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Bluetongue (BT) is a Culicoides midge-borne hemorrhagic disease affecting cervids and ruminant livestock species, resulting in significant economic losses from animal production and trade restrictions. Experimental animal infections using the α/β interferon receptor knockout IFNAR mouse model and susceptible target species are critical for understanding viral pathogenesis, virulence, and evaluating vaccines. However, conducting experimental vector-borne transmission studies with the vector itself are logistically difficult and experimentally problematic. Therefore, experimental infections are induced by hypodermic injection with virus typically derived from baby hamster kidney (BHK) cells. Unfortunately, for many U.S. BTV serotypes, it is difficult to replicate the severity of the disease seen in natural, midge-transmitted infections by injecting BHK-derived virus into target host animals. Using the IFNAR BTV murine model, we compared the virulence of traditional BHK cell-derived BTV-17 with C. sonorensis midge (W8) cell-derived BTV-17 to determine whether using cells of the transmission vector would provide an in vitro virulence aspect of vector-transmitted virus. At both low and high doses, mice inoculated with W8-BTV-17 had an earlier onset of viremia, earlier onset and peak of clinical signs, and significantly higher mortality compared to mice inoculated with BHK-BTV-17. Our results suggest using a Culicoides W8 cell-derived inoculum may provide an in vitro vector-enhanced infection to more closely represent disease levels seen in natural midge-transmitted infections while avoiding the logistical and experimental complexity of working with live midges.

In October 2023, bluetongue virus serotype 3 (BTV-3) emerged in Germany, where Schmallenberg virus is enzootic. We detected BTV-3 in 1 pool of Culicoides biting midges collected at the time ruminant infections were reported. Schmallenberg virus was found in many vector pools. Vector trapping and analysis could elucidate viral spread.

Bluetongue virus (BTV) is a segmented, double-stranded RNA virus transmitted by Culicoides midges that infects ruminants. As global temperatures increase and geographical ranges of midges expand, there is increased potential for BTV outbreaks from incursions of novel serotypes into endemic regions. However, an understanding of the effect of temperature on reassortment is lacking. The objectives of this study were to compare how temperature affected Culicoides survival, virogenesis, and reassortment in Culicoides sonorensis coinfected with two BTV serotypes. Midges were fed blood meals containing BTV-10, BTV-17, or BTV serotype 10 and 17 and maintained at 20 °C, 25 °C, or 30 °C. Midge survival was assessed, and pools of midges were collected every other day to evaluate virogenesis of BTV via qRT-PCR. Additional pools of coinfected midges were collected for BTV plaque isolation. The genotypes of plaques were determined using next-generation sequencing. Warmer temperatures impacted traits related to vector competence in offsetting ways: BTV replicated faster in midges at warmer temperatures, but midges did not survive as long. Overall, plaques with BTV-17 genotype dominated, but BTV-10 was detected in some plaques, suggesting parental strain fitness may play a role in reassortment outcomes. Temperature adds an important dimension to host-pathogen interactions with implications for transmission and evolution.

Epizootic hemorrhagic disease (EHD) is a non-contagious arthropod-transmitted viral disease and a World Organization for Animal Health (WOAH)-listed disease of domestic and wild ruminants since 2008. EHDV is transmitted among susceptible animals by a few species of midges of genus Culicoides . During the fall of 2021, a large outbreak caused by the epizootic hemorrhagic disease virus (EHDV), identified as serotype 8, was reported in Tunisian dairy and beef farms with Bluetongue virus (BTV)-like clinical signs. The disease was detected later in the south of Italy, in Spain, in Portugal and, more recently, in France, where it caused severe infections in cattle. This was the first evidence of EHDV-8 circulation outside Australia since 1982. In this study, we analyzed the epidemiological situation of the 2021-2022 EHDV outbreaks reported in Tunisia, providing a detailed description of the spatiotemporal evolution of the disease. We attempted to identify the eco-climatic factors associated with infected areas using generalized linear models (GLMs). Our results demonstrated that environmental factors mostly associated with the presence of C. imicola , such as digital elevation model (DEM), slope, normalized difference vegetation index (NDVI), and night-time land surface temperature (NLST)) were by far the most explanatory variables for EHD repartition cases in Tunisia that may have consequences in neighboring countries, both in Africa and Europe through the spread of infected vectors. The risk maps elaborated could be useful for disease control and prevention strategies.

Background: Bluetongue is a non-contagious viral disease that affects both domestic and wild ruminants. It is transmitted primarily by small hematophagous Diptera belonging to the genus Culicoides (Diptera: Ceratopogonidae). The current study represents the first molecular investigation into the potential role of Culicoides imicola, Culicoides paolae, Culicoides newsteadi, Culicoides spp., and Culicoides circumscriptus as bluetongue virus (BTV) vectors in Morocco. Additionally, the study aimed to evaluate the vectorial activity of midges during the survey seasons., Methods: Parous females of these species were captured from several regions of Morocco (6 out of 12) from 2018 to 2021 using Onderstepoort Veterinary Institute (OVI) traps. A total of 2003 parous female specimens were grouped into 55 batches. The midge body of each batch was dissected into three regions (head, thorax, and abdomen), and these regions were analyzed separately using reverse transcription quantitative polymerase chain reaction (RT-qPCR)., Results: BTV RNA was detected in 45 out of the 55 batches tested, indicating a positivity rate of 81.8%. The RT-qPCR-positive pools of the studied Culicoides species exhibited high levels of BTV positivity in each body part (head, thorax, and abdomen), confirming the successful replication of the virus within midge bodies. The BTV circulation was substantial across all three survey seasons (spring, summer, and autumn). High infection rates, calculated using the minimum infection rate (MIR) and maximum likelihood estimation (MLE), were observed during the collection seasons, particularly in autumn and spring, and for all investigated Culicoides species, most notably for C. imicola and C. newsteadi. These increased infection rates underscore the significant risk of Culicoides transmitting the BTV in Morocco., Conclusions: The detection of BTV positivity in Culicoides spp. (lacking wing spots that allow their differentiation according to morphological identification keys) suggested that other Culicoides species are competent for BTV transmission in Morocco. The study results indicated, for the first time at the molecular level, that C. imicola and C. newsteadi are the primary potential vectors of BTV in Morocco and that C. paolae and C. circumscriptus are strongly implicated in the propagation of bluetongue at the national level., (© 2024. The Author(s).)

Bluetongue virus (BTV) is a segmented, double-stranded RNA orbivirus listed by the World Organization for Animal Health and transmitted by Culicoides biting midges. Segmented viruses can reassort, which facilitates rapid and important genotypic changes. Our study evaluated reassortment in Culicoides sonorensis midges coinfected with different ratios of BTV-10 and BTV-17. Midges were fed blood containing BTV-10, BTV-17, or a combination of both serotypes at 90:10, 75:25, 50:50, 25:75, or 10:90 ratios. Midges were collected every other day and tested for infection using pan BTV and cox1 (housekeeping gene) qRT-PCR. A curve was fit to the ∆Ct values (pan BTV Ct- cox1 Ct) for each experimental group. On day 10, the midges were processed for BTV plaque isolation. Genotypes of the plaques were determined by next-generation sequencing. Pairwise comparison of ∆Ct curves demonstrated no differences in viral RNA levels between coinfected treatment groups. Plaque genotyping indicated that most plaques fully aligned with one of the parental strains; however, reassortants were detected, and in the 75:25 pool, most plaques were reassortant. Reassortant prevalence may be maximized upon the occurrence of reassortant genotypes that can outcompete the parental genotypes. BTV reassortment and resulting biological consequences are important elements to understanding orbivirus emergence and evolution.

Introduction: Bluetongue virus (BTV) is an arthropod-borne Orbivirus that is almost solely transmitted by Culicoides biting midges and causes a globally important haemorrhagic disease, bluetongue (BT), in susceptible ruminants. Infection with BTV is characterised by immunosuppression and substantial lymphopenia at peak viraemia in the host., Methods: In this study, the role of cell-mediated immunity and specific T-cell subsets in BTV pathogenesis, clinical outcome, viral dynamics, immune protection, and onwards transmission to a susceptible Culicoides vector is defined in unprecedented detail for the first time, using an in vivo arboviral infection model system that closely mirrors natural infection and transmission of BTV. Individual circulating CD4 + , CD8 + , or WC1 + γδ T-cell subsets in sheep were depleted through the administration of specific monoclonal antibodies., Results: The absence of cytotoxic CD8 + T cells was consistently associated with less severe clinical signs of BT, whilst the absence of CD4 + and WC1 + γδ T cells both resulted in an increased clinical severity. The absence of CD4 + T cells also impaired both a timely protective neutralising antibody response and the production of IgG antibodies targeting BTV non-structural protein, NS2, highlighting that the CD4 + T-cell subset is important for a timely protective immune response. T cells did not influence viral replication characteristics, including onset/dynamics of viraemia, shedding, or onwards transmission of BTV to Culicoides . We also highlight differences in T-cell dependency for the generation of immunoglobulin subclasses targeting BTV NS2 and the structural protein, VP7., Discussion: This study identifies a diverse repertoire of T-cell functions during BTV infection in sheep, particularly in inducing specific anti-viral immune responses and disease manifestation, and will support more effective vaccination strategies., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Newbrook, Khan, Fisher, Chong, Gubbins, Davies, Sanders, Busquets, Cooke, Corla, Ashby, Flannery, Batten, Stokes, Sanz-Bernardo, Carpenter, Moffat and Darpel.)

Introduction: Culicoides plays a crucial role as an insect vector in the field of veterinary medicine. The transmission of significant viruses such as bluetongue virus (BTV) and African horse sickness virus (AHSV) by this insect poses a substantial threat, leading to the development of severe diseases in domestic animals. This study aimed to explore the Culicoides species, identify their blood-meal sources, and assess the presence of BTV and AHSV carried by Culicoides in Yuanyang County, Yunnan Province. The aim was to gain insights into the potential vectors of these two viruses and elucidate their potential roles in the transmission of pathogens., Methods: The midges were collected from cattle ( Bos indicus ), pig ( Sus scrofa ), and goat ( Capra hircus ) pens in Yuanyang County, Yunnan Province in June 2020. Initial identification of midges was conducted through morphological characteristics, followed by molecular identification using the cytochrome C oxidase subunit I (COI) gene. The determination of Culicoides blood-meal sources was accomplished using specific primers targeting the cytochrome b (Cyt b ) gene from potential hosts. BTV and AHSV RNA were identified in Culicoides pools through the application of reverse transcriptase PCR and quantitative real-time PCR. Nucleotide homology and phylogenetic analysis were performed using MegAlign (DNAStar) and Mega 6.0 software., Results: A total of 6,300 Culicoides , consisting of C. oxystoma, C. arakawai, C. imicola , and C. innoxius , were collected from cattle, pigs, and goat pens. The engorgement rates for these species were 30.2%, 54.6%, 75%, and 66.7%, respectively. In the cattle pen, the prevailing species is C. oxystoma (100%). In the pig pen, C. arakawai dominates (70%), with C. oxystoma following at 30%. In the goat pen, C. imicola holds the majority (45.45%), trailed by C. oxystoma (25%), C. innoxius (20.45%), and C. arakawai (9.09%). These Culicoides species were identified as feeding on cattle, pigs, goats, chickens ( Gallus gallus ), and humans ( Homo sapiens ). The positivity rates for BTV were 20.00% and 11.54% in blood-fed specimens of C. imicola and C. oxystoma , respectively. Conversely, the positivity rates for BTV in non-blood-fed specimens were 0.00% and 6.67% for C. imicola and C. oxystoma , respectively. BTV was not detected in C. arakawai and C. innoxius . The specimens (YY86) from C. imicola that tested positive for BTV had the closest genetic relationship to YTS-4 isolated from Mangshi, Yunnan Province in 1996. All test results for the nucleic acid of AHSV were negative., Conclusion: The study reveals variations in the species distribution, community composition, blood sucking rate, and blood-feeding sources of Culicoides across different habitats. Notably, C. imicola and C. oxystoma emerge as potential vectors for the transmission of BTV in local animals. Accordingly, this investigation provides crucial insights that can serve as a valuable reference for the prevention and control of BTV in local animals, particularly from the perspective of vector management., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Li, Meng, He, Wang and Wang.)

Habitat selection of Culicoides spp. (Diptera: Ceratopogonidae) is influenced by the physicochemical factors such as temperature, pH, salinity, moisture, conductivity, organic and inorganic compounds of substrates. These factors determine the life history traits of the vectors. We studied the influence of substrate salinity (0-40 parts per thousand, ppt) and pH (pH 1-13) on oviposition, egg hatching, larval survivability, and adult emergence of Culicoides peregrinus Kieffer under laboratory conditions. Most eggs (80.74%) were laid in 0 ppt and 95% in pH 7 but lowered with increased salinity and pH levels. It was observed that the females did not lay eggs in 30 ppt to 40 ppt salinity; pH 1 and pH 13 but interestingly up to 95% of the eggs were retained within the abdomen. Little effect of salinity and pH on egg hatching was observed up to 5 ppt and 10 ppt except at the extreme values of 40 ppt and pH 1, pH 13. Pupation did not occur in rearing plates with high salinities, 30 ppt and 40 ppt, although the few eggs hatched when exposed to such salinity. In low salinity (0 to 2 ppt), occurrence of adult emergence was more and then decreased with increasing salinity. Maximum emergence was seen when the rearing media was alkaline. This study deals with the suitability of breeding substrate of C. peregrinus when exposed to salinity and pH ranges. Our study suggests the ambient salinity and pH ranges to be maintained during laboratory rearing of this vector species.

BLUETONGUE VIRUS SEROTYPE 16 DETECTION IN NSW: In coastal New South Wales (NSW), bluetongue virus (BTV) serotypes 1 and 21 are endemic and transmitted in most years without evidence of disease. However, serotype 16 (BTV-16) infection was detected for the first time in NSW in November 2016 in cattle undergoing testing for export. Retrospective testing of blood samples collected from sentinel cattle as part of the National Arbovirus Monitoring Program (NAMP) established that the first detected transmission of BTV-16 in NSW occurred in April 2016 in sentinel cattle on the NSW North Coast. Subsequently, until 2022, BTV-16 has been transmitted in most years and was the predominant serotype in the 2018-2019 transmission season. The data available suggests that BTV-16 may have become endemic in NSW. EXPERIMENTAL STUDIES: During experimental infection studies with BTV-16, all sheep were febrile, with the peak of viremia occurring 6-10 days after inoculation. There was nasal and oral hyperaemia in most sheep with several animals developing a nasal discharge and nasal oedema. All sheep developed coronitis of varying severity, with most also developing haemorrhages along the coronary band. There was a high incidence of haemorrhage in the pulmonary artery, epicardial petechiae, extensive pericardial haemorrhages and moderate body cavity effusions including pericardial effusions. CONCLUSION: Overall, experimental pathogenicity findings suggest moderate disease may occur in sheep in the field. These findings, when combined with climatic variability that could result in an expansion of the range of Culicoides brevitarsis into major sheep-producing areas of the state, suggest that there is an increasing risk of bluetongue disease in NSW., (© 2023 The Authors. Australian Veterinary Journal published by John Wiley & Sons Australia, Ltd on behalf of Australian Veterinary Association.)

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., (© 2023. BioMed Central Ltd., part of Springer Nature.)

Gut bacterial communities in insects provide several beneficial roles like nutrition, digestion, fecundity, and survival of the host. The microbial communities of Culicoides spp. (Diptera: Ceratopogonidae) vary with parity, developmental stages, and environmental factors. Previous studies have revealed the presence of hemolytic bacteria in adult Culicoides peregrinus Kieffer (Diptera: Ceratopogonidae), an important vector of bluetongue virus (BTV). Our objectives were (i) to identify bacterial communities with hemolytic activities associated with all life stages and (ii) to compare between reared and field-collected adults including age graded females. Bacterial identification followed Sanger sequencing of 16S rRNA. In vitro biochemical characterizations including antibiotic sensitivity tests were also done. The majority of bacterial species were beta hemolytic with one, Alcaligenes faecalis, showing alpha hemolysis. Most bacterial species were observed in field-collected adults except Proteus spp. Throughout the life history of the vector, Bacillus cereus (CU6A, CU1E) and Paenibacillus sp. (CU9G) were detected indicating their possible role in blood digestion within the gut of this vector species. In vivo hemolytic activities of these culturable bacterial communities within this vector may be addressed in future. These hemolytic bacterial communities may be targeted to develop novel and effective strategies for vector control., (© The Author(s) 2023. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Culicoides biting midges (Diptera: Ceratopogonidae) are the main vectors of livestock diseases such as bluetongue (BT) which mainly affect sheep and cattle. In Spain, bluetongue virus (BTV) is transmitted by several Culicoides taxa, including Culicoides imicola , Obsoletus complex, Culicoides newsteadi and Culicoides pulicaris that vary in seasonality and distribution, affecting the distribution and dynamics of BT outbreaks. Path analysis is useful for separating direct and indirect, biotic and abiotic determinants of species' population performance and is ideal for understanding the sensitivity of adult Culicoides dynamics to multiple environmental drivers. Start, end of season and length of overwintering of adult Culicoides were analysed across 329 sites in Spain sampled from 2005 to 2010 during the National Entomosurveillance Program for BTV with path analysis, to determine the direct and indirect effects of land use, climate and host factor variables. Culicoides taxa had species-specific responses to environmental variables. While the seasonality of adult C. imicola was strongly affected by topography, temperature, cover of agro-forestry and sclerophyllous vegetation, rainfall, livestock density, photoperiod in autumn and the abundance of Culicoides females, Obsoletus complex species seasonality was affected by land-use variables such as cover of natural grassland and broad-leaved forest. Culicoides female abundance was the most explanatory variable for the seasonality of C. newsteadi , while C. pulicaris showed that temperature during winter and the photoperiod in November had a strong effect on the start of the season and the length of overwinter period of this species. These results indicate that the seasonal vector-free period (SVFP) in Spain will vary between competent vector taxa and geographic locations, dependent on the different responses of each taxa to environmental conditions.

Epizootic hemorrhagic disease (EHD) is an infectious viral disease caused by epizootic hemorrhagic disease virus (EHDV) and EHDV frequently circulates in wild and domestic ruminants. Sporadic outbreaks of EHD have caused thousands of deaths and stillbirths on cattle farms. However, not much is known about the circulating status of EHDV in Guangdong, southern China. To estimate the seroprevalence of EHDV in Guangdong province, 2886 cattle serum samples were collected from 2013 to 2017 and tested for antibodies against EHDV using a competitive ELISA. The overall seroprevalence of EHDV reached 57.87% and was highest in autumn (75.34%). A subset of positive samples were serotyped by a serum neutralization test, showing that EHDV serotypes 1 and 5-8 were circulating in Guangdong. In addition, EHDV prevalence always peaked in autumn, while eastern Guangdong had the highest EHDV seropositivity over the five-year period, displaying apparent temporal-spatial distribution of EHDV prevalence. A binary logistic model analysis indicated a significant association between cattle with BTV infections and seroprevalence of EHDV (OR = 1.70, p < 0.001). The co-infection of different serotypes of EHDV and BTV raises a high risk of potential genomic reassortment and is likely to pose a significant threat to cattle, thus urging more surveillance to monitor their circulating dynamics in China.

Documenting the host use of vector species is important for understanding the transmission dynamics of vector-borne pathogens. Biting midges (Diptera: Ceratopogonidae: Culicoides) are vectors of epizootic hemorrhagic disease virus (EHDV) and bluetongue virus (BTV) worldwide. However, relative to mosquitoes and many other vector groups, host associations of this group are poorly documented. In this study, we used PCR-based bloodmeal analysis to determine species-level host associations of 3,603 blood-engorged specimens belonging to 18 Culicoides species at 8 deer farms in Florida, USA. We used a binomial mixed model with a Bayesian framework to compare the effect of host composition on the feeding patterns of Culicoides spp. and employed the Morisita-Horn Index to investigate the similarity of host use between farms for Culicoides stellifer and Culicoides insignis. Results show that the estimated probability of Culicoides spp. feeding upon white-tailed deer depends on the availability of cattle or exotic game and demonstrates differences in host-feeding selection among species. Culicoides insignis had high host similarity across farms suggesting that its host-use patterns are somewhat conserved. Culicoides stellifer had lower host similarity across farms suggesting that it is a more opportunistic feeder. White-tailed deer are fed upon by many Culicoides species on deer farms in Florida, and while most Culicoides species feed on white-tailed deer, the ratio of white-tailed deer bloodmeals to other bloodmeals is likely influenced by host availability. Culicoides spp. taking a majority of their bloodmeals from farmed white-tailed deer should be assessed for their vector competence for EHDV and BTV., (© The Author(s) 2023. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

During larval rearing of Culicoides peregrinus Kieffer (Diptera: Ceratopogonidae) it was obligatory to add a small quantity of mud from larval habitat to nutrient broth in culture plates. This initiated microbial growth in rearing plates which facilitated growth and development of immature. The primary aim was to enumerate gut microbial communities across the different life stages of C. peregrinus. Amplicon sequencing of the V3-V4 hypervariable region (16S rDNA) was done on Illumina Miseq platform to detect gut bacterial communities at different life stages, while ITS regions (18S rRNA) were targeted for fungal communities of the 4th instar larvae. The major findings were: 1) Phylum Proteobacteria and Firmicutes were the most abundant throughout the life stages, along with the highest bacterial alpha diversity in the egg, 2) bacterial compositions were similar to laboratory reared and field collected adults, and 3) abundant fungal phyla associated with the larval gut were Ascomycota and Basidiomycota. Furthermore, analyses of the gut microbiome with METAGENassist might be indicative of their likely function in the natural habitat. Abundant gut-associated bacteria and/or fungal genera detected in the present study could be used as dietary supplements to establish laboratory colonies for further vectorial research. While, individual roles of the bacteria or fungi in paratransgenesis are warned for their possible utilization to frame the management strategy in upcoming works., (© The Author(s) 2023. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Bluetongue (BT), once considered a disease of sheep confined to the southern African region, has spread all over the world. BT is a viral disease caused by the bluetongue virus (BTV). BT is regarded as an economically important disease in ruminants of compulsory notification to OIE. BTV is transmitted by the bite of Culicoides species. Research over the years has led to a better understanding of the disease, the nature of the virus life cycle between ruminants and Culicoides species, and its distribution in different geographical regions. Advances have also been made in understanding the molecular structure and function of the virus, the biology of the Culicoides species, its ability to transmit the disease, and the persistence of the virus inside the Culicoides and the mammalian hosts. Global climate change has enabled the colonization of new habitats and the spread of the virus into additional species of the Culicoides vector. This review highlights some of the current findings on the status of BT in the world based on the latest research on disease aspects, virus-host-vector interactions, and the different diagnostic approaches and control strategies available for BTV., Competing Interests: No Conflict of Interest is declared, (Copyright (c) 2023 Subhra Subhadra, Daggupati Sreenivasulu, Ritesh Pattnaik, Basant Kumar Panda, Subrat Kumar.)

Knowledge gaps exist on the feeding pattern and host range of bluetongue virus vectors, Culicoides species, associated with livestock in India. Adult midges were trapped with ultraviolet light traps at 13 household farms adjacent to human biotope. Host DNA was isolated from individual females (n = 101; blood engorged-82, gravid-4 and parous-15) and subjected to PCR amplification targeting CytB and 16S rRNA gene fragments followed by sequencing of amplified DNA samples. However, DNA sequences from only 71 individuals (70.3%) comprising of 10 Culicoides species were obtained. Blood meal analysis revealed at least 10 species that fed on five mammalian hosts including humans, but surprisingly none tested positive for birds. Results revealed that Culicoides innoxius tested positive for four not previously recognized species indicating a potential role as a vector species. Likewise, Culicoides shortti and Culicoides hegneri preferred goat and cattle respectively as hosts, whereas Culicoides palpifer preferred cattle along with buffalo as hosts, which is being reported for the first time. This is the first document on DNA-based blood meal identification and feeding preference of Culicoides midges associated with livestock in India., (© 2022 Royal Entomological Society.)

In the southeastern United States, biting midges transmit agents of hemorrhagic diseases that are enzootic among white-tailed deer (Odocoileus virginianus (Zimmermann), Artiodactyla: Cervidae). Culicoides sonorensis Wirth and Jones (Diptera: Ceratopogonidae), the only confirmed vector of epizootic hemorrhagic disease virus (EHDV) and bluetongue virus (BTV) in the United States, is rarely collected in the Southeast, implying that other Culicoides Latreille species act as vectors. Despite multiple surveillance studies, the influence of trapping habitat and light wavelength on Culicoides sampling has yet to be investigated in Alabama. This study sampled Culicoides species at a deer research facility using CO2-baited CDC light traps with three distinct wavelengths. Traps were rotated within three habitats to examine impacts of habitat type and light wavelength on Culicoides abundance and parity status. For most species, midges were more abundant in a pine forest compared to a hardwood-forest riparian zone or a lightly wooded area adjacent to a seasonal pond. The pine forest generally had negative effects on parity status, suggesting that most females in this habitat were foraging for their first bloodmeal. Ultraviolet (UV) black-light (350 nm-360 nm) attracted more midges than incandescent light or UV LED light (385 nm-395 nm), but wavelength had less of an effect on parity than habitat. This study indicates that light wavelength and habitat significantly influence Culicoides sampling outcomes, and that when collecting parous females is desired (e.g., EHDV/BTV surveillance), targeting areas around oviposition sites may be a better strategy than trapping where midges are most abundant., (© The Author(s) 2022. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Objective: Bluetongue is an arthropod-borne disease of ruminants. Here, we investigated the seroprevalence of bluetongue virus (BTV) in livestock and performed the first genetic characterization of BTV isolated from sheep and Culicoides midges in the southeastern region of Kazakhstan., Methods: Blood samples were collected from 1241 asymptomatic livestock. In addition, 497 Culicoides midges were collected. Samples were analyzed for specific anti-BTV antibodies and BTV RNA by ELISA and conventional RT-PCR, respectively., Results: The overall seroprevalence of BTV antibodies was shown to be 4.3 % (46/1079) in small ruminant and 1 % (1/82) in cattle. Anti-BTV antibodies were not detected in camels (0/80). The minimum infection rate of BTV in Culicoides was shown to be 0.24 %. Seg-2 and Seg-10 sequence analysis demonstrated that all isolates belonged to the 'western' topotype of the BTV-9 strain., Conclusion: The present data confirm circulation of BTV in southeastern Kazakhstan., Competing Interests: Declaration of Competing Interest The authors declare no conflict of interest., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

Determination of host choice of Culicoides species (Diptera: Ceratopogonidae), the vectors of bluetongue virus (BTV), is pivotal to ascertain the role of each species in the transmission of pathogens, pest management and enumeration of disease prediction models. Host preference of livestock associated Culicoides midges was investigated in West Bengal, India with four replicates of a 3 × 3 Latin square design during August and September 2021. Adult Culicoides were mouth aspirated from three BTV hosts viz., cattle, sheep and goats. Mouth aspirating was validated by the sweep net collections. The host-baited collections recorded seven Culicoides species; with the highest landing rate on cattle (n = 5,667; 92.9%) followed by sheep (n = 365; 6.0%) and goat (n = 67; 1.1%). Based on the Jacob's selectivity index, all midge species, except for Culicoides fulvus Sen & Das Gupta, encountered, preferred cattle over other mammalian hosts. Culicoides oxystoma Kieffer, the subgenus Trithecoides Wirth & Hubert and Culicoides actoni Smith, predominated on the ventral region (belly/flank) of the cattle. However, Culicoides peregrinus Kieffer and C. actoni were observed to be prevalent in the leg region of sheep. A significantly higher percentage of female (99.9%) with only 0.3% of male were trapped in aspiration based animal baited collections. On the other hand sweep net and light trap catch comprises of 50.7%, 89.7% female and 49.2%, 10.2% male respectively. Surprisingly, DNA based blood meal analysis revealed human blood from the midges trapped in UV-LED light traps. Supplying the first evidence that Culicoides similis Carter, Ingram & Macfie, C. fulvus and Culicoides palpifer Das Gupta & Ghosh, feed on humans., Competing Interests: Declaration of Competing Interest There is no conflict of interests amongst authors., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Culicoides biting midges are well known biological vectors of several arboviruses causing more than 100 veterinary and medical diseases worldwide. In Algeria, bluetongue virus, which is transmitted by Culicoides midges, is responsible for one of the most critical insect-borne diseases of sheep. For example, this disease caused clinically severe morbidity of about 2,661 confirmed cases out of 21,175 susceptible sheep in Algeria. This study compared the abundance, richness, and diversity of Culicoides species in rural and suburban environments of semi-arid regions in North Africa. It examined the potential influence of the bio-climatic factors on the variation of population sizes and the elevation distribution of biting midges. Specimen collection was carried out from June to September during two successive years (2018 and 2019) using CDC light traps installed at 15 sites in different environments. Culicoides specimens were dissected, slide-mounted, and morphologically identified using the interactive identification key IIKC and various standard morphological criteria. A total of 1,046 Culicoides specimens (1,024 females and 22 males) were trapped and classified into 22 species, belonged to nine distinct subgenera. Two new species records for Algeria and even North Africa are reported: Culicoides albicans (Winnertz, 1852) and Culicoides nubeculosus (Meigen, 1830). Culicoides newsteadi Austen, 1921 (51.6%) was the dominant species, and it was followed by Culicoides punctatus (Meigen, 1804) (16.3%) and Culicoides odiatus Austen, 1921 (11.5%). These three species, comprising 80% of the collected Culicoides, were the most abundant both outside and inside livestock stables in rural and suburban environments. Species diversity was similar in the two settings, with a slight increase in suburban environments. None of the Culicoides species encountered correlated significantly with the climatic factors (mean temperature, precipitation, and relative humidity). Elevation was the most determinant environmental parameter that affected the abundance and distribution of Culicoides midges in the semi-arid and sub-humid areas studied. The maximum distribution of Culicoides species was detected at mid elevations (400‒800 m). Using a modeling approach, we explored for the first time the variation of composition and diversity in Culicoides communities within different climatic regions, environments and livestock settings in Algeria. This survey deepens our understanding of the relationships among environmental factors, abundance, diversity, and geographic distribution of Culicoides. This is a crucial step to assess the epidemiological situation of the diseases transmitted by these biting midges and to allow mitigation of the associated risks., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Culicoides Latreille (Diptera: Ceratopogonidae) biting midges are vectors of important animal pathogens including bluetongue virus (BTV) and epizootic hemorrhagic disease virus (EHDV). While some Culicoides species present in the southern California desert are implicated in the transmission of these viruses to ruminant animals, these species have not been extensively studied due in part to the challenge of identifying Culicoides to species and to the lack of published gene sequences for these species to support their molecular identification. In this study, Culicoides were captured using suction traps baited with either carbon dioxide or UV light from transitional habitat between the southern California peninsular mountain ranges and the Colorado desert of southeastern California. Captured midges were initially identified using traditional morphological methods, with species identification subsequently confirmed by sequence analysis of COI and 28S rDNA genes. Phylogenetic analyses support that some Culicoides subgenera are not monophyletic. Two recognized species (C. sitiens Wirth and Hubert and C. bakeri Vargas) shared the same COI and 28S sequences. An additional cryptic species may be present within C. sitiens. Two additional recognized species (C. cacticola Wirth and Hubert and C. torridus Wirth and Hubert) may be conspecific or cryptic to each other. In total, 19 Culicoides species (or species aggregate) were collected in this study, with genetic sequences published for the first time for 16 of them. Published genetic sequences will support future research on these species, including studies on the ecology and habits of their immature stages which are often tedious to identify using morphology., (© The Author(s) 2022. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

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., (© 2022 The Authors. Medical and Veterinary Entomology published by John Wiley & Sons Ltd on behalf of Royal Entomological Society.)

Bluetongue virus (BTV) and African horse sickness virus (AHSV) cause economically important diseases that are currently exotic to the United Kingdom (UK), but have significant potential for introduction and onward transmission. Given the susceptibility of animals kept in zoo collections to vector-borne diseases, a qualitative risk assessment for the introduction of BTV and AHSV to ZSL London Zoo was performed. Risk pathways for each virus were identified and assessed using published literature, animal import data and outputs from epidemiological models. Direct imports of infected animals, as well as wind-borne infected Culicoides , were considered as routes of incursion. The proximity of ongoing disease events in mainland Europe and proven capability of transmission to the UK places ZSL London Zoo at higher risk of BTV release and exposure (estimated as low to medium ) than AHSV (estimated as very low to low ). The recent long-range expansion of AHSV into Thailand from southern Africa highlights the need for vector competence studies of Palearctic Culicoides for AHSV to assess the risk of transmission in this region.

Bluetongue virus (BTV) is an arthropod-borne, segmented double-stranded RNA virus that can cause severe disease in both wild and domestic ruminants. BTV evolves via several key mechanisms, including the accumulation of mutations over time and the reassortment of genome segments.Additionally, BTV must maintain fitness in two disparate hosts, the insect vector and the ruminant. The specific features of viral adaptation in each host that permit host-switching are poorly characterized. Limited field studies and experimental work have alluded to the presence of these phenomena at work, but our understanding of the factors that drive or constrain BTV's genetic diversification remains incomplete. Current research leveraging novel approaches and whole genome sequencing applications promises to improve our understanding of BTV's evolution, ultimately contributing to the development of better predictive models and management strategies to reduce future impacts of bluetongue epizootics.

Culicoides (Diptera, Ceratopogonidae) are small biting midges, some of which are vectors for animal associated arboviruses such as bluetongue virus (BTV) and Akabane virus (AKAV). BTV and AKAV are both pathogenic for livestock, with BTV in particular posing a major threat to domestic ruminants. Ongoing problems with BTV in ruminants in Shizong County of Yunnan Province, China, promoted a year-long investigation of the Culicoides in Shizong to determine relative abundance and seasonality of midges and to attempt to identify species that might be acting as vectors in the area. Culicoides were collected by UV light trap for one night per week for most weeks between May 2020 and May 2021. More than 21,000 specimens consisting of at least 21 species belonging to six subgenera and one unplaced group, including 5 species previously associated with BTV and one associated with AKAV, were collected. Culicoides tainanus dominated most collections throughout the year although C. sumatrae was often the dominant species over summer. Most species were abundant between May and October. These results indicate that C. tainanus, C. jacobsoni and C. oxystoma are the major midge pests of livestock in Shizong and should be considered in any disease investigation., (© Y.L. Duan et al., published by EDP Sciences, 2022.)

We investigated the Culicoides fauna in Turkey during the years 2016-2019 in the process of entomological surveillance for arboviral diseases. The entomological survey was conducted at 104 sampling stations in 51 provinces in Turkey during four consecutive years. There were approximately 450,000 specimens and 59 identified species collected during the surveillance. Ten species were newly recorded for Turkey: C . chiopterus , C. grisescens , C. paradoxalis , C. santonicus , C. poperinghensis , C. sergenti , C. tbilisicus , C. comosioculatus , C. haranti , and C. univittatus . Identification of C . chiopterus and C. grisescens was confirmed using species-specific PCR and DNA sequencing. With our recent findings, previous data were critically reviewed and updated, and the number of Culicoides species has been increased to 71 for Turkey. The presence of C. chiopterus has particular importance due to its potential vector status for bluetongue virus (BTV) and Schmallenberg virus (SBV). This study presents result of the first large-scale integrated faunistic survey on Culicoides species in Turkey.

Biting midges of the genus Culicoides Latreille are biological vectors of bluetongue virus (BTV), a member of family Reoviridae, genus Orbivirus. About 30 species of Culicoides have been identified as competent BTV vectors worldwide. Even though high seroprevalence of BTV has been reported among livestock ruminants from western Thailand, the Culicoides species which contribute to BTV transmission remain unclear. In the present study, Culicoides were collected from eight sampling sites, located in two BTV prevalent provinces in western Thailand. Adult Culicoides were identified using wing morphology and cytochrome c oxidase subunit I (COI) mtDNA molecular marker. A total of 9,677 Culicoides specimens belonging to 7 subgenera, 3 species groups, and 23 species were identified. After comparing sequencing results with available data from GenBank, COI sequences of five species were reported for the first time from Thailand. The most abundant potential BTV vector species collected were C. peregrinus, followed by C. orientalis, C. imicola, C. oxystoma, and C. fulvus. Out of 72 Culicoides pools, 9 pools (4 from C. orientalis, 2 from C. imicola, 2 from C. oxystoma, and 1 from C. fulvus) were positive by BTV RT-PCR analyses. These results are new to Culicoides BTV vector knowledge in Thailand and will contribute to further BTV studies in this particular region., (Copyright © 2021. Published by Elsevier B.V.)

Background: Bluetongue is a serious disease of ruminants caused by the bluetongue virus (BTV). BTV is transmitted by biting midges (Culicoides spp.). Serological evidence from livestock and the presence of at least one competent vector species of Culicoides suggests that transmission of BTV is possible and may have occurred in Kazakhstan., Methods: We estimated the risk of transmission using a mathematical model of the reproduction number R 0 for bluetongue. This model depends on livestock density and climatic factors which affect vector density. Data on climate and livestock numbers from the 2466 local communities were used. This, together with previously published model parameters, was used to estimate R 0 for each month of the year. We plotted the results on isopleth maps of Kazakhstan using interpolation to smooth the irregular data. We also mapped the estimated proportion of the population requiring vaccination to prevent outbreaks of bluetongue., Results: The results suggest that transmission of bluetongue in Kazakhstan is not possible in the winter from October to March. Assuming there are vector-competent species of Culicoides endemic in Kazakhstan, then low levels of risk first appear in the south of Kazakhstan in April before spreading north and intensifying, reaching maximum levels in northern Kazakhstan in July. The risk declined in September and had disappeared by October., Conclusion: These results should aid in surveillance efforts for the detection and control of bluetongue in Kazakhstan by indicating where and when outbreaks of bluetongue are most likely to occur. The results also indicate where vaccination efforts should be focussed to prevent outbreaks of disease., (© 2021. The Author(s).)

Background: Culicoides (Diptera: Ceratopogonidae) are vectors for many arboviruses. At least 20 species are considered as vectors or potential vectors of bluetongue virus (BTV) which cause bluetongue disease in ruminants. A BTV prevalence of 30-50% among cattle and goats in tropical southern Yunnan Province, China, prompted an investigation of the potential BTV vectors in this area., Methods: Culicoides were collected by light trapping at three sites in the tropical region of Yunnan Province. Species were identified based on morphology and DNA sequences of cytochrome c oxidase subunit 1 (cox1). PCR and quantitative PCR following reverse transcription were used to test for the presence of BTV RNA in these specimens. Phylogenetic analysis was used to analyze the cox1 sequences of Culicoides specimens infected with BTV., Results: Approximately 67,000 specimens of Culicoides were collected, of which 748 were tested for the presence of BTV. Five specimens, including two of Culicoides jacobsoni, one of C. tainanus and two of C. imicola, were identified as infected with BTV. No specimens of C. (subgenus Trithecoides) or C. oxystoma tested were positive for BTV infection., Conclusions: To our knowledge this is the first report of C. jacobsoni as a potential BTV vector and the fourth report of an association between C. tainanus and BTV, as well as the first direct evidence of an association between BTV and C. imicola in Asia. A fourth potential cryptic species within C. tainanus was identified in this study. Further analysis is required to confirm the importance of C. jacobsoni and C. tainanus in BTV epidemiology in Asia.

Bluetongue virus (BTV) causes a disease that is endemic in Spain and its two major biological vector species, C. imicola and the Obsoletus complex species, differ greatly in their ecology and distribution. Understanding the seasonality of BTV transmission in risk areas is key to improving surveillance and control programs, as well as to better understand the pathogen transmission networks between wildlife and livestock. Here, monthly risk transmission maps were generated using risk categories based on well-known BTV R 0 equations and predicted abundances of the two most relevant vectors in Spain. Previously, Culicoides spp. predicted abundances in mainland Spain and the Balearic Islands were obtained using remote sensing data and random forest machine learning algorithm. Risk transmission maps were externally assessed with the estimated date of infection of BTV-1 and BTV-4 historical outbreaks. Our results highlight the differences in risk transmission during April-October, June-August being the period with higher R 0 values. Likewise, a natural barrier has been identified between northern and central-southern areas at risk that may hamper BTV spread between them. Our results can be relevant to implement risk-based interventions for the prevention, control and surveillance of BTV and other diseases shared between livestock and wildlife host populations.

Bluetongue virus (BTV) and epizootic hemorrhagic disease virus (EHDV) cause hemorrhagic disease (HD) in wild ruminants and bluetongue disease (BT) and epizootic hemorrhagic disease (EHD) in livestock. These viruses are transmitted by biting midges in the genus Culicoides (family Ceratopogonidae). Mortality from this disease can reach 90% in certain breeds of sheep and in white-tailed deer (Odocoileus virginianus). From January until December of 2012, we conducted a prospective study to determine the origin and routes of transmission of BTV and EHDV in captive deer and cattle. The objective was to determine the abundance of Culicoides spp. and BTV/EHDV infection prevalence in midges, cattle, and deer in an area experiencing an outbreak of BT and EHD. Agar gel immunodiffusion (AGID) tests to detect for EHDV and BTV antibodies were conducted on serum collected from cattle and deer, quantitative reverse transcriptase polymerase chain reaction (RT-qPCR) was utilized for BTV/EHDV RNA detection in tissues from dead deer, and CDC miniature black light traps baited with dry ice were deployed to capture insects. The AGID results showed 19 out of 29 cattle and 18 out of 58 white-tailed deer seroconverted for these viruses during the vector season. Tradition gel-based reverse transcriptase polymerase chain reaction was utilized to determine serotype. Sixteen cows were positive for EHDV-2, EHDV-6, or BTV-12 and 15 deer positive for EHDV-1, EHDV-6, or BTV-12. Specimens from 14 species of Culicoides (Dptera: Ceratopogonidae) (Culicoides arboricola Root and Hoffman, Culicoides biguttatus Coquillett, Culicoides crepuscularis Malloch, Culicoides debilipalpis Lutz, Culicoides furens Poey, Culicoides haematopotus Malloch, Culicoides hinmani Khalaf, Culicoides nanus Root and Hoffman, Culicoides neopulicaris Wirth, Culicoides paraensis Goeldi, Culicoides stellifer Coquillet, Culicoides variipennis Coquillet, Culicoides villosipennis Root and Hoffman, and Culicoides venustus Hoffman) were captured and tested for BTV and EHDV using RT-qPCR assays. BTV viral nucleic acid was detected in three pools from three different species of midges: C. crepuscularis, C. debilipalpis, and C. stellifer., (© The Author(s) 2020. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

We determined the temporal aspects of detecting bluetongue virus (BTV) and epizootic hemorrhagic disease virus (EHDV) in postmortem bone marrow samples of white-tailed deer ( Odocoileus virginianus ) using molecular and in vitro cell culture techniques. Bone marrow samples from carcasses were collected and assayed on the day of death and at intervals up to 16 wk after death. We recovered BTV and EHDV from fresh bone marrow collected at day 0 by isolation in Vero and BHK-21 cell cultures. However, attempts to replicate the viruses from aged bone marrow in Vero and BHK-21 cell cultures failed. The real-time quantitative reverse transcriptase PCR (qRT-PCR) results confirmed that EHDV and BTV can be detected in aged bone marrow for up to 12 and 16 wk, respectively, after death. The RNA of BTV and EHDV could be detected by qRT-PCR in white-tailed deer bone marrow for extended periods of time postmortem. This technique will provide a useful tool for retrospective determination of BTV or EHDV infection of white-tailed deer at the time of death.

Background: Vector-borne diseases are among the leading causes of morbidity and mortality in humans and animals. In the Afrotropical region, some are transmitted by Culicoides, such as Akabane, bluetongue, epizootic haemorrhagic fever and African horse sickness viruses. Bluetongue virus infection has an enormous impact on ruminant production, due to its high morbidity and mortality rates., Methods: A nationwide Culicoides trapping campaign was organized at the end of the 2012 rainy season in Senegal. A Maximum Entropy approach (MaxEnt), Boosted Regression Tree (BRT) method and Ecological Niche Factor Analysis (ENFA) were used to develop a predictive spatial model for the distribution of Culicoides, using bio-climatic variables, livestock densities and altitude., Results: The altitude, maximum temperature of the warmest month, precipitation of the warmest quarter, mean temperature of the wettest quarter, temperature seasonality, precipitation of the wettest quarter and livestock density were among the most important factors to predict suitable habitats of Culicoides. Culicoides occurrences were, in most of the cases, positively correlated to precipitation variables and livestock densities; and negatively correlated to the altitude and temperature indices. The Niayes area and the Groundnut basin were the most suitable habitats predicted., Conclusion: We present ecological niche models for different Culicoides species, namely C. imicola, C. oxystoma, C. enderleini and C. miombo, potential vectors of bluetongue virus, on a nationwide scale in Senegal. Through our modelling approach, we were able to determine the effect of bioclimatic variables on Culicoides habitats and were able to generate maps for the occurrence of Culicoides species. This information will be helpful in developing risk maps for disease outbreaks.

Background: Transmission of vector-borne virus by insects is a complex mechanism consisting of many different processes; viremia in the host, uptake, infection and dissemination in the vector, and delivery of virus during blood-feeding leading to infection of the susceptible host. Bluetongue virus (BTV) is the prototype vector-borne orbivirus (family Reoviridae). BTV serotypes 1-24 (typical BTVs) are transmitted by competent biting Culicoides midges and replicate in mammalian (BSR) and midge (KC) cells. Previously, we showed that genome segment 10 (S10) encoding NS3/NS3a protein is required for virus propagation in midges. BTV serotypes 25-27 (atypical BTVs) do not replicate in KC cells. Several distinct BTV26 genome segments cause this so-called 'differential virus replication' in vitro., Methods: Virus strains were generated using reverse genetics and their growth was examined in vitro. The midge feeding model has been developed to study infection, replication and disseminations of virus in vivo. A laboratory colony of C. sonorensis, a known competent BTV vector, was fed or injected with BTV variants and propagation in the midge was examined using PCR testing. Crossing of the midgut infection barrier was examined by separate testing of midge heads and bodies., Results: A 100 nl blood meal containing ±10 5.3 TCID 50 /ml of BTV11 which corresponds to ±20 TCID 50 infected 50% of fully engorged midges, and is named one Midge Alimentary Infective Dose (MAID 50 ). BTV11 with a small in-frame deletion in S10 infected blood-fed midge midguts but virus release from the midgut into the haemolymph was blocked. BTV11 with S1[VP1] of BTV26 could be adapted to virus growth in KC cells, and contained mutations subdivided into 'corrections' of the chimeric genome constellation and mutations associated with adaptation to KC cells. In particular one amino acid mutation in outer shell protein VP2 overcomes differential virus replication in vitro and in vivo., Conclusion: Small changes in NS3/NS3a or in the outer shell protein VP2 strongly affect virus propagation in midges and thus vector competence. Therefore, spread of disease by competent Culicoides midges can strongly differ for very closely related viruses.

Starting in 2006, bluetongue virus serotype 8 (BTV8) was responsible for a major epizootic in Western and Northern Europe. The magnitude and spread of the disease were surprisingly high and the control of BTV improved significantly with the marketing of BTV8 inactivated vaccines in 2008. During late summer of 2011, a first cluster of reduced milk yield, fever, and diarrhoea was reported in the Netherlands. Congenital malformations appeared in March 2012 and Schmallenberg virus (SBV) was identified, becoming one of the very few orthobunyaviruses distributed in Europe. At the start of both epizootics, little was known about the pathogenesis and epidemiology of these viruses in the European context and most assumptions were extrapolated based on other related viruses and/or other regions of the World. Standardized and repeatable models potentially mimicking clinical signs observed in the field are required to study the pathogenesis of these infections, and to clarify their ability to cross the placental barrier. This review presents some of the latest experimental designs for infectious disease challenges with BTV or SBV. Infectious doses, routes of infection, inoculum preparation, and origin are discussed. Particular emphasis is given to the placental crossing associated with these two viruses.

In October 2017, the first outbreak of bluetongue virus serotype 3 (BTV-3) began in Italy, specifically in western Sicily. The route of entrance remains unclear, although since 2016 the same strain had been circulating only 150 km away, on the Tunisian peninsula of Cape Bon. The present analysis assessed the feasibility that wind could have carried BTV-3-infected Culicoides spp. from Tunisia to Sicily. An advection-deposition-survival (ADS) model was used to estimate when and where Culicoides spp. were likely to be introduced prior to the first BTV-3 report in Italy. Additionally, the Hybrid Single-Particle Lagrangian Integrated Trajectory (HYSPLIT) model was used to support ADS outputs. The modelling suggests that during September 2017, strong wind currents and suitable climatic conditions could have allowed the transportation of Culicoides spp. from BTV-3-infected areas in Tunisia into Sicily. ADS simulations suggest that particles could have reached the province of Trapani in western Sicily on 2 and 12 September. These simulations suggest the feasibility of aerial transportation of infected Culicoides spp. from Tunisia into Sicily. They demonstrate the suitability of the ADS model for retrospective studies of long-range transportation of insects across large water bodies, which may enhance the early detection of vectorial disease introduction in a region., (© 2019 The Authors. Transboundary and Emerging Diseases published by Blackwell Verlag GmbH.)

Bluetongue is an infectious disease transmitted by Culicoides biting midges. Culicoides imicola is considered the main vector in the Mediterranean basin but other species have been implicated in the Bluetongue virus (BTV) transmission. During 2017, BTV serotype 4 re-occurred in Sardinia causing outbreaks in sheep farms. A survey was carried out on affected farms with the aim to detect the virus in field-collected Culicoides Biting midges were morphologically identified, pooled and then assayed with a real time RT-PCR. To evaluate BTV dissemination, some Culicoides were dissected and head, thorax and abdomen were tested singly by PCR. A total of 173,738 Culicoides adults were collected. Viral RNA was detected in 68 out of 77 pools and all species analysed resulted positive. Detection of BTV in parous female body regions (head, thorax and abdomen) confirmed the full dissemination of BTV in all species analysed. During this study, the vector competence of C imicola , C newsteadi s.l. and Obsoletus complex was confirmed. The authors found two new Culicoides species BTV positive, C paolae never associated with BTV transmission and C circumscriptus only recently found BTV positive in Turkey, which could be considered potential vectors., Competing Interests: Competing interests: None declared., (© British Veterinary Association 2019. No commercial re-use. See rights and permissions. Published by BMJ.)

Background: Several European countries suffered important economic losses during the past decade due to the emergence of bluetongue and Schmallenberg viruses. Both are viruses of veterinary importance and are spread by Culicoides spp. This triggered many European countries to start Culicoides population monitoring. Recently a one year monitoring study at 16 sites in Belgium revealed that important variation existed in Culicoides abundance and species diversity between collection sites. In order to analyze whether this variation is consistent over years, a detailed analysis of monitoring data collected at seven locations in Belgium between 2007 and 2011 was performed in this study. At all locations, biting midges were collected with OVI black light traps set-up in close proximity to livestock., Results: In total, 42 different Culicoides species were morphologically identified. Species of the subgenus Avaritia represented 83% of all collected midges. Nevertheless, important differences in species composition were found between sites. Furthermore, statistical differences between sites were found for the total and maximum annual abundance, showing that a consistent higher or lower number of Culicoides could be collected depending on the selected collection site. Yearly, up to 16 and 30-fold differences in total and maximum annual abundances between sites, respectively, were found. Also the month in which most Culicoides were collected varied greatly between years, both at local (from May to October) and country level [May (2008), June (2010), July (2009), August (2011), October (2007)]. Finally, the average vector-free period over all sites and years was 173 days and could roughly be defined between November and the end of April. Interestingly, important yearly variations of up to two months in the duration of the vector-free period were found between the studied collection sites. In contrast to the abundance parameters, no specific sites could however be identified where monitoring consistently showed shorter or longer vector-free periods., Conclusions: In conclusion, our results show that the selection of collection sites for Culicoides monitoring, even in a small country such as Belgium, strongly influences abundance parameters and that yearly variation in seasonality occurs. This emphasizes that care should be taken when using such parameters in risk assessments for transmission of Culicoides-borne diseases and that more clear and strict guidelines for Culicoides monitoring should be considered when monitoring data are used for legislative purposes.

Background: The new genomic technologies have provided novel insights into the genetics of interactions between vectors, viruses and hosts, which are leading to advances in the control of arboviruses of medical importance. However, the development of tools and resources available for vectors of non-zoonotic arboviruses remains neglected. Biting midges of the genus Culicoides transmit some of the most important arboviruses of wildlife and livestock worldwide, with a global impact on economic productivity, health and welfare. The absence of a suitable reference genome has hindered genomic analyses to date in this important genus of vectors. In the present study, the genome of Culicoides sonorensis, a vector of bluetongue virus (BTV) in the USA, has been sequenced to provide the first reference genome for these vectors. In this study, we also report the use of the reference genome to perform initial transcriptomic analyses of vector competence for BTV., Results: Our analyses reveal that the genome is 189 Mb, assembled in 7974 scaffolds. Its annotation using the transcriptomic data generated in this study and in a previous study has identified 15,612 genes. Gene expression analyses of C. sonorensis females infected with BTV performed in this study revealed 165 genes that were differentially expressed between vector competent and refractory females. Two candidate genes, glutathione S-transferase (gst) and the antiviral helicase ski2, previously recognized as involved in vector competence for BTV in C. sonorensis (gst) and repressing dsRNA virus propagation (ski2), were confirmed in this study., Conclusions: The reference genome of C. sonorensis has enabled preliminary analyses of the gene expression profiles of vector competent and refractory individuals. The genome and transcriptomes generated in this study provide suitable tools for future research on arbovirus transmission. These provide a valuable resource for these vector lineage, which diverged from other major Dipteran vector families over 200 million years ago. The genome will be a valuable source of comparative data for other important Dipteran vector families including mosquitoes (Culicidae) and sandflies (Psychodidae), and together with the transcriptomic data can yield potential targets for transgenic modification in vector control and functional studies.

Background: Bluetongue disease, caused by bluetongue virus serotype 8 (BTV-8), appeared for the first time in the northern part of Europe in 2006, and subsequently rapidly spread causing severe economic losses to the farming industry. The implicated vectors of BTV in Europe are Culicoides species within the subgenus Avaritia (C. chiopterus, C. dewulfi, C. obsoletus and C. scoticus). Epidemiological data from Switzerland have shown that BTV, whose spread was eliminated at an early stage by vaccination campaigns, had not been circulating among livestock at higher altitudes where other species dominate the Culicoides fauna. In this study, we investigated the extent that Culicoides spp. prevailing at higher altitudes (mainly C. grisescens) can act as vectors for BTV., Methods: Culicoides were collected at farms in the pre-alpine region (two sites at 1550 m above sea level, masl, referred to as pre-alpine I; one site at 2030 masl, pre-alpine II) and, for comparative purposes, from the Swiss Plateau (one site, 650 masl). They were fed on bovine blood/BTV suspensions (BTV-1, 4 or 8) and incubated for eight days under a fluctuating temperature regime (13-25 °C, mean 19 °C), reflecting a mid-summer warm spell in the pre-alpine region. Susceptibility to BTV transmission was assessed from head homogenates by RT-qPCR and virus isolation., Results: Overall, 9196 female Culicoides were exposed to the three BTV strains through an artificial membrane, with feeding rates of 14-27%. Survival rates of blood-engorged Culicoides females at eight days post-infection depended on both virus serotype and altitude of origin. Virus dissemination (C q ≤ the cut-off value as determined by serial virus dilutions) was confirmed only for BTV-1 in C. scoticus (dissemination efficiency 22.5%; 9/40) and C. obsoletus (5.6%; 1/18) from the Swiss Plateau area. There was no strong evidence of susceptibility to infection for Culicoides from the pre-alpine area when fed with all BTV strains (BTV-1, 4 and 8)., Conclusions: This study confirms the susceptibility of C. scoticus and C. obsoletus to BTV-1 infection, including under cooler temperatures. Culicoides grisescens, which is highly abundant at higher altitudes, cannot be considered a potential vector under these temperature conditions.

There is a solid theoretical basis for expecting climate change to have a considerable effect on the infectious diseases of humans, animals and plants. Vector-borne diseases are the most likely to be affected. It is, however, rare to observe such impacts, as diseases are also influenced by many other drivers, some of which may have stronger effects over shorter time scales than climate change. Nevertheless, there is evidence that our warming climate has already influenced some animal diseases, of which bluetongue is considered a prime example. Bluetongue emerged dramatically in southern Europe after 1998 and in northern Europe from 2006. While the speed and scale of this emergence is a challenge to explain, there is evidence, principally from the development of climate-driven models, that recent climate change has played a significant role. Climate-driven models point to an increase in the risk of bluetongue transmission in Europe in recent decades, caused by an increased suitability of parts of southern Europe for the Afro-tropical biting midge, Culicoides imicola, as well as an increase in the vectorial capacity of indigenous Culicoides vectors in northern Europe. Farm-to-farm transmission models of bluetongue in England and Wales under predicted climatic conditions further suggest that, under high-emission scenarios, the scale of future outbreaks could far exceed those experienced to date. The role of climate change in the developing threat of animal disease is, therefore, likely to be economically and socially costly, unless lower emission targets can be set and followed.

Palearctic species of Culicoides (Diptera, Ceratopogonidae), in particular of the Obsoletus and Pulicaris complexes, were identified as putative vectors of bluetongue virus serotype 8 (BTV-8) on ruminant farms during the epizootic in Germany from 2006 to 2009. BTV may cause severe morbidity and mortality in ruminants and sporadically in South American camelids (SAC). However, the fauna of Culicoides spp. on SAC farms has not been investigated. Therefore, the ceratopogonid fauna was monitored on three farms with BTV-seropositive SAC in Germany. Black-light traps were set up on pastures and in stables from summer 2008 to autumn 2009. Additionally, ceratopogonids were caught in emergence traps mounted on llama dung and dung-free pasture from spring to autumn 2009. After morphological identification, selected Culicoides samples were analysed for BTV-RNA by real-time RT-PCR. The effects of the variables 'location', 'temperature' and 'humidity' on the number of Culicoides caught in black-light traps were modelled using multivariable Poisson regression. In total, 26 species of Culicoides and six other genera of biting midges were identified. The most abundant Culicoides spp. collected both outdoors and indoors with black-light traps belonged to the Obsoletus (77.4%) and Pulicaris (16.0%) complexes. The number of Culicoides peaked in summer, while no biting midges were caught during the winter months. Daily collections of Culicoides were mainly influenced by the location and depended on the interaction of temperature and humidity. In the emergence traps, species of the Obsoletus complex predominated the collections. In summary, the absence of BTV-RNA in any of the analysed Culicoides midges and in the BTV-seropositive SAC on the three farms together with the differences in the pathogenesis of BTV-8 in SAC compared to ruminants suggests a negligible role of SAC in the spread of the virus. Although SAC farms may provide similar suitable habitats for putative Culicoides vectors than ruminant farms, the results suggest that geographic and meteorological factors had a stronger influence on Culicoides abundance than the animal species., (Copyright © 2015 Elsevier B.V. All rights reserved.)

Bluetongue virus (BTV) and epizootic hemorrhagic disease virus (EHDV) are arthropod-transmitted viruses in the genus Orbivirus of the family Reoviridae. These viruses infect a variety of domestic and wild ruminant hosts, although the susceptibility to clinical disease associated with BTV or EHDV infection varies greatly among host species, as well as between individuals of the same species. Since their initial detection in North America during the 1950s, these viruses have circulated in endemic and epidemic patterns, with occasional incursions to more northern latitudes. In recent years, changes in the pattern of BTV and EHDV infection and disease have forced the scientific community to revisit some fundamental areas related to the epidemiology of these diseases, specifically in relation to virus-vector-host interactions and environmental factors that have potentially enabled the observed changes. The aim of this review is to identify research and surveillance gaps that obscure our understanding of BT and EHD in North America.

Heritable bacteria have been highlighted as important components of vector biology, acting as required symbionts with an anabolic role, altering competence for disease transmission, and affecting patterns of gene flow by altering cross compatibility. In this paper, we tested eight U.K. species of Culicoides (Diptera: Ceratopogonidae) midge for the presence of five genera of endosymbiotic bacteria: Cardinium (Bacteroidales: Bacteroidaceae); Wolbachia (Rickettsiales: Rickettsiaceae); Spiroplasma (Entomoplasmatales: Spiroplasmataceae); Arsenophonus (Enterobacteriales: Enterobacteriaceae), and Rickettsia (Rickettsiales: Rickettsiaceae). Cardinium spp. were detected in both sexes of Culicoides pulicaris and Culicoides punctatus, two known vectors of bluetongue virus. Cardinium spp. were not detected in any other species, including the Culicoides obsoletus group, the main vector of bluetongue and Schmallenberg viruses in northern Europe. The other endosymbionts were not detected in any Culicoides species. The Cardinium strain detected in U.K. Culicoides species is very closely related to the Candidatus Cardinium hertigii group C, previously identified in Culicoides species in Asia. Further, we infer that the symbiont is not a sex ratio distorter and shows geographic variation in prevalence within a species. Despite its detection in several species of Culicoides that vector arboviruses worldwide, the absence of Cardinium in the C. obsoletus group suggests that infections of these symbionts may not be necessary to the arboviral vector competence of biting midges., (© 2014 The Royal Entomological Society.)

BACKGROUND: As a primary vector of bluetongue virus (BTV) in the US, seasonal abundance and diel flight activity of Culicoides sonorensis has been documented, but few studies have examined how time of host-seeking activity is impacted by environmental factors. This knowledge is essential for interpreting surveillance data and modeling pathogen transmission risk. METHODS: The diel host-seeking activity of C. sonorensis was studied on a California dairy over 3 years using a time-segregated trap baited with CO2. The relationship between environmental variables and diel host-seeking activity (start, peak, and duration of activity) of C. sonorensis was evaluated using multiple linear regression. Fishers exact test and paired-sample z-test were used to evaluate the seasonal difference and parity difference on diel host-seeking activity. RESULTS: Host-seeking by C. sonorensis began and reached an activity peak before sunset at a higher frequency during colder months relative to warmer months. The time that host-seeking activity occurred was associated low and high daily temperature as well as wind speed at sunset. Colder temperatures and a greater diurnal temperature range were associated with an earlier peak in host-seeking. Higher wind speeds at sunset were associated with a delayed peak in host-seeking and a shortened duration of host-seeking. Parous midges reached peak host-seeking activity slightly later than nulliparous midges, possibly because of the need for oviposition by gravid females before returning to host-seeking. CONCLUSIONS: This study demonstrates that during colder months C. sonorensis initiates host-seeking and reaches peak host-seeking activity earlier relative to sunset, often even before sunset, compared to warmer months. Therefore, the commonly used UV light-baited traps are ineffective for midge surveillance before sunset. Based on this study, surveillance methods that do not rely on light trapping would provide a more accurate estimate of host-biting risk across seasons. The association of environmental factors to host-seeking shown in this study can be used to improve modeling or prediction of host-seeking activity. This study identified diurnal temperature range as associated with host-seeking activity, suggesting that Culicoides may respond to a rapidly decreasing temperature by shifting to an earlier host-seeking time, though this association needs further study.

Biting midges of the genus Culicoides (Diptera: Ceratopogonidae) are of huge veterinary importance, mainly as vectors of pathogens, such as Bluetongue virus. Currently, there are no effective methods to protect animals against biting midges, as insecticides have limited or short‐lived efficacy. Biting midges are attracted to hosts by carbon dioxide and by their body odours, which are mainly produced by skin bacteria. In humans, it has been shown that differences between individuals in attractiveness to mosquitoes is mediated by these skin bacterial volatiles. This opens the possibility to protect individuals from biting insects by supplementing their skin microbiome with probiotics. In this study, we investigated this approach by culturing sheep skin bacteria on various media and assessing their effects against field‐caught Culicoides (overwhelmingly Obsoletus group species) as well as laboratory‐reared Culicoides nubeculosus (Meigen). Aerococcus urinaeequi, Bacillus safensis, Bacillus subtilis, Jeotgalicoccus psychrophilus, Micrococcus sp. and Staphylococcus equorum were selected to be tested in a dual‐choice Y‐tube olfactometer, assessing their behavioural effects on biting midges. We revealed an avoidance effect on laboratory‐reared C. nubeculosus when testing B. safensis and B. subtilis. Bacillus safensis and Micrococcus sp. yielded significant repellent potential towards field‐caught Culicoides. These two candidates were subsequently tested in a membrane blood‐feeding assay. When the bacterial species B. safensis was applied to the membrane, a feeding reduction of 83% was observed with field‐caught Culicoides. [ABSTRACT FROM AUTHOR]

Culicoides are responsible for a number of diseases transmitted to the animals causing severe health conditions and great economic losses. The outbreak of bluetongue virus in Albania in 2014 and lumpy skin disease in 2016 strongly indicated the establishment of a sentinel herd surveillance in the country ruminant farms. Considering all evidence for Albania, investigation of species composition and abundance of Culicoides on livestock farms was performed in 10 sentinel sites. Ultraviolet-light traps (OVI-traps) were placed in selected sites for collection of Culicoides starting from April 15 to July 30, 2021. A total of 37,111 Culicoides midges from 18 different species were collected during the investigation. The most abundant species identified were C. newsteadi and C. obsoletus. Moreover, three species were described for the first time in Albania (C. collessi, C. chiopterus and C. immaculatus). Presence and abundance of the collected Culicoides spp. are associated with potential outbreak of diseases (i.e. bluetongue, ephemeral fever) linked with their transmission from Culicoides. [ABSTRACT FROM AUTHOR]