18S rRNA Metabarcoding With Host DNA Depletion for Parasite Detection

Resolve Bottlenecks,
Find Innovative SolutionsGenerate Solutions

Solution Overview

Problem

Current parasite diagnostics for eukaryotic endosymbionts are laborious, expensive, and suffer from limited taxonomic coverage and high levels of interfering host signal, making them inefficient for complex clinical specimens.

Innovation Solution

Development of primer sets targeting the V4 region of 18S rRNA genes and guide RNAs for CRISPR-Cas9 host DNA depletion, along with mock communities, to enhance the accuracy and specificity of eukaryotic endosymbiont detection through metabarcoding.

Engineering Contradictions & Design Principles

VSEngineering Contradiction Analysis

1Adaptability or versatility

If conventional microscopy and single-target assays are used for parasite diagnostics, then detection of known agents is possible, but the process becomes laborious and expensive with limited taxonomic coverage

Engineering Contradiction:

Improvetaxonomic coverageVSAvoiddiagnostic process time

Core Design Contradiction:

Adaptability or versatilityVSLoss of time

Solution Approach 1:

The patent develops universal metabarcoding primer sets that can simultaneously detect multiple eukaryotic endosymbionts across diverse taxonomic groups (protozoa, helminths, arthropods) in a single assay, replacing numerous single-target assays and enabling comprehensive parasite diagnostics with broad taxonomic coverage

Inventive Principle:

Principle #6Universality (Multi-functionality)

Solution Approach 2:

The patent replaces manual microscopy examination with automated metabarcoding sequencing technology, substituting labor-intensive visual inspection with high-throughput molecular detection that provides objective, quantifiable results across multiple taxa simultaneously

Inventive Principle:

Principle #28Mechanics substitution (Replace mechanical system)

2Productivity

If metabarcoding-based approaches are used for parasite detection, then throughput is improved, but high levels of interfering host signal and lack of parasite taxonomic coverage occur

Engineering Contradiction:

Improvedetection throughputVSAvoidsignal specificity

Core Design Contradiction:

ProductivityVSMeasurement precision

Solution Approach 1:

The patent extracts and removes host DNA from complex clinical specimens through selective enrichment methods, isolating eukaryotic endosymbiont DNA from overwhelming host background signal to enable specific detection of parasite taxa with high sensitivity

Inventive Principle:

Principle #2Taking out (Extraction)

Solution Approach 2:

The patent designs primers with specific local sequence characteristics that selectively amplify eukaryotic endosymbiont 18S rRNA genes while avoiding host DNA, using tailored primer sequences optimized for binding to conserved regions in parasite genomes but not in host genomes

Inventive Principle:

Principle #3Local quality

3Adaptability or versatility

If existing metabarcoding methods are used, then simultaneous detection of multiple organisms is possible, but off-target amplification and host interference remain high

Engineering Contradiction:

Improvemulti-organism detection capabilityVSAvoidhost signal interference

Core Design Contradiction:

Adaptability or versatilityVSObject-affected harmful factors

Solution Approach 1:

The patent uses CRISPR-Cas9 guide RNAs as intermediary molecules that specifically bind to and eliminate host DNA sequences, acting as a selective filter that removes interfering host signal while preserving eukaryotic endosymbiont DNA for accurate metabarcoding detection

Inventive Principle:

Principle #24Intermediary (Mediator)

Solution Approach 2:

The patent optimizes multiple parameters including primer melting temperatures, annealing conditions, and CRISPR-Cas9 reaction conditions to maximize specificity for eukaryotic endosymbiont DNA while minimizing off-target amplification and host DNA interference

Inventive Principle:

Principle #35Parameter changes

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 method enables simultaneous, accurate, and comprehensive detection of multiple eukaryotic endosymbionts across various host and sample types, reducing off-target amplification and host interference, thus improving diagnostic efficiency and cost-effectiveness.

Implementation Method 1

amplifying the V4 region of 18S ribosomal RNA (rRNA) genes

Methodology Applied

Scientific EffectDNA amplification:

Implementation Method 2

guide RNAs (gRNAs) that target the V4 region of 18S rRNA genes found in vertebrate organisms

Methodology Applied

Scientific EffectCRISPR-Cas9 host DNA depletion:

Implementation Method 3

sequencing the amplicons to generate sequencing reads

Methodology Applied

Scientific EffectDNA sequencing:

Data Source

PatentUS20260002223A1Universal method for parasite and eukaryotic endosymbiont identification

Publication Date: 2026.01.01 WISCONSIN ALUMNI RES FOUND

AI summary

The present invention provides methods for detecting eukaryotic endosymbionts in a sample. Primer sets, guide RNAs, and mock communities of eukaryotic endosymbionts for use in these methods are also provided. Further, methods for diagnosing and treating a subject with a parasitic infection are also provided.