Thermal Shock Viral RNA Release in COVID-19 Testing

Introduction/Objective The public health emergency of the COVID-19 pandemic emphasized the crucial role of medical laboratory professionals and scientists in molecular diagnostics laboratories to ensure success in infection control strategies. The demand for laboratory testing using nucleic acid amplification tests to detect SARS-CoV-2 RNA imposed strains in laboratory supplies. Here, we explored an alternative cost-effective solution that will simplify the pre-PCR steps by using a simple heating method to release viral RNA. Methods/Case Report Samples tested using the reference automated extraction method were used:100 samples identified as positive for SARS-CoV-2 RNA and 500 samples tested negative for SARS-CoV-2 RNA were used for the study and sorted with equal distribution according to Ct values of (a) <20, (b) 20–30, and (c) >30.100 ul from swab preserved in Universal Transport Medium was treated with 30 μg of proteinase K, and another set was tested without proteinase K pre-treatment. All samples with or without proteinase K were diluted to minimize PCR inhibitors. The thermal shock protocol was set at (98°C, 5 minutes;4°C, 2 minutes) and screened for purity. Performance and method verification studies were performed. Internal extraction, positive template, and no template controls were markers used for testing quality. The experimental study was performed by qualified testing personnel and all under the same experimental conditions. Results (if a Case Study enter NA) The Ct values from the thermal shock RNA release were compared to the automated extraction method and statistically analyzed.The criteria for acceptability for validation of this new RNA extraction proceeding were set to 100% concordance compared to the commercial kit using an automated extraction. PCR efficiency was at 98% and a slope of -3.3. Within run precision of 2% and limits of detection from 200 to 20,000 copies/uL The method 100% (50/50) concordance on samples previously identified as negative by automated methods and identified 86% (86/100) with a mean difference of 3 Ct. Conclusion Our findings suggest that the thermal shock treatment of nasopharyngeal swabs in viral transport media can successfully extract viral nucleic acid for nucleic acid amplification and is a reasonable alternative for chemical extraction methods when molecular diagnostic laboratories persistently encounter supply chain issues.

Posttraumatic-stress disorder, and others psychological impairment in COVID-19 surviving patients

Background: The post-COVID syndrome generates physical and respiratory complications that can be accompanied by psychological impairments, which can affect long-term mental and physical health. Objective(s): Know the prevalence and severity of PTSD, anxiety, and depression in surviving COVID-19 patients in a follow-up evaluation. Method(s): A cross-sectional descriptive study was conducted. 227 survivors COVID-19 patients participated, were assessed three months following discharge hospital. The following questionnaires were used: The Brief Davidson Trauma Scale, the General Anxiety Disorder Questionnaire, and the Patient Health Questionnaire. A descriptive and statically analysis was performed using the statistical software SPSS version 26. Result(s): The 64.5% of the patients were men, 60.9% required of invasive mechanical ventilation (IMV) during the hospitalization, the average age was about 48.23+/-14.33 years old. The 40% of the patients showed symptoms associated with PTSD, 38.4% anxiety symptoms, 36.6% depression symptoms. There were statistically significant differences between the type of treatment during hospitalization (IMV vs without IMV), in PTSD (t=2.482, df=223, p=.014, XIMV = 5.21, XWIMV = 6.08) and anxiety (t= -2.006, df=223, p=.046, XIMV = 4.05, XWIMV =5.44). Conclusion(s): Survivors of COVID-19 experience a high prevalence of PTSD, anxiety, and depression even three months after discharge from hospital. Patients who did not require IMV during hospitalization experienced a high prevalence and severity of PTSD and anxiety symptoms. Screening for PTSD and other emotional disturbances should be considered in follow-up evaluations in patients discharged from the hospital.

Sleep disturbances in post-Covid patients as a function of sex

Introduction: Within the post-Covid syndrome, sleep disturbances are one of the most persistent symptoms, in which women are more 50% vulnerable than men to insomnia. Objective(s): To compare the prevalence of sleep disorders in post-Covid patients according to sex. Method(s): A cross-sectional observational study was carried out. The participants were 264 post-Covid patients evaluated 3 months after hospital discharge, divided into men (n=156, G1) and women (n=108, G2). A general sleep habits survey and the Berlin SAOS questionnaire were used for sleep assessment. Statistical analyses were performed using SPSS v25. Result(s): Mean age was G1: 54.16 +/-11.751 and G2: 54.23 +/- 13.319. There were differences (p<0.05) between both sexes in the following disorders (G1 vs G2): snoring (73.1% vs 58.3%), primary snoring (45.6% vs 28.6%), unrefreshing sleep more than 3 times a week (28.2% vs 43.5%), tiredness or fatigue during the day at least 3 times a week (30.1% vs 51.9%), sudden movements in extremities: (37.2% vs 22.2%), onset insomnia: (34.0% vs 53.7%), intermediate insomnia (36.5% vs 58.3%), final insomnia: (39.7% vs 55.6%). No differences (p>0.05) were found in pauses in breathing, teeth grinding, feeling of paralysis and feeling of choking, with a prevalence greater than 20% in all cases. Conclusion(s): In our study, the affectation of most sleep disorders are more frequent in the female sex, with the predominance of different types of insomnia (onset, intermediate and final);which could be related to affective disorders. In men, the most prevalent disorders were snoring and sudden movements, mostly linked to respiratory disorders.

Digital Morphology Analyzer in Hematology Laboratory Training

Introduction: The COVID-19 pandemic led educators to adopt non-traditional methods for delivery of education materials. In hematology laboratory, training in morphologic assessment of blood smears is an important skill that entry-level staff needs to show competency CellaVision DM 96 and its proficiency testing features have benefits that extend far beyond its use in clinical practice. For hematology laboratory training programs, the use of Cellavision Proficiency Software can be used in training laboratory staff and medical laboratory program students, assess competency and to ensure standardization in result reporting when evaluating peripheral blood smear reviews CellaVision Proficiency Software can be used as a simulation-based learning tool in pathology and laboratory science education. Method(s): Pilot, crosssectional study. Students matriculated at Rose State College's Medical Laboratory Technician Program in the Fall of 2021 enrolled in Hematology. The laboratory component of Hematology trains the students in the proper evaluation of peripheral blood smears The RSC MLT Class of 2022 cohort was introduced to the traditional microscopic evaluation of peripheral blood smears in the laboratory component of RSC MLT's Introduction to Medical Laboratory class and digital cell morphology assessment is the method used to train students in peripheral blood smear evaluations in the Hematology class. Laboratory component of Hematology follows the educational methodology that includes: Instructive, Self-study, Formative assessment , and Summative assessment Effectiveness in learning was measured by comparing the assessment scores of students who were taught peripheral blood smear evaluations using traditional microscopy. A comparison of student surveys was used to measure the effectiveness of student learning. Result(s): The use of the digital cell morphology software in the Medical Laboratory Technician Program has resulted in: Increased student participation. Improved student learning because of real-time feedback to students. Improved performance in morphological assessment and peripheral blood smear evaluations. 100% student satisfaction with the course. 100% of respondents stated the use of digital microscopy increased their knowledge of subject matter. Conclusion(s): In the delivery of content for hematology laboratory training, the use of Cellavision Proficiency Software improved morphology skill set in evaluation of peripheral blood smears. The digital morphology software provides a system that caters to different learner skills because of its asynchronous delivery and complements the didactic portion of the class to fully understand the materials discussed The use of digital morphology software to train entrylevel medical laboratory staff is an excellent option to supplement traditional microscopy and leads to improved competency, improved technical knowledge and improved learner participation.

GLUCOCORTICOIDS, RITUXIMAB and the PRESENCE of INTERSTITIAL LUNG DISEASE ARE ASSOCIATED with POOR OUTCOMES of the SARS-COV-2 INFECTION in PATIENTS with RHEUMATOID ARTHRITIS: DATA from the NATIONAL REGISTRY SAR-COVID

Background: High disease activity, treatment with glucocorticoids (GC) and rituximab (RTX), have been related to worse outcomes of COVID-19. Objectives: To assess the clinical characteristics and severity of the SARS-CoV-2 infection in patients with rheumatoid arthritis (RA) included in the SAR-COVID registry and to identify factors associated with poor outcomes. Methods: SAR-COVID is a national, longitudinal and observational registry. Patients of ≥18 years old, with diagnosis of RA (ACR-EULAR criteria 2010) who had confrmed SARS-CoV-2 infection (RT-PCR or positive serology) were included between 13-8-20 and 31-7-21. Sociodemographic and clinical data, comorbidities, disease activity and treatment at the moment of the SARS-CoV-2 infection were collected. Additionally, infection symptoms, complications, medical interventions and treatments for COVID-19 were registered. Infection severity was assessed using the WHO-ordinal scale (WHO-OS)1. A cut-off value of ≥5 identifed patients with severe COVID-19 and those who died. Statistical analysis: Descriptive statistics. Chi2 or Fischer test, Student T test or Mann-Whitney and Kruskal Wallis or ANOVA, as appropriate. Multiple logistic regression model. Results: A total of 801 patients were included, with a mean age of 53.1 ± 12.9 years, most of them were female (84.5%) and the median (m) disease duration was 8 years (IQR 4-14). One third were in remission and 46.4% had comor-bidities, being the most frequent, hypertension (26.9 %), dyslipidemia (13.5 %), obesity (13.4 %) and diabetes (8.9%). Moreover, 3.2% had interstitial lung disease (ILD) associated with RA. At SARS-CoV-2 diagnosis, 42.5% were receiving glucocorticoids (GC), 73.9% conventional (c) disease modifying antirheumatic drugs (DMARD), 24% biologic (b) DMARD and 9.1% targeted synthetic (ts) DMARD. Among bDMARD, the most frequently used were TNF inhibitors (17%), followed by abatacept (2.8%), IL-6 inhibitors (2.4%) and rituximab (RTX) (2.1%). During the SARS-CoV-2 infection, 95.8% had symptoms, 27% required hospital-ization, 7.9% presented complications and 4.4% died due to COVID-19. Severe disease and death (WHO-OS≥5) was present in 7.5% of the patients. They were older (62.9±12.5 vs 52.2±12.7, p<0.001), and they had more frequently ILD (18.5% vs 2%, p<0.001), comorbidities (82.5% vs 43.7%, p<0.001), ≥2 comor-bidities (60.3% vs 25.8%, p<0.001), treatment with GC (61% vs 40.7%, p=0.04) and RTX (8.3% vs 1.6%, p=0.007). Conversely, the use of cDMARD and TNF inhibitors was more frequent in patients with WHO-OS<5, nevertheless this difference was not signifcant. Disease activity was comparable between groups. In multivariable analysis, older age, the presence of diabetes, ILD, the use of GC and RTX were signifcantly associated with WHO-OS≥5 (Figure 1). Furthermore, older age (65.7±10.8 vs 52.4±12.8, p<0.001), the presence of comor-bidities (87.9% vs 44.7%, p<0.001), chronic obstructive pulmonary disease (21.9% vs 5.2%, p=0.002), diabetes (30.3% vs 7.9%, p<0.001), hypertension (57.6% vs 25.6%, p<0.001), cardiovascular disease (15.6% vs 3.2%, p=0.005), cancer (9.1% vs 1.3%, p=0.001), ILD (23.3% vs 2.4%, p<0.001) and the use of GC (61.8% vs 41.4%, p=0.02) were associated with mortality. Older age [OR 1.1 IC95% 1.06-1.13] and the use of GC 5-10 mg/day [OR 4.6 IC95% 1.8-11.6] remained signifcantly associated with death due to COVID-19. Conclusion: Treatment with RTX and GC, as well as older age, the presence of diabetes and ILD were associated with poor COVID-19 outcomes in this national cohort of patients with RA. Older patients and those taking GC had a higher mortality rate.

PREVALENCE of LONG COVID in RHEUMATIC DISEASE PATIENTS: ANALYSIS of SAR COVID REGISTRY

Background: Persistent symptoms after acute COVID have been described previously. Main symptoms reported are fatigue, arthralgias, myalgias and mental sickness. Defnition and methods vary widely.1 Objectives: To asses prevalence and related factors to long COVID in a retrospective cohort of patients with rheumatic diseases from Argentina. Methods: A total of 1915 patients were registered from August 18th, 2020 to July 29th, 2021. Patients > 18 years old, with rheumatic disease and confrmed infection by SARS-CoV-2 (antigen or RT-PCR) were included. Those dead, with unknown outcome, wrong date or missing data were excluded. Demographic data, comorbidities, rheumatic disease, and characteristics of SARS-CoV-2 infection were recorded. Long COVID was defned according to NICE guidelines (persistent symptoms for more than 4 weeks, without alternative diagnosis). Long COVID symptoms were defned by rheumatologist. Severity of infection was clas-sifed according to WHO ordinal scale. We used descriptive statistics, univariate model (Student's test, chi square test, ANOVA) and multivariate logistic regression analysis. Results: 230 (12%) had long COVID. Median age was 51 (IQR 40-61]) years, 82% were females, 51% were not caucasian. Median of education was 13.3 years (IQR 12-16), 79 % had private health insurance and 55 % were employed. Nearly half (n=762, 46%) had comorbidities, the most prevalent was hypertension (n=396, 24%). The most frequent rheumatic diseases were rheumatoid arthritis (n=719, 42%) and systemic lupus ery-thematosus (n=280, 16 %). Most were in low activity/remission (79%), used Conventional DMARD (n=773 patients, 45%) and steroids (n=588, 34%) at low dose (n=415, 71%). Main laboratory findings were abnormal D-di-mer (n=94, 28%) and leukopenia (n=93, 26%). Most patients had a WHO ordinal scale < 5 (n=1472, 86%). Median of hospitalization at intensive care unit (ICU) was 8 days [IQR 5, 13]. Treatment for SARS-CoV-2 infection (steroids, anticoagulation, azithromycin, convalescent plasma) was used in 461 (27%) patients. Most of long COVID (n= 152, 69%) reported 1 symptom, the most frequent was fatigue (n= 55, 22%). Figure 1. Univariate analysis is presented in Table 1. In multivariate logistic regression analysis non-caucasian ethnicity OR 1.44 (1.07-1.95), years of education OR 1.05 (1-1.09), treatment with cyclophosphamide OR 11.35 (1.56-112.97), symptoms of COVID-19 OR 13.26 (2.75-242.08), severity scale WHO ≥ 5 OR 2.46 (1.68-3.57), and ICU hospitalization days OR 1.09 (1.05-1.14) were factors associated to long COVID. Conclusion: Prevalence of long COVID was 12%. Non-caucasian ethnicity, higher education, treatment with cyclophosphamide, symptoms of COVID-19, severe disease and ICU hospitalization days were related to long COVID.

Allogeneic and Autologous Hematopoietic Transplants Can be Safely Performed after COVID-19 Infections

We are in the midst of a pandemic with the COVID-19 virus, a pathogen with potential severe manifestations. A major clinical question is whether it is safe to undergo hematopoietic stem cell transplantation (HSCT) shortly after COVID-19 infection. A total of 21 patients received HSCT following a diagnosis of COVID-19 infection at our institution between 7/30/2020 and 4/14/2021. The majority (n=13, 62%) received an allogeneic (ALLO) HSCT from an HLA-matched related (n=5), -matched unrelated (n=6), or haploidentical (n=2) donor. The remaining 8 patients received autologous (AUTO) HSCT. Among ALLO-HSCT recipients, 4 (31%), 5 (38%), 3 (23%), and 1 (8%) patients had grade 1, 2, 3, and 4, manifestations respectively, scored according to the WHO COVID-19 infection severity grading system. Among AUTO-HSCT recipients, 5 (62%), 1 (12%), and 2 (25%) patients had grade 0, 1, and 2 manifestations, respectively. All patients had resolution of COVID-19 symptoms before HSCT. In recipients of ALLO-SCT, the median time from diagnosis of the COVID infection to HSCT was 134 (range: 55-311) days. Median age of recipients was 53 (range: 17-71) years and the majority (69%) of patients were male. Only one patient was <18 years old, and 38% were >60 years. Patients received ALLO-HSCT for treatment of acute myeloid leukemia or myelodysplastic syndrome (n=7, 54%), acute lymphoblastic leukemia (n=2, 15%), chronic lymphoblastic leukemia (n=2, 15%), and Hodgkin's (n=1, 8%) or non-Hodgkin's lymphoma (n=1, 15%). Most (62%) patients were not in remission at the time of HSCT. The median hematopoietic cell transplant-co-morbidity index (HCT-CI) score was 3 (range 0-6);one patient had a history of diabetes and another of hypertension before HSCT. Conditioning regimen was myeloablative in 61%, and stem cell source was peripheral blood (PB) in 92% of transplants. Median time to neutrophils engraftment was 15 (range: 10-20) days. With a median follow-up of 3.5 (range: 0.4-8) months since ALLO-HSCT, two patients died and another two experienced progression of the underlying malignancy. Three patients were diagnosed with grade 2 and none with grade 3 or 4 acute graft-versus-host disease (GvHD). The deaths occurred among patients with COVID-19 infection grade 2 and 3. The primary cause of death was attributed to alveolar hemorrhage/pneumonitis (no organism identified) and acute GvHD, respectively. Overall survival was 89% (95% confidence interval [CI]:43-98) and 76% (95% CI 33-93) at 3 and 6 months, respectively. In recipients of AUTO-HSCT, the median time from diagnosis of the COVID-19 infection to HSCT was 55 (range: 20-157) days. Median age of recipients was 55 (range: 34-75) years, and the majority (62%) of patients were male. One (12%) patient was >60 years. Patients received AUTO-HSCT for treatment of Hodgkin's (n=1, 15%) or non-Hodgkin's (n=4, 50%) lymphoma, or multiple myeloma (n=3, 37%). Six (75%) patients were in remission at the time of HSCT. The median HCT-CI score was 2 (range 0-6). None of the patients had a history of diabetes or hypertension before transplant. Conditioning regimen was myeloablative and stem cell source was PB for all patients. Median time to neutrophils engraftment was 10 (range: 9-13) days. With a median follow-up of 4 (range: 0.8-9) months since AUTO-HSCT, one patient with grade 1 COVID infection died as a result of a candida/cytomegalovirus infection, and none of the patients experienced progression of the underlying malignancy. Overall survival was 100% and 75% (95% CI 13-96) at 3 and 6 months respectively. After HSCT, one ALLO and two AUTO asymptomatic patients had a positive nasal swab COVID-19 PCR assay possibly due to delayed shedding of the virus. None of the 21 patients developed active COVID infections post-transplant. In conclusion, allogeneic and autologous hematopoietic transplantation can be performed in patients after COVID-19 infection. Two of 13 allogeneic and one of 8 autologous recipients experienced non-relapse mortality, none directly related to COVID-19 infection. Patients recovering from COVID-19 in ection should be considered eligible for hematopoietic transplantation as clinically indicated. Disclosures: Shpall: Magenta: Honoraria;Affimed: Patents & Royalties;Novartis: Honoraria;Navan: Consultancy;Magenta: Consultancy;Axio: Consultancy;Adaptimmune: Consultancy;Bayer HealthCare Pharmaceuticals: Honoraria;Novartis: Consultancy;Takeda: Patents & Royalties. Chemaly: Other: Other: Compensation: I am a consultant and advisor on companies who are developing new agents such as Merck, Ansun, and Janssen.

Diagnostic accuracy of COVID-19 antibody tests authorized by FDA Philippines: a systematic review and meta-analysis

Introduction: Coronavirus Disease (COVID-19) is a highly infectious disease caused by Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) which has infected many people all over the world. One of the best ways to lessen its spread is through early detection and diagnosis. Various serological tests are now being used as a surveillance tool in the detection of antibodies as a response to SARS-CoV-2. The aim of this study is to evaluate the diagnostic accuracy and performance of the available COVID-19 antibody tests authorized by the Food and Drug Administration (FDA) Philippines that make use of Enzyme-Linked Immunosorbent Assay (ELISA), Chemiluminescence Immunoassay (CLIA) and Lateral Flow Immunoassay (LFIA). Method: Complete published journal articles relevant to the diagnostic accuracy of the three antibody tests were collected using trusted medical journal search engines. The quality of journals was assessed using QUADAS-2 to determine the risk of bias and assess the applicability judgments of diagnostic accuracy studies. Forest plots were used to summarize the performance of LFIA, ELISA and CLIA according to their specificity and sensitivity in detecting various antibodies. Pooled sensitivity and specificity were also done using bivariate random-effects models with its log-likelihood, a corresponding chi-square test statistic, and area under the summary Receiver-Operating Characteristic curve to see the potential heterogeneity in the data and to assess the diagnostic accuracy of the COVID-19 antibody tests.