Rapid and Accurate Identification of SARS-CoV-2 Omicron Variants Using Droplet Digital PCR (RT-ddPCR)

Background Mutations in the receptor binding domain of the SARS-CoV-2 Spike protein are associated with increased transmission or substantial reductions in vaccine efficacy, including in the recently described Omicron variant. The changing frequencies of these mutations combined with their differing susceptibility to available therapies have posed significant problems for clinicians and public health professionals. Objective To develop an assay capable of rapidly and accurately identifying variants including Omicron in clinical specimens to enable case tracking and/or selection of appropriate clinical treatment. Study Design Using three duplex RT-ddPCR reactions targeting four amino acids, we tested 419 positive clinical specimens from February to December 2021 during a period of rapidly shifting variant prevalences and compared genotyping results to genome sequences for each sample, determining the sensitivity and specificity of the assay for each variant. Results Mutation determinations for 99.7% of detected samples agree with NGS data for those samples, and are accurate despite wide variation in RNA concentration and potential confounding factors like transport medium, presence of additional respiratory viruses, and additional mutations in primer and probe sequences. The assay accurately identified the first 15 Omicron variants in our laboratory including the first Omicron in Washington State and discriminated against S-gene dropout Delta specimen. Conclusion We describe an accurate, precise, and specific RT-ddPCR assay for variant detection that remains robust despite being designed prior the emergence of Delta and Omicron variants. The assay can quickly identify mutations in current and past SARS-CoV-2 variants, and can be adapted to future mutations.

Improved detection of Little cherry virus-2 using a hydrolysis probe to manage the Pacific Northwest little cherry disease epidemic.

Little cherry virus-2 (LChV-2) is a viral pathogen that is reaching epidemic levels in Washington state. This virus is insect-vectored and has significant impacts on sweet cherry production. To aid growers in making informed management decisions we sought to develop a diagnostic assay to better detect isolates of LChV-2 currently found in Washington, allowing for more accurate estimations of disease occurrence. This study showed that there were two distinct genotypes of LChV-2 present in Washington state. This information was used to develop an up-to-date reverse transcription real-time quantitative polymerase chain reaction (RT-qPCR) assay, which was then optimized, validated, and compared to four previously published assays using a panel of field samples. This comparison demonstrated that the newly developed assay provided greater sensitivity, accurately detecting less than 10 copies per reaction and could detect both LChV-2 genotypes. Finally, we examined the effect of potential inhibitors in various tissue types from cherry, finding that young leaf tissue affected sensitivity of detection less than root tissues.

Differences in the transient responses of individual glaciers: a case study of the Cascade Mountains of Washington State, USA

Mountain glaciers have response times that govern retreat due to anthropogenic climate change. We use geometric attributes to estimate individual response times for 383 glaciers in the Cascade mountain range of Washington State, USA. Approximately 90% of estimated response times are between 10 and 60 years, with many large glaciers on the short end of this distribution. A simple model of glacier dynamics shows that this range of response times entails consequential differences in recent and ongoing glacier changes: glaciers with decadal response times have nearly kept pace with anthropogenic warming, but those with multi-decadal response times are far from equilibrium, and their additional committed retreat stands well beyond natural variability. These differences have implications for changes in glacier runoff. A simple calculation highlights that transient peaks in area-integrated melt, either at the onset of forcing or due to variations in forcing, depend on the glacier's response time and degree of disequilibrium. We conclude that differences in individual response times should be considered when assessing the state of a population of glaciers and modeling their future response. These differences in response can arise simply from a range of different glacier geometries, and the same basic principles can be expected in other regions as well.

Nested spatial and temporal modeling of environmental conditions associated with genetic markers of Vibrio parahaemolyticus in Washington state Pacific oysters

The Pacific Northwest (PNW) is one of the largest commercial harvesting areas for Pacific oysters (Crassostrea gigas) in the United States. Vibrio parahaemolyticus, a bacterium naturally present in estuarine waters, accumulates in shellfish and is a major cause of seafood-borne illness. Growers, consumers, and public-health officials have raised concerns about rising vibriosis cases in the region. V. parahaemolyticus genetic markers (tlh, tdh, trh) were estimated using an MPN-PCR technique in Washington State Pacific oysters regularly sampled between May and October from 2005 to 2019 (N=2,836); environmental conditions were also measured at each sampling event. Multilevel mixed-effects regression models were used to assess relationships between environmental measures and genetic markers as well as genetic marker ratios (trh:tlh, tdh:tlh, and tdh:trh), accounting for variation across space and time. Spatial and temporal dependence were also accounted for in the model structure. Model fit improved when including environmental measures from previous weeks (1-week lag for air temperature, 3-week lag for salinity). Positive associations were found between tlh and surface water temperature, specifically between 15°C and 26°C, and between trh and surface water temperature up to 26°C. tlh and trh were negatively associated with 3-week lagged salinity in the most saline waters (> 27 ppt). There was also a positive relationship between tissue temperature and tdh, but only above 20°C. The tdh:tlh ratio displayed analogous inverted non-linear relationships as tlh. The non-linear associations found between the genetic targets and environmental measures demonstrate the complex habitat suitability of V. parahaemolyticus. Additional associations with both spatial and temporal variables also suggest there are influential unmeasured environmental conditions that could further explain bacterium variability. Overall, these findings confirm previous ecological risk factors for vibriosis in Washington State, while also identifying new associations between lagged temporal effects and pathogenic markers of V. parahaemolyticus.

Improvement in Emotion Regulation While Detained Predicts Lower Juvenile Recidivism

The goal of the current study was to investigate the relationships between observer-rated skills related to emotional and cognitive regulation post-admission and pre-release in a secure facility and official records of juvenile felony recidivism up to 1 year after release. Data came from a sample of 599 youth in a residential facility in Washington state (84% male; 38% White). Latent change score models indicated that both initial level of emotional regulation skills and improvement in emotion regulation skills while incarcerated were significantly related to lower recidivism. This pattern of findings remained when controlling for length of stay, among other covariates. Follow-up analyses indicated that the results for emotion regulation skills might be driven primarily by monitoring internal and external triggers. Additional research should investigate the connection between emotion regulation skills and juvenile recidivism, with a special focus on trigger monitoring and how to improve those skills.

Supervisory alliance as a moderator of the effects of behavioral rehearsal on TF-CBT fidelity: results from a randomized trial of supervision strategies

The present study examined relations between supervisory alliance and fidelity to the trauma narrative component of Trauma-Focused Cognitive Behavioral Therapy, as well as how supervisory alliance might moderate the effect of behavioral rehearsals (i.e., role plays) on fidelity. Forty-two supervisors and 124clinicians from 28 different community-based mental health offices across Washington State participated.Clinicians were randomized to receive one of two supervision conditions—symptom and fidelity monitoring or symptom and fidelity monitoring with behavioral rehearsal. Supervisory alliance alone did not predict adherence or extensiveness of the trauma narrative. One aspect of alliance, client focus, significantly altered the effect of supervision condition on adherence (p=0.05); however, this effect was only seen in 43.5% of clinicians. A second aspect of alliance, rapport, altered the effect of condition on trauma narrative extensiveness with moderate significance (p=0.09). Future research should investigate strategies to improve supervisory alliance or match supervision strategies to specific supervisor-clinician dyads.