Microorganism Quantification via Nucleic Acid Detection

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Solution Overview

Problem

Current methods for detecting microorganisms in food samples are inaccurate and time-consuming, requiring long enrichment periods and limited sample sizes, which hinders real-time quantification and process intervention in the food industry.

Innovation Solution

A method involving inoculation of samples with logarithmic serial dilutions in a quantification medium, followed by incubation and detection of target nucleic acid sequences at various temperatures and time points to generate a linear best fit curve for precise microorganism quantification, allowing for real-time detection and improved accuracy.

Engineering Contradictions & Design Principles

VSEngineering Contradiction Analysis

1Measurement precision

If conventional plating or MPN methods are used for microorganism enumeration, then microorganism concentration can be estimated, but detection time is extended to two to five days and accuracy is limited due to statistical probability and broad overlapping ranges

Engineering Contradiction:

Improvemicroorganism quantification accuracyVSAvoiddetection time

Core Design Contradiction:

Measurement precisionVSLoss of time

Solution Approach 1:

The patent changes the detection parameter from traditional culture-based methods to nucleic acid detection methods (PCR, qPCR, LAMP), enabling detection within hours rather than days. This parameter change in detection methodology resolves the contradiction by achieving both rapid detection and improved accuracy through molecular-level identification.

Inventive Principle:

Principle #35Parameter changes

Solution Approach 2:

The patent replaces the mechanical/culture-based enumeration system with a molecular detection system. Instead of relying on microbial growth in culture media over days, the invention uses nucleic acid amplification and detection to achieve rapid quantification, thereby resolving the time-accuracy contradiction.

Inventive Principle:

Principle #28Mechanics substitution (Replace mechanical system)

2Reliability

If selective media are used to recover stressed cells, then microorganism detection sensitivity improves, but cell growth rate slows thereby increasing enrichment time to more than 24 hours

Engineering Contradiction:

Improvemicroorganism detection sensitivityVSAvoidenrichment time

Core Design Contradiction:

ReliabilityVSDuration of action of moving object

Solution Approach 1:

The patent replaces the biological enrichment process (which requires selective media and extended incubation) with a direct molecular detection approach. By detecting nucleic acids from stressed cells without requiring prolonged selective enrichment, the invention achieves both high sensitivity and rapid results, resolving the contradiction between detection sensitivity and enrichment time.

Inventive Principle:

Principle #28Mechanics substitution (Replace mechanical system)

Solution Approach 2:

The patent performs nucleic acid extraction and detection as a preliminary action that does not depend on extended microbial growth. By preparing detection reagents and setting up molecular assays in advance, the system achieves rapid detection without waiting for prolonged enrichment periods, thereby resolving the time-sensitivity contradiction.

Inventive Principle:

Principle #10Preliminary action

3Measurement precision

If MPN method with large sample to media ratio (approximately 1:10) is used, then statistical probability improves, but sample size is limited to about 25 g to about 100 g

Engineering Contradiction:

Improvestatistical probability accuracyVSAvoidsample size

Core Design Contradiction:

Measurement precisionVSQuantity of substance

Solution Approach 1:

The patent replaces the MPN statistical estimation method with direct molecular detection. This substitution eliminates the need for replicate liquid broth growth and statistical probability calculations, allowing accurate quantification of microorganisms in larger sample sizes without being constrained by the 1:10 sample-to-media ratio limitation.

Inventive Principle:

Principle #28Mechanics substitution (Replace mechanical system)

Solution Approach 2:

The molecular detection platform described in the patent is universally applicable to various sample types and sizes. Unlike the MPN method which is constrained to specific sample size ranges, the invention can process diverse sample matrices (foods, environmental samples, clinical specimens) in various quantities, resolving the contradiction between statistical accuracy and sample size flexibility.

Inventive Principle:

Principle #6Universality (Multi-functionality)

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 enables rapid and accurate quantification of microorganisms, reducing detection time and improving the food industry's ability to intervene in contamination processes, providing better food safety through precise enumeration and real-time monitoring.

Implementation Method 1

detecting a target nucleic acid sequence of the microorganism in each aliquot by the detection assay

Methodology Applied

Scientific EffectNucleic acid amplification:

Implementation Method 2

detecting, if present, a target nucleic acid sequence of the microorganism in the test sample by a detection assay

Methodology Applied

Scientific EffectPCR amplification:

Data Source

PatentUS11634782B2Quantification of microorganisms in samples and methods of determining quantification conditions thereof

Publication Date: 2023.04.25 HYGIENA LLC

AI summary

This disclosure relates to methods of determining quantification conditions for a microorganism and methods of quantifying microorganism concentration in a sample.