Seroprevalence and risk factors of Neospora caninum and Bovine Viral Diarrhoea Virus in smallholder dairy cattle in Kenya

Little is known of the risk factors associated with occurrence of Neospora caninum and Bovine Viral Diarrhoea Virus (BVDV) infection in Kenya. This cross-sectional study hypothesized that there are significant biosecurity measures associated with N. caninum and BVDV infections on smallholder dairy farms in Kenya that could be adopted to reduce seroprevalence and impacts. From 158 randomly selected farms in Meru County, Kenya, 470 serum samples were collected from dairy cattle (over six months of age and unvaccinated for these two pathogens). Sera were analyzed for antibodies to N. caninum and antibodies and antigens to BVDV. Data on risk factors were obtained through face-to-face interviews with the farmers. Multivariable logistic regression models were used to identify significant risk factors associated with seropositivity for the pathogens. The apparent seroprevalence of N. caninum, BVDV antibody, BVDV antigen, and co-infection with N. caninum and BVDV antibody and/or antigen were 35.1%, 47.1%, 36.2% and 18.5%, respectively. Risk factors associated with N. caninum antibody included: introducing milking cows into the farm, lending of cattle between farms, farm dogs having access to bovine aborted fetuses, and dogs whelping in the farm compound, with an interaction between the last two variables. BVDV antigen was associated with cattle having contact with pigs, and an interaction between cattle age and whether farms introduced new calves onto farms, and cattle age and whether visiting dairy farmers have access to the cow shed. Cows had higher odds of having BVDV antibodies compared to heifers. Factors associated with co-infection included cow parity, direct contact between dairy cattle, dogs and goats, and introducing new milking cows into the farms. Antibody and antigen results may be partly a function of classical swine fever virus or border disease virus interactions. Farmer education on these biosecurity measures is recommended, along with introduction of BVDV vaccination.

Identification and Genetic Characterization of Viral Pathogens in Ruminant Gestation Abnormalities, Israel, 2015–2019

Infectious agents including viruses are important abortifacients and can cause fetal abnormalities in livestock animals. Here, samples that had been collected in Israel from aborted or malformed ruminant fetuses between 2015 and 2019 were investigated for the presence of the following viruses: the reoviruses bluetongue virus (BTV) and epizootic hemorrhagic disease virus (EHDV), the flaviviruses bovine viral diarrhea virus (BVDV) and border disease virus (BDV), the peribunyaviruses Shuni virus (SHUV) and Akabane virus (AKAV), bovine herpesvirus type 1 (BoHV-1) and bovine ephemeral fever virus (BEFV). Domestic (cattle, sheep, goat) and wild/zoo ruminants were included in the study. The presence of viral nucleic acid or antigen could be confirmed in 21.8 % of abnormal pregnancies (213 out of 976 investigated cases), with peribunyaviruses, reoviruses and pestiviruses being the most prevalent. At least four different BTV serotypes were involved in abnormal courses of pregnancy in Israel. The subtyping of pestiviruses revealed the presence of two BDV and several distinct BVDV type 1 strains. The peribunyaviruses AKAV and SHUV were identified annually throughout the study period, however, variation in the extent of virus circulation could be observed between the years. In 2018, AKAV even represented the most detected pathogen in cases of small domestic ruminant gestation abnormalities. In conclusion, it was shown that various viruses are involved in abnormal courses of pregnancy in ruminants in Israel.

Benefit of Bovine Viral Diarrhoea (BVD) Eradication in Cattle on Pestivirus Seroprevalence in Sheep

Bovine viral diarrhoea virus (BVDV) and Border disease virus (BDV) are closely related pestiviruses of cattle and sheep, respectively. Both viruses may be transmitted between either species, but control programs are restricted to BVDV in cattle. In 2008, a program to eradicate bovine viral diarrhoea (BVD) in cattle was started in Switzerland. As vaccination is prohibited, the cattle population is now widely naïve to pestivirus infections. In a recent study, we determined that nearly 10% of cattle are positive for antibodies to BDV. Here, we show that despite this regular transmission of BDV from small ruminants to cattle, we could only identify 25 cattle that were persistently infected with BDV during the last 12 years of the eradication program. In addition, by determining the BVDV and BDV seroprevalence in sheep in Central Switzerland before and after the start of the eradication, we provide evidence that BVDV is transmitted from cattle to sheep, and that the BVDV seroprevalence in sheep significantly decreased after its eradication in cattle. While BDV remains endemic in sheep, the population thus profited at least partially from BVD eradication in cattle. Importantly, on a national level, BVD eradication does not appear to be generally derailed by the presence of pestiviruses in sheep. However, with every single virus-positive cow, it is necessary to consider small ruminants as a potential source of infection, resulting in costly but essential investigations in the final stages of the eradication program.

Non-Bovine Species and the Risk to Effective Control of Bovine Viral Diarrhoea (BVD) in Cattle

Bovine viral diarrhoea virus (BVDV) is an economically important and highly prevalent virus of domestic cattle. Infections with BVDV may lead to both, reproductive and immunological effects that can result in widespread calf losses and increased susceptibility to diseases, such as mastitis and respiratory disease. While BVDV is generally considered to be host specific, it and other Pestivirus species, such as Border disease virus (BDV) in sheep, have been shown to be infecting species other than those from which they were originally isolated from. Recently BVDV was placed on the OIE’s list of notifiable disease and control and eradication programmes for BVDV have been developed throughout much of Europe, the United States, and the United Kingdom. While some countries, including Sweden and Ireland have successfully implemented eradication programmes, other countries such as New Zealand and Australia are still in the early stages of BVDV control. Despite effective control methods, incursions of BVDV into previously cleared herds still occur. While the cause of these incursions is often due to lapses in control methods, the ability of ruminant pestiviruses to infect species other than cattle poses the question as to whether non-bovine species could be impeding the success of BVDV eradication and control. As such, the aim of this review is to make mention of what is known about the cross-species transmission of BVDV, BDV and other pestiviruses between cattle and non-bovine ungulate species and draw conclusions as to the risk non-bovine species pose to the successful control and eradication of BVDV from cattle.

Eradication of Bovine Viral Diarrhoea (BVD) in Cattle in Switzerland: Lessons Taught by the Complex Biology of the Virus

Bovine viral diarrhoea virus (BVDV) and related ruminant pestiviruses occur worldwide and cause considerable economic losses in livestock and severely impair animal welfare. Switzerland started a national mandatory control programme in 2008 aiming to eradicate BVD from the Swiss cattle population. The peculiar biology of pestiviruses with the birth of persistently infected (PI) animals upon in utero infection in addition to transient infection of naïve animals requires vertical and horizontal transmission to be taken into account. Initially, every animal was tested for PI within the first year, followed by testing for the presence of virus in all newborn calves for the next four years. Prevalence of calves being born PI thus diminished substantially from around 1.4% to <0.02%, which enabled broad testing for the virus to be abandoned and switching to economically more favourable serological surveillance with vaccination being prohibited. By the end of 2020, more than 99.5% of all cattle farms in Switzerland were free of BVDV but eliminating the last remaining PI animals turned out to be a tougher nut to crack. In this review, we describe the Swiss BVD eradication scheme and the hurdles that were encountered and still remain during the implementation of the programme. The main challenge is to rapidly identify the source of infection in case of a positive result during antibody surveillance, and to efficiently protect the cattle population from re-infection, particularly in light of the endemic presence of the related pestivirus border disease virus (BDV) in sheep. As a consequence of these measures, complete eradication will (hopefully) soon be achieved, and the final step will then be the continuous documentation of freedom of disease.

Identification of a Common Conformational Epitope on the Glycoprotein E2 of Classical Swine Fever Virus and Border Disease Virus

Classical swine fever virus (CSFV) shares high structural and antigenic homology with bovine viral diarrhea virus (BVDV) and border disease virus (BDV). Because all three viruses can infect swine and elicit cross-reactive antibodies, it is necessary to differentiate among them with regard to serological diagnosis of classical swine fever. To understand the mechanism of cross-reactivity, it is important to define common or specific epitopes of these viruses. For this purpose, epitope mapping of six monoclonal antibodies (mAbs) was performed using recombinant expressed antigenic domains of CSFV and BDV E2 proteins. One CSFV-specific conformational epitope and one CSFV and BDV common epitope within domain B/C of E2 were identified. Site-directed mutagenesis confirmed that residues G725 and V738/I738 of the CSFV-specific epitope and P709/L709 and E713 of the second epitope are important for mAbs binding. Infection of CSFV in porcine cells was significantly reduced after pre-incubation of the cells with the domain B/C of E2 or after pre-incubation of CSFV with the mAbs detecting domain B/C. 3D structural modeling suggested that both epitopes are exposed on the surface of E2. Based on this, the identified epitopes represent a potential target for virus neutralization and might be involved in the early steps of CSFV infection.

In vitro Inhibition of Border Disease Virus Replication With Lentivirus-Mediated shRNAs

Background: Border disease is believed to be one of the most important diseases in the animal husbandry industry, which has not yet been eradicated in Iran. The development of approaches based on the application of interfering RNA (RNAi) for antiviral therapy has attracted a great deal of attention over the recent years. The present research was conducted to design, construct, and apply shRNA against the NS3 gene of BDV to evaluate the prevention of BDV proliferation in the cell culture system. For this purpose, the suitable oligonucleotide sequence of NS3 gene coding was selected utilizing BDV- X818 strain. Afterwards, using shRNA design software, shRNA molecules were designed and synthesized. These shRNAs were cloned into the desired vectors and were finally transfected in HEK293T cells employing the third generation of lentiviral packaging system. Subsequently, these shRNA expressing lentiviruses were transduced to the MDBK cell line to challenge to border virus. In order to evaluate the efficacy of shRNAs, the viral infectious titer and RNA copy number were calculated with TCID50 and Real-time RT-PCR tests, respectively.Results: The results revealed that shRNAs 1, 2, and 3 decreased viral RNA by more than 90% compared to the control groups. BDV titer noticeably decreased after the challenge with shRNAs 1, 2, and 3 from ~88% up to 99% in comparison with the control groups.Conclusions: Overall, it could be concluded that RNAi may be considered as a strong treatment proposal against viruses, such as BDV.

Positively Charged Amino Acids in the Pestiviral Erns Control Cell Entry, Endoribonuclease Activity and Innate Immune Evasion

The genus Pestivirus, family Flaviviridae, includes four economically important viruses of livestock, i.e., bovine viral diarrhea virus-1 (BVDV-1) and -2 (BVDV-2), border disease virus (BDV) and classical swine fever virus (CSFV). Erns and Npro, both expressed uniquely by pestiviruses, counteract the host’s innate immune defense by interfering with the induction of interferon (IFN) synthesis. The structural envelope protein Erns also exists in a soluble form and, by its endoribonuclease activity, degrades immunostimulatory RNA prior to their activation of pattern recognition receptors. Here, we show that at least three out of four positively-charged residues in the C-terminal glycosaminoglycan (GAG)-binding site of BVDV-Erns are required for efficient cell entry, and that a positively charged region more upstream is not involved in cell entry but rather in RNA-binding. Moreover, the C-terminal domain on its own determines intracellular targeting, as GFP fused to the C-terminal amino acids of Erns was found at the same compartments as wt Erns. In summary, RNase activity and uptake into cells are both required for Erns to act as an IFN antagonist, and the C-terminal amphipathic helix containing the GAG-binding site determines the efficiency of cell entry and its intracellular localization.

Comparative Analysis of Tunisian Sheep-Like Virus, Bungowannah Virus and Border Disease Virus Infection in the Porcine Host

Apart from the established pestivirus species Pestivirus A to Pestivirus K novel species emerged. Pigs represent not only hosts for porcine pestiviruses, but are also susceptible to bovine viral diarrhea virus, border disease virus (BDV) and other ruminant pestiviruses. The present study focused on the characterization of the ovine Tunisian sheep-like virus (TSV) as well as Bungowannah virus (BuPV) and BDV strain Frijters, which were isolated from pigs. For this purpose, we performed genetic characterization based on complete coding sequences, studies on virus replication in cell culture and in domestic pigs, and cross-neutralization assays using experimentally derived sera. TSV forms a distinct phylogenetic group more closely related to Pestivirus C (classical swine fever virus, CSFV) than to Pestivirus D (BDV). In contrast to BDV and BuPV, TSV replicates by far more efficiently on ovine than on porcine cells. Nevertheless, pigs were susceptible to TSV. As a consequence of close antigenic relatedness of TSV to CSFV, cross-reactivity was detected in CSFV-specific antibody assays. In conclusion, TSV is genetically closely related to CSFV and can replicate in domestic pigs. Due to close antigenic relatedness, field infections of pigs with TSV and other ruminant pestiviruses can interfere with serological diagnosis of classical swine fever.