Symbiont polyphyly, co-evolution, and necessity in pentatomid stinkbugs from Costa Rica

Abstract

Interdomain symbioses with bacteria allow insects to take advantage of underutilized niches and provide the foundation for their evolutionary success in neotropical ecosystems. The gut microbiota of 13 micro-allopatric tropical pentatomid species, from a Costa Rican lowland rainforest, was characterized and compared with insect and host plant phylogenies. Like other families within the Pentatomomorpha, these insects (within seven genera—Antiteuchus, Arvelius, Edessa, Euschistus, Loxa, Mormidea, and Sibaria) house near-monocultures of gamma-proteobacteria in midgut crypts, comprising three distinct lineages within the family Enterobacteriaceae. Identity of the dominant bacteria (78–100% of the recovered 16S rRNA genes) was partially congruent with insect phylogeny, at the level of subfamily and tribe, with bacteria closely related to Erwinia observed in six species of the subfamily Pentatominae, and bacteria in a novel clade of Enterobacteriaceae for seven species within the subfamilies Edessinae and Discocephalinae. Symbiont replacement (i.e., bacterial “contamination” from the environment) may occur during maternal transmission by smearing of bacteria onto the egg surfaces during oviposition. This transmission strategy was experimentally confirmed for Sibaria englemani, and suspected for four species from two subfamilies, based on observation of egg probing by nymphs. Symbiont-deprived S. englemani, acquired via egg surface sterilization, exhibited significantly extended second instars (9.1 days compared with 7.9 days for symbiotic nymphs; p = 0.0001, Wilcoxon's rank with Bonferroni correction), slower linearized growth rates (p = 0.005, Welch 2-sample t-test), and qualitative differences in ceca morphology, including increased translucency of crypts, elongation of extracellular cavities, and distribution of symbionts, compared to symbiotic nymphs. Combined, these results suggest a role of the symbiont in host development, the reliable transference of symbionts via egg surfaces, and a suggestion of co-evolution between symbiont and tropical pentatomid host insects.

Female-Specific Specialization of a Posterior End Region of the Midgut Symbiotic Organ in Plautia splendens and Allied Stinkbugs

“…In the control group, most of the nymphs (44/48 ϭ 91.7%) were symbiont positive, which was in sharp contrast with the sterilized group, wherein all the nymphs (0/48 ϭ 0%) were symbiont negative. These results indicate that the symbiont is vertically transmitted to the next generation of P. splendens via egg surface contamination, as known for many other stinkbugs (1,(12)(13)(14)(15)(16)(17)(18)(19)(20)(21). Population dynamics of the symbiont.…”

“…Such obligate symbiotic bacteria associated with the midgut symbiotic organs have been described from a variety of stinkbugs of the families Pentatomidae (15,17,18,20,21,34,44), Scutelleridae (16,45,46), Cydnidae (19), Plataspidae (24,25,47,48), Acanthosomatidae (14), Parastrachiidae (22), Urostylididae (26), and others. As are the cases of the gut symbiotic bacteria associated with the stinkbugs representing the superfamily Pentatomoidea (21,49,50), the gut symbiont of P. splendens was placed within the Enterobacteriaceae of the Gammaproteobacteria, in contrast to the betaproteobacterial gut symbionts widely found across the superfamilies Lygaeoidea and Coreoidea (5). Although P. splendens is closely related and congenic to the brown-winged green stinkbug P. stali, the gut symbiont of P. splendens was phylogenetically distinct from the gut symbiont of P. stali (Fig.…”

“…Although P. splendens is closely related and congenic to the brown-winged green stinkbug P. stali, the gut symbiont of P. splendens was phylogenetically distinct from the gut symbiont of P. stali (Fig. 2) (49, 51), reflecting the polyphyletic and promiscuous nature of the host-symbiont associations in the Pentatomidae (21,49).…”

“…These results suggest that the enlarged end crypts represent a female-specific morphological trait specialized for vertical symbiont transmission commonly found among stinkbugs of the family Pentatomidae. The hypothesis that the symbiont transmission organ presumably evolved in the com- mon ancestor of extant pentatomid stinkbugs is, although further experimental works are needed to verify this idea, in impressive contrast with the polyphyletic evolutionary origins of the gut symbiotic bacteria in the Pentatomidae (21,49).…”

“…In these stinkbugs, adult females excrete symbiont-containing materials from the anus, which are orally acquired by their offspring for vertical symbiont transmission. Most commonly, as observed in the families Pentatomidae, Scutelleridae, Acanthosomatidae, and Cydnidae, vertical symbiont transmission is set up upon oviposition by egg surface contamination with symbiont-containing materials and establishes upon egg hatching by nymphal oral exploitation of the egg surface (1,(12)(13)(14)(15)(16)(17)(18)(19)(20)(21). In several presocial stinkbugs of the families Parastrachiidae and Cydnidae, adult females continuously attend and take care of their eggs and excrete symbiont-containing materials just before or after egg hatching, which newborns consume and establish vertical symbiont transmission (22,23).…”