Clinical tips: Food poisoning

Food poisoning is used to describe a range of illnesses caused by drinking or eating contaminated drink or food. Infectious pathogens include bacteria, viruses, parasites, or their toxins, though food poisoning can also be a result of eating poisonous plants e.g. some mushrooms, or animals e.g. puffer fish. Common symptoms include nausea, vomiting, watery diarrhoea, abdominal pain and cramps, and fevers, though these will vary depending on the causative pathogen or toxin. Symptoms can start within hours of eating contaminated food, or may begin days or weeks later. Most food poisoning is mild in nature, lasts for several hours to a few days, and generally resolves without treatment. However, some cases of food poisoning can also be extremely severe, with people requiring medical attention or admission to hospital...

Allelic variation at a single gene increases food value in a drought-tolerant staple cereal

The production of adequate agricultural outputs to support the growing human population places great demands on agriculture, especially in light of ever-greater restrictions on input resources. Sorghum is a drought-adapted cereal capable of reliable production where other cereals fail, and thus represents a good candidate to address food security as agricultural inputs of water and arable land grow scarce. A long-standing issue with sorghum grain is that it has an inherently lower digestibility. Here we show that a low-frequency allele type in the starch metabolic gene, pullulanase, is associated with increased digestibility, regardless of genotypic background. We also provide evidence that the beneficial allele type is not associated with deleterious pleiotropic effects in the modern field environment. We argue that increasing the digestibility of an adapted crop is a viable way forward towards addressing food security while maximizing water and land-use efficiency.

Developments and Microbiological applications in African foods: Emphasis on Nigerian Wara cheese

African indigenous foods have received limited research. Most of these indigenous foods are fermented and they form part of the rich nutritional culture of many groups in African countries. The industrialization and commercialisation of these indigenous African fermented foods should be preceded by a thorough scientific knowledge of their processing which can be vital in the elimination of hunger and poverty. This study highlighted emerging developments and the microbiology of cereal-based and cassava-based food products that constitute a major part of the human diet in most African countries. In addition, investigations were also carried out on the coagulant of the Calotropis procera plant used in traditional production of Nigerian Wara cheese and on the effects of adding a nisin producing Lactococcus lactis strain originating from human milk to Nigerian Wara cheese. Fermented cereal-based food such as ogi utilize popular African and readily available grains maize, millet or sorghum as substrates and is popular as a weaning diet in infants. In this study, the bulkiness caused by starch gelatinization was solved by amylase treatments in the investigation on cooked and fermented oat bran porridge. A similar treatment could reduce the viscosity of any cereal porridge. The properties of the Sodom apple leaves (Calotropis procera) extract in cheesemaking were studied. C. procera was affected by monovalent (K+ and Na+) and divalent (Mg2+ and Ca2+) cations during coagulation. The rennet strength of this coagulant was found to be 7 % compared to animal rennet at 35 °C. Increasing the incubation temperature to 70 °C increased the rennet strength 28-fold. The molecular weight of the partially purified protease was determined by SDS-PAGE and was confirmed by Zymography to be approximately 60 kilodaltons. The high proteolytic activity at 70 °C supported the suitability of the protease enzyme as a coagulant in future commercial production of Nigerian Wara cheese. It was also possible to extend the shelf life of Wara cheese by a nisin producing lactic acid bacteria Lactococcus lactis LAC309. The levels of nisin in both whey and curd fractions of Wara were investigated, results showed a 3 log reduction of toxicogenic Bacillus licheniformis spiked on Wara after 3 days. These studies are the first in Finland to promote the advancement of scientific knowledge in African foods. Recognizing these indigenous food products and an efficient transfer of technology from the developed countries to industrialize them are necessary towards a successful realization of the United Nations Millenium Development Program.

Prolamin degradation in sourdoughs

This thesis examines protein behaviours that occur during cereal fermentations. The focus is on the prolamin degradation in sourdoughs. The thesis also looks at what happens to the oat globulins during an oat bran acidification process. The cereal prolamins are unique proteins in many respects. The wheat prolamins (glutenins and gliadins) are responsible for the formation of the gluten that provides the viscoelastic properties to wheat doughs whereas the rye prolamins (secalins) are unable to develop gluten-like structures. In addition, many baking technological features, such as flavour, shelf-life and dough properties are affected by the protein degradation that might occur during processing. On the other hand, the prolamins contain protein structures that are harmful to gluten sensitive people. It is thus evident that the degradation of the prolamins in sourdough processes may be approached from various aspects. This thesis describes some of these approaches. Four different cereal fermentations were carried out. Wheat sourdough (WSD) and rye sourdough (RSD) fermentations represented traditional sourdoughs. A germinated-wheat sourdough (GWSD) was a novel sourdough type that was prepared using germinated wheat grains that had high and diverse proteolytic activities. The oat bran fermentation (OBF) represented a fermentation system that lacked functional cereal proteases. The high molecular weight glutenins and rye secalins were degraded during the WSD and RSD fermentations, respectively. It was noteworthy that in WSD only a very limited degradation of the gliadins occurred. The gliadins were, however, hydrolysed very extensively during the GWSD fermentation. No protein degradation was observable in the OBF system. Instead the acidification altered the solubility of the oat globulins and this finally led to their aggregation. This thesis confirms that the endogenous proteases of cereals hydrolyse cereal prolamins in sourdoughs. The thesis also shows that the proteolytic activity of the used cereal raw material determines the extent of proteolysis that occurs in sourdough. This means that bakers may adjust the protein degradation in their sourdoughs by selecting the raw material based on its proteolytic activity. The thesis also demonstrates that by using germinated grains, with high and diverse proteolytic activity in sourdough preparations, the prolamins can be extensively degraded. Whether such highly proteolytic food technology could be used to manufacture new gluten-free cereal-based products for gluten sensitive people remains to be solved.

Bacillus cereus spores and cereulide in food-borne illness

B. cereus is a gram-positive bacterium that possesses two different forms of life:the large, rod-shaped cells (ca. 0.002 mm by 0.004 mm) that are able to propagate and the small (0.001 mm), oval shaped spores. The spores can survive in almost any environment for up to centuries without nourishment or water. They are insensitive towards most agents that normally kill bacteria: heating up to several hours at 90 ºC, radiation, disinfectants and extreme alkaline (≥ pH 13) and acid (≤ pH 1) environment. The spores are highly hydrophobic and therefore make them tend to stick to all kinds of surfaces, steel, plastics and live cells. In favorable conditions the spores of B. cereus may germinate into vegetative cells capable of producing food poisoning toxins. The toxins can be heat-labile protein formed after ingestion of the contaminated food, inside the gastrointestinal tract (diarrhoeal toxins), or heat stable peptides formed in the food (emesis causing toxin, cereulide). Cereulide cannot be inactivated in foods by cooking or any other procedure applicable on food. Cereulide in consumed food causes serious illness in human, even fatalities. In this thesis, B. cereus strains originating from different kinds of foods and environments and 8 different countries were inspected for their capability of forming cereulide. Of the 1041 isolates from soil, animal feed, water, air, used bedding, grass, dung and equipment only 1.2 % were capable of producing cereulide, whereas of the 144 isolates originating from foods 24 % were cereulide producers. Cereulide was detected by two methods: by its toxicity towards mammalian cells (sperm assay) and by its peculiar chemical structure using liquid-chromatograph-mass spectrometry equipment. B. cereus is known as one of the most frequent bacteria occurring in food. Most foods contain more than one kind of B. cereus. When randomly selected 100 isolates of B. cereus from commercial infant foods (dry formulas) were tested, 11% of these produced cereulide. Considering a frequent content of 103 to 104 cfu (colony forming units) of B. cereus per gram of infant food formula (dry), it appears likely that most servings (200 ml, 30 g of the powder reconstituted with water) may contain cereulide producers. When a reconstituted infant formula was inoculated with >105 cfu of cereulide producing B. cereus per ml and left at room temperature, cereulide accumulated to food poisoning levels (> 0.1 mg of cereulide per serving) within 24 hours. Paradoxically, the amount of cereulide (per g of food) increased 10 to 50 fold when the food was diluted 4 - 15 fold with water. The amount of the produced cereulide strongly depended on the composition of the formula: most toxin was formed in formulas with cereals mixed with milk, and least toxin in formulas based on milk only. In spite of the aggressive cleaning practices executed by the modern dairy industry, certain genotypes of B. cereus appear to colonise the silos tanks. In this thesis four strategies to explain their survival of their spores in dairy silos were identified. First, high survival (log 15 min kill ≤ 1.5) in the hot alkaline (pH >13) wash liquid, used at the dairies for cleaning-in-place. Second, efficient adherence of the spores to stainless steel from cold water. Third, a cereulide producing group with spores characterized by slow germination in rich medium and well preserved viability when exposed to heating at 90 ºC. Fourth, spores capable of germinating at 8 ºC and possessing the psychrotolerance gene, cspA. There were indications that spores highly resistant to hot 1% sodium hydroxide may be effectively inactivated by hot 0.9% nitric acid. Eight out of the 14 dairy silo tank isolates possessing hot alkali resistant spores were capable of germinating and forming biofilm in whole milk, not previously reported for B. cereus. In this thesis it was shown that cereulide producing B. cereus was capable of inhibiting the growth of cereulide non-producing B. cereus occurring in the same food. This phenomenon, called antagonism, has long been known to exist between B. cereus and other microbial species, e.g. various species of Bacillus, gram-negative bacteria and plant pathogenic fungi. In this thesis intra-species antagonism of B. cereus was shown for the first time. This brother-killing did not depend on the cereulide molecule, also some of the cereulide non-producers were potent antagonists. Interestingly, the antagonistic clades were most frequently found in isolates from food implicated with human illness. The antagonistic property was therefore proposed in this thesis as a novel virulence factor that increases the human morbidity of the species B. cereus, in particular of the cereulide producers.

Fish mediate high food web connectivity in the lower reaches of a tropical floodplain river

High levels of hydrological connectivity during seasonal flooding provide significant opportunities for movements of fish between rivers and their floodplains, estuaries and the sea, possibly mediating food web subsidies among habitats. To determine the degree of utilisation of food sources from different habitats in a tropical river with a short floodplain inundation duration (similar to 2 months), stable isotope ratios in fishes and their available food were measured from three habitats (inundated floodplain, dry season freshwater, coastal marine) in the lower reaches of the Mitchell River, Queensland (Australia). Floodplain food sources constituted the majority of the diet of large-bodied fishes (barramundi Lates calcarifer, catfish Neoarius graeffei) captured on the floodplain in the wet season and for gonadal tissues of a common herbivorous fish (gizzard shad Nematalosa come), the latter suggesting that critical reproductive phases are fuelled by floodplain production. Floodplain food sources also subsidised barramundi from the recreational fishery in adjacent coastal and estuarine areas, and the broader fish community from a freshwater lagoon. These findings highlight the importance of the floodplain in supporting the production of large fishes in spite of the episodic nature and relatively short duration of inundation compared to large river floodplains of humid tropical regions. They also illustrate the high degree of food web connectivity mediated by mobile fish in this system in the absence of human modification, and point to the potential consequences of water resource development that may reduce or eliminate hydrological connectivity between the river and its floodplain.

Food and Indoor Air Isolated Bacillus Non-Protein Toxins: : Structures, Physico-Chemical Properties and Mechanisms of Effects on Eukaryotic Cells

We report here the structures and properties of heat-stable, non-protein, and mammalian cell-toxic compounds produced by spore-forming bacilli isolated from indoor air of buildings and from food. Little information is available on the effects and occurrence of heat-stable non-protein toxins produced by bacilli in moisture-damaged buildings. Bacilli emit spores that move in the air and can serve as the carriers of toxins, in a manner similar to that of the spores of toxic fungi found in contaminated indoor air. Bacillus spores in food cause problems because they tolerate the temperatures applied in food manufacture and the spores later initiate growth when food storage conditions are more favorable. Detection of the toxic compounds in Bacillus is based on using the change in mobility of boar spermatozoa as an indicator of toxic exposure. GC, LC, MS, and nuclear magnetic resonance NMR spectroscopy were used for purification, detection, quantitation, and analysis of the properties and structures of the compounds. Toxicity and the mechanisms of toxicity of the compounds were studied using boar spermatozoa, feline lung cells, human neural cells, and mitochondria isolated from rat liver. The ionophoric properties were studied using the BLM (black-lipid membrane) method. One novel toxin, forming ion channels permeant to K+ > Na+ > Ca2+, was found and named amylosin. It is produced by B. amyloliquefaciens isolated from indoor air of moisture-damaged buildings. Amylosin was purified with an RP-HPLC and a monoisotopic mass of 1197 Da was determined with ESI-IT-MS. Furthermore, acid hydrolysis of amylosin followed by analysis of the amino acids with the GS-MS showed that it was a peptide. The presence of a chromophoric polyene group was found using a NMR spectroscopy. The quantification method developed for amylosin based on RP-HPLC-UV, using the macrolactone polyene, amphotericin B (MW 924), as a reference compound. The B. licheniformis strains isolated from a food poisoning case produced a lipopeptide, lichenysin A, that ruptured mammalian cell membranes and was purified with a LC. Lichenysin A was identified by its protonated molecules and sodium- and potassium- cationized molecules with MALDI-TOF-MS. Its protonated forms were observed at m/z 1007, 1021 and 1035. The amino acids of lichenysin A were analyzed with ESI-TQ-MS/MS and, after acid hydrolysis, the stereoisomeric forms of the amino acids with RP-HPLC. The indoor air isolates of the strain of B. amyloliquefaciens produced not only amylosin but also lipopeptides: the cell membrane-damaging surfactin and the fungicidal fengycin. They were identified with ESI-IT-MS observing their protonated molecules, the sodium- and potassium-cationized molecules and analysing the MS/MS spectra. The protonated molecules of surfactin and fengycin showed m/z values of 1009, 1023, and 1037 and 1450, 1463, 1493, and 1506, respectively. Cereulide (MW 1152) was purified with RP-HPLC from a food poisoning strain of B. cereus. Cereulide was identified with ESI-TQ-MS according to the protonated molecule observed at m/z 1154 and the ammonium-, sodium- and potassium-cationized molecules observed at m/z 1171, 1176, and 1192, respectively. The fragment ions of the MS/MS spectrum obtained from the protonated molecule of cereulide at m/z 1154 were also interpreted. We developed a quantification method for cereulide, using RP-HPLC-UV and valinomycin (MW 1110, which structurally resembles cereulide) as the reference compound. Furthermore, we showed empirically, using the BLM method, that the emetic toxin cereulide is a specific and effective potassium ionophore of whose toxicity target is especially the mitochondria.