|VIBBERT, HUNTER - University Of Chicago|
|KU, SEOCKMO - University Of Chicago|
|LI, XUAN - University Of Chicago|
|LIU, XINGYA - University Of Chicago|
|KREKE, THOMAS - Purdue University|
|DEERING, AMMANDA - Purdue University|
|XIMENES, EDUARDO - Purdue University|
|LADISCH, MICHAEL - Purdue University|
Submitted to: Biotechnology Progress
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 9/11/2015
Publication Date: 10/7/2015
Citation: Vibbert, H.B., Ku, S., Li, X., Liu, X., Kreke, T., Deering, A.J., Gehring, A.G., Ximenes, E., Ladisch, M.R. 2015. Accelerating sample preparation through enzyme-assisted microfiltration of Salmonella in chicken extract. Biotechnology Progress. 31(6):1551-1562.
Interpretive Summary: Though fast, rapid methods for bacterial detection suffer from relatively poor detection performance. The detection of the harmful bacteria, Salmonella, in chicken extracts may be greatly enhanced when the bacterial cells are separated from any residual sample debris and then concentrated prior to testing. Such higher concentration levels are often achieved with bacterial growth, but that uses up time (at least 16-18 hrs) and the number of initial bacterial cells can no longer be determined. An automated hollow fiber filtration system was developed that continuously flows, separates, and concentrates bacteria from sample debris. However, to prevent premature clogging of the filter membrane by the debris (which is mainly protein), an enzyme that breaks down protein was used prior to filtration. Concentration and recovery of 1 to 10 cells of Salmonella/mL from 400 mL chicken rinse was possible in less than 4 hours, with the filtration step requiring less than 25 min. Producers may use this filtration approach so that rapid methods may be applied in the field in support of on-line food processing.
Technical Abstract: Microfiltration of chicken extracts has the potential to significantly decrease the time required to detect Salmonella, as long as the extract can be efficiently filtered and the pathogenic microorganisms kept in a viable state during this process. We present conditions that enable microfiltration by adding endopeptidase from Bacillus amyloliquefaciens to chicken extracts or chicken rinse, prior to microfiltration with fluid flow on both retentate and permeate sides of 0.2 um cutoff polysulfone and polyethersulfone hollow fiber membranes. After treatment with this protease the distribution of micron, submicron, and nanometer particles in chicken extracts changes so that the size of the remaining particles corresponds to 0.4 to 1 um. Together with alteration of dissolved proteins, this change helps to explain how membrane fouling might be minimized since the potential foulants are significantly smaller or larger than the membrane pore size. At the same time, we found that the presence of protein protects Salmonella from protease action, thus maintaining cell viability. Concentration and recovery of 1 to 10 CFU Salmonella/mL from 400 mL chicken rinse is possible in less than 4 hours, with the microfiltration step requiring less than 25 min at fluxes of 0.028 to 0.32 mL/(cm** • min). The entire procedure - from sample processing to detection by PCR - is completed in 8 hours.