Lack of stability of the infectivity of budded virus of Anticarsia gemmatalis multiple nucleopolyhedrovirus in serum-free medium supplemented with lipid microemulsions

EBERHARDT I.; GIORIA V.; MICHELOUD G.; CLAUS J. D.

Buenos Aires

Congreso; 2012 International Congress on Invertebrate Pathology and Microbial Control and  45th Annual Meeting of the Society for Invertebrate Pathology ; 2012

Biotechnological applications of baculoviruses are constantly expanding, from bioinsecticides to protein expression and gene therapy vectors. The development of feasible processes for the production of baculovirus and baculovirus-based products in insect cell cultures requires that high-titer stocks of budded virus (BVs) being prepared and stored in serum-free medium. It is known that the preservation of baculovirus BVs by freezing at ultra-low temperatures in serum-free medium is less efficient than in culture media added with serum, but the causes were not elucidated. The aim of this work was to study the stability of the infectivity of BVs of the Anticarsia gemmatalis multiple nucleopolyhedrovirus (AgMNPV) in the UNL-10 serum-free medium under different conditions of  supplementation with lipid microemulsions, freezing at -80°C, and time of exposure to 27°C, employing a 23 full factorial design in duplicate. The exposure to 27°C, as well as the freezing and thawing of an AgMNPV stock, did not affect the stability of the viral titer determined through an end-point dilution method. On the other hand, the exposure to lipid microemulsion provoked a significant negative effect on the stock titer. This deleterious effect was magnified when BV samples in medium supplemented with lipid microemulsion were frozen and thawed, losing more than 90% of the viral infectivity. These results strongly suggest that the reduced stability of AgMNPV BVs in serum-free media is associated to the presence of lipid microemulsions, and indicate the necessity to establish alternative protocols to store BVs stocks produced in serum-free medium supplemented with lipid microemulsions.