scholarly journals In silico evaluation of the impact of the Omicron variant on the sensitivity of RT-qPCR assays for SARS-CoV-2 detection using whole genome sequencing

2022 ◽  
Author(s):  
Divya Sharma ◽  
Chengjin Ye ◽  
Giusppe Lippi ◽  
Jordi B. Torrelles ◽  
Luis Martinez-Sobrido ◽  
...  
Keyword(s):  
In Silico ◽  
False Negative ◽  
Whole Genome ◽  
False Negatives ◽  
Negative Results ◽  
False Negative Results ◽  
The World ◽  
Reverse Primer ◽  
Primer Annealing ◽  
The Impact

Abstract Background The novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variant of concern (VoC) Omicron (B.1.1.529) has rapidly spread around the world presenting a new threat to global public human health. Due to the large number of mutations possessed by Omicron, concerns have emerged over potentially reduced diagnostic accuracy of reverse transcription polymerase chain reaction (RT-qPCR), the gold standard diagnostic test for SARS-CoV-2. Here, we aimed to assess the impact of Omicron on the integrity and sensitivity of RT-qPCR assays used for coronavirus disease-2019 (COVID-19) diagnosis via in silico analysis employing whole genome sequencing data and evaluated the potential for false negatives or test failure due to mismatches between primers/probes and viral genome. Methods In silico sensitivity of 12 RT-qPCR tests (containing 30 primers and probe sets) developed for detection of SARS-CoV-2 reported by the World Health Organization (WHO) or available in the literature, was assessed for use in detecting SARS-CoV-2 Omicron BA.1 and BA.2 sublineages, obtained after removing redundancy from publicly available genomes from National Center for Biotechnology Information (NCBI) and Global Initiative on Sharing Avian Influenza Data (GISAID) databases. The mismatches between the amplicon regions of the SARS-CoV-2 Omicron VoC and primers and probe sets were evaluated, and the clustering analysis of the corresponding amplicon sequences was carried out. Results From the 232 representative SARS-CoV-2 BA.1 Omicron sublineage genomes analyzed, 229 showed substitutions at the forward primer annealing site for assay China-CDC N, 226 showed mismatches in the reverse primer annealing site for assay Thai N, and all 232 had substitution at the 3’ end of the reverse primer annealing site for assay HKUniv RdRp/Hel. Therefore, the lowest sensitivity was observed for assay ChinaCDC N, Thai N and HKUniv RdRp/Hel for SARS-CoV-2 BA.1 sublineage genomes. For 5 SARS-CoV-2 BA.2 Omicron sublineage genomes, false negative results were observed for assays ChinaCDC N, Thai N, HKUniv RdRp/Hel, SigmAldr S5, SigmAldr S6 and HKUniv S. Conclusion In this study, we observed three (25%) assays (ChinaCDC N, Thai N, and HKUniv RdRp/Hel) demonstrated potential for false negatives for the SARS-CoV-2 Omicron BA.1 sublineage, while four (33.3%) assays (ChinaCDC N, Thai N, HKUniv RdRp/Hel, HKUniv S, SigmAldr S5 and SigmAldr S6) demonstrated potential false negative results for the for SARS-CoV-2 Omicron BA.2 sublineage, which also has the potential for Spike (S) gene dropout despite lacking 69-70 deletion in the S gene. Further, amplicon clustering and additional substitutions analysis along with the sensitivity analysis could be used for modification and development of RT-qPCR assays for detection of SARS-CoV-2 Omicron VoC lineages.

scholarly journals The 28 + 28 day design is an effective sampling time for analyzing mutant frequencies in rapidly proliferating tissues of MutaMouse animals

2021 ◽  
Vol 95 (3) ◽  
pp. 1103-1116
Author(s):  
Francesco Marchetti ◽  
Gu Zhou ◽  
Danielle LeBlanc ◽  
Paul A. White ◽  
Andrew Williams ◽  
...  
Keyword(s):  
Germ Cells ◽  
False Negative ◽  
Sampling Time ◽  
Male Germ Cells ◽  
Negative Results ◽  
False Negative Results ◽  
Detection Of Mutations ◽  
The Impact ◽  
Sampling Times

AbstractThe Organisation for Economic Co-Operation and Development Test Guideline 488 (TG 488) uses transgenic rodent models to generate in vivo mutagenesis data for regulatory submission. The recommended design in TG 488, 28 consecutive daily exposures with tissue sampling three days later (28 + 3d), is optimized for rapidly proliferating tissues such as bone marrow (BM). A sampling time of 28 days (28 + 28d) is considered more appropriate for slowly proliferating tissues (e.g., liver) and male germ cells. We evaluated the impact of the sampling time on mutant frequencies (MF) in the BM of MutaMouse males exposed for 28 days to benzo[a]pyrene (BaP), procarbazine (PRC), isopropyl methanesulfonate (iPMS), or triethylenemelamine (TEM) in dose–response studies. BM samples were collected + 3d, + 28d, + 42d or + 70d post exposure and MF quantified using the lacZ assay. All chemicals significantly increased MF with maximum fold increases at 28 + 3d of 162.9, 6.6, 4.7 and 2.8 for BaP, PRC, iPMS and TEM, respectively. MF were relatively stable over the time period investigated, although they were significantly increased only at 28 + 3d and 28 + 28d for TEM. Benchmark dose (BMD) modelling generated overlapping BMD confidence intervals among the four sampling times for each chemical. These results demonstrate that the sampling time does not affect the detection of mutations for strong mutagens. However, for mutagens that produce small increases in MF, sampling times greater than 28 days may produce false-negative results. Thus, the 28 + 28d protocol represents a unifying protocol for simultaneously assessing mutations in rapidly and slowly proliferating somatic tissues and male germ cells.

Phenothiazine Derivatives in Urine: "False Nagative Results" With the Forrest Color Test (FPN), and a Method for Their Elimination

1965 ◽  
Vol 11 (10) ◽  
pp. 914-919
Author(s):  
K N Campbell
Keyword(s):  
False Negative ◽  
False Positives ◽  
Phenothiazine Derivatives ◽  
Color Test ◽  
False Negatives ◽  
Delayed Reaction ◽  
Negative Results ◽  
False Negative Results ◽  
Phenothiazine Drugs ◽  
Urine Specimens

Abstract More than 100 Forrest color tests (FPN) were performed on urine specimens from patients taking phenothiazine drugs. The results obtained showed > 20% "false negatives" and 6% "false positives." A delayed reaction due to possible individual patient variation seems to have been the cause for false-negative results. These false negatives were shown to disappear when test readings were delayed for 1-5 min. Some false positives were due to liver dysfunction.

Onychomycosis Infections

2017 ◽  
Vol 107 (4) ◽  
pp. 280-286 ◽  
Author(s):  
Aditya K. Gupta ◽  
Kerry-Ann Nakrieko
Keyword(s):  
False Negative ◽  
Kappa Statistic ◽  
False Negatives ◽  
Negative Results ◽  
False Negative Results ◽  
Repeated Sampling ◽  
Polymerase Chain ◽  
Serial Samples ◽  
Repeated Samples ◽  
Infecting Organisms

Background: Mycological culture is the traditional method for identifying infecting agents of onychomycosis despite high false-negative results, slower processing, and complications surrounding nondermatophyte mold (NDM) infections. Molecular polymerase chain reaction (PCR) methods are faster and suited for ascertaining NDM infections. Methods: To measure agreement between culture and PCR methods for identification of infecting species of suspected onychomycosis, single toenail samples from 167 patients and repeated serial samples from 43 patients with suspected onychomycosis were processed by culture and PCR for identification of 16 dermatophytes and five NDMs. Agreement between methods was quantified using the kappa statistic (κ). Results: The methods exhibited fair agreement for the identification of all infecting organisms (single samples: κ = 0.32; repeated samples: κ = 0.38). For dermatophytes, agreement was moderate (single samples: κ = 0.44; repeated samples: κ = 0.42). For NDMs, agreement was poor with single samples (κ = 0.16) but fair with repeated samples (κ = 0.25). Excluding false-negative reports from analyses improved agreement between methods in all cases except the identification of NDMs from single samples. Conclusions: Culture was three or four times more likely to report a false-negative result compared with PCR. The increased agreement between methods observed by excluding false-negative reports statistically clarifies and highlights the major discord caused by false-negative cultures. The increased agreement of NDM identification from poor to fair with repeated sampling along with their poor agreement in the single samples, with and without false-negatives, affirms the complications of NDM identification and supports the recommendation that serial samples help confirm the diagnosis of NDM infections.

scholarly journals FALSE-NEGATIVE RESULTS IN SCREENING PROGRAMS

2001 ◽  
Vol 17 (2) ◽  
pp. 164-170 ◽  
Author(s):  
Mark Petticrew ◽  
Amanda Sowden ◽  
Deborah Lister-Sharp
Keyword(s):  
Negative Impact ◽  
Relevant Literature ◽  
False Negative ◽  
Quantitative Information ◽  
Public Confidence ◽  
Psychological Consequences ◽  
False Negatives ◽  
Screening Programs ◽  
Negative Results ◽  
False Negative Results

Objectives: Assessment of the appropriateness of screening programs involves consideration of the harms as well as the benefits. These harms include the risk of false-negative results, the consequences of which have remained underinvestigated. This paper reports the results of a systematic literature review that aimed to assess the medical psychological, economic, and legal consequences of false-negative results in national screening programs.Methods: The review included a comprehensive literature search and contact with experts to identify relevant literature. Most studies that were identified presented only anecdotal evidence. However, thirteen studies presented quantitative information on medical consequences of false negatives, eight studies presented information on psychological consequences, and two studies presented information on economic consequences.Results: The strength of evidence from most of the primary studies was low. There is some evidence, however, that false-negative results may have a large legal impact. There is also a consensus in the literature that false negatives may have a negative impact on public confidence on screening; evidence is however limited.Conclusions: False negatives are evident even in high-quality screening programs. They may have the potential to delay the detection of breast and cervical cancer, but there is little evidence to help in assessing their psychological consequences. They also may lead to legal action being taken by those affected and may reduce public confidence in screening. Their impact may be reduced by provision of full information to participants about the benefits and limitations of screening programs and by increasing public education on these issues.

scholarly journals Analysis and Prediction of COVID-19 Patients’ False Negative Results for SARS-CoV-2 Detection with Pharyngeal Swab Specimen: A Retrospective Study

2020 ◽  
Author(s):  
Hui Xu ◽  
Li Yan ◽  
Chun (Martin) Qiu ◽  
Bo Jiao ◽  
Yanyan Chen ◽  
...  
Keyword(s):  
Nucleic Acid ◽  
Time Window ◽  
Clinical Symptoms ◽  
False Negative ◽  
Ct Imaging ◽  
Nucleic Acid Detection ◽  
False Negatives ◽  
Negative Results ◽  
False Negative Results ◽  
Blood Routine

ABSTRACTBackgroundFalse negative results of SARS-CoV-2 nucleic acid detection pose threats to COVID-19 patients and medical workers alike.ObjectiveTo develop multivariate models to determine clinical characteristics that contribute to false negative results of SARS-CoV-2 nucleic acid detection, and use them to predict false negative results as well as time windows for testing positive.DesignRetrospective Cohort Study (Ethics number of Tongji Hospital: No. IRBID: TJ-20200320)SettingA database of outpatients in Tongji Hospital (University Hospital) from 15 January 2020 to 19 February 2020.Patients1,324 outpatients with COVID-19MeasurementsClinical information on CT imaging reports, blood routine tests, and clinic symptoms were collected. A multivariate logistic regression was used to explain and predict false negative testing results of SARS-CoV-2 detection. A multivariate accelerated failure model was used to analyze and predict delayed time windows for testing positive.ResultsOf the 1,324 outpatients who diagnosed of COVID-19, 633 patients tested positive in their first SARS-CoV-2 nucleic acid test (47.8%), with a mean age of 51 years (SD=14.9); the rest, which had a mean age of 47 years (SD=15.4), tested negative in the first test. “Ground glass opacity” in a CT imaging report was associated with a lower chance of false negatives (aOR, 0.56), and reduced the length of time window for testing positive by 26%. “Consolidation” was associated with a higher chance of false negatives (aOR, 1.57), and extended the length of time window for testing positive by 44%. In blood routine tests, basophils (aOR, 1.28) and eosinophils (aOR, 1.29) were associated with a higher chance of false negatives, and were found to extend the time window for testing positive by 23% and 41%, respectively. Age and gender also affected the significantly.LimitationData were generated in a large single-center study.ConclusionTesting outcome and positive window of SARS-CoV-2 detection for COVID-19 patients were associated with CT imaging results, blood routine tests, and clinical symptoms. Taking into account relevant information in CT imaging reports, blood routine tests, and clinical symptoms helped reduce a false negative testing outcome. The predictive AFT model, what we believe to be one of the first statistical models for predicting time window of SARS-CoV-2 detection, could help clinicians improve the accuracy and efficiency of the diagnosis, and hence, optimizes the timing of nucleic acid detection and alleviates the shortage of nucleic acid detection kits around the world.Primary Funding SourceNone.

scholarly journals Modelling the epidemiology of malaria and spread of HRP2-negative Plasmodium falciparum following the replacement of HRP2-detecting rapid diagnostic tests

2022 ◽  
Vol 2 (1) ◽  
pp. e0000106
Author(s):  
Alisha Chaudhry ◽  
Jane Cunningham ◽  
Qin Cheng ◽  
Michelle L. Gatton
Keyword(s):  
Plasmodium Falciparum ◽  
Diagnostic Tests ◽  
False Negative ◽  
Parasite Prevalence ◽  
Rapid Diagnostic Tests ◽  
Negative Results ◽  
False Negative Results ◽  
Malaria Rapid Diagnostic Tests ◽  
The Impact

Malaria rapid diagnostic tests (RDTs) are dominated by products which use histidine-rich protein 2 (HRP2) to detect Plasmodium falciparum. The emergence of parasites lacking the pfhrp2 gene can lead to high rates of false-negative results amongst these RDTs. One solution to restore the ability to correctly diagnose falciparum malaria is to switch to an RDT which is not solely reliant on HRP2. This study used an agent-based stochastic simulation model to investigate the impact on prevalence and transmission caused by switching the type of RDT used once false-negative rates reached pre-defined thresholds within the treatment-seeking symptomatic population. The results show that low transmission settings were the first to reach the false-negative switch threshold, and that lower thresholds were typically associated with better long-term outcomes. Changing the diagnostic RDT away from a HRP2-only RDT is predicted to restore the ability to correctly diagnose symptomatic malaria infections, but often did not lead to the extinction of HRP2-negative parasites from the population which continued to circulate in low density infections, or return to the parasite prevalence and transmission levels seen prior to the introduction of the HRP2-negative parasite. In contrast, failure to move away from HRP2-only RDTs leads to near fixation of these parasites in the population, and the inability to correctly diagnose symptomatic cases. Overall, these results suggest pfhrp2-deleted parasites are likely to become a significant component of P. falciparum parasite populations, and that long-term strategies are needed for diagnosis and surveillance which do not rely solely on HRP2.

scholarly journals Maternal mortality by COVID-19 in Brazil

2021 ◽  
Vol 21 (suppl 1) ◽  
pp. 253-256
Author(s):  
Alex Sandro Rolland Souza ◽  
Melania Maria Ramos Amorim
Keyword(s):  
Prognostic Factors ◽  
Mortality Rate ◽  
Maternal Mortality ◽  
Risk Group ◽  
False Negative ◽  
Health Policies ◽  
Negative Results ◽  
False Negative Results ◽  
The World ◽  
Complications And Mortality

Abstract SARS-CoV-2, the new coronavirus of severe acute respiratory syndrome, which causes a predominantly respiratory disease called COVID-19, quickly caused a pandemic, due to its high transmissibility, leaving a trail of deaths around the world. Initially, the pregnancy puerperal cycle was not associated with complications and mortality, only later was recognized as a risk group. As the disease progressed, the maternal mortality rate by COVID-19 increased, Brazil is responsible for an important portion. This rate may be even higher due to underreporting, difficulties in performing laboratorial tests and possible false negative results and depends on the health policies adopted by each region or country. It is important to carry out studies on maternal mortality so that the prognostic factors can be recognized and so avoid them.

scholarly journals Preventing False Negatives for Histochemical Detection of Phenolics and Lignins in PEG-Embedded Plant Tissues

2016 ◽  
Vol 65 (2) ◽  
pp. 105-116 ◽  
Author(s):  
Bruno G. Ferreira ◽  
Renan Falcioni ◽  
Lubia M. Guedes ◽  
Sofia C. Avritzer ◽  
Werner C. Antunes ◽  
...  
Keyword(s):  
Ferrous Sulfate ◽  
Low Cost ◽  
False Negative ◽  
Plant Tissues ◽  
False Negatives ◽  
Plant Materials ◽  
Negative Results ◽  
False Negative Results ◽  
Histochemical Tests ◽  
Embedding Medium

Polyethylene glycol (PEG) is a low-cost and advantageous embedding medium, which maintains the majority of cell contents unaltered during the embedding process. Some hard or complex plant materials are better embedded in PEG than in other usual embedding media. However, the histochemical tests for phenolics and lignins in PEG-embedded plant tissues commonly result in false negatives. We hypothesize that these false negatives should be prevented by the use of distinct fixatives, which should avoid the bonds between PEG and phenols. Novel protocols for phenolics and flavanols detection are efficiently tested, with fixation of the samples in ferrous sulfate and formalin or in caffeine and sodium benzoate, respectively. The differentiation of lignin types is possible in safranin-stained sections observed under fluorescence. The Maule’s test faultlessly distinguishes syringyl-rich from guaiacyl- and hydroxyphenyl-rich lignins in PEG-embedded material under light microscopy. Current hypothesis is corroborated, that is, the adequate fixation solves the false-negative results, and the new proposed protocols fill up some gaps on the detection of phenolics and lignins.

scholarly journals PANDAA intentionally violates conventional qPCR design to enable durable, mismatch-agnostic detection of highly polymorphic pathogens

2021 ◽  
Vol 4 (1) ◽  
Author(s):  
Iain J. MacLeod ◽  
Christopher F. Rowley ◽  
M. Essex
Keyword(s):  
Sequence Variation ◽  
False Negative ◽  
Site Directed Mutagenesis ◽  
Resistance Mutations ◽  
Emerging Pathogens ◽  
Degenerate Primers ◽  
Negative Results ◽  
False Negative Results ◽  
Mutation Assay ◽  
The Impact

AbstractSensitive and reproducible diagnostics are fundamental to containing the spread of existing and emerging pathogens. Despite the reliance of clinical virology on qPCR, technical challenges persist that compromise their reliability for sustainable epidemic containment as sequence instability in probe-binding regions produces false-negative results. We systematically violated canonical qPCR design principles to develop a Pan-Degenerate Amplification and Adaptation (PANDAA), a point mutation assay that mitigates the impact of sequence variation on probe-based qPCR performance. Using HIV-1 as a model system, we optimized and validated PANDAA to detect HIV drug resistance mutations (DRMs). Ultra-degenerate primers with 3’ termini overlapping the probe-binding site adapt the target through site-directed mutagenesis during qPCR to replace DRM-proximal sequence variation. PANDAA-quantified DRMs present at frequency ≥5% (2 h from nucleic acid to result) with a sensitivity and specificity of 96.9% and 97.5%, respectively. PANDAA is an innovative advancement with applicability to any pathogen where target-proximal genetic variability hinders diagnostic development.

Elimination of False Negative Results with the FPN Forrest Test for Phenothiazine Derivatives in Urine

1966 ◽  
Vol 12 (6) ◽  
pp. 379-384 ◽  
Author(s):  
Irene S Forrest ◽  
Fred M Forrest ◽  
Saul L Kanter
Keyword(s):  
Reading Time ◽  
False Negative ◽  
False Positives ◽  
Phenothiazine Derivatives ◽  
False Negatives ◽  
Negative Results ◽  
Color Development ◽  
False Negative Results ◽  
Color Reactions ◽  
Urine Specimens

Abstract With the FPN Forrest test for urinary phenothiazine derivatives, omission of drug intake was found to be the most frequent cause of negative color reactions; accordingly, these negative reactions are true negatives. Excessive dilution of urine specimens occasionally yielded "apparent false negatives" by producing traces of drug metabolites, which cannot be detected without modification of the test. When measures to eliminate these conditions are taken, the previously specified reading time (10 sec. after mixing urine and reagent) is reliable. Unspecific color development due to reaction between endogenous urinary constituents and reagent appeared with a delay of 30 sec. or more, and hence recent recommendations of delaying reading times for the test were found to be unsuitable and productive of false positives.

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