Strain Improvement Research Articles

Global transcriptomic response of Escherichia coli to p-coumaric acid

The aromatic compound p-coumaric acid (p-CA) is a secondary metabolite produced by plants. This aromatic acid and derived compounds have positive effects on human health, so there is interest in producing them in biotechnological processes with recombinant Escherichia coli strains. To determine the physiologic response of E. coli W3110 to p-CA, dynamic expression analysis of selected genes fused to a fluorescent protein reporter as well as RNA-seq and RT-qPCR were performed. The observed transcriptional profile revealed the induction of genes involved in functions related to p-CA active export, synthesis of cell wall and membrane components, synthesis of amino acids, detoxification of formaldehyde, phosphate limitation, acid stress, protein folding and degradation. Downregulation of genes encoding proteins involved in energy production, carbohydrate import and metabolism, as well as several outer and plasma membrane proteins was detected. This response is indicative of cell envelope damage causing the leakage of intracellular components including amino acids and phosphate-containing compounds. The cellular functions responding to p-CA that were identified in this study will help in defining targets for production strains improvement.

Isolating Lysobacter enzymogenes strains with enhanced protease activity via chemical mutagenesis

Proteases are the most important industrial enzymes which have attracted enormous attention due to their vast variety and well-defined specificity. Microbial proteases are superior to other sources like plant and animal proteases because of their desired characteristics for biotechnological application. In this regard, Lysobacter enzymogenes is a rich source for the production of antibiotics and proteases. However, strain improvement in order to obtain overproduced microorganisms is always demanded at an industrial scale. Therefore, in the present study in order to enhance L. enzymogenes protease production, random mutagenesis was applied using N-methyl-N’-nitro-N-nitrosoguanidine (NTG) as a chemical mutagen. Random mutagenesis was conducted on L. enzymogenes suspension cultivated on nutrient broth using different concentrations of NTG (100, 150, and 200 µg/ml) for 20 and 40 minutes. The treated bacteria were cultivated on nutrient agar containing casein as a selective media. Primary and secondary screenings were performed by measuring the diameter of the casein hydrolysis zones in the isolated bacteria and the related supernatants, respectively. Finally, the unit of protease activity was quantified by Anson’s method of examining bacterial supernatants. Among the total of 30 isolated mutants, two mutants showed the highest level of extracellular proteolytic activity which showed 2.65 and 1.86 fold increments in contrast to the wild type, respectively. In general, the effect of mutagenesis by NTG can be emphasized to increase protease activity.

Production, characterization, and applications of bacterial pigments- a decade of review

Synthetic pigments have been employed universally for decades, resulting in environmental pollution and human health risks. So, it was critical to find out novel natural pigments, such as microbial pigments, that were safe and alternative to synthetic pigments. Bacterial pigments were getting the importance and attention of both researchers and industries for the mass production of various colored pigments. Bacterial pigments were not only used for industrial applications but also have several pharmacological activities like an antibiotic, antioxidant, anti-cancer properties. For the production of natural pigments, bacterial sources are cheap and have large economic potential when compared to plant sources. To make high-end goods, strain improvement, genetic engineering, fermentation conditions, simple extraction and characterization procedures are required. The importance of bacterial pigments is highlighted in this review, which covers their synthesis, characterization, and biological uses.

Efficient multiplex CRISPR/Cpf1 (Cas12a) genome editing system in Aspergillus aculeatus TBRC 277

CRISPR/Cas technology is a versatile tool for genome engineering in many organisms, including filamentous fungi. Cpf1 is a multi-domain protein of class 2 (type V) RNA-guided CRISPR/Cas endonuclease, and is an alternative platform with distinct features when compared to Cas9. However, application of this technology in filamentous fungi is limited. Here, we present a single CRISPR/Cpf1 plasmid system in Aspergillus aculeatus strain TBRC 277, an industrially relevant cell factory. We first evaluated the functionality of three Cpf1 orthologs from Acidaminococcus sp. BV3L6 (AsCpf1), Francisella tularensis subsp. novicida U112 (FnCpf1), and Lachnospiraceae bacterium (LbCpf1), in RNA-guided site-specific DNA cleavage at the pksP locus. FnCpf1 showed the highest editing efficiency (93%) among the three Cpf1s. It was further investigated for its ability to delete a 1.7kb and a 0.5kb from pksP and pyrG genes, respectively, using two protospacers targeting these gene loci in a single crRNA array. Lastly, simultaneous editing of three sites within TBRC 277 genome was performed using three guide sequences targeting these two genes as well as an additional gene, kusA, which resulted in combined editing efficiency of 40%. The editing of the NHEJ pathway by targeting kusA to generate a NHEJ-deficient strain of A. aculeatus TBRC 277 improved gene targeting efficiency and yielded more precise gene-editing than that of using wild-type strain. This promising genome-editing system can be used for strain improvement in industrial applications such as production of valuable bioproducts.

Reverse Remodeling Following Valve Replacement in Coexisting Aortic Stenosis and Transthyretin Cardiac Amyloidosis.

Dual pathology of severe aortic stenosis (AS) and transthyretin cardiac amyloidosis (ATTR) is increasingly recognized. Evolution of symptoms, biomarkers, and myocardial mechanics in AS-ATTR following valve replacement is unknown. We aimed to characterize reverse remodeling in AS-ATTR and compared with lone AS. Consecutive patients referred for transcatheter aortic valve replacement (TAVR) underwent ATTR screening by blinded 99mTc-DPD bone scintigraphy (Perugini Grade-0 negative, 1-3 increasingly positive) before intervention. ATTR was diagnosed by DPD and absence of monoclonal protein. Reverse remodeling was assessed by comprehensive evaluation before TAVR and at 1 year. One hundred twenty patients (81.8±6.3 years, 51.7% male, 95 lone AS, 25 AS-ATTR) with complete follow-up were studied. At 12 months (interquartile range, 7-17) after TAVR, both groups experienced significant symptomatic improvement by New York Heart Association functional class (both P<0.001). Yet, AS-ATTR remained more symptomatic (New York Heart Association ≥III: 36.0% versus 13.8; P=0.01) with higher residual NT-proBNP (N-terminal pro-brain natriuretic peptide) levels (P<0.001). Remodeling by echocardiography showed left ventricular mass regression only for lone AS (P=0.002) but not AS-ATTR (P=0.5). Global longitudinal strains improved similarly in both groups. Conversely, improvement of regional longitudinal strain showed a base-to-apex gradient in AS-ATTR, whereas all but apical segments improved in lone AS. This led to the development of an apical sparing pattern in AS-ATTR only after TAVR. Patterns of reverse remodeling differ from lone AS to AS-ATTR, with both groups experiencing symptomatic improvement by TAVR. After AS treatment, AS-ATTR transfers into a lone ATTR cardiomyopathy phenotype.

Organic selenium fortification in edible marine microalga Nannochloropsis oceanica CASA CC201 for food and feed applications

Selenium is an essential trace element that performs various biological functions (act as an antioxidant) in our body via a set of proteins called selenoproteins. Organic selenium sources include meat, fish, wheat, etc., which lose their selenium content to a significant extent while cooking and processing. New food sources fortified with organic Selenium are viable to meet selenium deficiency and associated health benefits. In the current study, an edible marine microalga Nannochloropsis oceanica CASA CC201 was developed for over 99 % organic selenium conversion by sequential strain improvement with Sodium selenite over several generations. Inductively Coupled Plasma Mass Spectrometry (ICP-MS) analysis of these cultures has shown that there was a higher accumulation of total intracellular selenium content in the treated samples (565.4 μg Se/g DW) as compared to control (0.108 μg Se/g DW). Organic Se enriched N. ocecanica CASA CC201can withstand as high as 1000 μM sodium selenite without compromising its growth pattern and biomass yield. Collectively, the current study proposes that, an edible marine microalga N. oceanica CASA CC201 biomass with high organic Se content can serve as Selenium fortified food/feed additives with excellent antioxidant properties.

Random Mutagenesis as a Promising Tool for Microalgal Strain Improvement towards Industrial Production.

Microalgae have become a promising novel and sustainable feedstock for meeting the rising demand for food and feed. However, microalgae-based products are currently hindered by high production costs. One major reason for this is that commonly cultivated wildtype strains do not possess the robustness and productivity required for successful industrial production. Several strain improvement technologies have been developed towards creating more stress tolerant and productive strains. While classical methods of forward genetics have been extensively used to determine gene function of randomly generated mutants, reverse genetics has been explored to generate specific mutations and target phenotypes. Site-directed mutagenesis can be accomplished by employing different gene editing tools, which enable the generation of tailor-made genotypes. Nevertheless, strategies promoting the selection of randomly generated mutants avoid the introduction of foreign genetic material. In this paper, we review different microalgal strain improvement approaches and their applications, with a primary focus on random mutagenesis. Current challenges hampering strain improvement, selection, and commercialization will be discussed. The combination of these approaches with high-throughput technologies, such as fluorescence-activated cell sorting, as tools to select the most promising mutants, will also be discussed.

Finerenone Reduces Renal RORγt γδ T Cells and Protects against Cardiorenal Damage

Introduction: Chronic activation of the mineralocorticoid receptor (MR) leads to pathological processes like inflammation and fibrosis during cardiorenal disease. Modulation of immunological processes in the heart or kidney may serve as a mechanistic and therapeutic interface in cardiorenal pathologies. In this study, we investigated anti-inflammatory/-fibrotic and immunological effects of the selective nonsteroidal MR antagonists finerenone (FIN) in the deoxycorticosterone acetate (DOCA)-salt model. Methods: Male C57BL6/J mice were uninephrectomized and received a DOCA pellet implantation (2.4 mg/day) plus 0.9% NaCl in drinking water (DOCA-salt) or received a sham operation and were orally treated with FIN (10 mg/kg/day) or vehicle in a preventive study design. Five weeks after the procedure, blood pressure (BP), urinary albumin/creatinine ratio (UACR), glomerular and tubulointerstitial damage, echocardiographic cardiac function, as well as cardiac/renal inflammatory cell content by FACS analysis were assessed. Results: BP was significantly reduced by FIN. FACS analysis revealed a notable immune response due to DOCA-salt exposure. Especially, infiltrating renal RORγt γδ-positive T cells were upregulated, which was significantly ameliorated by FIN treatment. This was accompanied by a significant reduction of UACR in FIN-treated mice. In the heart, FIN reduced DOCA-salt-induced cardiac hypertrophy, cardiac fibrosis and led to an improvement of the global longitudinal strain. Cardiac actions of FIN were not associated with a regulation of cardiac RORγt γδ-positive T cells. Discussion/Conclusion: The present study shows cardiac and renal protective effects of FIN in a DOCA-salt model. The cardiorenal protection was accompanied by a reduction of renal RORγt γδ T cells. The observed actions of FIN may provide a potential mechanism of its efficacy recently observed in clinical trials.

A comprehensive review on microalgal biomass production and processing for biodiesel production

Biodiesel is considered a promising alternative to conventional fuels in response to increasing global energy demands, and it also contributes to reduced environmental emissions. Microalgae is a potential resource for qualitative biodiesel production due to its high specific growth rate, ability to accumulate significant quantities of intracellular lipids, potential to utilize wastewater as a cultivation medium and less cropland requirement compared to conventional oil crops. However, the commercialization of microalgae biodiesel largely depends on the energy and cost-efficiency of the microalgae cultivation system. Moreover, optimization of cultivation systems and strain improvement in microalgae are promising strategies to enhance the growth rate and lipid productivities. This review provides a comprehensive overview of the various microalgae cultivation systems, focusing on the recent developments, including the genetic engineering perspectives in improving microalgae growth and lipid productivity, techno-economic analysis, and real case studies for microalgal biodiesel production. The review paper also summarizes the microalgae harvesting and lipid extraction processes, transesterification methodologies, quality of microalgae biodiesel, and environmental concerns associated with the large-scale production of microalgae biodiesel.

Overproduction of a thermo-stable halo-alkaline protease on agro-waste-based optimized medium through alternate combinatorial random mutagenesis of Stenotrophomonas acidaminiphila

A strain of Stenotrophomonas acidaminiphila, isolated from fermenting bean-processing wastewater, produced alkaline protease in pretreated cassava waste-stream, but with low yield. Strain improvement by alternate combinatorial random mutagenesis and bioprocess optimization using comparative statistical and neural network methods enhanced yield by 17.8-fold in mutant kGy-04-UV-25. Kinetics of production by selected mutant modeled by logistic and modified Gompertz functions revealed higher specific growth rate in mutant than in the parent strain, likewise volumetric and specific productivities. Purification by PEG/Na+ citrate aqueous two-phase system recovered 73.87% yield and 52.55-fold of protease. Its activity was stable at 5–35% NaCl, 45–75°C, and was significantly enhanced by 1–15 mM sodium dodecyl sulfate (SDS). The protease was inhibited by low concentrations of phenyl-methyl-sulfonyl fluoride but was activated by 1–5 mM Mn2+ suggesting a manganese-dependent serine‑protease. The 45.7 kDa thermo-halo-stable alkaline protease demonstrated keratinolytic and blood-stain removal potentials showing prospects in textile and detergent industries, respectively.

The Potential of Sequential Fermentations in Converting C1 Substrates to Higher-Value Products.

Today production of (bulk) chemicals and fuels almost exclusively relies on petroleum-based sources, which are connected to greenhouse gas release, fueling climate change. This increases the urgence to develop alternative bio-based technologies and processes. Gaseous and liquid C1 compounds are available at low cost and often occur as waste streams. Acetogenic bacteria can directly use C1 compounds like CO, CO2, formate or methanol anaerobically, converting them into acetate and ethanol for higher-value biotechnological products. However, these microorganisms possess strict energetic limitations, which in turn pose limitations to their potential for biotechnological applications. Moreover, efficient genetic tools for strain improvement are often missing. However, focusing on the metabolic abilities acetogens provide, they can prodigiously ease these technological disadvantages. Producing acetate and ethanol from C1 compounds can fuel via bio-based intermediates conversion into more energy-demanding, higher-value products, by deploying aerobic organisms that are able to grow with acetate/ethanol as carbon and energy source. Promising new approaches have become available combining these two fermentation steps in sequential approaches, either as separate fermentations or as integrated two-stage fermentation processes. This review aims at introducing, comparing, and evaluating the published approaches of sequential C1 fermentations, delivering a list of promising organisms for the individual fermentation steps and giving an overview of the existing broad spectrum of products based on acetate and ethanol. Understanding of these pioneering approaches allows collecting ideas for new products and may open avenues toward making full use of the technological potential of these concepts for establishment of a sustainable biotechnology.

Amino Acid Production and Role of Genetic Engineering Approaches to Strain Improvement for Enhanced Production: Review

Amino acids, both proteogenic and non-proteogenic amino acids, are produced at industrial scale for diverse applications. The industrial amino acids production methods include extraction, enzymatic, chemical synthesis and microbial fermentation methods. However, microbial fermentation method has been found to be economically feasible and convenient production scheme for production of L-forms of amino acid in natural and safe way. The fermentation method has also been augmented by improvement of producing microbial strains to meet rapidly growing need for amino acids. The advanced high throughput technique in genetic engineering enables development of efficient strains for amino acid overproduction. With this review the historical development of amino acid production, different amino acid production methods, and strain improvement techniques are well reviewed and the way forward are recommended. Keywords: Amino acids, fermentation, microorganisms, strain improvement DOI: 10.7176/JNSR/13-12-04 Publication date: June 30 th 2022

Improving Stress-Strain Behavior of Waste Aggregate Concrete Using Affordable Glass Fiber Reinforced Polymer (GFRP) Composites

Several studies have highlighted the potential of crushed brick aggregates in non-structural concrete. This is because crushed brick aggregates offer substandard mechanical properties in comparison to natural stone aggregates. Synthetic Fiber Reinforced Polymer (FRP) sheets have been known to overcome this issue. However, enormous costs associated with synthetic FRPs may limit their use in several low-budget applications. This study recognizes this issue and propose a cost-effective solution in the form of low-cost glass fiber (LC-GFRP) sheets. Two types of brick aggregates (i.e., solid-clay and hollow-clay brick aggregates) were used to fabricate concrete by replacing 50% of natural aggregates. Experimental results of 32 non-circular specimens were reported in this study. To overcome the substandard mechanical properties of recycled brick aggregate concrete (RBAC), specimens were strengthened with 2, 4, and 6 layers of LC-GFRP sheets. Noticeable improvements in ultimate compressive stress and corresponding strain were observed and were found to correlate positively with the number of LC-GFRP sheets. It was found that 4 and 6 layers of LC-GFRP sheets imparted significant axial ductility irrespective of the brick aggregate type and inherent concrete strength. Several existing stress-strain models for confined concrete were considered to predict ultimate confined compressive stress and corresponding strain. Accuracy of existing models was assessed by mean of the ratio of analytical to experimental values and associated standard deviations. For ultimate stress predictions, the lowest mean value of the ratio of analytical to experimental ultimate compressive stress was 1.07 with a standard deviation of 0.10. However, none of the considered models was able to provide good estimates of ultimate strains.

Elderly Caregiving Quality Improvement: A Pilot Study of the Burdens of Vietnamese Caregivers in Taiwan.

Taiwan is expected to reach super-aged status by 2026, leading to an increased demand for elderly caregiving services. Low local unemployment and a dwindling working-age population mean the island’s care system relies heavily on female foreign domestic workers (FDWs) from Southeast Asian neighbors such as Vietnam to satisfy labor shortages. Although suggested by anecdotal evidence, limited research has been conducted on the link between the shortfall in FDW qualifications, training, preparedness, and expertise and their employment stressors. Therefore, this study aims to assist FDWs by evaluating their stressors and helping them better understand health care delivery by (1) administering the Modified Caregiver Strain Index (MCSI) revised 2003 questionnaire, (2) performing semi-structured in-depth one-on-one interviews, (3) classifying interview results according to thematic analysis, and (4) using these themes to devise and deliver a 12-week multilingual health education teach-back program. Our results indicate that Vietnamese FDWs face specific challenges, including language barriers, homesickness, intensive physical and psychological work demands, stress adaptation, and occupational exposures. Despite yielding no significant improvements in caregiving strain, our intervention, conducted at the height of the COVID-19 pandemic, pinpoints and classifies areas of grave concern and proposes recommendations that can assist long-term care (LTC) stakeholders in understanding and overcoming their respective challenges, thereby improving the quality of elderly care.

Enhancing proteolytic activity of Lysobacter enzymogenes using cold atmospheric plasma.

Cold atmospheric plasma (CAP) is being used recently as a modern technique for microbial random mutagenesis. In the present study, CAP was used to induce mutagenesis in L. enzymogenes which is the bacteria known for producing proteolytic enzymes especially lysyl endopeptidase (Lys C). Enhanced proteolytic activity was the main criteria to select mutant strains. Therefore, the cell suspension of L. enzymogenes strain (ATCC 29487), was exposed to CAP for 30, 45, 90, and 150s. The proteolytic activity of mutant strains was screened initially by radial caseinolytic assay and then by Ansons method in different phases of bacterial growth in the selected mutants. The purification process of Lysyl endopeptidase as the target enzyme was optimized and for enlightening molecular aspect of CAP mutagenesis, the sequences of the upstream and coding regions of lys C gene from 10 selected mutant strains were determined. The bacterial survival assessment showed that the more CAP treatment time, the less survival rate, however, in all exposure times, a number of survived mutants showed enhanced proteolytic activity. Among 38 out of 100 examined mutants which showed higher proteolytic activity than that of wild type, the M1-30s mutant exhibited the highest increment to 1.94 fold. The SDS-PAGE analysis showed expected size of purified Lys C from M1-30s.The Lys C gene from M14-150s mutant strain (1.4-fold increment) harbored three point mutations which can be effective in enhancing protease activity. In conclusion, the results highlighted the role of CAP for strain improvement process to obtain industrial strains.

P236 LONG TERM FOLLOW UP WITH A DRASTIC REDUCTION OF HOSPITALIZATION IN HEART FAILURE PATIENT WITH MULTIPLE AETIOLOGIES TREATED WITH CARDIAC CONTRACTILITY MODULATION THERAPY: A CASE REPORT

Abstract Aims Cardiac contractility modulation (CCM) shows to be a concrete therapeutic option in patients with symptomatic HF despite optimal medical therapy (OMT), with Left Ventricular Ejection Fraction (LVEF) between 25% and 45%, with narrow QRS complex (&amp;lt;130ms) for reduction of hospitalization and improvement Quality of life (QoL). This case aims to explore the effectiveness of CCM therapy in a patient affected by concomitant ischemic cardiomyopathy and cardiac amyloidosis (CA) Methods A 42–year–old man with Chronic HF secondary to both post–ischemic due to spontaneous coronary artery dissection (SCAD) and post alcoholic dilated cardiomyopathy was hospitalized at our department in February 2020 due to worsening HF (3rd HF hospitalization in the same year). The patient was a NYHA class III, with chronic kidney failure, a narrow QRS complex (100 ms) and a LVEF of 27% with familiar history of sudden death, already implanted with ICD. The patient resulted untreatable with Sacubitril/Valsartan, as it elicited strong hypotension. During current hospitalization the BNP value was 942,60 pg/ml, and the Quality of Life (QoL) evaluated from Minnesota Living with Heart Failure Questionnaire (MLHFQ) score was 72 points. Moreover, the patient underwent umbilical biopsy that confirmed the presence of amyloidosis. Thus, the CCM therapy device (Optimizer® Smart, Impulse Dynamics) was implanted to try to reduce HF symptoms and hospitalizations (Fig 1 A/B). Results The patient significantly improved as early as the first period after implantation. The 10–month in–office FU performed on December 2020 revealed in addition to the absence of new HF hospitalizations, a significant improvement in QoL and HF–symptoms, with a MLWHFQ score of 42, an enhancement to NHYA class II and even a slight decrease of BNP of 767 pg/ml. The echo exam revealed no significant changes in the EF, with an improvement of global longitudinal strain and no worsening of other hemodynamic parameters. A further FU performed in June 2021 showed continuous improvement of QoL (MLWHFQ = 25) e no HF hospitalizations. Conclusions In this patient affected by multiple cardiomyopathies, including CA, CCM therapy proved to improve its QoL with no HF hospital admissions since the implantation. The absence of significant echocardiographic worsening is a positive aspect, considering the patient’s status, the concomitant aetiologies, and the presence of amyloidosis, given its progressive and infiltrative nature.

Microfluidic screening and genomic mutation identification for enhancing cellulase production in Pichia pastoris

BackgroundPichia pastoris is a widely used host organism for heterologous production of industrial proteins, such as cellulases. Although great progress has been achieved in improving protein expression in P. pastoris, the potential of the P. pastoris expression system has not been fully explored due to unknown genomic impact factors. Recently, whole-cell directed evolution, employing iterative rounds of genome-wide diversity generation and high-throughput screening (HTS), has been considered to be a promising strategy in strain improvement at the genome level.ResultsIn this study, whole-cell directed evolution of P. pastoris, employing atmospheric and room temperature plasma (ARTP) mutagenesis and droplet-based microfluidic HTS, was developed to improve heterogenous cellulase production. The droplet-based microfluidic platform based on a cellulase-catalyzed reaction of releasing fluorescence was established to be suitable for methanol-grown P. pastoris. The validation experiment showed a positive sorting efficiency of 94.4% at a sorting rate of 300 droplets per second. After five rounds of iterative ARTP mutagenesis and microfluidic screening, the best mutant strain was obtained and exhibited the cellulase activity of 11,110 ± 523 U/mL, an approximately twofold increase compared to the starting strain. Whole-genome resequencing analysis further uncovered three accumulated genomic alterations in coding region. The effects of point mutations and mutant genes on cellulase production were verified using reconstruction of point mutations and gene deletions. Intriguingly, the point mutation Rsc1G22V was observed in all the top-performing producers selected from each round, and gene deletion analysis confirmed that Rsc1, a component of the RSC chromatin remodeling complex, might play an important role in cellulase production.ConclusionsWe established a droplet-based microfluidic HTS system, thereby facilitating whole-cell directed evolution of P. pastoris for enhancing cellulase production, and meanwhile identified genomic alterations by whole-genome resequencing and genetic validation. Our approaches and findings would provide guides to accelerate whole-cell directed evolution of host strains and enzymes of high industrial interest.

Unlocking the functional potential of polyploid yeasts

Breeding and domestication have generated widely exploited crops, animals and microbes. However, many Saccharomyces cerevisiae industrial strains have complex polyploid genomes and are sterile, preventing genetic improvement strategies based on breeding. Here, we present a strain improvement approach based on the budding yeasts’ property to promote genetic recombination when meiosis is interrupted and cells return-to-mitotic-growth (RTG). We demonstrate that two unrelated sterile industrial strains with complex triploid and tetraploid genomes are RTG-competent and develop a visual screening for easy and high-throughput identification of recombined RTG clones based on colony phenotypes. Sequencing of the evolved clones reveal unprecedented levels of RTG-induced genome-wide recombination. We generate and extensively phenotype a RTG library and identify clones with superior biotechnological traits. Thus, we propose the RTG-framework as a fully non-GMO workflow to rapidly improve industrial yeasts that can be easily brought to the market.

Carbon Fiber Reinforced Polymer (CFRP) Composite Spirals as Confinement Reinforcement for Concrete under Axial Compression

The use of fiber composites in structural strengthening applications has been implemented for many years. Among others, this type of material has been employed as means to confine concrete and achieve superior material behavior. Confinement can be achieved either by wrapping the concrete with FRP fabrics or by embedding FRP stirrups or spiral (helical) reinforcement in concrete. Although FRP spirals have been used as reinforcement in concrete members, this was primarily in the form of FRP bars used as shear reinforcement in RC members under flexure. Some studies have been conducted using FRP spiral strips that were bonded on the outside face of cylindrical specimens and their confinement effect on concrete under axial compression was evaluated. This study deals with the experimental testing of concrete cylindrical specimens that utilize embedded carbon FRP (CFRP) composite spiral reinforcement, a unique application with limited investigation as to its effect on concrete confinement. Standard (300 mm x 150 mm) concrete cylinders were cast both with and without the embedded CFRP spiral. The cylinders were tested in axial compression to determine the effect of the CFRP spiral on their strength and ultimate deformation. This study contributes to the field of concrete confinement and in an area that is not well investigated especially when the FRP spiral is embedded into concrete. Experimental results indicate very significant improvements in ultimate strength and strain with more the enhancement reaching 116% for the strength and 147% for the strain. Comparing energy absorption capacity of the CFRP spirally confined concrete with the unconfined concrete, the improvement is even higher and reaches 320%. Therefore, the use of embedded CFRP spiral to confine concrete is effective and provides significant confinement and enhances concrete properties.

Review of the structures and functions of algal photoreceptors to optimize bioproduct production with novel bioreactor designs for strain improvement.

Microalgae are important renewable feedstock to produce biodiesel and high-value chemicals. Different wavelengths of light influence the growth and metabolic activities of algae. Recent research has identified the light-sensing proteins called photoreceptors that respond to blue or red light. Structural elucidations of algal photoreceptors have gained momentum over recent years. These include channelrhodopsins, PHOT proteins, animal-like cryptochromes, and blue-light sensors utilizing flavin-adenine dinucleotide proteins. Pulsing light has also been investigated as a means to optimize energy inputs into bioreactors. This study summarizes the current structural and functional basis of photoreceptor modulation to optimize the growth, production of carotenoids and other high-value metabolites from microalgae. The review also encompasses novel photobioreactor designs that implement different light regimes including light wavelengths and time to optimize algal growth and desired metabolite profiles for high-value products.