REPORT
CHRONIC CUTANEOUS LEISHMANIASIS: LE/SHMANM PARASITES IN BLOOD NUHA NUWAYRI-SALTI, M.D., DIA MU'ATASSEM, M.D., SALAH SALMAN, M.D., NAZIK IZZEDINE, M.D., AND ABDEL GHANI KIBBI, M.D.
Patient 2: A 60-year-old woman presented witb a painless 20-year-old skin lesion over tbe mid-forebead. It started as a small abscess-like lesion and grew gradually to reach its present size. On examination, she bad a mid-forebead brownisb-red plaque measuring 8 x 4 cm and extending beyond tbe glabellar area to tbe naso-orbital angles. Tbe plaque was slightly scaly witb well-defined borders and a depressed center (Fig. IB). Tbe rest of tbe physical examination was negative. Her routine laboratory tests were all within normal limits except for a mild eosinophilia of 6%. Similar lesions were present at one time or another in some people in her neighborhood but most bad spontaneous recovery.
Abstract Two patients witb cbronic cutaneous leisbmaniasis had positive blood cultures. Tbe diagnosis was established by tbe clinical picture, skin biopsy, and culture for Leishmania mayor and tropica. Cutaneous and systemic (kala-azar) leisbmaniasis are well-defined entities in the Middle East.'"-* The former is usually an affection limited to the skin, and only in rare exceptions does it seem to spread along the lymphatic vessels draining the area.**'' The sporotrichoid variant of this disease has been reported to invade only locally.''''' Leishmania tropica and Leishmania major parasites are thought to be the only causative agents for all types of old world cutaneous affections (acute and chronic);'"" whereas infection with L. donovani infantum, prevalent in our area, is almost invariably expressed as kala-azar and exclusively in children.' So far the established facts have been that the former species do not cross beyond the skin and that L. donovani infantum is hardly ever dermotropic. We are reporting 2 out of 21 cases of cutaneous leishmaniasis in adults from whose blood Leishmania parasites were grown successfully in vitro.
Materials and Methods Skin biopsies were obtained from all 21 patients, and a portion of each was used for culture, while the rest was processed for routine bistopatbologic examination after fixation in 3% paraformaldehyde. For each specimen, some paraffin sections were stained with hematoxylin and eosin, Giemsa, or with periodic acid-Schiff." Peripheral blood smears were stained with Giemsa. A portion of the skin biopsy and a 5 mL sample of blood from eacb patient were cultured each on a modified Nicole-Novy-McNeal (NNN) medium with Lock's overlay and in McCoy's - 2a medium witb L-glutamine witbout sodium bicarbonate. The pH was adjusted to
Case Reports Patient 1: A 49-year-old woman presented witb skin lesions of 3 years duration over tbe forebead and tbe rigbt cheek. Both lesions started as small papules that grew gradually over 3 years to become plaques measuring 2 x 3.5 cm over tbe rigbt side of the forehead and a 1 x 1 cm lesion over tbe right cbeek. Botb were brownisb-red elevated plaques covered by an atrophic tbin shiny skin with mild scaliness at the well-defined perimeter (Fig. IA). The rest of her physical examination was normal.
From the Departments of Human Morphology and Dermatology, Faculty of Medicine, American University of Beirut, Lebanon. Supported by a grant from the Lebanese National Council for Scientific Research and from the Diana Tamari Foundation.
Figure 1. Photograph of the facial chronic lesions (note scaliness). A, patient (No. 1) with lesions on forehead and cheek (bar is 0.5 cm). B, patient (No. 2) with lesion on forebead (bar is 1 cm).
Address for correspondence: Dr. Nuha Nuwayri-Salti, Department of Human Morphology, American University of Beirut, 850 Third Avenue, New York, New York 10022. •sm.
chronic Cutatieous Leishmaniasis Nuwavri-Salti et al.
7.2 witb bicarbonate buffer. Fetal calf serum (10%) was added to the media before use. All culture media were supplemented with 100 |jg/mL of penicillin streptomycin. The culture flasks (75 cm^ growth area) were kept at 22 to 24°C in room air for 2 to 3 weeks. Growth was monitored by microscopic examination using an inverted microscope and by looking at fresb and permanent preparations (Giemsa stained) every 4 days.
RESULTS
Tbese two cases happen to be the only ones witb lesions more than 24 months old. Both their skin biopsies and cultures failed to demonstrate the presence of any parasite. All tbe other cases (19/21) had 3 to 8 montbs old lesions and bad positive skin biopsies upon microscopy, although only 15 out of 19 yielded positive cultures from these biopsies. Both our cases had typically slow spreading almost sterile lesions, and a section of each under the microscope revealed typical granulomatous aggregates in the dermis (Fig. 2A) witb an occasional giant cell (Fig. 2B). Only the blood samples grew promastigote forms of Leishmania within 17 to 28 days. The most suitable medium for our isolates proved to be the biphasic modified NNN medium with the overlay supplemented with l-CS.
Figure 2. A, photomicrograph of lesion from patient (1) typical tuberculoid granulomas in dermis (Giemsa Stain, bar 500 l-im). B, higher magnification of the above granuloma showing a giant cell ('') (bar 50 |.im). CONCLUSIONS
DISCUSSION
We would like to recommend the practice of having blood cultures, on appropriate media, carried out on every patient witb cutaneous leishmaniasis in an endeavor to uncover some unknown aspects in the bebavior of the different strains of this parasite.
Our findings on the skin biopsies from botb patients are not any different from what is establisbed in the literature.**"'^ As for the blood cultures, we are confident that our isolates are not a contaminant, Leishmania being a rather fastidious organism to grow. We have been using a double-blind tecbnique whereby the clinician obtains the samples and has them sent coded to our unit. Decoding is only done a month later wbetber the cultures, by that time, are positive or negative. During that time, a total of 19 patients, proven on clinical grounds and by skin cultures and biopsies to bave cutaneous leisbmaniasis, had negative blood cultures. Circulating parasites in tbe blood of patients witb cutaneous leisbmaniasis is not totally unbeard of. Brahmachari and Dutt (1927) reported L. donovani grown on culture media from the blood of a patient bearing skin lesions recurrent from a case of Indian kala-azar.** Several reports have appeared in the literature demonstrating invasion of deeper organs by parasites assumed to be primarily dermotropic."""' Tbis empbasizes the importance of identifying the causative parasite using objective characteristics rather than clinical criteria.'•''''^'"^' Several groups in the field have used monoclonal antibodies rather successfully to this end.'-'^
Acknowledgments: Dr. A.K. Kurban and Dr. S. Zeinoun referred the patients. Mrs. Samia Mroueh Abul Hassan and Ms. Fatmen Ja'afar provided technical help.
REEERENCES
1.
2.
3.
4.
563
Adler S, Katzenellenbogen 1. The problem of the association between particular strains of Leishmania tropica and the clinical manifestations produced by tbem. Ann Trop Med Parasitol 1952; 46:25-32. Azadeh B. Localized Leishmania lymphadenitis: a light and electron microscopic study. Am J Trop Med Hyg 1985; 34:447-455. Barral A, Badaro R, Barral-Netto M, et al. Isolation of Leishmania mexicana amazonensis from tbe bone marrow in a case of American visceral leishmaniasis. Am J Trop Med Hyg 1986; 35:732-734. Bellazoug S, Lanotte G, Maazoun R, et al. A new enzymatic variant of Leishmania species. In: Morel CM, ed. Genes and antigens of parasites: a laboratory manual.
International Journal of Dermatology Vol. 31, No. 8, August 1992
5.
2nd Ed. Rio de Janeiro: Fundacao Oswaldo Cruz, 1985:47. Berger TG, Meltzer MS, Oster CN. Lymphnode involvement in leishmaniasis. J Am Acad Dermatol 1985; 12:993-996.
6.
Brahmachari UN, Dutt MB. Dermal leishmanoid with positive flagellate culture from the peripheral blood, j Trop Med Hyg 1927; 30:158-159.
7.
Chance ML. The identification of Leishnania. In: Taylor AER, MuUer R, eds. Problems in the identification of parasites and their vectors. Oxford: Blackwell, 1979: 55-74. Drury RAB, Wallington EA. Carleton's histological technique. Oxford: Oxford University Press, 1980. Farah FS, Malak JA. Cutaneous leishmaniasis. Arch Dermatol 1971; 103:467-473. Grimaldi G Jr, Moriearty PL. Visceral infection of Leishmania mexicana mexicana in mice before, during and after active phase of primary cutaneous lesions. IX Reuniao Anna de Pesquisa Basica, 1982.
8. 9. 10.
11.
12.
13.
Hommei M. The genus Leishmania: biology of the parasites and clinical aspects. Bull Inst Pasteur 1978; 76: 5-102. Kibbi AG, Karam PG, Kurban AK. Sporotrichoid leishmaniasis in patients from Saudi Arabia: clinical and histologic features. J Am Acad Dermatol 1987; 17:754-764. Nadim A, Faghih M. The epidemiology of cutaneous leishmaniasis in the Isfahan province of Iran. Trans Roy Soc Trop Med Hyg 1968; 62:534-542.
14.
Parks A, Camisa C. Sporotrichoid leishmaniasis: report of a case in an American graduate student (letter). J Am Acad Dermatol 1987; 17:855-856. 15. Peters W, Evans DA, Lanhani SM. Importance of parasite identification in cases of leishmaniasis (editorial). J Roy Soc Med 1983; 76:540-542. 16. Schnur LF, Ghance ML, Ebert F, et al. The biochemical and serological taxonomy of visceralizing Leish,na,,ia. ' - Ann Trop Med Parasitol 1981; 75:131-144. 17. McMahon-Pratt D, David JR. Monoclonal antibodies that distinguish between New world species of Leishma,iia. Nature 1981; 291:581-583.
Scientific Review In 1662, when the Royal Society of London for Improving Natural Knowledge was incorporated, the members chose Robert Hooke for the post of Curator of Experiments, assigned to perform experiments suggested by tnembers. Daniel Boorstin has noted that at the time of this appointment, Hooke sounded a keynote that marked the beginning of a new scientific age: "The truth is [that] the science of Nature has been already too long made only a work of the brain and the fancy. It is now high titue that it should return to the plainness and soundness of observations on material and obvious things." The needs to disseminate the new information and to provide a public record documenting the details of the experiments in this scientific revolution were recognized at the very outset. Many of the early "natural scientists" recorded their new observations and concepts in letters sent to an international network of correspondents. The letters came to be called "epistolary discourses": they were the forerunners of a new written form—the scietitific paper. More than 25 years ago Sir Peter Medawar attacked the format of the modern scientific paper as fraudulent. Articles in the form in which they are communicated to learned journals are, he charged, notorious for misrepresenting the processes of thought that led to whatever discoveries they describe. More recently, scientific communications have been criticized because they project a misleading image of perfection. Bob Coleman, a novelist and a perceptive critic of the written word, has observed that scientific writing often seems too good to be true. He quoted a widely published biochemist who said, "Nowadays, journals don't even want you to discuss your negative experiments. The very hint that you might have erred makes you insufficiently Olympian to be in print." The modem artificiality is odd because an open confession of difficulties, of failures, and of errors was considered to be part and parcel of the earliest scientific reports of efforts to obtain synthetic knowledge ("covered with the fingerprints and grime of the material world"), as distinct from analytical knowledge ("glowing with the radiatice of disembodied thought"). ¥rom Silverman WA. Suspended judgment. Is the scientific paper a fratid? Controlled Clin Trials 1991; 12:273-276.
sm
CHRONIC CUTANEOUS LEISHMANIASIS: LE/SHMANM PARASITES IN BLOOD NUHA NUWAYRI-SALTI, M.D., DIA MU'ATASSEM, M.D., SALAH SALMAN, M.D., NAZIK IZZEDINE, M.D., AND ABDEL GHANI KIBBI, M.D.
Patient 2: A 60-year-old woman presented witb a painless 20-year-old skin lesion over tbe mid-forebead. It started as a small abscess-like lesion and grew gradually to reach its present size. On examination, she bad a mid-forebead brownisb-red plaque measuring 8 x 4 cm and extending beyond tbe glabellar area to tbe naso-orbital angles. Tbe plaque was slightly scaly witb well-defined borders and a depressed center (Fig. IB). Tbe rest of tbe physical examination was negative. Her routine laboratory tests were all within normal limits except for a mild eosinophilia of 6%. Similar lesions were present at one time or another in some people in her neighborhood but most bad spontaneous recovery.
Abstract Two patients witb cbronic cutaneous leisbmaniasis had positive blood cultures. Tbe diagnosis was established by tbe clinical picture, skin biopsy, and culture for Leishmania mayor and tropica. Cutaneous and systemic (kala-azar) leisbmaniasis are well-defined entities in the Middle East.'"-* The former is usually an affection limited to the skin, and only in rare exceptions does it seem to spread along the lymphatic vessels draining the area.**'' The sporotrichoid variant of this disease has been reported to invade only locally.''''' Leishmania tropica and Leishmania major parasites are thought to be the only causative agents for all types of old world cutaneous affections (acute and chronic);'"" whereas infection with L. donovani infantum, prevalent in our area, is almost invariably expressed as kala-azar and exclusively in children.' So far the established facts have been that the former species do not cross beyond the skin and that L. donovani infantum is hardly ever dermotropic. We are reporting 2 out of 21 cases of cutaneous leishmaniasis in adults from whose blood Leishmania parasites were grown successfully in vitro.
Materials and Methods Skin biopsies were obtained from all 21 patients, and a portion of each was used for culture, while the rest was processed for routine bistopatbologic examination after fixation in 3% paraformaldehyde. For each specimen, some paraffin sections were stained with hematoxylin and eosin, Giemsa, or with periodic acid-Schiff." Peripheral blood smears were stained with Giemsa. A portion of the skin biopsy and a 5 mL sample of blood from eacb patient were cultured each on a modified Nicole-Novy-McNeal (NNN) medium with Lock's overlay and in McCoy's - 2a medium witb L-glutamine witbout sodium bicarbonate. The pH was adjusted to
Case Reports Patient 1: A 49-year-old woman presented witb skin lesions of 3 years duration over tbe forebead and tbe rigbt cheek. Both lesions started as small papules that grew gradually over 3 years to become plaques measuring 2 x 3.5 cm over tbe rigbt side of the forehead and a 1 x 1 cm lesion over tbe right cbeek. Botb were brownisb-red elevated plaques covered by an atrophic tbin shiny skin with mild scaliness at the well-defined perimeter (Fig. IA). The rest of her physical examination was normal.
From the Departments of Human Morphology and Dermatology, Faculty of Medicine, American University of Beirut, Lebanon. Supported by a grant from the Lebanese National Council for Scientific Research and from the Diana Tamari Foundation.
Figure 1. Photograph of the facial chronic lesions (note scaliness). A, patient (No. 1) with lesions on forehead and cheek (bar is 0.5 cm). B, patient (No. 2) with lesion on forebead (bar is 1 cm).
Address for correspondence: Dr. Nuha Nuwayri-Salti, Department of Human Morphology, American University of Beirut, 850 Third Avenue, New York, New York 10022. •sm.
chronic Cutatieous Leishmaniasis Nuwavri-Salti et al.
7.2 witb bicarbonate buffer. Fetal calf serum (10%) was added to the media before use. All culture media were supplemented with 100 |jg/mL of penicillin streptomycin. The culture flasks (75 cm^ growth area) were kept at 22 to 24°C in room air for 2 to 3 weeks. Growth was monitored by microscopic examination using an inverted microscope and by looking at fresb and permanent preparations (Giemsa stained) every 4 days.
RESULTS
Tbese two cases happen to be the only ones witb lesions more than 24 months old. Both their skin biopsies and cultures failed to demonstrate the presence of any parasite. All tbe other cases (19/21) had 3 to 8 montbs old lesions and bad positive skin biopsies upon microscopy, although only 15 out of 19 yielded positive cultures from these biopsies. Both our cases had typically slow spreading almost sterile lesions, and a section of each under the microscope revealed typical granulomatous aggregates in the dermis (Fig. 2A) witb an occasional giant cell (Fig. 2B). Only the blood samples grew promastigote forms of Leishmania within 17 to 28 days. The most suitable medium for our isolates proved to be the biphasic modified NNN medium with the overlay supplemented with l-CS.
Figure 2. A, photomicrograph of lesion from patient (1) typical tuberculoid granulomas in dermis (Giemsa Stain, bar 500 l-im). B, higher magnification of the above granuloma showing a giant cell ('') (bar 50 |.im). CONCLUSIONS
DISCUSSION
We would like to recommend the practice of having blood cultures, on appropriate media, carried out on every patient witb cutaneous leishmaniasis in an endeavor to uncover some unknown aspects in the bebavior of the different strains of this parasite.
Our findings on the skin biopsies from botb patients are not any different from what is establisbed in the literature.**"'^ As for the blood cultures, we are confident that our isolates are not a contaminant, Leishmania being a rather fastidious organism to grow. We have been using a double-blind tecbnique whereby the clinician obtains the samples and has them sent coded to our unit. Decoding is only done a month later wbetber the cultures, by that time, are positive or negative. During that time, a total of 19 patients, proven on clinical grounds and by skin cultures and biopsies to bave cutaneous leisbmaniasis, had negative blood cultures. Circulating parasites in tbe blood of patients witb cutaneous leisbmaniasis is not totally unbeard of. Brahmachari and Dutt (1927) reported L. donovani grown on culture media from the blood of a patient bearing skin lesions recurrent from a case of Indian kala-azar.** Several reports have appeared in the literature demonstrating invasion of deeper organs by parasites assumed to be primarily dermotropic."""' Tbis empbasizes the importance of identifying the causative parasite using objective characteristics rather than clinical criteria.'•''''^'"^' Several groups in the field have used monoclonal antibodies rather successfully to this end.'-'^
Acknowledgments: Dr. A.K. Kurban and Dr. S. Zeinoun referred the patients. Mrs. Samia Mroueh Abul Hassan and Ms. Fatmen Ja'afar provided technical help.
REEERENCES
1.
2.
3.
4.
563
Adler S, Katzenellenbogen 1. The problem of the association between particular strains of Leishmania tropica and the clinical manifestations produced by tbem. Ann Trop Med Parasitol 1952; 46:25-32. Azadeh B. Localized Leishmania lymphadenitis: a light and electron microscopic study. Am J Trop Med Hyg 1985; 34:447-455. Barral A, Badaro R, Barral-Netto M, et al. Isolation of Leishmania mexicana amazonensis from tbe bone marrow in a case of American visceral leishmaniasis. Am J Trop Med Hyg 1986; 35:732-734. Bellazoug S, Lanotte G, Maazoun R, et al. A new enzymatic variant of Leishmania species. In: Morel CM, ed. Genes and antigens of parasites: a laboratory manual.
International Journal of Dermatology Vol. 31, No. 8, August 1992
5.
2nd Ed. Rio de Janeiro: Fundacao Oswaldo Cruz, 1985:47. Berger TG, Meltzer MS, Oster CN. Lymphnode involvement in leishmaniasis. J Am Acad Dermatol 1985; 12:993-996.
6.
Brahmachari UN, Dutt MB. Dermal leishmanoid with positive flagellate culture from the peripheral blood, j Trop Med Hyg 1927; 30:158-159.
7.
Chance ML. The identification of Leishnania. In: Taylor AER, MuUer R, eds. Problems in the identification of parasites and their vectors. Oxford: Blackwell, 1979: 55-74. Drury RAB, Wallington EA. Carleton's histological technique. Oxford: Oxford University Press, 1980. Farah FS, Malak JA. Cutaneous leishmaniasis. Arch Dermatol 1971; 103:467-473. Grimaldi G Jr, Moriearty PL. Visceral infection of Leishmania mexicana mexicana in mice before, during and after active phase of primary cutaneous lesions. IX Reuniao Anna de Pesquisa Basica, 1982.
8. 9. 10.
11.
12.
13.
Hommei M. The genus Leishmania: biology of the parasites and clinical aspects. Bull Inst Pasteur 1978; 76: 5-102. Kibbi AG, Karam PG, Kurban AK. Sporotrichoid leishmaniasis in patients from Saudi Arabia: clinical and histologic features. J Am Acad Dermatol 1987; 17:754-764. Nadim A, Faghih M. The epidemiology of cutaneous leishmaniasis in the Isfahan province of Iran. Trans Roy Soc Trop Med Hyg 1968; 62:534-542.
14.
Parks A, Camisa C. Sporotrichoid leishmaniasis: report of a case in an American graduate student (letter). J Am Acad Dermatol 1987; 17:855-856. 15. Peters W, Evans DA, Lanhani SM. Importance of parasite identification in cases of leishmaniasis (editorial). J Roy Soc Med 1983; 76:540-542. 16. Schnur LF, Ghance ML, Ebert F, et al. The biochemical and serological taxonomy of visceralizing Leish,na,,ia. ' - Ann Trop Med Parasitol 1981; 75:131-144. 17. McMahon-Pratt D, David JR. Monoclonal antibodies that distinguish between New world species of Leishma,iia. Nature 1981; 291:581-583.
Scientific Review In 1662, when the Royal Society of London for Improving Natural Knowledge was incorporated, the members chose Robert Hooke for the post of Curator of Experiments, assigned to perform experiments suggested by tnembers. Daniel Boorstin has noted that at the time of this appointment, Hooke sounded a keynote that marked the beginning of a new scientific age: "The truth is [that] the science of Nature has been already too long made only a work of the brain and the fancy. It is now high titue that it should return to the plainness and soundness of observations on material and obvious things." The needs to disseminate the new information and to provide a public record documenting the details of the experiments in this scientific revolution were recognized at the very outset. Many of the early "natural scientists" recorded their new observations and concepts in letters sent to an international network of correspondents. The letters came to be called "epistolary discourses": they were the forerunners of a new written form—the scietitific paper. More than 25 years ago Sir Peter Medawar attacked the format of the modern scientific paper as fraudulent. Articles in the form in which they are communicated to learned journals are, he charged, notorious for misrepresenting the processes of thought that led to whatever discoveries they describe. More recently, scientific communications have been criticized because they project a misleading image of perfection. Bob Coleman, a novelist and a perceptive critic of the written word, has observed that scientific writing often seems too good to be true. He quoted a widely published biochemist who said, "Nowadays, journals don't even want you to discuss your negative experiments. The very hint that you might have erred makes you insufficiently Olympian to be in print." The modem artificiality is odd because an open confession of difficulties, of failures, and of errors was considered to be part and parcel of the earliest scientific reports of efforts to obtain synthetic knowledge ("covered with the fingerprints and grime of the material world"), as distinct from analytical knowledge ("glowing with the radiatice of disembodied thought"). ¥rom Silverman WA. Suspended judgment. Is the scientific paper a fratid? Controlled Clin Trials 1991; 12:273-276.
sm
