Statistical Analyses of Complex Denaturing Gradient Gel Electrophoresis Profiles

Abstract

Studies using molecular techniques have demonstrated that a culture-based approach can severely underestimate the bacterial diversity in most environments. One of the molecular techniques that has been applied in microbial ecology is denaturing gradient gel electrophoresis (DGGE). The purpose of this study was to investigate differences in the microbiota of plaque, using a number of analysis techniques, from children without gingivitis (n ‫؍‬ 30) and from those with gingivitis (n ‫؍‬ 30). Extracted DNA from gingival margin plaque was subjected to PCR targeting the 16S rRNA gene using universal primers. DGGE profiles were analyzed in three ways. (i) Bacterial diversity was compared between cohorts by using the Shannon-Wiener index (also known as the Shannon-Weaver index). (ii) A hierarchical cluster analysis of the banding patterns was calculated and expressed as a dendrogram. (iii) Individual DGGE bands and their intensities for both cohorts were compared using a logistic regression analysis. The Shannon-Wiener indices demonstrated a greater bacterial diversity associated with no-gingivitis plaque (P ‫؍‬ 0.009). Dendrograms demonstrated that seven clades associated with gingivitis and five clades associated with no gingivitis. The logistic regression demonstrated that one band was significantly associated with no gingivitis (P ‫؍‬ 0.001), while two bands were significantly associated with gingivitis (P ‫؍‬ 0.005 and P ‫؍‬ 0.042). In conclusion, this study demonstrates that the development of gingivitis might be accompanied by a decrease in bacterial diversity. Furthermore, we have demonstrated that logistic regression is a good statistical method for analyzing and characterizing DGGE profiles.

Molecular and Culture-Based Assessment of the Microbial Diversity of Diabetic Chronic Foot Wounds and Contralateral Skin Sites

“…Diversity indices were determined for each individual sample and also for each specimen group (i.e., the presence or absence of cultured wound pathogens) and using the Shannon-Weaver index of diversity (H=) with the following equation, where s is the number of species and P i is the proportion of species in the sample i (13,16):…”

“…The gels were left to equilibrate at room temperature overnight in the tank containing 7 liters of 1ϫ Tris-acetate buffer solution. Gel electrophoresis was carried out at 140 V at a constant 60°C for 5.5 h (16,30). All gels were stained using SYBR Gold stain (Molecular Probes, Leiden, The Netherlands) for 30 min with occasional agitation, after which they were transferred to a UV transilluminator (UVP, Upland, CA), visualized under UV light at 312 nm, and photographed using a Canon D60 digital single lens reflex (DSLR) camera (Canon, Surrey, United Kingdom).…”

Drinking Habits Are Associated with Changes in the Dental Plaque Microbial Community

“…A number of these allow the qualitative or quantitative detection of single bacterial species (13), while others can detect and profile whole heterogeneous populations (25). In order to more fully study the complex oral microbiota, techniques such as denaturing gradient gel electrophoresis (DGGE) can be used (8,9,15,16,26). This technique separates PCR-amplified DNA fragments from a heterogeneous population according to the sequence information.…”

The Effect of Artificial Rearing on Gut Microbiota in a Mouse Pup-in-a-Cup Model

“…A matrix was constructed for the lanes, taking into account the presence or absence of individual bands and the relative contribution of each band to the total intensity of the lane. This matrix was applied to calculate biodiversity indices, such as the number of bands and Shannon index [6]. Comparison was carried out using the Dice similarity coefficient with a position tolerance setting of 1% optimization and a 1% position tolerance for band ingredient.…”

“…The AR pups weighed less than the MR pups at 8 d of age (4.56 ± 0.15 g vs. 4.67 ± 0.11 g; P<0.05) and more than their MR counterparts at 13 d of age (7.00 ± 0.25 g vs. 6.93 ± 0.18 g, P<0.05) and 14 d of age (7.68 ± 0.29 g vs. 7.10 ± 0.24 g, P<0.01). Pup weight is presented as a growth curve in Fig.…”