Use of pGFPuv Mutants to Study the Influence of Drying on the Survival of Cronobacter sakazakii in Corn

Abstract

Cronobacter sakazakii is the causative agent of meningitis and necrotizing enterocolitis in certain groups of infants. Transformation of local isolates of C. sakazakii with a Green Fluorescent Protein (GFP) plasmid has produced C.sakazakii pGFPuv mutants potential to be used for studying C. sakazakii behavior without the need to suppress other microorganisms or use of diagnostic media. This research aimsto study the effect of drying on water content and water activity of corn and to evaluate the survival of C. sakazakii pGFPuv as well as other naturally occurring microorganisms in the corn during drying. Corn with moisture content of 40% (d.b) were inoculated with C. sakazakii pGFPuv mutant to achieve an initial concentration of CFU/g and drying was performed at 42 ºC, 46 ºC and 50 ºC for ten days in a drying chamber. Corn samples were taken every day and water content were analysed using oven method while water activity by Aw meter.The number of C.sakazakii and total microorganisms surviving in the corn were enumerated and the survival rate of C. sakazakii was determined. The results showed that the decrease in water content occurred at a constant rate at the first day of drying followed by a falling rate decrease that correspond to the decline in Aw. Drying at all temperatures for 10 days resulted in the decrease of naturally occurring microorganisms to an undetectable level. However, at the end of the drying, C.sakazakii was still present at a concentration of 2 Log CFU/g. Scanning electron microscopy showed that C. sakazakii form colonies on the surfaces of corn and in the cavity of the tip cap. This study showed that C.sakazakiipGFPuv could be used to study the survival of C.sakazakii during corn drying and C. sakazakii displayed resistance to drying at 42, 46 and 50^oC for 10 days.