Cycle Threshold Research Articles

SARS-CoV-2 Polymerase Chain Reaction Cycle Threshold Trends in Patients Who Are Immunocompromised and Implications for Isolation Precautions.

Among 495 patients who were immunocompromised and tested positive for SARS-CoV-2, polymerase chain reaction cycle thresholds remained <33 beyond 20 days more frequently in patients with hematologic malignancies, particularly those receiving B-cell-depleting or Bruton tyrosine kinase inhibitor therapy, as compared with those with solid organ malignancy (26% vs 5%).

Saliva Is a Sensitive and Accessible Sample Both for SARS-CoV-2 Detection and for the Evaluation of Treatment Effectiveness in Follow-Up Studies.

Despite emerging evidence indicating that molecular SARS-CoV-2 tests performed on saliva have diagnostic sensitivity and specificity comparable to those observed with nasopharyngeal swabs (NPSs), most in vivo follow-up studies on the efficacy of drugs against SARS-CoV-2 have been performed on NPSs, not considering saliva as a possible alternative matrix. For this reason, in this study, we used, in parallel, saliva and NPS samples for the detection of SARS-CoV-2 by real-time RT-PCR in patients receiving Tixagevimab/Cilgavimab, Nirmatrelvir/Ritonavir, or Sotrovimab as a treatment against SARS-CoV-2. Our results showed a good correlation between the NPS and saliva samples for each drug; moreover, comparable changes in the cycle threshold (Ct) levels in saliva and NPSs were observed both 7 days and 30 days after treatment, thus confirming that the saliva represents a good matrix for in vivo follow-up studies verifying the effectiveness of treatments against SARS-CoV-2.

Clinical features and viral load variations of Mpox: a retrospective study in Chongqing, China

PurposeSince May 2022, Mpox has spread extensively outside of Africa, posing a serious threat to the health of people globally, and particularly to the men who have sex with men (MSM) population. Chongqing, a province in Southwest China, has relatively large MSM and people living with HIV (PLWH) populations, presenting conditions conducive to the wide dissemination of Mpox. In this study, we investigated the clinical characteristics of Mpox patients among MSM and PLWH in Chongqing, aiming to inform the development of targeted prevention, control, and treatment strategies for Mpox.MethodWe evaluated the clinical characteristics, travel history, time of onset, distribution and number of skin lesions of Mpox patients admitted to the Chongqing Public Health Medical Center between September 2022 and October 2023. Meanwhile, a series of clinical samples were collected and the pathogen of interest was identified as Mpox virus using quantitative polymerase chain reaction (qPCR). The results were presented in the form of cycle thresholds (Ct), which help to approximate the quantification of viral load.ResultsAs of October 11, 2023, the Chongqing Public Health Medical Center reported a total of nine Mpox virus infections. All the patients identified were male and belonged to the MSM population, among whom seven (77.8%) were living with HIV, and maintained a preserved immune system while achieving viral suppression via effective ART. We observed no discernible clinical differences between MSM with Mpox with or without HIV, and no fatalities were recorded. Viral loads were observed to be higher in samples taken from the skin than those from the throat, nasopharynx, blood, or semen.ConclusionIn this retrospective study, the clinical manifestations of MPXV infection appeared consistent among MSM patients, regardless of HIV status. Elevated MPXV viral loads in the skin and mucosal tissues, particularly at genital and anal sites, indicate that transmission is more likely to occur via direct physical contact as opposed to respiratory pathways or through exposure to bodily fluids.

Effectiveness of Lianhua Qingwen Granule and Jingyin Gubiao Prescription in Omicron BA.2 Infection and Hospitalization: A Real-World Study of 56,244 Cases in Shanghai, China.

To examine the effectiveness of Chinese medicine (CM) Lianhua Qingwen Granule (LHQW) and Jingyin Gubiao Prescription (JYGB) in asymptomatic or mild patients with Omicron infection in the shelter hospital. This single-center retrospective cohort study was conducted in the largest shelter hospital in Shanghai, China, from April 10, 2022 to May 30, 2022. A total of 56,244 asymptomatic and mild Omicron cases were included and divided into 4 groups, i.e., non-administration group (23,702 cases), LHQW group (11,576 cases), JYGB group (12,112 cases), and dual combination of LHQW and JYGB group (8,854 cases). The length of stay (LOS) in the hospital was used to assess the effectiveness of LHQW and JYGB treatment on Omicron infection. Patients aged 41-60 years, with nadir threshold cycle (CT) value of N gene <25, or those fully vaccinated preferred to receive CM therapy. Before or after propensity score matching (PSM), the multiple linear regression showed that LHQW and JYGB treatment were independent influence factors of LOS (both P<0.001). After PSM, there were significant differences in LOS between the LHQW/JYGB combination and the other groups (P<0.01). The results of factorial design ANOVA proved that the LHQW/JYGB combination therapy synergistically shortened LOS (P=0.032). Patients with a nadir CT value <25 were more likely to accept CM. The LHQW/JYGB combination therapy could shorten the LOS of Omicron-infected individuals in an isolated environment.

A Targeted LC-MRM3 Proteomic Approach for the Diagnosis of SARS-CoV-2 Infection in Nasopharyngeal Swabs

Since its first appearance, severe acute respiratory syndrome coronavirus 2 quickly spread around the world and the lack of adequate PCR testing capacities, especially during the early pandemic, led the scientific community to explore new approaches such as mass spectrometry (MS). We developed a proteomics workflow to target several tryptic peptides of the nucleocapsid protein. A highly selective multiple reaction monitoring-cubed (MRM3) strategy provided a sensitivity increase in comparison to conventional MRM acquisition. Our MRM3 approach was first tested on an Amsterdam public health cohort (alpha-variant, 760 participants) detecting viral nucleocapsid protein peptides from nasopharyngeal swabs samples presenting a cycle threshold value down to 35 with sensitivity and specificity of 94.2% and 100.0%, without immunopurification. A second iteration of the MS-diagnostic test, able to analyze more than 400 samples per day, was clinically validated on a Leiden-Rijswijk public health cohort (delta-variant, 2536 participants) achieving 99.9% specificity and 93.1% sensitivity for patients with cycle threshold values up to 35. In this manuscript, we also developed and brought the first proof of the concept of viral variant monitoring in a complex matrix using targeted MS.

The use of real-time polymerase chain reaction for detection of raw rat meat (Rattus norvegicus) with species-specific primer for halal authentication

Rat meat (RM) can be obtained freely from farmers and can be used as a potential adulterant in meat-based food products. The presence of RM in any food products is prohibited by the Muslim community and is considered non-halal meat. Therefore, an analytical method capable of analysing RM specifically is very urgent. In this study, a real -time polymerase chain reaction using a species-specific primer targeting NADH dehydrogenase subunit 6 (ND6) is applied to analyze RM. DNA extraction was carried out using FavorPrepTM Tissue Genomic DNA Extraction Mini Kit, and the extracted DNA was subjected to purity index using NANO-Quant SPARK TECAN. The annealing temperature (Ta) used for PCR analysis was optimized to get the best Ta capable of providing the optimum amplification reaction. Furthermore, some performance characteristics were evaluated for Real-Time PCR, including sensitivity, efficiency and repeatability. The results exhibited that the designed primer is specific to rat DNA extracted toward other DNAs from meats typically used in food products. For quantitative evaluation, the limit of detection found was 0.39 ng with an efficiency of amplification (E) of 94.4% with a coefficient of determination (R2 ) of 0.986 for the relationship between log concentrations of DNA and cycle threshold (Ct) values. The repeatability assay was acceptable with an RSD value ≤ 25%. The developed method was reliable, provided high sensitivity for the analysis of RM. Therefore, this method is suitable to be used as a standard method for halal authentication analysis.

Hepatitis C virus diversity and treatment outcomes in Benin: a prospective cohort study

10 million people are chronically infected with the hepatitis C virus (HCV) in sub-Saharan Africa. The assessment of viral genotypes and treatment response in this region is necessary to achieve the WHO target of worldwide elimination of viral hepatitis by 2030. We aimed to investigate the prevalence of HCV genotypes and outcomes of treatment with direct-acting antiviral agents in Benin, a country with a national HCV seroprevalence of 4%. This prospective cohort study was conducted at two referral hospitals in Benin. Individuals were eligible for inclusion if they were seropositive for HCV and willing to consent to participation in the study; exclusion criteria were an inability to give consent or incarceration. Viraemia was confirmed by PCR. The primary outcomes were to identify HCV genotypes and measure sustained virological response rates 12 weeks after completion of treatment (SVR12) with a 12-week course of sofosbuvir-velpatasvir or sofosbuvir-ledipasvir, with or without ribavirin. We conducted phylogenetic and resistance analyses after the next-generation sequencing of samples with a cycle threshold (Ct) value of 30 or fewer cycles. The in-vitro efficacy of NS5A inhibitors was tested using a subgenomic replicon assay. Between June 2, 2019, and Dec 30, 2020, 148 individuals were screened for eligibility, of whom 100 were recruited prospectively to the study. Plasma samples from 79 (79%) of the 100 participants were positive for HCV by PCR. At the time of the study, 52 (66%) of 79 patients had completed treatment, with an SVR12 rate of 94% (49 of 52). 57 (72%) of 79 samples had a Ct value of 30 or fewer cycles and were suitable for whole-genome sequencing, from which we characterised 29 (51%) samples as genotype 1 and 28 (49%) as genotype 2. Three new genotype 1 subtypes (1q, 1r, and 1s) and one new genotype 2 subtype (2xa) were identified. The most commonly detected subtype was 2d (12 [21%] of 57 samples), followed by 1s (eight [14%]), 1r (five [9%]), 1b (four [7%]), 1q (three [5%]), 2xa (three [5%]), and 2b (two [3%]). 20 samples (11 genotype 2 and nine genotype 1) were unassigned new singleton lineages. 53 (93%) of 57 sequenced samples had at least two resistance-associated substitutions within the NS5A gene. Subtype 2d was associated with a lower-than-expected SVR12 rate (eight [80%] of ten patients). For one patient, with subtype 2b, treatment was not successful. This study revealed a high SVR rate in Benin among individuals treated for HCV with sofosbuvir-velpatasvir, including those with highly diverse viral genotypes. Further studies of treatment effectiveness in genotypes 2d and 2b are indicated. Medical Research Council, Wellcome, Global Challenges Research Fund, Academy of Medical Sciences, and PHARMBIOTRAC.

Use of PCR Cycle Threshold and Clinical Interventions to Aid in the Management of Pediatric Clostridioides difficile Patients

Better diagnostic tools are needed to improve the diagnosis of Clostridioides difficile infections (CDI) and reduce the overtreatment of colonized children. In this study, we evaluated two polymerase chain reaction (PCR) assays (Cepheid GeneXpert C. difficile and the Gastroenteritis PCR Panel by QIAstat-Dx) as a standalone method in combination with the PCR cycle threshold (Ct) value in positive samples to predict the presence of free toxins. We also evaluated the clinical impact of reporting toxin production results and provided comments alongside the PCR results in our pediatric population. PCR-positive stool samples from pediatric patients (aged 2 to 18 years old) were included in our study and tested for the presence of toxins A and B using the C. difficile Quik Chek Complete kit. For the clinical intervention, the CDI treatment rates 6 months pre- and post-intervention were compared. The use of PCR Ct value showed excellent sensitivity (100%) at a Ct value cutoff of 26.1 and 27.2 using the Cepheid GeneXpert C. difficile and the Gastroenteritis PCR Panel by QIAstat-Dx, respectively, while the toxin test showed inferior sensitivity of 64% in the PCR-positive samples. In addition, CDI treatment rates were decreased by 23% post-intervention. The results of our study suggest that nucleic acid amplification test (NAAT) assays supplemented by the use of PCR Ct value for positive samples can be used as standalone tests to differentiate CDI from colonization. Furthermore, the reporting of toxin production along with the PCR results can help reduce the unnecessary treatment of colonized children.

Self-swabbing versus assisted swabbing for viral detection by qRT-PCR: the experience from SARS-CoV-2 based on a meta-analysis of six prospectively designed evaluations conducted in a UK setting

PurposeIn April 2020, the UK Government implemented NHS Test and Trace to provide SARS-CoV-2 quantitative reverse transcription polymerase chain reaction (qRT-PCR) testing for the public, with nose-and-throat swabbing for samples performed by trained staff. Self-swabbing (SS) would allow rapid scale-up of testing capacity and access. Six studies were undertaken to determine whether SS was as effective for detecting SARS-CoV-2 as swabbing performed by trained staff.MethodsSix prospective studies were conducted between April–October 2020, using six swab/media combinations. Differences between assisted swabbing (AS) and SS were evaluated for concordance, positivity, sensitivity, cycle threshold (Ct) values and void rates. Statistical analysis was performed using 95% confidence intervals (CIs), paired t-tests and model-based methods.ResultsOverall, 3,253 individuals were recruited (median age 37 years, 49% female), with 2,933 having valid paired qRT-PCR results. Pooled concordance rate was 98% (95% CI: 96%, 99%). Positivity rate differences for SS (8.1%) and AS (8.4%) and differences in pooled sensitivities between SS (86%; 95% CI: 78%, 92%) and AS (91%; 95% CI: 78%, 96%) were nonsignificant. Both types of swabbing led to pooled void rates below 2% and strongly correlated Ct values. Age, sex and previous swabbing experience did not have a significant impact on concordance or sensitivity.ConclusionThe UK adopted a policy to promote self-testing for SARS-CoV-2 based on data demonstrating equivalence of SS versus AS. Positive outcomes with SS are likely generalisable to testing for other respiratory pathogens, and we consider self-sampling and self-testing essential for future pandemic preparedness.

Evaluation of silica spin‑column and magnetic bead formats for rapid DNA methylation analysis in clinical and point‑of‑care settings.

Late-stage cancers lack effective treatment, underscoring the need for early diagnosis to improve prognosis and decrease mortality rates. Molecular markers, such as DNA methylation, offer promise in early cancer detection. The present study compared commercial kits for analyzing DNA from cervical liquid cytology samples in cancer screening. Rapid bisulfite conversion kits using silica spin-columns and magnetic beads were assessed against standard DNA extraction and bisulfite conversion methods for profiling DNA methylation using quantitative methylation-specific PCR. β-actin amplification indicated the suitability of small sample volumes for methylation studies using either the pellet or supernatant (cell-free DNA) parts. Comparison of Bisulfite Conversion Kit-Whole Cell (Abcam), Methylamp Bisulfite Modification (Epigentek), EpiTect Fast LyseAll Bisulfite Kit (Qiagen GmbH) and EZ DNA Methylation-Direct Kit (Zymo Research Corp.) showed no significant differences in β-actin cycle threshold values. EZ-96 DNA Methylation-Lightning MagPrep (Zymo Research Corp.), a hybrid kit in a 96-well plate format, exhibited swift turnaround time and similar amplification efficiency. Automation with magnetic bead kits increased throughput without compromising amplification efficiency in open PCR systems. Cost analysis favored direct kits over the gold standard manual protocol. This comparison aids in selecting cost-effective DNA methylation diagnostic tests. The present study confirmed comparable kit performance in methylation-based analysis, highlighting the adequacy of cytology samples and the potential of bodily fluids as alternatives for liquid biopsy.

Impact of Laboratory Biomarkers on COVID-19 Severity: First Cross-sectional Study in a Remote Area of Pakistan

Objectives: The study aimed to evaluate the performance of rapid antigen test (RAT) and reverse transcription polymerase chain reaction (RT-PCR) in detecting COVID-19 and the impact of laboratory biomarkers on the severity of the disease. Material and Methods: A total of 150 nasopharyngeal swabs and blood samples were collected from symptomatic COVID-19 patients in Tehsil head-quarter Hospital, Dargai from August 2021 to March 2022. Results: RAT revealed a sensitivity of 86.67% and specificity of 100%, while RT-PCR revealed a sensitivity of 93.33% and specificity of 100%. The highest infection rate was found in males, and the average age of patients was 53.87 years. The most common symptoms were fever and sore throat. Patients were categorized into four groups based on cycle threshold values and blood biomarkers: mild, moderate, severe, and critical. White blood cell count, platelet count, and C-reactive protein were significantly different between the groups. Lactate dehydrogenase, D-dimer, and serum ferritin were significantly increased in critical patients. The receiver operating characteristic curve showed that inflammatory biomarkers had a comparative performance in predicting disease severity in COVID-19 patients. Conclusion: Blood biomarkers are associated with the disease severity in COVID-19 patients, and further studies, such as metabolomics, are recommended to explore the immunological mechanisms behind these biomarkers.

Artificial intelligence-based radiographic extent analysis to predict tuberculosis treatment outcomes: a multicenter cohort study

Predicting outcomes in pulmonary tuberculosis is challenging despite effective treatments. This study aimed to identify factors influencing treatment success and culture conversion, focusing on artificial intelligence (AI)-based chest X-ray analysis and Xpert MTB/RIF assay cycle threshold (Ct) values. In this retrospective study across six South Korean referral centers (January 1 to December 31, 2019), we included adults with rifampicin-susceptible pulmonary tuberculosis confirmed by Xpert assay from sputum samples. We analyzed patient characteristics, AI-based tuberculosis extent scores from chest X-rays, and Xpert Ct values. Of 230 patients, 206 (89.6%) achieved treatment success. The median age was 61 years, predominantly male (76.1%). AI-based radiographic tuberculosis extent scores (median 7.5) significantly correlated with treatment success (odds ratio [OR] 0.938, 95% confidence interval [CI] 0.895–0.983) and culture conversion at 8 weeks (liquid medium: OR 0.911, 95% CI 0.853–0.973; solid medium: OR 0.910, 95% CI 0.850–0.973). Sputum smear positivity was 49.6%, with a median Ct of 26.2. However, Ct values did not significantly correlate with major treatment outcomes. AI-based radiographic scoring at diagnosis is a significant predictor of treatment success and culture conversion in pulmonary tuberculosis, underscoring its potential in personalized patient management.

The Impact of Cycle Threshold Value in Influencing the Performance of COVID-19 Antigen

COVID-19 antigen is an alternative test for detecting SARS-CoV-2 infection. Viral load represented by the Cycle Threshold (CT) in the Real-Time Reverse Transcriptase Polymerase Chain Reaction (rRT-PCR) affects the diagnostic performance of the test. Higher CT values result in reduced sensitivity of the SARS-CoV-2 antigen. The main objective of this study was to determine the highest CT value in rRT-PCR that still yielded reactive results in the COVID-19 antigen test. This cross-sectional study was conducted at the Fever Outpatient Clinic in Dr. Cipto Mangunkusumo Hospital from July 2020 to June 2021. Two hundred and thirty-five naso-oropharyngeal swabs were taken from patients with confirmed and suspected COVID-19 diagnoses. About 24.7% of subjects were tested positive. The median highest CT value giving reactive COVID-19 antigen results was 28.22 (13.33-39.16), while the median CT value for non-reactive antigen results was 34.45 (26.08-39.65). At a CT value &lt; 40, the COVID-19 antigen test demonstrated 63.8% sensitivity, 99.4% specificity, 89.3% Negative Predictive Value (NPV), and 97.4% Positive Predictive Value (PPV). At the CT value &lt; 25, the test showed 92.3% sensitivity, 99.4% specificity, 99.4% NPV, 92.3% PPV, 163.4 LR+, and 0.1 LR-. The identified cut-off point for the CT value was 29.82, with a sensitivity of 64.9% and specificity of 81%. In conclusion, COVID-19 antigen is a valuable test for screening patients with symptoms of SARS-CoV-2 infection. Understanding the influence of cycle threshold can enhance the interpretation and reliability of the antigen test.

Are Pathogenic Leptospira Species Ubiquitous in Urban Recreational Parks in Sydney, Australia?

Leptospirosis is a zoonotic disease caused by the spirochete bacteria Leptospira spp. From December 2017 to December 2023, a total of 34 canine leptospirosis cases were reported in urban Sydney, Australia. During the same spatio-temporal frame, one locally acquired human case was also reported. As it was hypothesised that human residents and companion dogs might both be exposed to pathogenic Leptospira in community green spaces in Sydney, an environmental survey was conducted from December 2023 to January 2024 to detect the presence of pathogenic Leptospira DNA in multipurpose, recreational public parks in the council areas of the Inner West and City of Sydney, Australia. A total of 75 environmental samples were collected from 20 public parks that were easily accessible by human and canine visitors. Quantitative PCR (qPCR) testing targeting pathogenic and intermediate Leptospira spp. was performed, and differences in detection of Leptospira spp. between dog-allowed and dog-prohibited areas were statistically examined. The global Moran's Index was calculated to identify any spatial autocorrelation in the qPCR results. Pathogenic leptospires were detected in all 20 parks, either in water or soil samples (35/75 samples). Cycle threshold (Ct) values were slightly lower for water samples (Ct 28.52-39.10) compared to soil samples (Ct 33.78-39.77). The chi-squared test and Fisher's exact test results were statistically non-significant (p > 0.05 for both water and soil samples), and there was no spatial autocorrelation detected in the qPCR results (p > 0.05 for both sample types). Although further research is now required, our preliminary results indicate the presence of pathogenic Leptospira DNA and its potential ubiquity in recreational parks in Sydney.

Estimating Prevalence and Infection Intensity of Soil-Transmitted Helminths Using Quantitative Polymerase Chain Reaction and Kato-Katz in School-Age Children in Angola.

Quantitative polymerase chain reaction (qPCR) is gaining recognition in soil-transmitted helminth (STH) diagnostics, especially for Strongyloides stercoralis and differentiating hookworm species. However, sample preservation and DNA extraction may influence qPCR performance. We estimated STH prevalence and infection intensity by using qPCR in schoolchildren from Huambo, Uige, and Zaire, Angola, and compared its performance with that of the Kato-Katz technique (here termed Kato-Katz). Stool samples from 3,063 children (219 schools) were preserved in 96% ethanol and analyzed by qPCR, of which 2,974 children (215 schools) had corresponding Kato-Katz results. Cluster-adjusted prevalence and infection intensity estimates were calculated by qPCR and Kato-Katz, with cycle threshold values converted to eggs per gram for qPCR. Cohen's kappa statistic evaluated agreement between qPCR and Kato-Katz. DNA extraction and qPCR were repeated on 191 (of 278) samples that were initially qPCR negative but Kato-Katz positive, of which 112 (58.6%) became positive. Similar prevalence for Ascaris lumbricoides (37.5% versus 34.6%) and Trichuris trichiura (6.5% versus 6.1%) were found by qPCR and Kato-Katz, respectively, while qPCR detected a higher hookworm prevalence (11.9% versus 2.9%). The prevalence of moderate- or high-intensity infections was higher by Kato-Katz than by qPCR. Agreement between qPCR and Kato-Katz was very good for A. lumbricoides, moderate for T. trichiura, and fair for hookworm. Strongyloides stercoralis prevalence was 4.7% (municipality range, 0-14.3%), and no Ancylostoma ceylanicum was detected by qPCR. Despite suboptimal performance, presumably due to fixative choice, qPCR was fundamental in detecting S. stercoralis and excluding zoonotic A. ceylanicum. Further evaluations on sample fixatives and DNA extraction methods are needed to optimize and standardize the performance of qPCR.

An Evaluation of the Sensitivity and Specificity of Three COVID-19 Rapid Immunochromatographic Test Kits Compared to Real-Time Reverse Transcriptase-Polymerase Chain Reaction (rRT-PCR) Among Clinical Samples.

Background and objective The coronavirus disease 2019 (COVID-19) pandemic has imposed a significant burden on healthcare systems worldwide. This highlights the need for simple, rapid, and affordable diagnostic tests that can serve as alternatives tothe existing costly and demanding polymerase chain reaction (PCR) assay, especially in resource-limited countries like Ghana. In light of this, we aimed to assess thediagnostic efficacy of threeCOVID-19 rapid immunochromatographic antigen test kits vs. real-time reverse transcriptase-PCR (rRT-PCR). Methods This study evaluated the sensitivity and specificity of threeCOVID-19 rapid immunochromatographic antigen test kits: DG Rapid, SD Rapid, and SS Rapid. They were compared with the gold standard RT-PCRfor the detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) nucleocapsid antigen in 75 randomly selected archived nasopharyngeal samples. Results Of the 75 samples tested, 38 (50.7%) were positive and 37 (49.3%) were negative for SARS-CoV-2 RNA by rRT-PCR assay. No false positives were recorded. On the other hand, the DG Rapid kit detected 30 (78.9%) true positives and eight (21.1%) false negatives. SD Rapid kit detected 28 (73.7%) true positives and 10 (26.3%) false negatives, while the SS Rapid kit detected 19 (50.0%) true positives and 19 (50.0%) false negatives. While the specificity of each test kit was 100% (95% CI),the sensitivity of the DG Rapid, SD Rapid, and SS Rapid kits was 79%, 74%, and 50% (95% CI), respectively. Higher sensitivitieswere recorded among samples with cycle threshold (Ct) values <29.99 for each kit. Also, the DG Rapid kit demonstrated 79% excellent agreement with rRT-PCR, while the SD Rapid and SS Rapid kits demonstrated good agreement with rRT-PCR with 73% and 50% Cohen's kappa values, respectively. Conclusions Based on our findings,DG Rapid and SD Rapid kits are reliable alternatives to rRT-PCR for the detection of SARS-CoV-2 infection, especially in resource-limited settings like Ghana.

Pattern of Mutations in Rifampicin Resistance (rpoB) Gene and Analysis of Rifampicin Indeterminate Result in Tuberculosis Detected by Xpert MTB/RIF Assay in Kalaburagi, India

Tuberculosis (TB) is a serious disease that has been observed since ancient times. In the mid-to late-20th century, the main clinical approach to this disease involved focusing on its diagnosis, prevention, and treatment. However, in the 21st century, the focus has shifted toward the diagnosis and treatment of drug-resistant TB. With the use of the Xpert MTB/RIF assay at the frontlines in India, interpreting indeterminate results to treatment with rifampicin, an antitubercular drug, can be challenging. This is further exacerbated by a lack of knowledge regarding mutation frequency in antitubercular drug-resistant genes in this region. Among antitubercular drugs, rifampicin is the most potent and effective drug for the treatment of tuberculosis; hence, understanding the pattern of rifampicin resistance (rpoB) gene mutations will provide insights into the genetic basis of this resistance, which may help in the prevention and treatment of TB. This retrospective observational study presents sociodemographic details, sample types, Mycobacterium tuberculosis load, types of probe mutations detected, and rifampicin indeterminate results from the Xpert MTB/RIF assay. Of the 314 samples analyzed, 258 showed rifampicin resistance as detected by MTB, with 56 samples of MTB-detected rifampicin indeterminate results. Type E probe mutation (58.9%) was the most common type, while the least frequent mutation was Type C probe (1.5%). No missing probe was observed in approximately 8.9% of samples. Among the 56 rifampicin indeterminate results, the maximum Cycle threshold value did not cross 34.5 in six samples.

Association of Symptoms and Viral Culture Positivity for SARS-CoV-2-Tennessee, April-July 2020.

Understanding how symptoms are associated with SARS-CoV-2 culture positivity is important for isolation and transmission control guidelines. Individuals acutely infected with SARS-CoV-2 in Tennessee and their household contacts were recruited into a prospective study. All participants self-collected nasal swabs daily for 14 days and completed symptom diaries from the day of illness onset through day 14 postenrollment. Nasal specimens were tested for SARS-CoV-2 using RT-qPCR. Positive specimens with cycle threshold values < 40 were sent to the Centers for Disease Control and Prevention (CDC) for viral culture. First, we modeled the association between symptoms and the risk of culture positivity using an age-adjusted generalized additive model (GAM) accounting for repeated measurements within participants and a symptom-day spline. Next, we investigated how timing of symptom resolution was associated with the timing of culture resolution. In a GAM restricted to follow-up days after symptoms began, the odds of a specimen being culture positive was significantly increased on days when wheezing, loss of taste or smell, runny nose, nasal congestion, sore throat, fever, or any symptom were reported. For all symptoms except sore throat, it was more common for participants to have culture resolution before symptom resolution than for culture to resolve after or on the same day as symptom resolution. Overall, symptomatic individuals were more likely to be SARS-CoV-2 viral culture positive. For most symptoms, culture positivity was more likely to end before symptoms resolved. However, a proportion of individuals remained culture positive after symptom resolved, across all symptoms.

Clinical Characteristics of 427 Unvaccinated Chinese Adult Patients with Maintenance Hemodialysis Infected with the SARS-CoV-2 Omicron Variant: A Retrospective Study

Background: Maintenance hemodialysis (MHD) patients constitute a distinctive population characterized by unique traits, such as hemodialysis-related immune disturbances, and increased vulnerability to infections, heart failure, particularly severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Despite considerable research on MHD patients with coronavirus disease 2019 (COVID-19), there is limited information available on the clinical characteristics and outcomes of unvaccinated adult MHD patients infected with the SARS-CoV-2 Omicron variant. Methods: A retrospective investigation was conducted in a cohort of 427 unvaccinated MHD patients diagnosed with SARS-CoV-2 Omicron infection. The clinical symptoms, laboratory results, and treatment regimens were comprehensively examined. Multivariate regression identified independent risk factors for disease severity, and two-month follow-up assessments were conducted. Results: The cohort exhibited a spectrum of disease severity, with 37.5% classified as mild, 35.3% as common, 22.5% as severe, and 4.7% as critical. Serum C-reactive protein level, procalcitonin level, and bicarbonate concentration are related to the severity of the disease. Tailored treatment strategies—including nutritional support, hemodialysis management, anti-bacterial interventions, and more—were implemented. There was a 5.6% mortality rate, with a higher prevalence of deaths in the critical subtype. Independent risk factors for disease severity included fever, age, cycle threshold values (Ct values) of ORF1ab gene, and arterial oxygen partial pressure. Follow-up assessments revealed a median duration of 11 days for the negative nucleic acid of SARS-CoV-2. Twelve patients presented a second positive ORF1ab gene during follow-up but remained asymptomatic. Conclusion: These findings suggest the intricate clinical characteristics and outcomes of MHD patients with the SARS-CoV-2 Omicron infection. Tailored treatment strategies, continuous surveillance, and understanding prognostic factors are imperative for managing this high-risk population. Further research is warranted to validate these findings.

Replication kinetics and infectivity of SARS-CoV-2 Omicron variant sublineages recovered in the Republic of Korea.

We analyzed the correlation between the infectivity and transmissibility of the severe acute respiratory syndrome coronavirus 2 Omicron sublineages BA.1, BA. 2, BA.4, and BA.5. We assessed viral replication kinetics and infectivity at the cellular level. Nasopharyngeal and oropharyngeal specimens were obtained from patients with coronavirus disease 2019, confirmed using whole-genome sequencing to be caused by the Omicron sublineages BA.1, BA.2, BA.4, or BA.5. These specimens were used to infect Vero E6 cells, derived from monkey kidneys, for the purpose of viral isolation. Viral stocks were then passaged in Vero E6 cells at a multiplicity of infection of 0.01, and culture supernatants were harvested at 12-hour intervals for 72 hours. To evaluate viral replication kinetics, we determined the cycle threshold values of the supernatants using real-time reverse transcription polymerase chain reaction and converted these values to genome copy numbers. The viral load was comparable between BA.2, BA.4, and BA.5, whereas BA.1 exhibited a lower value. The peak infectious load of BA.4 was approximately 3 times lower than that of BA.2 and BA.5, while the peak load of BA.2 and BA.5 was about 7 times higher than that of BA.1. Notably, BA.1 demonstrated the lowest infectivity over the entire study period. Our results suggest that the global BA.5 wave may have been amplified by the higher viral replication and infectivity of BA.5 compared to other Omicron sublineages. These analyses could support the rapid assessment of the impact of novel variants on case incidence.