BACKGROUND: Phlebotomus (Larroussius) perniciosus (Diptera: Psychodidae) is the most common and predominant vector of Leishmania infantum in the Western Mediterranean region. Volatile organic compounds (VOCs) produced by vertebrates are important cues affecting the behaviour of blood-feeding insects. Generally, the identification of putative behaviourally active VOCs involves three distinct phases: extraction, chemical characterization and chemoreceptivity evaluation using electrophysiological techniques. Here, we present a simplified gas chromatography-mass spectrometry-electroantennographic detection (GC-MS-EAD) setup adapted for screening bioactive compounds in sand flies, in which the chemical identification and antennal responses are recorded simultaneously. METHODS: The method integrates: (i) a flow-splitter that balances the flow rate of the two outgoing streams, (ii) GC columns with different lengths and diameters in the two sections splitter-MS and splitter-EAD and (iii) an antennal signal amplifier. The GC-MS-EAD analysis was applied to headspace solid-phase microextraction (HS-SPME) extracts from a healthy dog, and antennal responses were recorded in female P. perniciosus sand flies. RESULTS: The canine VOC profile was predominantly composed of aldehydes, with hexanal and nonanal eliciting the strongest antennal responses in P. perniciosus. CONCLUSIONS: This simplified GC-MS-EAD system shows promise for broader application in the study of host-vector interactions. Its use across different host-vector pairs may enhance our understanding of these relationships and inform the development of strategies for integrated vector monitoring and control.

• Keywords
• Leishmania infantum, Electroantennography, HS-SPME, Leishmaniasis, Phlebotominae, Psychodidae, Semiochemicals,
• MeSH
• Insect Vectors physiology   chemistry   MeSH
• Solid Phase Microextraction MeSH
• Phlebotomus * physiology   chemistry   MeSH
• Gas Chromatography-Mass Spectrometry methods   MeSH
• Dogs MeSH
• Volatile Organic Compounds * analysis   MeSH
• Arthropod Antennae * physiology   MeSH
• Animals MeSH
• Check Tag
• Dogs MeSH
• Female MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Names of Substances
• Volatile Organic Compounds * MeSH

The olfactory response of insect vectors such as phlebotomine sand flies is a key facet for investigating their interactions with vertebrate hosts and associated vector-borne pathogens. Such studies are mainly performed by assessing the electrophysiological response and the olfactory behaviour of these arthropods towards volatile organic compounds (VOCs) produced by hosts. Nonetheless, few studies are available for species of the subgenera Lutzomyia and Nyssomyia in South America, leaving a void for Old World sand fly species of the genus Phlebotomus. In this study, we evaluated the olfactory responses of Phlebotomus perniciosus, one of the most important vectors of Leishmania infantum in the Old World. To test the P. perniciosus behavioural response to VOCs, 28 compounds isolated from humans and dogs were assessed using electrophysiological (i.e., electroantennogram, EAG) and behavioural assays (i.e., Y-tube olfactometer). In the EAG trials, 14 compounds (i.e., acetic acid, nonanoic acid, 2-propanol, 2-butanol, pentanal, hexanal, nonanal, trans-2-nonenal, decanal, myrcene, p-cymene, verbenone, 2-ethyl-1-hexanol, and acetonitrile) elicited high antennal responses (i.e., ≥ 0.30 mV) in female sand flies, being those VOCs selected for the behavioural assays. From the 14 compounds tested in the Y-tube olfactometer, nonanal was significantly attractive for P. perniciosus females, whereas myrcene and p-cymene were significantly repellents (p < 0.05). The attraction indexes varied from 0.53 for nonanal (i.e., most attractive) to -0.47 to p-cymene (i.e., most repellent). Overall, our results shed light on the role of olfactory cues routing host seeking behaviour in P. perniciosus, with implications to develop sustainable sand fly monitoring as well as control in leishmaniasis endemic areas.

• MeSH
• Behavior, Animal drug effects   MeSH
• Insect Vectors physiology   drug effects   MeSH
• Leishmania infantum drug effects   physiology   MeSH
• Humans MeSH
• Phlebotomus * physiology   drug effects   MeSH
• Dogs MeSH
• Volatile Organic Compounds * pharmacology   chemistry   analysis   MeSH
• Animals MeSH
• Check Tag
• Humans MeSH
• Male MeSH
• Dogs MeSH
• Female MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Names of Substances
• Volatile Organic Compounds * MeSH