Host-Modifying CRISPR/Cas for Selective Bacterial Ratio Control
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Solution Overview
Problem
Existing technologies struggle to effectively inhibit bacterial population growth and alter the relative ratios of different bacterial species in mixed populations, particularly in environments such as human microbiota, leading to issues like antibiotic resistance and microbiologically influenced corrosion.
Innovation Solution
Utilizing a host modifying (HM) CRISPR/Cas system that harnesses endogenous Cas nuclease activity, comprising engineered CRISPR arrays and guide RNAs to selectively target and inhibit the growth of specific bacterial strains while sparing others, achieved through a combination of nucleic acid sequences and vectors that transform host cells.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If conventional methods are used to inhibit bacterial population growth, then some bacterial growth is suppressed, but antibiotic resistance develops and the ability to selectively target specific species is lost
Solution Approach 1:
The invention segments the bacterial population into targetable units by designing species-specific CRISPR guide RNAs that recognize unique sequences in each bacterial species. This allows selective inhibition of specific species (e.g., S. mutans) while sparing others (e.g., S. salivarius) in the same ecological niche, preventing the development of broad-spectrum resistance.
Solution Approach 2:
The CRISPR/Cas system acts as an intermediary mechanism between the administered guide RNA and the target bacterial population. The guide RNA serves as a specific mediator that directs the Cas nuclease to cleave only the DNA of target species, enabling precise control without the non-specific effects that lead to resistance.
2Quantity of substance
If broad-spectrum antibiotics are used to control bacterial infections, then overall bacterial load is reduced, but the relative ratios of different bacterial species cannot be precisely altered
Solution Approach 1:
The invention applies local quality by tailoring the CRISPR guide RNA sequence to match specific genomic regions of target bacterial species. Each guide RNA is designed with local sequence complementarity to the target species' DNA, enabling precise species-specific inhibition while maintaining the overall bacterial ecosystem balance.
3Adaptability or versatility
If genetic modification of host cells is performed to enable CRISPR activity, then selective targeting capability is achieved, but the complexity of the system increases
Solution Approach 1:
The invention leverages the host cell's own endogenous Cas nuclease activity rather than requiring introduction of exogenous Cas proteins. The host cell naturally produces the Cas enzyme, and the administered guide RNA simply needs to direct this existing enzyme to the target sequence, significantly simplifying the system compared to full CRISPR/Cas9 introduction.
4Ease of operation
If existing CRISPR systems are used without endogenous Cas utilization, then external control is achieved, but the efficiency and selectivity are reduced
Solution Approach 1:
The invention makes the CRISPR system universal by utilizing the host cell's own Cas nuclease that can recognize and process the administered guide RNA. This multi-functional approach allows the same endogenous Cas enzyme to target different bacterial species by simply changing the guide RNA sequence, achieving both ease of operation and high reliability.
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
The system achieves selective growth inhibition of up to 10-fold in mixed bacterial populations, allowing for the alteration of bacterial ratios and reducing pathogenic infections or microbiologically influenced corrosion without prior genetic modification of host cells.
Implementation Method 1
the CRISPR array and guide RNA hybridise to a target nucleic acid sequence
Implementation Method 2
a Cas nuclease in the host cell to modify the target sequence
Data Source
AI summary
The invention relates to methods, uses, systems, arrays, engineered nucleotide sequences and vectors for inhibiting bacterial population growth or for altering the relative ratio of sub-populations of first and second bacteria in a mixed population of bacteria. The invention is particularly useful, for example, for treatment of microbes such as for environmental, medical, food and beverage use. The invention relates inter alia to methods of controlling microbiologically influenced corrosion (MIC) or biofouling of a substrate or fluid in an industrial or domestic system.