Microbiome of psyllids of the family Aphalaridae, including Aphalara itadori, a biocontrol agent against Reynoutria spp.

Several European and North American countries have started releasing the Japanese knotweed psyllid, Aphalara itadori (Shinji) (Hemiptera: Aphalaridae), to control the Japanese knotweed, Reynoutria japonica Houtt. (Polygonaceae), and its relatives, which are among the worst invasive exotic plants. However, establishing populations of the currently released strains in the field has not been successful, desiring newly collected lineages. Moreover, little is known about the microbiome of the current strains, which potentially impacts properties as biocontrol agents. Hence, this study analyzed the microbiota of an A. itadori strain newly collected on Honshu Island, Japan, along with related species of the family Aphalaridae, using amplicon sequencing of 16S rRNA genes. The localization of symbionts identified in A. itadori was further analyzed using fluorescence in situ hybridization. The results demonstrated that the A. itadori bacteriome, a specialized organ for microbial symbiosis, maintains a dual symbiotic system with the primary symbiont “Candidatus Carsonella ruddii” (Gammaproteobacteria: Oceanospirillales: Halomonadaceae) and the secondary symbiont Sodalis sp. (Gammaproteobacteria: Enterobacterales: Pectobacteriaceae), suggesting that they are evolutionarily stable obligate mutualists for A. itadori. The central area of the bacteriome containing Sodalis comprised uninucleate bacteriocytes with nuclei larger than those of bacteriocytes harboring Carsonella. This observation contrasted previous reports on various psyllid lineages in which secondary symbionts are housed in a central syncytium with nuclei smaller than those of bacteriocytes for Carsonella. No known plant pathogens or parasitic manipulators of insect reproduction were identified in the analyzed A. itadori strain, indicating its suitability as a biocontrol agent, posing a minimum risk to the ecosystem. Besides distinct Carsonella lineages, Sodalis independently acquired by Craspedolepta miyatakeai Klimaszewski and an ambiguous Enterobacterales symbiont in Epheloscyta kalopanacis Loginova were identified. Only Carsonella was found in Togepsylla matsumurana Kuwayama. These results indicate repeated infections and replacements of bacterial symbionts during the evolution of Psylloidea, providing deeper insights into the microbe-psyllid interactions.