CRISPR-Based FELUDA Diagnostic Tool Detects Antibiotic Resistance in H. pylori Infections

H. pylori infections affect over 43 percent of the global population and are linked to a wide range of gastrointestinal disorders, including peptic ulcers, gastritis, dyspepsia, and gastric cancer.

In a significant advancement for global healthcare, researchers have developed a CRISPR-based diagnostic tool, FELUDA, to detect Helicobacter pylori (H. pylori) infections and its antibiotic resistance mutations with high accuracy and minimal cost. This tool is designed for point-of-care use, especially in resource-limited rural areas of India with minimal access to diagnostic laboratories.

H. pylori infections affect over 43 percent of the global population and are linked to a wide range of gastrointestinal disorders, including peptic ulcers, gastritis, dyspepsia, and gastric cancer. A growing concern is the bacterium’s resistance to clarithromycin, driven by mutations in the 23S ribosomal RNA gene, which complicates treatment by necessitating repeated diagnostic tests and multiple antibiotic regimens.

The Role of CRISPR in Diagnostics

CRISPR-based methodologies, known for their precision in DNA recognition and cleavage, are well-suited for identifying genetic mutations in pathogens. Researchers have harnessed these capabilities to target the specific mutation sites associated with clarithromycin resistance.

Dr. Shraddha Chakraborty (Department of Science and Technology INSPIRE Faculty Fellow at DBEB, IIT Delhi) and her team at CSIR-IGIB, in collaboration with gastroenterologists and microbiologists from AIIMS New Delhi and Bhubaneswar, have explored the diagnostic potential of an engineered Cas9 protein variant, en31-FnCas9. This variant is based on a Cas9 ortholog derived from Francisella novicida and modified to overcome the limitations of conventional Cas9 proteins, such as the dependency on specific PAM sequences for mutation detection.

Breakthrough Findings

In their study, published in the Microchemical Journal, the team demonstrated that en31-FnCas9 can accurately identify H. pylori infections and clarify the mutation status of the 23S rDNA gene in gastric biopsy samples. This was achieved using in vitro cleavage studies and lateral flow test strip assays (FELUDA).

The integration of en31-FnCas9 with FELUDA allowed for a sequencing-free molecular diagnosis, enabling rapid, visual readouts of infection and mutation status. This innovation holds the potential to transform clinical diagnostics, particularly in resource-constrained environments.

Advantages and Impact

This is the first report of en31-FnCas9 being used for the molecular diagnosis of H. pylori mutations linked to clarithromycin resistance. The tool’s key advantages include:

• Cost-Effectiveness: Ideal for rural and underprivileged regions with limited healthcare infrastructure.
• Speed: Provides rapid and accurate results without requiring complex sequencing.
• Accessibility: Enables decentralized diagnostics for timely and effective treatment.

Dr. Debojyoti Chakraborty and Dr. Souvik Maiti, who had previously developed Cas9-based biosensing techniques for detecting H. pylori mutations, emphasized the potential of en31-FnCas9 in addressing the global challenge of antibiotic resistance.

A Path Toward Tailored Treatment

By enabling the detection of antibiotic resistance patterns in H. pylori strains, this tool facilitates personalized treatment strategies, reducing the risk of recurrent infections and complications such as gastric cancer.

Successful implementation of this methodology in clinical settings could revolutionize the management of H. pylori-related conditions, particularly in remote and underserved areas. The study underscores the importance of sequencing-free molecular diagnostics in addressing public health challenges, paving the way for tailored treatments and improved patient outcomes worldwide.

Researchers hope this innovation will serve as a model for the application of advanced molecular tools in tackling other infectious diseases with similar diagnostic and treatment challenges.