Spherical Nucleic Acid Enhanced FO-SPR DNA Melting for Detection of Mutations inLegionella pneumophila

Abstract

A home-built fiber optic surface plasmon resonance platform (FO-SPR) was applied to directly screen PCR amplified DNA for mutations. The FO-SPR sensor was used for real-time monitoring of DNA duplex melting during high resolution temperature cycling. The signal of the DNA melting was enhanced by means of gold nanoparticle labels. This FO-SPR genetic assay allowed for detection of single-point mutations (SNP) in less than 20 min. The concept was demonstrated for the analysis of 9 different serogroups of the bacterium Legionella pneumophila, a common human pathogen responsible for atypical pneumonia. FO-SPR allowed us to detect genetic mutations inhibiting PCR, which could lead to amplification bias when molecular diagnostics are applied for L. pneumophila detection. All serogroups were found to display unique melting temperatures, indicating that mutations have accumulated in the target sequence. In a next step, clinical samples of L. pneumophila were analyzed using the FO-SPR sensor. This technology was proven to be reliable for the detection of mutations for those samples that previously displayed ambiguous qPCR quantification results. When these results were benchmarked, FO-SPR results were found to be consistent with Sanger sequencing but not with fluorescence based DNA melting. The presented results convincingly advocate the advantages of FO-SPR as a high resolution and fast genetic screening tool that can compete with the current standard techniques for SNP detection.

Current and Emerging Legionella Diagnostics for Laboratory and Outbreak Investigations

“…Recent studies have shown HRM analysis to be a powerful technique for characterizing antibiotic resistance and typing respiratory disease agents, including M. pneumoniae, C. pneumoniae, and Chlamydophila psittaci (453)(454)(455)(456)(457)(458)(459). To our knowledge, HRM analysis for intraspecies or interspecies Legionella discrimination has been included in only a few studies (460)(461)(462)(463); however, this approach seems promising for future Legionella diagnostics or typing.…”

Fiber optic-SPR platform for fast and sensitive infliximab detection in serum of inflammatory bowel disease patients

“…Standard optical fibers have emerged over the years as excellent alternatives that can compete with the sensitivity and specificity of prism-based SPR sensors, while offering the advantage of low cost and small size greatly desired for developing a POC tool. One such platform, namely fiber optic surface plasmon resonance (FO-SPR), has been previously described by our group as a promising biosensor for protein-and DNA-based assays (Delport et al 2012;Janssen et al 2012;Knez et al 2013;Knez et al 2014;Pollet et al 2011;Pollet et al 2009;Tran et al 2011). The FO-SPR biosensor makes use of multimode optical fibers that facilitate SPR generation and enable implementation of bioassays.…”

Thermal annealing of gold coated fiber optic surfaces for improved plasmonic biosensing

“…Furthermore, coupling SPR to fiber optic (FO) technology triggered notable advancements in the field, by offering attractive advantages over the classical prism-based SPR platforms, such as simplicity, portability, cost-effectiveness and miniaturization [2]. FO-SPR sensors were successfully used already in a variety of applications, such as screening for bacterial infections [3], monitoring of environmental water pollution [4] or checking for specific toxins [5] and allergens [6] in food, among others. However, their performance still needs to be improved in order to match traditional prismbased SPR sensors, and consequently, to switch from laboratory proofs-of-principle to commercial products.…”