Development of a risk assessment for BSE in the aquatic environment

Bovine spongiform encephalopathy (BSE) is believed to be transmitted by the ingestion of proteinaceous agents called prions which accumulate in the brain and spinal cord of infected bovines. Concern has been expressed about the risks of transmission of BSE to humans through BSE prions discharged to the aquatic environment from rendering plants, abattoirs and landfills. The disease-related form of the prion protein is relatively resistant to degradation, and infectivity decays rather slowly in the environment. Levels of disinfection used for drinking water treatment would have little effect. This paper presents the assumptions which were used to model the risks from a rendering plant disposing of cull cattle carcasses in the catchment of a chalk aquifer which is used for a drinking water abstraction. The risk assessment approach focused on identifying the hydrogeological and physical barriers which would contribute to preventing BSE infectivity gaining entry to the aquifer. These barriers included inactivation of BSE agent by the rendering process, removal from the effluent by treatment at the plant, filtration and adsorption in the clay and chalk, and dilution in the ground water. The importance in environmental risk assessment of the cow-to-man species barrier is considered. Two key conclusions about the environmental behaviour of the BSE agent are that prion proteins are 'sticky' and bind to particulates, and that the millions of BSE prion molecules comprising a human oral ID₅₀ are subject to some degree of dispersion and hence dilution in the environment. Assuming the rendering plant processes 2000 cull cattle carcasses per week, the risks to drinking water consumers were estimated to be remote. Indeed, even using worst case assumptions an individual would have to consume 2 l d⁻¹ of tap water for 45 million years to have a 50% chance of infection through drinking water drawn from the aquifer.     

Sediment analysis for the assessment of risk from organic pollutants in lakes

Sediment analysis in three Italian subalpine lakes, very close to each other and exposed to different anthropogenic pressure, was used to assess the risk from organic pollutants, including some very persistent and widespread micropollutants (PCB, DDT, PAH) as well as known local pollutants. Additionally, a wide-spectrum characterization of organic compounds by GLC-MS analysis allowed the detection of other classes of chemicals. The contamination levels were related to long distance transport and to local sources of pollution in the watershed. PAH, PCB and organochlorine pesticide levels agree with the load calculated from atmospheric depositions. Local sources of pollution are responsible for the contamination by aliphatic hydrocarbons and tris(monochloroisopropyl)-phosphate (TCIPP); these compounds were found only in the two most urbanized basins. Heavy contamination by TCIPP was found in both water and sediments; water concentrations are considered hazardous for drinking purposes. This study shows that local contaminants, not yet included in any priority list, may single; out as a major environmental concern.     

An assessment of actinobacterial diversity in the marine environment

The 16S rRNA gene sequence diversity within the Phylum Actinobacteria was assessed from four sources: PCR-generated V6 sequence tags derived from seawater samples, metagenomic data from the Global Ocean Sampling (GOS) expedition, marine-derived sequences maintained in the Ribosomal Database Project (RDP), and select cultured strains for which sequence data is not yet available in the RDP. This meta-analysis revealed remarkable levels of phylogenetic diversity and confirms the existence of major, deeply rooted, and as of yet uncharacterized lineages within the phylum. A dramatic incongruence among cultured strains and those detected using culture-independent techniques was also revealed. Redundancy among the actinobacteria detected using culture-independent techniques suggests that greater sequence coverage or improved DNA extraction efficiencies may be required to detect the rare phylotypes that can be readily cultured from marine samples. Conversely, new strategies need to be developed for the cultivation of frequently observed but yet to be cultured marine actinobacteria.     

Comparative proteomics as a new tool for exploring human mitochondrial tRNA disorders.

More than 70 different point mutations in human mitochondrial tRNA genes are correlated with severe disorders, including fatal cardiopathies, encephalopathies, myopathies, and others. So far, investigation of the molecular impact(s) of mutations has focused on the affected tRNA itself by seeking structural and/or functional perturbations capable of interfering with synthesis of the 13 mitochondrion-encoded subunits of respiratory chain complexes. Here, a proteomic approach was used to investigate whether such mutations would affect the pattern of mitochondrial proteins at a broader level. Analysis of several hundred mitochondrial proteins from sibling cybrid cell lines by two-dimensional electrophoresis, an approach that takes into account all regulatory steps of mitochondrial and nuclear gene expression, indeed reveals a number of up- and downregulated proteins when healthy and single-point-mutation-carrying mitochondria representative of either MELAS or MERRF syndrome were compared. Assignment by mass spectrometry of the two proteins which exhibit obvious large quantitative decreases in the levels of both pathologic mitochondria identified nuclear-encoded subunits of cytochrome c oxidase, a respiratory chain complex. This clearly shows a linkage between the effects of mutations in mitochondrial tRNA genes and the steady-state level of nuclear-encoded proteins in mitochondria. It opens new routes toward a large-scale exploration of potential proteic partners involved in the genotype-phenotype correlation of mitochondrial disorders.     

Randomly amplified polymorphic DNA PCR as a tool for assessment of marine viral richness

Recent discoveries have uncovered considerable genetic diversity among aquatic viruses and raised questions about the variability of this diversity within and between environments. Studies of the temporal and spatial dynamics of aquatic viral assemblages have been hindered by the lack of a common genetic marker among viruses for rapid diversity assessments. Randomly amplified polymorphic DNA (RAPD) PCR bypasses this obstacle by sampling at the genetic level without requiring viral isolation or previous sequence knowledge. In this study, the utility of RAPD-PCR for assessing DNA viral richness within Chesapeake Bay water samples was evaluated. RAPD-PCR using single 10-mer oligonucleotide primers successfully produced amplicons from a variety of viral samples, and banding patterns were highly reproducible, indicating that each band likely represents a single amplicon originating from viral template DNA. In agreement with observations from other community profiling techniques, resulting RAPD-PCR banding patterns revealed more temporal than spatial variability in Chesapeake Bay virioplankton assemblages. High-quality hybridization probes and sequence information were also easily generated from single RAPD-PCR products or whole reactions. Thus, the RAPD-PCR technique appears to be practical and efficient for routine use in high-resolution viral diversity studies by providing assemblage comparisons through fingerprinting, probing, or sequence information.     

The Hirsch-index: a simple, new tool for the assessment of scientific output of individual scientists

In this brief paper we explore the Hirsch-index together with a couple of other bibliometric parameters for the assessment of the scientific output of 29 Dutch professors in clinical cardiology. It appears that even within such a homogeneous group there is large interindividual variability. Although the differences are quite remarkable, it remains undetermined what they mean; at least it is premature to interpret them as differences in scientific quality. It goes without saying that even more prudence is required when different fields of medicine and life sciences are compared (for example within University Medical Centres). Recent efforts to produce an amalgam of scientific ‘productivity’, ‘relevance’ and ‘viability’ as a surrogate parameter for the assessment of scientific quality, as for example performed in the AMC in Amsterdam, should be discouraged in the absence of a firm scientific base. Unfortunately for politicians and ‘managers of science’ only reading papers and studying are suitable for quality assessment of scientific output. Citations analyses can't substitute that. (Neth Heart J 2009;17:145–54.)     

Olfactory assessment of predation risk in the aquatic environment

The aquatic environment is well suited for the transmission of chemical information. Aquatic animals have evolved highly sensitive receptors for detecting these cues. Here, I review behavioural evidence for the use of chemical cues by aquatic animals for the assessment of predation risk. Chemical cues are released during detection, attack, capture and ingestion of prey. The nature of the cue released depends on the stage of the predation sequence in which cues are released. Predator odours, disturbance pheromones, injury-released chemical cues and dietary cues all convey chemical information to prey Prey use these cues to minimize their probability of being taken on to the next stage of the sequence. The evolution of specialized epidermal alarm substance cells in fishes in the superorder Ostariophysi represent an amplification of this general phenomenon. These cells carry a significant metabolic cost. The cost is offset by the fitness benefit of the chemical attraction of predators. Attempts of piracy by secondary predators interrupt predation events allowing prey an opportunity for escape. In conclusion, chemical cues are widely used by aquatic prey for risk assessment and this has resulted in the evolution of specialized structures among some taxa.     

Stepwise quantitative risk assessment as a tool for characterization of microbiological food safety

This paper describes a system for the microbiological quantitative risk assessment for food products and their production processes. The system applies a stepwise risk assessment, allowing the main problems to be addressed before focusing on less important problems. First, risks are assessed broadly, using order of magnitude estimates. Characteristic numbers are used to quantitatively characterize microbial behaviour during the production process. These numbers help to highlight the major risk-determining phenomena, and to find negligible aspects. Second, the risk-determining phenomena are studied in more detail. Both general and/or specific models can be used for this and varying situations can be simulated to quantitatively describe the risk-determining phenomena. Third, even more detailed studies can be performed where necessary, for instance by using stochastic variables. The system for quantitative risk assessment has been implemented as a decision supporting expert system called SIEFE: Stepwise and Interactive Evaluation of Food safety by an Expert System. SIEFE performs bacterial risk assessments in a structured manner, using various information sources. Because all steps are transparent, every step can easily be scrutinized. In the current study the effectiveness of SIEFE is shown for a cheese spread. With this product, quantitative data concerning the major risk-determining factors were not completely available to carry out a full detailed assessment. However, this did not necessarily hamper adequate risk estimation. Using ranges of values instead helped identifying the quantitatively most important parameters and the magnitude of their impact. This example shows that SIEFE provides quantitative insights into production processes and their risk-determining factors to both risk assessors and decision makers, and highlights critical gaps in knowledge.     

Multidisciplinary rapid assessment of coastal areas as a tool for the design and management of marine protected areas

A method is described for rapid multidisciplinary environmental assessment of coastal areas within the conceptual framework of comprehensive management of Marine Protected Areas (MPAs). The aim is to provide tools for the selection, design and management of coastal MPAs when time, budget or potential human pressures, either alone or in combination, create an urgent need for prioritisation. Maximising results and minimising cost and time is the goal, using a methodology that re evaluates existing information on the area, allows use of physical, environmental and socio-economic indicators, and finally integrates information in a Geographic Information System capable of generating outputs in the form of thematic maps to support managers.The final products obtained inform planners and managers about the study areas, across multiple aspects that all need to be considered in integrated coastal management. Although originally proposed for widespread use in the Mediterranean, this methodology can be flexibly adapted, with minor modifications in the selection of indicators, for its use in other regions. The results show its potential for merging and synthesising information not only as a tool in Rapid Assessment Programs but also as a tool for facing management of wide coastal areas as social-ecological ecosystems.     

Anti-sulfonylbenzoate antibodies as a tool for the detection of nucleotide-binding proteins for functional proteomics

Proteins that bind ATP and GTP are important cellular components. We developed an immunological approach to selectively tag nucleotide-binding proteins based on the use of 5'-[4-(fluorosulfonyl)benzoyl]adenosine and 5'-[4-(fluorosulfonyl)benzoyl]guanosine affinity tags and an antibody against 4-(sulfonyl)benzoate. Detection follows affinity labeling, gel electrophoresis, and ester bond cleavage to expose the epitope. Trial analyses of labeled proteins from lymphoid cells identified multiple ATP-binding proteins, including chaperones, actin, kinases, an RNA splicing factor, a membrane ATPase, and ATP synthase.     

Medical simulation: the new tool for training and skill assessment

Medical simulation is a new method to facilitate skill training and assessment. Simulation has achieved a high degree of sophistication in aviation and other fields. However, the complexity of health care, the numerous stakeholders, and the lack of central control of medical education have been barriers to the development and broad implementation of medical simulation. Acceptance by the medical community is growing, with the publication of scientific validation studies, the development of economic models and funding, and the integration of simulation into existing curricula and training programs. The major forces for implementing simulation will most likely come from the medical device industry and from institutions with mandates to improve the quality of health care and enhance patient safety. Certification boards are expected to increase their utilization of simulation technology to objectively assess proficiency of skills relevant to physicians and the health care system. Medical simulation has made the transition from an experimental technology to the clinical world, and the next five to 10 years may be viewed as the golden age of medical simulation.     

Decorospora, a new genus for the marine ascomycete Pleospora gaudefroyi

In this paper, we investigate the phylogenetic placement of Pleospora gaudefroyi using partial SSU as well as ITS ribosomal DNA sequences. Both SSU and ITS data sets agreed in the placement of P. gaudefroyi. Parsimony and neighbor-joining analyses of each data set placed P. gaudefroyi within the Pleosporaceae with 100% bootstrap support. Pleospora gaudefroyi was sister taxon in the Pleosporaceae represented by Alternaria alternata, Cochliobolus sativus, Pleospora herbarum, Pyrenophora tritici-repentis and Setosphaeria rostrata. Pleospora gaudefroyi was separated from other genera in the Pleosporaceae in 94% of the bootstrap replicates in parsimony and neighbor-joining analyses. When P. gaudefroyi was constrained to monophyly with P. herbarum, all resulting trees were significantly worse than the optimal tree in both Kishino-Hasegawa and Shimodaira-Hasegawa tests. Pleospora gaudefroyi was therefore excluded from Pleospora, and transferred to the new genus Decorospora placed in the Pleosporaceae. Decorospora (Dothideomycetes) has characteristic ascospores enclosed in a sheath with 4-5 apical extensions. The distribution and substrate types for D. gaudefroyi are summarized and updated based on additional collections.     

Lipolysis index: evaluation of a new tool for metabolic assessment in epidemiological studies on obesity

Lipid mobilization through adipocyte lipolysis is central for energy metabolism and is decreased in obesity. However, the factors of importance for lipolytic activity in the general population are not known. To further examine this we performed a cross-sectional study on teenagers and adults. We constructed and evaluated a simple index of lipolytic activity (ratio of fasting p-glycerol and body fat %) in population based samples in 316 teenagers (BMI 16-51 kg/m (2)) and 3,039 adults (BMI 16-70 kg/m (2)). In the adults, multiple regression analysis showed that waist and BMI but not age, plasma insulin, plasma noradrenaline or waist-to-hip ratio contributed independently and inversely to lipolytic activity (partial r=-0.37 and -0.28, respectively, p<0.0001). Together waist and BMI explained about 45% of the variability of lipolysis. Waist was a stronger factor than BMI in stepwise regression. The same analysis in teenagers showed that only BMI contributed independently and inversely to lipolytic activity (partial r=-0.90, p<0.0001) and explained about 55% of lipolysis variation. BMI had the strongest effect on lipolysis in lean teenagers. The results were the same for men and women. At all levels of lipolytic activity plasma fatty acid levels were elevated in obese subjects (p<0.0001). We conclude that during adolescence BMI is the major factor negatively influencing lipolytic activity, in particular among lean young subjects. In adulthood central fat accumulation together with increasing BMI decreases lipolysis. In spite of low lipolytic activity circulating fatty acid levels are increased in obesity, probably due to an adipose mass effect.     

Analysis of protein expression in cell microarrays: a tool for antibody-based proteomics.

Tissue microarray (TMA) technology provides a possibility to explore protein expression patterns in a multitude of normal and disease tissues in a high-throughput setting. Although TMAs have been used for analysis of tissue samples, robust methods for studying in vitro cultured cell lines and cell aspirates in a TMA format have been lacking. We have adopted a technique to homogeneously distribute cells in an agarose gel matrix, creating an artificial tissue. This enables simultaneous profiling of protein expression in suspension- and adherent-grown cell samples assembled in a microarray. In addition, the present study provides an optimized strategy for the basic laboratory steps to efficiently produce TMAs. Presented modifications resulted in an improved quality of specimens and a higher section yield compared with standard TMA production protocols. Sections from the generated cell TMAs were tested for immunohistochemical staining properties using 20 well-characterized antibodies. Comparison of immunoreactivity in cultured dispersed cells and corresponding cells in tissue samples showed congruent results for all tested antibodies. We conclude that a modified TMA technique, including cell samples, provides a valuable tool for high-throughput analysis of protein expression, and that this technique can be used for global approaches to explore the human proteome.