Application of Nuclear Magnetic Resonance to Detect Toxigenic Clostridium difficile from Stool Specimens

Abstract

We evaluated the performance of an early prototype core molecular mirroring nuclear magnetic resonance detection platform (Mentor-100) to detect toxigenic Clostridium difficile from stool. This technology uses customized nanoparticles bound to target specific oligonucleotide probes that form binaries in the presence of nucleic acid from the target microorganism. Liquid patient stool specimens were seeded with C. difficile or other Clostridium species to determine the analytical sensitivity and specificity. Samples underwent nucleic acid extraction and target amplification with probes conjugated with iron nanoparticles. Signal from nuclear magnetic resonance spin-spin relaxation time was measured to detect the presence or absence of toxigenic C. difficile. The limit of detection was <180 colony forming units per reaction of toxigenic C. difficile. No cross-reactivity was observed with nontoxigenic C. difficile, Clostridium sordellii, Clostridium perfringens, Bacillus subtilis, or Paenibacillus polymyxa at 10 colony forming units/mL. Correlation studies using frozen stool samples yielded a sensitivity of 88.4% (61 of 69) and a specificity of 87.0% (40 of 46) as compared with a commercial PCR assay for C. difficile. The area under the curve in the receiver operating characteristic curve analysis was 0.922. The prototype molecular mirroring platform showed promising performance for pathogen detection from clinical specimens. The platform design has the potential to offer a novel, low-cost alternative to currently available nucleic acid-based tests.

Nuclear magnetic resonance biosensor for rapid detection of Vibrio parahaemolyticus

“…As demonstrated from a series of experiments on bioassay, specificity and sensitivity of the methodology using combined NMR, molecular biology and nanoparticle application, the NMR biosensor is capable of detecting microbial pathogens with high degree of sensitivity and specificity. In addition, this biosensor system can detect a wide range of microorganism DNAs in different matrices within a short period of time, compared with conventional microbiology techniques that require at least 24 h or with conventional PCR that requires electrophoresis of agarose gels preparation in order to obtain qualitative results [27], [28]. The time factor of rapidly detecting microbial pathogen in specimens for food production and consumption is crucial to ensure food safety [29].…”

Food safety in the 21st century

“…Nuclear Magnetic Resonance (NMR) nanotechnology platform detects multiple target microbials hybridizing to pathogen's DNA or protein in same the device chamber that runs assays using nucleic acid, antibodies, and other biomarkers [30] . Orthogonal confirmatory tests can be achieved via multiple biomarkers of single microbial in same detection device.…”

Clostridium difficile Infection Diagnosis by Biological Molecular Methods

“…For this short presentation, we have chosen the latest references that offered the most eloquent data regarding sensitivity, specificity, and, if available, positive predictive value and negative predictive value ( Table 3) [101][102][103][104][105][106][107][108][109][110][111].…”

“…The costs, the clinical consequences, and the impact on the nosocomial transmission of treating and/or isolating patients-positive for toxigenic C. difficile by PCR but negative for in vivo toxin production-worth further study. Diagnostic algorithms combining immunoassays and NAATs could also improve the specificity and reduce the global cost of this analysis [108][109][110].…”