Development of a low-cost, insect larvae-derived recombinant subunit vaccine against RHDV

Abstract

Vaccine antigens against rabbit hemorrhagic disease virus (RHDV) are currently derived from inactivated RHDV obtained from livers of experimentally infected rabbits. Several RHDV-derived recombinant immunogens have been reported. However, their application in vaccines has been restricted due to their high production costs. In this paper, we describe the development of an inexpensive, safe, stable vaccine antigen for RHDV. A baculovirus expressing a recombinant RHDV capsid protein (VP60r) was used to infect Trichoplusia ni insect larvae. It reached an expression efficiency of 12.5% of total soluble protein, i.e. approximately 2 mg of VP60r per larva. Preservation of the antigenicity and immunogenicity of the VP60r was confirmed by immunological and immunization experiments. Lyophilized crude larvae extracts, containing VP60r, were stable, at room temperature, for at least 800 days. In all cases, rabbits immunized with a single dose of VP60r by the intramuscular route were protected against RHDV challenge. Doses used were as low as 2 microg of VP60r in the presence of adjuvant or 100 microg without one. Orally administered VP60r in the absence of an adjuvant gave no protection. The potential costs of an RHDV vaccine made using this technology would be reduced considerably compared with producing the same protein in insect cells maintained by fermentation. In conclusion, the larva expression system may provide a broad-based strategy for production of recombinant subunit antigens (insectigens) for human or animal medicines, especially when production costs restrain their use.

Potential Use of the Hemagglutinin-Neuraminidase Glycoprotein of Newcastle Disease Virus Expressed in Rachiplusia nu Larvae as an Immunogen for Chickens

“…This study shows the efficacy of infected larval extracts as an NDV recombinant immunogen and shows that the extracts constitute an easier and less expensive approach for the production of recombinant antigens, since production in R. nu larvae was nearly 400 times more economical than the expression of the same protein in Sf9 cells (22). In our hands, a single infected larva yields quantities of recombinant HN protein similar to those yielded by 1.5 ϫ 10 6 infected Sf9 cells (data not shown).…”

“…Insect larvae infected with recombinant baculoviruses can be an extremely useful alternative due to the high levels of protein expression achieved, the lower costs for large-scale production, and a higher efficiency of certain types of posttranslational modifications (19). In the last 10 years, larvae have been successfully used to produce different recombinant proteins (3,5,7,13,16,21,22).…”

Rainbow trout surviving infections of viral haemorrhagic septicemia virus (VHSV) show lasting antibodies to recombinant G protein fragments

“…Their sequences were then chemically synthesized and cloned into pRSET expression vectors that add poly histidine tails. The G21-465 was expressed in Trichoplusia ni (cabbage looper) insect larvae [22,27] while the other fragments frg11 (56e110), frg15 (65e250) and frg16 (252e450) were expressed in Escherichia coli. Purification of the recombinant frgs was made by Ni 2þ affinity chromatography followed by Sephadex chromatography as described [28].…”

Expression and Immunological Studies of Classical Swine Fever Virus Glycoprotein E2 in the Bi-Cistronic Baculovirus/Larvae Expression System

“…17,21,22) To avoid rapid degradation of the recombinant protein by baculovirusderived proteases, an appropriate time point to collect the infected larvae is critical. 15,23,24) However, it is difficult to distinguish the degree of infection and/or the level of protein expression based merely on the larval appearance. 25) To remove these obstacles, a bi-cistronic expression vector was developed that allowed vAcP 10 -E2-infected larvae with GFP þ phenotype to be easily discriminated from uninfected larvae.…”