Testing of four-sample pools offers resource optimization without compromising diagnostic performance of real time reverse transcriptase-PCR assay for COVID-19.

Quick identification and isolation of SARS-CoV-2 infected individuals is central to managing the COVID-19 pandemic. Real time reverse transcriptase PCR (rRT-PCR) is the gold standard for COVID-19 diagnosis. However, this resource-intensive and relatively lengthy technique is not ideally suited for mass testing. While pooled testing offers substantial savings in cost and time, the size of the optimum pool that offers complete concordance with results of individualized testing remains elusive. To determine the optimum pool size, we first evaluated the utility of pool testing using simulated 5-sample pools with varying proportions of positive and negative samples. We observed that 5-sample pool testing resulted in false negativity rate of 5% when the pools contained one positive sample. We then examined the diagnostic performance of 4-sample pools in the operational setting of a diagnostic laboratory using 500 consecutive samples in 125 pools. With background prevalence of 2.4%, this 4-sample pool testing showed 100% concordance with individualized testing and resulted in 66% and 59% reduction in resource and turnaround time, respectively. Since the negative predictive value of a diagnostic test varies inversely with prevalence, we re-tested the 4-sample pooling strategy using a fresh batch of 500 samples in 125 pools when the prevalence rose to 12.7% and recorded 100% concordance and reduction in cost and turnaround time by 36% and 30%, respectively. These observations led us to conclude that 4-sample pool testing offers the optimal blend of resource optimization and diagnostic performance across difference disease prevalence settings.

Evaluation of pooled sample analysis strategy in expediting case detection in areas with emerging outbreaks of COVID-19: A pilot study.

Timely diagnosis of COVID-19 infected individuals and their prompt isolation are essential for controlling the transmission of SARS-CoV-2. Though quantitative reverse transcriptase PCR (qRT-PCR) is the method of choice for COVID-19 diagnostics, the resource-intensive and time-consuming nature of the technique impairs its wide applicability in resource-constrained settings and calls for novel strategies to meet the ever-growing demand for more testing. In this context, a pooled sample testing strategy was evaluated in the setting of emerging disease outbreak in 3 central Indian districts to assess if the cost of the test and turn-around time could be reduced without compromising its diagnostic characteristics and thus lead to early containment of the outbreak. From 545 nasopharyngeal and oropharyngeal samples received from the three emerging districts, a total of 109 pools were created with 5 consecutive samples in each pool. The diagnostic performance of qRT-PCR on pooled sample was compared with that of individual samples in a blinded manner. While pooling reduced the cost of diagnosis by 68% and the laboratory processing time by 66%, 5 of the 109 pools showed discordant results when compared with induvial samples. Four pools which tested negative contained 1 positive sample and 1 pool which was positive did not show any positive sample on deconvolution. Presence of a single infected sample with Ct value of 34 or higher, in a pool of 5, was likely to be missed in pooled sample analysis. At the reported point prevalence of 4.8% in this study, the negative predictive value of qRT-PCR on pooled samples was around 96% suggesting that the adoption of this strategy as an effective screening tool for COVID-19 needs to be carefully evaluated.

Zoledronate and Molecular Iodine Cause Synergistic Cell Death in Triple Negative Breast Cancer through Endoplasmic Reticulum Stress.

Women consuming molecular iodine (I2) through seaweeds suffer the least from breast cancers. Zoledronate (Zol) is in clinical use for alleviation of bone pain in cancer patients. Triple negative breast cancers exhibit high mortality due to lack of neoadjuvant chemotherapy. I2 and Zol independently cause weak antiproliferative and apoptotic effect. So far, their combined effects have not been tested. We analyzed the effect of combination of I2 with Zol as a potent adjuvant therapeutic agent for triple negative breast cancer cells (MDA-MBA-231) and in the mice model of breast cancer. Cell viability, terminal deoxynucleotidyl transferase dUTP nick end labeling staining, Western blotting, real-time PCR, flow cytometry, and other assays were performed for assessing cell death, calcium levels, and migration potential, respectively, in treated cells. The increased caspase 8, increased [Ca(2+)]c levels, and endoplasmic reticulum (ER) stress resulted in apoptosis. Real time and fluorescence-based analysis demonstrated that the combination treatment targets ER Ca(2+) homeostasis chaperons leading to apoptosis. Combination therapy reduces metalloproteinases 2 and 9, inhibits invasion/migration of cells, and prevents growth of tumor in mice. I2 + Zol combination treatment induces synergistic increase in ER-mediated apoptosis, reduces invasion/migration potential of MDA-MB-231 cells, and exhibits antiproliferative property in vivo demonstrating its potential as combination therapy.

Occurrence of phthalic acid esters in Gomti River Sediment, India.

Phthalic acid esters (PAEs) are well-known ubiquitous environmental pollutants and used as plasticizers for the manufacturing of plastic products. During this exploratory study, an attempt has been made to determine the concentration and distribution of five prominent PAEs, viz. di-methyl phthalate (DMP), di-ethyl phthalate (DEP), di-butyl phthalate (DBP), di-(2-ethylhexyl) phthalate (DEHP), and di-octyl phthalate (DOP) in the sediment samples of Gomti River collected from 30 different locations. Identification and quantification of PAEs were performed by high-performance liquid chromatography. The mean concentration values of DMP, DEP, DBP, DEHP, and DOP were found as 10.54, 4.57, 10.41, 31.61, and 5.16 microg/kg, respectively. Limit of detection and limit of quantification for each PAE were also calculated and found in the ranges of 0.09-0.55 and 0.28-1.67 microg/kg. DEHP was the most frequently detected PAE (present in 93.3% samples); however, DOP was found only in 36.7% samples.

Distribution, sources and characterization of polycyclic aromatic hydrocarbons in the sediment of the river Gomti, Lucknow, India.

Sediment samples were collected for PAHs analysis (upstream, midstream and downstream) from the bank of the river Gomti in Lucknow city, India during 2005-2007. Total concentration of the PAHs ranged from 0.068 to 3.153 microg/g dry weight. A correlation existed between the sediment organic carbon content (f(OC)) and the total PAHs concentration with a correlation coefficient (r(2)) of 0.788, suggesting that sediment organic carbon content played an important role in controlling the PAHs levels in the sediments. According to observed molecular indices, PAHs contamination in the river Gomti seems to be originated both from the high temperature pyrolytic process as well as from the petrogenic source, indicating a mixed PAH input pattern.