BACKGROUND: The Greek island of Crete is endemic for both visceral leishmaniasis (VL) and recently increasing cutaneous leishmaniasis (CL). This study summarizes published data on the sand fly fauna of Crete, the results of new sand fly samplings and the description of a new sand fly species. METHODS: All published and recent samplings were carried out using CDC light traps, sticky traps or mouth aspirators. The specific status of Phlebotomus (Adlerius) creticus n. sp., was assessed by morphological analysis, cytochrome b (cytb) sequencing and MALDI-TOF protein profiling. RESULTS: Published data revealed the presence of 10 Phlebotomus spp. and 2 Sergentomyia spp. During presented field work, 608 specimens of 8 species of Phlebotomus and one species of Sergentomyia were collected. Both published data and present samplings revealed that the two most common and abundant species were Phlebotomus neglectus, a proven vector of Leishmania infantum causing VL, and Ph. similis, a suspected vector of L. tropica causing CL. In addition, the field surveys revealed the presence of a new species, Ph. (Adlerius) creticus n. sp. CONCLUSIONS: The identification of the newly described species is based on both molecular and morphological criteria, showing distinct characters of the male genitalia that differentiate it from related species of the subgenus Adlerius as well as species-specific sequence of cytb and protein spectra generated by MALDI-TOF mass spectrometry.

BioCeV Institute of Microbiology The Czech Academy of Sciences Vestec Czech Republic

Département de Parasitologie Mycologie Hôpital Avicenne AP HP Bobigny France

Department of Parasitology Faculty of Science Charles University Prague Czech Republic

FEA7510 ESCAPE USC ANSES VECPAR Faculté de Pharmacie Université de Reims Champagne Ardenne 51 rue Cognacq Jay 51096 Reims cedex Reims France

Laboratory of Clinical Bacteriology Parasitology Zoonoses and Geographical Medicine Faculty of Medicine University of Crete Heraklion Greece

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Maroli M, Feliciangeli MD, Bichaud L, Charrel RN, Gradoni L. Phlebotomine sandflies and the spreading of leishmaniases and other diseases of public health concern. Med Vet Entomol. 2013;27:123–147. doi: 10.1111/j.1365-2915.2012.01034.x. PubMed DOI

Antoniou M, Gramiccia M, Molina R, Dvorak V, Volf P. The role of indigenous phlebotomine sandflies and mammals in the spreading of leishmaniasis agents in the Mediterranean region. Euro Surveill. 2013;18:20540. doi: 10.2807/1560-7917.ES2013.18.30.20540. PubMed DOI

Svobodova M, Votypka J, Peckova J, Dvorak V, Nasereddin A, Baneth G, et al. Distinct transmission cycles of Leishmania tropica in 2 adjacent foci, Northern Israel. Emerg Infect Dis. 2006;12:1860–1868. doi: 10.3201/eid1212.060497. PubMed DOI PMC

Jacobson RL. Leishmania tropica (Kinetoplastida: Trypanosomatidae) - a perplexing parasite. Folia Parasitol. 2003;50:241–250. doi: 10.14411/fp.2003.042. PubMed DOI

Antoniou M, Haralambous C, Mazeris A, Pratlong F, Dedet J-P, Soteriadou K. Leishmania donovani causing visceral or cutaneous leishmaniasis in Europe: Cyprus evidence is an alarm call. Lancet Infect Dis. 2008;8:6–7. doi: 10.1016/S1473-3099(07)70297-9. PubMed DOI

Ntais P, Christodoulou V, Tsirigotakis N, Dokianakis E, Dedet J-P, Pratlong F, et al. Will the introduction of Leishmania tropica MON-58, in the island of Crete, lead to the settlement and spread of this rare zymodeme? Acta Trop. 2014;132:125–130. doi: 10.1016/j.actatropica.2014.01.003. PubMed DOI

Dvorak V, Shaw J, Volf P. Parasite biology: the vectors. In: Bruschi F, Gradoni L, editors. The leishmaniases: old neglected tropical diseases. Cham: Springer International Publishing; 2018. pp. 31–77.

Moher D, Shamseer L, Clarke M, Ghersi D, Liberati A, Petticrew M, et al. Preferred reporting items for systematic review and meta-analysis protocols (PRISMA-P) 2015 statement. Syst Rev. 2015;4:1. doi: 10.1186/2046-4053-4-1. PubMed DOI PMC

Blavier A, Laroche L, Randrianambinintsoa FJ, Lucas V, Gantier J-C, Léger N, et al. Phlebotomine sandflies (Diptera, Psychodidae) from the Ankarana tsingy of northern Madagascar: inventory and description of new taxa. Parasite. 2019;26:38. doi: 10.1051/parasite/2019039. PubMed DOI PMC

Artemiev MM. A revision of sandflies of the subgenus Adlerius (Diptera, Phlebotominae, Phlebotomus) Zool Zhurnal. 1980;59:1177–1192.

Lewis DJ. A taxonomic review of the genus Phlebotomus (Diptera: Psychodidae) Bull Br Mus Nat Hist (Ent). 1982;45:121–209.

Léger N, Pesson B, Madulo-Leblond G. Les phlébotomes de Grèce. Biol Gallo-hell. 1986;11:165–191.

Killick-Kendrick R, Tang Y, Killick-Kendrick M, Sang DK, Sirdar MK, Ke L, et al. The identification of female sandflies of the subgenus Larroussius by the morphology of the spermathecal ducts. Parassitologia. 1991;33:335–347. PubMed

Depaquit J, Léger N, Ferté H, Rioux JA, Gantier JC, Michaelides A, et al. Les Phlébotomes de l’île de Chypre III. Inventaire faunistique. arasite. 2001;8:11–20. PubMed

Galati EAB, Galvis-Ovallos F, Lawyer P, Léger N, Depaquit J. An illustrated guide for characters and terminology used in descriptions of Phlebotominae (Diptera, Psychodidae) Parasite. 2017;24:26. doi: 10.1051/parasite/2017027. PubMed DOI PMC

Esseghir S, Ready PD, Killick-Kendrick R, Ben-Ismail R. Mitochondrial haplotypes and phylogeography of Phlebotomus vectors of Leishmania major. Insect Mol Biol. 1997;6:211–225. doi: 10.1046/j.1365-2583.1997.00175.x. PubMed DOI

Parvizi P, Naddaf SR, AlaeeNovin E. Molecular typing and phylogenetic analysis of some species belonging to Phlebotomus (Larroussius) and Phlebotomus (Adlerius) subgenera (Diptera: Psychodidae) from two locations in Iran. Iran J Arthropod Borne Dis. 2010;4:1–10. PubMed PMC

Mahamdallie SS, Pesson B, Ready PD. Multiple genetic divergences and population expansions of a Mediterranean sandfly, Phlebotomus ariasi, in Europe during the Pleistocene glacial cycles. Heredity. 2011;106:714–726. doi: 10.1038/hdy.2010.111. PubMed DOI PMC

Krüger A, Strüven L, Post RJ, Faulde M. The sandflies (Diptera: Psychodidae, Phlebotominae) in military camps in northern Afghanistan (2007–2009), as identified by morphology and DNA ‘barcoding’. Ann Trop Med Parasitol. 2011;105:163–176. doi: 10.1179/136485911X12899838683241. PubMed DOI PMC

Depaquit J, Bounamous A, Akhoundi M, Augot D, Sauvage F, Dvorak V, et al. A taxonomic study of Phlebotomus (Larroussius) perfiliewi s.l. Infect Genet Evol. 2013;20:500–508. doi: 10.1016/j.meegid.2013.10.006. PubMed DOI

Latrofa MS, Dantas-Torres F, Weigl S, Tarallo VD, Parisi A, Traversa D, et al. Multilocus molecular and phylogenetic analysis of phlebotomine sand flies (Diptera: Psychodidae) from southern Italy. Acta Trop. 2011;119:91–98. doi: 10.1016/j.actatropica.2011.04.013. PubMed DOI

Thompson JD, Higgins DG, Gibson TJ. CLUSTAL W: improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position-specific gap penalties and weight matrix choice. Nucleic Acids Res. 1994;22:4673–4680. doi: 10.1093/nar/22.22.4673. PubMed DOI PMC

Kumar S, Stecher G, Tamura K. MEGA7: molecular evolutionary genetics analysis version 7.0 for bigger datasets. Mol Biol Evol. 2016;33:1870–1874. doi: 10.1093/molbev/msw054. PubMed DOI PMC

Tamura K, Nei M. Estimation of the number of nucleotide substitutions in the control region of mitochondrial DNA in humans and chimpanzees. Mol Biol Evol. 1993;10:512–526. PubMed

Lanfear R, Frandsen PB, Wright AM, Senfeld T, Calcott B. PartitionFinder 2: new methods for selecting partitioned models of evolution for molecular and morphological phylogenetic analyses. Mol Biol Evol. 2016;34:772–773. PubMed

Ronquist F, Teslenko M, van der Mark P, Ayres DL, Darling A, Höhna S, et al. MrBayes 3.2: efficient Bayesian phylogenetic inference and model choice across a large model space. Syst Biol. 2012;61:539–542. doi: 10.1093/sysbio/sys029. PubMed DOI PMC

Guindon S, Dufayard J-F, Lefort V, Anisimova M, Hordijk W, Gascuel O. New algorithms and methods to estimate maximum-likelihood phylogenies: assessing the performance of PhyML 3.0. Syst Biol. 2010;59:307–321. doi: 10.1093/sysbio/syq010. PubMed DOI

Yang Z. Computational molecular evolution. Oxford: University Press; 2006.

Huelsenbeck JP, Ronquist F. MRBAYES: Bayesian inference of phylogenetic trees. Bioinformatics. 2001;17:754–755. doi: 10.1093/bioinformatics/17.8.754. PubMed DOI

Felsenstein J. Confidence limits on phylogenies: an approach using the bootstrap. Evolution. 1985;39:783–791. doi: 10.1111/j.1558-5646.1985.tb00420.x. PubMed DOI

Dvorak V, Halada P, Hlavackova K, Dokianakis E, Antoniou M, Volf P. Identification of phlebotomine sand flies (Diptera: Psychodidae) by matrix-assisted laser desorption/ionization time of flight mass spectrometry. Parasit Vectors. 2014;7:21. doi: 10.1186/1756-3305-7-21. PubMed DOI PMC

Halada P, Hlavackova K, Dvorak V, Volf P. Identification of immature stages of phlebotomine sand flies using MALDI-TOF MS and mapping of mass spectra during sand fly life cycle. Insect Biochem Mol Biol. 2018;93:47–56. doi: 10.1016/j.ibmb.2017.12.005. PubMed DOI

ICZN International Commission on Zoological Nomenclature: Amendment of articles 8, 9, 10, 21 and 78 of the International Code of Zoological Nomenclature to expand and refine methods of publication. Bull Zool Nomencl. 2012;69:161–169. doi: 10.21805/bzn.v69i3.a8.161. PubMed DOI

Parrot L. Notes sur les phlébotomes. XIX. Phlébotomes de Crète. Arch Inst Pasteur Alger. 1936;14:50–52.

Adler S, Theodor O, Witenberg G. Investigations on Mediterranean kala azar. XI. A study of leishmaniasis in Canea (Crete) Proc R Soc Lond. 1938;125:491–516.

Ristorcelli A. Sur les phlébotomes de l’île de Crète. Ann Parasitol Hum Comp. 1939;17:355–358. doi: 10.1051/parasite/1939-1940174355. DOI

Hertig M. Phlebotomus and residual DDT in Greece and Italy. Am J Trop Med Hyg. 1949;s1-29:773–809. doi: 10.4269/ajtmh.1949.s1-29.773. PubMed DOI

Hadjinicolaou J. Present status of Phlebotomus in certain areas of Greece. Bull World Health Organ. 1958;19:967–979. PubMed PMC

Pesson B, Wallon M, Floer MT, Kristensen AR. Isoenzymatic studies of Mediterranean populations of sandflies of the subgenus Larroussius. Parassitologia. 1991;33:471–476. PubMed

Léger N, Pesson B, Madulo-Leblond G, Ferté H, Tselentis Y, Antoniou M, Les phlébotomes de Crète M. Résultats d’une enquête entomologique effectuée en Juillet 1988 et Août 1989. Biol Gallo-hell. 1993;20:135–143.

Chaniotis B, Spyridaki I, Scoulika E, Antoniou M. Colonization of Phlebotomus neglectus (Diptera: Psychodidae), the major vector of visceral leishmaniasis in Greece. J Med Entomol. 2000;37:346–348. doi: 10.1603/0022-2585(2000)037[0346:COPNDP]2.0.CO;2. PubMed DOI

Ivović V, Patakakis M, Tselentis Y, Chaniotis B. Faunistic study of sandflies in Greece. Med Vet Entomol. 2007;21:121–124. doi: 10.1111/j.1365-2915.2006.00649.x. PubMed DOI

Ivović V, Chaniotis V, Vujanić M, Bobić B, Nikolić A, Klun I, et al. Life tables and reproductive parameters of Phlebotomus neglectus Tonnoir, 1921 (Diptera, Psychodidae) under laboratory conditions. Arch Biol Sci. 2010;62:153–157. doi: 10.2298/ABS1001153I. DOI

Christodoulou V, Antoniou M, Ntais P, Messaritakis I, Ivović V, Dedet J-P, et al. Re-emergence of visceral and cutaneous leishmaniasis in the Greek Island of Crete. Vector Borne Zoonotic Dis. 2012;12:214–222. doi: 10.1089/vbz.2011.0004. PubMed DOI PMC

Alten B, Maia C, Afonso MO, Campino L, Jiménez M, González E, et al. Seasonal dynamics of phlebotomine sand fly species proven vectors of Mediterranean leishmaniasis caused by Leishmania infantum. PLoS Negl Trop Dis. 2016;10:e0004458. doi: 10.1371/journal.pntd.0004458. PubMed DOI PMC

Langeron M. Phlébotomes capturés en Crète. Ann Parasitol Hum Comp. 1923;1:108. doi: 10.1051/parasite/1923011108. DOI

Aransay AM, Scoulica E, Chaniotis B, Tselentis Y. Typing of sandflies from Greece and Cyprus by DNA polymorphism of 18S rRNA gene. Insect Mol Biol. 1999;8:179–184. doi: 10.1046/j.1365-2583.1999.820179.x. PubMed DOI

Blanc G, Caminopetros J. Enquiry into Oriental sore in Crete. Reflections suggested on the etiology and mode of dispersion of the disease. Ann Inst Pasteur. 1921;35:151–166.

Kasap OE, Dvorak V, Depaquit J, Alten B, Votypka J, Volf P. Phylogeography of the subgenus Transphlebotomus Artemiev with description of two new species, Phlebotomus anatolicus n. sp. and Phlebotomus killicki n. sp. Infect Genet Evol. 2015;34:467–479. doi: 10.1016/j.meegid.2015.05.025. PubMed DOI

Birt C. Phlebotomus fever in Malta and Crete. J R Army Med Corps. 1910;14:236–258.

Depaquit J, Ferté H, Léger N, Lefranc F, Alves-Pires C, Hanafi H, et al. ITS 2 sequences heterogeneity in Phlebotomus sergenti and Phlebotomus similis (Diptera, Psychodidae): possible consequences in their ability to transmit Leishmania tropica. Int J Parasitol. 2002;32:1123–1131. doi: 10.1016/S0020-7519(02)00088-7. PubMed DOI

Depaquit J, Léger N, Ferté H. Le statut taxinomique de Phlebotomus sergenti Parrot, 1917, vecteur de Leishmania tropica (Wright, 1903) et Phlebotomus similis Perfiliev, 1963 (Diptera-Psychodidae). Approches morphologique et morphométrique. Corollaires biogéographiques et épidémiologiques. Bull Soc Pathol Exot. 1998;91:346–352. PubMed

Killick-Kendrick R. Phlebotomine vectors of the leishmaniases: a review. Med Vet Entomol. 1990;4:1–24. doi: 10.1111/j.1365-2915.1990.tb00255.x. PubMed DOI

Depaquit J, Grandadam M, Fouque F, Andry PE, Peyrefitte C. Arthropod-borne viruses transmitted by phlebotomine sandflies in Europe: a review. Euro Surveill. 2010;15:19507. PubMed

Alwassouf S, Christodoulou V, Bichaud L, Ntais P, Mazeris A, Antoniou M, et al. Seroprevalence of sandfly-borne phleboviruses belonging to three serocomplexes (Sandfly fever Naples, Sandfly fever Sicilian and Salehabad) in dogs from Greece and Cyprus using neutralization test. PLoS Negl Trop Dis. 2016;10:e0005063. doi: 10.1371/journal.pntd.0005063. PubMed DOI PMC

Sadlova J, Dvorak V, Seblova V, Warburg A, Votypka J, Volf P. Sergentomyia schwetzi is not a competent vector for Leishmania donovani and other Leishmania species pathogenic to humans. Parasit Vectors. 2013;6:186. doi: 10.1186/1756-3305-6-186. PubMed DOI PMC

Maia C, Depaquit J. Can Sergentomyia (Diptera, Psychodidae) play a role in the transmission of mammal-infecting Leishmania? Parasite. 2016;23:55. doi: 10.1051/parasite/2016062. PubMed DOI PMC

Maia C, Parreira R, Cristóvão JM, Freitas FB, Afonso MO, Campino L. Molecular detection of Leishmania DNA and identification of blood meals in wild caught phlebotomine sand flies (Diptera: Psychodidae) from southern Portugal. Parasit Vectors. 2015;8:173. doi: 10.1186/s13071-015-0787-4. PubMed DOI PMC

Artemiev MM, Neronov VM. Distribution and ecology of sandflies of the old world: genus Phlebotomus. Moscow: USSR Academy of Sciences; 1984.

Esseghir S, Ready PD, Ben-Ismail R. Speciation of Phlebotomus sandflies of the subgenus Larroussius coincided with the late Miocene-Pliocene aridification of the Mediterranean subregion. Biol J Linn Soc. 2000;70:189–219. doi: 10.1111/j.1095-8312.2000.tb00207.x. DOI

Yssouf A, Almeras L, Raoult D, Parola P. Emerging tools for identification of arthropod vectors. Future Microbiol. 2016;11:549–566. doi: 10.2217/fmb.16.5. PubMed DOI

Depaquit J, Ferte H, Leger N, Killick-Kendrick R, Rioux JA, Killick-Kendrick M, et al. Molecular systematics of the phlebotomine sandflies of the subgenus Paraphlebotomus (Diptera, Psychodidae, Phlebotomus) based on ITS2 rDNA sequences. Hypotheses of dispersion and speciation. Insect Mol Biol. 2000;9:293–300. doi: 10.1046/j.1365-2583.2000.00179.x. PubMed DOI

Combining phylogeography and climate models to track the diversification and spread of Phlebotomus simici

Phlebotomine sand fly survey, blood meal source identification, and description of Sergentomyia imihra n. sp. in the central Sahara of Algeria

Phlebotomine sand flies (Diptera: Psychodidae) of the Maghreb region: A systematic review of distribution, morphology, and role in the transmission of the pathogens

Phlebotomine sand fly survey in the Republic of Moldova: species composition, distribution and host preferences

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