Microbial Communities in Methane- and Short Chain Alkane-Rich Hydrothermal Sediments of Guaymas Basin

Abstract

The hydrothermal sediments of Guaymas Basin, an active spreading center in the Gulf of California (Mexico), are rich in porewater methane, short-chain alkanes, sulfate and sulfide, and provide a model system to explore habitat preferences of microorganisms, including sulfate-dependent, methane- and short chain alkane-oxidizing microbial communities. In this study, hot sediments (above 60°C) covered with sulfur-oxidizing microbial mats surrounding a hydrothermal mound (termed "Mat Mound") were characterized by porewater geochemistry of methane, C2-C6 short-chain alkanes, sulfate, sulfide, sulfate reduction rate measurements, in situ temperature gradients, bacterial and archaeal 16S rRNA gene clone libraries and V6 tag pyrosequencing. The most abundantly detected groups in the Mat mound sediments include anaerobic methane-oxidizing archaea of the ANME-1 lineage and its sister clade ANME-1Guaymas, the uncultured bacterial groups SEEP-SRB2 within the Deltaproteobacteria and the separately branching HotSeep-1 Group; these uncultured bacteria are candidates for sulfate-reducing alkane oxidation and for sulfate-reducing syntrophy with ANME archaea. The archaeal dataset indicates distinct habitat preferences for ANME-1, ANME-1-Guaymas, and ANME-2 archaea in Guaymas Basin hydrothermal sediments. The bacterial groups SEEP-SRB2 and HotSeep-1 co-occur with ANME-1 and ANME-1Guaymas in hydrothermally active sediments underneath microbial mats in Guaymas Basin. We propose the working hypothesis that this mixed bacterial and archaeal community catalyzes the oxidation of both methane and short-chain alkanes, and constitutes a microbial community signature that is characteristic for hydrothermal and/or cold seep sediments containing both substrates.

Keywords: ANME; Guaymas Basin; alkanes; hydrothermal vents; methane; sediment.

FIGURE 1

(Left and Top Right) Panoramic views of Mat Mound, discovered during Alvin dive 4483. The mound is ca. 2.5 m high and several meters across. (Bottom Right) The basis of Mat Mound with close-up of the principal microbial mat area to be cored, photographed during dive 4484 before cores were taken. The two red laser points mark a distance of 10 cm. Small venting holes in the mat are visible in the mat foreground.

FIGURE 2

(Left) Profiles of dissolved methane concentrations, δ¹³C-CH₄ isotopic signatures, and sulfate concentrations, obtained from core 4484-3. δ¹³C-CH₄ values span a narrow range of approximately 3‰, and are plotted at high δ resolution. (Right) In situ temperature gradient obtained by Heatflow Probe measurement #3 (see Supplementary Figure S2C for location). The hyphenated lines indicate the 4–5 cm sample interval and its interpolated temperature range.

FIGURE 3

Concentration profiles (in μM) and δ¹³C values (in ‰) for short-chain alkanes (C₁ to C₆) from Guaymas sediment cores 4484-6 and 4484-14. The concentration scale is discontinuous to accommodate methane as well as the short-chain alkanes within the same plots. When available, error bars from triplicate measurements are included.

FIGURE 4

Hydrogen concentrations, shown as averages with standard deviations, after >72 h of pre-incubation at 20°C for all core depths (two Left panels, and Center panel), at increasing temperatures downcore (second panel from the Right), and at 4°C for all core depths (Right). The warm cores 4483-3, 4484-14, and 4484-6 were obtained from the mat-covered periphery of Mat Mound; the cool background core 4485-1 represents bare sediment at approximately 1 m distance from Mat Mound. Prolonged incubation (134–213 h) of core 4484-6 at >50°C shown in the second panel from the right resulted in localized H₂ accumulation in the range of 100 to 300 nM, indicating that microbial H₂ production can exceed consumption under these conditions. Interpolated temperatures for every incubated sediment sample are shown in the temperature profiles below each hydrogen profile; they are based on the in situ positions of cores and temperature profiles in Supplementary Figure S1 (core 4483-3, profile HF7; and core 4485-1, profile HF1) and Supplementary Figure S2C (core 4484-6, profile HT2). Core 4484-14 has no nearby temperature profile, but its position near the base of Mat Mound suggests high temperatures.

FIGURE 5

(A,B) Bar diagrams of bacterial and archaeal clone library and V6-tag composition, for DNA-based PCR amplicons in the 0–1 and 4–5 cm sediment layers. The phylogenetic groups that are found only in the V6-tag survey but not in the clone libraries appear in different shades of gray.

FIGURE 6

Phylogeny of methanogens and ANME archaea in Guaymas Mat Mound hydrothermal sediments, based on partial 16S rRNA gene sequences (E. coli positions 28–915). The tree was inferred using neighbor joining based on Jukes–Cantor sequence distances, and was checked by 1000 bootstrap iterations, as implemented in ARB.

FIGURE 7

Phylogeny of Proteobacteria and the HotSeep-1 Group in Guaymas Mat Mound hydrothermal sediments, based on near-complete 16S rRNA gene sequences (E. coli positions 28–1491). The tree was inferred using neighbor joining based on Jukes–Cantor sequence distances, and was checked by 1000 bootstrap iterations, as implemented in ARB.