SARS-CoV-2 と COVID-19 に関する備忘録 Vol.4――佐藤研・G2P-Japan『新型コロナ変異株EG.5とXBBブレイクスルー感染の関連解析』…etc.

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SARS-CoV-2 と COVID-19 に関するメモ・備忘録

佐藤佳・東大教授が率いるG2P-Japanチームから、変異株BA.2.86（ピロラ）に続いてEG.5（エリス）の解析に関する論文が出されました。
佐藤研からはこれまで数々の研究成果が論文で発表されていて世界の医療に貢献しているのですが、お膝元の日本政府（永田町＆霞ヶ関）が政策にまったく活かしていないのはホンマなんなんでしょうね・・・


◆Antiviral efficacy of the SARS-CoV-2 XBB breakthrough infection sera against omicron subvariants including EG.5【The Lancet　2023年9月11日】

The SARS-CoV-2 XBB lineage is a recombinant omicron lineage that emerged in the summer of 2022. As of July, 2023, some XBB sublineages bearing the F486P substitution in the spike protein (S; S:F486P), such as XBB.1.5 and XBB.1.16, have rapidly spread and become predominant in the world according to Nextstrain. Because S:F486P significantly increased pseudovirus infectivity, it is assumed that the spread of F486P-bearing XBB subvariants is attributed by the increased infectivity by S:F486P.

As of July, 2023, EG.5.1 (also known as XBB.1.9.2.5.1), a XBB subvariant bearing the S:Q52H and S:F456L substitutions, alongside the S:F486P substitution, has rapidly spread in some Asian and North American countries. On Aug 9, 2023, WHO classified EG.5 as a variant of interest. In fact, our epidemic dynamics analyses showed that EG.5.1 exhibits a higher effective reproduction number (Re) compared with XBB.1.5, XBB.1.16, and its parental lineage (XBB.1.9.2), suggesting that EG.5.1 will spread globally and outcompete these XBB subvariants in the near future.

To assess the possibility that the enhanced infectivity of EG.5.1 contributes to its augmented Re, we prepared the lentivirus-based pseudoviruses with the S proteins of XBB.1.5/1.9.2 (note that the XBB.1.5 S is identical to the XBB.1.9.2 S), EG.5.1, and two XBB.1.5/1.9.2 derivatives, XBB.1.5/1.9.2+Q52H and XBB.1.5/1.9.2+F456L. The pseudovirus assay showed that both S:Q52H and S:F456L did not increase pseudovirus infectivity, and the pseudovirus infectivity of EG.5.1 was significantly lower than that of its parental lineage (XBB.1.9.2). These results suggest that the increased Re is not due to the increased infectivity caused by these substitutions.

We then performed a neutralisation assay using XBB breakthrough infection (BTI) sera (n=24) to address whether EG.5.1 evades the antiviral effect of the humoral immunity induced by BTI of XBB subvariants. As shown on appendix p 13, the 50% neutralisation titre (NT50) of XBB BTI sera against EG.5.1 was significantly (1·4-fold) lower than those against the parental XBB.1.5/1.9.2 (p<0·0001). The NT50 values of XBB BTI sera against XBB.1.5/1.9.2, XBB.1.16, and XBB.1.5/1.9.2+Q52H were comparable. However, the NT50 value of XBB BTI sera against XBB.1.5/1.9.2+F456L was significantly (1·9-fold) lower than that of the parental XBB.1.5/1.9.2 (p<0·0001, appendix p 13). These results suggest that the increased Re of EG.5.1 is partly attributed to the immune evasion from the humoral immunity elicited by XBB BTI, and S:F456L is a key mutation leading to this immune evasion. We previously demonstrated that omicron BTI cannot efficiently induce antiviral humoral immunity against the infecting variant. In fact, the NT50values of the BTI sera of omicron BA.1, BA.2, and BA.5 against the infecting variant were 3·0-fold, 2·2-fold, and 3·4-fold lower than that against the ancestral B.1.1 variant, respectively. Strikingly, however, we found that the NT50 value of the BTI sera of XBB1.5/1.9.2 and XBB.1.16 against the infecting variant were 6·6-fold and 6·3-fold lower than that against the B.1.1 variant. These results suggest that the BTI of XBB subvariants cannot efficiently induce antiviral humoral immunity against the infecting variants when compared with the BTI of previous omicron variants.

This work was supported in part by the Japan Agency for Medical Research and Development (AMED) Strategic Center of Biomedical Advanced Vaccine Research and Development for Preparedness and Response (SCARDA) Japan Initiative for World-leading Vaccine Research and Development Centers UTOPIA programme (JP223fa627001 to KSat); AMED SCARDA Program on R&D of New Generation Vaccine Including New Modality Application (JP223fa727002 to KSat); AMED Research Program on Emerging and Re-emerging Infectious Diseases (JP22fk0108146 to KSat; JP21fk0108494 to G2P-Japan Consortium and KSat; JP21fk0108425 to KSat; JP21fk0108432 to KSat; JP22fk0108511 to G2P-Japan Consortium and KSat; JP22fk0108516 to KSat; JP22fk0108506 to KSat); AMED Research Program on HIV/AIDS (JP22fk0410039 to KSat); Japan Science and Technology Agency (JST) Precursory Research for Embryonic Science and Technology (PRESTO; JPMJPR22R1 to JI); JST Core Research for Evolutional Science and Technology (CREST; JPMJCR20H4, to KSat); Japan Society for the Promotion of Science (JSPS) KAKENHI Grant-in-Aid for Early-Career Scientists (23K14526 to JI); JSPS Core-to-Core Program (A Advanced Research Networks; JPJSCCA20190008 to KSat); JSPS Research Fellow DC2 (22J11578 to KU); JSPS Research Fellow DC1 (23KJ0710 to YKo); The Tokyo Biochemical Research Foundation (to KSat); and The Mitsubishi Foundation (to KSat). JI has received consulting fees and honoraria for lectures from Takeda Pharmaceutical. KSat has received consulting fees from Moderna Japan and Takeda Pharmaceutical and honoraria for lectures from Gilead Sciences, Moderna Japan, and Shionogi & Co. All other authors declare no competing interests. YKa, YKo, KU, and JI contributed equally to this study. Members of The Genotype to Phenotype Japan (G2P-Japan) Consortium are listed in the appendix (p 15).

 

 


◆Quantity of SARS-CoV-2 RNA copies exhaled per minute during natural breathing over the course of COVID-19 infection【medRxiv　2023年9月8日】

ABSTRACT

SARS-CoV-2 is spread through exhaled breath of infected individuals. A fundamental question in understanding transmission of SARS-CoV-2 is how much virus an individual is exhaling into the environment while they breathe, over the course of their infection. Research on viral load dynamics during COVID-19 infection has focused on internal swab specimens, which provide a measure of viral loads inside the respiratory tract, but not on breath. Therefore, the dynamics of viral shedding on exhaled breath over the course of infection are poorly understood. Here, we collected exhaled breath specimens from COVID-19 patients and used RTq-PCR to show that numbers of exhaled SARS-CoV-2 RNA copies during COVID-19 infection do not decrease significantly until day 8 from symptom-onset. COVID-19-positive participants exhaled an average of 80 SARS-CoV-2 viral RNA copies per minute during the first 8 days of infection, with significant variability both between and within individuals, including spikes over 800 copies a minute in some patients. After day 8, there was a steep drop to levels nearing the limit of detection, persisting for up to 20 days. We further found that levels of exhaled viral RNA increased with self-rated symptom-severity, though individual variation was high. Levels of exhaled viral RNA did not differ across age, sex, time of day, vaccination status or viral variant. Our data provide a fine-grained, direct measure of the number of SARS-CoV-2 viral copies exhaled per minute during natural breathing—including 312 breath specimens collected multiple times daily over the course of infection—in order to fill an important gap in our understanding of the time course of exhaled viral loads in COVID-19.

INTRODUCTION

SARS-CoV-2, the causative agent of COVID-19, spreads through exhaled breath during coughing, talking, singing, and breathing. Levels of SARS-CoV-2 over the course of infection have been extensively characterized in upper and lower respiratory tract specimens such as nasopharyngeal and oropharyngeal swabs, whereas the dynamics of levels on breath over the course of infection remain virtually unexplored. This is despite the fact that quantifying levels of viral shedding on exhaled breath would allow for a direct approximation of the amount of virus an individual is shedding into the environment, thereby exposing others to risk of infection. We know particularly little about the dynamics of viral shedding on breath during unlabored natural breathing, which serves as a baseline for viral transmission on breath.

While the dynamics of viral load inside the host respiratory tract has direct relevance to viral pathology, dynamics of viral load on the host’s breath has direct relevance to infectiousness. Understanding the dynamics of viral shedding on breath is important for prevention of transmission of disease. Measuring viral load characteristics of the primary route of onward transmission is critical to inform isolation times in the clinic, where isolation consumes scarce resources, and to inform public health transmission control protocols. In addition, variables that impact level of viral shedding on breath remain unclear, but may vary by a multitude of factors including symptom severity, days since symptom onset, co-morbidities, viral genotype, and other unknowns. Understanding of these factors requires quantification of exhaled viral loads, which cannot be inferred from internal viral loads.

Current techniques for measuring viral load in exhaled breath have successfully detected SARS-CoV-2 in specimens, though with variable detection rates ranging from 26.9% to 86%. Recent work has also shown that SARS-CoV-2 can be isolated from exhaled breath, confirming that it contains replication-competent virus. However, prior work has focused on exertive breathing conditions (talking, singing, coughing), and we therefore have less understanding of viral loads in exhaled breath during natural breathing. Furthermore, prior work has not analyzed exhaled breath collected longitudinally over the course of infection relative to the day of symptom onset (potentially due to expense and lack of portability of breath collection devices), which would allow for a better understanding of the time course of changes in viral loads on breath. An inexpensive, portable device that allows patients to self-collect breath samples at home would facilitate research into factors that contribute to virus transmission from breath, how exhaled virus levels change over the course of infection, and whether therapeutics and other interventions reduce levels of exhaled virus.

Here, we developed a portable, disposable exhaled breath condensate collection device (EBCD) (Fig S1) and used it to collect 312 specimens from 60 COVID-19-tested outpatients who were treated at Northwestern Memorial Hospital (NMH). Specimens were analyzed using RT-qPCR. Our data set included breath specimens collected multiple times per day over the course of infection. We report numbers of SARS-CoV-2 RNA copies exhaled per minute, during natural breathing, over the course of infection and across a range of factors including self-reported symptom severity, age, sex, presence of co-morbidities, vaccination status and viral variant.

 

◆Long COVID: major findings, mechanisms and recommendations【nature：nature reviews microbiology　2023年1月13日】

Abstract

Long COVID is an often debilitating illness that occurs in at least 10% of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infections. More than 200 symptoms have been identified with impacts on multiple organ systems. At least 65 million individuals worldwide are estimated to have long COVID, with cases increasing daily. Biomedical research has made substantial progress in identifying various pathophysiological changes and risk factors and in characterizing the illness; further, similarities with other viral-onset illnesses such as myalgic encephalomyelitis/chronic fatigue syndrome and postural orthostatic tachycardia syndrome have laid the groundwork for research in the field. In this Review, we explore the current literature and highlight key findings, the overlap with other conditions, the variable onset of symptoms, long COVID in children and the impact of vaccinations. Although these key findings are critical to understanding long COVID, current diagnostic and treatment options are insufficient, and clinical trials must be prioritized that address leading hypotheses. Additionally, to strengthen long COVID research, future studies must account for biases and SARS-CoV-2 testing issues, build on viral-onset research, be inclusive of marginalized populations and meaningfully engage patients throughout the research process.

Introduction

Long COVID (sometimes referred to as ‘post-acute sequelae of COVID-19’) is a multisystemic condition comprising often severe symptoms that follow a severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. At least 65 million individuals around the world have long COVID, based on a conservative estimated incidence of 10% of infected people and more than 651 million documented COVID-19 cases worldwide; the number is likely much higher due to many undocumented cases. The incidence is estimated at 10–30% of non-hospitalized cases, 50–70% of hospitalized cases and 10–12% of vaccinated cases. Long COVID is associated with all ages and acute phase disease severities, with the highest percentage of diagnoses between the ages of 36 and 50 years, and most long COVID cases are in non-hospitalized patients with a mild acute illness, as this population represents the majority of overall COVID-19 cases. There are many research challenges, as outlined in this Review, and many open questions, particularly relating to pathophysiology, effective treatments and risk factors.

Hundreds of biomedical findings have been documented, with many patients experiencing dozens of symptoms across multiple organ systems (Fig. 1). Long COVID encompasses multiple adverse outcomes, with common new-onset conditions including cardiovascular, thrombotic and cerebrovascular disease, type 2 diabetes, myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS) and dysautonomia, especially postural orthostatic tachycardia syndrome (POTS) (Fig. 2). Symptoms can last for years, and particularly in cases of new-onset ME/CFS and dysautonomia are expected to be lifelong. With significant proportions of individuals with long COVID unable to return to work7, the scale of newly disabled individuals is contributing to labour shortages. There are currently no validated effective treatments.

Fig. 1: Long COVID symptoms and the impacts on numerous organs with differing pathology.

The impacts of long COVID on numerous organs with a wide variety of pathology are shown. The presentation of pathologies is often overlapping, which can exacerbate management challenges. MCAS, mast cell activation syndrome; ME/CFS, myalgic encephalomyelitis/chronic fatigue syndrome; POTS, postural orthostatic tachycardia syndrome.

Fig. 2: SARS-CoV-2 infection, COVID-19 and long COVID increases the risk of several medical conditions.

Because diagnosis-specific data on large populations with long COVID are sparse, outcomes from general infections are included and a large proportion of medical conditions are expected to result from long COVID, although the precise proportion cannot be determined. One year after the initial infection, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infections increased the risk of cardiac arrest, death, diabetes, heart failure, pulmonary embolism and stroke, as studied with use of US Department of Veterans Affairs databases. Additionally, there is clear increased risk of developing myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS) and dysautonomia. Six months after breakthrough infection, increased risks were observed for cardiovascular conditions, coagulation and haematological conditions, death, fatigue, neurological conditions and pulmonary conditions in the same cohort. The hazard ratio is the ratio of how often an event occurs in one group relative to another; in this case people who have had COVID-19 compared with those who have not. Data sources are as follows: diabetes, cardiovascular outcomes, dysautonomia, ME/CFS and breakthrough infections.

There are likely multiple, potentially overlapping, causes of long COVID. Several hypotheses for its pathogenesis have been suggested, including persisting reservoirs of SARS-CoV-2 in tissues; immune dysregulation with or without reactivation of underlying pathogens, including herpesviruses such as Epstein–Barr virus (EBV) and human herpesvirus 6 (HHV-6) among others; impacts of SARS-CoV-2 on the microbiota, including the virome; autoimmunity and priming of the immune system from molecular mimicry; microvascular blood clotting with endothelial dysfunction; and dysfunctional signalling in the brainstem and/or vagus nerve (Fig. 3). Mechanistic studies are generally at an early stage, and although work that builds on existing research from postviral illnesses such as ME/CFS has advanced some theories, many questions remain and are a priority to address. Risk factors potentially include female sex, type 2 diabetes, EBV reactivation, the presence of specific autoantibodies, connective tissue disorders, attention deficit hyperactivity disorder, chronic urticaria and allergic rhinitis, although a third of people with long COVID have no identified pre-existing conditions6. A higher prevalence of long Covid has been reported in certain ethnicities, including people with Hispanic or Latino heritage. Socio-economic risk factors include lower income and an inability to adequately rest in the early weeks after developing COVID-19. Before the emergence of SARS-CoV-2, multiple viral and bacterial infections were known to cause postinfectious illnesses such as ME/CFS, and there are indications that long COVID shares their mechanistic and phenotypic characteristics. Further, dysautonomia has been observed in other postviral illnesses and is frequently observed in long COVID.

Fig. 3: Hypothesized mechanisms of long COVID pathogenesis.

There are several hypothesized mechanisms for long COVID pathogenesis, including immune dysregulation, microbiota disruption, autoimmunity, clotting and endothelial abnormality, and dysfunctional neurological signalling. EBV, Epstein–Barr virus; HHV-6, human herpesvirus 6; SARS-CoV-2, severe acute respiratory syndrome coronavirus 2.

In this Review, we explore the current knowledge base of long COVID as well as misconceptions surrounding long COVID and areas where additional research is needed. Because most patients with long COVID were not hospitalized for their initial SARS-CoV-2 infection, we focus on research that includes patients with mild acute COVID-19 (meaning not hospitalized and without evidence of respiratory disease). Most of the studies we discuss refer to adults, except for those in Box 1.

Box 1 Long COVID in children

Long COVID impacts children of all ages. One study found that fatigue, headache, dizziness, dyspnoea, chest pain, dysosmia, dysgeusia, reduced appetite, concentration difficulties, memory issues, mental exhaustion, physical exhaustion and sleep issues were more common in individuals with long COVID aged 15–19 years compared with controls of the same age. A nationwide study in Denmark comparing children with a positive PCR test result with control individuals found that the former had a higher chance of reporting at least one symptom lasting more than 2 months. Similarly to adults with long COVID, children with long COVID experience fatigue, postexertional malaise, cognitive dysfunction, memory loss, headaches, orthostatic intolerance, sleep difficulty and shortness of breath. Liver injury has been recorded in children who were not hospitalized during acute severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infections, and although rare, children who had COVID-19 have increased risks of acute pulmonary embolism, myocarditis and cardiomyopathy, venous thromboembolic events, acute and unspecified renal failure, and type 1 diabetes. Infants born to women who had COVID-19 during pregnancy were more likely to receive a neurodevelopmental diagnosis in the first year after delivery. A paediatric long COVID centre’s experience treating patients suggests that adolescents with a moderate to severe form of long COVID have features consistent with myalgic encephalomyelitis/chronic fatigue syndrome. Children experiencing long COVID have hypometabolism in the brain similar to the patterns found in adults with long COVID. Long-term pulmonary dysfunction is found in children with long COVID and those who have recovered from COVID-19. Children with long COVID were more likely to have had attention deficit hyperactivity disorder, chronic urticaria and allergic rhinitis before being infected.

More research on long COVID in children is needed, although there are difficulties in ensuring a proper control group due to testing issues. Several studies have found that children infected with SARS-CoV-2 are considerably less likely to have a positive PCR test result than adults despite seroconverting weeks later, with up to 90% of cases being missed. Additionally, children are much less likely to seroconvert and, if they develop antibodies, are more likely to have a waning response months after infection compared with adults.