Population wide testing pooling strategy for SARS-CoV-2 detection using saliva.
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ABSTRACT:
SUBMITTER: Esteves E
PROVIDER: S-EPMC8797214 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Expanding testing capabilities is integral to managing the further spread of SARS-CoV-2 and developing reopening strategies, particularly in regards to identifying and isolating asymptomatic and pre-symptomatic individuals. Central to meeting testing demands are specimens that can be easily and reliably collected and laboratory capacity to rapidly ramp up to scale. We and others have demonstrated that high and consistent levels of SARS-CoV-2 RNA can be detected in saliva from COVID-19 inpatients, outpatients, and asymptomatic individuals. As saliva collection is non-invasive, extending this strategy to test pooled saliva samples from multiple individuals could thus provide a simple method to expand testing capacity. However, hesitation towards pooled sample testing arises due to the dilution of positive samples, potentially shifting weakly positive samples below the detection limit for SARS-CoV-2 and thereby decreasing the sensitivity. Here, we investigated the potential of pooling saliva samples by 5, 10, and 20 samples prior to RNA extraction and RT-qPCR detection of SARS-CoV-2. Based on samples tested, we conservatively estimated a reduction of 7.41%, 11.11%, and 14.81% sensitivity, for each of the pool sizes, respectively. Using these estimates we modeled anticipated changes in RT-qPCR cycle threshold to show the practical impact of pooling on results of SARS-CoV-2 testing. In tested populations with greater than 3% prevalence, testing samples in pools of 5 requires the least overall number of tests. Below 1% however, pools of 10 or 20 are more beneficial and likely more supportive of ongoing surveillance strategies.
| S-EPMC7480055 | biostudies-literature
Project description:As the battle against coronavirus disease 2019 pandemic continues, an increase in workload and medical expenses have been a concern to the health care system worldwide. Developing a measure that helps to conserve the health care resource is, therefore, highly desirable, and the pooling of the specimens for testing is one of the attractive strategies. Recently, we showed that saliva could be a potential alternative specimen for the detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) by real-time polymerase chain reaction (RT-PCR). In the present study, we performed the pooling of saliva specimens for testing by SARS-CoV-2 RT-PCR. We showed that the saliva pool of either 5 or 10 samples, by allowing the detection of either gene in the pool at an increased cycle threshold cutoff value, further performing individual sample testing in the positive pools did not compromise the detection of SARS-CoV-2.
| S-EPMC7461487 | biostudies-literature
Project description:BackgroundSensitive and high throughput molecular detection assays are essential during the coronavirus disease 2019 (COVID-19) pandemic, caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). The vast majority of the SARS-CoV-2 molecular assays use nasopharyngeal swab (NPS) or oropharyngeal swab (OPS) specimens collected from suspected individuals. However, using NPS or OPS as specimens has apparent drawbacks, e.g. the collection procedures for NPS or OPS specimens can be uncomfortable to some people and may cause sneezing and coughing which in turn generate droplets and/or aerosol particles that are of risk to healthcare workers, requiring heavy use of personal protective equipment. There have been recent studies indicating that self-collected saliva specimens can be used for molecular detection of SARS-CoV-2 and provides more comfort and ease of use for the patients. Here we report the performance of QuantiVirus™ SARS-CoV-2 test using saliva as the testing specimens with or without pooling.MethodsDevelopment and validation studies were conducted following FDA-EUA and molecular assay validation guidelines. Using SeraCare Accuplex SARS-CoV-2 reference panel, the limit of detection (LOD) and clinical performance studies were performed with the QuantiVirus™ SARS-CoV-2 test. For clinical evaluation, 85 known positive and 90 known negative clinical NPS samples were tested. Additionally, twenty paired NPS and saliva samples collected from recovering COVID-19 patients were tested and the results were further compared to that of the Abbott m2000 SARS-CoV-2 PCR assay. Results of community collected 389 saliva samples for COVID-19 screening by QuantiVirus™ SARS-CoV-2 test were also obtained and analyzed. Additionally, testing of pooled saliva samples was evaluated.ResultsThe LOD for the QuantiVirus™ SARS-CoV-2 test was confirmed to be 100-200 copies/mL. The clinical performance studies using contrived saliva samples indicated that the positive percentage agreement (PPA) of the QuantiVirus™ SARS-CoV-2 test is 100% at 1xLOD, 1.5xLOD and 2.5xLOD. No cross-reactivity was observed for the QuantiVirus™ SARS-CoV-2 test with common respiratory pathogens. Testing of clinical samples showed a positive percentage agreement (PPA) of 100% (95% CI: 94.6% to 100%) and a negative percentage agreement (NPA) of 98.9% (95% CI: 93.1% to 99.9%). QuantiVirus™ SARS CoV-2 test had 80% concordance rate and no significant difference (p = 0.13) between paired saliva and NPS specimens by Wilcoxon matched pairs signed rank test. Positive test rate was 1.79% for 389 saliva specimens collected from local communities for COVID-19 screening. Preliminary data showed that saliva sample pooling up to 6 samples (1:6 pooling) for SARS-CoV-2 detection is feasible (sensitivity 94.8% and specificity 100%).ConclusionThe studies demonstrated that the QuantiVirus™ SARS-CoV-2 test has a LOD of 200 copies/mL in contrived saliva samples. The clinical performance of saliva-based testing is comparable to that of NPS-based testing. Pooling of saliva specimens for SARS-CoV-2 detection is feasible. Saliva based and high-throughput QuantiVirus™ SARS-CoV-2 test offers a highly desirable testing platform during the ongoing COVID-19 pandemic.
| S-EPMC7901781 | biostudies-literature
Project description:ObjectivesDespite the success in developing COVID-19 vaccines, containment of the disease is obstructed worldwide by vaccine production bottlenecks, logistics hurdles, vaccine refusal, transmission through unvaccinated children, and the appearance of new viral variants. This underscores the need for effective strategies for identifying carriers/patients, which was the main aim of this study.MethodsWe present a bubble-based PCR testing approach using swab-pooling into lysis buffer. A bubble is a cluster of people who can be periodically tested for SARS-CoV-2 by swab-pooling. A positive test of a pool mandates quarantining each of its members, who are then individually tested while in isolation to identify the carrier(s) for further epidemiological contact tracing.ResultsWe tested an overall sample of 25 831 individuals, divided into 1273 bubbles, with an average size of 20.3 ± 7.7 swabs/test tube, obtaining for all pools (≤37 swabs/pool) a specificity of 97.5% (lower bound 96.6%) and a sensitivity of 86.3% (lower bound 78.2%) and a post hoc analyzed sensitivity of 94.6% (lower bound 86.7%) and a specificity of 97.2% (lower bound 96.2%) in pools with ≤25 swabs, relative to individual testing.DiscussionThis approach offers a significant scale-up in sampling and testing throughput and savings in testing cost, without reducing sensitivity or affecting the standard PCR testing laboratory routine. It can be used in school classes, airplanes, hospitals, military units, and workplaces, and may be applicable to future pandemics.
| S-EPMC8849906 | biostudies-literature
Project description:The rapid identification and isolation of infected individuals remains a key strategy for controlling the spread of SARS-CoV-2. Frequent testing of populations to detect infection early in asymptomatic or presymptomatic individuals can be a powerful tool for intercepting transmission, especially when the viral prevalence is low. However, RT-PCR testing-the gold standard of SARS-CoV-2 diagnosis-is expensive, making regular testing of every individual unfeasible. Sample pooling is one approach to lowering costs. By combining samples and testing them in groups the number of tests required is reduced, substantially lowering costs. Here we report on the implementation of pooling strategies using 3-d and 4-d hypercubes to test a professional sports team in South Africa. We have shown that infected samples can be reliably detected in groups of 27 and 81, with minimal loss of assay sensitivity for samples with individual Ct values of up to 32. We report on the automation of sample pooling, using a liquid-handling robot and an automated web interface to identify positive samples. We conclude that hypercube pooling allows for the reliable RT-PCR detection of SARS-CoV-2 infection, at significantly lower costs than lateral flow antigen (LFA) tests.
| S-EPMC8423848 | biostudies-literature
Project description:Due to increased demand for testing, as well as restricted supply chain resources, testing for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection continues to face many hurdles. Pooling several samples has been proposed as an alternative approach to address these issues. We investigated the feasibility of pooling nasopharyngeal swab (NPS) or saliva samples for SARS-CoV-2 testing with a commercial assay (Idylla SARS-CoV-2 test; Biocartis). We evaluated the 10-pool and 20-pool approaches for 149 subjects, with 30 positive samples and 119 negative samples. The 10-pool approach had sensitivity of 78.95% (95% confidence interval [CI], 54.43% to 93.95%) and specificity of 100% (95% CI, 71.51% to 100%), whereas the 20-pool approach had sensitivity of 55.56% (95% CI, 21.20% to 86.30%) and specificity of 100% (95% CI, 25% to 100%). No significant difference was observed between the results obtained with pooled NPS and saliva samples. Given the rapidity, full automation, and practical advantages of the Idylla SARS-CoV-2 assay, pooling of 10 samples has the potential to significantly increase testing capacity for both NPS and saliva samples, with good sensitivity. IMPORTANCE To control outbreaks of coronavirus disease 2019 (COVID-19) and to avoid reagent shortages, testing strategies must be adapted and maintained for the foreseeable future. We analyzed the feasibility of pooling NPS and saliva samples for SARS-CoV-2 testing with the Idylla SARS-CoV-2 test, and we found that sensitivity was dependent on the pool size. The SARS-CoV-2 testing capacity with both NPS and saliva samples could be significantly expanded by pooling 10 samples; however, pooling 20 samples resulted in lower sensitivity.
| S-EPMC8579845 | biostudies-literature
Project description:The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic has significantly increased demand on laboratory throughput and reagents for nucleic acid extraction and polymerase chain reaction (PCR). Reagent shortages may limit the expansion of testing required to scale back containment measures. The aims of this study were to investigate the viability of sample pooling as a strategy for increasing test throughput and conserving PCR reagents; and to report our early experience with pooling of clinical samples. A pre-implementation study was performed to assess the sensitivity and theoretical efficiency of two, four, and eight-sample pools in a real-time reverse transcription PCR-based workflow. A standard operating procedure was developed and implemented in two laboratories during periods of peak demand, inclusive of over 29,000 clinical samples processed in our laboratory. Sensitivity decreased (mean absolute increase in cycle threshold value of 0.6, 2.3, and 3.0 for pools of two, four, and eight samples, respectively) and efficiency increased as pool size increased. Gains from pooling diminished at high disease prevalence. Our standard operating procedure was successfully implemented across two laboratories. Increased workflow complexity imparts a higher risk of errors, and requires risk mitigation strategies. Turnaround time for individual samples increased, hence urgent samples should not be pooled. Pooling is a viable strategy for high-throughput testing of SARS-CoV-2 in low-prevalence settings.
| S-EPMC7508550 | biostudies-literature
Project description:The worldwide COVID-19 pandemic outburst has caused a serious public health issue with increasing needs of accurate and rapid diagnostic and screening testing. This situation requires an optimized management of the chemical reagents, the consumables, and the human resources, in order to respond accurately and effectively, controlling the spread of the disease. Testing on pooled samples maximizes the number of tested samples, by minimizing the time and the lab supplies needed. The general conceptualization of the pooling method is based on mixing samples together in a batch. Individual testing is needed only if a specific pool exhibits a positive result. The development of alternative hybrid methods, based on "in house" protocols, utilizing commercially available consumables, in combination with a reliable pooling method would provide a solution, focusing on the better exploitation of the personnel and the lab supplies, allowing for rapid screening of a population in a reasonably short time.
| S-EPMC7834440 | biostudies-literature
Project description:Molecular diagnostic testing has played a critical role in the global response to the novel Coronavirus disease (COVID-19) pandemic, since its first outbreak in late 2019. At the inception of the COVID-19 pandemic, nasopharyngeal swab sample analysis for COVID-19 diagnosis using the real-time polymerase chain reaction (RT-PCR) technique was the most widely used. However, due to the high cost and difficulty of sample collection, the number of available sample types for COVID-19 diagnosis is rapidly increasing, as is the COVID-19 diagnostic literature. The use of nasal swabs, saliva, and oral fluids as viable sample options for the effective detection of SARS-CoV-2 has been implemented successfully in different settings since 2020. These alternative sample type provides a plethora of advantages including decreasing the high exposure risk to frontline workers, enhancing the chances of home self-sampling, reducing the cost, and significantly increasing testing capacity. This study sought to ascertain the effectiveness of Saliva samples as an alternative for COVID-19 diagnosis in Nigeria. Demographic data, paired samples of Nasopharyngeal Swab and Drooling Saliva were obtained from 309 consenting individuals aged 8-83 years presenting for COVID-19 testing. All samples were simultaneously assayed for the detection of SARS-CoV-2 RdRp, N, and E genes using the GeneFinder™ COVID-19 Plus RT-PCR test kit. Out of 309 participants, only 299 with valid RT-PCR results comprising 159 (53.2%) males and 140 (46.8%) females were analyzed in this study using the R Statistical package. Among the 299 samples analyzed, 39 (13.0%) had SARS-CoV-2 detected in at least one specimen type. Both swabs and saliva were positive in 20 (51.3%) participants. Ten participants (25.6%) had swab positive/saliva-negative results and 9 participants (23.1%) had saliva positive/swab-negative results. The percentage of positive and negative agreement of the saliva samples with the nasopharyngeal swab were 67% and 97% respectively with positive and negative predictive values as 69% and 96% respectively. The findings indicate that drooling saliva samples have good and comparable diagnostic accuracy to the nasopharyngeal swabs with moderate sensitivities and high specificities.
| S-EPMC9518879 | biostudies-literature
Project description:We evaluated saliva (SAL) specimens for SARS-CoV-2 RT-PCR testing by comparison of 459 prospectively paired nasopharyngeal (NP) or mid-turbinate (MT) swabs from 449 individuals with the aim of using saliva for asymptomatic screening. Samples were collected in a drive-through car line for symptomatic individuals (N=380) and in the emergency department (ED) (N=69). The percent positive and negative agreement of saliva compared to nasopharyngeal swab were 81.1% (95% CI: 65.8% - 90.5%) and 99.8% (95% CI: 98.7% - 100%), respectively. The sensitivity increased to 90.0% (95% CI: 74.4% - 96.5%) when considering only samples with moderate to high viral load (Cycle threshold (Ct) for the NP <=34). Pools of five saliva specimens were also evaluated on three platforms: bioMérieux NucliSENS easyMAG with ABI 7500Fast (CDC assay), Hologic Panther Fusion, and Roche COBAS 6800. The median loss of signal upon pooling was 2-4 Ct values across the platforms. The sensitivity of detecting a positive specimen in a pool compared with testing individually was 100%, 93%, and 95% for CDC 2019-nCoV Real-Time RT-PCR, Panther Fusion® SARS-CoV-2 assay, and cobas® SARS-CoV-2 test respectively, with decreased sample detection trending with lower viral load. We conclude that although pooled saliva testing, as collected in this study, is not quite as sensitive as NP/MT testing, saliva testing is adequate to detect individuals with higher viral loads in an asymptomatic screening program, does not require swabs or viral transport media for collection, and may help to improve voluntary screening compliance for those individuals averse to various forms of nasal collections.
| S-EPMC7553188 | biostudies-literature