Lack of association between HLA and asymptomatic SARS-CoV-2 infection (preprint)

Human genetic studies of critical COVID-19 pneumonia have revealed the essential role of type I interferon-dependent innate immunity to SARS-CoV-2 infection. Conversely, an association between the HLA-B*15:01 allele and asymptomatic SARS-CoV-2 infection in unvaccinated individuals was recently reported, suggesting a contribution of pre-existing T cell-dependent adaptive immunity. We report a lack of association of classical HLA alleles, including HLA-B*15:01, with pre-omicron asymptomatic SARS-CoV-2 infection in unvaccinated participants in a prospective population-based study in the US (191 asymptomatic vs. 945 symptomatic COVID-19 cases). Moreover, we found no such association in the international COVID Human Genetic Effort cohort (206 asymptomatic vs. 574 mild or moderate COVID-19 cases and 1,625 severe or critical COVID-19 cases). Finally, in the Human Challenge Characterisation study, the three HLA-B*15:01 individuals infected with SARS-CoV-2 developed symptoms. As with other acute primary infections, no classical HLA alleles favoring an asymptomatic course of SARS-CoV-2 infection were identified. These findings suggest that memory T-cell immunity to seasonal coronaviruses does not strongly influence the outcome of SARS-CoV-2 infection in unvaccinated individuals.

Exome-wide association study to identify rare variants influencing COVID-19 outcomes: Results from the Host Genetics Initiative (preprint)

Host genetics is a key determinant of COVID-19 outcomes. Previously, the COVID-19 Host Genetics Initiative genome-wide association study used common variants to identify multiple loci associated with COVID-19 outcomes. However, variants with the largest impact on COVID-19 outcomes are expected to be rare in the population. Hence, studying rare variants may provide additional insights into disease susceptibility and pathogenesis, thereby informing therapeutics development. Here, we combined whole-exome and whole-genome sequencing from 21 cohorts across 12 countries and performed rare variant exome-wide burden analyses for COVID-19 outcomes. In an analysis of 5,048 severe disease cases and 571,009 controls, we observed that carrying a rare deleterious variant in the SARS-CoV-2 sensor toll-like receptor TLR7 (on chromosome X) was associated with a 5.3-fold increase in severe disease (95% CI: 2.75-10.05, p=5.41x10-7). These results further support TLR7 as a genetic determinant of severe disease and suggest that larger studies on rare variants influencing COVID-19 outcomes could provide additional insights.

Wastewater sequencing uncovers early, cryptic SARS-CoV-2 variant transmission (preprint)

As SARS-CoV-2 becomes an endemic pathogen, detecting emerging variants early is critical for public health interventions. Inferring lineage prevalence by clinical testing is infeasible at scale, especially in areas with limited resources, participation, or testing/sequencing capacity, which can also introduce biases. SARS-CoV-2 RNA concentration in wastewater successfully tracks regional infection dynamics and provides less biased abundance estimates than clinical testing. Tracking virus genomic sequences in wastewater would improve community prevalence estimates and detect emerging variants. However, two factors limit wastewater-based genomic surveillance: low-quality sequence data and inability to estimate relative lineage abundance in mixed samples. Here, we resolve these critical issues to perform a high-resolution, 295-day wastewater and clinical sequencing effort, in the controlled environment of a large university campus and the broader context of the surrounding county. We develop and deploy improved virus concentration protocols and deconvolution software that fully resolve multiple virus strains from wastewater. We detect emerging variants of concern up to 14 days earlier in wastewater samples, and identify multiple instances of virus spread not captured by clinical genomic surveillance. Our study provides a scalable solution for wastewater genomic surveillance that allows early detection of SARS-CoV-2 variants and identification of cryptic transmission.

HLA-A*03:01 is associated with increased risk of fever, chills, and more severe reaction to Pfizer-BioNTech COVID-19 vaccination (preprint)

COVID-19 vaccines are safe and highly effective, but some individuals experience unpleasant reactions to vaccination. As the majority of adults in the US have received a COVID-19 vaccine this year, there is an unprecedented opportunity to study the genetics of reactions to vaccination via surveys of individuals who are already part of genetic research studies. Here, we have queried 17,440 participants in the Helix DNA Discovery Project and Healthy Nevada Project about their reactions to COVID-19 vaccination. Our GWAS identifies an association between severe vaccine side effects and HLA-A*03:01. This association was statistically significant only for those who received the Pfizer-BioNTech vaccine (BNT162b2; p=4.70E-11), but showed a trending association in those who received the Moderna vaccine (mRNA-1273; p=0.005) despite similar sample sizes for study. In Pfizer-BioNTech recipients, HLA-A*03:01 was associated with a two-fold increase in risk of severe vaccine side effects. The effect was consistent across ages, sexes, and whether the person had previously had a COVID-19 infection. The reactions experienced by HLA-A*03:01 carriers were driven by associations with chills, fever, fatigue, and in general feeling unwell.

Genomic epidemiology identifies emergence and rapid transmission of SARS-CoV-2 B.1.1.7 in the United States (preprint)

As of January of 2021, the highly transmissible B.1.1.7 variant of SARS-CoV-2, which was first identified in the United Kingdom (U.K.), has gained a strong foothold across the world. Because of the sudden and rapid rise of B.1.1.7, we investigated the prevalence and growth dynamics of this variant in the United States (U.S.), tracking it back to its early emergence and onward local transmission. We found that the RT-qPCR testing anomaly of S gene target failure (SGTF), first observed in the U.K., was a reliable proxy for B.1.1.7 detection. We sequenced 212 B.1.1.7 SARS-CoV-2 genomes collected from testing facilities in the U.S. from December 2020 to January 2021. We found that while the fraction of B.1.1.7 among SGTF samples varied by state, detection of the variant increased at a logistic rate similar to those observed elsewhere, with a doubling rate of a little over a week and an increased transmission rate of 35-45%. By performing time-aware Bayesian phylodynamic analyses, we revealed several independent introductions of B.1.1.7 into the U.S. as early as late November 2020, with onward community transmission enabling the variant to spread to at least 30 states as of January 2021. Our study shows that the U.S. is on a similar trajectory as other countries where B.1.1.7 rapidly became the dominant SARS-CoV-2 variant, requiring immediate and decisive action to minimize COVID-19 morbidity and mortality.

S gene dropout patterns in SARS-CoV-2 tests suggest spread of the H69del/V70del mutation in the US. (preprint)

Recently, multiple novel strains of SARS-CoV-2 have been found to share the same deletion of amino acids H69 and V70 in the virus S gene. This includes strain B.1.1.7 / SARS-CoV-2 VUI 202012/01, which has been found to be more infectious than other strains of SARS-CoV-2, and its increasing presence has resulted in new lockdowns in and travel restrictions leaving the UK. Here, we analyze 2 million RT-PCR SARS-CoV-2 tests performed at Helix to identify the rate of S gene dropout, which has been recently shown to occur in tests from individuals infected with strains of SARS-CoV-2 that carry the H69del/V70del mutation. We observe a rise in S gene dropout in the US starting in early October, with 0.25% of our daily SARS-CoV-2-positive tests exhibiting this pattern during the first week. The rate of positive samples with S gene dropout has grown slowly over time, with last week exhibiting the highest level yet, at 0.5%. Focusing on the 14 states for which we have sufficient sample size to assess the frequency of this rare event (n>1000 SARS-CoV-2-positive samples), we see a recent expansion in the Eastern part of the US, concentrated in MA, OH, and FL. However, we cannot say from these data whether the S gene dropout samples we observe here represent the B.1.1.7. strain. Only with an expansion of genomic surveillance sequencing in the US will we know for certain the prevalence of the B.1.1.7 strain in the US.

Long-term COVID-19 symptoms in a large unselected population (preprint)

It is increasingly recognized that SARS-CoV-2 can produce long-term complications after recovery from the acute effects of infection. Here, we report the analysis of 32 self-reported short and long-term symptoms in a general adult population cohort comprised of 233 COVID-19+ cases, 3,652 SARS-CoV-2-negative controls, and 17,474 non-tested individuals. The majority of our COVID-19+ cases are mild, with only 8 of the 233 COVID-19+ cases having been hospitalized. Our results show that 43.4% of COVID-19+ cases have symptoms lasting longer than 30 days, and 24.1% still have at least one symptom after 90 days. These numbers are higher for COVID-19+ cases who were initially more ill, 59.4% at 30 days and 40.6% at 90 days, but even for very mild and initially asymptomatic cases, 14.3% have complications persist for 30 days or longer. In contrast, only 8.6% of participants from the general untested population develop new symptoms lasting longer than 30 days due to any illness during the same study period. The long-term symptoms most enriched in those with COVID-19 are anosmia, ageusia, difficulty concentrating, dyspnea, memory loss, confusion, headache, heart palpitations, chest pain, pain with deep breaths, dizziness, and tachycardia. We additionally observe that individuals who had an initial symptom of dyspnea are significantly more likely to develop long-term symptoms. Importantly, our study finds that the overall level of illness is an important variable to account for when assessing the statistical significance of symptoms that are associated with COVID-19. Our study provides a baseline from which to understand the frequency of COVID-19 long-term symptoms at the population level and demonstrates that, although those most likely to develop long-term COVID-19 complications are those who initially have more severe illness, even those with mild or asymptomatic courses of infection are at increased risk of long-term complications.

Using symptom-based case predictions to identify host genetic factors that contribute to COVID-19 susceptibility (preprint)

Epidemiological and genetic studies on COVID-19 are hindered by inconsistent and limited testing policies to confirm SARS-CoV-2 infection. Recently, it was shown that it is possible to predict potential COVID-19 cases using cross-sectional self-reported disease-related symptoms. Using a previously reported COVID-19 prediction model, we show that it is possible to conduct a GWAS on predicted COVID-19 which benefits from a larger sample size in order to gain new insights into the genetic susceptibility of the disease. Furthermore, we find suggestive evidence that genetic variants for other viral infectious diseases do not overlap with COVID-19 susceptibility and that severity of COVID-19 may have a different genetic architecture compared to COVID-19 susceptibility. Our findings demonstrate the added value of using self-reported symptom assessments to quickly monitor novel endemic viral outbreaks in a scenario of limited testing. Should there be another outbreak of a novel infectious disease, then we recommend repeatedly collecting data of disease-related symptoms.

Impaired cytotoxic CD8+ T cell response in elderly COVID-19 patients (preprint)

SARS-CoV-2 infection induces a T cell response that most likely contributes to virus control in COVID-19 patients, but may also induce immunopathology. Until now, the cytotoxic T cell response has not been very well characterized in COVID-19 patients. Here, we analyzed the differentiation and cytotoxic profile of T cells in 30 cases of mild COVID-19 during acute infection. SARS-CoV-2 infection induced a cytotoxic response of CD8+ T cells, but not CD4+ T cells, characterized by the simultaneous production of granzyme A and B, as well as perforin within different effector CD8+ T cell subsets. PD-1 expressing CD8+ T cells also produced cytotoxic molecules during acute infection indicating that they were not functionally exhausted. However, in COVID-19 patients over the age of 80 years the cytotoxic T cell potential was diminished, especially in effector memory and terminally differentiated effector CD8+ cells, showing that elderly patients have impaired cellular immunity against SARS-CoV-2. Our data provides valuable information about T cell responses in COVID-19 patients that may also have important implications for vaccine development. ImportanceCytotoxic T cells are responsible for the elimination of infected cells and are key players for the control of viruses. CD8+ T cells with an effector phenotype express cytotoxic molecules and are able to perform target cell killing. COVID-19 patients with a mild disease course were analyzed for the differentiation status and cytotoxic profile of CD8+ T cells. SARS-CoV-2 infection induced a vigorous cytotoxic CD8+ T cell response. However, this cytotoxic profile of T cells was not detected in COVID-19 patients over the age of 80 years. Thus, the absence of a cytotoxic response in elderly patients might be a possible reason for the more frequent severity of COVID-19 in this age group in comparison to younger patients.

Antiviral activity of lambda-carrageenan against influenza viruses in mice and severe acute respiratory syndrome coronavirus 2 in vitro (preprint)

Influenza virus and coronavirus, belonging to enveloped RNA viruses, are major causes of human respiratory diseases. The aim of this study was to investigate the broad spectrum antiviral activity of a naturally existing sulfated polysaccharide, lambda-carrageenan ({lambda}-CGN), purified from marine red algae. Cell culture-based assays revealed that the macromolecule efficiently inhibited both influenza A and B viruses, as well as currently circulating severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), with EC50 values ranging from 0.3-1.4 g/ml. No toxicity to host cells was observed at concentrations up to 300 g/ml. Plaque titration and western blot analysis verified that {lambda}-CGN reduced expression of viral proteins in cell lysates and suppressed progeny virus production in culture supernatants in a dose-dependent manner. This polyanionic compound exerts antiviral activity by targeting viral attachment to cell surface receptors and preventing entry. Moreover, intranasal administration to mice during influenza A viral challenge not only alleviated infection-mediated reductions in body weight but also protected 60% of mice from virus-induced mortality. Thus, {lambda}-CGN could be a promising antiviral agent for preventing infection by several respiratory viruses.