Genus Pestivirus Research Articles

Caracterización genética del virus de la diarrea viral bovina detectado en granjas de traspatio en México

Infection by bovine viral diarrhea virus (BVDV) in cattle remains a source of significant economic losses for livestock producers. This virus is classified within the Pestivirus genus, including three main species: Pestivirus A (BVDV-1), Pestivirus B (BVDV-2), and Pestivirus H (HoBi-like pestivirus). Here, we performed a molecular epidemiological investigation aiming to evaluate the genetic diversity of BVDV in cattle from backyard farms in a municipality in Mexico named Tepalcingo, Morelos, with records of reproductive disorders. RT-PCR was conducted in 111 serum samples from affected cattle. Viral RNA was detected in 47.74% of the samples analyzed. Sequencing and phylogenetic analysis based on 5'UTR showed that the circulating subgenotype was BVDV-1a in all positive samples. These findings reveal the prevalence of BVDV in the surveyed population; thus suggesting a possible association with the previous records of reproductive manifestation in the herd. However, further studies are needed to confirm BVDV as the causative agent. Additionally, our results represent a helpful tool for designing control and prevention strategies accurate to the current regional epidemiological situation. Moreover, obtained information from this type of epidemiological study will assist the implementation of biosafety measures on backyard farms with limited economic resources.

Classical swine fever (CSF): towards new challenges

Classical swine fever (CSF) is a highly contagious swine-specific disease which may have a huge economic impact for porcine production. CSF is caused by a virus belonging to the Pestivirus genus, which has expanded for the past 5 years with the discovery of new species whose genetic proximity to the CSF virus could further complicate laboratory diagnosis. The various forms of the disease, and in particular the increased frequency of attenuated forms, linked to an evolution of CSF virus strains towards a reduction in their virulence, delay clinical diagnosis. Thus, a long period may elapse before an outbreak is detected, allowing the virus to circulate longer, with the risk of spreading to distant geographical areas. Efforts must be maintained in terms of surveillance and diagnostic tools development in order to detect CSF virus infection early and thus limit the spread of the disease and facilitate control measures.

Involvement of the Wnt pathway in BVDV cytopathogenic strain replication in primary bovine cells

BackgroundBovine viral diarrhea virus 1 (BVDV-1) of the pestivirus genus is an economically crippling virus in the cattle industry; this positive RNA virus causes mucosal disease resulting in reproductive losses and other disease syndromes. The pathogenesis mechanism of the disease caused by BVDV infection is not well understood; for a better understanding of in vivo host BVDV-1 interactions, we conducted a transcriptomic study of infected cells at different times post-infection.MethodsWe compared the permissiveness and cellular response of a BVDV-1 cytopathogenic strain on Madin-Darby Bovine Kidney cells (MDBK) and bovine lung primary cells, a model closer to in vivo infection. Then a RNAseq analysis was realized on the infected bovine lung primary cells, at 10 hpi and 30 hpi (hours post-infection), to identify transcriptomic signatures.ResultsRNAseq analysis on BVDV-1 infected bovine primary cells showed 2,759 and 5,376 differentially expressed genes at respectively 10 hpi and 30 hpi with an absolute Fold Change ≥ 2. Among the different pathways deregulated, data analysis revealed a deregulation of Wnt signaling pathway, a conserved process that play a critical role in embryogenesis, cellular proliferation, and differentiation as well as in viral responses against viruses such as Influenza or Hepatitis C. We demonstrated here that the deregulation of the Wnt/βcatenin signaling pathway plays a role in viral replication of BVDV cp strain. Interestingly, we showed that the inhibition of this Wnt pathway using two inhibitors, FZM1 and iCRT14, induced a delay in onset of the establishment of a cytopathic effect of primary cells.ConclusionsThereby, this study highlighted a role of the Wnt signaling pathway in the BVDV-1 viral replication in bovine cells, suggesting an interesting option to explore as a new therapeutic target.

A Novel Competitive ELISA for Specifically Measuring and Differentiating Immune Responses to Classical Swine Fever C-Strain Vaccine in Pigs.

Classical swine fever can be controlled effectively by vaccination with C-strain vaccine. In this study, we developed a novel competitive enzyme-linked immunosorbent assay (cELISA) based on a C-strain Erns specific monoclonal antibody (mAb 1504), aiming to serologically measure immune responses to C-strain vaccine in pigs, and finally to make the C-strain become a DIVA-compatible vaccine. The cELISA system was established based on the strategy that mAb 1504 will compete with the C-strain induced antibodies in the pig serum to bind the C-strain Erns protein. The cELISA was optimized and was further evaluated by testing different categories of pig sera. It can efficiently differentiate C-strain immunized from wild-type CSFV-infected pigs and lacks cross-reaction with other common swine viruses and viruses in genus Pestivirus such as Bovine viral diarrhea virus (BVDV). The C-strain antibody can be tested in pigs 7–14 days post vaccination with this cELISA. The sensitivity and specificity of the established cELISA were 100% (95% confidence interval: 95.60 to 100%) and 100% (95% confidence interval: 98.30 to 100%), respectively. This novel cELISA is a reliable tool for specifically measuring and differentiating immune responses to C-strain vaccine in pigs. By combining with the wild-type CSFV-specific infection tests, it can make the C-strain have DIVA capability.

The NS4A Protein of Classical Swine Fever Virus Suppresses RNA Silencing in Mammalian Cells.

RNA interference (RNAi) is a significant posttranscriptional gene silencing mechanism and can function as an antiviral immunity in eukaryotes. However, numerous viruses can evade this antiviral RNAi by encoding viral suppressors of RNA silencing (VSRs). Classical swine fever virus (CSFV), belonging to the genus Pestivirus, is the cause of classical swine fever (CSF), which has an enormous impact on animal health and the pig industry. Notably, little is known about how Pestivirus blocks RNAi in their host. In this paper, we uncovered that CSFV NS4A protein can antagonize RNAi efficiently in mammalian cells by binding to double-stranded RNA and small interfering RNA. In addition, the VSR activity of CSFV NS4A was conserved among Pestivirus. Furthermore, the replication of VSR-deficient CSFV was attenuated but could be restored by the deficiency of RNAi in mammalian cells. In conclusion, our studies uncovered that CSFV NS4A is a novel VSR that suppresses RNAi in mammalian cells and shed new light on knowledge about CSFV and other Pestivirus. IMPORTANCE It is well known that RNAi is an important posttranscriptional gene silencing mechanism that is also involved in the antiviral response in mammalian cells. While numerous viruses have evolved to block this antiviral immunity by encoding VSRs. Our data demonstrated that the NS4A protein of CSFV exhibited a potent VSR activity through binding to dsRNA and siRNA in the context of CSFV infection in mammalian cells, which are a conservative feature among Pestivirus. In addition, the replication of VSR-deficient CSFV was attenuated but could be restored by the deficiency of RNAi, providing a theoretical basis for the development of other important attenuated Pestivirus vaccines.

The first study on analysis of the codon usage bias and evolutionary analysis of the glycoprotein envelope E2 gene of seven Pestiviruses.

Background and Aim:Pestivirus, a genus of the Flaviviridae family, comprises viruses that affect bovines, sheep, and pigs. Symptoms, including hemorrhagic syndromes, abortion, respiratory complications, and deadly mucosal diseases, are produced in infected animals, which cause huge economic losses to the farmers. Bovine viral diarrhea virus-1, bovine viral diarrhea virus-2, classical swine fever virus, border disease virus, Bungowannah, Hobi-like, and atypical porcine pestivirus belonging to the Pestivirus genus were selected for the study. This study aimed to estimate the codon usage bias and the rate of evolution using the glycoprotein E2 gene. Furthermore, codon usage bias analysis was performed using publicly available nucleotide sequences of the E2 gene of all seven Pestiviruses. These nucleotide sequences might elucidate the disease epidemiology and facilitate the development of designing better vaccines.Materials and Methods:Coding sequences of the E2 gene of Pestiviruses A (n = 89), B (n = 60), C (n = 75), D (n = 10), F (n = 07), H (n = 52), and K (n = 85) were included in this study. They were analyzed using different methods to estimate the codon usage bias and evolution. In addition, the maximum likelihood and Bayesian methodologies were employed to analyze a molecular dataset of seven Pestiviruses using a complete E2 gene region.Results:The combined analysis of codon usage bias and evolutionary rate analysis revealed that the Pestiviruses A, B, C, D, F, H, and K have a codon usage bias in which mutation and natural selection have played vital roles. Furthermore, while the effective number of codons values revealed a moderate bias, neutrality plots indicated the natural selection in A, B, F, and H Pestiviruses and mutational pressure in C, D, and K Pestiviruses. The correspondence analysis revealed that axis-1 significantly contributes to the synonymous codon usage pattern. In this study, the evolutionary rate of Pestiviruses B, H, and K was very high. The most recent common ancestors of all Pestivirus lineages are 1997, 1975, 1946, 1990, 2004, 1990, and 1990 for Pestiviruses A, B, C, D, F, H, and K, respectively. This study confirms that both mutational pressure and natural selection have played a significant role in codon usage bias and evolutionary studies.Conclusion:This study provides insight into the codon usage bias and evolutionary lineages of pestiviruses. It is arguably the first report of such kind. The information provided by the study can be further used to elucidate the respective host adaptation strategies of the viruses. In turn, this information helps study the epidemiology and control methods of pestiviruses.

Transcriptomic Analysis of MDBK Cells Infected with Cytopathic and Non-Cytopathic Strains of Bovine Viral Diarrhea Virus (BVDV).

Bovine viral diarrhea virus (BVDV) belongs to the Flaviviridae family and the Pestivirus genus. Infection with BVDV causes a disease with a wide spectrum of clinical symptoms, most often mild, although infections with this virus constitute a serious economic problem all over the world. The virus is characterized by a high genetic variability, while the accumulation of single mutations leads to the formation of its new variants. The aim of this study was to better understand the complicated pathogenesis of this disease at the molecular level via the analysis of the transcriptome of cells infected with this virus. The bovine kidney cell line (MDBK), the cytopathic (cp) reference strain, and two non-cytopathic (ncp) BVD virus field strains were used in transcriptomic studies. The cell transcriptome was tested 24 and 72 h after infection. The results of the microarray analysis revealed changes in the expression levels of numerous genes. Genes with changed expression as a result of infection with the cp strain caused changes in the expression levels of a large number of genes and enriched a number of pathways. Genes with increased expression levels were enriched among other pathways involved in the cell cycle, while genes with reduced expression levels enriched pathways mostly related to metabolism. Genes with increased expression levels as a result of infection with ncp strains enriched a much smaller number of pathways, among them, pathways related to signaling activity 24 h post-infection and serine biosynthetic pathways both 24 and 72 h post-infection. Pathways enriched by genes with reduced expression levels were related to the innate immune response (72 h post-infection) or metabolism (24 and 72 h post-infection). The results of microarray studies can help us to better understand the host’s response to BVDV infection.

Evaluation of antibody and antigen cross-reaction in Kenyan dairy cattle naturally infected with two pestiviruses: Bovine viral diarrhea virus and classical swine fever virus.

Background and Aim:Bovine viral diarrhea virus (BVDV) and classical swine fever virus (CSFV) are important pathogens of cattle and pigs, respectively, and belong to the genus Pestivirus. As CSFV has been shown to infect cattle, it can create diagnostic challenges of BVDV results through possible cross-reactivity where cattle could be exposed to pigs and CSFV. This study aimed to determine the possible cross-reactivity of BVDV and CSFV enzyme-linked immunosorbent assay (ELISA) results for antigen (Ag) and antibody (Ab) among smallholder dairy cattle in Kenya.Materials and Methods:This was a cross-sectional study based on a single visit to farms to collect serum samples and other descriptive farm-level and animal-level information. Testing for BVDV Ag and Ab was conducted on serum samples from 320 dairy cows and heifers, with CSFV Ag and Ab testing conducted on a subset of 133 and 74 serum samples, respectively. CSFV testing was based on BVDV test results and the availability of enough sample volume from farms that kept pigs. The Ag and Ab tests utilized IDEXX ELISA for both BVDV and CSFV.Results:For the 74 samples with Ab tests for both viruses, 40 (54.0%) were BVDV Ab positive, while 63 (85.1%) were CSFV Ab positive. Of the 40 BVDV Ab positive samples, 36 cattle (90.0%) tested positive for CSFV Ab. However, of the 34 BVDV Ab negative samples, 27 (79.4%) were CSFV Ab test-positive. For the 133 samples with Ag tests for both viruses, 125 (94.0%) were BVDV Ag positive, while 2 (1.5%) samples were CSFV Ag positive. None of the eight BVDV Ag negative samples was positive for CSFV Ag and only two (1.6%) of the 125 BVDV Ag positive samples were positive for CSFV Ag.Conclusion:The results indicate either substantial cross-reactivity of the two Ab ELISA tests, or reactivity with some other protein in the samples that led to the positive Ab test results. There was only limited evidence for cross-reactivity of the two Ag ELISA tests. We recommend that Pestivirus genus cross-reactivity be considered when interpreting BVDV ELISA results in cattle, more for Ab than Ag tests. Further research is needed to clarify the levels of cross-reactivity between BVDV and other Pestivirus Ag and Ab tests from animals on mixed-species farms.

Genetic characterization of atypical porcine pestivirus from neonatal piglets with congenital tremor in Hubei province, China

BackgroundAtypical porcine pestivirus (APPV) is a single-stranded RNA virus with high genetic variation that causes congenital tremor (CT) in newborn piglets, belonging to the genus Pestivirus of the family Flaviviridae. Increasing cases of APPV infection in China in the past few years would pose severe challenges to the development of pig production. In view of the high genetic variability of APPV, the genetic characteristics of APPV in Hubei province was determined.Methods52 tissue samples from 8 CT-affected newborn piglets were collected at two different periods in the same pig farm in Hubei province. Viral nucleic acid was extracted to detect pathogens that can cause CT in piglets or other common clinical pathogens by RT-PCR. Haematoxylin and eosin (HE) staining, immunohistochemical (IHC) analysis, and qRT-PCR were performed to observe histopathological changes and histological distribution, and detect the viral load of APPV in CT-affected piglets. The full-length genome of APPV was obtained and sequence analysis was conducted to determine the phylogenetic relationship.ResultsHistopathological observation and histological distribution analysis showed that the histological lesions and distribution of APPV were mainly in central nervous system (CNS) tissues and immune tissues. Viral load analysis revealed that the viral copy number was higher in the cerebellum, submaxillary lymph nodes, tonsil, and serum than in other tissues. Phylogenetic analysis showed that CH-HB2020 and CH-HB2021 belonged to Clade I.3, and is most closely related to APPV_CH-GX2016. Sequence alignment based on APPV encoding sequences (CDS) showed that the nucleotide identities of CH-HB2020 or CH-HB2021 with Clade I, Clade II, and Clade III strains were 83.5–98.6%, 83.1–83.5%, and 81.1–81.4%, respectively, while the amino acid identities were 91.9–99.2%, 91.2–95.3%, and 90.77–91.4%, respectively. No recombination event was observed in CH-HB2020 or CH-HB2021 strains.ConclusionsThese findings enhance our understanding of the pathogenesis of APPV and may provide potential molecular evidence for its prevalence and transmission.

Genetic diversity and distribution of bovine pestiviruses (Flaviviridae: Pestivirus) in the world and in the Russian Federation

The genus Pestivirus of the family Flaviviridae includes 11 species. Bovine pestiviruses are the causative agents of viral diarrhea/mucosal disease and include three genetically distinct species: pestivirus A (BVDV-1), B (BVDV-2), and H (BVDV-3). The number of BVDV-1 subtypes is 21, BVDV-2 - 4, and BVDV-3 - 4, which complicates the diagnosis of associated diseases, reduces the effectiveness of vaccination and control programs.We performed the search in the PubMed, Web of Science, Scopus, eLIBRARY.RU databases for articles published in 2000-2021.Pestivirus A is distributed everywhere, although the largest number of subtypes was found in cattle in Italy and China. The virus is widespread in the Central region of the Russia (subtypes 1a and 1m). In Siberia, eleven subtypes circulate among native and imported animals: 1a (5%), 1b (35%), 1c (5%), 1d (10%), 1f (20%), 1g, 1i (both 2.5%), 1j, 1k, 1p, and 1r (all for 5%). Pestivirus B subtype is more virulent, found less frequently and mainly in the North and South America, in some European countries, and in Asia. Three subtypes have been identified in Siberia: 2a (25%), 2b (10%), and 2c (5%). Pestivirus H circulates in Europe, Asia and South America. The main route of entry is contaminated biological products. In Russia, BVDV-3 of the Italian-Brazilian group (3a) was detected in 7 lots of fetal bovine serum.The role of the virus in the occurrence of respiratory diseases in calves, abortion, systemic infection and enteritis in calves and adult animals has been established. The source of the virus in such cases was a contaminated modified live vaccine.

Fifty Shades of Erns: Innate Immune Evasion by the Viral Endonucleases of All Pestivirus Species.

The genus Pestivirus, family Flaviviridae, includes four historically accepted species, i.e., bovine viral diarrhea virus (BVDV)-1 and -2, classical swine fever virus (CSFV), and border disease virus (BDV). A large number of new pestivirus species were identified in recent years. A common feature of most members is the presence of two unique proteins, Npro and Erns, that pestiviruses evolved to regulate the host’s innate immune response. In addition to its function as a structural envelope glycoprotein, Erns is also released in the extracellular space, where it is endocytosed by neighboring cells. As an endoribonuclease, Erns is able to cleave viral ss- and dsRNAs, thus preventing the stimulation of the host’s interferon (IFN) response. Here, we characterize the basic features of soluble Erns of a large variety of classified and unassigned pestiviruses that have not yet been described. Its ability to form homodimers, its RNase activity, and the ability to inhibit dsRNA-induced IFN synthesis were investigated. Overall, we found large differences between the various Erns proteins that cannot be predicted solely based on their primary amino acid sequences, and that might be the consequence of different virus-host co-evolution histories. This provides valuable information to delineate the structure-function relationship of pestiviral endoribonucleases.

Antiviral Effects of 5-Aminolevulinic Acid Phosphate against Classical Swine Fever Virus: In Vitro and In Vivo Evaluation.

The inhibitory effects of 5-aminolevulinic acid phosphate (5-ALA), an important amino acid for energy production in the host, against viral infections were previously reported. Here, the antiviral effects of 5-ALA against classical swine fever virus (CSFV) belonging to the genus Pestivirus in the Flaviviridae family and its possible mechanisms were investigated. CSFV replication was suppressed in swine cells supplemented with 5-ALA or its metabolite, protoporphyrin IX (PPIX). The infectivity titer of CSFV was decreased after mixing with PPIX extracellularly. In addition, the activities of the replication cycle were decreased in the presence of PPIX based on the CSFV replicon assay. These results showed that PPIX exerted antiviral effects by inactivating virus particles and inhibiting the replication cycle. To evaluate the in vivo efficacy of 5-ALA, pigs were supplemented daily with 5-ALA for 1 week before virus inoculation and then inoculated with a virulent CSFV strain at the 107.0 50% tissue culture infectious dose. The clinical scores of the supplemented group were significantly lower than those of the nonsupplemented group, whereas the virus growth was not. Taken together, 5-ALA showed antiviral effects against CSFV in vitro, and PPIX played a key role by inactivating virus particles extracellularly and inhibiting the replication cycle intracellularly.

Bungowannah Pestivirus Chimeras as Novel Double Marker Vaccine Strategy against Bovine Viral Diarrhea Virus.

Marker or DIVA (differentiation of infected from vaccinated animals) vaccines are beneficial tools for the eradication of animal diseases in regions with a high prevalence of the designated disease. Bovine viral diarrhea virus (BVDV)-1 (syn. Pestivirus A) is a flavivirus that infects predominantly cattle resulting in major economic losses. An increasing number of countries have implemented BVDV eradication programs that focus on the detection and removal of persistently infected cattle. No efficient marker or DIVA vaccine is yet commercially available to drive the eradication success, to prevent fetal infection and to allow serological monitoring of the BVDV status in vaccinated farms. Bungowannah virus (BuPV, species Pestivirus F), a related member of the genus Pestivirus with a restricted prevalence to a single pig farm complex in Australia, was chosen as the genetic backbone for a marker vaccine candidate. The glycoproteins E1 and E2 of BuPV were substituted by the heterologous E1 and E2, which are major immunogens, of the BVDV-1 strain CP7. In addition, the candidate vaccine was further attenuated by the introduction of a deletion within the Npro protein coding sequence, a major type I interferon inhibitor. Immunization of cattle with the chimeric vaccine virus BuPV_ΔNpro_E1E2 CP7 (modified live or inactivated) followed by a subsequent experimental challenge infection confirmed the safety of the prototype strain and provided a high level of clinical protection against BVDV-1. The serological discrimination of vaccinated cattle could be enabled by the combined detection of BVDV-1 E2- in the absence of both BVDV NS3- and BVDV Erns-specific antibodies. The study demonstrates for the first time the generation and application of an efficient BVDV-1 modified double marker vaccine candidate that is based on the genetic background of BuPV accompanied by commercially available serological marker ELISA systems.

Different impact of bovine complement regulatory protein 46 (CD46bov) as a cellular receptor for members of the species Pestivirus H and Pestivirus G

ABSTRACT The genus Pestivirus within the family Flaviviridae comprises highly relevant animal pathogens such as bovine viral diarrhoea virus 1 and 2 (BVDV-1 and -2) classified into the two species Pestivirus A and Pestivirus B, respectively. First described in 2004, HoBi-like pestiviruses (HoBiPeV) represent emerging bovine pathogens that belong to a separate species (Pestivirus H), but share many similarities with BVDV-1 and -2. Additionally, two giraffe pestivirus (GPeV) strains both originating from Kenya represent another distinct species (Pestivirus G), whose members replicate very efficiently in bovine cells. In this study, we investigated the role of bovine complement regulatory protein 46 (CD46bov), the receptor of BVDV-1 and -2, in the entry of HoBiPeV and GPeV. For this purpose, bovine CD46-knockout and CD46-rescue cell lines were generated by CRISPR/Cas9 technology and subsequent trans-complementation, respectively. Our results provide strong evidence that the impact of CD46bov differs between viruses belonging to Pestivirus H and viruses representing Pestivirus G: CD46bov revealed to be a major cellular entry factor for HoBiPeV strain HaVi-20. In contrast, GPeV strain PG-2 presented as largely independent of CD46bov, suggesting a different entry mechanism involving other molecular determinants which remain to be identified. In addition, we demonstrated that, similar to BVDV-1 and -2, virus isolates of both Pestivirus H and Pestivirus G are able to adapt to cell culture conditions by using heparan sulfate to enter the host cell. In conclusion, our findings show that different bovine pestiviruses use diverse mechanisms of host cell entry.

Использование перевиваемых культур клеток ПТ-80 и КСТ для постановки реакции нейтрализации на обнаружение антител к вирусу вирусной диареи крупного рогатого скота

The flavivirus family (from Lat. flavus – yellow) represents a significant group of viruses circulating among vertebrates and insects, consists of three genera, one of which is the genus Pestivirus (classical swine fever virus, viral diarrhea (BVD) of cattle, borderline sheep disease, etc.). Viral diarrhea of cattle is a widespread infection in the world, and It is one of the most difficult economic problems in animal husbandry. The clinical picture of BVD is diverse: respiratory organ damage, fever, leukopenia, thrombocytopenia, enteritis, immunosuppression, the virus passes through the placenta, which leads to the birth of persistently infected calves, abortions in cows at all stages of pregnancy are noted, fetal mummification, producing bulls can be virus carriers and secrete the virus with sperm. For serodiagnostics of viral diarrhea of cattle in laboratory practice, ELISA (enzyme immunoassay), RNGA (indirect hemagglutination reaction), PH (neutralization reaction) are used - the gold standard of diagnostics. The PH method is universal and makes it possible to detect the level of virus neutralizing antibodies for making a retrospective diagnosis, evaluating animals for virus transmission and confirming well-being. The PH method is unique and is the only correct one in assessing the effectiveness of vaccines, since only when it is used, the level of virus neutralizing (protective) antibodies protecting animals from direct infection is revealed. The aim of our research was to formulate a neutralization reaction to the detection of antibodies to the bovine viral diarrhea virus using transplanted cultures of PT-80 and KST cells.

Identification of differentially expressed gene pathways between cytopathogenic and non-cytopathogenic BVDV-1 strains by analysis of the transcriptome of infected primary bovine cells

The bovine viral diarrhea virus 1 (BVDV-1), belonging to the Pestivirus genus, is characterized by the presence of two biotypes, cytopathogenic (cp) or non-cytopathogenic (ncp). For a better understanding of the host pathogen interactions, we set out to identify transcriptomic signatures of bovine lung primary cells (BPCs) infected with a cp or a ncp strain. For this, we used both a targeted approach by reverse transcription droplet digital PCR and whole genome approach using RNAseq. Data analysis showed 3571 differentially expressed transcripts over time (Fold Change >2) and revealed that the most deregulated pathways for cp strain are signaling pathways involved in responses to viral infection such as inflammatory response or apoptosis pathways. Interestingly, our data analysis revealed a deregulation of Wnt signaling pathway, a pathway described in embryogenesis, that was specifically seen with the BVDV-1 cp but not the ncp suggesting a role of this pathway in viral replication.

STUDI KOMPARASI RESPON ANTIBODI PASCA VAKSINASI CLASSICAL SWINE FEVER PADA BABI LANDRACE DAN BABI LOKAL TIMOR DI KABUPATEN KUPANG, NUSA TENGGARA TIMUR

Classical Swine Fever (CSF) is an important infectious disease caused by the CSF virus, which belongs to the Pestivirus genus from the family Flaviviridae. This disease has a high fatality rate in swine. Stamping out the disease and vaccinations aimed at forming and strengthening antibodies against CSF are proven to be effective ways of disease control and prevention. Several factors, including age, immune status, health status, nutrition, and the environment, are believed to influence the success of the immunization program. Furthermore, it is assumed that different pig breeds may have impacts on vaccination success. This study aimed to compare antibody responses after CSF vaccination in two different pig breeds, Landrace and local Timor pigs. A total of 26 pigs (13 pigs for each group) was occupied in this study. All of the pigs used in this study had never been vaccinated against CSF. Antibody titers were quantified using an ELISA test, then the data were compared using an independent t-test to assess the difference of antibody responses between Landrace and local Timor pigs. Finally, the study found no significant difference in antibody response between Landrace and local Timor pigs, with a significant value of 0.07 (P > 0.05). Therefore, it is feasible to conclude that breed does not influence antibody response following CSF vaccination.

Antiviral Effect of Ginsenoside Rb2 and Rb3 Against Bovine Viral Diarrhea Virus and Classical Swine Fever Virus in vitro

Bovine viral diarrhea virus (BVDV) and classical swine fever virus (CSFV) are members of the genus Pestivirus that cause disease in wild and domestic animals and are responsible for extensive economic losses of livestock and biological industry. BVDV is also a significant laboratory contaminant. Currently, no effective antiviral therapeutics are available to control their infection. Ginsenosides, as major pharmacological ingredients in the plants of ginseng, have various biological activities. In the present work, the antiviral activity of 9 ginsenosides and 3 other saponins from Araliaceae plants was investigated against Pestivirus. Ginsenoside Rb2 and Rb3 showed low cytotoxicity and obvious antiviral effect. They were able to inhibit the replication and proliferation of BVDV and CSFV. In addition, our results suggest that the possible antiviral mechanism of Rb2 might be related to its ability to affect the translation of these viruses. Obtained results suggest that ginsenoside Rb2 and Rb3 have a potential for effective treatment against Pestivirus infection.

Detection and localization of atypical porcine pestivirus in the testicles of naturally infected, congenital tremor affected piglets.

Atypical porcine pestivirus (APPV) belongs to the genus Pestivirus within the family Flaviviridae. Recently, APPV has been identified as the causative agent of congenital tremor (CT) type AII. The disease is a neurological disorder that affects newborn piglets and is characterized by generalized trembling of the animals and often splay legs. CT is well known worldwide, and the virus seems to be highly prevalent in major swine producing areas. However, little is known about the epidemiology of the infection, transmission and spread of the virus between herds. Here, we show the high prevalence of APPV in processing fluid samples collected from Hungarian pig herds which led us to investigate the cellular targets of the virus in the testicles of newborn piglets affected by CT. By the development of an RNA in situ hybridization assay and the use of immunohistochemistry on consecutive slides, we identified the target cells of APPV in the testicle: interstitial Leydig cells, peritubular myoid cells and smooth muscle cells of medium‐sized arteries. Previous studies have shown that APPV can be found in the semen of sexually mature boars suggesting the role of infected boars and their semen in the transmission of the virus similar to many other members of the Flaviviridae family. As in our case, the virus has not been identified in cells beyond the Sertoli cell barrier, further studies on infected adult boars’ testicles and other reproductive glands are needed to analyze the possible changes in the cell tropism of APPV that might contribute to its prolonged extraction by the semen beyond the period of viraemia.

First molecular evidence of border disease virus in wild boars in Turkey

Molecular studies on viral diseases in wildlife are limited in Turkey. Pestiviruses infect domestic animals such as pig, cattle, sheep, goats and many other wild ungulates. Cross-species transmission of pestiviruses between wildlife and domestic livestock is a subject of recent concern where wild ungulates are in close contact with domestic ruminants. The International Committee on Virus Taxonomy (ICTV) has named the genus Pestivirus, which belongs to the Flaviviridae family, using the format Pestivirus A, Pestivirus B, Pestivirus C, and so on. Pestivirus A-D replaces Bovine viral diarrhea virus-1 (BVDV-1), Bovine viral diarrhea virus-2 (BVDV-2), Classical swine fever virus (CSFV) and Border disease virus (BDV) respectively. During the 2013–2014 hunting season, a total of 40 samples were collected from wild boars (Sus scrofa ferus) in the area of Western Mediterranean Turkey. In the samples, nucleic acids were investigated for pestivirus, Aujeszky’s disease virus, Borna disease virus, coronavirus, mastadenovirus and rotavirus. RT-PCR was performed using primary sets to detect specific partial gene region specific to each virus. Sequence analysis was performed on a positive sample. Phylogenetic analysis revealed that the positive sample, TR/Burdur/13/Boar3, belonged to BDV genotype 1 (Pestivirus D). The first molecular findings of BDV in wild boars in Turkey are reported in this study. This study highlights the importance of further research into diseases that might be transmitted from wild boars to ruminants in Turkey.