Identification of a novel enterotoxigenic activity associated with Bacillus cereus.

Abstract

SYNOPSIS A strain of Bacillus cereus isolated from a food poisoning outbreak characterized by vomiting has been shown to be capable of causing vomiting when cultures grown on rice, but not other media, were fed to Rhesus monkeys. In contrast, a strain isolated from a diarrhoeal outbreak produced diarrhoea, but not vomiting, when grown on various media in similar feeding trials. Furthermore, culture filtrates from the diarrhoeal strain caused fluid accumulation in ligated rabbit ileal loops whereas those from the vomiting strain did not. It is proposed that at least two enterotoxins are involved, one responsible for the vomiting and one for the diarrhoeal symptoms.Bacillus cereus has been implicated in food poisoning outbreaks since early this century (Lubenau, 1906) but it was not until after 1950 that the by now classic description of B. cereus gastroenteritis was made (Hauge, 1950;1955; Goepfert etal, 1972). The illness was characterized by symptoms of diarrhoea and abdominal pain between 8 and 16 hours after ingestion of food; vomiting was rare. A wide variety of foods has been involved including meat and vegetable soups, cooked meat and poultry, cooked vegetables, puddings, and sauces. An enterotoxin from B. cereus strains isolated from such incidents has been partially characterized (Spira and Goepfert, 1975).Since 1971, however, numerous incidents of food poisoning associated with consumption of cooked rice from Chinese restaurants and 'take-away' shops have been reported in this country as well as in Australia, Canada, and the Netherlands . Although these outbreaks were also attributed to B. cereus (Mortimer and McCann, 1974;Gilbert and Taylor, 1975), the characteristic features were acute nausea and vomiting, usually occurring between 1 and 5 hours after a meal; diarrhoea was not a common feature.Monkey feeding trials have therefore been carried out in an attempt to confirm that these recent riceassociated outbreaks were in fact caused by B. cereus and to determine whether or not a new enterotoxigenic material was involved.Received for publication 12 April 1976 Material and methods ORGANISMSThree strains of B. cereus were used. Strain 4810/73 (formerly strain 88) was originally isolated from vomitus and had been implicated in a typical riceassociated illness. Strain 4433/73 had been isolated from meat loaf and was implicated in a typical diarrhoeal food poisoning outbreak. The serology of these strains has been described . Strain 2532B/74 was isolated from raw rice and had not been implicated in any food poisoning outbreak and was non-typable according to the scheme of Taylor and Gilbert (1975). SEROTYPINGSerotyping was carried out by the method of Taylor and Gilbert (1975). CULTURE METHODSFor rice cultures 100 g of ordinary cooking rice(short grain) was soaked in 400 ml of 0 95 % saline in 2-litre flasks and autoclaved at 115°C for 15 minutes. The rice was inoculated with 30 ml of an 8-hour culture of B. cereus in tryptone soya broth and incubated at 30°C for [18][19][20] hours. The rice culture w...

Studies on the production of enterotoxins by Bacillus cereus.

“…The number of loops tied off in a single animal was usually five and never more than six after preliminary tests. Into each loop were injected 2 ml aliquots of cell-free filtrate, filtered ultrasonicated culture, concentrated cell-free filtrate or whole cells grown in rice which was subsequently homogenized by treatment with diastase for easy administration (Melling et al, 1976). Filtrates were concentrated by dialysis at 40C Studies on the production of enterotoxins by Bacillus cereus against 70% Carbowax (Union Carbide) in water after the addition of 100 000 units of penicillin and 10 mg of streptomycin to approximately 80 ml of culture filtrate.…”

“…Evidence for the existence of a vomiting toxin produced by B. cereus has resulted from preliminary monkey feeding tests (Melling et al, 1976). This paper describes studies undertaken (1) to establish whether B. cereus strains may be classified as diarrhoeal, vomiting or non-enterotoxigenic, and (2) in search of a reliable method for the detection and quantitation of the enterotoxin(s).…”

Evaluation of the presence of cereulide in food products collected from the Belgian market

“…Nevertheless, besides production of cereulide and inducing the emetic syndrome some other strains of B. cereus may be responsible for diarrhoeal syndrome (Agata et al 1995;Melling et al 1976). It is generally assumed that food containing doses <10 5 -10 6 CFU/g of B. cereus and <8 μg cereulide/kg bodyweight is safe for consumption and will not cause acute intoxication (Rajkovic 2014).…”

A review of analytical methods for the detection of bacterial toxins

“…Early biological assay methods for screening for B. cereus emetic toxin involved feeding to rhesus monkeys and were therefore laborious and very expensive (Melling et al 1976). These tests were by in-vitro tissue culture assays with Hep-2 (Hughes et al 1988) or CHO cells (Buchanan and Shultz 1992).…”