Microbial Biotechnology最新文献

Filamentous fungi with their diverse inventory of carbohydrate-active enzymes promise a holistic usage of lignocellulosic residues. A major challenge for application is the inherent repression of enzyme production by carbon catabolite repression (CCR). In the presence of preferred carbon sources, the transcription factor CreA/CRE-1 binds to specific but conserved motifs in promoters of genes involved in sugar metabolism, but the status of CCR is notoriously difficult to quantify. To allow for a real-time evaluation of CreA/CRE-1-mediated CCR at the transcriptional level, we developed a luciferase-based construct, representing a dynamic, highly responsive reporter system that is inhibited by monosaccharides in a quantitative fashion. Using this tool, CreA/CRE-1-dependent CCR triggered by several monosaccharides could be measured in Neurospora crassa, Aspergillus niger and Aspergillus nidulans over the course of hours, demonstrating distinct and dynamic regulatory processes. Furthermore, we used the reporter to visualize the direct impacts of multiple CreA truncations on CCR induction. Our reporter thus offers a widely applicable quantitative approach to evaluate CreA/CRE-1-mediated CCR across diverse fungal species and will help to elucidate the multifaceted effects of CCR on fungal physiology for both basic research and industrial strain engineering endeavours.

Agricultural yields are often limited by damage caused by pathogenic microorganisms, including plant-pathogenic bacteria. The chemical control options to cope with bacterial diseases in agriculture are limited, predominantly relying on copper-based products. These compounds, however, possess limited efficacy. Therefore, there is an urgent need to develop novel technologies to manage bacterial plant diseases and reduce food loss. In this study, a new antimicrobial agent was developed using a doping method that entraps small bioactive organic molecules inside copper as the metal matrix. The food preservative agent lauroyl arginate ethyl ester (ethyl lauroyl arginate; LAE) was chosen as the doped organic compound. The new composites were termed LAE@[Cu]. Bactericidal assays against Acidovorax citrulli, a severe plant pathogen, revealed that LAE and copper in the composites possess a synergistic interaction as compared with each component individually. LAE@[Cu] composites were further characterised in terms of chemical properties and in planta assays demonstrated their potential for further development as crop protection agents.

Agricultural yields are often limited by damage caused by pathogenic microorganisms, including plant-pathogenic bacteria. The chemical control options to cope with bacterial diseases in agriculture are limited, predominantly relying on copper-based products. These compounds, however, possess limited efficacy. Therefore, there is an urgent need to develop novel technologies to manage bacterial plant diseases and reduce food loss. In this study, a new antimicrobial agent was developed using a doping method that entraps small bioactive organic molecules inside copper as the metal matrix. The food preservative agent lauroyl arginate ethyl ester (ethyl lauroyl arginate; LAE) was chosen as the doped organic compound. The new composites were termed LAE@[Cu]. Bactericidal assays against Acidovorax citrulli, a severe plant pathogen, revealed that LAE and copper in the composites possess a synergistic interaction as compared with each component individually. LAE@[Cu] composites were further characterised in terms of chemical properties and in planta assays demonstrated their potential for further development as crop protection agents.

Carotenoids are natural pigments utilized as colourants and antioxidants across food, pharmaceutical and cosmetic industries. They exist in carbon chain lengths of C₃₀, C₄₀, C₄₅ and C₅₀, with C₄₀ variants being the most common. Bacterioruberin (BR) and its derivatives are part of the less common C₅₀ carotenoid group, synthesized primarily by halophilic archaea. This study analysed the compositional characteristics of BR extract (BRE) isolated from ‘Haloferax marinum’ MBLA0078, a halophilic archaeon isolated from seawater near Yeoungheungdo Island in the Republic of Korea, and investigated its antioxidant activity and protective effect on lipopolysaccharide (LPS)-induced C2C12 myotube atrophy. The main components of BRE included all-trans-BR, monoanhydrobacterioruberin, 2-isopentenyl-3,4-dehydrorhodopin and all-trans-bisanhydrobacterioruberin. BRE exhibited higher antioxidant activity and DNA nicking protection activity than other well-known C₄₀ carotenoids, such as β-carotene, lycopene and astaxanthin. In C2C12 myotubes, LPS treatment led to a reduction in myotube diameter and number, as well as the hypertranscription of the muscle-specific ubiquitin ligase MAFbx and MuRF1. BRE mitigated these changes by activating the Akt/mTOR pathway. Furthermore, BRE abolished the elevated cellular reactive oxygen species levels and the inflammation response induced by LPS. This study demonstrated that ‘Hfx. marinum’ is an excellent source of natural microbial C₅₀ carotenoids with strong antioxidant capacity and may offer potential protective effects against muscle atrophy.

Carotenoids are natural pigments utilized as colourants and antioxidants across food, pharmaceutical and cosmetic industries. They exist in carbon chain lengths of C₃₀, C₄₀, C₄₅ and C₅₀, with C₄₀ variants being the most common. Bacterioruberin (BR) and its derivatives are part of the less common C₅₀ carotenoid group, synthesized primarily by halophilic archaea. This study analysed the compositional characteristics of BR extract (BRE) isolated from ‘Haloferax marinum’ MBLA0078, a halophilic archaeon isolated from seawater near Yeoungheungdo Island in the Republic of Korea, and investigated its antioxidant activity and protective effect on lipopolysaccharide (LPS)-induced C2C12 myotube atrophy. The main components of BRE included all-trans-BR, monoanhydrobacterioruberin, 2-isopentenyl-3,4-dehydrorhodopin and all-trans-bisanhydrobacterioruberin. BRE exhibited higher antioxidant activity and DNA nicking protection activity than other well-known C₄₀ carotenoids, such as β-carotene, lycopene and astaxanthin. In C2C12 myotubes, LPS treatment led to a reduction in myotube diameter and number, as well as the hypertranscription of the muscle-specific ubiquitin ligase MAFbx and MuRF1. BRE mitigated these changes by activating the Akt/mTOR pathway. Furthermore, BRE abolished the elevated cellular reactive oxygen species levels and the inflammation response induced by LPS. This study demonstrated that ‘Hfx. marinum’ is an excellent source of natural microbial C₅₀ carotenoids with strong antioxidant capacity and may offer potential protective effects against muscle atrophy.

Mycobacterium abscessus (MABS) displays differential subspecies susceptibility to macrolides. Thus, identifying MABS's subspecies (M. abscessus, M. bolletii and M. massiliense) is a clinical necessity for guiding treatment decisions. We aimed to assess the potential of Machine Learning (ML)-based classifiers coupled to Matrix-Assisted Laser Desorption/Ionization Time-of-Flight (MALDI-TOF) MS to identify MABS subspecies. Two spectral databases were created by using 40 confirmed MABS strains. Spectra were obtained by using MALDI-TOF MS from strains cultivated on solid (Columbia Blood Agar, CBA) or liquid (MGIT®) media for 1 to 13 days. Each database was divided into a dataset for ML-based pipeline development and a dataset to assess the performance. An in-house programme was developed to identify discriminant peaks specific to each subspecies. The peak-based approach successfully distinguished M. massiliense from the other subspecies for strains grown on CBA. The ML approach achieved 100% accuracy for subspecies identification on CBA, falling to 77.5% on MGIT®. This study validates the usefulness of ML, in particular the Random Forest algorithm, to discriminate MABS subspecies by MALDI-TOF MS. However, identification in MGIT®, a medium largely used in mycobacteriology laboratories, is not yet reliable and should be a development priority.

Mycobacterium abscessus (MABS) displays differential subspecies susceptibility to macrolides. Thus, identifying MABS's subspecies (M. abscessus, M. bolletii and M. massiliense) is a clinical necessity for guiding treatment decisions. We aimed to assess the potential of Machine Learning (ML)-based classifiers coupled to Matrix-Assisted Laser Desorption/Ionization Time-of-Flight (MALDI-TOF) MS to identify MABS subspecies. Two spectral databases were created by using 40 confirmed MABS strains. Spectra were obtained by using MALDI-TOF MS from strains cultivated on solid (Columbia Blood Agar, CBA) or liquid (MGIT®) media for 1 to 13 days. Each database was divided into a dataset for ML-based pipeline development and a dataset to assess the performance. An in-house programme was developed to identify discriminant peaks specific to each subspecies. The peak-based approach successfully distinguished M. massiliense from the other subspecies for strains grown on CBA. The ML approach achieved 100% accuracy for subspecies identification on CBA, falling to 77.5% on MGIT®. This study validates the usefulness of ML, in particular the Random Forest algorithm, to discriminate MABS subspecies by MALDI-TOF MS. However, identification in MGIT®, a medium largely used in mycobacteriology laboratories, is not yet reliable and should be a development priority.

Previous studies have reported the functional role, biochemical features and synthesis pathway of podophyllotoxin (PTOX) in plants. In this study, we employed combined morphological and molecular techniques to identify an endophytic fungus and extract PTOX derivatives. Based on the analysis of ITS sequences and the phylogenetic tree, the isolate was classified as Penicillium herquei HGN12.1C, with a sequence identity of 98.58%. Morphologically, the HGN12.1C strain exhibits white colonies, short-branched mycelia and densely packed hyphae. Using PacBio sequencing at an average read depth of 195×, we obtained a high-quality genome for the HGN12.1C strain, which is 34.9 Mb in size, containing eight chromosomes, one mitochondrial genome and a GC content of 46.5%. Genome analysis revealed 10 genes potentially involved in PTOX biosynthesis. These genes include VdtD, Pinoresinollariciresinol reductase (PLR), Secoisolariciresinol dehydrogenase (SDH), CYP719A23, CYP71BE54, O-methyltransferase 1 (OMT1), O-methyltransferase 3 (OMT3), 2-ODD, CYP71CU and CYP82D61. Notably, the VdtD gene in fungi shares functional similarities with the DIR gene found in plants. Additionally, we identified peltatin, a PTOX derivative, in the HGN12.1C extract. Docking analysis suggests a potential role for the 2-ODD enzyme in converting yatein to deoxypodophyllotoxin. These findings offer invaluable insights into the synthesis mechanism of PTOX in fungi, shedding light on the relationship between host plants and endophytes.

Feedstock variability represents a challenge in lignocellulosic biorefineries, as it can influence both lignocellulose deconstruction and microbial conversion processes for biofuels and biochemicals production. The impact of feedstock variability on microbial performance remains underexplored, and predictive tools for microbial behaviour are needed to mitigate risks in biorefinery scale-up. Here, twelve batches of corn stover were deconstructed via deacetylation, mechanical refining, and enzymatic hydrolysis to generate lignin-rich and sugar streams. These batches and their derived streams were characterised to identify their chemical components, and the streams were used as substrates for producing muconate and butyrate by engineered Pseudomonas putida and wildtype Clostridium tyrobutyricum, respectively. Bacterial performance (growth, product titers, yields, and productivities) differed among the batches, but no strong correlations were identified between feedstock composition and performance. To provide metabolic insights into the origin of these differences, we evaluated the effect of twenty-three isolated chemical components on these microbes, including three components in relevant bioprocess settings in bioreactors, and we found that growth-inhibitory concentrations were outside the ranges observed in the streams. Overall, this study generates a foundational dataset on P. putida and C. tyrobutyricum performance to enable future predictive models and underscores their resilience in effectively converting fluctuating lignocellulose-derived streams into bioproducts.

In recent years, microbiomes and their potential applications for human, animal or plant health, food production and environmental management came into the spotlight of major national and international policies and strategies. This has been accompanied by substantial R&D investments in both public and private sectors, with an increasing number of products entering the market. Despite widespread agreement on the potential of microbiomes and their uses across disciplines, stakeholders and countries, there is no consensus on what defines a microbiome application. This often results in non-comprehensive communication or insufficient documentation making commercialisation and acceptance of the novel products challenging. To showcase the complexity of this issue we discuss two selected, well-established applications and propose criteria defining a microbiome application and their conditions of use for clear communication, facilitating suitable regulatory frameworks and building trust among stakeholders.