Double Deletion of EP402R and EP153R in the Attenuated Lv17/WB/Rie1 African Swine Fever Virus (ASFV) Enhances Safety, Provides DIVA Compatibility, and Confers Complete Protection Against a Genotype II Virulent Strain.

African swine fever virus (ASFV) is a devastating disease affecting domestic and wild suids and causing significant economic losses in the global pig industry. Attenuated modified live virus (MLV) vaccines are the most promising approaches for vaccine development. This study aimed to evaluate the safety and efficacy of four recombinant ASFV genotype II strains, derived from the non-hemadsorbing (non-HAD) attenuated isolate Lv17/WB/Rie1, through the single or simultaneous deletion of virulence-associated genes.

A broadly reactive ultralong bovine antibody that can determine the integrity of foot-and-mouth disease virus capsids.

Foot-and-mouth disease vaccination using inactivated virus is suboptimal, as the icosahedral viral capsids often disassemble into antigenically distinct pentameric units during long-term storage, or exposure to elevated temperature or lowered pH, and thus raise a response that is no longer protective. Furthermore, as foot-and-mouth disease virus (FMDV)'s seven serotypes are antigenically diverse, cross-protection from a single serotype vaccine is limited, and most existing mouse and bovine antibodies and camelid single-domain heavy chain-only antibodies are serotype-specific. For quality control purposes, there is a real need for pan-serotype antibodies that clearly distinguish between pentamer (12S) and protective intact FMDV capsid.

Demonstrating the importance of porcine reproductive and respiratory syndrome virus papain-like protease 2 deubiquitinating activity in viral replication by structure-guided mutagenesis.

Deubiquitination of cellular substrates by viral proteases is a mechanism used to interfere with host cellular signaling processes, shared between members of the coronavirus- and arterivirus families. In the case of Arteriviruses, deubiquitinating and polyprotein processing activities are accomplished by the virus-encoded papain-like protease 2 (PLP2). Several studies have implicated the deubiquitinating activity of the porcine reproductive and respiratory syndrome virus (PRRSV) PLP2 in the downregulation of cellular interferon production, however to date, the only arterivirus PLP2 structure described is that of equine arteritis virus (EAV), a distantly related virus.

Immune efficacy of a candidate porcine reproductive and respiratory syndrome vaccine rHN-NP49 administered by a Needle-free intradermal delivery system in comparison with intramuscular injection.

Porcine reproductive and respiratory syndrome (PRRS) is one of the major drivers of economic loss in the swine industry worldwide. In commercial pig production, vaccination is the first option in an attempt to control infectious diseases. Pigs are therefore often immunized with different vaccines, and almost all of them are delivered via the intramuscular (IM) route.

Assessing the Protective Dose of a Candidate DIVA Vaccine against Classical Swine Fever.

Classical swine fever is a highly contagious and deadly disease in swine. The disease can be controlled effectively by vaccination with an attenuated virus known as the "Chinese" (C)-strain. A single vaccination with the C-strain provides complete protection against highly virulent isolates within days after vaccination, making it one of the most efficacious veterinary vaccines ever developed.

An octavalent vaccine provides pregnant gilts protection against a highly virulent porcine parvovirus strain.

Background: Porcilis® Ery+Parvo+Lepto is an octavalent inactivated ready-to-use vaccine that contains Erysipelothrix rhusiopathiae (Ery), porcine parvovirus (PPV), and six serogroups of Leptospira (Lepto). The efficacy of Porcilis® Ery + Parvo+Lepto against reproductive problems associated with porcine parvovirus (PPV) infection was evaluated in pregnant gilts. For this, a group of ninegilts was vaccinated twice (at 5 and 6 months old) with Porcilis® Ery + Parvo+Lepto (Group 1), while a group of eight gilts was included as unvaccinated controls (Group 2).

Cell Attachment Domains of the Porcine Epidemic Diarrhea Virus Spike Protein Are Key Targets of Neutralizing Antibodies.

Porcine epidemic diarrhea virus (PEDV) causes enteric disease in pigs, resulting in significant economic losses to the swine industry worldwide. Current vaccination approaches against this emerging coronavirus are only partially effective, though natural infection protects pigs against reinfection and provides lactogenic immunity to suckling piglets. The viral spike (S) glycoprotein, responsible for receptor binding and cell entry, is the major target for neutralizing antibodies.

The B Cell Response to Foot-and-Mouth Disease Virus in Cattle following Sequential Vaccination with Multiple Serotypes.

Foot-and-mouth disease virus (FMDV) is a highly contagious viral disease. Antibodies are pivotal in providing protection against FMDV infection. Serological protection against one FMDV serotype does not confer interserotype protection.

Vaccination of pigs reduces Torque teno sus virus viremia during natural infection.

Anelloviruses are a group of single-stranded circular DNA viruses infecting several vertebrate species. Four species have been found to infect swine, namely Torque teno sus virus (TTSuV) 1a and 1b (TTSuV1a, TTSuV1b; genus Iotatorquevirus), TTSuVk2a and TTSuVk2b (genus Kappatorquevirus). TTSuV infection in pigs is distributed worldwide, and is characterized by a persistent viremia.

In vivo assessment of equine arteritis virus vaccine improvement by disabling the deubiquitinase activity of papain-like protease 2.

Arteriviruses are a family of positive-stranded RNA viruses that includes the prototypic equine arteritis virus (EAV) and porcine reproductive and respiratory syndrome virus (PRRSV). Although several vaccines against these viruses are commercially available there is room for improvement, especially in the case of PRRSV. The ability of arteriviruses to counteract the immune response is thought to decrease the efficacy of the current modified live virus vaccines.

Differential repair of etheno-DNA adducts by bacterial and human AlkB proteins.

AlkB proteins are evolutionary conserved Fe(II)/2-oxoglutarate-dependent dioxygenases, which remove alkyl and highly promutagenic etheno(ɛ)-DNA adducts, but their substrate specificity has not been fully determined. We developed a novel assay for the repair of ɛ-adducts by AlkB enzymes using oligodeoxynucleotides with a single lesion and specific DNA glycosylases and AP-endonuclease for identification of the repair products. We compared the repair of three ɛ-adducts, 1,N(6)-ethenoadenine (ɛA), 3,N(4)-ethenocytosine (ɛC) and 1,N(2)-ethenoguanine (1,N(2)-ɛG) by nine bacterial and two human AlkBs, representing four different structural groups defined on the basis of conserved amino acids in the nucleotide recognition lid, engaged in the enzyme binding to the substrate.

Protozoan ALKBH8 oxygenases display both DNA repair and tRNA modification activities.

The ALKBH family of Fe(II) and 2-oxoglutarate dependent oxygenases comprises enzymes that display sequence homology to AlkB from E. coli, a DNA repair enzyme that uses an oxidative mechanism to dealkylate methyl and etheno adducts on the nucleobases. Humans have nine different ALKBH proteins, ALKBH1-8 and FTO.

Discovery of a novel Torque teno sus virus species: genetic characterization, epidemiological assessment and disease association.

The study describes a novel Torque teno sus virus (TTSuV) species, provisionally named Torque teno sus virus k2b (TTSuVk2b), originally found in commercial pig sera by applying the rolling-circle amplification technique. Full-length sequences of TTSuVk2b were obtained, annotated and used in the phylogenetic analyses, which revealed that TTSuVk2b is a novel Anellovirus species within the genus Kappatorquevirus of the family Anelloviridae. Quantitative PCR techniques were developed to determine total TTSuV DNA quantities as well as the prevalence and viral DNA quantities of TTSuV1, TTSuVk2a and TTSuVk2b.

Roles of Trm9- and ALKBH8-like proteins in the formation of modified wobble uridines in Arabidopsis tRNA.

Uridine at the wobble position of tRNA is usually modified, and modification is required for accurate and efficient protein translation. In eukaryotes, wobble uridines are modified into 5-methoxycarbonylmethyluridine (mcm(5)U), 5-carbamoylmethyluridine (ncm(5)U) or derivatives thereof. Here, we demonstrate, both by in vitro and in vivo studies, that the Arabidopsis thaliana methyltransferase AT1G31600, denoted by us AtTRM9, is responsible for the final step in mcm(5)U formation, thus representing a functional homologue of the Saccharomyces cerevisiae Trm9 protein.

ALKBH8-mediated formation of a novel diastereomeric pair of wobble nucleosides in mammalian tRNA.

Mammals have nine different homologues (ALKBH1-9) of the Escherichia coli DNA repair demethylase AlkB. ALKBH2 is a genuine DNA repair enzyme, but the in vivo function of the other ALKBH proteins has remained elusive. It was recently shown that ALKBH8 contains an additional transfer RNA (tRNA) methyltransferase domain, which generates the wobble nucleoside 5-methoxycarbonylmethyluridine (mcm(5)U) from its precursor 5-carboxymethyluridine (cm(5)U).

Mammalian ALKBH8 possesses tRNA methyltransferase activity required for the biogenesis of multiple wobble uridine modifications implicated in translational decoding.

Uridines in the wobble position of tRNA are almost invariably modified. Modifications can increase the efficiency of codon reading, but they also prevent mistranslation by limiting wobbling. In mammals, several tRNAs have 5-methoxycarbonylmethyluridine (mcm5U) or derivatives thereof in the wobble position.

Bioinformatics and functional analysis define four distinct groups of AlkB DNA-dioxygenases in bacteria.

The iron(II)- and 2-oxoglutarate (2OG)-dependent dioxygenase AlkB from Escherichia coli (EcAlkB) repairs alkylation damage in DNA by direct reversal. EcAlkB substrates include methylated bases, such as 1-methyladenine (m(1)A) and 3-methylcytosine (m(3)C), as well as certain bulkier lesions, for example the exocyclic adduct 1,N(6)-ethenoadenine (epsilonA). EcAlkB is the only bacterial AlkB protein characterized to date, and we here present an extensive bioinformatics and functional analysis of bacterial AlkB proteins.

Biochemical characterization of arterivirus nonstructural protein 11 reveals the nidovirus-wide conservation of a replicative endoribonuclease.

Nidoviruses (arteriviruses, coronaviruses, and roniviruses) are a phylogenetically compact but diverse group of positive-strand RNA viruses that includes important human and animal pathogens. Nidovirus RNA synthesis is mediated by a cytoplasmic membrane-associated replication/transcription complex that includes up to 16 viral nonstructural proteins (nsps), which carry common enzymatic activities, like the viral RNA polymerase, but also unusual and poorly understood RNA-processing functions. Of these, a conserved endoribonuclease (NendoU) is a major genetic marker that is unique to nidoviruses.

Viral AlkB proteins repair RNA damage by oxidative demethylation.

Bacterial and mammalian AlkB proteins are iron(II)- and 2-oxoglutarate-dependent dioxygenases that reverse methylation damage, such as 1-methyladenine and 3-methylcytosine, in RNA and DNA. An AlkB-domain is encoded by the genome of numerous single-stranded, plant-infecting RNA viruses, the majority of which belong to the Flexiviridae family. Our phylogenetic analysis of AlkB sequences suggests that a single plant virus might have acquired AlkB relatively recently, followed by horizontal dissemination among other viruses via recombination.

New structure model for the packaging signal in the genome of group IIa coronaviruses.

A 190-nucleotide (nt) packaging signal (PS) located in the 3' end of open reading frame 1b in the mouse hepatitis virus, a group IIa coronavirus, was previously postulated to direct genome RNA packaging. Based on phylogenetic data and structure probing, we have identified a 95-nt hairpin within the 190-nt PS domain which is conserved in all group IIa coronaviruses but not in the severe acute respiratory syndrome coronavirus (group IIb), group I coronaviruses, or group III coronaviruses. The hairpin is composed of six copies of a repeating structural subunit that consists of 2-nt bulges and 5-bp stems.

An infectious recombinant equine arteritis virus expressing green fluorescent protein from its replicase gene.

Thus far, systems developed for heterologous gene expression from the genomes of nidoviruses (arteriviruses and coronaviruses) have relied mainly on the translation of foreign genes from subgenomic mRNAs, whose synthesis is a key feature of the nidovirus life cycle. In general, such expression vectors often suffered from relatively low and unpredictable expression levels, as well as genome instability. In an attempt to circumvent these disadvantages, the possibility to express a foreign gene [encoding enhanced green fluorescent protein (eGFP)] from within the nidovirus replicase gene, which encodes two large polyproteins that are processed proteolytically into the non-structural proteins (nsps) required for viral RNA synthesis, has now been explored.

Antiviral activity of morpholino oligomers designed to block various aspects of Equine arteritis virus amplification in cell culture.

The antiviral efficacy of ten antisense phosphorodiamidate morpholino oligomers (PMOs) directed against Equine arteritis virus (EAV), a nidovirus belonging to the family Arteriviridae, was evaluated in mammalian (Vero-E6) cells. Peptide-conjugated PMOs (P-PMOs) supplied in cell culture medium at micromolar concentrations were efficiently taken up by Vero-E6 cells and were minimally cytotoxic. The P-PMOs were designed to base pair to RNA sequences involved in different aspects of EAV amplification: genome replication, subgenomic mRNA synthesis, and translation of genome and subgenomic mRNAs.

Discontinuous subgenomic RNA synthesis in arteriviruses is guided by an RNA hairpin structure located in the genomic leader region.

Nidoviruses produce an extensive 3'-coterminal nested set of subgenomic (sg) mRNAs, which are used to express structural proteins and sometimes accessory proteins. In arteriviruses and coronaviruses, these mRNAs contain a common 5' leader sequence, derived from the genomic 5' end. The joining of the leader sequence to different segments derived from the 3'-proximal part of the genome (mRNA bodies) presumably involves a unique mechanism of discontinuous minus-strand RNA synthesis in which base pairing between sense and antisense transcription-regulating sequences (TRSs) plays an essential role.