Environmental Transmission of the Gut Symbiont Burkholderia to Phloem-Feeding Blissus insularis

Abstract

The plant-phloem-feeding Blissus insularis possesses specialized midgut crypts, which harbor a dense population of the exocellular bacterial symbiont Burkholderia. Most individual B. insularis harbor a single Burkholderia ribotype in their midgut crypts; however, a diverse Burkholderia community exists within a host population. To understand the mechanism underlying the consistent occurrence of various Burkholderia in B. insularis and their specific association, we investigated potential gut symbiont transmission routes. PCR amplification detected a low titer of Burkholderia in adult reproductive tracts; however, fluorescence in situ hybridization assays failed to produce detectable signals in these tracts. Furthermore, no Burkholderia-specific PCR signals were detected in eggs and neonates, suggesting that it is unlikely that B. insularis prenatally transmits gut symbionts via ovarioles. In rearing experiments, most nymphs reared on St. Augustinegrass treated with cultured Burkholderia harbored the cultured Burkholderia strains. Burkholderia was detected in the untreated host grass of B. insularis, and most nymphs reared on untreated grass harbored a Burkholderia ribotype that was closely related to a plant-associated Burkholderia strain. These findings revealed that B. insularis neonates acquired Burkholderia primarily from the environment (i.e., plants and soils), even though the possibility of acquisition via egg surface cannot be excluded. In addition, our study explains how the diverse Burkholderia symbiont community in B. insularis populations can be maintained.

Impacts of Antibiotic and Bacteriophage Treatments on the Gut-Symbiont-Associated Blissus insularis (Hemiptera: Blissidae)

“…Other insects, such as the seed-feeding Riptortus bug, can be maintained on sterilized soybean seeds for multiple generations [10], providing an avenue to examine axenic insects without antibiotic therapy. Unfortunately, B. insularis neonates require live St. Augustinegrass that is known to harbor Burkholderia , precluding access to an axenic food source [21]. …”

“…Due to the high specificity of bacteriophage to the target bacteria, B. insularis infected with target gut-symbiotic Burkholderia were generated by rearing neonates on live plants inoculated with cultured symbionts (Bi16MC_R_vitro isolate), as described previously [21]. After three weeks of rearing on Burkholderia -inoculated plants, fourth instars were collected, provided with surface-sterilized corn kernels in a plastic container, and held at 27 °C with a 14:10 (L:D) h photoperiod.…”

“…Samples producing ≥75% homology in the BOX pattern of that produced by the Bi16MC_R_vitro DNA were considered as positive infections [21]. …”

“…Unlike many Burkholderia , the Blissus -associated Burkholderia cannot be directly cultured on bacteriological media but require an initial organ culture to transition these bacteria to an in vitro environment [20]. Genomic analysis has revealed that most B. insularis individuals harbor a single Burkholderia ribotype while a chinch bug population harbors a diverse Burkholderia community [20,21]. A recent study reveals that the oral acquisition of Burkholderia from the environment plays a key role in maintaining ribotype diversity of gut symbiont in B. insularis populations [21].…”

“…Genomic analysis has revealed that most B. insularis individuals harbor a single Burkholderia ribotype while a chinch bug population harbors a diverse Burkholderia community [20,21]. A recent study reveals that the oral acquisition of Burkholderia from the environment plays a key role in maintaining ribotype diversity of gut symbiont in B. insularis populations [21]. In addition to B. insularis , exocellular Burkholderia symbionts are also found in the midgut lumen of heteropterans in other genera of Lygaeoidea and Coreoidea superfamilies [22,23,24,25].…”