A bulletin for the Australian Food Industry March 1997
Contents: How safe are fruit juices and acid foods? | Genetically modified soya | Foreign objects in foods
How safe are fruit juices and acid foods?
Acid tolerance of enterohaemorrhagic E. coli (EHEC) and Salmonella spp. in fruit juices and other acid foods
In the previous issue of Food Safety and Hygiene (December 1996) mention was made of a 1996 food poisoning incident in the USA and Canada caused by E. coli O157:H7 in which unpasteurised apple juice was implicated as the vehicle of infection. Unpasteurised apple juice, usually referred to as apple cider in North America, was the source of E. coli O157:H7 in a food poisoning incident in Massachusetts in 1991 (Besser et al. 1993). An earlier outbreak in Canada involving 14 cases of haemolytic uraemic syndrome in children was also associated with unpasteurised apple juice although the infectious agent was not identified (Steele, Murphy and Rance 1982). Apple cider was identified as the vehicle for a food poisoning outbreak in New Jersey in 1974 caused by Salmonella typhimurium. Samples of the juice recovered during this outbreak had pH values ranging from 3.4 - 3.9 (Goverd et al. 1979).
Effect of pH, temperature and acid type
This 1974 incident prompted Goverd and co-workers to study the influence of temperature and pH (and alcohol content) on the survival of S. typhimurium and other Salmonella serotypes in sterile apple juice. In an important paper, Goverd et al., 1979, showed that several Salmonella serotypes could survive in apple juice at pH 3.6 for 30 days at 4°C. The authors concluded that survival of the organisms was greater at 22°C than at 4°C when an initial population of 2x105 organisms/ml was used. They recommended that only top grade, well-washed fruit should be used for juice production and the juice should be pasteurised before sale.
There are a number of earlier reports of the survival of pathogens such as Salmonella spp in fruit juices and other acid foods. The most comprehensive of these is that by Mossel and De Bruin (1960). These authors noted that work published prior to their own investigation permitted two general conclusions. The survival of pathogenic Enterobacteriaceae at reduced pH values is strongly dependent not only on the pH value but also on:
a) the type of organic acid (and other compounds) present, and
b) the temperature of storage.
In an experimental investigation, Mossel and De Bruin studied the survival of a range of Enterobacteriaceae including E. coli and S. typhimurium in tomato, apple, orange and lemon juices as well as mayonnaise and yoghurt.
They concluded that with an initial population of 106 organisms/ml, pathogenic Enterobacteriaceae do not survive more than one day in lemon juice (pH 2.4 - 2.6). In the other fruit juices examined, survival occurred for up to 35 days at 5°C and up to 10 days at 24°C. The pH of the products studied is shown below:
| pH | Product | Predominant acid |
|---|---|---|
| 2.4 - 2.6 | lemon juice | citric |
| 3.0 - 3.1 | apple juice | malic |
| 3.1 - 3.5 | orange juice | citric and malic |
| 3.8 - 4.1 |
tomato juice mayonnaise yoghurt |
malic and citric acetic lactic |
On the basis of this study, Mossel and De Bruin concluded that fruit juices with a pH greater than 3.0 must be considered as potential sources of pathogenic bacteria.
Their findings on survival at refrigeration temperature versus ambient temperature are not in agreement with those of Goverd et al. but because of the long survival periods of potential pathogens at 4°C and 5°C reported in each study, this conflict is of marginal importance.
Mossel and De Bruin also concluded that at a given pH value, in this case pH 3.8 - 4.1, the order of lethal effectiveness of the organic acids in the study appeared to be: lactic > acetic > malic or citric. This conclusion is debatable given the conditions of the study but, as the authors noted, it confirmed the findings of Shillinglaw and Levine (1943) working under more defined laboratory conditions. Certainly the results obtained in these two studies with Gram negative bacteria are contrary to comprehensive studies with organic acids on inhibition of yeasts by Pitt (1974) and earlier workers including Fabian and Wadsworth (1939) and Levine and Fellers (1940) which indicated that acetic acid was more inhibitory than other food acids at the same pH level.
Studies with E. coli O157:H7 and apple juice
The association of E. coli O157:H7 infection with unpasteurised apple juice and mayonnaise has prompted a number of recent studies on the acid tolerance of this organism. Zhao, Doyle and Besser (1993) found that E. coli O157:H7 when initially present at a level of 105 organisms/ml survived up to 31 days in apple cider with a pH of 3.7 at 8°C. These authors concluded that since the expected refrigerated shelf life of apple cider is typically less than 3 weeks, the pathogen is likely to survive during most of the time that the cider would be consumed at numbers close to the size of the initial population introduced into the product. In similar studies at 25°C, survivors were detected at 2 to 3 days but not at 6 days after inoculation.
These authors also studied the effect on E. coli O157:H7 of 0.1% potassium sorbate, 0.1% sodium benzoate and a combination of 0.1% potassium sorbate and 0.1% sodium benzoate in apple juice. They found that at 8°C, the sodium benzoate and the combination benzoate/sorbate treatment reduced numbers of E. coli O157:H7 but the results obtained do not seem to justify the authors' conclusion that use of 0.1% sodium benzoate is an option processors have, to increase substantially the safety of apple cider.
Miller and Kaspar (1994) conducted further work in this area which confirmed the acid tolerance of two strains of E. coli O157:H7 in apple ciders with pH values of 3.7 - 4.1. Both strains examined were significantly more acid tolerant than a control strain of E. coli although the degree of tolerance differed between strains. Again low storage temperature, 4°C, enhanced survival at low pH levels. The strains investigated were able to survive 14 to 21 days at 4°C when the initial population level was 3.0 x 104 organisms/ml of cider. In studies similar to those of Zhao, Doyle and Besser (1993), these authors were unable to confirm that 0.1% sodium benzoate in cider was an effective preservative against the strains of E. coli O157:H7 tested. Both strains investigated survived for 21 days in the presence of the sodium benzoate at 4°C. Miller and Kaspar also conclude that the characteristics which are responsible for acid tolerance in E. coli O157:H7 must be present prior to introduction of the organism into the acid environment. This is in contrast to the situation with S. typhimurium where adaptation to the acid environment is reported to occur (Foster and Hall 1990, Leyer and Johnson 1993).
Studies with E. coli O157:H7 and mayonnaise
A series of outbreaks of food poisoning in the United States in 1993 were traced to a restaurant chain specialising in steaks and self-serve salad bar food. An estimated 300 people were affected and E. coli O157:H7 was found to be the causative organism (Weagent, Bryant and Bark 1994). Concern that low pH foods like mayonnaise and sauces could act as vehicles for infection prompted these authors to study the survival of E. coli O157:H7 in these products held at room and refrigerator temperatures. Temperatures chosen for this study were based on the temperature normally used for storage of retail mayonnaise (25°C) and the temperatures at which the mayonnaise and sauces implicated in the food poisoning incidents had been stored, 5°C and 7°C respectively. The mayonnaise pH was 3.65 and the pH of selected sauce formulations lay in the range 3.68 - 4.44. Population levels were greater than 107 organisms/ml and three different strains of E. coli O157:H7 were used.
E. coli O157:H7 became undetectable after three days storage in mayonnaise at 25°C but the same strains inoculated into mayonnaise and stored at 7°C were detectable up to 35 days. E. coli O157:H7 inoculated into the mayonnaise-based sauces and held at 5°C were detectable past 35 days in three of the four sauces. No organisms could be detected after three days in a mayonnaise and mustard sauce with a pH of 3.68.
The authors conclude that mayonnaise and possibly other acidic foods could serve as vehicles for E. coli O157:H7 infection when stored at refrigeration temperatures.
Conclusion
Many food associated outbreaks of haemorrhagic colitis and haemolytic uraemic syndrome caused by EHEC have now been reported. Undercooked, ground beef has been the principal vehicle of infection but a number of other foods have been implicated. Moderately acid fermented food including salami type sausage (Centers for Disease Control, 1995, Rowe 1995) and yoghurt (Morgan et al. 1993) are amongst those identified as sources of infection.
It is now clear from both epidemiological and laboratory investigations that EHEC can survive in fruit juices and some other acid foods with a pH in the range 3.5 - 4.0 for many days at refrigeration temperatures. A similar situation applies with Salmonella spp. although actual salmonellosis incidents associated with acid foods appear to be extremely rare.
These findings are of considerable importance given changing food markets and the growing emphasis on fresh, i.e. unprocessed foods.
In Australia until recently, most fruit juices would have undergone a pasteurisation procedure either at or immediately after the filling operation or at some earlier processing stage, e.g. prior to concentration. The growth of the fresh juice market has seen this change so that in the case of citrus juices in particular, a major share of the market is now occupied by juices stored under refrigeration that have received little or no heat treatment. Fresh juices in Australia are defined in the Australian Food Standards Code as those which:
- do not contain preservatives
- have not been subjected to a concentration process and
- have a durable life of not more than 21 days.
Thus pasteurisation of juices to be labelled as 'fresh' is permitted but is not a procedure exercised by most processors who are selling their product on the qualities associated with a product which has not been subject to any heating.
The only recent reports of food poisoning caused by fruit juices have been those associated with unpasteurised apple juice in North America. The harvesting and collection of different fruits for processing depends on the type of fruit involved. In the case of the apple juice incidents, it is generally believed that infection of 'drop' apples occurred in the field and this fruit was subsequently collected for processing. This is obviously a risk specifically associated with apples when collected in this manner and may have no relevance to the Australian situation.
However cross contamination of fruit juice with small numbers of pathogenic bacteria could conceivably have occurred at any point in production. It has been reported (Anon 1996) that the Food and Drug Administration in the USA is considering a requirement that apple juice and possibly other juices must be pasteurised to prevent further food poisoning incidents of the type described above.
Proper handling of food is, however, the preventative measure of choice (Doyle 1991). A correctly constructed HACCP plan for fruit juices and other fruit products should take into account the possibility that under certain conditions these products may act as vehicles for some food poisoning bacteria. In fact, prepared fruit salads, which often contain high pH fruits such as watermelon and rockmelon, may be seen to present a greater potential hazard than fresh juices. This is so because not only may the pH of the product be less inhibitory to pathogens but the opportunity for cross contamination during preparation and storage is far greater (Del Rosario and Beuchat 1995).
References
Anon 1996. The Food Institute Report, November 18, p9.
Besser, R.E., Lett, S.M., Webber, T.Jr., Doyle, M.P., Barrett, T.J., Wells, J.G. and Griffin, P.M. 1993. An outbreak of diarrhea and hemolytic uremic syndrome from Escherichia coli O157:H7 in fresh-pressed apple cider. Journal of the American Medical Association 169,2217.
Centers for Disease Control. 1995. Community outbreak of Haemolytic Uraemic Syndrome attributable to Escherichia coli O111:NM - South Australia. 1995. MMWR 44,550.
Del Rosario, B.A. and Beuchat, L.R. 1995. Survival and growth of enterohemorrhagic Escherichia coli O157:H7 in cantaloupe and watermelon. Journal of Food Protection 58,105.
Doyle, M.P. 1991. Escherichia coli O157:H7 and its significance in foods. International Journal of Microbiology 12,289.
Fabian, F.W. and Wadsworth, C.K. 1939. Experimental work on lactic acid in preserving pickles and pickle products II. Preserving value of acetic and lactic acids in presence of sucrose. Food Research 4,511.
Foster, J.W. and Hall, H.K. 1990. Adaptive acidification tolerance response of Salmonella typhimurium. Journal of Bacteriology 172,771
Goverd, K.A., Beech, F.W., Hobbs, R.P., and Shannon, R. 1979. The occurrence and survival of coliforms and salmonellas in apple juice and cider. Journal of Applied Bacteriology 46,521.
Leyer, G.J. and Johnson, E.A. 1993. Acid adaptation induces cross-protection against environmental stresses in Salmonella typhimurium. Applied and Environmental Microbiology 59,1842.
Levine, A.S. and Fellers, C.R. 1940. Action of acetic acid on food spoilage microorganisms. Journal of Bacteriology 39,499. Miller, L.G. and Kaspar, C.W. 1994. Escherichia coli O157:H7 acid tolerance and survival in apple cider. Journal of Food Protection 57,460.
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Rowe, P.M. 1995. Eradicating E. coli O157:H7. The Lancet 345,117.
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Steele, B.T., Murphy, N. and Rance, C.P. 1982. An outbreak of hemolytic uremic syndrome associated with ingestion of fresh apple juice. The Journal of Pediatrics 101,963.
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Zhao, T., Doyle, M.P. and Besser, R.E. 1993. Fate of enterohemorrhagic Escherichia coli O157:H7 in apple cider with and without preservatives. Applied and Environmental Microbiology 59,2526.
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