Clinical, Histopathologic, and Cytologic Diagnosis of Mucosal Leishmaniasis and Literature Review

Leishmaniasis is endemic in Iran. Leishmania can involve almost any organ.1 It can present as cutaneous leishmaniasis (CL), localized leishmania lymphadenitis, mucosal leishmaniasis (ML), or diffuse multiorgan involvement (kala azar). Fine-needle aspiration biopsy of lymph nodes, bone marrow aspiration, splenic puncture, and skin-scraping cytology for diagnosis of leishmaniasis have previously been described.2 Demonstration of the parasite is necessary for the diagnosis of leishmaniasis. Cytologically, the cytoplasm of a typical Leishman-Donovan body contains a nucleus and a kinetoplast.1 The complete form of a Leishman-Donovan body can be seen in aspiration of bone marrow, lymph nodes, and skin smears. However, in histology sections the kinetoplasts are not visible, and the organisms can be easily mistaken for tingible bodies, fungal elements, toxoplasmosis, histoplasmosis, or artifacts.1,2 Mucosal lesions usually appear as ulcerations in the nose and mouth. However, they can affect any site and also may present as exophytic, nodular, and indurated lesions that can mimic a malignant lesion.3,4 Reports on cytologic diagnosis of ML are limited.3,4 Herein 11 patients with ML for whom cytology helped determine the correct diagnosis are presented. The literature on ML is reviewed.

Records of 11 patients diagnosed with ML were retrieved from our files. Of these patients, 7 were patients with oral lesions, one of whom was known to have oral leishmaniasis with recurrence of oral lesions; 2 had laryngeal lesions; and 3 had nasal lesions. One case of laryngeal leishmaniasis was recurrence of prior oral lesions. The patients were from different urban areas of Fars and Kerman provinces in southern and southeastern Iran. Scraping cytology was performed using a scalpel in 7 cases of oral leishmaniasis. The cytobrush was used in 1 case of nasal mucosa and in 2 cases of laryngeal ML. In 2 patients with nasal lesions, exfoliative cytology was made by washing the nasal cavity.5 In 6 patients, cytology was carried out when biopsies yielded inconclusive findings. Multiple smears were made on slides and were both air dried and alcohol fixed and stained by Wright and Papanicolaou stains, respectively. Tissue samples from the lesions were fixed in 10% neutral buffered formalin, embedded in paraffin, sectioned at 5-µm thickness, and stained with hematoxylin-eosin. Review of histologic sections (7 cases) and cytologic smears (11 cases) was conducted blindly by 7 pathologists.

The clinical and cytopathologic data of 11 patients, 10 men and 1 woman, with an age range of 21 to 45 years are explained. Cytologic examination results of the lesions in 8 patients was reported as leishmaniasis and as suggestive for leishmaniasis for another patient reported. In 5 patients, because of either clinical (Figure 1, A) or histologic findings, a malignant tumor was suspected but cytology helped to diagnose leishmaniasis. In one of the cases, which was followed for 7 years in a rheumatology clinic, the lesions were not typical of ML (Figure 1, B). Scraping cytology showed atypical organisms, so diagnosis was reported as suggestive of leishmaniasis, and after treatment for leishmaniasis, the lesions disappeared. However, 6 months later this patient showed recurrence with typical oral lesions (Figure 1, C) and leishmaniasis was confirmed by cytology. In a known patient with ML who presented with hoarseness, direct laryngoscopy showed vocal cord paralysis (absence of abduction on phonation), with arytenoid and epiglottis swelling and erythema, and cytobrush of the lesions confirmed ML. Cytologic findings consisted of granuloma, macrophages loaded with Leishman-Donovan bodies (Figure 2, A), multinucleated giant cells (Figure 2, B), binucleated-histiocyte (Reed-Sternberg–like) cells loaded with Leishman-Donovan bodies, free Leishman-Donovan bodies, acute and chronic inflammatory cells, atypical organisms, histiocytes, plasma cells and mast cells (Figure 2, C), Leishman-Donovan bodies in the vicinity of respiratory epithelium in nasal smears, and atypical intracytoplasmic Leishman-Donovan bodies with ballooning changes. The cytologic findings are summarized in Table 1.

Of the 7 patients who underwent tissue biopsies, the biopsy of only 1 was initially reported as suggestive of leishmaniasis. The other 6 biopsies were inconclusive and reported as granuloma, inflammation, ulceration, and 1 as lymphoma. However, after retrospective review of these sections, Leishman-Donovan bodies were detected in 3 of them. The histologic findings after review were ulceration and granulation tissue formation, granuloma, multinucleated giant cells with many intracytoplasmic Leishman-Donovan bodies, numerous multinucleated giant cells, severe lymphoplasma cells, infiltrating perineural tissue and muscles, pseudoepitheliomatous hyperplasia with keratin pearls, lymphoid aggregation in the submucosal area, lymphoid aggregation between skeletal muscles, and massive infiltration of macrophages loaded with Leishman-Donovan bodies in submucosal areas of oral (Figure 3), nasal, and respiratory epithelium. Histologic findings are summarized in Table 2.

Leishmaniasis is a protozoan disease caused by the genus Leishmania and is transmitted by sandfly vectors.1–3 Leishmaniasis is found worldwide, and is considered to be endemic in 88 countries.6 It is estimated that 350 million people are at risk, 12 million are already infected, and 1.5 to 2 million are infected annually.7 Leishmaniasis is classified as CL, ML, localized leishmania lymphadenitis, and visceral or kala azar, with a wide spectrum of clinicopathologic manifestations.1,2 Most cases of CL occur in Iran, Afghanistan, Syria, Saudi Arabia, Brazil, and Peru.8 In Iran, leishmaniasis is endemic in the southern and southeastern regions in which Fars and Kerman provinces are located. Mucosal involvement in leishmaniasis is uncommon, and results from hematogenous or lymphatic dissemination of amastigotes from the skin of 5% of the patients affected with CL to the naso-oropharyngeal mucosa and upper respiratory tract.9 Mucosal leishmaniasis is a destructive disease that predominantly affects the nose and does not heal spontaneously; it usually evolves during the course of several years before the patient seeks medical attention.4 The reason why ML subsequently develops in only a few patients with CL is unclear. In most cases, ML becomes evident some years or even decades after resolution of the preceding cutaneous lesions, but it can develop while the skin lesions are still present.9 The mucous membrane over the cartilaginous septum inside the nares is a common site for these lesions; the affected nasal septum may subsequently be perforated. As the disease progress, extensive tissue destruction within the mucosal and submucosal tissues of the nose and mouth may occur.10 Oral lesions usually appear as ulceration in the hard or soft palate.11 However, they can affect any site, and may present as exophytic, nodular, and indurated lesions that can mimic a malignant lesion.7 Mucosal leishmaniasis is characterized by an intense inflammatory reaction and tissue damage with few parasites in the lesion.12 Compared with CL, amastigote forms of the parasite are characteristically sparse in ML, so that Giemsa staining might not detect the organisms.9,10 The paucity of parasites in the destructive mucosal lesions highlights the current poor understanding of the pathogenesis of ML. This form of leishmaniasis is highly prevalent in Latin America13 and its occurrence is rare in Iran.14 ,Leishmania tropica and L major are reported as the causative agents of leishmaniasis in Fars Province, Iran,14 and it is more likely that some of the patients with ML in this study were infected with the same organisms. However, the question remains whether the lesions described represent the primary site of the parasite inoculation or are due to migration of the parasite from elsewhere.4,14 Most of the patients in the present study showed only mucosal involvement (8/11; 72.7%), and the clinical and histopathologic features were similar to those reported earlier.3,4 

ML is a public health problem and causes significant morbidity and mortality. In recent years, economic globalization and increased travel have extended its reach to people in developed countries.15,16 Therefore, an easy and reliable method for diagnosis of ML may be valuable. Review of the literature shows numerous reports3,4,11,16–25 on ML (Table 3).

Several methods have been used for the diagnosis of leishmaniasis, including serology, culture, molecular methods, and inoculation into animals.26–28 Application of immunohistochemistry technique is a useful tool at the light microscopic level to confirm the diagnosis, because the histopathologic findings usually show nonspecific chronic inflammation and/or a granulomatous reaction with very few parasites that are difficult to confirm.23 In the tissue, leishmaniasis is characterized by a subepithelial nonnecrotizing granulomatous inflammatory reaction with lymphocytes, plasma cells, and histiocytes as the predominant cells. Characteristic features of the amastigote, the intracytoplasmic kinetoplast and a rod-shaped mitochondrial structure composed of DNA, are helpful diagnostic criteria in cytology sections of patients with leishmaniasis.9 These kinetoplasts are not appreciated in histology sections. Intracytoplasmic inclusions in the histiocytes seen in hematoxylin-eosin staining may suggest the possibility of leishmania; however, other organisms such as Toxoplasma and Histoplasma, artifacts, and tangible body macrophages must also be considered.1,2 The identification of leishmania is possible by Giemsa stain touch imprint cytology and/or specific antibodies on tissue.19 Nevertheless, ML can be difficult to diagnose, even when clinically active, because amastigotes usually are scarce. To our knowledge, there are only 2 reports in which cytology has been conducted as the diagnostic method for ML identification.3,4 

In conclusion, clinically or histologically ML can be mistaken for benign or malignant lesions. In this study, cytology is suggested as the first method for patients with suspected ML. It is an easy, cost-effective diagnostic method that can be conducted even after inconclusive or nonspecific pathologic findings from biopsies.

The authors thank Ricardo H. Bardales, MD, for his review and comments.