scholarly journals Sensitivity and specificity of metatranscriptomics as an arbovirus surveillance tool

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Jana Batovska ◽  
Peter T. Mee ◽  
Stacey E. Lynch ◽  
Tim I. Sawbridge ◽  
Brendan C. Rodoni
Keyword(s):  
Viral Load ◽  
Sensitivity And Specificity ◽  
High Specificity ◽  
Digital Pcr ◽  
Infected Mosquito ◽  
Clear Understanding ◽  
Acid Extraction ◽  
Analytical Sensitivity ◽  
Illumina Hiseq ◽  
Arbovirus Surveillance

AbstractThe ability to identify all the viruses within a sample makes metatranscriptomic sequencing an attractive tool to screen mosquitoes for arboviruses. Practical application of this technique, however, requires a clear understanding of its analytical sensitivity and specificity. To assess this, five dilutions (1:1, 1:20, 1:400, 1:8,000 and 1:160,000) of Ross River virus (RRV) and Umatilla virus (UMAV) isolates were spiked into subsamples of a pool of 100 Culex australicus mosquitoes. The 1:1 dilution represented the viral load of one RRV-infected mosquito in a pool of 100 mosquitoes. The subsamples underwent nucleic acid extraction, mosquito-specific ribosomal RNA depletion, and Illumina HiSeq sequencing. The viral load of the subsamples was also measured using reverse transcription droplet digital PCR (RT-ddPCR) and quantitative PCR (RT-qPCR). Metatranscriptomic sequencing detected both RRV and UMAV in the 1:1, 1:20 and 1:400 subsamples. A high specificity was achieved, with 100% of RRV and 99.6% of UMAV assembled contigs correctly identified. Metatranscriptomic sequencing was not as sensitive as RT-qPCR or RT-ddPCR; however, it recovered whole genome information and detected 19 other viruses, including four first detections for Australia. These findings will assist arbovirus surveillance programs in utilising metatranscriptomics in routine surveillance activities to enhance arbovirus detection.

scholarly journals Modelling RT-qPCR cycle-threshold using digital PCR data for implementing SARS-CoV-2 viral load studies

PLoS ONE ◽  
2021 ◽  
Vol 16 (12) ◽  
pp. e0260884
Author(s):  
Fabio Gentilini ◽  
Maria Elena Turba ◽  
Francesca Taddei ◽  
Tommaso Gritti ◽  
Michela Fantini ◽  
...  
Keyword(s):  
Viral Load ◽  
Regression Model ◽  
Case Fatality ◽  
High Specificity ◽  
Digital Pcr ◽  
Public Health Authority ◽  
Load Factor ◽  
Threshold Values ◽  
Cycle Threshold ◽  
Low Sensitivity

Objectives To exploit the features of digital PCR for implementing SARS-CoV-2 observational studies by reliably including the viral load factor expressed as copies/μL. Methods A small cohort of 51 Covid-19 positive samples was assessed by both RT-qPCR and digital PCR assays. A linear regression model was built using a training subset, and its accuracy was assessed in the remaining evaluation subset. The model was then used to convert the stored cycle threshold values of a large dataset of 6208 diagnostic samples into copies/μL of SARS-CoV-2. The calculated viral load was used for a single cohort retrospective study. Finally, the cohort was randomly divided into a training set (n = 3095) and an evaluation set (n = 3113) to establish a logistic regression model for predicting case-fatality and to assess its accuracy. Results The model for converting the Ct values into copies/μL was suitably accurate. The calculated viral load over time in the cohort of Covid-19 positive samples showed very low viral loads during the summer inter-epidemic waves in Italy. The calculated viral load along with gender and age allowed building a predictive model of case-fatality probability which showed high specificity (99.0%) and low sensitivity (21.7%) at the optimal threshold which varied by modifying the threshold (i.e. 75% sensitivity and 83.7% specificity). Alternative models including categorised cVL or raw cycle thresholds obtained by the same diagnostic method also gave the same performance. Conclusion The modelling of the cycle threshold values using digital PCR had the potential of fostering studies addressing issues regarding Sars-CoV-2; furthermore, it may allow setting up predictive tools capable of early identifying those patients at high risk of case-fatality already at diagnosis, irrespective of the diagnostic RT-qPCR platform in use. Depending upon the epidemiological situation, public health authority policies/aims, the resources available and the thresholds used, adequate sensitivity could be achieved with acceptable low specificity.

scholarly journals Viral Dynamics and Real-Time RT-PCR Ct Values Correlation with Disease Severity in COVID-19

Diagnostics ◽  
2021 ◽  
Vol 11 (6) ◽  
pp. 1091
Author(s):  
Ali A. Rabaan ◽  
Raghavendra Tirupathi ◽  
Anupam A Sule ◽  
Jehad Aldali ◽  
Abbas Al Mutair ◽  
...  
Keyword(s):  
Viral Load ◽  
Real Time ◽  
Disease Severity ◽  
False Negative ◽  
Influential Factors ◽  
Confirmatory Test ◽  
Acid Extraction ◽  
Rt Pcr ◽  
Viral Kinetics ◽  
Ct Value

Real-time RT-PCR is considered the gold standard confirmatory test for coronavirus disease 2019 (COVID-19). However, many scientists disagree, and it is essential to understand that several factors and variables can cause a false-negative test. In this context, cycle threshold (Ct) values are being utilized to diagnose or predict SARS-CoV-2 infection. This practice has a significant clinical utility as Ct values can be correlated with the viral load. In addition, Ct values have a strong correlation with multiple haematological and biochemical markers. However, it is essential to consider that Ct values might be affected by pre-analytic, analytic, and post-analytical variables such as collection technique, specimen type, sampling time, viral kinetics, transport and storage conditions, nucleic acid extraction, viral RNA load, primer designing, real-time PCR efficiency, and Ct value determination method. Therefore, understanding the interpretation of Ct values and other influential factors could play a crucial role in interpreting viral load and disease severity. In several clinical studies consisting of small or large sample sizes, several discrepancies exist regarding a significant positive correlation between the Ct value and disease severity in COVID-19. In this context, a revised review of the literature has been conducted to fill the knowledge gaps regarding the correlations between Ct values and severity/fatality rates of patients with COVID-19. Various databases such as PubMed, Science Direct, Medline, Scopus, and Google Scholar were searched up to April 2021 by using keywords including “RT-PCR or viral load”, “SARS-CoV-2 and RT-PCR”, “Ct value and viral load”, “Ct value or COVID-19”. Research articles were extracted and selected independently by the authors and included in the present review based on their relevance to the study. The current narrative review explores the correlation of Ct values with mortality, disease progression, severity, and infectivity. We also discuss the factors that can affect these values, such as collection technique, type of swab, sampling method, etc.

Prediction of encephalopathy in perinatal asphyxia score: reaching the unreached

2021 ◽  
Vol 0 (0) ◽  
Author(s):  
Srinivasa Murthy Doreswamy ◽  
Amulya Ramakrishnegowda
Keyword(s):  
Therapeutic Hypothermia ◽  
Sensitivity And Specificity ◽  
Perinatal Asphyxia ◽  
High Specificity ◽  
Test Sample ◽  
Therapeutic Benefit ◽  
Primary Outcome Measure ◽  
Primary Objective ◽  
Post Test ◽  
Test Probability

Abstract Objectives Neonates who develop moderate to severe encephalopathy following perinatal asphyxia will benefit from therapeutic hypothermia. Current National Institute of Child Health and Human Development (NICHD) criteria for identifying encephalopathic neonates needing therapeutic hypothermia has high specificity. This results in correctly identifying neonates who have already developed moderate to severe encephalopathy but miss out many potential beneficiaries who progress to develop moderate to severe encephalopathy later. The need is therefore not just to diagnose encephalopathy, but to predict development of encephalopathy and extend the therapeutic benefit for all eligible neonates. The primary objective of the study was to develop and validate the statistical model for prediction of moderate to severe encephalopathy following perinatal asphyxia and compare with current NICHD criteria. Methods The study was designed as prospective observational study. It was carried out in a single center Level 3 perinatal unit in India. Neonates>35 weeks of gestation and requiring resuscitation at birth were included. Levels of resuscitation and blood gas lactate were used to determine the pre-test probability, Thompson score between 3 and 5 h of life was used to determine post-test probability of developing encephalopathy. Primary outcome measure: Validation of Prediction of Encephalopathy in Perinatal Asphyxia (PEPA) score by Holdout method. Results A total of 55 babies were included in the study. The PEPA score was validated by Holdout method where the fitted receiver-operating characteristic (ROC) area for the training and test sample were comparable (p=0.758). The sensitivity and specificity of various PEPA scores for prediction of encephalopathy ranged between 74 and 100% in contrast to NICHD criteria which was 42%. PEPA score of 30 had a best combination of sensitivity and specificity of 95 and 89% respectively. Conclusions PEPA score has a higher sensitivity than NICHD criteria for prediction of Encephalopathy in asphyxiated neonates.

scholarly journals Evaluation of the New BD Max GC Real-Time PCR Assay, Analytically and Clinically as a Supplementary Test for the BD ProbeTec GC Qx Amplified DNA Assay, for Molecular Detection of Neisseria gonorrhoeae

2015 ◽  
Vol 53 (12) ◽  
pp. 3935-3937 ◽  
Author(s):  
Daniel Golparian ◽  
Stina Boräng ◽  
Martin Sundqvist ◽  
Magnus Unemo
Keyword(s):  
Neisseria Gonorrhoeae ◽  
Real Time ◽  
Sensitivity And Specificity ◽  
Real Time Pcr ◽  
Molecular Detection ◽  
Analytical Sensitivity ◽  
Pcr Assay ◽  
Content Type ◽  
Dna Assay ◽  
Supplementary Test

The new BD Max GC real-time PCR assay showed high clinical and analytical sensitivity and specificity. It can be an effective and accurate supplementary test for the BD ProbeTec GC Qx amplified DNA assay, which had suboptimal specificity, and might also be used for initial detection ofNeisseria gonorrhoeae.

Is Endoscopic Assessment of the Esophagus and Stomach Enough to Determine the Need for Biopsy at These Sites in Pediatric Patients Undergoing Endoscopy for Elevated TTG?

2021 ◽  
pp. 109352662199148
Author(s):  
M. Cristina Pacheco ◽  
Nicole Green ◽  
Jane Dickerson ◽  
Dale Lee
Keyword(s):  
Sensitivity And Specificity ◽  
Pediatric Patients ◽  
Tissue Transglutaminase ◽  
Immunoglobulin A ◽  
High Specificity ◽  
Visual Assessment ◽  
Pathology Report ◽  
Low Sensitivity ◽  
Endoscopic Assessment

Objectives The goal of our study was to determine whether visual assessment of the esophagus and stomach could predict abnormal histology and determine the frequency of interventions based on biopsies in patients undergoing endoscopy for elevated tissue transglutaminase immunoglobulin A antibody (TTG). Methods Pathology records were searched for patients with biopsy performed for elevated TTG. Pathology report, endoscopy report, and follow-up were obtained and slides from the duodenum reviewed. Pathology was considered gold standard for sensitivity and specificity calculations. Results 240 patients were included. 215 patients had esophageal biopsies performed. Esophageal endoscopic visual assessment had sensitivity of 47% and specificity of 93% for abnormal histology. 16(7%) patients had therapy or referral related to results and, of these, 6(38%) had visually normal endoscopy. 237 biopsies were performed of stomach. Gastric endoscopic visual assessment had a sensitivity and specificity of 20% and 87%. 24(10%) patients had therapy based on findings and, of these, 12 (50%) had visually normal endoscopy. Conclusions Endoscopic assessment of esophagus and stomach has low sensitivity and high specificity for pathologic abnormalities when indication for endoscopy is elevated TTG. When endoscopy is visually normal clinical interventions based on biopsy are rare, and foregoing biopsy may be considered.

scholarly journals Limited-Resource Preparable Chitosan Magnetic Particles for Extracting Amplification-Ready Zeptomole-Range Nucleic Acid from Complex Biofluid

2021 ◽  
Author(s):  
Sayantan Tripathy ◽  
Arunansu Talukdar ◽  
Goutam Pramanik ◽  
P. V. Rajesh ◽  
Souradyuti Ghosh
Keyword(s):  
Nucleic Acid ◽  
Real Time ◽  
Real Time Pcr ◽  
Magnetic Particles ◽  
Limited Resource ◽  
Nucleic Acid Amplification ◽  
Acid Extraction ◽  
Analytical Sensitivity ◽  
Nucleic Acid Extraction ◽  
Spin Column

<b>Layman Summary: </b>Nucleic acid extraction is a key prerequisite for any nucleic acid amplification test (NAAT) or isothermal NAAT (iNAAT) based molecular diagnosis assays.<b> </b>Existing methods utilizes spin column system for nucleic acid extraction which are unsuitable for limited resource settings. Our work explores two methods for chitosan coated magnetic particle preparation that can be executed within 6 h from commonly available chemicals with nothing but a magnetic stirrer and water bath and doable by a minimally trained person. We will also investigated the compatibility of the extracted nucleic acid with downstream NAATs such as real time LAMP, colorimetric LAMP, and real time PCR. In the process, we established the analytical sensitivity of the overall method.<div><br><div><b>Characterization methods</b>: SEM, XRD, EDX, FT-IR</div><div><br></div><div><b>Bioanalytical methods:</b> Real time LAMP, Colorimetric LAMP, Real time PCR</div></div>

scholarly journals Evaluation of the performance of the COVID-19 rapid antigen tests in a tertiary level hospital in Bangladesh.

2021 ◽  
Author(s):  
Mohammad Jahidur Rahman Khan ◽  
Md. Shahadat Hossain ◽  
Samshad Jahan Shumu ◽  
Md. Selim Reza ◽  
Farzana Mim ◽  
...  
Keyword(s):  
Viral Load ◽  
Real Time ◽  
Sensitivity And Specificity ◽  
Detection Rate ◽  
High Sensitivity ◽  
Rt Pcr ◽  
College Hospital ◽  
Medical College ◽  
Qrt Pcr ◽  
Antigen Tests

Abstract Background: While the COVID-19 pandemic is a worldwide crisis, tests with high sensitivity and specificity are essential for identifying and managing COVID-19 patients. Globally, several rapid antigen tests RATs for COVID-19 have been developed, but their clinical efficacy has not been well established. This study aimed to evaluate the performance of several rapid antigen tests (RATs) to diagnose SARS-CoV-2 infection.Methods: This prospective observational study was conducted at Shaheed Suhrawardy Medical College hospital from February 2021 to April 2021 in Dhaka, Bangladesh. This study included the patients admitted in this hospital at the COVID-19 isolation unit or referred from the triage facility of the outdoor department of this hospital suspected as COVID-19 case. Two nasopharyngeal samples were collected simultaneously. one sample was used on the spot for the RAT. The other was sent to the adjacent Shaheed Suhrawardy Medical College COVID-19 RT-PCR laboratory for real-time reverse transcription-polymerase chain reaction (qRT-PCR). The performance of the RAT was evaluated using the results of qRT-PCR as a reference.Results: A total of 223 patients were included in this study, and the real-time RT-PCR detected SARS-CoV-2 in 84 (37.7%) patients. Of these 84 patients, 9 (10.7%) were asymptomatic. The overall sensitivity and specificity of RATs were 78.6% and 99.3%, respectively. The sensitivity was 81.3% in symptomatic cases and 55.6% in asymptomatic cases. False-negatives were observed in 18 patients, 3 of whom were asymptomatic and had a low viral load (cycle threshold (Ct) > 30). The detection rate of RATs was 100% when the Ct value was up to 24. The detection rate was 42.3% when the Ct was >29. The detection rate of RATs was 92.3% when the onset of symptoms was within three days. The detection rate was 33.3% when the onset of symptoms was >7 days.Conclusions: RATs for COVID-19 used in this study delivered an acceptable performance in patients with high viral load and within the first week of the onset of symptoms. They can be used as a supplementary method to RT-PCR for the diagnosis of COVID-19 patients.

scholarly journals Clinical data and basal gonadotropins in the diagnostic of central precocious puberty in girls

2021 ◽  
Author(s):  
Teodoro Durá-Travé ◽  
Fidel Gallinas-Victoriano ◽  
María Malumbres-Chacon ◽  
Lotfi Ahmed-Mohamed ◽  
María Jesús Chueca Guindulain ◽  
...  
Keyword(s):  
Sensitivity And Specificity ◽  
Age At Onset ◽  
Area Under The Curve ◽  
High Specificity ◽  
Central Precocious Puberty ◽  
Bone Age ◽  
Height Velocity ◽  
Pubertal Status ◽  
Serum Lh ◽  
Before And After

Objective. The objective of this study was to analyze whether some auxological characteristics or a single basal gonadotropin measurement will be sufficient to distinguish the prepubertal from pubertal status. Methods. Auxologycal characteristics were recorded and serum LH and FSH were measured by immunochemiluminescence assays before and after GnRH stimulation test in a sample of 241 Caucasian girls with breast budding between 6- to 8- years old. Peak LH levels higher than 5 IU/L were considered a pubertal response. Area under the curve, cut-off points, sensitivity, and specificity for auxologycal variables and basal gonadotropins levels were determined by receiver operating curves. Results. There were no significant differences in age at onset, weight, height, BMI and height velocity between both groups. Bone age was significantly higher in pubertal girls (p<0.05), although with limited discriminatory capacity. The sensitivity and specificity for the basal LH levels were 89% and 82% respectively, for a cut off point of 0.1 IU/L. All girls in the pubertal group had a basal LH higher than 1.0 IU/L (positive predictive value of 100%). There was a wide overlap of basal FSH and LH/FSH ratio between prepubertal and pubertal girls. Conclusions. Auxologycal characteristics should not be used only in the differential diagnosis between prepubertal from pubertal status in 6- to 8-year-old girls. We found a high specificity of a single basal LH sample and it would be useful for establishing the diagnosis of puberty in this age group, eliminating the need for GnRH stimulation testing.

scholarly journals Interpretations of Antibody Responses toSalmonella enterica Serotype Enteritidis gm Flagellin in Poultry Flocks Are Enhanced by a Kinetics-Based Enzyme-Linked Immunosorbent Assay

1998 ◽  
Vol 5 (4) ◽  
pp. 550-555 ◽  
Author(s):  
Patrick L. McDonough ◽  
Richard H. Jacobson ◽  
John F. Timoney ◽  
Ahmed Mutalib ◽  
David C. Kradel ◽  
...  
Keyword(s):  
Immunosorbent Assay ◽  
High Sensitivity ◽  
High Specificity ◽  
Enzyme Linked Immunosorbent Assay ◽  
Analytical Sensitivity ◽  
Department Of Agriculture ◽  
Trace Back ◽  
Elisa Format ◽  
Commercial Poultry ◽  
Computer Controlled

ABSTRACT Many regulatory and diagnostic programs for the detection ofSalmonella enterica serotype Enteritidis infection in commercial poultry flocks have relied on rapid Pullorum agglutination tests to screen birds because of the shared antigens of S. enterica Enteritidis and S. enterica Pullorum and Gallinarum; however, the use of the enzyme-linked immunosorbent assay (ELISA) format affords better analytical sensitivity than crude agglutination tests. In this study, we adapted our earlier conventional indirect ELISA, using gm flagellin as the antigen, to a kinetics-based, computer-controlled ELISA (KELA). The KELA was used to screen for flagellin antibody from three commercial flocks: (i) a large flock involved in a U.S. Department of Agriculture trace back from a humanS. enterica Enteritidis foodborne outbreak (n = 3,209), (ii) a flock infected with the endemicS. enterica Enteritidis serotype but which also had multiple other salmonella serotypes (n = 65), and (iii) an S. enterica Pullorum-infected flock (n = 12). The first flock (S. entericaEnteritidis prevalence of 2.45% based on culture) provided a field test of the KELA and allowed the calculation of diagnostic sensitivity (D-Sn) and diagnostic specificity (D-Sp). With a cutoff of 10 (used for screening flocks [i.e., high sensitivity]), the KELA has a D-Sn of 95.2% and a D-Sp of 18.5%; with a cutoff of 140 (used in confirmatory flock testing [i.e., high specificity]), the KELA has a D-Sn of 28.0% and a D-Sp of 99.1%. We found that with a cutoff of 60 (D-Sn = 63.1%; D-Sp = 91.6%), we could eliminate reactions in the KELA caused by other non-S. enterica Enteritidis salmonellae. The KELA was also compared to two commercial rapid Pullorum tests, the Solvay (D-Sn = 94.9%; D-Sp = 55.5%) and the Vineland (D-Sn = 62.0%; D-Sp = 75.3%).

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