Genetic bottlenecks reduce population variation in an experimental RNA virus population

Abstract

Genetic bottlenecks are stochastic events that limit genetic variation in a population and result in founding populations that can lead to genetic drift. Evidence of past genetic bottlenecks in numerous biological systems, from mammals to viruses, has been described. In this study, we used an artificial population of Cucumber mosaic virus consisting of 12 restriction enzyme marker-bearing mutants. This population was inoculated onto young leaves of tobacco plants and monitored throughout the course of systemic infection. We show here that the genetic variation in a defined population of an RNA virus is significantly, stochastically, and reproducibly reduced during the systemic infection process, providing clear evidence of a genetic bottleneck.

FIG. 1.

Schematic representation of plant materials. (A) Tobacco plant showing the location of the inoculated leaf (5th) and the primary (8th) and secondary (15th) systemically infected leaves. (B) Cross-sectional view of a minor vein. The cell-to-cell pathway by which virus is transported via PD is shown by arrows moving from mesophyll cells and passing through BS, VP, and companion cells (C) before entering the sieve elements (S). The PD are abundant in the walls between the mesophyll cells but are limited at the interface of the VP cells and the CC-SE complex. X, xylem.

FIG. 2.

Reduction in virus population during systemic infection of Cucumber mosaic virus. Tobacco plants were inoculated with a mixture of 12 mutant viruses of Fny CMV, and the inoculated leaves and systemic leaves of these plants were harvested at 2, 10 (the 8th leaf), and 15 (the 15th leaf) dpi, respectively. Analysis for the presence of each mutant was carried out by RT-PCR followed by restriction enzyme digestion. Exp1, Exp2, and Exp3 are three independent experiments. In, inoculated leaves; 8th and 15th, systemically infected leaves. All values are expressed as the mean numbers of the recovered mutants ± one standard error. Bars with different letters (a, b, or c) are significantly different from each other (P < 0.05), as determined by least significant difference test.

FIG. 3.

Recovery frequency of each mutant in systemically infected leaves. The recovery percentage of systemic leaves for each mutant was calculated for each experiment described in the legend to Fig. 2. The mean recovery percentage of each mutant was obtained by treating each experiment as a replicate; a to l represent the 12 mutants used in this study. All values are expressed as the mean recovery percentages ± one standard error.