Histopathological spectrum in cutaneous leishmaniasis: A study in Oman

Introduction

Cutaneous leishmaniasis (CL) is an important public health problem in several parts of the world, including East. In Oman, as other parts of Middle East, Leishmania tropica has been reported to be the strain causing cutaneous leishmaniasis. I The disease presents a spectrum of manifestations clinically, as the lesion undergoes different morphological stages such as a nodule, plaque and ulcer, which may either persist as a chronic lesion or heal with a scar. The lesion may later relapse as a recidivans lesion.[2],[3] The confirmation of diagnosis, though obvious clinically to an experienced practitioner in an endemic area, is by examination of smears from the lesions, culture on NNN medium and histopathological examination. Several previous studies have established the value of histopathological examination in the diagnosis in CL and different histological reaction patterns have been reported.[2],[1] We report the results of our study of 40 cases from the Sultanate of Oman.

Materials and Methods

Biopsies from 40 cases of cutaneous leishmaniasis, diagnosed during the years 94-98 in the Dept of Skin, Al Nahdha hospital, the main referral hospital for skin diseases in the Sultanate of Oman were studied. All cases had been confirmed by clinical examination, smear studies, culture and response to specific antileishmania treatment. The patient population included 27 male and 13 female patients, in the age group 4 to 35 years. The duration of lesions varied from 3 weeks to 18 months. All cases had localized cutaneous leishmaniasis. The lesions were clinically grouped into early acute lesions (of duration less than 3 months), early subacute lesions (duration 3 months-12 months) and late lesions (duration more than 12 months). The different clinical lesions were noted as nodule, plaque, crusted plaque, noduloulcer and ulcer. Smears by slit and scrape method from the lesions for Leishmanoid forms were examined with Giemsa stain. Culture was performed on NNN medium. All cases were treated with injections of sodium stibogluconate, 10-20 mg/kg wt, with complete resolution of lesions. Biopsy specimens were examined by H & E for histopathology and Giemsa stains for LD bodies. Special stains such as Fite Faraco, and PAS stains for other organisms were also examined. The number of LD bodies were graded on a scale (modification of Ridley′s parasitic index 1983) as ; - (no organism), + (1-9 organisms per standard section); ++ (10-100 organisms per standard section);+ + + (>100 organisms per standard section).

Results

[Table - 1]

Dermal reactions: Four different histological types were identified;

1. Type-1 pattern consisted of a diffuse dermal infiltrate composed predominantly of macrophages with an admixture of few polymorphs and lymphocytes [Figure - 1]. Eosinophils were rare. Plasma cells were seen occasionally. The sections showed plenty (+ + +) of LD bodies, which were apparent with both H& E as well as Giemsa stains. The LD bodies, of size 2-4u, consisted of a nucleus and a smaller kinetoplast, were present within macrophages and occasionally extracellularly. This pattern was seen in 18 (45%) cases.

2. Type-2 pattern consisted of an admixture of focal infiltration of macrophages, polymorphs and plasma cells. There were few lymphocytes. The striking feature of this pattern was the presence of necrosis, seen as haematophilic necrotic material, nuclear debris and karryorrhexis [Figure - 2]. Number of LD bodies was much less than 1 pattern, with 10-100 (+ +) organisms seen in a standard section. This pattern was seen in 11/40 (27.5%) of patients.

3. Type.3 pattern consisted of early granuloma formation with focal collection of epitheloid cells, lymphocytes, and few plasma cells Figure:3. There were no giant cells. Necrosis was not prominent. There were only a few (+) LD bodies seen. This pattern was seen in 6/40 (15%) cases.

4. Type-4 pattern showed well formed epitheloid granuloma in dermis.There were plenty of lymphocytes, epitheloid cells and Langhan′s type of giant cells Figure:4. Caseation was not seen. No plasma cells or LD bodies were present. This pattern was seen in 2/40 (5%) cases.

In addition, a nonspecific pattern consisting of a patchy dermal mononuclear infiltrate of lymphocytes, few macrophages and plasma cells was seen in 3/40 (7.5%) cases. LD bodies Figure:5 could be demonstrated in the dermis in 65% of all cases. Other dermal features included oedema, vasodilatation, endothelial hyperplasia, and periadnexal infiltrate.

Epidermal changes Most common epidermal changes were hyperkeratosis and, acanthosis (14 cases). Parakeratosis was less common (6 cases). Other changes included (11 cases), basal cell degeneration (6 cases) and atrophy of epidermis (4 cases). Intraepidermal abscesses (consisting of polymorphs in hyperplastic rete ridges), were seen in lesions with types-1 and 2 patterns. Basal cell degeneration, contiguous to the dermal infiltrate was also seen in lesions with type-1 and type-2 patterns. Epidermal atrophy was noted in cases with type-4 and nonspecific patterns. Intraepidermal organisms were not seen in any of the cases.

Clinico-pathological correlation Clinical type of the lesion and the duration of lesion was correlated with the histological type of reaction [Table - 1].

Acute lesions (less than 3 months duration); Clinically these were plaques/ nodules, which were often furunculoid. There were 24 cases in this category. A majority of them (56%) showed type-1 histology pattern. However type-2 pattern was seen in 25% of cases. Interestingly, 2 cases showed early granuloma (type-3 pattern) and one case showed a well formed tuberculoid granuloma (type-4 pattern). In one case the picture was nonspecific.

Subacute lesions (duration 3-12 months). Clinically these lesions were crusted plaques/ noduloulcers/ulcers. A majority of them had either type-2 pattern (4 cases) or type-1 (3cases). Two cases had a granulomatous histology (type-3 pattern) and one case showed a nonspecific pattern.

Late lesions (duration more than 1 year); Clinically these lesions were persistent firm nodules. Three of six cases had a granulomatous histology (one case with type-4 and 2 cases with type-3 pattern). Interestingly, type-1 and type-2 patterns were also seen in one case each even in these late lesions. One case had nonspecific features.

Discussion

It has been well recognized that there is a clinical and histological spectrum in cutaneous leishmaniasis.[4] The spectrum and its variability are dependent on a number of factors such as the type and duration of clinical lesion, strain of organism, geographic location, parasitic load, host immunity etc.[3],[4] There is an evolution of lesions as they progress from a papule/nodule, into a soft, boggy, crusted plaque/nodule. The lesion then breaks open, after a varying period of 3-4 months into a well- circumscribed ulcer, which heals slowly over a period of 3-12 months. Occasionally, the healing does not take place and the lesion may then persist as a chronic yellow-brown firm nodule. Opinion has varied for the definition of chronic lesion, with different criteria (6 months to 2 years) being quoted by different authors.[2],[3] The healed lesion may recur after a variable time as a recidivans lesion.

The histological spectrum in CL has been variously classified into different types. Bryceson first attempted a classification of CL, into tuberculoid, macrophage (leptomonad) and intermediate forms.[1],[6] Kurban[7] reported two histological patterns- early lesions ( less than 1 year) showing a diffuse macrophage infiltrate, with plenty of organisms and late lesions (more than 1 year) with a granulomatous infiltrate. Mansour[8] in a more recent study reported similar pattern. Ridley[9] suggested a three-group classification for CL after a comparison with classification of Brazilian leishmaniasis. Gaffar et al[10] reported four different patterns depending on the various cell populations, granuloma, necrosis and parasite count.

In a series of significant publications, 11-13 Ridley elucidated the pathogenesis of CL and classified into 5 subtypes; diffuse macrophage involvement without necrosis (type- 1),diffuse macrophage necrosis (type-2), focal macrophage necrosis (type-3), early reactive tuberculoid granuloma (type-4) and tuberculoid (lupoid) granuloma (type-5). The author also emphasized the essential differences between CL and leprosy,in that self healing occurs in the middle of the spectrum in leishmaniasis and that, unlike leprosy CL is not primarily a granulomatous condition. It was further emphasized that intact macrophage granuloma without necrosis, macrophage lysis with necrosis and macrophage activation with tuberculoid granuloma formation represent the three important host responses against leishmania. Intact macrophage granuloma is the initial response in the early anergic phase of diffuse infiltration and is associated with antigen equilibrium, plasma cell lysis, causing necrosis and is associated with high parasitic load (antigen excess). This leads later to macrophage lysis, causing necrosis and is associated with antigen equilibrium, plasma cell infiltration and formation of immune complexes on macrophage cell surface. Necrosis occurs during ulceration of the lesion. The necrotic debris consists of nuclear debris as well as remnants of parasites. Necrosis is diffuse when individual macrophages are lysed, and focal when groups of cells are involved. Plasma cells are recognized as an important component of the histological reaction in CL, mostly occurring with necrosis, and decreasing with the onset of granuloma.

Our cases could be categorized into four categories, which corresponded to types 1, 2, 4 and 5 of Ridley. Though necrosis was prominent in 25% of the cases, only diffuse necrosis was observed and focal necrosis described as type 3 pattern by Ridley was not prominent. This variation in prevalence of the different pattern has been reported before. In a study of comparison of pathological pattern of cutaneous leishmaniasis in different geographical regions, El Hassan[14] et al demonstrated that type 2 pattern was more prominent in Nicaragua and Guyane, while type 3 was more common in Sudan and Saudi Arabia, and hence concluded that the histological types varied from one region to another. In the study by Kurban,[7] necrosis was rare and was seen in only one of the 27 cases studied.

Granuloma the other mechanism for parasitic elimination, results from macrophage activation into epitheloid cells and is thought to be associated with low antigen load.[11],[12] Though early reactive epitheloid granuloma is common (found in six cases in our study) true well organised tuberculoid granuloma (found in two cases) is rare. Interestingly granuloma was seen in acute lesions also, even in a lesion only four weeks old. In a study by Azadeh,[15] 25% of the acute lesions had epitheloid cell and lymphocytic infiltrate. It has been suggested that in CL, the epitheloid cell population can occur at any stage in the evolution of the disease.[11],[12] Even early lesions with high parasitic load can show granuloma at the edge of lesions and deeper sections of the biopsy. These cases may represent accelerated macrophage activation in a patient with high immunity.7.5% of cases had a nonspecific histological pattern. Similar nonspecific histological picture has been reported in previous studies.[11],[16]

Organisms were demonstrated in 65% of lesions. This figure has varied from 38-75%, in different studies.[7],[15] The success of demonstration of organism depends on a multitude of factors, including the site of biopsy and duration of lesion. Biopsy from floor or margin is best suitable for detection of parasites, though such a biopsy is difficult to obtain.[12] Parasites disappear from the biopsies after 5-7 months.[17] In late lesions less than 50% may show prasites.[7] Occasionally parasites may not be detected even in earlier lesions. [17] This may be related to the early onset of epitheloid response as described above. Organisms in the epidermis are reported to be rare[3] and were not detected in our study. Kurban[7] suggested that the presence of intraepidermal parasites was an example of transepidermal elimination of the organisms.

Several studies[8],[11],[12],[18],[19] on clinicopathological correlation in CL have been published. Our results show inconsistent correlation. Type 1 pattern was expectedly found mostly in acute lesions. However even some subacute and late lesions also showed this pattern. Type 2 pattern was mostly found in subacute lesions, but interestingly also in acute and late lesions. Type 3 and 4 patterns were found mostly in late lesions, but also in some acute and subacute lesions. Thus the correlation was not fully consistent either with type of lesion or with the duration of lesion and hence the spectrum is variable, a finding which is in conformity with previous reports.[8],[11],[12],[18],[19] It is possible that diferent parts of a lesion and of the same section may show different stages of evolution and hence different histological patterns.[12] Such a picture is well known in leprosy and has been reported CL in also.

Epidermal reactions were nonspecific and only confirmed the previously reported findings.[4],[7],[11],[12],[15] Findings such as basal cell degeneration and intraepidermal abscesses, hyperplastic rete ridges though nonspecific, may be helpful in differentiating CL from other lesions.

Histological differential diagnosis in cutaneous leishmaniasis is from other granulomatous conditions such as lupus vulgaris, leprosy, sarcoidosis, and acne rosacea. In early lesions showing type 1 and 2 patterns this is not difficult, as the changes of diffuse macrophage infiltration, necrosis, plasma cells and presence of parasites is sufficiently diagnostic. Difficulties arise when organisms are scanty and there is a granulomatous infiltrate.[4],[7],[20] Presence of plasma cells, necrosis and parasites may be detected after examination of several sections and can be useful clues for diagnosis.As stressed before, true well formed tuberculoid granuloma is uncommon in CL. Epidermal changes can differentiate CL from leprosy and sarcoidosis where such changes are uncommon. Other useful diagnostic features include presence of well organised granuloma and Schumann bodies in sarcoidosis, perineural infiltrate in leprosy and caseation in lupus vulgaris.