title: Recurrent evolution of gut symbiotic bacteria in pentatomid stinkbugs.
abstract: BACKGROUND
  Diverse animals are intimately associated with microbial symbionts. How such host-symbiont associations have evolved is a fundamental biological issue. Recent studies have revealed a variety of evolutionary relationships, such as obligatory, facultative, and free-living, of gut bacterial symbiosis within the stinkbug family Pentatomidae, although the whole evolutionary picture remains elusive.
  
  
  RESULTS
  Here we investigated a comprehensive assembly of Japanese pentatomid stinkbugs representing 28 genera, 35 species, and 143 populations. Polymerase chain reaction (PCR), cloning, and sequencing of bacterial 16S rRNA gene from their midgut symbiotic organ consistently detected a single bacterial species from each of the insect samples, indicating a general tendency toward monosymbiotic gut association. Bacterial sequences detected from different populations of the same species were completely or nearly identical, indicating that the majority of the gut symbiotic associations are stably maintained at the species level. Furthermore, bacterial sequences detected from different species in the same genus tended to form well-supported clades, suggesting that host-symbiont associations are often stable even at the genus level. Meanwhile, when we compared such sequences with published sequences available in DNA databases, we found a number of counter-examples to such stable host-symbiont relationships; i.e., symbionts from different host species in the same genus may be phylogenetically distant, and symbionts from the same host species may be phylogenetically diverse. Likewise, symbionts of diverse pentatomid species may be closely related to symbionts of other stinkbug families, and symbionts of diverse pentatomid species may even be allied to free-living bacteria. Molecular evolutionary analyses revealed that higher molecular evolutionary rates, higher AT nucleotide compositions, and smaller genome sizes tended to be associated with the pentatomid symbionts constituting the stable lineages, whereas these traits were rarely observed in the pentatomid symbionts of promiscuous type.
  
  
  CONCLUSIONS
  These results indicate that gut symbiotic bacteria have evolved repeatedly and dynamically in the stinkbug family Pentatomidae, which have plausibly entailed frequent symbiont acquisitions, losses, replacements and transfers, while establishing a number of relatively stable host-symbiont associations. The diverse host-symbiont relationships observed in the Pentatomidae will provide an ideal arena for investigating the evolution of symbiosis experimentally and theoretically.
authors.affiliation: National Institute of Advanced Industrial Science and Technology (AIST), Tsukuba, 305-8566 Japan.|Faculty of Science, Kyushu University, Fukuoka, 819-0395 Japan.
authors.forename: Takahiro
authors.initials: T
authors.lastname: Hosokawa
authors.affiliation: National Institute of Advanced Industrial Science and Technology (AIST), Tsukuba, 305-8566 Japan.|Tropical Biosphere Research Center, University of the Ryukyus, Okinawa, 903-0213 Japan.
authors.forename: Yu
authors.initials: Y
authors.lastname: Matsuura
authors.affiliation: National Institute of Advanced Industrial Science and Technology (AIST), Hokkaido Center, Sapporo, 062-8517 Japan.
authors.forename: Yoshitomo
authors.initials: Y
authors.lastname: Kikuchi
authors.affiliation: National Institute of Advanced Industrial Science and Technology (AIST), Tsukuba, 305-8566 Japan.|Department of Biological Sciences, Graduate School of Science, University of Tokyo, Tokyo, 113-0033 Japan.|Graduate School of Life and Environmental Sciences, University of Tsukuba, Tsukuba, 305-8572 Japan.
authors.forename: Takema
authors.identifier: 0000-0001-5987-2602
authors.initials: T
authors.lastname: Fukatsu
keywords: 16S rRNA gene; Gut symbiont; Molecular evolution; Pentatomidae; Reductive genome evolution; Stinkbug; γ-Proteobacteria