Lee Young Jun Korea University Lab of Food Microbiology 2007. 5. 16 Influence of acid tolerance response on survival, growth, and thermal cross-protection of Escherichia coli O157:H7 in acidified media and fruit juices Lee Young Jun Korea University Lab of Food Microbiology 2007. 5. 16

Table of Contents Introduction Objective Experimental design Phase Ⅰ Survival of control, acid-adapted, acid-shocked cells of E. coli O157:H7 inoculated into TSB acidified with acetic or lactic acid Phase Ⅱ Growth pattern of E. coli O157:H7 in apple cider and orange juice incubated at 5 and 25℃ Phase Ⅲ D-values of three type of E. coli O157:H7 cells in TSB, apple cider, and orange juice at 52 and 54℃ Conclusions

E. coli O157:H7 Characteristics Diseases Reservoirs Gram negative Motile rods Facultative aerobes Diseases Diarrhea, Hemorrhagic colitis, Hemolytic uremic syndrom Reservoirs Cattle, Domestic animals, Humans

Acid-adapted? Acid-shocked? Acid-adapted cells are those which have been exposed to a gradual decrease in environmental pH Acid-shocked? Acid-shocked cells are those which have been exposed to an abrupt shift from high pH to low pH

D-value Time required to reduce population of microorganism to 1/10 at certain temperature

Objective To determine if survival and growth patterns of acid-adapted and acid-shocked cells of E. coli O157:H7 differ in acidified TSB and in fruit juices To determine if acid adaptation of acid shock alter heat tolerance of cells in juices

Experimental design Phase Ⅰ Survival of control, acid-adapted, acid-shocked cells of E. coli O157:H7 inoculated into TSB acidified with acetic or lactic acid Phase Ⅱ Growth pattern of E. coli O157:H7 in apple cider and orange juice incubated at 5 and 25℃ Phase Ⅲ D-values of three type of E. coli O157:H7 cells in TSB, apple cider, and orange juice at 52 and 54℃

Phase Ⅰ Survival of control, acid-adapted, acid-shocked cells of E. coli O157:H7 inoculated into TSB acidified with acetic or lactic acid

Materials and Methods Strains Cell types Media pH & Incubation time(h) - E. coli O157:H7 Cell types - control, acid-adapted, acid-shocked cells Media - Tryptic soy broth(TSB) - Tryptic soy agar(TSA) - TSA containing 3% sodium chloride(TSAS) pH & Incubation time(h) - pH3.4 : 0, 1, 2(acetic) 0, 1, 3, 6, 8(lactic) - pH3.9 : 0, 5, 9, 12, 18, 24

Control cells Acid-adapted cells Acid-shocked cells - grown in TSB for 18h at 37℃ Acid-adapted cells - grown in TSB supplement with 1% glucose for 18h at 37℃ Acid-shocked cells - grown in TSB culture for 16h (pH6.0) - adjusted to pH4.8 by adding 2M lactic acid - incubating for 2h at 37℃

(adjusted to pH 3.9 or 3.4 with acetic or lactic acid) E. coli O157:H7 TSB (adjusted to pH 3.9 or 3.4 with acetic or lactic acid) TSA and TSAS Incubated at 37℃ for 24~48 h

Table 1. Number of control, acid-adapted, and acid-shocked cells of E Table 1. Number of control, acid-adapted, and acid-shocked cells of E. coli O157:H7 inoculated into TSB acidified at pH 3.9 or 3.4 with acetic or lactic acids, incubated for up to 24h, and recovered on TSA or TSAS

Fig. 1. Differences(log10cfu/ml) in populations of control cells(■), acid-adapted cells(●), and acid-shocked cells(▲) of E. coli O157:H7 recovered from TSB acidified at pH 3.9 with lactic acid incubated at 37℃ for up to 12h when plated on TSA and TSAS

Phase Ⅱ Growth pattern of E. coli O157:H7 in apple cider and orange juice incubated at 5 and 25℃

Materials and Methods Strains Cell types Juice types pH Temparature - E. coli O157:H7 Cell types - control, acid-adapted, acid-shocked cells Juice types - Apple cider (AC1, AC2, AC3) - Orange juice (OJ1, OJ2, OJ3) pH - AC1:3.56, AC2:3.98, AC3:3.73 - OJ1:3.82, OJ2:3.86, OJ3:3.86 Temparature - 5 and 25℃

Apple cider and orange juice E. coli O157:H7 Apple cider and orange juice TSA Incubated at 37℃ for 24~48 h

Table 2. Populations(log10cfu/ml) of E Table 2. Populations(log10cfu/ml) of E. coli O157:H7 in apple cider(AC1, AC2, AC3) and orange juice(OJ1, OJ2, OJ3) incubated at 5 and 25℃

Phase Ⅲ D-values of three type of E. coli O157:H7 cells in TSB, apple cider, and orange juice at 52 and 54℃

Materials and Methods Strains Cell types Juice types - E. coli O157:H7 Cell types - control, acid-adapted, acid-shocked cells Juice types - Apple cider (AC1, AC2) - Orange juice (OJ1, OJ2) Temparature (D-value) - 52 and 54℃

Apple cider and orange juice E. coli O157:H7 Apple cider and orange juice (adjusted to 52 and 54℃) Heating (10, 20, 30 min) TSA Incubated at 37℃ for 48 h Counting Colony

Table 3. D-values of control, acid-adapted, and acid-shocked cells of E. coli O157:H7 cells in TSB(pH7.2), apple cider(AC1, AC2), and orange juice(OJ1, OJ2) at 52 and 54℃

Conclusions Acid-adapted E. coli O157:H7 cells have enhanced tolerance to lactic acid but not acetic acid Regardless of test parameter, there is not a strong indication that control, acid-adapted, and acid-shocked cells differ in tolerance to the acidic environment in apple cider or orange juice D-values of acid-adapted cells were considerably high than those of acid-shocked or control cells