Effects of Various Acids on Growth and Survival of Yersinia enterocolitica

Abstract

The relative effects of several different acids on survival of Yersinia enterocolitica in tryptic soy broth after 24 h of incubation were compared. The acids compared included 25, 50, 75, and 100 mW concentrations of hydrochloric (HC1), citric (CIT), acetic (ACE), lactic (LAC), propionic (PRO), or phosphoric (PHO) acid. The decrease in viable Yersinia cells was compared based on concentration of acid added, pH, and the calculated concentration of the undissociated portion of the monoprotic acids. In addition, data were subjected to analysis of variance procedures to determine significant differences among the anti-microbial activities of the various acids. Results indicated that the relative activity of the acids differed depending on how the data were compared. When based on equal molar concentration of acid added, the overall hierarchy of antimicrobial activity appeared to be CIT & HC1 & LAC & PHO > PRO & ACE. The antimicrobial activity based on pH gave the apparent overall hierarchy PRO 3* LAC & ACE > CIT s PHO 3* HC1. The antimicrobial activity based on the concentration of the un-dissociated portion of monoprotic acids appeared to be HC1 > LAC > PRO 3* ACE. Although graphic representation of the data gave the appearance of differences in antimicrobial effects, statistical analysis revealed that often there was no significant difference (p=s0.05) between acids. Differences which did appear were concentration-dependent. Statistical analysis also revealed that pH alone did not have a significant effect on survival. Yersinia enterocolitica is a bacterium which has only in the last decade become established as a foodborne human pathogen. During this time a number of foodborne disease outbreaks have been attributed to this organism (2,3,17). The most common symptom of a Yersinia infection is gastroenteritis although other symptoms can include mesenteric lymphadenitis (18), terminal ileitis (8), and arthritis (18). In addition, this organism is unique among common foodborne pathogens in being able to mimic appendicitis (3). The ability of foodborne pathogens to survive in acidic environments is important because acidity is often used in foods to control these bacteria. Y. enterocolitica is one pathogen that has shown this ability. In one study (1), viable Y. enterocolitica were isolated from tartar sauce, an acid food. However, Brackett (4) could not recover viable Y. enterocolitica cells from artificially contaminated tartar sauce or spoonable salad dressing at any time, or from mayonnaise after 48 h. Stern and Pierson (15) reported that four separate strains of Y. en-terocolitica grew at pH 4.6. Kendall and Gilbert (7) demonstrated that this organism grew at pH 4.4 and above and survived for at least 72 h at pH 4.2. Brackett (4) obtained similar results, but also found viable Y. en-terocolitica cells after 21 days at pH 4.0 and 5°C. None of the reports mentioned the effects of different acids on Yersinia. The purpose of this study was to determine the relative effects of different acids on growth and survival of Y. enterocolitica. In addition, methods of presenting survival data were compared. MATERIALS AND METHODS Cultures Cultures used were Y. enterocolitica strains Y7P (serotype 0:8), C122-76 (serotype 0:3), and 2635 (serotype 0:8). All three cultures were provided by M. P. Doyle of the Food Research Institute, University of Wisconsin. Before use in experiments, cultures were checked for purity, and the genus and species were confirmed using the API 20E system (Analytab Products, Plainview, NY). Stock cultures were maintained on tryptic soy agar (TSA; Difco) slants incubated for 24 h at 25°C, and held at 5°C until use. Preparation of acid solutions Two molar stock solutions of acetic (ACE), hydrochloric (HCL), lactic (LAC), propionic (PRO), and phosphoric (PHO) acids, and a 1 M stock solution of citric (CIT) acid were prepared and sterilized by autoclaving at 121°C for 15 min as described by Minor and Marth (9). Appropriate amounts of the stock solutions were then aseptically added to sterile tryptic soy broth (TSB; Difco) to give 100 ml each of 0, 25, 50, 75 and 100 mW concentrations of the individual acids in the acidified TSB (TSBA) in 250-mI Erlenmeyer flasks. Duplicate flasks of TSBA solutions were used as growth media for all experiments.