Comparative Analysis of Microscopy, Leishmanin Skin Test (LST), and Direct Agglutination Test (DAT) for the Diagnosis of Human Cutaneous Leishmaniasis
-
Mohammad Hamedanipour
,
Mohammad Barati
,
Zahra Mirabedini
,
Mehdi Mohebali
,
Alireza Latifi
,
Zahra Kakooie
Abstract
Background: Cutaneous Leishmaniasis (CL) is a highly infectious parasitic disease in Iran. Although various diagnostic methods are available, identifying a sensitive and specific approach remains essential for effective treatment and disease management. We aimed to compare microscopy, the Leishmanin Skin Test (LST), and the Direct Agglutination Test (DAT) for diagnosis of human CL.
Methods: Fifty samples were obtained from clinically suspected CL cases. The results of conventional methods, including microscopy, the LST, and the DAT, were then compared among these samples. For microscopical examination considered a gold standard, smears were stained with Giemsa 10% and then examined for the observation of amastigote forms for the LST, 0.1 ml of standard Leishmanin solution was intradermally injected into positive patients, and then indurations were measured after 48-72 hours; an induration of ≥5 mm was considered positive. Additionally, anti-Leishmania antibodies were detected using DAT on positive collected serum samples.
Results: Out of 50 samples collected from individuals suspected of CL, 66% (33/50) of them showed positive results using microscopic examination. Among these 33 patients diagnosed with CL, 9% (3/33) tested positive in the LST, and only 1 (3%) patient showed specific antibodies against Leishmania using DAT.
Conclusion: The microscopy method appears to be more suitable for diagnosing CL. However, there is a clear need for additional diagnostic methods with more validity for CL.
1. Reithinger R, JC Dujardin, H Louzir, et al. Cutaneous leishmaniasis. Lancet Infect Dis. 2007;7(9):581-596.
2. Desjeux P. Leishmaniasis: current situation and new perspectives. Comp Immunol Microbiol Infect Dis. 2004;27(5):305-18.
3. Serafim TD, Coutinho-Abreu IV, Dey R, et al. Leishmaniasis: the act of transmission. Trends Parasitol. 2021;37(11):976-987.
4. Minodier P, Parola P. Cutaneous leishmaniasis treatment. Travel Med Infect Dis. 2007;5(3):150-158.
5. Barati M, Sharifi I, Sharififar F, et al. Anti-leishmanial activity of Gossypium hirsutum L., Ferula assa-foetida L. and Artemisia aucheri Boiss extracts by colorimetric assay. Anti-Infect Agents. 2014;12(2):159-164.
6. Oliveira LF, Schubach AO, Martins MM, et al. Systematic review of the adverse effects of cutaneous leishmaniasis treatment in the New World. Acta Trop. 2011;118(2):87-96.
7. Bailey F, Mondragon-Shem K, Hotez P, et al. A new perspective on cutaneous leishmaniasis—Implications for global prevalence and burden of disease estimates. PLoS Negl Trop Dis. 2017;11(8):0005739.
8. Karami M, Gorgani-Firouzjaee T, Chehrazi M. Prevalence of cutaneous Leishmaniasis in the Middle East: a systematic review and meta-analysis. Pathog Glob Health. 2023;117(4):356-365.
9. Hailu A. The use of direct agglutination test (DAT) in serological diagnosis of Ethiopian cutaneous leishmaniasis. Diagn Microbiol Infect Dis. 2002;42(4):251-256.
10. Rasti S, Ghorbanzadeh B, Kheirandish F, et al. Comparison of molecular, microscopic, and culture methods for diagnosis of cutaneous leishmaniasis. J Clin Lab Anal. 2016;30(5):610-615.
11. Vega-López F. Diagnosis of cutaneous leishmaniasis. Curr Opin Infect Dis. 2003;16(2):97-101.
12. Faber WR, Oskam L, van Gool T, et al. Value of diagnostic techniques for cutaneous leishmaniasis. J Am Acad Dermatol. 2003;49(1):70-74.
13. Aronson NE, Joya CA. Cutaneous leishmaniasis: updates in diagnosis and management. Infect Dis. Clin North Am. 2019;33(1):101-117.
14. Pourmohammadi B. Motazedian MH, Hatam GR, et al. Comparison of three methods for diagnosis of cutaneous leishmaniasis. Iranian J Parasitol. 2010;5(4):1-8.
15. Singh S. New developments in diagnosis of leishmaniasis. Indian J Med Res. 2006;123(3):311-30.
16. Carstens-Kass J, Paulini K, Lypaczewski P, et al. A review of the Leishmanin skin test: A neglected test for a neglected disease. PLoS Negl Trop Dis. 2021;15(7):e0009531.
17. Manzur A. Sensitivity of Leishmanin skin test in patients of acute cutaneous leishmaniasis. Dermatol Online J. 2006;12(4): 2.
18. Hashemi SN, Mohebali M, Mansouri P, et al. Comparison of Leishmanin skin test and direct smear for the diagnosis of cutaneous leishmaniasis. Acta Med Iran. 2011; 49(3):136-41.
19. Alimohammadian M, Hakimi H, Nikseresht M. The preparation and evaluation of reference Leishmanin from Leishmania major for use in man for diagnostic and experimental purposes. Med J Islam Repub Iran. 1993;7:23–8.
20. Mohebali M, Edrisian G. H, Nadim A, et al. Application of direct agglutination test (DAT) for the diagnosis and seroepidemiological studies of visceral leishmaniasis in Iran. Iran J Parasitol. 2006;1:15-25.
21. Mohebali M, Keshavarz H, Shirmoham-mad S, et al. The diagnostic accuracy of direct agglutination test for serodiagnosis of human visceral leishmaniasis: a systematic review with meta-analysis. BMC Infect Dis. 2020;20:946.
22. Molaie S, Mohebali M, Abaiet MR, et al. Alteration of Direct Agglutination Test (DAT) results in Iranian Kala-Azar patients: a case series. J Parasit Dis. 2017;41:446-449.
23. Mengistu G, Kiessling R, Akuffo H. The value of a direct agglutination test in the diagnosis of cutaneous and visceral leishmaniasis in Ethiopia. Trans R Soc Trop Med Hyg. 1990;84(3):359-362.
24. Sabzevari, S, Teshnizi SH, Shokri A, et al. Cutaneous leishmaniasis in Iran: a systematic review and meta-analysis. Microb Pathog. 2021;152:104721.
25. Feiz-Haddad MH, Kassiri H, Kasiri N. Prevalence and epidemiologic profile of acute cutaneous leishmaniasis in an endemic focus, Southwestern Iran. J Acute Dis. 2015;4:292-297.
26. Sharifi I, Khosravi A, Aflatoonian MR, et al. Cutaneous leishmaniasis situation analysis in the Islamic Republic of Iran in preparation for an elimination plan. Front Public Health. 2023; 11:1091709.
27. Miramin-Mohammadi A, Javadi A, Eskandari SE, et al. Immune response in cutaneous leishmaniasis patients with healing vs. non-healing lesions. Iran J Microbiol. 2020;12(3):249-255.
28. Alimohammadian MH, Jones SL, Darabi H, et al. Assessment of interferon-γ levels and Leishmanin skin test results in persons recovered for leishmaniasis. Am J Trop Med Hyg. 2012;87(1):70-5.
29. Sadeghian G, Ziaei H, Bidabadi LS, et al. Evaluation of Leishmanin skin test reaction in different variants of cutaneous leishmaniasis. Indian J Dermatol. 2013;58(3):239.
30. Norouzinezhad F, Ghaffari F, Norouzinejad A, et al. Cutaneous lei-shmaniasis in Iran: results from an epidemiological study in urban and rural provinces. Asian Pac J Trop Biomed. 2016;6(7):614-619.
31. Pouresmaeiliyan S, Sharifi I, Aflatoniyan MR, et al. A new focus of anthroponotic cutaneous leishmaniasis in Dehbakry region of Bam district, southeastern Iran 2008. J Kerman Univ Med Sci.. 2010;16(1):15-24.
32. Mehregan DR, Mehregan AH, Mehregan DA, et al. Histologic diagnosis of cutaneous leishmaniasis. Clin Dermatol. 1999;17(3):297-304.
33. Ramírez JR, Agudelo S, Muskus C, et al. Diagnosis of cutaneous leishmaniasis in Colombia: the sampling site within lesions influences the sensitivity of parasitologic diagnosis. J Clin Microbiol. 2000;38(10):3768-3773.
34. Saab MEl, Hage H, Charafeddine K, et al. Diagnosis of cutaneous leishmaniasis: why punch when you can scrape? Am J Trop Med Hyg. 2015;92(3):518-22.
35. Weigle KA, de Davalos M, Heredia, et al. Diagnosis of cutaneous and mucocutaneous leishmaniasis in Colombia: a comparison of seven methods. Am J Trop Med Hyg. 1987;36(3):489-496.
36. Navin TR., Arana FE, de Mérid AM, et al. Cutaneous leishmaniasis in Guatemala: comparison of diagnostic methods. Am J Trop Med Hyg.1990;42(1)36-42.
37. Pena HP, Belo VS, Xavier-Junior JCC, et al. Accuracy of diagnostic tests for American tegumentary leishmaniasis: a systematic literature review with meta-analyses. Trop Med Int Health. 2020;25(10):1168-1181.
38. Azmi K, Nasereddin A, Ereqat S, et al. Methods incorporating a polymerase chain reaction and restriction fragment length polymorphism and their use as a ‘gold standard’in diagnosing Old World cutaneous leishmaniasis. Diagn Microbiol Infect Dis. 2011;71(2):151-155.
39. de Vries HJ, Reedijk SH, Schallig HD, et al. Cutaneous leishmaniasis: recent developments in diagnosis and management. Am J Clin Dermatol. 2015;16:99-109.
40. Montenegro J. Cutaneous reaction in leishmaniasis. Arch Dermatol Syph, 1926;13(2):187-194.
41. Pampiglione S, Manson-Bahr PEC, La Placa M, et al. Studies on Mediterranean leishmaniasis IV. The Leishmanin skin test in cutaneous leishmaniasis. Trans R Soc Trop Med Hyg. 1976;70(1):62-65.
42. Manson-Bahr P. Clinical aspects and control: Diagnosis in: The leishmaniases in biology and medicine. 1987;Academic Press, London & New York.
43. Dostrovsky A, Sagher F. The intracutaneous test in cutaneous leishmaniasis. Ann Trop Med Parasitol. 1946;40(3-4):265-269.
44. Sadeghian G, Momeni A, Siadat AH, et al. Evaluation of Leishmanin skin test and its relationship with the clinical form and duration of cutaneous leishmaniasis. Dermatol Online J. 2006;12(7):3.
45. Alimohammadian MH, Fariba Kojori Z, Darabi H, et al. Soluble Leishmanin as an Ideal Reagent for Skin Testing in Human Leishmaniasis. Iran Biomed J. 1997;1(4):39-47.
46. Roberts T, Keddie SH, Rattanavong S, et al. Accuracy of the direct agglutination test for diagnosis of visceral leishmaniasis: a systematic review and meta-analysis. BMC Infect Dis. 2023;23(1):782.
47. El Safi S, Evans DA. A comparison of the direct agglutination test and enzyme-linked immunosorbent assay in the sero-diagnosis of leishmaniasis in the Sudan. Trans R Soc Trop Med Hyg. 1989;83(3):334-337.
2. Desjeux P. Leishmaniasis: current situation and new perspectives. Comp Immunol Microbiol Infect Dis. 2004;27(5):305-18.
3. Serafim TD, Coutinho-Abreu IV, Dey R, et al. Leishmaniasis: the act of transmission. Trends Parasitol. 2021;37(11):976-987.
4. Minodier P, Parola P. Cutaneous leishmaniasis treatment. Travel Med Infect Dis. 2007;5(3):150-158.
5. Barati M, Sharifi I, Sharififar F, et al. Anti-leishmanial activity of Gossypium hirsutum L., Ferula assa-foetida L. and Artemisia aucheri Boiss extracts by colorimetric assay. Anti-Infect Agents. 2014;12(2):159-164.
6. Oliveira LF, Schubach AO, Martins MM, et al. Systematic review of the adverse effects of cutaneous leishmaniasis treatment in the New World. Acta Trop. 2011;118(2):87-96.
7. Bailey F, Mondragon-Shem K, Hotez P, et al. A new perspective on cutaneous leishmaniasis—Implications for global prevalence and burden of disease estimates. PLoS Negl Trop Dis. 2017;11(8):0005739.
8. Karami M, Gorgani-Firouzjaee T, Chehrazi M. Prevalence of cutaneous Leishmaniasis in the Middle East: a systematic review and meta-analysis. Pathog Glob Health. 2023;117(4):356-365.
9. Hailu A. The use of direct agglutination test (DAT) in serological diagnosis of Ethiopian cutaneous leishmaniasis. Diagn Microbiol Infect Dis. 2002;42(4):251-256.
10. Rasti S, Ghorbanzadeh B, Kheirandish F, et al. Comparison of molecular, microscopic, and culture methods for diagnosis of cutaneous leishmaniasis. J Clin Lab Anal. 2016;30(5):610-615.
11. Vega-López F. Diagnosis of cutaneous leishmaniasis. Curr Opin Infect Dis. 2003;16(2):97-101.
12. Faber WR, Oskam L, van Gool T, et al. Value of diagnostic techniques for cutaneous leishmaniasis. J Am Acad Dermatol. 2003;49(1):70-74.
13. Aronson NE, Joya CA. Cutaneous leishmaniasis: updates in diagnosis and management. Infect Dis. Clin North Am. 2019;33(1):101-117.
14. Pourmohammadi B. Motazedian MH, Hatam GR, et al. Comparison of three methods for diagnosis of cutaneous leishmaniasis. Iranian J Parasitol. 2010;5(4):1-8.
15. Singh S. New developments in diagnosis of leishmaniasis. Indian J Med Res. 2006;123(3):311-30.
16. Carstens-Kass J, Paulini K, Lypaczewski P, et al. A review of the Leishmanin skin test: A neglected test for a neglected disease. PLoS Negl Trop Dis. 2021;15(7):e0009531.
17. Manzur A. Sensitivity of Leishmanin skin test in patients of acute cutaneous leishmaniasis. Dermatol Online J. 2006;12(4): 2.
18. Hashemi SN, Mohebali M, Mansouri P, et al. Comparison of Leishmanin skin test and direct smear for the diagnosis of cutaneous leishmaniasis. Acta Med Iran. 2011; 49(3):136-41.
19. Alimohammadian M, Hakimi H, Nikseresht M. The preparation and evaluation of reference Leishmanin from Leishmania major for use in man for diagnostic and experimental purposes. Med J Islam Repub Iran. 1993;7:23–8.
20. Mohebali M, Edrisian G. H, Nadim A, et al. Application of direct agglutination test (DAT) for the diagnosis and seroepidemiological studies of visceral leishmaniasis in Iran. Iran J Parasitol. 2006;1:15-25.
21. Mohebali M, Keshavarz H, Shirmoham-mad S, et al. The diagnostic accuracy of direct agglutination test for serodiagnosis of human visceral leishmaniasis: a systematic review with meta-analysis. BMC Infect Dis. 2020;20:946.
22. Molaie S, Mohebali M, Abaiet MR, et al. Alteration of Direct Agglutination Test (DAT) results in Iranian Kala-Azar patients: a case series. J Parasit Dis. 2017;41:446-449.
23. Mengistu G, Kiessling R, Akuffo H. The value of a direct agglutination test in the diagnosis of cutaneous and visceral leishmaniasis in Ethiopia. Trans R Soc Trop Med Hyg. 1990;84(3):359-362.
24. Sabzevari, S, Teshnizi SH, Shokri A, et al. Cutaneous leishmaniasis in Iran: a systematic review and meta-analysis. Microb Pathog. 2021;152:104721.
25. Feiz-Haddad MH, Kassiri H, Kasiri N. Prevalence and epidemiologic profile of acute cutaneous leishmaniasis in an endemic focus, Southwestern Iran. J Acute Dis. 2015;4:292-297.
26. Sharifi I, Khosravi A, Aflatoonian MR, et al. Cutaneous leishmaniasis situation analysis in the Islamic Republic of Iran in preparation for an elimination plan. Front Public Health. 2023; 11:1091709.
27. Miramin-Mohammadi A, Javadi A, Eskandari SE, et al. Immune response in cutaneous leishmaniasis patients with healing vs. non-healing lesions. Iran J Microbiol. 2020;12(3):249-255.
28. Alimohammadian MH, Jones SL, Darabi H, et al. Assessment of interferon-γ levels and Leishmanin skin test results in persons recovered for leishmaniasis. Am J Trop Med Hyg. 2012;87(1):70-5.
29. Sadeghian G, Ziaei H, Bidabadi LS, et al. Evaluation of Leishmanin skin test reaction in different variants of cutaneous leishmaniasis. Indian J Dermatol. 2013;58(3):239.
30. Norouzinezhad F, Ghaffari F, Norouzinejad A, et al. Cutaneous lei-shmaniasis in Iran: results from an epidemiological study in urban and rural provinces. Asian Pac J Trop Biomed. 2016;6(7):614-619.
31. Pouresmaeiliyan S, Sharifi I, Aflatoniyan MR, et al. A new focus of anthroponotic cutaneous leishmaniasis in Dehbakry region of Bam district, southeastern Iran 2008. J Kerman Univ Med Sci.. 2010;16(1):15-24.
32. Mehregan DR, Mehregan AH, Mehregan DA, et al. Histologic diagnosis of cutaneous leishmaniasis. Clin Dermatol. 1999;17(3):297-304.
33. Ramírez JR, Agudelo S, Muskus C, et al. Diagnosis of cutaneous leishmaniasis in Colombia: the sampling site within lesions influences the sensitivity of parasitologic diagnosis. J Clin Microbiol. 2000;38(10):3768-3773.
34. Saab MEl, Hage H, Charafeddine K, et al. Diagnosis of cutaneous leishmaniasis: why punch when you can scrape? Am J Trop Med Hyg. 2015;92(3):518-22.
35. Weigle KA, de Davalos M, Heredia, et al. Diagnosis of cutaneous and mucocutaneous leishmaniasis in Colombia: a comparison of seven methods. Am J Trop Med Hyg. 1987;36(3):489-496.
36. Navin TR., Arana FE, de Mérid AM, et al. Cutaneous leishmaniasis in Guatemala: comparison of diagnostic methods. Am J Trop Med Hyg.1990;42(1)36-42.
37. Pena HP, Belo VS, Xavier-Junior JCC, et al. Accuracy of diagnostic tests for American tegumentary leishmaniasis: a systematic literature review with meta-analyses. Trop Med Int Health. 2020;25(10):1168-1181.
38. Azmi K, Nasereddin A, Ereqat S, et al. Methods incorporating a polymerase chain reaction and restriction fragment length polymorphism and their use as a ‘gold standard’in diagnosing Old World cutaneous leishmaniasis. Diagn Microbiol Infect Dis. 2011;71(2):151-155.
39. de Vries HJ, Reedijk SH, Schallig HD, et al. Cutaneous leishmaniasis: recent developments in diagnosis and management. Am J Clin Dermatol. 2015;16:99-109.
40. Montenegro J. Cutaneous reaction in leishmaniasis. Arch Dermatol Syph, 1926;13(2):187-194.
41. Pampiglione S, Manson-Bahr PEC, La Placa M, et al. Studies on Mediterranean leishmaniasis IV. The Leishmanin skin test in cutaneous leishmaniasis. Trans R Soc Trop Med Hyg. 1976;70(1):62-65.
42. Manson-Bahr P. Clinical aspects and control: Diagnosis in: The leishmaniases in biology and medicine. 1987;Academic Press, London & New York.
43. Dostrovsky A, Sagher F. The intracutaneous test in cutaneous leishmaniasis. Ann Trop Med Parasitol. 1946;40(3-4):265-269.
44. Sadeghian G, Momeni A, Siadat AH, et al. Evaluation of Leishmanin skin test and its relationship with the clinical form and duration of cutaneous leishmaniasis. Dermatol Online J. 2006;12(7):3.
45. Alimohammadian MH, Fariba Kojori Z, Darabi H, et al. Soluble Leishmanin as an Ideal Reagent for Skin Testing in Human Leishmaniasis. Iran Biomed J. 1997;1(4):39-47.
46. Roberts T, Keddie SH, Rattanavong S, et al. Accuracy of the direct agglutination test for diagnosis of visceral leishmaniasis: a systematic review and meta-analysis. BMC Infect Dis. 2023;23(1):782.
47. El Safi S, Evans DA. A comparison of the direct agglutination test and enzyme-linked immunosorbent assay in the sero-diagnosis of leishmaniasis in the Sudan. Trans R Soc Trop Med Hyg. 1989;83(3):334-337.
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| Issue | Vol 20 No 1 (2025) | |
| Section | Original Article(s) | |
| Keywords | ||
| Cutaneous leishmaniasis Microscopy test Leishmanin skin test Direct agglutination test Human | ||
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How to Cite
1.
Hamedanipour M, Barati M, Mirabedini Z, Mohebali M, Latifi A, Kakooie Z. Comparative Analysis of Microscopy, Leishmanin Skin Test (LST), and Direct Agglutination Test (DAT) for the Diagnosis of Human Cutaneous Leishmaniasis. Iran J Parasitol. 2025;20(1):13-20.
