Assessment and control of Bacillus cereus emetic toxin in food

Despite of improving levels of hygiene, the incidence of registered food borne disease has been at the same level for many years: there were 40 to 90 epidemics in which 1000-9000 persons contracted food poisoning through food or drinking water in Finland. Until the year 2004 salmonella and campylobacter were the most common bacterial causes of food borne diseases, but in years 2005-2006 Bacillus cereus was the most common. Similar developement has been published i.e. in Germany already in the 1990´s. One reason for this can be Bacillus cereus and its emetic toxin, cereulide. Bacillus cereus is a common environmental bacterium that contaminates raw materials of food. Otherwise than salmonella and campylobacter, Bacillus cereus is a heat resistant bacterium, capable of surviving most cooking procedures due to the production of highly thermo resistant spores. The food involved has usually been heat treated and surviving spores are the source of the food poisoning. The heat treatment induces germination of the spore and the vegetative cells then produce toxins. This doctoral thesis research focuses on developing methods for assessing and eliminating risks to food safety by cereulide producing Bacillus cereus. The biochemistry and physiology of cereulide production was investigated and the results were targeted to offer tools for minimizing toxin risk in food during the production. I developed methods for the extraction and quantitative analysis of cereulide directly from food. A prerequisite for that is knowledge of the chemical and physical properties of the toxin. Because cereulide is practically insoluble in water, I used organic solvents; methanol, ethanol and pentane for the extraction. For extraction of bakery products I used high temperature (100C) and pressure (103.4 bars). Alternaties for effective extraction is to flood the plain food with ethanol, followed by stationary equilibration at room temperature. I used this protocol for extracting cereulide from potato puree and penne. Using this extraction method it is also possible also extract cereulide from liquid food, like milk. These extraction methods are important improvement steps for studying of Bacillus cereus emetic food poisonings. Prior my work, cereulide extraction was done using water. As the result, the yield was poor and variable. To investigate suspected food poisonings, it is important to show actual toxicity of the incriminated food. Many toxins, but not cereulide, inactivate during food processing like heating. The next step is to identify toxin by chemical methods. I developed with my colleague Maria Andesson a rapid assay for the detection of cereulide toxicity, within 5 to 15 minutes. By applying this test it is possible to rapidly detect which food was causing the food poisoning. The chemical identification of cereulide was achieved using mass spectrometry. I used cereulide specific molecular ions, m/z (+/-0.3) 1153.8 (M+H+), 1171.0 (M+NH4+), 1176.0 (M+Na+) and 1191.7 (M+K+) for reliable identification. I investigated foods to find out their amenability to accumulate cereulide. Cereulide was formed high amounts (0.3 to 5.5 microg/g wet wt) when of cereulide producing B. cereus strains were present in beans, rice, rice-pastry and meat-pastry, if stored at non refrigerated temperatures (21-23C). Rice and meat pastries are frequently consumed under conditions where no cooled storage is available e.g. picnics and outdoor events. Bacillus cereus is a ubiquitous spore former and is therefore difficult to eliminate from foods. It is therefore important to know which conditions will affect the formation of cereulide in foods. My research showed that the cereulide content was strongly (10 to 1000 fold differences in toxin content) affected by the growth environment of the bacterium. Storage of foods under nitrogen atmosphere (> 99.5 %) prevented the production of cereulide. But when also carbon dioxide was present, minimizing the oxygen contant (< 1%) did not protect the food from formation of cereulide in preliminary experiments. Also food supplements affected cereulide production at least in the laboratory. Adding free amino acids, leucine and valine, stimulated cereulide production 10 to 20 fold. In peptide bonded form these amino acids are natural constituents in all proteins. Interestingly, adding peptide bonded leucine and valine had no significant effect on cereulide production. Free amino acids leucine and valine are approved food supplements and widely used as flawour modifiers in food technology. My research showed that these food supplements may increase food poisoning risk even though they are not toxic themselves.

Modeling Efficient Classification as a Process of Confidence Assessment and Delegation

In visual object detection and recognition, classifiers have two interesting characteristics: accuracy and speed. Accuracy depends on the complexity of the image features and classifier decision surfaces. Speed depends on the hardware and the computational effort required to use the features and decision surfaces. When attempts to increase accuracy lead to increases in complexity and effort, it is necessary to ask how much are we willing to pay for increased accuracy. For example, if increased computational effort implies quickly diminishing returns in accuracy, then those designing inexpensive surveillance applications cannot aim for maximum accuracy at any cost. It becomes necessary to find trade-offs between accuracy and effort. We study efficient classification of images depicting real-world objects and scenes. Classification is efficient when a classifier can be controlled so that the desired trade-off between accuracy and effort (speed) is achieved and unnecessary computations are avoided on a per input basis. A framework is proposed for understanding and modeling efficient classification of images. Classification is modeled as a tree-like process. In designing the framework, it is important to recognize what is essential and to avoid structures that are narrow in applicability. Earlier frameworks are lacking in this regard. The overall contribution is two-fold. First, the framework is presented, subjected to experiments, and shown to be satisfactory. Second, certain unconventional approaches are experimented with. This allows the separation of the essential from the conventional. To determine if the framework is satisfactory, three categories of questions are identified: trade-off optimization, classifier tree organization, and rules for delegation and confidence modeling. Questions and problems related to each category are addressed and empirical results are presented. For example, related to trade-off optimization, we address the problem of computational bottlenecks that limit the range of trade-offs. We also ask if accuracy versus effort trade-offs can be controlled after training. For another example, regarding classifier tree organization, we first consider the task of organizing a tree in a problem-specific manner. We then ask if problem-specific organization is necessary.

Physical Properties of Deep Drill Cores : Implications for Meteorite Impact Effects and Crustal Structures

Physical properties provide valuable information about the nature and behavior of rocks and minerals. The changes in rock physical properties generate petrophysical contrasts between various lithologies, for example, between shocked and unshocked rocks in meteorite impact structures or between various lithologies in the crust. These contrasts may cause distinct geophysical anomalies, which are often diagnostic to their primary cause (impact, tectonism, etc). This information is vital to understand the fundamental Earth processes, such as impact cratering and associated crustal deformations. However, most of the present day knowledge of changes in rock physical properties is limited due to a lack of petrophysical data of subsurface samples, especially for meteorite impact structures, since they are often buried under post-impact lithologies or eroded. In order to explore the uppermost crust, deep drillings are required. This dissertation is based on the deep drill core data from three impact structures: (i) the Bosumtwi impact structure (diameter 10.5 km, 1.07 Ma age; Ghana), (ii) the Chesapeake Bay impact structure (85 km, 35 Ma; Virginia, U.S.A.), and (iii) the Chicxulub impact structure (180 km, 65 Ma; Mexico). These drill cores have yielded all basic lithologies associated with impact craters such as post-impact lithologies, impact rocks including suevites and breccias, as well as fractured and unfractured target rocks. The fourth study case of this dissertation deals with the data of the Paleoproterozoic Outokumpu area (Finland), as a non-impact crustal case, where a deep drilling through an economically important ophiolite complex was carried out. The focus in all four cases was to combine results of basic petrophysical studies of relevant rocks of these crustal structures in order to identify and characterize various lithologies by their physical properties and, in this way, to provide new input data for geophysical modellings. Furthermore, the rock magnetic and paleomagnetic properties of three impact structures, combined with basic petrophysics, were used to acquire insight into the impact generated changes in rocks and their magnetic minerals, in order to better understand the influence of impact. The obtained petrophysical data outline the various lithologies and divide rocks into four domains. Based on target lithology the physical properties of the unshocked target rocks are controlled by mineral composition or fabric, particularly porosity in sedimentary rocks, while sediments result from diverse sedimentation and diagenesis processes. The impact rocks, such as breccias and suevites, strongly reflect the impact formation mechanism and are distinguishable from the other lithologies by their density, porosity and magnetic properties. The numerous shock features resulting from melting, brecciation and fracturing of the target rocks, can be seen in the changes of physical properties. These features include an increase in porosity and subsequent decrease in density in impact derived units, either an increase or a decrease in magnetic properties (depending on a specific case), as well as large heterogeneity in physical properties. In few cases a slight gradual downward decrease in porosity, as a shock-induced fracturing, was observed. Coupled with rock magnetic studies, the impact generated changes in magnetic fraction the shock-induced magnetic grain size reduction, hydrothermal- or melting-related magnetic mineral alteration, shock demagnetization and shock- or temperature-related remagnetization can be seen. The Outokumpu drill core shows varying velocities throughout the drill core depending on the microcracking and sample conditions. This is similar to observations by Kern et al., (2009), who also reported the velocity dependence on anisotropy. The physical properties are also used to explain the distinct crustal reflectors as observed in seismic reflection studies in the Outokumpu area. According to the seismic velocity data, the interfaces between the diopside-tremolite skarn layer and either serpentinite, mica schist or black schist are causing the strong seismic reflectivities.

Assessment and mitigation of agricultural water pollution

Yhteenveto: Maatalouden aiheuttama vesistönkuormitus ja sen vähentäminen