Production and consumption of hydrogen in hot spring microbial mats dominated by a filamentous anoxygenic photosynthetic bacterium

Abstract

Microbial mats containing the filamentous anoxygenic photosynthetic bacterium Chloroflexus aggregans develop at Nakabusa hot spring in Japan. Under anaerobic conditions in these mats, interspecies interaction between sulfate-reducing bacteria as sulfide producers and C. aggregans as a sulfide consumer has been proposed to constitute a sulfur cycle; however, the electron donor utilized for microbial sulfide production at Nakabusa remains to be identified. In order to determine this electron donor and its source, ex situ experimental incubation of mats was explored. In the presence of molybdate, which inhibits biological sulfate reduction, hydrogen gas was released from mat samples, indicating that this hydrogen is normally consumed as an electron donor by sulfate-reducing bacteria. Hydrogen production decreased under illumination, indicating that C. aggregans also functions as a hydrogen consumer. Small amounts of hydrogen may have also been consumed for sulfur reduction. Clone library analysis of 16S rRNA genes amplified from the mats indicated the existence of several species of hydrogen-producing fermentative bacteria. Among them, the most dominant fermenter, Fervidobacterium sp., was successfully isolated. This isolate produced hydrogen through the fermentation of organic carbon. Dispersion of microbial cells in the mats resulted in hydrogen production without the addition of molybdate, suggesting that simultaneous production and consumption of hydrogen in the mats requires dense packing of cells. We propose a cyclic electron flow within the microbial mats, i.e., electron flow occurs through three elements: S (elemental sulfur, sulfide, sulfate), C (carbon dioxide, organic carbon) and H (di-hydrogen, protons).

Fig. 1

Neighbor-joining phylogenetic tree of the phylum Thermodesulfobacteria based on 16S rRNA gene sequences (E. coli positions 160–845). Clones from this study are in bold. Accession numbers are shown in parentheses. Bootstrap support values >50% are given. Scale bar shows 2% estimated sequence divergence.

Fig. 2

Hydrogen production by microbial mats in artificial hot spring water at 65°C. A, intact mats: ●, in the dark without molybdate; ○, in the light without molybdate; ▲, in the dark with molybdate; △, in the light with molybdate. B, dispersed mats: ◆, in the dark: ⋄, in the light. C, D, phase-contrast photomicrographs of mats. C, intact mats; D, dispersed mats. Scale bar = 20 μm.

Fig. 3

Growth and hydrogen production of isolated strain HO-65 under anaerobic conditions in the dark. ▲ ABS₆₆₀; ● amount of hydrogen. Values are expressed as the means of three experiments. Error bars indicate SD.

Fig. 4

Model of material and electron flow in anaerobic regions within Nakabusa hot spring microbial mats. Carbon, hydrogen and sulfur flows are represented by white, black and gray arrows, respectively.