Herein, silver and zinc oxide Nanoparticles (NPs) were synthesized by using W. coagulant fruit extract as reducing agent and capping agent. The green synthesized NP with distinct properties were used for novel application against fungal and bacterial pathogen of honey bee (A. mellifera). The UV-spectroscopy confirms the synthesis of silver and zinc oxide NPs at 420 nm and 350 nm respectively. Further, XRD evaluated the monoclinic structure of Ag NPs while ZnO NPs showed wurtzite hexagonalcrystlized structure. Resistant honey bee pathogens such Paenibacilluslarvae, Melissococcus plutonius and Ascosphaera apis were isolated, identified and cultured in vitro to assess the antimicrobial potentials of Ag and ZnO NPs. Additionally, different biomolecules provide access to achieve maximum and stable Ag and ZnO NPs. It was also observed that with increasing the concentration of zinc oxide NPs and sliver NPs, zone of inhibition was also increased. Thus, present findings show that plant extracts can be a useful natural resource to prepare functional nonmaterial for targeted applications especially in the field of apicultural research.

The baculovirus expression system (BEVS) is widely used in biomanufacturing. However, massive late and very late genes are expressed during baculovirus infection of the host cell, and the protein aggregation has a significant negative impact on the intercellular environment of cells and organelle function. Baculovirus particle contamination also hinders the purification of particulate products. In this study, a novel baculovirus vector of deficient-lef5 (Ac-Δlef5) was established to prevent the production of baculovirus particles. It could transduce cells and replicate viral DNA but the expression of the late and very late genes was reduced so that no virus was packed and budded. The Ac-Δlef5 expression system was successfully used to express a secreted protein, thyroid peroxidase (TPO). Compared to wild-type baculovirus (Ac-wt), Ac-Δlef5 delayed the decline in cell viability and prolonged the harvest period from 4 to 6 days after infection. When expressed in the Ac-Δlef5 system, 98% of TPO was secreted extracellularly, which was about 1.8 times that of the Ac-wt system. Meanwhile, the transcription levels of protein folding-related genes were significantly increased. The results show that the Ac-Δlef5 system is a potential novel viral-free baculovirus expression system, which omits the virion removal operation in biomanufacturing.

Bio-functionalized metal oxide nanoparticles (NPs) have been taken great importance in biomedical fields. The use of nanoparticles as delivery agents of therapeutic molecules led the researchers to emphasize the potential impact of these NPs on bio-macromolecules as protein-nanoparticle complexes, which also extended their importance as vehicles in targeted drug delivery systems due to increased ease of administration, firmness, reduced toxic side effects, and half-life of drugs. Since human serum albumin is the blood protein responsible for transporting materials in the blood system, the interaction of these particles with HSA is essential to be understood before considering the nanoparticles for any individual biomedical application. In the present study, we synthesized zinc-oxide nanorods (ZONRs) using a microwave-assisted synthesis technique, and characterized them by XRD, FTIR, Raman, SEM-EDX, UV-Vis spectroscopy, and photoluminescence (PL) spectroscopy methods. The interaction studies were carried out using fluorescence spectroscopy, and the change in secondary structure was analyzed using CD spectroscopy. The results of MTT cell viability assay demonstrated that the ZONRs has potential cytotoxic properties.

Rhizobial inoculants are sold either as rhizobia within a liquid matrix; or as rhizobia adhered to granules composed of peat prill or finely ground peat moss. During the production of peat-based inoculants, a series of physiological changes occur that result in an increased capability of the rhizobia to survive on the seeds. The number of viable rhizobia on preinoculated seeds at the point of sale, however, is often a limiting factor, as is the inefficiency of the inoculant bacteria to compete with the local rhizobia for the host colonization. In the present work, we used STM-seq for the genome-wide screening of Ensifer meliloti mutants affected in the survival during the maturation of peat-based inoculant formulations. Through this approach, we were able to identify a set of mutants whose behavior suggests that persistence in peat inoculants involves a complex phenotype that is connected to diverse cellular activities, mainly related to satisfying the requirements of bacterial nutrition (e.g., carbon sources, ions) and to coping with specific stresses (e.g., oxidative, mutational). These results also provide a base knowledge that could be used to more completely understand the survival mechanisms used by rhizobia during the maturation of peat-based inoculants, as well as for the design and implementation of practical strategies to improve inoculant formulations.

An acid polysaccharide, named HP, was produced by endophytic Penicillium javanicum MSC-R1 isolated from southern medicine Millettia speciosa Champ. The molecular weight of HP was 37.8 kDa and consisted of Ara f, Galр, Glcр, Manр, and GlcрA with a molar ratio of 1.09: 3.47: 68.48: 16.59: 8.85. The glycosidic linkage of HP was proven to be →3, 4)-α-D-Glcр-(1→6)-α-D-Manр-(1→, →3, 4)-α-D-Glcр-(1→4)-α-D-Glcр-(1→, →3), →6)-α-D-Manр-(1→4)-α-D-Glcр-(1→, →3), β-D-Galр-(1→3)-α-D-Glcр-(1→, →4), →5)-α-L-Ara f -(1→3)-α-D-Glcр-(1→, →4), →6)-α-D-Manр-(1→4)-α-D-GlcAр-(1→ and →4)-α-D-GlcAр-(1→4)-α-D-Glcр-(1→, →3). Additionally, 250 μg/mL of HP possessed nontoxicity to RAW 264.7 cells and exhibited anti-inflammation activity. HP could significantly restrain the amount of tumor necrosis factor-α, interleukin-6 and NO release in RAW264.7, which property is possibly associated with its abundant glucosidic linkage. These results indicated that HP could be regarded as a ponderable ingredient for the health-beneficial functional foods.

The growth-associated metabolites are produced during the exponential phase; however, this phase terminates due to substrate depletion or product inhibition. In the present study, a semicontinuous mode with a fill-and-draw strategy was applied to extend the exponential phase of the biosurfactant production to overcome the product inhibition and in turn, enhance the yield. Bioreactor studies were performed in batch mode, followed by the semicontinuous operation. A potential biosurfactant producer Bacillus subtilis RSL2 was used in this study at the previously optimized conditions of pH 6.6, temperature 41 °C and 5% (w/v) of molasses. A better mass transfer was achieved in the bioreactor as compared to the shake flask study. In the batch bioreactor study, 90% of sugar was utilized with simultaneous 13.7 g L-1 of biosurfactant production. The sugar utilization was further improved to > 98% in the case of semicontinuous operation employing a fill-and-draw strategy. The exponential phase got extended up to 18 days and a total of 13 L of media was fed in the semicontinuous operation of 21 days as compared to 1.5 L of working volume in the batch reactor. The biosurfactant yield was enhanced by 1.5 folds and was found to be 0.97 g g-1. The produced biosurfactant was identified as a lipopeptide. The interfacial properties of the biosurfactant along with colloidal and thermal stability have been investigated. The critical micelle concentration of the produced biosurfactant was 70 mg L-1. The present study highlighted the efficient utilization of molasses for the production of biosurfactant, an alternative metabolite, in a semicontinuous mode of bioreactor.

qRT-PCR is a globally accepted technique for assaying gene expression in relative terms which compares the difference between critical threshold (CT) values of a gene calculated form two independently isolated RNA samples. Independent RNA isolations, however, include error due to batch effect which must be normalized for error-free calculation of relative gene expression. Hence, CT values of internal control (IC) genes are used for normalization during the calculation of expression fold-change in gene expression analysis. The expression of ICs genes expected to be stable in all the experimental conditions. However, it is almost impossible to find such a gene which do not depict expression fluctuation in response to the changes in experimental conditions. Hence, it is necessary to identify suitable IC gene(s) for any given experimental condition before conducting any particular gene expression study. Here, we examined the suitability of eight candidate IC genes, namely glyceraldehyde 3-phosphate dehydrogenase (GAPDH), eukaryotic elongation factor-1 (eEF-1α), 25 S rRNA (25 S), 18 S rRNA (18 S), ubiquitin C E2 ligase (UBC), Actin (Act), ubiquitin 5 (UBQ5) and ubiquitin 10 (UBQ10), for assaying gene expression in rice during sheath blight infection. Our analysis suggest that GAPDH might be the IC of choice when expression studies include contrasting genotypes differing in their tolerance to sheath blight pathogen as well as progressive infection time. While if expression analysis have to be performed only in one genotype but under progressive sheath blight infection, UBQ5 might be chosen as IC because of its high expression stability under the proposed experimental setup.

A hexagonal mesoporous molecular sieve-like structure of MCM41 and SBA15 with a large surface area was used to immobilize protein L-ribose isomerase (L-RI) through covalent linkages. The amino group of APTES functionalized nanosilica support MCM41 and SBA15 interacted with glutaraldehyde to promote bidentate linkage and on other side with amino group of enzyme. The use of mesoporous silica matrix for immobilization was observed to conserve the distinctive properties of the protein. The various operational conditions optimized for covalent conjugation of protein with the silica support were found to be dependent on enzyme support ratio, immobilization temperature and time. The immobilization yield of L-RI on MCM41 and SBA15 was achieved to be 60 % (600 mg enzyme /g matrix) and 45 % (450 mg enzyme/g matrix), respectively under the optimized conditions. The immobilized biocatalyst was characterized by various analytical techniques like HR-TEM, EDS, FTIR, TGA and BET. Effects of different experimental conditions were optimized to study enzyme kinetics, pH, temperature, bioconversion, reusability, metal ion effect and storage stability. The biocatalytic efficiency (kcat/Km) was increased by 1.2 fold on immobilization with the catalytic activity of 39.64 IU. Increase in the catalytic efficiency after immobilization could be due to the suitable orientation of enzyme active site and improved accessibility for substrate binding. The immobilization of L-RI on mesoporous silica support could improve the biocatalytic activity, storage stability and reusability. The immobilized biocatalyst was found to be reusable for more than 4 cycles retaining more than 50 % of catalytic activity and promoting the synthesis of a rare sugar L-ribose from L-ribulose with a conversion yield of 22 % in 2 h time.

Acid-casein production generates waste streams that are rich in nitrogen (in the form of protein and nitrate) and phosphate. This makes this type of waste very difficult to treat using conventional techniques resulting in a high amount of operating cost and costly investment. In this research, the application of single culture or consortium of microalgae for uptake of nitrogen and phosphate in the wastewater of an acid-casein factory was investigated. The waste was a 1:1 mixture of nanofiltered whey permeate and dairy processing wastewater. Monocultures of Chlorella vulgaris, Tetradesmus obloquus, Nonnochlropsis ocenica and a consortium of the three microalgae were analyzed. The results showed that the consortium exhibited more efficient nitrogen and phosphate removal compared to the individual species. The consortium was able to rapidly hydrolyse exogenous protein present in the waste medium, removing 88% of protein and breaking down complex protein molecules into simpler compounds (such as nitrate) for assimilation into the biomass. In the first fourteen days of cultivation, the rate of nitrate assimilation by the consortium biomass was lower than that of nitrate formation from protein degradation, leading to a net increase in nitrate concentration in the medium. As protein source was depleted and biomass concentration increased, however, the rate of nitrate assimilation began to exceed that of nitrate formation allowing for net removal of nitrate. The microalgae consortium was shown to successfully bioremediate all nitrates by day 21. It was indicated that Chlorella and Nannochloropsis species were responsible for nitrogen removal in monocultures. Phosphate, on the other hand, was efficiently removed by Tetradesmus. The results indicated that a consortium cultivation of three species of microalgae led to effective elimination of both nitrogen and phosphate. Combined flow-cytometry and microscopy analyses revealed that Chlorella overtook Tetradesmus and Nannochloropsis to emerge as the dominant population in the consortium by the end of the cultivation cycle. It can be concluded that the application of microalgae consortium for simultaneous recovery of nitrogen and phosphate is a promising approach for treating acid-casein wastewater.

Human society cannot afford the luxury of the business-as-usual approach when dealing with the emerging challenges of the 21st century. The challenges of food production to meet the pace of population growth in an environmentally-sustainable manner have increased considerably, emphasizing the need to explore newer approaches to agriculture. Agrochemical-based agricultural practices are known to have serious environmental and health implications. Even conventional organic farming is not sustainable in the long run. Although some "age-old" practices are useful, these will not help feed more people on the same or less land more sustainably. Sustainable intensification is the way forward. There is a need to incorporate a customer-centric outlook and make the organic system sustainable. Here, we bring forth the necessity to enhance the efficiency of organic agriculture by the inclusion of robotics and agrochemical-free GM seeds. Such an organic-GM hybrid agriculture system integrated with the use of artificial intelligence (AI) based technologies will have better energy efficiency. The produce from such a system will offer consumers a 'third' choice and create a new food label, 'organically-grown GM produce'.

For tissue engineering applications, cell adhesion and proliferation are crucial factors, and blending polymers is one of the most effective ways to create a biocompatible scaffold with desired properties. In order to create new potential porous, biodegradable scaffolds using salt leaching technique, poly(3-hydroxybutyrate) (PHB) and poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) were blended in different ratios. SEM, BET, FTIR, and water contact angle measurements were used to analyze the scaffolds' porous surface, surface area, and roughness, chemical interaction, and hydrophilicity. Additionally, a hemolysis assay revealed that the mixtures were hemocompatible and had no impact on red blood cells. Different cells- Vero, Hela and MDBK cell lines cultured on the porous mats of these biopolymeric scaffolds exhibited significant increase in cell viability and attachment over time. The overall finding was that blended scaffolds exhibited reduced crystallinity, diverse porosity, higher surface area and hydrophilicity, and better cell viability, proliferation and adhesion. Our findings imply that a blended scaffold could be more suitable for use in tissue engineering applications.

In this report, in vitro doubled haploid (DH) plants were established in two tea (Camellia spp) cultivars, TV21 (Assam Type) and TV19 (Cambod Type). Androgenic globular stage haploid embryos, obtained via callusing from microspores at an early-to-late uninucleate stage in anther cultures, were diploidized by colchicine treatments at varying concentrations and durations under dark incubation at 25 ± 2 °C temperature. Thereafter, treated embryos were transferred to development medium, Murashige and Skoog (MS) medium supplemented with 6-benzylaminopurine (BAP; 1 μM) + gibberellic acid (GA3; 0.3 μM) + L-glutamine (80 mg l-1) + L-serine (20 mg l-1) and incubated in diffused light. Ploidy of germinating embryos was evaluated by flow-cytometry and cytological squash preparation. High chromosome doubling, 76.89% and 67.34%, was obtained in embryos of TV21 and TV19, respectively, at 0.2% colchicine treatment for 24 h. The DH plants were further multiplied via axillary-bud proliferation on multiplication medium, MS + glucose (30 g l-1) + BAP (5 μM) + GA3 (0.5 μM) + IBA (0.5 μM) + L- glutamine (80 mg l-1) + L-serine (20 mg l-1). Rooting of shoots was achieved on ⅓ MS basal medium within 50 days of inoculation when shoots were pre-treated with IBA (175 μM) for ten days. The rooted plants were acclimatized in field. Homozygosity in diploidized plants was validated by SSR marker.

An efficient capture step for human adenovirus type 5 from cell lysate was developed as an initial virus purification step from cell debris supernatant. Organosilane-based polymer particles were synthesized and experimental monomer screening allowed the selection of appropriate functionalities for the development of particles for virus binding. After elution, virus recoveries of 83 % were obtained with significant reduction of matrix proteins and residual host cell DNA. Therefore, the implemented capture strategy for adenovirus via polymer particles provides a scalable and reproducible approach to reduce time and cost during virus purification processes.

A convenient and nonradioactive method for quantifying in vitro NR biosynthesis is presented that is based upon the quantitation of substrate depletion by ultra-high performance liquid chromatography tandem mass spectrometry (UHPLC-MS/MS). NR oligomers could be in vitro biosynthesized with the enzyme source from Hevea brasiliensis (Hevea) or Taraxacum kok-saghyz (TKS) by exogenous monomers (IPP) and initiators (FPP). The IPP incorporation rate and FPP consumption rate were 62.24% and 51.14% respectively when the washed rubber particles (WRP) of Hevea was the enzyme source. The IPP incorporation rate and the FPP consumption rate were 74.49% and 95.90% respectively when the sediment bottom fraction (BF) of Hevea was the enzyme source. The in vitro NR biosynthesis can be divided into two stages:(1) the initiation reaction of FPP, which occurs more in BF, and (2) the growth reaction of IPP, which occurs more in WRP. In addition, the IPP incorporation and FPP consumption rates were 59.39% and 34.15% respectively when the BF of TKS was selected as an enzyme source.

Development of low-cost and economic cellulase production is among the key challenges due to its broad industrial applications. One of the main topics of research pertaining to sustainable biomass waste based biorefinaries is the development of economic cellulase production strategies. The main cause of the increase in cellulase production costs is the use of commercial substrates; as a result, the cost of any cellulase-based bioprocess can be decreased by employing a productive, low-cost substrate. The goal of the current study is to develop low-cost cellulase using the carbohydrate-rich, renewable, and widely accessible cyanobacteria algae Oscillatoria obscura as the production substrate. Maximum cellulase was produced utilising the fungus Rhizopus oryzae at substrate concentration of 7.0 g among various tested concentrations of algal biomass. Maximum production rates of 22 IU/gds FP, 105 IU/gds BGL, and 116 IU/gds EG in 72 h were possible under optimal conditions and substrate concentration. Further investigations on the crude enzyme's stability in the presence of iron oxide nanoparticles (IONPs) revealed that it was thermally stable at 60 °C for up to 8 h. Additionally, the crude enzyme demonstrated pH stability by maintaining its complete activity at pH 6.0 for 8 h in the presence of the optimal dose of 15 mg IONPs. The outcomes of this research may be used to investigate the possibility of producing such enzymes in large quantities at low cost for industrial use.

Bacillus subtilis is a robust industrial workhorse for the production of heterologous proteins. Chromosomal integration-based protein production has advantages over plasmid-based methods. Considering that the expression level of a gene is affected by its location in the chromosome, it is important to find an optimal integration site for the gene to be expressed. This work establishes a method for random insertion of a gene expression cassette into chromosomes, enabling the screening of optimal integration sites for high-level protein production. Specifically, a gene expression cassette and a chloromycetin-resistance marker are assembled into a transposon. This transposon is inserted between the promoter and the ribosomal binding site of the zeocin-resistance marker in the chromosome, which blocks the transcription of the zeocin-resistance gene. Transposase Himar1-mediated transposition of this transposon activates the zeocin-resistance marker, which can be selected on plates containing both chloromycetin and zeocin. The transposition frequency was over 10-5. This method was used to select proper insertion sites for the expression cassette of methyl parathion hydrolase (MPH). Compared with the common integration site amyE, the expression level of MPH was increased up to 50 % at the yjbH site.

One of the main factors affecting hydrogen production and growth of photofermentative microorganisms is light; low light penetration and utilization are significant bottlenecks in photofermentative hydrogen production systems. In this study, light distribution in a thermosiphon photobioreactor operated with Rhodopseudomonas palustris was investigated. Radiation fields were modelled and simulated using computational fluid dynamics (ANSYS® Fluent, 2019 R2) and a reflector system was evaluated for the enhancement of light distribution in a thermosiphon photobioreactor. The effect of the reflector system was investigated experimentally in terms of hydrogen production, carbon substrate consumption and biomass circulation in the reactor. With the addition of the reflector system, hydrogen production was increased by 48% while glycerol consumption was increased from approximately 24% to 32%. After 336 h, the concentration of R. palustris cells still in suspension ranged from 0.13 to 0.18 g∙L-1, with no discernible difference in concentration between the systems with and without reflectors. Collectively, the reflector system was shown to be a viable option in enhancing light distribution in photobioreactors, with an associated increase in both hydrogen production as well as glycerol consumption.

Laccase production by fungal growth on agrifood waste is still poorly studied. Trametes versicolor K1 isolated from palm bark produced a yellow non glycosylated laccase from tomato waste based medium (TMT) and a blue glycosylated laccase on glucose medium (GLU). Lignocellulosic biomass, such as pinecones (PIN), palm leaves (PLM), olive pomace (OLV), and alfa stems (ALF) have also been used as growth medium for T. versicolor K1. In these conditions, very low or no laccase production was observed. When peptone was supplied in TMT medium, the laccase activity increased from 4170 U/L to 8618 U/L. By increasing the culture volume up to 1 L, laccase production on TMT was 9929 U/L. The yellow laccase (TmtLac) was purified from the supernatant TMT medium and has shown similar characteristics with the blue laccase (GluLac) purified from the GLU medium. Their apparent protein size was 63 kDa. Catalytic activities of the yellow form were not very different from those of the blue form, but specific activity of the purified yellow laccase produced on tomato waste was much higher. The Km and Vm values for four substrates, ABTS, DMP, guaiacol, and pyrogallol were almost similar for both isoenzymes. The optimum pH and temperature were respectively 4.0 and 50 °C. Although the level of glycosylation is clearly different, the thermostability of TmtLac and GluLac are quite similar. TmtLac is even slightly more tolerant at 60 °C for 24 h than GluLac. Moreover TmtLac showed greater stability at alkaline pH after 24 h compared to that of GluLac.We demonstrate that activity of the yellow TmtLac is not significantly affected compared to the blue laccase and that tomato waste is a simple and interesting lignocellulosic substrate to the laccase producer Trametes sp.

Lacto-N-fucopentaose III (LNFP III) is a human milk oligosaccharide (HMO) with potential health benefits in infants, including in immune development and modulation of the intestinal environment. Low-cost fermentative production of various HMOs from lactose by engineered Escherichia coli has attracted attention, but few reports have investigated long-chain HMO production, such as of the pentasaccharide LNFP III. LNFP III is synthesized by α1,3-fucosyltransfer reaction to the glucosamine (GlcNAc) moiety in the lacto-N-neotetraose (LNnT) skeleton by α1,3-fucosyltransferase (α1,3-FucT). However, the known α1,3-FucTs also transfer fucose to the reducing terminal glucose moiety of LNnT or the starting material lactose, resulting in various byproducts. Here, we found a useful α1,3-FucT from Parabacteroides goldsteinii (PgsFucT), which is only reactive for GlcNAc in the N-acetyllactosamine (LacNAc) skeleton in vivo. On the basis of sequence alignment with a FucT of known structure, we also generated α1,3-FucT variants with altered reactivity for LacNAc or lactose. An E. coli strain heterologously expressing PgsFucT accumulated 3.84 g/L of LNFP III after 48 h of culture in a 3-L jar-fermenter. The amounts of various byproduct sugars were remarkably decreased compared with a strain expressing the previously characterized α1,3-fucT from Bacteroides fragilis.

Global warming is the major cause of abiotic and biotic stresses that reduce plant growth and productivity. Various stresses such as drought, low temperature, pathogen attack, high temperature and salinity all negatively influence plant growth and development. Due to sessile beings, they cannot escape from these adverse conditions. However, plants develop a variety of systems that can help them to tolerate, resist, and escape challenges imposed by the environment. Among them, anthocyanins are a good example of stress mitigators. They aid plant growth and development by increasing anthocyanin accumulation, which leads to increased resistance to various biotic and abiotic stresses. In the primary metabolism of plants, anthocyanin improves the photosynthesis rate, membrane permeability, up-regulates many enzyme transcripts related to anthocyanin biosynthesis, and optimizes nutrient uptake. Generally, the most important genes of the anthocyanin biosynthesis pathways were up-regulated under various abiotic and biotic stresses. The present review will highlight anthocyanin mediated stress tolerance in plants under various abiotic and biotic stresses. We have also compiled literature related to genetically engineer stress-tolerant crops generated using over-expression of genes belonging to anthocyanin biosynthetic pathway or its regulation. To sum up, the present review provides an up-to-date description of various signal transduction mechanisms that modulate or enhance anthocyanin accumulation under stress conditions.