7-day post-vaccination efficacy of the CSF CL strain produced on ovine cell line against a virulent classical swine fever (Hog cholera) challenge

Classical swine fever (CSF), also known as hog cholera, is a highly contagious viral disease classified as a notifiable (previously List A) pig disease by OIE. In an infected environment, vaccines are the basic tools for control and eradication of CSFV. This study aimed at assessing the efficacy of an attenuated CSF CL strain produced on ovine cells against a virulent CSF challenge performed 7 days post-vaccination. Material and methodsTwo groups of 8 CSF-negative crossbred pigs weighing 18 kg were either vaccinated with a live one dose of CL strain (>100 PD50/dose) or left unvaccinated. Seven days post-vaccination, they were challenged with 5.5log10TCID50 CSFV of Haiti-96 strain both intramuscularly and intranasally with separated aliquots. Clinical signs, rectal temperature were monitored for 28 days post-challenge (DPC) and necropsied. Blood samples, nasal swabs and tonsil scrapings were regularly collected and assayed for blood formulation, sera antibody titres (E2-Erms ELISAs, SN titrations), and viral loads in total blood, nasal and tonsil mucus. ResultsAll controls showed typical acute CSF justifying euthanasia on ethical ground 22 DPCH at the latest. They also developed severe leukopenia and lymphopenia. Necropsic lesions were evocative of chronic form of CSF. None of the vaccinates developed any sign of CSF.CSFV was detected in controls from 4DPC in blood plateauing close to 6log10TCID50/mL. CSFV was detected in nasal and tonsillar mucus from 8DPC reaching up to 4.8log10TCID50/mL. Vaccinates showed no detectable CSFV in any sample post-challenge.All pigs were antibody negative before challenge. A seroneutralizing anamestic reaction was evidenced as early as 7DPC in all vaccinates whereas ELISA antibody titres turned positive slower. Serological response to challenge was almost absent in controls. Discussion and conclusionViral circulation in herds relies mainly on direct nose-to-nose contacts and in-utero transmission. The ability of vaccines to limit CSFV transmission is vital for control and eradication strategies. Under the conditions of the study, the CL strain was able to totally prevent CSF and totally abolished both CSFV viremia and mucosal shedding as early as 7 days post-vaccination, thus showing its relevance for whole herd strategies. Additionally, it was noticed that ELISA correlated poorly with protection.Fil: Risatti, Guillermo R.. University of Connecticut; Estados UnidosFil: Perez, Sandra. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tandil. Centro de Investigación Veterinaria de Tandil. Universidad Nacional del Centro de la Provincia de Buenos Aires. Centro de Investigación Veterinaria de Tandil. Provincia de Buenos Aires. Gobernación. Comision de Investigaciones Científicas. Centro de Investigación Veterinaria de Tandil; Argentina25th International Pig Veterinary Society CongressChongqingChinaInternational Pig Veterinary Societ

Three-week post-vaccination efficacy of the CSF CL strain produced on ovine cell line against a virulent classical swine fever (Hog cholera)

Classical swine fever (CSF) is a highly contagious viral disease classified as a notifiable pig disease by OIE. In an infected environment, vaccines are the basic tools for control and eradication of CSFV. This study aimed at assessing the efficacy of an attenuated CSF CL strain produced on ovine cells against a virulent CSF challenge performed three weeks post-vaccination. Material and methodsTwo groups of 8 CSF-negative crossbred pigs weighing 18 kg were either vaccinated with a live one dose of CL strain (>100 PD50/dose) or left unvaccinated. Three weeks post-vaccination, they were challenged with 5.5log10TCID50 CSFV of Haiti-96 strain both intramuscularly and intranasally with separated aliquots. Clinical signs, rectal temperature were monitored for 28 days post-challenge (DPC) and necropsied. Blood samples, nasal swabs and tonsil scrapings were regularly collected and assayed for blood formulation, sera antibody titres (E2-Erms ELISAs, SN titrations), and viral loads in total blood, nasal and tonsil mucus. ResultsFollowing 5-6 incubation days, all controls showed typical acute CSF. Two of them were euthanized on ethical ground. They also developed severe leukopenia and lymphopenia. Necropsic lesions were evocative of chronic form of CSF. None of the vaccinates developed any sign of CSF.In controls, CSFV viremia was detected from 5DPC reaching levels above 6log10TCID50/mL from 6DPCH to 11DPCH. Tonsils were positive for CSF as well from 5DPCH and viral sheddind in nasal mucus could reach more than 4log10TCID50/mL. Vaccinates showed drastically (p<0.001) reduced viremia with overall mean titer of 1.7log10TCID50/mL and no detectable CSFV in tonsils or nasal mucus. Vaccinates showed low levels of seroneutralizing antibodies and neither E2 nor Erms antibiodies before challenge. Following challenge seroconversion was significantly (p<0.05) faster in vaccinates. Discussion and conclusionViral circulation in herds relies mainly on direct nose-to-nose contacts and in-utero transmission. The ability of vaccines to limit CSFV transmission is vital for control and eradication strategies. Under the conditions of the study, the CL strain was able to totally prevent CSF, drastically limit viremia and abolished CSFV shedding through oro-nasal route.Total protection despite absent to low antibody levels suggested a major contribution of cell-mediated immunity to protection.Fil: Risatti, Guillermo R.. University of Connecticut; Estados UnidosFil: Perez, Sandra. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tandil. Centro de Investigación Veterinaria de Tandil. Universidad Nacional del Centro de la Provincia de Buenos Aires. Centro de Investigación Veterinaria de Tandil. Provincia de Buenos Aires. Gobernación. Comision de Investigaciones Científicas. Centro de Investigación Veterinaria de Tandil; Argentina25th International Pig Veterinary Society CongressChongqingChinaInternational Pig Veterinary Society Congres

Identification of a novel virulence determinant within the E2 structural glycoprotein of classical swine fever virus

AbstractClassical swine fever virus (CSFV) E2 glycoprotein contains a discrete epitope (TAVSPTTLR, residues 829–837 of CSFV polyprotein) recognized by monoclonal antibody (mAb) WH303, used to differentiate CSFV from related ruminant pestiviruses, Bovine Viral Diarrhea Virus (BVDV) and Border Disease Virus (BDV), that infect swine without causing disease. Progressive mutations were introduced into mAb WH303 epitope in CSFV virulent strain Brescia (BICv) to obtain the homologous amino acid sequence of BVDV strain NADL E2 (TSFNMDTLA). In vitro growth of mutants T1v (TSFSPTTLR), T2v (TSFNPTTLR), T3v (TSFNMTTLR) was similar to parental BICv, while mutants T4v (TSFNMDTLR) and T5v (TSFNMDTLA) exhibited a 10-fold decrease in virus yield and reduced plaque size. In vivo, T1v, T2v or T3v induced lethal disease, T4v induced mild and transient disease and T5v induced mild clinical signs. Protection against BICv challenge was observed at 3 and 21 days post-T5v infection. These results indicate that E2 residues TAVSPTTLR play a significant role in CSFV virulence

Perceptions of pastoralist problems: A participatory study on animal management, disease spectrum and animal health priorities of small ruminant pastoralists in Georgia

Small ruminants support the livelihoods of millions of poor pastoralist and sedentary households around the world. While pastoralists are generally not amongst the poorest in terms of assets, they are frequently marginalised in terms of their access to political power, health and education. This study was undertaken among pastoralist households keeping small ruminants in four regions of the country of Georgia. Small ruminants are an important cultural, social and economic asset in Georgia and are mainly managed in a transhumant pastoralist system. Georgia suffered its first, and so far only outbreak of peste des petits ruminants (PPR) in 2016. This qualitative interview study was designed to acquire contextual understanding of local small ruminant husbandry and the livelihood situations of the participating pastoralists, and to detect historical, unreported PPR outbreaks. Focus group discussions comprising participatory epidemiology tools and other forms of interviews were used to explore small ruminant management, disease spectrum and management, and animal health priorities.The participants had experienced a wide variety of animal health constraints, with intestinal worms, braxy, piroplasmosis, pasture-related problems, predators and lameness emerging as priorities. No historic, unreported PPR outbreak was detected in this study, and PPR was not a priority for participants. Instead, the day-to-day reality of animal health for the pastoralists was characterised by co-infections of mainly endemic pathogens, and problems related to other challenges such as access to land, feed and genetic resources. The rationale behind the participants' prioritisation of animal health problems was supported by the need to pay extra attention to animals in order to avoid risk factors, keep animals healthy and minimise the negative impact of diseases or management problems; the various epidemiological and clinical parameters of the prioritised diseases; the economic impact of the specific problems and the zoonotic potential of diseases and predation. Even within regions, and within seemingly socially and culturally homogenous groups, there were important local differences in the problems faced by pastoralists that affect their livestock management. This study underlines the importance of a contextualised understanding of the local disease panorama and complexities in the livelihood situations of rural people when designing actions to improve animal health in general or, more specifically, passive surveillance as well as prevention or control measures. Finally, it is concluded that to achieve such an understanding, there is a need for participatory, scoping-style studies that specifically acknowledge diversity and power relations

Clustering of classical swine fever virus isolates by codon pair bias

<p>Abstract</p> <p>Background</p> <p>The genetic code consists of non-random usage of synonymous codons for the same amino acids, termed codon bias or codon usage. Codon juxtaposition is also non-random, referred to as codon context bias or codon pair bias. The codon and codon pair bias vary among different organisms, as well as with viruses. Reasons for these differences are not completely understood. For classical swine fever virus (CSFV), it was suggested that the synonymous codon usage does not significantly influence virulence, but the relationship between variations in codon pair usage and CSFV virulence is unknown. Virulence can be related to the fitness of a virus: Differences in codon pair usage influence genome translation efficiency, which may in turn relate to the fitness of a virus. Accordingly, the potential of the codon pair bias for clustering CSFV isolates into classes of different virulence was investigated.</p> <p>Results</p> <p>The complete genomic sequences encoding the viral polyprotein of 52 different CSFV isolates were analyzed. This included 49 sequences from the GenBank database (NCBI) and three newly sequenced genomes. The codon usage did not differ among isolates of different virulence or genotype. In contrast, a clustering of isolates based on their codon pair bias was observed, clearly discriminating highly virulent isolates and vaccine strains on one side from moderately virulent strains on the other side. However, phylogenetic trees based on the codon pair bias and on the primary nucleotide sequence resulted in a very similar genotype distribution.</p> <p>Conclusion</p> <p>Clustering of CSFV genomes based on their codon pair bias correlate with the genotype rather than with the virulence of the isolates.</p

Classical Swine Fever Virus p7 Protein Interacts with Host Protein CAMLG and Regulates Calcium Permeability at the Endoplasmic Reticulum

We have previously shown that Classical Swine Fever Virus (CSFV) p7 is an essential nonstructural protein with a viroporin activity, a critical function in the progression of virus infection. We also identified p7 domains and amino acid residues critical for pore formation. Here, we describe how p7 specifically interacts with host protein CAMLG, an integral ER transmembrane protein involved in intracellular calcium release regulation and signal response generation. Detection of interaction as well as the identification of p7 areas mediating interaction with CAMLG was performed by yeast two-hybrid. p7-CAMLG interaction was further confirmed by confocal microscopy in eukaryotic cells, co-expressing both proteins. Mutant forms of p7 having substituted native residues identified as mediating interaction with CAMLG showed a decreased co-localization compared with the native forms of p7. Furthermore, it is shown that native p7, but not the mutated forms of p7 that fail to interact with CAMLG, efficiently mediates calcium permeability in the ER. Interestingly, viruses harboring some of those mutated forms of p7 have been previously shown to have a significantly decreased virulence in swine.ARS/USDA-University of Connecticut SCA# 58-1940-1-190 and ARS/USDA-University of the Basque Country NACA#8064-32000-056-18S

Establishment and characterization of an infectious cDNA clone of a classical swine fever virus LOM strain

Classical swine fever virus (CSFV) causes a highly contagious disease among swine that has an important economic impact worldwide. CSFV strain LOM is an attenuated virus of low virulent strain of Miyagi isolated from Japan in 1956. Eight DNA fragments representing the genome of the CSFV strain LOM were obtained by RT-PCR. These were used to determine the complete nucleotide sequence and construct a full-length cDNA clone which was called Flc-LOM. Sequence analysis of the recombinant clone (Flc-LOM) revealed the presence of eight mutations, resulting in two amino acid substitutions, when compared to the parental sequence. RNA transcripts of both LOM and Flc-LOM were directly infectious in PK-15 cells. The rescued Flc-LOM virus grew more slowly than the parental virus, LOM, in the cells. Intramuscular immunization with Flc-LOM was safe and highly immunogenic in pigs; no clinical signs or virus transmission to sentinel animals were observed after 35 days. CSFV-specific neutralizing antibodies were detected 14 days post-infection. After challenge with the virulent CSFV strain SW03, pigs immunized with Flc-LOM were shown to be fully protected. Thus, our newly established infectious clone of CSFV, Flc-LOM, could serve as a vaccine candidate