M. tuberculosis Detection via NarGHJI SNP Analysis
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Solution Overview
Problem
Current methods for detecting Mycobacterium tuberculosis in biological samples are inefficient, requiring lengthy laboratory cultivation and cannot specifically distinguish M. tuberculosis from other members of the M. tuberculosis complex, such as M. bovis, M. africanum, and M. microti, in clinical settings.
Innovation Solution
A nucleic acid amplification method using primers and hybridization probes targeting the narGHJI nitrate reductase operon, specifically detecting a T→C polymorphism at position −215 upstream of the translation start codon, allows for rapid and specific identification of M. tuberculosis through melting curve analysis.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If traditional nitrate reductase testing and cultivation methods are used to identify M. tuberculosis complex members, then specific identification can be achieved, but the process requires specialized laboratory equipment, takes several weeks, and incurs high financial costs
Solution Approach 1:
The patent replaces traditional mechanical/biochemical cultivation and testing methods with a molecular biology-based PCR amplification system. By targeting specific DNA sequences (IS6110 region) and using temperature-controlled thermal cycling instead of weeks-long cultural cultivation, the method achieves rapid specific identification within hours while maintaining diagnostic precision.
Solution Approach 2:
The patent performs preliminary DNA extraction and sequence-specific amplification before final detection. By pre-selecting and amplifying only the relevant IS6110 genomic regions using tailored primers, the method prepares the sample in advance for rapid specific identification, eliminating the need for time-consuming conventional cultivation steps.
2Measurement precision
If traditional nitrate reductase testing is used to distinguish M. tuberculosis from other M. tuberculosis complex members, then specific distinction can be achieved, but it requires cultivation in specially equipped laboratories with high expenditure
Solution Approach 1:
The patent substitutes complex specialized laboratory equipment and biochemical testing systems with a simplified PCR-based molecular detection system. By using universal PCR instruments and sequence-specific primers/probes targeting IS6110 regions, the method achieves the same distinction capability with significantly reduced equipment complexity and infrastructure requirements.
Solution Approach 2:
The patent creates specific DNA copies of the IS6110 region through PCR amplification. These amplified DNA copies serve as specific markers for identifying and distinguishing M. tuberculosis complex members, replacing the need for complex biochemical testing equipment while maintaining high distinction precision through sequence-specific amplification.
3Productivity
If rapid detection methods are implemented to reduce diagnosis time, then detection speed improves, but specific identification capability may be compromised
Solution Approach 1:
The patent performs preliminary amplification of specific IS6110 DNA regions before detection. This pre-amplification step ensures that only relevant target sequences are present in sufficient quantity for rapid detection, maintaining high specific identification accuracy while enabling quick results within hours rather than weeks.
Solution Approach 2:
The patent uses location-specific primers and probes that target the unique IS6110 insertion sequences at specific genomic locations. This local sequence specificity ensures that rapid amplification and detection only occur for M. tuberculosis complex members containing these specific genetic markers, maintaining high identification accuracy despite the speed of the method.
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 method enables rapid, accurate detection of M. tuberculosis in clinical samples, distinguishing it from other members of the M. tuberculosis complex and non-tuberculous mycobacteria, reducing the need for specialized laboratory equipment and improving diagnostic efficiency.
Implementation Method 1
a hybridization probe which comprises a nucleotide sequence which is selected from the group consisting of the nucleotide sequence represented in SEQ ID NO: 5, the complementary sequence to SEQ ID NO: 5, the nucleotide sequence represented in SEQ ID NO: 6 and the complementary sequence to SEQ ID NO: 6
Implementation Method 2
the specific detection of M. tuberculosis vis-à-vis other members of the M. tuberculosis complex taking place by analysis of the melting temperature of the specific hybridization of the hybridization probe with the amplified DNA fragment of the narGHJI operon
Data Source
AI summary
The invention relates to a method and system for the specific detection of a Mycobacterium tuberculosis (M. tuberculosis) in a biological sample, a difference being made, in particular between M. tuberculosis and other elements of M. tuberculosis complex, i.e., Mycobacterium bovis (M. bovis), Mycobacterium bovis BCG (M. bovis BCG), Mycobacterium africanum (M. africanum) and Mycobacterium microti (M. microti) based on a SNP in a narGHJI promoter.