Abstract
The ubiquitous and opportunistic pathogen Cronobacter sakazakii is responsible for severe meningitis, sepsis, and necrotizing enterocolitis in neonates and infants associated with ingestion of contaminated powdered infant formula (PIF). The current ISO method for isolation and detection of Cronobacter spp. is laborious, time-consuming and expensive. In this study, a multiplexed lateral flow test strip was developed to rapidly detect and simultaneously serotype O1 and O2 C. sakazakii serotypes. The assay is based on two monoclonal antibodies (MAb) that specifically bind to the lipopolysaccharides (LPS) of these pathogens. The test strip provides results very quickly; C. sakazakii could be detected in pure culture within 15 min with a sensitivity of 107 CFU/ml. After non-selective enrichment for 18 h as low as one Cronobacter cell per g PIF could be detected. Moreover, the established lateral flow assay (LFA) offers excellent specificity showing no cross-reactivity with other C. sakazakii serotypes, Cronobacter species or Enterobacteriaceae tested. These characteristics, together with several advantages such as speed, simplicity in performance, low analysis cost, and no requirement of specialized skills or sophisticated equipment make the developed multiplexed LFA suitable for reliable detection and serotyping of C. sakazakii serotypes O1 and O2.
Highlights
Cronobacter spp. are opportunistic foodborne pathogens and can cause severe infections in all age groups, with neonates being the most susceptible and affected group for invasive disease (Lai, 2001)
In the case of C. sakazakii serotype O1, DIG, and DNP-labeled antibodies (MAbs 1C4) bind to the LPS structures on the bacterial surface, whereas in the case of C. sakazakii serotype O2 the sandwich complex is composed of the bacteria bound to a DIG- and a biotin-labeled antibodies (MAbs 2F8)
The excess fluid and any unreacted substances are absorbed by the absorbent pad. Observation of both a colored test line 1 or test line 2 and a colored control line on the membrane is a positive result indicating the presence of C. sakazakii serotype O1 or C. sakazakii serotype O2 in the sample
Summary
Cronobacter spp. are opportunistic foodborne pathogens and can cause severe infections in all age groups, with neonates being the most susceptible and affected group for invasive disease (Lai, 2001). The species C. sakazakii consists of five serotypes O1–O4 and O7 (Blažková et al, 2015). The serotypes O1 and O2 are isolated from clinical as well as from food products, suggesting that some serotypes of C. sakazakii might be more virulent than others (Blažková et al, 2015; Yan et al, 2015a,b; Scharinger et al, 2016). LFA for C. sakazakii Serotypes it is important for epidemiological studies to identify the genus and species and the serotype of the pathogen
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