at the outflow of an sulfide-rich hot spring.

Alkaline sulfide-rich hot springs provide a unique environment for microbial community and arsenic (As) biogeochemistry. In this study, a representative alkaline sulfide-rich hot spring, Zimeiquan in the Tengchong geothermal area, was chosen to study arsenic geochemistry and microbial community using Illumina MiSeq sequencing. Over 0.26 million 16S rRNA sequence reads were obtained from 5-paired parallel water and sediment samples along the hot spring’s outflow channel. High ratios of As(V)/ As Sum (total combined arsenate and arsenite concentrations) (0.59–0.78), coupled with high sulfide (up to 5.87 mg/L), were present in the hot spring’s pools, which suggested As(III) oxidation occurred. Along the outflow channel, As Sum increased from 5.45 to 13.86 μ mol/L, and the combined sulfide and sulfate concentrations increased from 292.02 to 364.28 μ mol/L. These increases were primarily attributed to thioarsenic transformation. Temperature, sulfide, As and dissolved oxygen significantly shaped the microbial communities between not only the pools and downstream samples, but also water and sediment samples. Results implied that the upstream Thermocrinis was responsible for the transformation of thioarsenic to As(III) and the downstream Thermus contributed to derived As(III) oxidation. This study improves our understanding of microbially-mediated As transformation in alkaline sulfide-rich hot springs. for hydrogen gas (H 2 ), sulfur (S), iron (Fe) and arsenic (As), as well as having high temperature and a wide range of pH values 1–5 . Arsenic biogeochemistry has been studied extensively in acidic hot springs and it is well-documented that high concentrations of sulfide can inhibit microbial As(III) oxidation in acidic springs by inactivating expressed As(III) oxidase (AioA) in microorganisms such as Hydrogenobacter 6 , Hydrogenobaculum and Acidicaldus 8 . In contrast, sulfide has been shown to enhance microbial As(III) oxidation in the alkaline Mono Lake water by stimulating growth of sulfur-oxidizing bacteria, the first demonstration of the different roles of sulfide on microbial As(III) oxidation in acidic and alkaline environments 9 . Previous studies have revealed the presence of thioarsenic in alkaline sulfidic geothermal waters, and As and S oxidations were closely correlated in this type of system 10,11 . Thioarsenate was identified as the dominant As species at the source of the alkaline sulfide-rich hot springs in Yellowstone National Park (YNP) and account for up to 89% of total As 10,11 . Upon discharge, thioarsenate is biologically transformed to As(III) along the outflow channels, and is followed by appear- ance of As(V) and sulfate as the final products 12,13 . Studies of microbially controlled As transformation in mats or sediments in these alkaline sulfide-rich hot springs have been investigated using enrichments, pure cultures, clone libraries and metagenome sequencing and found Thermocrinis , Thermus and Ectothiorhodospira to be likely involved in transformation of As species 14–17 . As an example, anoxic cultures from alkaline Mono Lake and Big Soda Lake, which were dominated by Ectothiorhodospira , were able to use monothioarsenate as the sole elec- tron donor for anoxygenic photosynthesis . Further, the hyperthermophilic Thermocrinis rubber isolated from and by inductively plasma-optical emission spectrometry and ion chromatography (ICS1100, respectively. Arsenic concentrations were determined using liquid chromatography-hydride generation-atomic fluorescence spectrometry (LC-HG-AFS, Beijing) 33 . Thioarsenic species were identified using Q Exactive, a high resolution quadrupole orbitrap mass spectrometer (Thermo Scientific, Germany), by detecting the accurate mass and matching the isotope abundance. DOC of water samples were determined using a TOC analyzer (TOC-V CPH , Shimadzu, Japan). As and Fe in the sediments were extracted by 1:1 aqua regia digestion in a water bath 45 . Extracted Fe from sediments was determined by a 1,10-Phenanthroline-based assay: 10 mL extracted solutions were mixed with 5 mL acetate-sodium acetate buffer (pH = 4.6), 2.5 mL 1% hydroxylamine hydrochloride and 5 mL 0.1% 1,10-phenanthroline solution in a 50 mL volumetric flask. The mixture was brought up to a volume of 50 mL with deionized water and allowed to stand for 10 min. The absorbance of each solution at 510 nm was measured with a spectrophotometer (UV1750, Shimadzu, Japan). TOC of sediment samples was measured with a Macro elemental analyzer (Multi EA 4000, Analytik Jena, Germany) after inorganic carbon was digested using HCl. and the experiments. and contributed to field sampling. and conducted the experiment. the first of the and