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Rowden, G.

Publications and source records attributed to Rowden, G..

4 recordsLinked to original sources

Detection and decontamination of chronic wasting disease prions during venison processing

Prion diseases, including chronic wasting disease (CWD), are caused by prions, which are misfolded aggregates of normal cellular prion protein. Prions possess many characteristics that distinguish them from conventional pathogens, in particular, an extraordinary recalcitrance to inactivation and a propensity to avidly bind to surfaces. In mid to late stages of CWD, prions begin accumulating in cervid muscle tissues. These features collectively create scenarios where occupational hazards arise for workers processing venison and pose risks to consumers through direct prion exposure via ingestion and cross-contamination of food products. In this work, we show that steel and plastic surfaces used in venison processing can be directly contaminated with CWD prions and that cross-contamination of CWD-negative venison can occur from equipment that had previously been used with CWD-positive venison. We also show that several decontaminant solutions (commercial bleach and potassium peroxymonosulfate) are efficacious for prion inactivation on these same surfaces.

microbiology↗

Microfluidic Quaking-Induced Conversion (Micro-QuIC) for Rapid On-Site Amplification and Detection of Misfolded Proteins

Protein misfolding diseases, such as prion diseases, Alzheimers, and Parkinsons, share a common molecular mechanism involving the misfolding and aggregation of specific proteins. There is an urgent need for point-of-care (POC) diagnostic technologies that can accurately detect these misfolded proteins, facilitating early diagnosis and intervention. Here, we introduce the Microfluidic Quaking Induced Conversion (Micro-QuIC), a novel acoustofluidic platform for the rapid and sensitive detection of protein misfolding diseases. We demonstrate the utility of our technology using chronic wasting disease (CWD) as a model system, as samples from wild white-tailed deer are readily accessible, and CWD shares similarities with human protein misfolding diseases. Acoustofluidic mixing enables homogeneous mixing of reagents in a high-Reynolds-number regime, significantly accelerating the turnaround time for CWD diagnosis. Our Micro-QuIC assay amplifies prions by an order of magnitude faster than the current gold standard, real-time quaking-induced conversion (RT-QuIC). Furthermore, we integrated Micro-QuIC with a gold nanoparticle-based, naked-eye detection method, which enables visual discrimination between CWD positive and negative samples without the need for a bulky fluorescence detection module. This integration creates a rapid, POC testing platform capable of detecting misfolded proteins associated with a variety of protein misfolding diseases. TOC graphic O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=109 SRC="FIGDIR/small/549283v1_ufig1.gif" ALT="Figure 1"> View larger version (39K): org.highwire.dtl.DTLVardef@885eeaorg.highwire.dtl.DTLVardef@10f820eorg.highwire.dtl.DTLVardef@12396eaorg.highwire.dtl.DTLVardef@188fe0f_HPS_FORMAT_FIGEXP M_FIG C_FIG

bioengineering↗

Standardization of Data Analysis for RT-QuIC-based Detection of Chronic Wasting Disease

Chronic wasting disease (CWD) is a disease affecting cervids and is caused by prions accumulating as pathogenic fibrils in lymphoid tissue and the central nervous system. Approaches for detecting CWD prions historically relied on antibody-based assays. However, recent advancements in protein amplification technology provided the foundation for a new class of CWD diagnostic tools. In particular, real-time quaking-induced conversion (RT-QuIC) has rapidly become a feasible option for CWD diagnosis. Despite its increased usage for CWD-focused research, there lacks consensus regarding the interpretation of RT-QuIC data for diagnostic purposes. It is imperative then to identify a standardized and replicable method for determining CWD status from RT-QuIC data. Here, we assessed variables that could impact RT-QuIC results and explored the use of maxpoint ratios (maximumRFU/backgroundRFU) to improve the consistency of RT-QuIC analysis. We examined a variety of statistical analyses to retrospectively analyze CWD status based on RT-QuIC and ELISA results from 668 white-tailed deer lymph nodes. Our results revealed an MPR threshold of 2.0 for determining the rate of amyloid formation, and MPR analysis showed excellent agreement with independent ELISA results. These findings suggest that the use of MPR is a statistically viable option for normalizing between RT-QuIC experiments and defining CWD status.

molecular biology↗

A Field-Deployable Diagnostic Assay for the Visual Detection of Misfolded Prions

Chronic Wasting Disease (CWD), a prion disease of cervids, has been identified across North America, Northern Europe and Korea. Diagnostic tools for the rapid and reliable detection of prion diseases are limited. Here, we combine gold nanoparticles (AuNPs) and quaking induced conversion (QuIC) technologies for the visual detection of amplified misfolded prion proteins from tissues of wild white-tailed deer infected with Chronic Wasting Disease (CWD). Our newly developed diagnostic test, MN-QuIC, enables both naked-eye and light-absorbance measurements for the detection of misfolded prions. The MN-QuIC assay leverages basic laboratory equipment that is cost-effective and portable, thus facilitating real-time prion diagnostics across a variety of settings. To test the portability of our assay, we deployed to a rural field station in southeastern Minnesota and tested for CWD on site. We successfully demonstrated that MN-QuIC is functional in a non-traditional laboratory setting by performing a blinded analysis in the field and correctly identifying all CWD positive and CWD not detected (independently confirmed with ELISA and/or IHC tests) animals at the field site, thus documenting the portability of the assay. Additionally, we show that electrostatic forces and concentration effects help govern the AuNP/prion interactions and contribute to the differentiation of CWD-prion positive and negative samples. We examined 17 CWD-positive and 24 CWD-not-detected white-tailed deer tissues that were independently tested using ELISA, IHC, and RT-QuIC technologies, and results secured with MN-QuIC were 100% consistent with these tests. We conclude that hybrid AuNP and QuIC assays, such as MN-QuIC, have great potential for sensitive, field-deployable diagnostics for a variety of protein misfolding diseases.

molecular biology↗