Search

12 pages, 10 figures, 6 tables.-- Data availability: Data will be made available on request, Microalgae are attractive for the development of environmentally sustainable processes with the option for carbon fixation, given the photosynthetic capacity of these microorganisms. Nitrogen and phosphorus are key nutrients for microalgal growth as well as carbon fixation and product formations. These are also important nutrients in agriculture, the sustainable supply of which is a challenge. They are often lost in waste streams but can be effectively recovered using microalgae and made available for agriculture. In this investigation, we systematically examined the nitrate and phosphate uptake response of two halotolerant strains of Chlorella vulgaris exposed to four carbon and three nitrogen and phosphorus supply regimes to develop an understanding of the relationship between uptake and supply in two microalgae strains from a similar habitat. Markedly differential response to changes in carbon and nutrient supply regimes were noted between the supply regimes and the strains. Changes in carbon supply was found to significantly influence nitrogen and phosphorus uptake in the two strains. Kinetics of nutrient depletion and carbon fixation were observed to be dynamic and non-linear for both the strains, under high carbon and nutrient supply regimes. This investigation provides for the first time a clear evidence base for the role of supply regimes in the management of nutrient uptake in microalgae, including the relationship between the two, and the significance of carbon supply in nitrogen and phosphorus uptake by microalgae. This will enable development of sustainable processes and predictable strategies involving microalgae towards net-zero solutions, as well as nitrogen and phosphorus recovery from waste streams, This work was conducted as part of the SuBB project, supported by the Department of Biotechnology (DBT), Govt. of India (BT/IN/Indo-UK/SuBB/28/LU/2013) and the UKRI Biotechnology and Biological Sciences Research Council (UKRI BBSRC) (BB/K020633/1). This project has also received funding from the European Union’s Horizon 2020 research and innovation programme under the Marie Sklodowska-Curie grant agreement No 661063 to M.S.-N, and the Ministry of Education Malaysia and Universiti Malaysia Perlis (UniMAP) for SLTPA scholarship to FS. S.V. also acknowledges financial support from UKRI EPSRC (EP/E036252/1) to Sheffield. The authors also thank Department of Biotechnology Govt of India for sponsoring the NRMC – Marine repository (BT/PR29901/PBD/26/694/2018) and Mission Innovation #5 programme (BT/ PR31336/PBD/26/726/2019) in connection to the proposed study, With the institutional support of the ‘Severo Ochoa Centre of Excellence’ accreditation (CEX2019-000928-S)

The monogenic autoimmune disease Immunedysregulation polyendocrynopathy entheropathy X-linked syndrome (IPEX) has elucidated the essential function of the transcription factor FOXP3 and of thymic-derived regulatory T (Treg) cells in controlling autoimmunity. However, the presence of autoreactive T cells in IPEX remains undetermined, thus representing a crucial gap in understanding the origin of autoimmunity in a FOXP3 deficient immune system. Combining epigenetic analysis as a lineage marker of Treg identity and TCR sequencing to assess the self-reactive clones, we showed that IPEX patients have two pools of expanded autoreactive T cells. The first originates from the expansion of autoreactive effector T cells (Teff), likely due to loss of Treg suppressive function since it is absent in carrier mothers, in whom Treg cells are functional. The second pool originates, unexpectedly, from Treg cells which lose their phenotypic markers, including CD25 and FOXP3. We call these loss of identity Treg cells and show that they are i) suppressed by healthy donor Treg in a patient post hematopoietic transplantation despite low donor chimerism, and ii) not detectable in patients with Autoimmune polyendocrinopathy-candidiasis-ectodermal dystrophy syndrome (APECED), a monogenic autoimmune disease of thymic origin. Moreover, we demonstrate that FOXP3 knock-out in Treg cells leads to increased Treg expansion and production of Th17 and Th2 cytokines, known to be increased in IPEX patients. These results suggest that the loss of identity Treg cells could directly contribute to immune dysregulation in IPEX. Collectively, we provide a better understanding of autoimmunity and novel ways to monitor the effects of Treg cell therapies in IPEX disease or other autoimmune diseases.One Sentence SummaryMutations of FOXP3 gene in humans cause expansion of autoreactive T cells originating from both effector T cells and regulatory T cells which gain effector function.

Forkhead box protein 3 (FOXP3) is the master transcription factor in CD4We sought to study the type and extent of immunologic abnormalities that remain ill-defined in IPEX, across genetic and clinical heterogeneity.We performed Treg-cell-specific epigenetic quantification and immunologic characterization of severe "typical" (n = 6) and "atypical" or asymptomatic (n = 9) patients with IPEX.Increased number of cells with Treg-cell-Specific Demethylated Region demethylation in FOXP3 is a consistent feature in patients with IPEX, with (1) highest values in those with typical IPEX, (2) increased values in subjects with pathogenic FOXP3 but still no symptoms, and (3) gradual increase over the course of disease progression. Large-scale profiling using Luminex identified plasma inflammatory signature of macrophage activation and TElevated TSDR-demethylated cells, combined with elevation of plasmatic and cellular markers of a polarized type 2 inflammatory immune response, extends our understanding of IPEX diagnosis and heterogeneity.

Patients who fail to adequately reconstitute the donor-derived immune system after allogeneic hematopoietic stem cell transplantation (HSCT) are at increased risk for infections and leukemia relapse. In the past, pan T-cell depleted haploidentical grafts were associated with delayed immune reconstitution (IR). Recently, the majority of patients receiving αβ T-cell/B-cell depleted haploidentical HSCT (αβhaplo-HSCT) reach a threshold of 200 CD3+ T cells/mcl by 100 days after HSCT (Bertaina A et al. Blood 2014 Jul 31;124(5):822-6). However, a proportion of patients experience a slower IR with consequent higher morbidity and mortality. Early prediction of delayed IR may permit prompt clinical intervention such as infusion of donor lymphocytes or of virus-specific cytotoxic T cells. Flow cytometry, the most widely applied approach for IR analysis, suffers from intrinsic limitations, such as high lymphocyte number requirement, degradation of samples, and insufficient standardization due to technical and operator variability. To overcome these limitations, we used a DNA methylation-based quantitative PCR that detects the epigenetic signature of different peripheral blood immune cell subsets (epigenetic quantification). This technique provides relative and absolute immune cell counts applicable to fresh, frozen, or paper-spotted dried blood. Epigenetic measurements are based on a per cell DNA copy number and provide a clear positive or negative signal rather than arbitrarily defined thresholds for "positivity" as in flow cytometry. We hypothesize that epigenetic quantification at day 15 after αβhaplo-HSCT could predict flow-based IR at day 100. Patients were consented at Lucile Packard Children's Hospital (Stanford, CA). Blood was collected between days 10-17 for epigenetic quantification and days 82-124 for flow cytometry. Bisulfite treated DNA underwent qPCR quantification of cell type-specific DNA regions of de-methylation (Baron U et al. Sci Transl Med 2018 Aug 1;10(452):eaan3508). Flow cytometry was performed using directly conjugated antibodies. Absolute cell counts were determined, plotted, and then analyzed using a linear regression model. In the first 5 αβhaplo-HSCT patients evaluated, we found a direct correlation between the epigenetic quantification at day 15 and flow cytometry at day 100 for CD4+ T cells (P=0.01), while the early epigenetic quantification of CD3+ and CD8+ T cells was not informative (Fig. 1). Preliminary data suggest that the use of epigenetic quantification early after αβhaplo-HSCT can predict the IR of CD4+ T cells at day 100 in αβhaplo-HSCT recipients. Ongoing analysis on a larger cohort of both αβhaplo-HSCT and unmanipulated HSCT recipients, will confirm if epigenetic quantification results obtained early post-HSCT can be used as a clinical biomarker of delayed IR and guide physicians in the use of post-HSCT adoptive immunotherapy.

This study reports the surface interaction of the chemically modified marine unicellular cyanobacterium Synechococcus elongatus BDU130911 with uranium. The selective functional groups of the control (dead biomass) for binding with uranium in unicellular marine cyanobacteria were identified as carboxyl groups. The adsorption capacity of the biomass in a 1 mM uranium solution was found to be 92% in the control, 85% in the amine-blocked treatments, and 20% in the carboxyl-blocked treatments. The Langmuir isotherm provided a good fit to the data, suggesting a monolayer of uranium adsorption on all the tested biomass. The functional groups involved in the adsorption of uranium by the control and modified biomass were assessed by Fourier transform infrared spectroscopy, energy dispersive X-ray fluorescence and X-ray diffractive analysis. The results of this study identify, carboxyl groups as the dominant anionic functional group involved in uranium adsorption, which validates an ionic interaction between the biomass and uranium, a cationic metal.

SynRio is a Shiny and R based web analysis portal for viewing Synechocystis PCC 6803 genome, a cyanobacterial genome with data analysis capabilities. The web based user interface is created using R programming language powered by Shiny package. This web interface helps in creating interactive genome visualization based on user provided data selection along with selective data download options. Availability SinRio is available to download freely from Github - https://github.com/NFMC/SynRio or from http://www.nfmc.res.in/synrio/. In addition an online version of the platform is also hosted at nfmc.res.in/synrio, using shiny server (open source edition) installation.

This study reports the surface interaction of the chemically modified marine unicellular cyanobacterium Synechococcus elongatus BDU130911 with uranium. The selective functional groups of the control (dead biomass) for binding with uranium in unicellular marine cyanobacteria were identified as carboxyl groups. The adsorption capacity of the biomass in a 1 mM uranium solution was found to be 92% in the control, 85% in the amine-blocked treatments, and 20% in the carboxyl-blocked treatments. The Langmuir isotherm provided a good fit to the data, suggesting a monolayer of uranium adsorption on all the tested biomass. The functional groups involved in the adsorption of uranium by the control and modified biomass were assessed by Fourier transform infrared spectroscopy, energy dispersive X-ray fluorescence and X-ray diffractive analysis. The results of this study identify, carboxyl groups as the dominant anionic functional group involved in uranium adsorption, which validates an ionic interaction between the biomass and uranium, a cationic metal.

Cyanobacterial diazotrophs play a significant role in environmental nitrogen economy despite their habitat either tropical or polar. However, the phenomenon by which it copes with temperature induced stress is poorly understood. Temperature response study of psychrophilic and mesophilic Nostoc strains explores their adaptive mechanisms. The selected psychrophilic and mesophilic strains were confirmed as Nostoc punctiforme and Nostoc calcicola respectively, by ultrastructure and 16S rDNA phylogeny. The psychrophilic strain has extensive glycolipid and polysaccharide sheath along with characteristic deposition of cyanophycin, polyhydroxybutyrate granules, and carboxysomes. This is possibly an adaptive strategy exhibited to withstand the freezing temperature and high intense of ultraviolet rays. The biomass measured in terms of dry weight, protein, and chlorophyll indicated a temperature dependant shift in both the psychrophilic and mesophilic strains and attained maximum growth in their respective temperature niches. At low temperature, psychrophilic organism exhibited nitrogenase activity, while mesophilic strains did not. The maximum glutamine synthetase activity was observed at 4 °C for psychrophilic and 37 °C for mesophilic strains. Activity at 4 °C in psychrophilic strains revealed their energetic mechanism even at low temperature. The nitrate and nitrite reductase of both psychrophilic and mesophilic strains showed maximum activity at 37 °C denoting their similar nitrogen assimilating mechanisms for combined nitrogen utilization. The activity studies of nitrogen fixation/assimilation enzymes have differential effects at varying temperatures, which provide valuable insights of physiological contribution and role of Nostoc strains in the biological nitrogen cycle.

The marine cyanobacterium Phormidium valderianum BDU 140441 exhibited the ability to grow at 0.25 mM tannic acid, a known hindering chemical for microbial growth. The tannic acid-degrading ability of the organism is evident from the UV–visible absorption spectrum. In addition, the existence of tannase has been localized by activity staining, and its induction in activity upon tannic acid exposure was confirmed in native gel. The critical tannic acid metabolization enzymes tested for are polyphenol oxidase and esterases; both are well known for tannic acid degradation. Upon tannic acid exposure, increased activity of polyphenol oxidase and expression of few new isoforms of esterase were identified by activity staining.

The marine cyanobacterium Phormidium valderianum BDU140441 was tested for its capability of decolorizing Poly R-478 in the presence of copper. The rate of dye decolorization was decreased when the concentration of copper was increased in the medium, whereas copper in low amount (10 μM) enhanced the decolorization. To analyse the fate of dye-decolorizing and stress-stabilizing enzymes upon copper and dye exposure, four different conditions were maintained. The conditions were (A) only the organism P. valderianum BDU140441, (B) organism + Poly R-478 (0.0075%), (C) organism + copper (10 μM) and (D) organism + Poly R-478 (0.0075%) + copper (10 μM). After 16 h of exposure to Poly R-478 and copper, an increased trend in the activity of laccase and peroxidase was observed. In contrast, polyphenol oxidase revealed only a mild increase in activity along with the induction of a new isoform. The tested cellular antioxidants carotenoid and reduced glutathione were found to be reduced in varying percentage in accordance to the degree of stress bared by the cyanobacterium. Dye and copper exposure divulged an increased pattern in the production of reactive oxygen species (ROS). Concomitant with ROS, esterase activity was also found to increase. Towards copper and dye exposure, the activity of the primary stress-stabilizing enzymes catalase and superoxide dismutase were found to be increased in varied degrees. Upon copper exposure glutathione S-transferase and glutathione peroxidase expressed new isoforms. The results suggest that copper at this juncture played a role as inducer for dye-decolorizing copper oxidases and subjugated the antioxidant system to the objective of Poly R-478 decolorization.

The various marine cyanobacterial strains tested showed wide variation in growth patterns and decolourization patterns of the lignin model polymeric dye Poly R-478. The study revealed the presence of laccases (LACs) and polyphenol oxidases (PPOs) in marine cyanobacteria. All the ten tested strains were found to possess constitutive PPOs, whereas only four strains showed the presence of constitutive laccases. Within 7 days of incubation the highest percentage of decolourization was shown by Phormidium valderianum BDU140441 (65%), and Oscillatoria chlorina BDU 140691 (12%) showed the least. Isoforms of LACs were found to be induced by the laccase elicitors veratryl aldehyde, caffeic acid, guaiacol and tannic acid. Cyanobacterial strains that possess both LACs and PPOs were relatively more efficient in decolourizing the dye. Altering the concentrations of nitrogen, phosphorus, potassium and sulphur from the basal medium influenced the efficiency of dye decolourization.

Cyanobacterial KnowledgeBase (CKB) is a free access database that contains the genomic and proteomic information of 74 fully sequenced cyanobacterial genomes belonging to seven orders. The database also contains tools for sequence analysis. The Species report and the gene report provide details about each species and gene (including sequence features and gene ontology annotations) respectively. The database also includes cyanoBLAST, an advanced tool that facilitates comparative analysis, among cyanobacterial genomes and genomes of E. coli (prokaryote) and Arabidopsis (eukaryote). The database is developed and maintained by the Sub-Distributed Informatics Centre (sponsored by the Department of Biotechnology, Govt. of India) of the National Facility for Marine Cyanobacteria, a facility dedicated to marine cyanobacterial research. CKB is freely available at http://nfmc.res.in/ckb/index.html.