Cycle Threshold Values Research Articles

Design, creation, and use of the Test Us Bank (TUB) COVID-19 sample biorepository.

Shortly after the first case of SARS-CoV-2 was diagnosed a public health emergency (PHE) was declared and a multi-agency response was initiated within the US federal government to create and propagate testing capacity. As part of this response, an unprecedented program designated Rapid Acceleration of Diagnostics (RADx) Tech was established by the National Institutes of Health (NIH) to facilitate the development of point-of-care tests for the COVID-19. The RADx Tech Clinical Studies Core (CSC), located at the University of Massachusetts Chan Medical School (UMass Chan), with partnering academic, private, and non-governmental organizations around the country, was tasked with developing clinical studies to support this work. This manuscript details development of a biorepository specifically focused on the collection and storage of samples designed for diagnostic platform development. It highlights the unified collection and annotation process that enabled gathering a diverse set of samples. This diversity encompasses the geography and backgrounds of the participants as well as sample characteristics such as variant type and RT-PCR cycle threshold (CT) value of the corresponding reference sample on a uniform clinical reference platform.

Molecular Detection of SARS-CoV-2 Viral Particles in Exhaled Breath Condensate via Engineered Face Masks

In this study, we present a novel face mask engineered for the collection of exhaled breath condensate (EBC) and its application and performance in a clinical study of COVID-19 infection status assessment versus the gold standard polymerase chain reaction (PCR) nasopharyngeal swab testing. EBC was collected within a clinical trial of COVID-19-infected and non-infected patients and analyzed by reverse transcription quantitative (RT-q) PCR, with the results being compared with nasopharyngeal sampling of the same patient. The cycle threshold (Ct) values of the nasopharyngeal samples were generally lower than those of EBC, with viral loads in EBC ranging from 1.2 × 104 to 5 × 108 viral particles mL−1 with 5 min of breathing. From the 60 clinical patients’ samples collected, 30 showed a confirmed SARS-CoV-2 infection. Of these 30 individuals, 22 (73%) had Ct values < 40 (representing the threshold for SARS-CoV-2 infectivity) using both amplification of ORF1a/b and the E-gene. The 30 EBC samples from non-infected participants were all identified as negative, indicating a 100% specificity. These first results encourage the use of the face mask as a noninvasive sampling method for patients with lung-related diseases, especially with a view to equipping the face mask with miniaturized sensing devices, representing a true point-of-care solution in the future.

Using polymerase chain reaction cycle threshold (PCR Ct) values for predicting and managing cholera outbreaks: potential, challenges, and future directions

Abstract Background Cholera continues to be a major public health threat, especially in regions with poor sanitation and limited access to clean water. Accurate prediction and management of cholera outbreaks are essential to reduce mortality and morbidity. This review focuses on the potential of polymerase chain reaction (PCR) cycle threshold (Ct) values as an innovative tool for early detection and control of cholera outbreaks, specifically highlighting its utility in predicting outbreaks and guiding public health responses. Main body of abstract PCR Ct values offer a significant advantage in the early detection of Vibrio cholerae, the pathogen responsible for cholera, in both clinical and environmental samples. By providing quantitative data on bacterial load, lower Ct values indicate a higher concentration of the pathogen, signaling the potential for increased disease transmission. These values allow for more precise, real-time monitoring of cholera outbreaks and aid in targeting intervention strategies such as water sanitation improvements, vaccination campaigns, and antibiotic treatments. Moreover, the application of Ct values in environmental surveillance, particularly in monitoring water sources, offers a proactive approach to prevent the spread of cholera by identifying contamination risks before human cases arise. However, the implementation of PCR in low-resource settings faces significant challenges, including high costs, the need for advanced laboratory infrastructure, and a lack of technical expertise. Addressing these barriers through cost-effective innovations and capacity building initiatives is crucial for optimizing the use of PCR Ct values in cholera management. Conclusion PCR Ct values hold great promise for improving cholera outbreak prediction and control. Despite challenges in adopting this technology in resource-limited settings, the integration of Ct value monitoring into public health frameworks can enhance early detection efforts and contribute to more effective cholera management strategies.

Comparison of QIAstat-Dx and BioFire FilmArray Gastrointestinal Panels in a Pediatric Population

Accurate laboratory diagnosis of gastroenteritis is important to ensure that patients receive appropriate treatment and proper isolation precautions. This study evaluated the performance of the QIAGEN QIAstat-Dx gastrointestinal panel (QGP) in comparison to the bioMerieux BioFire FilmArray gastrointestinal panel (BGP) for the detection of gastrointestinal pathogens in 110 pediatric patients being evaluated for gastroenteritis at our hospital. We compared 23 different bacterial, viral, and parasite enteropathogens detected by the QGP against the BGP. The overall positive percent agreement (PPA) for all compared targets was 96.2% and the overall negative percent agreement (NPA) for all compared targets was 99.7%. Our study shows that QIAstat-Dx QGP provides comparable results to the BioFire BGP in our pediatric population. Additionally, the PCR cycle threshold (Ct) value reported by the QGP is potentially a helpful tool in estimating the load of the detected pathogen in stool samples.

Effects of storage temperature on throat swabs preserved in different transport media for detection of SARS-CoV-2 by reverse transcriptase polymerase chain reaction

Objectives: The coronavirus disease 2019 (COVID-19) pandemic resulted in an increased need for molecular diagnostic testing. Delay in the specimen processing and storage of samples in laboratories leads to degradation of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) viral RNA. Inactivation transport medium (ITM) contains chaotropic agents that inactivate the virus and stabilize SARS-CoV-2 RNA for a longer duration, even at room temperature. The effect of different temperatures and duration of storage of samples in viral transport media (VTM) and ITM for detection of SARS-CoV-2 RNA was assessed. Materials and Methods: Samples from COVID-19 patients were aliquoted in ITM and VTM and kept at ambient temperature, 37°C and 45°C. SARS-CoV-2 viral RNA was extracted. Multiplex real-time polymerase chain reaction was done on days 0, 1, 3, and 5, and cycle threshold (Ct) values were noted. Statistical Analysis: Data were analyzed using the Statistical Package for the Social Sciences version 26.0. Linear variables were summarized as mean and standard deviations. One-way analysis of variance test with post hoc Tukey honestly significant difference was used to compare mean value between different loops and for pair-wise comparison. P < 0.05 was taken as significant. Results: The mean Ct values of both the Orf and E genes of the samples in VTM and ITM were stable across all temperature conditions on day 1. On day 5, the increase in Ct values for both E and Orf genes were significantly higher for VTM than ITM at ambient temperature, 37°C and 45°C. Ribonuclease P failure was significantly higher for VTM than ITM at ambient temperature and 37°C on day 3 and at all temperatures on day 5. Conclusions: ITM is a valuable transport media that can preserve SARS-CoV-2 for up to 5 days at ambient temperature and 37°C. As it renders the samples non-infectious, thus reducing the potential of biohazard events, this transport medium can be used effectively for the collection and transportation of SARS-CoV-2 samples, especially from remote or isolated healthcare facilities.

The Clinical Characteristics and Virus Shedding of Omicron Patients in a Shanghai Mobile Cabin Hospital

Background Information about characteristics of Omicron-infected patients and estimation of risk factors for the duration of viral shedding were unclear. Methods This retrospective study included asymptomatic patients and patients with mild symptoms, who were infected with Omicron and hospitalized in a Shanghai mobile cabin hospital. The clinical characteristics of those patients were summarized, and the influencing factors of severe acute respiratory syndrome coronavirus 2 virus shedding were explored. Results Multivariate logistic regression analysis indicated that comorbidities (odds ratio [OR], 1.991; 95% confidence interval [CI], 1.365–2.904; P = 0.000), 2 doses of vaccines (OR, 2.421; 95% CI, 1.636–3.584; P = 0.000), 3 doses of vaccine/booster shots (OR, 3.797; 95% CI, 2.607–5.529; P = 0.000), and virus shedding time (OR, 1.078; 95% CI, 1.053–1.103; P = 0.000) were associated with increased odds of mild symptoms in Omicron patients, whereas age (OR, 0.951; 95% CI, 0.940–0.963; P = 0.000) was associated with a reduced risk of mild symptoms. The median length of virus ribonucleic acid shedding time was 10 days. Being male (OR, 1.868; 95% CI, 1.488–2.345; P < 0.001), age (OR, 1.018; 95% CI, 1.008–1.028; P < 0.001), and a diagnosis of mild symptoms (OR, 1.508; 95% CI, 1.244–1.895; P < 0.001) were risk factors of prolonged shedding time in multivariate regression analysis. The cycle threshold values of nucleocapsid protein and open reading frame 1ab genes in Omicron patients with virus shedding time ≤10 days are significantly different from that in patients with virus shedding time >20 days (P < 0.001). Conclusions Being male, being older, and having a mild case were all risk factors for prolonged virus shedding time, and patients with high viral load may have long virus shedding time. Patients who are young, have complications, and have been vaccinated with 2 or 3 doses/enhanced vaccines with long virus shedding time increased the risk of clinical symptoms.

Effect of the renin-angiotensin-aldosterone system inhibitors on time to nucleic acid negative conversion in hypertensive patients with SARS-CoV-2 omicron infection: a propensity score matching study.

We examined the relationship between RAAS inhibitor use and Omicron infection in mildly symptomatic patients. This retrospective case-control observational study included 50,121 patients with mild Omicron infection from the largest "Fangcang" shelter hospital in Shanghai between April 9, 2022, and May 21, 2022. Using 1:1 propensity score matching (PSM), we classified 4394 COVID-19 patients into hypertension and non-hypertension groups, and 406 hypertensive patients into RAAS inhibitor and non-RAAS inhibitor groups. The risk of initial symptoms of infection, cumulative negative conversion rates, time to nucleic-acid negative conversion, and viral loads were compared. In the hypertension group, the median number of days for nucleic-acid negative conversion was 7.0 (IQR, 5.0-9.0), which was greater than non-hypertensive group (median (IQR) 6.0 (4.0-8.0), P < 0.001, Cohen's d = 0.29); the mean and minimum cycle threshold values (CT-values) were significantly lower (P < 0.001, Cohen's d = 0.23). In the RAAS inhibitors group, the median number of days for nucleic-acid negative conversion was 7.0 days (IQR, 5.0-9.0), which was shorter than the non-RAAS inhibitors group ([median (IQR)] 8.0 (6.0-10.0), P < 0.001, Cohen's d = 0.34), the cumulative negative conversion rates at 3-8 days being all higher than non-RAAS inhibitors groups (P < 0.05). The most significant difference in negative conversion rate between the RAAS inhibitor and non-RAAS inhibitor group was on the 4th day. No significant difference was observed in mean and minimum CT-values from RAAS inhibitor and non-RAAS inhibitor groups (P > 0.05). RAAS inhibitor use in patients with hypertension is associated with nucleic-acid negative conversion duration and negative conversion rate. RAAS inhibitors clearly do not aggravate or prolong COVID-19.

Implementation and Performance of a Point-of-Care COVID-19 Test Program in 4,000 California Schools

To evaluate the feasibility and accuracy of an unprecedented COVID-19 antigen testing program in schools, which required a healthcare provider order, laboratory director, a Clinical Laboratory Improvement Amendments certificate of waiver, as well as training of school personnel. Descriptive report of a point-of-care, school-based antigen testing program in California from August 1st, 2021 through May 30, 2022, in which participants grades K-12 self-swabbed and school personnel performed testing. Participants included 944 009 students, personnel, and community members from 4022 California kindergarten through high schools. Outcomes measured include sensitivity and specificity (with polymerase chain reaction [PCR] as comparator) of the Abbott BinaxNOW antigen test, number of tests performed, and active infections identified. Of 102 022 paired PCR/antigen tests, the overall sensitivity and specificity for the antigen test was 81.2% (95% CI: 80.5%-81.8%) and 99.6% (95% CI: 99.5%-99.6%), respectively, using cycle threshold values<30. During January through March 2022, the highest prevalence period, the positive predictive value of antigen testing was 94.7% and the negative predictive value was 94.2%. Overall, 4022 school sites were enrolled and 3 987 840 million antigen tests were performed on 944 009 individuals. A total of 162 927 positive antigen tests were reported in 135 163 individuals (14.3% of persons tested). Rapidly implementing a school-based testing program in thousands of schools is feasible. Self-swabbing and testing by school personnel can yield accurate results. On-site COVID-19 testing is no longer necessary in schools, but this model provides a framework for future infectious disease threats.

Role of admission rapid antigen testing (RATs) for COVID-19 on patients transferred from acute hospitals to a post-acute rehabilitation setting.

BackgroundRapid antigen tests (RATs) are suitable for point-of -care testing, require no laboratory time and give immediate results. However, are RATs useful for detecting asymptomatic COVID-19 infection when compared with polymerase chain reaction (PCR) testing in healthcare settings? AimThe aim of this study was to implement a reliable testing system utilising RATs to promptly detect COVID-19 infection in predominantly asymptomatic patients transferred from acute hospitals to a post-acute rehabilitation unit (PARU). MethodsRAT testing was carried out on all new admissions without a history of confirmed Covid-19 infection within three months of admission. PCR testing was carried out on all patients with a positive RAT for confirmation purposes. The cycle threshold (Ct) values of COVID-19 detected results on PCR testing were examined to determine the utility of the RATs. ResultsA total of 1,403 patients were transferred to the PARU from January to December 2023. The results of the study revealed an 85% accuracy of RATs with a 15% rate of false negative results at the time of admission. All patients that had a positive RAT at the time of admission also had a positive PCR test. ConclusionThis testing algorithm resulted in early detection and prompt isolation of positive cases reducing the likely spread of COVID-19 infection, hospital outbreaks and bed/ward closures.

Effects of energy-gathered and energy-divergent ultrasound on structure, DNA extraction and adulterated quantification of sweet potato vermicelli and its starch.

The identification of adulterated sweet potato vermicelli faces significant challenges, seriously hindering the development of the vermicelli industry. Herein, we investigate effects of energy-gathered ultrasound (EGU) and energy-divergent ultrasound (EDU) (30, 40 and 50 W L-1) on structure, DNA extraction and adulterated quantification of sweet potato vermicelli and its starch, thereby exploring their potential in adulteration of sweet potato vermicelli. EGU-assisted modified cetyltrimethylammonium bromide (CTAB) protocol with β-mercaptoethanol significantly improved DNA extraction from sweet potato vermicelli (223.7-249.2 ng μL-1) and its starch (133.4-186.4 ng μL-1), followed by EDU-assisted DNA extraction from sweet potato vermicelli (115.1-209.3 ng μL-1) and its starch (33.4-61.0 ng μL-1). Both EGU and EDU treatments resulted in the destruction of microstructure and crystalline structure, as well as changes in pasting and thermal properties of sweet potato vermicelli and its starch. Real-time polymerase chain reaction (PCR) amplification results revealed that EGU and EDU enhanced the efficiency of DNA amplification, and EDU showed smaller cycle threshold (Ct) values than EGU. In addition, EDU-assisted CTAB protocol combined with real-time PCR could detect levels of less than 1% of cassava and maize starches in sweet potato vermicelli. EDU-assisted CTAB protocol combined with real-time PCR shows promise as a sensitive and reliable analytical tool for vermicelli adulteration. © 2024 Society of Chemical Industry.

Using intrahost single nucleotide variant data to predict SARS-CoV-2 detection cycle threshold values.

Over the last four years, each successive wave of the COVID-19 pandemic has been caused by variants with mutations that improve the transmissibility of the virus. Despite this, we still lack tools for predicting clinically important features of the virus. In this study, we show that it is possible to predict the PCR cycle threshold (Ct) values from clinical detection assays using sequence data. Ct values often correspond with patient viral load and the epidemiological trajectory of the pandemic. Using a collection of 36,335 high quality genomes, we built models from SARS-CoV-2 intrahost single nucleotide variant (iSNV) data, computing XGBoost models from the frequencies of A, T, G, C, insertions, and deletions at each position relative to the Wuhan-Hu-1 reference genome. Our best model had an R2 of 0.604 [0.593-0.616, 95% confidence interval] and a Root Mean Square Error (RMSE) of 5.247 [5.156-5.337], demonstrating modest predictive power. Overall, we show that the results are stable relative to an external holdout set of genomes selected from SRA and are robust to patient status and the detection instruments that were used. This study highlights the importance of developing modeling strategies that can be applied to publicly available genome sequence data for use in disease prevention and control.

Accuracy of point-of-care SARS-CoV-2 detection using buccal swabs in pediatric emergency departments.

To optimize the identification of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-infected children, specimen collection and testing method are crucial considerations. Ideally, specimen collection is easy and causes minimal discomfort, and the laboratory approach is simple, accurate, and rapid. In this prospective cohort study we evaluated the accuracy of a point-of care nucleic acid device using caregiver/patient self-collected buccal swabs. Participants were recruited in 14 Canadian tertiary care pediatric emergency departments. Children <18 years of age deemed to require SARS-CoV-2 testing were eligible. Caregivers or the patient-collected buccal swabs which were tested on the ABBOTT ID NOW. The reference standard was nasopharyngeal swab specimens collected by a healthcare provider tested via laboratory reverse transcription PCR (RT-PCR). We enrolled 2,640 study participants and 14.4% (381/2,640) were SARS-CoV-2 RT-PCR-positive. Eight percent (223/2,640) and 85.0% (2,244/2,640) were concordant test-positive and concordant test-negative, respectively. Sensitivity and specificity of the investigational approach were 58.5% [95% confidence interval (CI): 53.4, 63.5] and 99.3% (95% CI: 98.9, 99.6), respectively. Cycle threshold values were lower among concordant [median 17 (15, 21)] relative to discordant [median 30 (22, 35)] swabs (P < 0.001). Sensitivity was greatest among children <4 years of age, when caregivers performed the swabs, among unvaccinated children, and those with shorter symptom duration. Across multiple pain measures, less pain was associated with buccal swab testing. Although accuracy of the buccal swab point-of-care SARS-CoV-2 test was good and negative agreement was excellent, sensitivity was only 58.5%. Concordance was greater among those with higher viral loads, and the approach involving buccal swabs was less painful.IMPORTANCETo optimize the identification of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-infected children, specimen collection and testing method are crucial considerations. Ideally, specimen collection is easy and causes minimal discomfort, and laboratory approach is simple, accurate, and rapid. We evaluated the accuracy and pain associated with buccal swab specimen collection by caregivers or children themselves who were tested using a point-of-care isothermal nucleic acid amplification SARS-CoV-2 test. This novel approach was compared to nasopharyngeal swab specimens tested using laboratory-based PCR tests. While negative agreement was excellent, positive percent agreement was less than 60%. Concordance was greater among those with higher viral loads, and thus, sensitivity is excellent when transmissibility is more likely to occur. Importantly, the approach involving buccal swabs was significantly less painful, and thus, children and their caregivers are more likely to agree to testing using such an approach.CLINICAL TRIALSRegistered at ClinicalTrials.gov (NCT05040763).

Examining Cycle Threshold values in Repeat SARS-CoV-2 Cases: A Retrospective Analysis in Ireland

Abstract Background Effective public health management during the COVID-19 pandemic requires accurate differentiation between re-infections and residual viral RNA detections in repeat positive cases. This study investigates cycle threshold (CT) trends in repeat SARS-CoV-2 cases to discern between persistent viral remnants and active infections. Collaboration between contact tracing and laboratories is essential for shaping robust public health strategies. Methods A retrospective analysis was conducted on 427 individuals with repeat positive PCR tests. Data was collected at a contact tracing centre from March to August 2021. Key variables including age, gender, and biomedical markers (test intervals, CT values) were examined. Regression and correlation techniques assessed CT trends and their implications. Results Younger individuals (&amp;lt;60 years) occasionally exhibited significantly lower CT values in subsequent tests, suggesting potential re-infections, while older adults generally displayed higher CT values indicative of residual RNA. These age-specific CT trends were statistically significant and instrumental in refining testing protocols and isolation guidelines. Further analysis is expected to strengthen these findings Conclusions Collaborative approaches between contact tracing and laboratories are vital in shaping robust public health strategies for distinguishing between types of repeat infections. Revision of health policies is imperative to prevent unnecessary isolation and optimise healthcare workforce allocation during ongoing transmission periods. Key messages • Decisive decision-making skills are crucial in managing repeat SARS-CoV-2 infections. • Revised health policies align testing and isolation practices with empirical CT data, enhancing public health efficacy.

Characterization of the viral genome of Omicron variants of SARS-CoV-2 circulating in Tripura, a remote frontier state in Northeastern India.

From 2020, with advent of COVID-19 pandemic, Tripura has experienced SARS-CoV-2 viral evolution in accordance with other parts of India. Since January 2022, the Omicron variant of SARS-CoV-2 virus became the predominant lineage circulating in India and neighboring countries. This study characterizes the viral genome of the omicron variant circulating in the state since its inception to June, 2023. The current study was performed on nasopharyngeal and oropharyngeal samples received from the various departments of AGMC, as well as eight district hospitals from Tripura. The positive samples with a cycle threshold value of 25 or less for the E and/or N gene were considered for whole genome sequencing using Illumina Miseq NGS platform. Majority of the sequences belonged to Clade 21L, with BA.5.2 being the major sub variant detected during the study period. Majority of the mutations were detected in the Spike protein region, including L24-, P25-, P26-, V213G, G339D, S371F, S373P, S375F, T376A, D405N, R408S, K417N, S477N, T478K, Q498R, Y505H, Q954H and N969K. All the sequences uniquely showed the mutations A27S and G142D in N terminal domain in Spike protein, not being reported from other Indian sequences like BA.5 variants. T9I, A63T and P13L were major substitutions in E, M and N protein regions respectively. Escape of mutants from vaccine induced immunity was mostly observed in BA.2 sub variants, majority endowed with the triplet mutation of K417N + E484K + N501Y. The current study indicates that Omicron variants circulating in the state of Tripura is comparable to other regions of India and the neighbouring country of Bangladesh. Genetic mutations increasing viral transmissibility have been identified in the circulating viral genomes.

Evaluation of Fecal Sample Pooling for Real-Time RT-PCR Testing SARS-CoV-2 in Animals

During the COVID-19 pandemic, veterinary diagnostic laboratories tested both human and animal samples and needed to ensure that they could accurately perform large numbers of diagnostic tests in a timely manner. Sample pooling, a methodology used effectively for over 80 years as a surveillance tool for screening large numbers of potentially infected individuals, was employed. Given its sensitivity, real-time polymerase chain reaction (PCR) is more suitable for employing this strategy, as compared to other less sensitive testing methods. In this study, we evaluated the capability of detecting SARS-CoV-2 in both 5-sample and 10-sample pools of feces using real-time reverse transcriptase polymerase chain reaction (rRT-PCR) as well as determined the level of sensitivity. A blinded method test (BMT) by an independent laboratory was conducted to assess the five-sample fecal pool. To complement detection capability, the stability of the genome within a PBS fecal suspension was measured under various time and temperature conditions across a 28-day period. Our results showed that the limit of detection for 5-sample and 10-sample fecal pools is 12.8 and 6.4 genome copies in a 25 µL PCR, respectively. The 5-sample and 10-sample pooling resulted in a cycle threshold (Ct) value loss of 2.35 and 3.45, as compared to Ct values of known positive individual samples, but consistent detection was still achieved in pools containing positive samples with an original Ct below 36 and 34, respectively. The simulation of clinical five-sample pooling showed that all positive samples could be detected regardless of the number (1–3) of positive samples in each pool. The BMT results demonstrated excellent sensitivity (100 copies/reaction) in five-sample pools for the detection of SARS-CoV-2 RNA even though a fecal matrix effect was observed. Finally, our results show that the SARS-CoV-2 genome remains stable over a wide range of time and temperature variations. Overall, our findings provide solid data to scale up SARS-CoV-2 testing capacity in veterinary diagnostic laboratories.

Molecular Detection of Anaplasma marginale in Capybara (Hydrochoerus hydrochaeris) from Corrientes, Argentina.

Monitoring wildlife health is essential for understanding global disease patterns, particularly as vector-borne infections extend the geographic ranges and thereby hosts due to environmental shifts. Anaplasma marginale, primarily impacting cattle, has economic implications and has been found in diverse hosts, yet its presence in capybaras (Hydrochoerus hydrochaeris), influential in tick-borne pathogen spread, lacks comprehensive understanding. From 2015 to 2022, 14 capybaras were surveyed across two different areas of northeastern Argentina. In 1 of 14 (7%) capybaras, the presence of A. marginale was confirmed through the amplification of specific genes, msp5 and msp1β. In addition, A. marginale DNA was detected in the capybara's blood sample through quantitative PCR, with a cycle threshold value of 30.81 (800 copies per reaction). Amplification of a fragment of the msp1α gene revealed PCR products of three different sizes, suggesting the presence of at least three coinfecting A. marginale variants in the capybara host. This study suggests that capybaras are wild hosts for A. marginale in the Ibera Wetlands in Argentina, potentially influencing the infection dynamics of both domestic and wild species. This finding highlights the necessity for thorough studies on the role of capybaras in disease dynamics, crucial for understanding wildlife health and the spread of disease.

Cycle Threshold Values of SARS-CoV-2 RT-PCR during Outbreaks in Nursing Homes: A Retrospective Cohort Study.

a retrospective cohort design with Poisson regression and multinomial logistic regression were used. We studied four nursing home SARS-CoV-2 outbreaks, and the average infection rate, reinfection rate, and case fatality were 72.7%, 19.9%, and 5.5%, respectively; 98.9% of residents were vaccinated with three doses of a mRNA SARS-CoV-2 vaccine. Ct values for first infections and reinfections were 27.1 ± 6.6 and 31.9 ± 5.4 (p = 0.000). Considering Ct values ≥ 30 versus <30, residents with reinfections had Ct values higher than residents with a first infection, an adjusted relative risk of 1.66 (95% Confidence interval 1.10-2.51). A sensitivity analysis confirmed these results. Reinfection and SARS-CoV-2 vaccination (hybrid immunity) could protect against severe disease better than vaccination alone. High Ct values suggest lower transmission and severity. Its value can be useful for surveillance and forecasting future SARS-CoV-2 epidemics.

Evaluation of the Seegene Allplex™ RV master assay for one-step simultaneous detection of eight respiratory viruses in nasopharyngeal specimens

BackgroundThe Seegene Allplex™ RV Master (RVM) assay is a one-step multiplex real-time reverse transcription polymerase chain reaction (RT-PCR) system for detecting eight viral respiratory pathogens from nasopharyngeal swab, aspirate, and bronchoalveolar lavage specimens. The eight RVM targets are: severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), Influenza A (Flu A), Influenza B (Flu B), Human respiratory syncytial virus (RSV), adenovirus (AdV), rhinovirus (HRV), parainfluenza virus (PIV), and metapneumovirus (MPV). The assay is based on Seegene’s multiple detection temperature (MuDT) technology and provides cycle threshold (Ct) values for each of its viral targets upon PCR completion. ObjectiveWe aimed to evaluate the diagnostic performance of the RVM assay by calculating sensitivity, specificity, accuracy, Positive Predictive Value (PPV), Negative Predictive Value (NPV), Positive Percent Agreement (PPA), Negative Percent Agreement (NPA), and Overall Percent Agreement (OPA) compared to definite diagnosis and analogous reference assays. Study designDiagnostic sensitivity, specificity, accuracy, PPV, and NPV were calculated by comparing the results of the RVM assay to that of definite diagnosis assays; while PPA, NPA, and OPA were calculated by comparing results of the RVM assay to that of analogous reference products. Definite diagnosis and reference methods comprised whole genome sequencing and PCR genotyping, the Allplex™ SARS-CoV-2/FluA/FluB/RSV and Respiratory Panels 1, 2, and 3 assays (Seegene), and the Xpert® Xpress SARS-CoV-2/FluA/FluB/RSV Plus assay (Cepheid). Reproducibility of the RVM assay using fully-automated and semi-automated nucleic acid (NA) extraction workflows and as performed by independent operators was also assessed. In total, 249 positive respiratory specimens and at least 50 negative specimens for each target tested were used for this evaluation study. ResultsSensitivity, specificity, accuracy, PPV, NPV, PPA, NPA, and OPA ranged from 95.7 % to 100 % for detecting all eight targets tested on the RVM assay. Reproducibility PPA, NPA, and OPA between automated and semi-automated NA extraction workflows were all >97.9 %, while the reproducibility PPA, NPA and OPA between independent operators were all 100 %. ConclusionThese results demonstrate a high level of sensitivity, specificity, accuracy and diagnostic predictive value of the RVM assay and high agreement with comparable reference assays in identifying all eight of its targets. Taken together, our study underscores the diagnostic utility of the RVM assay in detecting eight viral respiratory pathogens.

Correlation of COVID-19 vaccination and RT-PCR ct value among cases in Addis Ababa, Ethiopia: implication for future preparedness

BackgroundThe COVID-19 disease requires accurate diagnosis to effectively manage infection rates and disease progression. The study aims to assess the relationship between vaccination status and RT-PCR cycle threshold (Ct) values by comparing clinical, RDT and RT-PCR results.MethodsA total of 453 suspected COVID-19 cases were included in this study. Nasopharyngeal swabs were collected for both RDT and RT-PCR testing, with RDTs conducted on-site and RT-PCR at the Ethiopian Public Health Institute (EPHI) genomics laboratory. Detailed clinical, RDT, and RT-PCR results were analyzed. Data analysis included descriptive statistics, cross-tabulation, and Chi-Square tests to investigate the connections between diagnostic outcomes and vaccination status, with a focusing on Ct values.ResultsRDT results showed 34.0% negative and 65.8% positive, while RT-PCR results indicated 35.8% negative and 64.2% positive cases. The discrepancies between RDT and RT-PCR results emphasize the importance of thorough testing. No significant association was found between vaccination status and viral load, as indicated by Ct values. Among RT-PCR positive cases, 49.8% had been vaccinated, suggesting challenges in interpreting results among vaccinated individuals. Further analysis revealed that vaccination (first or second dose) had minimal impact on Ct values, indicating limited influence of vaccination status on viral load dynamics in infected individuals.ConclusionsThe study highlights the significant differences between RDT and RT-PCR outcomes, underscoring the need for a comprehensive testing approach. Additionally, the findings suggest that vaccination status does not significantly impact RT-PCR Ct values, complicating the interpretation of diagnostic results in vaccinated individuals, especially in breakthrough infections and potential false positives.

Assessing infectiousness and the impact of effective treatment to guide isolation recommendations for people with pulmonary tuberculosis.

Determining the extent and duration of infectiousness of individuals with pulmonary tuberculosis (TB) is critical for various aspects of TB care, including decisions regarding isolation. Studies suggest considerable heterogeneity in infectiousness of people with pulmonary TB. Pre-treatment, measures of bacillary burden including sputum smear microscopy, culture time-to-positivity, and Xpert MTB/RIF cycle threshold (Ct) value, predict the risk of transmission to contacts. Index patients with smear negative disease pose lower infectious risk than those who have smear-positive disease, and household contact infection is more likely with index patients who have lower Xpert Ct values. Newer tools that enable detection of Mycobacterium tuberculosis complex (Mtb complex) from cough aerosol sampling and face mask sampling may be better predictors of contact infection risk. Clinical factors such as cough strength and frequency, and presence of cavitation on chest imaging, may also assist with risk prediction. Post-treatment, smear and culture status are poor predictors of infectiousness. While the exact duration of infectiousness post treatment initiation remains uncertain, data from human-to-guinea pig transmission studies and clinical studies suggest effective treatment results in a rapid decline in infectiousness, irrespective of smear or culture conversion. This is largely supported by early bactericidal activity and transcriptomic studies, and cough aerosol sampling studies, although a subset of patients may have persistent cough aerosol positivity. These findings can enable a more nuanced approach to isolation decision making, while further research studies are awaited.