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.

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.

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.

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.

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.

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.

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.

Monitoring longitudinal immunological responses to bluetongue virus 17 in experimentally infected sheep.

Bluetongue virus (BTV) is an economically important pathogen of ruminant species with worldwide prevalence. While many BTV infections are asymptomatic, animals with symptomatic presentation deteriorate quickly with the sickest succumbing to disease within one week. Animals that survive the infection often require months to recover. The immune response to BTV infection is thought to play a central role in controlling the disease. Key to understanding BTV disease is profiling vertebrate host immunological cellular and cytokine responses. Studies to characterize immune responses in ruminants have been limited by a lack of species-specific reagents and assay technology. Here we assess the longitudinal immunological response to experimental BTV-17-California (CA) infection in sheep using the most up to date assays. We infected a cohort of sheep with BTV-17-CA and longitudinally monitored each animal for clinical disease, viremia and specific immunological parameters (B cells, T cells, monocytes) by RT-qPCR, traditional flow cytometry and/or fluorescent based antibody arrays. BTV-inoculated sheep exhibited clinical signs characteristic of bluetongue virus disease. Circulating virus was demonstrated after 8 days post inoculation (DPI) and remained detectable for the remainder of the time course (24 DPI). A distinct lymphopenia was observed between 7 and 14 DPI that rebounded to mock-inoculated control levels at 17 DPI. In addition, we observed increased expression of pro-inflammatory cytokines after 8 DPI. Taken together, we have established a model of BTV infection in sheep and have successfully monitored the longitudinal vertebrate host immunological response and viral infection progression using a combination of traditional methods and cutting-edge technology.

The potential for bluetongue virus serotype 16 to cause disease in sheep in New South Wales, Australia.

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.

Bluetongue outbreak in a sheep flock from Iran.

Bluetongue virus (BTV) is an arthropodborne Orbivirus that belongs to the Reoviridae family. Bluetongue is one of the most important diseases of sheep. A flock of 300 Lacon sheep just arrived from France, located in the countryside of Qazvin city, Iran, was examined, in August 2022. In history taking and clinical examination, submandibular oedema (216/300, 72%), fever (216/300, 72%), inappetence (216/300, 72%), stomatitis (216/300, 72%), nasal discharge (90/300, 30%) and lameness (30/300, 10%) were recorded. Foot-and-mouth disease, bluetongue (BT), contagious ecthyma and peste des petits ruminants were the most important differential diagnosis with reference to clinical signs. Tongue scraping samples from four clinically affected sheep were sent to the laboratory for PCR tests and, in all of them, BTV was detected. The affected flock had a history of vaccination with an attenuated live vaccine in the previous 4 months. The morbidity rate, mortality rate and case fatality rate were 72% (216/300), 7% (21/300) and 9.7% (21/216), respectively. This report is the first documented clinical form of BT in sheep from Iran.