A fluorescence-based sweat test sensor in a proof-of-concept clinical study for COVID-19 screening diagnosis.

During the corona virus disease 2019 (COVID-19) pandemic period, rapid screening of covid-19 patients has been of great interest by developing a fluorescent sensor for complexation with nonanal, which is a marker for Covid-19 detection in sweat. Solid phase micro-extraction gas chromatography-mass spectrometry (SPME GC-MS) was initially used to quantify nonanal in armpit sweat samples based on an external calibration curve. A sample containing a nonanal content above the threshold of 1.04 µL is expected to be COVID-19 positive with a sensitivity and specificity of 87% and 89%, respectively, validated by comparison with RT-PCR results. For more practical applications, helicene dye-encapsulated ethyl cellulose, namely EC@dyeNH, was applied to screen 140 sweat samples collected from the foreheads of volunteers. The mixed sensor and sweat solution droplets were then visualized and imaged under blacklight. The COVID-19 positive droplets exhibited yellow fluorescence emission, the brightness of which could be measured by using ImageJ in the grey scale. With the optimum color intensity of >73 for positive results, the screening performance was observed with a sensitivity and specificity of 96% and 93%, respectively. The overall test time of this method is approximately less than 15 min. This alternative method offers a promising practical screening approach for the diagnosis of COVID-19 in sweat.

Self-enhancement lateral flow immunoassay for COVID-19 diagnosis.

Equipment-free colorimetric-based lateral flow immunoassay (LFIA) is the most convenient and popular tool for various applications, including diagnostic tools requiring high sensitivity for the detection of pathogens. Thus, improvements and developments of LFIA are constantly being reported. Herein, we enriched the sensitivity of LFIA using the gold enhancement principle, emphasizing needlessly complicated apparatus, only one step for the strip test operation, and typical time incubation (15 min) process. Self-enhanced LFIA was then executed for subsequent flows by overlapping the additionally enhanced pad composed of gold ions and reducing agent on the conjugate pad and the sample pad. Self-enhanced LFIA was performed to detect SARS-CoV-2 antigens in saliva. The obtained result depicted that the achieved sensitivity was up to tenfold compared with that of conventional LFIA by visual measurements. The detection limits of self-enhanced LFIA detecting nucleocapsid protein antigens in the saliva sample was 0.50 and 0.10 ng/mL employed by naked eye detection and calibration curve-based calculation, respectively. When the proposed device was applied to 207 human saliva samples, the diagnostic performance presented a 96.10 % sensitivity and 99.23 % specificity. This self-enhanced LFIA could be implemented in large-scale production and demonstrates higher sensitivity with effortless use, which meets the requirements for point-of-care testing and on-field mass screening.

Telepharmacy during home isolation: drug-related problems and pharmaceutical care in COVID-19 patients receiving antiviral therapy in Thailand.

BACKGROUND: Home isolation has been proposed for coronavirus disease 2019 (COVID-19) patients with mild symptoms to avoid hospital overcrowding. This study aimed to describe the drug-related problems (DRPs) and the pharmaceutical care of home-isolating COVID-19 patients in Thailand. METHODS: Our cross-sectional study was undertaken from July 1 to September 30, 2021, at the King Chulalongkorn Memorial Hospital, Thailand. Patients who were ≥ 18 years old, were diagnosed with mild COVID-19 by real-time polymerase chain reaction (RT-PCR), and were able to isolate at home while receiving an antiviral agent and standard symptomatic treatment were enrolled. Infectious disease pharmacists provided a telepharmacy service on days 1 and 3 after the COVID-19 diagnosis. RESULTS: A total of 197 patients met the study criteria. Their median age was 45 years, and their most common underlying disease was hypertension (44.29%). All patients exhibited excellent anti-COVID-19 drug adherence. We identified 125 DRPs, including adverse reactions (68%), and the unnecessary use of products (62.40%). Moreover, 91 patients (46.19%) reported the use of supplements or herbs, with vitamin C being the main supplement (37.36%). Pharmacists provided 36 recommendations and received 33 questions from COVID-19 patients. CONCLUSIONS: Our study demonstrates that telepharmacy is an essential service for detecting and preventing DRPs in home-isolating COVID-19 patients.

Simultaneous detection of omicron and other SARS-CoV-2 variants by multiplex PCR MassARRAY technology.

The rapid emergence of SARS-CoV-2 variants with high severity and transmutability adds further urgency for rapid and multiplex molecular testing to identify the variants. A nucleotide matrix-assisted laser-desorption-ionization time-of-flight mass spectrophotometry (MALDI-TOF MS)-based assay was developed (called point mutation array, PMA) to identify four major SARS-CoV-2 variants of concern (VOCs) including Alpha, Beta, Delta, and Omicron (namely PMA-ABDO) and differentiate Omicron subvariant (namely PMA-Omicron). PMA-ABDO and PMA-Omicron consist of 24 and 28 mutation sites of the spike gene. Both PMA panels specifically identified VOCs with as low as 10 viral copies/µl. The panel has shown a 100% concordant with the Next Generation Sequencing (NGS) results testing on 256 clinical specimens with real-time PCR cycle threshold (Ct) values less than 26. It showed a higher sensitivity over NGS; 25/28 samples were positive by PMA but not NGS in the clinical samples with PCR Ct higher than 26. Due to the mass of nucleotide used to differentiate between wild-type and mutation strains, the co-infection or recombination of multiple variants can be determined by the PMA method. This method is flexible in adding a new primer set to identify a new emerging mutation site among the current circulating VOCs and the turnaround time is less than 8 h. However, the spike gene sequencing or NGS retains the advantage of detecting newly emerged variants.

First case series and literature review of coronavirus disease 2019 (COVID-19) associated pulmonary tuberculosis in Southeast Asia: Challenges and opportunities.

BACKGROUND: Subclinical tuberculosis (TB) is accidentally detected by radiologic and microbiologic findings. Transmission by those with subclinical TB could delay prevention effort. However, our study demonstrated positive aspect of COVID-19 outbreak as it could allow subclinical TB to be detected faster through a chest X-Ray (CXR). METHODS: This cross-sectional cohort study aimed to report demographics, comorbidities, and outcomes related to early detection of TB among COVID-19 patients, and to elaborate the association between SARS-CoV-2 and pulmonary TB. Data of patients with SARS-CoV-2 co-infection with Mycobacterium tuberculosis (MTB) diagnosed between March 2020 - March 2022 was collected. RESULTS: Out of 12,275 COVID-19 patients, 26 were definitively diagnosed with MTB infection (mean age 48.16 ± 20.17 years). All cases that had suspicious CXR that were not typical for COVID-19, were tested for MTB. On average, pulmonary TB was diagnosed after admission 5(3-10) days, the treatment initiation period was 3(1-5) days from the TB diagnosis. CONCLUSIONS: This suggests an early detection of tuberculosis among COVID-19 patients by quicker screening CXR and sputum comparing to previous symptom guided screening. Thereby reducing the chance of TB transmission demonstrated during COVID-19 pandemic. So, clinicians should be aware of pulmonary tuberculosis in COVID-19 patients with atypical radiologic findings.

Immunogenicity of SARS-CoV-2 vaccines in patients with multiple myeloma: a systematic review and meta-analysis.

Patients with multiple myeloma (MM) have a diminished immune response to coronavirus disease 2019 (COVID-19) vaccines. Risk factors for an impaired immune response are yet to be determined. We aimed to summarize the COVID-19 vaccine immunogenicity and to identify factors that influence the humoral immune response in patients with MM. Two reviewers independently conducted a literature search in MEDLINE, Embase, ISI Web of Science, Cochrane library, and Clinicaltrials.gov from existence until 24 May 24 2022. (PROSPERO: CRD42021277005). A total of 15 studies were included in the systematic review and 5 were included in the meta-analysis. The average rate (range) of positive functional T-lymphocyte response was 44.2% (34.2%-48.5%) after 2 doses of messenger RNA (mRNA) COVID-19 vaccines. The average antispike antibody response rates (range) were 42.7% (20.8%-88.5%) and 78.2% (55.8%-94.2%) after 1 and 2 doses of mRNA COVID-19 vaccines, respectively. The average neutralizing antibody response rates (range) were 25% (1 study) and 62.7% (53.3%-68.6%) after 1 and 2 doses of mRNA COVID-19 vaccines, respectively. Patients with high-risk cytogenetics or receiving anti-CD38 therapy were less likely to have a humoral immune response with pooled odds ratios of 0.36 (95% confidence interval [95% CI], 0.18, 0.69), I2 = 0% and 0.42 (95% CI, 0.22, 0.79), I2 = 14%, respectively. Patients who were not on active MM treatment were more likely to respond with pooled odds ratio of 2.42 (95% CI, 1.10, 5.33), I2 = 7%. Patients with MM had low rates of humoral and cellular immune responses to the mRNA COVID-19 vaccines. Further studies are needed to determine the optimal doses of vaccines and evaluate the use of monoclonal antibodies for pre-exposure prophylaxis in this population.

Impact of SARS-CoV-2 infection on the profiles and responses of innate immune cells after recovery.

BACKGROUNDS: SARS-CoV-2 infection results in a broad spectrum of clinical outcomes, ranging from asymptomatic to severe symptoms and death. Most COVID-19 pathogenesis is associated with hyperinflammatory conditions driven primarily by myeloid cell lineages. The long-term effects of SARS-CoV-2 infection post recovery include various symptoms. METHODS: We performed a longitudinal study of the innate immune profiles 1 and 3 months after recovery in the Thai cohort by comparing patients with mild, moderate, and severe clinical symptoms using peripheral blood mononuclear cells (n = 62). RESULTS: Significant increases in the frequencies of monocytes compared to controls and NK cells compared to mild and moderate patients were observed in severe patients 1-3 months post recovery. Increased polymorphonuclear myeloid-derived suppressor cells (PMN-MDSCs) were observed in all recovered patients, even after 3 months. Increased IL-6 and TNFα levels in monocytes were observed 1 month after recovery in response to lipopolysaccharide (LPS) stimulation, while decreased CD86 and HLA-DR levels were observed regardless of stimulation. A multiplex analysis of serum cytokines performed at 1 month revealed that most innate cytokines, except for TNFα, IL4/IL-13 (Th2) and IFNγ (Th1), were elevated in recovered patients in a severity-dependent manner. Finally, the myelopoiesis cytokines G-CSF and GM-CSF were higher in all patient groups. Increased monocytes and IL-6- and TNFα-producing cells were significantly associated with long COVID-19 symptoms. CONCLUSIONS: These results reveal that COVID-19 infection influences the frequencies and functions of innate immune cells for up to 3 months after recovery, which may potentially lead to some of the long COVID symptoms.

Impact of SARS-CoV2 Infection on the Profiles and Responses of Innate Immune Cells After Recovery

Backgrounds SARS-CoV-2 infection results in a broad spectrum of clinical outcomes, ranging from asymptomatic to severe symptoms and death. Most COVID-19 pathogenesis is associated with hyperinflammatory conditions driven primarily by myeloid cell lineages. The long-term effects of SARS-CoV-2 infection post recovery include various symptoms. Methods We performed a longitudinal study of the innate immune profiles 1 and 3 months after recovery in the Thai cohort by comparing patients with mild, moderate, and severe clinical symptoms using peripheral blood mononuclear cells (n=62). Results Significant increases in the frequencies of monocytes compared to controls and NK cells compared to mild and moderate patients were observed in severe patients 1-3 months post recovery. Increased polymorphonuclear myeloid-derived suppressor cells (PMN-MDSCs) were observed in all recovered patients, even after 3 months. Increased IL-6 and TNFα levels in monocytes were observed 1 month after recovery in response to lipopolysaccharide (LPS) stimulation, while decreased CD86 and HLA-DR levels were observed regardless of stimulation. A multiplex analysis of serum cytokines performed at 1 month revealed that most innate cytokines, except for TNFα, IL4/IL-13 (Th2) and IFNγ (Th1), were elevated in recovered patients in a severity-dependent manner. Finally, the myelopoiesis cytokines G-CSF and GM-CSF were higher in all patient groups. Increased monocytes and IL-6- and TNFα-producing cells were significantly associated with long COVID-19 symptoms. Conclusions These results reveal that COVID-19 infection influences the frequencies and functions of innate immune cells for up to 3 months after recovery, which may potentially lead to some of the long COVID symptoms.

Anti-Inflammatory Effects and Decreased Formation of Neutrophil Extracellular Traps by Enoxaparin in COVID-19 Patients.

Neutrophil Extracellular Traps (NETs) are a contributing factor of vascular thrombosis and alveolar damage in COVID-19 patients. As enoxaparin is currently used to inhibit vascular thrombosis, this study aimed to investigate whether enoxaparin also reduced inflammation and NETs in COVID-19 patients. Patients with COVID-19 infection were classified into three groups: mild, moderate, and severe (n = 10 for all groups). Plasma was collected from patients and healthy donors (n = 10). Neutrophils isolated from healthy controls were incubated with COVID-19 or healthy plasma, and with or without enoxaparin pretreatment in vitro. Neutrophils and plasma isolated from patients treated with enoxaparin were also investigated. The levels of inflammatory cytokines and NET products such as dsDNA, NE, MPO-DNA and Histone-DNA complexes in plasma and supernatants were measured using immunofluorescence staining and ELISA kits. The expression of inflammatory signaling genes by neutrophils (RELA, SYK, ERK and PKC) was measured using real-time qPCR. The levels of NET products were elevated in the plasma of COVID-19 patients, particularly in the severe group (p < 0.01). Moreover, plasma from the severe group enhanced NET formation (p < 0.01) from neutrophils in vitro. Enoxaparin pretreatment in vitro decreased plasma-induced NETs in a dose-dependent manner and down-regulated the expression of inflammatory genes (p < 0.05). Patients treated with prophylactic enoxaparin showed lower inflammatory cytokine levels and expression of inflammatory genes (p < 0.05). Increased NETs were associated with the severity of COVID-19 infection, particularly in patients with severe pneumonia, and could be used as biomarkers to assess disease severity. Enoxaparin pretreatment inhibited NETs and reduced the expression of inflammatory cytokines, and these effects mostly persisted in patients treated with prophylactic enoxaparin.

Immunogenicity and Risk Factors Associated With Poor Humoral Immune Response of SARS-CoV-2 Vaccines in Recipients of Solid Organ Transplant: A Systematic Review and Meta-Analysis.

Importance: Recipients of solid organ transplant (SOT) experience decreased immunogenicity after COVID-19 vaccination. Objective: To summarize current evidence on vaccine responses and identify risk factors for diminished humoral immune response in recipients of SOT. Data Sources: A literature search was conducted from existence of database through December 15, 2021, using MEDLINE, Embase, Web of Science, Cochrane Library, and ClinicalTrials.gov. Study Selection: Studies reporting humoral immune response of the COVID-19 vaccines in recipients of SOT were reviewed. Data Extraction and Synthesis: Two reviewers independently extracted data from each eligible study. Descriptive statistics and a random-effects model were used. This report was prepared following the Preferred Reporting Items for Systematic Reviews and Meta-analyses (PRISMA) reporting guideline. Data were analyzed from December 2021 to February 2022. Main Outcomes and Measures: The total numbers of positive immune responses and percentage across each vaccine platform were recorded. Pooled odds ratios (pORs) with 95% CIs were used to calculate the pooled effect estimates of risk factors for poor antibody response. Results: A total of 83 studies were included for the systematic review, and 29 studies were included in the meta-analysis, representing 11 713 recipients of SOT. The weighted mean (range) of total positive humoral response for antispike antibodies after receipt of mRNA COVID-19 vaccine was 10.4% (0%-37.9%) for 1 dose, 44.9% (0%-79.1%) for 2 doses, and 63.1% (49.1%-69.1%) for 3 doses. In 2 studies, 50% of recipients of SOT with no or minimal antibody response after 3 doses of mRNA COVID-19 vaccine mounted an antibody response after a fourth dose. Among the factors associated with poor antibody response were older age (mean [SE] age difference between responders and nonresponders, 3.94 [1.1] years), deceased donor status (pOR, 0.66 [95% CI, 0.53-0.83]; I2 = 0%), antimetabolite use (pOR, 0.21 [95% CI, 0.14-0.29]; I2 = 70%), recent rituximab exposure (pOR, 0.21 [95% CI, 0.07-0.61]; I2 = 0%), and recent antithymocyte globulin exposure (pOR, 0.32 [95% CI, 0.15-0.71]; I2 = 0%). Conclusions and Relevance: In this systematic review and meta-analysis, the rates of positive antibody response in solid organ transplant recipients remained low despite multiple doses of mRNA vaccines. These findings suggest that more efforts are needed to modulate the risk factors associated with reduced humoral responses and to study monoclonal antibody prophylaxis among recipients of SOT who are at high risk of diminished humoral response.

Neutrophil Extracellular Traps in Severe SARS-CoV-2 Infection: A Possible Impact of LPS and (1→3)-ß-D-glucan in Blood from Gut Translocation.

Due to limited data on the link between gut barrier defects (leaky gut) and neutrophil extracellular traps (NETs) in coronavirus disease 2019 (COVID-19), blood samples of COVID-19 cases-mild (upper respiratory tract symptoms without pneumonia; n = 27), moderate (pneumonia without hypoxia; n = 28), and severe (pneumonia with hypoxia; n = 20)-versus healthy control (n = 15) were evaluated, together with in vitro experiments. Accordingly, neutrophil counts, serum cytokines (IL-6 and IL-8), lipopolysaccharide (LPS), bacteria-free DNA, and NETs parameters (fluorescent-stained nuclear morphology, dsDNA, neutrophil elastase, histone-DNA complex, and myeloperoxidase-DNA complex) were found to differentiate COVID-19 severity, whereas serum (1→3)-ß-D-glucan (BG) was different between the control and COVID-19 cases. Despite non-detectable bacteria-free DNA in the blood of healthy volunteers, using blood bacteriome analysis, proteobacterial DNA was similarly predominant in both control and COVID-19 cases (all severities). In parallel, only COVID-19 samples from moderate and severe cases, but not mild cases, were activated in vitro NETs, as determined by supernatant dsDNA, Peptidyl Arginine Deiminase 4, and nuclear morphology. With neutrophil experiments, LPS plus BG (LPS + BG) more prominently induced NETs, cytokines, NFκB, and reactive oxygen species, when compared with the activation by each molecule alone. In conclusion, pathogen molecules (LPS and BG) from gut translocation along with neutrophilia and cytokinemia in COVID-19-activated, NETs-induced hyperinflammation.

Host genetic factors of COVID-19 susceptibility and disease severity in a Thai population.

Host genetic factors have been shown to play a role in SARs-CoV-2 infection in diverse populations. However, the genetic landscape differs among various ethnicities; therefore, we explored the host genetic factors associated with COVID-19 disease susceptivity and disease severity in a Thai population. We recruited and genotyped 212 unrelated COVID-19 Thai patients and 36 controls using AxiomTM Human Genotyping SARs-COV-2 array, including 847,384 single nucleotide polymorphisms related to SARs-COV-2 pathogenesis, immune response, and related comorbidity No SNPs passed the genome-wide significance threshold of p value <1 × 10-8. However, with a threshold of p value <1 × 10-5, a locus on chromosome 5q32 was found to have a suggestive association with COVID-19 disease susceptibility (p value 6.9 × 10-6; Q-Q plot λ = 0.805, odds ratio 0.02). Notably, IL17B is a gene located in this linkage disequilibrium block and is previously shown to play a part in inflammation and pneumonia. Additionally, a suggestive locus on chromosome 12q22, harboring EEA1 and LOC643339, was associated with COVID-19 disease severity (p value 1.3 × 10-6 - 4.4 × 10-6, Q-Q plot λ = 0.997, odds ratio 0.28-0.31). EEA1 is involved in viral entry into cells, while LOC643339 is a long non-coding RNA. In summary, our study suggested loci on chromosomes 5q32 and 12q22 to be linked to COVID-19 disease susceptibility and disease severity, respectively. The small sample size of this study may lessen the likelihood that the association found is real, but it could still be true. Further study with a larger cohort is required to confirm these findings.

Short-term immune response after inactivated SARS-CoV-2 (CoronaVac®, Sinovac) and ChAdOx1 nCoV-19 (Vaxzevria®, Oxford-AstraZeneca) vaccinations in health care workers.

BACKGROUND: Inactivated SARS-CoV-2 (CoronaVac®, Sinovac, or SV) and ChAdOx1 nCoV-19 (Vaxzevria®, Oxford-Astra Zeneca, or AZ) vaccines have been administered to the health care workers (HCWs). OBJECTIVE: To determine the short-term immune response after the SV and AZ vaccinations in HCWs. METHODS: In this prospective cohort study, HCWs who completed a 2-dose regimen of the SV or AZ were included. Immune response was evaluated by surrogate viral neutralization test (sVNT) and anti-SARS-CoV-2 total antibody. Blood samples were analyzed at 4 and 12 weeks after the complete vaccination. The primary outcome was the seroconversion rate at 4-weeks after complete immunization. RESULTS: Overall, 185 HCWs with a median (IQR) age of 40.5 (30.3-55.8) years (94 HCWs in the SV group and 91 in the AZ group) were included. At 4 weeks after completing the SV vaccination, 60.6% (95%CI: 50.0-70.6%) had seroconversion evaluated by sVNT (≥ 68% inhibition), comparable to the patients recovered from mild COVID-19 infection (69.0%), with a rapid reduction to 12.2% (95%CI: 6.3-20.8) at 12 weeks. In contrast, 85.7% (95%CI: 76.8-92.2%) HCWs who completed two doses of the AZ for 4 weeks had seroconversion, comparable to the COVID-19 pneumonia patients (92.5%), with a reduction to 39.2% (95%CI: 28.4-50.9%) at 12 weeks. When using the anti-SARS-CoV-2 total antibody level (≥ 132 U/ml) criteria, only 71.3% HCWs in the SV group had seroconversion, compared to 100% in the AZ group at 4 weeks. CONCLUSIONS: A rapid decline of short-term immune response in the HCWs after the SV vaccination indicates the need for a vaccine booster, particularly during the ongoing spreading of the SARS-CoV-2 variants of concern.

A case of successful treatment of severe COVID-19 pneumonia with favipiravir and tocilizumab in post-kidney transplant recipient.

We report a case of COVID-19 in kidney transplant patient in Thailand. A 58-year-old 2 years post-kidney transplant recipient, with maintenance immunosuppression of tacrolimus, mycophenolate mofetil (MMF), and prednisolone, presented with acute diarrhea which followed by fever on day 12. Symptoms of pneumonia together with lymphopenia from complete blood count were developed on day 7 after onset of fever with the x-ray finding of bilateral multifocal patchy infiltration. COVID-19 infection has been confirmed by reverse real-time polymerase chain reaction (PCR) in nasal swab as well as found in stool. Darunavir together with ritonavir, hydroxychloroquine, azithromycin, and favipiravir was initiated on the first day of admission at primary hospital. Patient has been transferred to our hospital on day 2 of admission in which tacrolimus together with MMF was discontinued. High-flow nasal cannula oxygen therapy was required on days 4-5 of hospitalization. Tocilizumab was administered after rising of serum IL-6 level. Symptoms of pneumonia were improved in which no oxygen treatment required from day 10 of hospitalization. Drug interaction between tacrolimus and anti-viral treatment leads to severely high level of tacrolimus which caused reversible acute kidney injury (AKI) after supportive treatment.