1. Intraspecific geographical variability of Phlebotomus perniciosus assessed by MALDI-TOF MS protein profilingVít Dvořák, Carla Maia, Rafael Marmé, José Risueño, Pedro Pérez-Cutillas, Eduardo Berriatua, Julie Sevila, Nalia Mekarnia, Jorian Prudhomme, Fano José Randriananambinintsoa, Vladimir Ivović, Katja Adam, 2026, original scientific article Abstract: Background Matrix‑assisted laser desorption/ionization time‑of‑flight mass spectrometry (MALDI‑TOF MS) pro‑ tein profiling has emerged over the last decade as a method of choice for species identification of many medically important arthropods. However, the influence of intraspecific variability on the performance of this popular technique has seldom been tested. This study provides the first standardized comparison of different geographical popula‑ tions of Phlebotomus perniciosus, a vector of Leishmania infantum and Toscana virus in the western Mediterranean, by MALDI‑TOF MS protein profiling. Methods Phlebotomus perniciosus males were collected in five countries (Portugal, Spain, France, Italy, Croa‑ tia) that represent most of its distribution in Europe. All samples were trapped, stored and processed according to a highly standardized protocol to avoid effects other than geographical origin on their protein spectra acquired by MALDI‑TOF MS protein profiling. The obtained protein spectra were compared with laboratory‑reared specimens of Ph. perniciosus. Results Twenty‑two analysed specimens from five geographical populations provided protein spectra that were highly similar, species‑specific and clustering according to their quality. No grouping according to geographical origin was observed, and the protein spectra of field‑collected specimens showed similar composition and complexity to spectra from Ph. perniciosus laboratory colony‑reared in captivity for several decades. Conclusions Our findings demonstrate that in samples of a same sex, with the same collection method and storage time, MALDI‑TOF MS protein profiling does not reflect the geographical origin of analysed specimens, confirming the value of this technique for high‑fidelity and reproducible species identification of sand flies regardless of their geographical origin.
Keywords: intraspecific geographical variability, Phlebotomus perniciosus, MALDI-TOF
Published in RUP: 29.06.2026; Views: 390; Downloads: 9
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2. Processing and mounting phlebotomine sand flies : a consensus guidelineFano José Randrianambinintsoa, Laure Augendre, Jorian Prudhomme, Jean-Philippe Martinet, Mathieu Loyer, Nalia Mekarnia, Hocine Kerkoub, Farzana Khan Perveen, Antoine Huguenin, Vladimir Ivović, 2026, review article Abstract: This article provides a comprehensive guide for the processing and mounting of phlebotomine sand fly specimens, which is crucial for species identification and pathogen detection and isolation. It discusses a range of techniques suitable for both field and laboratory settings. The guide includes detailed instructions on sand fly collec- tion, handling, covering, and euthanasia (recommending dry freezing or CO2 over chemicals) as well as conservation strategies, such as cold storage and preservation in ethanol. The quality of preparation of certain anatomical structures (genital organs, head and wings) is essential for their proper microscopic observation and is described in this work. The article also presents detailed sample processing, including the clearing process with agents such as potassium hydrox- ide then Marc-André solution. The mounting process compares different media, emphasizing their optical properties and preservation potential. Hoyer fluid (also known as chloral gum) is recommended for quick observation, particularly for spermathecae, due to its clarity, although it is not suitable for long-term storage. Other media discussed include polyvinyl alcohol, EuparalÒ (for limited water tolerance), and Canada balsam (a hydrocarbon-soluble medium), with the latter two offering long-term preservation capabilities. Innovative molecular biology approaches such as DNA sequencing and MALDI-ToF, which require particular attention to sample processing, are also addressed. Furthermore, short video clips illustrating various mounting techniques as well as translations in many different languages are provided, allowing the guideline to reach the diverse needs and expectations of the global scientific community.
Keywords: mounting, Phlebotomine sand fly, Hoyer fluid, Marc-André solution, Chloral gum, Polyvinyl alcohol, Euparal®, Canada balsam, Leishmania isolation, field conditions, culture, dissection, molecular biology, MALDI-ToF, type-specimens
Published in RUP: 03.04.2026; Views: 695; Downloads: 14
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