Multiplex PCR Assay for Borrelia Detection
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Solution Overview
Problem
Current methods for detecting Lyme disease face challenges such as co-infection complications, unspecific testing leading to false positives, and limited sensitivity due to low-level bacteremia, making rapid and accurate diagnosis difficult.
Innovation Solution
A method involving PCR amplification using primer pairs targeting specific regions of Borrelia species' DNA and RNA, such as 16S rRNA, flaB, ospA, ospB, ospC, and other genes, followed by analysis using next-generation sequencing to detect Borrelia species causing Lyme disease or tick-borne relapsing fever.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If conventional PCR methods are used for Lyme disease detection, then the testing process is simple, but the sensitivity is limited due to low-level bacteremia
Solution Approach 1:
The patent combines multiple detection targets (16S rRNA, flaB, ospA, ospB, ospC genes) into a single multiplex PCR assay, allowing simultaneous detection of different Borrelia species and stages in one test run. This merging approach increases detection sensitivity without proportionally increasing testing complexity, as all targets are amplified and detected in a unified workflow rather than requiring separate tests for each target.
Solution Approach 2:
The patent develops a universal detection system that can identify multiple Borrelia species (B. burgdorferi, B. garinii, B. afzelii, B. mayonii) and different gene targets using a single PCR platform. The primers and probes are designed to be universally applicable across different Borrelia strains and infection stages, making the test multi-functional and highly sensitive without requiring multiple specialized assays.
2Measurement precision
If conventional Lyme disease tests are used, then the testing is straightforward, but false positives occur due to unspecific testing
Solution Approach 1:
The patent employs highly specific primers and probes that target unique sequences within specific regions of Borrelia genes (16S rRNA, flaB, ospA, ospB, ospC). Each primer-probe combination is designed to bind only to its intended target sequence, providing local specificity at the molecular level. This ensures that only the intended Borrelia species or gene variant is detected, eliminating false positives from unspecific binding while maintaining a relatively simple PCR-based assay format.
3Adaptability or versatility
If conventional detection methods are used, then the procedure is simple, but co-infection complications cannot be resolved
Solution Approach 1:
The patent merges multiple detection capabilities into a single multiplex PCR assay that can simultaneously detect different Borrelia species (B. burgdorferi, B. garinii, B. afzelii, B. mayonii) and different gene targets (16S rRNA, flaB, ospA, ospB, ospC) in one reaction. This allows clinicians to identify co-infections with multiple Borrelia species or different stages of infection from a single patient sample, providing comprehensive diagnostic information without requiring multiple separate tests.
Applied Scientific Principles
This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.
Function Achieved in This Case
This approach enhances diagnostic accuracy and sensitivity, overcoming co-infection confounding and false positives, enabling rapid identification of Borrelia species in clinical samples.
Implementation Method 1
PCR amplification using primer pairs targeting specific regions of Borrelia species' DNA and RNA
Data Source
AI summary
The present invention relates to method of detecting and characterizing one or more Borrelia species causing Lyme Disease or tick-borne relapsing fever within a sample from a subject, the method comprising: a) subjecting DNA and/or RNA from the sample to a PCR amplification reaction using primer pairs targeting at least one region of Borrelia 16S rRNA and at least one region of flaB, ospA, ospB, ospC, glpQ, 16S-23S intergenic spacer (IGS1), 5S-23S intergenic spacer (IGS2), bbk32, dbpA, dbpB, and/or p66; and b) analyzing amplification products resulting from the PCR amplification reaction to detect the one or more Borrelia species.