The Inter-Valley Soil Comparative Survey: the ecology of Dry Valley edaphic microbial communities

Recent applications of molecular genetics to edaphic microbial communities of the McMurdo Dry Valleys and elsewhere have rejected a long-held belief that Antarctic soils contain extremely limited microbial diversity. The Inter-Valley Soil Comparative Survey aims to elucidate the factors shaping these unique microbial communities and their biogeography by integrating molecular genetic approaches with biogeochemical analyses. Although the microbial communities of Dry Valley soils may be complex, there is little doubt that the ecosystem''s food web is relatively simple, and evidence suggests that physicochemical conditions may have the dominant role in shaping microbial communities. To examine this hypothesis, bacterial communities from representative soil samples collected in four geographically disparate Dry Valleys were analyzed using molecular genetic tools, including pyrosequencing of 16S rRNA gene PCR amplicons. Results show that the four communities are structurally and phylogenetically distinct, and possess significantly different levels of diversity. Strikingly, only 2 of 214 phylotypes were found in all four valleys, challenging a widespread assumption that the microbiota of the Dry Valleys is composed of a few cosmopolitan species. Analysis of soil geochemical properties indicated that salt content, alongside altitude and Cu²⁺, was significantly correlated with differences in microbial communities. Our results indicate that the microbial ecology of Dry Valley soils is highly localized and that physicochemical factors potentially have major roles in shaping the microbiology of ice-free areas of Antarctica. These findings hint at links between Dry Valley glacial geomorphology and microbial ecology, and raise previously unrecognized issues related to environmental management of this unique ecosystem.     

Antarctic Dry Valley mineral soils contain unexpectedly high levels of microbial biomass

We have applied bioluminescent ATP detection methods to microbial enumeration in Antarctic Dry Valley mineral soils, and validated our ATP data by two independent methods. We have demonstrated that ATP measurement is a valid means of determining microbial biomass in such sites, and that the desiccated surface mineral soils of the Antarctic Dry Valleys contain cell numbers over four orders of magnitude higher than previously suggested.     

Distribution and abiotic influences on hypolithic microbial communities in an Antarctic Dry Valley

The Miers Valley within the McMurdo Dry Valleys of Antarctica supports abundant quartz and marble substrates for hypolithons—microbial colonists on the underside of these translucent rocks. Three physically distinct hypolithic community types have been identified: cyanobacteria dominated (Type I), fungus dominated (Type II) or moss dominated (Type III). The distribution of the three types was mapped across much of the ~75 km² area of the upper Miers Valley and correlated this with the measurements of selected micro-environmental variables. Type I hypolithons were most common and occurred at all altitudes up to 824 m, whilst Type II and Type III hypolithons were less abundant and restricted to lower altitudes on the valley floor (<415 m and <257 m, respectively). Whilst all colonized quartz effectively filtered incident UVB irradiance, transmittance levels for UVA and PAR varied markedly and were significant in determining hypolith type. Notably, the Type I hypolithons occurred under rocks with a significantly lower transmittance of photosynthetically active radiation than Type II and III hypolithons. Altitude and aspect were also significant factors determining hypolith type, and a role for altitude-related abiotic variables in determining the distribution of Type I, II and III hypolithons is proposed.     

A halotolerantPlanococcus from Antarctic Dry Valley soil

A halotolerantPlanococcus (strain A4a) was isolated from saline Antarctic Dry Valley soil.Planococcus strain A4a grew over wide ranges of temperature (0°C–40°C) and NaCl concentrations (0–2.0M). When the NaCl concentration of the growth medium was increased, the total intracellular free amino acid concentration increased; however, the intracellular potassium concentration did not increase. This result suggested that intracellular free amino acids functioned as compatible solutes for growth ofPlanococcus strain A4a at elevated NaCl concentrations. The halotolerant and psychrotolerant nature ofPlanococcus strain A4a would appear to provide it with the capacity for growth in the saline Antarctic Dry Valley soil environment from which it was isolated.     

Extraction of nematodes from Dry Valley Antarctic soils

Nematode density and taxonomic composition from Dry Valley soil processed by the sugar centrifugation (SC) method in Antarctica was compared to those extracted from soils shipped frozen to the USA and processed by either the SC or Baermann Funnel (BF) (at 5°C and 10°C) techniques. Soil selected for the extraction comparisons represented a wide range of soil properties found in the Dry Valleys. More nematodes were recovered from freshly collected Antarctic soil and from stored frozen soil using the SC technique than from BF at either temperature (P<0.05). Temperature had no effect on nematode densities extracted by the BF. Scottnema lindsayae was the most abundant species recovered by all extraction methods, but recovery was significantly lower from stored soils. Thus, nematodes can be extracted qualitatively following frozen storage using SC, but quantitative studies of nematode populations should be based on soils extracted following field sampling.     

Soil Carbon Dioxide Flux in Antarctic Dry Valley Ecosystems

The Antarctic dry valleys of southern Victoria Land are extreme desert environments where abiotic factors, such as temperature gradients, parent material, and soil water dynamics, may have a significant influence on soil carbon dioxide (CO₂) flux. Previous measurements of soil respiration have demonstrated very low rates of CO₂ efflux, barely above detection limits. We employed a modified infrared gas-analyzer system that enabled detection of smaller changes in CO₂ concentration in the field than previously possible. We measured diel CO₂ fluxes and monitored soil microclimate at three sites in Taylor Valley. Soil CO₂ flux ranged from –0.1 to 0.15 mol m^–2 s^–1. At two of the three sites, we detected a physically driven flux associated with diel variability in soil temperature. At these sites, CO₂ uptake (negative flux) was associated with dropping soil temperatures, whereas CO₂ evolution (positive flux) was associated with increases in soil temperature. These observations are corroborated by laboratory experiments that suggest that CO₂ flux is influenced by physically driven processes. We discuss four potential mechanisms that may contribute to physically driven gas exchange. Our results suggest there are strong interactions between biological and abiotic controls over soil CO₂ flux in terrestrial ecosystems of the Antarctic dry valleys, and that the magnitude of either may dominate depending on the soil environment and biological activity.     

Invertebrate Biodiversity in Antarctic Dry Valley Soils and Sediments

We studied invertebrate communities across a transition zone between soils and stream sediments in the cold desert landscape of Taylor Valley, Antarctica. We hypothesized that hydrological and biogeochemical linkages in the functionally important transition zone between streams and surrounding soils should be important in structuring invertebrate communities. We compared invertebrate communities along transects beginning in the saturated sediments under flowing stream water and extending laterally through the hyporheic zone to the dry soils that characterize most of the dry valley landscape. Nematodes, rotifers, and tardigrades assembled into different communities in soils and sediments, but there was no relationship between the total abundance of invertebrates and moisture. Community diversity was, however, influenced by the moisture and salinity gradients created with distance from flowing waters. The wet, low-salinity sediments in the center of the stream contained the most invertebrates and had the highest taxonomic diversity. Adjacent to the stream, communities in the hyporheic zone were influenced strongly by salt deposition. Abundance of invertebrates was low in the hyporheic zone, but this area contained the most co-occurring nematode species (three species). In dry soils, communities were composed almost entirely of a single species of nematode, Scottnema lindsayae, an organism not found in the stream center. These results suggest spatially-partitioned niches for invertebrates in soils and sediments in the dry valley landscape based on proximity to sources of moisture and the interactive effects of salinity. Received 22 September 1998; accepted 16 April 1999.     

Mechanism divergence in microbial rhodopsins

A fundamental design principle of microbial rhodopsins is that they share the same basic light-induced conversion between two conformers. Alternate access of the Schiff base to the outside and to the cytoplasm in the outwardly open “E” conformer and cytoplasmically open “C” conformer, respectively, combined with appropriate timing of pKa changes controlling Schiff base proton release and uptake make the proton path through the pumps vectorial. Phototaxis receptors in prokaryotes, sensory rhodopsins I and II, have evolved new chemical processes not found in their proton pump ancestors, to alter the consequences of the conformational change or modify the change itself. Like proton pumps, sensory rhodopsin II undergoes a photoinduced E → C transition, with the C conformer a transient intermediate in the photocycle. In contrast, one light-sensor (sensory rhodopsin I bound to its transducer HtrI) exists in the dark as the C conformer and undergoes a light-induced C → E transition, with the E conformer a transient photocycle intermediate. Current results indicate that algal phototaxis receptors channelrhodopsins undergo redirected Schiff base proton transfers and a modified E → C transition which, contrary to the proton pumps and other sensory rhodopsins, is not accompanied by the closure of the external half-channel. The article will review our current understanding of how the shared basic structure and chemistry of microbial rhodopsins have been modified during evolution to create diverse molecular functions: light-driven ion transport and photosensory signaling by protein–protein interaction and light-gated ion channel activity.     

Phylogenetic analysis of actinobacterial populations associated with Antarctic Dry Valley mineral soils

Despite the apparent severity of the environmental conditions in the McMurdo Dry Valleys, Eastern Antarctica, recent phylogenetic studies conducted on mineral soil samples have revealed the presence of a wide diversity of microorganisms, with actinobacteria representing one of the largest phylotypic groups. Previous metagenomic studies have shown that the majority of Antarctic actinobacterial populations are classified as 'uncultured'. In this study, we assessed the diversity of actinobacteria in Antarctic cold desert soils by complementing traditional culture-based techniques with a metagenomic study. Phylogenetic analysis of clones generated with actinobacterium- and streptomycete-specific PCR primers revealed that the majority of the phylotypes were most closely related to uncultured Pseudonocardia and Nocardioides species. Phylotypes most closely related to a number of rarer actinobacteria genera, including Geodermatophilus , Modestobacter and Sporichthya , were also identified. While complementary culture-dependent studies isolated a number of Nocardia and Pseudonocardia species, the majority of the cultured isolates (> 80%) were S treptomyces species – although phylotypes affiliated to the genus Streptomyces were detected at a low frequency in the metagenomic study. This study confirms that Antarctic Dry Valley desert soil harbours highly diverse actinobacterial communities and suggests that many of the phylotypes identified may represent novel, uncultured species.     

Controls on the distribution of productivity and organic resources in Antarctic Dry Valley soils

The Antarctic Dry Valleys are regarded as one of the harshest terrestrial habitats on Earth because of the extremely cold and dry conditions. Despite the extreme environment and scarcity of conspicuous primary producers, the soils contain organic carbon and heterotrophic micro-organisms and invertebrates. Potential sources of organic compounds to sustain soil organisms include in situ primary production by micro-organisms and mosses, spatial subsidies from lacustrine and marine-derived detritus, and temporal subsidies ('legacies') from ancient lake deposits. The contributions from these sources at different sites are likely to be influenced by local environmental conditions, especially soil moisture content, position in the landscape in relation to lake level oscillations and legacies from previous geomorphic processes. Here we review the abiotic factors that influence biological activity in Dry Valley soils and present a conceptual model that summarizes mechanisms leading to organic resources therein.     

Molecular and evolutionary aspects of microbial sensory rhodopsins

Retinal proteins (~ rhodopsins) are photochemically reactive membrane-embedded proteins, with seven transmembrane α-helices which bind the chromophore retinal (vitamin A aldehyde). They are widely distributed through all three biological kingdoms, eukarya, bacteria and archaea, indicating the biological significance of the retinal proteins. Light absorption by the retinal proteins triggers a photoisomerization of the chromophore, leading to the biological function, light-energy conversion or light-signal transduction. This article reviews molecular and evolutionary aspects of the light-signal transduction by microbial sensory receptors and their related proteins. This article is part of a Special Issue entitled: Retinal Proteins - You can teach an old dog new tricks.     

The role of protein-bound water molecules in microbial rhodopsins

Protein-bound internal water molecules are essential features of the structure and function of microbial rhodopsins. Besides structural stabilization, they act as proton conductors and even proton storage sites. Currently, the most understood model system exhibiting such features is bacteriorhodopsin (bR). During the last 20 years, the importance of water molecules for proton transport has been revealed through this protein. It has been shown that water molecules are as essential as amino acids for proton transport and biological function. In this review, we present an overview of the historical development of this research on bR. We furthermore summarize the recently discovered protein-bound water features associated with proton transport. Specifically, we discuss a pentameric water/amino acid arrangement close to the protonated Schiff base as central proton-binding site, a protonated water cluster as proton storage site at the proton-release site, and a transient linear water chain at the proton uptake site. We highlight how protein conformational changes reposition or reorient internal water molecules, thereby guiding proton transport. Last, we compare the water positions in bR with those in other microbial rhodopsins to elucidate how protein-bound water molecules guide the function of microbial rhodopsins. This article is part of a Special Issue entitled: Retinal Proteins — You can teach an old dog new tricks.     

Life cycle of the microbivorous Antarctic Dry Valley nematode Scottnema lindsayae (Timm 1971)

The life cycle of the Antarctic Dry Valley soil nematode, Scottnema lindsayae (Timm 1971) was studied in laboratory culture at two temperatures, 10°C and 15°C. Soil yeast and bacteria isolated with the nematodes were used as the food source. The species reproduced sexually. The higher temperature had a negative effect on the life cycle. The number of eggs per female and the number of juveniles developing per female were greater at 10°C than at 15°C. Juveniles developed faster at 10°C and four juvenile stages were observed outside of the egg at both temperatures. The unusually long life cycle (218 d at 10°C) suggests that more than one austral summer may be required for successful completion. An increase in Dry Valley soil temperatures associated with potential global environmental change may have detrimental effects on soil nematodes.     

Spectral Properties of the Green Alga Trebouxia,a Phycobiont of Cryptoendolithic Lichens in the Antarctic Dry Valley

An algologically pure culture of the green alga Trebouxia, a phycobiont of cryptoendolithic lichens, was isolated from sandstone samples collected in the high-altitude polar regions of Antarctica. The absorption and second-derivative absorption spectra of acetone extract of the Antarctic phycobiont cells were studied in comparison with those of a Trebouxia phycobiont isolated recently from a Parmeliaceae lichen in the Mid-European climatic zone. The cells of the Antarctic phycobiont were characterized by a lower content of chlorophyll a and a higher ratio of chlorophyll b and carotenoids to chlorophyll a as compared to the Mid-European phycobiont. Furthermore, the carotenoids of the Antarctic phycobiont were more diverse. The low-temperature fluorescence spectra of the Antarctic phycobiont were characterized by an increased intensity of the short-wavelength fluorescence peak of chlorophyll aand a diminished intensity of fluorescence in the long-wavelength spectral region.     

Hypolithic microbial communities of quartz rocks from Miers Valley, McMurdo Dry Valleys, Antarctica

The McMurdo Dry Valleys region of eastern Antarctica is a cold desert that presents extreme challenges to life. Hypolithic microbial colonisation of the subsoil surfaces of translucent quartz rocks represent a significant source of terrestrial biomass and productivity in this region. Previous studies have described hypoliths as dominated by cyanobacteria. However, hypoliths that occur in the lower Dry Valleys such as the Miers, Garwood and Marshall Valleys are unusual as they are not necessarily cyanobacteria-dominated. These hypoliths support significant eukaryal colonisation by fungi and mosses in addition to cyanobacteria-dominated bacterial assemblages and so have considerable ecological value in this barren landscape. Here, we characterise these novel hypoliths by analysis of environmental rRNA gene sequences. The hypolithic community was demonstrated to be distinct from the surrounding soil and non-translucent rocks. Hypoliths supported cyanobacterial signatures from the Oscillatoriales and Nostocales. Other heterotrophic bacterial signatures were also recovered, and these were phylogenetically diverse and spanned 8 other bacterial phyla. Archaeal phylotypes recovered were phylogenetically affiliated with the large group of unclassified, uncultured Crenarcheota. Eukaryal phylotypes indicated that free-living ascomycetous fungi, chlorophytes and mosses (Bryum sp.) were all supported by these hypoliths, and these are thought to be responsible for the extensive eukaryotic biomass that develops around quartz rocks.     

Chimeric microbial rhodopsins containing the third cytoplasmic loop of bovine rhodopsin

G-protein-coupled receptors transmit stimuli (light, taste, hormone, neurotransmitter, etc.) to the intracellular signaling systems, and rhodopsin (Rh) is the most-studied G-protein-coupled receptor. Rh possesses an 11-cis retinal as the chromophore, and 11-cis to all-trans photoisomerization leads to the protein structural changes in the cytoplasmic loops to activate G-protein. Microbial rhodopsins are similar heptahelical membrane proteins that function as bacterial sensors, light-driven ion-pumps, or light-gated channels. Microbial rhodopsins possess an all-trans retinal, and all-trans to 13-cis photoisomerization triggers protein structural changes for each function. Despite these similarities, there is no sequence homology between visual and microbial rhodopsins, and microbial rhodopsins do not activate G-proteins. However, it was reported that bacteriorhodopsin (BR) chimeras containing the third cytoplasmic loop of bovine Rh are able to activate G-protein, suggesting a common mechanism of protein structural changes. Here we design chimeric proteins for Natronomonas pharaonis sensory rhodopsin II (SRII, also called pharaonis phoborhodopsin), which has a two-orders-of-magnitude slower photocycle than BR. Light-dependent transducin activation was observed for most of the nine SRII chimeras containing the third cytoplasmic loop of bovine Rh (from Y223, G224, Q225 to T251, R252, and M253), but the activation level was 30,000–140,000 times lower than that of bovine Rh. The BR chimera, BR/Rh223-253, activates a G-protein transducin, whereas the activation level was 37,000 times lower than that of bovine Rh. We interpret the low activation by the chimeric proteins as reasonable, because bovine Rh must have been optimized for activating a G-protein transducin during its evolution. On the other hand, similar activation level of the SRII and BR chimeras suggests that the lifetime of the M intermediates is not the simple determinant of activation, because SRII chimeras have two-orders-of-magnitude's slower photocycle than the BR chimera. Activation mechanism of visual and microbial rhodopsins is discussed on the basis of these results.     

Genome sequence of temperate bacteriophage Psymv2 from Antarctic Dry Valley soil isolate Psychrobacter sp. MV2

A temperate phage, Psymv2, was isolated from an Antarctic soil bacterium, Psychrobacter sp. MV2. The morphology of Psymv2 was typical of the Siphoviridae, with an isometric head and non-contractile tail. The Psymv2 genome was found to be 35,725?bp in length, had a G?+?C content of 44.5?%, with 49 protein-coding genes and one tRNA gene predicted. Integration of Psymv2 occurred at an ssrA gene, with the last 27 bases of this gene directly repeated at the prophage ends. The genome was organised in a modular fashion: integration, regulation, packaging, head assembly, tail assembly, host specificity and lysis. While the genome sequence had little similarity on a nucleotide level to previously reported phage sequences, the genome architecture resembled that of Siphoviridae of low G?+?C Gram-positive bacteria. The closest relatives to Psymv2 were uncharacterized putative prophages within the P. arcticus 273-4 and Acinetobacter baumannii 6013113 genomes. Global alignment of the Psymv2 genome and these prophages revealed significant conservation of the structural modules despite the large spatial divergence of their hosts. A number of unique ORFs were identified in the Psymv2 genome that may contribute to phage and lysogen fitness.     

Enhancement of the long-wavelength sensitivity of optogenetic microbial rhodopsins by 3,4-dehydroretinal

Electrogenic microbial rhodopsins (ion pumps and channelrhodopsins) are widely used to control the activity of neurons and other cells by light (optogenetics). Long-wavelength absorption by optogenetic tools is desirable for increasing the penetration depth of the stimulus light by minimizing tissue scattering and absorption by hemoglobin. A2 retinal (3,4-dehydroretinal) is a natural retinoid that serves as the chromophore in red-shifted visual pigments of several lower aquatic animals. Here we show that A2 retinal reconstitutes a fully functional archaerhodopsin-3 (AR-3) proton pump and four channelrhodopsin variants (CrChR1, CrChR2, CaChR1, and MvChR1). Substitution of A1 with A2 retinal significantly shifted the spectral sensitivity of all tested rhodopsins to longer wavelengths without altering other aspects of their function. The spectral shift upon substitution of A1 with A2 in AR-3 was close to that measured in other archaeal rhodopsins. Notably, the shifts in channelrhodopsins were larger than those measured in archaeal rhodopsins and close to those in animal visual pigments with similar absorption maxima of their A1-bound forms. Our results show that chromophore substitution provides a complementary strategy for improving the efficiency of optogenetic tools.     

Light-induced intramolecular charge movements in microbial rhodopsins in intact E. coli cells.

Microbial rhodopsins undergo cyclic photochemical reactions (photocycles) in which proton transfers and conformational changes result in charge displacements during transitions between photocycle intermediates. We report a new photoelectric method to monitor charge movements during rhodopsin photocycling with fast kinetic resolution in suspensions of intact E. coli cells. The method monitors electrical currents resulting from asymmetric photoexcitation of microbial rhodopsins by a unilateral laser flash, and kinetically resolves intramolecular charge movements. We investigated E. coli-expressed proton-transporting rhodopsins, specifically green- and blue-absorbing proteorhodopsins (GPR and BPR, respectively) from uncultivated marine plankton, and sensory rhodopsins, namely receptors from Natronomonas pharaonis and Anabaena (Nostoc) sp. PCC7120. Kinetic components of the currents correlate with photochemical transformations of the pigments, and the integrated current measures net transport by the proton-pumping rhodopsins. The photoelectric measurements distinguish between known light-driven transporters and photosensors, and reveal differences in proton transfer reactions in the two tested proton pumps. Screening of nine newly identified proteorhodopsins reveals two with GPR-type charge movements, five with BPR-type, and two with the characteristics of the sensory rhodopsins. The approach developed in the present work provides a direct, rapid and informative method for studying electrogenic events in rhodopsin photocycles and also gives a clue to functions of newly found microbial rhodopsins in nature.