leishmaniasis is very rare in India and to our knowledge, only two cases have been reported, and this is the first case of mucocutaneous leishmaniasis presenting with mutilating rhinopathy reported from the Indian subcontinent. CASE REPORT: A 64-year-old man presented with a destructive ulceration of the central face of 23 years' duration, who was diagnosed to have mucocutaneous leishmaniasis, and showed dramatic response to intramuscular injections of sodium stibogluconate. RESULTS: Histopathologic examination of skin biopsy revealed a granulomatous infiltrate with the presence of leishmania donovani (LD) bodies. The clinical picture, plus the pathologic findings, and the response to sodium stibogluconate confirmed mucocutaneous leishmaniasis. CONCLUSION: Mucocutaneous leishmaniasis is a rare disease in the Indian subcontinent, and clinicians from this region should have a high index of suspicion on encountering mucocutaneous ulcerative lesions.