Development of an electrochemical sensor for the quantification of ascorbic acid and acetaminophen in pharmaceutical samples.

This work presents the modification of glassy carbon electrodes (GCE) by using a dispersion resulting from the non-covalent functionalization of multi-walled carbon nanotubes (MWCNT) with polyarginine (polyArg). MWCNT-polyArg is used for the quantification of ascorbic acid (AA) in the presence of acetaminophen (APAP) and viceversa. Since ascorbic acid and acetaminophen are strongly absorbed on GCE/MWCNT-polyArg, they can be detected in the presence of 4.0×10-5 M acetaminophen (and 3.0×10-5 M ascorbic acid) by using adsorptive stripping with media exchange and differential pulse voltammetry. Using water as the solvent for the MWCNT dispersion, the result was Z-potential of 0.053 ± 0.006 V. The developed sensor showed excellent specificity, sensitivity, stability and reproducibility compared to previously published sensors. The GCE/MWCNT-polyArg sensor shows a fast response time of ∼5 minutes, low limits of detection and quantification for AA (0.95 and 2.9 µM respectively) and APAP (0.27 and 0.82µM, respectively), high sensitivity of 0.0616 µA/M for AA or APAP 0.240µA/M. It was used to test its practicability by determining the concentration of AA or APAP (AA and APAP) in pharmaceutical samples. Finally, the simultaneous measurement of ascorbic acid and acetaminophen in pharmaceuticals showed a good correlation, with a maximum error and RSD of 4.5 and 5.1 %, respectively.

Pilot-scale microalgae cultivation and wastewater treatment using high-rate ponds: a meta-analysis.

Microalgae cultivation in wastewater has been widely researched under laboratory conditions as per its potential to couple treatment with biomass production. Currently, only a limited number of published articles consider outdoor and long-term microalgae-bacteria cultivations in real wastewater environmental systems. The scope of this work is to describe microalgal cultivation steps towards high-rate algal pond (HRAP) scalability and identify key parameters that play a major role for biomass productivity under outdoor conditions and long-term cultivations. Reviewed pilot-scale HRAP literature is analysed using multivariate analysis to highlight key productivity parameters within environmental and operational factors. Wastewater treatment analysis indicated that HRAP can effectively remove 90% of NH4+, 70% of COD, and 50% of PO43-. Mean reference values of 210 W m-2 for irradiation, 18 °C for temperature, pH of 8.2, and HRT of 7.7 are derived from pilot-scale cultivations. Microalgae biomass productivity at a large scale is governed by solar radiation and NH4+ concentration, which are more important than retention time variations within investigated studies. Hence, selecting the correct type of location and a minimum of 70 mg L-1 of NH4+ in wastewater will have the greatest effect in microalgae productivity. A high nutrient wastewater content increases final biomass concentrations but not necessarily biomass productivity. Pilot-scale growth rates (~ 0.54 day-1) are half those observed in lab experiments, indicating a scaling-up bottleneck. Microalgae cultivation in wastewater enables a circular bioeconomy framework by unlocking microalgal biomass for the delivery of an array of products.

Zika virus non-coding RNAs antagonize antiviral responses by PKR-mediated translational arrest.

Zika virus (ZIKV) is an emerging mosquito-borne flavivirus that causes severe outbreaks in human populations. ZIKV infection leads to the accumulation of small non-coding viral RNAs (known as sfRNAs) that are crucial for evasion of antiviral responses and for viral pathogenesis. However, the mechanistic understanding of how sfRNAs function remains incomplete. Here, we use recombinant ZIKVs and ribosome profiling of infected human cells to show that sfRNAs block translation of antiviral genes. Mechanistically, we demonstrate that specific RNA structures present in sfRNAs trigger PKR activation, which instead of limiting viral replication, enhances viral particle production. Although ZIKV infection induces mRNA expression of antiviral genes, translation efficiency of type I interferon and interferon stimulated genes were significantly downregulated by PKR activation. Our results reveal a novel viral adaptation mechanism mediated by sfRNAs, where ZIKV increases its fitness by repurposing the antiviral role of PKR into a proviral factor.

Present and Future Applications of Digital PCR in Infectious Diseases Diagnosis.

Infectious diseases account for about 3 million deaths per year. The advent of molecular techniques has led to an enormous improvement in their diagnosis, both in terms of sensitivity and specificity and in terms of the speed with which a clinically useful result can be obtained. Digital PCR, or 3rd generation PCR, is based on a series of technical modifications that result in more sensitive techniques, more resistant to the action of inhibitors and capable of direct quantification without the need for standard curves. This review presents the main applications that have been developed for the diagnosis of viral, bacterial, and parasitic infections and the potential prospects for the clinical use of this technology.

Neurons regulate the esterification of bioactive lipid mediators in the brain of acid sphingomyelinase deficient mice.

Acid sphingomyelinase deficiency is a neurodegenerative lysosomal storage disorder caused by mutations in the sphingomyelin-degrading enzyme acid sphingomyelinase (ASM) gene. Upregulated neuroinflammation has been well-characterized in an ASM knockout mouse model of acid sphingomyelinase deficiency disease, but lipid mediator pathways involved in 'mediating' inflammation and inflammation-resolution have yet to be characterized. In this study, we 1) measured free (bioactive) and esterified (inactive) lipid mediators involved in inflammation and inflammation resolution in cerebellum and neuronal cultures of ASM knockout (ASMko) mice and wildtype (WT) controls, and 2) quantified the esterification of labeled pro-resolving free d11-14(15)-epoxyeicosatrienoic acid in cultured neurons from ASMko and WT mice. We found elevated concentrations of esterified pro-resolving lipid mediators and hydroxyeicosatrienoic acids typically destined for pro-resolving lipid mediator synthesis (e.g. lipoxins) in the cerebellum and neurons of ASMko mice compared to controls. Free d11-14(15)-epoxyeicosatrienoic acid esterification within neurons of ASMko mice was significantly elevated compared to WT. Our findings show evidence of increased inactivation of free pro-resolving lipid mediators through esterification in ASMko mice, suggesting impaired resolution as a new pathway underlying ASM deficiency pathogenesis.

Heterologous booster with a novel formulation containing glycosylated trimeric S protein is effective against Omicron.

In this study, we evaluated the efficacy of a heterologous three-dose vaccination schedule against the Omicron BA.1 SARS-CoV-2 variant infection using a mouse intranasal challenge model. The vaccination schedules tested in this study consisted of a primary series of 2 doses covered by two commercial vaccines: an mRNA-based vaccine (mRNA1273) or a non-replicative vector-based vaccine (AZD1222/ChAdOx1, hereafter referred to as AZD1222). These were followed by a heterologous booster dose using one of the two vaccine candidates previously designed by us: one containing the glycosylated and trimeric spike protein (S) from the ancestral virus (SW-Vac 2µg), and the other from the Delta variant of SARS-CoV-2 (SD-Vac 2µg), both formulated with Alhydrogel as an adjuvant. For comparison purposes, homologous three-dose schedules of the commercial vaccines were used. The mRNA-based vaccine, whether used in heterologous or homologous schedules, demonstrated the best performance, significantly increasing both humoral and cellular immune responses. In contrast, for the schedules that included the AZD1222 vaccine as the primary series, the heterologous schemes showed superior immunological outcomes compared to the homologous 3-dose AZD1222 regimen. For these schemes no differences were observed in the immune response obtained when SW-Vac 2µg or SD-Vac 2µg were used as a booster dose. Neutralizing antibody levels against Omicron BA.1 were low, especially for the schedules using AZD1222. However, a robust Th1 profile, known to be crucial for protection, was observed, particularly for the heterologous schemes that included AZD1222. All the tested schedules were capable of inducing populations of CD4 T effector, memory, and follicular helper T lymphocytes. It is important to highlight that all the evaluated schedules demonstrated a satisfactory safety profile and induced multiple immunological markers of protection. Although the levels of these markers were different among the tested schedules, they appear to complement each other in conferring protection against intranasal challenge with Omicron BA.1 in K18-hACE2 mice. In summary, the results highlight the potential of using the S protein (either ancestral Wuhan or Delta variant)-based vaccine formulation as heterologous boosters in the management of COVID-19, particularly for certain commercial vaccines currently in use.

Effect of Nordic Sensi® Chair on Behavioral and Psychological Symptoms of Dementia in Nursing Homes Residents: A Randomized Controlled Trial.

BACKGROUND: Behavioral and psychological symptoms of dementia (BPSD) are present in most people with dementia (PwD), including Alzheimer's disease. There is consensus that non-pharmacological therapies represent the first line of treatment to address BPSD. OBJECTIVE: We explore the efficacy of the use of a rocking chair (Nordic Sensi® Chair, NSC) in the treatment of BPSD in nursing home residents with moderate and severe dementia. METHODS: We carried out a 16-week randomized, single-blind, controlled, clinical trial with PwD admitted to nursing homes. Participants were assigned to a treatment group (n = 40) that received three times a week one session per day of 20 minutes in the NSC and a control group (n = 37). The Neuropsychiatric Inventory-Nursing Home (NPI-NH) was used as primary efficacy outcome. Occupational distress for the staff was evaluated using the NPI-NH Occupational Disruptiveness subscale (NPI-NH-OD). Statistical analyses were conducted by means of a Mixed Effects Model Analysis. RESULTS: Treatment with the NSC was associated with a beneficial effect in most of BPSD, as reflected by differences between the treatment and control group on the NPI-NH total score (mean change score -18.87±5.56 versus -1.74±0.67, p = 0.004), agitation (mean change score -2.32±2.02 versus -0.78±1.44, p = 0.003) and irritability (mean change score -3.35±2.93 versus -1.42±1.31, p = 0.004). The NPI-NH-OD total score also improved the most in the treatment group (mean change score -9.67±7.67 versus -7.66±6.08, p = 0.003). CONCLUSIONS: The reduction in overall BPSD along with decreased caregiver occupational disruptiveness represent encouraging findings, adding to the potential of nonpharmacological interventions for nursing home residents living with dementia.

Modulation of Dietary Choline Uptake in a Mouse Model of Acid Sphingomyelinase Deficiency.

Acid sphingomyelinase deficiency (ASMD) is a lysosomal storage disorder caused by mutations in the gene-encoding acid sphingomyelinase (ASM). ASMD impacts peripheral organs in all patients, including the liver and spleen. The infantile and chronic neurovisceral forms of the disease also lead to neuroinflammation and neurodegeneration for which there is no effective treatment. Cellular accumulation of sphingomyelin (SM) is a pathological hallmark in all tissues. SM is the only sphingolipid comprised of a phosphocholine group linked to ceramide. Choline is an essential nutrient that must be obtained from the diet and its deficiency promotes fatty liver disease in a process dependent on ASM activity. We thus hypothesized that choline deprivation could reduce SM production and have beneficial effects in ASMD. Using acid sphingomyelinase knock-out (ASMko) mice, which mimic neurovisceral ASMD, we have assessed the safety of a choline-free diet and its effects on liver and brain pathological features such as altered sphingolipid and glycerophospholipid composition, inflammation and neurodegeneration. We found that the choline-free diet was safe in our experimental conditions and reduced activation of macrophages and microglia in the liver and brain, respectively. However, there was no significant impact on sphingolipid levels and neurodegeneration was not prevented, arguing against the potential of this nutritional strategy to assist in the management of neurovisceral ASMD patients.

Dinámicas socioculturales y alternativas económicas de mujeres en un contexto de economía minera extractivista

Este artículo describe las dinámicas socioculturales presentes en Puerto Claver, El Bagre, Antioquia, a causa del contexto económico minero. El enfoque fenomenològico permitió un acercamiento a las realidades sociales a partir de la cotidianidad de las personas, de sus experiencias vitales y relatos. Un resultado es que la práctica extractiva de oro, arraigada en el corregimiento como la principal alternativa económica, genera desarraigo frente a otras formas de sobrevivencia, como la cosecha y siembra de alimentos y plantas medicinales, la pesca y la apicultura. Al transformarse las prácticas económicas y productivas, así como la relación con el medio ambiente, cambian también las relaciones sociales, las dinámicas socioculturales preexistentes y el tejido social y comunitario. En este contexto, las asociaciones de mujeres tienen un papel significativo en la resignificación y dinamización de procesos económicos alternativos a la práctica minera, a través de los cuales puedan transformar las relaciones sociales y comunitarias.

This article describes the socio-cultural dynamics present in the village of Puerto Claver, municipality of El Bagre, Antioquia, as a result of the mining economic context. The phenomenological approach allowed an approach to the social realities from the daily life of the people, their life experiences and stories. One result is that the practice of gold mining, rooted in the township as the main economic alternative, generates uprooting compared to other forms of survival, such as harvesting and planting food and medicinal plants, fishing and beekeeping. As economic and productive practices are transformed, as well as the relationship with the environment, social relations, pre-existing socio-cultural dynamics and the social and community fabric also change. In this context, women's associations have a significant role to play in re-signifying and energizing alternative economic processes to mining practices, through which they can transform social and community relations.

Dengue virus NS5 degrades ERC1 during infection to antagonize NF-kB activation.

Dengue virus (DENV) is the most important human virus transmitted by mosquitos. Dengue pathogenesis is characterized by a large induction of proinflammatory cytokines. This cytokine induction varies among the four DENV serotypes (DENV1 to 4) and poses a challenge for live DENV vaccine design. Here, we identify a viral mechanism to limit NF-κB activation and cytokine secretion by the DENV protein NS5. Using proteomics, we found that NS5 binds and degrades the host protein ERC1 to antagonize NF-κB activation, limit proinflammatory cytokine secretion, and reduce cell migration. We found that ERC1 degradation involves unique properties of the methyltransferase domain of NS5 that are not conserved among the four DENV serotypes. By obtaining chimeric DENV2 and DENV4 viruses, we map the residues in NS5 for ERC1 degradation, and generate recombinant DENVs exchanging serotype properties by single amino acid substitutions. This work uncovers a function of the viral protein NS5 to limit cytokine production, critical to dengue pathogenesis. Importantly, the information provided about the serotype-specific mechanism for counteracting the antiviral response can be applied to improve live attenuated vaccines.

Sphingomyelin 16:0 is a therapeutic target for neuronal death in acid sphingomyelinase deficiency.

Acid sphingomyelinase deficiency (ASMD) is a lysosomal storage disorder caused by mutations in the SMPD1 gene encoding for the acid sphingomyelinase (ASM). While intravenous infusion of recombinant ASM is an effective treatment for the peripheral disease, the neurological complications of ASMD remain unaddressed. It has been shown that aberrantly high level of total brain sphingomyelin (SM) is a key pathological event leading to neurodegeneration. Using mice lacking ASM (ASMko), which mimic the disease, we here demonstrate that among the SM species, SM16:0 shows the highest accumulation and toxicity in ASMko neurons. By targeting lysosomes, SM16:0 causes permeabilization and exocytosis of these organelles and induces oxidative stress and cell death. We also show that genetic silencing of Ceramide Synthase 5, which is involved in SM16:0 synthesis and overexpressed in the ASMko brain, prevents disease phenotypes in ASMko cultured neurons and mice. The levels of SM16:0 in plasma also show a strong correlation with those in brain that is higher than in liver, even at early stages of the disease. These results identify SM16:0 both as a novel therapeutic target and potential biomarker of brain pathology in ASMD.

Ellagic acid and its metabolites urolithins A/B ameliorate most common disease phenotypes in cellular and mouse models for lysosomal storage disorders by enhancing extracellular vesicle secretion.

Niemann Pick diseases types A (NPDA) and C (NPDC) are lysosomal storage disorders (LSDs) leading to cognitive impairment, neurodegeneration, and early death. NPDA and NPDC have different genetic origins, being caused by mutations in the acid sphingomyelinase (ASM) or the cholesterol transport protein NPC1, respectively. However, they share a common pathological hallmark in the accumulation of lipids in the endolysosomal compartment. Here, we tested the hypothesis that polyphenols reduce lipid overload in NPD cells by enhancing the secretion of extracellular vesicles (ECVs). We show that among the polyphenols tested, the ellagic acid metabolites, urolithin A and B, were the safest and most efficient in increasing ECV secretion. They reduced levels of accumulating lipids and lysosomal size and permeabilization in cultured bone marrow-derived macrophages and neurons from ASMko and NPC1 mutant mice, which mimic NPDA and NPDC, respectively. Moreover, oral treatment with ellagic acid reduced lipid levels, ameliorated lysosomal alterations, and diminished microglia activation in the brain of NPD mice. These results support the therapeutic value of ECV secretion and polyphenols for NPDs, which may also help treat other LSDs characterized by intracellular lipid overload.

"MiR-7 controls cholesterol biosynthesis through posttranscriptional regulation of DHCR24 expression".

Dysregulation of cholesterol homeostasis is associated with several pathologies including cardiovascular diseases and neurological disorders such as Alzheimer's disease (AD). MicroRNAs (miRNAs) have emerged as key post-transcriptional regulators of cholesterol metabolism. We previously established the role of miR-7 in regulating insulin resistance and amyloidosis, which represents a common pathological feature between type 2 diabetes and AD. We show here an additional metabolic function of miR-7 in cholesterol biosynthesis. We found that miR-7 blocks the last steps of the cholesterol biosynthetic pathway in vitro by targeting relevant genes including DHCR24 and SC5D posttranscriptionally. Intracranial infusion of miR-7 on an adeno-associated viral vector reduced the expression of DHCR24 in the brain of wild-type mice, supporting in vivo miR-7 targeting. We also found that cholesterol regulates endogenous levels of miR-7 in vitro, correlating with transcriptional regulation through SREBP2 binding to its promoter region. In parallel to SREBP2 inhibition, the levels of miR-7 and hnRNPK (the host gene of miR-7) were concomitantly reduced in brain in a mouse model of Niemann Pick type C1 disease and in murine fatty liver, which are both characterized by intracellular cholesterol accumulation. Taken together, the results establish a novel regulatory feedback loop by which miR-7 modulates cholesterol homeostasis at the posttranscriptional level, an effect that could be exploited for therapeutic interventions against prevalent human diseases.

Prediction of tenderness in bovine longissimus thoracis et lumborum muscles using Raman spectroscopy.

OBJECTIVE: This study was conducted to evaluate Raman spectroscopy technique as a noninvasive tool to predict meat quality traits on Braford longissimus thoracis et lumborum muscle. METHODS: Thirty samples of muscle from Braford steers were analyzed by classical meat quality techniques and by Raman spectroscopy with 785 nm laser excitation. Water holding capacity (WHC), intramuscular fat content (IMF), cooking loss (CL), and texture profile analysis recording hardness, cohesiveness, and chewiness were determined, along with fiber diameter and sarcomere length by scanning electron microscopy. Warner-Bratzler shear force (WBSF) analysis was used to differentiate tender and tough meat groups. RESULTS: Higher values of cohesiveness and CL, together with lower values of WHC, IMF, and shorter sarcomere were obtained for tender meat samples than for the tougher ones. Raman spectra analysis allows tender and tough sample differentiation. The correlation between the quality attributes predicted by Raman and the physical measurements resulted in values of R2 = 0.69 for hardness and 0,58 for WBSF. Pearson's correlation coefficient of hardness (r = 0.84) and WBSF (r = 0.79) parameters with the phenylalanine Raman signal at 1,003 cm-1, suggests that the content of this amino acid could explain the differences between samples. CONCLUSION: Raman spectroscopy with 785 nm laser excitation is a suitable and accurate technique to identify beef with different quality attributes.

Crimean-Congo Hemorrhagic Fever, Spain, 2013-2021.

Crimean-Congo hemorrhagic fever (CCHF) is a viral infectious disease for which distribution of the main vector, Hyalomma spp. ticks, is expanding. We analyzed all 10 cases of CCHF diagnosed in Spain during 2013-2021; case-patient median age was 56.5 years, and 7 were men. We identified CCHF virus genotypes III and V. Six case-patients acquired the infection in urban areas. Sixty percent of patients were infected in summer and 40% in spring. Two patients met criteria for hemophagocytic syndrome. Seven patients survived. The epidemiologic pattern of CCHF in Spain is based on occasional cases with an elevated mortality rate. Genotype III and, to a less extent also genotype V, CCHF circulates in humans in a common geographic area in Spain. Those data suggest that the expansion pathways are complex and may change over time. Physicians should remain alert to the possibility of new CCHF cases.

Adult alcohol drinking and emotional tone are mediated by neutral sphingomyelinase during development in males.

Alcohol use, abuse, and addiction, and resulting health hazards are highly sex-dependent with unknown mechanisms. Previously, strong links between the SMPD3 gene and its coded protein neutral sphingomyelinase 2 (NSM) and alcohol abuse, emotional behavior, and bone defects were discovered and multiple mechanisms were identified for females. Here we report strong sex-dimorphisms for central, but not for peripheral mechanisms of NSM action in mouse models. Reduced NSM activity resulted in enhanced alcohol consumption in males, but delayed conditioned rewarding effects. It enhanced the acute dopamine response to alcohol, but decreased monoaminergic systems adaptations to chronic alcohol. Reduced NSM activity increased depression- and anxiety-like behavior, but was not involved in alcohol use for the self-management of the emotional state. Constitutively reduced NSM activity impaired structural development in the brain and enhanced lipidomic sensitivity to chronic alcohol. While the central effects were mostly opposite to NSM function in females, similar roles in bone-mediated osteocalcin release and its effects on alcohol drinking and emotional behavior were observed. These findings support the view that the NSM and multiple downstream mechanism may be a source of the sex-differences in alcohol use and emotional behavior.

Efficacy and safety clinical trial with efavirenz in patients diagnosed with adult Niemann-pick type C with cognitive impairment.

BACKGROUND: Niemann-Pick disease Type C (NPC) is a genetic, incurable, neurodegenerative disorder. This orphan disease is most frequently caused by mutations in the NPC1 protein, resulting in intralysossomal cholesterol accumulation. NPC1 is found in neuronal cell bodies, axon terminals and synaptosomes, suggesting it plays a role in lysosomal degradation pathway and in synaptic transmission. Neuronal function is especially vulnerable to NPC1 deficiency and synaptic changes seem a key element in disease development. Currently, Miglustat (Zavesca®) is the only approved treatment for NPC. However, preclinical evidence showed that low-dose Efavirenz reverted synaptic defects through pharmacological activation of the enzyme CYP46. METHODS: This is a single-center, phase II clinical trial to evaluate the efficacy and safety of Efavirenz in addition to standard of care in patients diagnosed with adult or late juvenile-onset NPC with cognitive impairment. All enrolled patients will be treated orally with 25 mg/d of Efavirenz for 52 weeks (1 year). Secondary objectives include evaluating clinical (neurological and neuropsychological questionnaires) and biological (imaging and biochemical biomarkers) parameters. DISCUSSION: NPC is still an unmet medical need. Although different therapeutic approaches are under study, this is the first clinical trial (to the best of our knowledge) studying the effects of Efavirenz in adult- and late-juvenile-onset NPC. Despite the small sample size and the single-arm design, we expect the results to show Efavirenz's capacity of activating the CYP46 enzyme to compensate for NPC1 deficiency and correct synaptic changes, therefore compensating cognitive and psychiatric changes in these patients. This study may provide direct benefit to enrolled patients in terms of slowing down the disease progression.

Immunological study of COVID-19 vaccine candidate based on recombinant spike trimer protein from different SARS-CoV-2 variants of concern.

The emergency of new SARS-CoV-2 variants that feature increased immune escape marks an urgent demand for better vaccines that will provide broader immunogenicity. Here, we evaluated the immunogenic capacity of vaccine candidates based on the recombinant trimeric spike protein (S) of different SARS-CoV-2 variants of concern (VOC), including the ancestral Wuhan, Beta and Delta viruses. In particular, we assessed formulations containing either single or combined S protein variants. Our study shows that the formulation containing the single S protein from the ancestral Wuhan virus at a concentration of 2µg (SW2-Vac 2µg) displayed in the mouse model the highest IgG antibody levels against all the three (Wuhan, Beta, and Delta) SARS-CoV-2 S protein variants tested. In addition, this formulation induced significantly higher neutralizing antibody titers against the three viral variants when compared with authorized Gam-COVID-Vac-rAd26/rAd5 (Sputnik V) or ChAdOx1 (AstraZeneca) vaccines. SW2-Vac 2µg was also able to induce IFN-gamma and IL-17, memory CD4 populations and follicular T cells. Used as a booster dose for schedules performed with different authorized vaccines, SW2-Vac 2µg vaccine candidate also induced higher levels of total IgG and IgG isotypes against S protein from different SARS-CoV-2 variants in comparison with those observed with homologous 3-dose schedule of Sputnik V or AstraZeneca. Moreover, SW2-Vac 2µg booster induced broadly strong neutralizing antibody levels against the three tested SARS-CoV-2 variants. SW2-Vac 2µg booster also induced CD4+ central memory, CD4+ effector and CD8+ populations. Overall, the results demonstrate that SW2-Vac 2 µg is a promising formulation for the development of a next generation COVID-19 vaccine.