Background and Objective Since the initial coronavirus disease outbreak in late 2019 (COVID-19), reverse-transcription real-time polymerase chain reaction (RT-qPCR) has become the gold standard test to detect severe acute respiratory syndrome-related coronavirus 2 (SARS-CoV-2). However, a more sensitive and accurate diagnostic tool was required. Therefore, droplet digital polymerase chain reaction (ddPCR) was suggested as an alternative method. Here, we evaluated the performance of ddPCR to detect SARS-CoV-2 and compared it to the performance of RT-qPCR. Methods The analytical performances, including limit of blank and limit of detection, were established using positive and negative SARS-CoV-2 reference materials. A total of 366 RNA extracts (173 positive and 193 negative by RT-qPCR) were collected from four institutions and tested with a Bio-Rad SARS-CoV-2 ddPCR kit that detects the SARS-CoV-2 genome using primers for N1 and N2. Results Limit of blank was set at 0, and the limits of detection of N1 and N2 were 1.99 copies/μL and 5.18 copies/μL, respectively. Linearity was evaluated using serial dilution samples, which demonstrated good results (R²: 0.999, linear range: 5.88–6825.25 copies/μL for N1 and R²: 0.999, 5.53–5855.47 copies/μL for N2). The results of ddPCR and RT-qPCR revealed substantial agreement (Cohen’s kappa: 0.639, p < 0.01). The 63 samples with positive ddPCR but negative RT-qPCR showed low copy numbers, and 55% of them had COVID-19-related symptoms. Conclusions Droplet digital polymerase chain reaction demonstrated excellent sensitivity for SARS-Cov-2 detection and consistently agreed with the results from conventional RT-qPCR. Furthermore, ddPCR provided quantitative data that can be used to monitor changes in the viral load of patients with COVID-19.

Background and objectives: Ovarian cancer is one of the leading causes of death among women worldwide. Most newly diagnosed ovarian cancer patients are diagnosed in advanced stages of the disease. Despite various treatments, most patients with advanced stage ovarian cancer, including serous ovarian cancer (the most common subtype of ovarian cancer), experience recurrence, which is associated with extremely poor prognoses. In the present study, we aimed to identify core genes involved in ovarian cancer and their associated molecular mechanisms, as well as to investigate related clinicopathological implications in ovarian cancer. Materials and methods: Three gene expression cohorts (GSE14407, GSE36668, and GSE38666) were obtained from the Gene Expression Omnibus databases to explore potential therapeutic biomarkers for ovarian cancer. Nine up-regulated and six down-regulated genes were screened. Three publicly available gene expression datasets (GSE14407, GSE36668, and GSE38666) were analyzed. Results: A total of 14 differently expressed genes (DEGs) were identified, among which nine genes were upregulated (BIRC5, CDCA3, CENPF, KIF4A, NCAPG, RRM2, UBE2C, VEGFA, and NR2F6) and were found to be significantly enriched in cell cycle regulation by gene ontology analysis. Further protein–protein interaction network analysis revealed seven hub genes among these DEGs. Moreover, Kaplan–Meier survival analysis showed that a higher expression of CDCA3 and UBE2C was associated with poor overall patient survival regardless of tumor stage and a higher tumor histologic grade. Conclusion: Altogether, our study suggests that CDCA3 and UBE2C may be valuable biomarkers for predicting the outcome of patients with advanced serous ovarian cancer.

Kyoung Bo Kim, M.D., Sunggyun Park, M.D., Jung Sook Ha, M.D., Ph.D., Namhee Ryoo, M.D., Ph.D., and Do-Hoon Kim, M.D., Ph.D.. Ann Lab Med 2022;42:497-9. https://doi.org/10.3343/alm.2022.42.4.497

Kim KB, et al. Ann Lab Med. 2015 May;35(3):379-381. http://dx.doi.org/10.3343/alm.2015.35.3.379

Several manufacturers have developed systems that automatically measure the amount of blood in culture bottles. We compared the volumes measured automatically by the Virtuo instrument (bioMérieux, France) (height-based volumes) and those calculated by weighing the bottles. In all, 150 pairs of aerobic and anaerobic blood culture bottles (BacT/ALERT FA/FN Plus, bioMérieux) were randomly selected over two periods to compare the height- and weight-based volumes and analyze the effect of foam. We also estimated the limit of detection (LOD) and the cut-off value for 5 mL equine blood. The mean height-based volume was approximately 0.67 mL greater than the weight-based volume, particularly in anaerobic culture bottles. Foam did not have a significant effect. The LOD for the automatic height-based volume of equine blood was 0.2–0.4 mL. The 5 mL cut-off was 4–4.2 mL. Therefore, when reporting or monitoring blood volume within culture bottles in the laboratory, these performance characteristics should be adequately considered.

Background: Ocular infections can cause severe complications, including blindness, and distinguishing bacterial from fungal keratitis based on clinical features alone is difficult. This study compared broad-range conventional PCR and real-time PCR methods targeting the 16S rRNA gene with traditional culture for diagnosing bacterial ocular infections. Methods: We analyzed 160 ocular specimens from 111 patients, categorizing them as septic or aseptic. The results of both conventional PCR and real-time PCR methods targeting the 16S rRNA gene were compared with traditional culture outcomes. Results: Real-time PCR demonstrated higher sensitivity than conventional PCR, and receiver operating characteristic analysis determined optimal ΔCT cutoff values of −2.13 and −4.09 for septic and aseptic specimens, respectively. Delays in specimen processing significantly affected real-time PCR accuracy. The 16S rRNA meta-taxonomic analysis using nanopore sequencing only validated the PCR results when the DNA concentration was sufficient. Conclusions: Broad-range real-time PCR proved to be a valuable diagnostic tool, particularly in aseptic specimens, with greater sensitivity and specificity than conventional PCR. The established ΔCT cutoff values improved diagnostic accuracy and showed that standardized specimen collection and processing are crucial for maximizing PCR efficacy.