Molecular test for COVID-19 diagnosis based on a colorimetric genomagnetic assay.

The world is in a long pandemic period caused by the SARS-CoV-2 virus and massive diagnostic tests to assist efforts to control the spread of the disease and also to avoid new coronavirus variants are still needed. Herein, we propose a simple and accurate saliva-based colorimetric test for the diagnosis of COVID-19. Magnetic beads (MBs) modified with a sequence of single-strand DNA (ssDNA) complementary to the N gene of the SARS-CoV-2 RNA were developed and used for magnetic capture and separation from a complex saliva sample. A second biotinylated ssDNA sequence was applied, and the colorimetric detection was carried out by adding streptavidin-horseradish peroxidase conjugate, H2O2, and tetramethylbenzidine (TMB) as chromogenic substrate. The test does not require viral RNA isolation, transcription, or amplification steps and can be performed at room temperature. The molecular assay test can be run using 96-well microplates, allowing the diagnosis of a large number of samples in 90 min. A simple support for magnets was designed and constructed using a 3D printer that allows the magnetic separations directly in the 96-well microplate. The colorimetric test showed an excellent ability to discriminate between healthy individuals and patients infected with SARS-CoV-2, with 92% and 100% of clinical sensitivity and specificity, respectively. This performance was similar to that achieved using the gold standard RT-PCR technique. The proposed genomagnetic assay offers an opportunity to greatly increase population testing, contribute to controlling the spread of the virus, and improve health equity in testing for COVID-19.

Molecular test for COVID-19 diagnosis based on a colorimetric genomagnetic assay

The world is in a long pandemic period caused by the SARS-CoV-2 virus and massive diagnostic tests to assist efforts to control the spread of the disease and also to avoid new coronavirus variants are still needed. Herein, we propose a simple and accurate saliva-based colorimetric test for the diagnosis of COVID-19. Magnetic beads (MBs) modified with a sequence of single-strand DNA (ssDNA) complementary to the N gene of the SARS-CoV-2 RNA were developed and used for magnetic capture and separation from a complex saliva sample. A second biotinylated ssDNA sequence was applied, and the colorimetric detection was carried out by adding streptavidin-horseradish peroxidase conjugate, H2O2, and tetramethylbenzidine (TMB) as chromogenic substrate. The test does not require viral RNA isolation, transcription, or amplification steps and can be performed at room temperature. The molecular assay test can be run using 96-well microplates, allowing the diagnosis of a large number of samples in 90 min. A simple support for magnets was designed and constructed using a 3D printer that allows the magnetic separations directly in the 96-well microplate. The colorimetric test showed an excellent ability to discriminate between healthy individuals and patients infected with SARS-CoV-2, with 92% and 100% of clinical sensitivity and specificity, respectively. This performance was similar to that achieved using the gold standard RT-PCR technique. The proposed genomagnetic assay offers an opportunity to greatly increase population testing, contribute to controlling the spread of the virus, and improve health equity in testing for COVID-19. Graphical abstract Image 1

Probabilistic survival modeling in health research: an assessment using cohort data from hospitalized patients with COVID-19 in a Latin American city.

Probabilistic survival methods have been used in health research to analyze risk factors and adverse health outcomes associated with COVID-19. The aim of this study was to employ a probabilistic model selected among three distributions (exponential, Weibull, and lognormal) to investigate the time from hospitalization to death and determine the mortality risks among hospitalized patients with COVID-19. A retrospective cohort study was conducted for patients hospitalized due to COVID-19 within 30 days in Londrina, Brazil, between January 2021 and February 2022, registered in the database for severe acute respiratory infections (SIVEP-Gripe). Graphical and Akaike Information Criterion (AIC) methods were used to compare the efficiency of the three probabilistic models. The results from the final model were presented as hazard and event time ratios. Our study comprised of 7,684 individuals, with an overall case fatality rate of 32.78%. Data suggested that older age, male sex, severe comorbidity score, intensive care unit admission, and invasive ventilation significantly increased risks for in-hospital mortality. Our study highlights the conditions that confer higher risks for adverse clinical outcomes attributed to COVID-19. The step-by-step process for selecting appropriate probabilistic models may be extended to other investigations in health research to provide more reliable evidence on this topic.

COVID-19 vaccination and case fatality rates: a case report in a Brazilian municipality.

Vaccination campaigns played a crucial role in reducing the incidence of COVID-19. However, a scant number of studies evaluated the impact of vaccination on case fatality rates (CFRs), including in Brazil. Our study aimed to compare CFRs according to vaccination status among subjects living in Arapongas (Paraná State, Brazil), considering the age composition of the population. Several strategies adopted by the Arapongas City Hall to minimize the spread of the virus were also elaborated upon. We accessed the 2021 database of the Arapongas Municipal Health Department, in which a total of 16,437 confirmed cases and 425 deaths were reported. The CFR was calculated as the ratio between COVID-19 deaths and the number of confirmed cases. Differences in age composition between unvaccinated and fully vaccinated individuals were observed in our study. Considering that CFR is a crude indicator and is highly sensitive to the age composition of the population, we adopted the average age distribution of confirmed cases among the three vaccination statuses (unvaccinated, partially, and fully) as a standard age distribution. The age-standardized CFR for unvaccinated and fully vaccinated groups were 4.55% and 2.42%, respectively. Fully vaccinated individuals showed lower age-specific CFRs in all age groups above 60 years than unvaccinated populations. Our findings strengthen the role of vaccination as a critical measure for preventing deaths among infected people and is particularly important to the ongoing reassessment of public health interventions and policies.

Vitamin D deficiency predicts 30-day hospital mortality of adults with COVID-19.

BACKGROUND & AIMS: Several studies have shown conflicting results for the relationship between vitamin D deficiency and COVID-19 outcomes. Here, we aimed to evaluate whether plasma 25(OH)D levels predict mortality in adults admitted with COVID-19, considering potential confounders. METHODS: We conducted a retrospective cohort study that included 115 adults (age 62.1 ± 17.6 years, 65 males) admitted to a Brazilian public hospital for severely symptomatic COVID-19. Subjects were classified into two groups according to their plasma levels of 25(OH)D: sufficiency (≥50 nmol/L) and the deficiency (<50 nmol/L). The diagnosis of COVID-19 was performed using real-time polymerase chain reaction (qPCR). In addition, direct competitive chemiluminescence immunoassay assessed serum 25(OH)D levels. RESULTS: The all-cause 30-day mortality was 13.8% (95% CI: 6.5%-21%) in the group of patients with sufficient plasma 25(OH)D levels and 32.1% (95% CI: 14.8%-49.4%) among those with deficient plasma 25(OH)D levels. Cox regression showed that plasma 25(OH)D levels remained a significant predictor of mortality even after adjusting for the covariates sex, age, length of the delay between symptom onset and hospitalization, and disease severity (HR = 0.98, 95% CI: 0.96-1.00; p = 0.02). CONCLUSION: Vitamin D deficiency predicts higher mortality risk in adults with COVID-19.

The impact of COVID-19 vaccination on case fatality rates in a city in Southern Brazil.

BACKGROUND: Recent studies have established that vaccination plays a significant role in reducing COVID-19-related deaths. Here, we investigated differences in COVID-19 case fatality rates (CFRs) among vaccinated and unvaccinated populations, and analyzed whether the age composition of confirmed cases has a significant effect on the variations in the observed CFRs across these groups. METHODS: The study considered 59,853 confirmed cases and 1,687 deaths from COVID-19, reported between January 1 to October 20, 2021, by the Health Department of Londrina, a city in Southern Brazil. We used Negative Binomial regression models to estimate CFRs according to vaccination status and age range. RESULTS: There are significant differences between the CFR for fully vaccinated and unvaccinated populations (IRR = 0.596, 95% CI [0.460 - 0.772], P < .001). Vaccinated populations experience fatality rates 40.4% lower than non-vaccinated. In addition, the age composition of confirmed cases explains more than two-thirds of the variation in the CFR between these 2 groups. CONCLUSIONS: Our novel findings reinforce the importance of vaccination as an essential public health measure for reducing COVID-19 fatality rates in all age groups. The results also provide means for accurately assessing differences in CFRs across vaccinated and unvaccinated populations. Such assessment is essential to inform and determine appropriate containment and mitigation interventions in Brazil and elsewhere.

COVID-19 diagnosis by SARS-CoV-2 Spike protein detection in saliva using an ultrasensitive magneto-assay based on disposable electrochemical sensor.

The outbreak of the COVID-19 pandemic, caused by Severe Acute Respiratory Syndrome of Coronavirus 2 (SARS-CoV-2), has fueled the search for diagnostic tests aiming at the control and reduction of the viral transmission. The main technique used for diagnosing the Coronavirus disease (COVID-19) is the reverse transcription-polymerase chain reaction (RT-PCR) technique. However, considering the high number of cases and the underlying limitations of the RT-PCR technique, especially with regard to accessibility and cost of the test, one does not need to overemphasize the need to develop new and less expensive testing techniques that can aid the early diagnosis of the disease. With that in mind, we developed an ultrasensitive magneto-assay using magnetic beads and gold nanoparticles conjugated to human angiotensin-converting enzyme 2 (ACE2) peptide (Gln24-Gln42) for the capturing and detection of SARS-CoV-2 Spike protein in human saliva. The technique applied involved the use of a disposable electrochemical device containing eight screen-printed carbon electrodes which allow the simultaneous analysis of eight samples. The magneto-assay exhibited an ultralow limit of detection of 0.35 ag mL-1 for the detection of SARS-CoV-2 Spike protein in saliva. The magneto-assay was tested in saliva samples from healthy and SARS-CoV-2-infected individuals. In terms of efficiency, the proposed technique - which presented a sensitivity of 100.0% and specificity of 93.7% for SARS-CoV-2 Spike protein-exhibited great similarity with the RT-PCR technique. The results obtained point to the application potential of this simple, low-cost magneto-assay for saliva-based point-of-care COVID-19 diagnosis.

Phenylalanine and COVID-19: Tracking disease severity markers.

BACKGROUND: Although there are several severity predictors for COVID-19, none are specific. Serum levels of phenylalanine were recently associated with increased inflammation, higher SOFA scores, ICU admission, and mortality rates among non-COVID-19 patients. Here, we investigated the relationship between phenylalanine and inflammatory markers in adults with COVID-19. METHODS: We assessed adults with COVID-19 at hospital admission for clinical and laboratory data. Nuclear magnetic resonance spectroscopy measured serum levels of phenylalanine and other amino acids of its metabolomic pathway. Flow Cytometry measured serum levels of IL-2, IL-4, IL-6, Il-10, TNF-α, and IFN-γ. Linear regression models adjusted for potential confounders assessed the relationship between serum levels of phenylalanine and inflammatory cytokines. RESULTS: Phenylalanine and tyrosine were significantly lower in mild disease as compared to moderate and severe groups. Linear regression models showed that phenylalanine is independently and positively associated with disease severity regardless of the cytokine analyzed and after adjustment for potential confounders. In addition, mild cases showed consistently lower serum phenylalanine levels within the first ten days from disease onset to hospital admission. CONCLUSIONS: Phenylalanine is a marker of disease severity. This association is independent of the time between the onset of symptoms and the magnitude of the inflammatory state.

Meteorological parameters and cases of COVID-19 in Brazilian cities: an observational study.

Meteorological parameters modulate transmission of the SARS-Cov-2 virus, the causative agent related to coronavirus disease-2019 (COVID-19) development. However, findings across the globe have been inconsistent attributed to several confounding factors. The aim of the present study was to investigate the relationship between reported meteorological parameters from July 1 to October 31, 2020, and the number of confirmed COVID-19 cases in 4 Brazilian cities: São Paulo, the largest city with the highest number of cases in Brazil, and the cities with greater number of cases in the state of Parana during the study period (Curitiba, Londrina and Maringa). The assessment of meteorological factors with confirmed COVID-19 cases included atmospheric pressure, temperature, relative humidity, wind speed, solar irradiation, sunlight, dew point temperature, and total precipitation. The 7- and 15-day moving averages of confirmed COVID-19 cases were obtained for each city. Pearson's correlation coefficients showed significant correlations between COVID-19 cases and all meteorological parameters, except for total precipitation, with the strongest correlation with maximum wind speed (0.717, <0.001) in São Paulo. Regression tree analysis demonstrated that the largest number of confirmed COVID-19 cases was associated with wind speed (between ≥0.3381 and <1.173 m/s), atmospheric pressure (<930.5mb), and solar radiation (<17.98e+3). Lower number of cases was observed for wind speed <0.3381 m/s and temperature <23.86°C. Our results encourage the use of meteorological information as a critical component in future risk assessment models.

Elevations in Liver Transaminases in COVID-19: (How) Are They Related?

Liver involvement in COVID-19 is not yet well-understood, but elevations in liver transaminases have been described to occur in 14-53% of the cases and are more frequently seen in severe disease. This cross-sectional study explored the relationship between the elevations in liver transaminases and inflammatory parameters in 209 adults with COVID-19. Demographic and clinical data, serum levels of inflammatory cytokines and liver aminotransferases were analyzed. Three groups were formed according to the liver transaminase abnormalities: (I) Normal transaminases, (II) Borderline transaminases elevation, and (III) Mild to severe transaminases elevation. Altered liver transaminases were directly related to disease severity, showing association with the NEWS2 score at admission and greater need for ICU or death. Moreover, higher levels of IL-2 and CRP were associated with borderline transaminases elevations, whereas higher levels of IL-10 and Neutrophil to Lymphocyte ratio were associated with mild to severe transaminases elevation. These results reinforce the importance of liver transaminases in patients with COVID-19 as a complementary marker for disease severity and also point to them as a parameter reflecting the continuous dynamics between viral infection and the immune response.

Comorbidities predict 30-day hospital mortality of older adults with COVID-19.

We evaluated whether comorbidities predict disease severity and mortality in a cohort of 147 older adults with COVID-19. Patients were divided into three groups according to the Charlson Comorbidity Index (CCI) score. Groups 2 (CCI 4 - 5) and 3 (CCI ≥ 6) had higher 30-day mortality rate as compared to group 1 (CCI ≤ 3). Cox regression showed that even after adding sex, National Early Warning Score (NEWS) 2 score and the need for intensive care unit admission to the model, no significant changes were found in the mortality risk predicted by the CCI score, showing that chronic pathologies are key determinants of short-term survival in COVID-19. This work is important for the geriatric nursing field as it demonstrates that alternative approaches for clinical decision-making that consider the comorbidities, rather than only chronological age, can be especially significant for the management of COVID-19 patients' hospitalization.

IL-6 and IL-10 are associated with disease severity and higher comorbidity in adults with COVID-19.

AIM: COVID-19 pandemic has caused extensive burden on public life and health care worldwide. This study aimed to assess circulating levels of inflammatory cytokines in adult patients who were hospitalized with COVID-19 and stratified according to age (older or younger than 65 years) aiming to explore associations between these markers of inflammation and comorbidities. METHODS: This was a cross-sectional study of 142 COVID-19 patients consecutively admitted to the University Hospital of the Federal University of São Carlos, from July to October 2020. Sociodemographic data, chronic comorbidities, and baseline NEWS2 and SOFA for clinical deterioration were obtained at hospital admission. Serum levels of inflammatory cytokines were determined by flow cytometry. RESULTS: Older adults with COVID-19 had higher serum levels of IL-6 and IL-10 as compared to those under 65 years of age (p < 0.001 and p = 0.003, respectively). IL-10 was independently associated with age (p = 0.04) and severity of the disease (p = 0.05), whereas serum levels of IL-6 were not directly associated with age (p = 0.5). The comorbidity index seems to be the main responsible for this, being significantly associated with IL-6 levels among those aged 65 and over (p = 0.007), in addition to the severity of the disease. CONCLUSIONS: Higher serum levels of IL-6 and IL-10 are associated with the severity of the disease and a higher comorbidity index among adults aged 65 and over with COVID-19. This should raise awareness of the importance of comorbidity index, rather than age, during risk stratification.

Use of ivermectin in the treatment of Covid-19: A pilot trial.

OBJECTIVES: In this randomized open-label trial pilot study we assessed the antiviral effects and safety of various doses of ivermectin in patients with mild clinical symptoms of COVID-19. METHODS: Patients were randomly assigned to receive standard of care (SOC) treatment at hospital admission; SOC plus ivermectin 100 mcg/kg; SOC plus ivermectin 200 mcg/kg; or SOC plus ivermectin 400 mcg/kg. The primary assessed endpoint was the proportion of patients who achieved two consecutive negative SARS-CoV-2 RT PCR tests within 7 days of the start of the dosing period. This study was registered at ClinicalTrials.gov (NCT04431466). RESULTS: A total of 32 patients were enrolled and randomized to treatment. SOC treatment together with ivermectin did not result in any serious adverse events. All patients exhibited a reduction in SARS-CoV-2 viral load within 7 days; however, those who received ivermectin had a more consistent decrease as compared to the SOC alone group, characterized by a shorter time for obtaining two consecutive negative SARS-CoV-2 RT PCR tests. CONCLUSIONS: Ivermectin is safe in patients with SARS-CoV-2, reducing symptomatology and the SARS-CoV-2 viral load. This antiviral effect appears to depend on the dose used, and if confirmed in future studies, it suggests that ivermectin may be a useful adjuvant to the SOC treatment in patients with mild COVID-19 symptoms.

Burden and psychological symptoms on informal caregivers of the elderly in the COVID-19 pandemic.

OBJECTIVE: To evaluate the burden and psychological symptoms of informal caregivers of the elderly during the COVID-19 pandemic. METHOD: Cross-sectional study with 50 caregivers from the Gerontology Outpatient Clinic in the interior of São Paulo, evaluated in 2021 via teleconsultation by the Hospital Anxiety and Depression Scale and the Zarit-Brief Burden Interview. Poisson's multivariate regression was applied to the two instruments scores according to the set of characteristics of the participants. RESULTS: Psychological symptoms were highlighted in caregivers with longer time in exercise (p=0.01; p=0.001) and who lived in the same environment with the elderly (p=0.04; p=0.02). Burden was associated with age (p<0.001) and living with the elderly (p=0.001). CONCLUSION: There is a need for interventions that attenuate psychological symptoms and burden in older caregivers, who live with the elderly and have been working for a longer time.