Advertisement

Applied Microbiology and Biotechnology

, Volume 103, Issue 19, pp 8179–8190 | Cite as

Multiplex flow cytometry serology to diagnosis of canine visceral leishmaniasis

  • Henrique Gama Ker
  • Wendel Coura-Vital
  • Diogo Garcia Valadares
  • Rodrigo Dian Oliveira Aguiar-Soares
  • Rory Cristiane Fortes de Brito
  • Patrícia Sampaio Tavares Veras
  • Deborah Bittencourt Mothé Fraga
  • Olindo Assis Martins-Filho
  • Andréa Teixeira-Carvalho
  • Alexandre Barbosa ReisEmail author
Methods and protocols

Abstract

An accurate diagnosis of visceral leishmaniasis is an essential tool for control of the disease. While serologic methods are very useful, these conventional methodologies still present limitations in terms of sensitivity and specificity. The use of flow cytometry is a worldwide trend in the development of high-performance diagnostic methods. Herein, we describe a new flow cytometry serology test, characterized by the employment of the Cytometric Bead Array microspheres A4 and E4 coated with the recombinant antigens rLci1A and rLci2B respectively, to improve the serodiagnosis of canine visceral leishmaniasis. The tests were conducted in a wide variety of sera groups (n = 140), where the diagnostics development would be optimized accounting not just the ability to identify infected dogs with different clinical status, but also to exclude cross-reaction and differentiate vaccinated dogs from dogs infected. Serological testing of the antigenic system A4–rLci1A showed a sensitivity of 90.0% and specificity of 75%, while the E4–rLci2B testing demonstrated a sensitivity of 95.0% and specificity of 82.5%. The use of a multiplex assay of A4–rLci1A and E4–rLci2B, resulted in a diagnostic improvement, with a sensitivity of 95.0% and specificity of 91.2%. Our results show that this novel flow cytometry serology test is a viable tool for sensitive and specific serodiagnosis. Notably, the combination of distinct antigenic systems allows us to test for antibodies to multiple recombinant antigens from a single serum sample. This benefit emphasizes the importance of this methodology as an alternative in the serological diagnosis.

Key words

Leishmania infantum Canine visceral leishmaniasis Multiplexed diagnosis Serological methods Flow cytometry 

Notes

Acknowledgments

A.B.R., A.T.C., O.A.M.F., P.S.T.V, H.G.K., and R.C.F.B. are grateful to Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) and CAPES for PQ fellowships. We also thank Megan M. Keller of the Leishmania Research Laboratory, of Internal Medicine, University of Iowa for the critical reading and review of the English of our article.

Funding information

This work was supported by Fundação de Amparo à Pesquisa do Estado de Minas Gerais (FAPEMIG, grants CBB - INV-00037-14 and CDS - APQ-03505-131), the Programa de Pesquisa para o SUS (PPSUS grant APQ-03505-13), and the Instituto Nacional de Ciência e Tecnologia em Doenças Tropicais (INCT-DT).

Compliance with ethical standards

Conflict of Interest

The authors declare that they have no conflict of interest.

Ethical statement

This study was approved by the Ethics Research Committee of the Universidade Federal de Ouro Preto (protocol number 083/2007). All procedures involving animals were performed in compliance with Brazilian federal law for animal experimentation (Law 11794/2008).

References

  1. Alvar J, Velez ID, Bern C, Herrero M, Desjeux P, Cano J, Jannin J, den Boer M, Team WHOLC (2012) Leishmaniasis worldwide and global estimates of its incidence. PLoS One 7(5):e35671.  https://doi.org/10.1371/journal.pone.0035671 CrossRefGoogle Scholar
  2. Andrade RA, Silva Araujo MS, Reis AB, Gontijo CM, Vianna LR, Mayrink W, Martins-Filho OA (2009) Advances in flow cytometric serology for canine visceral leishmaniasis: diagnostic applications when distinct clinical forms, vaccination and other canine pathogens become a challenge. Vet Immunol Immunopathol 128(1-3):79–86.  https://doi.org/10.1016/j.vetimm.2008.10.308 CrossRefGoogle Scholar
  3. Badaro R, Eulalio MC, Benson D, Freire M, Miranda JC, Pedral-Sampaio D, Burns JM, David JR Jr, Johnson WD, Reed SG (1993) Sensitivity and specificity of a recombinant Leishmania chagasi antigen in the serodiagnosis of visceral leishmaniasis. Arch Inst Pasteur Tunis 70(3-4):331–332Google Scholar
  4. Campos FMF, Repoles LC, de Araujo FF, Peruhype-Magalhaes V, Xavier MAP, Sabino EC, de Freitas Carneiro Proietti AB, Andrade MC, Teixeira-Carvalho A, Martins-Filho OA, Gontijo CMF (2018) Usefulness of FC-TRIPLEX Chagas/Leish IgG1 as confirmatory assay for non-negative results in blood bank screening of Chagas disease. J Immunol Methods 455:34–40.  https://doi.org/10.1016/j.jim.2018.01.006 CrossRefGoogle Scholar
  5. Carvalho Neta AV, Rocha RDR, Gontijo CMF, Reis AB, Martins-Filho OA (2006) Citometria de fluxo no diagnóstico da leishmaniose visceral canina. Arquivo Brasileiro de Medicina Veterinária e Zootecnia 58:480–488CrossRefGoogle Scholar
  6. Costa CH (2011) How effective is dog culling in controlling zoonotic visceral leishmaniasis? A critical evaluation of the science, politics and ethics behind this public health policy. Rev Soc Bras Med Trop 44(2):232–242CrossRefGoogle Scholar
  7. Coura-Vital W, Marques MJ, Veloso VM, Roatt BM, Aguiar-Soares RD, Reis LE, Braga SL, Morais MH, Reis AB, Carneiro M (2011) Prevalence and factors associated with Leishmania infantum infection of dogs from an urban area of Brazil as identified by molecular methods. PLoS Negl Trop Dis 5(8):e1291.  https://doi.org/10.1371/journal.pntd.0001291 CrossRefGoogle Scholar
  8. Courtenay O, Quinnell RJ, Garcez LM, Shaw JJ, Dye C (2002) Infectiousness in a cohort of brazilian dogs: why culling fails to control visceral leishmaniasis in areas of high transmission. J Infect Dis 186(9):1314–1320.  https://doi.org/10.1086/344312 CrossRefGoogle Scholar
  9. da Silva DA, Madeira Mde F, Abrantes TR, Filho CJ, Figueiredo FB (2013) Assessment of serological tests for the diagnosis of canine visceral leishmaniasis. Vet J 195(2):252–253.  https://doi.org/10.1016/j.tvjl.2012.06.010 CrossRefGoogle Scholar
  10. Dantas-Torres F, Solano-Gallego L, Baneth G, Ribeiro VM, de Paiva-Cavalcanti M, Otranto D (2012) Canine leishmaniosis in the Old and New Worlds: unveiled similarities and differences. Trends Parasitol 28(12):531–538.  https://doi.org/10.1016/j.pt.2012.08.007 CrossRefGoogle Scholar
  11. de Andrade HM, Reis AB, dos Santos SL, Volpini AC, Marques MJ, Romanha AJ (2006) Use of PCR-RFLP to identify Leishmania species in naturally-infected dogs. Vet Parasitol 140(3-4):231–238.  https://doi.org/10.1016/j.vetpar.2006.03.031 CrossRefGoogle Scholar
  12. de Andrade RA, Reis AB, Gontijo CM, Braga LB, Rocha RD, Araujo MS, Vianna LR, Martins-Filho OA (2007) Clinical value of anti-Leishmania (Leishmania) chagasi IgG titers detected by flow cytometry to distinguish infected from vaccinated dogs. Vet Immunol Immunopathol 116(1-2):85–97.  https://doi.org/10.1016/j.vetimm.2007.01.002 CrossRefGoogle Scholar
  13. de Carvalho FLN, Riboldi EO, Bello GL, Ramos RR, Barcellos RB, Gehlen M, Halon ML, Romao PRT, Dallegrave E, Rossetti MLR (2018) Canine visceral leishmaniasis diagnosis: a comparative performance of serological and molecular tests in symptomatic and asymptomatic dogs. Epidemiol Infect 146(5):571–576.  https://doi.org/10.1017/S0950268818000225 CrossRefGoogle Scholar
  14. de Oliveira IQ, Silva RA, Sucupira MV, da Silva ED, Reis AB, Grimaldi G Jr, Fraga DB, Veras PS (2015) Multi-antigen print immunoassay (MAPIA)-based evaluation of novel recombinant Leishmania infantum antigens for the serodiagnosis of canine visceral leishmaniasis. Parasit Vectors 8:45.  https://doi.org/10.1186/s13071-015-0651-6 CrossRefGoogle Scholar
  15. de Paiva-Cavalcanti M, de Morais RC, Pessoa ESR, Trajano-Silva LA, Goncalves-de-Albuquerque Sda C, Tavares Dde H, Brelaz-de-Castro MC, Silva Rde F, Pereira VR (2015) Leishmaniases diagnosis: an update on the use of immunological and molecular tools. Cell & bioscience 5:31.  https://doi.org/10.1186/s13578-015-0021-2 CrossRefGoogle Scholar
  16. Faria AR, de Castro VL, Coura-Vital W, Reis AB, Damasceno LM, Gazzinelli RT, Andrade HM (2015) Novel recombinant multiepitope proteins for the diagnosis of asymptomatic Leishmania infantum-infected dogs. PLoS Negl Trop Dis 9(1):e3429.  https://doi.org/10.1371/journal.pntd.0003429 CrossRefGoogle Scholar
  17. Ferreira C, de Lana M, Carneiro M, Reis AB, Paes DV, da Silva ES, Schallig H, Gontijo CM (2007) Comparison of serological assays for the diagnosis of canine visceral leishmaniasis in animals presenting different clinical manifestations. Vet Parasitol 146(3-4):235–241.  https://doi.org/10.1016/j.vetpar.2007.02.015 CrossRefGoogle Scholar
  18. Folgueira C, Carrion J, Moreno J, Saugar JM, Canavate C, Requena JM (2008) Effects of the disruption of the HSP70-II gene on the growth, morphology, and virulence of Leishmania infantum promastigotes. Int Microbiol 11(2):81–89Google Scholar
  19. Fraga DB, da Silva ED, Pacheco LV, Borja LS, de Oliveira IQ, Coura-Vital W, Monteiro GR, Oliveira GG, Jeronimo SM, Reis AB, Veras PS (2014) A multicentric evaluation of the recombinant Leishmania infantum antigen-based immunochromatographic assay for the serodiagnosis of canine visceral leishmaniasis. Parasit Vectors 7:136.  https://doi.org/10.1186/1756-3305-7-136 CrossRefGoogle Scholar
  20. Giunchetti RC, Mayrink W, Genaro O, Carneiro CM, Correa-Oliveira R, Martins-Filho OA, Marques MJ, Tafuri WL, Reis AB (2006) Relationship between canine visceral leishmaniosis and the Leishmania (Leishmania) chagasi burden in dermal inflammatory foci. J Comp Pathol 135(2-3):100–107.  https://doi.org/10.1016/j.jcpa.2006.06.005 CrossRefGoogle Scholar
  21. Giunchetti RC, Correa-Oliveira R, Martins-Filho OA, Teixeira-Carvalho A, Roatt BM, de Oliveira Aguiar-Soares RD, de Souza JV, das Dores Moreira N, Malaquias LC, Mota e Castro LL, de Lana M, Reis AB (2007) Immunogenicity of a killed Leishmania vaccine with saponin adjuvant in dogs. Vaccine 25(44):7674–7686.  https://doi.org/10.1016/j.vaccine.2007.08.009 CrossRefGoogle Scholar
  22. Grimaldi G Jr, Teva A, Ferreira AL, dos Santos CB, Pinto I, de-Azevedo CT, Falqueto A (2012) Evaluation of a novel chromatographic immunoassay based on Dual-Path Platform technology (DPP(R) CVL rapid test) for the serodiagnosis of canine visceral leishmaniasis. Trans R Soc Trop Med Hyg 106(1):54–59.  https://doi.org/10.1016/j.trstmh.2011.10.001 CrossRefGoogle Scholar
  23. Harhay MO, Olliaro PL, Costa DL, Costa CH (2011) Urban parasitology: visceral leishmaniasis in Brazil. Trends Parasitol 27(9):403–409.  https://doi.org/10.1016/j.pt.2011.04.001 CrossRefGoogle Scholar
  24. Ker HG, Aguiar-Soares RDO, Roatt BM, Moreira ND, Coura-Vital W, Carneiro CM, Teixeira-Carvalho A, Martins-Filho OA, Giunchetti RC, da Silveira-Lemos D, Reis AB (2013a) Effect of the preservative and temperature conditions on the stability of Leishmania infantum promastigotes antigens applied in a flow cytometry diagnostic method for canine visceral leishmaniasis. Diagn Microbiol Infect Dis 76(4):470–476.  https://doi.org/10.1016/j.diagmicrobio.2013.04.007 CrossRefGoogle Scholar
  25. Ker HG, Coura-Vital W, Aguiar-Soares RD, Roatt BM, das Dores Moreira N, Carneiro CM, Machado EM, Teixeira-Carvalho A, Martins-Filho OA, Giunchetti RC, Araujo MS, Coelho EA, da Silveira-Lemos D, Reis AB (2013b) Evaluation of a prototype flow cytometry test for serodiagnosis of canine visceral leishmaniasis. Clin Vaccine Immunol 20(12):1792–1798.  https://doi.org/10.1128/CVI.00575-13 CrossRefGoogle Scholar
  26. Laurenti MD, de Santana Leandro MV Jr, Tomokane TY, De Lucca HR, Aschar M, Souza CS, Silva RM, Marcondes M, da Matta VL (2014) Comparative evaluation of the DPP((R)) CVL rapid test for canine serodiagnosis in area of visceral leishmaniasis. Vet Parasitol 205(3-4):444–450.  https://doi.org/10.1016/j.vetpar.2014.09.002 CrossRefGoogle Scholar
  27. Lira RA, Cavalcanti MP, Nakazawa M, Ferreira AG, Silva ED, Abath FG, Alves LC, Souza WV, Gomes YM (2006) Canine visceral leishmaniosis: a comparative analysis of the EIE-leishmaniose-visceral-canina-Bio-Manguinhos and the IFI-leishmaniose-visceral-canina-Bio-Manguinhos kits. Vet Parasitol 137(1-2):11–16.  https://doi.org/10.1016/j.vetpar.2005.12.020 CrossRefGoogle Scholar
  28. Mancianti F, Gramiccia M, Gradoni L, Pieri S (1988) Studies on canine leishmaniasis control. 1. Evolution of infection of different clinical forms of canine leishmaniasis following antimonial treatment. Trans R Soc Trop Med Hyg 82(4):566–567.  https://doi.org/10.1016/0035-9203(88)90510-x CrossRefGoogle Scholar
  29. Marcondes M, Biondo AW, Gomes AA, Silva AR, Vieira RF, Camacho AA, Quinn J, Chandrashekar R (2011) Validation of a Leishmania infantum ELISA rapid test for serological diagnosis of Leishmania chagasi in dogs. Vet Parasitol 175(1-2):15–19.  https://doi.org/10.1016/j.vetpar.2010.09.036 CrossRefGoogle Scholar
  30. Marcondes M, de Lima VM, de Araujo MF, Hiramoto RM, Tolezano JE, Vieira RF, Biondo AW (2013) Longitudinal analysis of serological tests officially adopted by the Brazilian Ministry of Health for the diagnosis of canine visceral leishmaniasis in dogs vaccinated with Leishmune(R). Vet Parasitol 197(3-4):649–652.  https://doi.org/10.1016/j.vetpar.2013.07.013 CrossRefGoogle Scholar
  31. Menezes-Souza D, Mendes TA, Gomes Mde S, Bartholomeu DC, Fujiwara RT (2015) Improving serodiagnosis of human and canine leishmaniasis with recombinant Leishmania braziliensis cathepsin l-like protein and a synthetic peptide containing its linear B-cell epitope. PLoS Negl Trop Dis 9(1):e3426.  https://doi.org/10.1371/journal.pntd.0003426 CrossRefGoogle Scholar
  32. Mettler M, Grimm F, Capelli G, Camp H, Deplazes P (2005) Evaluation of enzyme-linked immunosorbent assays, an immunofluorescent-antibody test, and two rapid tests (immunochromatographic-dipstick and gel tests) for serological diagnosis of symptomatic and asymptomatic Leishmania infections in dogs. J Clin Microbiol 43(11):5515–5519.  https://doi.org/10.1128/JCM.43.11.5515-5519.2005 CrossRefGoogle Scholar
  33. Ministério da Saúde (2006) Manual de Vigilância e Controle da Leishmaniose Visceral, 1st edn. Secretaria de Vigilância em Saúde, Brasília Available: http://bvsms.saude.gov.br/bvs/publicacoes/manual_vigilancia_controle_leishmaniose_visceral.pdf.120 Google Scholar
  34. Ministério da Saúde (2011) Esclarecimento sobre substituição do protocolo diagnóstico da leihsmaniose visceral canina; Nota técnica conjunta nu 01/2011 - CGDT-CGLAB/DEVIT/SVS/MSGoogle Scholar
  35. Mohebali M, Javadian E, Yaghoobi-Ershadi MR, Akhavan AA, Hajjaran H, Abaei MR (2004) Characterization of Leishmania infection in rodents from endemic areas of the Islamic Republic of Iran. East Mediterr Health J 10(4-5):591–599Google Scholar
  36. Molina R, Amela C, Nieto J, San-Andres M, Gonzalez F, Castillo JA, Lucientes J, Alvar J (1994) Infectivity of dogs naturally infected with Leishmania infantum to colonized Phlebotomus perniciosus. Trans R Soc Trop Med Hyg 88(4):491–493.  https://doi.org/10.1016/0035-9203(94)90446-4 CrossRefGoogle Scholar
  37. Oliveira GG, Magalhaes FB, Teixeira MC, Pereira AM, Pinheiro CG, Santos LR, Nascimento MB, Bedor CN, Albuquerque AL, dos-Santos WL, Gomes YM, Moreira ED Jr, Brito ME, Pontes de Carvalho LC, de Melo Neto OP (2011) Characterization of novel Leishmania infantum recombinant proteins encoded by genes from five families with distinct capacities for serodiagnosis of canine and human visceral leishmaniasis. Am J Trop Med Hyg 85(6):1025–1034.  https://doi.org/10.4269/ajtmh.2011.11-0102 CrossRefGoogle Scholar
  38. Otranto D, Dantas-Torres F (2013) The prevention of canine leishmaniasis and its impact on public health. Trends Parasitol 29(7):339–345.  https://doi.org/10.1016/j.pt.2013.05.003 CrossRefGoogle Scholar
  39. PAHO (2017) Informe Epidemiológico das Américas. Leishmanioses. In: 5 (ed) Informe Leishmanioses. Organização de saúde Pan Americana, p 8Google Scholar
  40. Porrozzi R, Santos da Costa MV, Teva A, Falqueto A, Ferreira AL, dos Santos CD, Fernandes AP, Gazzinelli RT, Campos-Neto A, Grimaldi G Jr (2007) Comparative evaluation of enzyme-linked immunosorbent assays based on crude and recombinant leishmanial antigens for serodiagnosis of symptomatic and asymptomatic Leishmania infantum visceral infections in dogs. Clin Vaccine Immunol 14(5):544–548.  https://doi.org/10.1128/CVI.00420-06 CrossRefGoogle Scholar
  41. Quijada L, Requena JM, Soto M, Gomez LC, Guzman F, Patarroyo ME, Alonso C (1996) Mapping of the linear antigenic determinants of the Leishmania infantum hsp70 recognized by leishmaniasis sera. Immunol Lett 52(2-3):73–79CrossRefGoogle Scholar
  42. Quinnell RJ, Courtenay O (2009) Transmission, reservoir hosts and control of zoonotic visceral leishmaniasis. Parasitology 136(14):1915–1934.  https://doi.org/10.1017/S0031182009991156 CrossRefGoogle Scholar
  43. Quinnell RJ, Carson C, Reithinger R, Garcez LM, Courtenay O (2013) Evaluation of rK39 rapid diagnostic tests for canine visceral leishmaniasis: longitudinal study and meta-analysis. PLoS Negl Trop Dis 7(1):e1992.  https://doi.org/10.1371/journal.pntd.0001992 CrossRefGoogle Scholar
  44. Reis AB, Teixeira-Carvalho A, Giunchetti RC, Guerra LL, Carvalho MG, Mayrink W, Genaro O, Correa-Oliveira R, Martins-Filho OA (2006) Phenotypic features of circulating leucocytes as immunological markers for clinical status and bone marrow parasite density in dogs naturally infected by Leishmania chagasi. Clin Exp Immunol 146(2):303–311.  https://doi.org/10.1111/j.1365-2249.2006.03206.x CrossRefGoogle Scholar
  45. Riboldi E, Carvalho F, Romao PRT, Barcellos RB, Bello GL, Ramos RR, de Oliveira RT, Junior JPA, Rossetti ML, Dallegrave E (2018) Molecular method confirms Canine Leishmania infection detected by serological methods in non-endemic area of Brazil. Korean J Parasitol 56(1):11–19.  https://doi.org/10.3347/kjp.2018.56.1.11 CrossRefGoogle Scholar
  46. Roatt BM, Aguiar-Soares RD, Vitoriano-Souza J, Coura-Vital W, Braga SL, Correa-Oliveira R, Martins-Filho OA, Teixeira-Carvalho A, de Lana M, Figueiredo Gontijo N, Marques MJ, Giunchetti RC, Reis AB (2012) Performance of LBSap vaccine after intradermal challenge with L. infantum and saliva of Lu. longipalpis: immunogenicity and parasitological evaluation. PloS one 7(11):e49780.  https://doi.org/10.1371/journal.pone.0049780 CrossRefGoogle Scholar
  47. Romero GA, Boelaert M (2010) Control of visceral leishmaniasis in latin america-a systematic review. PLoS Negl Trop Dis 4(1):e584.  https://doi.org/10.1371/journal.pntd.0000584 CrossRefGoogle Scholar
  48. Silva DA, Madeira MF, Teixeira AC, de Souza CM, Figueiredo FB (2011) Laboratory tests performed on Leishmania seroreactive dogs euthanized by the leishmaniasis control program. Vet Parasitol 179(1-3):257–261.  https://doi.org/10.1016/j.vetpar.2011.01.048 CrossRefGoogle Scholar
  49. Silvestre R, Santarem N, Cunha J, Cardoso L, Nieto J, Carrillo E, Moreno J, Cordeiro-da-Silva A (2008) Serological evaluation of experimentally infected dogs by LicTXNPx-ELISA and amastigote-flow cytometry. Vet Parasitol 158(1-2):23–30.  https://doi.org/10.1016/j.vetpar.2008.09.001 CrossRefGoogle Scholar
  50. Solano-Gallego L, Koutinas A, Miro G, Cardoso L, Pennisi MG, Ferrer L, Bourdeau P, Oliva G, Baneth G (2009) Directions for the diagnosis, clinical staging, treatment and prevention of canine leishmaniosis. Vet Parasitol 165(1-2):1–18.  https://doi.org/10.1016/j.vetpar.2009.05.022 CrossRefGoogle Scholar
  51. Sousa S, Cardoso L, Reed SG, Reis AB, Martins-Filho OA, Silvestre R, Cordeiro da Silva A (2013) Development of a fluorescent based immunosensor for the serodiagnosis of canine leishmaniasis combining immunomagnetic separation and flow cytometry. PLoS Negl Trop Dis 7(8):e2371.  https://doi.org/10.1371/journal.pntd.0002371 CrossRefGoogle Scholar
  52. Souza CM, Silva ED, Ano Bom AP, Bastos RC, Nascimento HJ, da Silva Junior JG (2012) Evaluation of an ELISA for canine leishmaniasis immunodiagnostic using recombinant proteins. Parasite Immunol 34(1):1–7.  https://doi.org/10.1111/j.1365-3024.2011.01334.x CrossRefGoogle Scholar
  53. Teixeira-Carvalho A, Campos FM, Geiger SM, Rocha RD, de Araujo FF, Vitelli-Avelar DM, Andrade MC, Araujo MS, Lemos EM, de Freitas Carneiro Proietti AB, Sabino EC, Caldas RG, Freitas CR, Campi-Azevedo AC, Eloi-Santos SM, Martins-Filho OA (2015) FC-TRIPLEX Chagas/Leish IgG1: a multiplexed flow cytometry method for differential serological diagnosis of chagas disease and leishmaniasis. PLoS One 10(4):e0122938.  https://doi.org/10.1371/journal.pone.0122938 CrossRefGoogle Scholar
  54. WHO (2017) Integrating neglected tropical diseases into global health and development: fourth WHO report on neglected tropical diseasesGoogle Scholar
  55. Yamasaki M, Inokuma H, Sugimoto C, Shaw SE, Aktas M, Yabsley MJ, Yamato O, Maede Y (2007) Comparison and phylogenetic analysis of the heat shock protein 70 gene of Babesia parasites from dogs. Vet Parasitol 145(3-4):217–227.  https://doi.org/10.1016/j.vetpar.2007.01.003 CrossRefGoogle Scholar
  56. Zanette MF, Lima VM, Laurenti MD, Rossi CN, Vides JP, Vieira RF, Biondo AW, Marcondes M (2014) Serological cross-reactivity of Trypanosoma cruzi, Ehrlichia canis, Toxoplasma gondii, Neospora caninum and Babesia canis to Leishmania infantum chagasi tests in dogs. Rev Soc Bras Med Trop 47(1):105–107.  https://doi.org/10.1590/0037-8682-1723-2013 CrossRefGoogle Scholar

Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2019

Authors and Affiliations

  • Henrique Gama Ker
    • 1
  • Wendel Coura-Vital
    • 1
    • 2
  • Diogo Garcia Valadares
    • 1
  • Rodrigo Dian Oliveira Aguiar-Soares
    • 1
  • Rory Cristiane Fortes de Brito
    • 1
  • Patrícia Sampaio Tavares Veras
    • 3
    • 4
  • Deborah Bittencourt Mothé Fraga
    • 3
    • 4
  • Olindo Assis Martins-Filho
    • 5
  • Andréa Teixeira-Carvalho
    • 5
  • Alexandre Barbosa Reis
    • 1
    • 4
    Email author
  1. 1.Laboratório de Imunopatologia, Núcleo de Pesquisas em Ciências Biológicas, Instituto de Ciências Exatas e BiológicasUniversidade Federal de Ouro Preto (NUPEB/UFOP)Ouro PretoBrazil
  2. 2.Pós-Graduação em Ciências Farmacêuticas, Departamento de Análises Clínicas, Escola de FarmáciaUniversidade Federal de Ouro Preto (NUPEB/UFOP)Ouro PretoBrazil
  3. 3.Laboratório Laboratório de Patologia e Biointervenção, Centro de Pesquisas Gonçalo MonizFundação Oswaldo Cruz (FIOCRUZ)SalvadorBrazil
  4. 4.Instituto de Ciência e Tecnologia de Doenças Tropicais (INCT-DT)SalvadorBrazil
  5. 5.Grupo Integrado de Pesquisas em Biomarcadores, Instituto René RachouFundação Oswaldo Cruz (FIOCRUZ)Belo HorizonteBrazil

Personalised recommendations