Entomological Surveillance in Former Malaria-endemic Areas of Southern Italy

Malaria still represents a potential public health issue in Italy, and the presence of former Anopheles vectors and cases imported annually merit continuous surveillance. In areas no longer endemic, the concurrent presence of gametocyte carriers and competent vectors makes re-emergence of local transmission possible, as recently reported in Greece. In October 2017, due to the occurrence of four suspected introduced malaria cases in the province of Taranto (Apulia region), entomological investigations were performed to verify the involvement of local anopheline species. In 2019–2020 entomological surveys were extended to other areas historically prone to malaria between the provinces of Taranto and Matera and the province of Foggia (Gargano Promontory). Resting mosquitoes were collected in animal shelters and human dwellings, larvae were sampled in natural and artificial breeding sites, and specimens were both morphologically and molecularly identified. A total of 2228 mosquitoes were collected, 54.3% of which were anophelines. In all the investigated areas, Anopheles labranchiae was the most widespread species, while Anopheles algeriensis was predominant at the Gargano sites, and Anopheles superpictus and Anopheles plumbeus were recorded in the province of Matera. Our findings showed a potentially high receptivity in the surveyed areas, where the abundance of the two former malaria vectors, An. labranchiae and An. superpictus, is related to environmental and climatic parameters and to anthropic activities.

Molecular Characterization of Mosquito Diversity in the Balearic Islands

Several outbreaks of mosquito-borne diseases have taken place in Europe in recent years. In Spain, both active and passive surveillance have demonstrated that dengue and West Nile viruses are currently circulating, and seven autochthonous dengue cases have been reported in the last 2 yr. The effectiveness of vector control programs largely depends on the accuracy of the taxonomic identification of the species. However, in Spain, identification almost completely relies on the use of morphological keys to characterize the mosquito fauna. This study investigates the congruence between molecular and morphological species boundaries in 13 Spanish mosquito taxa. The Cytochrome c oxidase subunit I (COI) gene region was sequenced from 60 adult specimens collected in Mallorca, plus several representatives from other Spanish regions for comparative purposes. Phylogenetic relationships were established using Bayesian and maximum-likelihood approaches. Using three species delimitation algorithms (ABGD, mPTP, and GMYC), we found strong evidence for cryptic speciation within Anopheles algeriensis Theobald, a widespread mosquito in the Mediterranean basin. We also delimited the Mallorcan rock pool mosquito Aedes mariae (Sergent & Sergent), from mainland European populations. Finally, we found difficulties in the use of wing characters in species keys to distinguish Culiseta annulata (Schrk) from Culiseta subochrea (Edwards). Given that these species are vectors of pathogens of medical relevance and have veterinary importance, their accurate taxonomic identification is essential in European vector surveillance programs.

Opening the Pandora Box: DNA-barcoding evidence limitations of morphology to identify Spanish mosquitoes

Cryptic speciation is frequent in the medically important mosquitoes. While most findings have been reported in tropical regions, it is an unexplored topic in countries where mosquito-borne diseases are not endemic, like Spain. The occurrence of recent outbreaks in Europe has increased the awareness on the native and invasive mosquito fauna present in the continent. Therefore, the central question of this study is whether the typological approach is sufficient to identify Spanish mosquitoes. To address this problem, we confronted the results of the morphological identification of 62 adult specimens collected from four different regions of Spain (La Rioja, Navarra, Castellón and the Island of Majorca) with the results obtained through DNA-barcoding. We conducted a comprehensive phylogenetic analysis of the COI gene region and compared this with the results of four species delimitation algorithms (ABGD initial partition, ABGD P=0.46%, bPTP and TCS). We report strong evidence for cryptic speciation in Anopheles algeriensis and Aedes vexans and reproductive isolation of the rock pool mosquito Aedes mariae. In addition, we report that the character present in the wings is not efficient to distinguish species Culiseta annulata from Culiseta subochrea, which distribution in the country may be different than previously described.