The electron microscope continues to provide a unique contribution to the research program of the Laboratory of Infectious Diseases (LID). It has been employed in novel areas of research which have led to seminal discoveries that have resulted in the unveiling of new viral agents and the expansion and initiation of various programs. For example, application of the technique of immune electron microscopy led to the discovery of the 27nm Norwalk virus, which was the first major viral etiologic agent of epidemic gastroenteritis, and later the hepatitis A virus. In addition, the human rotavirus which was discovered in Australia and has now emerged as the single most important etiologic agent of severe diarrhea of infants and young children world-wide was visualized for the first time in the U.S. in LID studies that were performed by immune electron microscopy. Because the Norwalk and related caliciviruses have yet to be cultivated in any tissue culture system, along with the recent expression of 27nm virus-like particles belonging to some members of this group of agents, as well as studies of expressed viral proteins of rotavirus, the electron microscope continues to have an important role in the research program of the Epidemiology Section. It continues to be the only method (i) for direct detection of the Norwalk and related native viruses and derivative 27nm expressed virus-like particles and (ii) for unraveling the antigenic relationships among these fastidious agents. The continued need for electron microscopic support was evidenced by the over 30 individual experiments carried out by electron microscopy since last year?s annual report. The electron microscope was used in many capacities in the past year in individual experiments as an adjunct to other studies such as: (i) examination of clinical stool specimens by direct or by immune electron microscopy for pathogenic viruses in clinical trials in which individuals who were administered a candidate live rotavirus vaccine became ill; (ii) examination of various fractions of recombinant Hawaii or Snow Mountain virus preparations; (iii) examination of cell culture harvests following inoculation with Norwalk or Norwalk-like virus: (iv) support for molecular biologic studies of feline calicivirus by immune electron microscopy, (v) examination by direct or immune electron microscopy of specimens from studies aimed at expressing virus-like particles of the Snow Mountain virus; (vi) examination of specimens from a vaccinia virus transient system for expression of VP2, VP6, VP2/VP6 of SA11 rotavirus; (vii) comparison of three negative strains (phosphotungstic acid [PTA], ammonium molybdate, and uranyl acetate) for their ability to facilitate the visualization of expressed rotavirus VP2/VP6, (viii) comparison of ammonium molybdate, PTA, and glutaraldehyde plus PTA for facilitating the visualization of expressed rotavirus VP2; and (ix) examination of cell cultures for identification of a putative CPE producing agent derived from a hepatitis E patient.
