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Biofilm Jun 2024The ability of bacteria to adhere to and form biofilms on food contact surfaces poses serious challenges, as these may lead to the cross-contamination of food products....
The ability of bacteria to adhere to and form biofilms on food contact surfaces poses serious challenges, as these may lead to the cross-contamination of food products. Biomimetic topographic surface modifications have been explored to enhance the antifouling performance of materials. In this study, the topography of two plant leaves, var. (cauliflower, CF) and (white cabbage, WC), was replicated through wax moulding, and their antibiofilm potential was tested against single- and dual-species biofilms of and . Biomimetic surfaces exhibited higher roughness values ( = 4.0 ± 1.0 μm and = 3.3 ± 1.0 μm) than the flat control ( = 0.6 ± 0.2 μm), whilst the CF surface demonstrated a lower interfacial free energy () than the WC surface (-100.08 mJ m and -71.98 mJ m, respectively). The CF and WC surfaces had similar antibiofilm effects against single-species biofilms, achieving cell reductions of approximately 50% and 60% for and , respectively, compared to the control. Additionally, the biomimetic surfaces led to reductions of up to 60% in biovolume, 45% in thickness, and 60% in the surface coverage of single-species biofilms. For dual-species biofilms, only the strain growing on the WC surface exhibited a significant decrease in the cell count. However, confocal microscopy analysis revealed a 60% reduction in the total biovolume and surface coverage of mixed biofilms developed on both biomimetic surfaces. Furthermore, dual-species biofilms were mainly composed of , which reduced growth. Altogether, these results demonstrate that the surface properties of CF and WC biomimetic surfaces have the potential for reducing biofilm formation.
PubMed: 38444517
DOI: 10.1016/j.bioflm.2024.100185 -
PloS One 2024Fermentation-based biosynthesis in synthetic biology relies heavily on sugar-derived feedstocks, a limited and carbon-intensive commodity. Unconventional feedstocks from...
Fermentation-based biosynthesis in synthetic biology relies heavily on sugar-derived feedstocks, a limited and carbon-intensive commodity. Unconventional feedstocks from less-noble sources such as waste are being utilized to produce high-value chemical products. Azo dyes, a major pollutant commonly discharged by food, textile, and pharmaceutical industries, present significant health and environmental risks. We explore the potential of engineering Pseudomonas putida KT2440 to utilize azo dyes as a substrate to produce a polyketide, actinorhodin (ACT). Using the constrained minimal cut sets (cMCS) approach, we identified metabolic interventions that optimize ACT biosynthesis and compare the growth-coupling solutions attainable on an azo dye compared to glucose. Our results predicted that azo dyes could perform better as a feedstock for ACT biosynthesis than glucose as it allowed growth-coupling regimes that are unfeasible with glucose and generated an 18.28% higher maximum ACT flux. By examining the flux distributions enabled in different carbon sources, we observed that carbon fluxes from aromatic compounds like azo dyes have a unique capability to leverage gluconeogenesis to support both growth and production of secondary metabolites that produce excess NADH. Carbon sources are commonly chosen based on the host organism, availability, cost, and environmental implications. We demonstrated that careful selection of carbon sources is also crucial to ensure that the resulting flux distribution is suitable for further metabolic engineering of microbial cell factories.
Topics: Azo Compounds; Pseudomonas putida; Carbon; Glucose; Benzoisochromanequinones; Anthraquinones
PubMed: 38437212
DOI: 10.1371/journal.pone.0299128 -
Microbial Cell Factories Feb 2024We are interested in converting second generation feedstocks into styrene, a valuable chemical compound, using the solvent-tolerant Pseudomonas putida DOT-T1E as a...
We are interested in converting second generation feedstocks into styrene, a valuable chemical compound, using the solvent-tolerant Pseudomonas putida DOT-T1E as a chassis. Styrene biosynthesis takes place from L-phenylalanine in two steps: firstly, L-phenylalanine is converted into trans-cinnamic acid (tCA) by PAL enzymes and secondly, a decarboxylase yields styrene. This study focuses on designing and synthesizing a functional trans-cinnamic acid decarboxylase in Pseudomonas putida. To achieve this, we utilized the "wholesale" method, involving deriving two consensus sequences from multi-alignments of homologous yeast ferulate decarboxylase FDC1 sequences with > 60% and > 50% identity, respectively. These consensus sequences were used to design Pseudomonas codon-optimized genes named psc1 and psd1 and assays were conducted to test the activity in P. putida. Our results show that the PSC1 enzyme effectively decarboxylates tCA into styrene, whilst the PSD1 enzyme does not. The optimal conditions for the PSC1 enzyme, including pH and temperature were determined. The L-phenylalanine DOT-T1E derivative Pseudomonas putida CM12-5 that overproduces L-phenylalanine was used as the host for expression of pal/psc1 genes to efficiently convert L-phenylalanine into tCA, and the aromatic carboxylic acid into styrene. The highest styrene production was achieved when the pal and psc1 genes were co-expressed as an operon in P. putida CM12-5. This construction yielded styrene production exceeding 220 mg L. This study serves as a successful demonstration of our strategy to tailor functional enzymes for novel host organisms, thereby broadening their metabolic capabilities. This breakthrough opens the doors to the synthesis of aromatic hydrocarbons using Pseudomonas putida as a versatile biofactory.
Topics: Styrene; Pseudomonas; Carboxy-Lyases; Pseudomonas putida; Phenylalanine; Cinnamates
PubMed: 38419048
DOI: 10.1186/s12934-024-02341-0 -
Nature Communications Feb 2024Plants and microbes communicate to collaborate to stop pests, scavenge nutrients, and react to environmental change. Microbiota consisting of thousands of species...
Plants and microbes communicate to collaborate to stop pests, scavenge nutrients, and react to environmental change. Microbiota consisting of thousands of species interact with each other and plants using a large chemical language that is interpreted by complex regulatory networks. In this work, we develop modular interkingdom communication channels, enabling bacteria to convey environmental stimuli to plants. We introduce a "sender device" in Pseudomonas putida and Klebsiella pneumoniae, that produces the small molecule p-coumaroyl-homoserine lactone (pC-HSL) when the output of a sensor or circuit turns on. This molecule triggers a "receiver device" in the plant to activate gene expression. We validate this system in Arabidopsis thaliana and Solanum tuberosum (potato) grown hydroponically and in soil, demonstrating its modularity by swapping bacteria that process different stimuli, including IPTG, aTc and arsenic. Programmable communication channels between bacteria and plants will enable microbial sentinels to transmit information to crops and provide the building blocks for designing artificial consortia.
Topics: Arabidopsis; Microbiota; Crops, Agricultural; Solanum tuberosum; Pseudomonas putida
PubMed: 38418817
DOI: 10.1038/s41467-024-45897-6 -
Microorganisms Feb 2024A Special Issue of devoted to 'Microbial Biocatalysis and Biodegradation' would be incomplete without some form of acknowledgement of the many important roles that... (Review)
Review
A Special Issue of devoted to 'Microbial Biocatalysis and Biodegradation' would be incomplete without some form of acknowledgement of the many important roles that dioxygen-dependent enzymes (principally mono- and dioxygenases) play in relevant aspects of bio-oxygenation. This is reflected by the multiple strategic roles that dioxygen -dependent microbial enzymes play both in generating valuable synthons for chemoenzymatic synthesis and in facilitating reactions that help to drive the global geochemical carbon cycle. A useful insight into this can be gained by reviewing the evolution of the current status of 2,5-diketocamphane 1,2-monooxygenase (EC 1.14.14.108) from (+)-camphor-grown ATCC 17453, the key enzyme that promotes the initial ring cleavage of this natural bicyclic terpene. Over the last sixty years, the perceived nature of this monooxygenase has transmogrified significantly. Commencing in the 1960s, extensive initial studies consistently reported that the enzyme was a monomeric true flavoprotein dependent on both FMNH and nonheme iron as bound cofactors. However, over the last decade, all those criteria have changed absolutely, and the enzyme is currently acknowledged to be a metal ion-independent homodimeric flavin-dependent two-component mono-oxygenase deploying FMNH as a cosubstrate. That transition is a paradigm of the ever evolving nature of scientific knowledge.
PubMed: 38399793
DOI: 10.3390/microorganisms12020389 -
Life (Basel, Switzerland) Jan 2024Mn-oxidizing microorganisms oxidize environmental Mn(II), producing Mn(IV) oxides. MnB1 is a widely studied organism for the oxidation of manganese(II) to manganese(IV)...
Mn-oxidizing microorganisms oxidize environmental Mn(II), producing Mn(IV) oxides. MnB1 is a widely studied organism for the oxidation of manganese(II) to manganese(IV) by a multi-copper oxidase. The biogenic manganese oxides (BMOs) produced by MnB1 and similar organisms have unique properties compared to non-biological manganese oxides. Along with an amorphous, poorly crystalline structure, previous studies have indicated that BMOs have high surface areas and high reactivities. It is also known that abiotic Mn oxides promote oxidation of organics and have been studied for their water oxidation catalytic function. MnB1 was grown and maintained and subsequently transferred to culturing media containing manganese(II) salts to observe the oxidation of manganese(II) to manganese(IV). The structures and compositions of these manganese(IV) oxides were characterized using scanning electron microscopy, energy dispersive X-ray spectroscopy, inductively coupled plasma optical emission spectroscopy, and powder X-ray diffraction, and their properties were assessed regarding catalytic functionality towards water oxidation in comparison to abiotic acid birnessite. Water oxidation was accomplished through the whole-cell catalysis of MnB1, the results for which compare favorably to the water-oxidizing ability of abiotic Mn(IV) oxides.
PubMed: 38398680
DOI: 10.3390/life14020171 -
Frontiers in Bioengineering and... 2024Polyhydroxyalkanoates are a class of biodegradable, biocompatible polymers composed of polyesters of R-hydroxyalkanoic acids and deposited intracellularly by a variety...
Polyhydroxyalkanoates are a class of biodegradable, biocompatible polymers composed of polyesters of R-hydroxyalkanoic acids and deposited intracellularly by a variety of microorganisms which have potential to serve as alternative to commercial plastic. Bioplastics are gaining attention due to sustainability, biodegradability, biocompatibility, and lower carbon footprint. Nevertheless, the commercialization of PHA is predominantly hindered by the elevated production expenses arising primarily from the use of a pure sugar substrate. Our study has established a feasible method for bioplastic formation applying MTCC 2475 and periderm as a carbon source. To optimize the sugar yield response surface methodology was used, which released 69.34% ± 0.25% reducing sugar. PHA production experiments were performed in hydrolysate containing media as well as commercial sugar containing mineral salt media. After 48 h of fermentation of using this sugar, a biomass concentration of 2.19 gL, with a PHA production of 0.60 gL (28.71% ± 0.55%) was obtained which was comparatively similar with synthetic media (2.56 gL cell dry weight and 29.97% ± 0.45% PHA). Furthermore, the monomers of PHA produced by hydrolysate were characterized using Gas chromatography-mass spectrometry, Fourier transform infrared spectroscopy, differential scanning calorimetry, and nuclear magnetic resonance. This investigation has identified three distinct monomers of medium-chain PHAs, namely, methyl 3-Hydroxydodecanoate, 3-Hydroxytetradecanoate, and Hexadecanoic acid 3-Hydroxy methyl esters. Hence this study concludes a sustainable production of bioplastics from periderm waste.
PubMed: 38384435
DOI: 10.3389/fbioe.2024.1343540 -
BMC Plant Biology Feb 2024Early blight (EB), caused by Alternaria solani, is a serious problem in tomato production. Plant growth-promoting rhizobacteria promote plant growth and inhibit plant...
Early blight (EB), caused by Alternaria solani, is a serious problem in tomato production. Plant growth-promoting rhizobacteria promote plant growth and inhibit plant disease. The present study explored the bio-efficacy of synergistic effect of rhizobacterial isolates and ginger powder extract (GPE) against tomato EB disease, singly and in combination. Six fungal isolates from symptomatic tomato plants were identified as A. solani on the basis of morphological features i.e., horizontal septation (6.96 to 7.93 µm), vertical septation (1.50 to 2.22 µm), conidia length (174.2 to 187.6 µm), conidial width (14.09 to 16.52 µm), beak length (93.06 to 102.26 µm), and sporulation. Five of the twenty-three bacterial isolates recovered from tomato rhizosphere soil were nonpathogenic to tomato seedlings and were compatible with each other and with GPE. Out of five isolates tested individually, three isolates (St-149D, Hyd-13Z, and Gb-T23) showed maximum inhibition (56.3%, 48.3%, and 42.0% respectively) against mycelial growth of A. solani. Among combinations, St-149D + GPE had the highest mycelial growth inhibition (76.9%) over the untreated control. Bacterial strains molecularly characterized as Pseudomonas putida, Bacillus subtilis, and Bacillus cereus and were further tested in pot trials through seed bacterization for disease control. Seeds treated with bacterial consortia + GPE had the highest disease suppression percentage (78.1%), followed by St-149D + GPE (72.2%) and Hyd-13Z + GPE (67.5%). Maximum seed germination was obtained in the bacterial consortia + GPE (95.0 ± 2.04) followed by St-149D + GPE (92.5 ± 1.44) and Hyd-13Z + GPE (90.0 ± 2.04) over control (73.8 ± 2.39) and chemical control as standard treatment (90.0 ± 2). Ginger powder extracts also induce the activation of defence-related enzymes (TPC, PO, PPO, PAL, and CAT) activity in tomato plants. These were highly significant in the testing bacterial inoculants against A. solani infection in tomato crops.
Topics: Animals; Solanum lycopersicum; Agricultural Inoculants; Powders; Alternaria; Bacteria; Plant Diseases; Plant Extracts; Zingiber officinale
PubMed: 38383294
DOI: 10.1186/s12870-024-04789-z -
MicroLife 2024Pyoverdin is a water-soluble metal-chelator synthesized by members of the genus and used for the acquisition of insoluble ferric iron. Although freely diffusible in...
Pyoverdin is a water-soluble metal-chelator synthesized by members of the genus and used for the acquisition of insoluble ferric iron. Although freely diffusible in aqueous environments, preferential dissemination of pyoverdin among adjacent cells, fine-tuning of intracellular siderophore concentrations, and fitness advantages to pyoverdin-producing versus nonproducing cells, indicate control of location and release. Here, using time-lapse fluorescence microscopy to track single cells in growing microcolonies of SBW25, we show accumulation of pyoverdin at cell poles. Accumulation occurs on cessation of cell growth, is achieved by cross-feeding in pyoverdin-nonproducing mutants and is reversible. Moreover, accumulation coincides with localization of a fluorescent periplasmic reporter, suggesting that pyoverdin accumulation at cell poles is part of the general cellular response to starvation. Compatible with this conclusion is absence of non-accumulating phenotypes in a range of pyoverdin mutants. Analysis of the performance of pyoverdin-producing and nonproducing cells under conditions promoting polar accumulation shows an advantage to accumulation on resumption of growth after stress. Examination of pyoverdin polar accumulation in a multispecies community and in a range of laboratory and natural species of , including PAO1 and KT2440, confirms that the phenotype is characteristic of .
PubMed: 38370141
DOI: 10.1093/femsml/uqae001 -
Metabolic Engineering Mar 2024Sustainable aviation fuel (SAF) will significantly impact global warming in the aviation sector, and important SAF targets are emerging. Isoprenol is a precursor for a...
Sustainable aviation fuel (SAF) will significantly impact global warming in the aviation sector, and important SAF targets are emerging. Isoprenol is a precursor for a promising SAF compound DMCO (1,4-dimethylcyclooctane) and has been produced in several engineered microorganisms. Recently, Pseudomonas putida has gained interest as a future host for isoprenol bioproduction as it can utilize carbon sources from inexpensive plant biomass. Here, we engineer metabolically versatile host P. putida for isoprenol production. We employ two computational modeling approaches (Bilevel optimization and Constrained Minimal Cut Sets) to predict gene knockout targets and optimize the "IPP-bypass" pathway in P. putida to maximize isoprenol production. Altogether, the highest isoprenol production titer from P. putida was achieved at 3.5 g/L under fed-batch conditions. This combination of computational modeling and strain engineering on P. putida for an advanced biofuels production has vital significance in enabling a bioproduction process that can use renewable carbon streams.
Topics: Pseudomonas putida; Carbon; Metabolic Engineering
PubMed: 38369052
DOI: 10.1016/j.ymben.2024.02.004