Diel variations in carbon metabolism by green nonsulfur-like bacteria in alkaline siliceous hot spring microbial mats from Yellowstone National Park

Abstract

Green nonsulfur-like bacteria (GNSLB) in hot spring microbial mats are thought to be mainly photoheterotrophic, using cyanobacterial metabolites as carbon sources. However, the stable carbon isotopic composition of typical Chloroflexus and Roseiflexus lipids suggests photoautotrophic metabolism of GNSLB. One possible explanation for this apparent discrepancy might be that GNSLB fix inorganic carbon only during certain times of the day. In order to study temporal variability in carbon metabolism by GNSLB, labeling experiments with [13C]bicarbonate, [14C]bicarbonate, and [13C]acetate were performed during different times of the day. [14C]bicarbonate labeling indicated that during the morning, incorporation of label was light dependent and that both cyanobacteria and GNSLB were involved in bicarbonate uptake. 13C-labeling experiments indicated that during the morning, GNSLB incorporated labeled bicarbonate at least to the same degree as cyanobacteria. The incorporation of [13C]bicarbonate into specific lipids could be stimulated by the addition of sulfide or hydrogen, which both were present in the morning photic zone. The results suggest that GNSLB have the potential for photoautotrophic metabolism during low-light periods. In high-light periods, inorganic carbon was incorporated primarily into Cyanobacteria-specific lipids. The results of a pulse-labeling experiment were consistent with overnight transfer of label to GNSLB, which could be interrupted by the addition of unlabeled acetate and glycolate. In addition, we observed direct incorporation of [13C]acetate into GNSLB lipids in the morning. This suggests that GNSLB also have a potential for photoheterotrophy in situ.

FIG. 1.

H₂ and H₂S profiles from Mushroom Spring microbial mats measured in situ at different times around sunset and sunrise on 24 to 25 August 1999.

FIG. 2.

The degree of labeling of specific lipids expressed in Δδ¹³C (per mille) relative to natural-abundance stable carbon isotopic composition on a log scale. (A) 1997 afternoon and morning labeling experiments. (B) 2000 morning labeling experiments (error in wax ester data from duplicate cores of the preincubation [pre-inc.] experiments is too small to include in the figure [Table 2]).

FIG. 3.

¹⁴C-labeled acetate diffusion experiment into mat core in the light and in the dark followed by cryosectioning and autoradiography. Silver grains, which appear as tiny black dots, demonstrate where the labeled substrate has diffused during the incubation.

FIG. 4.

The degree of labeling of specific lipids expressed in Δδ¹³C (per mille) relative to natural-abundance stable carbon isotopic composition on a log scale. (A) 1997 pulse-labeling experiment. (B) 1999 morning [¹³C]acetate labeling experiment. T1, 5:20 p.m.; T2, 6:45 a.m.; T3, 10:55 a.m.

FIG. 5.

Working model for carbon flow in an alkaline, siliceous hot spring microbial mat over a 24-h period. Only metabolic pathways thought to be relevant to photoautotrophy and photoheterotrophy are shown.