Bioeconomy and white biotechnology as a basic pillar of the Lisbon strategy

The entrance into the new millennium is branded by intensive development of science and new technologies. Life science and biotechnologies are widely recognize to be, after ICT, the parallel wave of knowledge – based economy, creating new opportunities for our society and economies. This application is the basic object of Lisbon strategy in Europe. This new trends, to make full use of biotechnology for sustainable economy, is official titled as bioeconomy around the world. The characteristics of bioeconomy and utilization of industrial biotechnology are presented in this article.

Extremophiles as a Source of Biotechnological Products

Extremophile and extremozyme capabilities to uphold catalytic actions under extreme situations open up a varied array of biotechnological applications. Extremophiles are a rich supply of biocatalysts used for innumerable purposes. Bioactive molecules and enzymes isolated from organisms inhabiting risky environments being used in biological innovation pipelines and pharmaceutical have positive claims. The species biodiversity has favourable reservoir of the unexploited amalgams with biotechnological significance. Prospective solicitations of extremozymes, chiefly as catalysis of multistep progressions, quorum sensing, bioremediation, biofuel, biodiversity and prospecting, biomining, and genetic technology are explored. To boost the biotechnological uses of extremozymes, research and development efforts are needed to address hurdles such as extremophile culture, gene expression in host cells, and extremozyme bioprocessing. Extremophiles can be a resource for innovative biotechnological comprising industrial biotechnology, agriculture, medical, food, and environmental biotechnology.

Genome Editing and CRISPR/Cas System of Extremophiles and Its Applications

Extremophiles will be the choice of next generation industrial biotechnology (NGIB) as they are known to be contaminant resistant, but engineering their genomes has always been difficult and time consuming task. CRIPR/Cas (clustered regularly interspaced short palindromic repeat and CRISPR associated proteins) system can be employed for this reason. The genome of an industrially important halophile (i.e., Halomonas) was edited to study a combined effect of four different genes on glucose breakdown and production of poly (3-hydroxybutyrate-co-3-hydroxyvalerate). This editing has resulted in 16-fold increase of 3HV, and the mutants generated by CRIPR/Cas system were significantly effective in synthesizing PHBV. Unfortunately, this system does not always work, specifically in extremophilic microorganisms because Cas9 or Cpf1 are from mesophilic bacteria. Therefore, alternatively, the endogenous CRISPR/Cas system is used for editing the genomes of such organisms. This genome editing of extremophiles will open the doors for developing next generation industrial biotechnology (NGIB).

Sustainable Design Approach for Modeling Bioprocesses from Laboratory toward Commercialization: Optimizing Chitosan Production

Enhancing the biochemical supply chain towards sustainable development requires more efforts to boost technology innovation at early design phases and avoid delays in industrial biotechnology growth. Such a transformation requires a comprehensive step-wise procedure to guide bioprocess development from laboratory protocols to commercialization. This study introduces a process design framework to guide research and development (R&D) through this journey, bearing in mind the particular challenges of bioprocess modeling. The method combines sustainability assessment and process optimization based on process efficiency indicators, technical indicators, Life Cycle Assessment (LCA), and process optimization via Water Regeneration Networks (WRN). Since many bioprocesses remain at low Technology Readiness Levels (TRLs), the process simulation module was examined in detail to account for uncertainties, providing strategies for successful guidance. The sustainability assessment was performed using the geometric mean-based sustainability footprint metric. A case study based on Chitosan production from shrimp exoskeletons was evaluated to demonstrate the method’s applicability and its advantages in product optimization. An optimized scenario was generated through a WRN to improve water management, then compared with the case study. The results confirm the existence of a possible configuration with better sustainability performance for the optimized case with a sustainability footprint of 0.33, compared with the performance of the base case (1.00).

A simple RT-multiplex PCR method for diagnosis of L-A and M totiviruses in Saccharomyces cerevisiae

Killer yeasts and their toxins have many potential applications in environmental, medical and industrial biotechnology. The killer phenotype in Saccharomyces cerevisiae relies on the cytoplasmic persistence of two dsRNA viruses, L-A and M. M encodes the toxin, and L-A provides proteins for expression, replication, and capsids for both viruses. Yeast screening and characterization of this trait is usually performed phenotypically, on the basis of their toxin production and immunity. In this study, we describe a simple and specific RT-multiplex PCR assay for direct diagnosis of the dsRNA totivirus genomes associated to the killer trait in the S. cerevisiae yeast. This method obviates RNA purification steps and primers addition to the RT reaction. Using a mixture of specific primers at the PCR step, this RT-multiplex PCR protocol provides accurate diagnosis of both L-A and M totivirus in all its known variants L-A-1/M1, L-A-2/M2, L-A-28/M28 and L-A-lus/Mlus to be found in infected killer yeasts. By means of this method, expected L-A-2/M2 totivirus associations in natural wine yeasts cells were identified, but importantly, asymptomatic L-A-2/M2 infected cells, as well as unexpected L-A-lus/M2 totiviral associations, were also found. Importance The killer phenomenon in S. cerevisiae yeast cells provides the opportunity to study host-virus interactions in a eukaryotic model. Therefore, development of simple methods for their detection significantly facilitates their study. The simplified RT-multiplex PCR protocol described here provides a useful and accurate tool for the genotypic characterization of yeast totiviruses in killer yeast cells. The killer trait depends on two dsRNA totiviruses, L-A and M. Each M dsRNA depends on a specific helper L-A virus. Thus, direct genotyping by the described method also provides valuable insights into L-A/M viral associations and their coadaptional events in nature.

Filamentous fungal applications in biotechnology: a combined bibliometric and patentometric assessment

Background Processes and products employing filamentous fungi are increasing contributors to biotechnology. These organisms are used as cell factories for the synthesis of platform chemicals, enzymes, acids, foodstuffs and therapeutics. More recent applications include processing biomass into construction or textile materials. These exciting advances raise several interrelated questions regarding the contributions of filamentous fungi to biotechnology. For example, are advances in this discipline a major contributor compared to other organisms, e.g. plants or bacteria? From a geographical perspective, where is this work conducted? Which species are predominantly used? How do biotech companies actually use these organisms? Results To glean a snapshot of the state of the discipline, literature (bibliometry) and patent (patentometry) outputs of filamentous fungal applications and the related fields were quantitatively surveyed. How these outputs vary across fungal species, industrial application(s), geographical locations and biotechnological companies were analysed. Results identified (i) fungi as crucial drivers for publications and patents in biotechnology, (ii) enzyme and organic acid production as the main applications, (iii) Aspergillus as the most commonly used genus by biotechnologists, (iv) China, the United States, Brazil, and Europe as the leaders in filamentous fungal science, and (v) the key players in industrial biotechnology. Conclusions This study generated a summary of the status of filamentous fungal applications in biotechnology. Both bibliometric and patentometric data have identified several key trends, breakthroughs and challenges faced by the fungal research community. The analysis suggests that the future is bright for filamentous fungal research worldwide.

Increased carvone production in Escherichia coli by balancing limonene conversion enzyme expression via targeted quantification concatamer proteome analysis

Abstract(−)-Carvone is a monoterpenoid with a spearmint flavor. A sustainable biotechnological production process for (−)-carvone is desirable. Although all enzymes in (−)-carvone biosynthesis have been functionally expressed in Escherichia coli independently, the yield was low in previous studies. When cytochrome P450 limonene-6-hydroxylase (P450)/cytochrome P450 reductase (CPR) and carveol dehydrogenase (CDH) were expressed in a single strain, by-product formation (dihydrocarveol and dihydrocarvone) was detected. We hypothesized that P450 and CDH expression levels differ in E. coli. Thus, two strains independently expressing P450/CPR and CDH were mixed with different ratios, confirming increased carvone production and decreased by-product formation when CDH input was reduced. The optimum ratio of enzyme expression to maximize (−)-carvone production was determined using the proteome analysis quantification concatamer (QconCAT) method. Thereafter, a single strain expressing both P450/CPR and CDH was constructed to imitate the optimum expression ratio. The upgraded strain showed a 15-fold improvement compared to the initial strain, showing a 44 ± 6.3 mg/L (−)-carvone production from 100 mg/L (−)-limonene. Our study showed the usefulness of the QconCAT proteome analysis method for strain development in the industrial biotechnology field.

CONTROL AND OPERATIONS MANAGEMENT OF EXTRACTION PROCESS IN INDUSTRIAL BIOTECHNOLOGY OF BETA-CAROTINE FROM BLAKESLEA TRISPORA

The technological parameters, namely temperature and duration of β-carotene extraction process from the biomass of filamentous fungus Blakeslea trispora with vegetable oils of various fatty acid compositions and with various contents of natural antioxidants (refined deodorized sunflower, viso-oleic sunflower, corn and sesame) have been investigated. Statistical models of dependences of β-carotene, as well as analytical numbers, characterizing the content of free fatty acids (acid number) and primary products of lipid oxidation (peroxide number) content, in oil extracts of the specified refined deodorized oils, from temperature and extraction duration have been built. Rational parameters of β-carotene extraction from Blakeslea trispora biomass with selected extractants (refined deodorized sunflower, high oleic sunflower, corn and sesame oils) have been determined for the extracts technological properties control. It has been proven that the use of these refined deodorized oils as extractants practically does not affect the content of the target product in oil extracts of biomass, but it does affect the analytical numbers of extracts characterizing the content of free fatty acids, peroxides and hydroperoxides. The highest content of free fatty acids in β-carotene containing biomass extraction with sunflower oil has been observed. The minimum content of free fatty acids in extracts with corn and sesame oils using has been be achieved. The highest content of primary products of lipid oxidation (peroxides and hydroperoxides) during β-carotene containing biomass extraction by sunflower oil has been observed. The minimum content of free fatty acids in extracts with sesame oil using has been achieved. It is possible to predict β-carotene content, as well as acid and peroxide numbers of oil extracts from biomass in specified refined deodorized oils, depending on temperature and extraction process duration using the obtained approximation dependences.