Survival and growth of Enterobacter sakazakii in powdered and reconstituted infant formulas, performance of media for recovering stressed cells, and sensitivity of the pathogen to the lactoperoxidase system
Gurtler, Joshua B.
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Studies were done to examine the survival of Enterobacter sakazakii in powdered and reconstituted infant formulas and the performance of media for recovering stressed cells. Sensitivity of the pathogen to the lactoperoxidase system (LPOS) was determined. The ability of tryptic soy agar supplemented with 0.1% pyruvate (TSAP) (control medium), two new fluorogenic agars developed by Leuschner, Baird, Donald, and Cox (LBDC) and Oh and Kang (OK), fecal coliform agar (FCA), Druggan-Forsythe-Iversen medium (DFI), VRBG agar, and Enterobacteriaceae enrichment agar (EE), to support colony development by healthy and heat-, freeze-, acid-, alkaline-, and desiccation-stressed cells of Enterobacter sakazakii was examined. The general order of performance of media for recovering control and heat-, freeze-, acid-, and alkaline-stressed cells by spiral plating was TSAP > LBDC > FCA > OK > VRBG > DFI > EE, which is similar to results obtained from desiccated cells. Results indicate that differential, selective media vary greatly in their ability to support resuscitation and colony formation by stressed cells of E. sakazakii. The ability of E. sakazakii to survive in six commercially manufactured powdered infant formulas formulas (aw 0.25 - 0.86) for up to 12 months at 4, 21, and 30°C was determined. Populations decreased significantly in all formulas at aw 0.25 - 0.50 during storage for 1 month at 21 or 30°C, and again between 1 and 6 months in most formulas; significant reductions occurred between 6 and 12 months in some formulas. At all storage temperatures, reductions in populations tended to be greater in formulas at aw 0.43 - 0.86 than in formulas at aw 0.25 - 0.30. Survival of the pathogen was generally unaffected by composition of powdered infant formula. E. sakazakii, initially at a population of 0.02 CFU/ml, grew to populations of • 1 log CFU/ml in reconstituted infant formulas held at 12, 21, and 30°C for 48, 12, and 8 h, respectively. Initially at a population of 0.53 CFU/ml, the pathogen grew to populations of • 1 log CFU/ml in reconstituted infant formulas held at 12 and 21°C for 24 and 8 h, respectively, and to populations of > 3 log CFU/ml when held at 30°C for 8 h. Growth of E. sakazakii was not greatly influenced by the composition of formula. The lactoperoxidase system (LPOS), when applied to reconstituted infant formula using lactoperoxidase (LPO) at concentrations of 10 - 30 µg/ml, inhibited growth of E. sakazakii initially at 0.03 CFU/ml (” 25 CFU/100 g of dry powder) to less than 1 CFU/ml for up to 24 h at 21 - 37°C. Treatment of reconstituted infant formula with LPO at concentrations of 10 - 30 µg/ml prevented the growth of E. sakazakii in formulas held at 21, 30, and 37°C. This research provides information on the recovery of healthy and stressed E. sakazakii cells on selective/differential media, survival and growth of the pathogen in powdered or reconstituted infant formula, and the use of LPOS in preventing the growth of E. sakazakii in reconstituted infant formula.