162.1 - Using Droplet Digital PCR to Detect Cyanobacteria in Human Lung Tissue

Guohong Huang (Dartmouth-Hitchcock Medical Center), Rachael Barney (Dartmouth-Hitchcock Medical Center), Torrey Gallagher (Dartmouth-Hitchcock Medical Center), Maeve Tischbein (Dartmouth-Hitchcock Medical Center), Elijah Stommel (Dartmouth-Hitchcock Medical Center), Gregory Tsongalis (Dartmouth-Hitchcock Medical Center)

Background

With the spread of highly infectious strains such as the Delta variant of SARS-CoV-2, rapid antigen testing and variant tracking are gaining more attention from infectious disease specialists and the general public as essentials for disease control and prevention amidst the COVID-19 pandemic. The lateral flow based antigen tests provide fast turnaround of results within minutes, but it’s open -faced assay format means that the test operators are more prone to accidental exposure to pathogen-containing samples. The viral inactivation transport media (ITMs) provide much needed protection against exposure to highly transmissive live viruses during sample collection, transport, and storage. However, the current ITMs were designed for PCR tests and share a harsh chemical formulation that denatures protein analytes, making them incompatible with antigen tests.

We developed an innovative viral transport media that is designed for a variety of tests, such as antigen testing and PCR. The new formulation can be used to safely inactivate SARS-CoV-2 virus while maintaining compatibility with many different formats beyond rt-PCR.

Design

To evaluate the viral inactivation performance of the novel inactivation transport media formulation, a cytopathic effect assay was conducted using VERO E6 cells in presence of SARS-CoV-2 samples incubated in our novel formulation and log reduction were reported.

To assess its compatibility with different SARS-CoV-2 related assays, the novel formulation was used as inactivation transport media in antigen, PCR, and NGS based Swab-Seq tests with known controls.

Results

The results for the inactivation study confirmed gt; 3.0 log reduction in viral titer after 30 min exposure in novel transport media formulation and demonstrated 99.5% effectiveness in SARS-CoV-2 inactivation. The results for the PCR and antigen test compatibility studies showed comparable and/or superior performance in detection limits for the novel formulation when compared to other sample media. The Swab-Seq result offers preliminary support on the novel formulation’s compatibility with NGS based assays.

Conclusion

Using a novel viral transport medium, we were able to completely inactivate the SARS-CoV-2 virus. The new formulation was compatible with both nucleic acid and protein assays while protecting cells from infection. This viral transport media could allow more test types to be done from a single specimen, resulting in safer and reliable outcomes. Additionally, it would reduce the need for multiple samples collected from patients while supporting SARS-CoV-2 variant tracking via NGS.

This work has been supported by the Ramp;amp;D department of Truckee Applied Genomics, in an effort to advance pre-analytical sample handling for molecular diagnostics.





TAG-NGPM+ has a unique value offering in assay compatibility, which means one swab sample in 2ml of TAG regent can help with returning a rapid test result in 10-15mins, a PCR test in a few hours, and variant information within days. TAG-NGPM+ offers cold chain free sample stabilization, inactivates the live virus, and is cost-effective, making it an ideal solution for high volume, mobile test sites.; Clinically positive upper respiratory samples were diluted into TAG-NGPM+, or TAG-NGPM, and incubated at ambient temperature for 1-10 days, in triplicates.  The result indicates that both TAG-NGPM+ and TAG-NGPM are compatible with the NGS-based SWAB-Seq protocol, and this compatibility also demonstrates strong evidence that TAG transport media will be compatible with sequencing-based variant tracking workflow.