Applied Microbiology and Biotechnology最新文献

Acute-on-chronic liver failure (ACLF) is a severe condition arising from chronic liver disease, characterized by acute decompensation, organ failure, and high short-term mortality. Poor outcomes have also been observed in patients with ACLF after liver transplantation (LT). Emerging evidence, including a study from our center, suggests that gut microbiota plays an important role in ACLF. Patients who underwent LT at our center between October 2022 and June 2024 were included. Fecal samples were collected within 1 month post-LT for 16S rRNA and untargeted metabolomic sequencing. In this study, 144 samples from 69 patients with ACLF, cirrhosis, or hepatocellular carcinoma (HCC) were analyzed. Distinct microbiota and metabolic profiles were observed among the groups. ACLF patients exhibited significantly altered beta diversity, with notable depletion of g__Anaerostipes. Metabolomic analysis revealed substantial differences, including enrichment of tangeritin and depletion of candesartan in the ACLF group. Network analysis identified g__Anaerostipes as a key node linking differential taxa and metabolites. A random forest model based on these features effectively distinguished patient groups, with the highest classification accuracy observed in HCC. Multi-omic signatures were also associated with early allograft dysfunction (EAD), particularly g__Lachnoclostridium. Several microbial and metabolic features, including g__Lachnoclostridium, showed significant correlations with clinical indicators. The gut microbiome after LT is closely associated with ACLF. This study offers valuable insights for further investigation into the pathogenesis and post-LT prognosis.

• ACLF patients have a unique gut microbiota and metabolic profile after LT

• g__Anaerostipes is the prominent biomarker of ACLF's multi-omics signature

• g__Lachnoclostridium is a promising indicator of recovery after LT

Marine plastic litter, including microplastics, has a profound impact on the ocean and its wildlife, and strategies to remove/eliminate it are needed. Microbial biodegradation, particularly by bacteria, offers a potential solution, where a link between hydrocarbon and plastic-degradation has been hypothesized. This study screened the plastic-degrading potential of 18 bacterial strains isolated from 1-month-old biofilms developed in three submerged plastic fishing nets (braided polyethylene (PE), braided nylon, thin nylon). In addition, three highly efficient hydrocarbon-degrading strains were also tested. Strains were cultivated on solid minimal media with fishing net small pieces (new/unused nets) added as a carbon source for 1 month, followed by tributyrin-agar assays to assess esterase/lipase activity. Eleven bacteria exhibited enhanced growth with net polymers, mainly from the genera Sulfitobacter, Rhodococcus, Bacillus, and Pseudomonas, and eight of which bacteria also demonstrated esterase/lipase activity. Then, genes encoding hydrocarbon or plastic-degrading enzymes (alkB and almA homologs, PETase-like enzymes) were screened by PCR in the 21 mentioned bacteria and in ca.100 other strains found in submerged nets biofilm. Amplification of the investigated genes was predominantly observed in Actinomycetes strains. Genome mining of six promising strains revealed hits with enzymes linked to degradation of synthetic polymers like polyethylene terephthalate, low-density PE and nylon. The workflow developed here enabled the selection of marine bacteria with plastic-degrading potential, sourced from biofilms of submerged plastic fishing nets and hydrocarbon-enriched environments.

• A comprehensive lab workflow was developed to assess plastic-degrading potential.

• Genome mining in Rhodococcus and Pseudomonas strains revealed plastic-degrading enzymes.

• Hydrocarbon-degrading bacteria could hold plastic-degrading capabilities.

The metabolism of soy isoflavones by gut microbiota is critical for the bioactivation and bioavailability of these compounds, particularly daidzein, which is further metabolized by gut bacteria to produce S-equol. S-equol, an exclusive gut bacterial metabolite, is associated with health benefits such as reduced blood pressure, cardiovascular disease prevention, and protection against hormone-related cancers due to its estrogen-mimicking structure and antioxidant properties. However, the limited availability of S-equol-producing bacteria has hindered its production and utilization. This study investigates the isolation and characterization of S-equol-producing microbes from albino Wistar rats and explores the impact of dietary interventions on S-equol production. Preliminary tests showed that both dietary groups excreted more S-equol in feces than urine, with rats on fermented soy feed showing higher S-equol levels due to the presence of daidzein, a precursor. In this study, we isolated four anaerobic S-equol-producing bacteria — MG1 (PX459562), MG2 (PX459563), MG3 (PX459564), and MG4 (PX459565) from the intestine and feces of albino Wistar rats. High-Performance Thin-Layer Chromatography (HPTLC) and High-Performance Liquid Chromatography (HPLC) confirmed the presence of S-equol, with concentrations ranging from 5.90 to 7.56 µg/g of fermented soybean across different strains. Phylogenetic analysis revealed that the isolates belonged to the Enterobacteriaceae and Enterococcaceae families, identifying MG1 as C. freundii strain ATCC 8090, MG2 as Escherichia fergusonii strain NBRC 102419, and both MG3 and MG4 as Enterococcus faecalis strain NBRC 100480. Our findings underscore the significant role of gut microbiota in metabolizing daidzein into S-equol, highlighting the potential for utilizing these bacterial strains in functional food development and therapeutic applications. While the pathogenic nature of E. fergusonii (MG2) precludes its therapeutic use, strains MG1, MG3, and MG4, which match common commensal bacteria, show promise for commercial S-equol production and may serve as valuable resources for further investigation and utilization in promoting health and preventing associated diseases.

• Dietary intervention modulates gut microbiota in albino Wistar rats.

• Soybean fermentation enables efficient conversion of daidzin to bioactive S-equol.

• Novel S-equol–producing microbes were isolated and identified.

Short- and medium chain carboxylic acids (SCCA/MCCA) are natural antimicrobials produced by fermentation and chain elongation, but currently only a few SCCA/MCCA are used in food-related applications. With the aim to diversify the SCCA/MCCA profile of fermentates for biopreservation, we designed bioprocesses employing bacterial or multi-kingdom consortia to produce caproate-containing fermentates using a targeted cross-feeding strategy. We combined Limosilactobacillus reuteri, Clostridium kluyveri, and Saccharomyces cerevisiae, and quantified substrates utilization and metabolites. The antimicrobial activity of SCCA/MCCA and fermentates was analysed in vitro and in a meat model system. In a first bioprocess, the addition of ethanol (EtOH) initiated caproate formation by the bacterial consortium with levels of 63.9 mM (7.4 g L⁻¹) after 22 days. Next, we run two shorter bioprocesses (12 days) and more caproate (28.9 mM, 3.4 g L⁻¹) was produced if EtOH was regularly added than without EtOH supplementation (21.7 mM, 2.5 g L⁻¹). When S. cerevisiae was included, 37.9 ± 11.4 mM (4.4 g L⁻¹) caproate was formed without EtOH addition. Beyond the intended cross-feeding activity, consortia produced and re-metabolised mannitol and glycerol. Lactate and caproate were the major carboxylic acids in fermentates. Caproate inhibited bacteria, yeast and molds at pH 4.5 and 6.5 in vitro, while lactate and caproate acted synergistically in minced meat. Fermentates conferred antimicrobial activity against indicator microbes mostly at pH 4.5 and to a lesser extent at pH 6.5. In this study we successfully designed a consortium to produce caproate. We provide evidence that caproate-containing fermentates had strong antimicrobial activity, and our results indicate the complexity of metabolic interactions that occurred within consortium members.

• Design of a self-contained bioprocess with multi-kingdom consortium.

• Cross-feeding of bacteria and yeast to produce caproate fermentates.

• Fermentates containing caproate inhibited bacteria, yeast and mold growth.

Soil-borne microbial diseases constitute a significant constraint on crop production, causing substantial annual economic losses estimated at 40%–60% in agricultural sectors. This study aimed to identify the principal plant pathogens causing diseases in Brassica crops in Ningxia, China, and the factors and mechanisms of infection. Employing ITS and 16S rRNA gene high-throughput sequencing, we investigated the soil microbial community associated with both healthy and diseased Brassica crops, as well as other main crops. We detected significant abundances of the soil-borne pathogenic fungi Fusarium and Olpidium, with Fusarium markedly enriched in diseased soils, suggesting its potential role in pathogenesis. Furthermore, the disease presence induced substantial disturbance in the soil fungal community, whereas the prokaryotic community structure remained relatively unaffected. Spatial and environmental factors exerted a more pronounced influence on fungal communities than on prokaryotic communities, with disease incidence attenuating the impact of environmental variables on fungal assemblages. Further analysis revealed that soil properties positively drove Fusarium abundance, and this proliferation synergized with soil factors to reduce fungal evenness. This evenness loss, coupled with the pathogen’s direct effect, ultimately led to network simplification and a vulnerable “low evenness–high complexity” state. Network analyses confirmed this vulnerability, showing heightened complexity but diminished stability in fungal networks within diseased soils. Collectively, our findings provide a theoretical and technical framework for the rapid identification and effective management of pathogens affecting Brassica crops.

Listeriosis is a foodborne infection caused by Listeria monocytogenes that causes febrile gastroenteritis and central nervous system infections and that can often lead to fatality. Upon consumption of contaminated food, Listeria is able to survive a number of gastrointestinal stressors, including competition with the host microbiota. The emergence of antibiotic-resistant clones of L. monocytogenes, together with the side effects of antibiotic treatment, highlights the need for alternatives or additives for its treatment and prevention. Saccharomyces boulardii is a probiotic yeast that is often used alongside antibiotics to minimize side effects since it is not affected by them as a result of its eukaryotic nature. Furthermore, it can be engineered to produce a wide range of molecules. We previously engineered Saccharomyces cerevisiae through CRISPR-Cas9 integration to produce Ply511, a bacteriophage endolysin active against L. monocytogenes, showing the potential of engineered yeast to produce endolysins for biocontrol. In this study, we extended this approach to the probiotic yeast S. boulardii and directly compared the two yeasts as secretion hosts for Ply511. Using a simulated human gastrointestinal environment, we evaluated their ability to retain endolysin activity and reduce L. monocytogenes levels. We then tested the cell extracts from both yeasts in a bacterial consortium termed SImplified HUman intestinal MIcrobiota (SIHUMI), confirming a specificity for Listeria. Finally, we evaluated their activity in a simulated intestinal fermentation using fecal samples from human donors. Overall, this study demonstrates the potential of delivering endolysins to the gut via engineered probiotic S. boulardii.

The advent of nanotechnology has revolutionised approaches to identifying and combating bacterial infections. This review highlights the latest applications of nanoparticles (NPs) for bacterial detection and treatment, with a focus on their translational potential in clinical settings. We discuss advanced nanotechnology-enabled biosensing platforms that offer ultra-sensitive, rapid and precise diagnostics capabilities crucial for addressing antibiotic-resistant pathogens. In addition to detection, various nanoparticles demonstrate multiple antibacterial mechanisms and function as targeted drug-delivery vehicles. The review also examines current clinical trials involving nanoparticle-based therapeutics, underscoring their promise for overcoming antimicrobial resistance.

• Nanoparticles enable rapid and sensitive detection of bacterial infections.

• Multiple antibacterial mechanisms improve efficacy against resistant pathogens.

• Translational challenges must be addressed for successful clinical application.

Vibrio spp. are significant zoonotic pathogens in seafood, causing human gastroenteritis and posing challenges to the aquaculture industry. The misuse of antibiotics in aquaculture has led to the rise of antibiotic-resistant strains, necessitating alternative solutions like phages for food safety and pathogen control. Herein, we isolated and characterized a novel lytic Vibrio phage, BUCT787, from salmon aquaculture environments. Genomic analysis revealed that BUCT787 shared only 68% genome-wide coverage and 95.97% sequence identity with Vibrio phage 207E29.1, classifying it as a new member of an unclassified family within the Caudoviricetes order. The phage genome contained 86 predicted open reading frames (ORFs), with only 29 ORFs encoding proteins with known functions. BUCT787 exhibited stability across a wide range of temperatures (4°C–36°C) and pH conditions (4–12). To optimize infection efficiency, the optimal MOI for BUCT787 was determined to be 0.01, and the one-step growth curve revealed a latent period of 13 min, followed by a burst phase of 100 min, with a burst size of 42.1 PFU/cell. Furthermore, we developed a novel phage application platform using aerosolization technology to significantly reduce pathogen levels in closed environments and on seafood products, maintaining the freshness of salmon fillets and meeting GRAS standards. BUCT787 exhibited remarkable inhibitory activities against Vibrio spp., with an inhibition efficiency of 99.9%, and preserved the quality of salmon fillets. These findings establish BUCT787 as a promising biocontrol agent, offering a sustainable solution for enhancing seafood safety, reducing the spread of antibiotic-resistant pathogens, and supporting the sustainability of the aquaculture industry.

• Isolated a novel lytic Vibrio phage BUCT787 with 68% coverage and 95.97% identity.

• Phage BUCT787 showed efficient lytic activity and a wide pH and temperature tolerance range.

• Developed a phage aerosolization platform that reduced Vibrio contamination by 99.9% on salmon fillets.

Micafungin, an echinocandin antifungal agent, has been extensively utilized for the treatment of invasive fungal infections. It is semi-synthesized from the lipohexapeptide FR901379, a natural product from the filamentous fungus Coleophoma empetri. The genetic manipulation of C. empetri has been significantly hindered by the scarcity of both effective neutral sites and selectable markers, which has impeded the systematic metabolic engineering of the industrial strain. Here, a visualizable and marker-free gene editing platform was developed in the industrial strain C. empetri MEFC09. Firstly, the biosynthesis gene cluster of melanin in C. empetri MEFC09 was characterized by gene knockoutin vivo, which comprises a core polyketide synthase gene cemelA and a transcriptional regulator cemelR. Based on this, two neutral sites were designed at the genomic loci of cemelA and cemelR, both demonstrating high-intensity expression of the luciferase reporter. Furthermore, a CRISPR/Cas9-mediated marker-free gene editing platform was developed. When the protoplast concentration was adjusted to 10⁴ cells/mL, a positive transformation rate of 12.4% was achieved on antibiotic-free screening plates. Finally, the transcriptional activator McfJ was overexpressed at the neutral site cemelR using the visualizable and marker-free gene editing platform. It substantially increased the FR901379 titer to 1807.8 mg/L, corresponding to a 3.3-fold improvement over the parental strain. This genetic manipulation system is poised to serve as a versatile platform for accelerating metabolic engineering and functional genomics investigations in industrial strains C. empetri. It will also shed light on the development of genetic manipulation platforms in other filamentous fungi.

• Two visualizable neutral sites were characterized in Coleophoma empetri.

• The CRISPR/Cas9-mediated marker-free gene editing platform was developed.

• The transcriptional activator McfJ was efficiently expressed at the neutral site.