Rapid detection of pathogens and hazardous elements in fresh fruits and vegetables after harvest requires the use of advanced sensor technology at each step in the farm-to-consumer or farm-to-processing sequence. Fourier-transform infrared (FTIR) spectroscopy and the complementary Raman spectroscopy, an advanced optical technique based on light scattering will be investigated for rapid and on-site assessment of produce safety. Paving the way toward the development of this innovative methodology, specific original objectives were to (1) identify and distinguish different serotypes of Escherichia coli, Listeria monocytogenes, Salmonella typhimurium, and Bacillus cereus by FTIR and Raman spectroscopy, (2) develop spectroscopic fingerprint patterns and detection methodology for fungi such as Aspergillus, Rhizopus, Fusarium, and Penicillium (3) to validate a universal spectroscopic procedure to detect foodborne pathogens and non-pathogens in food systems. The original objectives proposed were very ambitious hence modifications were necessary to fit with the funding. Elaborate experiments were conducted for sensitivity, additionally, testing a wide range of pathogens (more than selected list proposed) was also necessary to demonstrate the robustness of the instruments, most crucially, algorithms for differentiating a specific organism of interest in mixed cultures was conceptualized and validated, and finally neural network and chemometric models were tested on a variety of applications. Food systems tested were apple juice and buffer systems. Pathogens tested include Enterococcus faecium, Salmonella enteritidis, Salmonella typhimurium, Bacillus cereus, Yersinia enterocolitis, Shigella boydii, Staphylococus aureus, Serratiamarcescens, Pseudomonas vulgaris, Vibrio cholerae, Hafniaalvei, Enterobacter cloacae, Enterobacter aerogenes, E. coli (O103, O55, O121, O30 and O26), Aspergillus niger (NRRL 326) and Fusarium verticilliodes (NRRL 13586), Saccharomyces cerevisiae (ATCC 24859), Lactobacillus casei (ATCC 11443), Erwinia carotovora pv. carotovora and Clavibacter michiganense. Sensitivity of the FTIR detection was 103CFU/ml and a clear differentiation was obtained between the different organisms both at the species as well as at the strain level for the tested pathogens. A very crucial step in the direction of analyzing mixed cultures was taken. The vector based algorithm was able to identify a target pathogen of interest in a mixture of up to three organisms. Efforts will be made to extend this to 10-12 key pathogens. The experience gained was very helpful in laying the foundations for extracting the true fingerprint of a specific pathogen irrespective of the background substrate. This is very crucial especially when experimenting with solid samples as well as complex food matrices. Spectroscopic techniques, especially FTIR and Raman methods are being pursued by agencies such as DARPA and Department of Defense to combat homeland security. Through the BARD US-3296-02 feasibility grant, the foundations for detection, sample handling, and the needed algorithms and models were developed. Successive efforts will be made in transferring the methodology to fruit surfaces and to other complex food matrices which can be accomplished with creative sampling methods and experimentation. Even a marginal success in this direction will result in a very significant breakthrough because FTIR and Raman methods, in spite of their limitations are still one of most rapid and nondestructive methods available. Continued interest and efforts in improving the components as well as the refinement of the procedures is bound to result in a significant breakthrough in sensor technology for food safety and biosecurity.
Rapid detection and differentiation of Salmonella species, Salmonella Typhimurium and Salmonella Enteritidis by multiplex quantitative PCR
