Microbial Cell Factories最新文献

Extensive anthropogenic activity has led to the accumulation of organic and inorganic contaminants in diverse ecosystems, which presents significant challenges for the environment and its inhabitants. Utilizing microalgae as a bioremediation tool can present a potential solution to these challenges. Microalgae have gained significant attention as a promising biotechnological solution for detoxifying environmental pollutants. This is due to their advantages, such as rapid growth rate, cost-effectiveness, high oil-rich biomass production, and ease of implementation. Moreover, microalgae-based remediation is more environmentally sustainable for not generating additional waste sludge, capturing atmospheric CO₂, and being efficient for nutrient recycling and sustainable algal biomass production for biofuels and high-value-added products generation. Hence, microalgae can achieve sustainability's three main pillars (environmental, economic, and social). Microalgal biomass can mediate contaminated wastewater effectively through accumulation, adsorption, and metabolism. These mechanisms enable the microalgae to reduce the concentration of heavy metals and organic contaminants to levels that are considered non-toxic. However, several factors, such as microalgal strain, cultivation technique, and the type of pollutants, limit the understanding of the microalgal removal mechanism and efficiency. Furthermore, adopting novel technological advancements (e.g., nanotechnology) may serve as a viable approach to address the challenge of refractory pollutants and bioremediation process sustainability. Therefore, this review discusses the mechanism and the ability of different microalgal species to mitigate persistent refractory pollutants, such as industrial effluents, dyes, pesticides, and pharmaceuticals. Also, this review paper provided insight into the production of nanomaterials, nanoparticles, and nanoparticle-based biosensors from microalgae and the immobilization of microalgae on nanomaterials to enhance bioremediation process efficiency. This review may open a new avenue for future advancing research regarding a sustainable biodegradation process of refractory pollutants.

Background: Functional foods and dairy products are gaining global attention due to their nutritional value and health-promoting characteristics. Lactic acid bacteria (LAB) are one of the promising components included in these products, thanks to their probiotic properties and ability to produce bioactive compounds such as bacteriocins. On the other hand, ectomycorrhizal wild mushrooms (truffles) are known for their ethnomycological importance. Hence, we aimed to develop a functional dairy product using a bacteriocin-producing LAB isolate that has probiotic potentials together with the bioactive extract of a truffle mushroom.

Results: Screening for bacteriocin-producing LAB led to the selection of four safe isolates that also showed promising probiotic potentials. Isolate No. 7 was selected due to its wider antimicrobial spectrum and was identified as Lactiplantibacillus plantarum strain GA7. Out of resulting bands from Tricine SDS-PAGE analysis, a band (its molecular mass was approximately 7 kDa) exhibited antimicrobial activity. Amino acid sequencing of this active band detected 62 amino acid residues with 100% identity to plantaricin ASM1 bacteriocin. Simultaneously, an ethyl acetate extract was prepared from a truffle sample identified as Tirmania pinoyi. Safety of this truffle was confirmed and its extract exerted promising antioxidant and hypocholesterolemic activity. Prepared functional dairy products (Labneh) fortified with L. plantarum GA7 and nano-encapsulated T. pinoyi extract exhibited superior physicochemical, sensory and antioxidant properties compared to control. Moreover, an increase in probiotic count was observed in presence of T. pinoyi extract. Furthermore, prepared Labneh using the bacteriocin-producing L. plantarum GA7 and nano-encapsulated T. pinoyi extract remained unspoiled for over 60 days, compared to control, which spoiled after 21 days.

Conclusion: Besides improving Labneh physicochemical, sensory and antioxidant properties, the presence of the bacteriocin-producing L. plantarum GA7 has contributed in significantly extending its shelf life, while T. pinoyi extract showed prebiotic influence on probiotic count. As far as we know this is the first study describing production of a functional synbiotic dairy product fortified with bacteriocin-producing probiotic LAB and bioactive T. pinoyi truffle extract.

Background: Streptomyces roseochromogenes NRRL 3504 produces clorobiocin, an aminocoumarin antibiotic that inhibits DNA replication. No other natural products have been isolated from this bacterium so far, despite the presence of a rich repertoire of specialized metabolite biosynthesis gene clusters (smBGCs) within its genome. Heterologous expression of smBGCs in suitable chassis speeds up the discovery of the natural products hidden behind these sets of genes.

Results: In this work we focus on one intriguing smBGC of NRRL 3504 bearing some similarity to gene clusters involved in production of manumycin family polyketides. Through heterologous expression in Streptomyces chassis strains S. albus Del14 and S. lividans ΔYA9, this smBGC (hereafter referred to as lim BGC) was shown to direct the production of unusual polyketide limocrocin (LIM) known for its ability to interfere with viral reverse transcriptases. The organization of lim BGC, data on the structures of revealed metabolites as well as manipulations of lim genes allowed us to put forward an initial hypothesis about a biosynthetic pathway leading to LIM. We provide initial data on two LIM derivatives as well as updated NMR spectra for the main product.

Conclusion: This study reveals the genetic control of biosynthesis of LIM that remained hidden for the last 70 years. This, in turn, opens the door to biological routes towards overproduction of LIM as well as generation of its derivatives.

Oleaginous yeasts are considered promising sources for lipid production due to their ability to accumulate high levels of lipids under appropriate growth conditions. The current study aimed to isolate and identify oleaginous yeasts having superior ability to accumulate high quantities of lipids; and enhancing lipid production using response surface methodology and repeated-batch fermentation. Results revealed that, twenty marine oleaginous yeasts were isolated, and the most potent lipid producer isolate was Candida parapsilosis Y19 according to qualitative screening test using Nile-red dye. Orange peels was used as substrate where C. parapsilosis Y19 produced 1.14 g/l lipids at 23.0% in batch fermentation. To enhance the lipid production, statistical optimization using Taguchi design through Response surface methodology was carried out. Total lipids were increased to 2.46 g/l and lipid content increased to 30.7% under optimal conditions of: orange peel 75 g/l, peptone 7 g/l, yeast extract 5 g/l, inoculum size 2% (v/v), pH 5 and incubation period 6 d. Furthermore, repeated-batch fermentation of C. parapsilosis Y19 enhanced lipid production where total lipids increased at 4.19 folds (4.78 g/l) compared to batch culture (before optimization). Also, the lipid content was increased at 1.7 folds (39.1%) compared to batch culture (before optimization). Fatty acid profile of the produced lipid using repeated-batch fermentation includes unsaturated fatty acids (USFAs) at 74.8% and saturated fatty acids (SFAs) at 25.1%. Additionally, in repeated-batch fermentation, the major fatty acid was oleic acid at 45.0%; followed by linoleic acid at 26.0%. In conclusion, C. parapsilosis Y19 is considered a promising strain for lipid production. Also, both statistical optimizations using RSM and repeated-batch fermentation are efficient methods for lipid production from C. parapsilosis Y19.

Background: Paenibacillus polymyxa, is a Gram-positive, plant growth promoting bacterium, known for producing 98% optically pure 2,3-butanediol, an industrially valuable chemical for solvents, plasticizers and resins. Immobilization of Paenibacillus polymyxa has been proposed to improve the cell stability and efficiency of the fermentation process, reduce contamination and provide easy separation of butanediol in the culture broth as compared to conventional bioprocesses. This research aimed to explore the potential of Paenibacillus polymyxa with immobilization technique to produce 2,3-butanediol.

Results: We investigated different immobilization methods with natural biopolymers like alginate, chitosan and carrageenan-chitosan-based immobilization. These methods were further investigated for their immobilization efficiency and yield in 2,3-butanediol production. Carrageenan-chitosan beads enabled a higher cell concentration and demonstrated superior cell retention to calcium-alginate-chitosan beads. Carrageenan-chitosan immobilization preserved 2,3-butanediol production in bacteria and increased the product formation rate.

Conclusion: Carrageenan-chitosan immobilization enables non-pathogenic Paenibacillus polymyxa to be a capable 2,3-butanediol producer with increased product formation rate, which has not been previously reported. This novel strategy offers promising alternative to traditional fermentation processes using pathogenic strains and can be further applied in co-cultivations for metabolite production, wastewater management and bioremediation.

Genome mining is a promising avenue for expanding the repertoire of microbial natural products, which are important for drug development. This approach involves predicting genetically encoded small molecules by examining bacterial genomes via accumulated knowledge of microbial biosynthesis. However, it is also important that the microbes produce the predicted molecule in practice. Here, we introduce an endophytic Streptomyces sp. N50, which was isolated from the medicinal plant Selaginella tamariscina. Upon sequencing its entire genome, 33 biosynthetic gene clusters (BGCs) were identified in a chromosome and a megaplasmid. Subsequent genome mining revealed that the new 15-deoxynaphthomycin could be produced due to the presence of an enoyl reductase domain, which is absent in the known BGC of naphthomycin, a type of ansamycin antibiotics. In addition, the engineered strain with the introduction of the global regulatory gene afsR2 into N50 successfully produced 15-deoxynaphthomycins. Furthermore, molecular network analysis via MS/MS selectively confirmed the presence of additional sulfur-containing 15-deoxynaphthomycin congeners. Eventually, six new 15-deoxynaphthomycins were isolated and elucidated from the engineered strain N50. This family of compounds is known to exhibit various biological activities. Also, the presence of quinone moieties in these compounds, which are known to activate NRF2, they were tested for their ability to activate NRF2. Among the new compounds, three (1, 5, and 6) activated the antioxidant NRF2-ARE signaling pathway. Treatment with these compounds significantly elevated NRF2 levels in HepG2 cells and further induced the expression of NRF2 target genes associated with the antioxidant response. This study suggests that the combination of genome mining, gene engineering and molecular networking is helpful for generating new small molecules as pharmaceutical candidates from microorganisms.

Background: The biosynthesis of the natural product family of the polycyclic tetramate macrolactams (PoTeMs) employs an uncommon iterative polyketide synthase/non-ribosomal peptide synthetase (iPKS/NRPS). This machinery produces a universal PoTeM biosynthetic precursor that contains a tetramic acid moiety connected to two unsaturated polyene side chains. The enormous structural and hence functional diversity of PoTeMs is enabled by pathway-specific tailoring enzymes, particularly cyclization-catalyzing oxidases that process the polyene chains to form distinct ring systems, and further modifying enzymes.

Results: Ikarugamycin is the first discovered PoTeM and is formed by the three enzymes IkaABC. Utilizing the iPKS/NRPS IkaA, we established a genetic plug-and-play system by screening eight different strong promoters downstream of ikaA to facilitate high-level heterologous expression of PoTeMs in different Streptomyces host systems. Furthermore, we applied the system on three different PoTeM modifying genes (ptmD, ikaD, and cftA), showing the general utility of this approach to study PoTeM post-PKS/NRPS processing of diverse tailoring enzymes.

Conclusion: By employing our plug-and-play system for PoTeMs, we reconstructed the ikarugamycin biosynthesis and generated five derivatives of ikarugamycin. This platform will generally facilitate the investigation of new PoTeM biosynthetic cyclization and tailoring reactions in the future.

Background: During the last decades, the advancements in synthetic biology opened the doors for a profusion of cost-effective, fast, and ecologically friendly medical applications priorly unimaginable. Following the trend, the genetic engineering of the baker's yeast, Saccharomyces cerevisiae, propelled its status from an instrumental ally in the food industry to a therapy and prophylaxis aid.

Main text: In this review, we scrutinize the main applications of engineered S. cerevisiae in the medical field focusing on its use as a cell factory for pharmaceuticals and vaccines, a biosensor for diagnostic and biomimetic assays, and as a live biotherapeutic product for the smart in situ treatment of intestinal ailments. An extensive view of these fields' academic and commercial developments as well as main hindrances is presented.

Conclusion: Although the field still faces challenges, the development of yeast-based medical applications is often considered a success story. The rapid advances in synthetic biology strongly support the case for a future where engineered yeasts play an important role in medicine.

Background: Ecotoxicology is essential for the evaluation and comprehension of the effects of emergency pollutants (EP) such as heavy metal ions on the natural environment. EPs pose a substantial threat to the health of humans and the proper functioning of the global ecosystem. The primary concern is the exposure of humans and animals to heavy metal ions through contaminated water. The presence of heavy metal ions in drinking water ought to be monitored in accordance with World Health Organization regulations. Among the numerous harmful metal ions, copper ions are responsible for a variety of human diseases.

Results: This study investigates the application of phage display as a screening method for heavy metal toxicological targets, with copper served as the main focus. To identify a variety of Cu-binding M13 phage clones with unique peptides and to assess their affinity for metal ions, the study utilized Escherichia coli as a factories producing recombinant bacteriophages, modified biopanning procedure and an ELISA assay. The research highlights the increasing importance of phage display as a screening tool in ecotoxicology. We synthesized and modified the selected peptide to enable the rapid optical detection of Cu(II) ions in aqueous solutions. By incorporating the dansyl group into a designated peptide sequence, we implemented fluorescence detection assays for real-time measurements. The Cu²⁺- binding peptide's efficacy was confirmed through spectroscopic measurements, which allowed for real-time detection with rapid response times with high selectivity.

Conclusions: The phage display technique was successfully applied to develop the fluorescent peptide-based chemosensor that exhibited high selectivity and sensitivity for Cu²⁺.

Background: In the soy sauce fermentation industry, Aspergillus oryzae (A. oryzae) plays an essential role and is frequently subjected to high salinity levels, which pose a significant osmotic stress. This environmental challenge necessitates the activation of stress response mechanisms within the fungus. The Zn(II)₂Cys₆ family of transcription factors, known for their zinc binuclear cluster-containing proteins, are key regulators in fungi, modulating various cellular functions such as stress adaptation and metabolic pathways.

Results: Overexpression of AozC decreased growth rates in the presence of salt, while its knockdown enhanced growth, the number of spores, and biomass, particularly under conditions of 15% salt concentration, doubling these metrics compared to the wild type. Conversely, the knockdown of AozC via RNA interference significantly enhanced spore density and dry biomass, particularly under 15% salt stress, where these parameters were markedly improved over the wild type strain. Moreover, the overexpression of AozC led to a downregulation of the FAD2 gene, a pivotal enzyme in the biosynthesis of unsaturated fatty acids (UFAs), which are essential for preserving cell membrane fluidity and integrity under saline conditions. Transcriptome profiling further exposed the influence of AozC on the regulation of UFA biosynthesis and the modulation of critical stress response pathways. Notably, the regulatory role of AozC in the mitogen-activated protein kinase (MAPK) signaling and ABC transporters pathways was highlighted, underscoring its significance in cellular osmotic balance and endoplasmic reticulum homeostasis. These findings collectively indicate that AozC functions as a negative regulator of salt tolerance in A. oryzae.

Conclusion: This research suggest that AozC acts as a negative regulator in salt tolerance and modulates fatty acid biosynthesis in response to osmotic stress. These results provide insights into the regulatory mechanisms of stress adaptation in A. oryzae.