Dendritic-cell-targeting virus-like particles as potent mRNA vaccine carriers.

Messenger RNA vaccines lack specificity for dendritic cells (DCs)-the most effective cells at antigen presentation. Here we report the design and performance of a DC-targeting virus-like particle pseudotyped with an engineered Sindbis-virus glycoprotein that recognizes a surface protein on DCs, and packaging mRNA encoding for the Spike protein of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) or for the glycoproteins B and D of herpes simplex virus 1. Injection of the DC-targeting SARS-CoV-2 mRNA vaccine in the footpad of mice led to substantially higher and durable antigen-specific immunoglobulin-G titres and cellular immune responses than untargeted virus-like particles and lipid-nanoparticle formulations. The vaccines also protected the mice from infection with SARS-CoV-2 or with herpes simplex virus 1. Virus-like particles with preferential uptake by DCs may facilitate the development of potent prophylactic and therapeutic vaccines.

Developing deep LSTMs with later temporal attention for predicting COVID-19 severity, clinical outcome, and antibody level by screening serological indicators over time.

OBJECTIVE: The clinical course of COVID-19, as well as the immunological reaction, is notable for its extreme variability. Identifying the main associated factors might help understand the disease progression and physiological status of COVID-19 patients. The dynamic changes of the antibody against Spike protein are crucial for understanding the immune response. This work explores a temporal attention (TA) mechanism of deep learning to predict COVID-19 disease severity, clinical outcomes, and Spike antibody levels by screening serological indicators over time. METHODS: We use feature selection techniques to filter feature subsets that are highly correlated with the target. The specific deep Long Short-Term Memory (LSTM) models are employed to capture the dynamic changes of disease severity, clinical outcome, and Spike antibody level. We also propose deep LSTMs with a TA mechanism to emphasize the later blood test records because later records often attract more attention from doctors. RESULTS: Risk factors highly correlated with COVID-19 are revealed. LSTM achieves the highest classification accuracy for disease severity prediction. Temporal Attention Long Short-Term Memory (TA-LSTM) achieves the best performance for clinical outcome prediction. For Spike antibody level prediction, LSTM achieves the best permanence. CONCLUSION: The experimental results demonstrate the effectiveness of the proposed models. The proposed models can provide a computer-aided medical diagnostics system by simply using time series of serological indicators.

Nanoplastics and chrysene pollution: Potential new triggers for nonalcoholic fatty liver disease and hepatitis, insights from juvenile Siniperca chuatsi.

Nanopolystyrene (NP) and chrysene (CHR) are ubiquitous contaminants in the natural environment; however, research on their hepatotoxicity and associated adverse effects remains relatively inadequate. The present study aimed to investigate the hepatotoxic effects of NP and/or CHR at environmentally relevant concentrations, as well as the underlying molecular mechanisms, in juvenile Siniperca chuatsi (mandarin fish). After a 21-day exposure period, the livers of exposed S. chuatsi exhibited macrostructural and microstructural damage accompanied by oxidative stress. Importantly, our study provides the first evidence that NP exposure leads to the development of nonalcoholic fatty liver disease (NAFLD) and hepatitis in S. chuatsi. Similarly, CHR exposure has also been found, for the first time, to cause hepatic sinusoidal dilatation (HSD) and hepatitis. Exposure to the combination of NP and CHR alleviated the symptoms of NAFLD, HSD, and hepatitis. Furthermore, our comprehensive multi-omic analysis revealed that the pathogenesis of NP-induced NAFLD was mainly due to induction of the triglyceride synthesis pathway and inhibition of the very-low-density lipoprotein secretion process. CHR induced HSD primarily through a reduction in vasoprotective ability and smooth muscle contractility. Hepatitis was induced by activation of the JAK-STAT/NF-kappa B signaling pathways, which upregulated the expression of inflammation-specific genes. Collectively, results of this study offer novel insight into the multiple hepatotoxicity endpoints of NP and/or CHR exposure at environmentally relevant concentrations in organisms, and highlight the importance of nanoplastic/CHR pollution for liver health.

Histopathological, physiological, and multi-omics insights into the hepatotoxicity mechanism of nanopolystyrene and/or diclofenac in Mylopharyngodon piceus.

Nanopolystyrene (NP) and diclofenac (DCF) are common environmental contaminants in the aquatic ecosystem; therefore, the present study aimed to investigate the hepatotoxicity of NP and/or DCF exposure on aquatic organisms and the underlying mechanisms. Juvenile Mylopharyngodon piceus were used as a model organism to study the effects of NP and/or DCF exposure at environmentally relevant concentrations for 21 days. Subchronic exposure to NP and/or DCF resulted in liver histological damage. In the NP group, the presence of large lipid droplets was observed, whereas the DCF group exhibited marked hepatic sinusoidal dilatation accompanied by inflammation. Additionally, this exposure induced liver oxidative stress, as evidenced by the changes in several physiological parameters, including catalase (CAT), glutathione peroxidase (GSH-Px), superoxide dismutase (SOD), total antioxidant capacity (T-AOC), reactive oxygen species (ROS), and malondialdehyde (MDA). Integrated transcriptomic and metabolomic analysis was performed to further investigate the molecular mechanism underlying hepatotoxicity. Multi-omics analysis demonstrated, for the first time to our knowledge, that NP induced hepatic steatosis mainly through activating the glycerol-3-phosphate pathway and inhibiting VLDL assembly by targeting several key enzyme genes including GPAT, DGAT, ACSL, APOB, and MTTP. Furthermore, NP exposure disrupted arachidonic acid metabolism, which induced the release of inflammatory factors and inhibited the release of anti-inflammatory factors, ultimately causing liver inflammation in M. piceus. In contrast, DCF induced interleukin production and downregulated KLF2, causing hepatic sinusoidal dilatation with inflammation in juvenile M. piceus, which is consistent with the finding of JAK-STAT signaling pathway activation. In addition, the upregulated AMPK signaling pathway in the DCF group suggested perturbation of energy metabolism. Collectively, these findings provide novel insights into the molecular mechanism of the multiple hepatotoxicity endpoints of NP and/or DCF exposure in aquatic organisms.

The fusion gene hsf5-rnf43 in Nile tilapia: A potential regulator in the maintenance of testis function and sexual differentiation.

We identified that the genes heat shock transcription factor 5 (hsf5) and ring finger protein 43 (rnf43) happened fusion in Nile tilapia (Oreochromis niloticus), called hsf5-rnf43, and provided the characteristic and functional analysis of hsf5-rnf43 gene in fish for the first time. Analysis of spatiotemporal expression showed that hsf5-rnf43 was specifically expressed in the testis and located in primary spermatocytes of adult Nile tilapia and gradually increased during testis development from 5 to 180 days after hatching. We also found DNA methylation regulated sex-biased expression of hsf5-rnf43 in the early development of Nile tilapia, and was affected by high temperature during the thermosensitive period of Nile tilapia sex differentiation. Therefore, we first reported that the fusion gene hsf5-rnf43 was sex-biased expressed in the testis regulated by DNA methylation and affected by high temperature, which may be involved in the maintenance of testis function and sex differentiation of Nile tilapia.

Comparative proteomics elucidates the potential mechanism of heritable carapace color of three strains Chinese mitten crab Eriocheir sinensis.

The carapace coloration is important for the environmental adaptation and reproductive behaviors of crustaceans. We selected red, green and white three carapace color strains of Chinese mitten crab (Eriocheir sinensis) strains. These three carapace colors have stable heritability, but the mechanism for their coloration remains unclear.Through histological observations, we have found significant differences in the composition of pigment cells and pigments within the inner membrane of the three color strains, which may be one of the reasons for the color variation. The levels of various carotenoids in both the shell and inner membrane tissues of red and green strains were significantly higher than those of the white strain, while there was no significant difference between the red and green strains. Proteomics studies have identified 2, 034 and 947 different proteins in the shell and inner membrane, respectively. In the shell, there were 18, 13 and 43 differential proteins between red and white strains, green and white strains and green and red strains, respectively. In the inner membrane, there were 44, 24 and 16 differential proteins between red and white strains, green and white strains and green and red strains, respectively. It is clear that the deposited quantity of carotenoids affects the shell formation of three color strains. Some members of the hemocyanin family showed significant variation among different strains. The study yielded two crustacyanin proteins, which were extracted from both the shell and membrane. Of the two proteins, only Crustacyanin-A1 expression showed a difference between the red and green shells strains. In conclusion, these results indicated that the carapace color formation of E. sinensis may be accomplished through pigment binding proteins (PBPs) and pigment cells, which enhance the understanding of color formation mechanism for crustacean.

Impacts of exposure to nanopolystyrene and/or chrysene at ambient concentrations on neurotoxicity in Siniperca chuatsi.

Health risks caused by widespread environmental pollutants such as nanopolystyrene (NP) and chrysene (CHR) in aquatic ecosystems have aroused considerable concern. The present study established juvenile Mandarin fish (Siniperca chuatsi) models of NP and/or CHR exposure at ambient concentrations for 21 days to systematically investigate the underlying neurotoxicity mechanisms. The results showed that single and combined exposure to NP and CHR not only reduced the density of small neuronal cells in the grey matter layer of the optic tectum, but also induced brain oxidative stress according to physiological parameters including CAT, GSH-Px, SOD, T-AOC, and MDA. The co-exposure alleviated the histopathological damage, compared to NP and CHR single exposure group. These results indicate that NP and/or CHR causes neurotoxicity in S. chuatsi, in accordance with decreased acetylcholinesterase activity and altered expression of several marker genes of nervous system functions and development including c-fos, shha, elavl3, and mbpa. Transcriptomics analysis was performed to further investigate the potential molecular mechanisms of neurotoxicity. We propose that single NP and co-exposure induced oxidative stress activates MMP, which degrades tight junction proteins according to decreased expression of claudin, JAM, caveolin and TJP, ultimately damaging the integrity of the blood-brain barrier in S. chuatsi. Remarkably, the co-exposure exacerbated the blood-brain barrier disruption. More importantly, single NP and co-exposure induced neuronal apoptosis mainly activates the expression of apoptosis-related genes through the death receptor apoptosis pathway, while CHR acted through both death receptor apoptosis and endoplasmic reticulum apoptosis pathways. Additionally, subchronic CHR exposure caused neuroinflammation, supported by activation of TNF/NF-κB and JAK-STAT signaling pathways via targeting-related genes, while the co-exposure greatly alleviated the neuroinflammation. Collectively, our findings illuminate the underlying neurotoxicity molecular mechanisms of NP and/or CHR exposure on aquatic organisms.

Identification of hub genes in digestive system of mandarin fish (Siniperca chuatsi) fed with artificial diet by weighted gene co-expression network analysis.

Mandarin fish (Siniperca chuatsi) is a carnivorous freshwater fish and an economically important species. The digestive system (liver, stomach, intestine, pyloric caecum, esophagus, and gallbladder) is an important site for studying fish domestication. In our previous study, we found that mandarin fish undergoes adaptive changes in histological morphology and gene expression levels of the digestive system when subjected to artificial diet domestication. However, we are not clear which hub genes are highly associated with domestication. In this study, we performed WGCNA on the transcriptomes of 17 tissues and 9 developmental stages and combined differentially expressed genes analysis in the digestive system to identify the hub genes that may play important functions in the adaptation of mandarin fish to bait conversion. A total of 31,657 genes in 26 samples were classified into 23 color modules via WGCNA. The modules midnightblue, darkred, lightyellow, and darkgreen highly associated with the liver, stomach, esophagus, and gallbladder were extracted, respectively. Tan module was highly related to both intestine and pyloric caecum. The hub genes in liver were cp, vtgc, c1in, c9, lect2, and klkb1. The hub genes in stomach were ghrl, atp4a, gjb3, muc5ac, duox2, and chia2. The hub genes in esophagus were mybpc1, myl2, and tpm3. The hub genes in gallbladder were dyst, npy2r, slc13a1, and slc39a4. The hub genes in the intestine and pyloric caecum were slc15a1, cdhr5, btn3a1, anpep, slc34a2, cdhr2, and ace2. Through pathway analysis, modules highly related to the digestive system were mainly enriched in digestion and absorption, metabolism, and immune-related pathways. After domestication, the hub genes vtgc and lect2 were significantly upregulated in the liver. Chia2 was significantly downregulated in the stomach. Slc15a1, anpep, and slc34a2 were significantly upregulated in the intestine. This study identified the hub genes that may play an important role in the adaptation of the digestive system to artificial diet, which provided novel evidence and ideas for further research on the domestication of mandarin fish from molecular level.

Specific pupylation as IDEntity reporter (SPIDER) for the identification of protein-biomolecule interactions.

Protein-biomolecule interactions play pivotal roles in almost all biological processes. For a biomolecule of interest, the identification of the interacting protein(s) is essential. For this need, although many assays are available, highly robust and reliable methods are always desired. By combining a substrate-based proximity labeling activity from the pupylation pathway of Mycobacterium tuberculosis and the streptavidin (SA)-biotin system, we developed the Specific Pupylation as IDEntity Reporter (SPIDER) method for identifying protein-biomolecule interactions. Using SPIDER, we validated the interactions between the known binding proteins of protein, DNA, RNA, and small molecule. We successfully applied SPIDER to construct the global protein interactome for m6A and mRNA, identified a variety of uncharacterized m6A binding proteins, and validated SRSF7 as a potential m6A reader. We globally identified the binding proteins for lenalidomide and CobB. Moreover, we identified SARS-CoV-2-specific receptors on the cell membrane. Overall, SPIDER is powerful and highly accessible for the study of protein-biomolecule interactions.

Longitudinal neutralization activities on authentic Omicron variant provided by three doses of BBIBP-CorV vaccination during one year.

The majority of people in China have been immunized with the inactivated viral vaccine BBIBP-CorV. The emergence of the Omicron variant raised the concerns about protection efficacy of the inactivated viral vaccine in China. However, longitudinal neutralization data describing protection efficacy against Omicron variant is still lacking. Here we present one-year longitudinal neutralization data of BBIBP-CorV on authentic Omicron, Delta, and wild-type strains using 224 sera collected from 14 volunteers who have finished three doses BBIBP-CorV. The sera were also subjected for monitoring the SARS-CoV-2 specific IgG, IgA, and IgM responses on protein and peptide microarrays. The neutralization titers showed different protection efficacies against the three strains. By incorporating IgG and IgA signals of proteins and Spike protein derived peptide on microarray, panels as potential surrogate biomarkers for rapid estimation of neutralization titers were established. These data support the necessity of the 3rd dose of BBIBP-CorV vaccination. After further validation and assay development, the panels could be used for reliable, convenient and fast evaluation of the efficacy of vaccination.

Age- and Severity-Associated Humoral Immunity Response in COVID-19 Patients: A Cohort Study from Wuhan, China.

Age has been found to be the single most significant factor in COVID-19 severity and outcome. However, the age-related severity factors of COVID-19 have not been definitively established. In this study, we detected SARS-CoV-2-specific antibody responses and infectious disease-related blood indicators in 2360 sera from 783 COVID-19 patients, with an age range of 1−92 years. In addition, we recorded the individual information and clinical symptoms of the patients. We found that the IgG responses for S1, N, and ORF3a and the IgM for NSP7 were associated with severe COVID-19 at different ages. The IgM responses for the S-protein peptides S1-113 (aa 673−684) and S2-97 (aa 1262−1273) were associated with severe COVID-19 in patients aged <60. Furthermore, we found that the IgM for S1-113 and NSP7 may play a protective role in patients aged <60 and >80, respectively. Regarding clinical parameters, we analyzed the diagnostic ability of five clinical parameters for severe COVID-19 in six age groups and identified three-target panel, glucose, IL-6, myoglobin, IL-6, and NT proBNP as the appropriate diagnostic markers for severe COVID-19 in patients aged <41, 41−50, 51−60, 61−70, 71−80, and >80, respectively. The age-associated severity factors revealed here will facilitate our understanding of COVID-19 immunity and diagnosis, and eventually provide meaningful information for combating the pandemic.

Identification of sitagliptin binding proteins by affinity purification mass spectrometry.

Type 2 diabetes mellitus (T2DM) is recognized as a serious public health concern with increasing incidence. The dipeptidyl peptidase-4 (DPP-4) inhibitor sitagliptin has been used for the treatment of T2DM worldwide. Although sitagliptin has excellent therapeutic outcome, adverse effects are observed. In addition, previous studies have suggested that sitagliptin may have pleiotropic effects other than treating T2DM. These pieces of evidence point to the importance of further investigation of the molecular mechanisms of sitagliptin, starting from the identification of sitagliptin-binding proteins. In this study, by combining affinity purification mass spectrometry (AP-MS) and stable isotope labeling by amino acids in cell culture (SILAC), we discover seven high-confidence targets that can interact with sitagliptin. Surface plasmon resonance (SPR) assay confirms the binding of sitagliptin to three proteins, i. e., LYPLAL1, TCP1, and CCAR2, with binding affinities (K D) ranging from 50.1 µM to 1490 µM. Molecular docking followed by molecular dynamic (MD) simulation reveals hydrogen binding between sitagliptin and the catalytic triad of LYPLAL1, and also between sitagliptin and the P-loop of ATP-binding pocket of TCP1. Molecular mechanics Poisson-Boltzmann Surface Area (MMPBSA) analysis indicates that sitagliptin can stably bind to LYPLAL1 and TCP1 in active sites, which may have an impact on the functions of these proteins. SPR analysis validates the binding affinity of sitagliptin to TCP1 mutant D88A is ~10 times lower than that to the wild-type TCP1. Our findings provide insights into the sitagliptin-targets interplay and demonstrate the potential of sitagliptin in regulating gluconeogenesis and in anti-tumor drug development.

Ultrasensitive monitoring of SARS-CoV-2-specific antibody responses based on a digital approach reveals one week of IgG seroconversion.

COVID-19 is still unfolding, while many people have been vaccinated. In comparison to nucleic acid testing (NAT), antibody-based immunoassays are faster and more convenient. However, its application has been hampered by its lower sensitivity and the existing fact that by traditional immunoassays, the measurable seroconversion time of pathogen-specific antibodies, such as IgM or IgG, lags far behind that of nucleic acids. Herein, by combining the single molecule array platform (Simoa), RBD, and a previously identified SARS-CoV-2 S2 protein derivatized 12-aa peptide (S2-78), we developed and optimized an ultrasensitive assay (UIM-COVID-19 assay). Sera collected from three sources were tested, i.e., convalescents, inactivated virus vaccine-immunized donors and wild-type authentic SARS-CoV-2-infected rhesus monkeys. The sensitivities of UIM-COVID-19 assays are 100-10,000 times higher than those of conventional flow cytometry, which is a relatively sensitive detection method at present. For the established UIM-COVID-19 assay using RBD as a probe, the IgG and IgM seroconversion times after vaccination were 7.5 and 8.6 days vs. 21.4 and 24 days for the flow cytometry assay, respectively. In addition, using S2-78 as a probe, the UIM-COVID-19 assay could differentiate COVID-19 patients (convalescents) from healthy people and patients with other diseases, with AUCs ranging from 0.85-0.95. In summary, the UIM-COVID-19 we developed here is a promising ultrasensitive biodetection strategy that has the potential to be applied for both immunological studies and diagnostics.

Complete mitochondrial genome of the common Cuora (Cuora mccordi), China.

We report the complete mitochondrial genome of Cuora mccordi. The complete genome is a closed circular molecule of 16,551 bp, with an overall base composition of 34.06% for A, 26.73% for T, 12.84% for G, and 26.37% for C. The A + T content is 60.79%. The full length consists of 13 protein-coding genes, 22 tRNA genes, two rRNA genes, and one control region (D-loop). Phylogenetic analysis results showed that the mitogenome of Cuora mccordi was the closest to Cuora pani. The complete mitochondrial genome of Cuora mccordi (GenBank accession number: OM327796) can aid in understanding evolutionary relationships within Cuora.

SARS-CoV-2-specific antibody response characteristics in COVID-19 patients of different ages.

Age has been found to be one of the main risk factors for the severity and outcome of COVID-19. However, differences in SARS-CoV-2 specific antibody responses among COVID-19 patients of different age groups remain largely unknown. In this study, we analyzed the IgG/IgM responses to 21 SARS-CoV-2 proteins and 197 peptides that fully cover the spike protein against 731 sera collected from 731 COVID-19 patients aged from 1 to We show that there is no overall difference in SARS-CoV-2 antibody responses in COVID-19 patients in the 4 age groups. By antibody response landscape maps, we find that the IgG response profiles of SARS-CoV-2 proteins are positively correlated with age. The S protein linear epitope map shows that the immunogenicity of the S-protein peptides is related to peptide sequence, disease severity and age of the COVID-19 patients. Furthermore, the enrichment analysis indicates that low S1 IgG responses are enriched in patients aged <50 and high S1 IgG responses are enriched in mild COVID-19 patients aged >60. In addition, high responses of non-structural/accessory proteins are enriched in severe COVID-19 patients aged >70. These results suggest the distinct immune response of IgG/IgM to each SARS-CoV-2 protein in patients of different age, which may facilitate a deeper understanding of the immune responses in COVID-19 patients.

Evaluation of Spike Protein Epitopes by Assessing the Dynamics of Humoral Immune Responses in Moderate COVID-19.

The coronavirus disease 2019 (COVID-19) pandemic is caused by a novel coronavirus called severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). The spike protein (S) of SARS-CoV-2 is a major target for diagnosis and vaccine development because of its essential role in viral infection and host immunity. Currently, time-dependent responses of humoral immune system against various S protein epitopes are poorly understood. In this study, enzyme-linked immunosorbent assay (ELISA), peptide microarray, and antibody binding epitope mapping (AbMap) techniques were used to systematically analyze the dynamic changes of humoral immune responses against the S protein in a small cohort of moderate COVID-19 patients who were hospitalized for approximately two months after symptom onset. Recombinant truncated S proteins, target S peptides, and random peptides were used as antigens in the analyses. The assays demonstrated the dynamic IgM- and IgG recognition and reactivity against various S protein epitopes with patient-dependent patterns. Comprehensive analysis of epitope distribution along the spike gene sequence and spatial structure of the homotrimer S protein demonstrated that most IgM- and IgG-reactive peptides were clustered into similar genomic regions and were located at accessible domains. Seven S peptides were generally recognized by IgG antibodies derived from serum samples of all COVID-19 patients. The dynamic immune recognition signals from these seven S peptides were comparable to those of the entire S protein or truncated S1 protein. This suggested that the humoral immune system recognized few conserved S protein epitopes in most COVID-19 patients during the entire duration of humoral immune response after symptom onset. Furthermore, in this cohort, individual patients demonstrated stable immune recognition to certain S protein epitopes throughout their hospitalization period. Therefore, the dynamic characteristics of humoral immune responses to S protein have provided valuable information for accurate diagnosis and immunotherapy of COVID-19 patients.

SARS-CoV-2 proteome microarray for COVID-19 patient sera profiling.

The immunogenicity of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) proteome is largely unknown. Here we describe a protocol for analyzing sera samples with SARS-CoV-2 proteome microarray. The proteins were expressed by either E. coli expression system or eukaryotic cell expression systems and obtained by affinity purification. The protocol includes microarray fabricating and sera profiling, which will be used to build an antibody response landscape for IgG and IgM. The protocol may help to facilitate a deeper understanding of immunity related to SARS-CoV-2. For complete details on the use and execution of this protocol, please refer to Li et al. (2021c).

Anti-SARS-CoV-2 IgG responses are powerful predicting signatures for the outcome of COVID-19 patients.

Introduction: The COVID-19 global pandemic is far from ending. There is an urgent need to identify applicable biomarkers for early predicting the outcome of COVID-19. Growing evidences have revealed that SARS-CoV-2 specific antibodies evolved with disease progression and severity in COIVD-19 patients. Objectives: We assumed that antibodies may serve as biomarkers for predicting the clinical outcome of hospitalized COVID-19 patients on admission. Methods: By taking advantage of a newly developed SARS-CoV-2 proteome microarray, we surveyed IgG responses against 20 proteins of SARS-CoV-2 in 1034 hospitalized COVID-19 patients on admission and followed till 66 days. The microarray results were further correlated with clinical information, laboratory test results and patient outcomes. Cox proportional hazards model was used to explore the association between SARS-CoV-2 specific antibodies and COVID-19 mortality. Results: Nonsurvivors (n = 955) induced higher levels of IgG responses against most of non-structural proteins than survivors (n = 79) on admission. In particular, the magnitude of IgG antibodies against 8 non-structural proteins (NSP1, NSP4, NSP7, NSP8, NSP9, NSP10, RdRp, and NSP14) and 2 accessory proteins (ORF3b and ORF9b) possessed significant predictive power for patient death, even after further adjustments for demographics, comorbidities, and common laboratory biomarkers for disease severity (all with p trend < 0.05). Additionally, IgG responses to all of these 10 non-structural/accessory proteins were also associated with the severity of disease, and differential kinetics and serum positive rate of these IgG responses were confirmed in COVID-19 patients of varying severities within 20 days after symptoms onset. The area under curves (AUCs) for these IgG responses, determined by computational cross-validations, were between 0.62 and 0.71. Conclusions: Our findings might have important implications for improving clinical management of COVID-19 patients.