Developing a population-state decision system for intelligently reprogramming extracellular electron transfer in Shewanella oneidensis.

The unique extracellular electron transfer (EET) ability has positioned electroactive bacteria (EAB) as a major class of cellular chassis for genetic engineering aimed at favorable environmental, energy, and geoscience applications. However, previous efforts to genetically enhance EET ability have often impaired the basal metabolism and cellular growth due to the competition for the limited cellular resource. Here, we design a quorum sensing-based population-state decision (PSD) system for intelligently reprogramming the EET regulation system, which allows the rebalanced allocation of the cellular resource upon the bacterial growth state. We demonstrate that the electron output from Shewanella oneidensis MR-1 could be greatly enhanced by the PSD system via shifting the dominant metabolic flux from initial bacterial growth to subsequent EET enhancement (i.e., after reaching a certain population-state threshold). The strain engineered with this system achieved up to 4.8-fold EET enhancement and exhibited a substantially improved pollutant reduction ability, increasing the reduction efficiencies of methyl orange and hexavalent chromium by 18.8- and 5.5-fold, respectively. Moreover, the PSD system outcompeted the constant expression system in managing EET enhancement, resulting in considerably enhanced electron output and pollutant bioreduction capability. The PSD system provides a powerful tool for intelligently managing extracellular electron transfer and may inspire the development of new-generation smart bioelectrical devices for various applications.

A designed plasmid-transition strategy enables rapid construction of robust and versatile synthetic exoelectrogens for environmental applications.

Genetic engineering is promising to expand the application scope of exoelectrogens in energy and environmental applications and plasmid vectors, as one type of fundamental tool, are intensively applied. Antibiotics are widely utilized for plasmid selection and maintenance; however, their utilization suffers from environmental concerns on the spread of resistance genes, elevated costs, inevitable genotypic instability and phenotypic heterogeneity. In this work, we establish an auxotrophic complementation system for stable plasmid maintenance without antibiotic association, in Shewanella oneidensis, an attractive model exoelectrogen. A plasmid-transition strategy is designed to facilitate the rapid and efficient construction of the auxotrophic complementation system. Such a system not only enables the same intensive gene expression as the conventional antibiotic-associated plasmid system but also exhibits remarkably superior robustness and stability. With this system, the menaquinone pool of the extracellular respiratory chain is enhanced first independently and further synergized by engineering the Mtr conduit, leading to significantly promoted extracellular electron transfer (EET) outputs (up to 10.33- and 2.97-fold improvement in the maximum current density and the maximum output voltage) and heavy metal Cr(VI) reduction ability (5.15-fold improvement). This work provides a robust and stable platform to engineer exoelectrogens for environmental applications.

Rapid and highly efficient genomic engineering with a novel iEditing device for programming versatile extracellular electron transfer of electroactive bacteria.

The advances in synthetic biology bring exciting new opportunities to reprogram microorganisms with novel functionalities for environmental applications. For real-world applications, a genetic tool that enables genetic engineering in a stably genomic inherited manner is greatly desired. In this work, we design a novel genetic device for rapid and efficient genome engineering based on the intron-encoded homing-endonuclease empowered genome editing (iEditing). The iEditing device enables rapid and efficient genome engineering in Shewanella oneidensis MR-1, the representative strain of the electroactive bacteria group. Moreover, combining with the Red or RecET recombination system, the genome-editing efficiency was greatly improved, up to approximately 100%. Significantly, the iEditing device itself is eliminated simultaneously when genome editing occurs, thereby requiring no follow-up to remove the encoding system. Then, we develop a new extracellular electron transfer (EET) engineering strategy by programming the parallel EET systems to enhance versatile EET. The engineered strains exhibit sufficiently enhanced electron output and pollutant reduction ability. Furthermore, this device has demonstrated its great potential to be extended for genome editing in other important microbes. This work provides a useful and efficient tool for the rapid generation of synthetic microorganisms for various environmental applications.

Enhanced Bioreduction of Radionuclides by Driving Microbial Extracellular Electron Pumping with an Engineered CRISPR Platform.

Dissimilatory metal-reducing bacteria (DMRB) with extracellular electron transfer (EET) capability show great potential in bioremediating the subsurface environments contaminated by uranium through bioreduction and precipitation of hexavalent uranium [U(VI)]. However, the low EET efficiency of DMRB remains a bottleneck for their applications. Herein, we develop an engineered CRISPR platform to drive the extracellular electron pumping of Shewanella oneidensis, a representative DMRB species widely present in aquatic environments. The CRISPR platform allows for highly efficient and multiplex genome editing and rapid platform elimination post-editing in S. oneidensis. Enabled by such a platform, a genomic promoter engineering strategy (GPS) for genome-widely engineering the EET-encoding gene network was established. The production of electron conductive Mtr complex, synthesis of electron shuttle flavin, and generation of NADH as intracellular electron carrier are globally optimized and promoted, leading to a significantly enhanced EET ability. Applied to U(VI) bioreduction, the edited strains achieve up to 3.62-fold higher reduction capacity over the control. Our work endows DMRB with an enhanced ability to remediate the radionuclides-contaminated environments and provides a gene editing approach to handle the growing environmental challenges of radionuclide contaminations.

Rediverting Electron Flux with an Engineered CRISPR-ddAsCpf1 System to Enhance the Pollutant Degradation Capacity of Shewanella oneidensis.

Pursuing efficient approaches to promote the extracellular electron transfer (EET) of extracellular respiratory bacteria is essential to their application in environmental remediation and waste treatment. Here, we report a new strategy of tuning electron flux by clustered regularly interspaced short palindromic repeat (CRISPR)-ddAsCpf1-based rediverting (namely STAR) to enhance the EET capacity of Shewanella oneidensis MR-1, a model extracellular respiratory bacterium widely present in the environment. The developed CRISPR-ddAsCpf1 system enabled approximately 100% gene repression with the green fluorescent protein (GFP) as a reporter. Using a WO3 probe, 10 representative genes encoding for putative competitive electron transfer proteins were screened, among which 7 genes were identified as valid targets for EET enhancement. Repressing the valid genes not only increased the transcription level of the l-lactate metabolism genes but also affected the genes involved in direct and indirect EET. Increased riboflavin production was also observed. The feasibility of this strategy to enhance the bioreduction of methyl orange, an organic pollutant, and chromium, a typical heavy metal, was demonstrated. This work implies a great potential of the STAR strategy with the CIRPSR-ddAsCpf1 system for enhancing bacterial EET to favor more efficient environmental remediation applications.

Abundance and diversity of iron reducing bacteria communities in the sediments of a heavily polluted freshwater lake.

Iron reduction mediated by Fe(III)-reducing bacteria (FeRB) occurs in aqueous environments and plays an essential role in removing contaminates in polluted freshwater lakes. Two model FeRB species, Shewanella and Geobacter, have been intensively studied because of their functions in bioremediation, iron reduction, and bioelectricity production. However, the abundance and community diversity of Shewanella and Geobacter in eutrophic freshwater lakes remain largely unknown. In this work, the distribution, abundance and biodiversity of Shewanella, Geobacter and other FeRB in the sediments of a heavily polluted lake, Chaohu Lake, China, across four successive seasons were investigated. Shewanella, Geobacter, and other FeRB were found to be widely distributed in the sediment of this heavily eutrophic lake. Geobacter was abundant with at least one order of magnitude more than Shewanella in cold seasons. Three Shewanella-related operational taxonomic units were detected and sixty one Geobacter-related operational taxonomic units were grouped into three phylogenetic clades. Thiobacillus, Desulfuromonas and Geobacter were identified as the main members of FeRB in the lake sediments. Interestingly, nutrients like carbon, nitrogen, and phosphorus were found to be the key factors governing the abundance and diversity of FeRB. Total FeRB, as well as Geobacter and Shewanella, were more abundant in the heavily eutrophic zone than those in the lightly eutrophic zone. The abundance and diversity of FeRB in the sediments of freshwater lakes were highly related with the degree of eutrophication, which imply that FeRB might have a great potential in alleviating the eutrophication and contamination in aqueous environments.

Identification of Varieties of Dried Red Jujubes with Near-Infrared Hyperspectral Imaging.

In order to realize rapid identification of dried red jujubes, this paper proposes a method based on near-infrared hyperspectral imaging technology. The near-infrared hyperspectral images (1 000～1 600 nm) of 240 samples in total from 4 cultivars of dried red jujubes will be acquired. The samples are to be divided into the calibration set and the prediction set in the ratio of 2∶1. 7, 8, 10 effective wavelengths are to be selected by principal component analysis(PCA), x-loading weight(x-LW)and successive projection algorithm(SPA) respectively. The dimensionality of original hyperspectral images will be reduced with PCA, and texture features of the first principal component image are to be extracted with gray-level co-occurrence matrix(GLCM).The partial least squares-discriminant analysis(PLS-DA), back propagation neural network(BPNN)and least square support vector machine(LS-SVM) are to be applied to build identification models with the selected effective wavelengths, texture features and fusion of the former two features. The identification rates of the models based on fusion features will be higher than those of models based on the spectral features or texture features respectively. The BPNN models based on the fusion features will obtain the best results, whose identification rates of prediction set are to be 100%. The results in this paper indicate that the near-infrared hyperspectral imaging technology has great potential to identify the dried red jujubes rapidly.

Non-suicidal self-injury in adolescents with mood disorders and the roles of self-compassion and emotional regulation.

Objective: We aimed to investigate the characteristics and psychological mechanism of non-suicidal self-injury (NSSI) in adolescents with mood disorders. We examined how self-compassion and emotional regulation affected NSSI and tested the mediating role of self-compassion in the link between emotional regulation and NSSI. Method: We recruited outpatient and inpatient adolescent patients with bipolar and related disorders or depressive disorders (DSM-5), with a focus on NSSI. We also recruited healthy controls from the community. We collected demographic and clinical data. The Adolescent Self-injury Questionnaire, Self-compassion Scale (SCS), and Emotion Regulation Questionnaire (ERQ) were used to assess the frequency and severity of NSSI, level of self-compassion, and emotional regulation. Results: In total, we recruited 248 adolescent patients with mood disorders (N = 196 with NSSI, and 52 without NSSI) and 212 healthy controls. NSSI was significantly associated with the female sex, lower levels of education and less use of cognitive reappraisal strategies, lower levels of self-warmth, and higher levels of self-coldness. Multivariate analysis of variance showed that there were significant differences in the scores of ERQ, cognitive reassessment score, and the scores of SCS among the three groups, but no statistical differences in expressive suppression score among the three groups. Self-warmth had a mediating effect between cognitive reappraisal and NSSI behavior. Conclusion: NSSI is prevalent among adolescent patients with mood disorders in clinical settings, especially among girls and those with lower levels of education and less cognitive reappraisal strategies. More clinical attention is needed. Self-compassion and its factors may mediate the association between emotional regulation and NSSI. Clinical implications and future research directions were discussed.

Pharmacokinetic and pharmacodynamic profiles of recombinant human erythropoietin-loaded poly(lactic-co-glycolic acid) microspheres in rats.

AIM: To characterize the pharmacokinetic and pharmacodynamic profiles of the recombinant human erythropoietin (rhEPO)-loaded poly(lactic-co-glycolic acid) (PLGA) microspheres in rats. METHODS: The rhEPO-loaded microspheres were prepared using a solid-in-oil-in-water emulsion method. Pharmacokinetics and pharmacodynamics of the rhEPO-loaded microspheres were evaluated in male Sprague-Dawley rats. The serum rhEPO level was determined with ELISA. The level of anti-rhEPO antibody in the serum was measured to assess the immunogenicity of rhEPO released from the microspheres. RESULTS: rhEPO was almost completely released from the PLGA microspheres in vitro, following zero-order release kinetics over approximately 30 d. After intramuscular injection (10,000 or 30,000 IU rhEPO/kg) in the rats, the serum rhEPO concentration reached maximum levels on d 1, then decreased gradually and was maintained at nearly steady levels for approximately 4 weeks. Furthermore, the release of rhEPO from the PLGA microspheres was found to be controlled mainly by a dissolution/diffusion mechanism. A good linear correlation (R(2)=0.98) was obtained between the in vitro and in vivo release data. A single intramuscular injection of the rhEPO-loaded PLGA microspheres (10,000 or 30,000 IU rhEPO/kg) in the rats resulted in elevated hemoglobin and red blood cell concentrations for more than 28 d. Moreover, the immunogenicity of rhEPO released from the PLGA microspheres was comparable with that of the unencapsulated rhEPO. CONCLUSION: The results prove the feasibility of using the PLGA-based microspheres to deliver rhEPO for approximately 1 month.

Extracellular pollutant degradation feedback regulates intracellular electron transfer process of exoelectrogens: Strategy and mechanism.

Exoelectrogens possess extraordinary degradation ability to various pollutants through extracellular electron transfer (EET). Compared with extracellular electron release process, intracellular electron transfer network is not yet fully recognized. Especially, controversy remains regarding the role of CymA, an essential electron-transfer hub of Shewanella oneidensis MR-1, in EET process. In this study, we thoroughly surveyed the intracellular transfer strategies during EET through dye decolorization. Loss of CymA severely impaired the reduction ability of S. oneidensis MR-1 to methyl orange (MO), but hardly affected the decolorization of aniline blue (AB). Complement of cymA fully restored the MO decolorization ability of ΔcymA mutant. The contribution of CymA to extracellular decolorization was subjected to MO concentrations. The defect in the decolorization ability of ΔcymA mutant was not evident at low MO concentration, but severe at high MO concentration. Further investigation revealed that EET rate determined the significance of CymA in the extracellular bioremediation by S. oneidensis MR-1. Coupled with MO concentrations increasing from 15 to 120 mg/L, the initial electron transfer rates of S. oneidensis MR-1 increased accordingly from 2.69 × 104 to 11.21 × 104 electrons CFU-1 s-1, which led to a gradual increase of the dependencyCymA. Thus, we first revealed that extracellular degradation performance could feedback regulate the intracellular electron transfer process of S. oneidensis MR-1. This work is helpful to fully understand the complex EET process of exoelectrogens and facilitates the application of exoelectrogens in bioremediation of environmental pollutants.

Re-evaluation of the environmental hazards of nZnO to denitrification: Performance and mechanism.

Numerous studies have shown that zinc oxide nanoparticles (nZnO) have an inhibitory effect on wastewater biotreatment, where doses exceeding ambient concentrations are used. However, the effect of ambient concentrations of ZnO (<1 mg/L) on anaerobic digestion processes is not clear. Herein, this study comprehensively explored the impact of nZnO on the denitrification performance and core microbial community of activated sludge under ambient concentrations. Results showed that only 0.075 mg/L nZnO had shown a beneficial effect on nitrogen removal by activated sludge. When nZnO concentration reached 0.75 mg/L, significant enhancement of nitrate reduction and mitigation of nitrite accumulation were observed, indicating a remarkable stimulatory effect on nitrogen removal. Simultaneously, nZnO could weaken the sludge surface charge and improve the secretion of extracellular polymeric substances, thus enhancing sludge flocculation for denitrification. Microbial community analysis revealed that nZnO exposure increased the relative abundance of denitrifying bacteria, which could contribute to the reinforcement of traditional denitrification. Furthermore, exogenous addition of NH4+ significantly inhibited the accumulation of nitrite, implying that nZnO had a potential to improve the denitrification process via a partial denitrification-anammox pathway. Considering current ambient concentration, the stimulatory effect shown in our work may better represent the actual behavior of ZnO in wastewater biotreatment.

[Modification and luminescence properties of transparent Lu2SiO5 : Ce3+ thin-film phosphors].

To achieve high-spatial-resolution for X-ray imaging and flat panel display, transparent thin-film phosphors have been attracted much attention in recent years. In comparison with conventional powder phosphors, the transparent thin-film phosphors have some outstanding advantages such as high contrast and resolution, superior thermal conductivity and better adhesion. Cerium-doped lutetium oxyorthosilicate Lu2 SiO5 (LSO) is one promising candidate due to its high density (7.4 g x cm(-3)), high light yield (27 300 photons x MeV(-1)), short decay time (40 ns), and excellent chemical stability. The sol-gel method is one of the most important techniques for deposition of functional thin films, because it possesses a number of advantages over conventional film formation techniques, such as low processing temperature, easy coating of large surfaces, homogenous multicomponent films, and cheap equipments. In X-ray imaging application, the thickness of the thin-film phosphor is the most important factor, which can increase X-ray absorption of the film and then strengthen its luminescence intensity. In the present work, transparent LSO : Ce film was successfully prepared using sol-gel method and spin-coating technique by using inorganic salts as raw materials, 2-methoxyethanol as solvent, and poly (ethylene glycol) (PEG) as modifier without inert atmosphere. The effect of PEG on the luminescence properties of the film was investigated in detail. The results indicated that PEG200 played an important role in the formation of LSO : Ce film, improving its quality and luminescent intensity. The film thickness of 0.9 microm was achieved after 5 times of coating. The luminescence properties of the film were studied. Their performances were good, which implied that the film would have promising applications in high-spatial-resolution X-ray imaging and flat panel display devices.

Factors Associated With Non-suicidal Self-Injury in Chinese Adolescents: A Meta-Analysis.

Non-suicidal self-injury (NSSI) in adolescents is an increasing public health problem in China. We conducted a meta-analysis of studies on NSSI in Chinese adolescents (between 10 and 19 years) to examine factors associated with NSSI. Twenty-five papers including 30 separate studies with 186,447 participants were included for analysis. The results from a random-effects model showed a weak, but significant overall prediction of NSSI (OR = 1.734). There were significant associations between the following seven factors and NSSI (ranking by the effect sizes, in descending order): adverse life events (OR = 2.284), negative coping style (OR = 2.040), problematic internet use (OR = 2.023), sleep disturbance (OR = 1.734), traumatic experiences (OR = 1.728), problematic parent-child relationship (OR = 1.585), mental health problems (OR = 1.578). Additionally, NSSI sample type moderated these effects. These findings highlight factors significantly associated with NSSI in Chinese adolescents. Parent-child relationship and mental health of the only children and left-behind children in China deserve more attention. Systematic Review Registration: https://www.crd.york.ac.uk/prospero/display_record.php?ID=CRD42019123508.

Evaluation of antibacterial activities of silver nanoparticles on culturability and cell viability of Escherichia coli.

Nanoparticles released into the environment are attracting increasing concern because of their potential toxic effects. Conventional methods for assessing the toxicity of nanoparticles are usually confined to cultivable cells, but not applicable to viable but non-culturable (VBNC) cells. However, it remains unknown whether silver nanoparticles (AgNPs), a typical antimicrobial agent, could induce bacteria into a VBNC state in natural environments. In this work, the viability of E. coli, an indicator bacterium widely used for assessing the antibacterial activity of AgNPs, was examined through coupling plate counting, fluorescence staining and adenosine triphosphate (ATP) production. AgNPs were found to have a considerable antibacterial ability, which resulted in less than 0.0004% of culturable cells on plates. However, more than 80% of the cells still maintained their cell membrane integrity under the stress of 80 mg/L AgNPs. Meanwhile, the residue of ATP production (0.6%) was 1500 times higher than that of the culturable cells (< 0.0004%). These results clearly demonstrate that when exposed to AgNPs, most of cells fell into a VBNC state, instead of dying. Environmental factors, e.g., Cl- and illumination, which could change the dissolution, hydrophilicity and zeta potential of AgNPs, eventually influenced the culturability of E. coli. Inhibition of dissolved Ag+ and reactive oxygen species was found to facilitate the mitigation of the strain into a VBNC state. Our findings suggest the necessity of re-evaluating the environmental effects and antibacterial activities of AgNPs.

Mercury/silver resistance genes and their association with antibiotic resistance genes and microbial community in a municipal wastewater treatment plant.

Municipal wastewater treatment plants (WWTPs) are an important reservoir for heavy metal (e.g., Hg and Ag) resistance genes and antibiotic resistance genes (ARGs). However, current knowledge on Hg/Ag resistance genes and their association with ARGs in WWTPs remains largely unknown. In this study, the fates of five Hg/Ag resistance genes (merB, merD, merR, silE, and silR), five ARGs (sulI, sulII, tetO, tetQ, tetW), and class 1 integrase (intI1) in a WWTP were investigated. Results show that the absolute abundances of all target genes were greatly reduced through the treatment systems. The dynamics of merB, merD and silE were significantly correlated with tetW and sulII. Based on network analysis, Hg/Ag resistance genes might share the same microbial hosts with tetQ and tetW, implying the potential importance of Hg/Ag in ARGs evolution and spread. These findings advanced our understanding of the occurrence of Hg/Ag resistance genes and ARGs in WWTPs.

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.

Ultrasound-Enhanced Subcritical Fluid Extraction of Essential Oil from Nymphaea alba var and Its Antioxidant Activity.

Background: The essential oil content of the water lily is extremely low; thus, finding a new method that can extract essential oil from water lilies with a high extraction rate and no residual organic solvents is essential. Objective: The optimal processing conditions for the ultrasound-enhanced subcritical fluid extraction of essential oil from Nymphaea alba var (red water lily) and the antioxidant activity of the essential oil in vitro are investigated to provide theoretical bases for identification and development. Methods: Single-factor experiments and orthogonal designs are performed to determine the effects of extraction conditions on essential oil yields. The chemical composition of essential oil is analyzed using GC-MS. Results: The optimum extraction parameters are established as follows: extraction temperature, 35°C; extraction time, 30 min/time for four times; ratio of material to liquid, 1:3; ultrasound power, 250 W/L; and ultrasonic frequency, 20 kHz. The extraction rate of essential oil is 0.315% under these conditions. Eleven components comprise more than 1% content. The main chemical constituents are 8-hexadecyne (31.04%) and 2,6,10-trimethyl-tetradecane (3.95%). The essential oil from N. alba var has an antioxidant activity in vitro; however, its antioxidant activity is weaker than that of butylated hydroxytoluene. Conclusions: Subcritical fluid is suitable for the extraction of essential oil from N. alba var, and the essential oil has a good antioxidant activity. Highlights: The essential oil content of N. alba var is 0.315%. Forty-seven chemical constituents are identified and isolated from N. alba var and analyzed by GC-MS.

An isolable catenane consisting of two Möbius conjugated nanohoops.

Besides its mathematical importance, the Möbius topology (twisted, single-sided strip) is intriguing at the molecular level, as it features structural elegance and distinct properties; however, it carries synthetic challenges. Although some Möbius-type molecules have been isolated by synthetic chemists accompanied by extensive computational studies, the design, preparation, and characterization of stable Möbius-conjugated molecules remain a nontrivial task to date, let alone that of molecular Möbius strips assembling into more complex topologies. Here we report the efficient synthesis, crystal structure, and theoretical study of a catenane consisting of two fully conjugated nanohoops exhibiting Möbius topology in the solid state. This work highlights that oligoparaphenylene-derived nanohoops, a family of highly warped and synthetically challenging conjugated macrocycles, can not only serve as building blocks for interlocked supermolecular structures, but also represent a new class of compounds with isolable Möbius conformations stabilized by non-covalent interactions.

Estimates of abundance and diversity of Shewanella genus in natural and engineered aqueous environments with newly designed primers.

Shewanella species have a diverse respiratory ability and wide distribution in environments and play an important role in bioremediation and the biogeochemical cycles of elements. Primers with more accuracy and broader coverage are required with consideration of the increasing number of Shewanella species and evaluation of their roles in various environments. In this work, a new primer set of 640F/815R was developed to quantify the abundance of Shewanella species in natural and engineered environments. In silico tools for primer evaluation, quantitative polymerase chain reaction (qPCR) and clone library results showed that 640F/815R had a higher specificity and coverage than the previous primers in quantitative analysis of Shewanella. Another newly developed primer pair of 211F/815cR was also adopted to analyze the Shewanella diversity and demonstrated to be the best candidate in terms of specificity and coverage. We detected more Shewanella-related species in freshwater environments and found them to be substantially different from those in marine environments. Abundance and diversity of Shewanella species in wastewater treatment plants were largely affected by the process and operating conditions. Overall, this study suggests that investigations of abundance and diversity of Shewanella in various environments are of great importance to evaluate their ecophysiology and potential ecological roles.

Haplotypes Phased from Population Transcriptomes Detecting Selection in the Initial Adaptation of Miscanthus lutarioriparius to Stressful Environments.

Adaptation is a characteristic that enhances the survival or reproduction of organisms; selection is the critical process leading to adaptive evolution. Therefore, detecting selection is important in studying evolutionary biology. Changes in allele frequency are fundamental to adaptive evolution. The allele frequency of entire genes at the genomic scale is more intensive and precise for analyzing selection effects, compared with simple sequence repeat and single nucleotide polymorphism (SNP) alleles from nuclear gene fragments. Here, we analyzed 29,094 SNPs derived from 80 individuals of 14 L. Liou ex S.L. Chen & Renvoize populations planted near their native habitat (Jiangxia, Hubei Province, JH) and a stressful environment (Qingyang, Gansu Province, QG) to detect selection during initial adaptation. The nucleotide diversity of over 60% of genes was decreased in QG compared with JH, suggesting that most genes were undergoing selection in the stressful environment. We explored a new approach based on haplotype data inferred from RNA-seq data to analyze the change in frequency between two sites and to detect selection signals. In total, 402 and 51 genes were found to be targets of positive and negative selection, respectively. Among these candidate genes, the enrichment of abiotic stress-response genes and photosynthesis-related genes might have been responsible for establishment in the stressful environment. This is the first study assessing the change in allele frequency at the genomic level during adaptation. The method in which allele frequency detects selection during initial adaptation using population RNA-seq data would be useful for developing evolutionary biology.