Optimizing light regimes for neutral lipid accumulation in Dunaliella salina MCC 43: a study on physiological status and carbon allocation.

Dunaliella salina is a favourable source of high lipid feedstock for biofuel and medicinal chemicals. Low biomass output from microalgae is a significant barrier to industrial-scale commercialisation. The current study aimed to determine how photosynthetic efficiency, carbon fixation, macromolecular synthesis, accumulation of neutral lipids, and antioxidative defence (ROS scavenging enzyme activities) of D. salina cells were affected by different light intensities (LI) (50, 100, 200, and 400 µmol m-2 s-1). The cells when exposed to strong light (400 µmol m-2 s-1) led to reduction in chlorophyll a but the carotenoid content increased by 19% in comparison to the control (LI 100). The amount of carbohydrate changed significantly under high light and in spite of stress inflicted on the cells by high irradiation, a considerable increase in activity of carbonic anhydrase and fixation rate of CO2 were recorded, thus, preserving the biomass content. The high light exposed biomass when subjected to nitrogen-deficient medium led to increase in lipid content (59.92% of the dry cell weight). However, neutral lipid made up 78.26% of the total lipid while other lipids like phospholipid and glycolipid content decreased, showing that the lipid was redistributed in these cells under nitrogen deprivation, making the organism more appropriate for biodiesel/jet fuel use. Although D. salina cells had a relatively longer generation time (3.5 d) than other microalgal cells, an economic analysis concluded that the amount of carotenoid they produced and the quality of their lipids made them more suited for commercialization.

A quest for universal anti-SARS-CoV-2 T cell assay: systematic review, meta-analysis, and experimental validation.

Measuring SARS-CoV-2-specific T cell responses is crucial to understanding an individual's immunity to COVID-19. However, high inter- and intra-assay variability make it difficult to define T cells as a correlate of protection against COVID-19. To address this, we performed systematic review and meta-analysis of 495 datasets from 94 original articles evaluating SARS-CoV-2-specific T cell responses using three assays - Activation Induced Marker (AIM), Intracellular Cytokine Staining (ICS), and Enzyme-Linked Immunospot (ELISPOT), and defined each assay's quantitative range. We validated these ranges using samples from 193 SARS-CoV-2-exposed individuals. Although IFNγ ELISPOT was the preferred assay, our experimental validation suggested that it under-represented the SARS-CoV-2-specific T cell repertoire. Our data indicate that a combination of AIM and ICS or FluoroSpot assay would better represent the frequency, polyfunctionality, and compartmentalization of the antigen-specific T cell responses. Taken together, our results contribute to defining the ranges of antigen-specific T cell assays and propose a choice of assay that can be employed to better understand the cellular immune response against viral diseases.

Trends of humoral immune responses to heterologous antigenic exposure due to vaccination & omicron SARS-CoV-2 infection: Implications for boosting.

Background & objectives: Vaccination and natural infection can both augment the immune responses against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), but how omicron infection has affected the vaccine-induced and hybrid immunity is not well studied in Indian population. The present study was aimed to assess the durability and change in responses of humoral immunity with age, prior natural infection, vaccine type and duration with a minimum gap of six months post-two doses with either ChAdOx1 nCov-19 or BBV152 prior- and post-emergence of the omicron variant. Methods: A total of 1300 participants were included in this observational study between November 2021 and May 2022. Participants had completed at least six months after vaccination (2 doses) with either ChAdOx1 nCoV-19 or an inactivated whole virus vaccine BBV152. They were grouped according to their age (≤ or ≥60 yr) and prior exposure of SARS-CoV-2 infection. Five hundred and sixteen of these participants were followed up after emergence of the Omicron variant. The main outcome was durability and augmentation of the humoral immune response as determined by anti-receptor-binding domain (RBD) immunoglobulin G (IgG) concentrations, anti-nucleocapsid antibodies and anti-omicron RBD antibodies. Live virus neutralization assay was conducted for neutralizing antibodies against four variants - ancestral, delta and omicron and omicron sublineage BA.5. Results: Before the omicron surge, serum anti-RBD IgG antibodies were detected in 87 per cent participants after a median gap of eight months from the second vaccine dose, with a median titre of 114 [interquartile range (IQR) 32, 302] BAU/ml. The levels increased to 594 (252, 1230) BAU/ml post-omicron surge (P<0.001) with 97 per cent participants having detectable antibodies, although only 40 had symptomatic infection during the omicron surge irrespective of vaccine type and previous history of infection. Those with prior natural infection and vaccination had higher anti-RBD IgG titre at baseline, which increased further [352 (IQR 131, 869) to 816 (IQR 383, 2001) BAU/ml] (P<0.001). The antibody levels remained elevated after a mean time gap of 10 months, although there was a decline of 41 per cent. The geometric mean titre was 452.54, 172.80, 83.1 and 76.99 against the ancestral, delta, omicron and omicron BA.5 variants in the live virus neutralization assay. Interpretation & conclusions: Anti-RBD IgG antibodies were detected in 85 per cent of participants after a median gap of eight months following the second vaccine dose. Omicron infection probably resulted in a substantial proportion of asymptomatic infection in the first four months in our study population and boosted the vaccine-induced humoral immune response, which declined but still remained durable over 10 months.

Ancestral SARS-CoV-2-Driven Antibody Repertoire Diversity in an Unvaccinated Individual Correlates with Expanded Neutralization Breadth.

Understanding the quality of immune repertoire triggered during natural infection can provide vital clues that form the basis for development of a humoral immune response in some individuals capable of broadly neutralizing pan-SARS-CoV-2 variants. In the present study, we report variations in neutralization potential against Omicron variants of two novel neutralizing monoclonal antibodies (MAbs), THSC20.HVTR11 and THSC20.HVTR55, isolated from an unvaccinated convalescent individual that represent distinct B cell lineage origins and epitope specificity compared to five MAbs we previously reported that were isolated from the same individual. In addition, we observed neutralization of Omicron variants by plasma antibodies obtained from this particular individual postvaccination with increased magnitude. Interestingly, this observation was found to be comparable with six additional individuals who initially were also infected with ancestral SARS-CoV-2 and then received vaccines, indicating that hybrid immunity can provide robust humoral immunity likely by antibody affinity maturation. Development of a distinct antigen-specific B cell repertoire capable of producing polyclonal antibodies with distinct affinity and specificities offers the highest probability of protecting against evolving SARS-CoV-2 variants. IMPORTANCE Development of robust neutralizing antibodies in SARS-CoV-2 convalescent individuals is known; however, it varies at the population level. We isolated monoclonal antibodies from an individual infected with ancestral SARS-CoV-2 in early 2020 that not only varied in their B cell lineage origin but also varied in their capability and potency to neutralize all the known variants of concern (VOCs) and currently circulating Omicron variants. This indicated establishment of unique lineages that contributed in forming a B cell repertoire in this particular individual immediately following infection, giving rise to diverse antibody responses that could complement each other in providing a broadly neutralizing polyclonal antibody response. Individuals who were able to produce polyclonal antibody responses with higher magnitude have a higher chance of being protected from evolving SARS-CoV-2 variants.

Inactivated whole-virion vaccine BBV152/Covaxin elicits robust cellular immune memory to SARS-CoV-2 and variants of concern.

BBV152 is a whole-virion inactivated vaccine based on the Asp614Gly variant. BBV152 is the first alum-imidazoquinolin-adjuvanted vaccine authorized for use in large populations. Here we characterized the magnitude, quality and persistence of cellular and humoral memory responses up to 6 months post vaccination. We report that the magnitude of vaccine-induced spike and nucleoprotein antibodies was comparable with that produced after infection. Receptor binding domain-specific antibodies declined against variants in the order of Alpha (B.1.1.7; 3-fold), Delta (B.1.617.2; 7-fold) and Beta (B.1.351; 10-fold). However, pseudovirus neutralizing antibodies declined up to 2-fold against the Delta followed by the Beta variant (1.7-fold). Vaccine-induced memory B cells were also affected by the Delta and Beta variants. The SARS-CoV-2-specific multicytokine-expressing CD4+ T cells were found in ~85% of vaccinated individuals. Only a ~1.3-fold reduction in efficacy was observed in CD4+ T cells against the Beta variant. We found that antigen-specific CD4+ T cells were present in the central memory compartment and persisted for at least up to 6 months post vaccination. Vaccine-induced CD8+ T cells were detected in ~50% of individuals. Importantly, the vaccine was capable of inducing follicular T helper cells that exhibited B-cell help potential. These findings show that inactivated vaccine BBV152 induces robust immune memory to SARS-CoV-2 and variants of concern that persists for at least 6 months after vaccination.

Hyperhemolysis in a patient with sickle cell disease and recent SARS-CoV-2 infection, with complex auto- and alloantibody work-up, successfully treated with tocilizumab.

BACKGROUND: Hyperhemolysis syndrome (HHS) is a severe delayed hemolytic transfusion reaction seen in sickle cell disease (SCD) patients, characterized by destruction of donor and recipient RBCs. It results in a drop in hemoglobin to below pretransfusion levels and frequently reticulocytopenia. CASE REPORT: We report a case of a man in his thirties with SCD with a recent hospitalization 2 weeks prior for COVID-19. His red cell antibody history included anti-Fy(a) and warm autoantibody. At that time, he was given 2 units of RBC and discharged with a hemoglobin of 10.2 g/dl. He returned to the hospital approximately 1.5 weeks later with hemoglobin 6.0 g/dl and symptoms concerning for acute chest syndrome. Pretransfusion testing now showed 4+ pan-agglutinin in both gel-based and tube-based testing. Alloadsorption identified an anti-N and a strong cold agglutinin. Three least incompatible units were transfused to this patient over several days, with evidence of hemolysis. Further reference lab work revealed anti-Fya , anti-Fyb , anti-Lea , anti-Leb , and an anti-KN system antibody. The patient's hemoglobin nadired at 4.4 g/dl. The patient was treated with a single dose of tocilizumab, his hemoglobin stabilized, and he was discharged. DISCUSSION: We present a case of HHS proximate to recent SARS-CoV-2 infection with multiple allo and autoantibodies identified. Information on the relationship between SARS-CoV-2 infection and HHS is limited; however, it is possible that inflammation related to COVID-19 could predispose to HHS. Tocilizumab is an approved treatment for COVID-19. Additionally, tocilizumab appears to be a promising treatment option for patients with HHS.

A combination of potently neutralizing monoclonal antibodies isolated from an Indian convalescent donor protects against the SARS-CoV-2 Delta variant.

Although efficacious vaccines have significantly reduced the morbidity and mortality of COVID-19, there remains an unmet medical need for treatment options, which monoclonal antibodies (mAbs) can potentially fill. This unmet need is exacerbated by the emergence and spread of SARS-CoV-2 variants of concern (VOCs) that have shown some resistance to vaccine responses. Here we report the isolation of five neutralizing mAbs from an Indian convalescent donor, out of which two (THSC20.HVTR04 and THSC20.HVTR26) showed potent neutralization of SARS-CoV-2 VOCs at picomolar concentrations, including the Delta variant (B.1.617.2). One of these (THSC20.HVTR26) also retained activity against the Omicron variant. These two mAbs target non-overlapping epitopes on the receptor-binding domain (RBD) of the spike protein and prevent virus attachment to its host receptor, human angiotensin converting enzyme-2 (hACE2). Furthermore, the mAb cocktail demonstrated protection against the Delta variant at low antibody doses when passively administered in the K18 hACE2 transgenic mice model, highlighting their potential as a cocktail for prophylactic and therapeutic applications. Developing the capacity to rapidly discover and develop mAbs effective against highly transmissible pathogens like coronaviruses at a local level, especially in a low- and middle-income country (LMIC) such as India, will enable prompt responses to future pandemics as an important component of global pandemic preparedness.

Sub-optimal neutralisation of omicron (B.1.1.529) variant by antibodies induced by vaccine alone or SARS-CoV-2 Infection plus vaccine (hybrid immunity) post 6-months.

BACKGROUND: Rapid spread of the omicron SARS-CoV-2 variant despite extensive vaccination suggests immune escape. The neutralising ability of different vaccines alone or with natural SARS-CoV-2 infection against omicron is not well-known. METHODS: In this cross-sectional study, we tested the ability of vaccine and natural infection induced antibodies to neutralise omicron variant in a live virus neutralisation assay in four groups of individuals: (i) ChAdOx1 nCoV-19 vaccination, (ii) ChAdOx1 nCoV-19 vaccination plus prior SARS-CoV-2 infection, (iii) vaccination with inactivated virus vaccine (BBV152), and (iv) BBV152 vaccination plus prior SARS-CoV-2 infection. Primary outcome was fold-change in virus neutralisation titre against omicron compared with ancestral virus. FINDINGS: We included 80 subjects. The geometric mean titre (GMT) of the 50% focus reduction neutralisation test (FRNT50) was 380·4 (95% CI: 221·1, 654·7) against the ancestral virus with BBV152 vaccination and 379·3 (95% CI: 185·6, 775·2) with ChAdOx1 nCov-19 vaccination alone. GMT for vaccination plus infection groups were 806·1 (95% CI: 478·5, 1357·8) and 1526·2 (95% CI: 853·2, 2730·0), respectively. Against omicron variant, only 5 out of 20 in both BBV152 and ChAdOx1 nCoV-19 vaccine only groups, 6 out of 20 in BBV152 plus prior SARS-CoV-2 infection group, and 9 out of 20 in ChAdOx1 nCoV-19 plus prior SARS-CoV-2 infection group exhibited neutralisation titres above the lower limit of quantification (1:20) suggesting better neutralisation with prior infection. A reduction of 26·6 and 25·7 fold in FRNT50 titres against Omicron compared to ancestral SARS-CoV-2 strain was observed for individuals without prior SARS-CoV-2 infection vaccinated with BBV152 and ChAdOx1 nCoV-19, respectively. The corresponding reduction was 57·1 and 58·1 fold, respectively, for vaccinated individuals with prior infection. The 50% neutralisation titre against omicron demonstrated moderate correlation with serum anti-RBD IgG levels [Spearman r: 0·58 (0·41, 0·71)]. INTERPRETATION: Significant reduction in the neutralising ability of both vaccine-induced and vaccine plus infection-induced antibodies was observed for omicron variant which might explain immune escape. FUNDING: Department of Biotechnology, India; Bill & Melinda Gates Foundation, USA.

Activation of epigenetic regulator KDM6B by Salmonella Typhimurium enables chronic infections.

Non-typhoidal Salmonella (NTS) infections result in self limiting gastroenteritis except in rare cases wherein manifestations of chronic infections can occur. Strategies employed by Salmonella to thrive in hostile environments of host during chronic infections are complex and multifaceted. In chronic state, a coordinated action of bacterial effectors allows reprogramming of macrophages to M2 subtype and thereby creating a permissible replicative niche. The mechanistic details of these processes are not fully known. In the current study we identified, histone H3-lysine 27 trimethylation (H3K27me3)-specific demethylase, KDM6B to be upregulated in both cell culture and in murine model of Salmonella infection. KDM6B recruitment upon infection exhibited an associated loss of overall H3K27me3 in host cells and was Salmonella SPI1 effectors coordinated. ChIP-qRT-PCR array analysis revealed several new gene promoter targets of KDM6B demethylase activity including PPARδ, a crucial regulator of fatty acid oxidation pathway and Salmonella-persistent infections. Furthermore, pharmacological inhibition of KDM6B demethylase activity with GSKJ4 in chronic Salmonella infection mice model led to a significant reduction in pathogen load and M2 macrophage polarization in peripheral lymphoid organs. The following work thus reveals Salmonella effector-mediated epigenetic reprogramming of macrophages responsible for its long-term survival and chronic carriage.

Non-neutralizing SARS CoV-2 directed polyclonal antibodies demonstrate cross-reactivity with the HA glycans of influenza virus.

The spike protein of the SARS-CoV-2 virus is the foremost target for the designing of vaccines and therapeutic antibodies and also acts as a crucial antigen in the assessment of COVID-19 immune responses. The enveloped viruses; such as SARS-CoV-2, Human Immunodeficiency Virus-1 (HIV-1) and influenza, often hijack host-cell glycosylation pathways and influence pathobiology and immune selection. These glycan motifs can lead to either immune evasion or viral neutralization by the production of cross-reactive antibodies that can lead to antibody-dependent enhancement (ADE) of infection. Potential cross-protection from influenza vaccine has also been reported in COVID-19 infected individuals in several epidemiological studies recently; however, the scientific basis for these observations remains elusive. Herein, we show that the anti-SARS-CoV2 antibodies cross-reacts with the Hemagglutinin (HA) protein. This phenomenon is common to both the sera from convalescent SARS-CoV-2 donors and spike immunized mice, although these antibodies were unable to cross-neutralize, suggesting the presence of a non-neutralizing antibody response. Epitope mapping suggests that the cross-reactive antibodies are targeted towards glycan epitopes of the SARS-CoV-2 spike and HA. Overall, our findings address the cross-reactive responses, although non-neutralizing, elicited against RNA viruses and warrant further studies to investigate whether such non-neutralizing antibody responses can contribute to effector functions such as antibody-dependent cellular cytotoxicity (ADCC) or ADE.

Longitudinal Serology of SARS-CoV-2-Infected Individuals in India: A Prospective Cohort Study.

Clinical and epidemiological characteristics of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) are now widely available, but there are few data regarding longitudinal serology in large cohorts, particularly those from low-income and middle-income countries. We established an ongoing prospective cohort of 3,840 SARS-CoV-2-positive individuals according to RT-PCR in the Delhi-National Capital Region of India to document clinical and immunological characteristics during illness and convalescence. The immunoglobulin G (IgG) responses to the receptor binding domain (RBD) and nucleocapsid were assessed at 0 to 7 days, 10 to 28 days, and 6 to 10 weeks after infection. The clinical predictors of seroconversion were identified by multivariable regression analysis. The seroconversion rates during the postinfection windows of 0 to 7 days, 10 to 28 days, and 6 to 10 weeks were 46%, 84.7%, and 85.3%, respectively (N = 743). The proportion with a serological response increased with the severity of coronavirus disease 2019 (COVID-19). All participants with severe disease, 89.6% with mild to moderate infection, and 77.3% of asymptomatic participants had IgG antibodies to the RBD antigen. The threshold values for the nasopharyngeal viral RNA RT-PCR of a subset of asymptomatic and symptomatic seroconverters were comparable (P = 0.48) to those of nonseroconverters (P = 0.16) (N = 169). This is the first report of longitudinal humoral immune responses to SARS-CoV-2 over a period of 10 weeks in South Asia. The low seropositivity of asymptomatic participants and differences between assays highlight the importance of contextualizing the understanding of population serosurveys.

Peroxidases from an invasive Mesquite species for management and restoration of fertility of phenolic-contaminated soil.

Phenolics drive the global economy, but they also pose threats to soil health and plant growth. Enzymes like peroxidase have the potential to remove the phenolic contaminants from the wastewater; however, their role in restoring soil health and improving plant growth has not yet been ascertained. We fractionated efficient peroxidases (MPx) from leaves of an invasive species of Mesquite, Prosopis juliflora, and demonstrated its superiority over horseradish peroxidase (HRP) in remediating phenol, 3-chlorophenol (3-CP), and a mixture of chlorophenols (CP-M), from contaminated soil. MPx removes phenolics over a broader range of pH (2.0-9.0) as compared with HRP (pH: 7.0-8.0). In soil, replacing H2O2 with CaO2 further increases the phenolic removal efficiency of MPx (≥90% of phenol, ≥ 70% of 3-CP, and ≥90% of CP-M). MPx maintains ~4-fold higher phenolic removal efficiency than purified HRP even in soils with extremely high contaminant concentration (2 g phenolics/kg of soil), which is desirable for environmental applications of enzymes for remediation. MPx treatment restores soil biological processes as evident by key enzymes of soil fertility viz. Acid- and alkaline-phosphatases, urease, and soil dehydrogenase, and improves potential biochemical fertility index of soil contaminated with phenolics. MPx treatment also assists the Vigna mungo test plant to overcome toxicant stress and grow healthy in contaminated soils. Optimization of MPx for application in the field environment would help both in the restoration of phenolic-contaminated soils and the management of invasive Mesquite.

Fibroblasts and Their Pathological Functions in the Fibrosis of Aortic Valve Sclerosis and Atherosclerosis.

Cardiovascular diseases, such as atherosclerosis and aortic valve sclerosis (AVS) are driven by inflammation induced by a variety of stimuli, including low-density lipoproteins (LDL), reactive oxygen species (ROS), infections, mechanical stress, and chemical insults. Fibrosis is the process of compensating for tissue injury caused by chronic inflammation. Fibrosis is initially beneficial and maintains extracellular homeostasis. However, in the case of AVS and atherosclerosis, persistently active resident fibroblasts, myofibroblasts, and smooth muscle cells (SMCs) perpetually remodel the extracellular matrix under the control of autocrine and paracrine signaling from the immune cells. Myofibroblasts also produce pro-fibrotic factors, such as transforming growth factor-ß1 (TGF-ß1), angiotensin II (Ang II), and interleukin-1 (IL-1), which allow them to assist in the activation and migration of resident immune cells. Post wound repair, these cells undergo apoptosis or become senescent; however, in the presence of unresolved inflammation and persistence signaling for myofibroblast activation, the tissue homeostasis is disturbed, leading to excessive extracellular matrix (ECM) secretion, disorganized ECM, and thickening of the affected tissue. Accumulating evidence suggests that diverse mechanisms drive fibrosis in cardiovascular pathologies, and it is crucial to understand the impact and contribution of the various mechanisms for the control of fibrosis before the onset of a severe pathological consequence.

Synthesis and Biological Evaluation of 8-Quinolinamines and Their Amino Acid Conjugates as Broad-Spectrum Anti-infectives.

In the search of therapeutic agents for emerging drug-resistant parasites, the synthesis of newer classes of 8-quinolinamines has emerged as a successful chemotherapeutic approach. We report synthesis of 8-quinolinamines bearing 5-alkoxy, 4-methyl, and 2-tert-butyl groups in the quinoline framework and their amino acid conjugates as broad-spectrum anti-infectives. 8-Quinolinamines exhibited potent in vitro antimalarial activity [IC50 = 20-4760 ng/mL (drug-sensitive Plasmodium falciparum D6 strain) and IC50 = 22-4760 ng/mL (drug-resistant P. falciparum W2 strain)]. The most promising analogues have cured all animals at 25 mg/kg/day against drug-sensitive Plasmodium berghei and at 50 mg/kg/day against multidrug-resistant Plasmodium yoelii nigeriensis infections in Swiss mice. The in vitro antileishmanial activities (IC50 = 0.84-5.0 µg/mL and IC90 = 1.95-7.0 µg/mL) comparable to standard drug pentamidine were exhibited by several of the synthesized 8-quinolinamines. At the same time, very promising antifungal activities (Candida albicans-IC50 = 4.93-19.38 µg/mL; Candida glabrata-IC50 = 3.96-19.22 µg/mL; Candida krusei-IC50 = 2.89-18.95 µg/mL; Cryptococcus neoformans-IC50 = 0.67-18.64 µg/mL; and Aspergillus fumigatus-IC50 = 6.0-19.32 µg/mL) and antibacterial activities (Staphylococcus aureus-IC50 = 1.33-18.9 µg/mL; methicillin-resistant S. aureus-IC50 = 1.38-15.34 µg/mL; and Mycobacterium intracellulare-IC50 = 3.12-20 µg/mL) were also observed. None of the 8-quinolinamines exhibited cytotoxicity and therefore are a promising structural class of compounds as antiparasitic and antimicrobials.

Halotolerant Exiguobacterium profundum PHM11 Tolerate Salinity by Accumulating L-Proline and Fine-Tuning Gene Expression Profiles of Related Metabolic Pathways.

Salinity stress is one of the serious factors, limiting production of major agricultural crops; especially, in sodic soils. A number of approaches are being applied to mitigate the salt-induced adverse effects in agricultural crops through implying different halotolerant microbes. In this aspect, a halotolerant, Exiguobacterium profundum PHM11 was evaluated under eight different salinity regimes; 100, 250, 500, 1000, 1500, 2000, 2500, and 3000 mM to know its inherent salt tolerance limits and salt-induced consequences affecting its natural metabolism. Based on the stoichiometric growth kinetics; 100 and 1500 mM concentrations were selected as optimal and minimal performance limits for PHM11. To know, how salt stress affects the expression profiles of regulatory genes of its key metabolic pathways, and total production of important metabolites; biomass, carotenoids, beta-carotene production, IAA and proline contents, and expression profiles of key genes affecting the protein folding, structural adaptations, transportation across the cell membrane, stress tolerance, carotenoids, IAA and mannitol production in PHM11 were studied under 100 and 1500 mM salinity. E. profundum PHM11 showed maximum and minimum growth, biomass and metabolite production at 100 and 1500 mM salinity respectively. Salt-induced fine-tuning of expression profiles of key genes of stress pathways was determined in halotolerant bacterium PHM11.

Warfarin alters vitamin K metabolism: a surprising mechanism of VKORC1 uncoupling necessitates an additional reductase.

The anticoagulant warfarin inhibits the vitamin K oxidoreductase (VKORC1), which generates vitamin K hydroquinone (KH2) required for the carboxylation and consequent activation of vitamin K-dependent (VKD) proteins. VKORC1 produces KH2 in 2 reactions: reduction of vitamin K epoxide (KO) to quinone (K), and then KH2 Our dissection of full reduction vs the individual reactions revealed a surprising mechanism of warfarin inhibition. Warfarin inhibition of KO to K reduction and carboxylation that requires full reduction were compared in wild-type VKORC1 or mutants (Y139H, Y139F) that cause warfarin resistance. Carboxylation was much more strongly inhibited (∼400-fold) than KO reduction (two- to threefold). The K to KH2 reaction was analyzed using low K concentrations that result from inhibition of KO to K. Carboxylation that required only K to KH2 reduction was inhibited much less than observed with the KO substrate that requires full VKORC1 reduction (eg, 2.5-fold vs 70-fold, respectively, in cells expressing wild-type VKORC1 and factor IX). The results indicate that warfarin uncouples the 2 reactions that fully reduce KO. Uncoupling was revealed because a second activity, a warfarin-resistant quinone reductase, was not present. In contrast, 293 cells expressing factor IX and this reductase activity showed much less inhibition of carboxylation. This activity therefore appears to cooperate with VKORC1 to accomplish full KO reduction. Cooperation during warfarin therapy would have significant consequences, as VKD proteins function in numerous physiologies in many tissues, but may be poorly carboxylated and dysfunctional if the second activity is not ubiquitously expressed similar to VKORC1.

Viscum articulatum Burm. f. aqueous extract exerts antiproliferative effect and induces cell cycle arrest and apoptosis in leukemia cells.

ETHNOPHARMACOLOGICAL RELEVANCE: Viscum articulatum Burm. f. (leafless mistletoe) has been used in traditional system of medicines in India, China, Taiwan, Cambodia, Laos, and Vietnam, to treat blood-related diseases and various inflammatory and degenerative diseases including cancer. Anticancer activities of some phytomolecules purified from Viscum articulatum Burm. f. have been tested. However scientific evidence for the anticancerous potential of aqueous extract of V. articularum (VAQE) used in traditional medicine is lacking. AIM OF THE STUDY: To study the antiproliferative and apoptotic effect of VAQE on Jurkat E6.1 and THP1 leukemia cells. MATERIALS AND METHODS: The aqueous extract of the whole plant of Viscum articulatum Burm. f. was prepared in phosphate buffer saline. In VAQE, total soluble protein was estimated using Bradford's dye-binding assay; flavonoid content was determined using aluminum chloride colorimetric assay; and phenolic content was estimated following Folin-Ciocalteu colorimetric assay. XTT cell viability assay was used to test VAQE induced cytotoxicity in Jurkat E6.1 and THP1 leukemia cells and peripheral blood mononuclear cells (PBMC). The effect of VAQE on cell cycle progression was analyzed by PI staining using flow cytometry. Annexin-V-FITC/PI differential staining method was used for detecting the onset of apoptosis in leukemia cells. Rhodamine 123 dye was used to detect the change in mitochondrial membrane potential (MMP) using flow cytometry. DCF-DA fluorescence dye was used to estimate the level of reactive oxygen species (ROS). The ROS inhibitors were used to evaluate the role of ROS in mediating DNA degradation in VAQE-treated leukemia cells. The molecular mechanisms underlying VAQE induced apoptosis induction was studied by analyzing the expression of anti-apoptotic (Bcl-2) and pro-apoptotic (Bax) proteins, caspase-8 and caspase-3 enzymes using western blot. Diphenylamine (DPA) assay was used to determine the DNA fragmentation and conclusion of apoptosis. RESULTS: VAQE triggered cytotoxic effect on Jurkat E6.1 (IC50-2.4 µg/ml; 24 h) and THP1 (IC50-1.0 µg/ml; 24 h) cells in a dose- and time-dependent manner. The apoptosis induction and G2/M arrest of the cell cycle are the cause of VAQE-induced cytotoxicity in leukemia cells. The apoptosis in VAQE-treated Jurkat E6.1 and THP1 cells was mediated via a reduction in MMP, elevation of intracellular ROS, decreased expression of the anti-apoptotic (Bcl-2) and increased expression of the pro-apoptotic (Bax) protein, activation of caspase-8 and caspase-3 and DNA fragmentation. CONCLUSION: VAQE has a high efficacy to exert a cytotoxic effect in Jurkat E6.1 and THP1 cells and to induce apoptosis and G2/M cell cycle arrest. VAQE induces extrinsic pathway of apoptosis in both the leukemia cell lines via disruption of MMP, intracellular ROS imbalance, increased ratio of Bax/Bcl-2, activation of caspase-8, caspase-3 and ROS-mediated DNA fragmentation. The knowledge gained from the outcomes of the study may encourage the identification of novel chemotherapeutic agent from Viscum articulatum Burm. f. to treat leukemia.

Guanylthiourea derivatives as potential antimalarial agents: Synthesis, in vivo and molecular modelling studies.

Guanylthiourea (GTU) derivatives were identified as possible anti-malarial agents, recently, using in vitro studies on Plasmodium falciparum. This article gives an account of the in vivo anti-malarial activity of GTU derivatives against experimental rodent malaria. A total of 20 synthesized GTU derivatives were evaluated for in vivo antimalarial activity, out of which six showed encouraging results; one compound appeared to have curative potential. Molecular docking and molecular dynamics analysis were carried out to understand the molecular level interactions.

Racial disparities: disruptive genes in prostate carcinogenesis.

Population specific studies in prostate cancer (PCa) reveal a unique heterogeneous etiology. Various factors, such as genetics, environment and dietary regimen seems to determine disease progression, therapeutic resistance and rate of mortality. Enormous disparity documented in disease incidences, aggressiveness and mortality in PCa among AAs (African Americans) and CAs (Caucasian Americans) is attributed to the variations in genetics, epigenetics and their association with metabolism. Scientific and clinical evidences have revealed the influence of variations in Androgen Receptor (AR), RNAse L, macrophage scavenger receptor 1 (MRS1), androgen metabolism by cytochrome P450 3A4, differential regulation of microRNAs, epigenetic alterations and diet in racial disparity in PCa incidences and mortality. Concerted efforts are needed to identify race specific prognostic markers and treatment regimen for a better management of the disease.

Phenol remediation by peroxidase from an invasive mesquite: Turning an environmental wound into wisdom.

The present study examines mesquite (Prosopis juliflora), an invasive species, to yield peroxidase that may reduce hazards of phenolics to living organisms. As low as 0.3U of low-purity mesquite peroxidase (MPx) efficiently remove phenol and chlorophenols (90-92%) compared with Horseradish peroxidase (HRP) (40-60%). MPx shows a very high removal efficiency (40-50%) at a wide range of pH (2-9) and temperature (20-80°C), as opposed to HRP (15-20%). At a high-level of the substrate (2.4mM) and without the addition of PEG, MPx maintains a significant phenolic removal (60-≥92%) and residual activity (∼25%). It proves the superiority of MPx over HRP, which showed insignificant removal (10-12%) under similar conditions, and no residual activity even with PEG addition. The root elongation and plant growth bioassays confirm phenolic detoxification by MPx. Readily availability of mesquite across the countries and easy preparation of MPx from leaves make this tree as a sustainable source for a low-technological solution for phenol remediation. This study is the first step towards converting a biological wound of invasive species into wisdom and strength for protecting the environment from phenol pollution.