Biology subjects

Xu, L.

Publications and source records attributed to Xu, L..

At least 325 records · Page 18Linked to original sources

bioRxiv2021

Extracellular loop 2 of G protein-coupled olfactory receptors is critical for odorant recognition

G protein-coupled olfactory receptors (ORs) enable us to detect innumerous odorants. They are also ectopically expressed, emerging as attractive drug targets. ORs can be promiscuous or highly specific, which is part of Natures strategy for odor discrimination. This work demonstrates that the extracellular loop 2 (ECL2) plays critical roles in OR promiscuity and specificity. Using site-directed mutagenesis and molecular modeling, we constructed 3D OR models in which ECL2 forms a lid of the orthosteric pocket. ECL2 controls the shape and the volume of the odorant-binding pocket, maintains the pocket hydrophobicity and acts as a gatekeeper of odorant binding. The interplay between the specific orthosteric pocket and the variable, less specific ECL2 controls OR specificity and promiscuity. The 3D models enabled virtual screening of new OR agonists and antagonists, exhibiting 78% hit rate in cell assays. This approach can be generalized to structure-based ligand screening for other GPCRs that lack high-resolution 3D structures.

molecular biology↗

bioRxiv2021

Mapping enzyme catalysis with metabolomic biosensing

Enzymes are represented across a vast space of protein sequences and structural forms and have activities that far exceed the best chemical catalysts; however, engineering them to have novel or enhanced activity is limited by technologies for sensing product formation. Here, we describe a general and scalable approach for characterizing enzyme activity that uses the metabolism of the host cell as a biosensor by which to infer product formation. Since different products consume different molecules in their synthesis, they perturb host metabolism in unique ways that can be measured by mass spectrometry. This provides a general way by which to sense product formation, to discover unexpected products and map the effects of mutagenesis.

bioengineering↗

bioRxiv2021

WUSCHEL-Related Homeobox Genes Cooperate with Cytokinin Signalling to Promote Bulbil Formation in Lilium lancifolium

The bulbil is an important vegetative reproductive organ in triploid Lilium lancifolium. Based on our previously obtained transcriptome data, we screened two WUSCHCEL-related homeobox (WOX) genes closely related to bulbil formation, LlWOX9 and LlWOX11. However, the biological functions and regulatory mechanisms of LlWOX9 and LlWOX11 are unclear. In this study, we cloned the full-length coding sequences of LlWOX9 and LlWOX11. Transgenic Arabidopsis showed increased branch numbers, and the overexpression of LlWOX9 and LlWOX11 in stem segments promoted bulbil formation, while the silencing of LlWOX9 and LlWOX11 inhibited bulbil formation, indicating that LlWOX9 and LlWOX11 are positive regulators of bulbil formation. Cytokinins acting through type-B response regulators (type-B RRs) could bind to the promoters of LlWOX9 and LlWOX11 and promote their transcription. LlWOX11 could enhance cytokinin pathway signalling by inhibiting the transcription of type-A LlRR9. Our study enriches the understanding of the regulation of plant development by the WOX gene family and lays a foundation for further research on the molecular mechanism of bulbil formation in lily.

developmental biology↗

bioRxiv2021

Proton export drives the Warburg Effect

Aggressive cancers commonly ferment glucose to lactic acid at high rates, even in the presence of oxygen. This is known as aerobic glycolysis, or the "Warburg Effect". It is widely assumed that this is a consequence of the upregulation of glycolytic enzymes. Oncogenic drivers can increase the expression of most proteins in the glycolytic pathway, including the terminal step of exporting H+ equivalents from the cytoplasm. Proton exporters maintain an alkaline cytoplasmic pH, which can enhance all glycolytic enzyme activities, even in the absence of oncogene-related expression changes. Based on this observation, we hypothesized that increased uptake and fermentative metabolism of glucose could be driven by the expulsion of H+ equivalents from the cell. To test this hypothesis, we stably transfected lowly-glycolytic MCF-7, U2-OS, and glycolytic HEK293 cells to express proton exporting systems: either PMA1 (yeast H+-ATPase) or CAIX (carbonic anhydrase 9). The expression of either exporter in vitro enhanced aerobic glycolysis as measured by glucose consumption, lactate production, and extracellular acidification rate. This resulted in an increased intracellular pH, and metabolomic analyses indicated that this was associated with an increased flux of all glycolytic enzymes upstream of pyruvate kinase. These cells also demonstrated increased migratory and invasive phenotypes in vitro, and these were recapitulated in vivo by more aggressive behavior, whereby the acid-producing cells formed higher grade tumors with higher rates of metastases. Neutralizing tumor acidity with oral buffers reduced the metastatic burden. Therefore, cancer cells with increased H+ export increase intracellular alkalization, even without oncogenic driver mutations, and this is sufficient to alter cancer metabolism towards a Warburg phenotype.

cancer biology↗

bioRxiv2021

Copper(II) Gluconate Boosts the Anti-SARS-CoV-2 Effect of Disulfiram in Vitro

Disulfiram is a 70-year-old anti-alcoholism drug, while copper(II) gluconate (Cu(Glu)2) is a commonly used food additive or copper supplement. Here we disclose that the combination of disulfiram and copper(II) gluconate drastically enhances the anti-SARS-CoV-2 activity at the cellular level as compared to disulfiram or copper(II) gluconate alone. A 1:1 mixture of disulfiram and copper(II) gluconate shows an EC50 value of 154 nM against SARS-CoV-2 at the cellular level, much lower than the 17.45 M reported for disulfiram alone. Notably, previous clinical trials have shown that a combination of 250 mg disulfiram (0.843 mmol) and 8 mg copper(II) gluconate (0.0176 mmol) oral capsules per day is well tolerated (NCT03034135, NCT00742911). A preliminary mechanism is proposed to rationalize the observed promotional effect.

biochemistry↗

bioRxiv2021

VGLL2-NCOA2 leverages developmental programs for pediatric sarcomagenesis

Clinical sequencing efforts are uncovering numerous fusion genes in childhood solid tumors, yet few methods exist to delineate fusion-oncogenes from structural changes of unknown significance. One such novel fusion gene is VGLL2-NCOA2, which was described by us and others in patients with congenital sarcomas but has lacked functional validation. To determine if this fusion is an oncogene, and how it is driving disease, we developed a vertebrate zebrafish model and mouse allograft model of human VGLL2-NCOA2 driven sarcomagenesis. We found that VGLL2-NCOA2 is indeed an oncogene and is sufficient to generate mesenchymal tumors that recapitulate the human disease at the histological and transcriptional level. Zebrafish VGLL2-NCOA2 tumors display features of arrested skeletal muscle development, and a subset transcriptionally cluster with somitogenesis in developing embryos. By comparing tumor and embryonic gene expression signatures, we identified developmentally regulated targets that VGLL2-NCOA2 potentially leverages for tumorigenesis. These targets highlight the core biology of the disease and could represent therapeutic opportunities. Specifically, a RAS family GTPase, arf6/ARF6, involved in actin remodeling and rapid cycling of endocytic vesicles at the plasma membrane, is highly expressed during zebrafish somitogenesis and in VGLL2-NCOA2 tumors. In zebrafish tumors, arf6 protein is highly expressed and is absent from mature skeletal muscle. In VGLL2-NCOA2 mouse allograft models and patient tumors, ARF6 mRNA is overexpressed as compared to skeletal muscle or normal controls. More broadly, ARF6 is overexpressed in adult and pediatric sarcoma subtypes as compared to mature skeletal muscle. Overall, our cross-species comparative oncology approach provides evidence that VGLL2-NCOA2 is an oncogene which leverages developmental programs for tumorigenesis, and that one of these programs, the reactivation or persistence of arf6/ARF6, could represent a therapeutic opportunity.

cancer biology↗

bioRxiv2021

Effect of ORF7 of SARS-CoV-2 on the chemotaxis of monocytes and neutrophils in vitro

Coronavirus disease 2019 (COVID-19) caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is currently the most significant public health threats in worldwide. Patients with severe COVID-19 usually have pneumonia concomitant with local inflammation and sometimes a cytokine storm. Specific components of the SARS-CoV-2 virus trigger lung inflammation, and recruitment of immune cells to the lungs exacerbates this process, although much remains unknown about the pathogenesis of COVID-19. Our study of lung type II pneumocyte cells (A549) demonstrated that ORF7, an open reading frame (ORF) in the genome of SARS-CoV-2, induced the production of CCL2, a chemokine that promotes the chemotaxis of monocytes, and decreased the expression of IL-8, a chemokine that recruits neutrophils. A549 cells also had an increased level of IL-6. The results of our chemotaxis transwell assay suggested that ORF7 augmented monocyte infiltration and reduced the number of neutrophils. We conclude that the ORF7 of SARS-CoV-2 may have specific effects on the immunological changes in tissues after infection. These results suggest that the functions of other ORFs of SARS-CoV-2 should also be comprehensively examined.

cell biology↗

bioRxiv2021

Genomic evidence for divergent co-infections of SARS-CoV-2 lineages

Recently, patients co-infected by two SARS-CoV-2 lineages have been sporadically reported. Concerns are raised because previous studies have demonstrated co-infection may contribute to the recombination of RNA viruses and cause severe clinic symptoms. In this study, we have estimated the compositional lineage(s), tendentiousness, and frequency of co-infection events in population from a large-scale genomic analysis for SARS-CoV-2 patients. SARS-CoV-2 lineage(s) infected in each sample have been recognized from the assignment of within-host site variations into lineage-defined feature variations by introducing a hypergeometric distribution method. Of all the 29,993 samples, 53 (~0.18%) co-infection events have been identified. Apart from 52 co-infections with two SARS-CoV-2 lineages, one sample with co-infections of three SARS-CoV-2 lineages was firstly identified. As expected, the co-infection events mainly happened in the regions where have co-existed more than two dominant SARS-CoV-2 lineages. However, co-infection of two sub-lineages in Delta lineage were detected as well. Our results provide a useful reference framework for the high throughput detecting of SARS-CoV-2 co-infection events in the Next Generation Sequencing (NGS) data. Although low in average rate, the co-infection events showed an increasing tendency with the increased diversity of SARS-CoV-2. And considering the large base of SARS-CoV-2 infections globally, co-infected patients would be a nonnegligible population. Thus, more clinical research is urgently needed on these patients.

microbiology↗

bioRxiv2021

Defective Desmosomal Adhesion Causes Arrhythmogenic Cardiomyopathy by involving an Integrin-αVβ6/TGF-β Signaling Cascade

BackgroundArrhythmogenic Cardiomyopathy (ACM) is characterized by progressive loss of cardiomyocytes with fibrofatty replacement, systolic dysfunction and life-threatening arrhythmias. A substantial proportion of ACM is caused by mutations in genes of the desmosomal cell-cell adhesion complex, but the underlying mechanisms are not well understood. So far, treatment options are only symptomatic. Here, we investigate the relevance of defective desmosomal adhesion for ACM development and progression. MethodsWe mutated the binding site of desmoglein-2 (DSG2), a crucial desmosomal adhesion molecule in cardiomyocytes. This DSG2-W2A mutation abrogates the tryptophan swap, a central interaction mechanism of DSG2 based on structural data. Impaired adhesive function of DSG2-W2A was confirmed by cell-cell dissociation assays and force spectroscopy measurements by atomic force microscopy. We next generated a DSG2-W2A knock-in mouse model, which was analyzed by echocardiography and histological and bio-molecular techniques including RNA sequencing, transmission electron and super-resolution microscopy. The results were compared to ACM patient samples and their relevance was confirmed in cardiac slice cultures. ResultsThe DSG2-W2A mutation induced impaired binding and desmosomal adhesion dysfunction on cellular and molecular level. Mice bearing this mutation develop a severe cardiac phenotype recalling the characteristics of ACM, including cardiac fibrosis, impaired systolic function and arrhythmia. A comparison of the transcriptome of mutant mice with ACM patient data suggested deregulated integrin-V{beta}6 and subsequent TGF-{beta} signaling as driver of cardiac fibrosis. Accordingly, blocking antibodies targeting integrin-V{beta}6 or inhibition of TGF-{beta} receptor signaling both led to reduced expression of pro-fibrotic markers in cardiac slice cultures. ConclusionsHere, we show that disruption of desmosomal adhesion is sufficient to induce ACM, which confirms the dysfunctional adhesion hypothesis. Mechanistically, deregulation of integrin-V{beta}6 signaling was identified as a central step towards fibrosis. This highlights the value of this model to discern mechanisms of cardiac fibrosis and to identify and test novel treatment options for ACM.

cell biology↗

bioRxiv2021

Innate and adaptive immunity to transfused allogenic red blood cells in mice requires MyD88

Red blood cell (RBC) transfusion therapy is essential for the survival of patients with hematological disorders such as sickle cell anemia. A potentially fatal complication of transfusion is development of non-ABO alloantibodies to polymorphic RBC antigens, yet mechanisms of alloantibody formation remain unclear. Human and mouse RBCs acquire a "storage lesion" prior to transfusion, which in mice contributes to immunogenicity. We previously reported that mouse splenic dendritic cells (DCs) are required for RBC alloimmunization and are activated by sterile and leukoreduced mouse RBCs after storage. Yet how syngeneic RBCs activate innate immune pathways to induce DC activation is unknown. We now show that DC activation to transfused RBCs occurs regardless of alloantigen presence, suggesting that RBC damage induced during storage triggers innate immune receptors. We discovered an unexpected dependence of RBC alloimmunization on the Toll-like receptor (TLR) signaling adaptor molecule MyD88. TLRs are a class of pattern recognition receptors (PRRs) that regulate DC activation and signal through two adaptor molecules, MyD88 and TRIF. We show that the inflammatory cytokine response, DC activation, and the subsequent alloantibody response to transfused syngeneic RBCs require MyD88 but not TRIF, suggesting a restricted set of PRRs are responsible for sensing RBCs and triggering alloimmunization.

immunology↗

bioRxiv2021

Coordinately regulated interbacterial antagonism defense pathways constitute a bacterial innate immune system

Bacterial survival is fraught with antagonism, including that deriving from viruses and competing bacterial cells1-3 4. It is now appreciated that bacteria mount complex antiviral responses; however, whether a coordinated defense against bacterial threats is undertaken is not well understood. Previously we showed that Pseudomonas aeruginosa possess a danger sensing pathway that is a critical fitness determinant during competition against other bacteria5, 6. Here, we conducted genome-wide screens in P. aeruginosa that reveal three conserved and widespread interbacterial antagonism resistance clusters (arc1-3). We find that although arc1-3 are coordinately activated by the Gac/Rsm danger sensing system, they function independently and provide idiosyncratic defense capabilities, distinguishing them from general stress response pathways. Our findings demonstrate that Arc3 family proteins provide specific protection against phospholipase toxins by preventing the accumulation of lysophospholipids in a manner distinct from previously characterized membrane repair systems. These findings liken the response of P. aeruginosa to bacterial threats to that of eukaryotic innate immunity, wherein threat detection leads to the activation of specialized defense systems.

microbiology↗

bioRxiv2021

Neuron-specific coding sequences are the most highly conserved in the mammalian brain

Neurons are unique in that they are the only cell type in the body to display massive diversity in cell size, morphology, phenotype, and function both within individuals and across species. Here we use datasets encompassing up to 92 mammalian and 31 sauropsidian species to examine whether neuron-specific diversity occurs with higher evolutionary variation of neuron-specific coding and regulatory sequences compared to non-neuronal cell-specific sequences. We find that the opposite is true: Neuronal diversity in mammalian and sauropsidian evolution arose despite extreme levels of negative selection on neuron-specific protein-coding sequences on par with ATPase coding sequences, the benchmark of evolutionary conservation. We propose that such strong evolutionary conservation is imposed by excitability, which continually exposes cells to the risk of excitotoxic death, and speculate that neuronal cell size diversity is a self-organized consequence of variability in levels of activity, possibly constrained by energy supply to the developing brain.

neuroscience↗

bioRxiv2021

Rice LEAFY COTYLEDON1 hinders photosynthesis in the embryo development to promote seed dormancy

LEAFY COTYLEDON1 (LEC1) is the central regulator of seed development. During seed development, rice embryo photosynthesis is completely blocked, which is different from Arabidopsis green embryo. However, effects of LEC1 on photosynthesis in developing seeds is largely elusive. We generated OsLEC1 mutants using the CRISPR/Cas9 technique. Oslec1 mutant seeds lost the ability of dormancy and triggered photosynthesis in embryos at the early developing stage. Transcriptome analysis demonstrated that Oslec1 mutation promoted photosynthesis and altered diverse hormonal pathways and stress response contributing to seed dormancy. Further, genome-wide identification of OsLEC1 binding sites demonstrated that OsLEC1 directly bound to genes involved in photosynthesis, photomorphogenesis, as well as abscisic acid (ABA) and gibberellin (GA) pathways, in seed maturation. We illustrated an OsLEC1-controlling gene network during seed development, including the interconnection between photosynthesis and ABA/GA biosynthesis/signalling. Our findings suggested that OsLEC1 is an inhibitor of photosynthesis during embryo development to promote rice seed maturation. This study would provide new understanding for the OsLEC1 regulatory mechanisms on photosynthesis in the monocot seed development.

plant biology↗

bioRxiv2021

Perinatal maternal chronic exposure to dibutyl phthalate promotes visceral obesity in adult female offspring

IntroductionMaternal exposure to dibutyl phthalate (DBP) may result in glucolipid dysfunction in female offspring. However, the underlying mechanisms remain elusive. We hypothesized that chronic maternal DBP exposure could induce abnormal metabolism of glucolipid. Materials and methodsSprague-Dawley rats were intraperitoneally injected with different doses of DBP, estradiol, and corn oil from gestational day 7 until the end of lactation. The weights, visceral fat percentage, serum lipid, insulin and glucose, protein levels of PI3K signal pathway in muscle were detected in F1 female offspring. ResultsAlthough the birth weight of F1 female offspring was not different among groups, the weights were heavier in DBP groups from postnatal day 7 to adult (P<0.001). The visceral adipose percentage in adult female offspring was increased by perinatal exposure to DBP (P<0.001). Decreased serum levels of triglyceride (P<0.0001), fasting glucose (P=0.004), prolactin (P=0.006), HOMA-IR (P=0.014) were found in female offspring exposed to DBP, but no difference for fasting insulin, total cholesterol, adiponectin. Increased protein levels of p-AKT, but decreased PTEN and GPR30 were observed in muscle of female offspring in DBP group, but without significant difference. None difference was observed for the protein levels of PI3K, AKT, GLUT4, InsR and IRS-1. ConclusionMaternal perinatal exposure to DBP induced obesity and accumulation of visceral adipose tissue for the adult female offspring. Serum glucolipid and local signal transduction of PTEN/PI3K/AKT pathway in muscle were not adversely affected by perinatal exposure to DBP for adult female offspring.

pharmacology and toxicology↗

bioRxiv2021

Bmmp influences wing shape by regulating anterior-posterior and proximal-distal axis development

Insect wings are subject to strong selective pressure, resulting in the evolution of remarkably diverse wing shapes that largely determine flight capacity. However, the genetic basis and regulatory mechanisms underlying wing shape development are not well understood. The silkworm Bombyx mori micropterous (mp) mutant exhibits shortened wing length and enlarged vein spacings, albeit without changes in total wing area. Thus, the mp mutant comprises a valuable genetic resource for studying wing shape development. In this study, we used molecular mapping to identify the gene responsible for the mp phenotype and designated it Bmmp. Phenotype-causing mutations were identified as indels and single nucleotide polymorphisms in non-coding regions. These mutations resulted in decreased Bmmp mRNA levels and changes in transcript isoform composition. Bmmp null mutants were generated by CRISPR/Cas9 and exhibited significantly smaller wings. By examining the expression of genes critical to wing development in wildtype and Bmmp null mutants, we found that Bmmp exerts its function by coordinately modulating anterior-posterior and proximal-distal axis development. We also studied a Drosophila mp mutant and found that Bmmp is functionally conserved in Drosophila. The Drosophila mp mutant strain exhibits curly wings of reduced size and a complete loss of flight capacity. Our results increase our understanding of the mechanisms underpinning insect wing development and reveal potential targets for pest control.

genetics↗

bioRxiv2021

Amplification-free detection of viral RNA by super resolution imaging-based CRISPR/Cas13a System

RNA detection is crucial for biological research and clinical diagnosis. The current methods include both direct and amplification-based RNA detection. These methods require complicated procedures, suffering from low sensitivity, slow turnaround, and amplification bias. The CRISPR/Cas13a system is a direct RNA detection method via target RNA induced collateral cleavage activity. However, to detect low concentration RNA with CRISPR/Cas13a, target amplification is always required. Herein, we optimize the components of the CRISPR/Cas13a assay to enhance the sensitivity of viral RNA detection which improve the detection limit from 1 pM up to 100 fM. In addition, the integration of CRISPR/Cas13a biosensing and single molecule super resolution imaging is a novel strategy for direct and amplification-free RNA detection. After surface modification, fluorescent RNA reporters are immobilized on the glass coverslip surface and fluorescent signals are captured by total internal reflection fluorescence microscopy (TIRFM), shifting the measurement from spectroscopy to imaging. We quantify the fluorescence signal intensity before and after collateral cleavage of the CRISPR system when viral RNA is present and achieve a detection limit of 10 fM. Therefore, we provide a novel TIRFM-based system to visualize the CRISPR trans-cleavage for direct and robust RNA detection.

bioengineering↗

bioRxiv2021

Multi-omics consensus ensemble refines the classification of muscle-invasive bladder cancer with stratified prognosis, tumour microenvironment and distinct sensitivity to frontline therapies

The molecular classification of muscle-invasive bladder cancer (MIBC) based on transcriptomic signatures has been extensively studied. The complementary nature of information provided by different molecular profiles motivated us to refine MIBC classification by aggregating multi-omics data. We generated a consensus ensemble through ten multi-omics integrative clustering approaches on 396 MIBCs from TCGA. A total of 701 MIBCs from different sequencing technologies were used for external validation. Associations between subtypes and prognosis, molecular profiles, the tumour microenvironment, and potential response to frontline therapies were further analyzed. Nearest template prediction and random forest classification were used to develop a predictive signature/classifier for MIBC refinement. We identified four integrative consensus subtypes of MIBC, which were further designated basal-inflamed, basal-noninflamed, luminal-excluded and luminal-desert by immune profiling. Of note, the refinement of basal-like MIBC classification adds to the literature by identifying a basal-noninflamed MIBC subtype presenting with a significantly poor outcome and a global immune-cold phenotype, which might be triggered by Chr4 deletion and high activation of the oncogenic NRF2 pathway. In contrast, basal-inflamed MIBC showed high immunocyte infiltration and high expression of potential targets for immunotherapy. Using an external metastatic MIBC cohort in which patients received anti-PD-L1 treatment, we suggested that basal-inflamed MIBC had a higher likelihood of responding to immunotherapy than other MIBCs. The R package "refineMIBC" was offered as a research tool to refine MIBC from a single-sample perspective (https://github.com/xlucpu/refineMIBC). This consensus ensemble refines the intrinsic MIBC subtypes, which provides a blueprint for the clinical development of rational targeted and immunotherapeutic strategies.

cancer biology↗

bioRxiv2021

An intranasal ASO therapeutic targeting SARS-CoV-2

The COVID-19 pandemic is exacting an increasing toll worldwide, with new SARS-CoV-2 variants emerging that exhibit higher infectivity rates and that may partially evade vaccine and antibody immunity1. Rapid deployment of non-invasive therapeutic avenues capable of preventing infection by all SARS-CoV-2 variants could complement current vaccination efforts and help turn the tide on the COVID-19 pandemic2. Here, we describe a novel therapeutic strategy targeting the SARS-CoV-2 RNA using locked nucleic acid antisense oligonucleotides (LNA ASOs). We identified an LNA ASO binding to the 5 leader sequence of SARS-CoV-2 ORF1a/b that disrupts a highly conserved stem-loop structure with nanomolar efficacy in preventing viral replication in human cells. Daily intranasal administration of this LNA ASO in the K18-hACE2 humanized COVID-19 mouse model potently (98-99%) suppressed viral replication in the lungs of infected mice, revealing strong prophylactic and treatment effects. We found that the LNA ASO also represses viral infection in golden Syrian hamsters, and is highly efficacious in countering all SARS-CoV-2 "variants of concern" tested in vitro and in vivo, including B.1.427, B.1.1.7, and B.1.351 variants3. Hence, inhaled LNA ASOs targeting SARS-CoV-2 represents a promising therapeutic approach to reduce transmission of variants partially resistant to vaccines and monoclonal antibodies, and could be deployed intranasally for prophylaxis or via lung delivery by nebulizer to decrease severity of COVID-19 in infected individuals. LNA ASOs are chemically stable and can be flexibly modified to target different viral RNA sequences4, and they may have particular impact in areas where vaccine distribution is a challenge, and could be stockpiled for future coronavirus pandemics.

microbiology↗