• View in gallery

    Leishmania RNA virus type 1 (LRV1) detection in different Leishmania isolates. MW (molecular weight marker 100 bp, Sigma®, St. Louis, MO); 1, 5, and 6: LRV1 positive Leishmania guyanensis isolates; 2: LRV1 negative L. guyanensis isolate; 3 and 4: LRV1 negative Leishmania amazonensis isolates; 7: LRV1 positive Leishmania braziliensis isolate; 8: LRV1 positive control (reference strain of L. guyanensis MHOM/GF/97/LBC6); 9: LRV1 negative control (water).

  • View in gallery

    Distribution of Leishmania isolates carrying or not the Leishmania RNA virus (LRV), in French Guiana. Pie charts full correspond to positive LRV and pie charts with patterns correspond to negative LRV.

  • 1.

    Miller JH, Schwartzwelter JC, 1960. Virus-like particles in an Entamoeba histolytica trophozoite. J Parasitol 46: 523524.

  • 2.

    Molyneux DH, 1974. Virus-like particles in Leishmania parasites. Nature 249: 588589.

  • 3.

    Diamond LS, Mattern CF, 1976. Protozoal viruses. Adv Virus Res 20: 87112.

  • 4.

    Wang AL, Wang CC, 1986. Discovery of a specific double-stranded RNA virus in Giardia lamblia. Mol Biochem Parasitol 21: 269276.

  • 5.

    Wang AL, Wang CC, 1986. The double-stranded RNA in Trichomonas vaginalis may originate from virus-like particles. Proc Natl Acad Sci USA 83: 79567960.

    • Search Google Scholar
    • Export Citation
  • 6.

    Tipper DJ, Bostian KA, 1984. Double-stranded ribonucleic acid killer systems in yeasts. Microbiol Rev 48: 125156.

  • 7.

    Tarr PI, Aline RF Jr, Smiley BL, Scholler J, Keithly J, Stuart K, 1988. LR1: a candidate RNA virus of Leishmania. Proc Natl Acad Sci USA 85: 95729575.

  • 8.

    Cadd TL, Keenan MC, Patterson JL, 1993. Detection of Leishmania RNA virus 1 proteins. J Virol 67: 56475650.

  • 9.

    Zangger H, Hailu A, Desponds C, Lye LF, Akopyants NS, Dobson DE, Ronet C, Ghalib H, Beverley SM, Fasel N, 2014. Leishmania aethiopica field isolates bearing an endosymbiontic dsRNA virus induce pro-inflammatory cytokine response. PLoS Negl Trop Dis 8: e2836.

    • Search Google Scholar
    • Export Citation
  • 10.

    Guilbride L, Myler PJ, Stuart K, 1992. Distribution and sequence divergence of LRV1 viruses among different Leishmania species. Mol Biochem Parasitol 54: 101104.

    • Search Google Scholar
    • Export Citation
  • 11.

    Widmer G, Dooley S, 1995. Phylogenetic analysis of Leishmania RNA virus and Leishmania suggests ancient virus-parasite association. Nucleic Acids Res 23: 23002304.

    • Search Google Scholar
    • Export Citation
  • 12.

    Stuart KD, Weeks R, Guilbride L, Myler PJ, 1992. Molecular organization of Leishmania RNA virus 1. Proc Natl Acad Sci USA 89: 85968600.

  • 13.

    Scheffter SM, Ro YT, Chung IK, Patterson JL, 1995. The complete sequence of Leishmania RNA virus LRV2-1, a virus of an old world parasite strain. Virology 212: 8490.

    • Search Google Scholar
    • Export Citation
  • 14.

    Scheffter S, Widmer G, Patterson JL, 1994. Complete sequence of Leishmania RNA virus 1–4 and identification of conserved sequences. Virology 199: 479483.

    • Search Google Scholar
    • Export Citation
  • 15.

    Rotureau B, Ravel C, Nacher M, Couppié P, Curtet I, Dedet JP, Carme B, 2006. Molecular epidemiology of Leishmania (Viannia) guyanensis in French Guiana. J Clin Microbiol 44: 468473.

    • Search Google Scholar
    • Export Citation
  • 16.

    Simon S, Veron V, Carme B, 2010. Leishmania spp. identification by polymerase chain reaction–restriction fragment length polymorphism analysis and its applications in French Guiana. Diagn Microbiol Infect Dis 66: 175180.

    • Search Google Scholar
    • Export Citation
  • 17.

    Ives A, Ronet C, Prevel F, Ruzzante G, Fuertes-Marraco S, Schutz F, Zangger H, Revaz-Breton M, Lye L, Hickerson SM, Beverley SM, Acha-Orbea H, Launois P, Fasel N, Masina S, 2011. Leishmania RNA virus controls the severity of mucocutaneous leishmaniasis. Science 331: 775778.

    • Search Google Scholar
    • Export Citation
  • 18.

    Salinas G, Zamora M, Stuart K, Saravia N, 1996. Leishmania RNA viruses in Leishmania of the Viannia subgenus. Am J Trop Med Hyg 54: 425429.

  • 19.

    Saiz M, Llanos-Cuentas A, Echevarria J, Roncal N, Cruz M, Muniz MT, Lucas C, Wirth DF, Scheffter S, Magill AJ, Patterson JL, 1998. Short report: detection of Leishmania virus in human biopsy samples of leishmaniasis from Peru. Am J Trop Med Hyg 58: 192194.

    • Search Google Scholar
    • Export Citation
  • 20.

    Ogg MM, Carrion R Jr, Botelho AC, Mayrink W, Correa-Oliveira R, Patterson JL, 2003. Short report: quantification of Leishmania virus RNA in clinical samples and its possible role in pathogenesis. Am J Trop Med Hyg 69: 309313.

    • Search Google Scholar
    • Export Citation
  • 21.

    Bourreau E, Ginouves M, Prévot G, Hartley M, Gangneux J, Robert-Gangneux F, Dufour J, Marie DS, Bertolotti A, Pratlong F, Martin R, Schütz F, Couppié P, Fasel N, Ronet C, 2015. Leishmania RNA virus presence in L. guyanensis parasites increases the risk of first-line treatment failure and symptomatic relapse. J Infect Dis. [epub ahead of print].

    • Search Google Scholar
    • Export Citation
  • 22.

    Pereira L de O, Maretti-Mira AC, Rodrigues KM, Lima RB, Oliveira-Neto MP, Cupolillo E, Pirmez C, de Oliveira MP, 2013. Severity of tegumentary leishmaniasis is not exclusively associated with Leishmania RNA virus 1 infection in Brazil. Mem Inst Oswaldo Cruz 108: 665667.

    • Search Google Scholar
    • Export Citation
  • 23.

    Rotureau B, Gaborit P, Issaly J, Carinci R, Fouque F, Carme B, 2006. Diversity and ecology of sand flies (Diptera: Psychodidae: Phlebotominae) in coastal French Guiana. Am J Trop Med Hyg 75: 6269.

    • Search Google Scholar
    • Export Citation
  • 24.

    Fouque F, Gaborit P, Issaly J, Carinci R, Gantier JC, Ravel C, Dedet JP, 2007. Phlebotomine sand flies (Diptera: Psychodidae) associated with changing patterns in the transmission of the human cutaneous leishmaniasis in French Guiana. Mem Inst Oswaldo Cruz 102: 3540.

    • Search Google Scholar
    • Export Citation
  • 25.

    Croft SL, Molyneux DH, 1979. Studies on the ultrastructure, virus-like particles and infectivity of Leishmania hertigi. Ann Trop Med Parasitol 73: 213226.

    • Search Google Scholar
    • Export Citation
  • 26.

    Ronet C, Beverley SM, Fasel N, 2011. Muco-cutaneous leishmaniasis in the new world: the ultimate subversion. Virulence 2: 547552.

  • 27.

    Ives A, Masina S, Castiglioni P, Prevel F, Revaz-Breton M, Hartley MA, Launois P, Fasel N, Ronet C, 2014. MyD88 and TLR9 dependent immune responses mediate resistance to Leishmania guyanensis infections, irrespective of Leishmania RNA virus burden. PLoS One 9: e96766.

    • Search Google Scholar
    • Export Citation
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Prevalence and Distribution of Leishmania RNA Virus 1 in Leishmania Parasites from French Guiana

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  • Ecosystèmes Amazoniens et Pathologie Tropicale–EA 3593–Labex CEBA–Medicine Department, University of French Guiana, Cayenne, French Guiana; Laboratoire Associé–Centre National de Référence Leishmania, Laboratory of Parasitology and Mycology, Centre Hospitalier Andrée Rosemon, Cayenne, French Guiana; Immunology Laboratory of Leishmaniasis, Pasteur Institute of French Guiana, Cayenne, French Guiana; Laboratory of Virus-Host Interactions, Pasteur Institute of French Guiana, Cayenne, French Guiana; Department of Biochemistry, University of Lausanne, Epalinges, Switzerland; Guianan Institute of Tropical Dermatology, Centre Hospitalier Andrée Rosemon, Cayenne, French Guiana; Centre d'Investigation Clinique Epidémiologie Clinique Antilles Guyane, Cayenne General Hospital, Cayenne, French Guiana; Laboratory of Parasitology and Mycology, Centre Hospitalier Andrée Rosemon, Cayenne, French Guiana

In South America, the presence of the Leishmania RNA virus type 1 (LRV1) was described in Leishmania guyanensis and Leishmania braziliensis strains. The aim of this study was to determine the prevalence distribution of LRV1 in Leishmania isolates in French Guiana given that, in this French overseas department, most Leishmania infections are due to these parasite species. The presence of the virus was observed in 74% of Leishmania spp. isolates, with a highest presence in the internal areas of the country.

Introduction

Virus-like particles (VLPs) discovery began in the 1960s with the first report in the parasitic protozoan Entamoeba histolytica.1 Since then similar structures have been identified in an ever-expanding list of unicellular eukaryotes: Leishmania,2 Plasmodium,3 Naegleria,3 Giardia,4 Trichomonas,5 as well as in other parasite species and fungi.6 The first discovery of VLPs in Leishmania was reported by Molyneux2 in Leishmania hertigi, a nonhuman parasite species. The presence of VLPs in another Leishmania species was then highlighted by Tarr and others,7 namely the Leishmania (braziliensis) guyanensis species. Leishmania RNA virus (LRV) is an ico…sahedral encapsidated double-stranded 5.3-kb RNA virus, belonging to the Totiviridae family. This virus has been observed in the New World Leishmania species: L. (Viannia) guyanensis and L. (Viannia) braziliensis, in one strain of the Old World parasite Leishmania major, and in Leishmania aethiopica.8,9 New World strains are grouped into the LRV1 type, which is divided in 14 subtypes (LRV1-1 to LRV1-14) that spread over South America.7,10,11 Old World strains belong to the LRV2 type.9

The LRV complementary DNA (cDNA) was first completely sequenced by Stuart and others12 for the prototype virus termed LRV1-1. Then, complete cDNA sequences were reported by Scheffter and others13,14 for the LRV1-4 and LRV2-1 isolates. The LRV1-1 and LRV1-4 sequences share 77% nucleotide identity between themselves.14

In French Guiana, Leishmania species predominantly diagnosed are L. guyanensis, L. braziliensis, and occasionally Leishmania amazonensis (Simon and others, submitted data).15 Leishmaniasis represents a significant risk for those in contact with the forest. Our study aimed to determine the prevalence of LRV1 in the circulating isolates of Leishmania spp. in French Guiana and to establish its repartition on the territory.

Materials and Methods

Parasites.

A total of 129 isolates of Leishmania spp. successfully cultured, from 424 patients infected in French Guiana between 2012 and 2014, were kindly supplied by the parasitology and mycology laboratory of the Cayenne General Hospital, with strict respect for patient anonymity. Promastigote parasites were cultured in Roswell Park Memorial Institute medium 1640 (Gibco, Paisley, Scotland, UK) containing l-glutamine, 20 mM N-2-hydroxyethylpiperazine-N-2-ethane sulfonic acid, phenol red and supplemented with 20% heat-inactivated fetal calf serum (Gibco), 50 IU/mL penicillin (Invitrogen), 0.05 mg/mL streptomycin (Invitrogen), and nonessential amino acids (Gibco).

Leishmania isolates identification was performed by polymerase chain reaction (PCR) restriction fragment length polymorphism as previously described.16

When parasites reached a stationary phase, counting was performed in a Malassez cell. Pellets containing 1 × 107 parasites were constituted after culture by centrifugation for 5 minutes at 514 g and removal of the supernatant. The pellets were stored at −80°C until RNA extraction.

Total RNA extraction.

Total RNA was extracted using Trizol® (Ambion, Life Technologies, Carlsbad, CA) according to the manufacturer's recommendations, with minor modifications. Pellets were thawed at room temperature (RT) and homogenized with 1 mL of Trizol. After 15–30 minutes of incubation at RT, 0.2 mL of chloroform per milliliter of Trizol was added, vortexed for 15 seconds, and incubated for 2–15 minutes at RT. Phase separation by centrifugation for 10 minutes at 12,000 g and 4°C allowed recovering the upper phase to which 0.5 mL of isopropanol was added. The mixture was then incubated for 10 minutes at RT and centrifuged for 10 minutes at 12,000 g at 4°C. The RNA pellet was then washed with 75% ethanol, dried, and dissolved in 10 μL of diethylpyrocarbonate water. The RNA was stored at −80°C.

Reverse transcription PCR.

cDNA was synthesized using the SuperScript II Reverse Transcriptase (Invitrogen), according to the manufacturer's recommendations with random hexamers (Invitrogen). Then, PCR amplification of a 124-bp LRV fragment located in the most conserved LRV1 region was done with the LRV1 forward primer, LRV1-F1: 5′-CTGACTGGACGGGGGGTAAT-3′ and LRV1 reverse primer, LRV1-R1: 5′-CAAAACACTCCCTTACGC-3′ enabling to amplify all LRV1 subtypes that have been slightly modified from those of Ives and others.17 These primers were based on LRV1-1 and LRV1-4 genome sequences (Genbank accession nos.: LRV1-1: NC002063 and LRV1-4: NC003601). A denaturation step at 94°C for 2 minutes was followed by 40 cycles at 94°C for 30 seconds, 54°C for 30 seconds, and 72°C for 1 minute. The PCR was completed by a final elongation at 72°C for 5 minutes. PCR products were analyzed on a 2% gel agarose to verify the presence of amplification products at the expected size. The reference strain of L. guyanensis (MHOM/GF/97/LBC6) and water were used as positive and negative controls, respectively, in each reverse transcriptase (RT) PCR experiment.

Ethical aspects.

The study was retrospective. All patients were informed (during consultation, with posters) that data and analysis of results may be used in research and that they have a right to refuse.

Statistical analysis.

Statistical significance between proportion of LRV from coastal communes and inland communes and proportion of LRV from coastal area (seaside) and the rest of the territory were determined by χ2 test; P < 0.05 was considered significant.

Results

Search for LRV1 was performed on 129 Leishmania isolates including 112 isolates of L. guyanensis, 11 isolates of L. braziliensis, and six isolates of L. amazonensis. Its presence was detected in 96 of the 129 isolates (74%).

Figure 1 shows RT-PCR products obtained from a panel of different LRV1 positive or negative Leishmania species. Table 1 groups the results per site and Leishmania species. Eighty percent (90/112) of L. guyanensis isolates and 55% (6/11) of L. braziliensis isolates were LRV positive. Sequencing of the PCR fragment obtained from an L. braziliensis isolate confirmed the detection of LRV1-4. No LRV1 was detected from L. amazonensis isolates.

Figure 1.
Figure 1.

Leishmania RNA virus type 1 (LRV1) detection in different Leishmania isolates. MW (molecular weight marker 100 bp, Sigma®, St. Louis, MO); 1, 5, and 6: LRV1 positive Leishmania guyanensis isolates; 2: LRV1 negative L. guyanensis isolate; 3 and 4: LRV1 negative Leishmania amazonensis isolates; 7: LRV1 positive Leishmania braziliensis isolate; 8: LRV1 positive control (reference strain of L. guyanensis MHOM/GF/97/LBC6); 9: LRV1 negative control (water).

Citation: The American Society of Tropical Medicine and Hygiene 94, 1; 10.4269/ajtmh.15-0419

Table 1

Leishmania LRV positive or negative summary isolates

SpeciesLeishmania guyanensisLeishmania braziliensisLeishmania amazonensisLeishmania spp.POS (%)
LRVPOSNEGPOSNEGPOSNEGPOSNEG
Presumed site of infection
 Cacao6200006275
 Camopi3100003175
 Grand-Santi3010034357
 Iracoubo010000010
 Kourou10000010100
 Macouria040000040
 Mana010000010
 Maripasoula122210014382
 Montsinery10000010100
 Nouragues10000010100
 Papaïchton6100006186
 Régina142010014382
 Rémire1100001150
 Roura40000040100
 Saint Elie1100001150
 Saint Georges de l'Oyapock2100002167
 Saint Laurent du Maroni1100001150
 Saül901000100100
 Unknown/outisde2542303271073
Total902265069633 
%80 55 0 74  

LRV = Leishmania RNA virus; NEG = negative; POS = positive.

Figure 2 shows the distribution of Leishmania isolates carrying or not LRV across French Guiana. There were no significant differences between the proportion of LRV from coastal and inland communes (P = 0.22), however, there was a significant difference between the seaside and the rest of the territory (P < 0.001). Indeed, apart from Kourou, where the only available isolate was positive to LRV, isolates from Mana, Iracoubo, and Macouria were negative. Half isolates from Rémire were positive to LRV. In all inland communes, more than half of the isolates were LRV positive. In communes with the highest sampling effort, LRV presence was predominant: Regina with 82% (14/17) of positive isolates, Maripasoula 82% (14/17), Saül 100% (10/10), Cacao 75% (6/8), and Papaïchton 86% (6/7).

Figure 2.
Figure 2.

Distribution of Leishmania isolates carrying or not the Leishmania RNA virus (LRV), in French Guiana. Pie charts full correspond to positive LRV and pie charts with patterns correspond to negative LRV.

Citation: The American Society of Tropical Medicine and Hygiene 94, 1; 10.4269/ajtmh.15-0419

The presumed site of infection was unknown, or outside French Guiana (N = 2), for 37 of 129 isolates. Among them, 73% proved to be LRV positive. The two isolates, whose suspected infection sites were outside French Guiana, were contracted in Manaus and Suriname and were LRV-positive L. guyanensis isolates.

Discussion

Leishmania RNA viruses can infect different Leishmania species. Surveys of New World parasites have identified LRV1 only in isolates that originated from the Amazon basin, such as L. guyanensis and L. braziliensis.7,10,1820 The average molecular prevalence of LRV that we report here in Leishmania spp. isolates circulating in French Guiana is equivalent to the one described by Bourreau and others,21 but substantially higher (74%) than those reported in other South American countries, which ranged from 5.8% in Colombia, 18% in Peru, and 25.5% in the Brazilian city of Caratinga, Minas Girais.1820 This difference can first be explained by the geographical origin of the Leishmania isolates and, second, by the type of biological material used for LRV detection. Indeed, in Colombia, the virus was detected from parasites originating from the Amazon area of the country whereas no virus was detected in other regions.18 In Brazil, Pereira and others22 reported two LRV1 positive isolates of 48 tested (4.1%). These two L. guyanensis positive isolates were from the Amazon region. In this study, we report the presence of LRV throughout the whole territory. This could be correlated to tropical climate conditions encountered in French Guiana. In contrast, the weaker presence of LRV along the coastal area was presumably due to the low density of vectors and reservoirs implying a low parasite transmission in this more urbanized area.23,24 Ecologic and epidemiologic factors related to Amazon region might be involved in the repartition of Leishmania parasites susceptible to harbor the LRV.18

Concerning the type of biological material used for the search of LRV in all previous studies, this search was performed on human biopsies containing low amounts of the amastigote form of the parasite.1820,22 In our study, we used cultured parasites corresponding to the promastigote form (107 cells) in which the quantity of LRV has been reported to be higher due to virus multiplication in culture25 (personal communication). This could thus explain the large difference of prevalence observed between our results and those previously published.1820,22

We here report the detection of LRV1 from different L. guyanensis and L. braziliensis isolates.7,10,1820 None of the six L. amazonensis isolates tested were positive for LRV1. With the exception of the Leishmania panamensis species for which no LRV1 has been identified,18 no information is currently available for the other New World Leishmania species. Therefore, LRV1 has so far only been detected in parasite species of the Leishmania Viannia subgenus.

LRV infection of parasites seems to affect their virulence in a murine model of mucosal cutaneous leishmaniasis.17,26,27 To determine if the clinical evolution of infected patients can be affected by the presence of the virus or can be correlated, at least in part, to the genetic variability of the virus for which at least 14 subtypes have been identified, we plan to, based on the present results, to characterize the genetic diversity of different LRV1 isolates at the genomic level. These results should enable us to gain insights into the role LRV1 plays in the evolution of leishmaniasis lesions and deepen the achievements of Bourreau and others21 describing the impact of LRV on first-line treatment failure and symptomatic relapse.

ACKNOWLEDGMENTS

We thank trainees Claudiane Flora and Yasmina Auguste for their participation in this research work.

  • 1.

    Miller JH, Schwartzwelter JC, 1960. Virus-like particles in an Entamoeba histolytica trophozoite. J Parasitol 46: 523524.

  • 2.

    Molyneux DH, 1974. Virus-like particles in Leishmania parasites. Nature 249: 588589.

  • 3.

    Diamond LS, Mattern CF, 1976. Protozoal viruses. Adv Virus Res 20: 87112.

  • 4.

    Wang AL, Wang CC, 1986. Discovery of a specific double-stranded RNA virus in Giardia lamblia. Mol Biochem Parasitol 21: 269276.

  • 5.

    Wang AL, Wang CC, 1986. The double-stranded RNA in Trichomonas vaginalis may originate from virus-like particles. Proc Natl Acad Sci USA 83: 79567960.

    • Search Google Scholar
    • Export Citation
  • 6.

    Tipper DJ, Bostian KA, 1984. Double-stranded ribonucleic acid killer systems in yeasts. Microbiol Rev 48: 125156.

  • 7.

    Tarr PI, Aline RF Jr, Smiley BL, Scholler J, Keithly J, Stuart K, 1988. LR1: a candidate RNA virus of Leishmania. Proc Natl Acad Sci USA 85: 95729575.

  • 8.

    Cadd TL, Keenan MC, Patterson JL, 1993. Detection of Leishmania RNA virus 1 proteins. J Virol 67: 56475650.

  • 9.

    Zangger H, Hailu A, Desponds C, Lye LF, Akopyants NS, Dobson DE, Ronet C, Ghalib H, Beverley SM, Fasel N, 2014. Leishmania aethiopica field isolates bearing an endosymbiontic dsRNA virus induce pro-inflammatory cytokine response. PLoS Negl Trop Dis 8: e2836.

    • Search Google Scholar
    • Export Citation
  • 10.

    Guilbride L, Myler PJ, Stuart K, 1992. Distribution and sequence divergence of LRV1 viruses among different Leishmania species. Mol Biochem Parasitol 54: 101104.

    • Search Google Scholar
    • Export Citation
  • 11.

    Widmer G, Dooley S, 1995. Phylogenetic analysis of Leishmania RNA virus and Leishmania suggests ancient virus-parasite association. Nucleic Acids Res 23: 23002304.

    • Search Google Scholar
    • Export Citation
  • 12.

    Stuart KD, Weeks R, Guilbride L, Myler PJ, 1992. Molecular organization of Leishmania RNA virus 1. Proc Natl Acad Sci USA 89: 85968600.

  • 13.

    Scheffter SM, Ro YT, Chung IK, Patterson JL, 1995. The complete sequence of Leishmania RNA virus LRV2-1, a virus of an old world parasite strain. Virology 212: 8490.

    • Search Google Scholar
    • Export Citation
  • 14.

    Scheffter S, Widmer G, Patterson JL, 1994. Complete sequence of Leishmania RNA virus 1–4 and identification of conserved sequences. Virology 199: 479483.

    • Search Google Scholar
    • Export Citation
  • 15.

    Rotureau B, Ravel C, Nacher M, Couppié P, Curtet I, Dedet JP, Carme B, 2006. Molecular epidemiology of Leishmania (Viannia) guyanensis in French Guiana. J Clin Microbiol 44: 468473.

    • Search Google Scholar
    • Export Citation
  • 16.

    Simon S, Veron V, Carme B, 2010. Leishmania spp. identification by polymerase chain reaction–restriction fragment length polymorphism analysis and its applications in French Guiana. Diagn Microbiol Infect Dis 66: 175180.

    • Search Google Scholar
    • Export Citation
  • 17.

    Ives A, Ronet C, Prevel F, Ruzzante G, Fuertes-Marraco S, Schutz F, Zangger H, Revaz-Breton M, Lye L, Hickerson SM, Beverley SM, Acha-Orbea H, Launois P, Fasel N, Masina S, 2011. Leishmania RNA virus controls the severity of mucocutaneous leishmaniasis. Science 331: 775778.

    • Search Google Scholar
    • Export Citation
  • 18.

    Salinas G, Zamora M, Stuart K, Saravia N, 1996. Leishmania RNA viruses in Leishmania of the Viannia subgenus. Am J Trop Med Hyg 54: 425429.

  • 19.

    Saiz M, Llanos-Cuentas A, Echevarria J, Roncal N, Cruz M, Muniz MT, Lucas C, Wirth DF, Scheffter S, Magill AJ, Patterson JL, 1998. Short report: detection of Leishmania virus in human biopsy samples of leishmaniasis from Peru. Am J Trop Med Hyg 58: 192194.

    • Search Google Scholar
    • Export Citation
  • 20.

    Ogg MM, Carrion R Jr, Botelho AC, Mayrink W, Correa-Oliveira R, Patterson JL, 2003. Short report: quantification of Leishmania virus RNA in clinical samples and its possible role in pathogenesis. Am J Trop Med Hyg 69: 309313.

    • Search Google Scholar
    • Export Citation
  • 21.

    Bourreau E, Ginouves M, Prévot G, Hartley M, Gangneux J, Robert-Gangneux F, Dufour J, Marie DS, Bertolotti A, Pratlong F, Martin R, Schütz F, Couppié P, Fasel N, Ronet C, 2015. Leishmania RNA virus presence in L. guyanensis parasites increases the risk of first-line treatment failure and symptomatic relapse. J Infect Dis. [epub ahead of print].

    • Search Google Scholar
    • Export Citation
  • 22.

    Pereira L de O, Maretti-Mira AC, Rodrigues KM, Lima RB, Oliveira-Neto MP, Cupolillo E, Pirmez C, de Oliveira MP, 2013. Severity of tegumentary leishmaniasis is not exclusively associated with Leishmania RNA virus 1 infection in Brazil. Mem Inst Oswaldo Cruz 108: 665667.

    • Search Google Scholar
    • Export Citation
  • 23.

    Rotureau B, Gaborit P, Issaly J, Carinci R, Fouque F, Carme B, 2006. Diversity and ecology of sand flies (Diptera: Psychodidae: Phlebotominae) in coastal French Guiana. Am J Trop Med Hyg 75: 6269.

    • Search Google Scholar
    • Export Citation
  • 24.

    Fouque F, Gaborit P, Issaly J, Carinci R, Gantier JC, Ravel C, Dedet JP, 2007. Phlebotomine sand flies (Diptera: Psychodidae) associated with changing patterns in the transmission of the human cutaneous leishmaniasis in French Guiana. Mem Inst Oswaldo Cruz 102: 3540.

    • Search Google Scholar
    • Export Citation
  • 25.

    Croft SL, Molyneux DH, 1979. Studies on the ultrastructure, virus-like particles and infectivity of Leishmania hertigi. Ann Trop Med Parasitol 73: 213226.

    • Search Google Scholar
    • Export Citation
  • 26.

    Ronet C, Beverley SM, Fasel N, 2011. Muco-cutaneous leishmaniasis in the new world: the ultimate subversion. Virulence 2: 547552.

  • 27.

    Ives A, Masina S, Castiglioni P, Prevel F, Revaz-Breton M, Hartley MA, Launois P, Fasel N, Ronet C, 2014. MyD88 and TLR9 dependent immune responses mediate resistance to Leishmania guyanensis infections, irrespective of Leishmania RNA virus burden. PLoS One 9: e96766.

    • Search Google Scholar
    • Export Citation

Author Notes

* Address correspondence to Marine Ginouvès, Ecosystèmes Amazoniens et Pathologie Tropicale–EA 3593–Labex CEBA–Medicine Department, University of French Guiana, Campus Saint Denis, Avenue d'Estrees, Cayenne 97300, French Guiana. E-mail: marine.ginouves@univ-guyane.fr

Financial support: This work was supported by the University of the French West Indies and French Guiana and the Ministère Français de l'Enseignement Supérieur et de la Recherche Scientifique. It has benefited from an Investissement d'Avenir grant managed by Agence Nationale de la Recherche (CEBA, reference no. ANR-10-LABX-25-01).

Conflict of interest: Conflicts that the editors consider relevant to the content of the manuscript have been disclosed.

Authors' addresses: Marine Ginouvès and Stéphane Simon, Ecosystèmes Amazoniens et Pathologie Tropicale–EA 3593–Labex CEBA–Medicine Department, University of French Guiana, Cayenne, French Guiana, and Laboratoire Associé–Centre National de Référence Leishmania, Laboratory of Parasitology and Mycology, Centre Hospitalier Andrée Rosemon, Cayenne, French Guiana, E-mails: marine.ginouves@univ-guyane.fr and stephane.simon@guyane.univ-ag.fr. Eliane Bourreau, Immunology Laboratory of Leishmaniasis, Pasteur Institute of French Guiana, Cayenne, French Guiana, E-mail: ebourreau@pasteur-cayenne.fr. Vincent Lacoste, Laboratory of Virus Hosts Interaction, Pasteur Institute of French Guiana, Cayenne, French Guiana, E-mail: vlacoste@pasteur-cayenne.fr. Catherine Ronet, Department of Biochemistry, University of Lausanne, Epalinges, Switzerland, E-mail: catherine.ronet@unil.ch. Pierre Couppié, Ecosystemes Amazoniens et Pathologie Tropicale–EA 3593–Labex CEBA–Medicine Department, University of French Guiana, Cayenne, French Guiana, and Guianan Institute of Tropical Dermatology, Centre Hospitalier Andrée Rosemon, Cayenne, French Guiana, E-mail: pierre.couppie@ch-cayenne.fr. Mathieu Nacher, Ecosystemes Amazoniens et Pathologie Tropicale–EA 3593–Labex CEBA–Medicine Department, University of French Guiana, Cayenne, French Guiana, and Centre d'Investigation Clinique Epidémiologie Clinique Antilles Guyane, Cayenne General Hospital, Cayenne, French Guiana, E-mail: mathieu.nacher@ch-cayenne.fr. Magalie Demar, Ecosystemes Amazoniens et Pathologie Tropicale–EA 3593–Labex CEBA–Medicine Department, University of French Guiana, Cayenne, French Guiana, Laboratoire Associé–Centre National de Référence Leishmania, Laboratory of Parasitology and Mycology, Centre Hospitalier Andrée Rosemon, Cayenne, French Guiana, and Laboratory of Parasitology and Mycology, Centre Hospitalier Andrée Rosemon, Cayenne, French Guiana, E-mail: magalie.demar@ch-cayenne.fr. Ghislaine Prévot, Ecosystemes Amazoniens et Pathologie Tropicale–EA 3593–Labex CEBA–Medicine Department, University of French Guiana, Cayenne, French Guiana, E-mail: fac.prevot@gmail.com.

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