Major risk factors and histopathological profile of treatment failure, relapse and chronic patients with anthroponotic cutaneous leishmaniasis: A prospective case-control study on treatment outcome and their medical importance

Abstract

Over the last years, there has been a remarkable increase in the number of unresponsive patients with anthroponotic cutaneous leishmaniasis (ACL) reported worldwide. The primary objective of this study was to explore the role of demographic, clinical and environmental risk related-factors in the development of treatment failure, relapse and chronic cases compared to responsive patients with ACL. Moreover, molecular, histopathological and immunohistochemical (IHC) findings between these forms were explored. This work was undertaken as a prospective and case-control study in southeastern Iran. Culture media and nested PCR were used to identify the causative agent. Univariate multinomial and multiple multinomial logistic regression models and the backward elimination stepwise method were applied to analyze the data. A P<0.05 was defined as significant. Also, for different groups, skin punch biopsies were used to study the histopathological and immunohistochemical (IHC) profile. All samples showed that L. tropica was the only etiological agent in all unresponsive and responsive patients with ACL. Data analysis represented that 8 major risk factors including nationality, age groups, occupation, marital status, history of chronic diseases, duration of the lesion, the lesion on face and presence of domestic animals in the house were significantly associated with the induction of unresponsive forms. The histopathological and immunohistochemical findings were different from one form to another. The present findings clearly demonstrated a positive relation between ACL and distinct demographic, clinical and environmental risk determinants. Knowledge of the main risk factors for ACL infection is crucial in improving clinical and public health strategies and monitor such perplexing factors.

Fig 1. Representative images of various clinical forms of anthroponotic cutaneous leishmaniasis from Kerman district.

A) a responsive form with erythematous nodular indurated plaque on the face, B) a responsive form with a crusted nodule on the hand, C) a responsive form with a crusted plaque on the face, D) a treatment failure form consisting of three ulcerated indurated erythematous plaques on the leg, E) a treatment failure form with an indurated crusted plaque on the leg, F) a treatment failure form with a crusted large plaque on the lip, G) a relapse (lupoid) form with erythematous apple jelly-like indurated papules around and within the scar on the cheek, H) a relapse (lupoid) form with apple jelly nodules around and within scar of the left cheek [21], I) a relapse (lupoid) form with popular lesion around the scar on the left cheek, J) a chronic form with a large erythematous crusted scaling indurated plaque on the right cheek, K) a chronic form with indurated erythematous exudation ulcer of the forearm, L) a chronic form with an ulcerated plaque of elbow area.

Fig 2. Representative agarose gel electrophoresis images.

All Leishmania isolates from patients with anthroponotic cutaneous leishmaniasis areas of Kerman district were identified to be L. tropica species by nested PCR (1 and 2: Leishmania isolates from responsive patients, 3 and 4: Leishmania isolates from treatment failure patients, 5 and 6: Leishmania isolates from relapse patients, 7 and 8: Leishmania isolates from chronic patients, standard Leishmania tropica and distilled water as positive and negative controls, respectively).

Fig 3. Histopathology and immunohistochemical findings in responsive form.

A) H & E staining shows follicular plugging (star) and acanthosis, dermis displaying dense histiocytic and lymphocytic infiltrate (empty square and arrow) and many intracytoplasmic Leishman bodies, B) aggregation of histiocytes by CD68 IHC staining (arrows), C) collection of Langerhans cells in epidermis mostly, and dispersedly upper dermis by CD1a (arrows), D) CD3 lymphocytes infiltrate between histiocytes (arrowheads), E) rarity of positive B lymphocytes by CD20 IHC staining (arrowhead).

Fig 4. Histopathological and immunohistochemical findings in the treatment failure form.

A) H & E staining shows histopathological changes of hyperkeratosis and acanthosis epidermis (double headed arrow) and dermis displaying histiocytic, lymphocytic and fibrohistiocytic cells infiltrate (empty stars), B) collection of Langerhans cells lattice in epidermis mostly, and dispersedly upper dermis by CD1a IHC staining, C) aggregation of histiocytes by CD68 IHC staining (star), D) lymphocytes infiltrate between histiocytes by CD3, E) rarity of positive B lymphocytes by CD20 IHC staining.

Fig 5. Histopathological and immunohistochemical findings in relapse (lupoid) form.

A) H & E staining shows histopathological changes of thin epidermis and non-caseating epithelioid granuloma (empty circle) in the dermis, B) aggregation of histiocytes and Langhans giant cell by CD68 IHC staining (empty star), C) S100 and E) CD1a, network lattice like collection of Langerhans cells in epidermis and dermis and even in the granuloma, D) CD3, T-lymphocytes infiltrate between the histiocytes.

Fig 6. Histopathological and immunohistochemical findings in chronic form.

A) H & E staining shows histopathological changes of dense and diffuse lymphohistiocytic infiltrate (arrowheads) in the dermis, B) aggregation of histiocytes by CD68 IHC staining, C) haphazard distribution of Langerhans cells in epidermis and dermis by CD1a IHC staining, D) presence of few infiltrate of positive B lymphocytes by CD 20 IHC staining, E) heavily infiltrate of CD3 T-lymphocytes between histiocytes.