Magnetic bead-based separation of pneumococcal serotypes

York, Anna; Huynh, Emily; Mbodj, Sidiya; Yolda-Carr, Devyn; Hislop, Maikel S; Echlin, Haley; Rosch, Jason; Weinberger, Daniel M; Wyllie, Anne L.

doi:10.1016/j.crmeth.2023.100410

Cited by 6 publications

(4 citation statements)

References 33 publications

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“…However, the introduction of capsular serotype-based pneumococcus vaccines was found to lead to the occurrence of serotype replacement 13 , 14 . Over 100 pneumococcal capsular serotypes have been identified 15 , while polysaccharide-based vaccines only target a small subset of clinical strains. To address the current dilemma, it is imperative to develop new types of serotype-independent pneumococcal vaccines by seeking relatively conserved surface proteins that are common to most or all pneumococcal strains as vaccine formulations 10 .…”

Section: Introductionmentioning

confidence: 99%

A bacterial vesicle-based pneumococcal vaccine against influenza-mediated secondary Streptococcus pneumoniae pulmonary infection

Majumder,

Li,

Das

et al. 2024

Mucosal Immunology

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Section: Introductionmentioning

confidence: 99%

A bacterial vesicle-based pneumococcal vaccine against influenza-mediated secondary Streptococcus pneumoniae pulmonary infection

Majumder,

Li,

Das

et al. 2024

Mucosal Immunology

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“…A serotype 19A pneumococcus strain was serially diluted into the pooled saliva, from 5X10 7 CFU/ml to 5X10 1 CFU/mL, in duplicate (Supplementary Figure 1). 21 Each dilution was tested in triplicate, using both the extraction-free method and culture-enrichment methods as described above. The limit of detection was determined as the bacterial concentration in which all the triplicates in both replicates were positive for pneumococcus (C T <40).…”

Section: Methodsmentioning

confidence: 99%

A low-cost culture- and DNA extraction-free method for the molecular detection of pneumococcal carriage in saliva

Peno,

Lin,

Hislop

et al. 2023

Preprint

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BackgroundMolecular methods have improved the sensitivity of detection of pneumococcal carriage in saliva. However, they typically require sample culture-enrichment and nucleic acid extraction, prior to performing the detection assay. These factors may limit scalability for extensive surveillance of pneumococcus, particularly in low-resource settings. In this study, we evaluated the performance of a DNA-extraction-free method for the detection of pneumococcus in saliva.MethodsWe developed a streamlined qPCR-based protocol for the detection of pneumococcus, omitting culture-enrichment and DNA extraction. Using saliva samples collected from children attending childcare centers (New Haven, CT, USA), we evaluated detection of pneumococcus using saliva lysates as compared to purified DNA extracted from culture-enriched aliquots of the paired samples using qPCR targeting the pneumococcalpiaBgene.ResultsOf 759 saliva samples tested from 92 children (median age 3.65 years; IQR (2.46-4.78), pneumococcus was detected in 358 (47.2%) saliva lysates prepared using the extraction-free protocol and in 369 (48.6%) DNA extracted from the culture-enriched samples. We observed a near-perfect agreement between the two protocols (Cohen’s kappa: 0.92; 95%CI: 0.90-0.95). While we also observed a high correlation between the qPCR CTvalues generated by the two methods (r=0.93,p<0.0001), the CTvalues generated from the extraction-free, saliva lysates were higher (lower concentration) than those obtained from DNA extracted from culture-enriched samples (ΔCT= 6.68,p<0.00001).ConclusionsFor pneumococcal carriage surveillance in children, our findings suggest that a DNA extraction-free approach may offer a cost-effective alternative to the resource-intensive culture-enrichment method. While, as expected, we observed higher qPCR CTvalues (lower bacterial load) in the absence of culture-enrichment, the overall rate of detection remained unaffected.

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“…Saliva samples that tested positive for piaB with a Ct <28 were re-visited by culture 6 and/or magnetic bead-based separation (MBS) 23 in an attempt to isolate pure pneumococci. Pneumococcal isolates were serotyped by latex agglutination and qPCR then stored at -80°C.…”

Section: Strain Isolationmentioning

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

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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.

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Magnetic bead-based separation of pneumococcal serotypes

Cited by 6 publications

References 33 publications

A bacterial vesicle-based pneumococcal vaccine against influenza-mediated secondary Streptococcus pneumoniae pulmonary infection

A bacterial vesicle-based pneumococcal vaccine against influenza-mediated secondary Streptococcus pneumoniae pulmonary infection

A low-cost culture- and DNA extraction-free method for the molecular detection of pneumococcal carriage in saliva

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

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