Development of a rapid method for determining the infectious dose (ID)50 of Orientia tsutsugamushi in a scrub typhus mouse model for the evaluation of vaccine candidates

The infectious dose (ID) of an inoculum for which 50% of susceptible mice will become infected (ID50) with Orientia tsutsugamushi is usually determined by rechallenging mice that have already been challenged with O. tsutsugamushi to determine the lethal dose (LD)(50) titer of the inoculum. Those mice not killed by the initial challenge and which survived a rechallenge with 1000 LD50 were considered immune and to have been initially infected with O. tsutsugamushi. Mice that succumbed to the rechallenge were considered not to have been initially infected. We have developed a more rapid method of determining the ID50 of inocula for use in our vaccine studies based upon the observation that mice surviving initial challenge and that go on to survive rechallenge produced detectable IgG to O. tsutsugamushi antigens by enzyme-linked immunosorbent assay (ELISA). Mice that did not survive rechallenge, and therefore did not receive an initial infectious inoculum did not produce detectable IgG to O. tsutsugamushi antigens. Both original LD50 and ID50 titers determinations require observation of mice for 21 days post-challenge. Our new ID50 determination does not require mice or the additional 21-day observation period for rechallenge and therefore is more rapid and cost-effective than the previous standard method of determining ID50 titer necessary for the evaluation of vaccine candidates. Published by Elsevier Ltd.