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SARS-CoV-2 Omicron subvariants such as BA.2.12.1, BA.4 and BA.5 have been spreading rapidly and become dominant worldwide. Here we report the homologous or heterologous booster effects of S-268019-b, a recombinant spike protein vaccine with the squalene-based adjuvant A-910823 in cynomolgus macaques. In macaques which had been primed with S-268019-b or mRNA vaccines, boosting with S-268019-b enhanced neutralizing antibodies (NAb) against ancestral SARS-CoV-2. Since boosting with the antigen without adjuvant did not efficiently restore NAb titers, adjuvant A-910823 was essential for the booster effect. Importantly, boosting with S-268019-b enhanced NAb against all of the Omicron subvariants we tested, including BA.2.12.1, BA.4 and BA.5, in comparison to two vaccine doses. Additionally, expansion of Omicron-specific B cells was confirmed after boosting with S-268019-b. These results indicate that a booster dose of S-268019-b with the adjuvant enhances the neutralization breadth.

Immunogenic cell death (ICD) plays an important role in sensitizing tumor cells to antigen-presenting cells followed by antitumor immunity. However, a successful antitumor response by ICD requires both apoptotic tumor microenvironments and activated immune systems. Ivermectin (IVM) has been shown to induce cell apoptosis through autophagy which can be a great candidate for ICD therapy. However, a single treatment of IVM-free drug is not an ideal anticancer therapy due to its anti-inflammatory effects and cytotoxicity. In the present study, IVM was shown to enhance the ICD process in addition to the treatment of resiquimod (R848), a TLR7/8 agonist, when co-loaded in a squalene-based nanoemulsion (NE). R848-IVM co-loaded NE was developed and characterized in vitro. Antitumor activity of R848-IVM NE was also evaluated in vitro and in vivo. R848-IVM NE exhibited long-term stability and reduced cytotoxicity by IVM. In vivo studies demonstrated that IVM significantly augments the ICD by upregulating Cd8a and releasing HMGB1 in tumor tissue, which could enhance R848-driven antitumor immunity. R848-IVM NE treatment showed strong antitumor activity with over 80% tumor growth inhibition, suggesting a potential combination therapy of systemic co-delivering IVM with TLR agonists against solid cancer.

The object of the study was oilseeds of winter camelina grown in regions with contrasting climatic conditions: the forest-steppe of the Middle Volga region (Penza Research Institute of Agriculture) and the steppe zone of Crimea (Research Institute of Agriculture of Crimea). The aim of the study is to determine the biochemical parameters of winter camelina, to study its fatty acid and amino acid composition, depending on the region of cultivation. The climate of the Middle Volga region is temperate continental. The amount of annual precipitation varies from 350 to 750 mm, the average annual temperature is 5.3°С. The climate of the steppe Crimea is continental, the average annual temperature reaches 10.2°С, the amount of precipitation per year is 350–450 mm. The largest share in camelina seed oil is linolenic acid, the percentage of which was 33.79 (Crimea) and 34.87 (Penza). The concentration of linoleic and oleic acids is 16.12–17.98 and 14.82–15.70%, depending on the growing region. The content of palmitic acid is 4.93 and 5.17%, eicosene – 15.46 and 15.84%, erucic 3.15 and 3.16%. The ratio of linolenic (ω-3) and linoleic (ω-6) acids is on average 2:1. In the seeds of camelina, 18 amino acids were identified, including 9 irreplaceable ones, the content of which was 7.28 and 8.53%. Camelina proteins contain a high amount of leucine (2.14–3.02%), valine (1.01–1.25%), lysine (1.07–1.28%), proline (1.02–1.45%), glycine (1.13–1.46%), alanine (0.93–1.22%), serine (1.02–1.26%) and methionine (0.34–0.37%). It was found that camelina oilseeds contain tocopherols 0.085-0.098%, carotenoids 0.002%, steroids 0.144–0.145% and squalene 0.09–0.10%, depending on the region., Объектом исследования являлись маслосемена рыжика озимого, выращенного в контрастных по климатическим условиям регионах: лесостепи Среднего Поволжья (Пензенский НИИСХ) и степной зоне Крыма (НИИСХ Крыма). Цель исследования – определение биохимических показателей рыжика озимого, изучение его жирнокислотного иаминокислотного состава в зависимости от региона возделывания. Климат Средневолжского региона умеренно континентальный. Сумма годовых осадков варьирует от 350 до 750мм, среднегодовая температура составляет 5.3°С. Климат степного Крыма континентальный, среднегодовая температура достигает 10.2°С, сумма осадков составляет 350–450 мм в год. Наибольшую долю в масле семян рыжика представляет линоленовая кислота, процент которой составил 33.79 (Крым) и 34.87 (Пенза). Концентрация линолевой и олеиновой кислот составляет 16.12–17.98% и 14.82–15.70% в зависимости от региона выращивания. Содержание пальмитиновой кислоты составляет 4.93 и 5.17%, эйкозеновой – 15.46 и 15.84%, эруковой – 3.15 и 3.16%. Соотношение линоленовой (ω-3) и линолевой (ω-6) кислот составляет в среднем 2:1. В семенах рыжика было выявлено 18 аминокислот, в том числе 9 незаменимых, содержание которых составило 7.28 и 8.53%. Белки рыжика содержат высокое количество лейцина (2.14–3.02%), валина (1.01–1.25%), лизина (1.07–1.28%), пролина (1.02–1.45%), глицина (1.13–1.46%), аланина (0.93–1.22%), серина (1.02–1.26%) и метионина (0.34–0.37%). Установлено, что маслосемена рыжика содержат токоферолы – 0.085–0.098%, каротиноиды – 0.002%, стероиды – 0.144–0.145% и сквален – 0.09–0.10% в зависимости от региона.

This critical review describes the squalene-ozone (SqOz) reaction, or squalene ozonolysis. Ambient ozone penetrates indoors and drives indoor air chemistry. Squalene, a component of human skin oil, contains six carbon-carbon double bonds and is very reactive with ozone. Bioeffluents from people contribute to indoor air chemistry and affect the indoor air quality, resulting in exposures because people spend the majority of their time indoors. The SqOz reaction proceeds through various formation pathways and produces compounds that include aldehydes, ketones, carboxylic acids, and dicarbonyl species, which have a range of volatilities. In this critical review of SqOz chemistry, information on the mechanism of reaction, reaction probability, rate constants, and reaction kinetics are compiled. Characterizations of SqOz reaction products have been done in laboratory experiments and real-world settings. The effect of multiple environmental parameters (ozone concentration, air exchange rate (AER), temperature, and relative humidity (RH)) in indoor settings are summarized. This critical review concludes by identifying the paucity of available exposure, health, and toxicological data for known reaction products. Key knowledge gaps about SqOz reactions leading to indoor exposures and adverse health outcomes are provided as well as an outlook on where the field is headed.

Transplantation of pancreatic islets is a promising approach to controlling glucose levels in type 1 diabetes mellitus (T1DM), but islet survival is still limited. To overcome this, islet co-culture with mesenchymal stromal cells (MSCs) together with safe immunosuppressive agents like squalene-gusperimus nanoparticles (Sq-GusNPs) may be applied. This could support islet survival and engraftment. Here, we studied how Sq-GusNPs and adipose-derived stem cells (ASCs) influence islets response under pro-inflammatory conditions. Through qRT-PCR, we studied the expression of specific genes at 24 hours in human and rat islets and ASCs in co-culture under indirect contact with or without treatment with Sq-GusNPs. We characterized how the response of islets and ASCs starts at molecular level before impaired viability or function is observed and how this response differs between species. Human islets and ASCs responses showed to be principally influenced by NF-kappa B activation, whereas rat islet and ASCs responses showed to be principally mediated by nitrosative stress. Rat islets showed tolerance to inflammatory conditions due to IL-1Ra secretion which was also observed in rat ASCs. Human islets induced the expression of cytokines and chemokines with pro-angiogenic, tissue repair, and anti-apoptotic properties in human ASCs under basal conditions. This expression was not inhibited by Sq-GusNPs. Our results showed a clear difference in the response elicited by human and rat islets and ASCs in front of an inflammatory stimulus and Sq-GusNPs. Our data support the use of ASCs and Sq-GusNP to facilitate engraftment of islets for T1DM treatment.

All known triterpenes are generated by triterpene synthases (TrTSs) from squalene or oxidosqualene1. This approach is fundamentally different from the biosynthesis of short-chain (C10–C25) terpenes that are formed from polyisoprenyl diphosphates2–4. In this study, two fungal chimeric class I TrTSs, Talaromyces verruculosus talaropentaene synthase (TvTS) and Macrophomina phaseolina macrophomene synthase (MpMS), were characterized. Both enzymes use dimethylallyl diphosphate and isopentenyl diphosphate or hexaprenyl diphosphate as substrates, representing the first examples, to our knowledge, of non-squalene-dependent triterpene biosynthesis. The cyclization mechanisms of TvTS and MpMS and the absolute configurations of their products were investigated in isotopic labelling experiments. Structural analyses of the terpene cyclase domain of TvTS and full-length MpMS provide detailed insights into their catalytic mechanisms. An AlphaFold2-based screening platform was developed to mine a third TrTS, Colletotrichum gloeosporioides colleterpenol synthase (CgCS). Our findings identify a new enzymatic mechanism for the biosynthesis of triterpenes and enhance understanding of terpene biosynthesis in nature.

Triterpenes are C30 organic compounds abundantly found in all living organisms. Although previously believed to be exclusively produced from squalene or oxidosqualene, a recent report by Tao and colleagues describes a new triterpene biosynthetic route involving the cyclization of the precursor hexaprenyl diphosphate (HexPP) by unprecedented bifunctional terpene synthase (TS) enzymes.

Mevalonate (MVA) pathway is the core for terpene and sterol biosynthesis, whose metabolic flux influences the synthesis efficiency of such compounds. Saccharomyces cerevisiae is an attractive chassis for the native active MVA pathway. Here, the truncated form of Enterococcus faecalis MvaE with only 3-Hydroxy-3-methylglutaryl coenzyme A reductase (HMGR) activity was found to be the most effective enzyme for MVA pathway flux using squalene as the metabolic marker, resulting in 431-fold and 9-fold increases of squalene content in haploid and industrial yeast strains respectively. Furthermore, a positive correlation between MVA metabolic flux and β-alanine metabolic activity was found based on a metabolomic analysis. An industrial strain SQ3-4 with high MVA metabolic flux was constructed by combined engineering HMGR activity, NADPH regeneration, cytosolic acetyl-CoA supply and β-alanine metabolism. The strain was further evaluated as the chassis for terpenoids production. Strain SQ3-4-CPS generated from expressing β-caryophyllene synthase in SQ3-4 produced 11.86 ± 0.09 mg l

There is an urgent need for improved influenza vaccines especially for older adults due to the presence of immunosenescence. It is therefore highly relevant to compare enhanced influenza vaccines with traditional influenza vaccines with respect to their effectiveness.To compare vaccine efficacy and effectiveness of adjuvanted influenza vaccines (aTIV/aQIV) vs. non-adjuvanted standard-dose (TIV/QIV) and high-dose (TIV-HD/QIV-HD) influenza vaccines regarding influenza-related outcomes in older adults, complementing findings from the European Centre for Disease Prevention and Control (ECDC)'s systematic review of enhanced seasonal influenza vaccines from February 2020.A systematic literature search was conducted in Embase and MEDLINE to identify randomised controlled trials, observational studies and systematic reviews, published since ECDC's systematic review (between 7 February 2020 and 6 September 2021). Included studies were appraised with either the Cochrane Risk of Bias tool, ROBINS-I or AMSTAR 2.Eleven analyses from nine real-world evidence (RWE) studies comprising ∼53 million participants and assessing the relative vaccine effectiveness (rVE) of aTIV vs. TIV, QIV and/or TIV-HD in adults aged ≥65 years over the 2006/07-2008/09 and 2011/12-2019/20 influenza seasons were identified. Nine analyses found that aTIV was significantly more effective than TIV and QIV in reducing influenza-related outcomes by clinical setting and suspected influenza outbreaks (rVE ranging from 7.5% to 25.6% for aTIV vs. TIV and 7.1% to 36.3% for aTIV vs. QIV). Seven analyses found similar effectiveness of aTIV vs. TIV-HD in reducing influenza-related medical encounters, inpatient stays and hospitalisations/emergency room visits. In three analyses, aTIV was significantly more effective than TIV-HD in reducing influenza-related medical encounters and office visits (rVE ranging from 6.6% to 16.6%). Risk of bias of identified studies was moderate to high.Our study suggests that both adjuvanted and high-dose vaccines are effective alternatives for vaccination programmes in older adults and preferable over conventional standard-dose vaccines.

BACKGROUND: Influenza A/H7N9 viruses have pandemic potential. METHODS: We conducted an open-label, randomized, controlled trial of AS03-adjuvanted 2017 inactivated influenza A/H7N9 vaccine (H7N9 IIV) in healthy adults. Group 1 received H7N9 IIV and seasonal quadrivalent influenza vaccine (IIV4) simultaneously, followed by H7N9 IIV three weeks later. Group 2 received IIV4 alone and then two doses of H7N9 IIV at three-week intervals. Group 3 received one dose of IIV4. We used hemagglutination inhibition (HAI) and microneutralization (MN) assays to measure geometric mean titers and seroprotection (≥1:40 titer) to vaccine strains and monitored for safety. RESULTS: Among 149 subjects, seroprotection by HAI three weeks after H7N9 IIV dose 2 was 51% (95 %CI 37%-65%) for Group 1 and 40% (95 %CI 25%-56%) for Group 2. Seroprotection by MN at the same timepoint was 84% (95 %CI 72%-93%) for Group 1 and 74% (95 %CI 60%-86%) for Group 2. By 180 days after H7N9 IIV dose 2, seroprotection by HAI or MN was low for Groups 1 and 2. Responses measured by HAI and MN against each IIV4 strain three weeks after IIV4 vaccination were similar in all groups. Solicited local and systemic reactions were similar after a single vaccination, while those receiving simultaneous H7N9 and IIV4 had slightly more reactogenicity. There were no serious adverse events or medically-attended adverse events related to study product receipt. CONCLUSIONS: Adjuvanted H7N9 IIV was modestly immunogenic whether administered simultaneously or sequentially with IIV4, though responses declined by 180 days. IIV4 was immunogenic regardless of schedule. CLINICAL TRIALS REGISTRATION: NCT03318315

The cholesterol metabolism in humans can be indirectly reflected by measuring cholesterol metabolism marker levels. We aimed to investigate the association of cholesterol homeostasis markers on standard lipid profiling components in familial hypercholesteremia and hyperlipidemia patients. A total of 69 hyperlipidemia patients, 25 familial hypercholesteremia (FHC) patients, and 64 healthy controls were enrolled in this study. We performed routine testing of blood lipid water. Gas chromatography was used to determine the changes in the concentration of cholesterol synthesis (squalene, desmosterol, and lathosterol) and absorption markers (campesterol, sitosterol, and stigmasterol) in the blood. Baseline hyperlipidemia patients displayed significantly higher total cholesterol (TC), triglyceride (TG), and low-density lipoprotein cholesterol (LDL-C) levels in comparison to the control group, which was reflected in the increased levels of squalene, desmosterol, campesterol, and sitosterol observed ( P < 0.05 ) in the hyperlipidemia patients. The desmosterol, lathosterol, campesterol, stigmasterol, and sitosterol were statistically different in the FHC group than the hyperlipidemic group ( P < 0.05 ). The proportions of squalene/cholesterol, lathosterol/cholesterol, stigmasterol/cholesterol, and sitosterol/cholesterol in the FHC group were lower than those in the hyperlipidemic group; only desmosterol/cholesterol was higher than that in the hyperlipidemic group. Correlation studies between lipid metabolic factors showed that the proportion of moderate and strong correlations was much higher in the FHC group than in the other two groups (76.92% vs. 32.50% and 31.25%). Logistic regression analysis showed that the concentrations of glucose, LDL-C, lactosterol, and sitosterol were all independent risk factors for developing hyperlipidemia. This result was further confirmed by the ROC curve. These results indicated that the study of cholesterol synthesis and decomposition markers can serve as a reference index for related diseases caused by changes in its concentration.

Over 800 known carotenoids are synthesized from phytoene or 4,4'-diapophytoene (dehydrosqualene) characterized by three conjugated double bonds. In this paper, we report that carotenoid desaturase CrtN from Staphylococcus aureus and Methylomonas can accept oxidosqualene, which is the precursor for plant- or animal-type triterpenoids, yielding the yellow carotenoid pigments with 8, 9, or 10 conjugated double bonds. The resulting pathway is the second nonnatural route for carotenoid pigments and the first pathway for carotenoid pigments not biosynthesized via (diapo)phytoene.

Squalene, as an important terpenoid, is extensively used in the medicine and health care fields owing to its functions of anti-oxidation, blood lipid regulation and cancer prevention. The marine microalgae, Schizochytrium sp., which acts as an excellent strain with potential of high squalene production was selected as the starting strain. The overexpressed strain with sqs gene got the reduced biomass and lipid, while the squalene titer was increased by 79.6% ± 4.7% to 12.8 ± 0.2 mg/L. In order to further increase squalene production, the recombinant strain (HS strain) with sqs and hmgr gene co-overexpression was further constructed. The biomass and squalene titer of the HS strain were increased by 13.6% ± 1.2% and 88.8% ± 5.3%, respectively, which indicated the carbon flux of the mevalonate pathway was enhanced for squalene accumulation. Regarding the squalene synthesis is completely coupled with cell growth, fermentation strategy to prolong the logarithmic growth phase was conducive to improve squalene production. Under the condition of optimal composition and concentrated medium, the squalene titer of HS strain was 27.0 ± 1.3 mg/L, which was 2.0 times that of the basal medium condition (13.5 ± 0.4 mg/L). This study which combined the metabolic engineering and fermentation strategy provides a new strategy for squalene production in Schizochytrium sp. KEY POINTS: •The overexpression of sqs and hmgr genes promoted carbon metabolism for squalene. •The optimal and concentrated media can increase squalene yield.

Background Ginseng, officially known as Panax ginseng Meyer, has been traditionally used as a medicinal herb, particularly in Asia. Ginseng is propagated from seeds; however, seed germination is challenging, especially in its natural environment on farms. The seeds typically exhibit morphophysiological dormancy and require release from both morphological and physiological dormancy before germination. Although some studies have proposed methods for increasing seed germination rates, the underlying mechanisms of its dormancy release process remain unclear. Here, we investigated metabolic alterations during dehiscence in P. ginseng to determine their potential roles in dormancy release. Methods We compared the ginseng seed metabolome before and after dehiscence and the ginsenoside and phytosterol compositions of the seeds in both periods in the presence of related enzymes. Results After seed dehiscence, the sugar, amino acid, and squalene concentrations were significantly altered, phytosterols associated with the stigmasterol biosynthesis pathway were increased, while ginsenoside and brassinosteroid levels were not significantly altered. In addition, squalene epoxidase, cycloartenol synthase, 24-methylenesterol C-methyltransferase, and the stigmasterol biosynthesis pathway were activated. Conclusion Overall, our findings suggest that morphological activities that facilitate ginseng seed growth are the primary phenomena occurring during the dehiscence process. This study improves the understanding of P. ginseng germination processes and promotes further research of its germination and cultivation.

Although antivirals are important tools to control severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection, effective vaccines are essential to control the current coronavirus disease 2019 (COVID-19) pandemic. Plant-derived virus-like particle (VLP) vaccine candidates have previously demonstrated immunogenicity and efficacy against influenza. Here, we report the immunogenicity and protection induced in rhesus macaques by intramuscular injections of a VLP bearing a SARS-CoV-2 spike protein (CoVLP) vaccine candidate formulated with or without Adjuvant System 03 (AS03) or cytidine-phospho-guanosine (CpG) 1018. Although a single dose of the unadjuvanted CoVLP vaccine candidate stimulated humoral and cell-mediated immune responses, booster immunization (at 28 days after priming) and adjuvant administration significantly improved both responses, with higher immunogenicity and protection provided by the AS03-adjuvanted CoVLP. Fifteen micrograms of CoVLP adjuvanted with AS03 induced a polyfunctional interleukin-2 (IL-2)-driven response and IL-4 expression in CD4 T cells. Animals were challenged by multiple routes (i.e., intratracheal, intranasal, and ocular) with a total viral dose of 106 plaque-forming units of SARS-CoV-2. Lower viral replication in nasal swabs and bronchoalveolar lavage fluid (BALF) as well as fewer SARS-CoV-2-infected cells and immune cell infiltrates in the lungs concomitant with reduced levels of proinflammatory cytokines and chemotactic factors in the BALF were observed in animals immunized with the CoVLP adjuvanted with AS03. No clinical, pathologic, or virologic evidence of vaccine-associated enhanced disease was observed in vaccinated animals. The CoVLP adjuvanted with AS03 was therefore selected for vaccine development and clinical trials.

Increases in complexity attainable in molecular self-assembly necessitates both advanced molecular design as well as microenvironmental control. Such control is offered by microfluidics, where precise chemical compositions and gradients can be readily established. A droplet microfluidic platform combining upstream step emulsification with downstream hydrodynamic microtraps has been designed to facilitate molecular self-assembly. The step emulsification rapidly generates uniform droplets which act as reaction chambers. The hydrodynamic microtraps hold droplets against the flow ensuring they are exposed to a continuous supply of fresh fluid for constant reagent extraction and/or delivery. Additionally, the droplet immobilization permits real-time droplet characterization and reaction monitoring. Subsequently, droplets can be released from the traps through flow reversal, allowing post-process characterization. The microfluidic system was demonstrated by the phase separation of lyotropic droplets. Ethanol/water droplets were created in a continuous ambient squalene/monoolein microflow, causing the continuous extraction of ethanol from the droplets and delivery of monoolein from the ambient microflow. Unlike conventional bulk techniques and continuous microfluidics, where finite microchannel lengths necessarily impose limits to the extent to which slow processes can proceed, this approach allows extended duration reactions whilst enabling real time process monitoring.

Cholesterol synthesis is both energy- and oxygen-intensive, yet relatively little is known of the regulatory effects of hypoxia on pathway enzymes. We previously showed that the rate-limiting and first oxygen-dependent enzyme of the committed cholesterol synthesis pathway, squalene monooxygenase (SM), can undergo partial proteasomal degradation that renders it constitutively active. Here, we show hypoxia is a physiological trigger for this truncation, which occurs through a two-part mechanism: (1) increased targeting of SM to the proteasome via stabilization of the E3 ubiquitin ligase MARCHF6 and (2) accumulation of the SM substrate, squalene, which impedes the complete degradation of SM and liberates its truncated form. This preserves SM activity and downstream pathway flux during hypoxia. These results uncover a feedforward mechanism that allows SM to accommodate fluctuating substrate levels and may contribute to its widely reported oncogenic properties.

11 Páginas.-- 5 Figuras, Carotenoids are isoprenoid lipids found across the tree of life with important implications in oxidative stress adaptations, photosynthetic metabolisms, as well as in membrane dynamics. The canonical view is that C40 carotenoids are synthesized from phytoene and C30 carotenoids from diapophytoene. Squalene is mostly associated with the biosynthesis of polycyclic triterpenes, although there have been suggestions that it could also be involved in the biosynthesis of C30 carotenoids. However, demonstration of the existence of this pathway in nature is lacking. Here, we demonstrate that C30 carotenoids are synthesized from squalene in the Planctomycetes bacteria and that this squalene route to C30 carotenoids is the most widespread in prokaryotes. Using the evolutionary history of carotenoid and squalene amino oxidases, we propose an evolutionary scenario to explain the origin and diversification of the different carotenoid and squalene-related pathways. We show that carotenoid biosynthetic pathways have been constantly transferred and neofunctionalized during prokaryotic evolution. One possible origin of the squalene pathway connects it with the one of C40 carotenoid synthesis of Cyanobacteria. The widespread occurrence of the squalene route to C30 carotenoids in Bacteria increases the functional repertoire of squalene, establishing it as a general hub of carotenoids and polycyclic triterpenes synthesis., This work was supported by the Spanish Ministry of Economy and Competitiveness (Grant No. BFU2016-78326-Pand and MDM-2016-0687). This research is funded in part by the Gordon and Betty Moore foundation through Grant GBMF9733 to D.P.D. We also thank Berend Snel and Anja Spang for financial support of C.S.-M. through a grant funded by Utrecht University and the Netherlands Organisation for Scientific Research (NWO).

Santalenes and santalols from Santalum album are the main components of the valuable spice sandalwood essential oil, which also has excellent pharmacological activities such as antibacterial, anti-inflammatory, and antitumor. Firstly, we constructed biosynthesis pathways of santalenes by synthetic biology strategy. The assembled biosynthetic cassettes were integrated into the multiple copy loci of δ gene in S. cerevisiae BY4742 with assistance of pDi-CRISPR, and 94.6 mg/L santalenes was obtained by shake flask fermentation of engineered yeast. Secondly, a selected optimized P450-CPR redox system was integrated into the chromosome of the santalenes-producing strain with a single copy, and 24.6 mg/L santalols were obtained. Finally, the yields of santalenes and santalols were increased to 164.7 and 68.8 mg/L, respectively, by downregulating ERG9 gene. This is the first report on the de novo synthesis of santalols by P450-CPR chimera in S. cerevisiae. Meanwhile, the optimized chimeric CYP736A167opt-46tATR1opt exhibits higher activity to oxidize santalenes into santalols. It would provide a feasible solution for the optimal biosynthesis of santalols. KEY POINTS: • First-time de novo synthesis of santalols by P450-CPR chimera in S. cerevisiae. • Truncated 46tATR1 has higher activity than that of CPR2. • Yields of santalenes and santalols were increased by downregulating ERG9 gene.

The profitability of the olive flounder (Paralichthys olivaceus) aquaculture industry in Korea depends on high production and maintenance of flesh quality, as consumers prefer to eat raw flounders from aquaria and relish the raw muscles as ‘sashimi’. For sustaining high production, easy-to-deliver and efficient vaccination strategies against serious pathogens, such as viral hemorrhagic septicemia virus (VHSV), is very important as it cause considerable losses to the industry. Whereas, a safe and non-invasive vaccine formulation that is free from unacceptable side-effects and does not devalue the fish is needed to maintain flesh quality. We previously developed a squalene–aluminium hydroxide (Sq + Al) adjuvanted VHSV vaccine that conferred moderate to high protection in flounder, without causing any side effects when administered through the intraperitoneal (IP) injection route. However, farmers often demand intramuscular (IM) injection vaccines as they are relatively easy to administer in small fishes. Therefore, we administered the developed vaccine via IP and IM routes and investigated the safety and persistency of the vaccine at the injection site. In addition, we conducted a comparative analysis of vaccine efficacy and serum antibody response. The clinical and histological observation of the IM and IP groups showed that our vaccine remained persistence at the injection sites for 10–17 weeks post vaccination (wpv), without causing any adverse effects to the fish. The relative percentage of survival were 100% and 71.4% for the IP group and 88.9% and 92.3% for the IM group at 3 and 17 wpv, respectively. Thus, considering the persistency period (24 wpv) and both short and long-term efficacy of our vaccine, the present study offers an option to flounder farmers in selecting either IM or IP delivery strategy according to their cultured fish size and harvesting schedule — IM vaccination for small-sized fish and IP vaccination for table-sized fish.

Background Vaccines are the main prophylactic measure against pandemic influenza. Adjuvanted, cell culture–derived vaccines, which are not subject to limitations of egg-based vaccine production, have the potential to elicit an antibody response against heterologous strains and may be beneficial in the event of an A/H5N1 pandemic. Methods A prespecified exploratory analysis of data from a phase 2, randomized, controlled, observer-blind multicenter trial (NCT01776554) to evaluate the immunogenicity of a MF59-adjuvanted, cell culture–based A/H5N1 influenza vaccine (aH5N1c), containing 7.5 µg hemagglutinin antigen per dose, in subjects 6 months through 17 years of age was conducted. Geometric mean titers (GMT) were determined using hemagglutination inhibition (HI) and microneutralization (MN) assays, and proportions of patients achieving seroconversion, HI and MN titers ≥ 1:40, and a 4-fold increase in MN titers against 5 heterologous strains (influenza A/H5N1 Anhui/2005, Egypt/2010, Hubei/2010, Indonesia/2005, and Vietnam/1203/2004) three weeks after administration of the second dose were assessed. Results After the second dose, HI GMTs against heterologous strains increased between 8- and 40-fold, and MN GMTs increased 13- to 160-fold on Day 43 vs Day 1. On Day 43, 32–72% of subjects had HI titers ≥ 1:40 and achieved seroconversion against the heterologous strains. Using the MN assay, 84–100% of subjects had MN titers ≥ 1:40 and 83–100% achieved an at least 4-fold increase in MN titers against the heterologous strains. The highest responses were consistently against A/H5N1 Egypt/2010. Conclusions When given to children aged 6 months through 17 years, aH5N1c resulted in increased immunogenicity from baseline against all 5 heterologous A/H5N1 strains tested, demonstrating the potential of an MF59-adjuvanted, cell-derived A/H5N1 vaccine to provide cross-protection against other A/H5N1 strains (NCT01776554).

Harnessing mitochondria is considered as a promising method for biosynthesis of terpenes due to the adequate supply of acetyl-CoA and redox equivalents in mitochondria. However, mitochondrial engineering often causes serious metabolic burden indicated by poor cell growth. Here, we systematically analyzed the metabolic burden caused by the compartmentalization of the MVA pathway in yeast mitochondria for squalene synthesis. The phosphorylated intermediates of the MVA pathway, especially mevalonate-5-P and mevalonate-5-PP, conferred serious toxicity within mitochondria, which significantly compromised its possible advantages for squalene synthesis and was difficult to be significantly improved by routine pathway optimization. These phosphorylated intermediates were converted into ATP analogues, which strongly inhibited ATP-related cell function, such as mitochondrial oxidative respiration. Fortunately, the introduction of a partial MVA pathway from acetyl-CoA to mevalonate in mitochondria as well as the augmentation of the synthesis of mevalonate in cytosol could significantly promote the growth of yeasts. Accordingly, a combinatorial strategy of cytoplasmic and mitochondrial engineering was proposed to alleviate the metabolic burden caused by the compartmentalized MVA pathway in mitochondria and improve cell growth. The strategy also displayed the superimposed effect of cytoplasmic engineering and mitochondrial engineering on squalene production. Through a two-stage fermentation process, the squalene titer reached 21.1 g/L with a specific squalene titer of 437.1 mg/g dcw, which was the highest at present. This provides new insight into the production of squalene and other terpenes in yeasts based on the advantages of mitochondrial engineering.

Transcriptional downregulation is widely used for metabolic flux control. Here, marO, a cis-element of Escherichia coli mar operator, was explored to engineer promoters of Saccharomyces cerevisiae for downregulation. First, the ADH1 promoter (PADH1) and its enhanced variant PUADH1 were engineered by insertion of marO into different sites, which resulted in decrease in both gfp5 transcription and GFP fluorescence intensity to various degrees. Then, marO was applied to engineer the native ERG1 and ERG11 promoters due to their importance for accumulation of value-added intermediates squalene and lanosterol. Elevated squalene content (4.9-fold) or lanosterol content (4.8-fold) and 91 or 28% decrease in ergosterol content resulted from the marO-engineered promoter PERG1(M5) or PERG11(M3), respectively, indicating the validity of the marO-engineered promoters in metabolic flux control. Furthermore, squalene production of 3.53 g/L from cane molasses, a cheap and bulk substrate, suggested the cost-effective and promising potential for squalene production.

Vaccination is the most effective approach to reduce the substantial morbidity and mortality caused by influenza infection. Vaccine efficacy is highly sensitive to antigenic changes causing differences between circulating and vaccine viruses. Adjuvants such as MF59 increase antibody-mediated cross-reactive immunity and therefore may provide broader seasonal protection. A recent clinical trial showed that an MF59-adjuvanted vaccine was more efficacious than a nonadjuvanted comparator in subjects < 2 years of age, although not in those ≥ 2 years, during influenza seasons in which the predominant circulating virus was an A/H3N2 strain that was antigenically different from the vaccine virus. This finding suggested that the increased efficacy of the adjuvanted vaccine in younger subjects may be mediated by strain cross-reactive antibodies. A subset of the trial population, representing subjects with distinct age and/or immunological history, was tested for antibody responses to the vaccine A/H3N2 strain as well as A/H3N2 drifted strains antigenically matching the viruses circulating during the trial seasons. The neutralizing tests showed that, compared with nonadjuvanted vaccine, the adjuvanted vaccine improved not only the neutralizing antibody response to the vaccine strain but also the cross-reactive antibody response to the drifted strains in subjects with lower preexisting antibody titers, regardless of their age or vaccine history. The results demonstrated an immunological benefit and suggested a potential efficacy benefit by adjuvanted vaccine in subjects with lower preexisting antibody responses.

The C-H and O-H oscillators on the surfaces of thin films of human-derived skin oil and squalene are probed under ambient conditions (300 K, 1 atm total pressure, 40% RH) using second-order vibrational spectroscopy and contact angle goniometry before and after exposure to ppb amounts of ozone. Skin oil and squalene are found to produce different vibrational sum frequency generation spectra in the C-H stretching region, while exposure to ozone results in surface spectra for both materials that is consistent with a loss of C-H oscillators. The measured contact angles show that the hydrophobicity of the films increases following exposure to ozone, consistent with the reduction in C═C···H

Wangmo red ball Camellia oleifera is the main Camellia species cultivated for oil in the low-heat valley of Guizhou, China. In this study, we evaluated the comprehensive nutritional value of Wangmo C. oleifera seed oil through fatty acid and nutritional component analyses. Twenty excellent Wangmo C. oleifera plants with stable yield and disease resistance were selected from the Camellia oleifera germplasm resource garden in the low-heat valley site of Guizhou University. The unit crown yield, fatty acid content of the seed oil, fatty acid composition and functional nutrients were determined, and the oil quality was comprehensively evaluated using principal component analysis. The fatty acid content of C. oleifera seed oil was 35.03–53.47%, suggesting likely popularization and wide application prospects. The fatty acids included SFAs, MUFAs and PUFAs, and the oleic acid content was 80%, indicating a highly stable and nutritious oil. The oil was also rich in carotenoids, polyphenols, flavonoids, β-sitosterol, squalene and α-Ve, with average content of 7.404 mg/kg, 16.062 mg/kg, 0.401 g/100 g, 265.087 mg/kg, 129.315 mg/kg and 21.505 mg/100 g, respectively. However, the correlations among the nutritional indices were weak. PCA showed that germplasms GH7, GH43, GH28, GH8 and GH31 exhibited the top five nutritional qualities. The rankings in this study provide data for identifying excellent Wangmo C. oleifera plants with high nutritional quality. Additionally, this study provides a valuable reference for the research and development of high-end edible oil and a theoretical basis for the development of economic forest species in low-heat valley areas across the world.

Holm oak (

Heavy metal toxicity and drought stresses are two growing concerns of the global population as these have a disastrous effect on the agriculture sector, creating oxidative stress in plants and leads to deleterious effects, which end up causing a major decline in yield. Hence, carving out the best candidate for combating both these stresses have become the prime objective of researchers. Plectranthus amboinicus (Lour.) Spreng., a medicinal plant, is regarded as one such candidate that can tolerate both drought and zinc stressors by the elicitation of metabolic changes. Variation in the composition of primary and secondary metabolites of P. amboinicus was evaluated in the leaf tissues of the plants subjected to drought and ZnSO4 (4 mM) treatments. Drought stress resulted in the accumulation of amino acids and sugars in the leaves of plants exposed to this stress. Similarly, zinc stress exhibited a remarkable impact on the synthesis of secondary metabolites like alkaloids, phenols, and flavonoids. Further, on carrying out GC–MS profiling, the compositional variation of secondary metabolites produced in P. amboinicus implicated its inherent potential to survive environments of these two stresses. Compounds like 2-methoxy-4-ethyl-6-methylphenol, gamma-sitosterol, hexadecanoic acid, alpha-amyrin, and ethyl linalool were some of the major secondary metabolites developed in leaves of plants under drought, whereas during ZnSO4 treatment, the major compounds developed were trans-alpha-bergamotene, squalene, 1,5-dimethyl-1-vinyl-4-hexenyl isovalerate, and spathulenol. This in turn makes the plant more compatible with the stressful environmental conditions, aiding it with better survival and protection. At the same time, the enhancement in the content of these metabolites in this medicinal plant under the influence of these stressors may have applications in the pharmaceutical industry, necessary for the development of novel drugs.

PURPOSE: The enzyme Cyclooxygenases (COX-1 and COX-2) catalyze the formation of prostaglandin, a mediator of the inflammatory pathway. Inflammation related pathological conditions may be alleviated by targeting the Cox enzymes.COX-2 inhibitors that are currently available in the market causes undesirable side effects. Our present study focuses on the in-silico inhibition of COX -2 enzyme by the phytocompounds from Albizia amara and Phyla nodiflora. METHODS: The phytochemicals present in Albizia amara and Phyla nodiflora were analyzed for their COX-2 inhibition potential. Eight compounds from Albizia amara and eleven compounds from Phyla nodiflora obtained from GC–MS analysis was used for the current study. Molecular docking was performed using AutoDock vina. The crystal structure of COX-2 (PDB ID: 5IKR) was obtained from Protein data bank. PyMol was used to remove any solvent, organic and inorganic molecules. Energy minimization of the protein was carried out using SPDBV software. Geometrical optimizations of the ligands were performed using Avogadro software. Celecoxib was used as the positive control. ADMET properties of the compounds were analyzed using SwissADME and ProtoxII online servers. Molecular mechanics/generalized born surface area (MM/GBSA) calculations were performed to evaluate the binding efficiency. Molecular dynamics of the protein and protein–ligand complex was studied for about 100 ns using Desmond package of Schrodinger suite. RESULTS: Among the eighteen compounds, Squalene present in both the plants showed a better binding energy of -7.7 kcal/mol, when compare to other phytocompounds present in the extract. The control celecoxib showed a binding energy of about – 9.4 kcal/mol. The toxicity and ADMET properties of squalene indicated that it is non-toxic and followed Lipinski’s rule. Molecular Dynamics (MD) analysis showed that the binding of squalene to the enzyme was stable. CONCLUSION: Squalene could potentially inhibit COX2 and o wing to its properties, squalene can be formulated in gels/creams and could be possibly used for external edema and inflammation GRAPHICAL ABSTRACT: [Image: see text] SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s40199-021-00408-6.

Clerodendrum inerme is a perennial shrub belonging to the family Lamiaceae and is used in many of the herbal preparations of Siddha and Ayurveda in India. Ethnomedicinally, the leaves of this plant are used to cure various ailments. In spite of enormous ethnomedicinal implications, proper chemical profiling and characterization of isolated compounds of C. inerme still remains unaddressed. Present study involves the characterization and chemical profiling of phytoconstituents from leaf extract which promise to be of immense therapeutic significance. Two compounds namely squalene and linolenic acid methyl ester were isolated from the leaves of C. inerme employing column chromatography technique. The structures were elucidated by spectroscopic methods including FTIR, 1H NMR and 13C NMR. Subsequently, antioxidant and antimicrobial activities of the isolated compounds were estimated in-vitro. Pertaining results were validated with in silico molecular docking-based investigations which revealed that both the compounds displayed high binding affinities with the target proteins of the antioxidant pathway commonly targeted for drug development.

Summary Background The absolute degree of protection from influenza vaccines in older adults has not been studied since 2001. This study aimed to show the clinical efficacy of an MF59-adjuvanted quadrivalent influenza vaccine (aQIV) in adults 65 years or older compared with adults not vaccinated to prevent influenza. Methods We did a randomised, stratified, observer-blind, controlled, multicentre, phase 3 study at 89 sites in 12 countries in 2016–17 northern hemisphere and 2017 southern hemisphere influenza seasons. We enrolled community-dwelling male and female adults aged 65 years and older who were healthy or had comorbidities that increased their risk of influenza complications. We stratified eligible participants by age (cohorts 65–74 years and ≥75 years) and risk of influenza complications (high and low) and randomly assigned them (1:1) via an interactive response technology to receive either aQIV or a non-influenza comparator vaccine. We masked participants and outcome assessors to the administered vaccine. Personnel administering the vaccines did not participate in endpoint assessment. The primary outcome was absolute vaccine efficacy assessed by RT-PCR-confirmed influenza due to any influenza strain in the overall study population (full analysis set) from day 21 to 180 or the end of the influenza season. Vaccine efficacy was calculated on the basis of a Cox proportional hazard regression model for time to first occurrence of RT-PCR-confirmed influenza due to any strain of influenza. Safety outcomes were assessed in the overall study population. This trial was registered with ClinicalTrials.gov , NCT02587221 . Findings Northern hemisphere enrolment occurred between Sept 30, 2016, and Feb 28, 2017, and southern hemisphere enrolment between May 26, 2017, and 30 June 30, 2017. aQIV was administered to 3381 participants, who subsequently had 122 (3·6%) RT-PCR-confirmed influenza cases, and the comparator was administered to 3380 participants, who subsequently had 151 (4·5%) influenza cases. The majority, 214 (78·4%) of 273, were caused by influenza A H3N2. Most antigenically characterised isolates were mismatched to the vaccine strain (118 [85%] of 139). Vaccine efficacy was 19·8% (multiplicity-adjusted 95% CI −5·3 to 38·9) against all influenza and 49·9% (−24·0 to 79·8) against antigenically matched strains, when the protocol definition of influenza-like illness was used. The most common local solicited adverse event was injection site pain, reported by 102 (16·3%) of 624 participants in the aQIV group and 71 (11·2%) of 632 of participants in the comparator group. Deaths were evenly distributed; none were considered related to study vaccines. The safety profile for aQIV was similar to previously reported trials. Interpretation The prespecified criterion for showing the efficacy of aQIV in older adults was not met during the influenza seasons with high amounts of vaccine strain mismatch. Vaccine efficacy was higher against influenza cases associated with higher fever, which represent more clinically significant disease. Funding Seqirus UK.

The constituent and fatty-acid compositions of neutral lipids (NL), glyco-, and phospholipids and the squalene contents in seeds and the aerial part of Amaranthus retroflexus were studied for the first time. Elaidic acid was observed in all lipid groups. The squalene content was 4.3% of the NL mass in seeds and 3.15% in the aerial part. Preliminary wetting of wheat and cucumber seeds in unsaponified substances, fatty acids, and their methyl esters obtained from A. retroflexus seeds at concentrations 0.001 and 0.0001% was found to reduce the inhibiting action of salt stress. Neutral lipids did not exhibit a protective effect.

The oil palm (Elaeis guineensis) fruit is a source of vegetable oil and various phytonutrients. Phytochemical compounds present in palm oil include tocotrienols, carotenoids, phytosterols, squalene, coenzyme Q10, and phospholipids. Being a fruit, the oil palm is also a rich source of water-soluble phytonutrients, including phenolic compounds. Extraction of phytonutrients from the oil palm vegetation liquor of palm oil milling results in a phenolic acid-rich fraction termed Water-Soluble Palm Fruit Extract (WSPFE). Pre-clinical in vitro, ex vivo, and in vivo studies carried out using various biological models have shown that WSPFE has beneficial bioactive properties, while clinical studies in healthy volunteers showed that it is safe for human consumption and confers antioxidant and anti-inflammatory effects. The composition, biological properties, and relevant molecular mechanisms of WSPFE discovered thus far are discussed in the present review, with a view to offer future research perspectives on WSPFE for health and non-health applications.

Red-skin disease (RSD) is a common soil-borne disease encountered during ginseng cultivation. We found that RSD was closely related to soil properties such as soil acidification and metal stress. The levels of active ingredients, especially ginsenoside, in RSD ginseng were reduced, thereby reducing its medicinal value. We investigated the inducing factors and pathological processes leading to RSD in ginseng and the soil properties responsible for the inhibition of ginsenoside biosynthesis. Healthy and RSD ginseng were compared for properties including the planting slope, soil moisture, soil pH, and metal content. Changes in metal absorption and ginsenoside synthesis at different pH were screened as inducers of RSD using ginseng cells. The activities of nitrate reductase, glutamate dehydrogenase, glutamate synthase, glutamine synthase, and squalene synthase were determined using enzyme-linked immunosorbent assay. The effect of nitrogen on ginsenoside synthesis was analyzed by culturing ginseng cells in a nitrogen-deficient environment. Field data showed that RSD ginseng grew at a slope of about 13 degrees and at a slower rate than healthy ginseng. RSD was caused when ginseng was grown in soils having higher water content and acidity. Acidic conditions inhibited the ginsenoside biosynthesis, whereas an increase in the pH of the medium could alleviate this inhibition. The overabsorption of aluminum ions in ginseng cells in acidic conditions was most significant. Aluminum stress not only inhibited the ginsenoside synthesis but also decreased the levels of nitrogen-containing compounds and enzyme activities related to nitrogen metabolism. Moreover, nitrogen concentration was positively correlated with ginsenoside content and squalene synthase activity. Our findings indicated that RSD was observed in ginseng when the planting slope was slow, there was water accumulation in the soil, and when the soil was acidic. Acidic conditions appeared to promote aluminum overabsorption in ginseng cells. The interference in aluminum stress-induced nitrogen uptake was found to inhibit ginsenoside synthesis in RSD. Our findings may contribute to a better understanding of the susceptibility factors and pathological processes in the RSD of ginseng.

Panax quinquefolius L., a medicinal plant of Araliaceae family, is used in traditional Chinese medicine. Arbuscular mycorrhizal fungi (AMF) has been shown to promote plant growth and the accumulation of secondary metabolites in many plants. Our group has found a significant positive correlation between AMF infection rate and the accumulation of ginsenoside, while the effects and underlying mechanism of AMF on the growth and accumulation of ginsenoside in P. quinquefolius have been poorly studied. In this study, the effects on the biomass, photosynthesis and expressions of genes involved in biosynthesis of ginsenoside: hydroxymethylglutaryl-CoA reductase (HMGR), geranylgeranyl pyrophosphate synthase (GPS), farnesyl diphosphate synthase (FPS), squalene synthase (SS), squalene epoxidase (SE), dammarendiol synthase (DS) and cytochrome P450 enzyme (CYP716A47, CYP716A53v2) in P. quinquefolius during AMF inoculation were investigated. Plant biomass and photosynthesis of P. quinquefolius were stimulated by AMF treatment. Chlorophyll concentration in P. quinquefolius leaves treated by R. intraradices was higher than those in non-inoculated leaves. The concentration of Rb1 and Re was markedly increased by 26.15% and 22.38% (P

The transition from batch catalytic processes to continuous flow processes requires highly active and stable catalysts that still need to be developed. The preparation and characterization of catalysts where palladium single atoms and nanoparticles are simultaneously present on carbon nanotubes were recently reported by us. These catalysts are considerably more active than commercial or previously described catalysts for the liquid phase hydrogenation of terpenes. Herein is shown that under solvent-free conditions, squalene (SQE) could be converted into squalane (SQA,98 %) using only 300 ppm of Pd in less than 1.4 h at 20 bar H

The higher effectiveness of adjuvanted trivalent influenza vaccine (aTIV) versus non-adjuvanted (na) formulations in preventing all-cause hospitalization has been demonstrated for a single influenza season and in institutionalized elderly only. This study evaluated the relative vaccine effectiveness for aTIV vs. non-adjuvanted trivalent (naTIV) and/or quadrivalent (naQIV) influenza vaccines in preventing all-cause hospitalizations across 18 influenza seasons in primary care.Using Health Search Database, a nested case-control analysis was conducted in a cohort of older adults being vaccinated with aTIV or naTIV/naQIV. Conditional logistic regression was adopted to estimate the odds ratio (OR) of all-cause hospitalizations occurred during the epidemic period.Of 58,252 patients vaccinated with aTIV and naTIV/naQIV for the first time, 2,504 cases of all-cause hospitalization (3.46 per 1,000 person-weeks) during the 18 influenza seasons were identified. Compared with naTIV/naQIV, aTIV was associated with a 12% reduced the odds of all-cause hospitalizations (OR 0.88; 95% CI: 0.80-0.97).In an 18-season cohort of older adults, aTIV reduced the risk of all-cause hospitalizations when compared with naTIV/naQIV. Our findings confirm additional benefits for adjuvanted influenza vaccines in older adults.

Two key amino acids of isomultiflorenol synthase, Y125 and M254, were first proposed. They could be associated with the production of isomultiflorenol. Oxidosqualene cyclases (OSCs) are the first committed enzymes in the triterpenoid biosynthesis by converting 2,3-oxidosqualene to specific triterpenoid backbones. Thus, these enzymes are potential targets for developing plant-active compounds through the study of triterpenoid biosynthesis. We applied transcriptome information and metabolite profiling from Trichosanthes cucumerina L. to define the diversity of triterpenoids in this plant through OSCs. Isomultiflorenol synthase and cucurbitadienol synthase were previously identified in this plant. Here, three new OSCs, TcBAS, TcLAS, and TcCAS, were cloned and functionally characterized as β-amyrin synthase, lanosterol synthase, and cycloartenol synthase activities, respectively. We also took advantage of the multiple sequence alignment and molecular docking of OSCs exhibiting in this plant and other plant OSCs to identify key residues associated with isomultiflorenol synthase specificity. Two novel key amino acids, referred to the Y125 and M254, were first discovered. These results provide information on a possible catalytic mechanism for plant OSCs that produce specific products.

The ongoing COVID-19 pandemic has contributed largely to the global vaccine disparity. Development of protein subunit vaccines can help alleviate shortages of COVID-19 vaccines delivered to low-income countries. Here, we evaluated the efficacy of a three-dose virus-like particle (VLP) vaccine composed of hepatitis B surface antigen (HBsAg) decorated with the receptor binding domain (RBD) from the Wuhan or Beta SARS-CoV-2 strain adjuvanted with either aluminum hydroxide (alum) or squalene in water emulsion (SWE). RBD HBsAg vaccines were compared to the standard two doses of Pfizer mRNA vaccine. Alum-adjuvanted vaccines were composed of either HBsAg conjugated with Beta RBD alone (β RBD HBsAg+Al) or a combination of both Beta RBD HBsAg and Wuhan RBD HBsAg (β/Wu RBD HBsAg+Al). RBD vaccines adjuvanted with SWE were formulated with Beta RBD HBsAg (β RBD HBsAg+SWE) or without HBsAg (β RBD+SWE). Both alum-adjuvanted RBD HBsAg vaccines generated functional RBD IgG against multiple SARS-CoV-2 variants of concern (VOC), decreased viral RNA burden, and lowered inflammation in the lung against Alpha or Beta challenge in K18-hACE2 mice. However, only β/Wu RBD HBsAg+Al was able to afford 100% survival to mice challenged with Alpha or Beta VOC. Furthermore, mice immunized with β RBD HBsAg+SWE induced cross-reactive neutralizing antibodies against major VOC of SARS-CoV-2, lowered viral RNA burden in the lung and brain, and protected mice from Alpha or Beta challenge similarly to mice immunized with Pfizer mRNA. However, RBD+SWE immunization failed to protect mice from VOC challenge. Our findings demonstrate that RBD HBsAg VLP vaccines provided similar protection profiles to the approved Pfizer mRNA vaccines used worldwide and may offer protection against SARS-CoV-2 VOC.

Mycoses are a global problem that affects humans and animals. In the present study, the entomopathogenic soil fungus Conidiobolus coronatus (Entomophthorales), infecting in tropics also humans, sheep and horses, was cultivated with the addition of insect cuticular compounds (CCs) previously detected in the cuticle of C. coronatus—resistant fly species (C10–C30 fatty alcohols, butyl oleate, butyl stearate, glycerol oleate, squalene, tocopherol acetate). Our findings indicate that CCs have diversified and complex effects on the growth and sporulation of C. coronatus and its ability to infect the larvae of Galleria mellonella (Lepidoptera). The CCs affected protein content and cuticle-degrading enzymes (CDEs) activity in the conidia. Some CCs inhibited fungal growth (0.1% C10), decreased sporulation (C12, C16, C24, C28, C30, butyl stearate, squalene), virulence (C12, C14, butyl oleate, butyl stearate) and protein content (C18). They also reduced conidial CDE activity: elastase (C24, butyl oleate, butyl stearate, squalene, tocopherol acetate), chitobiosidase (C12, C14, C20) and lipase (C12, C18, C26, squalene, tocopherol acetate). Several CCs enhanced sporulation (C14, C18, C22, C26, C30), virulence (C18, C26, squalene), conidial protein content (C16, C24, C30, squalene) and CDE activity: elastase (C10, C16, C18), NAGase (C16, C20), chitobiosidase (C16) and lipase (C10, C14, C16, C20, butyl oleate). Our findings indicate that C. coronatus colonies grown on media supplemented with CCs employ various compensation strategies: colonies grown with C16 alcohol demonstrated reduced sporulation but greater conidial protein accumulation and increased elastase, NAGase, chitobiosidase and lipase activity, thus preserving high virulence. Also, colonies supplemented with C18 alcohol demonstrated high virulence and enhanced sporulation and elastase activity but slightly decreased conidial protein content. CCs that inhibit the activity of lipases and proteases show promise in the fight against conidiobolomycosis.

(1) Background: An earlier study on the hypoglycemic activity of

The use of alfalfa is a promising response to the increasing demand for squalene. Ensiling could enhance the squalene content of fresh alfalfa and silage. To investigate and exploit the anaerobic fermentation of forage as a new squalene source, alfalfa was ensiled without (CON) or with molasses (ML) and sunflower seed oil (SSL) for 10, 40, and 70 days.Naturally ensiled alfalfa was of poor quality but had up to 1.93 times higher squalene content (P 0.001) than fresh alfalfa. The squalene-producing bacteria were found to be cocci lactic acid bacteria (LAB). Adding ML and SSL decreased squalene content (P = 0.002 and P 0.001) by 6.89% and 11.6%, respectively. Multiple linear regression models and correlation analysis indicated that squalene synthase was the key enzyme for squalene synthesis. The addition of ML and SSL altered the structure of LAB communities, mainly decreasing the relative abundance of cocci LAB, which was responsible for squalene synthesis, and changing the fermentation products (lactic acid, propionic acid, and ammonia-N) influencing the squalene-related enzymes, thereby decreasing squalene production. Compared with squalene production from the reference bacteria (Pediococcus acidilactici Ch-2, Rhodopseudomonas palustris, Bacillus subtilis, engineered Escherichia coli), alfalfa silage had the potential to be a new squalene source.Natural ensiled alfalfa was a promising source for squalene, and ensiling was a potential pathway to obtain novel high-yield squalene bacteria. © 2022 Society of Chemical Industry.

SARS-CoV-2 variants have posed significant challenges to the hopes of using ancestral strain-based vaccines to address the risk of breakthrough infection by variants. We designed and developed a bivalent vaccine based on SARS-CoV-2 Alpha and Beta variants (named SCTV01C). SCTV01C antigens were stable at 25