Advancements in the Growth and Construction of Recombinant Lumpy Skin Disease Virus (LSDV) for Use as a Vaccine Vector

Attenuated vaccine strains of lumpy skin disease virus (LSDV) have become increasingly popular as recombinant vaccine vectors, to target both LSDV, as well as other pathogens, including human infectious agents. Historically, these vaccine strains and recombinants were generated in primary (lamb) testis (LT) cells, Madin–Darby bovine kidney (MDBK) cells or in eggs. Growth in eggs is a laborious process, the use of primary cells has the potential to introduce pathogens and MDBK cells are known to harbor bovine viral diarrhea virus (BVDV). In this study, data is presented to show the growth of an attenuated LSDV strain in baby hamster kidney (BHK-21) cells. Subsequently, a recombinant LSDV vaccine was generated in BHK-21 cells. Partial growth was also observed in rabbit kidney cells (RK13), but only when the vaccinia virus host range gene K1L was expressed. Despite the limited growth, the expression of K1L was enough to serve as a positive selection marker for the generation of recombinant LSDV vaccines in RK13 cells. The simplification of generating (recombinant) LSDV vaccines shown here should increase the interest for this platform for future livestock vaccine development and, with BHK-21 cells approved for current good manufacturing practice, this can be expanded to human vaccines as well.

Successful Male Rabbit Whole Kidney Decellularization For Tissue Engineering

Background Renal transplantation represents the only cure for end stage renal disease. Shortage of donated organs, immune compatibility and post-transplantation complications urges the search for another radical and revolutionary treatment . Whole kidney decellularization is a promising technique, hopefully offering a threedimensional natural kidney scaffold for engineering a patient’s compatible kidney after recellularization with patient’s own cells. The current study aimed at successfully decellularizing the male rabbit kidney while preserving the histological profile of the kidney decellularized extracellular matrix. Materials and Methods Kidneys of ten male New Zealand White Rabbits weighing (1000- 1500 gm) were harvested and sorted into two groups. Control Group I included the ten right kidneys. Decellularization Group II included the ten left kidneys; harvested carefully and kept frozen until decellularization. They were decellularized using 0.5% sodium dodecyl sulfate for 5-6 hours. Kidneys from both groups were processed similarly for histological examination. Results Decellularized kidney Scaffolds showed complete removal of cellular materials. Meanwhile, the structural profile of the decellularized kidney scaffolds was apparently well preserved. Conclusion The current study yielded an efficient decellularization of male rabbit kidney paving the way for future kidney recellularization.

Role of Vesicle-Associated Membrane Protein-Associated Proteins (VAP) A and VAPB in Nuclear Egress of the Alphaherpesvirus Pseudorabies Virus

The molecular mechanism affecting translocation of newly synthesized herpesvirus nucleocapsids from the nucleus into the cytoplasm is still not fully understood. The viral nuclear egress complex (NEC) mediates budding at and scission from the inner nuclear membrane, but the NEC is not sufficient for efficient fusion of the primary virion envelope with the outer nuclear membrane. Since no other viral protein was found to be essential for this process, it was suggested that a cellular machinery is recruited by viral proteins. However, knowledge on fusion mechanisms involving the nuclear membranes is rare. Recently, vesicle-associated membrane protein-associated protein B (VAPB) was shown to play a role in nuclear egress of herpes simplex virus 1 (HSV-1). To test this for the related alphaherpesvirus pseudorabies virus (PrV), we mutated genes encoding VAPB and VAPA by CRISPR/Cas9-based genome editing in our standard rabbit kidney cells (RK13), either individually or in combination. Single as well as double knockout cells were tested for virus propagation and for defects in nuclear egress. However, no deficiency in virus replication nor any effect on nuclear egress was obvious suggesting that VAPB and VAPA do not play a significant role in this process during PrV infection in RK13 cells.