Hybrid Barcoding for Multi-Platform mNGS Pathogen Detection
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Solution Overview
Problem
Current diagnostic methods for detecting causative microorganisms in severe infections are challenging due to indistinguishable clinical manifestations, slow growth of certain pathogens, and limitations in molecular detection techniques, leading to delayed and inaccurate diagnosis.
Innovation Solution
The development of a metagenomic next-generation sequencing (mNGS) method using cell-free DNA from body fluids, which employs hybrid protocols and barcoding schemes compatible with multiple sequencing platforms, enabling rapid and accurate identification of pathogens through Illumina and nanopore sequencing.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Loss of time
If traditional diagnostic methods (culture, PCR) are used, then pathogen detection can be performed, but diagnosis is delayed and accuracy is limited due to slow growth and hypothesis-driven approach
Solution Approach 1:
The patent applies preliminary action by performing metagenomic sequencing on cell-free DNA from body fluids before pathogen isolation or culture growth is complete. This allows detection of pathogens in their environmental DNA form, eliminating the waiting period for slow-growing organisms to multiply to detectable levels in culture.
Solution Approach 2:
The patent replaces traditional mechanical culture-based detection systems with molecular sequencing technology. Instead of relying on physical growth and visual identification, the system uses next-generation sequencing to directly detect and identify pathogen DNA sequences, dramatically reducing detection time and expanding detectable pathogen range.
2Adaptability or versatility
If single-platform sequencing is used, then protocol complexity is reduced, but adaptability to different sequencing technologies is limited
Solution Approach 1:
The patent implements universality by designing a single barcoding scheme and library preparation protocol that is compatible with multiple sequencing platforms (Illumina, Oxford Nanopore, PacBio). The universal barcode design allows the same prepared library to be sequenced on different platforms without requiring platform-specific protocol modifications, enabling multi-platform adaptability.
Solution Approach 2:
The patent applies segmentation by dividing the barcode into distinct functional domains: a first domain compatible with bridge amplification technologies and a second domain compatible with nanopore sequencing. This segmented barcode structure allows each domain to be optimized for its respective platform while maintaining overall compatibility across multiple sequencing systems.
3Adaptability or versatility
If barcodes are designed for single platform, then barcode design is simplified, but cross-platform compatibility is lost leading to barcode crosstalk
Solution Approach 1:
The patent segments the barcode sequence into distinct functional domains: a first domain (4-12 nucleotides) optimized for bridge amplification compatibility and a second domain (14-35 nucleotides) optimized for nanopore sequencing compatibility. This segmentation allows each domain to be independently designed for its target platform while maintaining overall barcode functionality across multiple platforms.
Solution Approach 2:
The patent applies parameter changes by systematically varying the nucleotide sequences of different barcode domains to achieve optimal compatibility with respective platforms. The first domain uses parameters optimized for Illumina bridge amplification (shorter length, specific sequence characteristics) while the second domain uses parameters optimized for nanopore sequencing (longer length, different sequence properties), with minimum Levenshtein distance constraints applied to prevent crosstalk.
Data Source
AI summary
The disclosure provides for hybrid protocols and barcoding schemes that allow for sequencing of targeted polynucleotides in multiple types of sequencing platforms, and applications thereof, including for metagenomic analysis.