Bovine Viral Diarrhea Virus Genome Research Articles

Bovine viral diarrhoea viruses from New Zealand belong predominantly to the BVDV-1a genotype

Aim To determine which genotypes of bovine viral diarrhoea virus (BVDV) circulate among cattle in New Zealand. Methods Samples comprised BVDV-1-positive sera sourced from submissions to veterinary diagnostic laboratories in 2019 (n = 25), 2020 (n = 59) and 2022 (n = 74) from both beef and dairy herds, as well as archival BVDV-1 isolates (n = 5). Fragments of the 5′ untranslated region (5′ UTR) and glycoprotein E2 coding sequence of the BVDV genome were amplified and sequenced. The sequences were aligned to each other and to international BVDV-1 sequences to determine their similarities and phylogenetic relationships. The 5′ UTR sequences were also used to create genetic haplotype networks to determine if they were correlated with selected traits (location, type of farm, and year of collection). Results The 5′ UTR sequences from New Zealand BVDV were closely related to each other, with pairwise identities between 89% and 100%. All clustered together and were designated as BVDV-1a (n = 144) or BVDV-1c (n = 5). There was no evidence of a correlation between the 5′ UTR sequence and the geographical origin within the country, year of collection or the type of farm. Partial E2 sequences from New Zealand BVDV (n = 76) showed 74–100% identity to each other and clustered in two main groups. The subtype assignment based on the E2 sequence was the same as based on the 5′ UTR analysis. This is the first comprehensive analysis of genomic variability of contemporary New Zealand BVDV based on the analysis of the non-coding (5′ UTR) and coding (E2) sequences. Conclusions and clinical relevance Knowledge of the diversity of the viruses circulating in the country is a prerequisite for the development of effective control strategies, including a selection of suitable vaccines. The data presented suggest that New Zealand BVDV are relatively homogeneous, which should facilitate eradication efforts including selection or development of the most suitable vaccines.

New Insights into the Role and Therapeutic Potential of Heat Shock Protein 70 in Bovine Viral Diarrhea Virus Infection.

Bovine viral diarrhea virus (BVDV), a positive-strand RNA virus of the genus Pestivirus in the Flaviviridae family, is the causative agent of bovine viral diarrhea-mucosal disease (BVD-MD). BVDV's unique virion structure, genome, and replication mechanism in the Flaviviridae family render it a useful alternative model for evaluating the effectiveness of antiviral drugs used against the hepatitis C virus (HCV). As one of the most abundant and typical heat shock proteins, HSP70 plays an important role in viral infection caused by the family Flaviviridae and is considered a logical target of viral regulation in the context of immune escape. However, the mechanism of HSP70 in BVDV infection and the latest insights have not been reported in sufficient detail. In this review, we focus on the role and mechanisms of HSP70 in BVDV-infected animals/cells to further explore the possibility of targeting this protein for antiviral therapy during viral infection.

Molecular Investigation of Bovine Viral Diarrhea Virus, Bovine Herpes Virus-1 and Bovine Herpes Virus-4 Infections in Abortion Cases of Cattle in Van District, Turkey

Abortions, fetal mummification, calf anomalies, and infertility problems constitute most of the reproductive problems in cattle. Viruses play a significant role in the cause of these cases. In cattle, these agents are known as primary abortion agents and the most common of these agents are Bovine Viral Diarrhea Virus (BVDV), Bovine Herpes Virus Type 1 (BoHV-1), and Bovine Herpes Virus Type 4 (BoHV-4). The objective of this research is to determine the potential role of BVDV, BoHV-1, and BoHV-4 as viral abortion agents in cattle housed in the Van district. For this, a total of 115 animal specimens (blood, serum, vaginal swab, vaginal fluid discharge, nasal swab, and abortion material) from 100 abortion, early embryonic deaths, and infertility cases in cattle over the age of 2-5 years old were collected. All samples for detection of BVDV, BoHV-1 and BoHV-4 genomes were tested by the Polymerase Chain Reaction (PCR) technique using specific primers encoding Panpesti 5'-UTR, Glycoprotein C (gC) and Glycoprotein B (gB) genes, respectively. Result out of the samples tested, 41.73% were positive for BVDV and all samples were negative for BoHV-1 and BoHV-4. In conclusion, the presence of BVDV in cattle in the Van region and its role in the occurrence of abortion cases was emphasized for the first time. It is necessary to the consideration of viral abortions and determine the etiology of abortion cases and genital system problems. According to this, we need to focus on the detection of persistently infected (PI) animals for prevention and control of infection and the most effective way of vaccinating susceptible populations.

Bovine viral diarrhea virus in China: A comparative genomic and phylogenetic analysis with complete genome sequences.

Bovine viral diarrhea virus (BVDV), causing bovine viral diarrhea (BVD) in cattle, is one of the highly contagious and devastating diseases of cattle. Since 1980, BVDV has been identified all-over China in a variety of animal species including cattle, camels, yaks, sheep, water buffalo, goats, Sika deer and pigs. In this study, 31 BVDV complete genomes reported in China (from 2004 to 2020) with other 112 genomes reported around the world were comparatively analyzed. Phylogenetic analysis shows that BVDV genomes reported worldwide clustered in three major clades i.e., BVDV-1, BVDV-2, and BVDV-3. The BVDV-1 is genetically the most diverged genotype and phylogenetically classified into 7 sub-clades in our study based on full-length genomes. The China BVDV genomes fall into all three major clades, e.g., BVDV-1, BVDV-2 and BVDV-3. China BVDV-1 clustered into five sub-clades, e.g., 1, 2, 3, 6 and 7, where sub-clade 7 clustered as a separate sub-clade. Full-length genome recombination analysis reveals that the BVDV-1 reported in China appears to be mainly involved in recombination events. In addition, comparative analysis of E2 proteins between BVDV-1, BVDV-2, and BVDV-3 reveals that the amino acid variations could affect 12 potential linear B cell epitopes, demonstrating a dramatic antigen drift in the E2 protein. These results provide a thorough view of the information about the genetic and antigenic diversity of BVDVs circulating in China and therefore could benefit the development of suitable strategies for disease control.

Cattle connection: molecular epidemiology of BVDV outbreaks via rapid nanopore whole-genome sequencing of clinical samples

BackgroundAs a global ruminant pathogen, bovine viral diarrhea virus (BVDV) is responsible for the disease Bovine Viral Diarrhea with a variety of clinical presentations and severe economic losses worldwide. Classified within the Pestivirus genus, the species Pestivirus A and B (syn. BVDV-1, BVDV-2) are genetically differentiated into 21 BVDV-1 and four BVDV-2 subtypes. Commonly, the 5’ untranslated region and the Npro protein are utilized for subtyping. However, the genetic variability of BVDV leads to limitations in former studies analyzing genome fragments in comparison to a full-genome evaluation.ResultsTo enable rapid and accessible whole-genome sequencing of both BVDV-1 and BVDV-2 strains, nanopore sequencing of twelve representative BVDV samples was performed on amplicons derived through a tiling PCR procedure. Covering a multitude of subtypes (1b, 1d, 1f, 2a, 2c), sample matrices (plasma, EDTA blood and ear notch), viral loads (Cq-values 19–32) and species (cattle and sheep), ten of the twelve samples produced whole genomes, with two low titre samples presenting 96 % genome coverage.ConclusionsFurther phylogenetic analysis of the novel sequences emphasizes the necessity of whole-genome sequencing to identify novel strains and supplement lacking sequence information in public repositories. The proposed amplicon-based sequencing protocol allows rapid, inexpensive and accessible obtainment of complete BVDV genomes.

Recent Advances on the Bovine Viral Diarrhea Virus Molecular Pathogenesis, Immune Response, and Vaccines Development.

The bovine viral diarrhea virus (BVDV) consists of two species and various subspecies of closely related viruses of varying antigenicity, cytopathology, and virulence-induced pathogenesis. Despite the great ongoing efforts to control and prevent BVDV outbreaks and the emergence of new variants, outbreaks still reported throughout the world. In this review, we are focusing on the molecular biology of BVDV, its molecular pathogenesis, and the immune response of the host against the viral infection. Special attention was paid to discuss some immune evasion strategies adopted by the BVDV to hijack the host immune system to ensure the success of virus replication. Vaccination is one of the main strategies for prophylaxis and contributes to the control and eradication of many viral diseases including BVDV. We discussed the recent advances of various types of currently available classical and modern BVDV vaccines. However, with the emergence of new strains and variants of the virus, it is urgent to find some other novel targets for BVDV vaccines that may overcome the drawbacks of some of the currently used vaccines. Effective vaccination strategy mainly based on the preparation of vaccines from the homologous circulating strains. The BVDV-E2 protein plays important role in viral infection and pathogenesis. We mapped some important potential neutralizing epitopes among some BVDV genomes especially the E2 protein. These novel epitopes could be promising targets against the currently circulating strains of BVDV. More research is needed to further explore the actual roles of these epitopes as novel targets for the development of novel vaccines against BVDV. These potential vaccines may contribute to the global eradication campaign of the BVDV.

Detection of bovine viral diarrhoea virus nucleic acid, but not infectious virus, in bovine serum used for human vaccine manufacture

Bovine viral diarrhoea virus (BVDV) is a cattle pathogen that has previously been reported to be present in bovine raw materials used in the manufacture of biological products for human use. Seven lots of trivalent measles, mumps and rubella (MMR) vaccine and 1 lot of measles vaccine from the same manufacturer, together with 17 lots of foetal bovine serum (FBS) from different vendors, 4 lots of horse serum, 2 lots of bovine trypsin and 5 lots of porcine trypsin were analysed for BVDV using recently developed techniques, including PCR assays for BVDV detection, a qRT-PCR and immunofluorescence-based virus replication assays, and deep sequencing to identify and genotype BVDV genomes.All FBS lots and one lot of bovine-derived trypsin were PCR-positive for the presence of BVDV genome; in contrast all vaccine lots and the other samples were negative. qRT-PCR based virus replication assay and immunofluorescence-based infection assay detected no infectious BVDV in the PCR-positive samples. Complete BVDV genomes were generated from FBS samples by deep sequencing, and all were BVDV type 1. These data confirmed that BVDV nucleic acid may be present in bovine-derived raw materials, but no infectious virus or genomic RNA was detected in the final vaccine products.

Identification of Conserved Amino Acid Substitutions During Serial Infection of Pregnant Cattle and Sheep With Bovine Viral Diarrhea Virus.

Bovine viral diarrhea virus (BVDV) is an economically important pathogen of cattle that can also infect a wide range of domestic and wild species including sheep, goats, deer, camelids, and pigs. BVDV isolates are genetically highly diverse and previous work demonstrated that many substitutions were introduced in the viral genome during acute infections in cattle. In contrast, only limited information exists regarding changes occurring during BVDV infections in species other than cattle. The purpose of this study was to determine the changes introduced in the open reading frame (ORF) of the BVDV genome during serial infection of pregnant cattle and sheep with an isolate of bovine origin. Serial experimental inoculations were performed in six pregnant heifers and six pregnant ewes using BVDV-1b isolate AU526 in the first heifer and ewe, and serum from the preceding acutely infected dam thereafter. Complete ORF sequences were determined for 23 BVDV-1b isolates including AU526, one isolate from each pregnant dam, and one isolate from each BVDV-positive offspring born to these dams. Sequence comparison revealed that greater numbers of substitutions occurred during serial infection of pregnant sheep than of pregnant cattle. Furthermore, multiple host-specific amino acid changes were gradually introduced and conserved. These changes were more abundant in ovine isolates and occurred primarily in the E2 coding region. These results suggest that BVDV infections in heterologous species may serve as a significant source of viral genetic diversity and may be associated with adaptive changes.

A reverse transcriptase-loop mediated isothermal amplification assay (RT-LAMP) for rapid detection of bovine viral diarrhea virus 1 and 2

Bovine viral diarrhea virus (BVDV) is a pathogen that infects cattle, and is globally important. It causes substantial financial losses to the livestock industry. In the current study, a one-step reverse transcriptase-loop-mediated isothermal amplification (RT-LAMP) assay was set up for rapid and efficient detection of BVDV. For this purpose, four primers were designed to recognize six distinct regions on the target RNA based on a highly conserved sequence in the 5΄ UTR of the BVDV genome. Eighty blood specimens were collected from bovines suspected to suffer from BVDV infection, and were tested in parallel by RT-LAMP and RT-PCR. Twenty four of these samples were positive by RT-LAMP, while twenty were positive by RT-PCR. The RT-LAMP detection limit was estimated to be approximately 70PFU /mL of virus. Comparison of RT-PCR with RT-LAMP in this study revealed the recent developed RT-LAMP a highly sensitive and specific for BVD virus detection in the clinical samples.

First Results in the Use of Bovine Ear Notch Tag for Bovine Viral Diarrhoea Virus Detection and Genetic Analysis.

BackgroundInfection due to bovine viral diarrhoea virus (BVDV) is endemic in most cattle-producing countries throughout the world. The key elements of a BVDV control programme are biosecurity, elimination of persistently infected animals and surveillance. Bovine viral diarrhoea (BVD) is a notifiable disease in Belgium and an official eradication programme started from January 2015, based on testing ear notches sampled during the official identification and registration of calves at birth. An antigen-capture ELISA test based on the detection of BVDV Erns protein is used. Ear notch sample may also be used to characterize the genotype of the calf when appropriate elution/dilution buffer is added. Both BVDV antigen-ELISA analysis and animal traceability could be performed.MethodologyWith regards to the reference protocol used in the preparation of ear notch samples, alternative procedures were tested in terms of BVDV analytic sensitivity, diagnostic sensitivity and specificity, as well as quality and purity of animal DNA.Principal Findings/SignificanceThe Allflex DNA Buffer D showed promising results in BVDV diagnosis and genome analyses, opening new perspectives for the livestock industry by the exploitation of the animal genome. Due to the high number of cattle involved in the Belgian official BVDV eradication programme based on ear notch tags sample, a large database on both BVDV status of newborn calves and cattle genome could be created for subsequent different uses (e.g. traceability, determination of parentage, genetic signatures throughout the genome associated with particular traits) evolving through a more integrated animal health.

Pelacakan Virus Bovine Viral Diarrhea pada Darah yang Dikoleksi dengan Kertas Saring Flinders Technology AssociatesTM (DETECTION OF BOVINE VIRAL DIARRHEA VIRUS FROM BLOOD SAMPLES COLLECTED USING FLINDERS TECHNOLOGY ASSOCIATESTM CARDS)

Bovine viral diarrhea virus (BVDV) is a common pathogen that causes major economic losses to dairyindustry worldwide. Amplification of 5’-UTR regions of BVDV genome from blood samples usingreversetranscriptase polymerase chain reaction (RT-PCR) is one of sensitive and accurate techniques forBVDV detection. A common limitation for molecular detection of virus is the ability to obtain high qualityof nucleic acid in the samples. Inappropriate storage conditions will result in a false negative test. FiveEDTA-blood samples collected from BVDV persistently infected cattle were used in the present study.Ten repetitions were done for each sample. The blood were then dripped in the middle part of the FlindersTechnology AssociatesTM (FTA) cards and let it dried for approximately 2 hours. The effectiveness of theFTATMcards for storage and retrieval of BVDV RNA from blood samples molecularly using RT-PCRtechnique were tested and evaluated. The results proved that FTATM cards are effective and safe for thestorage of BVDV genome for a long period at room temperature. Based on the result of the present study,it can be concluded that in veterinary medicine field, the application of FTATM technology needs to bedeveloped to various type of samples and should be used routinely for collecting samples containinginfectious agents.

Molecular detection and characterization of bovine viral diarrhea virus in Mongolian cattle and yaks.

Bovine viral diarrhea virus (BVDV) is classified into two species, namely, Bovine viral diarrhea virus 1 and Bovine viral diarrhea virus 2, and affects cattle worldwide, resulting in significant economic loss. The prevalence of BVDV-1 and BVDV-2 infections and its genotypes in Mongolian animals has not been studied. In this study, we surveyed BVDV infection in dairy cattle and yaks from Bornuur and Bulgan counties by RT-PCR, and the average infection rate in the sampling sites was 15.8% and 20.0%, respectively. In addition, molecular features of the 5'-UTR region of the BVDV genome in Mongolian cattle and yaks were identified as belonging to the subtypes BVDV-1a and BVDV-2a, respectively. Determining the prevalence, geographical distribution, and molecular diversity of BVDV-1 and BVDV-2 in various host species in Mongolia is important for further studies and process control programs.

Simultaneous detection of five notifiable viral diseases of cattle by single-tube multiplex real-time RT-PCR

Multiplexed real-time PCR (qPCR) assays enable the detection of several target genes in a single reaction, which is applicable for simultaneous testing for the most important viral diseases in samples obtained from ruminants with unspecific clinical symptoms. Here, reverse transcription qPCR (RT-qPCR) systems for the detection of bovine viral diarrhoea virus (BVDV) and bluetongue virus (BTV) were combined with an internal control system based on the beta-actin gene. Additionally, a background screening for three further major pathogens of cloven-hoofed animals reportable to the World Organisation for Animal Health, namely foot-and-mouth disease virus, epizootic haemorrhagic disease virus, and Rift Valley fever virus, was integrated using the identical fluorophore for the respective RT-qPCR assays.Every pathogen-specific assay had an analytical sensitivity of at least 100genome copies per reaction within the multiplex approach, and a series of reference samples and clinical specimens obtained from cattle, but also from small ruminants, were detected reliably. The qPCR systems integrated in the background screening were even not influenced by the simultaneous amplification of very high BVDV and BTV genome copy numbers.The newly developed multiplex qPCR allows the specific and sensitive detection of five of the most important diseases of ruminants and could be used in the context of monitoring programs or for differential diagnostics.

First report of Bovine Viral Diarrhea Virus antigen from pneumonic cattle in Sudan

To explore the expected role of Bovine Viral Diarrhea Virus (BVDV) in pneumonia in cattle, cattle lungs (n=242) showing signs of pneumonia were collected from slaughter houses of three different localities located at Northern, Central and Western Sudan during 2010–2013. The collected samples were tested for the presence of BVDV antigen using Enzyme-Linked Immunosorbent Assay (ELISA), and Fluorescent Antibody Test (FAT). Twenty six (10.7%) out of 242 samples were found to be positive for BVDV. Positive results were seen in all the three studied areas, with the highest prevalence (16.7%; n=4/24) at Gezira State in Central Sudan. BVDV genome could be detected in all ELISA positive samples. The results indicated the existence of BVDV infection in cattle in different areas in Sudan, and its possible association with respiratory infections in cattle. Analysis using BLAST indicated that the sequence was identical to the previously reported BVDV-1 (GenBank accession AF220247.1.); nucleotide A was found in our study at position 9 of our sequence, whereas T was present instead in the reference virus. This is the first report of detecting BVDV antigen, genome, and its sequence analysis collected from cattle lungs in Sudan.

The first study of bovine immunodeficiency virus (BIV) and bovine viral diarrhea virus (BVDV) co- infection in industrial herds of cattle in two provinces of Iran

BACKGROUND: BIV is a well-known bovine immunosup- pressive cause, but its pathogenesis has not been well characterized. It seems that it is possible that cofactors such as co- infection with other bovine viral pathogens may play a role in enhancing the pathogenesis of BIV infection; BVDV also has immunosuppressive effects. OBJECTIVE: The aim of this study was determination of possible correlation between BIV and BVDV infections. METHODS: Blood samples were randomly collected from a total of 1800 cattle in dairy industrial farms in Isfahan and Chaharmahal va Bakhtiari provinces of Iran. First BIV or BVDV positive sera were screened by ELISA, and then samples were analyzed to detect BIV proviral DNA or BVDV RNA, using PCR. RESULTS: Out of 1800 blood samples, 19 (1.06%) samples were BVDV positive, while BIV positive samples were 10 (0.55%). Nine (0.5%) samples contained both BIV and BVDV genomes and were positive in ELISA, while one of the samples (0.05%) was only BIV positive. CONCLUSIONS: In this study, there was a statistically significant relationship between BIV status and BVDV infection using Chi square and Pearson's correlation coefficient test (p=0, r=0.65).

Construction of chimeric bovine viral diarrhea viruses containing glycoprotein Erns of heterologous pestiviruses and evaluation of the chimeras as potential marker vaccines against BVDV

Bovine viral diarrhea virus (BVDV) infections are enzootic in the cattle population and continue to cause significant economic losses to the beef and dairy industries worldwide. Extent of the damages has stimulated increasing interest in control programs directed at eradicating BVDV infections. Use of a BVDV marker vaccine would facilitate eradication efforts as a negatively marked vaccine would enable differentiation of infected from vaccinated animals (DIVA). We describe here the construction of three chimeric BVDVs containing glycoprotein Erns of heterologous pestiviruses and the evaluation of the chimera viruses as potential marker vaccines against BVDV infections. Chimeric NADL/G-Erns, NADL/R-Erns, and NADL/P-Erns were constructed by replacing the Erns gene of the full-length BVDV (NADL strain) genome with the Erns genes of giraffe (G-Erns), reindeer (R-Erns), or pronghorn antelope (P-Erns) pestiviruses, respectively. Each chimeric NADL virus was viable and infectious in RD 420 (bovine testicular) and BK-6 (bovine kidney) cells. By immunohistochemistry assays, NADL/G-Erns and NADL/R-Erns chimeric viruses reacted to BVDV Erns specific monoclonal antibody (mAb) 15C5, whereas the NADL/P-Erns chimeric virus did not. In an animal vaccination study, inactivated vaccines made from two chimeric viruses and the wild type NADL BVDV induced similar neutralizing antibody responses. NADL/P-Erns-vaccinated animals were distinguished from animals vaccinated with the wild type virus by means of a companion serological DIVA assay. These results show that chimeric NADL/P-Erns virus containing the Erns gene of pronghorn antelope pestivirus could be a potential marker vaccine candidate for use in a BVDV control and eradication program.

Analysis of codon usage in bovine viral diarrhea virus

Bovine viral diarrhea virus (BVDV) is a widespread virus in beef and dairy herds. BVDV has been grouped into two genotypes, genotype 1 and genotype 2. In this study, the relative synonymous codon usage (RSCU) values, effective number of codon (ENC) values and nucleotide content were investigated, and a comparative analysis of codon usage patterns for open reading frames (ORFs) of 22 BVDV genomes, including 14 of genotype 1 and 8 of genotype 2, was carried out. A high A+U content and low codon bias were found in BVDV genomes. Depending on the RSCU data, it was found that there was a significant variation in bias of codon usage between the two genotypes, and a geographic factor exists only in genotype-1 of BVDV. The RSCU data have a negative correlation with general average hydrophobicity (GRAVY), aromaticity and nucleotide content. Furthermore, the overall abundance of C and U has no effect on the synonymous codon usage patterns. In contrast, the A and G content showed a significant correlation with the nucleotide content at the third position. In addition, the codon usage patterns of BVDV are similar to those of 22 conserved genes of Bos taurus. Taken together, the genetic characteristics of BVDV possibly result from interactions between natural seclection and mutation pressure.Electronic supplementary materialThe online version of this article (doi:10.1007/s00705-010-0848-0) contains supplementary material, which is available to authorized users.

A replicon trans-packaging system reveals the requirement of nonstructural proteins for the assembly of bovine viral diarrhea virus (BVDV) virion

A selective trans-packaging system was developed to produce and isolate bovine viral diarrhea virus (BVDV) pseudo-particles with complementing reporter replicons and their packaging proteins expressed in trans with recombinant vaccinia virus. The encapsidation of replicon rNS3-5B was dependent not only on the in trans expression of structural proteins C, E rns, E1 and E2, but also the nonstructural proteins, p7 and contiguous precursor NS2-3-4A. Nonstructural p7, NS4B, NS5A or NS5B could be expressed in cis and in trans with precursor NS2-3-4A without significantly affecting virion assembly efficiency. NS2-3-4A was identified as an in trans functional precursor in virion assembly. BVDV genomes with mutant NS5B, which did not undergo active replication, were packaged 5-fold less efficiently than the intact genomes demonstrating the importance of replication in virion packaging. These results suggest that genome replication and assembly are closely associated, consistent with a model in which these two steps are coupled for maximum efficiency.

Prevalence of bovine viral diarrhea virus infections in alpacas in the United States

To determine the prevalence of bovine viral diarrhea virus (BVDV)-infected alpaca herds in the United States and investigate factors associated with seropositive herd status and, subsequently, determine the proportion of animals within seropositive alpaca herds that are persistently infected (PI) carriers for BVDV, obtain information regarding previous herd exposure to BVDV, determine titers of anti-BVDV antibodies of dams, and ascertain whether individual seropositive crias had received supplemental colostrum at birth. Prevalence study. 63 alpaca herds with >or= 12 registered female alpacas. 250 alpaca breeders were randomly selected from 562 eligible herds listed in the Alpaca Owner and Breeders Association membership directory and mailed a voluntary participation request. Sixty-three alpaca breeders participated in the study. From each herd, blood samples from >or= 4 crias were tested for BVDV, BVDV RNA, and serum neutralizing antibodies against BVDV. A region of the genome of BVDV recovered from PI crias was sequenced to determine genetic homology. Among the 63 herds, 16 (25.4%) had seropositive crias and 4 (6.3%) had PI crias. Infections in 3 of the 4 herds with PI crias were linked as evidence by the genetic homologies of viruses. In addition to PI crias, feeding supplemental colostrum was associated with herd seropositivity. Results confirmed the importance of BVDV infections in alpacas in the United States and highlighted the importance of determining the BVDV infection status of animals before they are commingled to limit exposure of herds to BVDV infection.

Bovine viral diarrhea virus is a suitable viral vector for stable expression of heterologous gene when inserted in between N pro and C genes

Bovine viral diarrhea virus (BVDV) is a group of small enveloped viruses with a single-stranded, positive-oriented RNA genome of ∼12.3 kb. BVDV genome directs the production of a viral polyprotein that is subsequently cleaved to release the mature viral proteins. To explore the potential of using BVDV as viral vector for stable expression of heterologous genes, eGFP2A was inserted in between N pro and C genes of a noncytopathic type-I BVDV strain SD1. eGFP2A was designed with eGFP protein in frame fused to the N terminus of the foot-and-mouth disease virus 2A protease. This strategy promised not only the correct processing of both viral N pro and C protein but also releasing of the chimeric protein from the nascent viral polyprotein. The recombinant reporter virus was successfully rescued in MDBK cells. In vitro study showed that eGFP2A protein, as expected, was expressed and processed properly from the nascent viral polyprotein. The reporter virus was similar to wt SD1 in viral RNA replication and protein expression and comparable to wt SD1 in growth kinetics except that this virus had a peak virus titer approximately 0.5 log 10 lower and a maximum yield about 4 h later than wt SD1. In summary, these results indicated that BVDV is a suitable viral vector for stable expression of heterologous genes when inserted in between N pro and C genes.