Leishmaniasis is widely regarded as a vaccine-preventable disease, but the costs required to reach pivotal Phase 3 studies and uncertainty about which candidate vaccines should be progressed into human studies significantly limits progress in vaccine development for this neglected tropical disease. Controlled human infection models (CHIMs) provide a pathway for accelerating vaccine development and to more fully understand disease pathogenesis and correlates of protection. Here, we describe the isolation, characterization and GMP manufacture of a new clinical strain of Leishmania major. Two fresh strains of L. major from Israel were initially compared by genome sequencing, in vivo infectivity and drug sensitivity in mice, and development and transmission competence in sand flies, allowing one to be selected for GMP production. This study addresses a major roadblock in the development of vaccines for leishmaniasis, providing a key resource for CHIM studies of sand fly transmitted cutaneous leishmaniasis.

Department of Microbiology and Immunology McGill University Montreal Quebec Canada

Department of Parasitology Faculty of Science Charles University Viničná 7 Prague Czech Republic

The Center for Geographic Medicine and Tropical Diseases Chaim Sheba Medical Center Sackler School of Medicine Tel Aviv University Tel Aviv Israel

The Hebrew University Hadassah Medical School Jerusalem Israel

Vibalogics GmbH Cuxhaven Germany

York Biomedical Research Institute Hull York Medical School University of York York UK

See more in PubMed

Alvar J, et al. Leishmaniasis worldwide and global estimates of its incidence. PLoS ONE. 2012;7:e35671. doi: 10.1371/journal.pone.0035671. PubMed DOI PMC

Hotez PJ. The rise of leishmaniasis in the twenty-first century. Trans. R. Soc. Trop. Med. Hyg. 2018;112:421–422. doi: 10.1093/trstmh/try075. PubMed DOI

Dostalova A, Volf P. Leishmania development in sand flies: parasite-vector interactions overview. Parasit. Vectors. 2012;5:276. doi: 10.1186/1756-3305-5-276. PubMed DOI PMC

Kamhawi S. Phlebotomine sand flies and Leishmania parasites: friends or foes? Trends Parasitol. 2006;22:439–445. doi: 10.1016/j.pt.2006.06.012. PubMed DOI

McGwire BS, Satoskar AR. Leishmaniasis: clinical syndromes and treatment. QJM. 2014;107:7–14. doi: 10.1093/qjmed/hct116. PubMed DOI PMC

Burza S, Croft SL, Boelaert M. Leishmaniasis. Lancet. 2018;392:951–970. doi: 10.1016/S0140-6736(18)31204-2. PubMed DOI

Mondal D, et al. Quantifying the infectiousness of post-kala-azar dermal leishmaniasis toward sand flies. Clin. Infect. Dis. 2019;69:251–258. doi: 10.1093/cid/ciy891. PubMed DOI PMC

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. 2017;11:e0005877. doi: 10.1371/journal.pntd.0005877. PubMed DOI PMC

Bailey F, et al. Cutaneous leishmaniasis and co-morbid major depressive disorder: a systematic review with burden estimates. PLoS Negl. Trop. Dis. 2019;13:e0007092. doi: 10.1371/journal.pntd.0007092. PubMed DOI PMC

Bennis I, De Brouwere V, Belrhiti Z, Sahibi H, Boelaert M. Psychosocial burden of localised cutaneous Leishmaniasis: a scoping review. BMC Public Health. 2018;18:358. doi: 10.1186/s12889-018-5260-9. PubMed DOI PMC

Pires M, Wright B, Kaye PM, da Conceicao V, Churchill RC. The impact of leishmaniasis on mental health and psychosocial well-being: a systematic review. PLoS ONE. 2019;14:e0223313. doi: 10.1371/journal.pone.0223313. PubMed DOI PMC

Alvar J, et al. Case study for a vaccine against leishmaniasis. Vaccine. 2013;31:B244–B249. doi: 10.1016/j.vaccine.2012.11.080. PubMed DOI

Bethony JM, et al. Vaccines to combat the neglected tropical diseases. Immunol. Rev. 2011;239:237–270. doi: 10.1111/j.1600-065X.2010.00976.x. PubMed DOI PMC

Gillespie PM, et al. Status of vaccine research and development of vaccines for leishmaniasis. Vaccine. 2016;34:2992–2995. doi: 10.1016/j.vaccine.2015.12.071. PubMed DOI

Khamesipour A. Therapeutic vaccines for leishmaniasis. Expert Opin. Biol. Ther. 2014;14:1641–1649. doi: 10.1517/14712598.2014.945415. PubMed DOI

Velez R, et al. Evaluation of canine leishmaniosis vaccine CaniLeish(R) under field conditions in native dog populations from an endemic Mediterranean area-A randomized controlled trial. Acta Trop. 2020;205:105387. doi: 10.1016/j.actatropica.2020.105387. PubMed DOI

Palatnik-de-Sousa CB, Nico D. The delay in the licensing of protozoal vaccines: a comparative history. Front Immunol. 2020;11:204. doi: 10.3389/fimmu.2020.00204. PubMed DOI PMC

Chapman, N. et al. Neglected Disease Research and Development: Uneven Progress. Policy Cures Research (2019). Available from: https://s3-ap-southeast-2.amazonaws.com/policy-cures-website-assets/app/uploads/2020/02/11150341/G-Finder2019.pdf.

Duthie, M. S. & Reed, S. G. Not all antigens are created equally: progress, challenges, and lessons associated with developing a vaccine for leishmaniasis. Clin. Vaccine Immunol. 24, 10.1128/CVI.00108-17 (2017). PubMed PMC

Aslan H, et al. A new model of progressive visceral leishmaniasis in hamsters by natural transmission via bites of vector sand flies. J. Infect. Dis. 2013;207:1328–1338. doi: 10.1093/infdis/jis932. PubMed DOI PMC

Duthie MS, et al. A defined subunit vaccine that protects against vector-borne visceral leishmaniasis. NPJ Vaccines. 2017;2:23. doi: 10.1038/s41541-017-0025-5. PubMed DOI PMC

Mo AX, Pesce J, Fenton Hall B. Visceral leishmaniasis control and elimination: is there a role for vaccines in achieving regional and global goals? Am. J. Tropical Med. Hyg. 2016;95:514–521. doi: 10.4269/ajtmh.16-0184. DOI

Bahrami F, Harandi AM, Rafati S. Biomarkers of cutaneous leishmaniasis. Front Cell Infect. Microbiol. 2018;8:222. doi: 10.3389/fcimb.2018.00222. PubMed DOI PMC

Singh OP, Hasker E, Sacks D, Boelaert M, Sundar S. Asymptomatic Leishmania infection: a new challenge for Leishmania control. Clin. Infect. Dis. 2014;58:1424–1429. doi: 10.1093/cid/ciu102. PubMed DOI PMC

Working Group on Research Priorities for Development of Leishmaniasis, V. et al. Vaccines for the leishmaniases: proposals for a research agenda. PLoS Negl. Trop. Dis. 2011;5:e943. doi: 10.1371/journal.pntd.0000943. PubMed DOI PMC

Darton TC, et al. Using a human challenge model of infection to measure vaccine efficacy: a randomised, controlled trial comparing the typhoid vaccines M01ZH09 with placebo and Ty21a. PLoS Negl. Trop. Dis. 2016;10:e0004926. doi: 10.1371/journal.pntd.0004926. PubMed DOI PMC

Eyal, N., Lipsitch, M. & Smith, P. G. Human challenge studies to accelerate coronavirus vaccine licensure. J. Infect. Dis. 10.1093/infdis/jiaa152 (2020). PubMed PMC

Gibani MM, et al. Investigation of the role of typhoid toxin in acute typhoid fever in a human challenge model. Nat. Med. 2019;25:1082–1088. doi: 10.1038/s41591-019-0505-4. PubMed DOI PMC

Gibani MM, et al. The impact of vaccination and prior exposure on stool shedding of Salmonella typhi and Salmonella paratyphi in 6 controlled human infection studies. Clin. Infect. Dis. 2019;68:1265–1273. doi: 10.1093/cid/ciy670. PubMed DOI PMC

Meiring JE, Giubilini A, Savulescu J, Pitzer VE, Pollard AJ. Generating the evidence for typhoid vaccine introduction: considerations for global disease burden estimates and vaccine testing through human challenge. Clin. Infect. Dis. 2019;69:S402–S407. doi: 10.1093/cid/ciz630. PubMed DOI PMC

Osowicki J, et al. Controlled human infection for vaccination against Streptococcus pyogenes (CHIVAS): establishing a group A Streptococcus pharyngitis human infection study. Vaccine. 2019;37:3485–3494. doi: 10.1016/j.vaccine.2019.03.059. PubMed DOI

Roestenberg M, Mo A, Kremsner PG, Yazdanbakhsh M. Controlled human infections: a report from the controlled human infection models workshop, Leiden University Medical Centre 4–6 May 2016. Vaccine. 2017;35:7070–7076. doi: 10.1016/j.vaccine.2017.10.092. PubMed DOI

Roestenberg M, et al. The frontline of controlled human malaria infections: A report from the controlled human infection models Workshop in Leiden University Medical Centre 5 May 2016. Vaccine. 2017;35:7065–7069. doi: 10.1016/j.vaccine.2017.10.093. PubMed DOI

Mohebali M, Nadim A, Khamesipour A. An overview of leishmanization experience: a successful control measure and a tool to evaluate candidate vaccines. Acta Trop. 2019;200:105173. doi: 10.1016/j.actatropica.2019.105173. PubMed DOI

Khamesipour A, et al. Leishmanization: use of an old method for evaluation of candidate vaccines against leishmaniasis. Vaccine. 2005;23:3642–3648. doi: 10.1016/j.vaccine.2005.02.015. PubMed DOI

Osman M, et al. A third generation vaccine for human visceral leishmaniasis and post kala azar dermal leishmaniasis: First-in-human trial of ChAd63-KH. PLoS Negl. Trop. Dis. 2017;11:e0005527. doi: 10.1371/journal.pntd.0005527. PubMed DOI PMC

Banerjee A, et al. Live attenuated Leishmania donovani centrin gene-deleted parasites induce IL-23-dependent IL-17-protective immune response against visceral leishmaniasis in a murine model. J. Immunol. 2018;200:163–176. doi: 10.4049/jimmunol.1700674. PubMed DOI PMC

Bhattacharya P, et al. Live attenuated Leishmania donovani centrin knock out parasites generate non-inferior protective immune response in aged mice against visceral leishmaniasis. PLoS Negl. Trop. Dis. 2016;10:e0004963. doi: 10.1371/journal.pntd.0004963. PubMed DOI PMC

Ismail N, Kaul A, Bhattacharya P, Gannavaram S, Nakhasi HL. Immunization with live attenuated Leishmania donovani Centrin(−/−) parasites is efficacious in asymptomatic infection. Front Immunol. 2017;8:1788. doi: 10.3389/fimmu.2017.01788. PubMed DOI PMC

Zhang WW, et al. A second generation leishmanization vaccine with a markerless attenuated Leishmania major strain using CRISPR gene editing. Nat. Commun. 2020;11:3461. doi: 10.1038/s41467-020-17154-z. PubMed DOI PMC

Coler RN, et al. From mouse to man: safety, immunogenicity and efficacy of a candidate leishmaniasis vaccine LEISH-F3+GLA-SE. Clin. Transl. Immunol. 2015;4:e35. doi: 10.1038/cti.2015.6. PubMed DOI PMC

Duthie MS, et al. Heterologous immunization with defined RNA and subunit vaccines enhances T Cell responses that protect against Leishmania donovani. Front Immunol. 2018;9:2420. doi: 10.3389/fimmu.2018.02420. PubMed DOI PMC

Atayde VD, et al. Exosome secretion by the parasitic protozoan leishmania within the sand fly midgut. Cell Rep. 2015;13:957–967. doi: 10.1016/j.celrep.2015.09.058. PubMed DOI PMC

Dey R, et al. Gut microbes egested during bites of infected sand flies augment severity of leishmaniasis via inflammasome-derived IL-1beta. Cell Host Microbe. 2018;23:134–143 e136. doi: 10.1016/j.chom.2017.12.002. PubMed DOI PMC

Serafim TD, et al. Sequential blood meals promote Leishmania replication and reverse metacyclogenesis augmenting vector infectivity. Nat. Microbiol. 2018;3:548–555. doi: 10.1038/s41564-018-0125-7. PubMed DOI PMC

Peters NC, et al. Vector transmission of leishmania abrogates vaccine-induced protective immunity. PLoS Pathog. 2009;5:e1000484. doi: 10.1371/journal.ppat.1000484. PubMed DOI PMC

Bufman H, et al. A retrospective study on demographic and clinical characteristics of cutaneous leishmaniasis suspected cases in southern Israel, 2013–2016: comparison between confirmed and negative cases. J. Vector Borne Dis. 2019;56:159–165. doi: 10.4103/0972-9062.263723. PubMed DOI

Orshan L, et al. Distribution and dispersal of Phlebotomus papatasi (Diptera: Psychodidae) in a zoonotic cutaneous leishmaniasis focus, the Northern Negev, Israel. PLoS Negl. Trop. Dis. 2016;10:e0004819. doi: 10.1371/journal.pntd.0004819. PubMed DOI PMC

Nalcaci M, et al. Detection of Leishmania RNA virus 2 in Leishmania species from Turkey. Trans. R. Soc. Trop. Med. Hyg. 2019;113:410–417. doi: 10.1093/trstmh/trz023. PubMed DOI

Sacks DL, Hieny S, Sher A. Identification of cell surface carbohydrate and antigenic changes between noninfective and infective developmental stages of Leishmania major promastigotes. J. Immunol. 1985;135:564–569. PubMed

Gradoni, L., Lopez-Velz, R. & Mokni, M. Manual on Case Management and Surveillance of the Leishmaniases in the WHO European Region (Copenhagen, 2017).

Cihakova J, Volf P. Development of different Leishmania major strains in the vector sandflies Phlebotomus papatasi and P. duboscqi. Ann. Trop. Med. Parasitol. 1997;91:267–279. doi: 10.1080/00034983.1997.11813140. PubMed DOI

Academy of Medical Sciences. Controlled Human Infection Model Studies. Academy of Medical Sciences (2018). Available at: https://acmedsci.ac.uk/file-download/55062331.

Adler S, Zuckerman A. Observations on a strain of Leishmania tropica after prolonged cultivation; notes on infectivity and immunity. Ann. Trop. Med. Parasitol. 1948;42:178–183. doi: 10.1080/00034983.1948.11685360. PubMed DOI

Sadlova J, Svobodova M, Volf P. Leishmania major: effect of repeated passages through sandfly vectors or murine hosts. Ann. Trop. Med. Parasitol. 1999;93:599–611. doi: 10.1080/00034983.1999.11813463. PubMed DOI

Kaye P, Scott P. Leishmaniasis: complexity at the host-pathogen interface. Nat. Rev. Microbiol. 2011;9:604–615. doi: 10.1038/nrmicro2608. PubMed DOI

McCall LI, Zhang WW, Matlashewski G. Determinants for the development of visceral leishmaniasis disease. PLoS Pathog. 2013;9:e1003053. doi: 10.1371/journal.ppat.1003053. PubMed DOI PMC

Rossi M, Fasel N. The criminal association of Leishmania parasites and viruses. Curr. Opin. Microbiol. 2018;46:65–72. doi: 10.1016/j.mib.2018.07.005. PubMed DOI

Kleschenko, Y. et al. Molecular Characterization of Leishmania RNA virus 2 in Leishmania major from Uzbekistan. Genes10, 10.3390/genes10100830 (2019). PubMed PMC

Saberi R, Fakhar M, Mohebali M, Anvari D, Gholami S. Global status of synchronizing Leishmania RNA virus in Leishmania parasites: a systematic review with meta-analysis. Transbound. Emerg. Dis. 2019;66:2244–2251. doi: 10.1111/tbed.13316. PubMed DOI

Abtahi M, et al. Relationship of Leishmania RNA Virus (LRV) and treatment failure in clinical isolates of Leishmania major. BMC Res. Notes. 2020;13:126. doi: 10.1186/s13104-020-04973-y. PubMed DOI PMC

Caridha D, et al. Route map for the discovery and pre-clinical development of new drugs and treatments for cutaneous leishmaniasis. Int J. Parasitol. Drugs Drug Resist. 2019;11:106–117. doi: 10.1016/j.ijpddr.2019.06.003. PubMed DOI PMC

Sullivan, D. J. & Agre, P. Human Plasmodium vivax mosquito experimental transmission. J. Clin. Invest. 10.1172/JCI135794 (2020). PubMed PMC

Rogers ME, Ilg T, Nikolaev AV, Ferguson MA, Bates PA. Transmission of cutaneous leishmaniasis by sand flies is enhanced by regurgitation of fPPG. Nature. 2004;430:463–467. doi: 10.1038/nature02675. PubMed DOI PMC

Giraud E, Svobodova M, Muller I, Volf P, Rogers ME. Promastigote secretory gel from natural and unnatural sand fly vectors exacerbate Leishmania major and Leishmania tropica cutaneous leishmaniasis in mice. Parasitology. 2019;146:1796–1802. doi: 10.1017/S0031182019001069. PubMed DOI PMC

Oliveira F, et al. A sand fly salivary protein vaccine shows efficacy against vector-transmitted cutaneous leishmaniasis in nonhuman primates. Sci. Transl. Med. 2015;7:290ra290. PubMed

Doehl JSP, et al. Skin parasite landscape determines host infectiousness in visceral leishmaniasis. Nat. Commun. 2017;8:57. doi: 10.1038/s41467-017-00103-8. PubMed DOI PMC

Zijlstra EE, el-Hassan AM, Ismael A, Ghalib HW. Endemic kala-azar in eastern Sudan: a longitudinal study on the incidence of clinical and subclinical infection and post-kala-azar dermal leishmaniasis. Am. J. Trop. Med Hyg. 1994;51:826–836. doi: 10.4269/ajtmh.1994.51.826. PubMed DOI

Romano A, Doria NA, Mendez J, Sacks DL, Peters NC. Cutaneous infection with Leishmania major mediates heterologous protection against visceral infection with Leishmania infantum. J. Immunol. 2015;195:3816–3827. doi: 10.4049/jimmunol.1500752. PubMed DOI PMC

Schroeder J, Brown N, Kaye P, Aebischer T. Single dose novel Salmonella vaccine enhances resistance against visceralizing L. major and L. donovani infection in susceptible BALB/c mice. PLoS Negl. Trop. Dis. 2011;5:e1406. doi: 10.1371/journal.pntd.0001406. PubMed DOI PMC

Tonui WK, Titus RG. Cross-protection against Leishmania donovani but not L. braziliensis caused by vaccination with L. major soluble promastigote exogenous antigens in BALB/c mice. Am. J. Trop. Med Hyg. 2007;76:579–584. doi: 10.4269/ajtmh.2007.76.579. PubMed DOI

Lypaczewski, P. & Matlashewski, G. Evidence that interspecies Leishmania hybrids contribute to changes in disease pathology. bioRxiv, 10.1101/2020.06.29.177667 (2020).

Aslett M, et al. TriTrypDB: a functional genomic resource for the Trypanosomatidae. Nucleic Acids Res. 2010;38:D457–D462. doi: 10.1093/nar/gkp851. PubMed DOI PMC

Li H, Durbin R. Fast and accurate short read alignment with Burrows-Wheeler transform. Bioinformatics. 2009;25:1754–1760. doi: 10.1093/bioinformatics/btp324. PubMed DOI PMC

Koboldt DC, et al. VarScan 2: somatic mutation and copy number alteration discovery in cancer by exome sequencing. Genome Res. 2012;22:568–576. doi: 10.1101/gr.129684.111. PubMed DOI PMC

Krzywinski M, et al. Circos: an information aesthetic for comparative genomics. Genome Res. 2009;19:1639–1645. doi: 10.1101/gr.092759.109. PubMed DOI PMC

Robinson JT, et al. Integrative genomics viewer. Nat. Biotechnol. 2011;29:24–26. doi: 10.1038/nbt.1754. PubMed DOI PMC

Hajjaran H, et al. Detection and molecular identification of Leishmania RNA virus (LRV) in Iranian Leishmania species. Arch. Virol. 2016;161:3385–3390. doi: 10.1007/s00705-016-3044-z. PubMed DOI

Volf P, Volfova V. Establishment and maintenance of sand fly colonies. J. Vector Ecol. 2011;36:S1–9. doi: 10.1111/j.1948-7134.2011.00106.x. PubMed DOI

Myskova J, Votypka J, Volf P. Leishmania in sand flies: comparison of quantitative polymerase chain reaction with other techniques to determine the intensity of infection. J. Med. Entomol. 2008;45:133–138. doi: 10.1093/jmedent/45.1.133. PubMed DOI

Sadlova J, et al. The stage-regulated HASPB and SHERP proteins are essential for differentiation of the protozoan parasite Leishmania major in its sand fly vector, Phlebotomus papatasi. Cell Microbiol. 2010;12:1765–1779. doi: 10.1111/j.1462-5822.2010.01507.x. PubMed DOI PMC

Rodgers MR, Popper SJ, Wirth DF. Amplification of kinetoplast DNA as a tool in the detection and diagnosis of Leishmania. Exp. Parasitol. 1990;71:267–275. doi: 10.1016/0014-4894(90)90031-7. PubMed DOI

Safety and reactogenicity of a controlled human infection model of sand fly-transmitted cutaneous leishmaniasis

The development of L. major, L. donovani and L. martiniquensis, Leishmania currently emerging in Europe, in the sand fly species Phlebotomus perniciosus and P. tobbi

A clinical study to optimise a sand fly biting protocol for use in a controlled human infection model of cutaneous leishmaniasis (the FLYBITE study)