Rapid Detection of Escherichia coli O157 and Shiga Toxins by Lateral Flow Immunoassays

Abstract

Shiga toxin-producing Escherichia coli O157:H7 (STEC) cause food-borne illness that may be fatal. STEC strains enumerate two types of potent Shiga toxins (Stx1 and Stx2) that are responsible for causing diseases. It is important to detect the E. coli O157 and Shiga toxins in food to prevent outbreak of diseases. We describe the development of two multi-analyte antibody-based lateral flow immunoassays (LFIA); one for the detection of Stx1 and Stx2 and one for the detection of E. coli O157 that may be used simultaneously to detect pathogenic E. coli O157:H7. The LFIA strips were developed by conjugating nano colloidal gold particles with monoclonal antibodies against Stx1 and Stx2 and anti-lipid A antibodies to capture Shiga toxins and O157 antigen, respectively. Our results indicate that the LFIA for Stx is highly specific and detected Stx1 and Stx2 within three hours of induction of STEC with ciprofloxacin at 37 °C. The limit of detection for E. coli O157 LFIA was found to be 105 CFU/mL in ground beef spiked with the pathogen. The LFIAs are rapid, accurate and easy to use and do not require sophisticated equipment or trained personnel. Following the assay, colored bands on the membrane develop for end-point detection. The LFIAs may be used for screening STEC in food and the environment.

Three Dimensional Vero Cell-Platform for Rapid and Sensitive Screening of Shiga-Toxin Producing Escherichia coli

“…Previously, a similar 3D cell culture system using Ped-2E9 hybridoma B cells significantly improved toxin detection from Listeria monocytogenes , and Bacillus cereus (Banerjee et al, 2008; Banerjee and Bhunia, 2010). Overall, a 6 h 3D Vero cell-based assay is more competitive than the previously modified 2D Vero cell-based assay (12–16 h) (Roberts et al, 2001), and compatible with immunoassays (Tokarskyy and Marshall, 2008; Il-Hoon et al, 2015; Wang et al, 2016; Armstrong et al, 2018), PCR (Cui et al, 2003; Amani et al, 2015; Parsons et al, 2016), and light scattering sensor (Tang et al, 2014), thus provides an attractive alternative approach for the screening of food samples for the presence of STEC.…”

“…Biochemical and physiological characteristics can be used to differentiate STEC O157 from non-pathogenic E. coli ; however, such methods may not be able to distinguish non-O157 STEC from non-pathogenic E. coli (FDA, 2001). Molecular assay tools such as PCR and immunoassays are widely used (Tate and Ward, 2004; Medina et al, 2012; Schrader et al, 2012; Wang et al, 2016; Armstrong et al, 2018) but they may fail to differentiate viable from dead cells or active from inactive toxins. Moreover, these methods may have limited specificity due to the cross-reactivity of antibodies and performance may be hampered by inhibitors from complex food matrices.…”

Lateral flow based immunobiosensors for detection of food contaminants

“…Biochemical characterization and microbiological identification are used as conventional methods for bacteria detection (Byrne, Gilmartin et al 2015). With the emphasis on fast bacterial pathogen detection, lateral flow test has been confirmed to be a rapid and sensitive method for the detection of food-borne pathogens (Keiser and Utzinger 2005;Law, Ab Mutalib et al 2015;Tallent, Hait et al 2015;Wang, Katani et al 2016).…”

Yersinia pseudotuberculosis-derived adhesins

“…Несмотря на то что липид А является наиболее глубоко расположенным компонентом ЛПС, многие грамотрицательные бактерии могут взаимодействовать с объектами внешней среды путем прямого участия липида А. Так, показано, что химический состав липида A ЛПС (степень ацилирования) не только живых, но также и инактивированных формалином клеток Salmonella enterica серовар Typhimurium определяет уровень продукции провоспалительных цитокинов клетками U937 человека [42]. Более того, на основе моноклональных антител к липиду А разработана иммунохимическая тест-система, позволяющая с удовлетворительной чувствительностью (10 5 клеток/ мл) выявлять бактерии Escherichia coli O157 [69]. Вместе с тем многочисленные исследования, направленные на разработку терапевтических средств на основе моноклональных антител к липиду А ряда грамотрицательных бактерий, к настоящему времени не завершились успешными клиническими испытаниями [26].…”