Arbovirus screening of mosquitoes collected in 2022 in Emilia-Romagna, Italy, with the implementation of a real-time PCR for the detection of Tahyna virus.

Several Arboviruses (Arthropod-borne virus) are a concrete health risk. While some arboviruses, such as the West Nile virus (WNV) and the Usutu virus (USUV) are actively surveyed, others are neglected, including the Tahyna virus (TAHV). In this work, we tested - searching for all the three viruses - 37,995 mosquitoes collected in 95 attractive traps, baited by carbon dioxide, distributed in the lowlands of Emilia-Romagna, northern Italy, between 19 July and 12 August 2022. Among the 668 pools obtained, WNV was detected in 45 pools of Culex (Cx.) pipiens and USUV was recorded in 24 pools of the same mosquito; ten of these Cx. pipiens pools tested positive for both WNV and USUV. Interestingly, we recorded a significant circulation of both WNV lineage 1 (WNV-L1) and lineage 2 (WNV-L2): WNV-L1 strains were detected in 40 pools, WNV-L2 strains in three pools and both lineages were detected in two pools. TAHV was detected in 8 different species of mosquitoes in a total of 37 pools: Aedes (Ae.) caspius (25), Ae. albopictus (5), Ae. vexans (3), Cx. pipiens (2), Ae. cinereus (1) and Anopheles maculipennis sl (1). The significant number of Ae. caspius-pools tested positive and the estimated viral load suggest that this mosquito is the principal vector in the surveyed area. The potential involvement of other mosquito species in the TAHV cycle could usefully be the subject of further experimental investigation. The results obtained demonstrate that, with adequate sampling effort, entomological surveillance is able to detect arboviruses circulating in a given area. Further efforts must be made to better characterise the TAHV cycle in the surveyed area and to define health risk linked to this virus.

Monitoring of Insecticide Resistance Mutations and Pathogen Circulation in Sand Flies from Emilia-Romagna, a Leishmaniasis Endemic Region of Northern Italy.

The continuously expanding distribution of sand flies, proven vectors of Leishmania and of several phleboviruses, is a growing public health issue in Europe. Especially in Italy, visceral leishmaniasis (VL) is occurring with increasing incidence northward, in previously non-endemic provinces. Around the globe, disease elimination efforts largely focus on sand fly vector insecticidal control, often leading to the development of resistance. In Emilia-Romagna (ER), northern Italy, insecticides are heavily applied for agricultural and mosquito control, but not specifically against sand flies. Here, we investigated the sand fly species composition in certain environmental settings in ER provinces and monitored the presence of pyrethroid resistance mutations and pathogen circulation. Phlebotomus perfiliewi, a dominant vector of Leishmania infantum, was detected almost exclusively in the region. No mutations in the voltage-gated sodium channel gene, e.g., knock-down resistance mutations I1011M, L1014F/S, V1016G, or F1020S, were recorded. Pathogen monitoring revealed that almost 40% of the tested sand fly pools were positive for Leishmania, while the presence of Toscana and Fermo phleboviruses was also observed in much lower frequencies (≤3% positive pools). Regular epidemiological and entomological monitoring, alongside resistance surveillance, is highly recommended to ensure the sustainability and efficiency of vector control interventions.

Evaluation of Alternative Methods for Testing the Bioefficacy of Household Ambient Insecticide Products Against Aedes albopictus.

Ambient insecticides are receiving increasing attention in many developed countries because of their value in reducing mosquito nuisance. As required by the European Union Biocidal Products Regulation 528/2012, these devices require appropriate testing of their efficacy, which is based on estimating the knockdown and mortality rates of free-flying (free) mosquitoes in a test room. However, evaluations using free mosquitoes present many complexities. The performances of 6 alternative methods with mosquitoes held in 2 different cage designs (steel wire and gauze/plastic) with and without an operating fan for air circulation were monitored in a test room through a closed-circuit television system and were compared with the currently recommended method using free mosquitoes. Results for caged mosquitoes without a fan showed a clearly delayed knockdown effect, whereas outcomes for caged mosquitoes with a fan recorded higher mortality at 24 h, compared to free mosquitoes. Among the 6 methods, cages made of gauze and plastic operating with fan wind speed at 2.5-2.8 m/sec was the only method without a significant difference in results for free mosquitoes, and therefore appears as the best alternative to assess knockdown by ambient insecticides accurately.

Active surveillance for avian influenza viruses in wild birds and backyard flocks in Northern Italy during 2004 to 2006.

Following the avian influenza epidemics that occurred in Italy between 1997 and 2003, the Italian Ministry of Health in collaboration with veterinary authorities promoted, funded and implemented a national surveillance programme. The main objectives of the surveillance effort were to identify avian influenza viruses circulating in wild birds and to investigate the role of backyard poultry flocks in the dynamics of infection in a densely populated poultry area. Over 2 years (2004 to 2006), 164 backyard flocks and 4083 wild birds (mainly migratory Anseriformes and Charadriiformes) were sampled in three regions in the North of Italy. Samples collected were screened by means of real-time reverse transcriptase-polymerase chain reaction and the positive samples were processed for attempted virus isolation in embryonated fowl's specific pathogen free eggs. At the end of the study period, 27 low-pathogenic avian influenza viruses had been isolated from backyard flocks and 49 strains obtained from wild birds. Of these, 26 belonged to the H5 or H7 subtype and were closely related to contemporary low-pathogenic strains of Eurasian lineage. The findings confirm that backyard free-range farming is at high risk for avian influenza virus introduction, and confirm the role of wild waterfowl in the introduction and perpetuation of low-pathogenic avian influenza viruses during the winter season in Southern Europe.