Bacterial Identification via 16S rRNA Mass Spectrometry
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Solution Overview
Problem
Current methods for identifying and characterizing infectious agents, particularly bacteria, are time-consuming and lack the breadth to handle the vast variety of organisms that can cause human disease, including bioterrorism threats and naturally emerging pathogens, due to reliance on culturing and sequencing, which are labor-intensive and not suited for rapid detection.
Innovation Solution
The development of compositions and kits containing oligonucleotide primers that amplify bacterial bioagent identifying amplicons, allowing for identification and quantification by molecular mass and base composition analysis, using primer pairs with specific sequences to produce amplification products that can be matched against known profiles for rapid identification and quantification of bacteria like Staphylococcus aureus.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If PCR-based methods with specific primers and probes are used to detect pathogenic organisms, then detection specificity is improved, but the ability to handle the vast variety of organisms is worsened
Solution Approach 1:
The patent employs universal 16S rRNA primers that can amplify DNA from all bacteria, making the system multi-functional for detecting any bacterial pathogen rather than requiring separate assays for each organism. This universal approach allows simultaneous detection of diverse bacteria including bioterrorism threats and naturally emerging pathogens.
Solution Approach 2:
The patent segments the bacterial identification process into two stages: first amplifying universal 16S rRNA regions to obtain broad coverage, then using variable region analysis and molecular mass spectrometry to differentiate specific species and strains. This segmentation enables both broad versatility and precise identification.
2Measurement precision
If culturing and sequencing methods are used for bacterial identification, then identification accuracy is improved, but time consumption and labor intensity are worsened
Solution Approach 1:
The patent replaces the mechanical culturing process with molecular amplification (PCR) and mass spectrometry detection. Instead of waiting for bacterial growth on culture media, the system directly amplifies and analyzes bacterial DNA, eliminating the time-consuming culturing step while maintaining high identification accuracy.
Solution Approach 2:
The patent performs preliminary amplification of the 16S rRNA gene region before detection, preparing the target DNA in advance to enable rapid subsequent analysis by mass spectrometry. This preliminary action allows the system to bypass the lengthy culturing process while ensuring sufficient target material is available for accurate identification.
3Speed
If DNA microarrays with specific probes are used for bacterial diagnosis, then detection speed is improved, but the breadth of function is worsened
Solution Approach 1:
The patent uses universal primers that target conserved regions of the 16S rRNA gene present in all bacteria, enabling a single assay to detect any bacterial pathogen. This universal approach provides broad functional coverage that DNA microarrays with organism-specific probes cannot achieve, while maintaining rapid detection through PCR amplification and mass spectrometry.
Solution Approach 2:
The patent changes the detection parameter from probe hybridization (used in DNA microarrays) to molecular mass spectrometry of amplified products. This parameter change enables rapid detection while maintaining the ability to identify diverse bacteria through their unique 16S rRNA gene sequences, providing both speed and versatility.
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
Enables rapid and accurate identification and quantification of bacteria, overcoming the limitations of existing methods by providing a rapid, automated process for distinguishing between various bacterial species and strains, including those with similar sequences, facilitating timely public health and biodefense responses.
Implementation Method 1
The present invention provides compositions and kits containing oligonucleotide primers, which define bacterial bioagent identifying amplicons and, upon amplification, produce corresponding amplification products
Implementation Method 2
determining the molecular mass of said amplification product
Data Source
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AI summary
The present invention provides compositions, kits and methods for rapid identification and quantification of bacteria by molecular mass and base composition analysis.