Carbon nanoparticles in lateral flow methods to detect genes encoding virulence factors of Shiga toxin-producing Escherichia coli

• DOI: 10.1007/s00216-010-4334-z
• Publication Date: 2010-11-02T16:44:04Z
• Authors: P. Noguera; G. A. Posthuma-Tr...; M. van Tuil; F. J. van der Wal; A. de Boer; A. P. H. A. Moers; A. van Amerongen
• Keywords: Carbon nanoparticles; Verotoxin; Epidemiology; Economics; Nucleic acid lateral flow immunoassay; 02 engineering and technology; Biomedische technologie en medicijnen; Screening method; Biochemistry; Polymerase Chain Reaction; Analytical Chemistry; Farmacie/Biofarmaceutische wetenschappen (FARM); Nanoparticle; samples; Bos; Instrumentation; Ziekenhuisstructuur en organisatie van de gezondheidszorg; Immunoassay; Toxic materials; Shiga-Toxigenic Escherichia coli; Bacterial gene; Farmacie(FARM); Virulence factor; Polymerase chain reaction; 3. Good health; Nucleic acids; Chemistry; Sensitivity and specificity; statistics; Public Health; 0210 nano-technology; agreement; DNA, Bacterial; Carbon nano particles; Virulence Factors; Shiga toxin producing Escherichia coli; Immunology; Sensitivity and Specificity; Article; pcr; SDG 3 - Good Health and Well-being; molecular diagnosis; QUIMICA ANALITICA; Escherichia coli; Genetics; Animals; immunoassay; particles; Biomolecules; Original Paper; Animal; food; E. coli; Methodology; pathogenic bacteria; biosensors; Shiga toxin; Bacterial DNA; Carbon; Isolation and purification; Genes, Bacterial; Nanoparticles; Cattle

The use of carbon nanoparticles is shown for the detection and identification of different Shiga toxin-producing Escherichia coli virulence factors (vt1, vt2, eae and ehxA) and a 16S control (specific for E. coli) based on the use of lateral flow strips (nucleic acid lateral flow immunoassay, NALFIA). Prior to the detection with NALFIA, a rapid amplification method with tagged primers was applied. In the evaluation of the optimised NALFIA strips, no cross-reactivity was found for any of the antibodies used. The limit of detection was higher than for quantitative PCR (q-PCR), in most cases between 10(4) and 10(5) colony forming units/mL or 0.1-0.9 ng/μL DNA. NALFIA strips were applied to 48 isolates from cattle faeces, and results were compared to those achieved by q-PCR. E. coli virulence factors identified by NALFIA were in very good agreement with those observed in q-PCR, showing in most cases sensitivity and specificity values of 1.0 and an almost perfect agreement between both methods (kappa coefficient larger than 0.9). The results demonstrate that the screening method developed is reliable, cost-effective and user-friendly, and that the procedure is fast as the total time required is <1 h, which includes amplification.