Evolution and virulence of porcine epidemic diarrhea virus following in vitro and in vivo propagation.

Practice of inoculating porcine epidemic diarrhea virus (PEDV) in piglets generating feedback material might influence the genetic evolution and attenuation of PEDV. The study was conducted to evaluate evolutionary rate and attenuation following serial in vitro and in vivo propagation. In the study, PED-JPFP0-PJ, Passage 0 (P0), was isolated from infected pigs and serially passaged in Vero cells for 5 consecutive times, P1-P5. P0, P2 and P5 were then subjected to orally inoculate 3-day-old piglets. At 24 h post inoculation, intestines of each passage (F1), were collected, and subsequently sub-passaged in piglets for 2 additional passages (F2-F3). Virus titration, PEDV genomic copies number, VH:CD ratios, and immunohistochemistry were evaluated. S and ORF3 genes were characterized. The results of the study demonstrated that virus titer and virulence were negatively correlated with increased passages, both in vitro and in vivo. Increased substitution rate was observed in higher passages. The evolutionary rate of S gene was higher than that of ORF3. Seven aa changes at positions 223, 291, 317, 607, 694, 1114 and 1199, with reduced N-linked glycan were observed in P5F3. In conclusion, serial passage of PEDV, both in vitro and in vivo, influence the genetic development and the attenuation of PEDV.

Intradermal needle-free injection prevents African Swine Fever transmission, while intramuscular needle injection does not.

Shared needles are a possible iatrogenic and hematogenous inanimate vector of African Swine Fever virus (ASFV) in farm conditions. To evaluate that possible transmission, sixty, 4-week-old pigs were procured from an ASF free herd free. Upon arrival, pigs were randomly divided into two sets. Set 1 served as seeder pigs, and were randomly allocated to 4 groups. The other pigs were divided into 8 groups, and served as sentinels. Seeder pigs were oronasally challenged with ASFV at high (108 copy numbers/mL), moderate (106 copy numbers/mL) or low (101 copy numbers/mL) challenge titer, except a subgroup that remained unchallenged (negative control). At 7 days post challenge (peak viremia), all four seeder groups were intradermally and intramuscularly (IM) injected with a vaccine adjuvant (Diluvac Forte, MSD Animal Health, The Netherlands) using a needle-free device (IDAL 3G, MSD Animal Health, The Netherlands) and conventional needles, respectively. The same needle or needle-free device was then used to inject the same volume of adjuvant into set 2 (n = 48) pigs. All pigs were observed for clinical disease daily and assayed for the presence of ASFV DNA by quantitative PCR. All seeder groups developed viremia (except the control pigs). ASFV viremia was detected in all sentinel groups injected via the intramuscular route. Transmission rate from the IM route via conventional needles was positively correlated with virus titer in blood circulation of seeders. Sentinels intramuscularly exposed to needles from high titer challenged seeders displayed more severe and acute clinical disease compared to that of exposed to low titer challenged seeders. No viremia nor clinical signs were observed in the sentinel groups injected via the intradermal route. This study confirmed the hematogenous transmission of ASFV between pigs through needle-sharing.

Retrospective study, full-length genome characterization and evaluation of viral infectivity and pathogenicity of chimeric porcine deltacoronavirus detected in Vietnam.

Increased evidence of porcine deltacoronavirus (PDCoV) causing diarrhoea in pigs has been reported in several countries worldwide. The virus has currently evolved into three separated groups including US, China and Southeast Asia (SEA) groups. In Vietnam, PDCoV was first reported in 2015. Based on phylogenetic analyses of spike, membrane and nucleocapsid genes, it is suggested that Vietnam PDCoV is chimeric virus. In the present study, we retrospectively investigated the presence of PDCoV in Vietnam and the full-length genomes of six PDCoV isolates identified in 2014-2016 were further characterized. The results demonstrated that Vietnam PDCoV was first detected as early as 2014. All six Vietnam PDCoV are in the SEA group and further divided into two separated subgroups including SEA-1 and SEA-2. Vietnam PDCoV in SEA-2 was closely related to Thai and Lao PDCoV. Recombination analysis demonstrated that three isolates in SEA-1 were a chimeric virus of which P12_14_VN_0814, the first Vietnam isolate, and US PDCoV isolates were major and minor parents, respectively. The recombination was further evaluated by phylogenetic construction based on 3 recombinant fragments. The first and third fragments, closely related to P12_14_VN_0814, were associated with ORF1a/1b and N genes, respectively. The second fragment, associated with S, E, and M genes, was closely related to US PDCoV isolates. High antigenic and hydrophobic variations were detected in S1 protein. Three-day-old pigs challenged with the chimeric virus displayed clinical diseases and villus atrophy. In conclusion, Vietnam PDCoV is genetically diverse influenced by an external introduction from neighbouring countries. The chimeric Vietnam PDCoV can induce a disease similar to Thai PDCoV.