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.
The hydrothermal mats, mounds, and chimneys of the southern Guaymas Basin are the surface expression of complex subsurface hydrothermal circulation patterns. In this overview, we document the most frequently visited features of this hydrothermal area with photographs, temperature measurements, and selected geochemical data; many of these distinct habitats await characterization of their microbial communities and activities. Microprofiler deployments on microbial mats and hydrothermal sediments show their steep geochemical and thermal gradients at millimeter-scale vertical resolution. Mapping these hydrothermal features and sampling locations within the southern Guaymas Basin suggest linkages to underlying shallow sills and heat flow gradients. Recognizing the inherent spatial limitations of much current Guaymas Basin sampling calls for comprehensive surveys of the wider spreading region.
On page 2568 of this issue, Hatzenpichler and colleagues describe the application of BONCAT (bioorthogonal non-canonical amino acid tagging) (Hinz et al., 2013), a click-chemistry method originally developed to study protein synthesis and localization in neuronal cells, to pure-culture and environmental bacteria and archaea. Click chemistry and other bioorthogonal reactions have been intensively studied by chemists and some biologists for the past 15 years but have been little used as yet by environmental microbiologists. The authors show that click-chemistry amino acid analogues can be taken up by and detected in a range of pure-culture and environmental bacteria and archaea; that cells identified as translationally active by BONCAT are generally also metabolically active by the independent criterion of ammonia incorporation; and that production of proteins induced by an environmental change (heat shock) can be followed over time. BONCAT and other click-chemistry methods offer a promising route towards minimally invasive, cultivation-free investigations of the in situ enzymatic capabilities of microbes in diverse communities.
Detection of bacterial and archaeal cells, and isolation and analysis of their DNA and RNA, is challenging where populations are sparse and activities low. In “click” chemistry, so far applied mainly in eukaryotes, target molecules incorporate a label that can be specifically tagged for capture or microscopic detection under gentle reaction conditions. Ideally the labeling, tagging, and capture or detection efficiencies are high, and the label does not affect growth or metabolism. We have been investigating whether bacteria can incorporate the “clickable” RNA analog 5-ethynyl uridine (EU) for subsequent attachment of fluorescent label (Alexa Fluor) for microscopic detection or a chemical tag (desthiobiotin) for streptavidin-mediated RNA capture. The results for both aspects so far are a qualified “yes”. Both Escherichia coli and Bacillus subtilis grown with EU can subsequently be detected by fluorescence microscopy, but E. coli becomes considerably more brightly labeled. It also exhibits a slight but reproducible growth inhibition, and a distribution of fluorescence intensity across the population that becomes increasingly uneven over a one-day incubation. This might be due to normal physiological variation, or to the slower growth and hence slower label dilution rate of labeled cells. If the latter, the implication for natural samples is that slowly growing cells might actually become more strongly labeled. For the desthiobiotin labeling, both pure culture and environmental RNA has been captured, as shown on gels, but sequencing has been unsuccessful. We expect that this is a solvable methodological problem, and should not discourage others from exploring these methods.