Mealybugs with distinct endosymbiotic systems living on the same host plant

Mealybugs (Homoptera: Coccoidea: Pseudococcidae) possess a large bacteriome consisting of a number of bacteriocytes whose cytoplasm is populated by endosymbiotic bacteria. In many mealybugs of the subfamily Pseudococcinae, a peculiar endosymbiotic configuration has been identified: within the bacteriocytes, the primary betaproteobacterial endosymbiont Tremblaya princeps endocellularly harbor secondary gammaproteobacterial endosymbionts in a nested manner. Meanwhile, some mealybugs of the subfamily Phenacoccinae are associated only with a betaproteobacterial endosymbiont, designated as Tremblaya phenacola, which constitutes a distinct sister clade of T. princeps. However, cytological configuration of the endosymbiotic system in the phenacoccine mealybugs has not been established. Here, we investigated the endosymbiotic systems of the azalea mealybugs Crisicoccus azaleae (Pseudococcinae) and Phenacoccus azaleae (Phenacoccinae) living on the same host plants. Crisicoccus azaleae possessed a nested endosymbiotic system with T. princeps within the bacteriocyte cytoplasm and itself endocellularly harboring gammaproteobacterial cells, whereas P. azaleae exhibited a simple endosymbiotic system in which T. phenacola cells are localized within the bacteriocytes without additional gammaproteobacterial associates. Considering that these mealybugs live on the identical plant phloem sap, these different endosymbiotic consortia likely play similar biological roles for their host insects. The findings presented here should be helpful for future functional and comparative genomics toward elucidating evolutionary pathways of mealybugs and their endosymbionts.