DETECTION OF LISTERIA MONOCYTOGENES BY ADSORPTION AND ELUTION FROM HYDROPHOBIC MATRICES

Abstract

ABSTRACTThe binding of L. monocytogenes Scott A strain to three hydrophobic matrices, octyl, phenyl and butyl Sepharose, was investigated. Optimal adsorption of L. monocytogenes to octyl Sepharose was obtained at pH 3.5 and 4 M NaCl. However, it was difficult to elute the bacteria from octyl Sepharose, even after changing the pH and lowering the salt concentration. Good adsorption of L. monocytogenes to phenyl Sepharose at pH 3.5 and 4 M NaCl was also observed. L. monocytogenes was found to adsorb weakly to butyl Sepharose, which is less hydrophobic than phenyl Sepharose. Bacteria were eluted under various conditions. The best elution was obtained with 10 mM sodium phosphate, followed by an increasing gradient of ethylene glycol. To test the potential application of hydrophobic chromatography for separating L. monocytogenes from food matrices, milk was inoculated with L. monocytogenes and then passed through a column of phenyl Sepharose at pH 3.5 and 4 M NaCl. Nearly all L. monocytogenes were bound to the hydrophobic gel and were eluted in a pure and viable form by changing the pH and lowering the salt concentration, and by using a polar reducing agent, ethylene glycol. This study shows that hydrophobic interaction chromatography can be used to separate L. monocytogenes from milk and may be applicable to other food suspensions. It is a gentle method that makes use of the hydrophobic surface properties of Listeria for attachment to hydrophobic gels, as well as using mild elution conditions to avoid inactivation of the organism.