Development of an Indirect Fluorescent Antibody (IFA) Assay for the Detection of Leishmania RNA Virus 2 (LRV2) in Leishmania Parasites
-
Homa Hajjaran
,
Maryam Ebadizadeh
,
Angila Ataei-Pirkooh
,
Mehdi Mohebali
,
Katayoun Samimi-Rad
,
Reza Saberi
,
Saied Reza Naddaf
Abstract
Background: Detection of Leishmania RNA virus (LRV) in Old World Leishmania species and their possible role in the disease prognosis requires sensitive and specific methods, preferably independent of the viral genome. We aimed to develop an indirect immunofluorescence antibody (IFA) assay to detect LRV in the Old World Leishmania parasites.
Methods: Clinical samples were collected from 86 cutaneous leishmaniasis (CL) patients in different endemic areas of CL in Iran, during 2017-2019. For antibody preparation, the viruses were obtained from sediment of an LRV-infected L. major culture-using freeze and thaw cycles followed by gradient cesium chloride centrifugation. The purified viruses were used to immunize a male 3-4 months rabbit. Various dilutions of the LRV-immunized rabbit's serum and a conjugated antibody were deployed to detect LRV in 48 isolates by IFA assay.
Results: LRV virus was detected in four of the 48 CL cases using IFA method. Amplification of a partial fragment of RNA-dependent RNA polymerase (RdRp) gene from the isolates confirmed the IFA results. In phylogeny, the generated RdRp sequences from four isolates were grouped with the other Old World LRVs, but separate from L. aethiopica LRVs, which appeared as a highly supported distinct clade.
Conclusion: Further optimization of this approach to detect the LRV directly in lesion scrapings can make it a more reliable tool for field studies and disclosing the virus's possible role in disseminating and unusual clinical features.
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10. Hajjaran H, Mahdi M, Mohebali M, et al. Detection and molecular identification of Leishmania RNA virus (LRV) in Iranian Leishmania species. Arch Virol. 2016;161:3385-3390.
11. Nalçacı M, Karakuş M, Yılmaz B, Demir S, Özbilgin A, Özbel Y, Töz S. Detection of Leishmania RNA virus 2 in Leishmania species from Turkey. Trans R Soc Trop Med Hyg. 2019;113:410-417.
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13. Hartley M-A, Ronet C, Zangger H, Beverley SM, Fasel N. Leishmania RNA virus: When the host pays the toll. Front Cell Infect Microbiol. 2012;2:99.
14. 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.
15. Ives A, Ronet C, Prevel F, et al. Leishmania RNA virus controls the severity of mucocutaneous leishmaniasis. Science. 2011;331:775-778.
16. Ives A, Masina S, Castiglioni P, et al. Myd88 and TLR9 dependent immune responses mediate resistance to Leishmania guyanensis infections, irrespective of Leishmania RNA virus burden. PLoS One. 2014;9
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19. Hajjaran H, Saberi R, Borjian A, Fakhar M, Hosseini SA, Ghodrati S, Mohebali M. The Geographical Distribution of Human Cutaneous and Visceral Leishmania Species Identified by Molecular Methods in Iran: A Systematic Review with Meta-Analysis. Front Public Health. 2021;9:661674.
20. Hajjaran H, Mohebali M, Teimouri A, et al. Identification and phylogenetic relationship of Iranian strains of various Leishmania species isolated from cutaneous and visceral cases of leishmaniasis based on N-acetylglucosamine-1-phosphate transferase gene. Infect Genet Evol. 2014;26:203-212.
21. Hajjaran H, Mohebali M, Mamishi S, et al. Molecular identification and polymorphism determination of cutaneous and visceral leishmaniasis agents isolated from human and animal hosts in Iran. Biomed Res Int. 2013; 789326.
22. Kumar S, Stecher G, Tamura K. Mega7: Molecular evolutionary genetics analysis version 7.0 for bigger datasets. Mol Biol Evol. 2016;33:1870-1874.
23. Hamidi-Fard M, Ataei-Pirkooh A, Aghasadeghi M, Kazemi R. Purification of human reovirus without the need for equipment of suspension cell culture. Iran J Virol.2019;13:35-40.
24. Ataei Pirkooh A, Shahrabadi MS. Development of a latex agglutination method for diagnosis of rotavirus infections. Iran J Med Sci. 2007; 32(2): 100-104.
25. De Carlo S, Harris JR. Negative staining and cryo-negative staining of macromolecules and viruses for tem. Micron. 2011; 42(2):117-31.
26. Croft S, Molyneux D. Studies on the ultrastructure, virus-like particles and infectivity of Leishmania hertigi. Ann Trop Med Parasitol. 1979;73:213-226.
27. Mikaeili F, Fakhar M, Sarkari B, Motazedian MH, Hatam G. Comparison of serological methods (ELISA, DAT and IFA) for diagnosis of visceral leishmaniasis utilizing an endemic strain. Iran J Immunol. 2007;4:116-121.
28. World Health Organization, 2018. Leishmaniasis. http://www.who.int/mediacentre/fact sheets/fs375/en/. (Accessed 15 May 2018).
29. Shirzadi M, Esfahania S, Mohebalia M, et al. Epidemiological status of leishmaniasis in the Islamic Republic of Iran, 1983-2012. East Mediterr Health J. 2015;21:736-42.
30. Hajjaran H, Mohebali M, Akhavan A, Taheri A, Barikbin B, Soheila NS. Unusual presentation of disseminated cutaneous leishmaniasis due to Leishmania major: Case reports of four Iranian patients. Asian Pac J Trop Med. 2013; 6(4):333-6.
31. Hashemi SA, Badirzadeh A, Sabzevari S, Nouri A, Seyyedin M. First case report of atypical disseminated cutaneous leishmaniasis in an opium abuser in Iran. Rev Inst Med Trop Sao Paulo. 2018 ;60:e5.
32. Atayde VD, da Silva Lira Filho A, Chaparro V, et al. Exploitation of the Leishmania exosomal pathway by Leishmania RNA virus 1. Nat Microbio. 2019;4:714-723 .
2. Tamiru HF, Mashalla YJ, Mohammed R, Tshweneagae GT. Cutaneous leishmaniasis a neglected tropical disease: community knowledge, attitude and practices in an endemic area, Northwest Ethiopia. BMC Infect Dis. 2019;19(1):855.
3. Grieve S, Steffen R. Epidemiology: Morbidity and mortality in travelers. Travel Med. 2019;3-14.
4. Burza S, Croft SL, Boelaert M. Leishmaniasis. Lancet. 2018;392:951-970.
5. Bussotti G, Gouzelou E, Boité MC, et al. Leishmania genome dynamics during environmental adaptation reveal strain-specific differences in gene copy number variation, karyotype instability, and telomeric amplification. mBio.2018; 9:e01399-01318.
6. Gómez-Arreaza A, Haenni A-L, Dunia I, Avilan L. Viruses of parasites as actors in the parasite-host relationship: A “ménage à trois”. Acta Trop. 2017;166:126-132.
7. Kariyawasam R, Mukkala AN, Lau R, Valencia BM, Llanos-Cuentas A, Boggild AK. Virulence factor RNA transcript expression in the Leishmania viannia subgenus: Influence of species, isolate source, and Leishmania RNA virus-1. Trop Med Health. 2019;47:25.
8. Zangger H, Ronet C, Desponds C, et al. Detection of Leishmania RNA virus in Leishmania parasites. PLoS Negl Trop Dis. 2013;7
9. Kleschenko Y, Grybchuk D, Matveeva NS, et al. Molecular characterization of Leishmania RNA virus 2 in Leishmania major from Uzbekistan. Genes. 2019;10:830.
10. Hajjaran H, Mahdi M, Mohebali M, et al. Detection and molecular identification of Leishmania RNA virus (LRV) in Iranian Leishmania species. Arch Virol. 2016;161:3385-3390.
11. Nalçacı M, Karakuş M, Yılmaz B, Demir S, Özbilgin A, Özbel Y, Töz S. Detection of Leishmania RNA virus 2 in Leishmania species from Turkey. Trans R Soc Trop Med Hyg. 2019;113:410-417.
12. Saberi R, Fakhar M, Hajjaran H, et al. Presence and diversity of Leishmania RNA virus in an old zoonotic cutaneous leishmaniasis focus, northeastern Iran: Haplotype and phylogenetic based approach. Int J Infect Dis. 2020;101:6-13.
13. Hartley M-A, Ronet C, Zangger H, Beverley SM, Fasel N. Leishmania RNA virus: When the host pays the toll. Front Cell Infect Microbiol. 2012;2:99.
14. 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.
15. Ives A, Ronet C, Prevel F, et al. Leishmania RNA virus controls the severity of mucocutaneous leishmaniasis. Science. 2011;331:775-778.
16. Ives A, Masina S, Castiglioni P, et al. Myd88 and TLR9 dependent immune responses mediate resistance to Leishmania guyanensis infections, irrespective of Leishmania RNA virus burden. PLoS One. 2014;9
17. Robinson JI. Antiviral nucleoside inhibitors of Leishmania RNA virus 1: Discovery and mechanism. Washington University in St. Louis; 2017.
18. de Carvalho RV, Lima-Junior DS, da Silva MVG, et al. Leishmania RNA virus exacerbates leishmaniasis by subverting innate immunity via tlr3-mediated nlrp3 inflammasome inhibition. Nat Commun. 2019;10:1-17.
19. Hajjaran H, Saberi R, Borjian A, Fakhar M, Hosseini SA, Ghodrati S, Mohebali M. The Geographical Distribution of Human Cutaneous and Visceral Leishmania Species Identified by Molecular Methods in Iran: A Systematic Review with Meta-Analysis. Front Public Health. 2021;9:661674.
20. Hajjaran H, Mohebali M, Teimouri A, et al. Identification and phylogenetic relationship of Iranian strains of various Leishmania species isolated from cutaneous and visceral cases of leishmaniasis based on N-acetylglucosamine-1-phosphate transferase gene. Infect Genet Evol. 2014;26:203-212.
21. Hajjaran H, Mohebali M, Mamishi S, et al. Molecular identification and polymorphism determination of cutaneous and visceral leishmaniasis agents isolated from human and animal hosts in Iran. Biomed Res Int. 2013; 789326.
22. Kumar S, Stecher G, Tamura K. Mega7: Molecular evolutionary genetics analysis version 7.0 for bigger datasets. Mol Biol Evol. 2016;33:1870-1874.
23. Hamidi-Fard M, Ataei-Pirkooh A, Aghasadeghi M, Kazemi R. Purification of human reovirus without the need for equipment of suspension cell culture. Iran J Virol.2019;13:35-40.
24. Ataei Pirkooh A, Shahrabadi MS. Development of a latex agglutination method for diagnosis of rotavirus infections. Iran J Med Sci. 2007; 32(2): 100-104.
25. De Carlo S, Harris JR. Negative staining and cryo-negative staining of macromolecules and viruses for tem. Micron. 2011; 42(2):117-31.
26. Croft S, Molyneux D. Studies on the ultrastructure, virus-like particles and infectivity of Leishmania hertigi. Ann Trop Med Parasitol. 1979;73:213-226.
27. Mikaeili F, Fakhar M, Sarkari B, Motazedian MH, Hatam G. Comparison of serological methods (ELISA, DAT and IFA) for diagnosis of visceral leishmaniasis utilizing an endemic strain. Iran J Immunol. 2007;4:116-121.
28. World Health Organization, 2018. Leishmaniasis. http://www.who.int/mediacentre/fact sheets/fs375/en/. (Accessed 15 May 2018).
29. Shirzadi M, Esfahania S, Mohebalia M, et al. Epidemiological status of leishmaniasis in the Islamic Republic of Iran, 1983-2012. East Mediterr Health J. 2015;21:736-42.
30. Hajjaran H, Mohebali M, Akhavan A, Taheri A, Barikbin B, Soheila NS. Unusual presentation of disseminated cutaneous leishmaniasis due to Leishmania major: Case reports of four Iranian patients. Asian Pac J Trop Med. 2013; 6(4):333-6.
31. Hashemi SA, Badirzadeh A, Sabzevari S, Nouri A, Seyyedin M. First case report of atypical disseminated cutaneous leishmaniasis in an opium abuser in Iran. Rev Inst Med Trop Sao Paulo. 2018 ;60:e5.
32. Atayde VD, da Silva Lira Filho A, Chaparro V, et al. Exploitation of the Leishmania exosomal pathway by Leishmania RNA virus 1. Nat Microbio. 2019;4:714-723 .
| Files | ||
| Issue | Vol 17 No 3 (2022) | |
| Section | Original Article(s) | |
| DOI | https://doi.org/10.18502/ijpa.v17i3.10625 | |
| Keywords | ||
| Leishmania RNA virus Indirect fluorescence antibody (RdRp) gene | ||
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How to Cite
1.
Hajjaran H, Ebadizadeh M, Ataei-Pirkooh A, Mohebali M, Samimi-Rad K, Saberi R, Naddaf SR. Development of an Indirect Fluorescent Antibody (IFA) Assay for the Detection of Leishmania RNA Virus 2 (LRV2) in Leishmania Parasites. Iran J Parasitol. 2022;17(3):349-357.
