High-throughput identification of regulatory elements and functional assays to uncover susceptibility genes for nasopharyngeal carcinoma.

Large-scale genetic association studies have identified multiple susceptibility loci for nasopharyngeal carcinoma (NPC), but the underlying biological mechanisms remain to be explored. To gain insights into the genetic etiology of NPC, we conducted a follow-up study encompassing 6,907 cases and 10,472 controls and identified two additional NPC susceptibility loci, 9q22.33 (rs1867277; OR = 0.74, 95% CI = 0.68-0.81, p = 3.08 × 10-11) and 17q12 (rs226241; OR = 1.42, 95% CI = 1.26-1.60, p = 1.62 × 10-8). The two additional loci, together with two previously reported genome-wide significant loci, 5p15.33 and 9p21.3, were investigated by high-throughput sequencing for chromatin accessibility, histone modification, and promoter capture Hi-C (PCHi-C) profiling. Using luciferase reporter assays and CRISPR interference (CRISPRi) to validate the functional profiling, we identified PHF2 at locus 9q22.33 as a susceptibility gene. PHF2 encodes a histone demethylase and acts as a tumor suppressor. The risk alleles of the functional SNPs reduced the expression of the target gene PHF2 by inhibiting the enhancer activity of its long-range (4.3 Mb) cis-regulatory element, which promoted proliferation of NPC cells. In addition, we identified CDKN2B-AS1 as a susceptibility gene at locus 9p21.3, and the NPC risk allele of the functional SNP rs2069418 promoted the expression of CDKN2B-AS1 by increasing its enhancer activity. The overexpression of CDKN2B-AS1 facilitated proliferation of NPC cells. In summary, we identified functional SNPs and NPC susceptibility genes, which provides additional explanations for the genetic association signals and helps to uncover the underlying genetic etiology of NPC development.

Deciphering antifungal and antibiofilm mechanisms of isobavachalcone against Cryptococcus neoformans through RNA-seq and functional analyses.

Cryptococcus neoformans has been designated as critical fungal pathogens by the World Health Organization, mainly due to limited treatment options and the prevalence of antifungal resistance. Consequently, the utilization of novel antifungal agents is crucial for the effective treatment of C. neoformans infections. This study exposed that the minimum inhibitory concentration (MIC) of isobavachalcone (IBC) against C. neoformans H99 was 8 µg/mL, and IBC dispersed 48-h mature biofilms by affecting cell viability at 16 µg/mL. The antifungal efficacy of IBC was further validated through microscopic observations using specific dyes and in vitro assays, which confirmed the disruption of cell wall/membrane integrity. RNA-Seq analysis was employed to decipher the effect of IBC on the C. neoformans H99 transcriptomic profiles. Real-time quantitative reverse transcription PCR (RT-qPCR) analysis was performed to validate the transcriptomic data and identify the differentially expressed genes. The results showed that IBC exhibited various mechanisms to impede the growth, biofilm formation, and virulence of C. neoformans H99 by modulating multiple dysregulated pathways related to cell wall/membrane, drug resistance, apoptosis, and mitochondrial homeostasis. The transcriptomic findings were corroborated by the antioxidant analyses, antifungal drug sensitivity, molecular docking, capsule, and melanin assays. In vivo antifungal activity analysis demonstrated that IBC extended the lifespan of C. neoformans-infected Caenorhabditis elegans. Overall, the current study unveiled that IBC targeted multiple pathways simultaneously to inhibit growth significantly, biofilm formation, and virulence, as well as to disperse mature biofilms of C. neoformans H99 and induce cell death.

Discovery of psoralen as a quorum sensing inhibitor suppresses Pseudomonas aeruginosa virulence.

Pseudomonas aeruginosa is a common opportunistic pathogen with growing resistance and presents heightened treatment challenges. Quorum sensing (QS) is a cell-to-cell communication system that contributes to the production of a variety of virulence factors and is also related to biofilm formation of P. aeruginosa. Compared to traditional antibiotics which kill bacteria directly, the anti-virulence strategy by targeting QS is a promising strategy for combating pseudomonal infections. In this study, the QS inhibition potential of the compounds derived from the Traditional Chinese Medicines was evaluated by using in silico, in vitro, and in vivo analyses. The results showed that psoralen, a natural furocoumarin compound derived from Psoralea corylifolia L., was capable of simultaneously inhibiting the three main QS regulators, LasR, RhlR, and PqsR of P. aeruginosa. Psoralen had no bactericidal activity but could widely inhibit the production of extracellular proteases, pyocyanin, and biofilm, and the cell motilities of the model and clinical P. aeruginosa strains. RNA-sequencing and quantitative PCR analyses further demonstrated that a majority of QS-activated genes in P. aeruginosa were suppressed by psoralen. The supplementation of psoralen could protect Caenorhabditis elegans from P. aeruginosa challenge, especially for the hypervirulent strain PA14. Moreover, psoralen showed synergistic antibacterial effects with polymyxin B, levofloxacin, and kanamycin. In conclusions, this study identifies the anti-QS and antibiofilm effects of psoralen against P. aeruginosa strains and sheds light on the discovery of anti-pseudomonal drugs among Traditional Chinese Medicines. KEY POINTS: • Psoralen derived from Psoralea corylifolia L. inhibits the virulence-related phenotypes of P. aeruginosa. • Psoralen simultaneously targets the three core regulators of P. aeruginosa QS system and inhibits the expression of a large part of downstream genes. • Psoralen protects C. elegans from P. aeruginosa challenge and enhances the susceptibility of P. aeruginosa to antibiotics.

Deciphering the role of micro/nano-hydroxyapatite in aerobic granular sludge system: Effects on treatment performance and enhancement mechanism.

Hydroxyapatite (HAP), a mineral nucleus identified within aerobic granular sludge (AGS), plays a vital role in enhancing the AGS systems. However, the microscopic mechanism underlying their roles remains largely unexplored. Herein, a systematic investigation was carried out to elucidate the impact and enhanced mechanisms associated with HAP of different sizes, i.e. micro-HAP (mHAP) and nano-HAP (nHAP), on the aerobic granulation, nutrient removal and microbial diversity of AGS. Results showed that the presence of nHAP and mHAP significantly shortened the granulation process to 15 and 20 days, respectively. This might be ascribed to the fact that the large specific surface area of nHAP aggregates was conducive to microbial adhesion, biomass accumulation and sludge granulation. Compared with mHAP, the granules with nHAP showed better settlement performance, mechanical strength and larger diameter. The X-ray diffraction (XRD) and Raman spectrometer analysis confirmed the presence of HAP within the granules, which was found to stimulate the secretion of extracellular polymeric substance, improve the compactness of granule structure and suppress the growth of filamentous bacteria, thereby contributing to a stable AGS system. The presence of HAP, especially nHAP, effectively enriched the functional microorganisms, such as nitrifying and denitrifying bacteria (e.g. Candidatus_Competibacter) and phosphorus accumulating organisms (e.g. Flavobacterium), leading to the improved nutrient removal efficiencies (COD > 96%, TN > 76%, and TP > 74%). Further analysis revealed the up-regulation of functional enzymes (e.g. nitrite oxidoreductase and polyphosphate kinase) involved in nutrient metabolism, underlying the inherent mechanisms for the excellent nutrient removal. This study deepens the understanding of granulation mechanisms from the perspective of mineral cores, and proposes an economically feasible strategy for rapid initiation and stabilization of AGS reactors.

A deep transcriptome meta-analysis reveals sex differences in multiple sclerosis.

BACKGROUND: Multiple sclerosis (MS), a chronic auto-immune, inflammatory, and degenerative disease of the central nervous system, affects both males and females; however, females suffer from a higher risk of developing MS (2-3:1 ratio relative to males). The precise sex-based factors influencing risk of MS are currently unknown. Here, we explore the role of sex in MS to identify molecular mechanisms underlying observed MS sex differences that may guide novel therapeutic approaches tailored for males or females. METHODS: We performed a rigorous and systematic review of genome-wide transcriptome studies of MS that included patient sex data in the Gene Expression Omnibus and ArrayExpress databases following PRISMA statement guidelines. For each selected study, we analyzed differential gene expression to explore the impact of the disease in females (IDF), in males (IDM) and our main goal: the sex differential impact of the disease (SDID). Then, for each scenario (IDF, IDM and SDID) we performed 2 meta-analyses in the main tissues involved in the disease (brain and blood). Finally, we performed a gene set analysis in brain tissue, in which a higher number of genes were dysregulated, to characterize sex differences in biological pathways. RESULTS: After screening 122 publications, the systematic review provided a selection of 9 studies (5 in blood and 4 in brain tissue) with a total of 474 samples (189 females with MS and 109 control females; 82 males with MS and 94 control males). Blood and brain tissue meta-analyses identified, respectively, 1 (KIR2DL3) and 13 (ARL17B, CECR7, CEP78, IFFO2, LOC401127, NUDT18, RNF10, SLC17A5, STMP1, TRAF3IP2-AS1, UBXN2B, ZNF117, ZNF488) MS-associated genes that differed between males and females (SDID comparison). Functional analyses in the brain revealed different altered immune patterns in females and males (IDF and IDM comparisons). The pro-inflammatory environment and innate immune responses related to myeloid lineage appear to be more affected in females, while adaptive responses associated with the lymphocyte lineage in males. Additionally, females with MS displayed alterations in mitochondrial respiratory chain complexes, purine, and glutamate metabolism, while MS males displayed alterations in stress response to metal ion, amine, and amino acid transport. CONCLUSION: We found transcriptomic and functional differences between MS males and MS females (especially in the immune system), which may support the development of new sex-based research of this disease. Our study highlights the importance of understanding the role of biological sex in MS to guide a more personalized medicine.

Synergism, Bifunctionality, and the Evolution of a Gradual Sensory Trade-off in Hummingbird Taste Receptors.

Sensory receptor evolution can imply trade-offs between ligands, but the extent to which such trade-offs occur and the underlying processes shaping their evolution is not well understood. For example, hummingbirds have repurposed their ancestral savory receptor (T1R1-T1R3) to detect sugars, but the impact of this sensory shift on amino acid perception is unclear. Here, we use functional and behavioral approaches to show that the hummingbird T1R1-T1R3 acts as a bifunctional receptor responsive to both sugars and amino acids. Our comparative analyses reveal substantial functional diversity across the hummingbird radiation and suggest an evolutionary timeline for T1R1-T1R3 retuning. Finally, we identify a novel form of synergism between sugars and amino acids in vertebrate taste receptors. This work uncovers an unexplored axis of sensory diversity, suggesting new ways in which nectar chemistry and pollinator preferences can coevolve.

Microbiome assembly mechanism and functional potential in enhanced biological phosphorus removal system enriched with Tetrasphaera-related polyphosphate accumulating organisms.

Tetrasphaera-related polyphosphate accumulating organisms (PAOs) are the key functional guilds for enhanced biological phosphorus removal (EBPR) systems. Their biomass enrichment can be enhanced by the nitrification inhibitor allylthiourea (ATU). However, the underlying assembly mechanism and the functional potential of the EBPR microbiome regulated by ATU are unclear. This study investigates the effect of ATU on microbiome assembly and functional potential by closely following the microbiota dynamics in an EBPR system enriched with Tetrasphaera-related PAOs for 288-days before, during and after ATU addition. The results showed that ATU addition increased microbiota structural similarity and compositional convergence, and enhanced determinism in the assembly of EBPR microbiome. During exposure to ATU, Tetrasphaera-related PAOs were governed by homogeneous selection and the dominant species revealed by 16S rRNA gene-based phylogenetic analysis shifted from clade III to clade I. Meanwhile, ATU supply promoted significant enrichment of functional genes involved in phosphate transport (pit) and polyphosphate synthesis and degradation (ppk1 and ppk2), whereas both Nitrosomonas and ammonia monooxygenase-encoding genes (amoA/B/C) assignable to this group of nitrifying bacteria decreased. Moreover, ATU addition relieved the significant abundance correlation between filamentous bacteria Ca. Promineofilum and denitrifying Brevundimonas (FDR-adjusted P < 0.01), damaging their potential synergic or cooperative interactions, thus weakening their competitiveness against Tetrasphaera-related PAOs. Notably, ATU withdrawn created opportunistic conditions for the unexpected explosive growth and predominance of Thiothrix filaments, leading to a serious bulking event. Our study provides new insights into the microbial ecology of Tetrasphaera-related PAOs in EBPR system, which could guide the establishment of an efficient microbiota for EBPR.

Prediction and validation of microbial community function from normal pulp to pulpitis caused by deep dentinal caries.

INTRODUCTION: Microbial function changes may be responsible for dental pulp transformation from normal to diseased. However, studies on the prediction and verification of the function of the microbial community in the deep dentine and pulp of caries-induced pulpitis are lacking. METHODS: This study included 171 cases of deep dentinal caries divided into normal pulp (NP), reversible pulpitis (RP), and irreversible pulpitis (IRP). In Experiment I, the microbial community composition was identified in 111 samples using 16S ribosomal DNA. Function prediction was performed through phylogenetic investigation of communities by Phylogenetic Investigation of Communities by Reconstruction of Unobserved States prediction and qPCR. In Experiment II, different microbiome functions were confirmed in 60 samples using liquid chromatography-tandem mass spectrometry. RESULTS: In Experiment I, microbial abundance significantly differed in the IRP group compared to the other two groups. The RP and NP groups had the same microbiome composition, but the predicted functional difference between the RP and NP groups pertained to membrane transport (p < .010). The predicted functional difference between the IRP and NP groups pertained to amino-acid, co-factor, and vitamin metabolism (p < .010). In Experiment II, Kyoto Encyclopedia of Genes and Genomes functional annotation revealed that the differential metabolites between the RP and NP groups did not participate in membrane transport; however, the differential metabolites between the IRP and NP groups participated in amino-acid metabolism. CONCLUSIONS: The near-pulp microbiome in RP and NP with deep dentinal caries had the same differential function. However, amino acid metabolism in near the pulp microbial community differed between IRP and NP with deep dentinal caries.

Germline-Activating RRAS2 Mutations Cause Noonan Syndrome.

Noonan syndrome (NS) is characterized by distinctive craniofacial appearance, short stature, and congenital heart disease. Approximately 80% of individuals with NS harbor mutations in genes whose products are involved in the RAS/mitogen-activating protein kinase (MAPK) pathway. However, the underlying genetic causes in nearly 20% of individuals with NS phenotype remain unexplained. Here, we report four de novo RRAS2 variants in three individuals with NS. RRAS2 is a member of the RAS subfamily and is ubiquitously expressed. Three variants, c.70_78dup (p.Gly24_Gly26dup), c.216A>T (p.Gln72His), and c.215A>T (p.Gln72Leu), have been found in cancers; our functional analyses showed that these three changes induced elevated association of RAF1 and that they activated ERK1/2 and ELK1. Notably, prominent activation of ERK1/2 and ELK1 by p.Gln72Leu associates with the severe phenotype of the individual harboring this change. To examine variant pathogenicity in vivo, we generated zebrafish models. Larvae overexpressing c.70_78dup (p.Gly24_Gly26dup) or c.216A>T (p.Gln72His) variants, but not wild-type RRAS2 RNAs, showed craniofacial defects and macrocephaly. The same dose injection of mRNA encoding c.215A>T (p.Gln72Leu) caused severe developmental impairments and low dose overexpression of this variant induced craniofacial defects. In contrast, the RRAS2 c.224T>G (p.Phe75Cys) change, located on the same allele with p.Gln72His in an individual with NS, resulted in no aberrant in vitro or in vivo phenotypes by itself. Together, our findings suggest that activating RRAS2 mutations can cause NS and expand the involvement of RRAS2 proto-oncogene to rare germline disorders.

Gut Microbiota Perturbation in IgA Deficiency Is Influenced by IgA-Autoantibody Status.

BACKGROUND & AIMS: IgA exerts its primary function at mucosal surfaces, where it binds microbial antigens to regulate bacterial growth and epithelial attachment. One third of individuals with IgA deficiency (IgAD) suffers from recurrent mucosal infections, possibly related to an altered microbiota. We aimed to delineate the impact of IgAD and the IgA-autoantibody status on the composition and functional capacity of the gut microbiota. METHODS: We performed a paired, lifestyle-balanced analysis of the effect of IgA on the gut microbiota composition and functionality based on fecal samples from individuals with IgAD and IgA-sufficient household members (n = 100), involving quantitative shotgun metagenomics, species-centric functional annotation of gut bacteria, and strain-level analyses. We supplemented the data set with 32 individuals with IgAD and examined the influence of IgA-autoantibody status on the composition and functionality of the gut microbiota. RESULTS: The gut microbiota of individuals with IgAD exhibited decreased richness and diversity and was enriched for bacterial species encoding pathogen-related functions including multidrug and antimicrobial peptide resistance, virulence factors, and type III and VI secretion systems. These functional changes were largely attributed to Escherichia coli but were independent of E coli strain variations and most prominent in individuals with IgAD with IgA-specific autoreactive antibodies. CONCLUSIONS: The microbiota of individuals with IgAD is enriched for species holding increased proinflammatory potential, thereby potentially decreasing the resistance to gut barrier-perturbing events. This phenotype is especially pronounced in individuals with IgAD with IgA-specific autoreactive antibodies, thus warranting a screening for IgA-specific autoreactive antibodies in IgAD to identify patients with IgAD with increased risk for gastrointestinal implications.

Bioprospecting uncultivable microbial diversity in tannery effluent contaminated soil using shotgun sequencing and bio-reduction of chromium by indigenous chromate reductase genes.

The tannery industry generates a consequential threat to the environment by producing a large amount of potentially toxic metal-containing waste. Bioremediation has been a promising approach for treating potentially toxic metals, but the efficiency of remediation in microbes is one of the factors limiting their application in tanneries waste treatment. The motivation behind the present work was to explore the microbial diversity and chromate reductase genes present in the tannery effluent-contaminated soil using metagenomics approach. The use of shotgun sequencing enabled the identification of operational parameters that influence microbiome composition and their ability to reduce Chromium (Cr) concentration. The Cr concentration in Kanpur tannery effluent contaminated soil sample was 700 ppm which is many folds than the approved permissible limit by World Health Organisation (WHO) for Cr is 100 ppm. Metagenomic Deoxyribo Nucleic Acid (DNA) was extracted to explore taxonomic community structure, phylogenetic linkages, and functional profile. With a Guanine-Cytosine (GC) abundance of 54%, total of 45,163,604 high-quality filtered reads were obtained. Bacteria (83%), Archaebacteria (14%), and Viruses (3%) were discovered in the structural biodiversity. Bacteria were classified to phylum level, with Proteobacteria (52%) being the dominant population, followed by Bacteriodetes (15%), Chloroflexi (15%), Spirochaetes (7%), Thermotogae (5%), Actinobacteria (4%), and Firmicutes (1%). The OXR genes were cloned and checked for their efficiency to reduce Cr concentration. Insitu validation of OXR8 gene showed a reduction of Cr concentration from 700 ppm to 24 ppm in 72 h (96.51% reduction). The results of this study suggests that there is a huge reservoir of microbes and chromate reductase genes which are unexplored yet.

Gut milieu shapes the bacterial communities of invasive silver carp.

Silver carp is an invasive fish present in the Gobindsagar reservoir, India and has a profound impact on aquaculture. Understanding taxonomic diversity and functional attributes of gut microbiota will provide insights into the important role of bacteria in metabolism of silver carp that facilitated invasion of this exotic species. Microbial composition in foregut, midgut, hindgut and water samples was analysed using 16S rRNA gene amplicon sequencing. The bacterial communities of water samples were distinct from gut microbiota, and unique microbial assemblages were present in different regions of gut depicting profound impact of gut environment on microflora. Proteobacteria was the most abundant phyla across all samples. Ecological network analysis showed dominance of competitive interactions within posteriors region of the gut, promoting niche specialization. Predictive functional profiling revealed the microbiota specialized in digestive functions in different regions of the gut, which also reflects the dietary profile of silver carp.

Functional Dysregulation of CDC42 Causes Diverse Developmental Phenotypes.

Exome sequencing has markedly enhanced the discovery of genes implicated in Mendelian disorders, particularly for individuals in whom a known clinical entity could not be assigned. This has led to the recognition that phenotypic heterogeneity resulting from allelic mutations occurs more commonly than previously appreciated. Here, we report that missense variants in CDC42, a gene encoding a small GTPase functioning as an intracellular signaling node, underlie a clinically heterogeneous group of phenotypes characterized by variable growth dysregulation, facial dysmorphism, and neurodevelopmental, immunological, and hematological anomalies, including a phenotype resembling Noonan syndrome, a developmental disorder caused by dysregulated RAS signaling. In silico, in vitro, and in vivo analyses demonstrate that mutations variably perturb CDC42 function by altering the switch between the active and inactive states of the GTPase and/or affecting CDC42 interaction with effectors, and differentially disturb cellular and developmental processes. These findings reveal the remarkably variable impact that dominantly acting CDC42 mutations have on cell function and development, creating challenges in syndrome definition, and exemplify the importance of functional profiling for syndrome recognition and delineation.

Ultrahigh-throughput functional profiling of microbiota communities.

Microbiome spectra serve as critical clues to elucidate the evolutionary biology pathways, potential pathologies, and even behavioral patterns of the host organisms. Furthermore, exotic sources of microbiota represent an unexplored niche to discover microbial secondary metabolites. However, establishing the bacterial functionality is complicated by an intricate web of interactions inside the microbiome. Here we apply an ultrahigh-throughput (uHT) microfluidic droplet platform for activity profiling of the entire oral microbial community of the Siberian bear to isolate Bacillus strains demonstrating antimicrobial activity against Staphylococcus aureus Genome mining allowed us to identify antibiotic amicoumacin A (Ami) as responsible for inhibiting the growth of S. aureus Proteomics and metabolomics revealed a unique mechanism of Bacillus self-resistance to Ami, based on a subtle equilibrium of its deactivation and activation by kinase AmiN and phosphatase AmiO, respectively. We developed uHT quantitative single-cell analysis to estimate antibiotic efficacy toward different microbiomes and used it to determine the activity spectra of Ami toward human and Siberian bear microbiota. Thus, uHT microfluidic droplet platform activity profiling is a powerful tool for discovering antibiotics and quantifying external influences on a microbiome.

Comprehensive functional profiling of long non-coding RNAs through a novel pan-cancer integration approach and modular analysis of their protein-coding gene association networks.

BACKGROUND: Long non-coding RNAs (lncRNAs) are emerging as crucial regulators of cellular processes in diseases such as cancer, although the functions of most remain poorly understood. To address this, here we apply a novel strategy to integrate gene expression profiles across 32 cancer types, and cluster human lncRNAs based on their pan-cancer protein-coding gene associations. By doing so, we derive 16 lncRNA modules whose unique properties allow simultaneous inference of function, disease specificity and regulation for over 800 lncRNAs. RESULTS: Remarkably, modules could be grouped into just four functional themes: transcription regulation, immunological, extracellular, and neurological, with module generation frequently driven by lncRNA tissue specificity. Notably, three modules associated with the extracellular matrix represented potential networks of lncRNAs regulating key events in tumour progression. These included a tumour-specific signature of 33 lncRNAs that may play a role in inducing epithelial-mesenchymal transition through modulation of TGFß signalling, and two stromal-specific modules comprising 26 lncRNAs linked to a tumour suppressive microenvironment and 12 lncRNAs related to cancer-associated fibroblasts. One member of the 12-lncRNA signature was experimentally supported by siRNA knockdown, which resulted in attenuated differentiation of quiescent fibroblasts to a cancer-associated phenotype. CONCLUSIONS: Overall, the study provides a unique pan-cancer perspective on the lncRNA functional landscape, acting as a global source of novel hypotheses on lncRNA contribution to tumour progression.

Composition and predictive functional analysis of bacterial communities inhabiting Chinese Cordyceps insight into conserved core microbiome.

BACKGROUND: Over the past few decades, most attention to Chinese Cordyceps-associated endogenous microorganism was focused on the fungal community that creates critical bioactive components. Bacterial community associated with Chinese Cordyceps has been previously described; however, most studies were only presenting direct comparisons in the Chinese Cordyceps and its microenvironments. In the current study, our objectives were to reveal the bacterial community structure composition and predict their function. RESULTS: We collected samples of Chinese Cordyceps from five sites located in the Qinghai-Tibet Plateau and used a high throughput sequencing method to compare Chinese Cordyceps-associated bacterial community composition and diversity quantitatively across sites. The results indicated that for the Chinese Cordyceps-associated bacterial community there is no single core microbiome, which was dominated by the both Proteobacteria and Actinobacteria. Predictive functional profiling suggested a location specific function pattern for Chinese Cordyceps and bacteria in the external mycelial cortices involved in the biosynthesis of active constituents. CONCLUSIONS: This study is firstly used high throughput sequencing method to compare the bacterial communities inhabiting Chinese Cordyceps and its microhabitat and to reveal composition functional capabilities of the bacteria, which will accelerate the study of the functions of bacterial communities in the micro-ecological system of Chinese Cordyceps.

Underlying mechanisms of ANAMMOX bacteria adaptation to salinity stress.

Dealing with nitrogen-rich saline wastewater produced by industries remains challenging because of the inhibition of functional microorganisms by high salinity. The underlying mechanisms of anaerobic ammonium-oxidizing bacteria (AnAOB) exposed to salinity stress should be studied to investigate the potential of anaerobic ammonium oxidation (ANAMMOX) for applications in such wastewater. In this study, the total DNA from granular sludge was extracted from an expanded granular sludge bed (EGSB) reactor operated at 0, 15 and 30 g/L salinity and subjected to high-throughput sequencing. The nitrogen removal performance in the reactor could be maintained from 86.2 to 88.0% at less than 30 g/L salinity level. The microbial diversity in the reactor under saline conditions was lower than that under the salt-free condition. Three genera of AnAOB were detected in the reactor, and Candidatus Kuenenia was the most abundant. The predictive functional profiling based on the Clusters of Orthologous Groups of proteins (COGs) database showed that the inhibition of AnAOB under saline conditions was mainly characterised by the weakening of energy metabolism and intracellular repair. AnAOB might adapt to salinity stress by increasing their rigidity and intracellular osmotic pressure. The predictive functional profiling based on the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway database revealed that the inhibition of AnAOB was mainly manifested by the weakening of intracellular carbohydrate and lipid metabolism, the blockage of intracellular energy supply and the reduction of membrane transport capacity. AnAOB might adapt to salinity stress by strengthening wall/membrane synthesis, essential cofactors (porphyrins) and energy productivity, enhancing intracellular material transformation and gene repair and changing its structure and group behaviour. The stability of the nitrogen removal performance could be maintained via the adaptation of AnAOB to salinity and their increased abundance.

Cell-based screens and phenomics with fission yeast.

Next-generation sequencing approaches have considerably advanced our understanding of genome function and regulation. However, the knowledge of gene function and complex cellular processes remains a challenge and bottleneck in biological research. Phenomics is a rapidly emerging area, which seeks to rigorously characterize all phenotypes associated with genes or gene variants. Such high-throughput phenotyping under different conditions can be a potent approach toward gene function. The fission yeast Schizosaccharomyces pombe (S. pombe) is a proven eukaryotic model organism that is increasingly used for genomewide screens and phenomic assays. In this review, we highlight current large-scale, cell-based approaches used with S. pombe, including computational colony-growth measurements, genetic interaction screens, parallel profiling using barcodes, microscopy-based cell profiling, metabolomic methods and transposon mutagenesis. These diverse methods are starting to offer rich insights into the relationship between genotypes and phenotypes.

Species Diversity and Functional Prediction of Surface Bacterial Communities on Aging Flue-Cured Tobaccos.

Microbes on aging flue-cured tobaccos (ATFs) improve the aroma and other qualities desirable in products. Understanding the relevant organisms would picture microbial community diversity, metabolic potential, and their applications. However, limited efforts have been made on characterizing the microbial quality and functional profiling. Herein, we present our investigation of the bacterial diversity and predicted potential genetic capability of the bacteria from two AFTs using 16S rRNA gene sequences and phylogenetic investigation of communities by reconstruction of unobserved states (PICRUSt) software. The results show that dominant bacteria from AFT surfaces were classified into 48 genera, 36 families, and 7 phyla. In addition, Bacillus spp. was found prevalent on both ATFs. Furthermore, PICRUSt predictions of bacterial community functions revealed many attractive metabolic capacities in the AFT microbiota, including several involved in the biosynthesis of flavors and fragrances and the degradation of harmful compounds, such as nicotine and nitrite. These results provide insights into the importance of AFT bacteria in determining product qualities and indicate specific microbial species with predicted enzymatic capabilities for the production of high-efficiency flavors, the degradation of undesirable compounds, and the provision of nicotine and nitrite tolerance which suggest fruitful areas of investigation into the manipulation of AFT microbiota for AFT and other product improvements.

A bioinformatics pipeline integrating predictive metagenomics profiling for the analysis of 16S rDNA/rRNA sequencing data originated from foods.

The recent advances in molecular biology, such as the advent of next-generation sequencing (NGS) platforms, have paved the way to new exciting tools which rapidly transform food microbiology. Nowadays, NGS methods such as 16S rDNA/rRNA metagenomics or amplicon sequencing are used for the taxonomic profiling of the food microbial communities. Although 16S rDNA/rRNA NGS-based microbial data are not suited for the investigation of the functional potential of the identified operational taxonomic units as compared to shotgun metagenomics, advances in the bioinformatics discipline allow now the performance of such studies. In this paper, a bioinformatics workflow is described integrating predictive metagenomics profiling with specific application to food microbiology data. Bioinformatics tools pertinent to each sub-module of the pipeline are suggested as well. The published 16S rDNA/rRNA amplicon data originated from an Italian Grana-type cheese, using an NGS platform, was employed to demonstrate the predictive metagenomics profiling approach. The pipeline identified the microbial community and the changes that occurred in the microbial profile during manufacture of the food product studied (taxonomic profiling). The workflow also indicated significant changes in the functional profiling of the community. The tool may help to investigate the functional potential, alterations, and interactions of a microbial community. The proposed workflow may also find an application in the investigation of the ecology of foodborne pathogens encountered in various food products.