ABSTRACT
Lignocellulose digestion by wood-feeding termites depends on the mutualistic interaction of unusual, flagellate protists located in their hindgut. Most of the flagellates harbor numerous prokaryotic endosymbionts of so-far-unknown identity and function. Using a full-cycle molecular approach, we show here that the endosymbionts of the larger gut flagellates of
Reticulitermes santonensis
belong to the so-called termite group 1 (TG-1) bacteria, a group of clones previously obtained exclusively from gut homogenates of
Reticulitermes speratus
that are only distantly related to other bacteria and are considered a novel bacterial phylum based on their 16S rRNA gene sequences. Fluorescence in situ hybridization with specifically designed oligonucleotide probes confirmed that TG-1 bacteria are indeed located within the flagellate cells and demonstrated that
Trichonympha agilis
(Hypermastigida) and
Pyrsonympha vertens
(Oxymonadida) harbor phylogenetically distinct populations of symbionts (<95% sequence similarity). Transmission electron microscopy revealed that the symbionts are small, spindle-shaped cells (0.6 μm in length and 0.3 μm in diameter) surrounded by two membranes and located within the cytoplasm of their hosts. The symbionts of the two flagellates are described as candidate species in the candidate genus “
Endomicrobium
.” Moreover, we provide evidence that the members of the TG-1 phylum, for which we propose the candidate name “
Endomicrobia
,” are phylogenetically extremely diverse and are present in and also restricted to the guts of all lower termites and wood-feeding cockroaches of the genus
Cryptocercus
, the only insects that are in an exclusive, obligately mutualistic association with such unique cellulose-fermenting protists.