Viral potential to modulate microbial methane metabolism varies by habitat.

Methane is a potent greenhouse gas contributing to global warming. Microorganisms largely drive the biogeochemical cycling of methane, yet little is known about viral contributions to methane metabolism (MM). We analyzed 982 publicly available metagenomes from host-associated and environmental habitats containing microbial MM genes, expanding the known MM auxiliary metabolic genes (AMGs) from three to 24, including seven genes exclusive to MM pathways. These AMGs are recovered on 911 viral contigs predicted to infect 14 prokaryotic phyla including Halobacteriota, Methanobacteriota, and Thermoproteota. Of those 24, most were encoded by viruses from rumen (16/24), with substantially fewer by viruses from environmental habitats (0-7/24). To search for additional MM AMGs from an environmental habitat, we generate metagenomes from methane-rich sediments in Vrana Lake, Croatia. Therein, we find diverse viral communities, with most viruses predicted to infect methanogens and methanotrophs and some encoding 13 AMGs that can modulate host metabolisms. However, none of these AMGs directly participate in MM pathways. Together these findings suggest that the extent to which viruses use AMGs to modulate host metabolic processes (e.g., MM) varies depending on the ecological properties of the habitat in which they dwell and is not always predictable by habitat biogeochemical properties.

Magnetic-field-induced splitting of Rydberg Electromagnetically Induced Transparency and Autler-Townes spectra in 87Rb vapor cell.

We theoretically and experimentally investigate the Rydberg electromagnetically induced transparency (EIT) and Autler-Townes (AT) splitting of 87Rb vapor under the combined influence of a magnetic field and a microwave field. In the presence of static magnetic field, the effect of the microwave field leads to the dressing and splitting of each mF state, resulting in multiple spectral peaks in the EIT-AT spectrum. A simplified analytical formula was developed to explain the EIT-AT spectrum in a static magnetic field, and the theoretical calculations agree qualitatively with experimental results. The Rydberg atom microwave electric field sensor performance was enhanced by making use of the splitting interval between the two maximum absolute mF states separated by the static magnetic field, which was attributed to the stronger Clebsch-Gordon coefficients between the extreme mF states and the frequency detuning of the microwave electric field under the static magnetic field. The traceable measurement limit of weak electric field by EIT-AT splitting method was extended by an order of magnitude, which is promising for precise microwave electric field measurement.

Lower viral evolutionary pressure under stable versus fluctuating conditions in subzero Arctic brines.

BACKGROUND: Climate change threatens Earth's ice-based ecosystems which currently offer archives and eco-evolutionary experiments in the extreme. Arctic cryopeg brine (marine-derived, within permafrost) and sea ice brine, similar in subzero temperature and high salinity but different in temporal stability, are inhabited by microbes adapted to these extreme conditions. However, little is known about their viruses (community composition, diversity, interaction with hosts, or evolution) or how they might respond to geologically stable cryopeg versus fluctuating sea ice conditions. RESULTS: We used long- and short-read viromics and metatranscriptomics to study viruses in Arctic cryopeg brine, sea ice brine, and underlying seawater, recovering 11,088 vOTUs (~species-level taxonomic unit), a 4.4-fold increase of known viruses in these brines. More specifically, the long-read-powered viromes doubled the number of longer (≥25 kb) vOTUs generated and recovered more hypervariable regions by >5-fold compared to short-read viromes. Distribution assessment, by comparing to known viruses in public databases, supported that cryopeg brine viruses were of marine origin yet distinct from either sea ice brine or seawater viruses, while 94% of sea ice brine viruses were also present in seawater. A virus-encoded, ecologically important exopolysaccharide biosynthesis gene was identified, and many viruses (~half of metatranscriptome-inferred "active" vOTUs) were predicted as actively infecting the dominant microbial genera Marinobacter and Polaribacter in cryopeg and sea ice brines, respectively. Evolutionarily, microdiversity (intra-species genetic variations) analyses suggested that viruses within the stable cryopeg brine were under significantly lower evolutionary pressures than those in the fluctuating sea ice environment, while many sea ice brine virus-tail genes were under positive selection, indicating virus-host co-evolutionary arms races. CONCLUSIONS: Our results confirmed the benefits of long-read-powered viromics in understanding the environmental virosphere through significantly improved genomic recovery, expanding viral discovery and the potential for biological inference. Evidence of viruses actively infecting the dominant microbes in subzero brines and modulating host metabolism underscored the potential impact of viruses on these remote and underexplored extreme ecosystems. Microdiversity results shed light on different strategies viruses use to evolve and adapt when extreme conditions are stable versus fluctuating. Together, these findings verify the value of long-read-powered viromics and provide foundational data on viral evolution and virus-microbe interactions in Earth's destabilized and rapidly disappearing cryosphere. Video Abstract.

Precise measurement of microwave polarization using a Rydberg atom-based mixer.

A Rydberg atom-based mixer has opened up a new method to characterize microwave electric fields such as the precise measurement of their phase and strength. This study further demonstrates, theoretically and experimentally, a method to accurately measure the polarization of a microwave electric field based on a Rydberg atom-based mixer. The results show that the amplitude of the beat note changes with the polarization of the microwave electric field in a period of 180 degrees, and in the linear region a polarization resolution better than 0.5 degree can be easily obtained which reaches the best level by a Rydberg atomic sensor. More interestingly, the mixer-based measurements are immune to the polarization of the light field that forms the Rydberg EIT. This method considerably simplifies theoretical analysis and the experimental system required for measuring microwave polarization using Rydberg atoms and is of interest in microwave sensing.

Evolutionary Divergence of Marinobacter Strains in Cryopeg Brines as Revealed by Pangenomics.

Marinobacter spp. are cosmopolitan in saline environments, displaying a diverse set of metabolisms that allow them to competitively occupy these environments, some of which can be extreme in both salinity and temperature. Here, we introduce a distinct cluster of Marinobacter genomes, composed of novel isolates and in silico assembled genomes obtained from subzero, hypersaline cryopeg brines, relic seawater-derived liquid habitats within permafrost sampled near Utqiagvik, Alaska. Using these new genomes and 45 representative publicly available genomes of Marinobacter spp. from other settings, we assembled a pangenome to examine how the new extremophile members fit evolutionarily and ecologically, based on genetic potential and environmental source. This first genus-wide genomic analysis revealed that Marinobacter spp. in general encode metabolic pathways that are thermodynamically favored at low temperature, cover a broad range of organic compounds, and optimize protein usage, e.g., the Entner-Doudoroff pathway, the glyoxylate shunt, and amino acid metabolism. The new isolates contributed to a distinct clade of subzero brine-dwelling Marinobacter spp. that diverged genotypically and phylogenetically from all other Marinobacter members. The subzero brine clade displays genomic characteristics that may explain competitive adaptations to the extreme environments they inhabit, including more abundant membrane transport systems (e.g., for organic substrates, compatible solutes, and ions) and stress-induced transcriptional regulatory mechanisms (e.g., for cold and salt stress) than in the other Marinobacter clades. We also identified more abundant signatures of potential horizontal transfer of genes involved in transcription, the mobilome, and a variety of metabolite exchange systems, which led to considering the importance of this evolutionary mechanism in an extreme environment where adaptation via vertical evolution is physiologically rate limited. Assessing these new extremophile genomes in a pangenomic context has provided a unique view into the ecological and evolutionary history of the genus Marinobacter, particularly with regard to its remarkable diversity and its opportunism in extremely cold and saline environments.

MetaPop: a pipeline for macro- and microdiversity analyses and visualization of microbial and viral metagenome-derived populations.

BACKGROUND: Microbes and their viruses are hidden engines driving Earth's ecosystems from the oceans and soils to humans and bioreactors. Though gene marker approaches can now be complemented by genome-resolved studies of inter-(macrodiversity) and intra-(microdiversity) population variation, analytical tools to do so remain scattered or under-developed. RESULTS: Here, we introduce MetaPop, an open-source bioinformatic pipeline that provides a single interface to analyze and visualize microbial and viral community metagenomes at both the macro- and microdiversity levels. Macrodiversity estimates include population abundances and α- and ß-diversity. Microdiversity calculations include identification of single nucleotide polymorphisms, novel codon-constrained linkage of SNPs, nucleotide diversity (π and θ), and selective pressures (pN/pS and Tajima's D) within and fixation indices (FST) between populations. MetaPop will also identify genes with distinct codon usage. Following rigorous validation, we applied MetaPop to the gut viromes of autistic children that underwent fecal microbiota transfers and their neurotypical peers. The macrodiversity results confirmed our prior findings for viral populations (microbial shotgun metagenomes were not available) that diversity did not significantly differ between autistic and neurotypical children. However, by also quantifying microdiversity, MetaPop revealed lower average viral nucleotide diversity (π) in autistic children. Analysis of the percentage of genomes detected under positive selection was also lower among autistic children, suggesting that higher viral π in neurotypical children may be beneficial because it allows populations to better "bet hedge" in changing environments. Further, comparisons of microdiversity pre- and post-FMT in autistic children revealed that the delivery FMT method (oral versus rectal) may influence viral activity and engraftment of microdiverse viral populations, with children who received their FMT rectally having higher microdiversity post-FMT. Overall, these results show that analyses at the macro level alone can miss important biological differences. CONCLUSIONS: These findings suggest that standardized population and genetic variation analyses will be invaluable for maximizing biological inference, and MetaPop provides a convenient tool package to explore the dual impact of macro- and microdiversity across microbial communities. Video abstract.

Glacier ice archives nearly 15,000-year-old microbes and phages.

BACKGROUND: Glacier ice archives information, including microbiology, that helps reveal paleoclimate histories and predict future climate change. Though glacier-ice microbes are studied using culture or amplicon approaches, more challenging metagenomic approaches, which provide access to functional, genome-resolved information and viruses, are under-utilized, partly due to low biomass and potential contamination. RESULTS: We expand existing clean sampling procedures using controlled artificial ice-core experiments and adapted previously established low-biomass metagenomic approaches to study glacier-ice viruses. Controlled sampling experiments drastically reduced mock contaminants including bacteria, viruses, and free DNA to background levels. Amplicon sequencing from eight depths of two Tibetan Plateau ice cores revealed common glacier-ice lineages including Janthinobacterium, Polaromonas, Herminiimonas, Flavobacterium, Sphingomonas, and Methylobacterium as the dominant genera, while microbial communities were significantly different between two ice cores, associating with different climate conditions during deposition. Separately, ~355- and ~14,400-year-old ice were subject to viral enrichment and low-input quantitative sequencing, yielding genomic sequences for 33 vOTUs. These were virtually all unique to this study, representing 28 novel genera and not a single species shared with 225 environmentally diverse viromes. Further, 42.4% of the vOTUs were identifiable temperate, which is significantly higher than that in gut, soil, and marine viromes, and indicates that temperate phages are possibly favored in glacier-ice environments before being frozen. In silico host predictions linked 18 vOTUs to co-occurring abundant bacteria (Methylobacterium, Sphingomonas, and Janthinobacterium), indicating that these phages infected ice-abundant bacterial groups before being archived. Functional genome annotation revealed four virus-encoded auxiliary metabolic genes, particularly two motility genes suggest viruses potentially facilitate nutrient acquisition for their hosts. Finally, given their possible importance to methane cycling in ice, we focused on Methylobacterium viruses by contextualizing our ice-observed viruses against 123 viromes and prophages extracted from 131 Methylobacterium genomes, revealing that the archived viruses might originate from soil or plants. CONCLUSIONS: Together, these efforts further microbial and viral sampling procedures for glacier ice and provide a first window into viral communities and functions in ancient glacier environments. Such methods and datasets can potentially enable researchers to contextualize new discoveries and begin to incorporate glacier-ice microbes and their viruses relative to past and present climate change in geographically diverse regions globally. Video Abstract.

Expanding standards in viromics: in silico evaluation of dsDNA viral genome identification, classification, and auxiliary metabolic gene curation.

BACKGROUND: Viruses influence global patterns of microbial diversity and nutrient cycles. Though viral metagenomics (viromics), specifically targeting dsDNA viruses, has been critical for revealing viral roles across diverse ecosystems, its analyses differ in many ways from those used for microbes. To date, viromics benchmarking has covered read pre-processing, assembly, relative abundance, read mapping thresholds and diversity estimation, but other steps would benefit from benchmarking and standardization. Here we use in silico-generated datasets and an extensive literature survey to evaluate and highlight how dataset composition (i.e., viromes vs bulk metagenomes) and assembly fragmentation impact (i) viral contig identification tool, (ii) virus taxonomic classification, and (iii) identification and curation of auxiliary metabolic genes (AMGs). RESULTS: The in silico benchmarking of five commonly used virus identification tools show that gene-content-based tools consistently performed well for long (≥3 kbp) contigs, while k-mer- and blast-based tools were uniquely able to detect viruses from short (≤3 kbp) contigs. Notably, however, the performance increase of k-mer- and blast-based tools for short contigs was obtained at the cost of increased false positives (sometimes up to ∼5% for virome and ∼75% bulk samples), particularly when eukaryotic or mobile genetic element sequences were included in the test datasets. For viral classification, variously sized genome fragments were assessed using gene-sharing network analytics to quantify drop-offs in taxonomic assignments, which revealed correct assignations ranging from ∼95% (whole genomes) down to ∼80% (3 kbp sized genome fragments). A similar trend was also observed for other viral classification tools such as VPF-class, ViPTree and VIRIDIC, suggesting that caution is warranted when classifying short genome fragments and not full genomes. Finally, we highlight how fragmented assemblies can lead to erroneous identification of AMGs and outline a best-practices workflow to curate candidate AMGs in viral genomes assembled from metagenomes. CONCLUSION: Together, these benchmarking experiments and annotation guidelines should aid researchers seeking to best detect, classify, and characterize the myriad viruses 'hidden' in diverse sequence datasets.

Lower-Grade Gliomas: Predicting DNA Methylation Subtyping and its Consequences on Survival with MR Features.

RATIONALE AND OBJECTIVES: To explore associations between MR imaging features, DNA methylation subtyping, and survival in lower-grade gliomas (LGG). MATERIALS AND METHODS: The MR data from 170 patients generated with the Cancer Imaging Archive were reviewed. The correlation was evaluated by Fisher's Exact Test, Pearson Chi-Square and binary regression analysis. Survival analysis was conducted by using time-dependent ROC analysis and the Kaplan-Meier method (the worst prognosis subgroup). RESULTS: Identified were 9 (5.3%) M1-subtype, 18 (10.6%) M2-subtype, 48 (28.2%) M3-subtype, 31 (18.2%) M4-subtype and 64 (37.6%) M5-subtype. Patients with M4-subtype had the shortest median OS (49.3 vs. 28.4) months(p < 0.05). The time-dependent ROC for the M4-subtype was 0.83 (95% confidence interval 0.72-0.95) for survival at 12 months, 0.82 (95% confidence interval 0.70-0.94) for survival at 24 months, and 0.74 (95% confidence interval 0.62-0.86) for survival at 36 months. After uni- and multivariate analysis, a nomogram was built based on proportion contrast-enhanced (CE) tumor, extranodular growth, volume_cutoff_median, and location. For the prediction of M4-subtype, the nomogram showed good discrimination, with an area under the curve (AUC) of 0.886 (95% CI: 0.820-952) and was well calibrated. On multivariate logistic regression analysis, volume ≥60cm3 (OR: 0.200; p < 0.001; 95%CI: 0.048-0.834) was associated with M1-subtype (AUC: 0.690). Hemorrhage (OR: 5.443; p = 0.002; 95%CI: 1.844-16.069) and volume > median (OR: 3.256; p = 0.05; 95%CI: 0.992-10.686) were associated with M2-subtype (AUC: 0.733). Proportion CE tumor<=5% (OR: 3.968; P=0.002; 95%CI: 1.634-9.635) was associated with M3-subtype (AUC: 0.632). Poorly-defined (OR: 2.258; p = 0.05; 95%CI: 1.000-5.101) and volume > median (OR: 2.447; p = 0.01; 95%CI: 1.244-4.813) were associated with M5-subtype (AUC: 0.645). Decision curve analysis indicated predictions for all models were clinically useful. CONCLUSION: This preliminary radiogenomics analysis of lower-grade gliomas demonstrated associations between MR features and DNA methylation subtyping. The shortest survival was observed in patients with M4-subtype. And we have constructed nomogram that enables more accurate predictions of M4-subtype.

Dispersive microwave electrometry using Zeeman frequency modulation spectroscopy of electromagnetically induced transparency in Rydberg atoms.

We herein developed and demonstrated a Zeeman frequency modulation scheme for improving the signal-to-noise ratio of microwave electric field measurement using Rydberg atoms. The spectra of the electromagnetically induced transparency (EIT) and Autler-Townes splitting of Rydberg atoms is frequency modulated by an alternating current magnetic field. The signal-to-noise ratio of the corresponding dispersive error signal is enhanced more than 10 times than that of the original spectrum. Furthermore, we show that the slope of the dispersive error signal near the resonance of the Rydberg EIT can be used to characterize the weak microwave electric field amplitudes. The more intuitive and simpler structure compared with other existing frequency modulation technologies greatly reduces the difficulties of experiments and experimental data analysis.

Viral Ecogenomics of Arctic Cryopeg Brine and Sea Ice.

Arctic regions, which are changing rapidly as they warm 2 to 3 times faster than the global average, still retain microbial habitats that serve as natural laboratories for understanding mechanisms of microbial adaptation to extreme conditions. Seawater-derived brines within both sea ice (sea-ice brine) and ancient layers of permafrost (cryopeg brine) support diverse microbes adapted to subzero temperatures and high salinities, yet little is known about viruses in these extreme environments, which, if analogous to other systems, could play important evolutionary and ecosystem roles. Here, we characterized viral communities and their functions in samples of cryopeg brine, sea-ice brine, and melted sea ice. Viral abundance was high in cryopeg brine (1.2 × 108 ml-1) and much lower in sea-ice brine (1.3 × 105 to 2.1 × 105 ml-1), which roughly paralleled the differences in cell concentrations in these samples. Five low-input, quantitative viral metagenomes were sequenced to yield 476 viral populations (i.e., species level; ≥10 kb), only 12% of which could be assigned taxonomy by traditional database approaches, indicating a high degree of novelty. Additional analyses revealed that these viruses: (i) formed communities that differed between sample type and vertically with sea-ice depth; (ii) infected hosts that dominated these extreme ecosystems, including Marinobacter, Glaciecola, and Colwellia; and (iii) encoded fatty acid desaturase (FAD) genes that likely helped their hosts overcome cold and salt stress during infection, as well as mediated horizontal gene transfer of FAD genes between microbes. Together, these findings contribute to understanding viral abundances and communities and how viruses impact their microbial hosts in subzero brines and sea ice.IMPORTANCE This study explores viral community structure and function in remote and extreme Arctic environments, including subzero brines within marine layers of permafrost and sea ice, using a modern viral ecogenomics toolkit for the first time. In addition to providing foundational data sets for these climate-threatened habitats, we found evidence that the viruses had habitat specificity, infected dominant microbial hosts, encoded host-derived metabolic genes, and mediated horizontal gene transfer among hosts. These results advance our understanding of the virosphere and how viruses influence extreme ecosystems. More broadly, the evidence that virally mediated gene transfers may be limited by host range in these extreme habitats contributes to a mechanistic understanding of genetic exchange among microbes under stressful conditions in other systems.

Frequency stabilization method for transition to a Rydberg state using Zeeman modulation.

We herein develop and demonstrate a stable frequency-locking scheme for Rydberg atomic experiments. We use the Zeeman effect to modulate the three-level ladder-type Rydberg electromagnetically induced transparency (EIT) signal to lock the laser frequency of the coupling light for transition from its intermediate state to a Rydberg state. The effects of polarization of the probe and coupling lights, and the amplitude of the AC modulated magnetic field ${{\boldsymbol B}_0}$B0 on the EIT and the corresponding dispersive error signal, are both analyzed. The results show that both the EIT signal and dispersive error signal are the strongest when the polarizations of coupling and probe fields are circular and equal. The signal-to-noise ratio of the dispersive error signal increases with ${{\boldsymbol B}_0}$B0. The slope of the dispersive error signal increases first and then decreases with ${{\boldsymbol B}_0}$B0, which is related to the increase of the EIT linewidth caused by the higher ${{\boldsymbol B}_0}$B0. The linewidth of the laser is significantly less than 500 kHz after frequency locking, which satisfies the requirements of most experiments involving Rydberg atoms. The method proposed herein can generally be applied to any cascade system of Rydberg atoms.

Clean Low-Biomass Procedures and Their Application to Ancient Ice Core Microorganisms.

Microorganisms in glacier ice provide tens to hundreds of thousands of years archive for a changing climate and microbial responses to it. Analyzing ancient ice is impeded by technical issues, including limited ice, low biomass, and contamination. While many approaches have been evaluated and advanced to remove contaminants on ice core surfaces, few studies leverage modern sequencing to establish in silico decontamination protocols for glacier ice. Here we sought to apply such "clean" sampling techniques with in silico decontamination approaches used elsewhere to investigate microorganisms archived in ice at ∼41 (D41, ∼20,000 years) and ∼49 m (D49, ∼30,000 years) depth in an ice core (GS3) from the summit of the Guliya ice cap in the northwestern Tibetan Plateau. Four "background" controls were established - a co-processed sterile water artificial ice core, two air samples collected from the ice processing laboratories, and a blank, sterile water sample - and used to assess contaminant microbial diversity and abundances. Amplicon sequencing revealed 29 microbial genera in these controls, but quantitative PCR showed that the controls contained about 50-100-times less 16S DNA than the glacial ice samples. As in prior work, we interpreted these low-abundance taxa in controls as "contaminants" and proportionally removed them in silico from the GS3 ice amplicon data. Because of the low biomass in the controls, we also compared prokaryotic 16S DNA amplicons from pre-amplified (by re-conditioning PCR) and standard amplicon sequencing, and found the resulting microbial profiles to be repeatable and nearly identical. Ecologically, the contaminant-controlled ice microbial profiles revealed significantly different microorganisms across the two depths in the GS3 ice core, which is consistent with changing climate, as reported for other glacier ice samples. Many GS3 ice core genera, including Methylobacterium, Sphingomonas, Flavobacterium, Janthinobacterium, Polaromonas, and Rhodobacter, were also abundant in previously studied ice cores, which suggests wide distribution across glacier environments. Together these findings help further establish "clean" procedures for studying low-biomass ice microbial communities and contribute to a baseline understanding of microorganisms archived in glacier ice.

[Bacterial community characterization of rearing water of marine recirculating aquaculture systems for yellow grouper (Epinephelus awoara)].

OBJECTIVE: This study was aimed to reveal the bacterial community characteristics of rearing water of marine recirculating aquaculture system for yellow grouper (Epinephelus awoara), and compare the differences between bacterial community structure of healthy rearing water and sick rearing water in order to elucidate the relationship between bacterial community and fish disease. METHODS: The next generation sequencing method was used, and the bacterial community structure and α-diversity indices (species richness, species evenness and phylogenetic diversity) between the rearing water of healthy and diseased groups were studied and compared. In addition, the traditional cultivation method was used to isolate suspected pathogens from the niduses of diseased yellow groupers. RESULTS: There was no significant difference between the α-diversities of healthy and diseased rearing water bodies, however, the results of principal coordinates analysis (PCoA) and the sample clustering of heatmap showed that the bacterial communities of healthy and diseased rearing water bodies were quite different. Although phyla Proteobacteria, Verrucomicrobia and Bacteroidetes were all the predominant ones in both communities, their relative abundance varied greatly. In diseased community, the relative abundances of α-Proteobacteria (25.07%) and γ-Protebacteria (22.74%) were similar, whereas the proportion of γ-Protebacteria (40.49%) was much higher than α-Proteobacteria (10.87%) in the healthy community. The differences of Verrucomicrobia and Bacteroidetes between the healthy and the diseased rearing water were also significant with relative abundances of 10.9% and 26.4%, and 20.9% and 12.3%, respectively. The significantly different families were Rhodobacteraceae and Rhodospirillaceae within class α-Proteobacteria; Alteromonadaceae and HTCC2188 within class γ-Protebacteria; Verrucomicrobiaceae within Verrucomicrobia; Cryomorphaceae within Bacteroidetes. The healthy and the diseased communities owned specific core microbes themselves. Glaciecola, HTCC, Sediminicola and Prevotella were the core genera in healthy rearing water, and Vibrio, Rubritalea and Oleibacter in diseased rearing water. Twenty strains of Vibrio spp. and one of Acinetobacter haemolyticus were isolated from skin, liver and spleen of diseased yellow grouper. CONCLUSION: The shift of bacterial community structure and relative abundance of rearing water will help monitor the healthy status of recirculating aquaculture system. Our study provides theory and experimental basis to diagnosis and monitor of Vibrio disease for yellow grouper recirculating aquaculture system.

Planktosalinus lacus gen. nov., sp. nov., a member of the family Flavobacteriaceae isolated from a salt lake.

A Gram-staining-negative bacterium, strain X14M-14T, was isolated from a salt lake (Lake Xiaochaidan) in Qaidam basin, Qinghai Province, China. Its taxonomic position was determined by using a polyphasic approach. Cells of strain X14M-14T were non-spore-forming, non-motile rods. Strain X14M-14T was strictly heterotrophic and aerobic, catalase-positive and oxidase-negative. Phylogenetic analyses based on 16S rRNA gene sequences showed that strain X14M-14T belonged to the family Flavobacteriaceae and formed a distinct lineage that was independent of the most closely related genera: Aequorivita (16S rRNA gene sequence similarities, 91.8-93.1 %) and Salinimicrobium (91.5-92.4 %). Strain X14M-14T contained MK-6 as the major respiratory quinone, and phosphatidylethanolamine and two unknown lipids as the major polar lipids. The major cellular fatty acids were iso-C15 : 0, iso-C15 : 1 G and iso-C17 : 0 3-OH. The presence of iso-C15 : 1 G as a predominant fatty acid could distinguish this strain clearly from the most closely related genera in the family Flavobacteriaceae. The DNA G+C content was 36.6 mol%. On the basis of phenotypic, chemotaxonomic and phylogenetic data, strain X14M-14T represents a novel genus and species of the family Flavobacteriaceae, for which the name Planktosalinus lacus gen. nov., sp. nov. is proposed. The type strain is X14M-14T ( = CGMCC 1.12924T = KCTC 42675T).

Aquisalinus flavus gen. nov., sp. nov., a member of the family Parvularculaceae isolated from a saline lake.

A Gram-stain-negative bacterium, strain D11M-2T, was isolated from a saline lake (Lake Dasugan) in Qaidam basin, Qinghai Province, China. Its taxonomic position was determined by using a polyphasic approach. Cells were non-spore-forming rods, 0.5-0.7 µm wide and 1.2-1.6 µm long, and motile by means of a single subpolar or lateral flagellum. Strain D11M-2T was strictly heterotrophic and aerobic, and catalase- and oxidase-positive. Growth was observed in the presence of 0-14.0% (w/v) NaCl (optimum, 2.0%), and at 10-35 °C (optimum, 30 °C) and pH 6.0-10.5 (optimum, pH 8.0). Strain D11M-2T contained Q-10 and Q-11 as the respiratory quinones and three unknown glycolipids as the major polar lipids. The major cellular fatty acids (>10.0%) were summed feature 8 (C18:1ω7c and/or C18:1ω6c) and C16:0. Phylogenetic analysis based on 16S rRNA gene sequences showed that strain D11M-2T belonged to the family Parvularculaceae and formed a separate lineage that was independent of the two genera within the family Parvularculaceae. Strain D11M-2T exhibited 92.8-93.4% 16S rRNA gene sequence similarity to members of the genus Parvularcula (highest to Parvularcula bermudensis HTCC 2503T), and 90.2% to a member of the genus Amphiplicatus. The DNA G+C content was 59 mol% (Tm). Based on the phenotypic, chemotaxonomic and phylogenetic data, strain D11M-2T is considered to represent a novel species of a new genus in the family Parvularculaceae, for which the name Aquisalinus flavus gen. nov., sp. nov. is proposed. The type strain of Aquisalinus flavus is D11M-2T (=CGMCC 1.12921T=KCTC 42673T).

Prokaryotic Community Structure Driven by Salinity and Ionic Concentrations in Plateau Lakes of the Tibetan Plateau.

The prokaryotic community composition and diversity and the distribution patterns at various taxonomic levels across gradients of salinity and physiochemical properties in the surface waters of seven plateau lakes in the Qaidam Basin, Tibetan Plateau, were evaluated using Illumina MiSeq sequencing. These lakes included Lakes Keluke (salinity, <1 g/liter), Qing (salinity, 5.5 to 6.6 g/liter), Tuosu (salinity, 24 to 35 g/liter), Dasugan (salinity, 30 to 33 g/liter), Gahai (salinity, 92 to 96 g/liter), Xiaochaidan (salinity, 94 to 99 g/liter), and Gasikule (salinity, 317 to 344 g/liter). The communities were dominated by Bacteria in lakes with salinities of <100 g/liter and by Archaea in Lake Gasikule. The clades At12OctB3 and Salinibacter, previously reported only in hypersaline environments, were found in a hyposaline lake (salinity, 5.5 to 6.6 g/liter) at an abundance of ∼1.0%, indicating their ecological plasticity. Salinity and the concentrations of the chemical ions whose concentrations covary with salinity (Mg(2+), K(+), Cl(-), Na(+), SO4 (2-), and Ca(2+)) were found to be the primary environmental factors that directly or indirectly determined the composition and diversity at the level of individual clades as well as entire prokaryotic communities. The distribution patterns of two phyla, five classes, five orders, five families, and three genera were well predicted by salinity. The variation of the prokaryotic community structure also significantly correlated with the dissolved oxygen concentration, pH, the total nitrogen concentration, and the PO4 (3-) concentration. Such correlations varied depending on the taxonomic level, demonstrating the importance of comprehensive correlation analyses at various taxonomic levels in evaluating the effects of environmental variable factors on prokaryotic community structures. Our findings clarify the distribution patterns of the prokaryotic community composition in plateau lakes at the levels of individual clades as well as whole communities along gradients of salinity and ionic concentrations.

Lacimicrobium alkaliphilum gen. nov., sp. nov., a member of the family Alteromonadaceae isolated from a salt lake.

A Gram-stain-negative, facultatively aerobic bacterium, strain X13M-12T, was isolated from a salt lake (Lake Xiaochaidan) in the Qaidam basin, Qinghai Province, PR China. Cells of strain X13M-12T were slightly curved, rod-shaped, 0.5-0.8 µm wide and 1.2-2.3 µm long, and motile by means of a polar flagellum. Strain X13M-12T was catalase- and oxidase-positive. Growth was observed in the presence of 0-15.0 % (w/v) NaCl (optimum 3.0-5.0 %), and at 4-40 °C (optimum 25-30 °C) and pH 6.0-11.0 (optimum pH 8.5). Strain X13M-12T contained Q-8 as the sole respiratory quinone, and phosphatidylglycerol and phosphatidylethanolamine as the major polar lipids. The major cellular fatty acids (>10 % of totals) were C16 : 0, C16 : 1ω7c and/or C16 : 1ω6c, and C18 : 1ω7c and/or C18 : 1ω6c. Phylogenetic analysis, based on 16S rRNA gene sequences, showed that strain X13M-12T belonged to the family Alteromonadaceae and formed a distinct lineage, showing low gene sequence similarities to closely related genera: Bowmanella, Aestuariibacter and Salinimonas (16S rRNA gene sequence similarities, 93.0-93.1 %, 92.3-93.1 % and 92.6-92.7 %, respectively). In addition, strain X13M-12T showed < 92.7 % gene sequence similarities to other species of the family Alteromonadaceae. The DNA G+C content of strain X13M-12T was 49 mol% (Tm). Based on the data presented above, strain X13M-12T is considered to represent a novel genus and species of the family Alteromonadaceae, for which the name Lacimicrobium alkaliphilum gen. nov., sp. nov. is proposed. The type strain is X13M-12T ( = CGMCC 1.12923T = KCTC 42674T).

Psychroflexus salis sp. nov. and Psychroflexus planctonicus sp. nov., isolated from a salt lake.

Two Gram-stain-negative, catalase- and oxidase-positive, strictly aerobic, non-motile, moderately halophilic bacteria (strains X15M-6T and X15M-8T) were isolated from Lake Xiaochaidan, a salt lake in Qaidam basin, Qinghai Province, China. Cells of X15M-6T were rod-like or coccoid, 0.5-0.9 µm wide and 0.9-1.5 µm long; cells of X15M-8T were rods, 0.3-0.6 µm wide and 1.2-2.2 µm long. Growth was observed in the presence of 0.5-14.0 % (w/v) NaCl (optimum, 3.0 %) and at pH 6.5-10.0 (optimum, pH 7.0-7.5) for both. X15M-6T and X15M-8T grew at 10-35 °C (optimum, 20-25 °C) and 4-35 °C (optimum, 25 °C), respectively. Both contained iso-C15 : 0, anteiso-C15 : 0 and iso-C17 : 0 3-OH as the major fatty acids, phosphatidylethanolamine and an unknown lipid as the major polar lipids, and menaquinone MK-6 as the major respiratory quinone. The DNA G+C contents were 32.8 and 35.0 mol% for X15M-6T and X15M-8T, respectively. Phylogenetic trees based on 16S rRNA gene sequences showed that both strains belonged to the genus Psychroflexus and formed a separate lineage. In addition, strains X15M-6T and X15M-8T shared 96.8 % 16S rRNA gene sequence similarity and showed highest similarities to members of the genus Psychroflexus (92.7-93.5 and 91.8-93.1 %, respectively). Based on the above data, it is concluded that strains X15M-6T and X15M-8T represent two novel species of the genus Psychroflexus, for which the names Psychroflexus salis sp. nov. (type strain X15M-6T = CGMCC 1.12925T = JCM 30615T) and Psychroflexus planctonicus sp. nov. (type strain X15M-8T = CGMCC 1.12931T = JCM 30616T) are proposed.

Lacimonas salitolerans gen. nov., sp. nov., isolated from surface water of a saline lake.

A Gram-stain-negative bacterium, strain TS-T30T, was isolated from a saline lake (Lake Tuosu) in Qaidam basin, Qinghai province, China, and its taxonomic position was determined by using a polyphasic approach. Cells were non-spore-forming rods, non-motile, 0.8-1.4 µm wide and 1.9-4.0 µm long. Strain TS-T30T was strictly heterotrophic and aerobic. Catalase- and oxidase-positive. Growth was observed in the presence of 0.5-11.0 % (w/v) NaCl (optimum 3.0 %), and at 10-35 °C (optimum 25 °C) and pH 6.5-10.0 (optimum pH 8.5). Strain TS-T30T contained C18 : 1ω7c as the only predominant fatty acid. The major respiratory quinone was Q-10. The DNA G+C content was 62 mol% (Tm). Phylogenetic trees based on 16S rRNA gene sequences showed that strain TS-T30T formed a distinct lineage that was independent of other most closely related genera: Lutimaribacter (95.2-95.9 % 16S rRNA gene sequence similarities), Poseidonocella (95.4 %), Ruegeria (92.8-94.9 %), Marivita (93.6-94.9 %), Seohaeicola (94.7 %), Sediminimonas (94.7 %), Shimia (93.9-94.7 %), Oceanicola (92.6-94.5 %) and Roseicyclus (94.5 %). The major polar lipids were phosphatidylglycerol, one unidentified phospholipid and an unknown aminolipid; phosphatidylcholine was not detected. These data demonstrated that strain TS-T30T represents a novel species of a new genus in the family Rhodobacteraceae, for which the name Lacimonas salitolerans gen. nov., sp. nov. is proposed. The type strain of the type species is TS-T30T ( = CGMCC 1.12477T = NBRC 110969T).