Mpox vaccination for men who have sex with men and their differential risk of exposure and infection.

Following the post-COVID-19 reopening of the society with enhanced traveling between countries, people at risk of mpox infection, notably men who have sex with men (MSM) and people living with HIV, are facing increasing threat of virus exposure. Mpox vaccination is an important public health strategy which is provided free in Hong Kong to people at higher risk of infection. Between October 2022 and January 2023, 326 and 184 MSM vaccinees from vaccination sites and HIV specialist clinics in Hong Kong, respectively, were recruited for assessing their infection risks. Apart from the urge to protect one's significant others (68%), 45% were worried about the stigmatizing mpox symptoms if infected. Compared with MSM vaccinees at vaccination site, a lower proportion of MSM vaccinees in HIV care were sexually active in the past 6 months (88% vs 97%), but a higher proportion had recent sexually transmitted infection diagnoses (19% vs 10%) and perceived considerable exposure risk in the following 6 months (40% vs 22%). There were no differences in the perceived effectiveness of mpox vaccination. If optimal supplies of mpox vaccines can be secured, a low threshold approach at vaccination site could enable MSM with different levels of behavioral risks to become protected.

The differential impacts of early detection and accelerated antiretroviral therapy on the epidemiologic trend of sexually acquired HIV infection in Hong Kong.

OBJECTIVES: To assess impacts of early detection and prompt antiretroviral therapy (ART) on the latest epidemiologic situation to inform intervention strategy. METHODS: We analysed data from two clinical cohorts in Hong Kong where sexual transmission accounted for the majority of HIV infections. The two cohorts comprised patients newly diagnosed in 2007-2008 and 2016-2018 respectively. Secular trend and differences between men who have sex with men (MSM) and heterosexual patients were examined. Predictors of late presentation (defined as CD4 ≤350 or AIDS-defining illness within 3 months of diagnosis) and prolonged interval between diagnosis and ART initiation were assessed by multivariable regressions. RESULTS: There were 1,136 newly diagnosed HIV patients with 644 in the first and 492 in the second cohort, a majority (91.7%) presented with sexually acquired infection. There were less MSM in the first than the second cohort (50.3%% vs 87.8%, χ2 = 117.05, p<0.001). The mean (SD) number of days between diagnosis and ART initiation decreased from 514.3 (516.1) to 61.8 (94.2) days across the two cohorts. Younger age, non-Chinese, outpatient-based service and lower CD4 count were predictors of faster ART initiation in the first but not in the second cohort. Interval between diagnosis and ART initiation became highly uniform among groups in the second cohort. Nearly 60% were classified as late presenters in both cohorts. Heterosexuals (aOR 1.58, 95% CI 1.13-2.19) had a higher risk of late presentation. CONCLUSIONS: There was remarkable improvement in acceleration of ART initiation. Clinical implementation of accelerated ART recommendations has been effective for both MSM and heterosexuals. Late presentation was more marked among heterosexuals and remained a problem. The continued phenomenon of late presentation could offset the epidemiologic gains from accelerated ART initiation.

Mitochondrial regulation of acute extrafollicular B-cell responses to COVID-19 severity.

BACKGROUND: Patients with COVID-19 display a broad spectrum of manifestations from asymptomatic to life-threatening disease with dysregulated immune responses. Mechanisms underlying the detrimental immune responses and disease severity remain elusive. METHODS: We investigated a total of 137 APs infected with SARS-CoV-2. Patients were divided into mild and severe patient groups based on their requirement of oxygen supplementation. All blood samples from APs were collected within three weeks after symptom onset. Freshly isolated PBMCs were investigated for B cell subsets, their homing potential, activation state, mitochondrial functionality and proliferative response. Plasma samples were tested for cytokine concentration, and titer of Nabs, RBD-, S1-, SSA/Ro- and dsDNA-specific IgG. RESULTS: While critically ill patients displayed predominantly extrafollicular B cell activation with elevated inflammation, mild patients counteracted the disease through the timely induction of mitochondrial dysfunction in B cells within the first week post symptom onset. Rapidly increased mitochondrial dysfunction, which was caused by infection-induced excessive intracellular calcium accumulation, suppressed excessive extrafollicular responses, leading to increased neutralizing potency index and decreased inflammatory cytokine production. Patients who received prior COVID-19 vaccines before infection displayed significantly decreased extrafollicular B cell responses and mild disease. CONCLUSION: Our results reveal an immune mechanism that controls SARS-CoV-2-induced detrimental B cell responses and COVID-19 severity, which may have implications for viral pathogenesis, therapeutic interventions and vaccine development.

Boosting of serum neutralizing activity against the Omicron variant among recovered COVID-19 patients by BNT162b2 and CoronaVac vaccines.

BACKGROUND: SARS-CoV-2 Omicron variant evades immunity from past infection or vaccination and is associated with a greater risk of reinfection among recovered COVID-19 patients. We assessed the serum neutralizing antibody (NAb) activity against Omicron variant (Omicron NAb) among recovered COVID-19 patients with or without vaccination. METHODS: In this prospective cohort study with 135 recovered COVID-19 patients, we determined the serum NAb titers against ancestral virus or variants using a live virus NAb assay. We used the receiver operating characteristic analysis to determine the optimal cutoff for a commercially-available surrogate NAb assay. FINDINGS: Among recovered COVID-19 patients, the serum live virus geometric mean Omicron NAb titer was statistically significantly higher among BNT162b2 recipients compared to non-vaccinated individuals (85.4 vs 5.6,P < 0.0001). The Omicron seropositive rates in live virus NAb test (NAb titer ≥10) were statistically significantly higher among BNT162b2 (90.6% [29/32];P < 0.0001) or CoronaVac (36.7% [11/30]; P = 0.0115) recipients when compared with non-vaccinated individuals (12.3% [9/73]). Subgroup analysis of CoronaVac recipients showed that the Omicron seropositive rates were higher among individuals with two doses than those with one dose (85.7% vs 21.7%; P = 0.0045). For the surrogate NAb assay, a cutoff of 109.1 AU/ml, which is 7.3-fold higher than the manufacturer's recommended cutoff, could achieve a sensitivity and specificity of 89.5% and 89.8%, respectively, in detecting Omicron NAb. INTERPRETATION: Among individuals with prior COVID-19, one dose of BNT162b2 or two doses of CoronaVac could induce detectable serum Omicron NAb. Our result would be particularly important for guiding vaccine policies in countries with COVID-19 vaccine shortage. FUNDING: Health and Medical Research Fund, Richard and Carol Yu, Michael Tong (see acknowledgments for full list).

Case series of HIV SARS-CoV-2 co-infection in Chinese adults.

Objectives: Little is known whether differences exist in virus shedding, immune and inflammatory response related to SARS-CoV-2 in people living with human immunodeficiency virus (PLWH). We assessed viral RNA and cytokine profiles of HIV and SARS-CoV-2 coinfection in Hong Kong. Methods: PLWH hospitalized with SARS-CoV-2 infection in Hong Kong were included, compared with age-matched and disease severity-matched SARS-CoV-2 infected controls (ratio of 1:5) from February 1st 2020 to July 31st 2020. SARS-CoV-2 infection was confirmed by public health laboratory and virus concentration was quantified by an in-house real-time reverse transcription-quantitative polymerase chain reaction. A panel of cytokines and chemokines were performed. Results: HIV patients had a similar respiratory shedding profile compared to controls. Duration of faecal shedding of patient A, B, C and D were at least 9, 10, 33, and 11 days, respectively. HIV patients had lower plasma levels of IL-10 and NT-pro-BNP. All 4 PLWH cases showed seroconversion to SARS-CoV-2 with anti-SARS-CoV-2 S antibodies detected in serum collected between day 18 and 30 after symptom onset. Conclusions: PLWH behaves similarly with HIV-negative controls in respiratory viral load, but with decrease in IL-10 and NT-proBNP. PLWH may have a lower risk of immunostimulatory effect due to lower IL-10.

Probable Animal-to-Human Transmission of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) Delta Variant AY.127 Causing a Pet Shop-Related Coronavirus Disease 2019 (COVID-19) Outbreak in Hong Kong.

BACKGROUND: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) can infect human and other mammals, including hamsters. Syrian (Mesocricetus auratus) and dwarf (Phodopus sp.) hamsters are susceptible to SARS-CoV-2 infection in the laboratory setting. However, pet shop-related Coronavirus Disease 2019 (COVID-19) outbreaks have not been reported. METHODS: We conducted an investigation of a pet shop-related COVID-19 outbreak due to Delta variant AY.127 involving at least 3 patients in Hong Kong. We tested samples collected from the patients, environment, and hamsters linked to this outbreak and performed whole genome sequencing analysis of the reverse transcription polymerase chain reaction (RT-PCR)-positive samples. RESULTS: The patients included a pet shop keeper (Patient 1), a female customer of the pet shop (Patient 2), and the husband of Patient 2 (Patient 3). Investigation showed that 17.2% (5/29) and 25.5% (13/51) environmental specimens collected from the pet shop and its related warehouse, respectively, tested positive for SARS-CoV-2 RNA by RT-PCR. Among euthanized hamsters randomly collected from the storehouse, 3% (3/100) tested positive for SARS-CoV-2 RNA by RT-PCR and seropositive for anti-SARS-CoV-2 antibody by enzyme immunoassay. Whole genome analysis showed that although all genomes from the outbreak belonged to the Delta variant AY.127, there were at least 3 nucleotide differences among the genomes from different patients and the hamster cages. Genomic analysis suggests that multiple strains have emerged within the hamster population, and these different strains have likely transmitted to human either via direct contact or via the environment. CONCLUSIONS: Our study demonstrated probable hamster-to-human transmission of SARS-CoV-2. As pet trading is common around the world, this can represent a route of international spread of this pandemic virus.

Hepatitis E Virus Species C Infection in Humans, Hong Kong.

BACKGROUND: Hepatitis E virus (HEV) variants belonging to Orthohepevirus species A (HEV-A) are the primary cause of human hepatitis E. However, we previously reported that Orthohepevirus species C genotype 1 (HEV-C1), a divergent HEV variant commonly found in rats, also causes hepatitis in humans. Here, we present a clinical-epidemiological investigation of human HEV-C1 infections detected in Hong Kong, with an emphasis on outcomes in immunocompromised individuals. METHODS: A surveillance system for detecting human HEV-C1 infections was established in Hong Kong. Epidemiological and clinical characteristics of HEV-C1 cases identified via this system between 1 August 2019 and 31 December 2020 were retrieved. Phylogenetic analysis of HEV-C1 strain sequences was performed. Infection outcomes of immunocompromised individuals with HEV-A and HEV-C1 infections were analyzed. RESULTS: HEV-C1 accounted for 8 of 53 (15.1%) reverse-transcription polymerase chain reaction (RT-PCR)-confirmed HEV infections in Hong Kong during the study period, raising the total number of HEV-C1 infections detected in the city to 16. Two distinct HEV-C1 strain groups caused human infections. Patients were elderly and/or immunocompromised; half tested negative for HEV immunoglobulin M. Cumulatively, HEV-C1 accounted for 9 of 21 (42.9%) cases of hepatitis E recorded in immunocompromised patients in Hong Kong. Immunocompromised HEV-C1 patients progressed to persistent hepatitis at similar rates (7/9 [77.8%]) as HEV-A patients (10/12 [75%]). HEV-C1 patients responded to oral ribavirin, although response to first course was sometimes poor or delayed. CONCLUSIONS: Dedicated RT-PCR-based surveillance detected human HEV-C1 cases that evade conventional hepatitis E diagnostic testing. Immunosuppressed HEV-C1-infected patients frequently progress to persistent HEV-C1 infection, for which ribavirin is a suitable treatment option.

Neutralization of Severe Acute Respiratory Syndrome Coronavirus 2 Omicron Variant by Sera From BNT162b2 or CoronaVac Vaccine Recipients.

BACKGROUND: The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) omicron variant, designated as a variant of concern by the World Health Organization, carries numerous spike mutations that are known to evade neutralizing antibodies elicited by coronavirus disease 2019 (COVID-19) vaccines. A deeper understanding of the susceptibility of omicron variant to vaccine-induced neutralizing antibodies is urgently needed for risk assessment. METHODS: Omicron variant strains HKU691 and HKU344-R346K were isolated from patients using TMPRSS2-overexpressing VeroE6 cells. Whole genome sequence was determined using nanopore sequencing. Neutralization susceptibility of ancestral lineage A virus and the omicron, delta and beta variants to sera from 25 BNT162b2 and 25 CoronaVac vaccine recipients was determined using a live virus microneutralization assay. RESULTS: The omicron variant strain HKU344-R346K has an additional spike R346K mutation, which is present in 8.5% of strains deposited in the GISAID database. Only 20% and 24% of BNT162b2 recipients had detectable neutralizing antibody against the omicron variant HKU691 and HKU344-R346K, respectively, whereas none of the CoronaVac recipients had detectable neutralizing antibody titer against either omicron isolate. For BNT162b2 recipients, the geometric mean neutralization antibody titers (GMTs) of the omicron variant isolates (5.43 and 6.42) were 35.7-39.9-fold lower than that of the ancestral virus (229.4), and the GMTs of both omicron variant isolates were significantly lower than those of the beta and delta variants. There was no significant difference in the GMTs between HKU691 and HKU344-R346K. CONCLUSIONS: Omicron variant escapes neutralizing antibodies elicited by BNT162b2 or CoronaVac. The additional R346K mutation did not affect the neutralization susceptibility. Our data suggest that the omicron variant may be associated with lower COVID-19 vaccine effectiveness.

The impact of spike N501Y mutation on neutralizing activity and RBD binding of SARS-CoV-2 convalescent serum.

BACKGROUND: Several SARS-CoV-2 lineages with spike receptor binding domain (RBD) N501Y mutation have spread globally. We evaluated the impact of N501Y on neutralizing activity of COVID-19 convalescent sera and on anti-RBD IgG assays. METHODS: The susceptibility to neutralization by COVID-19 patients' convalescent sera from Hong Kong were compared between two SARS-CoV-2 isolates (B117-1/B117-2) from the α variant with N501Y and 4 non-N501Y isolates. The effect of N501Y on antibody binding was assessed. The performance of commercially-available IgG assays was determined for patients infected with N501Y variants. FINDINGS: The microneutralization antibody (MN) titers of convalescent sera from 9 recovered COVID-19 patients against B117-1 (geometric mean titer[GMT],80; 95% CI, 47-136) were similar to those against the non-N501Y viruses. However, MN titer of these serum against B117-2 (GMT, 20; 95% CI, 11-36) was statistically significantly reduced when compared with non-N501Y viruses (P < 0.01; one-way ANOVA). The difference between B117-1 and B117-2 was confirmed by testing 60 additional convalescent sera. B117-1 and B117-2 differ by only 3 amino acids (nsp2-S512Y, nsp13-K460R, spike-A1056V). Enzyme immunoassay using 272 convalescent sera showed reduced binding of anti-RBD IgG to N501Y or N501Y-E484K-K417N when compared with that of wild-type RBD (mean difference: 0.1116 and 0.5613, respectively; one-way ANOVA). Of 7 anti-N-IgG positive sera from patients infected with N501Y variants (collected 9-14 days post symptom onset), 6 (85.7%) tested negative for a commercially-available anti-S1-IgG assay. FUNDING: Richard and Carol Yu, Michael Tong, and the Government Consultancy Service (see acknowledgments for full list). INTERPRETATION: We highlighted the importance of using a panel of viruses within the same lineage to determine the impact of virus variants on neutralization. Furthermore, clinicians should be aware of the potential reduced sensitivity of anti-RBD IgG assays.

Homologous and heterologous serological response to the N-terminal domain of SARS-CoV-2 in humans and mice.

The increasing numbers of infected cases of coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) poses serious threats to public health and the global economy. Most SARS-CoV-2 neutralizing antibodies target the receptor binding domain (RBD) and some the N-terminal domain (NTD) of the spike protein, which is the major antigen of SARS-CoV-2. While the antibody response to RBD has been extensively characterized, the antigenicity and immunogenicity of the NTD protein are less well studied. Using 227 plasma samples from COVID-19 patients, we showed that SARS-CoV-2 NTD-specific antibodies could be induced during infection. As compared to the results of SARS-CoV-2 RBD, the serological response of SARS-CoV-2 NTD is less cross-reactive with SARS-CoV, a pandemic strain that was identified in 2003. Furthermore, neutralizing antibodies are rarely elicited in a mice model when NTD is used as an immunogen. We subsequently demonstrate that NTD has an altered antigenicity when expressed alone. Overall, our results suggest that while NTD offers a supplementary strategy for serology testing, it may not be suitable as an immunogen for vaccine development.

Dynamics of B cell repertoires and emergence of cross-reactive responses in patients with different severities of COVID-19.

Individuals with the 2019 coronavirus disease (COVID-19) show varying severity of the disease, ranging from asymptomatic to requiring intensive care. Although monoclonal antibodies specific to the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) have been identified, we still lack an understanding of the overall landscape of B cell receptor (BCR) repertoires in individuals with COVID-19. We use high-throughput sequencing of bulk and plasma B cells collected at multiple time points during infection to characterize signatures of the B cell response to SARS-CoV-2 in 19 individuals. Using principled statistical approaches, we associate differential features of BCRs with different disease severity. We identify 38 significantly expanded clonal lineages shared among individuals as candidates for responses specific to SARS-CoV-2. Using single-cell sequencing, we verify the reactivity of BCRs shared among individuals to SARS-CoV-2 epitopes. Moreover, we identify the natural emergence of a BCR with cross-reactivity to SARS-CoV-1 and SARS-CoV-2 in some individuals. Our results provide insights important for development of rational therapies and vaccines against COVID-19.

Mass Screening Is Associated with Low Rates of Acute Kidney Injury among COVID-19 Patients in Hong Kong.

INTRODUCTION: Renal involvement in COVID-19 is less well characterized in settings with vigilant public health surveillance, including mass screening and early hospitalization. We assessed kidney complications among COVID-19 patients in Hong Kong, including the association with risk factors, length of hospitalization, critical presentation, and mortality. METHODS: Linked electronic records of all patients with confirmed COVID-19 from 5 major designated hospitals were extracted. Duplicated records due to interhospital transferal were removed. Primary outcome was the incidence of in-hospital acute kidney injury (AKI). Secondary outcomes were AKI-associated mortality, incident renal replacement therapy (RRT), intensive care admission, prolonged hospitalization and disease course (defined as >90th percentile of hospitalization duration [35 days] and duration from symptom onset to discharge [43 days], respectively), and change of estimated glomerular filtration rate (GFR). Patients were further stratified into being symptomatic or asymptomatic. RESULTS: Patients were characterized by young age (median: 38.4, IQR: 28.4-55.8 years) and short time (median: 5, IQR: 2-9 days) from symptom onset to admission. Among the 591 patients, 22 (3.72%) developed AKI and 4 (0.68%) required RRT. The median time from symptom onset to in-hospital AKI was 15 days. AKI increased the odds of prolonged hospitalization and disease course by 2.0- and 3.5-folds, respectively. Estimated GFR 24 weeks post-discharge reduced by 7.51 and 1.06 mL/min/1.73 m2 versus baseline (upon admission) in the AKI and non-AKI groups, respectively. The incidence of AKI was comparable between asymptomatic (4.8%, n = 3/62) and symptomatic (3.7%, n = 19/519) patients. CONCLUSION: The overall rate of AKI among COVID-19 patients in Hong Kong is low, which could be attributable to a vigilant screening program and early hospitalization. Among patients who developed in-hospital AKI, the duration of hospitalization is prolonged and kidney function impairment can persist for up to 6 months post-discharge. Mass surveillance for COVID-19 is warranted in identifying asymptomatic subjects for earlier AKI management.

Mitochondrial Dysfunction Associates With Acute T Lymphocytopenia and Impaired Functionality in COVID-19 Patients.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection results in rapid T lymphocytopenia and functional impairment of T cells. The underlying mechanism, however, remains incompletely understood. In this study, we focused on characterizing the phenotype and kinetics of T-cell subsets with mitochondrial dysfunction (MD) by multicolor flow cytometry and investigating the association between MD and T-cell functionality. While 73.9% of study subjects displayed clinical lymphocytopenia upon hospital admission, a significant reduction of CD4 or CD8 T-cell frequency was found in all asymptomatic, symptomatic, and convalescent cases. CD4 and CD8 T cells with increased MD were found in both asymptomatic and symptomatic patients within the first week of symptom onset. Lower proportion of memory CD8 T cell with MD was found in severe patients than in mild ones at the stage of disease progression. Critically, the frequency of T cells with MD in symptomatic patients was preferentially associated with CD4 T-cell loss and CD8 T-cell hyperactivation, respectively. Patients bearing effector memory CD4 and CD8 T cells with the phenotype of high MD exhibited poorer T-cell responses upon either phorbol 12-myristate-13-acetate (PMA)/ionomycin or SARS-CoV-2 peptide stimulation than those with low MD. Our findings demonstrated an MD-associated mechanism underlying SARS-CoV-2-induced T lymphocytopenia and functional impairment during the acute phase of infection.

Transmission of Rat Hepatitis E Virus Infection to Humans in Hong Kong: A Clinical and Epidemiological Analysis.

BACKGROUND AND AIMS: Hepatitis E virus (HEV) variants causing human infection predominantly belong to HEV species A (HEV-A). HEV species C genotype 1 (HEV-C1) circulates in rats and is highly divergent from HEV-A. It was previously considered unable to infect humans, but the first case of human HEV-C1 infection was recently discovered in Hong Kong. The aim of this study is to further describe the features of this zoonosis in Hong Kong. APPROACH AND RESULTS: We conducted a territory-wide prospective screening study for HEV-C1 infection over a 31-month period. Blood samples from 2,860 patients with abnormal liver function (n = 2,201) or immunosuppressive conditions (n = 659) were screened for HEV-C1 RNA. In addition, 186 captured commensal rats were screened for HEV-C1 RNA. Sequences of human-derived and rat-derived HEV-C1 isolates were compared. Epidemiological and clinical features of HEV-C1 infection were analyzed. HEV-C1 RNA was detected in 6/2,201 (0.27%) patients with hepatitis and 1/659 (0.15%) immunocompromised persons. Including the previously reported case, eight HEV-C1 infections were identified, including five in patients who were immunosuppressed. Three patients had acute hepatitis, four had persistent hepatitis, and one had subclinical infection without hepatitis. One patient died of meningoencephalitis, and HEV-C1 was detected in cerebrospinal fluid. HEV-C1 hepatitis was generally milder than HEV-A hepatitis. HEV-C1 RNA was detected in 7/186 (3.76%) rats. One HEV-C1 isolate obtained from a rat captured near the residences of patients was closely related to the major outbreak strain. CONCLUSIONS: HEV-C1 is a cause of hepatitis E in humans in Hong Kong. Immunosuppressed individuals are susceptible to persistent HEV-C1 infection and extrahepatic manifestations. Subclinical HEV-C1 infection threatens blood safety. Tests for HEV-C1 are required in clinical laboratories.

Unique Clusters of Severe Acute Respiratory Syndrome Coronavirus 2 Causing a Large Coronavirus Disease 2019 Outbreak in Hong Kong.

After 2 months of relative quiescence, a large coronavirus disease 2019 outbreak occurred in Hong Kong in July 2020 after gradual relaxation of social distancing policy. Unique severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) phylogenetic clusters have been identified among locally acquired cases, with most genomes belonging to cluster HK1, which is phylogenetically related to SARS-CoV-2 reported overseas.

Dynamics of B-cell repertoires and emergence of cross-reactive responses in COVID-19 patients with different disease severity.

COVID-19 patients show varying severity of the disease ranging from asymptomatic to requiring intensive care. Although a number of SARS-CoV-2 specific monoclonal antibodies have been identified, we still lack an understanding of the overall landscape of B-cell receptor (BCR) repertoires in COVID-19 patients. Here, we used high-throughput sequencing of bulk and plasma B-cells collected over multiple time points during infection to characterize signatures of B-cell response to SARS-CoV-2 in 19 patients. Using principled statistical approaches, we determined differential features of BCRs associated with different disease severity. We identified 38 significantly expanded clonal lineages shared among patients as candidates for specific responses to SARS-CoV-2. Using single-cell sequencing, we verified reactivity of BCRs shared among individuals to SARS-CoV-2 epitopes. Moreover, we identified natural emergence of a BCR with cross-reactivity to SARS-CoV-1 and SARS-CoV-2 in a number of patients. Our results provide important insights for development of rational therapies and vaccines against COVID-19.

Dynamics of B-cell repertoires and emergence of cross-reactive responses in COVID-19 patients with different disease severity.

COVID-19 patients show varying severity of the disease ranging from asymptomatic to requiring intensive care. Although a number of SARS-CoV-2 specific monoclonal antibodies have been identified, we still lack an understanding of the overall landscape of B-cell receptor (BCR) repertoires in COVID-19 patients. Here, we used high-throughput sequencing of bulk and plasma B-cells collected over multiple time points during infection to characterize signatures of B-cell response to SARS-CoV-2 in 19 patients. Using principled statistical approaches, we determined differential features of BCRs associated with different disease severity. We identified 38 significantly expanded clonal lineages shared among patients as candidates for specific responses to SARS-CoV-2. Using single-cell sequencing, we verified reactivity of BCRs shared among individuals to SARS-CoV-2 epitopes. Moreover, we identified natural emergence of a BCR with cross-reactivity to SARS-CoV-1 and SARS-CoV-2 in a number of patients. Our results provide important insights for development of rational therapies and vaccines against COVID-19.

Intra-host non-synonymous diversity at a neutralizing antibody epitope of SARS-CoV-2 spike protein N-terminal domain.

OBJECTIVES: SARS-CoV-2 has evolved rapidly into several genetic clusters. However, data on mutations during the course of infection are scarce. This study aims to determine viral genome diversity in serial samples of COVID-19 patients. METHODS: Targeted deep sequencing of the spike gene was performed on serial respiratory specimens from COVID-19 patients using nanopore and Illumina sequencing. Sanger sequencing was then performed to confirm the single nucleotide polymorphisms. RESULTS: A total of 28 serial respiratory specimens from 12 patients were successfully sequenced using nanopore and Illumina sequencing. A 75-year-old patient with severe disease had a mutation, G22017T, identified in the second specimen. The frequency of G22017T increased from ≤5% (nanopore: 3.8%; Illumina: 5%) from the first respiratory tract specimen (sputum) to ≥60% (nanopore: 67.7%; Illumina: 60.4%) in the second specimen (saliva; collected 2 days after the first specimen). The difference in G22017T frequency was also confirmed by Sanger sequencing. G22017T corresponds to W152L amino acid mutation in the spike protein which was only found in <0.03% of the sequences deposited into a public database. Spike amino acid residue 152 is located within the N-terminal domain, which mediates the binding of a neutralizing antibody. DISCUSSION: A spike protein amino acid mutation W152L located within a neutralizing epitope has appeared naturally in a patient. Our study demonstrated that monitoring of serial specimens is important in identifying hotspots of mutations, especially those occurring at neutralizing epitopes which may affect the therapeutic efficacy of monoclonal antibodies.

Acute SARS-CoV-2 Infection Impairs Dendritic Cell and T Cell Responses.

The SARS-CoV-2 pandemic has resulted in millions of infections, yet the role of host immune responses in early COVID-19 pathogenesis remains unclear. By investigating 17 acute and 24 convalescent patients, we found that acute SARS-CoV-2 infection resulted in broad immune cell reduction including T, natural killer, monocyte, and dendritic cells (DCs). DCs were significantly reduced with functional impairment, and ratios of conventional DCs to plasmacytoid DCs were increased among acute severe patients. Besides lymphocytopenia, although neutralizing antibodies were rapidly and abundantly generated in patients, there were delayed receptor binding domain (RBD)- and nucleocapsid protein (NP)-specific T cell responses during the first 3 weeks after symptoms onset. Moreover, acute RBD- and NP-specific T cell responses included relatively more CD4 T cells than CD8 T cells. Our findings provided evidence that impaired DCs, together with timely inverted strong antibody but weak CD8 T cell responses, could contribute to acute COVID-19 pathogenesis and have implications for vaccine development.