Optimal conditions of algal breeding using neutral beam and applying it to breed Euglena gracilis strains with improved lipid accumulation.

Microalgae are considered to be more useful and effective to use in biomass production than other photosynthesis organisms. However, microalgae need to be altered to acquire more desirable traits for the relevant purpose. Although neutron radiation is known to induce DNA mutations, there have been few studies on its application to microalgae, and the optimal relationship between irradiation intensity and mutation occurrence has not been established. In this study, using the unicellular red alga Cyanidioschyzon merolae as a model, we analyzed the relationship between the absorbed dose of two types of neutrons, high-energy (above 1 MeV) and thermal (around 25 meV) neutrons, and mutation occurrence while monitoring mutations in URA5.3 gene encoding UMP synthase. As a result, the highest mutational occurrence was observed when the cells were irradiated with 20 Gy of high-energy neutrons and 13 Gy of thermal neutrons. Using these optimal neutron irradiation conditions, we next attempted to improve the lipid accumulation of Euglena gracilis, which is a candidate strain for biofuel feedstock production. As a result, we obtained several strains with a maximum 1.3-fold increase in lipid accumulation compared with the wild-type. These results indicate that microalgae breeding by neutron irradiation is effective.

Arterial desaturation rate does not influence self-selected knee extension force but alters ventilatory response to progressive hypoxia: A pilot study.

The absolute magnitude and rate of arterial desaturation each independently impair whole-body aerobic exercise. This study examined potential mechanisms underlying the rate-dependent relationship. Utilizing an exercise protocol involving unilateral, intermittent, isometric knee extensions (UIIKE), we provided sufficient reperfusion time between contractions to reduce the accumulation of intramuscular metabolic by-products that typically stimulate muscle afferents. The objective was to create a milieu conducive to accentuating any influence of arterial desaturation rate on muscular fatigue. Eight participants completed four UIIKE sessions, performing one 3 s contraction every 30s at a perceived intensity of 50% MVC for 25 min. Participants voluntarily adjusted their force generation to maintain perceptual effort at 50% MVC without feedback. Reductions in inspired oxygen fraction (FI O2 ) decreased arterial saturation from >98% to 70% with varying rates in three trials: FAST (5.3 ± 1.3 min), MED (11.8 ± 2.7 min), and SLOW (19.9 ± 3.7 min). FI O2 remained at 0.21 during the control trial. Force generation and muscle activation remained at baseline levels throughout UIIKE trials, unaffected by the magnitude or rate of desaturation. Minute ventilation increased with hypoxia (p < 0.05), and faster desaturation rates magnified this response. These findings demonstrate that arterial desaturation magnitude and rate independently affect ventilation, but do not influence fatigue development during UIIKE.

Development of engineered Candida tropicalis strain for efficient corncob-based xylitol-ethanol biorefinery.

BACKGROUND: Xylitol has a wide range of applications in the pharmaceuticals, cosmetic, food and beverage industry. Microbial xylitol production reduces the risk of contamination and is considered as environment friendly and sustainable compared to the chemical method. In this study, random mutagenesis and genetic engineering approaches were employed to develop Candida tropicalis strains with reduced xylitol dehydrogenase (XDH) activity to eliminate co-substrate requirement for corn cob-based xylitol-ethanol biorefinery. RESULTS: The results suggest that when pure xylose (10% w/v) was fermented in bioreactor, the Ethyl methane sulfonate (EMS) mutated strain (C. tropicalis K2M) showed 9.2% and XYL2 heterozygous (XYL2/xyl2Δ::FRT) strain (C. tropicalis K21D) showed 16% improvement in xylitol production compared to parental strain (C. tropicalis K2). Furthermore, 1.5-fold improvement (88.62 g/L to 132 g/L) in xylitol production was achieved by C. tropicalis K21D after Response Surface Methodology (RSM) and one factor at a time (OFAT) applied for media component optimization. Finally, corncob hydrolysate was tested for xylitol production in biorefinery mode, which leads to the production of 32.6 g/L xylitol from hemicellulosic fraction, 32.0 g/L ethanol from cellulosic fraction and 13.0 g/L animal feed. CONCLUSIONS: This work, for the first time, illustrates the potential of C. tropicalis K21D as a microbial cell factory for efficient production of xylitol and ethanol via an integrated biorefinery framework by utilising lignocellulosic biomass with minimum waste generation.

Can Genome Sequencing Coupled to Flux Balance Analyses Offer Precision Guidance for Industrial Strain Development? The Lessons from Carbon Trafficking in Corynebacterium glutamicum ATCC 21573.

Systems biology tools offer new prospects for industrial strain selection. For bacteria that are significant for industrial applications, whole-genome sequencing coupled to flux balance analysis (FBA) can help unpack the complex relationships between genome mutations and carbon trafficking. This work investigates the l-tyrosine (l-Tyr) overproducing model system Corynebacterium glutamicum ATCC 21573 with an eye to more rational and precision strain development. Using genome-wide mutational analysis of C. glutamicum, we identified 27,611 single nucleotide polymorphisms and 479 insertion/deletion mutations. Mutations in the carbon uptake machinery have led to phosphotransferase system-independent routes as corroborated with FBA. Mutations within the central carbon metabolism of C. glutamicum impaired the carbon flux, as evidenced by the lower growth rate. The entry to and flow through the tricarboxylic acid cycle was affected by mutations in pyruvate and α-ketoglutarate dehydrogenase complexes, citrate synthase, and isocitrate dehydrogenase. FBA indicated that the estimated flux through the shikimate pathway became larger as the l-Tyr production rate increased. In addition, protocatechuate export was probabilistically impossible, which could have contributed to the l-Tyr accumulation. Interestingly, aroG and cg0975, which have received previous attention for aromatic amino acid overproduction, were not mutated. From the branch point molecule, prephenate, the change in the promoter region of pheA could be an influential contributor. In summary, we suggest that genome sequencing coupled with FBA is well poised to offer rational guidance for industrial strain development, as evidenced by these findings on carbon trafficking in C. glutamicum ATCC 21573.

Designing gene manipulation schedules for high throughput parallel construction of objective strains.

Recent advances in biofoundries have enabled the construction of a large quantity of strains in parallel, accelerating the design-build-test-learn (DBTL) cycles for strain development. However, the construction of a large number of strains by iterative gene manipulation is still time-consuming and costly, posing a challenge for the development of commercial strains. Common gene manipulations among different objective strains open up the possibility of reducing cost and time for strain construction in biofoundries by optimizing genetic manipulation schedules. A method is introduced consisting of two complementary algorithms for designing optimal parent-children manipulation schedules for strain construction: greedy search of common ancestor strains (GSCAS) and minimizing total manipulations (MTM). By reusing common ancestor strains, the number of strains to be constructed can be effectively reduced, resulting in a tree-like structure of descendants instead of linear lineages for each strain. The GSCAS algorithm can quickly find common ancestor strains and clusters them together based on their genetic makeup, and the MTM algorithm subsequently minimize the genetic manipulations required, resulting in a further reduction in the total number of genetic manipulations. The effectiveness of our method is demonstrated through a case study of 94 target strains, where GSCAS reduces an average of 36% of the total gene manipulations, and MTM reduces an additional 10%. The performance of both algorithms is robust among case studies with different average occurrences of gene manipulations across objective strains. Our method potentially improves cost efficiency and accelerate the development of commercial strains significantly. The implementation of the methods can be freely accessed via https://gscas-mtm.biodesign.ac.cn/.

Implant-Supported Overdentures: Current Status and Preclinical Testing of a Novel Attachment System.

Numerous attachment systems exist for implant-supported overdentures, with each having specific limitations in terms of retention, cost, wear, maintenance and cleanability. A retrospective analysis of patients restored with implant-supported overdentures using bars, telescopic crowns and Locator-type attachments was performed and the patients were interviewed. An in vitro strain gauge study compared telescopic crowns, Locator-type attachments and a novel flexible attachment system employing a shape memory alloy (NiTi) with respect to peri-implant strain development during insertion, loading and removal of an overdenture. A significantly lower number of attachment-related complications was observed in bars as compared to telescopic crowns (p = 0.00007) and Locator-type attachments (p = 0.00000), respectively. Greater overall patient satisfaction was noted in bar-retained restorations while Locator-type attachments led to lower levels of satisfaction regarding prosthesis retention. In vitro, telescopic crowns caused maximum strain development during prosthesis insertion and loading, while during removal this was observed in Locators with white retentive inserts. NiTi attachments caused significantly lower strain development during insertion as compared to telescopic crowns (p = 0.027). During loading, NiTi attachments caused significantly lower strain development than Locators with blue retentive inserts (p = 0.039). During removal, NiTi attachments caused significantly less strain development as compared to Locators with white retentive inserts (p = 0.027). Positional discrepancies between male and female attachment parts affected the retention and reaction force between both components, which may be minimized by using the novel NiTi attachment system. This may be beneficial in terms of component wear and implant loading.

Strain Development in Microalgal Biotechnology-Random Mutagenesis Techniques.

Microalgal biomass and metabolites can be used as a renewable source of nutrition, pharmaceuticals and energy to maintain or improve the quality of human life. Microalgae's high volumetric productivity and low impact on the environment make them a promising raw material in terms of both ecology and economics. To optimize biotechnological processes with microalgae, improving the productivity and robustness of the cell factories is a major step towards economically viable bioprocesses. This review provides an overview of random mutagenesis techniques that are applied to microalgal cell factories, with a particular focus on physical and chemical mutagens, mutagenesis conditions and mutant characteristics.

Compressive Fatigue Investigation on High-Strength and Ultra-High-Strength Concrete within the SPP 2020.

The influence of the compressive strength of concrete on fatigue resistance has not been investigated thoroughly and contradictory results can be found in the literature. To date, the focus of concrete fatigue research has been on the determination of the numbers of cycles to failure. Concerning the fatigue behaviour of high-strength concrete (HPC) and, especially, ultra-high-strength concrete (UHPC), which is described by damage indicators such as strain and stiffness development, little knowledge is available, as well as with respect to the underlying damage mechanisms. This lack of knowledge has led to uncertainties concerning the treatment of high-strength and ultra-high-strength concretes in the fatigue design rules. This paper aims to decrease the lack of knowledge concerning the fatigue behaviour of concrete compositions characterised by a very high strength. Within the priority programme SPP 2020, one HPC and one UHPC subjected to monotonically increasing and cyclic loading were investigated comparatively in terms of their numbers of cycles to failure, as well as the damage indicators strain and stiffness. The results show that the UHPC reaches a higher stiffness and a higher ultimate strain and strength than the HPC. The fatigue investigations reveal that the UHPC can resist a higher number of cycles to failure than the HPC and the damage indicators show an improved fatigue behaviour of the UHPC compared to the HPC.

MICROBIAL isoprene production: an overview.

Isoprene, a volatile C5 hydrocarbon, is a precursor of synthetic rubber and an important building block for a variety of natural products, solely being produced by petrochemical routes. To mitigate the ever-increasing contribution of petrochemical industry to global warming through significant carbon (CO2) evolution, bio-based process for isoprene production using microbial cell factories have been explored. Highly efficient fermentation-based processes have been studied for little over a decade now with extensive research on the rational strain development for creating robust strains for commercial isoprene production. Most of these studies involved sugars as feedstocks and using naturally occurring isoprene pathways viz., mevalonate and methyl erythritol pathway in E. coli. Recent advances, driven by efforts in reducing environmental pollution, have focused on utilization of inorganic CO2 by cyanobacteria or syngas from waste gases by acetogens for isoprene production. This review endeavors to capture the latest relevant progress made in rational strain development, metabolic engineering and synthetic biology strategies used, challenges in fermentation process development at lab and commercial scale production of isoprene along with a future perspective pertaining to this area of research.

Advances and prospects in metabolic engineering of Escherichia coli for L-tryptophan production.

As an important raw material for pharmaceutical, food and feed industry, highly efficient production of L-tryptophan by Escherichia coli has attracted a considerable attention. However, there are complicated and multiple layers of regulation networks in L-tryptophan biosynthetic pathway and thus have difficulty to rewrite the biosynthetic pathway for producing L-tryptophan with high efficiency in E. coli. This review summarizes the biosynthetic pathway of L-tryptophan and highlights the main regulatory mechanisms in E. coli. In addition, we discussed the latest metabolic engineering strategies achieved in E. coli to reconstruct the L-tryptophan biosynthetic pathway. Moreover, we also review a few strategies that can be used in E. coli to improve robustness and streamline of L-tryptophan high-producing strains. Lastly, we also propose the potential strategies to further increase L-tryptophan production by systematic metabolic engineering and synthetic biology techniques.

Exploiting unconventional prokaryotic hosts for industrial biotechnology.

Developing cost-efficient biotechnological processes is a major challenge in replacing fossil-based industrial production processes. The remarkable progress in genetic engineering ensures efficient and fast tailoring of microbial metabolism for a wide range of bioconversions. However, improving intrinsic properties such as tolerance, handling, growth, and substrate consumption rates is still challenging. At the same time, synthetic biology tools are becoming easier applicable and transferable to nonmodel organisms. These trends have resulted in the exploitation of new and unconventional microbial systems with sophisticated properties, which render them promising hosts for the bio-based industry. Here, we highlight the metabolic and cellular capabilities of representative prokaryotic newcomers and discuss the potential and drawbacks of these hosts for industrial application.

Efficient breeding of industrial brewing yeast strains using CRISPR/Cas9-aided mating-type switching.

Yeast breeding is a powerful tool for developing and improving brewing yeast in a number of industry-relevant respects. However, breeding of industrial brewing yeast can be challenging, as strains are typically sterile and have large complex genomes. To facilitate breeding, we used the CRISPR/Cas9 system to generate double-stranded breaks in the MAT locus, generating transformants with a single specified mating type. The single mating type remained stable even after loss of the Cas9 plasmid, despite the strains being homothallic, and these strains could be readily mated with other brewing yeast transformants of opposite mating type. As a proof of concept, we applied this technology to generate yeast hybrids with an aim to increase ß-lyase activity for fermentation of beer with enhanced hop flavour. First, a genetic and phenotypic pre-screening of 38 strains was carried out in order to identify potential parent strains with high ß-lyase activity. Mating-competent transformants of eight parent strains were generated, and these were used to generate over 60 hybrids that were screened for ß-lyase activity. Selected phenolic off-flavour positive (POF +) hybrids were further sporulated to generate meiotic segregants with high ß-lyase activity, efficient wort fermentation, and lack of POF, all traits that are desirable in strains for the fermentation of modern hop-forward beers. Our study demonstrates the power of combining the CRISPR/Cas9 system with classic yeast breeding to facilitate development and diversification of brewing yeast. KEY POINTS: • CRISPR/Cas9-based mating-type switching was applied to industrial yeast strains. • Transformed strains could be readily mated to form intraspecific hybrids. • Hybrids exhibited heterosis for a number of brewing-relevant traits.

Fatigue-Induced Damage in High-Strength Concrete Microstructure.

A high-strength concrete subjected to compressive fatigue loading with two maximum stress levels was investigated and the behaviour was evaluated using the macroscopic damage indicators, strain and acoustic emission hits (AE-hits), combined with microstructural analyses utilising light microscopy and scanning electron microscopy (SEM). A clustering technique using Gaussian mixture modelling combined with a posterior probability of 0.80 was firstly applied to the AE-hits caused by compressive fatigue loading, leading to two clusters depending on the maximum stress level. Only a few cracks were visible in the microstructure using light microscopy and SEM, even in phase III of the strain development, which is shortly before failure. However, bluish impregnated areas in the mortar matrix of higher porosity or defects, changing due to the fatigue loading, were analysed. Indications were found that the fatigue damage process is continuously ongoing on a micro- or sub-microscale throughout the mortar matrix, which is difficult to observe on a mesoscale by imaging. Furthermore, the results indicate that two different damage mechanisms take place, which are pronounced depending on the maximum stress level. This might be due to diffuse and widespread compressive damage and localised tensile damage, as the findings documented in the literature suggest.

Investigation of the Influence of Moisture Content on Fatigue Behaviour of HPC by Using DMA and XRCT.

High-performance concrete (HPC) is a topic of current research and construction projects, due to its outstanding compressive strength and durability. In particular, its behaviour under high-cycle fatigue loading is the focus of current investigations, to further pave the way to highly challenging long-lasting constructions; e.g., bridges or offshore buildings. In order to investigate the behaviour of HPC with different moisture contents in more detail, a mixture of silica sand and basalt aggregate with a maximum grain size of 8 mm was investigated with three different moisture contents. For this purpose, cyclic compressive fatigue tests at a loading frequency of 10 Hz and different maximum stress levels were performed. The main focus was the moisture influence on the number of cycles to failure and the development of concrete temperature and strain. In a further step, only the mortar matrix was investigated. For this purpose, the mixture was produced without basalt, and the moisture influence was investigated on smaller-sized test specimens using dynamic mechanical analysis (DMA) and X-ray computed tomography (XRCT). It was shown that the moisture content of HPC had a significant influence on the fatigue damage behaviour due to the number of cycles to failure decreasing significantly with increased moisture. In addition, there was also an influence on the temperature development, as well as on the strain development. It was shown that increasing moisture content was associated with an increase in strain development. XRCT scans, in the course of the damage phases, showed an increase in internal cracks, and made their size visible. With the help of DMA as a new research method in the field of concrete research, we were also able to measure damage development related to a decrease in sample stiffness. Both methods, XRCT and DMA, can be listed as nondestructive methods, and thus can complement the known destructive test methods, such as light microscopy.

Agrobacterium strains and strain improvement: Present and outlook.

Almost 40 years ago the first transgenic plant was generated through Agrobacterium tumefaciens-mediated transformation, which, until now, remains the method of choice for gene delivery into plants. Ever since, optimized Agrobacterium strains have been developed with additional (genetic) modifications that were mostly aimed at enhancing the transformation efficiency, although an optimized strain also exists that reduces unwanted plasmid recombination. As a result, a collection of very useful strains has been created to transform a wide variety of plant species, but has also led to a confusing Agrobacterium strain nomenclature. The latter is often misleading for choosing the best-suited strain for one's transformation purposes. To overcome this issue, we provide a complete overview of the strain classification. We also indicate different strain modifications and their purposes, as well as the obtained results with regard to the transformation process sensu largo. Furthermore, we propose additional improvements of the Agrobacterium-mediated transformation process and consider several worthwhile modifications, for instance, by circumventing a defense response in planta. In this regard, we will discuss pattern-triggered immunity, pathogen-associated molecular pattern detection, hormone homeostasis and signaling, and reactive oxygen species in relationship to Agrobacterium transformation. We will also explore alterations that increase agrobacterial transformation efficiency, reduce plasmid recombination, and improve biocontainment. Finally, we recommend the use of a modular system to best utilize the available knowledge for successful plant transformation.

Genome shuffling improves pigment and other bioactive compound production in Monascus purpureus.

Monascus purpureus, a pigment-producing ascomycetous fungus, has been traditionally used for red rice preparation using solid-state fermentation. The objective of this study was to develop an improved pigment-producing strain of M. purpureus MTCC 1090 through genome shuffling followed by detailed analytical estimations of pigments and other bioactive compounds produced by the fusant. Protoplast formation was optimum with 12 h-old mycelia incubated at 30 °C with cellulase, lyticase, and chitinase (40:1:1) for 5 h. Four UV-induced mutants that produced 13.1-39.5% higher amount of yellow, orange, and red pigments in fermented low-grade (cheap) broken rice were used as parents for genome shuffling. After the first round of fusion, four fusants with 35.9-60.52% higher pigment production capabilities were fused again, and finally the fusant F2-19 with distinct culture characteristic was selected under multi-selection pressure. It consistently produced 67%, 70%, and 76% higher content of yellow, orange, and red pigments respectively as compared to the wild-type. High-performance liquid chromatography (HPLC) analysis also reveals clear variation in pigment productions between wild-type and the fusant. Furthermore, HPLC analysis of F2-19 fermented rice extract confirms the production of 186 ± 8.71 and 3810 ± 29.81 mg/kg mevinolin and gamma-aminobutyric acid respectively. Citrinin was not detected. F2-19 fermented rice also has high antioxidant activity (7.92 ± 0.32 mg/g trolox equivalent), with good amount of phenolics (18.0 ± 0.95 mg/g gallic acid equivalent) and flavonoids (2.7 ± 0.26 mg/g quercetin equivalent). Thus, genome shuffling was successfully implemented on M. purpureus for the first time to develop a citrinin-free, better-performing fusant that holds future biotechnological potential. KEY POINTS: • Genome shuffling was performed by recursive protoplast fusion in Monascus purpureus. • The selected fusant, F2-19, was used in solid-state fermentation using low-grade rice. • It produced 67-76% higher content of yellow, orange, and red pigments than the wild-type. • HPLC detected 186 mg/kg mevinolin and 3810 mg/kg γ-aminobutyric acid, but no citrinin. • F2-19 shows high antioxidant activity with good amount of phenolics and flavonoids. Graphical abstract.

Current Status and Applications of Adaptive Laboratory Evolution in Industrial Microorganisms.

Adaptive laboratory evolution (ALE) is an evolutionary engineering approach in artificial conditions that improves organisms through the imitation of natural evolution. Due to the development of multi-level omics technologies in recent decades, ALE can be performed for various purposes at the laboratory level. This review delineates the basics of the experimental design of ALE based on several ALE studies of industrial microbial strains and updates current strategies combined with progressed metabolic engineering, in silico modeling and automation to maximize the evolution efficiency. Moreover, the review sheds light on the applicability of ALE as a strain development approach that complies with non-recombinant preferences in various food industries. Overall, recent progress in the utilization of ALE for strain development leading to successful industrialization is discussed.

Novel mutagenesis and screening technologies for food microorganisms: advances and prospects.

Microorganisms are indispensable in the food industry, but wild-type strains hardly meet the current industrial demands due to several undesirable traits. Therefore, microbial strain improvement offers a critical solution to enhance the food industry. Traditional techniques for food microbial improvement, such as the use of chemical mutagens and manual isolation/purification, are inefficient, time-consuming, and laborious, restricting further progress in the area of food fermentation. In this review, the applications of novel mutagenesis and screening technologies used for the improvement of food microbes were summarized, including random mutagenesis based on physical irradiation, microbial screening facilitated by a microtiter plate, fluorescence-activated cell or droplet sorting, and microscaled fermentation in a microtiter plate or microbioreactor. In comparison with conventional methods, these new tools have the potential in accelerating microbial strain improvement and their combined applications could create a new trend for strain development. However, several problems that could affect its potential application may include the following: the lack of specific mutagenesis devices and biosensing systems, the insufficient improvement of the mixed culture system, the low efficiency when using filamentous fungi and flocculating bacteria, and the insufficient safety assessment on harnessing genome-editing technology. Therefore, future works on strain improvement remain challenging for the food industry.

Improving lipid production by strain development in microalgae: Strategies, challenges and perspectives.

Over the past decade, the number of original articles and reviews presenting microalgae as a promising feedstock for biodiesel has increased tremendously. Many improvements of microalgae have been achieved through selection and strain development for industrial applications. However, the large-scale production of lipids for commercialization is not yet realistic because the production is still much more expensive than that of agricultural products. This review summarizes recent research on the induction of lipid biosynthesis in microalgae and the various strategies of genetic and metabolic engineering for enhancing lipid production. Strain engineering targets are proposed based on these strategies. To address current limitations of strain engineering for lipid production, this review provides insights on recent engineering strategies based on molecular tools and methods, and also discusses further perspectives.

Evaluation of surfactin synthesis in a genome reduced Bacillus subtilis strain.

Strain engineering is often a method of choice towards increasing the yields of the biosurfactant surfactin which is naturally synthesized by many Bacillus spp., most notably Bacillus subtilis. In the current study, a genome reduced B. subtilis 168 strain lacking 10% of the genome was established and tested for its suitability to synthesize surfactin under aerobic and anaerobic conditions at 25 °C, 30 °C, 37 °C and 40 °C. This genome reduced strain was named IIG-Bs20-5-1 and lacks, amongst others, genes synthesizing the lipopeptide plipastatin, the antibiotic bacilysin, toxins and prophages, as well as genes involved in sporulation. Amongst all temperatures tested, 37 °C was overall superior. In comparison to the reference strain JABs24, a surfactin synthesizing variant of B. subtilis 168, strain IIG-Bs20-5-1 was both aerobically and anaerobically superior with respect to specific growth rates µ and yields YX/S. However, in terms of surfactin production, strain JABs24 reached higher absolute concentrations with up to 1147.03 mg/L and 296.37 mg/L under aerobic and anaerobic conditions, respectively. Concomitant, strain JABs24 reached higher YP/S and YP/X. Here, an outstanding YP/X of 1.541 g/g was obtained under anaerobic conditions at 37 °C. The current study indicates that the employed genome reduced strain IIG-Bs20-5-1 has several advantages over the strain JABs24 such as better conversion from glucose into biomass and higher growth rates. However, regarding surfactin synthesis and yields, the strain was overall inferior at the investigated temperatures and oxygen conditions. Further studies addressing process development and strain engineering should be performed combining the current observed advantages of the genome reduced strain to increase the surfactin yields and to construct a tailor-made genome reduced strain to realize the theoretically expected advantages of such genome reduced strains.