Topography of variola smallpox virus inverted terminal repeats

We examined the nucleotide sequences of the inverted terminal repeat (ITR) regions adjacent to the covalently closed hairpin end sequences of three variola major and four minor strains from smallpox outbreaks in Europe, Asia, Africa, and South America. The ITR regions ranged in size from 581 to 1051 base pairs (bp) and contained no apparent open reading frames. Two nonrepetitive sequence elements, NR1 and NR2, were conserved and resembled nonrepetitive elements in the ITRs of other orthopoxviruses. Depending on strain, the terminally positioned NR1 and the more internal NR2 flanked a direct repeat region containing from none to four copies of a 69-bp sequence and one copy of a 54-bp related sequence partial repeat. A distinctive pattern of ITR topography of NR1 and NR2 flanking a single copy of the 69-bp unit characterized each of three examined alastrim variola minor strains. A nonalastrim African minor strain from the last natural case of smallpox in Somalia in 1977 showed the largest ITR region of the examined viruses because of a second direct repeat cluster following NR2.