COLD-ADAPATION OF HUMAN ROTAVIRUSES

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Acute nonbacterial gastroenteritis has long been recognized throughout the world as one of the most frequent and important human diseases. Within this broad disease complex, group A rotaviruses are the single most important etiologic agents of severe and often life-threatening diarrhea of infants and young children worldwide. Three candidate live oral rotavirus vaccines that have been evaluated in various parts of the world were developed using the "Jennerian" approach. In this approach, an antigenically-related live virus derived from a non-human host is used as a vaccine for immunization against its human virus counterpart. The protective efficacy of the two monovalent bovine or the simian rotavirus vaccines has proved to be variable. In addition, the target population of two-to-five-month-old infants characteristically developed only a homotypic immune response following vaccination. As a consequence, a modified "Jennerian" approach was developed for the formulation of a quadrivalent vaccine. This involved replacing the VP7 gene of the animal (rhesus or bovine) rotavirus by the corresponding gene of each of the human rotaviruses of major clinical importance. Group A rotaviruses possess two outer capsid proteins that function as independent neutralization antigens, namely VP4 (encoded by genome segment 4) and VP7 (encoded by genome segment 7, 8, or 9 depending on the strain). Although initially VP7 was thought to be the dominant neutralization antigen, later studies showed that VP4 is as effective as VP7 in inducing neutralizing antibodies following experimental infection of experimental animals or vaccine induced infection of susceptible infants or young children. Also, antibodies to VP4 or VP7 are independently associated with resistance of gnotobiotic piglets to experimental challenge with virulent rotavirus. However, VP7 is the only relevant rotavirus protective antigen that is present in candidate live attenuated vaccines that are currently under evaluation for protective efficacy in humans; these vaccines contain the VP4 of an animal rotavirus that is not related antigenically to the VP4 of any of the epidemiologically important human rotaviruses. In an attempt to maximize immunogenicity of live rotavirus vaccines by incorporating both protective antigens (i.e., VP4 and VP7) of clinically important rotaviruses, we pursued another approach to vaccine development in which cold-adapted (ca) and temperature-sensitive (ts) mutants of human rotaviruses were sought. This strategy, which entails the selection of such mutants during adaptation of virus to grow efficiently at suboptimal temperature, has been successfully employed by others to select various attenuated candidate human viral vaccine strains for the polioviruses, influenza A viruses, parainfluenza type 3 virus, and respiratory syncytial virus. The objectives of this project are: (i) to cold-adapt selected wild-type human rotaviruses, (ii) to analyze genetically various phenotypes exhibited by such cold-adapted viruses including temperature sensitivity, cold-adaption, and attenuation, and (iii) to evaluate these ca rotaviruses for their potential usefulness in a live rotavirus vaccine for humans.