Enhancing the quality of fermented plant leaves: the role of metabolite signatures and associated fungi.

Fungi play a pivotal role in fermentation processes, influencing the breakdown and transformation of metabolites. However, studies focusing on the effects of fungal-metabolite correlations on leaf fermentation quality enhancement are limited. This study investigated specific metabolites and fungi associated with high- and low-quality fermented plant leaves. Their changes were monitored over fermentation periods of 0, 8, 16, and 24 days. The results indicated that organoheterocyclic compounds, lipids, lipid-like molecules, organic nitrogen compounds, phenylpropanoids, and polyketides were predominant in high-quality samples. The fungi Saccharomyces (14.8%) and Thermoascus (4.6%) were predominantly found in these samples. These markers exhibited significant changes during the 24-day fermentation period. The critical influence of fungal community equilibrium was demonstrated by interspecies interactions (e.g., between Saccharomyces and Eurotium). A co-occurrence network analysis identified Saccharomyces as the primary contributor to high-quality samples. These markers collectively enhance the quality and sensory characteristics of the final product.

What role do biocontrol agents with Mg2+ play in the fate of antibiotic resistome and pathogenic bacteria in the phyllosphere?

The contamination of the plant phyllosphere with antibiotics and antibiotic resistance genes (ARGs), caused by application of antibiotics, is a significant environmental issue in agricultural management. Alternatively, biocontrol agents are environmentally friendly and have attracted a lot of interest. However, the influence of biocontrol agents on the phyllosphere resistome remains unknown. In this study, we applied biocontrol agents to control the wildfire disease in the Solanaceae crops and investigated their effects on the resistome and the pathogen in the phyllosphere by using metagenomics. A total of 250 ARGs were detected from 15 samples, which showed a variation in distribution across treatments of biocontrol agents (BA), BA with Mg2+ (T1), BA with Mn2+ (T2), and kasugamycin (T3) and nontreated (CK). The results showed that the abundance of ARGs under the treatment of BA-Mg2+ was lower than that in the CK group. The abundance of cphA3 (carbapenem resistance), PME-1 (carbapenem resistance), tcr3 (tetracycline antibiotic resistance), and AAC (3)-VIIIa (aminoglycoside antibiotic resistance) in BA-Mg2+ was significantly higher than that in BA-Mn2+ (P < 0.05). The abundance of cphA3, PME_1, and tcr3 was significantly negatively related to the abundance of the phyllosphere pathogen Pseudomonas syringae (P < 0.05). We also found that the upstream and downstream regions of cphA3 were relatively conserved, in which rpl, rpm, and rps gene families were identified in most sequences (92%). The Ka/Ks of cphA3 was 0 in all observed sequences, indicating that under the action of purifying selection, nonsynonymous substitutions are often gradually eliminated in the population. Overall, this study clarifies the effect of biocontrol agents with Mg2+ on the distribution of the phyllosphere resistome and provides evolutionary insights into the biocontrol process. IMPORTANCE: Our study applied metagenomics analysis to examine the impact of biocontrol agents (BAs) on the phyllosphere resistome and the pathogen. Irregular use of antibiotics has led to the escalating dissemination of antibiotic resistance genes (ARGs) in the environment. The majority of BA research has focused on the effect of monospecies on the plant disease control process, the role of the compound BA with nutrition elements in the phyllosphere disease, and the resistome is still unknown. We believe BAs are eco-friendly alternatives for antibiotics to combat the transfer of ARGs. Our results revealed that BA-Mg2+ had a lower relative abundance of ARGs compared to the CK group, and the phyllosphere pathogen Pseudomonas syringae was negatively related to three specific ARGs, cphA3, PME-1, and tcr3. These three genes also present different Ka/Ks. We believe that the identification of the distribution and evolution modes of ARGs further elucidates the ecological role and facilitates the development of BAs, which will attract general interest in this field.

Metagenomic insights into the response of soil microbial communities to pathogenic Ralstonia solanacearum.

Understanding the response of soil microbial communities to pathogenic Ralstonia solanacearum is crucial for preventing bacterial wilt outbreaks. In this study, we investigated the soil physicochemical and microbial community to assess their impact on the pathogenic R.solanacearum through metagenomics. Our results revealed that certain archaeal taxa were the main contributors influencing the health of plants. Additionally, the presence of the pathogen showed a strong negative correlation with soil phosphorus levels, while soil phosphorus was significantly correlated with bacterial and archaeal communities. We found that the network of microbial interactions in healthy plant rhizosphere soils was more complex compared to diseased soils. The diseased soil network had more linkages, particularly related to the pathogen occurrence. Within the network, the family Comamonadaceae, specifically Ramlibacter_tataouinensis, was enriched in healthy samples and showed a significantly negative correlation with the pathogen. In terms of archaea, Halorubrum, Halorussus_halophilus (family: Halobacteriaceae), and Natronomonas_pharaonis (family: Haloarculaceae) were enriched in healthy plant rhizosphere soils and showed negative correlations with R.solanacearum. These findings suggested that the presence of these archaea may potentially reduce the occurrence of bacterial wilt disease. On the other hand, Halostagnicola_larseniia and Haloterrigena_sp._BND6 (family: Natrialbaceae) had higher relative abundance in diseased plants and exhibited significantly positive correlations with R.solanacearum, indicating their potential contribution to the pathogen's occurrence. Moreover, we explored the possibility of functional gene sharing among the correlating bacterial pairs within the Molecular Ecological Network. Our analysis revealed 468 entries of horizontal gene transfer (HGT) events, emphasizing the significance of HGT in shaping the adaptive traits of plant-associated bacteria, particularly in relation to host colonization and pathogenicity. Overall, this work revealed key factors, patterns and response mechanisms underlying the rhizosphere soil microbial populations. The findings offer valuable guidance for effectively controlling soil-borne bacterial diseases and developing sustainable agriculture practices.

Transcriptomics of Lactobacillus paracasei: metabolism patterns and cellular responses under high-density culture conditions.

Lactobacillus paracasei has significant potential for development and application in the environmental field, particularly in addressing malodor pollution. This study aims to investigate the cellular response of L. paracasei B1 under high-density culture conditions. The selected strain has previously shown effective deodorizing and bacteriostatic abilities. Transcriptomics techniques are employed to dissect the nutrient metabolism pattern of L. paracasei B1 and its response mechanism under environmental stress. The study characterizes the functions of key differentially expressed genes during growth before and after optimizing the culture conditions. The optimization of fermentation culture conditions provides a suitable growth environment for L. paracasei B1, inducing an enhancement of its phosphotransferase system for sugar source uptake and maintaining high levels of glycolysis and pyruvate metabolism. Consequently, the strain is able to grow and multiply rapidly. Under acid stress conditions, glycolysis and pyruvate metabolism are inhibited, and L. paracasei B1 generates additional energy through aerobic respiration to meet the energy demand. The two-component system and quorum sensing play roles in the response and regulation of L. paracasei B1 to adverse environments. The strain mitigates oxygen stress damage through glutathione metabolism, cysteine and methionine metabolism, base excision repair, and purine and pyrimidine metabolism. Additionally, the strain enhances lysine synthesis, the alanine, aspartate, and glutamate metabolic pathways, and relies on the ABC transport system to accumulate amino acid-compatible solutes to counteract acid stress and osmotic stress during pH regulation. These findings establish a theoretical basis for the further development and application of L. paracasei B1 for its productive properties.

AI-driven pan-proteome analyses reveal insights into the biohydrometallurgical properties of Acidithiobacillia.

Microorganism-mediated biohydrometallurgy, a sustainable approach for metal recovery from ores, relies on the metabolic activity of acidophilic bacteria. Acidithiobacillia with sulfur/iron-oxidizing capacities are extensively studied and applied in biohydrometallurgy-related processes. However, only 14 distinct proteins from Acidithiobacillia have experimentally determined structures currently available. This significantly hampers in-depth investigations of Acidithiobacillia's structure-based biological mechanisms pertaining to its relevant biohydrometallurgical processes. To address this issue, we employed a state-of-the-art artificial intelligence (AI)-driven approach, with a median model confidence of 0.80, to perform high-quality full-chain structure predictions on the pan-proteome (10,458 proteins) of the type strain Acidithiobacillia. Additionally, we conducted various case studies on de novo protein structural prediction, including sulfate transporter and iron oxidase, to demonstrate how accurate structure predictions and gene co-occurrence networks can contribute to the development of mechanistic insights and hypotheses regarding sulfur and iron utilization proteins. Furthermore, for the unannotated proteins that constitute 35.8% of the Acidithiobacillia proteome, we employed the deep-learning algorithm DeepFRI to make structure-based functional predictions. As a result, we successfully obtained gene ontology (GO) terms for 93.6% of these previously unknown proteins. This study has a significant impact on improving protein structure and function predictions, as well as developing state-of-the-art techniques for high-throughput analysis of large proteomic data.

Sequence similarity network and protein structure prediction offer insights into the evolution of microbial pathways for ferrous iron oxidation.

IMPORTANCE: Microbial Fe(II) oxidation is a crucial process that harnesses and converts the energy available in Fe, contributing significantly to global element cycling. However, there are still many aspects of this process that remain unexplored. In this study, we utilized a combination of comparative genomics, sequence similarity network analysis, and artificial intelligence-driven structure modeling methods to address the lack of structural information on Fe(II) oxidation proteins and offer a comprehensive perspective on the evolution of Fe(II) oxidation pathways. Our findings suggest that several microbial Fe(II) oxidation pathways currently known may have originated within classes Gammaproteobacteria and Betaproteobacteria.

Community ecological study on the reduction of soil antimony bioavailability by SRB-based remediation technologies.

Sulfate-reducing bacteria (SRB) were effective in stabilizing Sb. However, the influence of electron donors and acceptors during SRB remediation, as well as the ecological principles involved, remained unclear. In this study, Desulfovibrio desulfuricans ATCC 7757 was utilized to stabilize soil Sb within microcosm. Humic acid (HA) or sodium sulfate (Na2SO4) were employed to enhance SRB capacity. The SRB+HA treatment exhibited the highest Sb stabilization rate, achieving 58.40%. Bacterial community analysis revealed that SRB altered soil bacterial diversity, community composition, and assembly processes, with homogeneous selection as the predominant assembly processes. When HA and Na2SO4 significantly modified the stimulated microbial community succession trajectories, shaped the taxonomic composition and interactions of the bacterial community, they showed converse effect in shaping bacterial community which were both helpful for promoting dissimilatory sulfate reduction. Na2SO4 facilitated SRB-mediated anaerobic reduction and promoted interactions between SRB and bacteria involved in nitrogen and sulfur cycling. The HA stimulated electron generation and storage, and enhanced the interactions between SRB and bacteria possessing heavy metal tolerance or carbohydrate degradation capabilities.

Genome-resolved metagenomics provides insights into the ecological roles of the keystone taxa in heavy-metal-contaminated soils.

Microorganisms that exhibit resistance to environmental stressors, particularly heavy metals, have the potential to be used in bioremediation strategies. This study aimed to explore and identify microorganisms that are resistant to heavy metals in soil environments as potential candidates for bioremediation. Metagenomic analysis was conducted using microbiome metagenomes obtained from the rhizosphere of soil contaminated with heavy metals and mineral-affected soil. The analysis resulted in the recovery of a total of 175 metagenome-assembled genomes (MAGs), 73 of which were potentially representing novel taxonomic levels beyond the genus level. The constructed ecological network revealed the presence of keystone taxa, including Rhizobiaceae, Xanthobacteraceae, Burkholderiaceae, and Actinomycetia. Among the recovered MAGs, 50 were associated with these keystone taxa. Notably, these MAGs displayed an abundance of genes conferring resistance to heavy metals and other abiotic stresses, particularly those affiliated with the keystone taxa. These genes were found to combat excessive accumulation of zinc/manganese, arsenate/arsenite, chromate, nickel/cobalt, copper, and tellurite. Furthermore, the keystone taxa were found to utilize both organic and inorganic energy sources, such as sulfur, arsenic, and carbon dioxide. Additionally, these keystone taxa exhibited the ability to promote vegetation development in re-vegetated mining areas through phosphorus solubilization and metabolite secretion. In summary, our study highlights the metabolic adaptability and ecological significance of microbial keystone taxa in mineral-affected soils. The MAGs associated with keystone taxa exhibited a markedly higher number of genes related to abiotic stress resistance and plant growth promotion compared to non-keystone taxa MAGs.

Dissecting the HGT network of carbon metabolic genes in soil-borne microbiota.

The microbiota inhabiting soil plays a significant role in essential life-supporting element cycles. Here, we investigated the occurrence of horizontal gene transfer (HGT) and established the HGT network of carbon metabolic genes in 764 soil-borne microbiota genomes. Our study sheds light on the crucial role of HGT components in microbiological diversification that could have far-reaching implications in understanding how these microbial communities adapt to changing environments, ultimately impacting agricultural practices. In the overall HGT network of carbon metabolic genes in soil-borne microbiota, a total of 6,770 nodes and 3,812 edges are present. Among these nodes, phyla Proteobacteria, Actinobacteriota, Bacteroidota, and Firmicutes are predominant. Regarding specific classes, Actinobacteria, Gammaproteobacteria, Alphaproteobacteria, Bacteroidia, Actinomycetia, Betaproteobacteria, and Clostridia are dominant. The Kyoto Encyclopedia of Genes and Genomes (KEGG) functional assignments of glycosyltransferase (18.5%), glycolysis/gluconeogenesis (8.8%), carbohydrate-related transporter (7.9%), fatty acid biosynthesis (6.5%), benzoate degradation (3.1%) and butanoate metabolism (3.0%) are primarily identified. Glycosyltransferase involved in cell wall biosynthesis, glycosylation, and primary/secondary metabolism (with 363 HGT entries), ranks first overwhelmingly in the list of most frequently identified carbon metabolic HGT enzymes, followed by pimeloyl-ACP methyl ester carboxylesterase, alcohol dehydrogenase, and 3-oxoacyl-ACP reductase. Such HGT events mainly occur in the peripheral functions of the carbon metabolic pathway instead of the core section. The inter-microbe HGT genetic traits in soil-borne microbiota genetic sequences that we recognized, as well as their involvement in the metabolism and regulation processes of carbon organic, suggest a pervasive and substantial effect of HGT on the evolution of microbes.

Inoculation with thermophiles enhanced the food waste bio-drying and complicated interdomain ecological networks between bacterial and fungal communities.

Bio-drying is a practical approach for treating food waste (FW). However, microbial ecological processes during treatment are essential for improving the dry efficiency, and have not been stressed enough. This study analyzed the microbial community succession and two critical periods of interdomain ecological networks (IDENs) during FW bio-drying inoculated with thermophiles (TB), to determine how TB affects FW bio-drying efficiency. The results showed that TB could rapidly colonize in the FW bio-drying, with the highest relative abundance of 5.13%. Inoculating TB increased the maximum temperature, temperature integrated index and moisture removal rate of FW bio-drying (55.7 °C, 219.5 °C, and 86.11% vs. 52.1 °C, 159.1 °C, and 56.02%), thereby accelerating the FW bio-drying efficiency by altering the succession of microbial communities. The structural equation model and IDEN analysis demonstrated that TB inoculation complicated the IDENs between bacterial and fungal communities by significantly and positively affecting bacterial communities (b = 0.39, p < 0.001) and fungal communities (b = 0.32, p < 0.01), thereby enhancing interdomain interactions between bacteria and fungi. Additionally, inoculation TB significantly increased the relative abundance of keystone taxa, including Clostridium sensu stricto, Ochrobactrum, Phenylobacterium, Microvirga and Candida. In conclusion, the inoculation of TB could effectively improve FW bio-drying, which is a promising technology for rapidly reducing FW with high moisture content and recovering resources from it.

A bi-steric mTORC1-selective inhibitor overcomes drug resistance in breast cancer.

Activation of the PI3K-mTOR pathway is central to breast cancer pathogenesis including resistance to many targeted therapies. The mTOR kinase forms two distinct complexes, mTORC1 and mTORC2, and understanding which is required for the survival of malignant cells has been limited by tools to selectively and completely impair either subcomplex. To address this, we used RMC-6272, a bi-steric molecule with a rapamycin-like moiety linked to an mTOR active-site inhibitor that displays >25-fold selectivity for mTORC1 over mTORC2 substrates. Complete suppression of mTORC1 by RMC-6272 causes apoptosis in ER+/HER2- breast cancer cell lines, particularly in those that harbor mutations in PIK3CA or PTEN, due to inhibition of the rapamycin resistant, mTORC1 substrate 4EBP1 and reduction of the pro-survival protein MCL1. RMC-6272 reduced translation of ribosomal mRNAs, MYC target genes, and components of the CDK4/6 pathway, suggesting enhanced impairment of oncogenic pathways compared to the partial mTORC1 inhibitor everolimus. RMC-6272 maintained efficacy in hormone therapy-resistant acquired cell lines and patient-derived xenografts (PDX), showed increased efficacy in CDK4/6 inhibitor treated acquired resistant cell lines versus their parental counterparts, and was efficacious in a PDX from a patient experiencing resistance to CDK4/6 inhibition. Bi-steric mTORC1-selective inhibition may be effective in overcoming multiple forms of therapy-resistance in ER+ breast cancers.

Community- and genome-based evidence for a shaping influence of redox potential on bacterial protein evolution.

Despite deep interest in how environments shape microbial communities, whether redox conditions influence the sequence composition of genomes is not well known. We predicted that the carbon oxidation state (ZC) of protein sequences would be positively correlated with redox potential (Eh). To test this prediction, we used taxonomic classifications for 68 publicly available 16S rRNA gene sequence data sets to estimate the abundances of archaeal and bacterial genomes in river & seawater, lake & pond, geothermal, hyperalkaline, groundwater, sediment, and soil environments. Locally, ZC of community reference proteomes (i.e., all the protein sequences in each genome, weighted by taxonomic abundances but not by protein abundances) is positively correlated with Eh corrected to pH 7 (Eh7) for the majority of data sets for bacterial communities in each type of environment, and global-scale correlations are positive for bacterial communities in all environments. In contrast, archaeal communities show approximately equal frequencies of positive and negative correlations in individual data sets, and a positive pan-environmental correlation for archaea only emerges after limiting the analysis to samples with reported oxygen concentrations. These results provide empirical evidence that geochemistry modulates genome evolution and may have distinct effects on bacteria and archaea. IMPORTANCE The identification of environmental factors that influence the elemental composition of proteins has implications for understanding microbial evolution and biogeography. Millions of years of genome evolution may provide a route for protein sequences to attain incomplete equilibrium with their chemical environment. We developed new tests of this chemical adaptation hypothesis by analyzing trends of the carbon oxidation state of community reference proteomes for microbial communities in local- and global-scale redox gradients. The results provide evidence for widespread environmental shaping of the elemental composition of protein sequences at the community level and establish a rationale for using thermodynamic models as a window into geochemical effects on microbial community assembly and evolution.

RNA methylation pattern and immune microenvironment characteristics mediated by m6A regulator in ischemic stroke.

Background: Ischemic stroke (IS) is a highly heterogeneous disease. Recent studies have shown that epigenetic variables affect the immune response. However, only a few studies have examined the relationship between IS and m6A immunoregulation. Therefore, we aim to explore the methylation of RNA mediated by m6A regulatory factor and the immune microenvironment characteristics of IS. Methods: Differentially expressed m6A regulators were detected in IS microarray datasets GSE22255 and GSE58294. We used a series of machine learning algorithms to identify key IS-related m6A regulators and validated them on blood samples of IS patients, oxygen-glucose deprivation/reoxygenation (OGD/R) microglia and GSE198710 independent data sets. Different m6A modification modes were determined and the patients were classified. In addition, we systematically associate these modification patterns with the characteristics of immune microenvironment, including infiltrating immune cells, immune function genes and immune response genes. Then we developed a model of m6A score to quantify the m6A modification in IS samples. Results: Through the analysis of the differences between the control group and IS patients, METTL16, LRPPRC, and RBM15 showed strong diagnostic significance in three independent data sets. In addition, qRT-PCR and Western blotting also confirmed that the expression of METTL16 and LRPPRC was downregulated and the expression of RBM15 was upregulated after ischemia. Two m6A modification modes and two m6A gene modification modes were also identified. m6A gene cluster A (high m6A value group) was positively correlated with acquired immunity, while m6A gene cluster B (low m6A value group) was positively correlated with innate immunity. Similarly, five immune-related hub genes were significantly associated with m6Acore (CD28, IFNG, LTF, LCN2, and MMP9). Conclusion: The modification of m6A is closely related to the immune microenvironment. The evaluation of individual m6A modification pattern may be helpful for future immunomodulatory therapy of anti-ischemic response.

Chlorella sorokiniana FK-montmorillonite interaction enhanced remediation of heavy metals in tailings.

Non-ferrous metal mining activities are known to cause ecological irreversible damage in the tailings and surrounding areas as well as heavy metal (HM) contamination. The enhancement of Chlorella-montmorillonite interaction on the remediation of HM contaminated tailings was verified from the lab to the tailings in Daye City, Hubei Province, China. The results showed a positive correlation between the quantity of montmorillonite and the transformation of Pb and Cu into residual and carbonate-binding states, which resulted in a considerable decrease in the leaching ratio. The buildup of tailings fertility throughout this process benefited from montmorillonite's ability to buffer environmental changes and store water. This further offers a required environmental foundation for the rebuilding of microbial community and the growth of herbaceous plants. The structural equation model demonstrated that the interaction between Chlorella and montmorillonite directly affected the stability of HM, and that this interaction also had an impact on the accumulation of organic carbon, total nitrogen, and available phosphorus, which improved the immobilization of Pb, Cu, Cd, and Zn. This work made the first attempt to apply Chlorella-montmorillonite composite to in-situ tailings remediation, and proposed that the combination of inorganic clay minerals and organic microorganisms was an eco-friendly, long-lasting, and efficient method for immobilizing multiple-HMs in mining areas.

Low MHR Is Associated with Hemorrhagic Transformation in Acute Large Artery Atherosclerosis Ischemic Stroke Patients with Intravenous Thrombolysis.

Background: Hemorrhagic transformation (HT) is a common complication of intravenous thrombolysis (IVT) in patients with acute ischemic stroke (AIS) and may lead to neurological deterioration. This article discusses whether monocyte count to high-density lipoprotein ratio (MHR) level is associated with HT in AIS patients. Materials and methods: The clinical data of AIS patients who underwent rt-PA IVT treatment were continuously collected. According to whether HT occurred, patients were divided into HT group and non-HT group. Potential association between MHR and HT in different subtypes AIS was explored by using logistic regression. Results: A total of 444 AIS patients were retrospective analyzed. The MHR level was lower in HT group compared with the non-HT group in all AIS patients (0.28 vs 0.36, P = .031) and in large-artery atherosclerosis (LAA) type AIS patients (0.31 vs 0.37, P = .032). Low MHR was independently related to HT (OR:0.035, 95%CI:0.003-0.390, P = .006). Among all TOAST subtypes, low MHR was only independently associated with HT in patients of LAA-type AIS after adjusting for confounding factors (OR:0.01, 95%CI:0.00-0.62, P = .031), with an optimal cut-off value of 0.41, sensitivity of 85.7%, and specificity of 43.1%. MHR was not correlated with SVO, VE, and CE subtype AIS. Conclusion: Low MHR may be an independent predictor of HT in patients with AIS and this conclusion only existed in LAA-type AIS.

Biogeochemical behavior and pollution control of arsenic in mining areas: A review.

Arsenic (As) is one of the most toxic metalloids that possess many forms. As is constantly migrating from abandoned mining area to the surrounding environment in both oxidation and reducing conditions, threatening human health and ecological safety. The biogeochemical reaction of As included oxidation, reduction, methylation, and demethylation, which is closely associated with microbial metabolisms. The study of the geochemical behavior of arsenic in mining areas and the microbial remediation of arsenic pollution have great potential and are hot spots for the prevention and remediation of arsenic pollution. In this study, we review the distribution and migration of arsenic in the mining area, focus on the geochemical cycle of arsenic under the action of microorganisms, and summarize the factors influencing the biogeochemical cycle of arsenic, and strategies for arsenic pollution in mining areas are also discussed. Finally, the problems of the risk control strategies and the future development direction are prospected.

Viruses Regulate Microbial Community Assembly Together with Environmental Factors in Acid Mine Drainage.

Viruses are widespread in various ecosystems, and they play important roles in regulating the microbial community via host-virus interactions. Recently, metagenomic studies showed that there are extremely diverse viruses in different environments from the ocean to the human gut, but the influences of viral communities on microbial communities are poorly understood, especially in extreme environments. Here, we used metagenomics to characterize microbial communities and viral communities in acid mine drainage (AMD) and evaluated how viruses shape microbial community constrained by the harsh environments. Our results showed that AMD viral communities are significantly associated with the microbial communities, and viral diversity has positive correlations with microbial diversity. Viral community explained more variations of microbial community composition than environmental factors in AMD of a polymetallic mine. Moreover, we found that viruses harboring adaptive genes regulate a relative abundance of hosts under the modulation of environmental factors, such as pH. We also observed that viral diversity has significant correlations with the global properties of microbial cooccurrence networks, such as modularity. In addition, the results of null modeling analyses revealed that viruses significantly affect microbial community phylogeny and play important roles in regulating ecological processes of community assembly, such as dispersal limitation and homogenous dispersal. Together, these results revealed that AMD viruses are critical forces driving microbial network and community assembly via host-virus interactions. IMPORTANCE Viruses as mobile genetic elements play critical roles in the adaptive evolution of their hosts in extreme environments. However, how viruses further influence microbial community structure and assembly is still unclear. A recent metagenomic study observed diverse viruses unexplored in acid mine drainage, revealing the associations between the viral community and environmental factors. Here, we showed that viruses together with environmental factors can constrain the relative abundance of host and microbial community assembly in AMD of copper mines and polymetallic mines. Our results highlight the importance of viruses in shaping the microbial community from the individual host level to the community level.

MiR-140-3p directly targets Tyro3 to regulate OGD/R-induced neuronal injury through the PI3K/Akt pathway.

BACKGROUND AND PURPOSE: MicroRNAs (miRNAs) are highly expressed in the central nervous system and play important roles in ischaemic stroke pathogenesis. However, the role of miRNAs in cerebral ischaemia-reperfusion injury remains unclear. Here, we investigated the role of miR-140-3p in regulating oxygen-glucose deprivation/reoxygenation (OGD/R)-induced neuronal injury in vitro to identify a new biomarker for research on ischaemic stroke. METHODS: The differential expression of miR-140-3p and Tyro3 in OGD/R-exposed N2a cells was verified by qRT-PCR. N2a cells were transfected with miR-140-3p mimic, miR-140-3p inhibitor, Tyro3 or siTyro3, and qRT-PCR, Western blotting, the Cell counting kit-8 (CCK-8) assay, Hoechst 33342/PI staining and flow cytometry analyses were performed to measure miRNA, mRNA and protein expression; cell viability; and apoptosis. RESULTS: OGD/R-exposed N2a cells exhibited increased miR-140-3p expression, decreased viability, reduced Bcl-2 protein expression and increased Bax and Caspase-3 protein expression and apoptosis; the miR-140-3p mimic markedly amplified these changes, exacerbating OGD/R-induced injury to N2a cells, while the miR-140-3p inhibitor reversed these changes and alleviated OGD/R-induced injury. OGD/R-exposed N2a cells expressed less Tyro3, and Tyro3 overexpression increased cell viability and Bcl-2 protein expression, reduced Bax and Caspase-3 protein expression, and alleviated OGD/R-induced injury. However, silencing Tyro3 reversed these changes and exacerbated OGD/R-induced injury. MiR-140-3p directly bound the Tyro3 mRNA 3'UTR. Rescue experiments indicated that the miR-140-3p mimic-induced changes in cell viability and protein expression were alleviated by Tyro3 overexpression and that the miR-140-3p inhibitor-induced changes in cell viability and protein expression were alleviated by silencing Tyro3. Tyro3 overexpression increased cell viability and PI3K and p-Akt protein expression, but these effects were weakened by the addition of LY294002. CONCLUSIONS: MiR-140-3p directly targets Tyro3 to regulate cell viability and apoptosis of OGD/R-exposed N2a cells through the PI3K/Akt pathway, suggesting that miR-140-3p is a novel biomarker and therapeutic target for ischaemic stroke.

The lncRNA NEAT1 Mediates Neuronal Cell Autophagy and Related Protein Expression After Cerebral Ischemia‒Reperfusion Injury.

The present study focuses on the role of the long noncoding RNA (lncRNA) NEAT1 in regulating autophagy during the ischemia‒reperfusion (I/R) injury process and its possible regulatory mechanism based on the results of laboratory experiments. Neuro-2a (N2a) cells and BV-2 microglial cells were cultured separately, and oxygen-glucose deprivation/reoxygenation (OGD/R) was induced in vitro to mimic cerebral I/R injury. The expression of lncRNA NEAT1 was measured after reoxygenation for different durations, and the results showed that NEAT1 expression was significantly different after OGD/R for 12 h; thus, cell models of NEAT1 overexpression and knockdown were constructed. Knockdown of NEAT1 effectively relieved reperfusion injury. In an N2a and BV-2 cell coculture system, knockdown of NEAT1 reduced autophagic flow in neuronal cells after reperfusion. To clarify the mechanism of NEAT1 after neuronal I/R injury, label-free quantitative proteomics (LFQ) was used to identify the differentially expressed proteins (DEPs) in NEAT1 knockdown neurons after OGD/R for 12 h. Additionally, Gene Ontology (GO) enrichment, protein‒protein interaction (PPI) network and parallel-reaction monitoring (PRM) quantitative analyses were carried out; the results showed that the expression levels of the autophagy-related proteins Gaa, Glb1, Prkaa1, Kif23, Sec24a and Vps25 were significantly reduced and that these proteins interact. In summary, this study shows that NEAT1 can regulate the interactions between autophagy-related proteins after neuronal I/R injury, reducing the level of autophagy and relieving neuronal reperfusion injury.