Waning cellular immune responses and predictive factors in maintaining cellular immunity against SARS-CoV-2 six months after BNT162b2 mRNA vaccination.

Several clinical trials have shown that the humoral response produced by anti-spike antibodies elicited by coronavirus disease 2019 (COVID-19) vaccines gradually declines. The kinetics, durability and influence of epidemiological and clinical factors on cellular immunity have not been fully elucidated. We analyzed cellular immune responses elicited by BNT162b2 mRNA vaccines in 321 health care workers using whole blood interferon-gamma (IFN-γ) release assays. IFN-γ, induced by CD4 + and CD8 + T cells stimulated with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike epitopes (Ag2), levels were highest at 3 weeks after the second vaccination (6 W) and decreased by 37.4% at 3 months (4 M) and 60.0% at 6 months (7 M), the decline of which seemed slower than that of anti-spike antibody levels. Multiple regression analysis revealed that the levels of IFN-γ induced by Ag2 at 7 M were significantly correlated with age, dyslipidemia, focal adverse reactions to full vaccination, lymphocyte and monocyte counts in whole blood, Ag2 levels before the second vaccination, and Ag2 levels at 6 W. We clarified the dynamics and predictive factors for the long-lasting effects of cellular immune responses. The results emphasize the need for a booster vaccine from the perspective of SARS-CoV-2 vaccine-elicited cellular immunity.

A case report of fatal COVID-19 complicated by rapidly progressive sepsis caused by Klebsiella variicola.

BACKGROUND: There is a growing interest in Klebsiella variicola as a causative pathogen in humans, though its clinical features and the impact of co-infection or secondary infection with COVID-19 remain unknown. CASE PRESENTATION: A 71-year-old man presented with fever, altered mental status and generalized weakness and was admitted to ICU due to severe COVID-19 pneumonia. He was newly diagnosed with type II diabetes mellitus upon admission. On hospital day 3, his respiratory status deteriorated, requiring invasive mechanical ventilation. On hospital day 10, superimposed bacterial pneumonia was suspected and subsequently, broad-spectrum antibiotics were administered for the associated bloodstream infection. On hospital day 13, despite administration of active antibiotics and appropriate source control, he decompensated and died. The causative organism isolated from blood cultures was initially reported as K. pneumoniae, but it was identified as K. variicola by a genetic analysis. A representative isolate (FUJ01370) had a novel multilocus sequence typing allelic profile (gapA-infB-mdh-pgi-phoE-rpoB-tonB: 16-24-21-27-52-17-152), to which sequence type 5794 was assigned (GenBank assembly accession: GCA_019042755.1). CONCLUSIONS: We report a fatal case of respiratory and bloodstream infection due to K. variicola complicating severe COVID-19. Co-infection or secondary infection of K. variicola in COVID-19 is likely under-recognized and can be fulminant as in this case.

Multicenter, single-blind, randomized controlled study of the efficacy and safety of favipiravir and nafamostat mesilate in patients with COVID-19 pneumonia.

OBJECTIVES: To evaluate the efficacy and safety of nafamostat combined with favipiravir for the treatment of COVID-19. METHODS: We conducted a multicenter, randomized, single-blind, placebo-controlled, parallel assignment study in hospitalized patients with mild-to-moderate COVID-19 pneumonia. Patients were randomly assigned to receive favipiravir alone (n = 24) or nafamostat with favipiravir (n = 21). The outcomes included changes in the World Health Organization clinical progression scale score, time to improvement in body temperature, and improvement in oxygen saturation (SpO2). RESULTS: There was no significant difference in the changes in the clinical progression scale between nafamostat with favipiravir and favipiravir alone groups (median, -0.444 vs -0.150, respectively; least-squares mean difference, -0.294; P = 0.364). The time to improvement in body temperature was significantly shorter in the combination group (5.0 days; 95% confidence interval, 4.0-7.0) than in the favipiravir group (9.0 days; 95% confidence interval, 7.0-18.0; P =0.009). The changes in SpO2 were greater in the combination group than in the favipiravir group (0.526% vs -1.304%, respectively; least-squares mean difference, 1.831; P = 0.022). No serious adverse events or deaths were reported, but phlebitis occurred in 57.1% of the patients in the combination group. CONCLUSION: Although our study showed no differences in clinical progression, earlier defervescence, and recovery of SpO2 were observed in the combination group.

Dynamic modulations of urinary sphingolipid and glycerophospholipid levels in COVID-19 and correlations with COVID-19-associated kidney injuries.

BACKGROUND: Among various complications of coronavirus disease 2019 (COVID-19), caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), renal complications, namely COVID-19-associated kidney injuries, are related to the mortality of COVID-19. METHODS: In this retrospective cross-sectional study, we measured the sphingolipids and glycerophospholipids, which have been shown to possess potent biological properties, using liquid chromatography-mass spectrometry in 272 urine samples collected longitudinally from 91 COVID-19 subjects and 95 control subjects without infectious diseases, to elucidate the pathogenesis of COVID-19-associated kidney injuries. RESULTS: The urinary levels of C18:0, C18:1, C22:0, and C24:0 ceramides, sphingosine, dihydrosphingosine, phosphatidylcholine, lysophosphatidylcholine, lysophosphatidic acid, and phosphatidylglycerol decreased, while those of phosphatidylserine, lysophosphatidylserine, phosphatidylethanolamine, and lysophosphatidylethanolamine increased in patients with mild COVID-19, especially during the early phase (day 1-3), suggesting that these modulations might reflect the direct effects of infection with SARS-CoV-2. Generally, the urinary levels of sphingomyelin, ceramides, sphingosine, dihydrosphingosine, dihydrosphingosine L-phosphate, phosphatidylcholine, lysophosphatidic acid, phosphatidylserine, lysophosphatidylserine, phosphatidylethanolamine, lysophosphatidylethanolamine, phosphatidylglycerol, lysophosphatidylglycerol, phosphatidylinositol, and lysophosphatidylinositol increased, especially in patients with severe COVID-19 during the later phase, suggesting that their modulations might result from kidney injuries accompanying severe COVID-19. CONCLUSIONS: Considering the biological properties of sphingolipids and glycerophospholipids, an understanding of their urinary modulations in COVID-19 will help us to understand the mechanisms causing COVID-19-associated kidney injuries as well as general acute kidney injuries and may prompt researchers to develop laboratory tests for predicting maximum severity and/or novel reagents to suppress the renal complications of COVID-19.

Dynamic modulations of sphingolipids and glycerophospholipids in COVID-19.

BACKGROUND: A heterogeneous clinical phenotype is a characteristic of coronavirus disease 2019 (COVID-19). Therefore, investigating biomarkers associated with disease severity is important for understanding the mechanisms responsible for this heterogeneity and for developing novel agents to prevent critical conditions. This study aimed to elucidate the modulations of sphingolipids and glycerophospholipids, which have been shown to possess potent biological properties. METHODS: We measured the serum sphingolipid and glycerophospholipid levels in a total of 887 samples from 215 COVID-19 subjects, plus 115 control subjects without infectious diseases and 109 subjects with infectious diseases other than COVID-19. RESULTS: We observed the dynamic modulations of sphingolipids and glycerophospholipids in the serum of COVID-19 subjects, depending on the time course and severity. The elevation of C16:0 ceramide and lysophosphatidylinositol and decreases in C18:1 ceramide, dihydrosphingosine, lysophosphatidylglycerol, phosphatidylglycerol and phosphatidylinositol were specific to COVID-19. Regarding the association with maximum severity, phosphatidylinositol and phosphatidylcholine species with long unsaturated acyl chains were negatively associated, while lysophosphatidylethanolamine and phosphatidylethanolamine were positively associated with maximum severity during the early phase. Lysophosphatidylcholine and phosphatidylcholine had strong negative correlations with CRP, while phosphatidylethanolamine had strong positive ones. C16:0 ceramide, lysophosphatidylcholine, phosphatidylcholine and phosphatidylethanolamine species with long unsaturated acyl chains had negative correlations with D-dimer, while phosphatidylethanolamine species with short acyl chains and phosphatidylinositol had positive ones. Several species of phosphatidylcholine, phosphatidylethanolamine and sphingomyelin might serve as better biomarkers for predicting severe COVID-19 during the early phase than CRP and D-dimer. Compared with the lipid modulations seen in mice treated with lipopolysaccharide, tissue factor, or histone, the lipid modulations observed in severe COVID-19 were most akin to those in mice administered lipopolysaccharide. CONCLUSION: A better understanding of the disturbances in sphingolipids and glycerophospholipids observed in this study will prompt further investigation to develop laboratory testing for predicting maximum severity and/or novel agents to suppress the aggravation of COVID-19.

The association between adverse reactions and immune response against SARS-CoV-2 spike protein after vaccination with BNT162b2 among healthcare workers in a single healthcare system: a prospective observational cohort study.

Adverse reactions after vaccination with COVID-19 mRNA vaccines are common; however, the association between adverse reactions and humoral responses is uncertain. To determine whether humoral immune responses after BNT162b2 vaccine administration were associated with local and systemic adverse reactions, we conducted a prospective observational cohort study in a single tertiary referral center. Healthcare workers who received the first dose of BNT162b2 vaccine were recruited. SARS-CoV-2 anti-spike IgG antibody titers were measured three weeks after the second dose and information about adverse reactions after vaccination was collected. Among the 887 participants, 641 (72.3%) were women. The median age was 38 (range, 22-74) years. All but one showed anti-spike IgG levels well above the cutoff, with a median level of 13,600 arbitrary units/mL. Overall, 800 (92.2%) participants reported some reactions after the first dose and 822 (96.3%) after the second dose. Significantly more participants reported systemic reactions after the second dose than after the first dose (P < .01), and 625 (73.6%) reported that reactions were stronger after the second dose. Factors positively associated with elevation of anti-spike IgG levels were history of asthma (24% higher if present, P = .01) and stronger reactions after the second dose (19% higher if experienced, P = .02). The majority of participants showed good humoral responses and reported some adverse reactions after vaccination. Anti-spike IgG levels were significantly higher if adverse reactions after the second dose were stronger than those after the first dose. These findings may help inform current and future vaccine recipients.

Urine sediment findings were milder in patients with COVID-19-associated renal injuries than in those with non-COVID-19-associated renal injuries.

BACKGROUND: Acute renal injury is an important complication of coronavirus disease 2019 (COVID-19). Both COVID-19-specific mechanisms, such as damage to the renal parenchyma by direct infection, and non-specific mechanisms, such as the pre-renal injury factors, have been proposed to be involved in COVID-19-associated renal injuries. In this study, we aimed to elucidate the characteristics of COVID-19-associated renal injuries, focusing mainly on urine sediment findings. METHODS: We compared the urine sediment findings and their associations with renal functions or urinary clinical parameters between subjects with COVID-19 and subjects without COVID-19 with acute renal injuries. RESULTS: We found that the number of urine sediment particles and the levels of N-acetyl-ß-D-glucosaminidase, α1-microglobulin, liver type fatty acid-binding protein, and neutrophil gelatinase-associated lipocalin were associated with the severity of COVID-19. In addition, we observed that the number of granular casts, epithelial casts, waxy casts, and urinary chemical marker levels were lower in the subjects with COVID-19 than subjects without COVID-19 with acute renal injuries when the subjects were classified according to their renal function. CONCLUSIONS: These results suggest that pre-renal injury factors might be largely involved in the pathogenesis of COVID-19-associated renal injuries compared with non-COVID-19-associated renal injuries arising from surgery or sepsis.

Association of the Serum Levels of the Nucleocapsid Antigen of SARS-CoV-2 With the Diagnosis, Disease Severity, and Antibody Titers in Patients With COVID-19: A Retrospective Cross-Sectional Study.

Background: Several types of laboratory tests for COVID-19 have been established to date; however, the clinical significance of the serum SARS-CoV-2 nucleocapsid (N) antigen levels remains to be fully elucidated. In the present study, we attempted to elucidate the usefulness and clinical significance of the serum N antigen levels. Methods: We measured the serum N antigen levels in 391 serum samples collected from symptomatic patients with a confirmed diagnosis of COVID-19 and 96 serum samples collected from patients with non-COVID-19, using a fully automated chemiluminescence immunoassay analyzer. Results: Receiver operating characteristic analysis identified the optimal cutoff value of the serum N antigen level (cutoff index, based on Youden's index) as 0.255, which yielded a sensitivity and specificity for the diagnosis of COVID-19 of 91.0 and 81.3%, respectively. The serum N antigen levels were significantly higher in the patient groups with moderate and severe COVID-19 than with mild disease. Moreover, a significant negative correlation was observed between the serum N antigen levels and the SARS-CoV-2 IgG antibody titers, especially in patients with severe COVID-19. Conclusion: Serum N antigen testing might be useful both for the diagnosis of COVID-19 and for obtaining a better understanding of the clinical features of the disease.

Standardized reporting systems of chest computed tomography in a population with low coronavirus disease 2019 prevalence: A retrospective comparative study.

PURPOSE: To compare the diagnostic performance and interobserver agreement of three reporting systems for computed tomography findings in coronavirus disease 2019 (COVID-19), namely the COVID-19 Reporting and Data System (CO-RADS), COVID-19 Imaging Reporting and Data System (COVID-RADS), and Radiological Society of North America (RSNA) expert consensus statement, in a low COVID-19 prevalence area. METHOD: This institutional review board approval single-institutional retrospective study included 154 hospitalized patients between April 1 and May 21, 2020; 26 (16.9 %; 63.2 ± 14.1 years, 21 men) and 128 (65.7 ± 16.4 years, 87 men) patients were diagnosed with and without COVID-19 according to reverse transcription-polymerase chain reaction results, respectively. Written informed consent was waived due to the retrospective nature of the study. Six radiologists independently classified chest computed tomography images according to each reporting system. The area under receiver operating characteristic curves, sensitivity, specificity, positive predictive value, negative predictive value, accuracy, and interobserver agreements were calculated and compared across the systems using paired t-test and kappa analysis. RESULTS: Mean area under receiver operating characteristic curves were as follows: CO-RADS, 0.89 (95 % confidence interval [CI], 0.87-0.90); COVID-RADS, 0.78 (0.75-0.80); and RSNA expert consensus statement, 0.88 (0.86-0.90). Average kappa values across observers were 0.52 (95 % CI: 0.45-0.60), 0.51 (0.41-0.61), and 0.57 (0.49-0.64) for CO-RADS, COVID-RADS, and RSNA expert consensus statement, respectively. Sensitivity, specificity, positive predictive value, negative predictive value, and accuracy were the highest at 0.71, 0.53, 0.72, 0.96, and 0.56 in the CO-RADS; 0.56, 0.31, 0.54, 0.95, and 0.35 in the COVID-RADS; 0.83, 0.49, 0.61, 0.96, and 0.55 in the RSNA expert consensus statement, respectively. CONCLUSIONS: The CO-RADS exhibited the highest specificity, positive predictive value, which are especially important in a low-prevalence population, while maintaining high accuracy and negative predictive value, demonstrating the best performance in a low-prevalence population.

Comparative performance and cycle threshold values of 10 nucleic acid amplification tests for SARS-CoV-2 on clinical samples.

BACKGROUND: A number of nucleic acid amplification tests (NAATs) for SARS-CoV-2 with different reagents have been approved for clinical use in Japan. These include research kits approved under emergency use authorization through simplified process to stabilize the supply of the reagents. Although these research kits have been increasingly used in clinical practice, limited data is available for the diagnostic performance in clinical settings. METHODS: We compared sensitivity, specificity, and cycle threshold (Ct) values obtained by NAATs using 10 kits approved in Japan including eight kits those receiving emergency use authorization using 69 frozen-stored clinical samples including 23 positive samples with various Ct values and 46 negative samples. RESULTS: Viral copy number of the frozen-stored samples determined with LightMix E-gene test ranged from 0.6 to 84521.1 copies/µL. While no false-positive results were obtained by any of these tests (specificity: 100% [95% CI, 88.9%-100%]), sensitivity of the nine tests ranged from 68.2% [95% CI, 45.1%-86.1%] to 95.5% [95% CI, 77.2%-99.9%] using LightMix E-gene test as the gold standard. All tests showed positive results for all samples with ≥100 copies/µL. Significant difference of Ct values even among tests amplifying the same genetic region (N1-CDC, N2) was also observed. CONCLUSION: Difference in the diagnostic performance was observed among NAATs approved in Japan. Regarding diagnostic kits for emerging infectious diseases, a system is needed to ensure both rapidity of reagent supply and accuracy of diagnosis. Ct values, which are sometimes regarded as a marker of infectivity, are not interchangeable when obtained by different assays.

Diagnostic prediction of COVID-19 based on clinical and radiological findings in a relatively low COVID-19 prevalence area.

BACKGROUND: Distinguishing coronavirus disease 2019 (COVID-19) pneumonia from other lung diseases is often difficult, especially in a highly comorbid patient population in a low prevalence region. We aimed to distinguish clinical data and computed tomography (CT) images between COVID-19 and other lung diseases in an advanced care hospital. METHODS: We assessed clinical characteristics, laboratory data, and chest CT images of patients with COVID-19 and non-COVID-19 patients who were suspected of having COVID-19 between February 20 and May 21, 2020, at the University of Tokyo Hospital. RESULTS: Typical appearance for COVID-19 on CT images were found in 24 of 29 COVID-19 cases and 21 of 168 non-COVID-19 cases, according to the Radiological Society of North America Expert Consensus Statement (for predicting COVID-19, sensitivity 0.828, specificity 0.875, positive predictive value 0.533, negative predictive value 0.967). When we focused on cases with typical CT images, loss of taste or smell, and close contact with COVID-19 patients were exclusive characteristics for the COVID-19 cases. Among laboratory data, high fibrinogen (P < 0.01) and low white blood cell count (P < 0.01) were good predictors for COVID-19 with typical CT images in multivariate analysis. CONCLUSIONS: In a relatively low prevalence region, CT screening has high sensitivity to COVID-19 in patients with suspected symptoms. When chest CT findings are typical for COVID-19, close contact, loss of taste or smell, lower white blood cell count, and higher fibrinogen are good predictors for COVID-19.

Clot waveform of APTT has abnormal patterns in subjects with COVID-19.

In Coronavirus disease 2019 (COVID-19) subjects, recent evidence suggests the presence of unique coagulation abnormalities. In this study, we performed clot waveform analyses to investigate whether specific modulations are observed in COVID-19 subjects. We analyzed the second derivative of the absorbance in routine APTT tests performed using an ACL-TOP system. We observed high frequencies of abnormal patterns in APTT second-derivative curves that could be classified into an early shoulder type, a late shoulder type, or a biphasic type, high maximum first-derivative and second-derivative peak levels, and a low minimum second-derivative peak level in COVID-19 subjects. These modulations were not observed in subjects with disseminated intravascular coagulation. These abnormal patterns are also observed in patients with lupus anticoagulant, hemophilia, or factor IX deficiency. The plasma fibrinogen levels might also be involved in the abnormal APTT waveforms, especially the high maximum first-derivative and second-derivative peak levels. The abnormal patterns in the APTT second-derivative curves appear with highest frequency at around 2 weeks after the onset of COVID-19 and were not associated with the severity of COVID-19. These results suggest the possible presence of a specific abnormal coagulopathy in COVID-19.

COVID-19 and Fatal Sepsis Caused by Hypervirulent Klebsiella pneumoniae, Japan, 2020.

A patient in Japan with coronavirus disease and hypervirulent Klebsiella pneumoniae K2 sequence type 86 infection died of respiratory failure. Bacterial and fungal co-infections caused by region-endemic pathogens, including hypervirulent K. pneumoniae in eastern Asia, should be included in the differential diagnosis of coronavirus disease patients with acutely deteriorating condition.

Dynamic changes in fibrinogen and D-dimer levels in COVID-19 patients on nafamostat mesylate.

Critical illnesses associated with coronavirus disease 2019 (COVID-19) are attributable to a hypercoagulable status. There is limited knowledge regarding the dynamic changes in coagulation factors among COVID-19 patients on nafamostat mesylate, a potential therapeutic anticoagulant for COVID-19. First, we retrospectively conducted a cluster analysis based on clinical characteristics on admission to identify latent subgroups among fifteen patients with COVID-19 on nafamostat mesylate at the University of Tokyo Hospital, Japan, between April 6 and May 31, 2020. Next, we delineated the characteristics of all patients as well as COVID-19-patient subgroups and compared dynamic changes in coagulation factors among each subgroup. The subsequent dynamic changes in fibrinogen and D-dimer levels were presented graphically. All COVID-19 patients　were classified into three subgroups: clusters A, B, and C, representing low, intermediate, and high risk of poor outcomes, respectively. All patients were alive 30 days from symptom onset. No patient in cluster A required mechanical ventilation; however, all patients in cluster C required mechanical ventilation, and half of them were treated with venovenous extracorporeal membrane oxygenation. All patients in cluster A maintained low D-dimer levels, but some critical patients in clusters B and C showed dynamic changes in fibrinogen and D-dimer levels. Although the potential of nafamostat mesylate needs to be evaluated in randomized clinical trials, admission characteristics of patients with COVID-19 could predict subsequent coagulopathy.