Sandwich-Based Immunosensor for Dual-Mode Detection of Pathogenic F17–Positive Escherichia coli Strains

Abstract

Bacterial diseases cause tremendous economic losses due to high morbidity and mortality in livestock animals. F17A protein, the major subunit of F17 fimbriae, is one of the most prevalent and crucial virulence factors among the pathogenic Escherichia coli (E. coli) isolated from diarrheic and septicemic animals of various species. Purification and detection of this protein is regarded as an interesting field of investigation due to its important role as a therapeutic target, such as vaccines, and as a diagnostic tool. In this context, polyclonal rabbit antibodies recognizing F17A protein (anti−F17A antibody) were developed and used for its detection. In fact, sandwich biosensor using anti−F17A/gold nanoparticles conjugates as capture probe and anti−F17A antibody labelled with horseradish peroxidase as signal amplification probe was developed for electrochemical and fluorescent detection of purified F17A protein and live F17–positive E. coli bacteria. Good specificity and sensitivity for detection of F17–positive E. coli strains were obtained. The dynamic range for the biosensor varies from 1 × 102 to 1 × 109 CFU·mL−1 (R2 = 0.998) and the detection limit (LOD) and the IC50 value were estimated to be 37 CFU·mL−1 and 75 CFU·mL−1, respectively.

A Study of the Resistance of Hu Sheep Lambs to Escherichia coli F17 Based on Whole Genome Sequencing

“…The rapid spread of sheep diseases has led to an increase in the culling of sheep and losses to breeding farms. Escherichia coli infections are a major threat to young animals and can be fatal in severe cases [ 51 , 52 ]. Therefore, the breeding of high quality disease-resistant sheep is an important issue, and the use of key genes or candidate sites that influence the occurrence of sheep diseases can improve the economic benefits of breeding enterprises and promote the healthy development of the sheep industry.…”

Nanobody-Based Sandwich Immunoassay for Pathogenic Escherichia coli F17 Strain Detection

“…Two microplates were coated overnight at 4 • C with 100 µL per well of each of the purified nanobodies diluted in PBS to a final concentration of 10 µg/mL. Post blocking with 5% skimmed milk in PBS for 2 h at RT, different concentrations (10 4 to 10 9 cells diluted in PBS buffer) of the E. coli F17 strain [23] and the control strain BL-21(DE3) were added and incubated for 1 h at 37 • C. A rabbit anti-F17A polyclonal antibody [12] diluted to 1/1000 was applied for 1 h at 37 • C. An anti-rabbit-HRP conjugated antibody (Invitrogen, Carlsbad, CA, USA) diluted to 1/5000 in PBS was used for detection. Three washes with PBS buffer were performed after each incubation.…”

“…As a result, researchers have suggested that this approach can reduce antibiotic use and mortality rates [11]. To develop biosensors for pathogenic bacteria, various signal transmission techniques have been investigated, including electrochemiluminescence, piezoelectric quartz crystal microbalance, and electrochemical approaches [12].…”

A Novel Multiepitope Fusion Antigen as a Vaccine Candidate for the Prevention of Enterotoxigenic E. coli-Induced Calf Diarrhea

“…For a long time, F5 and F41 fimbriae have been recognized as the predominant pathogenic 2 of 11 factors in diarrheal ETEC, affecting different domestic animals [8,9]. In contrast, F17 fimbriae are associated with diarrhea and septicemia, commonly found in ETEC strains from ruminants [10]. Furthermore, recent epidemiological data have highlighted significant shifts in the prevalence of fimbrial types associated with calf diarrhea, shown by a decline in F5 and F41 fimbriae, while the emergence of F17 fimbriae was observed as the most common fimbriae in calf ETEC infection [11,12].…”