Applied and Environmental Microbiology最新文献

{"title":"Hydration conditions as a critical factor in antibiotic-mediated bacterial competition outcomes.","authors":"Yana Beizman-Magen, Tomer Orevi, Nadav Kashtan","doi":"10.1128/aem.02004-24","DOIUrl":"10.1128/aem.02004-24","url":null,"abstract":"<p><p>Antibiotic secretion plays a pivotal role in bacterial interference competition; yet, the impact of environmental hydration conditions on such competition is not well understood. Here, we investigate how hydration conditions affect interference competition among bacteria, studying the interactions between the antibiotic-producing <i>Bacillus velezensis</i> FZB42 and two bacterial strains susceptible to its antibiotics: <i>Xanthomonas euvesicatoria</i> 85-10 and <i>Pseudomonas syringae</i> DC3000. Our results show that wet-dry cycles significantly modify the response of the susceptible bacteria to both the supernatant and cells of the antibiotic-producing bacteria, compared to constantly wet conditions. Notably<i>, X. euvesicatoria</i> shows increased protection against both the cells and supernatants of <i>B. velezensis</i> under wet-dry cycles, while <i>P. syringae</i> cells become more susceptible under wet-dry cycles. In addition, we observed a reciprocal interaction between <i>P. syringae</i> and <i>B. velezensis</i>, where <i>P. syringae</i> inhibits <i>B. velezensis</i> under wet conditions. Our findings highlight the important role of hydration conditions in shaping bacterial interference competition, providing valuable insights into the microbial ecology of water-unsaturated surfaces, with implications for applications such as biological control of plant pathogens and mitigating antibiotic resistance.IMPORTANCEOur study reveals that hydration conditions, particularly wet-dry cycles, significantly influence antibiotic-mediated competition between bacterial species. We revealed that the effectiveness of antibiotics produced by <i>Bacillus velezensis</i> against two susceptible bacterial species: <i>Xanthomonas</i> and <i>Pseudomonas</i> varies based on these hydration conditions. Unlike traditional laboratory environments, many real-world habitats, such as soil, plant surfaces, and even animal skin, undergo frequent wet-dry cycles. These conditions affect bacterial competition dynamics and outcomes, with wet-dry cycles providing increased protection for some bacteria while making others more susceptible. Our findings highlight the importance of considering environmental hydration when studying microbial interactions and developing biological control strategies. This research has important implications for improving agricultural practices and understanding natural microbial ecosystems.</p>","PeriodicalId":8002,"journal":{"name":"Applied and Environmental Microbiology","volume":" ","pages":"e0200424"},"PeriodicalIF":3.9,"publicationDate":"2025-01-31","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11784440/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142875972","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Positive regulation of a LuxR family protein, MilO, in mildiomycin biosynthesis.","authors":"Zhiyu Li, Yuli Wang, Chen Lin, Yu Wen, Zixin Deng, Ming Jiang, Xinyi He","doi":"10.1128/aem.01654-24","DOIUrl":"10.1128/aem.01654-24","url":null,"abstract":"<p><p>Mildiomycin is a representative peptidyl nucleoside antibiotic and was first isolated from <i>Streptoverticillium rimofaciens</i>, which has been used as an important biological agent to control powdery mildew in plants. Despite its importance, the biosynthetic pathways and regulatory mechanisms remain to be fully elucidated. In this study, we identified MilO as a positive pathway-specific regulator of mildiomycin biosynthesis in the heterologous host <i>Streptomyces avermitilis</i>. Gene disruption of <i>milO</i> resulted in almost loss of mildiomycin production, and it was restored to the level comparable to that in the wild-type strain in complemented strain. Overexpression of <i>milO</i> using host native promoter <i>rpsJ</i>p, engineered promotor <i>SP44,</i> and <i>kasO</i>p* led to a 50%, 6.5-fold, and 9.2-fold increase in mildiomycin production compared with the wild-type strain, respectively. Quantitative real-time PCR and electrophoretic mobility shift assay (EMSA) experiments revealed that MilO directly enhances the transcription of the <i>milA</i> gene by 20 folds after 48 h fermentation and indirectly regulates the transcription levels of other genes from <i>milB</i> to <i>milM</i>. Using DNase I footprinting assays, <i>milO</i> was revealed to bind to a 44 bp DNA sequence of the <i>milA</i> promoter region. The binding region consists of three imperfect direct repeats of TGTC(N)₃CGGT separated by two-nucleotide spacers and each repeat is important to efficient binding to MilO. In addition, we identified two related compounds by overexpressing <i>milO</i> in a structural gene <i>milN</i>-deficient mutant. Taken together, this study indicates that pathway-specific regulator MilO is essential for mildiomycin biosynthesis and provides an effective strategy to improve the production of mildiomycin and its intermediates.IMPORTANCEAs an important biological agent to control powdery mildew on plants, mildiomycin has been commercialized and used in various plants. However, its regulatory mechanisms and biosynthetic pathways remain unknown. This study provides new insights into the regulation of mildiomycin biosynthesis through MilO, a LuxR family protein that modulates mildiomycin production by directly enhancing the transcription of <i>milA</i>. The yield of mildiomycin was significantly improved by overexpressing <i>milO</i> in a heterologous host. In addition, the positive regulatory effect of <i>milO</i> helped to discover two related compounds, which provide important clues for the timing of uploading of two amino acid side chains during mildiomycin biosynthesis for the first time. In brief, our findings on transcriptional regulation of mildiomycin biosynthesis by <i>milO</i> will be valuable to further increase the yield of mildiomycin and explore its biosynthetic pathways.</p>","PeriodicalId":8002,"journal":{"name":"Applied and Environmental Microbiology","volume":" ","pages":"e0165424"},"PeriodicalIF":3.9,"publicationDate":"2025-01-31","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11784345/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142876000","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Temporal dynamics of soil microbial C and N cycles with GHG fluxes in the transition from tropical peatland forest to oil palm plantation.","authors":"Frazer Midot, Kian Mau Goh, Kok Jun Liew, Sharon Yu Ling Lau, Mikk Espenberg, Ülo Mander, Lulie Melling","doi":"10.1128/aem.01986-24","DOIUrl":"10.1128/aem.01986-24","url":null,"abstract":"&lt;p&gt;&lt;p&gt;Tropical peatlands significantly influence local and global carbon and nitrogen cycles, yet they face growing pressure from anthropogenic activities. Land use changes, such as peatland forests conversion to oil palm plantations, affect the soil microbiome and greenhouse gas (GHG) emissions. However, the temporal dynamics of microbial community changes and their role as GHG indicators are not well understood. This study examines the dynamics of peat chemistry, soil microbial communities, and GHG emissions from 2016 to 2020 in a logged-over secondary peat swamp forest in Sarawak, Malaysia, which transitioned to an oil palm plantation. This study focuses on changes in genetic composition governing plant litter degradation, methane (CH&lt;sub&gt;4&lt;/sub&gt;), and nitrous oxide (N&lt;sub&gt;2&lt;/sub&gt;O) fluxes. Soil CO&lt;sub&gt;2&lt;/sub&gt; emission increased (doubling from approximately 200 mg C m&lt;sup&gt;-2&lt;/sup&gt; h&lt;sup&gt;-1&lt;/sup&gt;), while CH&lt;sub&gt;4&lt;/sub&gt; emissions decreased (from 200 µg C m&lt;sup&gt;-2&lt;/sup&gt; h&lt;sup&gt;-1&lt;/sup&gt; to slightly negative) following land use changes. The N&lt;sub&gt;2&lt;/sub&gt;O emissions in the oil palm plantation reached approximately 1,510 µg N m&lt;sup&gt;-2&lt;/sup&gt; h&lt;sup&gt;-1&lt;/sup&gt;, significantly higher than previous land uses. The CH&lt;sub&gt;4&lt;/sub&gt; fluxes were driven by groundwater table, humification levels, and C:N ratio, with &lt;i&gt;Methanomicrobia&lt;/i&gt; populations dominating methanogenesis and &lt;i&gt;Methylocystis&lt;/i&gt; as the main CH&lt;sub&gt;4&lt;/sub&gt; oxidizer. The N&lt;sub&gt;2&lt;/sub&gt;O fluxes correlated with groundwater table, total nitrogen, and C:N ratio with dominant &lt;i&gt;nirK&lt;/i&gt;-type denitrifiers (13-fold &lt;i&gt;nir&lt;/i&gt; to &lt;i&gt;nosZ&lt;/i&gt;) and a minor role by nitrification (a threefold increase in &lt;i&gt;amoA&lt;/i&gt;) in the plantation. &lt;i&gt;Proteobacteria&lt;/i&gt; and &lt;i&gt;Acidobacteria&lt;/i&gt; encoding incomplete denitrification genes potentially impact N&lt;sub&gt;2&lt;/sub&gt;O emissions. These findings highlighted complex interactions between microbial communities and environmental factors influencing GHG fluxes in altered tropical peatland ecosystems.IMPORTANCETropical peatlands are carbon-rich environments that release significant amounts of greenhouse gases when drained or disturbed. This study assesses the impact of land use change on a secondary tropical peat swamp forest site converted into an oil palm plantation. The transformation lowered groundwater levels and changed soil properties. Consequently, the oil palm plantation site released higher carbon dioxide and nitrous oxide compared to previous land uses. As microbial communities play crucial roles in carbon and nitrogen cycles, this study identified environmental factors associated with microbial diversity, including genes and specific microbial groups related to nitrous oxide and methane emissions. Understanding the factors driving microbial composition shifts and greenhouse gas emissions in tropical peatlands provides baseline information to potentially mitigate environmental consequences of land use change, leading to a broader impact on climate change mitigation ef","PeriodicalId":8002,"journal":{"name":"Applied and Environmental Microbiology","volume":" ","pages":"e0198624"},"PeriodicalIF":3.9,"publicationDate":"2025-01-31","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11784229/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142876004","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Papain expression in the <i>Escherichia coli</i> cytoplasm by T7-promoter engineering and co-expression with human protein disulfide isomerase (PDI) and thiol peroxidase (GPx7) genes.","authors":"Md Anarul Hoque, Richard A Gross, Mattheos A G Koffas","doi":"10.1128/aem.02119-24","DOIUrl":"10.1128/aem.02119-24","url":null,"abstract":"&lt;p&gt;&lt;p&gt;Difficulties exist in obtaining full-length, correctly folded, and soluble papain or papain-like proteases that necessitate the exploration of alternative strategies. This study describes the development of an &lt;i&gt;Escherichia coli&lt;/i&gt; strain capable of producing soluble papain without the need for complex and time-consuming &lt;i&gt;in vitro&lt;/i&gt; refolding steps. To enhance the production of soluble papain, engineered T7 promoters and a recombinant papain translationally fused with varying tags were constructed. The tags investigated include the maltose-binding protein, small ubiquitin modifier protein, and glutathione transferase. An &lt;i&gt;E. coli&lt;/i&gt; SHuffle strain was engineered to accumulate hydrogen peroxide (H&lt;sub&gt;2&lt;/sub&gt;O&lt;sub&gt;2&lt;/sub&gt;) by disruption of the redox pathway. This was accomplished by co-expression of the fusion constructs with two human endoplasmic reticulum-resident proteins, thiol peroxidase glutathione peroxidase-7 (GPx7), and protein disulfide isomerase (PDI). The oxidizing capacity of H&lt;sub&gt;2&lt;/sub&gt;O&lt;sub&gt;2&lt;/sub&gt; was used to improve disulfide bond formation in papain. The GPx7-PDI fusion dyad played a significant role in consuming harmful H&lt;sub&gt;2&lt;/sub&gt;O&lt;sub&gt;2&lt;/sub&gt; generated by the SHuffle cells. This consumption of H&lt;sub&gt;2&lt;/sub&gt;O&lt;sub&gt;2&lt;/sub&gt; helped provide the necessary oxidizing conditions for the efficient production of soluble papain. In shake-flask experiments, the recombinant strain produced ~110 mg/L of papain. Moreover, in batch fermentation, the volumetric yield reached ~349 mg/L. This work provides insights into recombinant papain microbial production that can lead to an industrial viable production strain.&lt;/p&gt;&lt;p&gt;&lt;strong&gt;Importance: &lt;/strong&gt;Papain, a cysteine-like protease, has extensive applications across various industries including food, chemical, pharmaceutical, drug, and polymer. However, the traditional isolation of papain from &lt;i&gt;Carica papaya&lt;/i&gt; plants results in a complex mixture of proteases. Such protease mixtures result in an inability to understand which component enzyme contributed to substrate conversions. Concentrations of constituent enzymes likely differ based on the ripeness of the papaya fruit. Also, constituent enzymes from papaya differ in optimal activity as a function of temperature and pH. Thus, by using papain-like enzymes from papaya fruit, valuable information on component enzyme activity and specificity is lost. Numerous methods have been reported to purify papain and papain-like enzymes from the crude mixture. Often, methods involve at least three steps including column chromatography to separate five cysteine proteases. Such procedures represent tedious processes to manufacture the pure enzymes in &lt;i&gt;Carica papaya&lt;/i&gt; extracts. The numerous uses of papain for industrial processes, as well as the probability that certain components of papain crude mixtures will be preferred for specific applications, necessitate alternative methods such as recombinant expression from microbial production ","PeriodicalId":8002,"journal":{"name":"Applied and Environmental Microbiology","volume":" ","pages":"e0211924"},"PeriodicalIF":3.9,"publicationDate":"2025-01-31","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11784408/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142715077","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Postdocs should receive relocation benefits from the universities that hire them.","authors":"Zakee L Sabree, Kayla Cross, James Gentry, Katie McAfee","doi":"10.1128/aem.01483-24","DOIUrl":"10.1128/aem.01483-24","url":null,"abstract":"<p><p>Postdocs are essential to microbial science and STEM academic workforces but are underpaid and receive little-to-no relocation benefits. PhDs foregoing postdoctoral training for lucrative industry and government jobs exit the academic pipeline, which imperils current scholarship and the future professoriate. Relocation to postdoc jobs is expensive, especially for recent graduates and international scholars, but academia rarely provides support. Solving this short-term liquidity pressure can increase productivity, job satisfaction, and the likelihood they remain in academia.</p>","PeriodicalId":8002,"journal":{"name":"Applied and Environmental Microbiology","volume":" ","pages":"e0148324"},"PeriodicalIF":3.9,"publicationDate":"2025-01-31","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11784396/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142708529","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Ecological features of microbial community linked to stochastic and deterministic assembly processes in acid mine drainage.","authors":"Zhenghua Liu, Chengying Jiang, Zhuzhong Yin, Ibrahim Ahmed Ibrahim, Teng Zhang, Jing Wen, Lei Zhou, Guoping Jiang, Liangzhi Li, Zhendong Yang, Ye Huang, Zhaoyue Yang, Yabing Gu, Delong Meng, Huaqun Yin","doi":"10.1128/aem.01028-24","DOIUrl":"10.1128/aem.01028-24","url":null,"abstract":"<p><p>Ecological processes greatly shape microbial community assembly, but the driving factors remain unclear. Here, we compiled a metagenomic data set of microbial communities from global acid mine drainage (AMD) and explored the ecological features of microbial community linked to stochastic and deterministic processes from the perspective of species niche position, interaction patterns, gene functions, and viral infection. Our results showed that dispersal limitation (DL) (48.5%~93.5%) dominated the assembly of phylogenetic bin in AMD microbial community, followed by homogeneous selection (HoS) (3.1%~39.2%), heterogeneous selection (HeS) (1.4%~22.2%), and drift (DR) (0.2%~2.7%). The dominant process of dispersal limitation was significantly influenced by niche position in temperature (<i>r</i> = -0.518, <i>P</i> = 0.007) and dissolved oxygen (<i>r</i> = 0.471, <i>P</i> = 0.015). Network stability had a significantly negative correlation with the relative importance of dispersal limitation, while it had a positive correlation with selection processes, implying changes in network properties could be mediated by ecological processes. Furthermore, we found that ecological processes were mostly related to the gene functions of energy production and conversion (C), and amino acid transport and metabolism (E). Meanwhile, our results showed that the number of proviruses and viral genes involved in arsenic (As) resistance is negatively associated with the relative importance of ecological drift in phylogenetic bin assembly, implying viral infection might weaken ecological drift. Taken together, these results highlight that ecological processes are associated with ecological features at multiple levels, providing a novel insight into microbial community assembly in extremely acidic environments.</p><p><strong>Importance: </strong>Unraveling the forces driving community assemblage is a core issue in microbial ecology, but how ecological constraints impose stochasticity and determinism remains unknown. This study presents a comprehensive investigation to uncover the association of ecological processes with species niche position, interaction patterns, microbial metabolisms, and viral infections, which provides novel insights into community assembly in extreme environments.</p>","PeriodicalId":8002,"journal":{"name":"Applied and Environmental Microbiology","volume":" ","pages":"e0102824"},"PeriodicalIF":3.9,"publicationDate":"2025-01-31","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11784436/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142827224","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Mitigating genetic instability caused by the excision activity of the <i>phi</i>C31 integrase in <i>Streptomyces</i>.","authors":"Yadan Duan, Zhangliang Liu, Xiaofang Huang, Lu Xu, Xianxue Wang, Hao Liu, Zhoujie Xie","doi":"10.1128/aem.01812-24","DOIUrl":"10.1128/aem.01812-24","url":null,"abstract":"<p><p>Over the past three decades, the integrase (Int) from <i>Streptomyces</i> phage <i>phi</i>C31 has become a valuable genome engineering tool across various species. <i>phi</i>C31 Int was thought to mediate unidirectional site-specific integration (<i>attP</i> × <i>attB</i> to <i>attL</i> and <i>attR</i>) in the absence of the phage-encoded recombination directionality factor (RDF). However, we have shown in this study that Int can also catalyze reverse excision (<i>attL</i> × <i>attR</i> to <i>attP</i> and <i>attB</i>) at low frequencies in <i>Streptomyces lividans</i> and <i>Escherichia coli</i>, causing genetic instability in engineered strains. To address this issue, we developed a two-plasmid co-conjugation (TPC) system. This system consists of an <i>attP</i>-containing integration vector and an Int expression suicide plasmid, both carrying <i>oriT</i> to facilitate efficient conjugation transfer from <i>E. coli</i> to <i>Streptomyces</i>. Using the TPC system, genetically stable integrants free of Int can be generated quickly and easily. The indigoidine-producing strains generated by the TPC system exhibited higher genetic stability and production efficiency compared to the indigoidine-producing strain generated by the conventional integration system, further demonstrating the utility of the TPC system in the field of biotechnology. We anticipate that the strategies presented here will be widely adopted for stable genetic engineering of industrial microbes using phage integrase-based integration systems.IMPORTANCELarge serine recombinases (LSRs), including the bacteriophage <i>phi</i>C31 integrase, were previously thought to allow only unidirectional site-specific integration (<i>attP</i> × <i>attB</i> to <i>attL</i> and attR). Our study is the first to show that the <i>phi</i>C31 integrase can also catalyze a low-efficiency reverse excision reaction in <i>Streptomyces</i> and <i>E. coli</i> without the involvement of the phage-encoded recombination directionality factor (RDF). The genetic instability caused by the low <i>in vivo</i> excisionase activity of the <i>phi</i>C31 integrase is a major challenge for biotechnological applications. Our study addresses this issue by developing a two-plasmid co-conjugation (TPC) system that facilitates the construction of Int-deficient genomic engineering strains. The Int-deficient integrants produced by this TPC system exhibit strong genetic stability for introduced genes and maintain stable production traits even in the absence of selection pressure, making them highly valuable for industrial applications.</p>","PeriodicalId":8002,"journal":{"name":"Applied and Environmental Microbiology","volume":" ","pages":"e0181224"},"PeriodicalIF":3.9,"publicationDate":"2025-01-31","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11784100/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142863172","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Regulation mechanism of the long-chain <i>n</i>-alkane monooxygenase gene <i>almA</i> in <i>Acinetobacter venetianus</i> RAG-1.","authors":"Shuai Chen, Lu Cao, Tianhua Lv, Jia Liu, Ge Gao, Mingchang Li, Liyuan Sun, Wenzhuo Tian, Yutong Tian, Guoqiang Li, Ting Ma","doi":"10.1128/aem.02050-24","DOIUrl":"10.1128/aem.02050-24","url":null,"abstract":"&lt;p&gt;&lt;p&gt;As toxic pollutants, &lt;i&gt;n&lt;/i&gt;-alkanes are pervasively distributed in most environmental matrices. Although the alkane monooxygenase AlmA plays a critical role in the metabolic pathway of solid long-chain &lt;i&gt;n&lt;/i&gt;-alkanes (≥C&lt;sub&gt;20&lt;/sub&gt;) that are extremely difficult to degrade, the mechanism regulating this process remains unclear. Here, we characterized the function of AlmA in &lt;i&gt;Acinetobacter venetianus&lt;/i&gt; RAG-1, which was mainly involved in the degradation of long-chain &lt;i&gt;n&lt;/i&gt;-alkanes (C&lt;sub&gt;26&lt;/sub&gt;-C&lt;sub&gt;38&lt;/sub&gt;), among which, &lt;i&gt;n&lt;/i&gt;-C&lt;sub&gt;32&lt;/sub&gt; induced the &lt;i&gt;almA&lt;/i&gt; promoter activity most. APR1 (&lt;u&gt;A&lt;/u&gt;lmA &lt;u&gt;P&lt;/u&gt;ositive &lt;u&gt;R&lt;/u&gt;egulator) that it is an AraC/XylS-type transcription regulator, a potential transcriptional regulator of &lt;i&gt;almA&lt;/i&gt;, was screened by DNA-pull down, which was determined by conserved domain analysis. The deletion of &lt;i&gt;apR1&lt;/i&gt; severely reduced the capacity of strain RAG-1 to utilize long-chain &lt;i&gt;n&lt;/i&gt;-alkanes (C&lt;sub&gt;22&lt;/sub&gt;-C&lt;sub&gt;38&lt;/sub&gt;), indicating the involvement of APR1 in &lt;i&gt;n&lt;/i&gt;-alkanes degradation. The results of the APR1-dependent reporter system, electrophoretic mobility shift assay, and microscale thermophoresis further proved that APR1 was able to directly bind to the &lt;i&gt;almA&lt;/i&gt; promoter region, thus activating the &lt;i&gt;almA&lt;/i&gt; transcription. Furthermore, APR1 could inhibit self-expression through autoregulation in the absence of long-chain &lt;i&gt;n&lt;/i&gt;-alkanes. &lt;i&gt;n&lt;/i&gt;-C&lt;sub&gt;32&lt;/sub&gt; acted as a ligand of APR1, and the amino acid residues Val10, Gln50, Ala99, and Ile106 at the N-terminus of APR1 were necessary for binding &lt;i&gt;n&lt;/i&gt;-C&lt;sub&gt;32&lt;/sub&gt;. In addition, the key amino acid residues of APR1 within the C-terminal helix-turn-helix motif that bound to the downstream promoter region were confirmed by multiple sequence alignment and site-directed mutagenesis. The homologs of AlmA and APR1 shared a similar evolutionary course in the &lt;i&gt;Proteobacteria&lt;/i&gt;; thus, this mode of regulation might be relatively conserved.&lt;/p&gt;&lt;p&gt;&lt;strong&gt;Importance: &lt;/strong&gt;The extreme hydrophobicity of long-chain &lt;i&gt;n&lt;/i&gt;-alkanes ({greater than or equal to}C&lt;sub&gt;20&lt;/sub&gt;) presents a significant challenge to their degradation in natural environments. It is, therefore, imperative to elucidate the regulatory mechanisms of the metabolic pathways of long-chain &lt;i&gt;n&lt;/i&gt;-alkanes, which will be of great significance for the future application of hydrocarbon-degrading bacteria to treat oil spills. However, the majority of current studies have focused on the regulatory mechanisms of short- and medium-chain &lt;i&gt;n&lt;/i&gt;-alkanes, with long-chain &lt;i&gt;n&lt;/i&gt;-alkanes receiving comparatively little attention. In this study, we identified APR1, a transcriptional regulator of the alkane monooxygenase AlmA in &lt;i&gt;Acinetobacter venetianus&lt;/i&gt; RAG-1, and characterized its function and regulatory mechanism. In the presence of ligand &lt;i&gt;n&lt;/i&gt;-C&lt;sub&gt;32&lt;/sub&gt;, APR1 could directly activate the transcription of &lt;i&gt;almA&lt;/i&gt;, which was involved","PeriodicalId":8002,"journal":{"name":"Applied and Environmental Microbiology","volume":" ","pages":"e0205024"},"PeriodicalIF":3.9,"publicationDate":"2025-01-31","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11784139/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142891487","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"A microaerobically induced small heat shock protein contributes to <i>Rhizobium leguminosarum</i>/<i>Pisum sativum</i> symbiosis and interacts with a wide range of bacteroid proteins.","authors":"Lucía Domingo-Serrano, Claudia Sanchis-López, Carla Alejandre, Joanna Soldek, José Manuel Palacios, Marta Albareda","doi":"10.1128/aem.01385-24","DOIUrl":"10.1128/aem.01385-24","url":null,"abstract":"<p><p>During the establishment of the symbiosis with legume plants, rhizobia are exposed to hostile physical and chemical microenvironments to which adaptations are required. Stress response proteins including small heat shock proteins (sHSPs) were previously shown to be differentially regulated in bacteroids induced by <i>Rhizobium leguminosarum</i> bv. viciae UPM791 in different hosts. In this work, we undertook a functional analysis of the host-dependent sHSP RLV_1399. A <i>rlv_1399-</i>deleted mutant strain was impaired in the symbiotic performance with peas but not with lentil plants. Expression of <i>rlv_1399</i> gene was induced under microaerobic conditions in a FnrN-dependent manner consistent with the presence of an anaerobox in its regulatory region. Overexpression of this sHSP improves the viability of bacterial cultures following exposure to hydrogen peroxide and to cationic nodule-specific cysteine-rich (NCR) antimicrobial peptides. Co-purification experiments have identified proteins related to nitrogenase synthesis, stress response, carbon and nitrogen metabolism, and to other relevant cellular functions as potential substrates for RLV_1399 in pea bacteroids. These results, along with the presence of unusually high number of copies of <i>shsp</i> genes in rhizobial genomes, indicate that sHSPs might play a relevant role in the adaptation of the bacteria against stress conditions inside their host.IMPORTANCEThe identification and analysis of the mechanisms involved in host-dependent bacterial stress response is important to develop optimal <i>Rhizobium</i>/legume combinations to maximize nitrogen fixation for inoculant development and might have also applications to extend nitrogen fixation to other crops. The data presented in this work indicate that sHSP RLV_1399 is part of the bacterial stress response to face specific stress conditions offered by each legume host. The identification of a wide diversity of sHSP potential targets reveals the potential of this protein to protect essential bacteroid functions. The finding that nitrogenase is the most abundant RLV_1399 substrate suggests that this protein is required to obtain an optimal nitrogen-fixing symbiosis.</p>","PeriodicalId":8002,"journal":{"name":"Applied and Environmental Microbiology","volume":" ","pages":"e0138524"},"PeriodicalIF":3.9,"publicationDate":"2025-01-31","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11784457/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142875968","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Patterns of spontaneous and induced genomic alterations in <i>Yarrowia lipolytica</i>.","authors":"Yuan-Ru Xiong, Yuan-Chun Fang, Min He, Ke-Jing Li, Lei Qi, Yang Sui, Ke Zhang, Xue-Chang Wu, Liang Meng, Ou Li, Dao-Qiong Zheng","doi":"10.1128/aem.01678-24","DOIUrl":"10.1128/aem.01678-24","url":null,"abstract":"<p><p>This study explored the genomic alterations in <i>Yarrowia lipolytica</i>, a key yeast in industrial biotechnology, under both spontaneous and mutagen-induced conditions. Our findings reveal that spontaneous mutations occur at a rate of approximately 4 × 10^-10 events per base pair per cell division, primarily manifesting as single-nucleotide variations (SNVs) and small insertions and deletions (InDels). Notably, C-to-T/G-to-A transitions and C-to-A/G-to-T transversions dominate the spontaneous SNVs, while 1 bp deletions, likely resulting from template slippage, are the most frequent InDels. Furthermore, chromosomal aneuploidy and rearrangements occur, albeit at a lower frequency. Exposure to ultraviolet (UV) light, methylmethane sulfonate (MMS), and Zeocin significantly enhances the rates of SNVs and alters their mutational spectra in distinct patterns. Notably, Zeocin-induced SNVs are predominantly T-to-A and T-to-G substitutions, often occurring within the 5'-TGT^*-3' motif (^* denotes the mutated base). Additionally, Zeocin exhibits a higher potency in stimulating InDels compared to UV and MMS. Translesion DNA synthesis is implicated as the primary mechanism behind most Zeocin-induced SNVs and some InDels, whereas non-homologous end joining serves as the main pathway for Zeocin-mediated InDels. Intriguingly, the study identifies the gene <i>YALI1_E21053g</i>, encoding a protein kinase, as negatively associated with Zeocin resistance. Overall, our results not only deepened our knowledge about the genome evolution in <i>Y. lipolytica</i> but also provided reference to develop innovative strategies to harness its genetic potential.IMPORTANCE<i>Yarrowia lipolytica</i> exhibits high environmental stress tolerance and lipid metabolism capabilities, making it a microorganism with significant industrial application potential. In this study, we investigated the genomic variation and evolutionary patterns of this yeast under both spontaneous and induced mutation conditions. Our results reveal distinctive mutation spectra induced by different mutagenic conditions and elucidate the underlying genetic mechanisms. We further highlight the roles of non-homologous end joining and translesion synthesis pathways in Zeocin-induced mutations, demonstrating that such treatments can rapidly confer drug resistance to the cells. Overall, our research enhances the understanding of how yeast genomes evolve under various conditions and provides guidance for developing more effective mutagenesis and breeding techniques.</p>","PeriodicalId":8002,"journal":{"name":"Applied and Environmental Microbiology","volume":" ","pages":"e0167824"},"PeriodicalIF":3.9,"publicationDate":"2025-01-31","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11784153/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142875996","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}