Bacteria Associated With Piezodorus guildinii (Hemiptera: Pentatomidae), With Special Reference to Those Transmitted by Feeding.

The redbanded stink bug, Piezodorus guildinii (Westwood) (Hemiptera: Heteroptera: Pentatomidae), is a rapidly growing pest damaging southern US agriculture. Pentatomid stink bugs are known to vector bacterial, fungal, and viral plant diseases. However, bacteria associated with redbanded stink bugs and their vector potential have not yet been assessed. In this study, we 1) cultured and identified bacteria transmitted by feeding of redbanded stink bug and 2) described bacteria from guts of redbanded stink bug individuals using next-generation sequencing of 16S rRNA genes. Nineteen bacteria transmitted by feeding of redbanded stink bug on soybean agar were isolated and identified via Sanger sequencing of near full length 16S RNA genes. The transmitted bacteria belonged to at least a dozen species in eight genera and included potential plant pathogens (Phaseolibacter flectens), plant beneficials (Bacillus atropheus), and possible insect beneficials (Acinetobacter sp. and Citrobacter farmeri). A total of 284,448 reads were captured from Illumina MiSeq sequencing of the uncultured gut bacteria community. Fifty-one putative bacteria species (74% of the estimated total species richness) were identified via matches to NCBI databases. The bacteria metagenome contained potential plant and insect pathogens (Erwinia persicina, E. rhaponici, Brenneria nigrifluens, Ralstonia picketti, and Serratia marcescens) and beneficials (Pantoea dispersa, Klebsiella oxytoca, Clostridium butyricum, and Citrobacter farmeri).

Assessment of genetically engineered Trabulsiella odontotermitis as a 'Trojan Horse' for paratransgenesis in termites.

BACKGROUND: The Formosan subterranean termite, Coptotermes formosanus is an invasive urban pest in the Southeastern USA. Paratransgenesis using a microbe expressed lytic peptide that targets the termite gut protozoa is currently being developed for the control of Formosan subterranean termites. In this study, we evaluated Trabulsiella odontotermitis, a termite-specific bacterium, for its potential to serve as a 'Trojan Horse' for expression of gene products in termite colonies. RESULTS: We engineered two strains of T. odontotermitis, one transformed with a constitutively expressed GFP plasmid and the other engineered at the chromosome with a Kanamycin resistant gene using a non- disruptive Tn7 transposon. Both strains were fed to termites from three different colonies. Fluorescent microscopy confirmed that T. odontotermitis expressed GFP in the gut and formed a biofilm in the termite hindgut. However, GFP producing bacteria could not be isolated from the termite gut after 2 weeks. The feeding experiment with the chromosomally engineered strain demonstrated that T. odontotermitis was maintained in the termite gut for at least 21 days, irrespective of the termite colony. The bacteria persisted in two termite colonies for at least 36 days post feeding. The experiment also confirmed the horizontal transfer of T. odontotermitis amongst nest mates. CONCLUSION: Overall, we conclude that T. odontotermitis can serve as a 'Trojan Horse' for spreading gene products in termite colonies. This study provided proof of concept and laid the foundation for the future development of genetically engineered termite gut bacteria for paratransgenesis based termite control.