Diversi Lab Analysis

The DiversiLab System is integrated with the Web-based software that is used for rep-PCR fingerprint analysis. Data from the microfluidics chip are viewed

FIGURE 26.2. An example DiversiLab Report. Each report provides an interactive dendrogram and similarity matrix. For each sample, up to four demographic fields can be displayed, and any one of the fields can be color-coded to aid data interpretation. An overlay of any two samples is generated by clicking the corresponding box in the similarity matrix.

FIGURE 26.2. An example DiversiLab Report. Each report provides an interactive dendrogram and similarity matrix. For each sample, up to four demographic fields can be displayed, and any one of the fields can be color-coded to aid data interpretation. An overlay of any two samples is generated by clicking the corresponding box in the similarity matrix.

in real-time and are automatically uploaded to a personalized, secured customer Web site. The data can then be accessed either from the DiversiLab System or any Internet-connected computer. Once imported into the software, each sample pattern is normalized based on marker position and viewed for quality control. Acceptable rep-PCR results can then be used to infer relationships among microorganisms based on the similarities of the fingerprint patterns. The similarities between all possible pairs of samples in a data set are calculated by two methods: the Pearson correlation coefficient or a modified Kullback-Leibler coefficient. Both methods compare fluorescent intensity at each data point between a pair of fingerprint patterns.

The DiversiLab software facilitates sample comparisons by including a number of interpretation tools including a dendrogram, an interactive similarity matrix, a scatterplot, electropherograms, overlays, virtual gel images, and selectable demographic fields (Fig. 26.2). The dendrogram is a tree-like diagram that uses branching and branch length to indicate average percentages of similarities between samples and sample clusters that show like fingerprints. Each dendrogram can include up to four demographic fields (such as species, strain, location, and date), one of which can be color-coded to reveal sample clustering. The similarity matrix also provides percentages of similarity, but the matrix format allows the direct comparison of each sample pair. The similarity matrix also allows an overlay of any two sample graphs to easily be generated for direct examination of the fingerprint pattern differences. When comparing new data to a user-defined or DiversiLab reference library, a top-match module allows comparisons to be seen as a table instead of in a more complicated dendrogram. Improved assay repro-ducibility and Web-based software make it possible to share user-defined libraries between collaborators or multisite facilities, if so desired.

Automation of the DiversiLab System allows sample processing, analysis, and report generation to be completed for 13 samples in approximately 4 h with an additional hour for every 13 extra samples (Healy et al., 2005). Strain-level discrimination using the DiversiLab System has been seen with mycobacteria (Cangelosi et al., 2004; Gira et al., 2004), Staphylococcus aureus (Shutt et al., 2005), Lactobacillus (Land et al., 2005), Aspergillus (Healy et al., 2004), zygomycetes (D. Kontoyiannis et al., 2005), dermatophytes (Pounder et al., 2005), and Candida (Chau et al., 2004; Li et al., 2004). The DiversiLab System is also applicable to atypical organisms, including archeabacteria, mycobacteria (ATCC, Technical Bulletin, 2005), and anaerobes (Spigaglia and Mastrantonio, 2003). This wide utility makes DiversiLab an excellent tool for the clinical microbiology laboratory.

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