Prevalence of Foodborne Bacterial Pathogens and Antibiotic Resistance Genes in Sweets from Local Markets in Iran.

BACKGROUND: This study aimed to investigate the prevalences of some important antibiotic-resistance genes (ARGs) and foodborne bacterial pathogens in sweet samples collected from local markets in Iran. METHODS: Forty sweet samples were collected. Foodborne pathogens and ARGs were detected in the sweet samples by conventional and multiplex PCR assays using species-specific primers. RESULTS: Staphylococcus aureus, Cronobacter sakazakii, Shigella spp., Campylobacter jejuni, and Campylobacter coli were detected and identified in 47.5%, 20%, 45%, 5%, and 30% of the sweet samples, respectively. We found S. aureus and Shigella spp. were the most prevalent bacterial pathogens. S. aureus was found to be the most frequent pathogenic bacteria profiled in these samples. We also found a significant correlation between the presence of C. coli and Cr. sakazakii. We detected the blaSHV resistance gene in 97.5% of the sweet samples; however, blaTEM was detected in only one sample (2.5%). CONCLUSIONS: Regarding these results, we suggest preventive strategies such as implementing automation of food processing; monitoring the personal hygiene and health of food handlers, and testing regularly for antibiotic resistance in raw materials and products.

Expanding the genetic toolbox for Cutaneotrichosporon oleaginosus employing newly identified promoters and a novel antibiotic resistance marker.

BACKGROUND: Cutaneotrichosporon oleaginosus is an oleaginous yeast that can produce up to 80% lipid per dry weight. Its high capacity for the biosynthesis of single cell oil makes it highly interesting for the production of engineered lipids or oleochemicals for industrial applications. However, the genetic toolbox for metabolic engineering of this non-conventional yeast has not yet been systematically expanded. Only three long endogenous promoter sequences have been used for heterologous gene expression, further three dominant and one auxotrophic marker have been established. RESULTS: In this study, the structure of putative endogenous promoter sequences was analyzed based on more than 280 highly expressed genes. The identified motifs of regulatory elements and translational initiation sites were used to annotate the four endogenous putative promoter sequences D9FADp, UBIp, PPIp, and 60Sp. The promoter sequences were tested in a construct regulating the known dominant marker hygromycin B phosphotransferase. The four newly described promoters and the previously established GAPDHp successfully initiated expression of the resistance gene and PPIp was selected for further marker development. The geneticin G418 resistance (aminoglycoside 3'-phosphotransferase, APH) and the nourseothricin resistance gene N-acetyl transferase (NAT) were tested for applicability in C. oleaginosus. Both markers showed high transformation efficiency, positive rate, and were compatible for combined use in a successive and simultaneous manner. CONCLUSIONS: The implementation of four endogenous promoters and one novel dominant resistance markers for C. oleaginosus opens up new opportunities for genetic engineering and strain development. In combination with recently developed methods for targeted genomic integration, the established toolbox allows a wide spectrum of new strategies for genetic and metabolic engineering of the industrially highly relevant yeast.

Proteomic and Transcriptomic Analyses to Decipher the Chitinolytic Response of Jeongeupia spp.

In nature, chitin, the most abundant marine biopolymer, does not accumulate due to the action of chitinolytic organisms, whose saccharification systems provide instructional blueprints for effective chitin conversion. Therefore, discovery and deconstruction of chitinolytic machineries and associated enzyme systems are essential for the advancement of biotechnological chitin valorization. Through combined investigation of the chitin-induced secretome with differential proteomic and transcriptomic analyses, a holistic system biology approach has been applied to unravel the chitin response mechanisms in the Gram-negative Jeongeupia wiesaeckerbachi. Hereby, the majority of the genome-encoded chitinolytic machinery, consisting of various glycoside hydrolases and a lytic polysaccharide monooxygenase, could be detected extracellularly. Intracellular proteomics revealed a distinct translation pattern with significant upregulation of glucosamine transport, metabolism, and chemotaxis-associated proteins. While the differential transcriptomic results suggested the overall recruitment of more genes during chitin metabolism compared to that of glucose, the detected protein-mRNA correlation was low. As one of the first studies of its kind, the involvement of over 350 unique enzymes and 570 unique genes in the catabolic chitin response of a Gram-negative bacterium could be identified through a three-way systems biology approach. Based on the cumulative data, a holistic model for the chitinolytic machinery of Jeongeupia spp. is proposed.

Isolation, biochemical characterization, and genome sequencing of two high-quality genomes of a novel chitinolytic Jeongeupia species.

Chitin is the second most abundant polysaccharide worldwide as part of arthropods' exoskeletons and fungal cell walls. Low concentrations in soils and sediments indicate rapid decomposition through chitinolytic organisms in terrestrial and aquatic ecosystems. The enacting enzymes, so-called chitinases, and their products, chitooligosaccharides, exhibit promising characteristics with applications ranging from crop protection to cosmetics, medical, textile, and wastewater industries. Exploring novel chitinolytic organisms is crucial to expand the enzymatical toolkit for biotechnological chitin utilization and to deepen our understanding of diverse catalytic mechanisms. In this study, we present two long-read sequencing-based genomes of highly similar Jeongeupia species, which have been screened, isolated, and biochemically characterized from chitin-amended soil samples. Through metabolic characterization, whole-genome alignments, and phylogenetic analysis, we could demonstrate how the investigated strains differ from the taxonomically closest strain Jeongeupia naejangsanensis BIO-TAS4-2T (DSM 24253). In silico analysis and sequence alignment revealed a multitude of highly conserved chitinolytic enzymes in the investigated Jeongeupia genomes. Based on these results, we suggest that the two strains represent a novel species within the genus of Jeongeupia, which may be useful for environmentally friendly N-acetylglucosamine production from crustacean shell or fungal biomass waste or as a crop protection agent.

Effects of Probiotic Saccharomyces boulardii Supernatant on Viability, Nano-Mechanical Properties of Cytoplasmic Membrane and Pro-Inflammatory Gene Expression in Human Gastric Cancer AGS Cells.

BACKGROUND: Gastric cancer has been recognized as the second most probable cause of death in humans from cancer diseases around the world. Postbiotics, supernatant, and metabolites from probiotic microorganisms have recently been used widely to prevent and treat cancer diseases in humans, without any undesirable side effects. This study explores the antiproliferative and antitumor activities of the probiotic Saccharomyces cerevisiae var. boulardii supernatant (SBS) against AGS cancer cells, a human gastric adenocarcinoma cell line. METHODS: We evaluated cell growth inhibitory and mechanical properties of the cytoplasmic membrane and the downregulation of survivin and proinflammatory genes in AGS cells treated with SBS after 24 and 48 h. RESULTS: SBS significantly inhibits the AGS cell growth, and the concentrations with IC50 values after 24 and 48 h treatments are measured as 2266 and 1956 µg/mL, respectively. Regarding the AFM images and Young`s modulus analysis, SBS significantly induces morphological changes in the cytoplasmic membrane of the treated AGS cells. Expression of survivin, NFƙB, and IL-8 genes is significantly suppressed in AGS cells treated with SBS. CONCLUSIONS: Considering the antitumor activities of SBS on AGS cell line, it can be regarded as a prospective therapeutic and preventive strategy against human stomach cancer disease.

Rare earths stick to rare cyanobacteria: Future potential for bioremediation and recovery of rare earth elements.

Biosorption of metal ions by phototrophic microorganisms is regarded as a sustainable and alternative method for bioremediation and metal recovery. In this study, 12 cyanobacterial strains, including 7 terrestrial and 5 aquatic cyanobacteria, covering a broad phylogenetic diversity were investigated for their potential application in the enrichment of rare earth elements through biosorption. A screening for the maximum adsorption capacity of cerium, neodymium, terbium, and lanthanum was conducted in which Nostoc sp. 20.02 showed the highest adsorption capacity with 84.2-91.5 mg g-1. Additionally, Synechococcus elongatus UTEX 2973, Calothrix brevissima SAG 34.79, Desmonostoc muscorum 90.03, and Komarekiella sp. 89.12 were promising candidate strains, with maximum adsorption capacities of 69.5-83.4 mg g-1, 68.6-83.5 mg g-1, 44.7-70.6 mg g-1, and 47.2-67.1 mg g-1 respectively. Experiments with cerium on adsorption properties of the five highest metal adsorbing strains displayed fast adsorption kinetics and a strong influence of the pH value on metal uptake, with an optimum at pH 5 to 6. Studies on binding specificity with mixed-metal solutions strongly indicated an ion-exchange mechanism in which Na+, K+, Mg2+, and Ca2+ ions are replaced by other metal cations during the biosorption process. Depending on the cyanobacterial strain, FT-IR analysis indicated the involvement different functional groups like hydroxyl and carboxyl groups during the adsorption process. Overall, the application of cyanobacteria as biosorbent in bioremediation and recovery of rare earth elements is a promising method for the development of an industrial process and has to be further optimized and adjusted regarding metal-containing wastewater and adsorption efficiency by cyanobacterial biomass.

Molecular Mechanisms of Shigella Pathogenesis; Recent Advances.

Shigella species are the main cause of bacillary diarrhoea or shigellosis in humans. These organisms are the inhabitants of the human intestinal tract; however, they are one of the main concerns in public health in both developed and developing countries. In this study, we reviewed and summarised the previous studies and recent advances in molecular mechanisms of pathogenesis of Shigella Dysenteriae and non-Dysenteriae species. Regarding the molecular mechanisms of pathogenesis and the presence of virulence factor encoding genes in Shigella strains, species of this bacteria are categorised into Dysenteriae and non-Dysenteriae clinical groups. Shigella species uses attachment, invasion, intracellular motility, toxin secretion and host cell interruption mechanisms, causing mild diarrhoea, haemorrhagic colitis and haemolytic uremic syndrome diseases in humans through the expression of effector delivery systems, protein effectors, toxins, host cell immune system evasion and iron uptake genes. The investigation of these genes and molecular mechanisms can help us to develop and design new methods to detect and differentiate these organisms in food and clinical samples and determine appropriate strategies to prevent and treat the intestinal and extraintestinal infections caused by these enteric pathogens.

A quantitative prevalence of Escherichia coliO157 in different food samples using real-time qPCR method.

Escherichia coli serogroup O157 is the main causative agent of several intestinal and extra-intestinal foodborne diseases in humans through consumption of low-dose contaminated foods such as milk, beef, and vegetables. To date, studies regarding the quantitative prevalence of E. coli O157 in foods are so limited. Therefore, this study aimed to evaluate the quantitative prevalence rate of E. coli serogroup O157 in raw milk (n = 144), vegetable salad (n = 174), and minced beef samples (n = 108) using the real-time qPCR SYBR green melting curve method targeting the rfbA gene. First, we evaluated the method and found a sensitive and specific qPCR assay with 1 log of CFU/ml detection limit to detect E. coli O157 (Tm = 80.3 ± 0.1°C). About 2.77%, 10.18%, and 9.19% of raw milk, minced beef, and vegetable salad samples, respectively, were contaminated with E. coli O157. Minced beef and vegetable salad samples were significantly more contaminated than raw milk samples. Population average of E. coli O157 in raw milk, minced beef, and vegetable salad samples were 2.22 ± 0.57, 3.30 ± 0.40, and 1.65 ± 0.44 log CFU/ml or gr, respectively. Significantly higher levels of population of E. coli O157 were observed in minced beef samples. Minced beef can be regarded as the main food in the transmission of this foodborne pathogen. Routine quantitative rapid monitoring is strongly suggested to be carried out to prevent foodborne diseases caused by E. coli O157.

Structural and Functional Insight into the Mechanism of the Fe-S Cluster-Dependent Dehydratase from Paralcaligenes ureilyticus.

Enzyme-catalyzed reaction cascades play an increasingly important role for the sustainable manufacture of diverse chemicals from renewable feedstocks. For instance, dehydratases from the ilvD/EDD superfamily have been embedded into a cascade to convert glucose via pyruvate to isobutanol, a platform chemical for the production of aviation fuels and other valuable materials. These dehydratases depend on the presence of both a Fe-S cluster and a divalent metal ion for their function. However, they also represent the rate-limiting step in the cascade. Here, catalytic parameters and the crystal structure of the dehydratase from Paralcaligenes ureilyticus (PuDHT, both in presence of Mg2+ and Mn2+ ) were investigated. Rate measurements demonstrate that the presence of stoichiometric concentrations Mn2+ promotes higher activity than Mg2+ , but at high concentrations the former inhibits the activity of PuDHT. Molecular dynamics simulations identify the position of a second binding site for the divalent metal ion. Only binding of Mn2+ (not Mg2+ ) to this site affects the ligand environment of the catalytically essential divalent metal binding site, thus providing insight into an inhibitory mechanism of Mn2+ at higher concentrations. Furthermore, in silico docking identified residues that play a role in determining substrate binding and selectivity. The combined data inform engineering approaches to design an optimal dehydratase for the cascade.

Stripped: contribution of cyanobacterial extracellular polymeric substances to the adsorption of rare earth elements from aqueous solutions.

The transformation of modern industries towards enhanced sustainability is facilitated by green technologies that rely extensively on rare earth elements (REEs) such as cerium (Ce), neodymium (Nd), terbium (Tb), and lanthanum (La). The occurrence of productive mining sites, e.g., is limited, and production is often costly and environmentally harmful. As a consequence of increased utilization, REEs enter our ecosystem as industrial process water or wastewater and become highly diluted. Once diluted, they can hardly be recovered by conventional techniques, but using cyanobacterial biomass in a biosorption-based process is a promising eco-friendly approach. Cyanobacteria can produce extracellular polymeric substances (EPS) that show high affinity to metal cations. However, the adsorption of REEs by EPS has not been part of extensive research. Thus, we evaluated the role of EPS in the biosorption of Ce, Nd, Tb, and La for three terrestrial, heterocystous cyanobacterial strains. We cultivated them under N-limited and non-limited conditions and extracted their EPS for compositional analyses. Subsequently, we investigated the metal uptake of a) the extracted EPS, b) the biomass extracted from EPS, and c) the intact biomass with EPS by comparing the amount of sorbed REEs. Maximum adsorption capacities for the tested REEs of extracted EPS were 123.9-138.2 mg g-1 for Komarekiella sp. 89.12, 133.1-137.4 mg g-1 for Desmonostoc muscorum 90.03, and 103.5-129.3 mg g-1 for Nostoc sp. 20.02. A comparison of extracted biomass with intact biomass showed that 16% (Komarekiella sp. 89.12), 28% (Desmonostoc muscorum 90.03), and 41% (Nostoc sp. 20.02) of REE adsorption was due to the biosorption of the extracellular EPS. The glucose- rich EPS (15%-43% relative concentration) of all three strains grown under nitrogen-limited conditions showed significantly higher biosorption rates for all REEs. We also found a significantly higher maximum adsorption capacity of all REEs for the extracted EPS compared to cells without EPS and untreated biomass, highlighting the important role of the EPS as a binding site for REEs in the biosorption process. EPS from cyanobacteria could thus be used as efficient biosorbents in future applications for REE recycling, e.g., industrial process water and wastewater streams.

Dietary Modulation of the Human Gut Microbiota and Metabolome with Flaxseed Preparations.

Flaxseeds are typically consumed either as whole flaxseed, ground flaxseed, flaxseed oil, partially defatted flaxseed meal, or as a milk alternative. They are considered a rich source of vitamins, minerals, proteins and peptides, lipids, carbohydrates, lignans, and dietary fiber, which have shown hypolipidemic, antiatherogenic, anticholesterolemic, and anti-inflammatory property activity. Here, an in vitro batch culture model was used to investigate the influence of whole milled flaxseed and partially defatted milled flaxseed press cake on the gut microbiota and the liberation of flaxseed bioactives. Microbial communities were profiled using 16S rRNA gene-based high-throughput sequencing with targeted mass spectrometry measuring lignan, cyclolinopeptide, and bile acid content and HPLC for short-chain fatty acid profiles. Flaxseed supplementation decreased gut microbiota richness with Firmicutes, Proteobacteria, and Bacteroidetes becoming the predominant phyla. Secoisolariciresinol, enterodiol, and enterolactone were rapidly produced with acetic acid, butyric acid, and propionic acid being the predominant acids after 24 h of fermentation. The flaxseed press cake and whole flaxseed were equivalent in microbiota changes and functionality. However, press cake may be superior as a functional additive in a variety of foods in terms of consumer acceptance as it would be more resistant to oxidative changes.

Efficient Green Light Acclimation of the Green Algae Picochlorum sp. Triggering Geranylgeranylated Chlorophylls.

In analogy to higher plants, eukaryotic microalgae are thought to be incapable of utilizing green light for growth, due to the "green gap" in the absorbance profiles of their photosynthetic pigments. This study demonstrates, that the marine chlorophyte Picochlorum sp. is able to grow efficiently under green light emitting diode (LED) illumination. Picochlorum sp. growth and pigment profiles under blue, red, green and white LED illumination (light intensity: 50-200 µmol m-2 s-1) in bottom-lightened shake flask cultures were evaluated. Green light-treated cultures showed a prolonged initial growth lag phase of one to 2 days, which was subsequently compensated to obtain comparable biomass yields to red and white light controls (approx. 0.8 gDW L-1). Interestingly, growth and final biomass yields of the green light-treated sample were higher than under blue light with equivalent illumination energies. Further, pigment analysis indicated, that during green light illumination, Picochlorum sp. formed unknown pigments (X1-X4). Pigment concentrations increased with illumination intensity and were most abundant during the exponential growth phase. Mass spectrometry and nuclear magnetic resonance data indicated, that pigments X1-X2 and X3-X4 are derivatives of chlorophyll b and a, which harbor C=C bonds in the phytol side chain similar to geranylgeranylated chlorophylls. Thus, for the first time, the natural accumulation of large pools (approx. 12 mg gDW -1) of chlorophyll intermediates with incomplete hydrogenation of their phytyl chains is demonstrated for algae under monochromatic green light (Peak λ 510 nm, full width at half maximum 91 nm). The ability to utilize green light offers competitive advantages for enhancing biomass production, particularly under conditions of dense cultures, long light pathways and high light intensity. Green light acclimation for an eukaryotic microalgae in conjunction with the formation of new aberrant geranylgeranylated chlorophylls and high efficiency of growth rates are novel for eukaryotic microalgae. Illumination with green light could enhance productivity in industrial processes and trigger the formation of new metabolites-thus, underlying mechanisms require further investigation.

Isolation and Investigation of Natural Rare Earth Metal Chelating Agents From Calothrix brevissima - A Step Towards Unraveling the Mechanisms of Metal Biosorption.

In this study water soluble compounds that form complexes with Rare Earth Elements (REE) and other metals were isolated from Calothrix brevissima biomass with chromatographic methods for the first time. Molecular characterization showed that the isolated compounds are most likely polysaccharides comprised of arabinose, xylose, mannose, galactose and glucose. FT-IR analysis revealed functional groups involved in the binding mechanism of Tb are likely sulfate- and to a lesser extend hydroxyl-groups. The binding specificity of the isolated compounds was investigated with different metal solutions. Here, ions of the alkali and alkaline earth metals Na, K, Mg and Ca showed no competition for Tb-binding even at 10-fold excess concentration. Ions of the elements Co and Pb on the other hand replaced Tb at higher concentrations. Addition of the isolated compounds significantly reduced the precipitation of Eu at pH-values between 6.7 and 9.5, indicating that the interaction between the isolated chelators and Rare Earth Metals is stable even at high pH-values.

FTIR differentiation based on genomic DNA for species identification of Shigella isolates from stool samples.

Shigellosis is one of the major public health concerns in developing and low-income countries caused by four species of Shigella. There is an apparent need to develop rapid, cost-effective, sensitive and specific methods for differentiation of Shigella species to be used in outbreaks and health surveillance systems. We developed a sensitive and specific Fourier-transform infrared spectroscopy (FTIR) based method followed by principal component analysis (PCA) and hierarchical clustering analysis (HCA) assays to differentiate four species of Shigella isolates from stool samples. The FTIR based method was evaluated by differentiation of 91 Shigella species from each other in clinical samples using both gold standards (culture-based and agglutination methods) and developed FTIR assay; eventually, the sensitivity and specificity of the developed method were calculated. In summary, four distinct FTIR spectra associated with four species of Shigella were obtained with wide variations in three definite regions, including 1800-1550 cm-1, 1550-1100 cm-1, and 1100-800 cm-1 distinguish these species from each other. In this study, we found the FTIR method followed by PCA analysis with specificity, sensitivity, differentiation error and correct differentiation rate values of 100, 100, 0 and 100%, respectively, for identification and differentiation of all species of the Shigella in stool samples.

A Newly Designed Automatically Controlled, Sterilizable Flat Panel Photobioreactor for Axenic Algae Culture.

In context of the global climate change, microalgae processes are gaining momentum as a biotechnological tool for direct fixation and valorization of greenhouse gases. Algae have the metabolic capacity to photosynthetically convert CO2 into high value products, such as food additives, under economic boundary conditions. High cost, commercial flat panel gas-lift bioreactors for microalgae cultivation at laboratory scale provide either small volumes or no sterile operation, which limits academic research. This brief report presents initial data for a new type of sterile operating flat panel gas-lift bioreactor with a unique asymmetrical U-shape. It utilizes automatable process control technologies that adhere to industrial standards to enhance data reproducibility and aid industrial scale up. The practicability was demonstrated using a Chlorella sorokiniana cultivation, which showed the typical growth behavior. Due to the sophisticated implemented control engineering technology, pivotal parameters as pH and temperature can be determined within a range of ±0.1 units, which was confirmed experimentally. The new flat panel gas-lift photobioreactor presented in this brief report fills the technology gap at laboratory scale with an autoclavable volume of 7.2 L. Moreover, it is easy to rebuild by means of the hereby provided blueprint, while exhibiting a six-fold cost reduction compared to commercially available flat panel photobioreactors.

Kinetic and Structural Characterization of the First B3 Metallo-ß-Lactamase with an Active-Site Glutamic Acid.

The structural diversity in metallo-ß-lactamases (MBLs), especially in the vicinity of the active site, has been a major hurdle in the development of clinically effective inhibitors. Representatives from three variants of the B3 MBL subclass, containing either the canonical HHH/DHH active-site motif (present in the majority of MBLs in this subclass) or the QHH/DHH (B3-Q) or HRH/DQK (B3-RQK) variations, were reported previously. Here, we describe the structure and kinetic properties of the first example (SIE-1) of a fourth variant containing the EHH/DHH active-site motif (B3-E). SIE-1 was identified in the hexachlorocyclohexane-degrading bacterium Sphingobium indicum, and kinetic analyses demonstrate that although it is active against a wide range of antibiotics, its efficiency is lower than that of other B3 MBLs but has increased efficiency toward cephalosporins relative to other ß-lactam substrates. The overall fold of SIE-1 is characteristic of the MBLs; the notable variation is observed in the Zn1 site due to the replacement of the canonical His116 by a glutamate. The unusual preference of SIE-1 for cephalosporins and its occurrence in a widespread environmental organism suggest the scope for increased MBL-mediated ß-lactam resistance. Thus, it is relevant to include SIE-1 in MBL inhibitor design studies to widen the therapeutic scope of much needed antiresistance drugs.

Terbium Excitation Spectroscopy as a Detection Method for Chromatographic Separation of Lanthanide-Binding Biomolecules.

Studies of biosorption and bioaccumulation of heavy metals deal mostly with challenging, inhomogeneous, and complex materials. Therefore, most reports describe only application studies, while fundamental research is limited to indirect methods and speculations on the binding mechanisms. In this study, we describe a method for detecting and isolating heavy metal-binding biomolecules directly from crude extracts. The underlying principle is terbium sensitization and fluorescence excitation spectroscopy used offline after a chromatographic run. Compounds interacting with metal ions inevitably change the coordination sphere of terbium, which is reflected in the excitation spectrum leading to metal-specific luminescence. Main advantages of our approach include simple, fast, and inexpensive experiment design, nondestructive measurements, and detection limits far below 1 mg. Here, we have applied our method for three promising biosorbents (green algae, moss, and cyanobacterium) and obtained first information on the character of active compounds isolated from each species.

Additive Analytics: Easy Transformation of Low-Cost Fused Deposition Modeling Three-Dimensional Printers for HPTLC Sample Application.

Additive manufacturing, known as three-dimensional (3D) printing technologies, has revolutionized production in all domains of science and technology. Although 3D printing has a high impact on research and development, its capacity to implement low-cost, flexible, and robust sample handling automation has not been exploited in full. To this end, we have created a low-cost, robust, and easy-to-utilize kit to transform an off-the-shelf fused deposition modeling 3D printer to a thin layer chromatography (TLC) sample application device. Our technology solution improves TLC convenience when higher throughput of the established method is required. The developed dual-needle sprayer allows simple and exceptionally robust automatic sample application. The device is especially well-suited for high-performance TLC-assisted method selection in counter-current chromatography. A step-by-step guide and list of required parts, including 3D printable files with instruction, can be obtained from the Supporting Information for research usage and open development.

Enzymatic Modification of Native Chitin and Conversion to Specialty Chemical Products.

: Chitin is one of the most abundant biomolecules on earth, occurring in crustacean shells and cell walls of fungi. While the polysaccharide is threatening to pollute coastal ecosystems in the form of accumulating shell-waste, it has the potential to be converted into highly profitable derivatives with applications in medicine, biotechnology, and wastewater treatment, among others. Traditionally this is still mostly done by the employment of aggressive chemicals, yielding low quality while producing toxic by-products. In the last decades, the enzymatic conversion of chitin has been on the rise, albeit still not on the same level of cost-effectiveness compared to the traditional methods due to its multi-step character. Another severe drawback of the biotechnological approach is the highly ordered structure of chitin, which renders it nigh impossible for most glycosidic hydrolases to act upon. So far, only the Auxiliary Activity 10 family (AA10), including lytic polysaccharide monooxygenases (LPMOs), is known to hydrolyse native recalcitrant chitin, which spares the expensive first step of chemical or mechanical pre-treatment to enlarge the substrate surface. The main advantages of enzymatic conversion of chitin over conventional chemical methods are the biocompability and, more strikingly, the higher product specificity, product quality, and yield of the process. Products with a higher Mw due to no unspecific depolymerisation besides an exactly defined degree and pattern of acetylation can be yielded. This provides a new toolset of thousands of new chitin and chitosan derivatives, as the physio-chemical properties can be modified according to the desired application. This review aims to provide an overview of the biotechnological tools currently at hand, as well as challenges and crucial steps to achieve the long-term goal of enzymatic conversion of native chitin into specialty chemical products.

Biogas yields and composition from oil-extracted halophilic algae residues in conventional biogas plants operated at high salinities.

CO2-induced climate change drives the development of renewable processes for industrial products. Algae processes can actively fix and convert CO2 into value adding products, such as oils. Algae lipids hence counteract climate change and provide access to renewable commodities. In this context, valorization of algal residues remaining after oil extraction is a challenge for the emerging cyclic bioeconomy. This study focuses on the valorization of oil-extracted algae residues derived from the halophilic strain Scenedesmus obliquus via anaerobic digestion. We examined the effect of prior oil extraction on microbial digestibility and increasing salt content in the substrate with regard to biogas yield and composition. Our cumulative data demonstrate that the supercritical CO2 oil extraction acts as a physical pretreatment that facilitates enhanced hydrolysis of both polymeric call wall carbohydrates and cellular proteins, providing methane yields of 213.2 LN kg-1 VS day-1. Methane yields were 20% higher than literature values obtained with the same algae strain in the absence of prior oil extraction. We obtained these superior results albeit all lipids and nonpolar proteins had been extracted from the biogas substrate. Our data indicate that continuous anaerobic digestion without loss of fermentation efficiency is feasible up to a salt concentration of 2% w/v, if conventional, agricultural biogas plants are gradually adapted to the salt content of the substrate. Monofermentation of the investigated oil-extracted algae residue is technically feasible at loading rates of 1.5 kg VS m-3 day-1, but a supplementation with carbohydrate rich biomass would prove beneficial to alleviate ammonia inhibition.