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Use of the polymerase chain reaction for the diagnosis of American tegumentary leishmaniasis*

Use of the polymerase chain reaction for the diagnosis of American tegumentary leishmaniasis* Letters 113 in tissue samples in which the parasite forms could not be found. In 2017, two cases were investigated in Lavras. Case 1 was Use of the polymerase chain reaction for the a 39-year-old female patient who presented with a 5-month his- diagnosis of American tegumentary leishmaniasis tory of a lesion on the right arm. Clinical examination revealed a painful ulcer with pruritus measuring 2cm in diameter, with raised, hyperemic, and irregular edges, and central granulation tissue and Thales Augusto Barçante¹ serous exudate (Figure 1). Due to the unavailability of Montenegro Marcos Vilela de Souza¹ Allana Cristina Baltazar Fernandes² antigen for intradermal reaction, we performed a biopsy to collect Joziana Muniz de Paiva Barçante¹ lesional specimens. The biopsy was performed after the lesion was cleaned with soap and water and disinfected with ethyl alcohol 70%. We used 2% lidocaine for local anesthesia. Two punch biop- DOI: http://dx.doi.org/10.1590/abd1806-4841.20197929 sies were taken from the ulcer edge for histopathological analysis, which showed no amastigote forms. The second sample was used for imprint with staining, which also revealed no amastigote forms of the parasite. Subsequently, the same material was preserved in sterile saline solution, macerated with 300µg of extraction buffer, Dear Editor, and frozen in a properly identified flask for PCR. After analyzing Leishmaniasis is an infectious and parasitic disease with a the electrophoretic pattern of the sample with the positive control, variety of clinical forms, ranging from dermatological lesions (teg- the presence of Leishmania spp. DNA was confirmed. umentary leishmaniasis) to systemic manifestations (visceral leish- Case 2 was a 32-year-old female patient who had a lesion on 1-4 maniasis). American tegumentary leishmaniasis (ATL) represents the right thigh since October 2016. She was receiving antibiotic treat- a serious public health problem. Due to the high number of cases ment under medical supervision with transient clinical improve- of ATL in Brazil, differential diagnosis is necessary to understand ment. After 6 months, clinical examination revealed the presence of the epidemiological profile of ATL, both in areas previously con- a painful ulcer of approximately 2cm in diameter with hyperemic sidered non-endemic and in endemic areas. The diagnosis depends edges, and center covered with a meliceric crust (Figure 2). Simi- on the patient’s clinical history, epidemiology, and laboratory tests. larly, as described for case 1, we also performed histopathological Lavras, state of Minas Gerais, had 9 suspected ATL cases reported analysis and imprint with staining from five different slides and in the last 7 years, 8 of which were confirmed. Out of them, 7 cases rapid panoptic staining (Romanowsky), also with no identification were treated and considered cured, and 1 patient died from treat- of amastigote forms of the parasite. After the PCR, the presence of ment-associated toxicity. Furthermore, studies on the phlebotomine Leishmania spp. DNA was confirmed. fauna evidenced the presence of the main vectors in the munici- In both cases, after the detection of Leishmania DNA (Figure pality. Considering that epidemiological knowledge and clinical 3) and diagnostic confirmation of ATL, the treatment chosen was characterization of ATL are fundamental precepts of dermatological N-methylglucamine antimoniate at a daily intravenous dose of 13.7 practices, this case report aims to suggest polymerase chain reac- ml for 20 days. Both patients evolved to clinical cure after therapy. tion (PCR) as a possible diagnostic tool for cases where the Monte- negro antigen is not available and/or histopathological analysis is inconclusive. The PCR is capable of detecting the DNA of parasite Received 04 December 2017. Accepted 16 May 2018. * Work conducted at the Universidade Federal de Lavras, Lavras (MG), Brazil. Financial Support: FAPEMIG - APQ 02553-14. Conflict of interest: None. Department of Health Sciences, Universidade Federal de Lavras, Lavras (MG), Brazil. Medical Student, Medicine Course, Department of Health Sciences, Universidade Federal de Lavras, Lavras (MG), Brazil. Mailing a ddress : Joziana Muniz de Paiva Barçante E-mail: joziana@dsa.ufla.br FIgure 1: Ulcer measuring 2cm in diameter, with raised, red, and irregular edges, with central granulation tissue and serous exudate ©2019 by Anais Brasileiros de Dermatologia An Bras Dermatol. 2019;94(1):105-21. 114 Letters REFERENCES 1. Brasil. Ministério da Saúde. Secretaria de Vigilância em Saúde. Manual de Vigilância da Leishmaniose Tegumentar Americana. 2nd ed. Atual. Brasília: Ministério da Saúde; 2017. 189 p. 2. Kevric I, Cappel MA, Keeling JH. New World and Old World Leishmania Infections: A Practical Review. Dermatol Clin. 2015;33:579-93. 3. Murback NDN, Hans-Filho G, Nascimento RAF, Nakazato KRO, Dorval MEMC. American cutaneous leishmaniasis: clinical, epidemiological and laboratory studies conducted at a university teaching hospital in Campo Grande, Mato Grosso do Sul, Brazil. An Bras Dermatol. 2011;86:55-63. 4. Barçante TA, Botelho MC, Freitas HF, Soares GD, Barçante JM. First report of the main vector of visceral leishmaniasis in America, Lutzomyia longipalpis (Lutz, Neiva, 1912) (Diptera: Psychodidae: Phlebotominae), in southern Minas Gerais State, Brazil. J Vector Ecol. 2015;40:412-4. 5. Gomes CM, de Paula NA, de Morais OO, Soares KA, Roselino AM, Sampaio RNR. Complementary exams in the diagnosis of American tegumentary leishmaniasis. An Bras Dermatol. 2014;89:701-11. FIgure 2: Ulcer of approximately 2cm in diameter, red edges with center covered with meliceric crusts FIgure 3: Gel eletrophoresis of DNA products using polymerase chain reaction (PCR): 100pb - DNA Ladder; CN – Negative control; P1 - Case 1; P2 - Case 2; CP – Positive control with Leishmania DNA AUTHORS’CONTRIBUTIONS Early diagnosis of ATL is a difficult, but essential, task for Thales Augusto Barçante 0000-0002-2506-4065 the clinician, considering the toxicity of the drugs. Since the discov- Approval of the final version of the manuscript; Conception and planning of the study; ery of Leishmania parasite as causative agents of leishmaniasis, sev- Elaboration and writing of the manuscript; Effective participation in research orienta- tion; Critical review of the literature; Critical review of the manuscript eral tests have been developed. However, none of the tests available Marcos Vilela de Souza 0000-0001-7187-5283 today can be considered as the gold standard due to the lack of ac- Approval of the final version of the manuscript; Intellectual participation in propaedeu- curacy to detect the disease. The present study reports two cases of tic and/or therapeutic conduct of the cases studied; Critical review of the manuscript Allana Cristina Baltazar Fernandes 0000-0003-2630-9551 ATL with diagnosis confirmed by PCR and clinical cure after treat- Approval of the final version of the manuscript; Elaboration and writing of the manu- ment. These data point to the importance of systematized studies script; Intellectual participation in propaedeutic and/or therapeutic conduct of the cases studied about alternative methodologies to the intradermal Montenegro Joziana Muniz de Paiva Barçante 0000-0002-7963-5775 test, in view of the unavailability of the antigen used. Approval of the final version of the manuscript; Conception and planning of the study; The authors consider it important to report these cases of Elaboration and writing of the manuscript; Obtaining, analyzing and interpreting the data; Effective participation in research orientation; Critical review of the literature; Crit- ATL with unusual presentation in this municipality and to draw at- ical review of the manuscript tention to the fact that, although PCR is an exam that is still poorly accessed and expensive, in special situations, it can be very useful. How to cite this article: Barçante TA, Souza MV, Fernandes ACB, Furthermore, our findings show the importance of continuous health Barçante JMP. Use of the polymerase chain reaction for the diagnosis of epidemiological surveillance and emphasizes the need for permanent American tegumentary leishmaniasis. An Bras Dermatol. 2019;94(1):113-4. health education to avoid misdiagnosis or underreporting. q An Bras Dermatol. 2019;94(1):105-21. http://www.deepdyve.com/assets/images/DeepDyve-Logo-lg.png Anais Brasileiros de Dermatologia Pubmed Central

Use of the polymerase chain reaction for the diagnosis of American tegumentary leishmaniasis*

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Publisher
Pubmed Central
ISSN
0365-0596
eISSN
1806-4841
DOI
10.1590/abd1806-4841.20197929
Publisher site
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Abstract

Letters 113 in tissue samples in which the parasite forms could not be found. In 2017, two cases were investigated in Lavras. Case 1 was Use of the polymerase chain reaction for the a 39-year-old female patient who presented with a 5-month his- diagnosis of American tegumentary leishmaniasis tory of a lesion on the right arm. Clinical examination revealed a painful ulcer with pruritus measuring 2cm in diameter, with raised, hyperemic, and irregular edges, and central granulation tissue and Thales Augusto Barçante¹ serous exudate (Figure 1). Due to the unavailability of Montenegro Marcos Vilela de Souza¹ Allana Cristina Baltazar Fernandes² antigen for intradermal reaction, we performed a biopsy to collect Joziana Muniz de Paiva Barçante¹ lesional specimens. The biopsy was performed after the lesion was cleaned with soap and water and disinfected with ethyl alcohol 70%. We used 2% lidocaine for local anesthesia. Two punch biop- DOI: http://dx.doi.org/10.1590/abd1806-4841.20197929 sies were taken from the ulcer edge for histopathological analysis, which showed no amastigote forms. The second sample was used for imprint with staining, which also revealed no amastigote forms of the parasite. Subsequently, the same material was preserved in sterile saline solution, macerated with 300µg of extraction buffer, Dear Editor, and frozen in a properly identified flask for PCR. After analyzing Leishmaniasis is an infectious and parasitic disease with a the electrophoretic pattern of the sample with the positive control, variety of clinical forms, ranging from dermatological lesions (teg- the presence of Leishmania spp. DNA was confirmed. umentary leishmaniasis) to systemic manifestations (visceral leish- Case 2 was a 32-year-old female patient who had a lesion on 1-4 maniasis). American tegumentary leishmaniasis (ATL) represents the right thigh since October 2016. She was receiving antibiotic treat- a serious public health problem. Due to the high number of cases ment under medical supervision with transient clinical improve- of ATL in Brazil, differential diagnosis is necessary to understand ment. After 6 months, clinical examination revealed the presence of the epidemiological profile of ATL, both in areas previously con- a painful ulcer of approximately 2cm in diameter with hyperemic sidered non-endemic and in endemic areas. The diagnosis depends edges, and center covered with a meliceric crust (Figure 2). Simi- on the patient’s clinical history, epidemiology, and laboratory tests. larly, as described for case 1, we also performed histopathological Lavras, state of Minas Gerais, had 9 suspected ATL cases reported analysis and imprint with staining from five different slides and in the last 7 years, 8 of which were confirmed. Out of them, 7 cases rapid panoptic staining (Romanowsky), also with no identification were treated and considered cured, and 1 patient died from treat- of amastigote forms of the parasite. After the PCR, the presence of ment-associated toxicity. Furthermore, studies on the phlebotomine Leishmania spp. DNA was confirmed. fauna evidenced the presence of the main vectors in the munici- In both cases, after the detection of Leishmania DNA (Figure pality. Considering that epidemiological knowledge and clinical 3) and diagnostic confirmation of ATL, the treatment chosen was characterization of ATL are fundamental precepts of dermatological N-methylglucamine antimoniate at a daily intravenous dose of 13.7 practices, this case report aims to suggest polymerase chain reac- ml for 20 days. Both patients evolved to clinical cure after therapy. tion (PCR) as a possible diagnostic tool for cases where the Monte- negro antigen is not available and/or histopathological analysis is inconclusive. The PCR is capable of detecting the DNA of parasite Received 04 December 2017. Accepted 16 May 2018. * Work conducted at the Universidade Federal de Lavras, Lavras (MG), Brazil. Financial Support: FAPEMIG - APQ 02553-14. Conflict of interest: None. Department of Health Sciences, Universidade Federal de Lavras, Lavras (MG), Brazil. Medical Student, Medicine Course, Department of Health Sciences, Universidade Federal de Lavras, Lavras (MG), Brazil. Mailing a ddress : Joziana Muniz de Paiva Barçante E-mail: joziana@dsa.ufla.br FIgure 1: Ulcer measuring 2cm in diameter, with raised, red, and irregular edges, with central granulation tissue and serous exudate ©2019 by Anais Brasileiros de Dermatologia An Bras Dermatol. 2019;94(1):105-21. 114 Letters REFERENCES 1. Brasil. Ministério da Saúde. Secretaria de Vigilância em Saúde. Manual de Vigilância da Leishmaniose Tegumentar Americana. 2nd ed. Atual. Brasília: Ministério da Saúde; 2017. 189 p. 2. Kevric I, Cappel MA, Keeling JH. New World and Old World Leishmania Infections: A Practical Review. Dermatol Clin. 2015;33:579-93. 3. Murback NDN, Hans-Filho G, Nascimento RAF, Nakazato KRO, Dorval MEMC. American cutaneous leishmaniasis: clinical, epidemiological and laboratory studies conducted at a university teaching hospital in Campo Grande, Mato Grosso do Sul, Brazil. An Bras Dermatol. 2011;86:55-63. 4. Barçante TA, Botelho MC, Freitas HF, Soares GD, Barçante JM. First report of the main vector of visceral leishmaniasis in America, Lutzomyia longipalpis (Lutz, Neiva, 1912) (Diptera: Psychodidae: Phlebotominae), in southern Minas Gerais State, Brazil. J Vector Ecol. 2015;40:412-4. 5. Gomes CM, de Paula NA, de Morais OO, Soares KA, Roselino AM, Sampaio RNR. Complementary exams in the diagnosis of American tegumentary leishmaniasis. An Bras Dermatol. 2014;89:701-11. FIgure 2: Ulcer of approximately 2cm in diameter, red edges with center covered with meliceric crusts FIgure 3: Gel eletrophoresis of DNA products using polymerase chain reaction (PCR): 100pb - DNA Ladder; CN – Negative control; P1 - Case 1; P2 - Case 2; CP – Positive control with Leishmania DNA AUTHORS’CONTRIBUTIONS Early diagnosis of ATL is a difficult, but essential, task for Thales Augusto Barçante 0000-0002-2506-4065 the clinician, considering the toxicity of the drugs. Since the discov- Approval of the final version of the manuscript; Conception and planning of the study; ery of Leishmania parasite as causative agents of leishmaniasis, sev- Elaboration and writing of the manuscript; Effective participation in research orienta- tion; Critical review of the literature; Critical review of the manuscript eral tests have been developed. However, none of the tests available Marcos Vilela de Souza 0000-0001-7187-5283 today can be considered as the gold standard due to the lack of ac- Approval of the final version of the manuscript; Intellectual participation in propaedeu- curacy to detect the disease. The present study reports two cases of tic and/or therapeutic conduct of the cases studied; Critical review of the manuscript Allana Cristina Baltazar Fernandes 0000-0003-2630-9551 ATL with diagnosis confirmed by PCR and clinical cure after treat- Approval of the final version of the manuscript; Elaboration and writing of the manu- ment. These data point to the importance of systematized studies script; Intellectual participation in propaedeutic and/or therapeutic conduct of the cases studied about alternative methodologies to the intradermal Montenegro Joziana Muniz de Paiva Barçante 0000-0002-7963-5775 test, in view of the unavailability of the antigen used. Approval of the final version of the manuscript; Conception and planning of the study; The authors consider it important to report these cases of Elaboration and writing of the manuscript; Obtaining, analyzing and interpreting the data; Effective participation in research orientation; Critical review of the literature; Crit- ATL with unusual presentation in this municipality and to draw at- ical review of the manuscript tention to the fact that, although PCR is an exam that is still poorly accessed and expensive, in special situations, it can be very useful. How to cite this article: Barçante TA, Souza MV, Fernandes ACB, Furthermore, our findings show the importance of continuous health Barçante JMP. Use of the polymerase chain reaction for the diagnosis of epidemiological surveillance and emphasizes the need for permanent American tegumentary leishmaniasis. An Bras Dermatol. 2019;94(1):113-4. health education to avoid misdiagnosis or underreporting. q An Bras Dermatol. 2019;94(1):105-21.

Journal

Anais Brasileiros de DermatologiaPubmed Central

Published: Feb 1, 2019

References