Seasonal change and influence of environmental variables on host-seeking activity of the biting midge Culicoides sonorensis at a southern California dairy, USA.

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. Fisher's 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.

Multi-Gene Recombinant Baculovirus Expression Systems: From Inception to Contemporary Applications.

Many protein expression systems are primarily utilised to produce a single, specific recombinant protein. In contrast, most biological processes such as virus assembly rely upon a complex of several interacting proteins rather than the activity of a sole protein. The high complexity of the baculovirus genome, coupled with a multiphase replication cycle incorporating distinct transcriptional steps, made it the ideal system to manipulate for high-level expression of a single, or co-expression of multiple, foreign proteins within a single cell. We have developed and utilised a series of recombinant baculovirus systems to unravel the sequential assembly process of a complex non-enveloped model virus, bluetongue virus (BTV). The high protein yields expressed by the baculovirus system not only facilitated structure-function analysis of each viral protein but were also advantageous to crystallography studies and supported the first atomic-level resolution of a recombinant viral protein, the major BTV capsid protein. Further, the formation of recombinant double-shelled virus-like particles (VLPs) provided insights into the structure-function relationships among the four major structural proteins of the BTV whilst also representing a potential candidate for a viral vaccine. The baculovirus multi-gene expression system facilitated the study of structurally complex viruses (both non-enveloped and enveloped viruses) and heralded a new generation of viral vaccines.

Spatial Transmission Characteristics of the Bluetongue Virus Serotype 3 Epidemic in The Netherlands, 2023.

A devastating bluetongue (BT) epidemic caused by bluetongue virus serotype 3 (BTV-3) has spread throughout most of the Netherlands within two months since the first infection was officially confirmed in the beginning of September 2023. The epidemic comes with unusually strong suffering of infected cattle through severe lameness, often resulting in mortality or euthanisation for welfare reasons. In total, tens of thousands of sheep have died or had to be euthanised. By October 2023, more than 2200 locations with ruminant livestock were officially identified to be infected with BTV-3, and additionally, ruminants from 1300 locations were showing BTV-associated clinical symptoms (but not laboratory-confirmed BT). Here, we report on the spatial spread and dynamics of this BT epidemic. More specifically, we characterized the distance-dependent intensity of the between-holding transmission by estimating the spatial transmission kernel and by comparing it to transmission kernels estimated earlier for BTV-8 transmission in Northwestern Europe in 2006 and 2007. The 2023 BTV-3 kernel parameters are in line with those of the transmission kernel estimated previously for the between-holding spread of BTV-8 in Europe in 2007. The 2023 BTV-3 transmission kernel has a long-distance spatial range (across tens of kilometres), evidencing that in addition to short-distance dispersal of infected midges, other transmission routes such as livestock transports probably played an important role.

RNA genome packaging and capsid assembly of bluetongue virus visualized in host cells.

Unlike those of double-stranded DNA (dsDNA), single-stranded DNA (ssDNA), and ssRNA viruses, the mechanism of genome packaging of dsRNA viruses is poorly understood. Here, we combined the techniques of high-resolution cryoelectron microscopy (cryo-EM), cellular cryoelectron tomography (cryo-ET), and structure-guided mutagenesis to investigate genome packaging and capsid assembly of bluetongue virus (BTV), a member of the Reoviridae family of dsRNA viruses. A total of eleven assembly states of BTV capsid were captured, with resolutions up to 2.8 Å, with most visualized in the host cytoplasm. ATPase VP6 was found underneath the vertices of capsid shell protein VP3 as an RNA-harboring pentamer, facilitating RNA packaging. RNA packaging expands the VP3 shell, which then engages middle- and outer-layer proteins to generate infectious virions. These revealed "duality" characteristics of the BTV assembly mechanism reconcile previous contradictory co-assembly and core-filling models and provide insights into the mysterious RNA packaging and capsid assembly of Reoviridae members and beyond.

Evaluating Temperature Effects on Bluetongue Virus Serotype 10 and 17 Coinfection in Culicoides sonorensis.

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.

Epidemiological Analyses of the First Incursion of the Epizootic Hemorrhagic Disease Virus Serotype 8 in Tunisia, 2021-2022.

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.

Assessing Reassortment between Bluetongue Virus Serotypes 10 and 17 at Different Coinfection Ratios in Culicoides sonorenesis.

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.

Specific T-cell subsets have a role in anti-viral immunity and pathogenesis but not viral dynamics or onwards vector transmission of an important livestock arbovirus.

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.

The Study of Bluetongue Virus (BTV) and Epizootic Hemorrhagic Disease Virus (EHDV) Circulation and Vectors at the Municipal Parks and Zoobotanical Foundation of Belo Horizonte, Minas Gerais, Brazil (FPMZB-BH).

Bluetongue Virus (BTV) and Epizootic Hemorrhagic Disease Virus (EHDV) are Orbiviruses primarily transmitted by their biological vector, Culicoides spp. Latreille, 1809 (Diptera: Ceratopogonidae). These viruses can infect a diverse range of vertebrate hosts, leading to disease outbreaks in domestic and wild ruminants worldwide. This study, conducted at the Belo Horizonte Municipal Parks and Zoobotany Foundation (FPMZB-BH), Minas Gerais, Brazil, focused on Orbivirus and its vectors. Collections of Culicoides spp. were carried out at the FPMZB-BH from 9 December 2021 to 18 November 2022. A higher prevalence of these insects was observed during the summer months, especially in February. Factors such as elevated temperatures, high humidity, fecal accumulation, and proximity to large animals, like camels and elephants, were associated with increased Culicoides capture. Among the identified Culicoides spp. species, Culicoides insignis Lutz, 1913, constituted 75%, and Culicoides pusillus Lutz, 1913, 6% of the collected midges, both described as competent vectors for Orbivirus transmission. Additionally, a previously unreported species in Minas Gerais, Culicoides debilipalpis Lutz, 1913, was identified, also suspected of being a transmitter of these Orbiviruses. The feeding preferences of some Culicoides species were analyzed, revealing that C. insignis feeds on deer, Red deer (Cervus elaphus) and European fallow deer (Dama dama). Different Culicoides spp. were also identified feeding on humans, raising concerns about the potential transmission of arboviruses at the site. In parallel, 72 serum samples from 14 susceptible species, including various Cervids, collected between 2012 and 2022 from the FPMZB-BH serum bank, underwent Agar Gel Immunodiffusion (AGID) testing for BTV and EHDV. The results showed 75% seropositivity for BTV and 19% for EHDV. Post-testing analysis revealed variations in antibody presence against BTV in a tapir and a fallow deer and against EHDV in a gemsbok across different years. These studies confirm the presence of BTV and EHDV vectors, along with potential virus circulation in the zoo. Consequently, implementing control measures is essential to prevent susceptible species from becoming infected and developing clinical diseases.

Engineering recombinant replication-competent bluetongue viruses expressing reporter genes for in vitro and non-invasive in vivo studies.

Bluetongue virus (BTV) is the causative agent of the important livestock disease bluetongue (BT), which is transmitted via Culicoides bites. BT causes severe economic losses associated with its considerable impact on health and trade of animals. By reverse genetics, we have designed and rescued reporter-expressing recombinant (r)BTV expressing NanoLuc luciferase (NLuc) or Venus fluorescent protein. To generate these viruses, we custom synthesized a modified viral segment 5 encoding NS1 protein with the reporter genes located downstream and linked by the Porcine teschovirus-1 (PTV-1) 2A autoproteolytic cleavage site. Therefore, fluorescent signal or luciferase activity is only detected after virus replication and expression of non-structural proteins. Fluorescence or luminescence signals were detected in cells infected with rBTV/Venus or rBTV/NLuc, respectively. Moreover, the marking of NS2 protein confirmed that reporter genes were only expressed in BTV-infected cells. Growth kinetics of rBTV/NLuc and rBTV/Venus in Vero cells showed replication rates similar to those of wild-type and rBTV. Infectivity studies of these recombinant viruses in IFNAR(-/-) mice showed a higher lethal dose for rBTV/NLuc and rBTV/Venus than for rBTV indicating that viruses expressing the reporter genes are attenuated in vivo. Interestingly, luciferase activity was detected in the plasma of viraemic mice infected with rBTV/NLuc. Furthermore, luciferase activity quantitatively correlated with RNAemia levels of infected mice throughout the infection. In addition, we have investigated the in vivo replication and dissemination of BTV in IFNAR (-/-) mice using BTV/NLuc and non-invasive in vivo imaging systems.IMPORTANCEThe use of replication-competent viruses that encode a traceable fluorescent or luciferase reporter protein has significantly contributed to the in vitro and in vivo study of viral infections and the development of novel therapeutic approaches. In this work, we have generated rBTV that express fluorescent or luminescence proteins to track BTV infection both in vitro and in vivo. Despite the availability of vaccines, BTV and other related orbivirus are still associated with a significant impact on animal health and have important economic consequences worldwide. Our studies may contribute to the advance in orbivirus research and pave the way for the rapid development of new treatments, including vaccines.

Seven-year prospective study on yearly incidence of Orbivirus infection of captive white-tailed deer and potential Culicoides vectors.

Bluetongue virus (BTV) and epizootic hemorrhagic disease virus (EHDV) are arthropod-borne viruses that are transmitted by biting midges in the genus Culicoides (Diptera: Ceratopogonidae) and can cause hemorrhagic disease in certain ruminants. The objectives of this study were to measure the incidence of BTV and EHDV infections in captive white-tailed deer herd as well as tissues and corresponding presence of Culicoides midges at a location near Clinton, LA. During a 7-yr study with yearly outbreaks of hemorrhagic disease in the deer herd, 15 species of Culicoides were captured using Centers for Disease Control (CDC) black light traps. Reverse transcriptase quantitative polymerase chain reaction (PCR) was performed to screen for BTV and EHDV in pools of midges and tissues of deer. From 2012 to 2018, 1,711 pools of midges representing 24,859 specimens were tested, and specimens from 5 of the 15 collected species (Culicoides debilipalpis, Culicoides stellifer, Culicoides venustus, Culicoides haematopotus, and Culicoides crepuscularis) were found to be PCR positive for BTV and EHDV. Most of the BTV-positive pools of biting midges were from specimens of C. debilipalpis and C. stellifer, and most of the EHDV-positive pools were from specimens of C. venustus and C. stellifer. During the 7-yr period, 112 white-tailed deer that died at the study location were PCR positive for BTV or EHDV: detected BTV serotypes were 10 and 12 and EHDV serotypes were 1, 2, and 6. There was a significant increase in BTV/EHDV antibody prevalence in white-tailed deer during the study; antibody-positive rates increased from 15% to 78% in the deer herd of approximately 100 animals.

First molecular evidence of potential Culicoides vectors implicated in bluetongue virus transmission in Morocco.

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.

A novel bluetongue virus serotype 2 strain isolated from a farmed Florida white-tailed deer (Odocoileus virginianus) arose from reassortment of gene segments derived from co-circulating serotypes in the Southeastern USA.

Bluetongue disease is a reportable animal disease that affects wild and farmed ruminants, including white-tailed deer (WTD). This report documents the clinical findings, ancillary diagnostics, and genomic characterization of a novel reassortant bluetongue virus serotype 2 (BTV-2) strain isolated from a dead Florida farmed WTD in 2022. Our analyses support that this BTV-2 strain likely stemmed from the acquisition of genome segments from co-circulating BTV strains in Florida and Louisiana. In addition, our analyses also indicate that genetically uncharacterized BTV strains may be circulating in the Southeastern USA; however, the identity and reassortant status of these BTV strains cannot be determined based on the VP2 and VP5 genome sequences. Hence, continued surveillance based on complete genome characterization is needed to understand the genetic diversity of BTV strains in this region and the potential threat they may pose to the health of deer and other ruminants.

Detection, Characterization and Sequencing of BTV Serotypes Circulating in Cuba in 2022.

In Cuba, despite a high sero-prevalence of bluetongue virus (BTV), circulating serotypes remain unknown. The aim of this study was to identify circulating BTV serotypes in farms throughout the western region of Cuba. Blood samples were collected from 200 young cattle and sheep between May and July 2022 for virological analyses (PCR, viral isolation and virus neutralization) and genome sequencing. The results confirmed viral circulation, with viro-prevalence of 25% for BTV. The virus was isolated from 18 blood samples and twelve BTV serotypes were identified by sequencing RT-PCR products targeting the segment 2 of the BTV genome (BTV-1, 2, 3, 6, 10, 12, 13, 17, 18, 19, 22 and 24). Finally, the full genome sequences of 17 Cuban BTV isolates were recovered using a Sequence Independent Single Primer Amplification (SISPA) approach combined to MinION Oxford Nanopore sequencing technology. All together, these results highlight the co-circulation of a wide diversity of BTV serotypes in a quite restricted area and emphasize the need for entomological and livestock surveillance, particularly in light of recent changes in the global distribution and nature of BTV infections.

From forests to fields: investigating Culicoides (Diptera: Ceratopogonidae) abundance and diversity in cattle pastures and adjacent woodlands.

Culicoides Latreille (Diptera: Ceratopogonidae) biting midges are hematophagous flies that feed on wild and domestic ruminants. They can transmit arboviruses, such as bluetongue virus (BTV) and epizootic hemorrhagic disease virus (EHDV), which circulate in the United States. Larvae occupy a range of aquatic and semiaquatic habitats, and disperse short distances from their development sites. In the southeastern United States, there are limited studies on the abundance and diversity of Culicoides in wooded and adjacent livestock pasture habitats. In this study, we characterized Culicoides diversity and abundance within these distinct habitat types. BG-Sentinel and CDC miniature suction traps baited with CO2 or UV-light were placed in wooded and pasture habitats at 2 locations on a university beef farm in Savoy, Arkansas. Traps were set once per week for 9 wk during August-October of 2021 and 2022. Fifteen species were collected during this study, and the 2 most abundant species were Culicoides haematopotus Malloch and Culicoides stellifer Coquillett. There was a significant effect of site and location on C. haematopotus collections, and a significant effect and interaction of site and trap on C. stellifer collections. In the woods, significantly more C. stellifer were collected from CDC-UV traps, while in the pasture significantly more were collected in CDC-CO2 traps. These data suggest that C. stellifer, a putative vector of BTV and EHDV in the southeast, may be traveling into the pasture to host-seek, while C. haematopotus remains primarily in wooded areas. This study reveals community differences between these habitat types and implications for Culicoides control.

Preliminary investigation reveals novel pathological consequences of bluetongue virus-1 infection in the endocrine glands of pregnant Indian sheep.

Bluetongue virus (BTV), a major peril to the sheep industry, infects a wide range of the cells in the infected animals including mononuclear, dendritic and epithelial cells. However, little is known about its tropism for the secretory epithelial cells of endocrine glands and the pathogenesis it induces. The aim of the study was to assess the BTV load, antigen distribution in the tissue of the pituitary, thyroid as well as adrenal glands and associated histopathological consequences. BTV antigens were localized using immunohistochemistry in the thyroid's epithelial cells, zona fasciculata and zona reticularis cells and the anterior pituitary epithelial cells. The real-time PCR portrayed the high viral load in adrenals at 7th days postinoculation (DPI) and in thyroid and pituitary glands at 15th DPI. Serum examination revealed variation in the T-3 and T-4 of infected animals in comparison to the control group. Caspase-3 immunolocalization revealed BTV-1 induces apoptosis in the affected cells of endocrine gland of infected animals. Further, this study signifies the tropism of BTV in the novel sites (endocrine glands) of the host that might be one of the reasons for the poor performance of infected animals.

Bluetongue disease in sheep in New South Wales - April 2023.

In 2016, bluetongue virus (BTV), serotype 16 (BTV-16), was detected in New South Wales (NSW) in sentinel cattle for the first time. Over the next 6 years, BTV-16 has been detected regularly and over an increasing area of the BTV zone in NSW. In April 2023, disease was reported in sheep on two farms on the Northern Tablelands of NSW. The consistent clinical signs included reduced exercise tolerance, facial swelling, serous nasal discharges with encrustation of the nasal plane, subcutaneous oedema of the neck and brisket and variable congestion of the coronary band. Affected sheep were mainly mature ewes and rams, with an estimated morbidity of 20% over a period of 6-8 weeks. Although there were several unexpected deaths, no veterinary examination was sought. Predominantly BTV-16 RNA was detected in sick sheep, with an incidence of infection of approximately 40% in a cross section of one flock. These events represent the first confirmation of disease due to bluetongue virus in NSW. As these cases occurred in a region with a high density of sheep, if there is ongoing transmission of BTV-16 during subsequent summers, further disease might be expected.

Bluetongue virus seroprevalence and risk factor analysis in cattle and water buffalo in southern Italy (Campania region).

Bluetongue is an arthropod-borne viral infection that is notifiable in several countries and causes significant economic losses and major concerns for ruminant trade. In this study, we investigated bluetongue 1seroprevalence in the Campania region, southern Italy, in cattle and buffalo populations, and assessed which factors were correlated with a high risk of exposure. The infection was widespread, as evidenced by the high individual (43.6%) and herd prevalence (85.4%). The highest prevalence was found in adult animals. Among the climatic factors analyzed, average temperature played a prominent role, being capable of affecting the probability of being positive for this infection. Surprisingly, exposure to Schmallenberg virus did not predispose animals to be positive for bluetongue virus, even though these infections share the same vector (Culicoides). Our data, consistent with those in the literature, suggest the transversal spread of bluetongue virus in the Mediterranean area, and indicate a limited co-exposure rate between Bluetongue and Schmallenberg viruses.

Clinical presentation, medical management, and outcomes in 35 hospitalized sheep diagnosed with bluetongue virus disease.

BACKGROUND: There is only limited information on the clinical presentation, medical management, and outcomes of hospitalized sheep diagnosed with bluetongue virus (BTV) disease. OBJECTIVES: To describe the signalment, history, clinical signs, clinicopathological findings, medical management, and clinical outcomes of sheep diagnosed with BTV disease. ANIMALS: Thirty-five hospitalized sheep with BTV disease. METHODS: Retrospective case series. Medical records from 1989 to 2021 were evaluated. History, signalment, clinical signs, laboratory test results, treatments, and outcomes were recorded. RESULTS: BTV disease was diagnosed from July to December, with a peak proportion (43%; 15/35) of diagnoses recorded in October. Pyrexia and anorexia, respiratory disease, vasculitis, coronitis and lameness, and ulcerative mucosal lesions were present in 71%, 71%, 66%, 49%, and 22% of sheep, respectively. BTV serotypes 10, 11, 13, and 17 were identified, with serotype 17 (75%) being the most frequent. Management of cases included administration of antimicrobials (89%), anti-inflammatories (77%), IV fluids (60%), vitamins (20%), proton-pump inhibitors (14%), diuretics (9%), and antioxidants (9%). Six ewes were pregnant on presentation, but none aborted. Six (17%) sheep died or were euthanized because of clinical deterioration, whereas 83% were discharged. CONCLUSIONS AND CLINICAL IMPORTANCE: The proportion of sheep that survived BTV disease after treatment was relatively high. Serotyping of BTV is recommended because of the mismatch between frequently identified serotypes and the serotype present in the vaccine.

Recovery of multireassortant bluetongue virus serotype 6 sequences from a mule deer (Odocoileus hemionus) and Dorset sheep (Ovis aries) in Colorado.

We report the discovery of two bluetongue virus serotype 6 (BTV-6) reassortants recovered from a domestic sheep and a free-ranging mule deer in northern Colorado. At the time of this publication, whole-genome sequencing of BTV-6 isolates in the Western U.S. have not been undertaken. These findings reflect the incursive movement of geographically distinct BTV serotypes into important agricultural areas of the U.S. and demonstrate reassortment with regionally circulating serotypes.