Loss of genetic diversity increases the risk of extinction of a population through inbreeding depression. In addition, the number of deleterious genetic variations, which might accumulate in a small population through genetic drift, can also make the population vulnerable. Rhododendron boninense is a critically endangered azalea species, which grows on the Bonin Islands, a UNESCO World Heritage Site; its population size in the wild dropped to only a single individual. Although a governmental program for species conservation has been conducted, no successful natural regeneration or population growth was achieved. We hypothesized that the reduced genetic diversity and accumulated deleterious genetic variation might cause the vulnerability of the species, and conducted comparative genetic and genomic analyses between R. boninense and its congeners. Genetic analysis using microsatellite markers indicated that the genetic diversity of R. boninense was ultimately low; microsatellite loci of this species were all fixed, whereas the congeners maintain high allelic diversity. Based on comprehensive transcriptome analysis, the amount of deleterious variation of R. boninense was significantly greater than that of R. amanoi. Repeated generation transitions of the small population of R. boninense on the oceanic islands is likely to have resulted in low genetic diversity as well as more deleterious variations in the genome, and we speculated that the accumulated deleterious variation might result in vulnerability of R. boninense. This study indicates the possibility of evaluating the feasibility of effective conservation programs based on the genetic diversity and the amount of accumulated deleterious variation in the genome of critically endangered species.