Bovine viral diarrhea virus (BVDV) is a widespread pathogen of cattle and other mammals that causes major economic losses in the livestock industry. N4-TSC and 6NO2-TSC are two thiosemicarbazones derived from 1-indanone that exhibit anti-BVDV activity in vitro. These compounds selectively inhibit BVDV and are effective against both cytopathic and non-cytopathic BVDV-1 and BVDV-2 strains. We confirmed that N4-TSC acts at the onset of viral RNA synthesis, as previously reported for 6NO2-TSC. Moreover, resistance selection and characterization showed that N4-TSCR mutants were highly resistant to N4-TSC but remained susceptible to 6NO2-TSC. In contrast, 6NO2-TSCR mutants were resistant to both compounds. Additionally, mutations N264D and A392E were found in the viral RNA-dependent RNA polymerase (RdRp) of N4-TSCR mutants, whereas I261 M was found in 6NO2-TSCR mutants. These mutations lay in a hydrophobic pocket within the fingertips region of BVDV RdRp that has been described as a \"hot spot\" for BVDV non-nucleoside inhibitors.

Bovine viral diarrhea virus (BVDV) infections cause USD 1.5-2 billion in losses annually. Maternal BVDV after 150 days of gestation causes transient fetal infection (TI) in which the fetal immune response clears the virus. The impact of fetal TI BVDV infections on postnatal growth and white blood cell (WBC) methylome as an index of epigenetic modifications was examined by inoculating pregnant heifers with noncytopathic type 2 BVDV or media (sham-inoculated controls) on Day 175 of gestation to generate TI (n = 11) and control heifer calves (n = 12). Fetal infection in TI calves was confirmed by virus-neutralizing antibody titers at birth and control calves were seronegative. Both control and TI calves were negative for BVDV RNA in WBCs by RT-PCR. The mean weight of the TI calves was less than that of the controls (p < 0.05). DNA methyl seq analysis of WBC DNA demonstrated 2349 differentially methylated cytosines (p ≤ 0.05) including 1277 hypomethylated cytosines, 1072 hypermethylated cytosines, 84 differentially methylated regions based on CpGs in promoters, and 89 DMRs in islands of TI WBC DNA compared to controls. Fetal BVDV infection during late gestation resulted in epigenomic modifications predicted to affect fetal development and immune pathways, suggesting potential consequences for postnatal growth and health of TI cattle.

Wild pigs (Sus scrofa) are among the most detrimental invasive species in the USA. They are damaging to crops and agriculture, pose a public health risk as reservoirs of zoonotic pathogens, and may also spread disease to livestock. One pathogen identified in wild pigs is bovine viral diarrhea virus (BVDV), a virus that causes an economically important disease of cattle (Bos taurus and Bos indicus). We sought to determine the BVDV seroprevalence in wild pigs in 17 states across the US and to determine whether age category, sex, or location were associated with a positive antibody titer. Serum samples from 945 wild pigs were collected from 17 US states. Virus neutralization assays were performed to determine antibody titers against BVDV-1b and BVDV-2a. Total BVDV seroprevalence for the study area was 5.8% (95% confidence interval [CI], 4.11-8.89). Seroprevalence across all evaluated states was determined to be 4.4% (95% CI, 2.48-6.82) for BVDV-1b and 3.6% (95% CI, 1.54-5.60) for BVDV-2a. The seroprevalence for individual states varied from 0% to 16.7%. There was no statistical difference in median antibody titer for BVDV-1b or BVDV-2a by sex or age category. State seroprevalences for both BVDV-1b and BVDV-2a were associated with wild pig population estimates for those states.

Bovine viral diarrhea virus (BVDV) induces immunosuppression and thymus depletion in calves. This study explores the impact of prior BVDV-2 exposure on the subsequent immune response to influenza D virus (IDV). Twenty 3-week-old calves were divided into four groups. Calves in G1 and G3 were mock-treated on day 0, while calves in G2 and G4 received BVDV. Calves in G1 (mock) and G2 (BVDV) were necropsied on day 13 post-infection. IDV was inoculated on day 21 in G3 calves (mock + IDV) and G4 (BVDV + IDV) and necropsy was conducted on day 42. Pre-exposed BVDV calves exhibited prolonged and increased IDV shedding in nasal secretions. An approximate 50% reduction in the thymus was observed in acutely infected BVDV calves (G2) compared to controls (G1). On day 42, thymus depletion was observed in two calves in G4, while three had normal weight. BVDV-2-exposed calves had impaired CD8 T cell proliferation after IDV recall stimulation, and the α/β T cell impairment was particularly evident in those with persistent thymic atrophy. Conversely, no difference in antibody levels against IDV was noted. BVDV-induced thymus depletion varied from transient to persistent. Persistent thymus atrophy was correlated with weaker T cell proliferation, suggesting correlation between persistent thymus atrophy and impaired T cell immune response to subsequent infections.

We characterized bovine viral diarrhea virus (BVDV)-related abortions in cattle and identified the species and subgenotypes in the state of Santa Catarina, southern Brazil. Our RT-PCR assay was positive for BVDV in 5 fetuses from different farms; however, 3 of the 5 fetuses were also PCR-positive for Neospora caninum. In the 5 BVDV-positive fetuses, gross lesions included fetal mummification (1), hepatomegaly (1), subcutaneous edema (1), and perirenal edema (1). Predominant histologic lesions included epicarditis and mild-to-moderate lymphoplasmacytic myocarditis (5), mild multifocal lymphoplasmacytic interlobular pneumonia (4), nephrosis associated with moderate multifocal interstitial nephritis (1), moderate multifocal lymphoplasmacytic necrotic hepatitis (1), and mild multifocal lymphoplasmacytic meningitis (1). The amplification products from the Pestivirus 5\'UTR region of 4 of the 5 fetuses had 96.3-100% similarity between fetal strains and reference strains. The samples were distributed into 2 branches of the phylogenetic tree; strains UDESC:01, UDESC:02, and UDESC:05 clustered in the BVDV-1e branch, uncommon in the Americas, and strain UDESC:04 clustered in the BVDV-2b branch. The three 1e strains had 96.9-97.4% similarity.

Keds are hematophagous ectoparasites of animals belonging to the family Hippoboscidae (Diptera: Hippoboscoidea). Because of their importance as vectors of some pathogens of medical and veterinary importance, they have received special attention. There are numerous studies demonstrating the presence of various parasites and pathogenic bacteria in keds. At the same time, there are very few reports on ked-related viruses. The aim of this study was to perform a molecular survey of viral pathogens in the forest fly (Hippobosca equina) from southern Kazakhstan. In this study, 104H. equina were collected from livestock in Turkistan oblast (southern region of Kazakhstan), which has the largest concentration of livestock in the country. Insect homogenates were screened by PCR for pestiviruses, orbiviruses, flaviviruses, orthobunyaviruses, phleboviruses, orthopoxviruses, capripoxviruses, parapoxviruses, and asfiviruses. The causative agents of two livestock diseases, bovine viral diarrhea virus (BVDV) (3/104; 2.88%; 95% confidence interval (CI): 0.6-8.2%) and bluetongue virus (BTV) (1/104; 0.96%; 95% CI: 0.02-5.24%), were identified and subjected to further analysis. The BTV strain was isolated and all ten genomic RNA segments were sequenced using the Sanger technique. The isolated BTV strain showed >99.6% identity in all genomic segments with the BTV-9 strains belonging to the \'western\' topotype. Partial analysis of the 5\'-untranslated region demonstrated that both BVDV strains are closely related to Pestivirus B. Flaviviruses, phleboviruses, orthobunyaviruses, poxviruses, and asfiviruses were not detected. This is the first report describing BVDV type 2 in Kazakhstan. The study also confirms the presence of BTV serotype 9 in southern Kazakhstan. The data presented here can help improve preventive measures to control the spread of viral diseases in livestock by using forest flies as an object of epidemiological studies. However, further studies are needed to investigate the vector capacity of H. equina and its suitability for xenodiagnosis of veterinary relevant pathogens.

Whole-genome phylogenetic analysis, the most suitable strategy for subtyping bovine viral diarrhea virus 1 (BVDV-1) and BVDV-2, is not feasible for many laboratories. Consequently, BVDV isolates/strains have been frequently subtyped based on analysis of single genomic regions, mainly the 5\' untranslated region (UTR). This approach, however, may lead to inaccurate and/or poorly statistically supported viral classification. Herein, we describe novel primer sets whose amplicons may be easily sequenced and used for BVDV subtyping. Initially, genomic regions previously described as the most suitable targets for BVDV subtyping were analyzed for design of high-coverage primers. The putative amplicons were analyzed in silico for their suitability to reproduce the phylogenetic classification of 118 BVDV-1 and 88 BVDV-2 complete/near-complete genomes (CNCGs) (GenBank). This analysis was also performed considering the region amplifiable by primers HCV90-368, 324-326 and BP189-389 (5\'UTR), which have been used for BVDV diagnosis and/or classification. After confirming the agreement between the analyses of our primers\' amplicon versus the CNCGs, we optimized the RT-PCRs and evaluated their performance for amplification of BVDV isolates/strains (n = 35 for BVDV-1; n = 33 for BVDV-2). Among the potential targets for BVDV subtyping, we designed high-coverage primers for NS3-NS4A (BVDV-1) (526 bp amplicon) and NS5B (BVDV-2) (728 bp). The classification based on these regions fully reproduced the subtyping of all CNCGs. On the other hand, subtyping based on the putative amplicons from primers HCV90-368, 324-326 and BP189-389 showed disagreements in relation the CNCG analysis. The NS3-NS4A and NS5B primers also allowed the amplification of all BVDV isolates/strains tested. Finally, we suggest the use of these primers in future phylogenetic and epidemiological studies of BVDVs.

The aim of this study was to estimate the occurrence of bovine viral diarrhea virus (BVDV) infection and to assess the population immunity in cattle vaccinated against BVDV in different regions of Kazakhstan. Cattle samples were collected in 12 oblasts (43 districts) of Kazakhstan. A total of 2477 cattle from 114 herds and 21 Bukhara deer (Cervus elaphus bactrianus) were examined by ELISA and conventional RT-PCR. Univariate and multivariate logistic regression analysis was performed to identify risk factors associated with BVDV infection in the country. In total, antibodies against BVDV were found in 79.3% (1965/2477) of all the animals and 92.1% (105/114) of all the herds examined. Seroprevalence in unvaccinated and vaccinated animals was 48.6% (447/920) and 98.7% (1391/1410), respectively. Seroprevalence in deer was 19.1% (4/21). The BVDV RNA was detected in six unvaccinated cattle (0.2%). Sequence analysis of the 5\'-untranslated region demonstrated that four of the detected strains belonged to BVDV-1 and two strains to BVDV-2. Regression analysis revealed that age, production type, housing method, farm size, and geographic location were risk factors for BVDV infection in cattle in Kazakhstan. The present data confirm circulation of BVDV-1 and BVDV-2 in Kazakhstan and highlight the need to improve strategies for prevention and control of BVDV infection in the country.

BACKGROUND: Little is known about the effects of trace mineral supplementation on the stress response in beef calves.
OBJECTIVE: To investigate the effect of injectable trace mineral supplementation (ITM) on the stress response in beef calves exposed to different types of stress.
METHODS: Thirty weaned Angus and Angus crossbred calves.
METHODS: The enrolled calves were randomly assigned to 2 groups: ITM, 15 calves received modified-live virus vaccine (MLV) and ITM SC and 15 calves received MLV and saline SC (CONT). The calves were exposed to 3 types of stress: the stress of MLV vaccination (d0), nasal aerosol with bovine viral diarrhea virus-2 (BVDV-2) challenge (d5), and liver biopsy (d26). The calves\' body weights and health status were monitored. Leukocyte counts, serum cortisol concentration ([cort]), BVDV-2 serum neutralizing antibodies (SNA), and percentages of CD4+ , CD8+ , WC1+ , and CD25+ T-lymphocytes were measured.
RESULTS: Serum cortisol concentration ([cort]) showed strong associations with the percentage of CD8+ (rs  = .50), BVDV2-SNA (rs  = -.43), and WC1CD25+ (rs  = .41) cells, and rectal temperature (rs  = .40). The highest [cort] was reported 3 days after aerosol BVDV-2 challenge. Serum [cort] was decreased in ITM-treated calves 3 days post-BVDV-2 challenge, compared with CONT calves, with an average decrease of 18.5 ng/μL (95% confidence interval [CI], -6.07 to -31.3). The ITM-treated calves were heavier and healthier (P < .01) than the CONT calves.
CONCLUSIONS: Trace mineral supplementation appears to have stress mitigation effects in beef cattle that may reflect positively on growth and health performance. Viral exposure is associated with a high degree of stress, which is considered a major welfare concern.

This study compares immune and clinical responses of bovine viral diarrhea virus (BVDV)-maternal antibody (MatAb)-positive beef calves primed with intranasal modified-live virus vaccine (MLV) and differentially boosted with a systemic MLV or an inactivated vaccine (KV).
Eighteen commercial Black Angus steers.
Calves were mucosally primed at ~24 h of age with an MLV and boosted by injection of a MLV (IN-MLV) or inactivated vaccine (IN-KV) at an average age of 54 d. Challenge occurred at weaning with a virulent non-cytopathic BVDV-2 strain, 24515.
Clinically, the IN-KV group had a longer duration of fever, leukopenia, and viremia, whereas the IN-MLV group had greater BVDV Types-1 and -2 heterospecific antibody responses.
Altogether, these data indicated that systemic MLV boosting resulted in a more robust protection to BVDV Type-2 challenge at weaning.
Mucosal prime-boosting of neonatal calves provided protection against BVDV Type-2 challenge at weaning.
Efficacité comparative des vaccins vivants modifiés et inactivés pour stimuler les réponses épargnant la maladie à la provocation par le virus de la diarrhée virale bovine chez des veaux de boucherie sevrés sensibilisés par voie mucosale en période néo-natale.
Cette étude compare les réponses immunitaires et cliniques des veaux de boucherie positifs au virus de la diarrhée virale bovine (BVDV) dus aux anticorps maternels (MatAb), sensibilisés avec un vaccin intranasal à virus vivant modifié (MLV) et différentiellement stimulés avec un vaccin MLV systémique ou un vaccin inactivé (KV).
Dix-huit bouvillons commerciaux Black Angus.
Les veaux ont été sensibilisés par voie mucosale à environ 24 h d’âge avec un MLV et ont reçu un rappel par injection d’un MLV (IN-MLV) ou d’un vaccin inactivé (IN-KV) à un âge moyen de 54 jours. L’épreuve a eu lieu au sevrage avec une souche virulente non cytopathique de BVDV-2, 24515.
Cliniquement, le groupe IN-KV présentait une durée plus longue de fièvre, de leucopénie et de virémie, tandis que le groupe IN-MLV présentait des réponses en anticorps hétérospécifiques BVDV de types 1 et 2 plus importantes.
Dans l’ensemble, ces données ont indiqué que le renforcement par le vaccin MLV systémique entraînait une protection plus robuste contre la provocation par le BVDV de type 2 au sevrage.
La stimulation mucosale des veaux nouveau-nés a fourni une protection contre la provocation par le BVDV de type 2 au sevrage.(Traduit par Dr Serge Messier).