Sulfate-reducing prokaryotic communities in two deep hypersaline anoxic basins in the Eastern Mediterranean deep sea

In the Eastern Mediterranean Sea, deep hypersaline anoxic basins (DHABs) and deep-sea sediment contain anoxic environments where sulfate reduction is an important microbial metabolic process. The objective of this study was to characterize the sulfate-reducing community in the brine and interface of the DHABs L'Atalante and Urania based on a phylogenetic analysis of the dissimilatory sulfite reductase gene (dsrA). Results demonstrated that the sulfate-reducing community was diverse, except for the sulfidogenic brine of the Urania basin. The similarity of the dsrA sequences between different environments was very low demonstrating that each environment had a unique sulfate-reducing community. Sequences had 67.6-93.3% similarity to dsrA sequences from GenBank database and were mostly related to the delta-proteobacteria. Each environment was dominated by a different family within the delta-proteobacteria except for the Urania interface, which was dominated by sequences related to the Gram-positive Peptococcaceae. We conclude that sulfate-reducing communities inhabiting the L'Atalante and Urania basins are highly diverse with low similarities to each other and contain a sulfate-reducing species composition that is very different from sulfate-reducing species compositions in previously studied ecosystems.     

Current state of athalassohaline deep-sea hypersaline anoxic basin research—recommendations for future work and relevance to astrobiology

Deep-sea hypersaline anoxic basins (DHABs) are uniquely stratified polyextreme environments generally found in enclosed seas. These environments select for elusive and widely uncharacterized microbes that may be living below the currently recognized window of life on Earth. Still, there is strong evidence of highly specialized active microbial communities in the Kryos, Discovery, and Hephaestus basins located in the Eastern Mediterranean Sea; the only known athalassohaline DHABs. Life is further constrained in these DHABs as near-saturated concentrations of magnesium chloride significantly reduces water activity (a_w) and exerts extreme chaotropic stress, the tendency of a solution to disorder biomolecules. In this review, we provide an overview of microbial adaptations to polyextremes focusing primarily on chaotropicity, summarize current evidence of microbial life within athalassohaline DHABs and describe the difficulties of life detection approaches and sampling within these environments. We also reveal inconsistent measurements of chaotropic activity in the literature highlighting the need for a new methodology. Finally, we generate recommendations for future investigations and discuss the importance of athalassohaline DHAB research to help inform extraterrestrial life detection missions.     

Spatial and Seasonal Variation in a Reservoir Sedimentary Microbial Community as Determined by Phospholipid Analysis

Sediment samples were collected monthly from Acton Lake, a eutrophic reservoir located in an agricultural region of southwestern Ohio, from three stations (River, Middle, and Dam) during the period May 1995 through January 1997. Sedimentary microbial biomass and community structures from these stations were studied using phospholipid analysis. At the River and Middle stations, the water column remained aerobic throughout the year, whereas the water overlying the Dam station sediments became anaerobic during summer stratification. Sedimentary microbial biomass at the River and Middle stations, as measured by the phospholipid phosphate (PLP) method, ranged from 225 to 450 nmol PLP g?1 d.w. (dry weight). Sedimentary microbial biomass at the Dam station was typically greater and ranged from 500 to 1,500 nmol PLP g?1 d.w. Principal component analysis of phospholipid fatty acid (PLFA) profiles indicated that the sedimentary microbial communities at all three stations displayed seasonal patterns of change. Among these patterns of change was a shift from aerobic microorganisms during times of cold water to anaerobic microorganisms during times of warm water. The Dam station differed from the River and Middle stations in that sediments from this station had disproportionately more polyenoic fatty acids, whereas sediments from the River and Middle stations had disproportionately more bacterial fatty acids. These data suggest that the Dam station may be a depositional zone for microeukaryotic phytoplankton produced in the overlying water column. These findings have implications for the understanding of carbon flux in reservoirs and preservation of organic matter in aquatic systems.     

Variation in Microbial Biomass and Community Structure in Sediments of Eutrophic Bays as Determined by Phospholipid Ester-Linked Fatty Acids

The distribution of phospholipid ester-linked fatty acids (PLFA) in sediments of eutrophic bays (Hiroshima Bay and Aki Nada) was studied to quantify the microbial biomass, community structure, and nutritional status. A total of 63 fatty acids in the range of C₁₀ to C₂₄ were determined. They consist of saturated fatty acids, branched fatty acids, monounsaturated fatty acids, and polyunsaturated fatty acids, and variation was revealed in the relative proportions of these fatty acids in sediments. On the basis of the PLFA concentration in sediments, the calculated microbial biomass showed variation (mean ± standard deviation = 0.70 × 10⁸ ± 0.53 × 10⁸ cells per g [dry weight] of sediment) in the eutrophic bays. In sediments, a higher amount of biomass was observed in the coastal area of Hiroshima Bay than that observed in the rest of the bay and adjacent Aki Nada. The microbial community structure of the present study area, as characterized by the PLFA profiles, showed very low percentages of polyunsaturated fatty acids and long-chain fatty acids characteristic of microeukary-otes and terrestrial input, respectively, and high percentages of fatty acids characteristic of bacteria. The distribution of PLFA profiles also showed the relative contribution of both aerobic and anaerobic bacteria, especially sulfate-reducing bacteria, in the study area. The relative proportions of PLFA revealed distinctive differences among the stations of the study area, as is evidenced from six clusters obtained for the PLFA profiles. The results of Tukey's honestly significant difference test further confirmed that the sediments in the coastal area of Hiroshima Bay were significantly enriched by a number of fatty acids when compared with other areas investigated where relatively few fatty acids were present in significant quantities. No marked variation in environmental parameters in the surface- and bottom-water samples was observed, indicating the absence of any water movement in the study area. Furthermore, low redox potential and the levels of sulfide in the sediment revealed the reduced condition of the sediment. The existing environmental conditions and pollution of the study area were attributed to the observed microbial community structure in the sediments.     

Metazoans of redoxcline sediments in Mediterranean deep-sea hypersaline anoxic basins

Background

The deep-sea hypersaline anoxic basins (DHABs) of the Mediterranean (water depth ~3500 m) are some of the most extreme oceanic habitats known. Brines of DHABs are nearly saturated with salt, leading many to suspect they are uninhabitable for eukaryotes. While diverse bacterial and protistan communities are reported from some DHAB haloclines and brines, loriciferans are the only metazoan reported to inhabit the anoxic DHAB brines. Our goal was to further investigate metazoan communities in DHAB haloclines and brines.

Results

We report observations from sediments of three DHAB (Urania, Discovery, L’Atalante) haloclines, comparing these to observations from sediments underlying normoxic waters of typical Mediterranean salinity. Due to technical difficulties, sampling of the brines was not possible. Morphotype analysis indicates nematodes are the most abundant taxon; crustaceans, loriciferans and bryozoans were also noted. Among nematodes, Daptonema was the most abundant genus; three morphotypes were noted with a degree of endemicity. The majority of rRNA sequences were from planktonic taxa, suggesting that at least some individual metazoans were preserved and inactive. Nematode abundance data, in some cases determined from direct counts of sediments incubated in situ with CellTracker^TM Green, was patchy but generally indicates the highest abundances in either normoxic control samples or in upper halocline samples; nematodes were absent or very rare in lower halocline samples. Ultrastructural analysis indicates the nematodes in L’Atalante normoxic control sediments were fit, while specimens from L’Atalante upper halocline were healthy or had only recently died and those from the lower halocline had no identifiable organelles. Loriciferans, which were only rarely encountered, were found in both normoxic control samples as well as in Discovery and L’Atalante haloclines. It is not clear how a metazoan taxon could remain viable under this wide range of conditions.

Conclusions

We document a community of living nematodes in normoxic, normal saline deep-sea Mediterranean sediments and in the upper halocline portions of the DHABs. Occurrences of nematodes in mid-halocline and lower halocline samples did not provide compelling evidence of a living community in those zones. The possibility of a viable metazoan community in brines of DHABs is not supported by our data at this time.

     

Sediment Microbial Community Structure and Mercury Methylation in Mercury-Polluted Clear Lake, California

Spatial and temporal variations in sediment microbial community structure in a eutrophic lake polluted with inorganic mercury were identified using polar lipid fatty acid (PLFA) analysis. Microbial community structure was strongly related to mercury methylation potential, sediment organic carbon content, and lake location. Pore water sulfate, total mercury concentrations, and organic matter C/N ratios showed no relationships with microbial community structure. Seasonal changes and changes potentially attributable to temperature regulation of bacterial membranes were detectable but were less important influences on sediment PLFA composition than were differences due to lake sampling location. Analysis of biomarker PLFAs characteristic of Desulfobacter and Desulfovibrio groups of sulfate-reducing bacteria suggests that Desulfobacter-like organisms are important mercury methylators in the sediments, especially in the Lower Arm of Clear Lake.     

Sediment microbial community structure and mercury methylation in mercury-polluted Clear Lake, California

Spatial and temporal variations in sediment microbial community structure in a eutrophic lake polluted with inorganic mercury were identified using polar lipid fatty acid (PLFA) analysis. Microbial community structure was strongly related to mercury methylation potential, sediment organic carbon content, and lake location. Pore water sulfate, total mercury concentrations, and organic matter C/N ratios showed no relationships with microbial community structure. Seasonal changes and changes potentially attributable to temperature regulation of bacterial membranes were detectable but were less important influences on sediment PLFA composition than were differences due to lake sampling location. Analysis of biomarker PLFAs characteristic of Desulfobacter and Desulfovibrio groups of sulfate-reducing bacteria suggests that Desulfobacter-like organisms are important mercury methylators in the sediments, especially in the Lower Arm of Clear Lake.     

Microbial Diversity in Water and Sediment of Lake Chaka, an Athalassohaline Lake in Northwestern China

We employed culture-dependent and -independent techniques to study microbial diversity in Lake Chaka, a unique hypersaline lake (32.5% salinity) in northwest China. It is situated at 3,214 m above sea level in a dry climate. The average water depth is 2 to 3 cm. Halophilic isolates were obtained from the lake water, and halotolerant isolates were obtained from the shallow sediment. The isolates exhibited resistance to UV and gamma radiation. Microbial abundance in the sediments ranged from 10⁸ cells/g at the water-sediment interface to 10⁷ cells/g at a sediment depth of 42 cm. A major change in the bacterial community composition was observed across the interface. In the lake water, clone sequences affiliated with the Bacteroidetes were the most abundant, whereas in the sediments, sequences related to low G+C gram-positive bacteria were predominant. A similar change was also present in the archaeal community. While all archaeal clone sequences in the lake water belonged to the Halobacteriales, the majority of the sequences in the sediments were related to those previously obtained from methanogenic soils and sediments. The observed changes in the microbial community structure across the water-sediment interface were correlated with a decrease in salinity from the lake water (32.5%) to the sediments (approximately 4%). Across the interface, the redox state also changed from oxic to anoxic and may also have contributed to the observed shift in the microbial community.     

Characterization of Microbial Community Structure in the Surface Sediment of Osaka Bay, Japan, by Phospholipid Fatty Acid Analysis

Twenty-eight sediment samples collected from Osaka Bay, Japan, were analyzed for phospholipid ester-linked fatty acids (PLFA) to determine regional differences in microbial community structure of the bay. The abundance of three major groups of C₁₀ to C₁₉ PLFA (saturated, branched, and monounsaturated PLFA), which accounted for 84 to 97% of the total PLFA, indicated the predominance of prokaryotes in the sediment. The distribution of six clusters obtained by similarity analysis in the bay revealed a marked regional distribution in the PLFA profiles. Total PLFA concentrations (0.56 to 2.97 μg/g [dry weight] of the sediment) in sediments also showed marked variation among the stations, with higher concentrations of total PLFA in the central part of the bay. The biomass, calculated on the basis of total PLFA concentration, ranged from 0.25 × 10⁸ to 1.35 × 10⁸ cells per g (dry weight) of the sediment. The relative dominance of microbial groups in sediments was described by using the reported bacterial biomarker fatty acids. Very small amounts of the characteristic PLFA of microeukaryotes in sediments indicated the restricted distribution of microeukaryotes. By examining the distribution of clusters and groups of microorganisms in the bay, there were two characteristics of the distribution pattern: (i) the predominance of anaerobic bacteria and gram-positive prokaryotes, characterized by the high proportions of branched PLFA in the eastern and northeastern sides of the bay, where the reported concentrations of pollutants were also high, and (ii) the predominance of aerobic prokaryotes and eukaryotes, except for a few stations, in the western and southwestern sides of the bay, as evidenced by the large amounts of monounsaturated PLFA. Such significant regional differences in microbial community structure of the bay indicate shifts in microbial community structure.     

Microbial diversity in water and sediment of Lake Chaka, an athalassohaline lake in northwestern China

We employed culture-dependent and -independent techniques to study microbial diversity in Lake Chaka, a unique hypersaline lake (32.5% salinity) in northwest China. It is situated at 3,214 m above sea level in a dry climate. The average water depth is 2 to 3 cm. Halophilic isolates were obtained from the lake water, and halotolerant isolates were obtained from the shallow sediment. The isolates exhibited resistance to UV and gamma radiation. Microbial abundance in the sediments ranged from 10(8) cells/g at the water-sediment interface to 10(7) cells/g at a sediment depth of 42 cm. A major change in the bacterial community composition was observed across the interface. In the lake water, clone sequences affiliated with the Bacteroidetes were the most abundant, whereas in the sediments, sequences related to low G+C gram-positive bacteria were predominant. A similar change was also present in the archaeal community. While all archaeal clone sequences in the lake water belonged to the Halobacteriales, the majority of the sequences in the sediments were related to those previously obtained from methanogenic soils and sediments. The observed changes in the microbial community structure across the water-sediment interface were correlated with a decrease in salinity from the lake water (32.5%) to the sediments (approximately 4%). Across the interface, the redox state also changed from oxic to anoxic and may also have contributed to the observed shift in the microbial community.     

Microbial community structure and ecology of subglacial sediments in two polythermal Svalbard glaciers characterized by epifluorescence microscopy and PLFA

Biological and physico-chemical characteristics of subglacial sediments were studied in Svalbard. Sediment from close proglacial and supraglacial environments was used for a comparison. Viable bacteria, cyanobacteria and microalgae were detected in subglacial sediments from two polythermal glaciers using epifluorescence microscopy and phospholipid fatty acid (PLFA) analyses. The subglacial samples were generally of higher pH values, coarser texture and lower water content, organic matter, organic carbon, and nitrogen compared to proglacial and supraglacial sediments). Bacterial counts of 1.6 × 10⁷ cells mg^{− 1} OM (organic matter) were found. Cyanobacteria and algae were also of low abundance [4.2 cells mg^{− 1} DW (dry weight)]. Cyanobacteria comprised the major proportion of the autophotothrophic assemblages of subglacial soils. Deglaciated soils were similar to subglacial sediment in physico-chemical properties and microbial structure and numbers, unlike soil from vegetated sites or cryoconite sediment. In subglacial and deglaciated soil, relatively low diversity of microorganisms and low substrate availability was detected by PLFA analyses. Good accordance in microbial community structure assessments between epifluorescence microscopy and PLFA analyses was found. Our results suggest that the subglacial microbial populations can be divided into two groups: autochthonous microorganims (chemoheterotrophic bacteria) and allochthonous that retain the ability to proliferate and give rise to active population when conditions become favorable. Electronic Supplementary Material Supplementary material is available to authorised users in the online version of this article at .     

Microbial Community of a Hydrothermal Mud Vent Underneath the Deep-Sea Anoxic Brine Lake Urania (Eastern Mediterranean)

The composition of a metabolically active prokaryotic community thriving in hydrothermal mud fluids of the deep-sea hypersaline anoxic Western Urania Basin was characterized using rRNA-based phylogenetic analysis of a clone library. The physiologically active prokaryotic assemblage in this extreme environment showed a great genetic diversity. Most members of the microbial community appeared to be affiliated to yet uncultured organisms from similar ecosystems, i.e., deep-sea hypersaline basins and hydrothermal vents. The bacterial clone library was dominated by phylotypes affiliated with the epsilon-Proteobacteria subdivision recognized as an ecologically significant group of bacteria inhabiting deep-sea hydrothermal environments. Almost 18% of all bacterial clones were related to delta-Proteobacteria, suggesting that sulfate reduction is one of the dominant metabolic processes occurring in warm mud fluids. The remaining bacterial phylotypes were related to alpha- and beta-Proteobacteria, Actinobacteria, Bacteroides, Deinococcus-Thermus, KB1 and OP-11 candidate divisions. Moreover, a novel monophyletic clade, deeply branched with unaffiliated 16S rDNA clones was also retrieved from deep-sea sediments and halocline of Urania Basin. Archaeal diversity was much lower and detected phylotypes included organisms affiliated exclusively with the Euryarchaeota. More than 96% of the archaeal clones belonged to the MSBL-1 candidate order recently found in hypersaline anoxic environments, such as endoevaporitic microbial mats, Mediterranean deep-sea mud volcanoes and anoxic basins. Two phylotypes, represented by single clones were related to uncultured groups DHVE-1 and ANME-1. Thus, the hydrothermal mud of hypersaline Urania Basin seems to contain new microbial diversity. The prokaryotic community was significantly different from that occurring in the upper layers of the Urania Basin since 60% of all bacterial and 40% of all archaeal phylotypes were obtained only from mud fluids. The uniqueness of the composition of the active prokaryotic community could be explained by the complex environmental conditions at the site. The interaction of oxygenated warm mud fluids with the cold hypersaline brine of the Urania Basin seems to simultaneously select for various metabolic processes, such as aerobic and anaerobic heterotrophy, sulfide- and methane-dependent chemotrophy along with anaerobic oxidation of methane, sulfate- and metal-reduction.     

Methanogenesis in the hypersaline Solar Lake (Sinai)

Abstract Enrichment studies on microbial mat sediments (potential stromatolites) from the hypersaline Solar Lake (Sinai) indicated high numbers of methanogenic bacteria (up to 10⁵ ml⁻¹ sediment) in spite of the high sulfate reduction rate, sulfate concentration and salinity. Among H₂/CO₂, acetate and monomethylamine, the methylated amine was the preferred substrate. The predominant species enriched was a Methanosarcina sp. The findings indicate that methanogenic bacteria play an important role in hypersaline sulfate-enriched anoxic sediments and stromatolithic microbial mats.     

Microbial Diversity in Sediments of Saline Qinghai Lake, China: Linking Geochemical Controls to Microbial Ecology

Saline lakes at high altitudes represent an important and extreme microbial ecosystem, yet little is known about microbial diversity in such environments. The objective of this study was to examine the change of microbial diversity from the bottom of the lake to sediments of 40 cm in depth in a core from Qinghai Lake. The lake is saline (12.5 g/L salinity) and alkaline (pH 9.4) and is located on the Qinghai–Tibetan Plateau at an altitude of 3196 m above sea level. Pore water chemistry of the core revealed low concentrations of sulfate and iron (<1 mM), but high concentrations of acetate (40–70 mM) and dissolved organic carbon (1596–5443 mg/L). Total organic carbon and total nitrogen contents in the sediments were ∼2 and <0.5%, respectively. Acridine orange direct count data indicated that cell numbers decreased from 4 × 10⁹ cells/g at the water–sediment interface to 6× 10⁷ cells/g wet sediment at the 40-cm depth. This change in biomass was positively correlated with acetate concentration in pore water. Phospholipid fatty acid (PLFA) community structure analyses determined decrease in the proportion of the Proteobacteria and increase in the Firmicutes with increased depth. Characterization of small subunit (SSU) rRNA genes amplified from the sediments indicated a shift in the bacterial community with depth. Whereas the α-, β-, and γ-Proteobacteria and the Cytophaga/Flavobacterium/Bacteroides (CFB) were dominant at the water–sediment interface, low G + C gram-positive bacteria (a subgroup of Firmicutes) became the predominant group in the anoxic sediments. Both PLFA and the sequence data showed similar trend. The Proteobacteria, CFB, and gram-positive bacteria are present in other saline lakes, but thepresence of Actinobacteria and Acidobacteria/Holophaga in significant proportions in the Qinghai Lake sediments appears to be unique. The archaeal diversity was much lower, and clone sequences could be grouped inthe Euryarchaeota and Crenarchaeota domains. The archaeal clones were not related to any known cultures but to sequences previously found in methane-rich sediments. Acetate-utilizing methanogens were isolated from sediment incubations, and α- and γ-proteobacterial isolates were obtained from a water sample from the lakebottom (23 m). Our data collectively showed that the observed diversity and shift in the community structure with depth was correlated with geochemical parameters (the redox state and availability of electron acceptor and donor). Heterotrophic methanogenesis is possibly adominant metabolic process in the Qinghai Lake sediments. These results reinforce the importance of geochemical controls on microbial ecology in saline and alkaline lake environments.     

Microbial community composition and function in permanently cold seawater and sediments from an arctic fjord of svalbard

Heterotrophic microbial communities in seawater and sediments metabolize much of the organic carbon produced in the ocean. Although carbon cycling and preservation depend critically on the capabilities of these microbial communities, their compositions and capabilities have seldom been examined simultaneously at the same site. To compare the abilities of seawater and sedimentary microbial communities to initiate organic matter degradation, we measured the extracellular enzymatic hydrolysis rates of 10 substrates (polysaccharides and algal extracts) in surface seawater and bottom water as well as in surface and anoxic sediments of an Arctic fjord. Patterns of enzyme activities differed between seawater and sediments, not just quantitatively, in accordance with higher cell numbers in sediments, but also in their more diversified enzyme spectrum. Sedimentary microbial communities hydrolyzed all of the fluorescently labeled polysaccharide and algal extracts, in most cases at higher rates in subsurface than surface sediments. In seawater, in contrast, only 5 of the 7 polysaccharides and 2 of the 3 algal extracts were hydrolyzed, and hydrolysis rates in surface and deepwater were virtually identical. To compare bacterial communities, 16S rRNA gene clone libraries were constructed from the same seawater and sediment samples; they diverged strongly in composition. Thus, the broader enzymatic capabilities of the sedimentary microbial communities may result from the compositional differences between seawater and sedimentary microbial communities, rather than from gene expression differences among compositionally similar communities. The greater number of phylum- and subphylum-level lineages and operational taxonomic units in sediments than in seawater samples may reflect the necessity of a wider range of enzymatic capabilities and strategies to access organic matter that has already been degraded during passage through the water column. When transformations of marine organic matter are considered, differences in community composition and their different abilities to access organic matter should be taken into account.     

Variability in carbon uptake and (re)cycling in Antarctic cryptoendolithic microbial ecosystems demonstrated through radiocarbon analysis of organic biomarkers

Cryptoendolithic lichens and cyanobacteria living in porous sandstone in the high‐elevation McMurdo Dry Valleys are purported to be among the slowest growing organisms on Earth with cycles of death and regrowth on the order of 10³–10⁴ years. Here, organic biomarker and radiocarbon analysis were used to better constrain ages and carbon sources of cryptoendoliths in University Valley (UV; 1,800 m.a.s.l) and neighboring Farnell Valley (FV; 1,700 m.a.s.l). Δ¹⁴C was measured for membrane component phospholipid fatty acids (PLFA) and glycolipid fatty acids, as well as for total organic carbon (TOC). PLFA concentrations indicated viable cells comprised a minor (<0.5%) component of TOC. TOC Δ¹⁴C values ranged from ?272‰ to ?185‰ equivalent to calibrated ages of 1,100–2,550 years old. These ages may be the result of fractional preservation of biogenic carbon and/or sudden large‐scale community death and extended period(s) of inactivity prior to slow recolonization and incorporation of ¹⁴C‐depleted fossil material. PLFA Δ¹⁴C values were generally more modern than the corresponding TOC and varied widely between sites; the FV PLFA Δ¹⁴C value (+40‰) was consistent with modern atmospheric CO₂, while UV values ranged from ?199‰ to ?79‰ (calibrated ages of 1,665–610 years). The observed variability in PLFA Δ¹⁴C depletions is hypothesized to reflect variations in the extent of fixation of modern atmospheric CO₂ and the preservation and recycling of older organic carbon by the community in various stages of sandstone recolonization. PLFA profiles and microbial community compositions as determined by molecular genetic characterizations and microscopy differed between the two valleys (e.g., predominance of biomarker 18:2 [>50%] in FV compared to UV), representing microbial communities that may reflect distinct stages of sandstone recolonization and/or environmental conditions. It is thus proposed that Dry Valley cryptoendolithic microbial communities are faster growing than previously estimated.     

Effects of salinity on microbial tolerance to drying and rewetting

Soil salinity and fluctuations in soil matric potential are stressors for soil microorganisms which, in turn, may affect soil organic matter turnover. In response to salinity and low soil water content, many microorganisms accumulate osmolytes. Therefore, it is conceivable that microorganisms in saline soils are more tolerant to drying and rewetting (DRW) stress than those in non-saline soils. An experiment was carried out with three different salinity levels: electrical conductivity (EC_1:5) 0, 2 and 4 dS m^?1 (EC0, EC2, EC4), and two water treatments: a constantly moist control or two DRW cycles. Respiration as an indicator of microbial activity was measured throughout the 59 days of incubation. At the end of the second dry period (day 35) and at the end of the following moist incubation (day 59), microbial biomass and microbial community structure were determined by phospholipid fatty acid (PLFA) analysis. Increasing salinity decreased microbial activity but did not affect its resistance to DRW. On day 59, cumulative respiration decreased in the order EC0 > EC2 > EC4 with no differences between water treatments. Fungal biomass was negatively affected by salinity at the end of the experiment, while bacterial biomass was unaffected. Microbial community structure in moist treatments differed between salinity levels, with EC4 influencing microbial community structure earlier than EC2. The resistance of microbial communities to DRW stress was salt level dependent; only beyond a critical salinity level adaptation to salt stress was able to reduce the impact of water stress on microbial community structure.     

Effect of stand age on soil microbial community structure in wolfberry (Lycium barbarum L.) fields

Soil physicochemical properties and microbes are essential in terrestrial ecosystems through their role in cycling mineral compounds and decomposing organic matter. This study examined the effect of stand age on soil physicochemical properties and microbial community structure in wolfberry (Lycium barbarum L.) fields, in order to reveal the mechanism of soil degradation due to long-term stand of L. barbarum. The objective of the study was achieved by phospholipid fatty acid (PLFA) biomarker analysis of soil samples from L. barbarum fields in Zhongning County, Ningxia Province—the origin of L. barbarum. Five stand ages of L. barbarum were selected, < 1, 3, 6, 9, and 12 years (three plots each). The results showed that soil bulk density increased slightly with increasing stand age, while no clear trend was observed in soil pH or total salinity. As the stand age increased, soil organic matter and nutrients first increased before decreasing, with the highest levels being found in year 9. There was an amazing variety of PLFA biomarkers in soil samples at different stand ages. The average concentrations of total, bacterial, fungal, and actinomycete PLFAs in the surface soil initially decreased and then increased, before decreasing with the stand age in summer. The PLFA concentrations of major microbial groups were highest in year 9, with the total PLFA concentrations being 32.97% and 10.67% higher than those in years < 1 and 12, respectively. Higher microbial PLFA concentrations were detected in summer relative to autumn and in the surface relative to the subsurface soil. The highest ratios of Gram-positive to Gram-negative bacterial (G^?/G⁺) and fungal to bacterial (F/B) PLFAs were obtained in year 6, on average 76.09% higher than those at the other four stand ages. The soil environment was most stable in year 6, with no differences between other stand ages. Therefore, soil microbial community structure was strongly influenced by the stand age in year 6 only. The effect of stand age on soil G^?/G⁺ and microbial community structure varied with season and depth; there was little effect for F/B in the 20–40 cm soil layer. Principal component analysis revealed no correlations between microbial PLFA concentrations and total salinity in the soil; negative correlations were noted between soil pH and F/B in summer (P < 0.01), as well as between soil pH and fungal PLFA in autumn (P < 0.05). Moreover, microbial PLFA concentrations were correlated with soil organic matter (mean R = 0.7725), total nitrogen (mean R = 0.8296), total phosphorus (mean R = 0.8175), available nitrogen (mean R = 0.7458), and available phosphorus (mean R = 0.7795) (P < 0.01). On the whole, the soil ecosystem was most stable in year 6, while soil organic matter, nutrients, and microbial PLFA concentrations were maximal in year 9; thereafter, soil fertility indices and microbial concentrations decreased and soil quality declined gradually as the stand age increased. Therefore, farmers should reduce the application rate of fertilizers, especially compound or mixed fertilizers, in L. barbarum fields; organic or bacterial manure can be applied increasingly to improve the soil environment and prolong the economic life of L. barbarum.     

Comparison of Whole-Cell Fatty Acid (MIDI) or Phospholipid Fatty Acid (PLFA) Extractants as Biomarkers to Profile Soil Microbial Communities

The whole-cell lipid extraction to profile microbial communities on soils using fatty acid (FA) biomarkers is commonly done with the two extractants associated with the phospholipid fatty acid (PLFA) or Microbial IDentification Inc. (MIDI) methods. These extractants have very different chemistry and lipid separation procedures, but often shown a similar ability to discriminate soils from various management and vegetation systems. However, the mechanism and the chemistry of the exact suite of FAs extracted by these two methods are poorly understood. Therefore, the objective was to qualitatively and quantitatively compare the MIDI and PLFA microbial profiling methods for detecting microbial community shifts due to soil type or management. Twenty-nine soil samples were collected from a wide range of soil types across Oregon and extracted FAs by each method were analyzed by gas chromatography (GC) and GC-mass spectrometry. Unlike PLFA profiles, which were highly related to microbial FAs, the overall MIDI-FA profiles were highly related to the plant-derived FAs. Plant-associated compounds were quantitatively related to particulate organic matter (POM) and qualitatively related to the standing vegetation at sampling. These FAs were negatively correlated to respiration rate normalized to POM (Resp_POM), which increased in systems under more intensive management. A strong negative correlation was found between MIDI-FA to PLFA ratios and total organic carbon (TOC). When the reagents used in MIDI procedure were tested for the limited recovery of MIDI-FAs from soil with high organic matter, the recovery of MIDI-FA microbial signatures sharply decreased with increasing ratios of soil to extractant. Hence, the MIDI method should be used with great caution for interpreting changes in FA profiles due to shifts in microbial communities.