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Introduction: Culicoides Latreille biting midges are vectors of high concern as they can transmit serious veterinary diseases such as bluetongue virus or epizootic haemorrhagic disease virus, among others. Little is known about these vectors in Galicia, so a comprehensive literature review and an intensive monitoring were carried out in the region. Material and methods: The Autonomous Community of Galicia was sampled through three different vector surveillance projects between 2004 and 2023. A total of 239 sampling points were deployed alongside the Galician territory. In addition, a literature review of Culicoides in Galicia related content was made by consulting several digital repositories. Results: A total of 33 species of Culicoides belonging to 8 subgenera were identified. Among them, 15 are considered or suspected to be potential vectors of several pathogens of medical and/or veterinary interest. In addition, 20 of them are reported for the first time in the region. Updated distribution maps of the Culicoides biting midges of Galicia were provided, including several notes regarding their ecology and relevance for both public health and animal welfare. Conclusions: The present work is one of the most complete works made at regional level in Spain to date. As Galicia's economy relies heavily on livestock farming, this work will provide a solid baseline in order to develop new research lines in the future regarding prevention to vector‐borne diseases. [ABSTRACT FROM AUTHOR]

Sexual differences in pathogen prevalence in wildlife often arise from varying susceptibility influenced by factors such as sex hormones and exposure to pathogens. In the case of vector‐borne pathogens, host selection by insect vectors determines the exposure of hosts to infections, largely affecting the transmission of these infectious diseases. We identify the blood‐feeding patterns of insect vectors in Blue Tit (Cyanistes caeruleus) nestlings in a 3‐year study. Blood from both nestlings and insect vectors (Culicoides spp. and Simuliidae) captured inside nest‐boxes were used to molecularly determine the sex of the host. We then compared the sex‐ratios of the nestlings that had been bitten and those of the complete brood in each nest. We found that males were bitten more frequently than females in 2021, when males weighed less in comparison to other years. Additionally, we molecularly identified bitten nestlings individually by genotyping the DNA of blood obtained from both, the vector's abdomen and nestlings of each brood in 2022. Nestlings more frequently bitten by vectors were males, weighed less and were closest to the nest entrance. To our knowledge this is the first study identifying the nestling selection by insect vectors in bird nests under natural conditions. These results contribute to understanding the mechanisms of host selection by insect vectors, shedding light on pathogen transmission and offering insights into the observed sex‐biased infections in wildlife populations. [ABSTRACT FROM AUTHOR]

Culicoides biting midge species (Diptera: Ceratopogonidae) of the Obsoletus Group and the Pulicaris Complex are considered the major vectors of bluetongue and Schmallenberg viruses in Europe. Overwintering strategies of these arboviruses are controversially discussed, with the ongoing activity of vector species and a non‐disrupted transmission cycle during winter being a plausible explanation. Although data on Culicoides winter activity are relatively scant, a seasonal vector‐free period (SVFP), during which adult Culicoides are not or hardly active, is questionable. To determine winter activity and define SVFPs according to the EU Commission Regulation No 1266/2007, adult Culicoides were trapped weekly with UV‐light traps from October to April 2019/2020 and 2020/2021 inside and outside stables on 16 farms throughout Germany. Temperature measurements were taken regularly at each trapping site since the temperature is a known driver of biting midge activity. In 960 indoor and outdoor catches, 32,377 Culicoides were trapped, with 90.9% of them belonging to the Obsoletus Group, 6.1% to the Pulicaris Complex and 3.0% to ‘other Culicoides’ according to morphological identification. The majority (61.3%) of Culicoides were trapped indoors, with substantial numbers of specimens collected from October to December, in March and in April, and only a few or no specimens in January and February. Obsoletus Group biting midges were active indoors for almost the entire winter. Outdoors, Culicoides numbers decreased from October to December, few or no specimens were caught from January to March, and high numbers were captured in April. Of the collected Culicoides, 2028 were blood‐fed, of which 94.6% were trapped in the stables. The indoor SVFP, although calculated for blood‐fed instead of parous females, lasted for almost 4 months (late November until mid‐March) in winter 2019/2020 and 2 months (January and February) in winter 2020/2021. The outdoor SVFPs covered almost the entire study period in both winters, with slight differences between the onsets and the ends. The Culicoides activity significantly depended on temperature. Specimens of the Obsoletus Group were caught at an average temperature of 7.4°C (minimum 0.3°C) and of the Pulicaris Complex at an average temperature of 10.3°C (minimum 1.2°C). These temperatures were reached inside the stables over more extended periods than outside. The average indoor temperatures were 1.2 K higher than the average outdoor temperatures, although absolute temperature differences of up to 9.0 K were recorded. Based on Culicoides activity, the results of the present study indicate an almost continuous potential for virus transmission in winter within livestock houses. [ABSTRACT FROM AUTHOR]

Background: Culicoides midges have been well-studied in Spain, particularly over the last 20 years, mainly because of their role as vectors of arboviral diseases that affect livestock. Most studies on Culicoides are conducted using suction light traps in farmed environments, but studies employing alternative trapping techniques or focusing on natural habitats are scarce. Methods: In the present study, we analyze Culicoides captured in 2023 at 476 sites in western Andalusia (southern Spain) using carbon dioxide-baited Biogents (BG)-sentinel traps across different ecosystems. Results: We collected 3,084 Culicoides midges (3060 females and 24 males) belonging to 23 species, including the new species Culicoides grandifovea sp. nov. and the first record of Culicoides pseudolangeroni for Europe. Both species were described with morphological and molecular methods and detailed data on spatial distribution was also recorded. The new species showed close phylogenetic relations with sequences from an unidentified Culicoides from Morocco (92.6% similarity) and with Culicoides kurensis. Culicoides imicola was the most abundant species (17.4%), followed by Culicoides grandifovea sp. nov. (14.6%) and Culicoides kurensis (11.9%). Interestingly, Culicoides montanus was the only species of the obsoletus and pulicaris species complexes captured, representing the first record of this species in southern Spain. A total of 53 valid Culicoides species have been reported in the area, with 48 already reported in literature records and 5 more added in the present study. Information on the flight period for the most common Culicoides species is also provided. Conclusions: To the best of our knowledge, our study represents the most comprehensive effort ever done on nonfarmland habitats using carbon-dioxide baited suction traps for collecting Culicoides. Our data suggests that using carbon dioxide traps offers a completely different perspective on Culicoides communities compared with routinely used light traps, including the discovery of previously unrecorded species. [ABSTRACT FROM AUTHOR]

Abstract Background Culicoides midges have been well-studied in Spain, particularly over the last 20 years, mainly because of their role as vectors of arboviral diseases that affect livestock. Most studies on Culicoides are conducted using suction light traps in farmed environments, but studies employing alternative trapping techniques or focusing on natural habitats are scarce. Methods In the present study, we analyze Culicoides captured in 2023 at 476 sites in western Andalusia (southern Spain) using carbon dioxide-baited Biogents (BG)-sentinel traps across different ecosystems. Results We collected 3,084 Culicoides midges (3060 females and 24 males) belonging to 23 species, including the new species Culicoides grandifovea sp. nov. and the first record of Culicoides pseudolangeroni for Europe. Both species were described with morphological and molecular methods and detailed data on spatial distribution was also recorded. The new species showed close phylogenetic relations with sequences from an unidentified Culicoides from Morocco (92.6% similarity) and with Culicoides kurensis. Culicoides imicola was the most abundant species (17.4%), followed by Culicoides grandifovea sp. nov. (14.6%) and Culicoides kurensis (11.9%). Interestingly, Culicoides montanus was the only species of the obsoletus and pulicaris species complexes captured, representing the first record of this species in southern Spain. A total of 53 valid Culicoides species have been reported in the area, with 48 already reported in literature records and 5 more added in the present study. Information on the flight period for the most common Culicoides species is also provided. Conclusions To the best of our knowledge, our study represents the most comprehensive effort ever done on nonfarmland habitats using carbon-dioxide baited suction traps for collecting Culicoides. Our data suggests that using carbon dioxide traps offers a completely different perspective on Culicoides communities compared with routinely used light traps, including the discovery of previously unrecorded species. Graphical Abstract

Abstract Schmallenberg virus (SBV) and bluetongue virus (BTV) are both transmitted by Culicoides biting midges and infect predominantly ruminants. To investigate the extent of virus spread in the 2022 and 2023 vector seasons, we serologically tested wild ruminants from western Germany. While antibodies against BTV were not detected in any animal, regardless of age or sampling time, numerous wild ruminants tested positive for antibodies to SBV. In 2022, a low seroprevalence of 4.92% was measured. In sharp contrast, 40.15% of the animals tested positive in 2023. Of the young animals, about 31.82% were seropositive, clearly indicating large-scale SBV circulation in summer and autumn 2023.

Because the vertical distribution and diversity of blood-sucking flies are poorly known, we determined the diversity, structure, and composition of culicids between vertical vegetation strata. We evaluated the influence of microclimatic variables during different times of the day over a year. We used eight CDC traps baited with CO2 at a height of 1.5 m and 12- 15 m. We conducted rank-abundance curves, similarity analysis (ANOSIM and SIMPER), and multivariate clustering with incidence and abundance data. We used GAM models to analyze the influence of strata (understory vs canopy), humidity, and temperature on insect richness and abundance. During the day, the difference between strata was mainly due to higher abundance of Wyeomyia arthrostigma and Wyeomyia ca. adelpha in the understory. During the night, the differences were mainly due to higher abundance of Culex stigmatosoma, Culex salinarius, and Aedes allotecnon in the canopy, and Wyeomyia arthrostigma in the understory. Seasonality played a role in the similarity between the strata. Diversity during the day was positively related to humidity and temperature, and nocturnal diversity increased with temperature but decreased with higher humidity. The effects of environmental factors on the spatiotemporal distribution of fly species are essential for epidemiological surveillance. [ABSTRACT FROM AUTHOR]

The collection of insects of medical importance from the Instituto Nacional de Salud, INS (Bogotá, Colombia: https://www.ins.gov.co/Paginas/Inicio.aspx), was started in 1934 with the aim of being an institutional and national repository of the biodiversity of insects involved in vector-borne diseases of importance in public health. Today, the entomological collection includes more than 7,500 specimens. The ceratopogonid insects are one group of Diptera that are represented in this collection. Within the Ceratopogonidae, the genus Culicoides Latreille, 1809 is relevant in public health because of the nuisance caused by their bites when they are presented in great abundance and because of their role as vectors of several agents (virus, protozoa and nematodes) that cause diseases to humans and to animals (Mellor et al. 2000, Mullen 2002). An overview of the Ceratopogonidae, represented in this collection, is presented here. A total of 801 individuals, mainly adults of the genus Culicoides (90%) are represented. The collection is the result of the effort of several researchers of the Group of Entomology at INS. These researchers collected ceratopogonids when they went to different transmission scenarios of vector-borne diseases in Colombia, with the purpose of making entomological characterisations including the processing, assembly and identification of the specimens in the laboratory. New information: New information about the geographical distribution of 39 species of the genus Culicoides in Colombia. All data have been uploaded to GBIF and are publicly available there. [ABSTRACT FROM AUTHOR]

Abstract Introduction Culicoides Latreille biting midges are vectors of high concern as they can transmit serious veterinary diseases such as bluetongue virus or epizootic haemorrhagic disease virus, among others. Little is known about these vectors in Galicia, so a comprehensive literature review and an intensive monitoring were carried out in the region. Material and methods The Autonomous Community of Galicia was sampled through three different vector surveillance projects between 2004 and 2023. A total of 239 sampling points were deployed alongside the Galician territory. In addition, a literature review of Culicoides in Galicia related content was made by consulting several digital repositories. Results A total of 33 species of Culicoides belonging to 8 subgenera were identified. Among them, 15 are considered or suspected to be potential vectors of several pathogens of medical and/or veterinary interest. In addition, 20 of them are reported for the first time in the region. Updated distribution maps of the Culicoides biting midges of Galicia were provided, including several notes regarding their ecology and relevance for both public health and animal welfare. Conclusions The present work is one of the most complete works made at regional level in Spain to date. As Galicia's economy relies heavily on livestock farming, this work will provide a solid baseline in order to develop new research lines in the future regarding prevention to vector‐borne diseases.

Schmallenberg virus (SBV) and bluetongue virus (BTV) are both transmitted by Culicoides biting midges and infect predominantly ruminants. To investigate the extent of virus spread in the 2022 and 2023 vector seasons, we serologically tested wild ruminants from western Germany. While antibodies against BTV were not detected in any animal, regardless of age or sampling time, numerous wild ruminants tested positive for antibodies to SBV. In 2022, a low seroprevalence of 4.92% was measured. In sharp contrast, 40.15% of the animals tested positive in 2023. Of the young animals, about 31.82% were seropositive, clearly indicating large-scale SBV circulation in summer and autumn 2023. [ABSTRACT FROM AUTHOR]

Two new species of biting midges are described: Forcipomyia (Lepidohelea) buccina sp. n. from Lepchakha (Queen of Dooars), the northern part of West Bengal at the Indo-Bhutan border and Forcipomyia (Lepidohelea) biharinathensis sp. n. from Biharinath Hill, highest peak of Bankura district of West Bengal and a remnant of Chota Nagpur Plateau. The discovery of these two new species elevates the total number of species in India to 80. A key to the Oriental species of the subgenus Lepidohelea Kieffer is also designed. [ABSTRACT FROM AUTHOR]

ABSTRACT An updated list of the biting midges recorded from the 26 Brazilian states and the Federal District is provided based on the data available in the “Taxonomic Catalog of the Brazilian Fauna” (Portuguese CTFB). The Brazilian Ceratopogonidae fauna is represented by 529 known species, corresponding to 40% of the Neotropical fauna. A table showing the number of species of Ceratopogonidae genera in the Neotropical region, Brazil, and Brazilian Amazon is included. In accordance with the family knowledge worldwide, the subfamily Ceratopogoninae is the best represented in the number of species due to the study of Culicoides, which has sanitary importance. The analysis of the diversity in each region and in the Brazilian states indicates the need for further studies in the five Brazilian geopolitical regions, especially in the Midwest and Northeast, which are home to the natural biomes of Cerrado, Pantanal, and Caatinga.

Simple Summary: Vesicular stomatitis virus (VSV) infects cows, horses, and pigs, economically impacting livestock producers due to animal production losses, quarantines, and animal movement/trade restrictions. Typically, VSV is transmitted from animal to animal by direct contact, but it is also transmitted by insects such as Culicoides biting midges. These tiny flies can ingest virus particles when blood feeding on an infected animal, multiply them inside their bodies, and then transmit them to other animals the next time they feed. In addition, midges are also able to pass the virus from one to another with extremely high efficiency when they mate, even though they carry very little virus in their bodies. Through this mechanism, VSV may be maintained and overwinter in midges and appear again in livestock the next summer once the insects start feeding on blood again. Our research shows that one reason midges can transmit VSV to other midges so efficiently is because viruses that come from insect cells have an increased ability to infect more insect cells. This helps explain the midge-to-midge infection efficiency and highlights the importance of Culicoides midges in VSV maintenance and transmission. Vesicular stomatitis virus (VSV) is an arthropod-borne virus affecting livestock. In the United States, sporadic outbreaks result in significant economic losses. During epizootics, Culicoides biting midges are biological vectors and key to the geographic expansion of outbreaks. Additionally, Culicoides may play a role in VSV overwintering because females and males are capable of highly efficient venereal transmission, despite their relatively low virus titers. We hypothesized that VSV propagated within a midge has increased fitness for subsequent midge infections. To evaluate the potential host-specific fitness increase, we propagated three viral isolates of VSV in porcine skin fibroblasts and Culicoides cell lines. We then evaluated the viral infection dynamics of the different cell-source groups in Culicoides sonorensis. Our results indicate that both mammalian- and insect-derived VSV replicate well in midges inoculated via intrathoracic injection, thereby bypassing the midgut barriers. However, when the virus was required to infect and escape the midgut barrier to disseminate after oral acquisition, the insect-derived viruses had significantly higher titers, infection, and dissemination rates than mammalian-derived viruses. Our research suggests that VSV replication in Culicoides cells increases viral fitness, facilitating midge-to-midge transmission and subsequent replication, and further highlights the significance of Culicoides midges in VSV maintenance and transmission dynamics. [ABSTRACT FROM AUTHOR]

The results of long-term studies of biting midges (Diptera, Ceratopogonidae) in macrozoobenthos communities of five saline rivers of the hyperhaline Lake Elton basin are presented. The fauna of Ceratopogonidae includes five halophilic and euryhaline species, among which representatives of the tribe Sphaeromiini predominate. The taxonomic composition of the family differs in rivers with different levels of salinity. Species Sphaeromias miricornis and Culicoides riethi are widely distributed in the mesohaline Khara, Lantsug, and Bolshaya Samoroda rivers, and Palpomyia schmidti is widely distributed in the polyhaline Chernavka and Solyanka rivers. Two generations of S. miricornis and three generations of P. schmidti are recorded. The emergence of the first generation of P. schmidti occurs in mid-May; S. miricornis emerges in late May to early June. There is an increase in the mean annual abundance and biomass of biting midges in polyhaline rivers when compared with mesohaline rivers. [ABSTRACT FROM AUTHOR]

Abstract Background After several years without bluetongue disease, a ruminant illness caused by Culicoides-borne bluetongue virus (BTV), two new autochthonous cases were reported in 2018 in Germany. By contrast, Schmallenberg virus (SBV), another Culicoides-borne virus pathogenic to ruminants, has continuously circulated in Germany since its first emergence in 2011. The disease outbreaks have triggered numerous studies on the biology of the Culicoides vectors, but many ecological details are still obscure. Methods Culicoides biting midge species were collected with UV-light traps on 10 farms in Germany, with one trap inside and one trap outside barns on each of the farms. Traps were run once a week for 24 h from January to December 2019. Collected biting midges were morphologically identified, counted and statistically evaluated, with a focus on the Obsoletus Group and the Pulicaris Complex of the ceratopogonid genus Culicoides, which are believed to contain the major virus vectors. Temperature and relative humidity recorded at each trap were linked to the quantity of caught Culicoides. Correlations between relative Culicoides abundance and presence of livestock or type of husbandry were also investigated. Results A total of 38,886 Culicoides biting midges were trapped, with most of them belonging to the Obsoletus Group (51.0%) and the Pulicaris Complex (38.8%). The majority of captured specimens were collected in traps inside the barns. Obsoletus Group individuals were caught from late January to the last week of December while Pulicaris Complex individuals were captured from the end of March to early December. The lowest average temperatures at which members of the two groups were collected were 10.7 °C and 12.8 °C, respectively. While temperature had a statistically significant effect on the activity of both the Obsoletus Group and the Pulicaris Complex, relative humidity only significantly affected the activity of the latter. The presence of livestock significantly influenced the number of captured Obsoletus Group, but not of Pulicaris Complex specimens. Inside the barns, no statistical difference was found between numbers of caught Obsoletus Group and Pulicaris Complex specimens in livestock holdings with deep litter and manure scraper or slatted floor husbandry systems. Conclusions The almost year-round presence of Obsoletus Group biting midges and the demonstrated high relative abundance of other potential Culicoides vector species inside barns suggest a high risk of indoor virus transmission to ruminants should BTV or SBV circulate locally. Appropriate structural, organisational and vector control measures to reduce biting midge exposure should be implemented. Graphical Abstract

A new fossil ceratopogonid genus and species from Lower Cretaceous Lebanese amber, Baskintoconops maaloufi Pielowska-Ceranowska gen. et sp. nov., is described and illustrated. The studied material originates from a newly discovered amber site in the Lebanese village Baskinta at a locality dubbed Qanat Bakish. The described genus is typified by its wing venation pattern combining characters of genera Fossileptoconops and Jordanoconops belonging to the subfamily Leptoconopinae.

The prevalence of autochthonous leishmaniasis in Thailand is increasing but the natural vectors that are responsible for transmission remain unknown. Experimental in vivo infections in Culicoides spp. with Leishmania (Mundinia) martiniquensis and Leishmania (Mundinia) orientalis, the major causative pathogens in Thailand, have demonstrated that biting midges can act as competent vectors. Therefore, the isolation and detection of Leishmania and other trypanosomatids were performed in biting midges collected at a field site in an endemic area of leishmaniasis in Tha Ruea and a mixed farm of chickens, goats, and cattle in Khuan Phang, Nakhon Si Thammarat province, southern Thailand. Results showed that Culicoides peregrinus was the abundant species (>84%) found in both locations and only cow blood DNA was detected in engorged females. Microscopic examination revealed various forms of Leishmania promastigotes in the foregut of several C. peregrinus in the absence of bloodmeal remnants, indicating established infections. Molecular identification using ITS1 and 3'UTR HSP70 type I markers showed that the Leishmania parasites found in the midges were L. martiniquensis. The infection rate of L. martiniquensis in the collected flies was 2% in Tha Ruea and 6% in Khuan Phang, but no L. orientalis DNA or parasites were found. Additionally, organisms from two different clades of Crithidia, both possibly new species, were identified using SSU rRNA and gGAPDH genes. Choanomastigotes and promastigotes of both Crithidia spp. were observed in the hindgut of the dissected C. peregrinus. Interestingly, midges infected with both L. martiniquensis and Crithidia were found. Moreover, four strains of Crithidia from one of the clades were successfully isolated into culture. These parasites could grow at 37°C in the culture and infect BALB/c mice macrophages but no multiplication was observed, suggesting they are thermotolerant monoxenous trypanosomatids similar to Cr. thermophila. These findings provide the first evidence of natural infection of L. martiniquensis in C. peregrinus supporting it as a potential vector of L. martiniquensis. [ABSTRACT FROM AUTHOR]

Background: After several years without bluetongue disease, a ruminant illness caused by Culicoides-borne bluetongue virus (BTV), two new autochthonous cases were reported in 2018 in Germany. By contrast, Schmallenberg virus (SBV), another Culicoides-borne virus pathogenic to ruminants, has continuously circulated in Germany since its first emergence in 2011. The disease outbreaks have triggered numerous studies on the biology of the Culicoides vectors, but many ecological details are still obscure. Methods: Culicoides biting midge species were collected with UV-light traps on 10 farms in Germany, with one trap inside and one trap outside barns on each of the farms. Traps were run once a week for 24 h from January to December 2019. Collected biting midges were morphologically identified, counted and statistically evaluated, with a focus on the Obsoletus Group and the Pulicaris Complex of the ceratopogonid genus Culicoides, which are believed to contain the major virus vectors. Temperature and relative humidity recorded at each trap were linked to the quantity of caught Culicoides. Correlations between relative Culicoides abundance and presence of livestock or type of husbandry were also investigated. Results: A total of 38,886 Culicoides biting midges were trapped, with most of them belonging to the Obsoletus Group (51.0%) and the Pulicaris Complex (38.8%). The majority of captured specimens were collected in traps inside the barns. Obsoletus Group individuals were caught from late January to the last week of December while Pulicaris Complex individuals were captured from the end of March to early December. The lowest average temperatures at which members of the two groups were collected were 10.7 °C and 12.8 °C, respectively. While temperature had a statistically significant effect on the activity of both the Obsoletus Group and the Pulicaris Complex, relative humidity only significantly affected the activity of the latter. The presence of livestock significantly influenced the number of captured Obsoletus Group, but not of Pulicaris Complex specimens. Inside the barns, no statistical difference was found between numbers of caught Obsoletus Group and Pulicaris Complex specimens in livestock holdings with deep litter and manure scraper or slatted floor husbandry systems. Conclusions: The almost year-round presence of Obsoletus Group biting midges and the demonstrated high relative abundance of other potential Culicoides vector species inside barns suggest a high risk of indoor virus transmission to ruminants should BTV or SBV circulate locally. Appropriate structural, organisational and vector control measures to reduce biting midge exposure should be implemented. [ABSTRACT FROM AUTHOR]

Dasyhelea bifida Zilahi-Sebess is fully redescribed, photographed and drawn from adults collected in Mexico City by Alfonso Dampf in 1944. This species is compared with the American congeners D. crassiseta Borkent and Forster, D. furva Remm and D. pollex Borkent and Forster. This is a new record of this species from Mexico. [ABSTRACT FROM AUTHOR]

Simple Summary: Culicoides sonorensis midges are able to transmit pathogenic viruses to livestock, resulting in significant economic losses. Virus transmission is linked to blood feeding, where female C. sonorensis midges require blood to produce eggs. Due to the relationship between egg development, blood feeding, and pathogen transmission, there is a need to understand midge physiology to control virus transmission. One aspect of midge physiology that has yet to be explored is microRNAs (miRNAs), which display different expression patterns related to egg development and pathogen infection. miRNAs degrade messenger RNAs in a sequence-specific manner which stops associated protein production. To determine the Culicoides miRNA catalog, we sequenced small RNA molecules (small RNA-Seq) and used bioinformatic analyses (miRDeep2) of whole female midges and digestive tissues before and after blood feeding. Our analyses characterized 76 miRNAs within C. sonorensis. Based on our findings, we suggest an interesting evolutionary relationship between miRNA expression and blood meal requirements across blood-feeding insects. We also identified miRNAs with expression patterns regulated by blood meal ingestion and/or tissue. Lastly, we identified potential miRNAs regulated by virus infection. Overall, our data provide a foundation for future studies to determine the mechanisms of gene regulation associated with midge physiology and virus transmission. Culicoides sonorensis midges vector multiple livestock arboviruses, resulting in significant economic losses worldwide. Due to the tight association between virus transmission, blood feeding, and egg development, understanding midge physiology is paramount to limiting pathogen transmission. Previous studies have demonstrated the importance of small non-coding RNAs (ncRNAs), specifically microRNAs (miRNAs), in multiple aspects of vector physiology. These small ncRNAs regulate gene expression at the post-transcriptional level and display differential expression during pathogen infection. Due to the lack of annotated miRNAs in the biting midge and associated expression profiles, we used small RNA-Seq and miRDeep2 analyses to determine the Culicoides miRNAs in whole females and midgut tissues in response to blood feeding. Our analyses revealed 76 miRNAs within C. sonorensis composed of 73 orthologous and three candidate novel miRNAs, as well as conserved miRNA clusters. miRNA conservation suggests an interesting evolutionary relationship between miRNA expression and hematophagy in the infraorder Culicomorpha. We also identified multiple blood meal-regulated and tissue-enriched miRNAs. Lastly, we further identified miRNAs with expression patterns potentially associated with virus infection by probing publicly available datasets. Together, our data provide a foundation for future ncRNA work to untangle the dynamics of gene regulation associated with midge physiology. [ABSTRACT FROM AUTHOR]

An updated catalogue of Culicoides of Mexico is presented. It includes 86 species with their regional distribution and corresponding record references, known immature stages and associated pathogens. In addition, a taxonomic key for subgenera and species groups for Mexico is presented and an index of species by state is included. [ABSTRACT FROM AUTHOR]

The new species Atrichopogon (Atrichopogon) tolfensis n. sp. from Tolfa Mountains (Italy, Latium, and Rome) is described and illustrated. Both sexes of the species were collected using cantharidin-baited traps from June 2020 to January 2021. The new species is an atypical member of the genus Atrichopogon.https://urn:lsid:zoobank.org:pub:3A012331-57D7-4532-B3F9-FDEE7C988FFD

Abstract Background Mitochondrial genomes are the most sequenced genomes after bacterial and fungal genomic DNA. However, little information on mitogenomes is available for multiple metazoan taxa, such as Culicoides, a globally distributed, megadiverse genus containing 1,347 species. Aim Generating novel mitogenomic information from single Culicoides sonorensis and C. biguttatus specimens, comparing available mitogenome mapping and de novo assembly tools, and identifying the best performing strategy and tools for Culicoides species. Results We present two novel and fully annotated mitochondrial haplotypes for two Culicoides species, C. sonorensis and C. biguttatus. We also annotated or re-annotated the only available reference mitogenome for C. sonorensis and C. arakawae. All species present a high similarity in mitogenome organization. The general gene arrangement for all Culicoides species was identical to the ancestral insect mitochondrial genome. Only short spacers were found in C. sonorensis (up to 30 bp), contrary to C. biguttatus (up to 114 bp). The mitochondrial genes ATP8, NAD2, NAD6, and LSU rRNA exhibited the highest nucleotide diversity and pairwise interspecific p genetic distance, suggesting that these genes might be suitable and complementary molecular barcodes for Culicoides identification in addition to the commonly utilized COI gene. We observed performance differences between the compared mitogenome generation strategies. The mapping strategy outperformed the de novo assembly strategy, but mapping results were partially biased in the absence of species-specific reference mitogenome. Among the utilized tools, BWA performed best for C. sonorensis while SPAdes, MEGAHIT, and MitoZ were among the best for C. biguttatus. The best-performing mitogenome annotator was MITOS2. Additionally, we were able to recover exogenous mitochondrial DNA from Bos taurus (biting midges host) from a C. biguttatus blood meal sample. Conclusions Two novel annotated mitogenome haplotypes for C. sonorensis and C. biguttatus using High-Throughput Sequencing are presented. Current results are useful as the baseline for mitogenome reconstruction of the remaining Culicoides species from single specimens to HTS and genome annotation. Mapping to a species-specific reference mitogenome generated better results for Culicoides mitochondrial genome reconstruction than de novo assembly, while de novo assembly resulted better in the absence of a closely related reference mitogenome. These results have direct implications for molecular-based identification of these vectors of human and zoonotic diseases, setting the basis for using the whole mitochondrial genome as a marker in Culicoides identification.

A newly described species, Dasyhelea mesophylla Dominiak and Borkent, from Costa Rica is the first species in this large and diverse family known to be a leaf-miner. The species is described as third and fourth instar larvae, pupa, and male and female adults. The biology of larvae mining the floating leaves of Salvinia minima Baker is also described. The pupa has a distinctive, sharply pointed respiratory organ, shared with some other species of Dasyhelea, which pierces the surface of the leaf to breathe. The species belongs in the grisea group and its close relatives within that group are noted. [ABSTRACT FROM AUTHOR]

The New World species of the genus Ceratoculicoides Wirth & Ratanaworabhan are described, illustrated and keyed in both sexes. Ceratoculicoides borkenti sp. nov., C. confusus sp. nov., C. grogani sp. nov., C. pacificus sp. nov. and C. propinquus sp. nov. are described, with C. confusus being the first record of the genus from South America (Colombia). Ceratoculicoides blantoni is a junior synonym of C. virginianus and the identity of C. longipennis remains unclear as the male morphospecies associated in taxonomic literature with the female type series is not conspecific. A morphological phylogeny of all extant species in the genus is presented and characters discussed. The moravicus species group is recognized for a clade of species with the lateral margins of the aedeagus straight or concave.

Onchocerca lupi (Rodonaja, 1967) is an understudied, vector-borne, filarioid nematode that causes ocular onchocercosis in dogs, cats, coyotes, wolves, and is also capable of infecting humans. Onchocercosis in dogs has been reported with increasing incidence worldwide. However, despite the growing number of reports describing canine O. lupi cases as well as zoonotic infections globally, the disease prevalence in endemic areas and vector species of this parasite remains largely unknown. Here, our study aimed to identify the occurrence of O. lupi infected dogs in northern Arizona, New Mexico, and Utah, United States and identify the vector of this nematode. A total of 532 skin samples from randomly selected companion animals with known geographic locations within the Navajo Reservation were collected and molecularly surveyed by PCR for the presence of O. lupi DNA (September 2019–June 2022) using previously published nematode primers (COI) and DNA sequencing. O. lupi DNA was detected in 50 (9.4%) sampled animals throughout the reservation. Using positive animal samples to target geographic locations, pointed hematophagous insect trapping was performed to identify potential O. lupi vectors. Out of 1,922 insects screened, 38 individual insects and 19 insect pools tested positive for the presence of O. lupi, all of which belong to the Diptera family. This increased surveillance of definitive host and biological vector/intermediate host is the first large scale prevalence study of O. lupi in companion animals in an endemic area of the United States, and identified an overall prevalence of 9.4% in companion animals as well as multiple likely biological vector and putative vector species in the southwestern United States. Furthermore, the identification of these putative vectors in close proximity to human populations coupled with multiple, local zoonotic cases highlight the One Health importance of O. lupi.

Blood-feeding arthropods are involved in the transmission of several pathogens that have a major impact on public health. Entomological investigations highlighted the composition, abundance, and diversity of flying hematophagous arthropods at four dog shelters located in central Morocco during an eight-month study, with the aim of discussing their vectorial roles and assessing the risk of these shelters as foci for zoonotic diseases. Monitoring of the arthropod fauna for 64 catch nights resulted in the collection of 2,321 biting midges (Ceratopogonidae), 570 mosquitoes (Culicidae), and 475 sand flies (Psychodidae). Fourteen Culicoides species were recorded and dominant species were Culicoides imicola (55.96%), C. paolae (16.07%), C. circumscriptus (10.29%), and C. newsteadi (5.77%). Three mosquito species were collected, including Culex pipiens s.l. (96.84%), Culiseta longiareolata (2.80%), and Cx. perexiguus (0.36%). Ten sand fly species were collected, including seven Phlebotomus species (62.70%) and three Sergentomyia species (37.30%); Sergentomyia minuta was the most dominant species (34.31%), followed by Phlebotomus sergenti (32.42%), typical Ph. perniciosus (8.63%), Ph. alexandri (6.94%), and Ph. riouxi (6.52%). The coexistence of several vectors in these study areas indicates the potential circulation of a wide range of pathogens, including zoonotic ones, thus requiring the implementation of surveillance and control programs to prevent the emergence and spread of disease outbreaks.

Vesicular stomatitis virus (VSV) is an arthropod-borne virus affecting livestock. In the United States, sporadic outbreaks result in significant economic losses. During epizootics, Culicoides biting midges are biological vectors and key to the geographic expansion of outbreaks. Additionally, Culicoides may play a role in VSV overwintering because females and males are capable of highly efficient venereal transmission, despite their relatively low virus titers. We hypothesized that VSV propagated within a midge has increased fitness for subsequent midge infections. To evaluate the potential host-specific fitness increase, we propagated three viral isolates of VSV in porcine skin fibroblasts and Culicoides cell lines. We then evaluated the viral infection dynamics of the different cell-source groups in Culicoides sonorensis. Our results indicate that both mammalian- and insect-derived VSV replicate well in midges inoculated via intrathoracic injection, thereby bypassing the midgut barriers. However, when the virus was required to infect and escape the midgut barrier to disseminate after oral acquisition, the insect-derived viruses had significantly higher titers, infection, and dissemination rates than mammalian-derived viruses. Our research suggests that VSV replication in Culicoides cells increases viral fitness, facilitating midge-to-midge transmission and subsequent replication, and further highlights the significance of Culicoides midges in VSV maintenance and transmission dynamics.

Abstract Background Aquatic ecosystems provide breeding sites for blood-sucking insects such as Culicoides biting midges (Diptera: Ceratopogonidae), but factors affecting their distribution and host choice are poorly understood. A study was undertaken at two nature reserves in northern Spain to examine the abundance, species composition, population dynamics and feeding patterns of biting midges between 2018 and 2019. Methods Culicoides were captured by light suction traps baited with CO2 and by sweep netting vegetation. Blood meals and species identification of blood-fed specimens were determined using cytochrome c oxidase I subunit (COI) DNA barcoding. Multivariate generalized linear models were used to evaluate the associations between the abundance of Culicoides, the species richness and other parameters. Results The 4973 identified specimens comprised 28 species of Culicoides. These included two species reported for the first time in northern Spain, thus raising to 54 the number of Culicoides species described in the region. Specimens of all 28 species and 99.6% of the total specimens collected were caught in suction traps, while sweep netting vegetation revealed just 11 species and 0.4% of the total specimens. Midge abundance peaked in June/early July, with five species comprising > 80% of the captures: Culicoides alazanicus (24.9%), Culicoides griseidorsum (20.3%), Culicoides poperinghensis (16.2%), Culicoides kibunensis (10.7%) and Culicoides clastrieri (9.6%). DNA barcode analysis of blood meals from eight Culicoides species revealed that they fed on 17 vertebrate species (3 mammals and 14 birds). Species in the subgenus Avaritia were primarily ornithophilic, except for C. griseidorsum and C. poperinghensis. Host DNA from blood meals was successfully amplified from 75% of blood-fed females. A pictorial blood meal digestion scale is provided to accurately assess the blood-fed status of female Culicoides. Conclusions The large number of different blood meal sources identified in the midges captured in this study signals the likely importance of wild birds and mammals (e.g. red deer and wild boar) as reservoir/amplifying hosts for pathogens. Available hosts are more exposed to being bitten by biting midge populations in aquatic ecosystems in late spring and early summer.

The new species Atrichopogon (Atrichopogon) tolfensisn. sp. from Tolfa Mountains (Italy, Latium, and Rome) is described and illustrated. Both sexes of the species were collected using cantharidin-baited traps from June 2020 to January 2021. The new species is an atypical member of the genus Atrichopogon. [ABSTRACT FROM AUTHOR]

The territory of the Tyumen region is located in the West Siberian Lowland and is characterized by conditions favorable for the development of blood-sucking diptera insects. The fauna of the blood-sucking diptera insects of the Tyumen region is represented by 109 species and 4 subspecies: horseflies-35 species and 4 subspecies, mosquitoes-41 species, blackflies-16 species, biting midges-17 species. In the subzone of the southern taiga, Hybomitra ciureai, H. bimaculata, H. lundbecki, H. muehlfeldi, Haematopota pluvialis, Aedes cantans, A. punctor, Simulium maculatum, Culicoides punctatus dominate; in the subzone of aspenbirch forests-Tabanus bromius, Hybomitra ciureai, H. bimaculata, H. lundbecki, Atylotus fulvus, A. rusticus, Haematopota subcylindrica, H. pluvialis, Aedes cinereus, A. punctor, A. cantans, Simulium pusillum, Culicoides punctatus; in the forest-steppe zone-Tabanus bovinus, T. bromius, Hybomitra ciureai, H. bimaculata, H. lundbecki, Haematopota pluvialis, H. subcylindrica, Coquillettidia richiardii, Aedes caspius, A. albescens, A. cantans, Simulium maculatum, Culicoides punctatus. The total summer period of insects of this group ranges from 140 days in the subzone of the southern taiga to 170 days in the forest-steppe zone. Depending on the meteorological conditions of the spring-summer season, the start and end dates of summer may shift by 1-2 decades. The activity of horseflies and blackflies is observed during the daytime, and mosquitoes and biting midges are twilight insects and attack mainly in the morning and evening hours. The main abiotic factors determining the nature of the diurnal rhythm of the vital activity of diptera insects in the conditions of the Tyumen region are air temperature and illumination. [ABSTRACT FROM AUTHOR]

Abstract Background Proper vector surveillance relies on the ability to identify species of interest accurately and efficiently, though this can be difficult in groups containing cryptic species. Culicoides Latreille is a genus of small biting flies responsible for the transmission of numerous pathogens to a multitude of vertebrates. Regarding pathogen transmission, the C. variipennis species complex is of particular interest in North America. Of the six species within this group, only C. sonorensis Wirth & Jones is a proven vector of bluetongue virus and epizootic hemorrhagic disease virus. Unfortunately, subtle morphological differences, cryptic species, and mitonuclear discordance make species identification in the C. variipennis complex challenging. Recently, single-nucleotide polymorphism (SNP) analysis enabled discrimination between the species of this group; however, this demanding approach is not practical for vector surveillance. Methods The aim of the current study was to develop a reliable and affordable way of distinguishing between the species within the C. variipennis complex, especially C. sonorensis. Twenty-five putative microsatellite markers were identified using the C. sonorensis genome and tested for amplification within five species of the C. variipennis complex. Machine learning was then used to determine which markers best explain the genetic differentiation between species. This led to the development of a subset of four and seven markers, which were also tested for species differentiation. Results A total of 21 microsatellite markers were successfully amplified in the species tested. Clustering analyses of all of these markers recovered the same species-level identification as the previous SNP data. Additionally, the subset of seven markers was equally capable of accurately distinguishing between the members of the C. variipennis complex as the 21 microsatellite markers. Finally, one microsatellite marker (C508) was found to be species-specific, only amplifying in the vector species C. sonorensis among the samples tested. Conclusions These microsatellites provide an affordable way to distinguish between the sibling species of the C. variipennis complex and could lead to a better understanding of the species dynamics within this group. Additionally, after further testing, marker C508 may allow for the identification of C. sonorensis with a single-tube assay, potentially providing a powerful new tool for vector surveillance in North America. Graphical Abstract

Within the scope of our research, we have performed 59 trapping sessions and collected 15,756 biting midges from 20 species at four farms (Kluknava, Ostrov, Turňa nad Bodovou and Zemplínska Teplica), The most frequent types of captured insects were representatives of the Avaritia subgenus, C. obsoletus/C. scoticus, representing on average 85.1 % (13,295 individuals) of the fauna of the biting midges, with the exception of the farm in Ostrov where this group represented only 41.7 % of the fauna. At this particular farm, the most frequently trapped insects belonged to the Culicoides subgenus (54.1 %), in particular the C. bysta, C. lupicaris, C. newsteadi, C. Pulicaris and C. Punctatus species. During the trapping sessions, we monitored factors affecting the number of trapped biting midges, such as the temperature, relative air humidity and airflow rate: the air temperature during the trapping of the biting midges ranged from 9.8 to 26.2 °C; the relative air humidity ranged from 35.1 to 100 %; and the air flow rate ranged from no wind to a wind velocity of 8.2 m.s-1. However, in the final evaluation, we failed to observe a statistically significant correlation between the air flow and the number of trapped biting midges. The largest amounts of biting midges were trapped at temperatures ranging from 15.8 to 24.6 °C and at a relative air humidity ranging from 54.2 % to 68.6 %. While monitoring the seasonal dynamics of the physiological conditions of biting midge females at the selected farms, we confirmed that during the period from June to August, the most frequently trapped females were parous (50.1 %; 7,826 individuals). In addition, nulliparous females comprised 43.8 % (6,842 individuals) and were continuously trapped throughout the season (April— November).

Simple Summary: Biting midges, which feed on a variety of hosts, transmit a wide range of harmful human and animal viruses and parasites. According to recent studies in Thailand, biting midges may act as potential vectors for leishmaniasis and trypanosomiasis. The purpose of this study is to look for Leishmania and Trypanosoma DNA in biting midges obtained in the field in a leishmaniasis-endemic area in Phatthalung province, Southern Thailand. In addition, we analysed engorged midges for host blood DNA and screened the collected midges for avian haemosporidian parasites. According to our findings, biting midges have a diversified feeding habit and may be able to transmit various pathogens, including trypanosomatid and avian haemosporidian parasites. Five hundred and fifty-nine female biting midges were collected, and seventeen species in six subgenera (Avaritia, Haemophoructus, Hoffmania, Meijerehelea, Remmia, and Trithecoides) and two groups (Clavipalpis and Shortti) were identified. The dominant Culicoides species was C. peregrinus (30.94%), followed by C. subgenus Trithecoides. From blood meal analysis of engorged biting midges, they were found to feed on cows, dogs, pigs, and avians. The majority of blood preferences of biting midges (68%; 49/72) displayed a mixed pattern of host blood DNA (cow and avian). The overall non-engorged biting midge field infectivity rate was 1.44 % (7/487). We detected Leucocytozoon sp. in three Culicoides specimens, one from each species: C. fulvus, C. oxystoma, and C. subgenus Trithecoides. Crithidia sp. was found in two C. peregrinus specimens, and Trypanosoma sp. and P. juxtanucleare were separately found in two C. guttifer. More consideration should be paid to the capacity of biting midges to transmit pathogens such as avian haemosporidian and trypanosomatid parasites. To demonstrate that these biting midges are natural vectors of trypanosomatid parasites, additional research must be conducted with a greater number of biting midges in other endemic regions. [ABSTRACT FROM AUTHOR]

Culicoides midges are hematophagous insects that transmit arboviruses of veterinary importance. These viruses include bluetongue virus (BTV) and epizootic hemorrhagic fever virus (EHDV). The endosymbiont Wolbachia pipientis Hertig spreads rapidly through insect host populations and has been demonstrated to inhibit viral pathogen transmission in multiple mosquito vectors. Here, we have demonstrated a replication inhibitory effect on BTV and EHDV in a Wolbachia (wAlbB strain)‐infected Culicoides sonorensis Wirth and Jones W8 cell line. Viral replication was significantly reduced by day 5 for BTV and by day 2 for EHDV as detected by real‐time polymerase chain reaction (RT‐qPCR) of the non‐structural NS3 gene of both viruses. Evaluation of innate cellular immune responses as a cause of the inhibitory effect showed responses associated with BTV but not with EHDV infection. Wolbachia density also did not play a role in the observed pathogen inhibitory effects, and an alternative hypothesis is suggested. Applications of Wolbachia‐mediated pathogen interference to impact disease transmission by Culicoides midges are discussed. [ABSTRACT FROM AUTHOR]

Background: Mitochondrial genomes are the most sequenced genomes after bacterial and fungal genomic DNA. However, little information on mitogenomes is available for multiple metazoan taxa, such as Culicoides, a globally distributed, megadiverse genus containing 1,347 species. Aim: Generating novel mitogenomic information from single Culicoides sonorensis and C. biguttatus specimens, comparing available mitogenome mapping and de novo assembly tools, and identifying the best performing strategy and tools for Culicoides species. Results: We present two novel and fully annotated mitochondrial haplotypes for two Culicoides species, C. sonorensis and C. biguttatus. We also annotated or re-annotated the only available reference mitogenome for C. sonorensis and C. arakawae. All species present a high similarity in mitogenome organization. The general gene arrangement for all Culicoides species was identical to the ancestral insect mitochondrial genome. Only short spacers were found in C. sonorensis (up to 30 bp), contrary to C. biguttatus (up to 114 bp). The mitochondrial genes ATP8, NAD2, NAD6, and LSU rRNA exhibited the highest nucleotide diversity and pairwise interspecific p genetic distance, suggesting that these genes might be suitable and complementary molecular barcodes for Culicoides identification in addition to the commonly utilized COI gene. We observed performance differences between the compared mitogenome generation strategies. The mapping strategy outperformed the de novo assembly strategy, but mapping results were partially biased in the absence of species-specific reference mitogenome. Among the utilized tools, BWA performed best for C. sonorensis while SPAdes, MEGAHIT, and MitoZ were among the best for C. biguttatus. The best-performing mitogenome annotator was MITOS2. Additionally, we were able to recover exogenous mitochondrial DNA from Bos taurus (biting midges host) from a C. biguttatus blood meal sample. Conclusions: Two novel annotated mitogenome haplotypes for C. sonorensis and C. biguttatus using High-Throughput Sequencing are presented. Current results are useful as the baseline for mitogenome reconstruction of the remaining Culicoides species from single specimens to HTS and genome annotation. Mapping to a species-specific reference mitogenome generated better results for Culicoides mitochondrial genome reconstruction than de novo assembly, while de novo assembly resulted better in the absence of a closely related reference mitogenome. These results have direct implications for molecular-based identification of these vectors of human and zoonotic diseases, setting the basis for using the whole mitochondrial genome as a marker in Culicoides identification. [ABSTRACT FROM AUTHOR]

We provide new records of Culicoides paraensis from the Yucatan Peninsula. The anthropophilic biting midge C. paraensis is the main vector of Oropouche fever virus in South and Central America. We also report Culicoides poikilonotus for the 1st time in the Yucatan Peninsula and a key to identify the species in this region of Mexico., (Copyright © 2024 by The American Mosquito Control Association, Inc.)

Abstract Background Culicoides insignis is a confirmed vector of bluetongue virus (BTV) throughout the American tropics and a possible vector of epizootic hemorrhagic disease virus (EHDV) in Florida. Despite its importance, fundamental information on the biology and ecology of this vector species is lacking. In this study, we examined the oviposition of C. insignis under laboratory conditions, monitored the development of immature stages and attempted colonization of this species. Methods Live C. insignis females were collected from the field using CDC-UV-LED traps, allowed to blood-feed on live chicken and given various natural substrates for oviposition in two-choice assays. The eggs deposited were transferred to 0.3% agar slants, and the hatched larvae were provided a diet of Panagrellus redivivus Linnaeus nematodes and the development of all immature stages was monitored. Results Culicoides insignis females exhibited an overall oviposition preference for dishes containing mud from their larval habitat as gravid females deposited a significantly higher number of eggs on these dishes (35.3 eggs/female) than on controls (17.7 eggs/female). The ovipositing females also deposited a higher percentage of eggs on substrates with habitat mud and other organically enriched muds (≥ 75.2%) compared to controls (31.0%). The larvae developed successfully to adulthood on the nematode diet, exhibiting high overall larval survival rates (85.0%). Sex ratios of the F1 generation were male biased, approximately 3:1 (male:female). Captive mating could not be induced in the F1 adults. Conclusions Mud from the larval habitat and other organically enriched muds provide strong oviposition cues to C. insignis under laboratory conditions. Further studies will be needed to identify the key biotic/abiotic factors influencing midge oviposition in the field. The agar/nematode method is effective for the rearing of C. insignis larvae. However, further studies will be needed to address the issue of male-biased sex ratios in the progeny and to examine the mating habits/cues of C. insignis in nature, which may provide clues towards inducing captive mating in the F1 adults. Graphical abstract

Two new species and a new record of blood sucking biting midges of the subgenus Lasiohelea Kieffer, 1921 of the genus Forcipomyia Meigen, 1818 are described after the morphological and molecular data. The new species. Forcipomyia (Lasiohelea) peditata and F. (L.) falcata were fetched from the Sub-Himalayan region, and F. (L.) parvitas (Liu and Yu, 1996) from both the Gangetic plain and western plateau regions of West Bengal. The DNA barcoding of Mitochondrial COX I gene has also been used as molecular evidence.

Culicoides sonorensis midges vector multiple livestock arboviruses, resulting in significant economic losses worldwide. Due to the tight association between virus transmission, blood feeding, and egg development, understanding midge physiology is paramount to limiting pathogen transmission. Previous studies have demonstrated the importance of small non-coding RNAs (ncRNAs), specifically microRNAs (miRNAs), in multiple aspects of vector physiology. These small ncRNAs regulate gene expression at the post-transcriptional level and display differential expression during pathogen infection. Due to the lack of annotated miRNAs in the biting midge and associated expression profiles, we used small RNA-Seq and miRDeep2 analyses to determine the Culicoides miRNAs in whole females and midgut tissues in response to blood feeding. Our analyses revealed 76 miRNAs within C. sonorensis composed of 73 orthologous and three candidate novel miRNAs, as well as conserved miRNA clusters. miRNA conservation suggests an interesting evolutionary relationship between miRNA expression and hematophagy in the infraorder Culicomorpha. We also identified multiple blood meal-regulated and tissue-enriched miRNAs. Lastly, we further identified miRNAs with expression patterns potentially associated with virus infection by probing publicly available datasets. Together, our data provide a foundation for future ncRNA work to untangle the dynamics of gene regulation associated with midge physiology.

Models on climate change have predicted an increase of temperature over the earth's surface with potential drastic effects on living organisms. We analyzed the relationships between climatic conditions (temperature, rainfall, and wind speed) and the abundance of blood‐sucking flying insects (biting midges and blackflies) and nest‐dwelling ectoparasites (mites, fleas, and blowflies) collected from blue tit nests during bird breeding seasons for a period of 10 years. Average temperature, rainfall, and wind speed showed significant differences among years. Temperature and wind speed increased during the period of study while rainfall decreased. Biting midge, blackfly, and blowfly abundances increased across years but not flea and mite abundances. Hatching date decreased and brood size increased across years. Independently of year variation, parasites were related to climatic variables. For example, biting midge, blowfly, mite, and flea abundances were positive and significantly related to average temperature. We also found a positive and significant relationship between abundances of Haemoproteus infections and biting midge abundances during the first year of life of birds out of nests. However, abundance and prevalence of Lankesterella infections in yearlings were positive and significantly related to mite abundances during the year of birth of birds. Leucocytozoon and Lankesterella infections were also significantly related to climatic variables and Haemoproteus and Lankesterella infections increased across years. In addition, body condition of adult females and males were negatively related to flea larvae and blowfly abundance respectively. Nestling body condition was also negatively related to biting midge abundance. Changes in climatic conditions across years could therefore affect several parasites of birds but also to birds themselves. [ABSTRACT FROM AUTHOR]

Background: Aquatic ecosystems provide breeding sites for blood-sucking insects such as Culicoides biting midges (Diptera: Ceratopogonidae), but factors affecting their distribution and host choice are poorly understood. A study was undertaken at two nature reserves in northern Spain to examine the abundance, species composition, population dynamics and feeding patterns of biting midges between 2018 and 2019. Methods: Culicoides were captured by light suction traps baited with CO2 and by sweep netting vegetation. Blood meals and species identification of blood-fed specimens were determined using cytochrome c oxidase I subunit (COI) DNA barcoding. Multivariate generalized linear models were used to evaluate the associations between the abundance of Culicoides, the species richness and other parameters. Results: The 4973 identified specimens comprised 28 species of Culicoides. These included two species reported for the first time in northern Spain, thus raising to 54 the number of Culicoides species described in the region. Specimens of all 28 species and 99.6% of the total specimens collected were caught in suction traps, while sweep netting vegetation revealed just 11 species and 0.4% of the total specimens. Midge abundance peaked in June/early July, with five species comprising > 80% of the captures: Culicoides alazanicus (24.9%), Culicoides griseidorsum (20.3%), Culicoides poperinghensis (16.2%), Culicoides kibunensis (10.7%) and Culicoides clastrieri (9.6%). DNA barcode analysis of blood meals from eight Culicoides species revealed that they fed on 17 vertebrate species (3 mammals and 14 birds). Species in the subgenus Avaritia were primarily ornithophilic, except for C. griseidorsum and C. poperinghensis. Host DNA from blood meals was successfully amplified from 75% of blood-fed females. A pictorial blood meal digestion scale is provided to accurately assess the blood-fed status of female Culicoides. Conclusions: The large number of different blood meal sources identified in the midges captured in this study signals the likely importance of wild birds and mammals (e.g. red deer and wild boar) as reservoir/amplifying hosts for pathogens. Available hosts are more exposed to being bitten by biting midge populations in aquatic ecosystems in late spring and early summer. [ABSTRACT FROM AUTHOR]

Two new species and a new record of blood sucking biting midges of the subgenus Lasiohelea Kieffer, 1921 of the genus Forcipomyia Meigen, 1818 are described after the morphological and molecular data. The new species. Forcipomyia (Lasiohelea) peditata and F. (L.) falcata were fetched from the Sub-Himalayan region, and F. (L.) parvitas (Liu and Yu, 1996) from both the Gangetic plain and western plateau regions of West Bengal. The DNA barcoding of Mitochondrial COX I gene has also been used as molecular evidence. [ABSTRACT FROM AUTHOR]

Abstract Background Culicoides obsoletus (s.l.) is the most abundant Culicoides species in northern Europe and an important vector of bluetongue virus and Schmallenberg virus. Nevertheless, information on its subadult life stages remains scarce and no laboratory-reared colony exists. Methods C. obsoletus (s.l.) adults were collected in Belgium and transferred to the laboratory in an attempt to establish a laboratory-reared colony. C. obsoletus (s.l.) were reared from eggs to adults at different temperatures (28 °C, 24 °C, 20/16 °C) and under different food regimes. Results The most suitable temperature for rearing seemed to be 24 °C for most developmental parameters, but resulted in a biased 3:1 male/female sex ratio. The latter could be optimized to a 1:1 sex ratio when a 20/16 °C day/night temperature gradient was applied, but rearing at these low temperature conditions resulted in significantly lower egg hatching and pupation rates and a longer subadult development time. Independent of the rearing temperature, adding dung as an additional food source during larval development resulted in a significantly higher adult emergence rate and a decrease in subadult development time. Furthermore, blood-feeding rates of field-collected C. obsoletus (s.l.) were compared for different blood sources and feeding systems. The overall blood-feeding success was low and only successful with cotton pledgets (2.7% blood-fed midges) and through a membrane system with chicken skin (3.5% blood-fed midges). Higher feeding rates were obtained on cattle blood compared to sheep blood. Conclusions These results will help us to determine the necessary conditions to rear a viable laboratory colony of this important vector species, although further optimization is still required.

Ceratopogonidae (biting midges) species are of medical and veterinary importance and play a role as disease vectors caused by viruses, protozoa, and filarial nematodes. Besides their vectorial importance, they are harmful due to painful bites that cause discomfort in humans and animals. Identifying possible disease vectors is of significant epidemiological importance, therefore, the aim of this study was to carry out a survey of the species of Ceratopogonids that occur in Santa Maria, Rio Grande do Sul, to verify the frequency of occurrence of adults of Ceratopogonidae within one year of capture. Insects were collected weekly (three night a week) using a Captor® suction light trap, from March 2020 to March 2021, between 6 p.m. and 6 a.m. Collections of biting midges were separated from other insects, divided into genera and morphologically identified at the species level. A total of 16,054 specimens (71.55% female to 28.44% male) of Ceratopogonidae, were collected and identified, comprising thirteen species. Data presented show that several species of biting midges, including seven new records, have variable frequency and occur year-round at the study site, with the highest number in the warm season. The dominant species were Culicoides insignis and Culicoides venezuelensis. [ABSTRACT FROM AUTHOR]

Background: Proper vector surveillance relies on the ability to identify species of interest accurately and efficiently, though this can be difficult in groups containing cryptic species. Culicoides Latreille is a genus of small biting flies responsible for the transmission of numerous pathogens to a multitude of vertebrates. Regarding pathogen transmission, the C. variipennis species complex is of particular interest in North America. Of the six species within this group, only C. sonorensis Wirth & Jones is a proven vector of bluetongue virus and epizootic hemorrhagic disease virus. Unfortunately, subtle morphological differences, cryptic species, and mitonuclear discordance make species identification in the C. variipennis complex challenging. Recently, single-nucleotide polymorphism (SNP) analysis enabled discrimination between the species of this group; however, this demanding approach is not practical for vector surveillance. Methods: The aim of the current study was to develop a reliable and affordable way of distinguishing between the species within the C. variipennis complex, especially C. sonorensis. Twenty-five putative microsatellite markers were identified using the C. sonorensis genome and tested for amplification within five species of the C. variipennis complex. Machine learning was then used to determine which markers best explain the genetic differentiation between species. This led to the development of a subset of four and seven markers, which were also tested for species differentiation. Results: A total of 21 microsatellite markers were successfully amplified in the species tested. Clustering analyses of all of these markers recovered the same species-level identification as the previous SNP data. Additionally, the subset of seven markers was equally capable of accurately distinguishing between the members of the C. variipennis complex as the 21 microsatellite markers. Finally, one microsatellite marker (C508) was found to be species-specific, only amplifying in the vector species C. sonorensis among the samples tested. Conclusions: These microsatellites provide an affordable way to distinguish between the sibling species of the C. variipennis complex and could lead to a better understanding of the species dynamics within this group. Additionally, after further testing, marker C508 may allow for the identification of C. sonorensis with a single-tube assay, potentially providing a powerful new tool for vector surveillance in North America. [ABSTRACT FROM AUTHOR]

Abstract Background Culicoides biting midges (Diptera, Ceratopogonidae) are known vectors of avian Haemoproteus parasites. These parasites cause diseases, pathology and even mortality in birds. The diversity of biting midges in Europe is great, but only four Culicoides species are known to be vectors of avian Haemoproteus parasites. In general, our knowledge about the role of the particular Culicoides species in the transmission of Haemoproteus parasites remains insufficient. Information gaps hinder a better understanding of parasite biology and the epizootiology of parasite-caused diseases. The aim of this study was to determine new Culicoides species involved in the transmission of Haemoproteus parasites. Methods Biting midges were collected using a UV trap as well as sticky traps installed in bird nest boxes. Individual parous females were diagnosed for the presence of haemoproteids using both PCR-based and microscopic methods. Results We collected and dissected 232 parous Culicoides females from 9 species using a UV trap and 293 females from 11 species from bird nest boxes. Culicoides obsoletus was the dominant species collected using a UV trap, and Culicoides kibunensis dominated among midges collected in nest boxes. PCR-based screening showed that 5.2% of parous biting midges collected using a UV trap and 4.4% of midges collected from nest boxes were infected with avian haemosporidian parasites. Haemoproteid DNA was detected in C. kibunensis, Culicoides pictipennis, Culicoides punctatus, Culicoides segnis and Culicoides impunctatus females. The sporozoites of Haemoproteus minutus (genetic lineages hTURDUS2 and hTUPHI01) were detected in the salivary glands of two C. kibunensis females using microscopy, and this finding was confirmed by PCR. Conclusions Culicoides kibunensis was detected as a new natural vector of Haemoproteus minutus (hTURDUS2 and hTUPHI01). Haemoproteid DNA was detected in females from five Culicoides species. This study contributes to the epizootiology of avian Haemoproteus infections by specifying Culicoides species as vectors and species that are likely to be responsible for the transmission of haemoproteids in Europe.

Culicoides is a genus of biting midges in the family Ceratopogonidae, representing a big threat, giving that several species serve as biological vectors of pathogens of veterinary importance. BTV distribution and persistence is thought to be almost entirely dependent upon the presence or absence of Culicoides adults vector. In this study, biting midges were collected during 2015-2016, once a week, from April to December. In the first year, 2617 BTV vector specimens were collected and identified as Culicoides obsoletus (41.2%; 95% CI: 39-43), Culicoides pulicaris (58%; 95% CI: 56-59) and Culicoides nubeculosus (0.8%; 95% CI: 0.4-1.1). In the second year, 165 BTV vector specimens were collected and identified as Culicoides obsoletus (44.25%; 95% CI: 36-51), Culicoides pulicaris (50.3%; 95% CI: 42-57), Culicoides dewulfi (4.84%; 95% CI: 1.5-8.1) and Culicoides nubeculosus (0.61%; 95% CI: 0.5-1.7). Culicoides pulicaris and Culicoides obsoletus complex were especially abundant within the two collection sites in Iași County. Our data on post-epidemic occurrence and composition of BTV vector species in the studied area will facilitate a better understanding of Culicoides vectors distribution and the possible role in bluetongue virus overwintering.