Abstract
Clostridium difficile (C. diff) infection is one of the most contagious diseases associated with high morbidity and mortality rates in hospitalised patients. Accurate diagnosis can slow its spread by determining the most effective treatment. Herein, we report a novel testing platform as a proof-of-concept for the selective, sensitive, rapid and cost-effective diagnosis of C. diff infection (CDI) based on a duplex measurement. This was achieved by detecting two specific biomarkers, surface layer protein A (SlpA) and toxin B (ToxB), using a surface enhanced Raman scattering-based lateral flow assay (SERS-based LFA). The simultaneous duplex detection of SlpA with ToxB has not been described for the clinical diagnosis of CDI previously. The SlpA biomarker "AKDGSTKEDQLVDALA" was first reported by our group in 2018 as a species-specific identification tool. The second biomarker, ToxB, is the essential virulence biomarker of C. diff pathogenic strains and is required to confirm true infection pathogenicity. The proposed SERS-based LFA platform enabled rapid duplex detection of SlpA and ToxB on separate test lines using a duplex LF test strip within 20 minutes. The use of a handheld Raman spectrometer to scan test lines allowed for the highly sensitive quantitative detection of both biomarkers with a lowest observable concentration of 0.01 pg μL-1. The use of a handheld device in this SERS-based LFA instead of benchtop machine paves the way for rapid, selective, sensitive and cheap clinical evaluation of CDI at the point of care (POC) with minimal sample backlog.
Highlights
Clostridium difficile (C. diff ) is a Gram-positive species of spore-forming bacteria, that is considered the main cause of infectious diarrhoea in hospitalised patients.[1]
We have successfully demonstrated that surface layer protein A (SlpA) is considered as a unique biomarker for C. diff species only which presents in all C. diff strains sequenced to-date
We presented a novel duplex surface enhanced Raman scattering (SERS)-based lateral flow assay (LFA) platform for the selective and sensitive quantification of the C. diff biomarkers, SlpA and toxin B (ToxB) within 20 minutes
Summary
Isolates, so toxin B must be detected to cover all pathogenic isolates.[3,4,5] The detection of the toxins level during CDI to evaluate the infection severity remains controversial.[6]. Due to their complexity and delayed turnaround time, both methods are unsuitable for the clinical testing of CDI at POC.[6] In 2013, a large UK-based study was carried out on 12 420 faecal samples to validate these two methods The study considered both methods as having inadequate specificity, insufficient sensitivity and noted that most UK diagnostic laboratories do not use these tests.[14] The study indicated that the toxin detection is the most crucial step in the diagnosis of CDI, and concluded that the deficiencies of existing tests should drive further assay development.[14] In the UK, the current testing and diagnosis protocol for CDI is a two-step testing algorithm, that consists of a GDH enzyme immunoassay (EIA) or molecular assay to screen samples, followed by toxins A/B EIA if the first test in the algorithm is positive.[15] simple and user-friendly membrane EIA16 and commercially available lateral flow (LF)[17] tests have been introduced to the diagnostic market for the simultaneous testing of GDH and toxins A/B in a single system (3 in 1). The use of a handheld Raman spectrometer with this novel duplex LFA, paves the way to move the C. diff diagnostic test from localised laboratories to POC application.[30,31,32] This would allow hospital staff to implement infectious disease control measures in a timely manner, which could lead to more cost-effective systems.[33]
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
