Table 1.

Properties of wt DEN2 and 3′-SL mutant viruses derived by transfection of LLC-MK2 cells

VirusResult of IFAa at dayb:Plaque size (mm) (day)c
310
wt DEN2++++++2.0 (8)
D2/WN-SL+1.5 (20)
D2/WN-SL(mutA)++++2.0 (20)
D2/WN-SL(mutB)NA
D2/WN-SL(mutC)NA
D2/WN-SL(mutD)+++3.0 (20)
D2/WN-SL(mutE)+1.5 (20)
D2/WN-SL(mutF)+++4.0 (20)
D2-SL(a)++++4.0 (20)
D2-SL(b)+++1.5 (20)
D2-SL(c)NA
  • a Cells were stained on the days indicated with DEN2 HMAF (see Materials and Methods). The percentage of DEN antigen-positive cells was determined by examination of at least 100 cells. −, no positive cells; +, <10% positive cells; ++, 10 to 40% positive cells; +++, 40 to 70% positive cells; ++++, 70 to 100% positive cells.

  • b Days p.e. of LLC-MK2 cells. One-half milligram of RNA was transfected into 106 cells. On day 0, 105 transfected cells were seeded to a 1-cm2chamber for IFA.

  • c Viable mutant viruses were passaged three times in C6/36 cells or in LLC-MK2 cells [D2/WN-SL(mutE)]. The sequence of the 3′-terminal 242 nt in viral RNA was then verified, and the diameter of plaques was determined in LLC-MK2 cells. For all viable mutant viruses, plaques were not evident at day 8 postinfection. NA, not applicable; the mutation was lethal.