Applied Microbiology and Biotechnology最新文献

{"title":"Targeted genome editing of the non-model cyanobacterium Cyanothece PCC 7425 via CRISPR/Cas12a","authors":"Monis Athar Khan,&nbsp;Anne Durand,&nbsp;Fériel Skouri-Panet,&nbsp;Karim Benzerara,&nbsp;Corinne Cassier-Chauvat,&nbsp;Franck Chauvat,&nbsp;Soufian Ouchane","doi":"10.1007/s00253-026-13959-y","DOIUrl":"10.1007/s00253-026-13959-y","url":null,"abstract":"<p>Cyanobacteria are diverse photosynthetic microorganisms of great interest for fundamental science and sustainable biotechnological applications. However, their polyploidy makes genetic manipulation challenging and time-consuming. The development of CRISPR/Cas tools has greatly accelerated genome editing and metabolic engineering of some cyanobacterial model species. In this work, we extend the CRISPR/Cas12a system for targeted gene deletion in the non-model cyanobacterium <i>Cyanothece</i> sp. PCC 7425, interesting for its ability to perform intracellular calcium carbonate (CaCO₃) biomineralization, nitrogen fixation, etc. We demonstrate for the first time its tractability to gene knockout by generating deletion mutants of four genes (<i>cax3-cax4</i>, <i>gor</i>, and <i>sodB</i>) acting in metabolism and/or response to stresses, using Cas12a-mediated homologous recombination. Importantly, full chromosome segregation was rapidly achieved after a single round of selection in all cases. All mutants were genotypically and phenotypically characterised. Moreover, biochemical analysis in the case of the <i>ΔsodB</i> mutant further confirmed its targeted deletion. Overall, CRISPR/Cas12a provides a rapid and efficient system for genome editing in <i>Cyanothece</i> sp. PCC 7425, establishing this organism as a versatile model for studying oxidative stress pathways, metal toxicity, and moreover, the still poorly known mechanism(s) of intracellular CaCO₃ biomineralization.</p><p>• <i>Rapid and efficient CRISPR/Cas12a editing established in Cyanothece sp. PCC 7425.</i></p><p>• <i>Fully segregated knockout mutants obtained after a single selection round.</i></p><p>• <i>Platform for exploring the biotechnological potential of Cyanothece sp. PCC 7425.</i></p>","PeriodicalId":8342,"journal":{"name":"Applied Microbiology and Biotechnology","volume":"110 1","pages":""},"PeriodicalIF":4.7,"publicationDate":"2026-07-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://link.springer.com/content/pdf/10.1007/s00253-026-13959-y.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148418255","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Improving autotrophic D-lactate production by heterologous expression of a pyruvate formate lyase in an LDHD-expressing Acetobacterium woodii strain","authors":"Kira S. Baur,&nbsp;Anna Stock,&nbsp;Dirk Weuster-Botz,&nbsp;Frank R. Bengelsdorf","doi":"10.1007/s00253-026-13952-5","DOIUrl":"10.1007/s00253-026-13952-5","url":null,"abstract":"<div><p>The anaerobic acetogen <i>Acetobacterium woodii</i> is a promising candidate producing chemicals while converting the greenhouse gas CO₂. In this study, <i>A. woodii</i> was genetically engineered to express a lactate dehydrogenase (LDHD) from <i>Leuconostoc mesenteroides</i>, and a pyruvate formate lyase (PFL) as well as its activating enzyme (PFL-AE) originated from <i>Clostridium pasteurianum</i> to autotrophically produce D-lactate next to acetate, using H₂ + CO₂. In this study, an improved D-lactate production was observed compared to the control strain without PFL and PFL-AE. Especially, during anabolism, positive effects were determined due to the recombinant expression of <i>C. pasteurianum</i>’s PFL. During this work, the promoter system, used to control the expression of the recombinant <i>ldhD</i> gene, was exchanged to the lactate-inducible P_<i>lctA</i>-promoter originated from <i>A. woodii</i>, which also caused an increase in D-lactate production. Cultivation of these P_<i>lctA</i> promoter strains in a stirred-tank bioreactor (STR) resulted in elevated D-lactate and acetate concentrations compared to experiments in serum bottles and showed increased D-lactate accumulation during expression of the PFL compared to the strain carrying only the <i>ldhD</i> gene. A maximum D-lactate concentration of 21.0 ± 2.4 mM was reached in bottle-scale cultivation. Controlled STR batch cultivations with continuous gassing resulted in D-lactate concentrations of up to 34.9 mM. This finding demonstrates that <i>A. woodii</i> overexpressing LDHD, PFL, and PFL-AE is a robust strain for the autotrophic production of D-lactate.</p><h3>Graphical Abstract</h3>\u0000<div><figure><div><div><picture><source><img></source></picture></div></div></figure></div></div>","PeriodicalId":8342,"journal":{"name":"Applied Microbiology and Biotechnology","volume":"110 1","pages":""},"PeriodicalIF":4.7,"publicationDate":"2026-07-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://link.springer.com/content/pdf/10.1007/s00253-026-13952-5.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148418248","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Endophytic Aspergillus glaucus as a novel epothilone B producer: Antiproliferative activity and in silico analysis","authors":"Ashraf S. A. El-Sayed,&nbsp;Sara Refaat,&nbsp;May A. El-Sayed,&nbsp;Eman Fikry,&nbsp;Nora Tawfeek,&nbsp;Maher M. El-Domiaty,&nbsp;Azza M. El-Shafae","doi":"10.1007/s00253-026-13954-3","DOIUrl":"10.1007/s00253-026-13954-3","url":null,"abstract":"<p>Epothilones are among the most potent anticancer agents, particularly against drug-resistant tumors, owing to their high affinity for β-tubulin stabilization, inhibiting the microtubule disassembly, ultimately leading to cell cycle arrest. However, their limited natural availability has restricted broader clinical and industrial exploitation. Accordingly, this study aimed to screen novel fungal isolates with sustainable and high metabolic capacity for epothilone production. <i>Aspergillus glaucus</i> OR342865.1, an endophyte isolated from the palm tree <i>Latania lontaroides</i>, exhibited the highest epothilone productivity (240 µg/L) upon response surface methodology optimization. The chemical identity of the purified compound was resolved by different chromatographic and spectroscopic analyses, revealing a structural profile consistence with authentic epothilone B. The putative epothilone B displayed potent cytotoxic activity against HepG-2, MCF-7, and HCT-116 cell lines, with half-maximal inhibitory concentration (IC₅₀) values of 0.028, 0.034, and 0.13 µM, with selectivity indices 12.5, 10.3, and 2.7, respectively, relative to Vero cells. The anti-invasive assay demonstrates a significant suppression (35.5–40.2%) of migratory capacity in MCF-7 and HepG-2 cells following treatment, indicating antimetastatic potential. Cell cycle analysis revealed that epothilone treatment induced G1 phase arrest in HepG-2 cells, with a 55.03% increase compared to the control. The total and early apoptotic populations increased to 32.2% and 22.03%, respectively, while late apoptosis exhibited a 57.6-fold elevation relative to untreated cells. The biological productivity of <i>A. glaucus</i> for epothilone B was stably maintained for up to 6 months when preserved as a slope culture at 4 °C. Molecular docking analysis demonstrated that epothilone B exhibits coherent multitarget interactions with key oncogenic kinases, including SRC, EGFR, JAK2, PIK3CA, mTOR, and HSP90, in addition to binding the β-tubulin taxane site, highlighting its capacity to modulate multiple cancer-related signaling pathways. To our knowledge, this is the first report demonstrating the high metabolic potential of <i>A. glaucus</i> for epothilone B production, positioning this fungus as a promising platform for sustainable industrial-scale biosynthesis of this clinically valuable anticancer agent.</p><p>• <i>Aspergillus glaucus, an endophyte of Latania lontaroides, is a potential epothilone B producer.</i></p><p>• <i>The purified epothilone has plausible antiproliferative, apoptotic, and antimetastatic activities.</i></p><p>• <i>The epothilone had coherent multitarget interactions with key oncogenic kinases and β-tubulin taxane site.</i></p>","PeriodicalId":8342,"journal":{"name":"Applied Microbiology and Biotechnology","volume":"110 1","pages":""},"PeriodicalIF":4.7,"publicationDate":"2026-07-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://link.springer.com/content/pdf/10.1007/s00253-026-13954-3.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148418233","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Multi-omics profiling reveals antagonistic strategies of endophytic Streptomyces sp. MEPP0209 against Lasiodiplodia theobromae causing Dendrobium officinale stem rot","authors":"Zhili Jiang,&nbsp;Hailan Fu,&nbsp;Yujie Chen,&nbsp;Yongjing Zhang,&nbsp;Jing Shi,&nbsp;Jun Dai,&nbsp;Chenyang Qiao,&nbsp;Jie Gu,&nbsp;Jihong Jiang,&nbsp;Xiaoying Cao,&nbsp;Ludan Li","doi":"10.1007/s00253-026-13950-7","DOIUrl":"10.1007/s00253-026-13950-7","url":null,"abstract":"<div><p>Stem rot disease frequently occurs during the artificial cultivation of <i>Dendrobium officinale</i> Kimura et Migo, which has adverse effects on its yield and quality. Endophytic bacteria present in plants represented potential sources of biocontrol agents. This study isolated 43 distinct microbial strains from wild <i>Dendrobium moniliforme</i> (L.) Sw., among which strain MEPP0209 exhibited significant inhibition of <i>Lasiodiplodia theobromae</i> hyphal growth. Based on the 16S rDNA sequences, the strain was identified as <i>Streptomyces</i> sp. MEPP0209. The cell-free supernatant (CFS) of MEPP0209 significantly inhibited <i>L. theobromae</i> growth, and effectively reduced the disease index of stem rot in <i>D. officinale</i> tissue-cultured seedlings. Transcriptome analysis showed that differentially expressed genes (DEGs) related to cell division and cell cycle were significantly downregulated in <i>L. theobromae</i> hyphae after MEPP0209 CFS treatment, such as <i>MCM2</i>/<i>3</i>/<i>4</i>/<i>5</i>/<i>6</i>/<i>7</i>, <i>NDT80</i>, <i>POLA1</i>, and <i>MBP1</i>, while DEGs related to energy metabolism and oxidative stress were significantly upregulated. Metabolomics analysis and in vitro antagonistic assays indicated that the candidate antagonistic substances, 5,6-dimethylbenzimidazole and pyrrole-2-carboxylic acid, can effectively inhibit the growth of <i>L. theobromae</i>. And qRT-PCR results obtained from the pathogen treated with these two metabolites further indicated the downregulation of genes implicated in cell division and cell cycle. Overall, our findings suggested that the inhibition of fungal growth by MEPP0209 may be associated with the downregulation of genes related to cell division and proliferation in <i>L. theobromae</i>. The metabolites produced by MEPP0209, including 5,6-dimethylbenzimidazole and pyrrole-2-carboxylic acid, likely contributed to this inhibitory effect. This study provides a theoretical basis for future investigations into the role of MEPP0209 in biological control of <i>D. officinale</i> stem rot.</p></div>","PeriodicalId":8342,"journal":{"name":"Applied Microbiology and Biotechnology","volume":"110 1","pages":""},"PeriodicalIF":4.7,"publicationDate":"2026-07-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13638884/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148418199","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Evaluation of targeted Massively Parallel Sequencing methods for forensic metagenomics","authors":"Kamila Marszałek,&nbsp;Michał B. Kowalski,&nbsp;Agata Jagiełło,&nbsp;Anna Woźniak,&nbsp;Kinga Herda,&nbsp;Rafał Płoski,&nbsp;Andrzej Ossowski,&nbsp;Manuela Oliveira,&nbsp;Renata Zbieć-Piekarska,&nbsp;Paweł P. Łabaj,&nbsp;Wojciech Branicki","doi":"10.1007/s00253-026-13944-5","DOIUrl":"10.1007/s00253-026-13944-5","url":null,"abstract":"<p>Massively Parallel Sequencing (MPS) is effective for monitoring the microbial composition of environmental samples. Soil microbial signatures are critical for pinpointing the geographic location of forensic evidence, but standard 16S rRNA methods lack species-level resolution. Targeted sequencing panels, consisting of informative DNA fragments, can overcome this shortcoming and are highly desirable for forensic investigations. To address this, we evaluated three target enrichment methods for metagenomic analysis. First, we used Whole Metagenome Sequencing (WMS) data from 134 soil samples across 46 locations in Poland to extract a set of 200 markers. Using these markers, we created prototype targeted sequencing panels to compare two amplicon capture-based methods (Thermo Fisher AmpliSeq™ and Integrated DNA Technologies xGen™) and one hybridization capture-based method (Roche KAPA HyperPlus). The comparison of the technologies was guided by the results of classification of sample origin by machine learning classifier trained on feature profiles from WMS. The methods were assessed on technical parameters including data quality, reproducibility, sensitivity, and practical implementation for forensic laboratories. The performance and precision varied depending on technology and DNA concentration. The Roche KAPA HyperPlus hybridization capture-based method consistently demonstrated superior performance. Across various DNA input quantities, it showed the highest correlation with WMS data and achieved an exceptional F1 score of 0.94 at 5 ng, significantly outperforming the amplicon-based methods. This indicates that hybridization capture is a more robust and accurate approach for forensic soil microbiome profiling, particularly for low-template evidence, providing a highly reliable tool for predicting geographic origin.</p><p><i>• Targeted Massively Parallel Sequencing methods for forensic soil microbial analysis</i></p><p><i>• Targeted sequencing allowed the determination of the place of origin of soil samples</i></p><p><i>• Roche KAPA HyperPlus: the most accurate classification of the soil samples origin.</i>\u0000</p>","PeriodicalId":8342,"journal":{"name":"Applied Microbiology and Biotechnology","volume":"110 1","pages":""},"PeriodicalIF":4.7,"publicationDate":"2026-07-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13635074/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148418278","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Harnessing the potential of lignocellulosic biomass for biofuels production","authors":"Nutan Mhetras,&nbsp;Digambar Gokhale","doi":"10.1007/s00253-026-13947-2","DOIUrl":"10.1007/s00253-026-13947-2","url":null,"abstract":"<p>Currently, fossil fuels are the main and dominant sources for producing fuels and commodity chemicals. The rising demand for fossil fuels continues to entrench global dependency on non-renewable sources creating the climate risk, the economic instability and sustainability challenges. Hence, there is a need to shift towards globally available, sustainable and renewable resources such as lignocellulosic biomass (LCB) that offers a sustainable pathway for producing biofuels and chemicals. Though various technologies are available for LCB conversion to biofuels and chemicals, scaling up of biorefineries remains stifled by its recalcitrance nature and volatile supply chain economics. LCB processing often requires pretreatment to disrupt the rigid lignin–hemicellulose barrier and decrystallize cellulose. This structural opening is essential to maximize the enzymatic hydrolysis and sugar yields for biofuel production. The pretreatment process is energy-intensive and expensive accounting for 40% of the overall biofuel production cost followed by hydrolysis using expensive enzyme cocktails. These economic barriers currently limit the adoption of LCB as a cost-competitive fuel resource. The promising strategy for low cost LCB derived ethanol production is to adopt integrated biorefinery approach utilizing physical, chemical and biological processes. The integrated biorefinery approach tackles the high costs of second generation ethanol production by mimicking traditional petroleum refineries. It valorizes all three LCB components to marketable fuels and high-value chemicals maximizing the overall process profitability. This review discusses on the latest developments in biofuels production processes especially in relation to ethanol and butanol production.</p>","PeriodicalId":8342,"journal":{"name":"Applied Microbiology and Biotechnology","volume":"110 1","pages":""},"PeriodicalIF":4.7,"publicationDate":"2026-07-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13635065/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148418268","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Whey protein isolate enhances the growth and probiotic features of Lacticaseibacillus rhamnosus GG","authors":"Adrienne B. Narrowe,&nbsp;LinShu Liu,&nbsp;Venkateswari J. Chetty,&nbsp;Karley K. Mahalak,&nbsp;Jenni Firrman,&nbsp;Johanna M. S. Lemons","doi":"10.1007/s00253-026-13945-4","DOIUrl":"10.1007/s00253-026-13945-4","url":null,"abstract":"<p><i>Lacticaseibacillus rhamnosus</i> strain GG (LGG) is a broadly used probiotic with several unique features that help it provide beneficial effects to its host. Key among the probiotic features of LGG is the production of bioactive metabolites and secreted proteins such as p40 and p75. The ability of LGG to persist in the gastrointestinal tract depends primarily on its ability to adhere to the gut mucosa via the generation of adhesion pili. While LGG is already used as a probiotic, potential still exists for optimization of the metabolic state of LGG to further enhance its probiotic capacity. Here, we evaluated the ability of whey protein isolate to enhance the cell growth and probiotic effects of LGG. RNA sequencing and untargeted metabolomics revealed that WPI supplementation enhanced LGG growth and metabolic activity and increased the transcription of genes for the production of adhesion pili, key secreted proteins, and beneficial metabolites. These results indicate that whey protein is a viable supplement option for use with LGG and may help to boost the probiotic activity and growth of LGG within the gastrointestinal tract.</p><p>• <i>Whey protein isolate supplementation increases Lacticaseibacillus rhamnosus GG growth.</i></p><p>• <i>Transcription of genes for probiotic features is amplified by the addition of WPI.</i></p><p>• <i>Transcriptomics and metabolomics suggest the protein produces the beneficial effects.</i></p>","PeriodicalId":8342,"journal":{"name":"Applied Microbiology and Biotechnology","volume":"110 1","pages":""},"PeriodicalIF":4.7,"publicationDate":"2026-07-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13635073/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148418215","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Metagenomics comparison identifies shared pathogenic microbiome in humans, pigs and chickens","authors":"Sung J. Yu,&nbsp;Dragana Stanley,&nbsp;Thi T. H. Van,&nbsp;Jason C. Steel,&nbsp;Yadav S. Bajagai","doi":"10.1007/s00253-026-13948-1","DOIUrl":"10.1007/s00253-026-13948-1","url":null,"abstract":"<p>Integrating human, animal, and environmental health is crucial for combating infectious diseases, as an estimated 60 to 75% of emerging infectious diseases originate from zoonotic sources globally. In this study, we analysed 1274 shotgun metagenomic faecal samples of humans, pigs, and chickens collected across multiple countries to estimate levels of microbial sharing at the species-level genome bins (SGBs) resolution. We confirm that host species, rather than geography, significantly structures the gut microbial community, as shown by alpha and beta diversity analyses. Despite this high host specificity, we identified substantial cross-host sharing of SGBs, including taxa recognised as pathogens such as <i>Escherichia coli</i>, <i>Clostridium perfringens</i>, <i>Clostridium innocuum</i>, <i>Clostridium disporicum</i>, <i>Enterococcus</i> species, and<i> Streptococcus alactolyticus.</i> Core taxa were predominantly host-specific, while non-core taxa were more frequently shared across humans, pigs, and chickens. LEfSe analysis identified distinctive microbial signatures for each host, further supporting differences in community composition. These findings demonstrate that unrelated and geographically distant humans and livestock can harbour highly similar microbial populations with pathogenic potential. This work provides molecular evidence supporting the need for integrated One Health surveillance to better detect, manage, and prevent zoonotic and reverse zoonotic transmission events across interconnected human, animal, and environmental systems.</p><p>• <i>There is substantial cross-host sharing of species-level genome bins, including potential pathogens</i></p><p>• <i>Core taxa are predominantly </i><i>host-specific</i></p><p>• <i>Non-core taxa are more likely to be shared across humans, pigs, and chickens</i></p>","PeriodicalId":8342,"journal":{"name":"Applied Microbiology and Biotechnology","volume":"110 1","pages":""},"PeriodicalIF":4.7,"publicationDate":"2026-07-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13631169/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148403555","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Biocontrol of mushroom crop mycoparasites by novel Bacillus velezensis strains","authors":"William Kay,&nbsp;Jaime Carrasco,&nbsp;Souvik Kusari,&nbsp;Marjon Krijger,&nbsp;M. José Carpio,&nbsp;Thomas Barnes,&nbsp;M. Sonia Rodríguez Cruz,&nbsp;Jan van der Wolf,&nbsp;Till Bebenroth,&nbsp;Gail M. Preston","doi":"10.1007/s00253-026-13938-3","DOIUrl":"10.1007/s00253-026-13938-3","url":null,"abstract":"<p>The cultivation of button mushroom (<i>Agaricus bisporus</i>) requires the design of tailor-made substrates that nourish the crop and promote morphology changes from mycelium to basidiome. The agronomic stages of mushroom development are also influenced by the microbiota present in the mushroom crop microcosm. These microbes can have a beneficial impact on mushroom growth, development and quality, or a detrimental impact through reduction of yield or quality (parasites, competitors or disease vectors). In this report, we describe the isolation of multiple strains of <i>Bacillus velezensis</i> from mushroom casing material and basidiomes. We show that these strains exhibit antifungal activity towards major mushroom mycoparasites in vitro and further characterise their mode of action. Full genomes of <i>B. velezensis</i> CM5, CM19, CM35, EM5 and EM39 were sequenced and annotated, which together with metabolic profiling of specialised metabolites produced by CM5, CM19 and CM35 suggested that the antifungal activity of these strains is likely to be linked to the production of the lipopeptide fengycin. The addition of these <i>B. velezensis</i> strains to a growth chamber trial with crops infected by <i>Zarea fungicola</i> strain 150/1 did not result in a statistically significant reduction in disease incidence compared to the chemical fungicide prochloraz-Mn. Despite in vitro results, no negative effect on mushroom yield was observed. The analysis of the quantitative microbiome during this trial suggests that microbial dynamics is consistent with a regular crop cycle. Additionally, the application of <i>B. velezensis</i> strain CM5 resulted in increased Gram-negative and Gram-positive bacteria. Genomic and analytical tools were designed and used to evaluate <i>B. velezensis</i> persistence in casing soil when the selected strains were artificially applied. <i>B. velezensis</i> population levels decreased significantly after application, potentially contributing to the lack of biocontrol activity observed in growth chamber crop trials.</p><p>• <i>Bacillus velezensis strains isolated from peat-based microcosms show significant inhibitory effects against major fungal parasites of mushrooms in vitro.</i></p><p>• <i>Genome sequencing and metabolic profiling correlated antifungal activity with the production of lipopeptides, particularly fengycin.</i></p><p>• <i>Novel strains did not significantly limit dry bubble disease under growth chamber crop trials—potentially due to low bacterial persistence.</i></p>","PeriodicalId":8342,"journal":{"name":"Applied Microbiology and Biotechnology","volume":"110 1","pages":""},"PeriodicalIF":4.7,"publicationDate":"2026-07-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://link.springer.com/content/pdf/10.1007/s00253-026-13938-3.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148395549","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Total fungal RNA extraction from complex solid- state fermentation products","authors":"Mikael Terp,&nbsp;Christian Enrico Rusbjerg-Weberskov,&nbsp;Teis E. Sondergaard,&nbsp;Mette Lübeck","doi":"10.1007/s00253-026-13946-3","DOIUrl":"10.1007/s00253-026-13946-3","url":null,"abstract":"<p>Investigating fungal gene expression in complex solid-state fermentation (SSF) systems is essential for process optimization but is hindered by challenges in RNA extraction. RNA extraction from fungal samples rich in plant biomass is a special challenge that is not met by the commercial RNA extraction kits and conventional RNA extraction methods we have tested. This study presents an optimized RNA extraction method, suitable for RT-qPCR from agricultural side streams fermented by filamentous fungi. The method was applied to investigate the temporal expression of the class II hydrophobin NC2 in <i>Neurospora intermedia</i> grown on four different sources of brewer’s spent grain (BSG): IPA, Pilsner, Stout, and a microbrewery batch. Relative NC2 expression showed a clear temporal increase, peaking on day 6, with the microbrewery BSG batch having the highest peak expression. It demonstrates the applicability of the presented RNA extraction method to monitor expression of a gene of interest during solid-state fermentation of a complex substrate. Furthermore, the RNA extraction protocol was successfully validated on other challenging substrates, including rapeseed press cake and oat hulls, and with other fungi (<i>Aspergillus oryzae</i> and <i>Trichoderma asperellum</i>), consistently yielding high-purity RNA. This work provides a reliable method for RNA extraction from complex SSF products that can be used for gene expression analysis.</p><p>• <i>Reliable RNA extraction protocol developed for complex fungal SSF substrates</i></p><p>• <i>Hydrophobin NC2 expression in N. intermedia might be temporally regulated on BSG</i></p><p>• <i>Peak NC2 expression (day 6) was observed on BSG from a microbrewery</i></p>","PeriodicalId":8342,"journal":{"name":"Applied Microbiology and Biotechnology","volume":"110 1","pages":""},"PeriodicalIF":4.7,"publicationDate":"2026-07-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13620088/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148389770","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}