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Microbial electrolysis cells (MECs) are emerging sustainable power generation technologies that can be used for biohydrogen production and wastewater treatment. The prospect for hydrogen gas as a vehicle fuel suitable for the environment is enormous. MEC serves as a reactor for producing biohydrogen by mixing MFC with electrolysis. MECs are the most sustainable, clean, and effective ways to produce biohydrogen. Effective aspects impacting H 2 production, including microbial culture, electrodes, membrane, and substrate, have been explored for greater comprehension of creating MEC technology-based hydrogen generation. The present issues with effective scale-up include the costly production expenses of microbial electrolysis cells, their significant internal resistance, methanogenesis, and membrane/cathode biofouling. This review examines the various operational parameters in optimization studies that influence overall hydrogen generation. This paper also presents strategies to overcome the constraints listed above by creating effective architecture, including several reactor designs, biological, and physical properties, and the use of various operational conditions for improved hydrogen production. This study summarizes recent developments and several other operating factors (organic loading, ionic strength, hydraulic retention time, applied voltage, etc.) that affect hydrogen production in an MEC system. The paper outlines the present technical difficulties and offers practical future ideas for addressing the constraints. [Display omitted] • MEC has great efficiency in hydrogen recovery and concurrent waste conversion. • Microbial culture, electrodes, membrane, and substrate affect H 2 production in MEC. • Scale-up constraints: high costs, internal resistance, methanogenesis, and biofouling. • Modifying strain, nanoparticles, and reactor design can help in overcoming this. [ABSTRACT FROM AUTHOR]

Excessive alcohol intake will aggravate the health risk between the liver and intestine and affect the multi-directional information exchange of metabolites between host cells and microbial communities. Because of the side effects of clinical drugs, people tend to explore the intervention value of natural drugs on diseases. As a flavor substance, spices have been proven to have medicinal value, but they are still rare in treating hepatointestinal diseases caused by alcohol. This paper summarized the metabolic transformation of alcohol in the liver and intestine and summarized the potential value of various perfume active substances in improving liver and intestine diseases caused by alcohol. It is also found that bioactive substances in spices can exert antioxidant activity in the liver and intestine environment and reduce the oxidative stress caused by diseases. These substances can interfere with fatty acid synthesis, promote sugar and lipid metabolism, and reduce liver injury caused by steatosis. They can effectively regulate the balance of intestinal flora, promote the production of SCFAs, and restore the intestinal microenvironment. [ABSTRACT FROM AUTHOR]

At present, atmospheric and room‐temperature plasma (ARTP) is regarded as a new and powerful mutagenesis technology with the advantages of environment‐friendliness, operation under mild conditions, and fast mutagenesis speed. Compared with traditional mutagenesis strategies, ARTP is used mainly to change the structure of microbial DNA, enzymes, and proteins through a series of physical, chemical, and electromagnetic effects with the organisms, leading to nucleotide breakage, conversion or inversion, causing various DNA damages, so as to screen out the microbial mutants with better biological characteristics. As a result, in recent years, ARTP mutagenesis and the combination of ARTP with traditional mutagenesis have been widely used in microbiology, showing great potential for application. In this review, the recent progress of ARTP mutagenesis in different application fields and bottlenecks of this technology are systematically summarized, with a view to providing a theoretical basis and technical support for better application. Finally, the outlook of ARTP mutagenesis is presented, and we identify the challenges in the field of microbial mutagenesis by ARTP. Significance statement: The aim of this review is to analyze the beneficial mutagenic effects induced by ARTP on microbial cells and the related mutagenic mechanisms. Subsequently, bottleneck issues and the recent research progress of ARTP on mutagenesis are systematically summarized, and the application prospect is presented. This review may provide valuable insights and research ideas for future atmospheric and room‐temperature plasma mutagenesis of cells. [ABSTRACT FROM AUTHOR]

Biofilm, being an aggregate of microbial cells play a pivotal role in the development of chronic diseases by making the conventional antibiotics ineffective in killing the pathogenetic microbes. Such incomplete bactericidal activities of antibiotics on the other hand, gives rise to antibiotic resistance. In order to find out some alternative therapeutic approaches the flowers of Clitoria ternatea, an ethnobotanical plant with several medicinal attributes were tested for its antibiofilm activities both by in vitro as well as by in silico analyses in detail. Floral extract of Clitoria ternatea containing various bioactive compounds were found to 85% and 82% biofilm produced by Pseudomonas aeruginosa and Staphylococcus aureus respectively, whereas it was only 25% after antibiotic treatment. FTIR analysis indicated significant alterations in the active residues in the spectral regions of lipids, polysaccharide, nucleic acids and proteins after phytoextract treatment. Remarkable eradication of biofilm bound sessile cells of both P. aeruginosa and S. aureus was brought about by the treatment of petal extracts, as evident from the scanning electron micrographs. Docking interactions between biofilm forming proteins of S. aureus and P. aeruginosa and the bioactive compounds of C. ternatea clearly indicate the antibiofilm efficacies of the bioactive compounds present in the floral extract of C. ternatea. Hence the floral extract can be used as an herbal and nontoxic source of prospective drug to combat biofilm mediated diseases without generating the risk of antibiotic resistance. [ABSTRACT FROM AUTHOR]

In recent years, research on Microbial Electrolytic Cell (MEC) systems, such as wastewater treatment, has shown growing potential. The present research work is based on a study of water purification potential, microbial sustainability, and electrochemical characterization of a two-chambered MEC system, with Aluminium-PANI (Polyaniline) -graphene-oxide based electrode and salt-bridge based proton exchange membrane (PEM)., using the process parameters, optimized for corresponding MFC system. The present research indicates the purification of rice mill wastewater with a MEC-based treatment system with controlled voltage drop (<0.8 Volt) and augmented active microbial activity, as supported by the spectroscopic, biochemical, and carbon-dioxide emission profile assessment. [ABSTRACT FROM AUTHOR]

Aromatic compounds are used in pharmaceutical, food, textile and other industries. Increased demand has sparked interest in exploring biotechnological approaches for their sustainable production as an alternative to chemical synthesis from petrochemicals or plant extraction. These aromatic products may be toxic to microorganisms, which complicates their production in cell factories. In this study, we analysed the toxicity of multiple aromatic compounds in common production hosts. Next, we screened a subset of toxic aromatics, namely 2‐phenylethanol, 4‐tyrosol, benzyl alcohol, berberine and vanillin, against transporter deletion libraries in Escherichia coli and Saccharomyces cerevisiae. We identified multiple transporter deletions that modulate the tolerance of the cells towards these compounds. Lastly, we engineered transporters responsible for 2‐phenylethanol tolerance in yeast and showed improved 2‐phenylethanol bioconversion from L‐phenylalanine, with deletions of YIA6, PTR2 or MCH4 genes improving titre by 8–12% and specific yield by 38–57%. Our findings provide insights into transporters as targets for improving the production of aromatic compounds in microbial cell factories. [ABSTRACT FROM AUTHOR]

Fructooligosaccharides (FOS) are fructose oligomers beneficial to human health and nutrition for prebiotic sugars. Their production occurs by a transfructosylation reaction in sucrose molecules catalyzed by fructosyltransferase enzymes (FTase, E.C.2.4.1.9) adhered to microbial cells. The purpose of this work was to study the preparation, enzymatic activity, and stability of glutaraldehyde-crosslinked Aspergillus oryzae IPT- 301 cells used as a biocatalyst for the transfructosylation reaction of sucrose in a packed bed reactor (PBR), aiming at FOS production. The highest transfructosylation activity (AT) was presented by the biocatalyst prepared by cross-linking at 200 rpm and 45 min. The highest AT in the PBR was obtained at 50 °C, with flow rates from 3 mL min-1 to 5 mL min-1 and sucrose concentrations of 473 g L-1 and 500 g L-1. The enzymatic kinetics was described using the Michaelis-Menten model. Finally, the biocatalyst showed constant AT of approximately 75 U g-1 and 300 U g-1 for 12 h of reaction in the PBR operating in continuous and discontinuous flow, respectively. These results demonstrate a high potential of glutaraldehyde- crosslinked A. oryzae IPT-301 cells as heterogeneous biocatalysts for the continuous production of FOS in PBR reactors. [ABSTRACT FROM AUTHOR]

Patchoulol is a natural sesquiterpene alcohol with extensive applications in cosmetics and pharmaceuticals. In this study, we first constructed the synthesis pathway of patchoulol in Saccharomyces cerevisiae by expressing the patchoulol synthase PTS gene using the strong promoter GAL1. Afterward, the metabolic flux of the precursor was enhanced by strengthening the mevalonate pathway and balancing the precursor competition pathway, resulting in a 32.74-fold increase in patchoulol production. Subsequently, the supply of acetyl-CoA in yeast was increased by modifying transcriptional regulators and modulating the acetyl-CoA pathway, and the titer of patchoulol reached 155.94 mg/L. Finally, optimization of the fermentation conditions resulted in a titer of 195.96 mg/L in the shake flasks. Further, batch-fed fermentation in a 5 L bioreactor yielded 1.95 g/L. This work accelerated the development of a microbial cell factory for the production of patchoulol. [ABSTRACT FROM AUTHOR]

Aerobic granules are small, dense aggregates of microbial cells that form naturally in aerobic wastewater treatment systems. They are characterized by their spherical shape, strong structural integrity, and ability to rapidly settle. These granules are formed through a self-immobilization process where different microbial species coalesce to degrade organic and inorganic compounds in wastewater. This study summarizes the development of aerobic granulation technology in wastewater treatment and the mechanism of aerobic granules' formation, analyzes the characteristics and the factors affecting the aerobic granules' formation, and presents practical engineering examples of its application from pilot-scale to full-scale operation. [ABSTRACT FROM AUTHOR]

Raman flow cytometry (RFC) uniquely integrates the "label-free" capability of Raman spectroscopy with the "high-throughput" attribute of traditional flow cytometry (FCM), offering exceptional performance in cell characterization and sorting. Unlike conventional FCM, RFC stands out for its elimination of the dependency on fluorescent labels, thereby reducing interference with the natural state of cells. Furthermore, it significantly enhances the detection information, providing a more comprehensive chemical fingerprint of cells. This review thoroughly discusses the fundamental principles and technological advantages of RFC and elaborates on its various applications in the biomedical field, from identifying and characterizing cancer cells for in vivo cancer detection and surveillance to sorting stem cells, paving the way for cell therapy, and identifying metabolic products of microbial cells, enabling the differentiation of microbial subgroups. Moreover, we delve into the current challenges and future directions regarding the improvement in sensitivity and throughput. This holds significant implications for the field of cell analysis, especially for the advancement of metabolomics. [ABSTRACT FROM AUTHOR]

There is an urgent need to discover and develop novel antibacterial agents. Accordingly, we synthesised 2-(piperazin-1-yl)naphtho[2,3-d]thiazole-4,9-dione (PNT), which exhibits antimicrobial activity. The aim of this study was to characterise PNT as an effective antimicrobial agent. Fluorescence microscopy was used to measure PNT's uptake into microbial cells (strains of Staphylococcus epidermidis, Staphylococcus aureus, and methicillin-resistant S. aureus (MRSA)), transmission electron microscopy (TEM) was used to investigate the influence of PNT on the configuration of microbial cells, and a DNA gyrase supercoiling assay was used to investigate whether PNT inhibits DNA gyrase. PNT was taken up by more than 50% of microbial cells within 30 min. Using TEM, hollowed-out bacterial cytoplasms were observed in the specimen treated with PNT, although there was no disintegration of the bacterial membrane. In the DNA gyrase supercoiling assay, a dose-dependent reduction in fluorescence intensity was observed as the concentration of PNT increased. This suggests that PNT is taken up by microbial cells, resulting in cell disruption, and it reveals that one of the mechanisms underlying the antimicrobial activity of PNT is the inhibition of DNA gyrase. [ABSTRACT FROM AUTHOR]

Background: Microorganisms are responsible for nutrient removal and resource recovery in wastewater treatment plants (WWTPs), and their diversity is often studied by 16S rRNA gene amplicon sequencing. However, this approach underestimates the abundance and diversity of Patescibacteria due to the low coverage of commonly used PCR primers for this highly divergent bacterial phylum. Therefore, our current understanding of the global diversity, distribution, and ecological role of Patescibacteria in WWTPs is very incomplete. This is particularly relevant as Patescibacteria are considered to be associated with microbial host cells and can therefore influence the abundance and temporal variability of other microbial groups that are important for WWTP functioning. Results: Here, we evaluated the in silico coverage of widely used 16S rRNA gene-targeted primer pairs and redesigned a primer pair targeting the V4 region of bacterial and archaeal 16S rRNA genes to expand its coverage for Patescibacteria. We then experimentally evaluated and compared the performance of the original and modified V4-targeted primers on 565 WWTP samples from the MiDAS global sample collection. Using the modified primer pair, the percentage of ASVs classified as Patescibacteria increased from 5.9 to 23.8%, and the number of detected patescibacterial genera increased from 560 to 1576, while the detected diversity of the remaining microbial community remained similar. Due to this significantly improved coverage of Patescibacteria, we identified 23 core genera of Patescibacteria in WWTPs and described the global distribution pattern of these unusual microbes in these systems. Finally, correlation network analysis revealed potential host organisms that might be associated with Patescibacteria in WWTPs. Interestingly, strong indications were found for an association between Patescibacteria of the Saccharimonadia and globally abundant polyphosphate-accumulating organisms of the genus Ca. Phosphoribacter. Conclusions: Our study (i) provides an improved 16S rRNA gene V4 region-targeted amplicon primer pair inclusive of Patescibacteria with little impact on the detection of other taxa, (ii) reveals the diversity and distribution patterns of Patescibacteria in WWTPs on a global scale, and (iii) provides new insights into the ecological role and potential hosts of Patescibacteria in WWTPs. 9DpTXjuT--dysnV-FwKdBe Video Abstract [ABSTRACT FROM AUTHOR]

Reprogramming metabolic flux is a promising approach for constructing efficient microbial cell factories (MCFs) to produce chemicals. However, how to boost the transmission efficiency of metabolic flux is still challenging in complex metabolic pathways. In this study, metabolic flux is systematically reprogrammed by regulating flux size, flux direction, and flux rate to build an efficient MCF for chondroitin production. The ammoniation pool for UDP‐GalNAc synthesis and the carbonization pool for UDP‐GlcA synthesis are first enlarged to increase flux size for providing enough precursors for chondroitin biosynthesis. Then, the ammoniation pool and the carbonization pool are rematched using molecular valves to shift flux direction from cell growth to chondroitin biosynthesis. Next, the adaptability of polymerization pool with the ammoniation and carbonization pools is fine‐tuned by dynamic and static valve‐based adapters to accelerate flux rate for polymerizing UDP‐GalNAc and UDP‐GlcA to produce chondroitin. Finally, the engineered strain E. coli F51 is able to produce 9.2 g L−1 chondroitin in a 5‐L bioreactor. This strategy shown here provides a systematical approach for regulating metabolic flux in complex metabolic pathways for efficient biosynthesis of chemicals. [ABSTRACT FROM AUTHOR]

Inorganic phosphate (Pi) concentration modulates polyphosphate (polyP) levels in diverse bacteria, affecting their physiology and survival. Lactiplantibacillus paraplantarum CRL 1905 is a lactic acid bacterium isolated from quinoa sourdough with biotechnological potential as starter, for initiating fermentation processes in food, and as antimicrobial-producing organism. The aim of this work was to evaluate the influence of the environmental Pi concentration on different physiological and molecular aspects of the CRL 1905 strain. Cells grown in a chemically defined medium containing high Pi (CDM + P) maintained elevated polyP levels up to late stationary phase and showed an enhanced bacterial survival and tolerance to oxidative stress. In Pi sufficiency condition (CDM-P), cells were ~ 25% longer than those grown in CDM + P, presented membrane vesicles and a ~ 3-fold higher capacity to form biofilm. Proteomic analysis indicated that proteins involved in the “carbohydrate transport and metabolism” and “energy production and conversion” categories were up-regulated in high Pi stationary phase cells, implying an active metabolism in this condition. On the other hand, stress-related chaperones and enzymes involved in cell surface modification were up-regulated in the CDM-P medium. Our results provide new insights to understand the CRL 1905 adaptations in response to differential Pi conditions. The adjustment of environmental Pi concentration constitutes a simple strategy to improve the cellular fitness of L. paraplantarum CRL 1905, which would benefit its potential as a microbial cell factory. [ABSTRACT FROM AUTHOR]

One central goal of bioprocess engineering is to maximize the production of specific chemicals using microbial cell factories. Many bioprocesses are one-stage (batch) processes (OSPs), in which growth and product synthesis are coupled. However, OSPs often exhibit low volumetric productivities due to the competition for substrate for biomass and product synthesis implying trade-offs between biomass and product yields. Two-stage or, more generally, multi-stage processes (MSPs) offer the potential to tackle this trade-off for improved efficiency of bioprocesses, for example, by separating growth and production. MSPs have recently gained much attention, also because of a rapidly growing toolbox for the dynamic control of metabolic fluxes. Despite these promising advancements, computational tools specifically tailored for the optimal design of MSPs in the field of biotechnology are still lacking. Here, we present OptMSP, a new Python-based toolbox for identifying optimal MSPs maximizing a user-defined process metrics (such as volumetric productivity, yield, and titer or combinations thereof) under given constraints. In contrast to other methods, our framework starts with a set of well-defined modules representing relevant stages or sub-processes. Experimentally determined parameters (such as growth rates, substrate uptake and product formation rates) are used to build suitable ODE models describing the dynamic behavior of each module. OptMSP finds then the optimal combination of those modules, which, together with the optimal switching time points, maximize a given objective function. We demonstrate the applicability and relevance of the approach with three different case studies, including the example of lactate production by E. coli in a batch setup, where an aerobic growth phase can be combined with anaerobic production phases with or without growth and with or without enhanced ATP turnover. • The work addresses the design of multi-stage processes (MSPs) to improve the efficiency of bioprocesses. • OptMSP: a new Python-based toolbox for identifying MSPs maximizing a given process metrics. • Input: ODE (or analytical) models for each stage (modules). • Output: optimal combination of modules together with switching times. • Application example: lactate production by E. coli with up to six combinable modules. [ABSTRACT FROM AUTHOR]

Numerous bacterial species associate with plants through commensal, mutualistic, or parasitic association, affecting host physiology and health. The mechanism for such association is intricate and involves the secretion of multiple biochemical substances through dedicated protein systems called secretion systems SS. Eleven SS pathways deliver protein factors and enzymes in their immediate environment or host cells, as well as in competing microbial cells in a contact-dependent or independent fashion. These SS are instrumental in competition, initiation of infection, colonization, and establishment of association (positive or negative) with host organisms. The role of SS in infection and pathogenesis has been demonstrated for several phytopathogens, including Agrobacterium, Xanthomonas, Ralstonia, and Pseudomonas. Since there is overlap in mechanisms of establishing association with host plants, several studies have investigated the role of SSs in the interaction of plant and beneficial bacteria, including symbiotic rhizobia and plant growth bacteria (PGPB). Therefore, the present review updates the role of different SSs required for the colonization of beneficial bacteria such as rhizobia, Burkholderia, Pseudomonas, Herbaspirillum, etc., on or inside plants, which can lead to a long-term association. Most SS like T3SS, T4SS, T5SS, and T6SS are required for the antagonistic activity needed to prevent competing microbes, including phytopathogens, ameliorate biotic stress in plants, and produce substances for successful colonization. Others are required for chemotaxis, adherence, niche formation, and suppression of immune response to establish mutualistic association with host plants. [ABSTRACT FROM AUTHOR]

Product secretion from an engineered cell can be advantageous for microbial cell factories. Extensive work on nucleotide manufacturing, one of the most successful microbial fermentation processes, has enabled Corynebacterium stationis to transport nucleotides outside the cell by random mutagenesis; however, the underlying mechanism has not been elucidated, hindering its applications in transporter engineering. Herein, we report the nucleotide-exporting major facilitator superfamily (MFS) transporter from the C. stationis genome and its hyperactive mutation at the G64 residue. Structural estimation and molecular dynamics simulations suggested that the activity of this transporter improved via two mechanisms: (1) enhancing interactions between transmembrane helices through the conserved "RxxQG" motif along with substrate binding and (2) trapping substrate-interacting residue for easier release from the cavity. Our results provide novel insights into how MFS transporters change their conformation from inward- to outward-facing states upon substrate binding to facilitate efflux and can contribute to the development of rational design approaches for efflux improvements in microbial cell factories. Keypoints: • An MFS transporter from C. stationis genome and its mutation at residue G64 were assessed • It enhanced the transporter activity by strengthening transmembrane helix interactions and trapped substrate-interacting residues • Our results contribute to rational design approach development for efflux improvement [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

The microbial biosynthesis of proteins, primary metabolites, and chemicals is gaining extraordinary momentum and is presently viewed as an advancing approach in the industrial research sector. Increased threats to the environment and the possibility of declining petroleum assets have switched the spotlight to microbial cell factories (MCFs). Aside from possessing various advantages over chemical synthesis, such as less toxicity, cheaper methodologies, and an environmentally benign nature, microbes can be cultivated in fermenters, resulting in an effective bioprocessing approach in terms of industrial relevance. As the overwhelming majority of biodiversity is microbial, this review first highlights the microbial biodiversity of industrially vital microorganisms. Then, the paper delineates the production pathways for generating valuable bioproducts via microbial workhorses. Many host cells synthesize bio-compounds as a part of their natural mechanism; however, several techniques have also been developed to attain the desired end product from non-native microbes with selected properties. The microbial biosynthetic pathways can be categorized as native-existing pathways, heterologous pathways, and artificial de novo pathways. Systems metabolic engineering, which integrates metabolic engineering with evolutionary engineering, synthetic biology, and systems biology, has further revolutionized the field of engineering robust phenotypes. The employment of these strategies improves the performance of the strain, eventually achieving high titer and productivity rates of bio-chemicals. Modern trends and tools for exploiting native pathways and designing non-native-created pathways are also briefly discussed in this paper. Finally, the review discusses the use of microbial workhorses for producing a myriad of materials and chemicals, including carboxylic acids, amino acids, plant natural products (PNPs), carotenoids, flavors, and fragrances, unveiling the efficacy of utilizing microbial species to generate sustainable bio-based products. [ABSTRACT FROM AUTHOR]

Microbial cell factories, renowned for their economic and environmental benefits, have emerged as a key trend in academic and industrial areas, particularly in the fermentation of natural compounds. Among these, plant-derived terpenes stand out as a significant class of bioactive natural products. The large-scale production of such terpenes, exemplified by artemisinic acid—a crucial precursor to artemisinin—is now feasible through microbial cell factories. In the fermentation of terpenes, two-phase fermentation technology has been widely applied due to its unique advantages. It facilitates in situ product extraction or adsorption, effectively mitigating the detrimental impact of product accumulation on microbial cells, thereby significantly bolstering the efficiency of microbial production of plant-derived terpenes. This paper reviews the latest developments in two-phase fermentation system applications, focusing on microbial fermentation of plant-derived terpenes. It also discusses the mechanisms influencing microbial biosynthesis of terpenes. Moreover, we introduce some new two-phase fermentation techniques, currently unexplored in terpene fermentation, with the aim of providing more thoughts and explorations on the future applications of two-phase fermentation technology. Lastly, we discuss several challenges in the industrial application of two-phase fermentation systems, especially in downstream processing. [ABSTRACT FROM AUTHOR]

Melanins are complex, polymeric pigments with interesting properties like UV-light absorbance ability, metal ion chelation capacity, antimicrobial action, redox behaviors, and scavenging properties. Based on these characteristics, melanins might be applied in different industrial fields like food packaging, environmental bioremediation, and bioelectronic fields. The actual melanin manufacturing process is not environmentally friendly as it is based on extraction and purification from cuttlefish. Synthetic melanin is available on the market, but it is more expensive than animal-sourced pigment and it requires long chemical procedures. The biotechnological production of microbial melanin, instead, might be a valid alternative. Streptomycetes synthesize melanins as pigments and as extracellular products. In this review, the melanin biotechnological production processes by different Streptomyces strains have been revised according to papers in the literature. The different fermentation strategies to increase melanin production such as the optimization of growth conditions and medium composition or the use of raw sources as growth substrates are here described. Diverse downstream purification processes are also reported as well as all the different analytical methods used to characterize the melanin produced by Streptomyces strains before its application in different fields. [ABSTRACT FROM AUTHOR]

Transcription factor engineering has unique advantages in improving the performance of microbial cell factories due to the global regulation of gene transcription. Omics analyses and reverse engineering enable learning and subsequent incorporation of novel design strategies for further engineering. Here, we identify the role of the global regulator IhfA for overproduction of free fatty acids (FFAs) using CRISPRi-facilitated reverse engineering and cellular physiological characterization. From the differentially expressed genes in the ihfALL- strain, a total of 14 beneficial targets that enhance FFAs production by above 20 % are identified, which involve membrane function, oxidative stress, and others. For membrane-related genes, the engineered strains obtain lower cell surface hydrophobicity and increased average length of membrane lipid tails. For oxidative stress-related genes, the engineered strains present decreased reactive oxygen species (ROS) levels. These gene modulations enhance cellular robustness and save cellular resources, contributing to FFAs production. This study provides novel targets and strategies for engineering microbial cell factories with improved FFAs bioproduction. [ABSTRACT FROM AUTHOR]

Lactoferricin, a multifunctional peptide located in the N-terminal region of lactoferrin, has a broad-spectrum bacteriostatic activity. It is a promising candidate as a food additive and immune fortification agent and does not have the risks associated with drug residues and drug resistance. First, we performed promoter and host cell screening to achieve the recombinant expression of lactoferricin in Pichia pastoris, showing an initial titer of 19.5 mg/L in P. pastoris X-33 using PAOX1 promoter. Second, we constructed a 0030-a hybrid signal peptide by fusing the 0030 signal peptide with the pro-sequence of a-factor secretory signal peptide. This further increased the production of lactoferricin, with a titer of 28.8 mg/L in the fermentation supernatant in the shaking flask. Next, we increased the expression of lactoferricin by fusing it with anionic antioxidant peptides. The neutralization of positive charges yielded a titer of 55.3 mg/L in the shaking flask, and a highest titer of 193.9 mg/L in a 3-L bioreactor. The antimicrobial activity analysis showed that recombinant-expressed lactoferricin exhibited potent antibacterial activity against Escherichia coli, Bacillus subtilis, and Staphylococcus aureus. This study provides a reference for the construction of microbial cell factories capable of efficiently synthesizing antimicrobial peptides. [ABSTRACT FROM AUTHOR]

Promoter sequences are important genetic control elements. Through their interaction with RNA polymerase they determine transcription strength and specificity, thereby regulating the first step in gene expression. Consequently, they can be targeted as elements to control predictability and tuneability of a genetic circuit, which is essential in applications such as the development of robust microbial cell factories. This review considers the promoter elements implicated in the three stages of transcription initiation, detailing the complex interplay of sequence-specific interactions that are involved, and highlighting that DNA sequence features beyond the core promoter elements work in a combinatorial manner to determine transcriptional strength. In particular, we emphasize that, aside from promoter recognition, transcription initiation is also defined by the kinetics of open complex formation and promoter escape, which are also known to be highly sequence specific. Significantly, we focus on how insights into these interactions can be manipulated to lay the foundation for a more rational approach to promoter engineering. [ABSTRACT FROM AUTHOR]

The review summarizes and analyzes information regarding the effect of nanoparticles (NPs) of metals, metal oxides and carbon on the biofilm formation and mature biofilms of microorganisms. The viability of individual microbial cells, including direct disruption of cell surface structures and oxidative stress associated with the formation of reactive oxygen species (ROS), as well as the effect on the production of the exopolymer matrix and the quorum sensing system are considered as the mechanisms of NPs action on biofilms. The effects of silver NPs, gold NPs, some metal oxides, and carbon nanomaterials on microbial biofilms have been described in more detail. The effects of metal and carbon NPs on microbial biofilms are compared. Both antibiofilm and probiofilm effects of NPs are noted, depending on their nature, and the prospect of their use as antimicrobial agents and carriers for the production of microbial biofilms of biotechnological significance are considered. [ABSTRACT FROM AUTHOR]

• γδ T cells regulate the gut microbiota. • In the absence of γδ T cells, mice develop a microbial dysbiosis with the growth of microbes involved in the metabolism of aromatic amino acids. • The gut microbiota from γδ T cell deficient mice produces high amounts of hippurate. • Hippurate reaches the brain where it activates striatal neurons expressing the dopamine type 1 receptor (D1R). • Overactivation of striatal D1R + neurons lead to repetitive/compulsive behavior. Intestinal γδ T cells play an important role in shaping the gut microbiota, which is critical not only for maintaining intestinal homeostasis but also for controlling brain function and behavior. Here, we found that mice deficient for γδ T cells (γδ-/-) developed an abnormal pattern of repetitive/compulsive (R/C) behavior, which was dependent on the gut microbiota. Colonization of WT mice with γδ-/- microbiota induced R/C behavior whereas colonization of γδ-/- mice with WT microbiota abolished the R/C behavior. Moreover, γδ-/- mice had elevated levels of the microbial metabolite 3-phenylpropanoic acid in their cecum, which is a precursor to hippurate (HIP), a metabolite we found to be elevated in the CSF. HIP reaches the striatum and activates dopamine type 1 (D1R)-expressing neurons, leading to R/C behavior. Altogether, these data suggest that intestinal γδ T cells shape the gut microbiota and their metabolites and prevent dysfunctions of the striatum associated with behavior modulation. [ABSTRACT FROM AUTHOR]

Medium‐chain‐length α,ω‐diols (mcl‐diols) play an important role in polymer production, traditionally depending on energy‐intensive chemical processes. Microbial cell factories offer an alternative, but conventional strains like Escherichia coli and Saccharomyces cerevisiae face challenges in mcl‐diol production due to the toxicity of intermediates such as alcohols and acids. Metabolic engineering and synthetic biology enable the engineering of non‐model strains for such purposes with P. putida emerging as a promising microbial platform. This study reviews the advancement in diol production using P. putida and proposes a four‐module approach for the sustainable production of diols. Despite progress, challenges persist, and this study discusses current obstacles and future opportunities for leveraging P. putida as a microbial cell factory for mcl‐diol production. Furthermore, this study highlights the potential of using P. putida as an efficient chassis for diol synthesis. [ABSTRACT FROM AUTHOR]

The long-range transport of bioaerosols by dust events significantly impacts ecological and meteorological networks of the atmosphere, biosphere, and anthroposphere. Bioaerosols not only cause significant public health risks, but also act as efficient ice nuclei for inducing cloud formation and precipitation in the hydrological cycle. To establish risk management for bioaerosol impacts on the Earth system, a large-scale investigation of bioaerosols must be performed under different environmental conditions. For this purpose, a Dust–Bioaerosol (DuBi) field campaign was conducted to investigate the distribution of bioaerosols by collecting ∼950 samples at 39 sites across East Asia from 2016 to 2021. Concentrations and community structures of bioaerosols were further analyzed using fluorescence microscopic observations and high-throughput DNA sequencing, and these factors were compared to environmental factors, such as PM10 and aridity. The results indicated that microbial concentrations at dryland sites were statistically higher than those at humid sites, while the microbe-to-total-particle ratio was statistically lower in drylands than in humid regions. Microbial cells per microgram of PM10 decreased when PM10 increased. The proportion of airborne particles at each site did not vary substantially with season. The richness and diversity of airborne bacteria were significantly higher in drylands than in semiarid regions, while the community structures were stable among all sampling sites. The DuBi field campaign improves our understanding of bioaerosol characteristic variations along the dust transport pathway in East Asia and the changes of bioaerosols under the trend of climate warming, supporting the efforts to reduce public health risks. SIGNIFICANCE STATEMENT: We first conducted large-scale Dust–Bioaerosol (DuBi) field observations in East Asia from 2016 to 2021. We used unified sampling and analysis methods to collect 950 samples at 39 sites and analyzed bioaerosols' concentration and community structure. This study aims to find the relationship between bioaerosols with atmospheric particles and aridity, which fills gaps in the field. These results improve our understanding of bioaerosols' impacts on ecological and meteorological networks of the atmosphere, biosphere, and anthroposphere. The comprehensive observations will promote prediction of bioaerosol change under the trend of climate warming, supporting the efforts to reduce global climate, ecological security, and public health risks in the future. [ABSTRACT FROM AUTHOR]

Metabolic fluxes and their control mechanisms are fundamental in cellular metabolism, offering insights for the study of biological systems and biotechnological applications. However, quantitative and predictive understanding of controlling biochemical reactions in microbial cell factories, especially at the system level, is limited. In this work, we present ARCTICA, a computational framework that integrates constraint-based modelling with machine learning tools to address this challenge. Using the model cyanobacterium Synechocystis sp. PCC 6803 as chassis, we demonstrate that ARCTICA effectively simulates global-scale metabolic flux control. Key findings are that (i) the photosynthetic bioproduction is mainly governed by enzymes within the Calvin–Benson–Bassham (CBB) cycle, rather than by those involve in the biosynthesis of the end-product, (ii) the catalytic capacity of the CBB cycle limits the photosynthetic activity and downstream pathways and (iii) ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBisCO) is a major, but not the most, limiting step within the CBB cycle. Predicted metabolic reactions qualitatively align with prior experimental observations, validating our modelling approach. ARCTICA serves as a valuable pipeline for understanding cellular physiology and predicting rate-limiting steps in genome-scale metabolic networks, and thus provides guidance for bioengineering of cyanobacteria. • A workflow for metabolic flux control analysis in Synechocystis sp. PCC 6803. • Machine learning with features derived from genome-scale metabolic modelling. • Identification of potential key reactions for metabolic adaptations and cell bioengineering. [ABSTRACT FROM AUTHOR]

The integration of a microbial electrolysis cell (MEC) is an effective strategy for enhancing the efficiency and stability of an anaerobic digestion (AD) system for energy recovery from waste-activated sludge (WAS). Typically, electrodes are arranged as separate components, potentially disrupting mixing and complicating the reactor configuration, posing challenges for the scaling up of AD-MEC coupling systems. In this study, electrodes were introduced into a continuous stirring tank reactor (CSTR) in a "stealth" manner by integrating them with the inner wall and stirring paddle. This electrode arrangement approach was validated through a sequential batch digestion experiment, resulting in a remarkable 1.5-fold increase in cumulative methane production and a shortened lag period compared to the traditional CSTR with a nonconductive inner wall and stirring paddle. Both the conductive materials (CMs) employed in the electrodes and the electrochemical processes equally contributed to the observed enhancement effect of the electrodes by regulating the evolution of the microbial community within the electrode biofilms, with a specific emphasis on the enrichment of methanogens (primarily Methanobacterium). This research offers a potential avenue to solve the contradiction between the electrode introduction and the mixing operation in AD-MEC coupling systems and to contribute to its future commercial application. [ABSTRACT FROM AUTHOR]

In this study, nickel–cobalt co-modified stainless steel mesh (Ni-Co@SSM) was prepared and used as the biocathode in microbial electrolysis cell (MEC) for sulfamethazine (SMT) degradation. The optimal electrochemical performance of the Ni-Co@SSM was obtained at the electrodeposition time of 600 s, electrodeposition current density of 20 mA cm−2, and nickel–cobalt molar ratio of 1:2. The removal of SMT in MEC with the Ni-Co@SSM biocathode (MEC-Ni-Co@SSM) was 82%, which increased by 30% compared with the conventional anaerobic reactor. Thirteen intermediates were identified and the potential degradation pathways of SMT were proposed. Proteobacteria, Firmicutes, Patescibacteria, Chloroflexi, Bacteroidetes, and Euryarchaeota are the dominant bacteria at the phylum level in the MEC-Ni-Co@SSM, which are responsible for SMT metabolism. Due to the electrical stimulation, there was an increase in the abundance of the metabolic function and the genetic information processing. This work provides valuable insight into utilizing MECs for effective treatment of antibiotic-containing wastewater. [ABSTRACT FROM AUTHOR]

The manipulation of single particles remains a topic of interest with many applications. Here we characterize the impact of selected parameters on the motion of single particles thanks to dielectrophoresis (DEP) induced by visible light, in a technique called Light-induced Dielectrophoresis, or LiDEP, also known as optoelectronic tweezers, optically induced DEP, and image-based DEP. Baker's yeast and Candida cells are exposed to an electric field gradient enabled by shining a photoconductive material with a specific pattern of visible light, and their response is measured in terms of the average cell velocity towards the gradient. The impact on cell velocity when varying the shape and color of the light pattern, as well as the distance from the cell to the pattern, is presented. The experimental setup featured a commercial light projector featuring digital light processing (DLP) technology but mechanically modified to accommodate a 40× microscope objective lens. The minimal resolution achieved on the light pattern was 8 µm. Experimental results show the capability for single cell manipulation and the possibility of using different shapes, colors, and distances to determine the average cell velocity. [ABSTRACT FROM AUTHOR]

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As the most abundant renewable aromatic resource, lignin is an ideal precursor for the production of aromatic fine chemicals. Biotransformation of lignin into 4-vinylphenol (4VP) derivatives holds the promise to contribute to a lignin-based economy. However, the limited bioavailability of lignin and the absence of an effective biotransformation route for lignin to 4VPs have hindered the valorization of lignin and the production efficiency of 4VPs. This review aims to explore an atom-economic biotransformation route for lignin to 4VPs by summarizing the state-of-the-art technologies involved in the entire process, from upstream to downstream. Effective depolymerization technologies have enabled the generation of favorable lignin-derived aromatic precursors for the biosynthesis of 4VPs. Additionally, these technologies also yielded oligomers or monomers that can serve as carbon sources for the cell growth of host strains. Furthermore, the exploration of key enzymes and the construction of effective microbial cell factories have facilitated the assimilation of lignin derivatives and resulted in improved yields and titers of 4VPs. Emerging technologies have further enhanced the biotransformation of lignin by regulating metabolism efficiency, improving the stability and activity of key enzymes, and mitigating inhibitory effects. Overall, by harnessing the inherent aromatic value of lignin, the biotransformation of lignin-derived aromatic precursors into 4VPs offers a promising and sustainable route for 4VPs biosynthesis and lignin valorization. This approach aligns with the atom-economic concept and holds great potential. [ABSTRACT FROM AUTHOR]

Measurements during episodes of African dust, made with two Wideband Integrated Bioaerosol Spectrometers (WIBS), one on the northeastern coast of Puerto Rico and the other in the city of León, Spain, show unmistakable, bioaerosol-like fluorescing aerosol particles (FAP) that can be associated with these dust episodes. The Puerto Rico events occurred during a major incursion of African dust during June 2020. The León events occurred in the late winter and spring of 2022 when widespread, elevated layers of dust inundated the Iberian Peninsula. Satellite and back trajectory analyses confirm that dust from Northern Africa was the source of the particles during both events. The WIBS measures the size of individual particles in the range from 0.5 µm to 30 µm, derives a shape factor and classifies seven types of fluorescence from the FAP. In general it is not possible to directly determine the specific biological identity from fluorescence signatures, however, measurements of these types of bioaerosols in laboratory studies allow us to compare ambient fluorescence patterns with whole microbial cells measured under controlled conditions. Here we introduce some new metrics that offer a more quantitative approach for comparing FAP characteristics derived from particles measured under different environmental conditions. The analysis highlights the similarities and differences at the two locations and reveals differences that can be attributed to the age and history of the dust plumes, e.g., the amount of time that the air masses were in the mixed layer and the frequency of precipitation along the air mass trajectory. [ABSTRACT FROM AUTHOR]

Background: The demand for melatonin is increasing due to its health-promoting bioactivities such as antioxidant and sleep benefits. Although melatonin is present in various organisms, its low content and high extraction cost make it unsustainable. Biosynthesis is a promising alternative method for melatonin production. However, the ectopic production of melatonin in microorganisms is very difficult due to the low or insoluble expression of melatonin synthesis genes. Hence, we aim to explore the biosynthesis of melatonin using Escherichia coli as a cell factory and ways to simultaneously coordinated express genes from different melatonin synthesis pathways. Results: In this study, the mXcP4H gene from Xanthomonas campestris, as well as the HsAADC, HsAANAT and HIOMT genes from human melatonin synthesis pathway were optimized and introduced into E. coli via a multi-monocistronic vector. The obtained strain BL7992 successfully synthesized 1.13 mg/L melatonin by utilizing L-tryptophan (l-Trp) as a substrate in a shake flask. It was determined that the rate-limiting enzyme for melatonin synthesis is the arylalkylamine N-acetyltransferase, which is encoded by the HsAANAT gene. Targeted metabolomics analysis of l-Trp revealed that the majority of l-Trp flowed to the indole pathway in BL7992, and knockout of the tnaA gene may be beneficial for increasing melatonin production. Conclusions: A metabolic engineering approach was adopted and melatonin was successfully synthesized from low-cost l-Trp in E. coli. This study provides a rapid and economical strategy for the synthesis of melatonin. [ABSTRACT FROM AUTHOR]

Candida albicans remains the predominant cause of fungal infections, where adhered microbial cells form biofilms - densely packed communities. The central feature of C. albicans biofilms is the production of an extracellular matrix (ECM) consisting of polymers and extracellular nucleic acids (eDNA, eRNA), which significantly impedes the infiltration of host cells. Neutrophils, as crucial players in the innate host defense, employ several mechanisms to eradicate the fungal infection, including NETosis, endocytosis, or the release of granules containing, among others, antimicrobial peptides (AMPs). The main representative of these is the positively charged peptide LL-37 formed from an inactive precursor (hCAP18). In addition to its antimicrobial functions, this peptide possesses a propensity to interact with negatively charged molecules, including nucleic acids. Our in vitro studies have demonstrated that LL-37 contacting with C. albicans nucleic acids, isolated from biofilm, are complexed by the peptide and its shorter derivatives, as confirmed by electrophoretic mobility shift assays. We indicated that the generation of the complexes induces discernible alterations in the neutrophil response to fungal nucleic acids compared to the effects of unconjugated molecules. Our analyses involving fluorescence microscopy, flow cytometry, and Western blotting revealed that stimulation of neutrophils with DNA:LL-37 or RNA:LL-37 complexes hamper the activation of pro-apoptotic caspases 3 and 7 and fosters increased activation of anti-apoptotic pathways mediated by the Mcl-1 protein. Furthermore, the formation of complexes elicits a dual effect on neutrophil immune response. Firstly, they facilitate increased nucleic acid uptake, as evidenced by microscopic observations, and enhance the proinflammatory response, promoting IL-8 production. Secondly, the complexes detection suppresses the production of reactive oxygen species and attenuates NETosis activation. In conclusion, these findings may imply that the neutrophil immune response shifts toward mobilizing the immune system as a whole, rather than inactivating the pathogen locally. Our findings shed new light on the intricate interplay between the constituents of the C. albicans biofilm and the host's immune response and indicate possible reasons for the elimination of NETosis from the arsenal of the neutrophil response during contact with the fungal biofilm. [ABSTRACT FROM AUTHOR]

Aquifers harbor unique and highly adapted species, contributing to critical ecological processes and services. Understanding the key factors driving invertebrate assemblages in aquifers is a challenging task that, traditionally, has primarily been achieved in karst aquifers. This study aimed to uncover the factors influencing the composition and functionality of groundwater crustaceans in a volcanic aquifer of central Italy. We adopted a multidisciplinary approach, combining hydrogeology, geology, microbiology, and ecology, and found that the aquifer consisted of three adjacent groundwater bodies (GWBs) with different geochemistry (i.e., sulfate-depleted, K-rich and earth-alkaline) and microbial characteristics that remained consistent over the study period. We also unveiled significant differences in both the taxonomic and functional composition of groundwater crustaceans across the three GWBs and these patterns were consistent over time. Notably, the sulfate-depleted GWB lacked groundwater-obligate species, burrowers, stenothermal and moderately stenothermal species, while the K-rich and earth-alkaline GWBs had different species with similar functions related to locomotion, diet, and feeding habits. Stenothermal and moderately stenothermal crustacean species were only found in the K-rich GWB, which lacked epigean species. Major ions (SO4, Ca, NO3, and K), trace elements (B, Al, V, Se, and Ba), and microbial factors related to microbial cells with low nucleic acid (LNA cells) and carbohydrate catabolic profiles were the main descriptors of groundwater-obligate species abundances. Our findings revealed a significant correlation between the abundances of groundwater-obligate crustaceans and LNA cells, thus suggesting a selective feeding of groundwater invertebrate species on the aquatic microbial community. Our research emphasizes the need to consider diverse hydrogeological contexts within individual aquifers. Potential avenues for future research should further consider food web dynamics in groundwater communities and their impact on carbon and nutrient cycling. [ABSTRACT FROM AUTHOR]

The total rate of plastic production is anticipated to surpass 1.1 billion tons per year by 2050. Plastic waste is non-biodegradable and accumulates in natural ecosystems. In 2020, the total amount of plastic waste was estimated to be 367 million metric tons, leading to unmanageable waste disposal and environmental pollution issues. Plastics are produced from petroleum and natural gases. Given the limited fossil fuel reserves and the need to circumvent pollution problems, the focus has shifted to biodegradable biopolymers, such as polyhydroxyalkanoates (PHAs), polylactic acid, and polycaprolactone. PHAs are gaining importance because diverse bacteria can produce them as intracellular inclusion bodies using biowastes as feed. A critical component in PHA production is the downstream processing procedures of recovery and purification. In this review, different bioengineering approaches targeted at modifying the cell morphology and synchronizing cell lysis with the biosynthetic cycle are presented for product separation and extraction. Complementing genetic engineering strategies with conventional downstream processes, these approaches are expected to produce PHA sustainably. [ABSTRACT FROM AUTHOR]

Polymetallic exposure causes complex toxicity to microorganisms. In this study, we investigated the responses of Escherichia coli under co-existence of cadmium (Cd) and lead (Pb), primarily based on biochemical analysis and RNA sequencing. Cd completely inhibited bacterial growth at a concentration of 2.41 mmol/L, with its removal rate as low as <10%. In contrast, the Pb removal rate was >95% under equimolar sole Pb stress. In addition, the Raman analysis confirmed the loss of proteins for the bacterial cells. Under the co-existence of Cd and Pb, the Cd toxicity to E. coli was alleviated. Meanwhile, the biosorption of Pb cations was more intense during the competitive sorption with Cd. Transmission electron microscopy images showed that a few cells were elongated during incubation, i.e., the average cellular length increased from 1.535 ± 0.407 to 1.845 ± 0.620 µm. Moreover, NanoSIMS imaging showed that the intracellular distribution of Cd and Pb was coupled with sulfur. Genes regulating sulfate transporter were also upregulated to promote sulfate assimilation. Then, the subsequent production of biogenic sulfide and sulfur-containing amino acids was enhanced. Although this strategy based on S enrichment could resist the polymetallic stress, not all related genes were induced to upregulate under sole Cd stress. Therefore, the S metabolism might remodel the microbial resistance to variable occurrence of heavy metals. Furthermore, the competitive sorption (in contrast to sole Cd stress) could prevent microbial cells from strong Cd toxicity. [ABSTRACT FROM AUTHOR]

In microbiological studies, a common goal is to link environmental factors to microbial activities. Both environmental factors and microbial activities are typically derived from bulk samples. It is becoming increasingly clear that such bulk environmental parameters poorly represent the microscale environments microorganisms experience. Using infrared (IR) microspectroscopy, the spatial distribution of chemical compound classes can be visualized, making it a useful tool for studying the interactions between microbial cells and their microenvironments. The spatial resolution of conventional IR microspectroscopy has been limited by the diffraction limit of IR light. The recent development of optical photothermal infrared (O-PTIR) microspectroscopy has pushed the spatial resolution of IR microspectroscopy beyond this diffraction limit, allowing the distribution of chemical compound classes to be visualized at sub-micrometer spatial scales. To examine the potential and limitations of O-PTIR microspectroscopy to probe the interactions between fungal cells and their immediate environments, we imaged the decomposition of cellulose films by cells of the ectomycorrhizal fungus Paxillus involutus and compared O-PTIR results using conventional IR microspectroscopy. Whereas the data collected with conventional IR microspectroscopy indicated that P. involutus has only a very limited ability to decompose cellulose films, O-PTIR data suggested that the ability of P. involutus to decompose cellulose was substantial. Moreover, the O-PTIR method enabled the identification of a zone located outside the fungal hyphae where the cellulose was decomposed by oxidation. We conclude that O-PTIR can provide valuable new insights into the abilities and mechanisms by which microorganisms interact with their surrounding environments. [ABSTRACT FROM AUTHOR]

Bifidobacteria are among the first microbial colonizers of the human gut, being frequently associated with human health-promoting activities. In the current study, an in silico methodology based on an ecological and phylogenomic-driven approach allowed the selection of a Bifidobacterium adolescentis prototype strain, i.e., B. adolescentis PRL2023, which best represents the overall genetic content and functional features of the B. adolescentis taxon. Such features were confirmed by in vitro experiments aimed at evaluating the ability of this strain to survive in the gastrointestinal tract of the host and its ability to interact with human intestinal cells and other microbial gut commensals. In this context, co-cultivation of B. adolescentis PRL2023 and several gut commensals revealed various microbe-microbe interactions and indicated co-metabolism of particular plant-derived glycans, such as xylan. [ABSTRACT FROM AUTHOR]

(2S)-eriodictyol (ERD) is a flavonoid widely found in citrus fruits, vegetables, and important medicinal plants with neuroprotective, cardioprotective, antidiabetic, and anti-obesity effects. However, the microbial synthesis of ERD is limited by complex metabolic pathways and often results in a low production performance. Here, we engineered Saccharomyces cerevisiae by fine-tuning the metabolism of the ERD synthesis pathway. The results showed that the ERD titer was effectively increased, and the intermediate metabolites levels were reduced. First, we successfully reconstructed the de novo synthesis pathway of p-coumaric acid in S. cerevisiae and fine-tuned the metabolic pathway using promoter engineering and terminator engineering for the high-level production of (2S)-naringenin. Subsequently, the synthesis of ERD was achieved by introducing the ThF3′H gene from Tricyrtis hirta. Finally, by multiplying the copy number of the ThF3′H gene, the production of ERD was further increased, reaching 132.08 mg L−1. Our work emphasizes the importance of regulating the metabolic balance to produce natural products in microbial cell factories. [ABSTRACT FROM AUTHOR]

The interdisciplinary nature of synthetic biology merges engineering principles with biology and provides innovative solutions for issues in the biomanufacturing industry. To develop industrially applicable biocatalysts and/or microbial cell factories, a design-build-test-learn cycle-based iterative process is necessary, which is often time-consuming and labor-intensive. The integration of microfluidic technologies into synthetic biology can accelerate these processes, particularly for achieving high-throughput phenotyping and screening. In this review, we examine the potential of microfluidic technologies to revolutionize synthetic biology. Although commercial microfluidics demonstrate superior throughput for single-cell assays, their application can be limited, for example, in cases where products are retained inside the cells. Droplet microfluidics, on the other hand, is a rather flexible platform and shows high diversity in single-cell, cell-to-cell interaction-based, and cell-free reaction-based analyses. By examining previous studies, we have summarized the potential of microfluidic technologies to foster sustainable biomanufacturing and advanced biological engineering. [ABSTRACT FROM AUTHOR]

Microbial electrochemical systems have shown great value as a means of enhancing the efficiency of fermentation reactions, but at present, there is no reliable means to balance the extracellular electron supply and corresponding intracellular demands in these systems. The current work describes the unique use of an oxidation–reduction-potential (ORP)-level-controlled microbial electrolysis cell (MEC) system to successfully balance the extracellular electron supply and succinic acid fermentation via A. succinogenes (130Z). The ORP-controlled MEC system with neutral red (NR) yielded a significant increase in succinic acid production (17.21%). The utilization of NR in this MEC system improved the ORP regulatory sensitivity. The optimal approach to the ORP level control was the use of a −400 mV high-voltage electric pulse-based strategy, which increased the yield of succinic acid by 13.08% compared to the control group, and reduced the energy consumption to 52.29% compared to the potentiostatic method. When compared to the −1 V constant potential MEC system, the high-voltage electric pulse-based ORP strategy for the MEC system control provided sufficient electrons to this system while using less electricity (11.96%) and producing 12.48% (74.43 g/L) more succinic acid during fed-batch fermentation. The electronic utilization efficiency of the ORP-controlled MEC system was 192.02%, which was 15.19 times that of the potentiostatic system. The electronic utilization efficiency is significantly increased in the ORP-controlled MEC system. Succinic acid production is ensured by a high-voltage electric pulse-based method, while the influence on cell growth and power consumption are minimized. Fed-batch fermentation with the high-voltage electric pulse-based ORP strategy for MEC system control is noted to be ideal to achieve a further increase in succinic acid concentration and electronic utilization efficiency. [ABSTRACT FROM AUTHOR]

Bio-electrochemical systems have increasingly become the focus of research due to their potential in environmental biotechnology, particularly in the domains of waste utilization and energy recovery. A prominent method within this domain is the transformation of organic matter into hydrogen via microbial electrolysis cells (MECs). This study offers a thorough analysis of MEC performance, employing exergy analysis and incorporating relevant data from the existing literature. The findings of this research indicate a relationship between process efficiency and effective electron transfer originating from biological oxidation to the cathode reaction, facilitating hydrogen generation. The assessment performed revealed that the exergy efficiency of the process varies by a wide range, depending on conditions such as substrate type and concentration, applied external voltage, and the presence of specific inhibitors. This interplay between substrate concentration, overall efficiency, and energy requirement underlines the complex dynamics of optimizing MEC performance. Our insights provide understanding of the challenges in bio-electrochemical systems, offering implications for their sustainable and efficient use in environmental biotechnology. The theoretical analysis involved assessing the utilization of glucose and glycerol, along with the evaluation of electrical energy consumption and hydrogen yield. Our results demonstrate that a higher applied voltage is associated with greater exergy efficiency. Furthermore, after comparing the use of glucose and glycerol as substrates, our study supports the preferential application of glucose for enhanced efficiency. [ABSTRACT FROM AUTHOR]

The abuse of antibiotics leads to the production of antibiotic-resistant bacteria (ARB) and antibiotic resistance genes (ARGs). Microbial electrolysis cells (MECs) have been widely applicated in the field of degrading antibiotics. ARGs were increased via horizontal transfer in single and two-chamber MECs. As one of the critical parameters in MECs, voltage has a particular impact on the ARGs transfer via horizontal transfer. However, there have been few studies of ARGs transfer under the exposure of antibiotics and voltage in MECs. In this study, five concentrations of tetracycline (0, 1, 5, 10, 20 mg/L) were selected to explore the conjugative transfer frequency of plasmid-encoded the ARGs from the donor (E. coli RP4) to receptor (E. coli HB101) in MECs, two voltages (1.5 and 2.0 V) were used to explore the conjugative transfer frequency of ARGs in MECs, then, the transfer of ARGs in MECs under the co-effect of tetracycline and voltage was explored. The results showed that the conjugative transfer frequency of ARGs was significantly increased with the increase of tetracycline concentration and voltage, respectively (p < 0.05). Under the pressure of tetracycline and voltage, the conjugative transfer frequency of ARGs is significantly enhanced with the co-effect of tetracycline and voltage (p < 0.05). The oxidative response induced by electrical stimulation promotes the overproduction of reactive oxygen species and the enhancement of cell membrane permeability of donor and recipient bacteria in MECs. These findings provide insights for studying the spread of ARGs in MECs. [ABSTRACT FROM AUTHOR]

The microbial production of polyhydroxyalkanoate (PHA) block copolymers has attracted research interests because they can be expected to exhibit excellent physical properties. Although post-polymerization conjugation and/or extension have been used for PHA block copolymer synthesis, the discovery of the first sequence-regulating PHA synthase, PhaCAR, enabled the direct synthesis of PHA–PHA type block copolymers in microbial cells. PhaCAR spontaneously synthesizes block copolymers from a mixture of substrates. To date, Escherichia coli and Ralstonia eutropha have been used as host strains, and therefore, sequence regulation is not a host-specific phenomenon. The monomer sequence greatly influences the physical properties of the polymer. For example, a random copolymer of 3-hydroxybutyrate and 2-hydroxybutyrate deforms plastically, while a block copolymer of approximately the same composition exhibits elastic deformation. The structure of the PHA block copolymer can be expanded by in vitro evolution of the sequence-regulating PHA synthase. An engineered variant of PhaCAR can synthesize poly(d-lactate) as a block copolymer component, which allows for greater flexibility in the molecular design of block copolymers. Therefore, creating sequence-regulating PHA synthases with a further broadened substrate range will expand the variety of properties of PHA materials. This review summarizes and discusses the sequence-regulating PHA synthase, analytical methods for verifying block sequence, properties of block copolymers, and mechanisms of sequence regulation. Key points: • Spontaneous monomer sequence regulation generates block copolymers • Poly(D-lactate) segment can be synthesized using a block copolymerization system • Block copolymers exhibit characteristic properties [ABSTRACT FROM AUTHOR]