Synchronization of cowpea chlorotic mottle virus replication in cowpea leaves

A technique that 'systemically inoculates' cowpea leaves with cowpea chlorotic mottle virus (CCMV) is described. The upper leaves of cowpea plants are maintained at 10 degrees and allowed to become infected with inoculum from lower leaves maintained at 32 degrees; when the whole plant is shifted to 25 degrees, synchronous virus replication starts in the upper leaves. Following the shift to 25 degrees, no infectivity could be detected during the first 6 h, after which the infectivity increased exponentially between 8 and 20 h and linearly between 20 and 72 h. The time-courses of effectiveness of different inhibitors occurred sequentially and at different times during the infection. An actinomycin-D-sensitive step occurred at 10 degrees, prior to the shift to 25 degrees. A 2.0 mM guanidine-sensitive step occurred between 4 and 20 h and a cycloheximide-sensitive step occurred between 12 and 48 h after the shift to 25 degrees. When the upper leaves were maintained at 5 degrees while the infected lower leaves were at 32 degrees, viral inoculum did not move into them. However, rapid systemic infection occurred during the first 6 h after a shift of the whole plant to 25 degrees.