A Multiplex PCR Assay for Identifying All Major SARS-CoV-2 Variants

Background Variants of Concern (VOC) of SARS-CoV-2, including the Alpha, Beta, Gamma, and Delta, threaten to prolong the pandemic leading to more global morbidity and mortality. Genome sequencing is the mainstay of tracking the development and evolution of the virus, but is costly, slow, and not easily accessible. Methods A multiplex qRT-PCR assay for SARS-CoV-2 was developed, which identifies all VOC as well as other mutations of interest in the viral genome, eight mutations total, using single nucleotide discriminating molecular beacons in a two-tube assay. The sensitivity and specificity of the assay was tested using in vitro-transcribed targets. Twenty-six SARS-CoV-2 positive patient samples were blinded, then tested using this assay and compared with deep sequencing results. Findings The presented variant molecular beacon assay showed high accuracy when testing in vitro-transcribed targets, down to a limit of detection of five copies of the viral RNA, with 100% specificity. When testing patient samples, the assay was in full agreement with results from deep sequencing with a sensitivity and specificity of 100% (26/26). Using this accurate genotyping, the SARS-CoV-2 samples were classified as the appropriate variants, and of the twenty-six samples two were identified as VOC Alpha, eight as VOC Delta, and two as Epsilon. Interpretation We have developed a qRT-PCR assay for the identification of currently circulating VOC of SARS-CoV-2 as well as other important mutations in its Spike protein coding sequence. This assay can be easily implemented on broadly available five-color thermal cyclers and will help track the spread of these variants.

Overview of Idiopathic Normal Pressure Hydrocephalus: Diagnosis and Improvement

Idiopathic normal pressure hydrocephalus (iNPH) is an uncommon yet potentially reversible cause of dementia in the elderly. It is characterized by cognitive deterioration, urinary incontinence, ventriculomegaly (enlarged cerebral ventricles) and gait apraxia. The diagnosis and management of iNPH requires an organized approach, starting with a comprehensive history and neurologic examination, a review of neuroimaging, and an evaluation of diagnosis. It is important to treat comorbidities in patients prior to specific iNPH testing, which includes testing patient responses to temporary CSF removal and assessing CSF hydrodynamics. In carefully selected patients, all physiological and neurological symptoms improved with a ventriculoperitoneal (VP) shunt surgery, which allows for a gradual adjustment in pressure to prevent complications caused by over-drainage.

Socioeconomic and comorbid factors affecting mortality and length of stay in COVID-19

Background The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic exposed and exacerbated health disparities between socioeconomic groups. Our purpose was to determine which disparities are most prevalent and their impact on length of stay (LoS) and in hospital mortality in patients diagnosed with Covid-19. Methods De-Identified data for patients who tested positive for COVID-19 was abstracted from the HCA enterprise database. Data was binned into summary tables. A negative binomial regression with LoS as the dependent variable and a logistic regression of in-hospital mortality data, using age, insurance status, sex, comorbidities as the dependent variables, were performed. Results From March 1, 2020 to August 23, 2020, of 111,849 covid testing patient records, excluding those with missing data (n=7), without confirmed COVID-19 (n=27,225), and those from a carceral environment (n=1,861), left 84,624 eligible patients. Compared to the US population, the covid cohort had more black patients (23.17% vs 13.4%). Compared to the white cohort, the black cohort had higher private insurance rates (28.52% vs. 23.68%), shorter LoS (IRR=0.97 CI=0.95-0.99, P<0.01) and lower adjusted mortality (OR 0.81, 95% CI 0.75-0.97). Increasing age was associated with increased mortality and LoS. Patients with Medicare or Medicaid had longer LoS (IRR=1.07, 95% CI=1.04-1.09) and higher adjusted mortality rates (OR=1.11, 95% CI=1-1.23) than those with private insurance Conclusion Conclusions We found that when blacks have higher rates of private insurance, they have shorter hospitalizations and lower mortality than whites, when diagnosed with Covid-19. Some other psychiatric and medical conditions also significantly impacted outcomes in patients with Covid-19.

The long road to recovery: at six months since the first COVID-19 wave, elective orthopedic care has still not fully recovered in Belgium

Purpose The primary aim of our study was to investigate elective orthopaedic care during the first wave government-imposed COVID-19 lockdown and at four weeks and 21 weeks after resuming elective care. The secondary aim of our study was to evaluate the implementation of the European Society of Sports Traumatology, Knee Surgery and Arthroscopy (ESSKA) COVID-19 Guidelines and Recommendations for Resuming Elective Surgery in the clinical practice of Belgian knee surgeons. Methods We sent three anonymous online surveys to 102 Belgian Knee Society members (BKS) at times mentioned above. Addressed topics were: (1) participant demographics, (2) elective surgeries, (3) outpatient visits, (4) ESSKA Guidelines, (5) patient and surgeon safety. Results During the COVID-19 lockdown, there was a decrease of 97% in elective knee surgeries and 91% in outpatient visits. At four and 21 weeks after resuming elective care, volumes were respectively 67% and 89% for elective surgeries and 81% and 91% for outpatient visits. Regarding ESSKA guidelines, 91% of surgeons had no COVID-19 testing prior to resuming elective care. Ninety-two per cent reported preoperative (< 72 h) patient PCR testing, and 45% gave preference to young patients without comorbidities. Seventy-two per cent did not use additional personal protective equipment (PPE) if a patient PCR test was negative. Forty-nine per cent continued to give preference to general anaesthesia. Conclusion Our study shows that elective surgeries and outpatient visits were almost completely interrupted during the COVID-19 lockdown and were still below normal at four and 21 weeks after resuming elective care. Regarding ESSKA COVID-19 guidelines, our study observes good compliance in preoperative patient COVID-19 testing, but lower compliance for preoperative health care personnel testing, patient selection, use of PPE, and locoregional anaesthesia. Level of Evidence V

Lessons from South Korea Regarding the Early Stage of the COVID-19 Outbreak

South Korea has experienced difficulty in controlling the spread of the novel coronavirus disease (COVID-19) during the early stages of the outbreak. South Korea remains passionately determined to protect Koreans against COVID-19 and through trial and error hopes to improve the strategies used to limit the outbreak. Here, we review how COVID-19 spread and what prevention strategies were implemented during the early stages of the outbreak in South Korea. We investigated online newspapers published in South Korea from 21 January 2020 to 20 March 2020, and reviewed academic medical articles related to COVID-19. Additionally, we acquired data on COVID-19 cases through the official website for COVID-19 in South Korea. To date, numerous measures have been applied by the government and the medical community during the early stages of the COVID-19 outbreak including the reporting of methods for diagnostic testing, patient classification, the introduction of drive-through screening centers, COVID-19 preventive measures, implementation of government policies for the shortage of face masks, and entry restrictions. Here, we present data from the early stages of the COVID-19 outbreak and measures to prevent its spread in South Korea. We believe that sharing the experience of South Korea during the COVID-19 outbreak can help other countries to implement strategies to prevent its rapid transmission.

Performance characteristics of Fungitell STAT™, a rapid (1→3)-β-D-glucan single patient sample in vitro diagnostic assay

Serum (1→3)-β-D-glucan (BDG), is an adjunct test in the diagnosis of invasive fungal disease (IFD). Fungitell STAT™, a facile, rapid, single patient option, executable for one or more patient specimens in approximately an hour, has been developed to address a need for rapid in-house testing. This method presents qualitative information concerning serum BDG levels, using an index value that allows the rapid categorization of patients as positive, negative, or indeterminate relative to serum BDG titer. The categorical and analytical performance of Fungitell STAT was evaluated. The categorical agreement between methods was established by testing patient samples which had been previously categorized with Fungitell. Receiver Operating Characteristic curves were used to identify cut-offs using 93 de-identified patient specimens. Subsequently, using these cutoffs, an independent group of 488 patient specimens was analyzed. Positive percent agreement (PPA) with, and without, indeterminate results was 74% and 99%, respectively. Negative percent agreement (NPA) was 91% and 98% with, and without, indeterminate results, respectively. Additionally, commercially available normal off-the-clot sera were spiked with Saccharomyces cerevisiae-derived (1→3)-β-D-glucan to produce analytical samples. Analytical reproducibility using spiked samples was excellent with 94% of the CV (coefficient of variation) values ≤10% among three independent laboratories. Good correlation with the predicate method was demonstrated with correlation coefficients of 0.90 or better with patient samples and 0.99 with spiked samples. The Fungitell STAT index assay provides a rapid and suitable method for serum BDG testing.

The Potential Use of Unprocessed Sample for RT-qPCR Detection of COVID-19 without an RNA Extraction Step

ABSTRACTQuantitative reverse transcription polymerase chain reaction (RT-qPCR) assay is the gold standard recommended to test for acute SARS-CoV-2 infection.1–4 It has been used by the Centers for Disease Control and Prevention (CDC) and several other companies in their Emergency Use Authorization (EUA) assays. With many PCR-based molecular assays, an extraction step is routinely used as part of the protocol. This step can take up a significant amount of time and labor, especially if the extraction is performed manually. Long assay time, partly caused by slow sample preparation steps, has created a large backlog when testing patient samples suspected of COVID-19. Using flu and RSV clinical specimens, we have collected evidence that the RT-qPCR assay can be performed directly on patient sample material from a nasal swab immersed in virus transport medium (VTM) without an RNA extraction step. We have also used this approach to test for the direct detection of SARS-CoV-2 reference materials spiked in VTM. Our data, while preliminary, suggest that using a few microliters of these untreated samples still can lead to sensitive test results. If RNA extraction steps can be omitted without significantly affecting clinical sensitivity, the turn-around time of COVID-19 tests and the backlog we currently experience can be reduced drastically. Next, we will confirm our findings using patient samples.

Age-stratified burden of pneumococcal community acquired pneumonia in hospitalised Canadian adults from 2010 to 2015

BackgroundIn Canada, 13-valent pneumococcal conjugate vaccine (PCV13) is recommended in childhood, in individuals at high risk of invasive pneumococcal disease (IPD) and in healthy adults aged ≥65 years for protection against vaccine-type IPD and pneumococcal community-acquired pneumonia (pCAP). Since vaccine recommendations in Canada include both age-based and risk-based guidance, this study aimed to describe the burden of vaccine-preventable pCAP in hospitalised adults by age.MethodsSurveillance for community-acquired pneumonia (CAP) in hospitalised adults was performed prospectively from 2010 to 2015. CAP was radiologically confirmed, and pCAP was identified using blood and sputum culture and urine antigen testing. Patient demographics and outcomes were stratified by age (16–49, 50–64, ≥65 and ≥50 years).ResultsOf 6666/8802 CAP cases tested, 830 (12.5%) had pCAP, and 418 (6.3%) were attributed to a PCV13 serotype. Of PCV13 pCAP, 41% and 74% were in adults aged ≥65 and ≥50 years, respectively. Compared with non-pCAP controls, pCAP cases aged ≥50 years were more likely to be admitted to intensive care units (ICUs) and to require mechanical ventilation. Older adults with pCAP were less likely to be admitted to ICU or required mechanical ventilation, given their higher mortality and goals of care. Of pCAP deaths, 67% and 90% were in the ≥65 and ≥50 age cohorts, respectively.ConclusionsAdults hospitalised with pCAP in the age cohort of 50–64 years contribute significantly to the burden of illness, suggesting that an age-based recommendation for adults aged ≥50 years should be considered in order to optimise the impact of pneumococcal vaccination programmes in Canada.

Comparison of Accuracy Guidelines for Hospital Glucose Meters

When used in hospital settings, glucose meter performance issues involve analytic comparability to lab-based testing, patient and sample variables, and clinical affects such as insulin treatment protocol outcomes and morbidity or outcome risk factors. Different tools are available to assess these issues, including accuracy and precision statistics along with clinical risk measures such as error grids or simulation testing. Regulatory, guidance, and professional bodies have advocated a number of varying recommendations for glucose meter performance in different situations and under different patient conditions. These are summarized and compared, but reconciling these guidelines can be confusing or difficult for providers. Blood glucose meters are useful in the management of patients in acute or assisted care facilities, but users must appreciate the variables that affect measurements and provide for oversight that can manage risk factors and maintain meter performance expectations.