A lack of predatory interaction between rumen ciliate protozoa and Shiga-toxin producing Escherichia coli L.C. Burow 1 , K.S. Gobius 2 , B.A. Vanselow 3 and A.V. Klieve 4 1 Department of Microbiology, The University of Queensland, Brisbane, Qld, Australia, 2 Food Science Australia, Cannon Hill, Qld, Australia, 3 Beef Industry Centre, New South Wales Agriculture, Armidale, NSW, Australia, and 4 Department of Primary Industries and Fisheries, Animal Research Institute, Moorooka, Qld, Australia 2004/0555: received 17 May 2004, revised 6 September 2004 and accepted 27 September 2004 ABSTRACT L.C. BUROW, K.S. GOBIUS, B.A. VANSELOW AND A.V. KLIEVE. 2004. Aims: To investigate interactions between rumen protozoa and Shiga toxin-producing Escherichia coli (STEC) and to ascertain whether it is likely that rumen protozoa act as ruminant hosts for STEC. Methods and Results: The presence of stx genes in different microbial fractions recovered from cattle and sheep rumen contents and faeces was examined using PCR. In animals shedding faecal STEC, stx genes were not detected in the rumen bacterial or rumen protozoal fractions. Direct interactions between ruminal protozoa and STEC were investigated by in vitro co-incubation. Rumen protozoa did not appear to ingest STEC, a STEC lysogen or non-STEC E. coli populations when co-incubated. Conclusions: The ruminal environment is unlikely to be a preferred habitat for STEC. Bacterial grazing by rumen protozoa appears to have little, if any, effect on STEC populations. Significance and Impact of the Study: This study indicates that ruminal protozoa are unlikely to be a major factor in the survival of STEC in ruminants. They appear as neither a host that protects STEC from the ruminal environment nor a predator that might reduce STEC numbers. Keywords: cattle, E. coli, PCR, rumen, sheep. INTRODUCTION Shiga-toxin producing Escherichia coli (STEC), causative organisms in outbreaks of food-borne illness, are widely distributed in domestic ruminants, which are considered an important route for transmission to humans (Hancock et al. 2001). There is a need to understand the maintenance of STEC populations in ruminants and their survival in the rumen following ingestion and prior to colonizing the intestines. This knowledge may contribute to understanding STEC ecology and to possible strategies to limit their presence in ruminants. Free-living protozoa are found in a diverse range of habitats, from water and soils, as well as within the gastrointestinal tract (GIT) of a range of vertebrate hosts. These protozoa can act as hosts for other micro-organisms, especially bacteria (Barker and Brown 1994). It has been shown that increased survival of E. coli O157 occurs in association with the soil protist Acanthamoeba polyphaga (Barker et al. 1999). Ciliate protozoa are common inhabitants of the rumen of domestic livestock that may constitute a potential reservoir for the survival of E. coli O157 and other STEC within the rumen and provide a mechanism for survival and transit through the rumen to the lower digestive tract in ruminants. Rumen protozoa are known to engulf a range of bacteria, kill and digest some of them and excrete their digestion products (Coleman 1989). However, other bacteria maintain commensal relationships with ruminal protozoans (Lloyd et al. 1996). To date, little is known about the predatory interactions between ruminal protozoa and bacterial pathogens (Williams and Coleman 1992) and a symbiotic association of STEC with rumen protozoa has not been reported. Correspondence to: Athol Klieve, Animal Research Institute, Locked Mail Bag No 4, Moorooka, Qld 4105, Australia (e-mail: athol.klieve@dpi.qld.gov.au). ª 2004 The Society for Applied Microbiology Letters in Applied Microbiology 2005, 40, 117–122 doi:10.1111/j.1472-765X.2004.01642.x