Prediction of fungal growth and ochratoxin A production by Aspergillus ochraceus on irradiated barley grain as influenced by temperature and water activity
Introduction
Aspergillus ochraceus is important in stored grain because of its ability to produce ochratoxin A (OTA) (Van der Merwe et al., 1965), a potent nephrotoxin known also to be teratogenic, immunosuppressive and carcinogenic. It has been classified by the International Agency for Research on Cancer (IARC, 1993) as a possible human carcinogen (group 2B) based on sufficient evidence of carcinogenicity for animals and inadequate evidence in humans (Kuiper-Goodman, 1996).
Barley grain is commonly colonized by toxigenic A. ochraceus together with various species of Penicillium, Fusarium, and yeasts Clarke and Hill, 1981, Hill and Lacey, 1983, Sala, 1993. Some studies have shown the presence of OTA-producing isolates of A. ochraceus in stored barley grains contained by OTA (Cvetnin and Pepeljnjak, 1990).
Individual fungal species differ in their growth responses to the water activity (aw) and temperature of the grain Ayerst, 1969, Mislivec and Tuite, 1970, Magan and Lacey, 1988. Fusarium spp. are generally considered to be field fungi with a high water requirement for growth, although they can sometimes grow in stored grain, while Aspergillus and Penicillium spp. are typical storage species that are able to thrive at relatively low water activities (Ramakrishna et al., 1996).
Exposure of grains to high moisture levels is most likely to occur during harvest, but may occur at other stages between harvest and final consumption. Mycotoxin contamination of grain is difficult to predict because it depends on a complex interaction of factors, such a temperature, moisture, kind of grain, endogenous fungal species, storage history, storage time, type of transit and transit time (Chelack et al., 1991).
According to recommendations by FAO (JECFA, 2002) studies should be conducted to improve the understanding of the occurrence and ecology of the fungi that produce ochratoxin A. Growth of fungi has frequently been studied on nutrient media adjusted to different aw with glycerol and incubated at different temperatures. However, results obtained on culture media cannot necessarily be extrapolated to natural systems, and colonization patterns may be modified in grain ecosystems Magan and Lacey, 1984, Magan and Lacey, 1985. The moisture content and temperature are the most important variables in determining growth and rate of mycotoxin production by fungi (Magan and Lacey, 1988).
The objectives of this study were (a) to determine the effects of water activity, temperature and time on growth and ochratoxin A production of three ochratoxigenic isolates of A. ochraceus on barley grains and (b) to obtain models for prediction of growth and OTA production as a function of aw and temperature which may be useful for predicting safe storage conditions for cereals.
Section snippets
Fungal isolates
Three ochratoxigenic isolates of A. ochraceus Wilhem (=Aspergillus alutaceus Berk & Curt) isolated from cereals were used: isolate NRRL 3174 (coded as 3.94) and isolates 3.113 and 3.38, which are deposited in the Food Technology Department Collection of the University of Lleida, Spain.
Source of barley grain and gamma-irradiation treatment
Spanish barley grain (variety Grafic) harvested in 2001 was used and exposed to 12 kGy gamma irradiation and stored at 4 °C. This dose was sufficient to kill all fungi, bacteria and yeasts on or within the grain
Growth study
Optimal conditions for growth on barley grain of the three isolates of A. ochraceus tested were 30 °C and 0.99 aw (Fig. 1). At 0.80 aw none of the isolates grew. Minimum aw for growth was found at 0.85 at levels of temperature of 30 and 20 °C, and at 0.95 aw at 10 °C (Table 1).
Factors assayed, water activity, temperature and isolates, as well as their two- and three-way interactions significantly affected colony radius, during incubation for 28 days (Table 2).
At 30 °C, the decrease at water
Discussion
This study has shown a significant influence of water activity and temperature on growth and ochratoxin A production on barley grains of three isolates of A. ochraceus.
Minimum water activity level found to be 0.85 for growth of isolates of A. ochraceus and 0.90 aw for OTA production. Previous studies found a minimum water activity values of 0.77 for growth of isolates of A. ochraceus Pitt and Christian, 1968, Kozakievicz and Smith, 1994 and of 0.80–0.87 aw for OTA production Northolt et al.,
Acknowledgements
The authors are grateful to the Catalonian government (Direcció General de Recerca) and to the Spanish government (CICYT, AGL 2001-2974-C05-02) for their financial support.
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