Surveillance of cutaneous leishmaniasis in clinical samples: distribution of Leishmania guyanensis in the state of Amapá, Brazil, 2018**This research received financial support from the Amapá State Research Support Foundation (FAPEAP) - Process No. 250.203.039/2016 - and a doctoral Research Grant from the Pará State Research Support Foundation (FAPESPA) - Process No. 003/2014.

Ariely Nunes Ferreira de Almeida Luciana de Cássia Silva do Nascimento Edith Silvia Moura de Moura Sousa Afonso José Diger de Oliveira Maria Gorete de Sena Breno Maués de Resende Raimunda Cleide Gonçalves Chaves Lourdes Maria Garcez About the authors

Abstract

Objective:

to investigate Leishmania species in a series of autochthonous cutaneous leishmaniasis (CL) cases in Amapá State, Brazilian Amazon.

Methods:

this was a descriptive ecological study carried out from January-October/2018 at a reference center for CL diagnosis in Amapá; individuals with CL receiving care from January-May/2018 were recruited; clinical data and skin biopsies were obtained; from extracted DNA (phenol-chloroform) we amplified the hsp70-234 gene region (PCR) for nucleotide sequencing (Applied Biosystems: ABI3500XL).

Results:

38 individuals were interviewed, examined and diagnosed; men predominated (28/38; mean age=32.5±11.3); lesions (most ulcers: 37/38) measuring 0,4-10mm (34/38) and ≥11mm (4/38) were multiple in 20/38 individuals; diagnosis of L. braziliensis (1), L. naiffi (1), L. infantum (1), L. (Viannia) sp. (1), L. amazonensis (2) and L. guyanensis (32); individuals infected with L. guyanensis (32/38) lived in 9/10 municipalities represented in the sample, and 17/32 of these had multiple lesions.

Conclusion:

presence of Leishmania guyanensis predominated and was frequently associated with multiple lesions.

Keywords:
Leishmaniasis, Cutaneous; Leishmania guyanensis; Molecular Epidemiology; Amazonian Ecosystem; Epidemiology, Descriptive

Introduction

Leishmaniasis is a neglected tropical disease caused by protozoa of the Leishmania genus and has two main clinical forms: tegumentary leishmaniasis (TL), which can be cutaneous and/or mucousal; and visceral leishmaniasis (VL).11. Ministério da Saúde (BR). Secretaria de Vigilância das Doenças Transmissíveis. Manual de vigilância da leishmaniose tegumentar [Internet]. Brasília: Ministério da Saúde; 2017 [citado 2018 nov 21]. 190 p. Disponível em: Disponível em: http://bvsms.saude.gov.br/bvs/publicacoes/manual_vigilancia_leishmaniose_tegumentar.pdf
http://bvsms.saude.gov.br/bvs/publicacoe...

Between 2007 and 2015, 6,801 cases of cutaneous leishmaniasis (CL) were registered in the state of Amapá.22. Almeida ANF, Garcez LM, Araújo OCL. Leishmaniose tegumentar americana no estado do Amapá, Brasil: 2007 a 2015 [Internet]. In: Anais do 53º Congresso da Sociedade Brasileira de Medicina Tropical. 2017 ago 27-30; Campinas: Galoá; 2018 [citado 2018 nov 21]. Disponível em: Disponível em: https://proceedings.science/medtrop/papers/leishmaniose-tegumentar-americana-no-estado-do-amapa%2C-brasil%3A-2007-a-2015
https://proceedings.science/medtrop/pape...
CL can progress to severe although not fatal forms. Notwithstanding, between 2017 and 2018, the Amapá State Health Surveillance Superintendency recorded four deaths of patients with CL being treated with Glucantime®33. Superintendência de Vigilância em Saúde (AP). Situação epidemiológica da leishmaniose tegumentar no estado do Amapá: período de 2017 a novembro de 2018. Bol Epidemiol [Internet]. 2018 [citado 2019 mar 12];2:1-5. Disponível em: Disponível em: https://editor.amapa.gov.br/arquivos_portais/publicacoes/SVS_4d77b443923909a984f01e74bf38240f.pdf
https://editor.amapa.gov.br/arquivos_por...
(meglumine antimoniate) and 19 cases of canine VL.44. Superintendência de Vigilância em Saúde (AP). Diretoria Executiva de Vigilância em Saúde. Nota informativa: confirmado primeiro caso autoctone de leishmaniose visceral humana no estado do Amapá [Internet]. Amapá: Superintendência de Vigilância em Saúde; 2018 [citado 2019 mar 12]. Disponível em: Disponível em: https://drive.google.com/file/d/1DA51roHaaHUQpKj00O3RveDO6nPbgJB7/view
https://drive.google.com/file/d/1DA51roH...
Also in 2018 the first autochthonous case of human VL in the state was confirmed: a riverside dweller aged 82 who was infected in the municipality of Mazagão and died from complications caused by cirrhosis of the liver.44. Superintendência de Vigilância em Saúde (AP). Diretoria Executiva de Vigilância em Saúde. Nota informativa: confirmado primeiro caso autoctone de leishmaniose visceral humana no estado do Amapá [Internet]. Amapá: Superintendência de Vigilância em Saúde; 2018 [citado 2019 mar 12]. Disponível em: Disponível em: https://drive.google.com/file/d/1DA51roHaaHUQpKj00O3RveDO6nPbgJB7/view
https://drive.google.com/file/d/1DA51roH...

Surveillance of VL, which is caused by the etiological agent Leishmania infantum, favors early diagnosis and treatment, thus reducing risk of death. Nevertheless, it is uncommon for people with CL to die and death usually results from comorbidities exacerbated by the toxicity of Glucantime®.55. Sundar S, Chakravarty J. Leishmaniasis: an update of current pharmacotherapy. Expert Opin Pharmacother [Internet]. 2013 Jan [cited 2019 Jul 16];14(1):53-63. Available from: Available from: https://www.tandfonline.com/doi/full/10.1517/14656566.2013.755515 . doi: 10.1517/14656566.2013.755515
https://www.tandfonline.com/doi/full/10....
Among the agents causing CL, Leishmania braziliensis and, principally, Leishmania guyanensis, show resistance to Glucantime®, the prolonged use of which increases the risk of adverse events.66. Romero GA, Guerra MV, Paes MG, Macêdo VO. Comparison of cutaneous leishmaniasis due to Leishmania (Viannia) braziliensis and L. (V.) guyanensis in Brazil: therapeutic response to meglumine antimoniate. Am J Trop Med Hyg [Internet]. 2001 Nov [cited 2019 Jul 16];65(5):456-65. Available from: Available from: http://www.ajtmh.org/docserver/fulltext/14761645/65/5/11716098.pdf?expires=1563806586&id=id&accname=guest&checksum=521DD0993B0869A3D5A44A65ECE28098
http://www.ajtmh.org/docserver/fulltext/...
L. guyanensis occurs in the North of Brazil11. Ministério da Saúde (BR). Secretaria de Vigilância das Doenças Transmissíveis. Manual de vigilância da leishmaniose tegumentar [Internet]. Brasília: Ministério da Saúde; 2017 [citado 2018 nov 21]. 190 p. Disponível em: Disponível em: http://bvsms.saude.gov.br/bvs/publicacoes/manual_vigilancia_leishmaniose_tegumentar.pdf
http://bvsms.saude.gov.br/bvs/publicacoe...
,77. Araujo-Pereira T, Pita-Pereira D, Moreira RB, Silva-Galdino T, Duarte MPO, Brazil RP, et al . Molecular diagnosis of cutaneous leishmaniasis in an endemic area of Acre State in the Amazonian Region of Brazil. Rev Soc Bras Med Trop [Internet]. 2018 May-Jun [cited 2019 Jul 16];51(3):376-81. Available from: Available from: http://www.scielo.br/pdf/rsbmt/v51n3/1678-9849-rsbmt-51-03-376.pdf . doi: 10.1590/0037-8682-0232-2017
http://www.scielo.br/pdf/rsbmt/v51n3/167...
,88. Lainson R. Espécies neotropicais de Leishmania: uma breve revisão histórica sobre sua descoberta, ecologia e taxonomia. Rev Pan-Amaz Saúde [Internet]. 2010 jun [citado 2019 jul 16];1(2):13-32. Disponível em: Disponível em: http://scielo.iec.gov.br/pdf/rpas/v1n2/pt_v1n2a02.pdf . doi: 10.5123/S2176-62232010000200002
http://scielo.iec.gov.br/pdf/rpas/v1n2/p...
but its frequency and distribution in Amapá are unknown.

The objective of this study was to investigate Leishmania species in a series of autochthonous cutaneous leishmaniasis cases in Amapá in 2018.

Methods

We conducted a descriptive study of a series of CL cases. The Brazilian state of Amapá has 670,000 inhabitants, distributed over 16 municipalities.9 One side of the state faces the Atlantic Ocean, while to the North it borders with Suriname and French Guiana, and to the south it borders with the Brazilian state of Pará. In 2018, the CL detection coefficient in Amapá was 68.7/100,000 inhabitants, with autochthonous cases in 14 of the state’s 16 municipalities.33. Superintendência de Vigilância em Saúde (AP). Situação epidemiológica da leishmaniose tegumentar no estado do Amapá: período de 2017 a novembro de 2018. Bol Epidemiol [Internet]. 2018 [citado 2019 mar 12];2:1-5. Disponível em: Disponível em: https://editor.amapa.gov.br/arquivos_portais/publicacoes/SVS_4d77b443923909a984f01e74bf38240f.pdf
https://editor.amapa.gov.br/arquivos_por...
Phlebotomine insects, which are potential CL vectors, are abundant in this Amazon subregion,88. Lainson R. Espécies neotropicais de Leishmania: uma breve revisão histórica sobre sua descoberta, ecologia e taxonomia. Rev Pan-Amaz Saúde [Internet]. 2010 jun [citado 2019 jul 16];1(2):13-32. Disponível em: Disponível em: http://scielo.iec.gov.br/pdf/rpas/v1n2/pt_v1n2a02.pdf . doi: 10.5123/S2176-62232010000200002
http://scielo.iec.gov.br/pdf/rpas/v1n2/p...
,1010. Saraiva JF, Souto RNP, Ferreira RMA. Flebotomineos (diptera: psychodidae) coletados em um assentamento rural no estado do Amapá, Brasil. Biota Amazônia [Internet]. 2011 [citado 2019 jul 16];1(1):58-62. Disponível em: Disponível em: https://periodicos.unifap.br/index.php/biota/article/view/145/v1n1p58-62.pdf . doi: 10.18561/2179-5746/biotaamazonia.v1n1p58-62
https://periodicos.unifap.br/index.php/b...
,1111. Galardo AKR, Galardo CD, Santana AA, Mendes JCC, Souza FRA, Duarte JP, et al. Primeira ocorrência de Lutzomyia (Lutzomyia) longipalpis LUTZ & NEIVA, 1912 (Diptera: Psychodidae: Phlebotominae) no estado do Amapá, Brasil. Biota Amazônia [Internet]. 2013 [citado 2019 jul 16];3(2):179-83. Disponível em: Disponível em: https://periodicos.unifap.br/index.php/biota/article/view/688/v3n2p179-183.pdf . doi: 10.18561/2179-5746/biotaamazonia.v3n2p179-183
https://periodicos.unifap.br/index.php/b...
where CL is endemic.22. Almeida ANF, Garcez LM, Araújo OCL. Leishmaniose tegumentar americana no estado do Amapá, Brasil: 2007 a 2015 [Internet]. In: Anais do 53º Congresso da Sociedade Brasileira de Medicina Tropical. 2017 ago 27-30; Campinas: Galoá; 2018 [citado 2018 nov 21]. Disponível em: Disponível em: https://proceedings.science/medtrop/papers/leishmaniose-tegumentar-americana-no-estado-do-amapa%2C-brasil%3A-2007-a-2015
https://proceedings.science/medtrop/pape...
,33. Superintendência de Vigilância em Saúde (AP). Situação epidemiológica da leishmaniose tegumentar no estado do Amapá: período de 2017 a novembro de 2018. Bol Epidemiol [Internet]. 2018 [citado 2019 mar 12];2:1-5. Disponível em: Disponível em: https://editor.amapa.gov.br/arquivos_portais/publicacoes/SVS_4d77b443923909a984f01e74bf38240f.pdf
https://editor.amapa.gov.br/arquivos_por...

In order to collect data, we recruited individuals attending the Amapá Tropical Diseases Reference Center between January and May 2018. The inclusion criteria were: males and females over 12 years old, with primary lesions suggestive of CL for at least two weeks. Those with medical contraindication to anesthesia and/or biopsy or with some other laboratory diagnosis were excluded.

The Leishmania species identified in the study were assessed in relation to the following variables:

  1. a) sociodemographic variables

  2. - sex;

  3. - municipality of infection;

  4. - mean age; and

  5. - age groups;

  6. b) clinical variables

  7. - number of cases;

  8. - type of lesion (ulcer; verrucous);

  9. - diameter of lesion;

  10. - site of lesions on the body; and

  11. - predominant clinical manifestations (crusting, secondary infection, satellite lesions etc.).

Leishmania variable measurement and diagnosis were based on leishmaniasis surveillance criteria defined in the Ministry of Health technical manual,1 as well as being based on molecular techniques.1212. Graça GC, Volpini AC, Romero GAS, Oliveira Neto MP, Hueb M, Porrozzi R, et al. Development and validation of PCR-based assays for diagnosis of American cutaneous leishmaniasis and identification of the parasite species. Mem Inst Oswaldo Cruz [Internet]. 2012 Aug [cited 2019 Jul 16];107(5):664-74. Available from: Available from: http://www.scielo.br/pdf/mioc/v107n5/14.pdf . doi: 10.1590/S0074-02762012000500014
http://www.scielo.br/pdf/mioc/v107n5/14....
,1313. Altschul SF, Gish W, Miller W, Myers EW, Lipman DJ. Basic local alignment search tool. J Mol Biol [Internet]. 1990 Oct [cited 2019 Jul 16];215(3):403-10. Available from: Available from: https://www.sciencedirect.com/science/article/pii/S0022283605803602?via%3Dihub . doi: 10.1016/S0022-2836(05)80360-2
https://www.sciencedirect.com/science/ar...

Lesion exudates were used to confirm infection, obtained via swabs for smearing on glass slides, using Giemsa stain to examine for amastigotes under the microscope.11. Ministério da Saúde (BR). Secretaria de Vigilância das Doenças Transmissíveis. Manual de vigilância da leishmaniose tegumentar [Internet]. Brasília: Ministério da Saúde; 2017 [citado 2018 nov 21]. 190 p. Disponível em: Disponível em: http://bvsms.saude.gov.br/bvs/publicacoes/manual_vigilancia_leishmaniose_tegumentar.pdf
http://bvsms.saude.gov.br/bvs/publicacoe...
Leishmania were characterized using DNA extracted from the skin (phenol-chloroform). Skin was collected via lesion biopsy using local anesthesia (2% lidocaine). The sample was kept in a flask containing 0.5mL of NET (NaCl 0.15mM; EDTA 50mM; Tris-HCl 0.1M/pH 7.5). In order to ensure the quality of the extracted skin/DNA samples, refrigerator temperature was controlled daily (2-8oC). Polymerase chain reaction (PCR) was performed with subsequent sequencing of the 234 base pair target, i.e. Leishmania DNA heat shock-protein 70 gene coding region (hsp70-234).1212. Graça GC, Volpini AC, Romero GAS, Oliveira Neto MP, Hueb M, Porrozzi R, et al. Development and validation of PCR-based assays for diagnosis of American cutaneous leishmaniasis and identification of the parasite species. Mem Inst Oswaldo Cruz [Internet]. 2012 Aug [cited 2019 Jul 16];107(5):664-74. Available from: Available from: http://www.scielo.br/pdf/mioc/v107n5/14.pdf . doi: 10.1590/S0074-02762012000500014
http://www.scielo.br/pdf/mioc/v107n5/14....

The following assays were performed:

  1. a) Polymerase chain reaction - PCR

  2. PCR was based on Leishmania hsp70-234, highly sensitive for protozoan DNA detection and species discrimination.12 Assay conditions: 50mL Mix (Taq DNA polymerase 0.03U/µL; MgCl2 1.5mM; Invitrogen® KCl buffer; dNTPs 0.25mM; F- 5’ GGA CGA GAT CGA GCG CAT GGT 3’ e R- 5’ TCC TTC GAC GCC TCC TGG TTG 3’ initiators, 0.2pM each; DNA 3.0µL); denaturation, annealing and extension (1x 94°C/5’; 32x 94°C/0,5’, 61oC/1’ and 72oC/1’); and final extension (72°C/10’). A positive result confirms the presence of Leishmania DNA in the lesion, since it signifies amplification of the hsp70-234 gene fragment.1212. Graça GC, Volpini AC, Romero GAS, Oliveira Neto MP, Hueb M, Porrozzi R, et al. Development and validation of PCR-based assays for diagnosis of American cutaneous leishmaniasis and identification of the parasite species. Mem Inst Oswaldo Cruz [Internet]. 2012 Aug [cited 2019 Jul 16];107(5):664-74. Available from: Available from: http://www.scielo.br/pdf/mioc/v107n5/14.pdf . doi: 10.1590/S0074-02762012000500014
    http://www.scielo.br/pdf/mioc/v107n5/14....

  3. b) DNA sequencing

  4. This procedure identifies the sequence of nucleotide bases of the hsp70-234 gene region, confirming the distinction between Leishmania species by comparing the sequence with well characterized sequences available in the GenBank. The PCR-hsp70-234 products were therefore purified (illustra ExoProStar; GE Healthcare Life Science) and sequenced (forward and reverse) in the automatic ABI3500XL DNA analyzer, using the BigDye® Terminator v3.1 Cycle Sequencing Kit (Applied Biosystems). BioEdit and Basic Local Alignment Search Tool (BLAST) were used to analyze sequences.13 Control samples were comprised of DNA from L. braziliensis (MHOM/BR/1975/M2904), L. guyanensis (MHOM/BR/1975/M4147), L. lainsoni (MHOM/BR1881/M6426), L. shawi (MCEB/BR/1984/M8408), L. naiffi (MDAS/BR/1979/M533) and L. amazonensis (IFLA/BR/1968/PH8).

We used BioStat 5.0 to produce frequency tables and to calculate means and standard deviations (available at: www.mamiraua.org.br).

The study project was approved by the Federal University of Amapá Research Ethics Committee (Certification of Submission for Ethical Appraisal [CAAE] No. 57036716.0.0000.0003, approved on 22/08/2016) and by the Evandro Chagas Institute Research Ethics Committee (CAAE No. 57036716.0.3001.0019, approved on 03/10/2016).

Results

Thirty-eight individuals with CL were included. They were 28 males and 10 females aged between 13 and 81 with a mean age of 34.4 years (standard deviation: 14.2), resident in ten municipalities in the state of Amapá. The largest number of cases (n=13) came from the municipality of Mazagão. The most frequent etiological agent among the participants was Leishmania guyanensis (32/38). Clinical/demographic aspects and etiology of the CL cases are shown in Table 1.

Table 1
- Sociodemographic aspects and etiology of cutaneous leishmaniasis in males and females in a series of leishmaniasis cases in the state of Amapá, Brazil, 2018

With regard to the number of lesions, 20 individuals had multiple lesions while 18 had a single lesion. In 37/38 cases, lesions were of the ulcer type, mostly with a 5-6cm diameter, predominantly located on the upper limbs (n=9) and lower limbs (n=11). Predominant clinical manifestations were crust, secondary infection and satellite lesions. The characteristics of the lesions of the individuals with CL and respective etiological agents identified following sequencing and analysis of the Leishmania hsp70-234 gene region are described in Table 2.

Table 2
- Characteristics of lesions found in individuals with cutaneous leishmaniasis (n=38) and respective etiologic agents, identified following sequencing and analysis of Leishmania hsp70-234 gene region, Amapá, Brasil, 2018

This study contains the first report of L. infantum associated with CL in Amapá: a 35year-old woman infected in the municipality of Porto Grande, with multiple varied size verrucous cutaneous lesions on her body, including her thighs and legs, without visceral disease

Discussion

The most frequent etiological agent among the five species identified in individuals with CL in Amapá was L. guyanensis: 32 of the 38 cases analyzed. The hsp70-234 Leishmania marker discriminates six pathogens in the Amazon.12,14 The specificity of hsp70-234 has not been previously evaluated for L. lindenbergi, the seventh pathogen that causes human CL in the Amazon.8 Some species identified in this study are worthy of note: (i) Leishmania amazonensis, in addition to localized CL it can cause diffuse (anergic) leishmaniasis which is rarely curable; and (ii) Leishmania guyanensis and L. braziliensis, which induce cutaneous/mucous lesions that are hard to deal with,1,15 having a high degree of hypersensitivity and which may appear months or years after clinical cure of the primary lesion.1 Only long-term medical follow-up will ensure effective diagnosis and treatment of relapses.15 Even in the absence of mucous involvement, knowing what the pathogen is enables a prognosis to be defined.

Those who had CL caused by L. braziliensis and L. naiffi had a single ulcer. Leishmania braziliensis is widely distributed in Brazil.11. Ministério da Saúde (BR). Secretaria de Vigilância das Doenças Transmissíveis. Manual de vigilância da leishmaniose tegumentar [Internet]. Brasília: Ministério da Saúde; 2017 [citado 2018 nov 21]. 190 p. Disponível em: Disponível em: http://bvsms.saude.gov.br/bvs/publicacoes/manual_vigilancia_leishmaniose_tegumentar.pdf
http://bvsms.saude.gov.br/bvs/publicacoe...
L. naiffi infection in humans is limited to the states of Amazonas, Acre, Pará, Rondônia and Mato Grosso,1616. Figueira LP, Soares FV, Naiff Junior RD, Vinhote Silva AC, Silva SS, Epir TT, et al. New human case reports of cutaneous leishmaniasis by Leishmania (Viannia) naiffi in the Amazon region, Brazil. Acta Amaz [Internet]. 2017 Jan-Mar [cited 2019 Jul 16];47(1):47-52. Available from: Available from: http://www.scielo.br/pdf/aa/v47n1/1809-4392-aa-47-01-00047.pdf . doi: 10.1590/1809-4392201601484
http://www.scielo.br/pdf/aa/v47n1/1809-4...
with no complications having been reported until our study revealed the first case of CL caused by L. naiffi in Amapá.

L. infantum is phylogenetically close to Leishmania donovani, the agent that causes VL and post-kala-azar dermal leishmaniasis in the Old World (Europe);1717. Zijlstra EE, Alves F, Rijal S, Arana B, Alvar J. Post-kala-azar dermal leishmaniasis in the Indian subcontinent: a threat to the South-East Asia Region Kala-azar Elimination Programme. PLoS Negl Trop Dis [Internet]. 2017 Nov [cited 2019 Jul 16];11(11):e0005877. Available from: Available from: https://journals.plos.org/plosntds/article?id=10.1371/journal.pntd.0005877 . doi: 10.1371/journal.pntd.0005877
https://journals.plos.org/plosntds/artic...
occurrence of cutaneous lesions caused by L. infantum in immunocompetent individuals is uncommon in the Americas, especially in Brazil.1414. Garcez LM, Hueb M, Cordies N, Sanchez L, Nascimento L, Santos R, et al. utaneous Leishmaniasis in Brazil: undeniable diversity of species is still poorly known [Internet]. In: Abstracts book: proceedings of the 6th World Congress on Leishmaniasis; 2017 16-20 may; Toledo, Spain; 2017 [citado 2018 nov 24]. p. 175. Available from: Available from: http://worldleish2017.org/documentos/Abstracts_Book_WL6_2017.pdf
http://worldleish2017.org/documentos/Abs...
,1818. Lyra MR, Pimentel MIF, Madeira MF, Antonio LF, Lyra JPM, Fagundes A, et al. First report of cutaneous leishmaniasis caused by Leishmania (Leishmania) infantum chagasi in an urban area of Rio de Janeiro, Brazil. Rev Inst Med Trop S. Paulo [Internet]. 2015 Sep-Oct [cited 2019 Jul 16];57(5):451-4. Available from: Available from: http://www.scielo.br/pdf/rimtsp/v57n5/0036-4665-rimtsp-57-05-00451.pdf . doi: 10.1590/S0036-46652015000500016
http://www.scielo.br/pdf/rimtsp/v57n5/00...

L. infantum has been isolated in the skin of Africans with CL, producing a lesion histologically distinct from that caused by L. major.1919. Boussoffara T, Boubaker MS, Ben Ahmed M, Mokni M, Guizani I, Salah AB, et al. Histological and immunological differences between zoonotic cutaneous leishmaniasis due to Leishmania major and sporadic cutaneous leishmaniasis due to Leishmania infantum. Parasite [Internet]. 2019 Feb [cited 2019 Jul 16];26:9. Available from: Available from: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6391896/ . doi: 10.1051/parasite/2019007
https://www.ncbi.nlm.nih.gov/pmc/article...
Since 2015, cases of CL caused by L. infantum have been recorded in different Brazilian states.1414. Garcez LM, Hueb M, Cordies N, Sanchez L, Nascimento L, Santos R, et al. utaneous Leishmaniasis in Brazil: undeniable diversity of species is still poorly known [Internet]. In: Abstracts book: proceedings of the 6th World Congress on Leishmaniasis; 2017 16-20 may; Toledo, Spain; 2017 [citado 2018 nov 24]. p. 175. Available from: Available from: http://worldleish2017.org/documentos/Abstracts_Book_WL6_2017.pdf
http://worldleish2017.org/documentos/Abs...
,1818. Lyra MR, Pimentel MIF, Madeira MF, Antonio LF, Lyra JPM, Fagundes A, et al. First report of cutaneous leishmaniasis caused by Leishmania (Leishmania) infantum chagasi in an urban area of Rio de Janeiro, Brazil. Rev Inst Med Trop S. Paulo [Internet]. 2015 Sep-Oct [cited 2019 Jul 16];57(5):451-4. Available from: Available from: http://www.scielo.br/pdf/rimtsp/v57n5/0036-4665-rimtsp-57-05-00451.pdf . doi: 10.1590/S0036-46652015000500016
http://www.scielo.br/pdf/rimtsp/v57n5/00...
In Africa and the Americas, the clinical characteristics of CL vary according to the etiological agent.11. Ministério da Saúde (BR). Secretaria de Vigilância das Doenças Transmissíveis. Manual de vigilância da leishmaniose tegumentar [Internet]. Brasília: Ministério da Saúde; 2017 [citado 2018 nov 21]. 190 p. Disponível em: Disponível em: http://bvsms.saude.gov.br/bvs/publicacoes/manual_vigilancia_leishmaniose_tegumentar.pdf
http://bvsms.saude.gov.br/bvs/publicacoe...
,1919. Boussoffara T, Boubaker MS, Ben Ahmed M, Mokni M, Guizani I, Salah AB, et al. Histological and immunological differences between zoonotic cutaneous leishmaniasis due to Leishmania major and sporadic cutaneous leishmaniasis due to Leishmania infantum. Parasite [Internet]. 2019 Feb [cited 2019 Jul 16];26:9. Available from: Available from: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6391896/ . doi: 10.1051/parasite/2019007
https://www.ncbi.nlm.nih.gov/pmc/article...
Our study highlights the pleomorphism of CL caused by L. infantum, alerting as to the need for early diagnosis2020. Meireles CB, Maia LC, Soares GC, Teodoro IPP, Gadelha MDSV, Silva CGL, et al. Atypical presentations of cutaneous leishmaniasis: a systematic review. Acta Trop [Internet]. 2017 Aug [cited 2019 Jul 16];172:240-54. Available from: Available from: https://www.sciencedirect.com/science/article/abs/pii/S0001706X17303406?via%3Dihub . doi: 10.1016/j.actatropica.2017.05.022
https://www.sciencedirect.com/science/ar...
and, in this sense, the importance of surveillance for preventing also human VL in Amapá.

CL hotspots have been identified in Amapá, principally in areas of agricultural, plant extractivism and prospecting/mining settlements,22. Almeida ANF, Garcez LM, Araújo OCL. Leishmaniose tegumentar americana no estado do Amapá, Brasil: 2007 a 2015 [Internet]. In: Anais do 53º Congresso da Sociedade Brasileira de Medicina Tropical. 2017 ago 27-30; Campinas: Galoá; 2018 [citado 2018 nov 21]. Disponível em: Disponível em: https://proceedings.science/medtrop/papers/leishmaniose-tegumentar-americana-no-estado-do-amapa%2C-brasil%3A-2007-a-2015
https://proceedings.science/medtrop/pape...
where males of productive age are most affected. The majority of the individuals we analyzed had multiple primary lesions, suggesting diverse vector bites. This is a recognized behavior of Nyssomyia umbratilis, the agent that transmits L. guyanensis.11. Ministério da Saúde (BR). Secretaria de Vigilância das Doenças Transmissíveis. Manual de vigilância da leishmaniose tegumentar [Internet]. Brasília: Ministério da Saúde; 2017 [citado 2018 nov 21]. 190 p. Disponível em: Disponível em: http://bvsms.saude.gov.br/bvs/publicacoes/manual_vigilancia_leishmaniose_tegumentar.pdf
http://bvsms.saude.gov.br/bvs/publicacoe...
,88. Lainson R. Espécies neotropicais de Leishmania: uma breve revisão histórica sobre sua descoberta, ecologia e taxonomia. Rev Pan-Amaz Saúde [Internet]. 2010 jun [citado 2019 jul 16];1(2):13-32. Disponível em: Disponível em: http://scielo.iec.gov.br/pdf/rpas/v1n2/pt_v1n2a02.pdf . doi: 10.5123/S2176-62232010000200002
http://scielo.iec.gov.br/pdf/rpas/v1n2/p...
In these cases, prolonged treatment with Glucantime® would be necessary; however, there is considerable morbidity and even deaths associated with the toxicity of this chemotherapy.33. Superintendência de Vigilância em Saúde (AP). Situação epidemiológica da leishmaniose tegumentar no estado do Amapá: período de 2017 a novembro de 2018. Bol Epidemiol [Internet]. 2018 [citado 2019 mar 12];2:1-5. Disponível em: Disponível em: https://editor.amapa.gov.br/arquivos_portais/publicacoes/SVS_4d77b443923909a984f01e74bf38240f.pdf
https://editor.amapa.gov.br/arquivos_por...
,55. Sundar S, Chakravarty J. Leishmaniasis: an update of current pharmacotherapy. Expert Opin Pharmacother [Internet]. 2013 Jan [cited 2019 Jul 16];14(1):53-63. Available from: Available from: https://www.tandfonline.com/doi/full/10.1517/14656566.2013.755515 . doi: 10.1517/14656566.2013.755515
https://www.tandfonline.com/doi/full/10....
,2121. Oliveira MC, Amorim RFB, Freitas RA, Costa ALL. A fatal case of mucocutaneous leishmaniasis after pentavalent antimonial use. Rev Soc Bras Med Trop [Internet]. 2005 May-Jun [cited 2019 Jul 16];38(3):258-60. Available from: Available from: http://www.scielo.br/pdf/rsbmt/v38n3/24006.pdf . doi: 10.1590/S0037-86822005000300011
http://www.scielo.br/pdf/rsbmt/v38n3/240...
,2222. Lima MVN, Oliveira RZ, Lima AP, Cerino DA, Silveira TGV. Leishmaniose cutânea com desfecho fatal durante tratamento com antimonial pentavalente. An Bras Dermatol [Internet]. 2007 May-Jun [cited 2019 Jul 16];82(3):269-71. Available from: Available from: http://www.scielo.br/pdf/abd/v82n3/v82n03a10.pdf . doi: 10.1590/S0365-05962007000300010
http://www.scielo.br/pdf/abd/v82n3/v82n0...

Although Glucantime® is the first drug option for treating CL in Brazil, Pentamidine Isethionate® is recommended where L. guyanensis is predominant.11. Ministério da Saúde (BR). Secretaria de Vigilância das Doenças Transmissíveis. Manual de vigilância da leishmaniose tegumentar [Internet]. Brasília: Ministério da Saúde; 2017 [citado 2018 nov 21]. 190 p. Disponível em: Disponível em: http://bvsms.saude.gov.br/bvs/publicacoes/manual_vigilancia_leishmaniose_tegumentar.pdf
http://bvsms.saude.gov.br/bvs/publicacoe...
Study participants infected with L. guyanensis came from nine of the ten municipalities considered in the sample, suggesting widespread distribution of this species in Amapá, where vector fauna are abundant.88. Lainson R. Espécies neotropicais de Leishmania: uma breve revisão histórica sobre sua descoberta, ecologia e taxonomia. Rev Pan-Amaz Saúde [Internet]. 2010 jun [citado 2019 jul 16];1(2):13-32. Disponível em: Disponível em: http://scielo.iec.gov.br/pdf/rpas/v1n2/pt_v1n2a02.pdf . doi: 10.5123/S2176-62232010000200002
http://scielo.iec.gov.br/pdf/rpas/v1n2/p...
,1010. Saraiva JF, Souto RNP, Ferreira RMA. Flebotomineos (diptera: psychodidae) coletados em um assentamento rural no estado do Amapá, Brasil. Biota Amazônia [Internet]. 2011 [citado 2019 jul 16];1(1):58-62. Disponível em: Disponível em: https://periodicos.unifap.br/index.php/biota/article/view/145/v1n1p58-62.pdf . doi: 10.18561/2179-5746/biotaamazonia.v1n1p58-62
https://periodicos.unifap.br/index.php/b...
,1111. Galardo AKR, Galardo CD, Santana AA, Mendes JCC, Souza FRA, Duarte JP, et al. Primeira ocorrência de Lutzomyia (Lutzomyia) longipalpis LUTZ & NEIVA, 1912 (Diptera: Psychodidae: Phlebotominae) no estado do Amapá, Brasil. Biota Amazônia [Internet]. 2013 [citado 2019 jul 16];3(2):179-83. Disponível em: Disponível em: https://periodicos.unifap.br/index.php/biota/article/view/688/v3n2p179-183.pdf . doi: 10.18561/2179-5746/biotaamazonia.v3n2p179-183
https://periodicos.unifap.br/index.php/b...

L. braziliensis and L. guyanensis infection caused by Leishmania RNA virus subtype 1, found in Peru, Bolivia, French Guiana, Suriname and Brazil, would explain the high virulence and resistance of these species to the drug.2323. Cantanhêde LM, Fernandes FG, Ferreira GEM, Porrozzi R, Ferreira RGM, Cupolillo E. New insights into the genetic diversity of Leishmania RNA Virus 1 and its species-specific relationship with Leishmania parasites. PLoS One [Internet]. 2018 Jun [cited 2019 Jul 16];13(6):e0198727. Available from: Available from: https://journals.plos.org/plosone/article?id=10.1371/journal.pone.0198727 . doi: 10.1371/journal.pone.0198727
https://journals.plos.org/plosone/articl...
Biological factors of CL need to be monitored, especially in frontier regions.

With regard to treatment, with effect from 2017 Pentamidine Isethionate® has been considered preferential to Glucantime® for systemic treatment of CL in areas where L. guyanensis is predominant in Brazil. In cases of (i) kidney, heart and liver failure, (ii) people who have had kidney, heart and liver transplants, (iii) pregnant women and people more than 50 years old, the use of liposomal Amphotericin B® is recommended. Glucantime® would be the first option only for intralesional use in these areas.11. Ministério da Saúde (BR). Secretaria de Vigilância das Doenças Transmissíveis. Manual de vigilância da leishmaniose tegumentar [Internet]. Brasília: Ministério da Saúde; 2017 [citado 2018 nov 21]. 190 p. Disponível em: Disponível em: http://bvsms.saude.gov.br/bvs/publicacoes/manual_vigilancia_leishmaniose_tegumentar.pdf
http://bvsms.saude.gov.br/bvs/publicacoe...
The results found by our study support the new guidelines for treatment of CL in Amapá adopted with effect from 2018.2424. Superintendência de Vigilância em Saúde (AP). Nota informativa nº 02/2018: atualização do protocolo terapêutico de pacientes de leishmaniose tegumentar (LT) no estado do Amapá, com a substituição da droga de primeira escolha para o Isotionato de Pentamidina [Internet]. Amapá: Superintendência de Vigilância em Saúde ; 2018 [citado 2019 mar 12]. 6 p. Disponível em: Disponível em: https://editor.amapa.gov.br/arquivos_portais/publicacoes/SVS_4cb5d682eda3b9611eef9ebccdf63e4a.pdf
https://editor.amapa.gov.br/arquivos_por...

Our study confirmed the predominance of L. guyanensis infection in Amapá, frequently associated with multiple lesions, as was as providing the first reports of the presence of Leishmania naiffi and L. infantum in clinical samples of CL in the state of Amapá.

Acknowledgements

Our thanks go to Dr. Inês Celeste Ribeiro Martins, Director of the Amapá Tropical Diseases Reference Center (CRDT-AP), and to Dr. Aldo Aparecido Proietti Júnior, Coordinator of the Federal University of Amapá Special Applied Microbiology Laboratory (UNIFAP), for their support for our study, making available the necessary space and conditions for our work at CRDT/AP and for proper storage of samples at UNIFAP.

References

  • *
    This research received financial support from the Amapá State Research Support Foundation (FAPEAP) - Process No. 250.203.039/2016 - and a doctoral Research Grant from the Pará State Research Support Foundation (FAPESPA) - Process No. 003/2014.

Publication Dates

  • Publication in this collection
    17 Feb 2020
  • Date of issue
    2020

History

  • Received
    09 Feb 2019
  • Accepted
    29 June 2019
Secretaria de Vigilância em Saúde - Ministério da Saúde do Brasil Brasília - Distrito Federal - Brazil
E-mail: leilapgarcia@gmail.com