Untargeted metabolomics of the alkaliphilic cyanobacterium Plectonema terebrans elucidated novel stress-responsive metabolic modulations.

Alkaliphilic cyanobacteria are suitable candidates to study the effect of alkaline wastewater cultivation on molecular metabolic responses. In the present study, the impact of wastewater, alkalinity, and alkaline wastewater cultivation was studied on the biomass production, biochemical composition, and the alkalinity responsive molecular mechanism through metabolomics. The results suggested a 1.29 to 1.44-fold higher biomass production along with improved lipid, carbohydrate, and pigment production under alkaline wastewater cultivation. The metabolomics analysis showed 1.2-fold and 5.54-fold increase in the indole-acetic acid and phytoene biosynthesis which contributed to overall enhanced cell differentiation and photo-protectiveness. Furthermore, lower levels of Ribulose-1,5-bisphosphate (RuBP), and higher levels of 2-phosphoglycerate and 3-phosphoglycerate suggested the efficient fixation of CO2 into biomass, and storage compounds including polysaccharides, lipids, and sterols. Interestingly, except L-histidine and L-phenylalanine, all the metabolites related to protein biosynthesis were downregulated in response to wastewater and alkaline wastewater cultivation. The cells protected themselves from alkalinity and nutrient stress by improving the biosynthesis of sterols, non-toxic antioxidants, and osmo-protectants. Alkaline wastewater cultivation regulated the activation of carbon concentration mechanism (CCM), glycolysis, fatty-acid biosynthesis, and shikimate pathway. The data revealed the importance of alkaline wastewater cultivation for improved CO2 fixation, wastewater treatment, and producing valuable bioproducts including phytoene, Lyso PC 18:0, and sterols. These metabolic pathways could be future targets of metabolic engineering for improving biomass and metabolite production. SIGNIFICANCE: Alkalinity is an imperative factor, responsible for the contamination control and biochemical regulation in cyanobactera, especially during the wastewater cultivation. Currently, understanding of alkaline wastewater responsive molecular mechanism is lacking and most of the studies are focused on transcriptomics of model organisms for this purpose. In this study, untargeted metabolomics was employed to analyze the impact of wastewater and alkaline wastewater on the growth, CO2 assimilation, nutrient uptake, and associated metabolic modulations of the alkaliphilic cyanobacterium Plectonema terebrans BERC10. Results unveiled that alkaline wastewater cultivation regulated the activation of carbon concentration mechanism (CCM), glycolysis, fatty-acid biosynthesis, and shikimate pathway. It indicated the feasibility of alkaline wastewater as promising low-cost media for cyanobacterium cultivation. The identified stress-responsive pathways could be future genetic targets for strain improvement.

Impact of wastewater cultivation on pollutant removal, biomass production, metabolite biosynthesis, and carbon dioxide fixation of newly isolated cyanobacteria in a multiproduct biorefinery paradigm.

The impact of wastewater cultivation was studied on pollutant removal, biomass production, and biosynthesis of high-value metabolites by newly isolated cyanobacteria namely Acaryochloris marina BERC03, Oscillatoria sp. BERC04, and Pleurocapsa sp. BERC06. During cultivation in urabn wastewater, its pH used to adjust from pH 8.0 to 11, offering contamination-free cultivation, and flotation-based easy harvesting. Besides, wastewater cultivation improved biomass production by 1.3-fold when compared to control along with 3.54-4.2 gL-1 of CO2 fixation, concomitantly removing suspended organic matter, total nitrogen, and phosphorus by 100%, 53%, and 88%, respectively. Biomass accumulated 26-36% carbohydrates, 15-28% proteins, 38-43% lipids, and 6.3-9.5% phycobilins, where phycobilin yield was improved by 1.6-fold when compared to control. Lipids extracted from the pigment-free biomass were trans-esterified to biodiesel where pigment extraction showed no negative impact on quality of the biodiesel. These strains demonstrated the potential to become feedstock of an integrated biorefinery using urban wastewater as low-cost growth media.

Assessment of insertion/deletion polymorphism of ACE gene as a genetic risk marker for preeclampsia in pregnant women.

OBJECTIVE: To investigate the possible associations of angiotensin converting enzyme insertion or deletion genotypes and alleles with the risk of preeclampsia in Arab women. METHODS: The case-control study was conducted from January 2016 to December 2017 at King Abdulaziz University Hospital and Maternity & Children Hospital, Jeddah, Saudi Arabia, and comprised pregnant women withpreeclampsia as cases and normal pregnant women as controls. Deoxyribonucleic acid was extracted and angiotensin-converting enzyme gene was amplified by polymerase chain reaction analysis and characterised through gel electrophoresis. RESULTS: Of the 162 women, 68(42%) were cases and 94(58%) controls. The mean values of age, body mass index, and systolic and diastolic blood pressure were significantly different among the cases than the controls (p<0.05), but mean gestational age did not significantly differ between the groups (p>0.05). The distribution of the polymorphic variants of the angiotensin converting enz yme gene insertion/deletion was not significantly different between the groups (p>0.05). Also, genotype distribution and allelic frequencies were not significantly different between the groups (p>0.05). CONCLUSIONS: For insertion/deletion polymorphism, no significant differences were detected in the genotype and allele frequencies or any of the inheritance models between preeclampsia patients and controls.

Microalgal flocculation: Global research progress and prospects for algal biorefinery.

Microalgal research has made significant progress due to versatile and high-value industrial applications of microalgal biomass or its derivatives. However, to explore their full potential and to achieve commercial robustness, microalgal biorefinery needs cost-effective technologies to produce, harvest, and process the microalgal biomass on large scale as higher production and harvesting cost is one of the key hindrances in the commercialization of algae-based products. Among several other algal biomass harvesting technologies, self-flocculation seems to be an attractive, low-cost, and eco-friendly harvesting technology. This review covers various flocculation-based methods that have been employed to harvest microalgal biomass with a special emphasis on self-flocculation in microalgae. Moreover, genetic engineering approaches to induce self-flocculation in non-flocculating microalgae along with the factors affecting self-flocculation and recent research trends have also been discussed. It is concluded that self-flocculation is the most desired approach for the energy- and environment-efficient harvesting of microalgal biomass. However, its poorly understood genetic basis needs to be deciphered through detailed studies to harness its potential for the algal biorefinery.

Bioenergy potential of the residual microalgal biomass produced in city wastewater assessed through pyrolysis, kinetics and thermodynamics study to design algal biorefinery.

The suitability of integrating biological and thermal transformation of microalgal biomass to design a biorefinery was studied. The mixed cultivation of Chlorella sp. and Bracteacoccus sp. in city wastewater produced 12 g L-1 of biomass (0.77 g L-1 day-1) and removed nitrates and phosphates by 68% and 75%, respectively. Microalgae outcompeted the contaminating microbes by raising the pH of wastewater to 9.93. The lipid-free residual biomass was pyrolyzed at four heating rates (10, 20, 30, 40 °C min-1) which showed a three-stage pyrolysis. The activation energies (182-256 kJ mol-1) and their corresponding lower enthalpies at the conversional fractions from 0.2 to 0.6 indicated that product formation was being favored. The values of pre-exponential factors (1015-17 s-1), Gibbs free energy (159-190 kJ mol-1) and entropy (43-81 J mol-1) showed efficient pyrolysis. The data may lead to establish a robust microalgal biorefinery to produce biomass and energy along with primary treatment of city wastewater.

Bioenergy potential of Wolffia arrhiza appraised through pyrolysis, kinetics, thermodynamics parameters and TG-FTIR-MS study of the evolved gases.

This study evaluated the bioenergy potential of Wolffia arrhiza via pyrolysis. The biomass was collected from the pond receiving city wastewater. Oven dried powdered biomass was exposed to thermal degradation at three heating rates (10, 30 and 50°â€¯C min-1) using Thermogravimetry-Differential Scanning Calorimetry analyzer in an inert environment. Data obtained were subjected to the isoconversional models of Kissenger-Akahira-Sunose (KSA) and Flynn-Wall-Ozawa (FWO) to elucidate the reaction chemistry. Kinetic parameters including, Ea (136-172 kJmol-1) and Gibb's free energy (171 kJmol-1) showed the remarkable bioenergy potential of the biomass. The average enthalpies indicated that the product formation is favored during pyrolysis. Advanced coupled TG-FTIR-MS analyses showed the evolved gases to contain the compounds containing CO functional groups (aldehydes, ketones), aromatic and aliphatic hydrocarbons as major pyrolytic products. This low-cost abundant biomass may be used to produce energy and chemicals in a cost-efficient and environmentally friendly way.

Thermodynamics and Kinetics Parameters of Eichhornia crassipes Biomass for Bioenergy.

BACKGROUND: Eichhornia crassipes is an aquatic plant well known for its role in soil reclamation due to the containment of valuable nutrients. Moreover, its biomass is an abundant and low-cost biological resource. Pyrolysis of a biomass offers one of the cleanest methods to harness the bioenergy stored in the biomass. OBJECTIVE: The present study was focused on evaluating the bioenergy potential of Eichhornia crassipes via pyrolysis. METHODS: Biomass of E. crassipes was collected from a municipal wastewater pond. Oven dried powdered biomass of E. crassipes was subjected to pyrolysis at three heating rates including 10, 30 and 50 °C min-1 in a simultaneous Thermogravimetry-Differential Scanning Calorimetry analyzer under an inert environment containing nitrogen. Data obtained were subjected to isoconversional models of Kissenger-Akahira-Sunose (KSA) and Flynn-Wall-Ozawa (FWO) to understand the reaction chemistry. RESULTS: Kinetic parameters have shown that the pyrolysis followed first-order reaction kinetics. The average values of activation energies (129.71-133.03 kJ mol-1) and thermodynamic parameters including high heating values (18.12 MJ kg-1), Gibb's free energies (171-180 kJ mol-1) and enthalpy of reaction (124-127 kJ mol-1) have shown the remarkable bioenergy potential of this biomass. CONCLUSION: This low-cost biomass may be used to produce liquids, gases, and biochar in a costefficient and environmentally friendly way via pyrolysis or co-pyrolysis in the future.

Heterologous Synthesis and Recovery of Advanced Biofuels from Bacterial Cell Factories.

BACKGROUND: Microbial engineering to produce advanced biofuels is currently the most encouraging approach in renewable energy. Heterologous synthesis of biofuels and other useful industrial chemicals using bacterial cell factories has radically diverted the attentions from the native synthesis of these compounds. However, recovery of biofuels from the media and cellular toxicity are the main hindrances to successful commercialization of advanced biofuels. Therefore, membrane transporter engineering is gaining increasing attentions from all over the world. OBJECTIVE: The main objective of this review is to explore the ways to increase the microbial production of biofuels by counteracting the cellular toxicity and facilitating their easier recovery from media. CONCLUSION: Microbial synthesis of industrially viable compounds such as biofuels has been increased due to genomic revolution. Moreover, advancements in protein engineering, gene regulation, pathway portability, metabolic engineering and synthetic biology led the focus towards the development of robust and cost-effective systems for biofuel production. The most convenient way to combat cellular toxicity and to secrete biofuels is the use of membrane transport system. The use of membrane transporters is currently a serious oversight as do not involve chemical changes and contribute greatly to efflux biofuels in extracellular milieu. However, overexpression of transport systems can also be detrimental to cell, so, in future, structure-based engineering of transporters can be employed to evaluate optimum expression range, to increase biofuel specificity and transport rate through structural studies of biofuel molecules.

Pyrolysis, kinetics analysis, thermodynamics parameters and reaction mechanism of Typha latifolia to evaluate its bioenergy potential.

This work was focused on understanding the pyrolysis of Typha latifolia. Kinetics, thermodynamics parameters and pyrolysis reaction mechanism were studied using thermogravimetric data. Based on activation energies and conversion points, two regions of pyrolysis were established. Region-I occurred between the conversion rate 0.1-0.4 with peak temperatures 538K, 555K, 556K at the heating rates of 10Kmin-1, 30Kmin-1, and 50Kmin-1, respectively. Similarly, the Region-II occurred between 0.4 and 0.8 with peak temperatures of 606K, 621K, 623K at same heating rates. The best model was diffusion mechanism in Region-I. In Region-II, the reaction order was shown to be 2nd and 3rd. The values of activation energy calculated using FWO and KAS methods (134-204kJmol-1) remained same in both regions reflecting that the best reaction mechanism was predicted. Kinetics and thermodynamic parameters including E, ΔH, ΔS, ΔG shown that T. latifolia biomass is a remarkable feedstock for bioenergy.

Diversity, Physiochemical and Phylogenetic Analyses of Bacteria Isolated from Various Drinking Water Sources.

OBJECTIVE: To evaluate the indigenous bacterial strains of drinking water from the most commercial water types including bottled and filtered water that are currently used in Saudi Arabia. METHODS: Thirty randomly selected commercial brands of bottled water were purchased from Saudi local markets. Moreover, samples from tap water and filtered water were collected in sterilized glass bottles and stored at 4°C. Biochemical analyses including pH, temperature, lactose fermentation test (LAC), indole test (IND), methyl red test (MR), Voges-Proskauer test (VP), urease test (URE), catalase test (CAT), aerobic and anaerobic test (Ae/An) were measured. Molecular identification and comparative sequence analyses were done by full length 16S rRNA gene sequences using gene bank databases and phylogenetic trees were constructed to see the closely related similarity index between bacterial strains. RESULTS: Among 30 water samples tested, 18 were found positive for bacterial growth. Molecular identification of four selected bacterial strains indicated the alarming presence of pathogenic bacteria Bacillus spp. in most common commercial types of drinking water used in Saudi Arabia. CONCLUSION: The lack of awareness about good sanitation, poor personal hygienic practices and failure of safe water management and supply are the important factors for poor drinking water quality in these sources, need to be addressed.

Heterologous expression of the antifungal ß-chitin binding protein CBP24 from bacillus thuringiensis and its synergistic action with bacterial chitinases.

The genome sequence analysis of Bacillus thuringiensis serovar konkukian S4 has shown to contain two chitinases (Chi74, Chi39) and two chitin-binding proteins (CBP50 and CBP24). The Chi74, Chi39 and CBP50 have been characterized previously. The chitin-binding protein CBP24 was cloned and heterologously expressed in Escherichia coli. The recombinant protein was purified using a Ni-NTA purification system. The purified protein was used to study its substrate binding activity using crystalline chitin variants as substrates. The Bmax and Kd values have shown that it preferably binds to ß-type of the crystalline chitin at a range of pH with peak activity between 5.5-7.5. To elucidate the role of CBP24 in the chitin degradation system of S4, the purified chitinases Chi74, Chi39 along with the ChiA from Serratia proteamcualans were used in different combinations with the CBP24 and chitinolytic activity was assayed. It was shown that the CBP24 acts synergistically with chitin degradation activity of bacterial chitinases non-specifically. Moreover, the CBP24 has shown antifungal activity against plant pathogenic fungi Fusarium oxysporum and Rhizoctonia solani. The present study will lead us to develop a technology for environmental friendly conversion of chitin to valuable products.

Domain wise docking analyses of the modular chitin binding protein CBP50 from Bacillus thuringiensis serovar konkukian S4.

This paper presents an in silico characterization of the chitin binding protein CBP50 from B. thuringiensis serovar konkukian S4 through homology modeling and molecular docking. The CBP50 has shown a modular structure containing an N-terminal CBM33 domain, two consecutive fibronectin-III (Fn-III) like domains and a C-terminal CBM5 domain. The protein presented a unique modular structure which could not be modeled using ordinary procedures. So, domain wise modeling using MODELLER and docking analyses using Autodock Vina were performed. The best conformation for each domain was selected using standard procedure. It was revealed that four amino acid residues Glu-71, Ser-74, Glu-76 and Gln-90 from N-terminal domain are involved in protein-substrate interaction. Similarly, amino acid residues Trp-20, Asn-21, Ser-23 and Val-30 of Fn-III like domains and Glu-15, Ala-17, Ser-18 and Leu-35 of C-terminal domain were involved in substrate binding. Site-directed mutagenesis of these proposed amino acid residues in future will elucidate the key amino acids involved in chitin binding activity of CBP50 protein.