scholarly journals Comparison of DNA Extracted from Pediatric Saliva, Gingival Crevicular Fluid and Site-Specific Biofilm Samples

2020 ◽  
Vol 3 (3) ◽  
pp. 48
Author(s):  
Jason Emett ◽  
Roxanne David ◽  
Jaydene McDaniel ◽  
Steven McDaniel ◽  
Karl Kingsley
Keyword(s):  
Pediatric Patients ◽  
Gingival Crevicular Fluid ◽  
Limit Of Detection ◽  
Low Cost ◽  
Molecular Screening ◽  
Site Specific ◽  
Paper Point ◽  
Unstimulated Saliva ◽  
Oral Microbes ◽  
Crevicular Fluid

(1) Introduction: Due to the non-invasive nature of saliva, many methods have been used to isolate and collect DNA from saliva samples for microbial screening. Many oral microbes also inhabit the oral biofilm, which may represent significantly different microbial constituents that may contribute to oral health and disease, including caries and periodontal disorders. Moreover, the biofilm may vary within the same patient at different sites. Few studies have evaluated the comparison between DNA isolated from saliva and DNA from site-specific biofilm, with virtually no studies addressing this analysis among pediatric patients. (2) Methods: An existing repository of paper point derived biofilm, gingival crevicular fluid (GCF), and unstimulated saliva samples previously collected from pediatric patients (n = 47) was identified. DNA was isolated from biofilm sites (tongue, upper buccal molar, mandibular lingual incisor), and GCF and saliva were used for quantitative DNA comparison using a phenol:chloroform extraction. A quantitative and qualitative analysis was performed using the NanoDrop 2000 spectrophotometer using absorbance readings at A230 nm, A260 nm and A280 nm. (3) Results: These data demonstrated the successful isolation of DNA from all of the patient samples, with the highest concentrations observed among unstimulated saliva (4264.1 ng/μL) and the lowest derived from GCF (1771.5 ng/μL). No differences were observed between males and females or minorities and non-minority patients. In addition, comparison of the overall concentrations of DNA obtained from adult samples was slightly higher than, but not significantly different from, the concentrations obtained from pediatric samples (p = 0.2827). A real-time quantitative qPCR screening revealed that all of the samples evaluated harbored bacterial and human DNA of sufficient quantity and quality for a molecular screening greater than the limit of detection (ΔRn = 0.01). (4) Conclusions: Many methods are currently available to provide the sampling and screening of saliva and specific sites within the oral cavity, but the validation and comparison of simple and low-cost methods, that include paper point sampling and unstimulated saliva collection, may suggest these methods and protocols provide sufficient DNA quality and quantity for molecular screening and other comparison applications. In addition, although heterogeneity will be a constant and consistent feature between patient samples, standardized methods that provide similar and consistent DNA from various oral sites may provide needed consistency for screening and molecular analysis.

scholarly journals Quantitative Comparison of Oral Site-specific DNA Isolates Reveals Differential Outcomes

2019 ◽  
pp. 1-7 ◽  
Author(s):  
Graydon Carr ◽  
Arvin Alexander ◽  
Kevin Dionisio ◽  
Karl Kingsley
Keyword(s):  
Gingival Crevicular Fluid ◽  
Low Cost ◽  
Direct Analysis ◽  
Lower Incisor ◽  
Differential Outcomes ◽  
Paper Point ◽  
Significant Difference ◽  
Unstimulated Saliva ◽  
Saliva Collection ◽  
Crevicular Fluid

Introduction: More and more evidence has accumulated that suggests salivary sampling may provide direct analysis of oral conditions and microbial constituents, but may also be useful in the diagnosis and early detection of other chronic diseases. Although multiple methods of oral sampling currently exist, some methods are prohibitively expensive or based upon technologies not ubiquitously available at public health centers or state-funded colleges. This study provides a comparative analysis of DNA concentrations and quality from five specific oral sites derived using sterile paper points, including the gingival crevice between the upper central incisors, biofilm of the upper first molar, lingual incisor, and the dorsum of the tongue for comparison with unstimulated saliva collection. Methods: This study analyzed previously collected unstimulated saliva and paper point samples. In brief, DNA was isolated from each using TRIzol (phenol: Chloroform) extraction and DNA quantification and quality was measured using a NanoDrop spectrophotometer at 260 and 280 nm. Results: Analysis of Paper Point (PP) biofilm sampling sites from upper first molar, lower incisor, and dorsum of the tongue revealed similar average DNA concentrations, ranging between 14,342 ng/uL and 14,402 ng/uL (p=0.9851). Although variations were observed between different patients, samples from different oral sites within the same patient were strikingly similar, R=0.8355. Comparison of DNA isolated from fluids, gingival crevicular fluid (GCF) and unstimulated saliva revealed average DNA concentrations that were similar to the biofilm sampling sites (14,686 ng/uL and 13,743 ng/uL, respectively), which were not significantly different from one another (p=0.7893). DNA concentrations ranged considerably between patients (low = 4,410 ng/uL; high = 48,783 ng/uL), but were most similar with different samples (GCF, saliva) from the same patient (Pearson’s R=0.6979). In addition, DNA purity measured by A260:A280 nm absorbance did not reveal any significant difference among sampling sites (range 1.62 – 1.70; p=0.427). Discussion: Although many methods are available to provide oral sampling, simple and low-cost methods such as paper point sampling, unstimulated saliva collection and buccal swabs may represent tools that provide sufficient DNA quality and quantity for molecular screening. In addition, although heterogeneity between patient samples will always be present – samples from various oral sites within the same patient may provide roughly equivalent DNA samples for further screening and molecular analysis.

A new ultrasensitive chemiluminescent assay for the site-specific quantification of alkaline phosphatase in gingival crevicular fluid

1993 ◽  
Vol 28 (4) ◽  
pp. 266-273 ◽  
Author(s):  
I. L. C. Chapple ◽  
J. B. Matthews ◽  
G. H. G. Thorpe ◽  
H. D. Glenwright ◽  
J. M. Smith ◽  
...  
Keyword(s):  
Alkaline Phosphatase ◽  
Gingival Crevicular Fluid ◽  
Site Specific ◽  
Crevicular Fluid

scholarly journals Microbial Screening Reveals Oral Site-Specific Locations of the Periodontal Pathogen Selenomonas noxia

2021 ◽  
Vol 43 (1) ◽  
pp. 353-364
Author(s):  
Jaydene McDaniel ◽  
Steven McDaniel ◽  
Beanca Jhanine Samiano ◽  
Matthew Marrujo ◽  
Karl Kingsley ◽  
...  
Keyword(s):  
Low Socioeconomic Status ◽  
Gingival Crevicular Fluid ◽  
Clinical Information ◽  
Periodontal Pathogen ◽  
Site Specific ◽  
Low Socioeconomic ◽  
Oral Site ◽  
Crevicular Fluid ◽  
Selenomonas Noxia ◽  
Positive Results

Introduction: Selenomonas noxia (SN) is an important periodontal pathogen, associated with gingivitis and periodontitis. Many studies have found associations between SN and indicators of poor health outcomes, such as smoking, low socioeconomic status and obesity. However, less is known about the prevalence of this organism and more specifically about other oral site-specific locations that may harbor this organism. Methods: Using an existing patient repository (n = 47) of DNA isolated from saliva and other oral sites (n = 235), including the dorsum of the tongue, lower lingual incisor, upper buccal molar and gingival crevicular fluid (GCF), molecular screening for SN was performed. Screening results were analyzed for associations between demographic variables (age, sex, race/ethnicity) and clinical information (body mass index or BMI, presence of orthodontic brackets, primary/mixed/permanent dentition). Results: qPCR screening revealed a total of n = 62/235 sites or 26.3% harboring SN with saliva and GCF (either alone or in combination with one or more sites) most often observed (Saliva, n = 23/27 or 85.18%, GCF, n = 14/27 or 51%). Analysis of site-specific data revealed most positive results were found among saliva and GCF alone or in combination, with fewer positive results observed among the tongue (33.3%), lower lingual incisor (29.6%), and upper buccal molar (25.9%). No significant associations were found between demographic or clinical variables and presence of SN at any site. Conclusions: These results may be among the first to describe site-specific locations of S. noxia among various additional oral biofilm sites. These data may represent a significant advancement in our understanding of the sites and locations that harbor this organism, which may be important for our understanding of the prevalence and distribution of these organisms among patients of different ages undergoing different types of oral treatments, such as orthodontic treatment or therapy.

Matrix metalloproteinases and myeloperoxidase in gingival crevicular fluid provide site-specific diagnostic value for chronic periodontitis

2014 ◽  
Vol 41 (4) ◽  
pp. 348-356 ◽  
Author(s):  
Jussi M. Leppilahti ◽  
Patricia A. Hernández-Ríos ◽  
Jorge A. Gamonal ◽  
Taina Tervahartiala ◽  
Romina Brignardello-Petersen ◽  
...  
Keyword(s):  
Matrix Metalloproteinases ◽  
Chronic Periodontitis ◽  
Gingival Crevicular Fluid ◽  
Diagnostic Value ◽  
Site Specific ◽  
Crevicular Fluid
Sign in / Sign up

Export Citation Format

Share Document