Bacterial symbionts of termite gut flagellates: cospeciation and nitrogen fixation in the gut of dry-wood termites
The subject of this thesis is the symbiosis between flagellates and bacteria in the gut of dry-wood termites (Kalotermitidae). In a series of studies, the evolution of devescovinid flagellates and their bacterial symbionts was elucidated, and the physiological basis of the symbiosis was investigated...
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|Summary:||The subject of this thesis is the symbiosis between flagellates and bacteria in the gut of dry-wood termites (Kalotermitidae). In a series of studies, the evolution of devescovinid flagellates and their bacterial symbionts was elucidated, and the physiological basis of the symbiosis was investigated, with a focus on nitrogen fixation.
Devescovinid flagellates are the dominant flagellates in the gut of Kalotermitidae. Species-pure suspensions of devescovinid flagellates (Devescovina and Metadevescovina species) from a wide range of termite species in the family Kalotermitidae were isolated with micropipettes. Ribosomal RNA gene sequences of the host flagellates and their bacterial symbionts were obtained using a full-cycle-rRNA approach.
Phylogenetic analysis showed that Devescovina spp. present in many species of Kalotermitidae form a monophyletic group. They were consistently associated with a distinct lineage of ectosymbionts, which form a monophyletic group among the Bacteroidales. The well-supported congruence of their phylogenies documented strict cospeciation of flagellates and their ectosymbionts, which were temporarily classified as “Candidatus Armantifilum devescovinae”. Nevertheless, the complete incongruence between the phylogenies of devescovinid flagellates and Kalotermitidae (COII genes) demonstrated horizontal transfer of flagellates among several species of Kalotermitidae.
The presence of filamentous “A. devescovinae” on the surface of Devescovina spp. was corroborated with scanning electron microscopy and fluorescent in situ hybridization. However, several Metadevescovina species, which form a sister group of Devescovina spp., did not possess Bacteroidales ectosymbionts. Moreover, a combination of molecular analysis and electron microscopy led to a correction of the previously overestimated diversity of Metadevescovina species in the gut of termite Incisitermes marginipennis.
In contrast to the Bacteroidales ectosymbionts, the endosymbionts of Devescovina spp., which belong to the so-called “Endomicrobia” (TG-1 phylum) and consistently colonized the cytoplasm of all flagellates of this group, were clearly polyphyletic. This suggested that they were acquired independently by each host species. The same seems to be true for the Bacteroidales ectosymbionts of the Oxymonas flagellates present in several Kalotermitidae. These ectosymbionts form several distantly related novel lineages in the phylogenetic tree, underscoring the notion that evolutionary histories of flagellate–bacteria symbioses in the termite gut are complex.
Kalotermitidae are known to fix large amounts of atmospheric nitrogen, and acetylene reduction assay showed the presence of nitrogenase activity in the gut of these termites. Community fingerprinting of the nitrogenase genes (homologs of nifH) by T-RFLP analysis revealed that a gene encoding an alternative nitrogenase (anfH) of unknown origin was most highly expressed homolog in mRNA-based profiles. Cloning of the nifH homologs from capillary-picked suspensions of Devescovina arta and Snyderella tabogae gave strong evidence that the “A. devescovinae” are the putative carriers of the anfH gene and therefore responsible for most of the nitrogen-fixing activity in the guts of Neotermes castaneus and Cryptotermes longicollis.
Despite a high diversity of nifH homologs in gut homogenates, the only other homologs that were expressed belonged to Treponema, Bacteroidales (nifH), and the proteo-cyano group. The gene expression profiles were specific for the termites. The anfH genes were not expressed in termite species that accumulated large amounts of hydrogen (35–45 kPa, microsensor measurements), suggesting a repression of gene expression by high hydrogen partial pressure.|