Sediment Microbes Research Articles

Mediation of functional gene and bacterial community profiles in the sediments of eutrophic Chaohu Lake by total nitrogen and season

Microbes in sediments contribute to nutrient release and play an important role in lake eutrophication. However, information about the profiles of functional genes and bacterial communities and the most important environmental factor affecting them in the sediments of eutrophic lake remains unrevealed. In this work, the real-time fluorescent quantitative polymerase chain reaction (qPCR) assay and 16S ribosomal RNA gene next generation sequencing analysis were used to explore the profiles of functional genes and bacterial communities in the sediments of Chaohu Lake. The selected 18 functional genes involved in C, N and P cycles were detected in most of samples. Seasonal variation and sediment variables were found to affect the profiles of functional genes and bacterial communities, and total nitrogen was the dominant environmental factor to drive the formation of bacterial community structure. Proteobacteria and Firmicutes were observed to be the two dominant phyla in the sediments with relative abundance ranging from 10.8% to 36.0% and 7.7%–46.7%, respectively. Three bacterial phyla, i.e., Actinobacteria, Proteobacteria, and Spirochaetes, were found to be significantly positively correlated with the C, N and P-cycle related functional genes. Bacterial community structure was the most important driver to shape the profiles of functional genes. Seasonal variation also influenced the co-occurrence patterns between functional genes and bacterial taxa as revealed by network analysis. The findings from this work facilitate a better understanding about the C, N, and P cycles in the sediments of eutrophic lakes.

Comparative metagenomics study reveals pollution induced changes of microbial genes in mangrove sediments

Mangrove forests are widespread along the subtropical and tropical coasts. They provide a habitat for a wide variety of plants, animals and microorganisms, and act as a buffer zone between the ocean and land. Along with other coastal environments, mangrove ecosystems are under increasing pressure from human activities, such as excessive input of nutrients and toxic pollutants. Despite efforts to understand the diversity of microbes in mangrove sediments, their metabolic capability in pristine and contaminated mangrove sediments remains largely unknown. By using metagenomic approach, we investigated the metabolic capacity of microorganisms in contaminated (CMS) and pristine (PMS) mangrove sediments at subtropical and tropical coastal sites. When comparing the CMS with PMS, we found that the former had a reduced diazotroph abundance and nitrogen fixing capability, but an enhanced metabolism that is related to the generation of microbial greenhouse gases via increased methanogenesis and sulfate reduction. In addition, a high concentration of heavy metals (mainly Zn, Cd, and Pb) and abundance of metal/antibiotic resistance encoding genes were found in CMS. Together, these data provide evidence that contamination in mangrove sediment can markedly change microbial community and metabolism; however, no significant differences in gene distribution were found between the subtropical and tropical mangrove sediments. In summary, contamination in mangrove sediments might weaken the microbial metabolisms that enable the mangrove ecosystems to act as a buffer zone for terrestrial nutrients deposition, and induce bioremediation processes accompanied with an increase in greenhouse gas emission.

Screening and Immobilizing the Denitrifying Microbes in Sediment for Bioremediation

In this study, immobilized microbial beads were proposed as a solution for excessive nitrogen concentration of the river sediment. The predominant denitrifying microbes were screened from the river sediment. The optimized production of immobilized microbial beads and long-term nitrogen removal efficiency were investigated. 16S rRNA gene sequencing analysis showed that denitrifying bacteria such as Pseudomonas, Alcaligenes, Proteiniclasticum, Achromobacter and Methylobacillus were dominant microflora in the enriched microbial agent, which accounted for 94.43% of the total microbes. Pseudomonas belongs to Gammaproteo bacteria, accounting for 49.22% and functioned as the most predominant denitrifying bacteria. The material concentration of 8% polyvinyl alcohol, 0.5% sodium alginate and 12.5% microbial biomass were found to be the optimal immobilizing conditions. The NH4+-N and total nitrogen (TN) removal rates in sediment with dosing immobilized microbial beads were estimated as 68.1% and 67.8%, respectively, when compared to the dosing liquid microbial agent were 50.5% and 49.3%. Meanwhile, the NH4+-N and TN removal rates in overlying water went up from 53.14% to 59.69% and from 68.03% to 78.13%, respectively, by using immobilized microbial beads.

The Speciation and Mobility of Mn and Fe in Estuarine Sediments

Dissolved and solid-phase speciation of Mn and Fe was measured in the porewaters of sediments recovered from three sites in the Greater St. Lawrence Estuary: the Saguenay Fjord, the Lower St. Lawrence Estuary (LSLE) and the Gulf of St. Lawrence (GSL). At all sites and most depths, metal organic ligand complexes (Mn(III)–L and Fe(III)–L) dominated the sedimentary porewater speciation, making up to 100% of the total dissolved Mn or Fe. We propose that these complexes play a previously underestimated role in maintaining oxidized soluble metal species in sedimentary systems and in stabilizing organic matter in the form of soluble metal–organic complexes. In the fjord porewaters, strong (log KCOND > 13.2) and weak (log KCOND < 13.2) Mn(III)–L complexes were detected, whereas only weak Mn(III)–L complexes were detected at the pelagic and hemipelagic sites of the GSL and LSLE, respectively. At the fjord site, Mn(III)–L complexes were kinetically stabilized against reduction by Fe(II), even when Fe(II) concentrations were as high as 57 μM. Only dissolved Mn(II) was released from the sediments to overlying waters, suggesting that Mn(III) may be preferentially oxidized by sedimentary microbes at or near the sediment–water interface. We calculated the dissolved Mn(II) fluxes from the sediments to the overlying waters to be 0.24 µmol cm−2 year−1 at the pelagic site (GSL), 11 µmol cm−2 year−1 at the hemipelagic site (LSLE) and 2.0 µmol cm−2 year−1 in the fjord. The higher benthic flux in the LSLE reflects the lower oxygen concentrations (dO2) of the bottom waters and sediments at this site, which favor the reductive dissolution of Mn oxides as well as the decrease in the oxidation rate of dissolved Mn(II) diffusing through the oxic layer of the sediment and its release to the overlying water.

Comparison of Microbial Communities in the Sediments and Water Columns of Frozen Cryoconite Holes in the McMurdo Dry Valleys, Antarctica

Although cryoconite holes, sediment-filled melt holes on glacier surfaces, appear small and homogenous, their microbial inhabitants may be spatially partitioned. This partitioning could be particularly important for maintaining biodiversity in holes that remain isolated for many years, such as in Antarctica. We hypothesized that cryoconite holes with greater species richness and biomass should exhibit greater partitioning between the sediments and water, promoting greater biodiversity through spatial niche partitioning. We tested this hypothesis by sampling frozen cryoconite holes along a gradient of biomass and biodiversity in the Taylor Valley, Antarctica, where ice-lidded cryoconite holes are a ubiquitous feature of glaciers. We extracted DNA and chlorophyll a from the sediments and water of these samples to describe biodiversity and quantify proxies for biomass. Contrary to our expectation, we found that cryoconite holes with greater richness and biomass showed less partitioning of phylotypes by the sediments versus the water, perhaps indicating that the probability of sediment microbes being mixed into the water is higher from richer sediments. Another explanation may be that organisms from the water were compressed by freezing down to the sediment layer, leaving primarily relic DNA of dead cells to be detected higher in the frozen water. Further evidence of this explanation is that the dominant sequences unique to water closely matched organisms that do not live in cryoconite holes or the Dry Valleys (e.g., vertebrates); so this cryptic biodiversity could represent unknown microbial animals or DNA from atmospheric deposition of dead biomass in the otherwise low-biomass water. Although we cannot rule out spatial niche partitioning occurring at finer scales or in melted cryoconite holes, we found no evidence of partitioning between the sediments and water in frozen holes. Future work should include more sampling of cryoconite holes at a finer spatial scale, and characterizing the communities of the sediments and water when cryoconite holes are melted and active.

Electrode Potential Dependency of Single-Cell Activity Identifies the Energetics of Slow Microbial Electron Uptake Process.

Electrochemical measurements have been widely applied to study microbial extracellular electron transport processes. However, because electrochemistry detects not only microbial electron transport but also other reactions, background signals comparable to or larger than microbial ones hamper the identification of microbial electrochemical properties. This problem is crucial especially for the detection of electron uptake processes by slow-growing microbes in low-energy subsurface sediments, as the environmental samples contain electrochemically active humus and mineral particles. In this study, we report a cell-specific stable isotope analysis to quantify the electrode potential dependency of anabolic activity in individual cells for identifying the electron uptake energetics of slow-growing bacteria. Followed by the incubation of Desulfovibrio ferrophilus IS5 cells with isotopic 15N-ammonium as the sole N source on electrodes poised at potentials of -0.2, -0.3, -0.4, and -0.5 V [vs. standard hydrogen electrode (SHE)], we conducted nanoscale secondary ion mass spectroscopy (NanoSIMS) to quantify 15N assimilation in more than 100 individual cells on the electrodes. We observed significant 15N assimilation at potentials of -0.4 and more 15N assimilation at -0.5 V, which is consistent with the onset potential for electron uptake via outer-membrane cytochromes (OMCs). The activation of cell energy metabolism was further examined by transcriptome analysis. Our results showed a novel methodology to study microbial electron uptake energetics. The results also serve as the first direct evidence that energy acquisition is coupled to the electron uptake process in sulfate-reducing bacteria that are ubiquitous in the subsurface environments, with implications on the electron-fueled subsurface biosphere hypothesis and other microbial processes, such as anaerobic iron corrosion and anaerobic methane oxidation.

Seasonal and spatial variation in the sediment bacterial community and diversity of Lake Bosten, China.

To explore the influence of seasonal and regional environmental factors on the bacterial community composition (BCC) and diversity in the sediments of Lake Bosten, the 16S rRNA gene of sediment bacteria in four samples from the lake center area and the macrophyte-dominated area were sequenced using 454 pyrosequencing. According to the operational taxonomic units (OTUs), diversity index, relative abundance, and redundancy analysis (RDA) of the bacteria, the results showed that (i) the bacterial diversity of the lake center area was lower than that of the macrophyte-dominated area, and it was higher in winter than that in summer as a whole; (ii) seasonal factors and geographical changes had obvious effects on the abundance of dominant bacteria, including Proteobacteria and Firmicutes; (iii) a large number of unclassified bacteria were detected in this study, and the dominant unclassified genera in both lake areas included unclassified Sva0485, unclassified Anaerolineaceae, and unclassified Nitrospiraceae; and (iv) TN and TOC were the main environmental factors influencing the sediment bacterial community in Lake Bosten, as determined by RDA analysis. The study provides a reference for the in-depth understanding the impact with the change of time and space on sediment microbes in Lake Bosten.

Intracellular nitrate in sediments of an oxygen-deficient marine basin is linked to pelagic diatoms.

Intracellular nitrate is an important electron acceptor in oxygen-deficient aquatic environments, either for the nitrate-storing microbes themselves, or for ambient microbial communities through nitrate leakage. This study links the spatial distribution of intracellular nitrate with the abundance and identity of nitrate-storing microbes in sediments of the Bornholm Basin, an environmental showcase for severe hypoxia. Intracellular nitrate (up to 270 nmol cm-3 sediment) was detected at all 18 stations along a 35-km transect through the basin and typically extended as deep as 1.6 cm into the sediment. Intracellular nitrate contents were particularly high at stations where chlorophyll contents suggested high settling rates of pelagic primary production. The depth distribution of intracellular nitrate matched that of the diatom-specific photopigment fucoxanthin in the upper 1.6 cm and calculations support that diatoms are the major nitrate-storing microbes in these sediments. In contrast, other known nitrate-storing microbes, such as sulfide-oxidizing bacteria and foraminifers, played only a minor role, if any. Strikingly, 18S rRNA gene sequencing revealed that the majority of the diatoms in the sediment were pelagic species. We conclude that intracellular nitrate stored by pelagic diatoms is transported to the seafloor by settling phytoplankton blooms, implying a so far overlooked 'biological nitrate pump'.

Bacterial Communities in Riparian Sediments: A Large-Scale Longitudinal Distribution Pattern and Response to Dam Construction.

Sediment microbes play major roles in riparian ecosystems; however, little is known about their longitudinal distribution pattern and their responses to dam construction, the most severe human disturbance in river basins. Here, we investigated the variability of sediment bacterial communities along a large-scale longitudinal gradient and between dam-controlled and dam-affected sites in riparian zone of the Lancang River, China. The abundance, activity and diversity of sediment bacteria gradually increased in a downstream direction, but were significantly lower in the dam-affected sites than in the dam-controlled sites. The bacterial community compositions differed significantly between the upper-middle-reach and downstream sites at all control sites, and also between the dam-affected and dam-controlled sites. In the cascade dam area, the relative importance of spatial distance and environmental heterogeneity for bacterial distribution differed between the dam-controlled and dam-affected sites. Spatial distance was the primary cause of variations in bacterial community in dam-controlled site. By contrast, the environmental heterogeneity had more control over the bacterial communities than did the spatial distance in dam-affected site. Network analysis showed that the bacterial community in the dam-affected sites had lower connectivity and stability when compared with that in dam-controlled sites. These results suggest the distinct variations in sediment bacterial community in dam-affected sites, which could enhance our understanding of potential ecological effects caused by dam construction.

Effects of different salinity levels on nitrification processes in sediments of Minjiang River Estuary, China

Two estuary wetlands in Minjiang River, Shanyutan and Daoqingzhou, were selected as the research objects. Wetland sediments were collected to examine the effects of different salinity levels on nitrification processes in the wetland with a culture experiment. The results showed that the nitrification rate of sediment in Minjiang River estuary wetland was generally low. The highest nitrification rate of sediment in the Shanyutan wetland was 0.193 mg·kg-1·d-1, while that in the Daoqingzhou wetland did not exceed 0.050 mg·kg-1·d-1. In the low salinity level (5), the decrease of nitrification rate was attributed to the restraint of nitrifying bacteria activities. The nitrification rate slightly increased with the increases of salinity (10), but was still lower than the initial value. This would be owed to the fact that the inhibitory effect of salinity on the activities of aerobic bacteria was strengthened, which reduced the rate of NH4+-N production, and thus resulted in a decrease of the contribution of aerobic ammonification bacteria to the apparent nitrification rate. There were regional variations in the responses of sediment nitrifying activity to salinity. In saltwater wetland (Shanyutan wetland), the adaptability of microbes in sediment to salinity was stronger, so that the nitrification activity in high salinity conditions was still higher. While in the freshwater wetland (Daoqingzhou wetland), the adaptability of the sediment to the salinity change was relatively lower, with a lower nitrification activity of the sediment in high salinity than in middle salinity. Acidic condition was the main reason for the low nitrification in the sediments of Minjiang estuary wetland. The nitrification rate and nitrification activity of the two wetlands increased first and then decreased with the duration of the culture experiment, which was driven by combined effects of initial NH4+-N concentration, oxygen content and denitrifying.

Sediment diffusion method improves wastewater nitrogen removal in the receiving lake sediments

Sediment microbes have a great potential to transform reactive N to harmless N2, thus decreasing wastewater nitrogen load into aquatic ecosystems. Here, we examined if spatial allocation of the wastewater discharge by a specially constructed sediment diffuser pipe system enhanced the microbial nitrate reduction processes. Full-scale experiments were set on two Finnish lake sites, Keuruu and Petäjävesi, and effects on the nitrate removal processes were studied using the stable isotope pairing technique. All nitrate reduction rates followed nitrate concentrations, being highest at the wastewater-influenced sampling points. Complete denitrification with N2 as an end-product was the main nitrate reduction process, indicating that the high nitrate and organic matter concentrations of wastewater did not promote nitrous oxide (N2O) production (truncated denitrification) or ammonification (dissimilatory nitrate reduction to ammonium; DNRA). Using 3D simulation, we demonstrated that the sediment diffusion method enhanced the contact time and amount of wastewater near the sediment surface especially in spring and in autumn, altering organic matter concentration and oxygen levels, and increasing the denitrification capacity of the sediment. We estimated that natural denitrification potentially removed 3–10% of discharged wastewater nitrate in the 33 ha study area of Keuruu, and the sediment diffusion method increased this areal denitrification capacity on average 45%. Overall, our results indicate that sediment diffusion method can supplement wastewater treatment plant (WWTP) nitrate removal without enhancing alternative harmful processes.

Effects of Ag and Ag2S nanoparticles on denitrification in sediments

The widespread use of commercial silver nanoparticles (Ag NPs) inevitably results in their increased release into natural waters and subsequent deposition in sediments, requiring the environmental impact of such deposition to be closely investigated. Hence, the effects of Ag NPs, polyvinylpyrrolidone (PVP)-Ag NPs, and sliver sulfide nanoparticles (Ag2S NPs) on denitrification-induced gas production (N2O and 15N-N2), and denitrifying microbes in freshwater sediments were investigated. Slurry experiments (8 h) combined with a 15NO3− addition technique were performed to determine the gaseous production. The abundance of relative functional genes (nirK, nirS and nosZ) and the composition of functional community were determined through RT-PCR and high-throughput sequencing, respectively. The obtained results showed that the toxicity of NPs on denitrification depended on their type (Ag+ > Ag NPs > PVP-Ag NPs > Ag2S NPs) and concentration, e.g., all 1 mg/L NPs exhibited no effects on denitrification, whereas evident hormesis effect–induced acceleration was observed in the case of Ag+. Conversely, 10 mg/L Ag+ and Ag NPs significantly inhibited the release rates of N2O and N2 by decreasing the abundance of functional genes (nirK and/or nirS) and the predominant bacteria Paracoccus. PVP-Ag and Ag2S NPs had no effects on N2 release rates and the composition of denitrifiers, however, inhibited the emission of N2O (by reducing the abundance of nirK), suggesting that normal denitrification-induced N2 formation in sediments could still be sustained when the N2O production decrease lied within a certain range. Further, the inhibiting ability of Ag-containing NPs was caused by their intrinsic nanotoxicity to functional microbes rather than by the general toxicity of Ag+. Besides, Ag2S NPs (as a main detoxification form of AgNPs) were revealed to be intrinsically nanotoxic to denitrifiers, albeit showing the lowest inhibitory effect among the three tested NPs. Thus, this study demonstrated that the inhibitory effect of Ag-containing NPs on denitrification in sediments depends on their morphology and type, implying that the stability and toxicity of Ag-containing NPs should be considered with caution.

Diversity of Gene Clusters for Polyketide and Nonribosomal Peptide Biosynthesis Revealed by Metagenomic Analysis of the Yellow Sea Sediment.

Polyketides (PKs) and nonribosomal peptides (NRPs) are widely applied as drugs in use today, and one potential source for novel PKs and NRPs is the marine sediment microbes. However, the diversities of microbes and their PKs and NRPs biosynthetic genes in the marine sediment are rarely reported. In this study, 16S rRNA gene fragments of the Yellow Sea sediment were analyzed, demonstrating that Proteobacteria and Bacteroidetes accounted for 62% of all the bacterial species and Actinobacteria bacteria which were seen as the typical PKs and NRPs producers only accounted for 0.82% of all the bacterial species. At the same time, PKs and NRPs diversities were evaluated based on the diversity of gene fragments of type I polyketide synthase (PKS) ketosynthase domain (KS), nonribosomal peptide synthetase (NRPS) adenylation domain (AD), and dTDP-glucose-4,6-dehydratase (dTGD). The results showed that AD genes and dTGD genes were abundant and some of them had less than 50% identities with known ones; By contrast, only few KS genes were identified and most of them had more than 60% identities with known KS genes. Moreover, one 70,000-fosmid clone library was further constructed to screen for fosmid clones harboring PKS or NRPS gene clusters of the Yellow Sea sediment. Nine selected fosmid clones harboring KS or AD were sequenced, and three of the clones were assigned to Proteobacteria. Though only few Actinobacteria 16S rRNA gene sequences were detected in the microbial community, five of the screened fosmid clones were assigned to Actinobacteria. Further assembly of the 9 fosmid clones resulted in 11 contigs harboring PKS, NRPS or hybrid NPRS-PKS gene clusters. These gene clusters showed less than 60% identities with the known ones and might synthesize novel natural products. Taken together, we revealed the diversity of microbes in the Yellow Sea sediments and found that most of the microbes were uncultured. Besides, evaluation of PKS and NRPS biosynthetic gene clusters suggested that the marine sediment might have the ability to synthesize novel natural products and more NRPS gene clusters than PKS gene clusters distributed in this environment.

Homoacetogenesis in Deep-Sea Chloroflexi, as Inferred by Single-Cell Genomics, Provides a Link to Reductive Dehalogenation in Terrestrial Dehalococcoidetes.

ABSTRACTThe deep marine subsurface is one of the largest unexplored biospheres on Earth and is widely inhabited by members of the phylum Chloroflexi. In this report, we investigated genomes of single cells obtained from deep-sea sediments of the Peruvian Margin, which are enriched in such Chloroflexi. 16S rRNA gene sequence analysis placed two of these single-cell-derived genomes (DscP3 and Dsc4) in a clade of subphylum I Chloroflexi which were previously recovered from deep-sea sediment in the Okinawa Trough and a third (DscP2-2) as a member of the previously reported DscP2 population from Peruvian Margin site 1230. The presence of genes encoding enzymes of a complete Wood-Ljungdahl pathway, glycolysis/gluconeogenesis, a Rhodobacter nitrogen fixation (Rnf) complex, glyosyltransferases, and formate dehydrogenases in the single-cell genomes of DscP3 and Dsc4 and the presence of an NADH-dependent reduced ferredoxin:NADP oxidoreductase (Nfn) and Rnf in the genome of DscP2-2 imply a homoacetogenic lifestyle of these abundant marine Chloroflexi. We also report here the first complete pathway for anaerobic benzoate oxidation to acetyl coenzyme A (CoA) in the phylum Chloroflexi (DscP3 and Dsc4), including a class I benzoyl-CoA reductase. Of remarkable evolutionary significance, we discovered a gene encoding a formate dehydrogenase (FdnI) with reciprocal closest identity to the formate dehydrogenase-like protein (complex iron-sulfur molybdoenzyme [CISM], DET0187) of terrestrial Dehalococcoides/Dehalogenimonas spp. This formate dehydrogenase-like protein has been shown to lack formate dehydrogenase activity in Dehalococcoides/Dehalogenimonas spp. and is instead hypothesized to couple HupL hydrogenase to a reductive dehalogenase in the catabolic reductive dehalogenation pathway. This finding of a close functional homologue provides an important missing link for understanding the origin and the metabolic core of terrestrial Dehalococcoides/Dehalogenimonas spp. and of reductive dehalogenation, as well as the biology of abundant deep-sea Chloroflexi.

Estimating Population Turnover Rates by Relative Quantification Methods Reveals Microbial Dynamics in Marine Sediment.

The difficulty involved in quantifying biogeochemically significant microbes in marine sediments limits our ability to assess interspecific interactions, population turnover times, and niches of uncultured taxa. We incubated surface sediments from Cape Lookout Bight, North Carolina, USA, anoxically at 21°C for 122 days. Sulfate decreased until day 68, after which methane increased, with hydrogen concentrations consistent with the predicted values of an electron donor exerting thermodynamic control. We measured turnover times using two relative quantification methods, quantitative PCR (qPCR) and the product of 16S gene read abundance and total cell abundance (FRAxC, which stands for "fraction of read abundance times cells"), to estimate the population turnover rates of uncultured clades. Most 16S rRNA reads were from deeply branching uncultured groups, and ∼98% of 16S rRNA genes did not abruptly shift in relative abundance when sulfate reduction gave way to methanogenesis. Uncultured Methanomicrobiales and Methanosarcinales increased at the onset of methanogenesis with population turnover times estimated from qPCR at 9.7 ± 3.9 and 12.6 ± 4.1 days, respectively. These were consistent with FRAxC turnover times of 9.4 ± 5.8 and 9.2 ± 3.5 days, respectively. Uncultured Syntrophaceae, which are possibly fermentative syntrophs of methanogens, and uncultured Kazan-3A-21 archaea also increased at the onset of methanogenesis, with FRAxC turnover times of 14.7 ± 6.9 and 10.6 ± 3.6 days. Kazan-3A-21 may therefore either perform methanogenesis or form a fermentative syntrophy with methanogens. Three genera of sulfate-reducing bacteria, Desulfovibrio, Desulfobacter, and Desulfobacterium, increased in the first 19 days before declining rapidly during sulfate reduction. We conclude that population turnover times on the order of days can be measured robustly in organic-rich marine sediment, and the transition from sulfate-reducing to methanogenic conditions stimulates growth only in a few clades directly involved in methanogenesis, rather than in the whole microbial community.IMPORTANCE Many microbes cannot be isolated in pure culture to determine their preferential growth conditions and predict their response to changing environmental conditions. We created a microcosm of marine sediments that allowed us to simulate a diagenetic profile using a temporal analog for depth. This allowed for the observation of the microbial community population dynamics caused by the natural shift from sulfate reduction to methanogenesis. Our research provides evidence for the population dynamics of uncultured microbes as well as the application of a novel method of turnover rate analysis for individual taxa within a mixed incubation, FRAxC, which stands for "fraction of read abundance times cells," which was verified by quantitative PCR. This allows for the calculation of population turnover times for microbes in a natural setting and the identification of uncultured clades involved in geochemical processes.

The Enigmatic Genome of an Obligate Ancient Spiroplasma Symbiont in a Hadal Holothurian.

Protective symbiosis has been reported in many organisms, but the molecular mechanisms of the mutualistic interactions between the symbionts and their hosts are unclear. Here, we sequenced the 424-kbp genome of "Candidatus Spiroplasma holothuricola," which dominated the hindgut microbiome of a sea cucumber, a major scavenger captured in the Mariana Trench (6,140 m depth). Phylogenetic relationships indicated that the dominant bacterium in the hindgut was derived from a basal group of Spiroplasma species. In this organism, the genes responsible for the biosynthesis of amino acids, glycolysis, and sugar transporters were lost, strongly suggesting endosymbiosis. The highly decayed genome consists of two chromosomes and harbors genes coding for proteolysis, microbial toxin, restriction-methylation systems, and clustered regularly interspaced short palindromic repeats (CRISPRs), composed of three cas genes and 76 CRISPR spacers. The holothurian host is probably protected against invading viruses from sediments by the CRISPRs/Cas and restriction systems of the endosymbiotic spiroplasma. The protective endosymbiosis indicates the important ecological role of the ancient Spiroplasma symbiont in the maintenance of hadal ecosystems.IMPORTANCE Sea cucumbers are major inhabitants in hadal trenches. They collect microbes in surface sediment and remain tolerant against potential pathogenic bacteria and viruses. This study presents the genome of endosymbiotic spiroplasmas in the gut of a sea cucumber captured in the Mariana Trench. The extreme reduction of the genome and loss of essential metabolic pathways strongly support its endosymbiotic lifestyle. Moreover, a considerable part of the genome was occupied by a CRISPR/Cas system to provide immunity against viruses and antimicrobial toxin-encoding genes for the degradation of microbes. This novel species of Spiroplasma is probably an important protective symbiont for the sea cucumbers in the hadal zone.

Accumulation of deaminated peptides in anoxic sediments of Santa Barbara Basin

Proteins represent the most abundant class of biomolecules in marine sinking particles and microbial biomass, yet their cycling in marine sediments is not fully understood. To investigate whether some portion of hydrolyzed proteins escapes complete remineralization and accumulate in the pore waters, we analyzed dissolved organic matter from the anoxic sediments of Santa Barbara Basin, California, by Fourier Transform Ion Cyclotron Resonance Mass Spectrometry (FTICR-MS). The results showed an increase in the molecular diversity and abundance of dissolved organic nitrogen (DON) formulas with depth. A comparison of the detected DON formulas to a database of small peptides (2–4 amino acid sequences) returned 119 matches, and these formulas were most abundant near the sediment surface. When we compared our detected formulas to all possible structures that would result from deamination of peptides in the database, we found 680 formula matches. However, these molecular formulas can represent hundreds of different structural isomers (in the present case as many as 3257 different deaminated peptide structures), which cannot be distinguished by the FTICR-MS settings that were used. Analysis of amino acid sequences suggests that these deaminated peptides may be the products of selective degradation of source proteins in marine sediments. We hypothesize that these deaminated peptides accumulate in the pore waters due to extracellular proteinases being inhibited from completely hydrolyzing specific peptides to free amino acids. We suggest that anaerobic microbes deaminate peptides largely to produce H2, which is ultimately used as a reducing agent by other sediment microbes (e.g. CO2 reduction by methanogens). Simple calculations suggest that deaminated peptides may represent ∼25–45% of DOC accumulating in these sediment pore waters. Unlike rapid remineralization of free amino acids, peptide deamination leaves behind the peptide carbon skeleton. Molecular structures of these remnant carbon skeletons may hold important clues about specific microbial processes influencing organic matter remineralization and accumulation.

New Method for Isolation of DNA from River Sediment Microbes of Sungai Kelantan, Malaysia

Extraction of pure genomic DNA from microbes isolated from river sediments often hindered byco-isolated carryovers from the reagents used which reduces the yield and quality of DNA.Existing methods were lengthy, expensive and does not result in high yield of genomic DNA inshort time. This study describes a rapid and inexpensive DNA extraction method that involvedminimal purification step without phenol or other carcinogenic reagents. The microbial DNAextraction steps also does not require the use of special laboratory equipment. This method usesCetyltrimethylammonium bromide (CTAB) followed by ethanol based purification steps andwithin one hour the extraction of microbial DNA could be completed. The extracted genomicDNA from microbes isolated from river sediment of Sungai Kelantan, Malaysia using the methoddescribed in this study is of quality suitable for other downstream researches such asmetagenomics sequencing.

Nitrifier Gene Abundance and Diversity in Sediments Impacted by Acid Mine Drainage.

Extremely acidic and metal-rich acid mine drainage (AMD) waters can have severe toxicological effects on aquatic ecosystems. AMD has been shown to completely halt nitrification, which plays an important role in transferring nitrogen to higher organisms and in mitigating nitrogen pollution. We evaluated the gene abundance and diversity of nitrifying microbes in AMD-impacted sediments: ammonia-oxidizing archaea (AOA), ammonia-oxidizing bacteria (AOB), and nitrite-oxidizing bacteria (NOB). Samples were collected from the Iron Springs Mining District (Ophir, CO, United States) during early and late summer in 2013 and 2014. Many of the sites were characterized by low pH (<5) and high metal concentrations. Sequence analyses revealed AOA genes related to Nitrososphaera, Nitrosotalea, and Nitrosoarchaeum; AOB genes related to Nitrosomonas and Nitrosospira; and NOB genes related to Nitrospira. The overall abundance of AOA, AOB and NOB was examined using quantitative PCR (qPCR) amplification of the amoA and nxrB functional genes and 16S rRNA genes. Gene copy numbers ranged from 3.2 × 104 – 4.9 × 107 archaeal amoA copies ∗ μg DNA-1, 1.5 × 103 – 5.3 × 105 AOB 16S rRNA copies ∗ μg DNA-1, and 1.3 × 106 – 7.7 × 107 Nitrospira nxrB copies ∗ μg DNA-1. Overall, Nitrospira nxrB genes were found to be more abundant than AOB 16S rRNA and archaeal amoA genes in most of the sample sites across 2013 and 2014. AOB 16S rRNA and Nitrospira nxrB genes were quantified in sediments with pH as low as 3.2, and AOA amoA genes were quantified in sediments as low as 3.5. Though pH varied across all sites (pH 3.2–8.3), pH was not strongly correlated to the overall community structure or relative abundance of individual OTUs for any gene (based on CCA and Spearman correlations). pH was positivity correlated to the total abundance (qPCR) of AOB 16S rRNA genes, but not for any other genes. Metals were not correlated to the overall nitrifier community composition or abundance, but were correlated to the relative abundances of several individual OTUs. These findings extend our understanding of the distribution of nitrifying microbes in AMD-impacted systems and provide a platform for further research.

Performance Optimization of Microbes from Shrimp Pond Sediment by Adding EM4 In Nitrification Process for the Treatment of Wastewater Containing High Ammonia Concentration

In liquid wastes, especially domestic wastewater, many organic substances are mixed causing water quality degradation, one of them is ammonia. Liquid wastes containing ammonia can be treated using an activated sludge system. One of the active sludge that can be used is shrimp pond sediments. This experiment investigated the performance of microbes in shrimp pond sediments with the addition of EM4 in nitrification process for the treatment of wastewater with high ammonia concentration in a 8 L batch reactor capacity. The results show that the addition of shrimp pond sediment as the active sludge can remove high ammonia level almost completely and there is known interaction between time and variation of shrimp pond quantity (p value <0,05) to the decreasing of ammonia level. Efficiency of decreasing the concentration of ammonia up to 100% can be reached on the 15th day in each treatment. The addition of EM4 can shorten the decreasing of ammonia level by 50%. Keywords : Nitrification, Ammonia, Shrimp Pond Sediment, EM4, Activated Sludge