American cutaneous and mucocutaneous ...

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&al. PALMA

Tropical Doctor, Supplement 1, 1992

American cutaneous and mucocutaneous leishmaniasis (tegumentary): a diagnostic challenge Miguel A Escobar MD Fernando Martinez MD Fundación Centro Internacional de Entrenamiento e Investigaciones Médicas (CIDEIM) D Scott Smith MPH School of Medicine, University of Colorado, Denver, Colorado Gloria 1 Palma MD PhD Department of Microbiology, Universidad del Valle and Fundación Centro Internacional de Entrenamiento e Investigaciones Médicas (CIDEIM), AA 5390, Cali, Colombia, South America TROPICAL DOCTOR,

1992,22 (suppl 1),69-78

INTRODUCTION

The genus Leishmania comprises a yet undetermined number of different parasite species which infect a wide variety of mammalian hosts. Some of the parasites are associated with disease in man while others appear to be restricted to lower orders of mammals such as rodents'. Each of the different species found in the New World has a unique ecology and geographic distribution which ranges from southern Texas to northern Argentina. The basic maintenance cycle for all involves wild

or domestic mammals belonging to several orders, which serve as reservoirs, and phlebotomine sand flies of the genus Lutzomyia, which act as vectors. American tegumentary leishmaniasis (ATL) is the generic name for different cutaneous and mucocutaneous presentations of zoonoses caused by various species of Leishmania found in the New World (Table 1). Their transmission is highly dependent upon ecology, and consequently, the conditions under which man becomes infected, vary considerably with time and place. The intrusion of man into a sylvatic cycle, as frequently happens in the Americas, may result in a greater exposure to sand flies that are part of this cycle and, hence, in a higher risk of infection. However, disease transmission is not associated exclusively with tropical jungles. Given the appropriate conditions, the disease can be found in peridomestic or possibly intradomestic settings+, and secondary forests and farmlands, as evidenced by the finding of ATL in coffee growing areas in Venezuela' and Colombias-". The parasites are dimorphic protozoa belonging to the family Trypanosornatidae. Extracellularly, in the vector's gut or in' culture, they exist as flagellated motile organisms known as promastigotes. When man acquires the parasite through the bite of an infected female sand fly, these promastigotes enter reticuloendothelial cells through specific receptors". Here they transform into amastigotes, aflagellate oval or rounded organisms, which replicate through binary fission

Table 1. Geographical distribution and clinical forms of Leishmania species most commonly with American tegumentary leishmaniasis

~--

associated

Parasite

Clinical presenta/ion

Country

L. brazitiensis

cutaneous mucocutaneous

L. panamensis

cutaneous mucocutaneous .1 cutaneous mucocutaneous (rare) cutaneous lesions diffuse cutaneous leishmaniasis

Argentina, Belize, Bolivia, Brazil, Colombia, Costa Rica, French Guiana, Guatemala, Honduras, Mexico, Panama, Paraguay, Peru, Venezuela Colombia, Costa Rica, Ecuador, Honduras, Nicaragua, Panama, Venezuela Brazil, Colombia, French Guiana, Guyana, Peru, Suriname Belize, Colombia, Costa Rica, Guatemala, Honduras, Panama, United States (Texas) Bolivia, Brazil, Colombia, Ecuador, French Guiana, Panama, Peru, Venezuela Peru

L. guyanensis • -.

69

L. mexicana

L. amazonensis

L. peruviana

cutaneous mucocutaneous (rare) diffuse cutaneous leishmaniasis cutaneous

Adapted from Grimaldi et al. 19892, and WHO 19903


70 in the parasitophorus vacuole of the macrophage and perpetuate the infection in susceptible hosts. The cycle is completed when the insect vector takes a blood meal and ingests mononuclear cells containing amastigotes, which transform to promastigotes, multiply in the gut and later migrate to the insect's salivary glands. ATL is a serious health problem in endemic areas of the New World9,IO and can easily be confused with other granulomatous dermal diseases. Diagnosis is complicated by the high costs of laboratory supplies and reagents, superinfection of the lesions and the lack of adequately trained personnel in endemic areas. It is of utmost importance to establish a parasitologic diagnosis in order to administer the specific treatment. LEISHMANIA

AND THE HUMAN HOST

Clinical jeatures ATL is a highly prevalent tropical disease with a broad clinical spectrum 11 which ranges widely in severity and health impact. Infection with Leishmania may be inapparent, and the onset of mucocutaneous disease can be delayed months or years, while some individuals may never develop overt signs and symptoms. Indirect evidence of this is the high prevalence of skin test reactivity to Leishmania antigens in the absence of disease observed in some endemic areas 12, 13 • Patients can present with single or multiple, papular, nodular or ulcerated dermal lesions (Plate 17, P 63) which clinically may resemble many other skin diseases. Sand fly bites on exposed areas of the body produce a burning sensation at the inoculation site; single or multiple red papules of variable sizes, depending on the number of bites, will appear. After an incubation period of 2-8 weeks or sometimes more, a firm, small nodule develops which proceeds to open at the top and will ooze a serous material that develops a superficial crust. If this crust is removed, a rounded ulcer with raised and hyperaemic border is seen. It evolves slowly: an increase of a few centimetres in diameter requires several months. Usually these lesions are painless, unless they are secondarily-infected with bacteria, when a purulent discharge will be observed. Some patients may develop Iymphatic dissemination of the disease, and parasites may be recovered from the draining Iymph nodesl4,IS (personal experience). In its severest form, ATL involves the naso-oropharyngeal mucosae. These lesions can occur simultaneously with a primary cutaneous lesion or

can present months or years after a skin lesion has healed spontaneously or in response to treatment!". Usually lesions are found on the anterior nasal septum. They consist of hyperaemia, oedema and perforation of the nasal cartilage. At this stage, patients generally complain of pruritus, bloody nasal discharge and nasal obstruction. As the disease advances, the turbinates, soft palate and the uvula can become involved. Rarely, involvement of the larynx can be observed. Early diagnosis is important since chronic mucosal disease can be lifethreatening if upper respiratory airways are obstructed or secondary bacterial infection is present; in addition, mutilating sequelae can also be avoided '. Diffuse cutaneous leishmaniasis is a rare presentation of ATL associated mostly, if not exclusively, with organisms belonging to the L. mexicana, L. amazonensis or very closely related species. Lesions often present as disseminated non-ulcerated plaques or nodules with abundant amastigotes. The absence of spécific delayed type hypersensitivity as measured by intradermal testing with Leishmania antigens and the poor response to conventional antimonial treatment strongly suggests an immunological abnormality in the hostI7,18. Other diseases such as pulmonary tuberculosis can be found in patients with ATL, especially in areas where both diseases are highly prevalent. Tuberculosis is most often seen in leishmaniasis with chronic severe involvement of mucosal tissue'? (Escobar et al. unpublished); it must be treated before the leishmaniasis, because of its lifethreatening potential!". Malnutrition may also be associated with ATL, particularly in poorer countries. In patients who are thought to have malnutrition, hepatic, renal and cardiac function must be assessed carefully. Although visceral leishmaniasis has beenreported as an opportunistic infection in HIV positive patients-P-". reports of ATL being associated with AIDS patients have not yet been published. However, as the epidemiological distribution of AIDS extends to rural areas, overlap with American tegumentary leishmaniasis becomes more likely. DIAGNOSIS

Methods of parasitological diagnosis of ATL have been comparatively evaluated with variable results. Most researchers in the New World use three or more diagnostic methods (direct smear, aspirate, biopsy) for the Leishmania braziliensis species complex because of difficulty in detection and/or

Tropical Doctor, Supplement 1, 1992 isolation of the parasite22-25. However, many patients remain without a parasitological diagnosis in spite of having lesions clinically compatible with leishmaniasis22-24.26. Conventional diagnostic methods enable a definitive parasitological diagnosis in only 70-750/0 of cutaneous cases22-24. This figure is even lower in patients with mucosal (48% parasitologic diagnosis), chronic and recurrent lesions-v-". In endemic areas, presumptive diagnoses are often made using the clinical presentation and delayed cutaneous reactivity to the leishmanin skin test antigen'v". Others in the Old World use morphologic characteristics of lesions based on their knowledge of the natural history of the disease-". In order to determine the precision with which clinical criteria could be used for the diagnosis of leishmaniasis, our group has developed a clinical prediction rule. This rule was developed for patients in a specific endemic area with cutaneous lesions, based on the demographic, historical and physical characteristics that were closely associated with leishmaniasis. Our rule demonstrated a sensitivity of 96%, specificity of 92% and an efficacy of 95%, as compared with parasitological diagnosis in patients from the Pacific Coast region of Colombia (Weigle KA, et al. submitted for publication). Further studies will be needed to test this rule in other areas to determine its universality. DIAGNOSTIC METHODS

Medical or para-rnedical personnel may suspect mucocutaneous leishmaniasis based on the characteristics of the lesion and on the epidemiological history of the patient, who usually reports living in or visiting an endemic area. Parasitological diagnosis of leishmaniasis is defined as the visualization of amastigotes and/or isolation of replicating promastigotes from tissue samples. Diagnostic methods, described below, include establishing the presence of amastigotes in dermal scrapings, biopsy impressions, histological sections, growth of parasites following inoculation of lesion aspirates and/or biopsies into b'iphasic cultures and/or golden hamsters. The leishmanin skin test, which demonstrates delayed hypersensitivity to Leishmania-derived antigens, and the indirect fluorescent antibody test (IFAT) are indirect immunological approaches that are most useful for epidemiological studies of ATL. Usually the individual with cutaneous leishmaniasis has a single lesion, sometimes two or more. When multiple lesions are present, samples should

71 preferentially be taken from the most recent or active one-". If there is any evidence of bacterial contamination, oral antibiotics accompanied by daily debridement are recommended for at least 8 days before samples are taken. Dermal scraping Obtaining an optimal sample To obtain dermal scrapings, clean the margins of the lesion thoroughly with 70% alcohol and povidone-iodine. The outer border of the skin lesion is slit 3 mm in length and depth, with a number 15 surgical blade (Plate 18A, p 64). Dermal tissue is scraped from the walls of the slit with a sterile stainless steel spatula or a disposable scalpel and smeared gently onto a glass slide (Plate 18B,C, p 64), air dried, fixed in methanol, Giemsa-stained, and examined for the presence of amastigotes (Plate 18D, p 64). ldentification of parasites Stained smears are initially scanned under the 10 x objective for amastigotes in areas where granulomas and mononuclear leucocytes are seen. These granulomas are identified as clumps of ' cells (Iymphocytes, monocytes and macropháges) that stain darkpurple. Although observation of polymorphonuclear cells indicates that a good dermal scraping was obtained, granulomas formed by these cells are less unlikely to yield parasites. When the optimal field has been identified, it is focused with the 40 x . If present, amastigotes will be found in areas with an abundant mononuclear reaction or near granulomas. Sometimes they will be found around fibres that are formed when cell nuclei are destroyed due to the trauma of smearing the sample onto the glass slide. Finally, to confirm the identification of the parasite, use the 100 x objective. Under the microscope, amastigotes appear as ovoid bodies of about 4 ¡.tm long by 2-3 ¡.tm wide. They are usually found intracellularly, but can also be seen extracellularly when macrophages rupture during the smearing process. When stained with Giemsa, amastigotes have a pale blue cytoplasm, a large pinkish-red round nucleus and a darker rod-shaped red staining kinetoplast. The appearance of this three dimensional organism can vary in smears-"; vacuo les are sometimes seen in the cytoplasm and the nucleus or the kinetoplast cannot be visualized. However, care must be taken in visualizing both these structures in order to establish definitive diagnosis. Reading the slides requires some practice and, while a positive smear confirms the diagnosis, a negative smear does not rule out leíshmaniasis-".

•

72


Sensitivity varies frorn 22070to 47.7%23,24,30(see Table 3) depending on the time of evolution of the lesions. The number of amastigotes found in a lesion seems to be inversely praportional to the 'age' of the lesioné". The pro bability of seeing amastigotes is increased if 4 to 5 smears are prepared frorn each lesiono Be careful to avoid bleeding since smears with large amounts of blood are difficult to evaluate. The dermal scraping is an important diagnostic method because of its simplicity, rapid outcome, low cost-', ready availability of required materials in the field and minimal trauma to the patient-". Disadvantages inelude: low sensitivity in chronic lesions, the need for a micrascope with a 100x oil immersion objective and the need for training in preparing and evaluating the samples. Normally this procedure is not carried out in mucosal lesions because of the difficulty in obtaining adequate samples. Such lesions usually bleed abundantly and their location often makes it impossible to obtain a smear. For mucosallesions we recommend that the samples be taken by an ear, nose and throat specialist, who has equipment and experience - if one is available. Aspirate culture There is some contraversy regarding the usefulness of different culture media for the primary isolation of New World Leishmanior-". Basically there are two different kinds of media for culturing parasites: monophasic and biphasic media. We use Senekjie's medium-? (Table 2), a standard biphasic blood agar medium similar to Novy-MacNeal-Nicolle (NNN) medium. For isolation of Leishmania in the Colombian Pacific Coast, Senekjie's medium has a sensitivity of 89.8%30. Differences among culture media with regard to their ability to isolate Table 3. Comparison

Table 2. Senekjie's medium-': Difco-Bactobeef Bacto-agar Bactopeptone NaCI Defibrinated rabbit blood

Disadvantages

50 g 20 g 20g

5g 150ml

Leishmania inelude differing techniques for obtaining tissue samples, different human populations and different species of parasítes+-". To obtain lesion aspirates a disposable tuberculin syringe, fitted with a 26-gauge needle and containing 100 ¡.tI of 0.01 M phosphate buffered saline (PBS), pH 7.2, is inserted intradermally into the outer border of the lesion (Plate 19A, p 64). The syringe is ratated, and tissue fluid is gently aspirated into the needle hub at withdrawal. We recommend that four or five aspirates per patient be performed. One is inoculated subcutaneously into the nose of a golden hamster (aspirate-hamster); the others are seeded into different tubes of Senekjie's media (aspirate-culture). Inoculated culture tubes, maintained at 27°C, are exam:ined thrice weekly using an inverted micrascope for a period of one month or until positive for the presence of pramastigotes (Plate 19B, p 64). Sensitivity varies frorn 58% to 89.8% in lesions with less than 12 months of evolution=-". Advantages inelude its high sensitivity, .especially in chronic lesions, and the possibility of isolating the parasite for identification and taxonomic elassification. The disadvantages of this method are high costs, susceptibility to contamination, and the need for laboratory support. Parasites can also be isolated by in vivo inoculation of susceptible laboratory animals with elinical samples containing Leishmania. Golden

of diagnostic methods* Histological

Sensitivity' Advantages

1 Iitre

Dermal scraping

Aspira¡e culture

biopsy

44070

90% Useful in chronic cutaneous lesions; Identification of species High costs Susceptible to contamination

35% Diagnosis of other aetiologies

Low cost Simple Quick diagnosis Limited use in chronic or mucosal lesions

.r

Skill required in observing parasites High cost

Immunoperoxidase 61% Easier visualization of amastigotes Requirement for primary antisera and conjugates

Biopsy imprints

,.

A

19% Simple Quick diagnosis Low sensitivity High cost reJated to biopsy

*Modified from Salinas el al. 19893°, and Weigle el al. 198723 "Percentage 'sensitivity' is based on parasitological diagnosis by any one of the multiple methods employed in cited studies

~

J


;

hamsters (Mesocricetus auratus) are the most suitable animals because, unlike mice, their susceptibility to different Leishmania species is considered to be uniformé. Triturated biopsies or tissue fluid aspirates are inoculated intradermally in the nose or footpads. Parasites can then be isolated into culture or visualized in direct smears from the hamster when a lesion becomes apparent usually weeks or months later at the inoculation site. The costs of maintaining an animal colony and the delay in the appearance of clinical manifestations in animal s make this method unsuitable for diagnosis of patients in the usual clinical setting. HISTOPATHOLOGY

Most reports are consistent with the fact that histopathology in ATL has limited sensitivity, varying from 14070to 3507022-24 for the L. braziliensis and L. panamensis species. This is probably due to the scarcity of parasites or distortion of amastigotes during fixing22.29.34.Biopsies are mainly useful in the diagnosis of non-leishmanial diseases and should be reserved for selected patients, because they are high cost, of low sensitivity and require expertise and equlpment-t->. Lesions and lesion borders are cleaned and debrided with povidone iodine. After injecting 2070 xylocaine into the dermis, a 4-mm punch biopsy is taken at the margin of the lesiono Impression smears (touch preparations) are carried out, Giemsastained, and examined with the 100 x eyepiece for the presence of amastigotes using the same technique as for the dermal scraping. Disadvantages of impressions smears are their low sensitivity and the probability of jeopardizing the physical integrity of the biopsy sample. The biopsy is then fixed in 10070 buffered formalin, pH 7.0, imbedded in paraffin, and sectioned. As with dermal scrapings, the probability of finding amastigotes increases with the number of samples examined. When examining the histological sections stained with Giemsa, tissue sections should be scanned in the derrnal-epiderrnal junction for intracellular amastigotes. The presence of necrosis and histiocytes has been .highly correlated with the finding of amastigotes iri lesions of less than 6 months' duration-". The sensitivity of histopathology can be increased using an indirect immunoperoxidase technique, which is more specific in demonstrating amastigotes in tissues-". Immunoperoxidase method

Paraffin embedded histological slides from biopsies are evaluated using an indirect immunoperoxidase

73 assay employing a hyperimmune rabbit antileishmanial primary antibody as previously described-". In our experience, its sensitivity is 61.3070;efficiency being greatest in recent lesions. Positivity ranges from 75070in cases with less than 3 months of evolution to 21.1070in patients with lesions of 12 or more months duration. The combined use of dermal scraping and immunoperoxidase allowed a diagnosis in 72070 of cases. Disadvantages of this method are the need to prepare and/or obtain monoclonal or polyclonal antibodies. Although cross-reactivity with Paracoccidioides braziliensis may be seen with a polyclonal primary antiserum, morphology readily allows discrimination-". Montenegro skin test The Montenegro skin test is a highly sensitive and specific intradermal test that cannot distinguish between current and previous leishmanial infecnon-'-". In some endemic areas, skin test reactivity is very frequent in the general population, hence it is not helpful in the diagnosis of active lesionslO.23.36.However, if a patient from a nonendemic country presents with suggestive lesions after travelling to an endemic area, the Montenegro skin test can be used as a diagnostic aid. But this test depends on antigen being available and on the patient's epidemiological history. The sensitivity and potency (measured by size of induration) of the skin test can be influenced by some host characteristics such as: lesion type (cutaneous vs mucocutaneous); lesion stage (active vs healed); duration of active lesions and time since healing'". To apply the leishmanin skin test, 100 ¡.tI of Montenegro skin test antigen containing heat-killed sonicated promastigotes are injected intradermally into the volar surface ofthe forearm. The diameter of induration is measured in millimetres 40-56 h later by outlining the indurated border with a ballpoint pen and transferring it to paper for permanent record'". An indurated area of 5 mm or more is considered positive-". Appropriate handling of the antigen in the field includes adequate storage at temperatures ranging between 2°C and goC; harsh conditions with fluctuating temperatures (4°C to 32°C) can decrease potency although, if allowance is made for this, sensitivity is apparently unaffected. For field use we recommend that each vial of antigen be dated on arrival, and once taken out of refrigeration, be used only during one day in the field".

74 Indirect jluorescent antibody test The presence of parasites in tissues can be detected indirectly through evaluation of circulating antiLeishmania antibodies by various methods39-42• The indirect fluorescent antibody test (IF A T) has been widely used to detect circulating antibodies in leishrnaniasis+". In cutaneous leishmaniasis however, parasitologically proven cases may have negative serology'", while positive serological results may be seen in patients with no apparent cutaneous disease '. In early lesions neither humoral antibody response nor delayed hypersensitivity, alone, are sufficiently sensitive parameters for detection of infection. Specific IgM is detected in sera of patients having recent lesions, usually of less than 2 months' duration. On the other hand, IgG titres are usually detectable in sera of patients with lesions more than one month old and for several months after treatment has ended. If Montenegro skin test reactivity and antiLeishmania IgM titres are used together, sensitivity can be increased and specificity can reach 100070in patients with lesions of less than 2 months. This compares with a specificity of 81.2070 when IgG titres and Montenegro skin test results are used together. Therefore, combined use of IgM titre and Montenegro reactivity is of potential utility in the clinical diagnosis of early lesions+. Mucocutaneous disease is associated with greater antibody response". It is often impossible to demonstrate parasites in these patients because amastigotes are frequently scarce. For lack of better diagnostic evidence, clinical diagnosis supported by positive serology is considered significant in patients presenting mucosal lesions". Other diagnostic techniques for detection of parasite DNA sequences using labelled probes together with an amplification system such as the polymerase chain reaction (PCR) offer highly specific and sensitive methods that can theoretically overcome the diagnostic constraint imposed by the scarcity of parasites in chronic and mucosal disease45,46. This promising experimental approach is being tested for application on clinical specímens and awaits further assessment in the field. DIAGNOSTIC CRITERIA

Although definitive diagnosis of ATL is established only when Leishmania are observed or isolated, in endemic are as a clinical diagnosis is sometimes the only option available for a patient with tegumentary lesions. However, certain minimal criteria must be met. In our experience at least three of the following

Tropical Doctor, Supplement 1, 1992 parameters should be fulfilled in patients from proven endemic areas: (1) The presence of a characteristic lesion; that is, ulcers with a sharp, raised border with a rim of oedema and hyperaemia. The base is clean with healthy pink granulation tissue with hyperaemia and petechiaef"; (2) a compatible histological picture, including a granulomatous lesion with predominantly histiocytic infiltrate and a variable number of lymphocytes, polymorphonuclear leucocytes, and plasma cells48; (3) a positive res pon se to treatment, ie patients who received parenteral pentavalent antimonials in doses recommended by the W orld Health Organization= and have had an obvious clinical response; (4) specific antileishmanial immune response, either serological or delayed-type cutaneous hypersensitivity-s; (5) exclusion of other possible diseases, for example bacterial and fungal infections, leprosy, syphilis, non-venereal treponematoses, vascular lesions, skin cancer and local trauma.

Difjerential diagnosis

-,

For patients with negative leishmania tests and who do not fulfil the parameters for a clinical diagnosis, it is important to establish the aetiology of their lesions. Epidemiological considerations are important in these cases, since the prevalence of specific di seas es can vary from one region to another. The individual's occupation will also aid in establishing the aetiology; for example, agricultural workers and wood cutters will be more exposed to Sporothrix schenckii and environmental dematiaceous fungi that cause chromoblastomycosis and related dermal disorders. Fishermen can acquire infections with atypical mycobacteria whereas children tend to acquire common pyogenic bacterial infections. Underlying diseases like diabetes mellitus, vascular abnormalities in the lower limbs, sickle cell disease, arterial hypertension among others, must be ruled out. In our own experience, sporotrichosis is the commonest differential diagnosis, accounting for 36070 of all patients presenting with non-Ieishmanian dermallesions. Its laboratory diagnosis is made by culturing material from within subcutaneous nodules, and/or exudate from draining ulcerated lesions in a medium of Mycosel agar containing cycloheximide and chloramphenicor". AIso a biopsy from an ulcerated les ion or a firm subcutaneous nodule can be taken for histopathological study.


!

The second most common diagnostic problem is simple bacterial infection found in 22070 of the cases. Culture in appropriate media and/or biopsy from the border of the lesion can give a diagnosis. Some patients may have more than one bacterium in the same lesion, hence the advantage of taking cultures. In areas where these methods are not available, the physician can prescribe oral, wide spectrum antibiotics for 10-14 days, based on clinical criteria. Patients must be followed closely in order to document an adequate res pon se to this therapy. Diagnosis may then be made retrospectively. Skin cancers are the third most common problem occurring in populations that are constantly exposed to the sun's ultraviolet radiation. In our series of patients skin cancer ranks third in the list of differential diagnoses. Histopathology is the only definitive diagnostic method in such cases. Other rarer skin disorders that must be differentiated are ulcers associated with sickle cell disease, Hansen's disease, yaws, secondary syphilis, local trauma and lesions secondary to vascular disease. Mucosal involvement due to leishmaniasis can be clinically confused with diseases such as: tuberculosis, leprosy, syphilis, yaws, rhinoscleroma, paracoccidioidomycosis, and carcinoma among others. Adequate examination of biopsy material generally allows these diseases to be differentiated. TREATMENT Treatment of mucocutaneous leishmaniasis in the New World in the last decade has included multiple drugs. Pentavalent antimony (Sb5+) is the drug of choice recommended by the W orld Health Organization-". Two presentations are available: meglumine antimoniate (Glucantime'') which is generally used in French- and Spanish-speaking countries and sodium stibogluconate (Pentostam'') used in English-speaking countries. Most reports mention the many disadvantages of the antimonials, particularly toxicity, inconvenient dosage schedules, route of administration and high costs3.50-52. For all disease forms, except those produced by L. braziliensis complex, 10-20 mg of systemic pentavalent antimonials per kilogram of body weight given once daily, with a maximum total dose of 850 mg per day is the regimen of choice->. Give this regimen until clinical and parasitological cure is achieved, and then continue for a few days after healing. Patients with cutaneous leishmaniasis due to L. braziliensis, L. panamensis or L. guyanesis are at risk of developing mucosal dissemination; therefore give

75 20 mg of Sb5 + per kg of body weight per day for a minimum of 4 weeks. This regimen is also appropriate in areas where the presence of L. braziliensis is suspected. In mucocutaneous lesions, give 20 mg of parenteral Sb5+ per kg of body weight every 12 h until clinical and parasitological cure has been achieved. There is no rule to determine when to discontinue the treatment in these patients '. Most physicians make a decision based on clinical judgement, but many patients can relapse and this is often associated with lower doses of antimonials or interrupted treatments+'. Patients with relapses should receive the same drug and daily dose, but for a duration of twice the original treatment. When there is no response to antimony therapy, amphotericin B is the second line drug of choice- (20-40 doses at 1 mg/kg of body weight). Amphotericin B is highly toxic and requires parenteral administration-? in a hospital setting. In patients whose mucosa is diseased, severe inflammation, erythema and sometimes haemorrhage and life-threatening laryngeal obstruction may occur during the first week of treatment with antimonials-l->'. This seems to be due to a local reaction to antigens released from killed parasites and not a side effect of the treatment itself+>'. Administration