High concentrations of dissolved biogenic methane associated with cyanobacterial blooms in East African lake surface water

The contribution of oxic methane production to greenhouse gas emissions from lakes is globally relevant, yet uncertainties remain about the levels up to which methanogenesis can counterbalance methanotrophy by leading to CH4 oversaturation in productive surface waters. Here, we explored the biogeochemical and microbial community variation patterns in a meromictic soda lake, in the East African Rift Valley (Kenya), showing an extraordinarily high concentration of methane in oxic waters (up to 156 µmol L−1). Vertical profiles of dissolved gases and their isotopic signature indicated a biogenic origin of CH4. A bloom of Oxyphotobacteria co-occurred with abundant hydrogenotrophic and acetoclastic methanogens, mostly found within suspended aggregates promoting the interactions between Bacteria, Cyanobacteria, and Archaea. Moreover, aggregate sedimentation appeared critical in connecting the lake compartments through biomass and organic matter transfer. Our findings provide insights into understanding how hydrogeochemical features of a meromictic soda lake, the origin of carbon sources, and the microbial community profiles, could promote methane oversaturation and production up to exceptionally high rates.

The rationale of sample choice was guided by the need to depict water column stratification, as a compromise of the number of samples that could be properly collected, handled and transported from such a remote area. Based on a preliminary profile of physicochemical characteristics (by multiple probe) we defined the sampling depth, based on the water column stratification. Another challenge for the number of samples, was related to the possibility to collect aliquots for dissolved gases analysis by a wooden artisanal raft, submerging a Rilsan® tube to the desired depth and purging it through a syringe.
Aliquots of water were defined based on current analytical methods for chemical and microbiological analysis. The amount of water filtered for DNA extraction was until filter clogging.
Data collection procedure is detailed in the method section. Sampling campaign was carried out by S. Fazi, S. Venturi, N. Pacini, E. Vazquez, Lydia A. Olaka and A. Butturini, with the kind logistic support of Mr. Silas W. Wanjala of the Naivasha Riparian Association and Mr. Lawi Kiplimo, Head Manager of the Crater Lake Sanctuary. During sampling campaign, the physical-chemical parameters were recorded in situ by a probe. Samples were fixed, filtered, stored at the Lake Naivasha Riparian Association Camp, were a field laboratory was available on the shore of the nearby Lake Naivasha. After that, aliquots were distributed among the different institutions that carried out the different analysis.
Samples were collected in ! one day at the spatial scale of meters No data were excluded No attempt to repeat the data collection was done, except for oxygen measurements that was repeated twice at 11.30 AM and 3.15 PM. The different results were interpreted as impact of primary production.
Being collected along a vertical profile, our data were not randomised. We grouped the sampling point by depth.
The extent of blinding during sampling depended on the fact that we defined the sampling depth a priori, based on the water column stratification. Moreover, during data acquisition samples were identified by numbers and the operators did not have direct access to correspondence with sample name.
As described in the method section, local climate at Lake Sonachi is warm and semiarid, with evaporation exceeding precipitation on an annual basis. Protection from wind by steep crater walls (rising up from 30 to 115 m above the lake surface) and vegetation (mainly Vachellia xanthophloea) limit water mixing. The hydrological balance is maintained by precipitation (~680 mm/year in the crater catchment) and evaporation (~1,870 mm/year). Furthermore, the occurrence of subsurface inflow from the near Lake Naivasha was proposed according to synchronous lake-level changes among the two lakes and other hydrological evidences. Chemical stratification and meromixis were documented across 8 years of periodic measurements and attributed to several local factors, including basin morphometry, diurnal periodicity of winds and thermal stratification, seasonal/yearly rainfall variations, and biological decomposition. Lake Sonachi is located at about 90 km NW of Nairobi at 1,884 m a.s.l., within the Eastern Rift Valley in central Kenya (0°46'57.68''S; 36°16''E).
The study was performed under research clearance permit NACOSTI/P/16/23342/10489 Biodiversity studies in Kenya's Rift Valley, granted to David M. Harper by the Government of Kenya. The study was conceived during the 2nd African International Symposium and Advanced Training Course on Ecohydrology for Water, Biodiversity, Ecosystem Services and Resilience in Africa November 2016organized by the UNESCO Ecohydrology Programme. Lake Naivasha basin is a demosite for the implementation of the UNESCO Ecohydrology Programme (Ecohydrology as the framework for sustainable utilization of water in the Naivasha basin -Kenya). During that symposium the study was conceived and discussed among different authors and Mr. Silas W. Wanjala of the Lake Naivasha Riparian Association. Moreover, Dr. Lydia A. Olaka from the Department of Geology (University of Nairobi, Kenya) joined the international team during the sampling campaign, collaborating on exploring hot springs adjacent to Lake Sonachi.
No disturbances need to be reported. Tick this box to confirm that the raw and calibrated dates are available in the paper or in Supplementary Information.

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Field-collected samples
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Preprocessing
Preprocessing software n/a n/a n/a n/a n/a Unfiltered and GFF-filtered water samples (2 mL) were fixed with a formaldehyde solution (final concentration 1%) and stored at 4°C until the analyses.

A50-micro from Apogee Flow Systems
Apogee Histogram (v89.0 -Apogee Flow System) Absolute volumetric counts were performed by staining with SYBR Green I (1:10000 dilution). A threshold was set to the green channel and samples were run at low flow rate (< 1000 events per s-1).
Fixed gates were designed to discriminate between free-living cells and aggregates according to their signatures in a side scatter vs. green fluorescence plot. Microbial aggregates were back-gated on a forward scatter histogram plot and divided into putative submicrometric and micrometric particles, respectively showing forward scatter signal intensities lower and higher than that of 1-µm size calibration beads used as internal standard. The .fcs files will be freely available at the Flow Repository identifier: https://flowrepository.org/id/[...].
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