Exploring the potential of a saliva-based, RNA-extraction-free PCR test for the multiplexed detection of key respiratory pathogens

Allicock, Orchid M.; Lin, Tzu-Yi; Fajardo, Katherine T.; Yolda-Carr, Devyn; Hislop, Maikel S.; Wang, Jianhui; Zuniga, Denora; Platt, William; Tuohy, Beth; Wyllie, Anne L.

doi:10.1101/2023.10.04.23296240

Cited by 4 publications

(1 citation statement)

References 38 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Lysates were prepared from 50 µl of each saliva sample which was heated at 95°C for 5 minutes 24 before testing in a modified “SalivaDirect” PCR assay, 25 expanded for multiplexed detection of these viruses. 26…”

Section: Methodsmentioning

confidence: 99%

Contact with young children is a major risk factor for pneumococcal colonization in older adults

Wyllie,

Yolda-Carr,

Hislop

et al. 2024

Preprint

Self Cite

View full text Add to dashboard Cite

BackgroundImportant questions remain about the sources of transmission of pneumococcus to older adults in the community. This is a critical question for understanding the potential indirect effects of using pneumococcal conjugate vaccines (PCVs) in children and older adults. For non-institutionalized individuals, the most likely source of adult-to-adult transmission is in the household. The goal of this study was to characterize the dynamics and risk factors for acquisition of pneumococcus in older adults.MethodsWe designed a longitudinal study to sample adults >60 years of age living in the same household (New Haven, CT, USA), and without younger contacts residing in the household. Saliva samples and questionnaires regarding social behaviors and health status were obtained every 2 weeks for a period of 10 weeks. DNA extracted from culture-enriched saliva was tested using qPCR for pneumococcus genespiaBandlytA.ResultsAcross two study seasons (November 2020-August 2021, November 2021-September 2022), 121 individuals from 61 households were followed for 6 study visits; 62 individuals were enrolled in both seasons. Overall, 52/1088 (4.8%) samples tested positive for pneumococcus based onpiaB, with 27/121 (22.3%) individuals colonized on at least one time point. Several individuals were colonized at multiple timepoints including two individuals who were colonized throughout the 10-week sampling period; two others were colonized at 5 of 6 time points. In 5 instances, both members of the household were carriers in the same season, though not necessarily at the same time point. Pneumococcal carriage was substantially higher among individuals who had contact with children (10.0% vs 1.6%). Participants who reported recent contact with <5-year-olds and 5-9-year-olds had particularly elevated prevalence (13.8%; 14.1%, respectively).ConclusionsContact with young children was the most important factor that influenced pneumococcal acquisition rates. While there were several instances where both adult household members were colonized at the same time or at sequential visits, these individuals also both typically had contact with children.

show abstract

Section: Methodsmentioning

confidence: 99%

Contact with young children is a major risk factor for pneumococcal colonization in older adults

Wyllie,

Yolda-Carr,

Hislop

et al. 2024

Preprint

Self Cite

View full text Add to dashboard Cite

show abstract

Contact with young children is a major risk factor for pneumococcal colonization in older adults

Wyllie,

Yolda-Carr,

Hislop

et al. 2024

FEMS Microbes

View full text Add to dashboard Cite

Background Important questions remain about the sources of transmission of pneumococcus to older adults in the community. This is critical for understanding the potential effects of using pneumococcal conjugate vaccines (PCVs) in children and older adults. For non-institutionalized individuals, we hypothesized that the most likely source of adult-to-adult transmission is within the household. Methods We designed a longitudinal study to sample adults ≥60 years of age living in the same household (New Haven, CT, USA), without younger residents in the household. Saliva samples and social and health questionnaires were obtained every 2 weeks for a period of 10 weeks. DNA extracted from culture-enriched saliva was tested using qPCR for pneumococcus genes piaB, lytA, and serotype. Results Across two study seasons (November 2020-August 2021, November 2021-September 2022), 121 individuals from 61 households completed all 6 visits; 62 individuals were enrolled in both seasons. Overall, 52/1088 (4.8%) samples tested positive for pneumococcus, with 27/121 (22.3%) individuals colonized at least once. Several individuals were colonized at multiple timepoints; two individuals were colonized at 5/6 time points and two at all six. In 5 instances, both household members were carriers in the same season, though not necessarily at the same time. Pneumococcal carriage was substantially higher among individuals who had contact with children (10.0% vs 1.6%). Conclusions Contact with young children was the most important factor that influenced pneumococcal acquisition rates. While there were several instances where both adult household members were colonized at the same time or at sequential visits, these individuals typically had contact with children. As such, PCV immunization can directly protect older adults who have contact with children.

show abstract

Expansion of a low-cost, saliva-based PCR test for the detection of mpox virus

Thomas,

Allicock,

Yolda-Carr

et al. 2023

Preprint

View full text Add to dashboard Cite

Background. Current recommendations for the diagnosis of mpox rely on lesion-swabs as the gold-standard specimen type even though many patients experience symptoms prior to lesion-onset. Earlier detection could bolster the mpox response by mitigating transmission and facilitating access to antiviral treatments. Methods. We first compared five PCR assays for their detection of mpox DNA extracted from 30 saliva specimens in collection devices with a stabilizing buffer. Next, we investigated the stability of mpox detection in five raw, unsupplemented saliva samples diluted 1:10 in mpox-negative saliva, after storage at 4°C, room temperature (~19°C), 30°C, and 40°C for 72 hours. We also investigated the stability of virus detection through simulated shipping conditions. Lastly, we performed amplicon sequencing on seven saliva samples and assessed concordance of the PCR assays against mpox virus sequences. Results. Despite identifying three different substitutions in the CDC's Monkeypox Virus Generic Real-Time PCR Test's forward and reverse primers, we observed no difference in the mean cycle threshold values generated between assays. However, one gene target for one assay performed better for overall detection when validated. Detection following storage at 4°C, ~19°C, and 30°C remained relatively stable for 24-48 hours but this declined by 72 hours. At 40°C, detection was stable at 24 hours but declined by 48 hours. Detection following simulated summer and winter shipping temperature profiles also remained stable. Conclusions. Findings of this pilot investigation support a flexible, saliva-based, extraction-free PCR test as a promising approach for the low-cost detection of mpox virus. With stability observed for 24-48 hours as well as over simulated shipping temperatures, saliva-based sampling and simplified testing could reduce mpox diagnostic costs, increase access to testing and address hurdles in low- and middle-income countries. Future studies should build upon this and assess the temporal dynamics of mpox virus in saliva.

show abstract

Exploring the potential of a saliva-based, RNA-extraction-free PCR test for the multiplexed detection of key respiratory pathogens

Cited by 4 publications

References 38 publications

Contact with young children is a major risk factor for pneumococcal colonization in older adults

Contact with young children is a major risk factor for pneumococcal colonization in older adults

Contact with young children is a major risk factor for pneumococcal colonization in older adults

Expansion of a low-cost, saliva-based PCR test for the detection of mpox virus

Contact Info

Product

Resources

About