Plant-sucking stinkbugs are especially associated with mutualistic gut bacterial symbionts. Here, we explored the symbiotic relationship of a pistachio stinkbug, Acrosternum heegeri Fieber by histological, fluorescence in situ hybridization (FISH), real-time PCR and molecular phylogenetic techniques. Furthermore, the effects of the symbiont on the resting/wandering behaviors of the newborn nymphs, pre-adult survival rates, and stage compositions were investigated. Transmission electron microscopy and real-time PCR analyses showed that a rod-shaped gammaproteobacterium was persistently located within the posterior midgut crypts. Molecular phylogenetic and FISH techniques strongly suggested that this symbiont should be placed in the genus Pantoea of the Enterobacteriales. Scanning electron microscopy confirmed the presence of the bacterial cells on the egg surface which the surface sterilization of the eggs resulted in the successful removal of the symbiont from the eggs. Symbiotic and aposymbiotic A. heegeri showed no significant differences in the wandering behaviors of the first nymphal stages, while the symbiont-free insects suffered retarded growth and lower survivability. Together, the results highlight the habitat and acquisition features of Pantoea symbiont and its contribution in A. heegeri biology that might help us for better pest management in the future.

The galatheid crab Shinkaia crosnieri, is the sole member of the subfamily Shinkaiinae. It is abundant and forms dense beds around active hydrothermal vents in the Okinawa Trough. Thousands of filamentous bacteria attached to the plumose setae on the ventral surface of this crab were observed using field-emission scanning electron microscopy and transmission electron microscopy. Nucleic acids were extracted from the filamentous bacteria, and the phylotypes of 16S rRNA genes were identified from 81 clones. These phylotypes were divided into three groups: Epsilonproteobacteria (74%); Gammaproteobacteria (20%); and Bacteroidetes (6%). Gamma- and major phylotypes of Epsilonproteobacteria were also detected using fluorescence in situ hybridization analysis. These Epsilon- and Gammaproteobacteria were closely related to cultured and uncultured bacteria from hydrothermal vent fields including episymbionts of vent-associated invertebrates such as Rimicaris exoculata, Alvinella pompejana, the scaly-foot snail, Kiwa hirsuta etc. The carbon isotopic compositions of the muscle of S. crosnieri and in filamentous bacteria were similar. The muscle of S. crosnieri contained monounsaturated C16 and C18 fatty acids, which are known to be characteristic of sulphur-oxidizing bacteria in H2S-rich marine habitats. Through the video images transmitted by a submersible and a remotely operated vehicle, S. crosnieri was observed to comb out its ventral setae using the third maxilliped and appeared to consume the contents. These evidences suggest the epibiotic association between S. crosnieri and the filamentous bacteria attached to the ventral setae of the crab, but the details of role and function are still unclear at the present study.