ABSTRACT
The USDA-FSIS has zero tolerance for E. coli O157:H7 in raw ground beef. Currently, FSIS collects samples from beef processing facilities and ships them overnight to regional testing laboratories. Pathogen detection requires robust methods that employ an initial 15-24 h culture enrichment. This study assessed the potential of using the ΦV10nluc phage-based luminescence detection assay during enrichment while the sample is in transit. Parameters including phage concentrations, temperature, and media-to-sample ratios were evaluated. Results in liquid media showed that 1.73× 103 pfu/mL of ΦV10nluc was able to detect 2 CFU in 10 h. The detection of E. coli O157:H7 was further evaluated in kinetic studies using ratios of 1:3, 1:2, and 1:1 ground beef sample to enrichment media, yielding positive results for as little as 2-3 CFU in 325 g ground beef in about 15 h at 37 °C. These results suggest that this approach is feasible, allowing the detection of a presumptive positive upon arrival of the sample to the testing lab. As the current cargo hold controlled temperature is required to be 15-25 °C, the need for elevated temperature should be easily addressed. If successful, this approach could be expanded to other pathogens and foods.
ABSTRACT
Rapid detection of the foodborne pathogen Escherichia coli O157:H7 is of vital importance for public health worldwide. Among detection methods, reporter phages represent unique and sensitive tools for the detection of E. coli O157:H7 from food as they are host-specific and able to differentiate live cells from dead ones. Upon infection, target bacteria become identifiable since reporter genes are expressed from the engineered phage genome. The E. coli O157:H7 bacteriophage ΦV10 was modified to express NanoLuc luciferase (Nluc) derived from the deep-sea shrimp Oplophorus gracilirostris. Once infected by the ΦV10 reporter phage, E. coli O157:H7 produces a strong bioluminescent signal upon addition of commercial luciferin (Nano-Glo(®)). Enrichment assays using E. coli O157:H7 grown in LB broth with a reporter phage concentration of 1.76 × 10(2) pfu ml(-1) are capable of detecting approximately 5 CFU in 7 hours. Comparable detection was achieved within 9 hours using 9.23 × 10(3) pfu ml(-1) of phage in selective culture enrichments of ground beef as a representative food matrix. Therefore we conclude that this NanoLuc reporter phage assay shows promise for detection of E. coli O157:H7 from food in a simple, fast and sensitive manner.
Subject(s)
Bacteriophages/genetics , Escherichia coli O157/isolation & purification , Food Microbiology/methods , Luciferases/chemistry , Animals , Cattle , Colony Count, Microbial , Escherichia coli O157/pathogenicity , Food Contamination/analysis , Luciferases/genetics , Meat/microbiologyABSTRACT
Banana starch was esterified with octenylsuccinic anhydride (OSA) at different degree substitution (DS) and used to stabilize emulsions. Morphology, emulsion stability, emulsification index, rheological properties and particle size distribution of the emulsions were tested. Emulsions dyed with Solvent Red 26 showed affinity for the oil phase. Backscattering light showed three regions in the emulsion where the emulsified region was present. Starch concentration had higher effect in the emulsification index (EI) than the DS used in the study because similar values were found with OSA-banana and native starches. However, OSA-banana presented greater stability of the emulsified region. Rheological tests in emulsions with OSA-banana showed G'>G" values and low dependence of G' with the frequency, indicating a dominant elastic response to shear. When emulsions were prepared under high-pressure conditions, the emulsions with OSA-banana starch with different DS showed a bimodal distribution of particle size. The emulsion with OSA-banana starch and the low DS showed similar mean droplet diameter than its native counterpart. In contrast, the highest DS led to the highest mean droplet diameter. It is concluded that OSA-banana starch with DS can be used to stabilize specific emulsion types.
