Search

Op 27 september 2004 promoveerde Marco Kruijt aan Wageningen Universiteit op het proefschrift getiteld 'Molecular evolution of Cladosporium fulvum disease resistance genes in wild tomato'. De evolutie van het Cf-9 gen in de wilde tomatensoort L.pimpinellifolium is in detail bestudeerd. Naast het Cf-9 gen blijkt nog een resistentiegen, het 9DCgen, aanwezig. Het Cf-9 gen is ouder dan het 9DC gen en komt in verschillende wilde tomatensoorten voor. Hieruit blijkt ondermeer dat C. fulvum waarschijnlijk al een pathogeen van de 'oertomaat' was

Een aantal fokprogramma's met gebruikmaking van superovulatie en embryotransplantatie (MOET) is vergeleken wat betreft de te verwachten genetische vooruitgang t.o.v. een programma zonder MOET bij nucleusmoeders

Op basis van literatuurgegevens is de stof carbendazim geodentificeerd als de meest milieukritische stof die naast enkele andere stoffen waarschijnlijk hoofdzakelijk verantwoordelijk is voor bodemverontreiniging onder bezinkbassins. Berekeningen latenzien dat de zone waarin bodemverontreiniging onder bassins optreedt, beperkt blijft door biologische afbraak van bestrijdingsmiddelen in de bodem. De dikte van deze zone is maximaal 6-7 m zonder bodemafdichting en minder dan 1 m met bodemafdichting. Emissies vanuit bezinkbassins naar de bodem zijn geringer dan emissies via drainage naar het oppervlaktewater, maar niet verwaarloosbaar en bovendien geconcentreerd op een relatief gering oppervlak. Emissies kunnen met 90-99% worden gereduceerd. De goedkoopste oplossingen, die echter geen absolute bescherming van de bodem bieden, lijken afdichting met plaatselijk aanwezige klei, hydrologiche isolatie al dan niet in combinatie met een afdichtingslaag, en adsorptielagen. Praktijkervaring met deze oplossingen ontbreekt en nader onderzoek naar de effectiviteit is aanbevolen.

Litter size is an important characteristic in pig breeding. Apart from selection within available lines, also the development of a synthetic line with the Chinese Meishan breed could be an interesting approach to obtain a line with an increased level of litter size. To investigate genetic aspects of traits of interest in such a synthetic line, Dutch pig breeding companies have produced F 1 and F 2 Meishan x Western crossbreds. This thesis focusses on one important genetic aspect, the presence of major genes. In Chapters 2 to 4, statistical methodology to model a major gene inheritance is investigated and developed Chapters 5 and 6 consider analysis of data collected on the produced Meishan crossbreds for presence of major genes. To develop a synthetic line with Meishan, presence of major genes affecting litter size, growth and fatness is of interest. Additionally, the presence of major genes is investigated for meat quality traits.Statistical methodologyIn Chapter 2, the possibility to detect major genes by use of F 1 and F 2 is investigated. Here, special attention is paid to the situation where alleles at the major locus are fixed in the founder populations. Using 1000 F 2 observations, the power to detect major genes reaches more than 95% for additive and completely dominant effects (difference between homozygotes) of 4 and 2 residual standard deviations, respectively. When F, data is included, any increase in variance from F 1 to F 2 biases parameter estimates and leads to putative detection of a major gene. Also when in reality alleles at the major locus segregate in the founder populations, parameter estimates become biased, unless the average allele frequency in the founder populations is exactly 0.5. Use of data and use of a model in which alleles segregate in parents, e.g. F 3 data, is concluded to give better robustness and larger power. The latter is confirmed in a separate study, as referenced in Chapter 7, which shows that effects up to 4 times as small can be detected when alleles at the major locus segregate in the founder lines. Based on the findings in Chapter 2, Chapters 3 and 4 focus on the development of general models for a mixed inheritance. Use of such models is referred to as 'segregation analysis'.In Chapter 3, an advancement is made for use of analytical approaches to segregation analysis. It is noted that animal breeding pedigrees, as opposed to human pedigrees, generally contain many loops, such that exact computation of likelihoods isinfeasible. Loops in animal breeding pedigrees arise due to multiple matings, i.e. sires are generally mated to several dams, and due to inbreeding. Multiple matings generally already create many loops when considering 3-generation pedigrees. In Chapter 3, 'iterative peeling' is introduced, a method equivalent to the traditional recursive peeling method to compute exact likelihoods in non-looped pedigrees, but which also can be used to obtain approximate likelihoods in looped pedigrees. In simulations, hypothesis testing and parameter estimation are compared based on approximated likelihoods in looped pedigrees and exact likelihoods in non-looped pedigrees. This shows that no biases are introduced by the approximation in looped pedigrees. Iterative peeling is developed and investigated using a monogenic model, but could be extended to compute likelihoods for a mixed inheritance model. Such extension, however, was not made because an alternative non-analytical approach became available and was developed in Chapter 4. 0In Chapter 4, the application of Gibbs sampling is considered for inference in a mixed inheritance model. Gibbs sampling is a Markov chain Monte Carlo procedure which does not require analytical approximation. The approximation in such an approach is of a different nature: a marginal posterior distribution, or a feature thereof, is estimated based on a finite sample from the true posterior distribution. To generate such a sample, a Markov chain is constructed with an equilibrium distribution equal to the posterior distribution to be approximated. For application of Gibbs sampling to a mixed inheritance model, an implementation on scalar components, as used for human populations, appears not efficient because mixing of parameters in the Markov chain is slow. Therefore, an approach with blockwise sampling of genotypes is proposed for use in animal populations. The blockwise sampling, by which genotypes of a sire and its final progeny were sampled jointly, is effective to improve mixing. In Chapter 4 it is concluded that further measures to improve mixing could be looked for. In later Chapters such a further improvement is found in the additional use of a relaxation technique. In Chapter 4, inferences are made from a single Gibbs chain. In later Chapters, this approach is improved by use of multiple chains from which convergence of the Gibbs sampler is assessed by comparison of between- and within chain variances in an analysis- of-variance. The use of Bayesian estimators, which is feasible when using Gibbs sampling, is found preferable over the use of classical maximum likelihood estimators. In Chapter 7, it is discussed that the use of Bayeslian procedures fits in a general trend to better account for uncertainty in statistical estimation procedures.Analysis of dataIn Chapters 5 and 6, analysis of data obtained on the Meishan crossbreds is presented. In Chapter 5, presence of major genes affecting meat quality traits is investigated using data from F 2 individuals. Cooking loss, drip loss, two pH measurements, intramuscular fat, shearforce and back-fat thickness (by HGP measurement) are found to be likely influenced by a major gene. In all cases, a recessive allele is found, which originates from one of the founder lines, likely the Meishan breed. By studying associations between genotypes for major genes affecting the various traits, it is concluded that cooking loss, two pH measurements and possibly backfat thickness are influenced by one gene, and that a second gene influences intramuscular fat and possibly shearforce and drip loss. The statistical findings are supported by demonstrating marked differences in vanances of families of fathers inferred as carriers and families of fathers inferred as non-carriers.In Chapter 6, presence of major genes is investigated for two growth traits, backfat thickness (by" ultrasonic measurement) and litter size at first and second parity, using data from F 1 and F 2 crossbreds. Here, two analyses are performed for each trait. In a first analysis, joint analysis of F, and F 2 crossbred data is performed, in which different error variances are fitted for F 1 and F 2 observations. In this first analysis, significant contributions of major-gene variance are found for the two growth traits, for backfat, and for litter size at first parity. In a second analysis, analysis of F 2 data only is performed to check whether no biases are introduced in the joint analysis of F 1 and F 2 data. In the second analysis, no major genes are found for growth traits. Major genes affecting backfat and litter size at first parity are confirmed. Effects of the gene affecting backfat are similar to the effects of the gene affecting backfat identified in Chapter 5, and this likely is the same gene. The major genes affecting backfat and litter size are dominant genes, of which the recessive alleles can be considered unfavourable.. the recessive alleles of these genes cause an increase of backfat and a decrease of litter size.General results from the statistical analyses indicate that further molecular genetic research effort to map these genes will have a high probability of success. III Chapter 7 benefits are discussed from selection against the recessive alleles of the genes influencing backfat and litter size, as well as use of the gene affecting intramuscular fat to produce extra-tasty quality meat.ConclusionsIn this thesis, segregation analysis (SA) is made applicable for use in animal populations. SA will be a valuable addition to linkage analysis, where SA will be more typically applied to large amounts of data which are routinely collected. In the search for genes affecting quantitative traits, SA can directly identify functional genes, and can estimate genotypes of animals for such a functional gene. In combination with linkage analyses, this could supply important aids for molecular geneticists to locate functional genes. In this thesis, a number of major genes was identified to affect traits in the Meishan crosses. Further genetic analyses could generate more knowledge on the regulation of the quantitative traits involved and will aid in assessing the value of these genes for practical breeding. Chapter 8 additionally describes expected variance changes in a synthetic line, which could aid to optimise selection in such a line.

Litter size is an important characteristic in pig breeding. Apart from selection within available lines, also the development of a synthetic line with the Chinese Meishan breed could be an interesting approach to obtain a line with an increased level of litter size. To investigate genetic aspects of traits of interest in such a synthetic line, Dutch pig breeding companies have produced F 1 and F 2 Meishan x Western crossbreds. This thesis focusses on one important genetic aspect, the presence of major genes. In Chapters 2 to 4, statistical methodology to model a major gene inheritance is investigated and developed Chapters 5 and 6 consider analysis of data collected on the produced Meishan crossbreds for presence of major genes. To develop a synthetic line with Meishan, presence of major genes affecting litter size, growth and fatness is of interest. Additionally, the presence of major genes is investigated for meat quality traits.Statistical methodologyIn Chapter 2, the possibility to detect major genes by use of F 1 and F 2 is investigated. Here, special attention is paid to the situation where alleles at the major locus are fixed in the founder populations. Using 1000 F 2 observations, the power to detect major genes reaches more than 95% for additive and completely dominant effects (difference between homozygotes) of 4 and 2 residual standard deviations, respectively. When F, data is included, any increase in variance from F 1 to F 2 biases parameter estimates and leads to putative detection of a major gene. Also when in reality alleles at the major locus segregate in the founder populations, parameter estimates become biased, unless the average allele frequency in the founder populations is exactly 0.5. Use of data and use of a model in which alleles segregate in parents, e.g. F 3 data, is concluded to give better robustness and larger power. The latter is confirmed in a separate study, as referenced in Chapter 7, which shows that effects up to 4 times as small can be detected when alleles at the major locus segregate in the founder lines. Based on the findings in Chapter 2, Chapters 3 and 4 focus on the development of general models for a mixed inheritance. Use of such models is referred to as 'segregation analysis'.In Chapter 3, an advancement is made for use of analytical approaches to segregation analysis. It is noted that animal breeding pedigrees, as opposed to human pedigrees, generally contain many loops, such that exact computation of likelihoods isinfeasible. Loops in animal breeding pedigrees arise due to multiple matings, i.e. sires are generally mated to several dams, and due to inbreeding. Multiple matings generally already create many loops when considering 3-generation pedigrees. In Chapter 3, 'iterative peeling' is introduced, a method equivalent to the traditional recursive peeling method to compute exact likelihoods in non-looped pedigrees, but which also can be used to obtain approximate likelihoods in looped pedigrees. In simulations, hypothesis testing and parameter estimation are compared based on approximated likelihoods in looped pedigrees and exact likelihoods in non-looped pedigrees. This shows that no biases are introduced by the approximation in looped pedigrees. Iterative peeling is developed and investigated using a monogenic model, but could be extended to compute likelihoods for a mixed inheritance model. Such extension, however, was not made because an alternative non-analytical approach became available and was developed in Chapter 4. 0In Chapter 4, the application of Gibbs sampling is considered for inference in a mixed inheritance model. Gibbs sampling is a Markov chain Monte Carlo procedure which does not require analytical approximation. The approximation in such an approach is of a different nature: a marginal posterior distribution, or a feature thereof, is estimated based on a finite sample from the true posterior distribution. To generate such a sample, a Markov chain is constructed with an equilibrium distribution equal to the posterior distribution to be approximated. For application of Gibbs sampling to a mixed inheritance model, an implementation on scalar components, as used for human populations, appears not efficient because mixing of parameters in the Markov chain is slow. Therefore, an approach with blockwise sampling of genotypes is proposed for use in animal populations. The blockwise sampling, by which genotypes of a sire and its final progeny were sampled jointly, is effective to improve mixing. In Chapter 4 it is concluded that further measures to improve mixing could be looked for. In later Chapters such a further improvement is found in the additional use of a relaxation technique. In Chapter 4, inferences are made from a single Gibbs chain. In later Chapters, this approach is improved by use of multiple chains from which convergence of the Gibbs sampler is assessed by comparison of between- and within chain variances in an analysis- of-variance. The use of Bayesian estimators, which is feasible when using Gibbs sampling, is found preferable over the use of classical maximum likelihood estimators. In Chapter 7, it is discussed that the use of Bayeslian procedures fits in a general trend to better account for uncertainty in statistical estimation procedures.Analysis of dataIn Chapters 5 and 6, analysis of data obtained on the Meishan crossbreds is presented. In Chapter 5, presence of major genes affecting meat quality traits is investigated using data from F 2 individuals. Cooking loss, drip loss, two pH measurements, intramuscular fat, shearforce and back-fat thickness (by HGP measurement) are found to be likely influenced by a major gene. In all cases, a recessive allele is found, which originates from one of the founder lines, likely the Meishan breed. By studying associations between genotypes for major genes affecting the various traits, it is concluded that cooking loss, two pH measurements and possibly backfat thickness are influenced by one gene, and that a second gene influences intramuscular fat and possibly shearforce and drip loss. The statistical findings are supported by demonstrating marked differences in vanances of families of fathers inferred as carriers and families of fathers inferred as non-carriers.In Chapter 6, presence of major genes is investigated for two growth traits, backfat thickness (by" ultrasonic measurement) and litter size at first and second parity, using data from F 1 and F 2 crossbreds. Here, two analyses are performed for each trait. In a first analysis, joint analysis of F, and F 2 crossbred data is performed, in which different error variances are fitted for F 1 and F 2 observations. In this first analysis, significant contributions of major-gene variance are found for the two growth traits, for backfat, and for litter size at first parity. In a second analysis, analysis of F 2 data only is performed to check whether no biases are introduced in the joint analysis of F 1 and F 2 data. In the second analysis, no major genes are found for growth traits. Major genes affecting backfat and litter size at first parity are confirmed. Effects of the gene affecting backfat are similar to the effects of the gene affecting backfat identified in Chapter 5, and this likely is the same gene. The major genes affecting backfat and litter size are dominant genes, of which the recessive alleles can be considered unfavourable.. the recessive alleles of these genes cause an increase of backfat and a decrease of litter size.General results from the statistical analyses indicate that further molecular genetic research effort to map these genes will have a high probability of success. III Chapter 7 benefits are discussed from selection against the recessive alleles of the genes influencing backfat and litter size, as well as use of the gene affecting intramuscular fat to produce extra-tasty quality meat.ConclusionsIn this thesis, segregation analysis (SA) is made applicable for use in animal populations. SA will be a valuable addition to linkage analysis, where SA will be more typically applied to large amounts of data which are routinely collected. In the search for genes affecting quantitative traits, SA can directly identify functional genes, and can estimate genotypes of animals for such a functional gene. In combination with linkage analyses, this could supply important aids for molecular geneticists to locate functional genes. In this thesis, a number of major genes was identified to affect traits in the Meishan crosses. Further genetic analyses could generate more knowledge on the regulation of the quantitative traits involved and will aid in assessing the value of these genes for practical breeding. Chapter 8 additionally describes expected variance changes in a synthetic line, which could aid to optimise selection in such a line.

Een aantal fokprogramma's met gebruikmaking van superovulatie en embryotransplantatie (MOET) is vergeleken wat betreft de te verwachten genetische vooruitgang t.o.v. een programma zonder MOET bij nucleusmoeders

Since the foundation of colloid chemistry as a branch of science, much attention has been paid to the subject of colloid stability, i.e. the stability of colloid systems against aggregation. Gradually, our knowledge of the mechanisms involved has improved and models were developed, comprised in the DUO theory, which form the basis of a quantitative description of the stability of a colloidal system. There is plenty of experimental evidence which substantiate the correctness of the principles of the DUO theory, and hence, this theory is regarded as one of the fundaments of colloid chemistry. However, in one respect the theory is not confirmed by experiments: calculations predict pronounced size effects, but in practice stability seems to be little affected by particle size.It was the purpose of this study to gain insight in this contradictory matter. Chapter 1 offers a more extended introduction to the problem as well as the outline of this study.Chapter 2 focusses attention on a model system which meets the outlined requirements regarding surface charge, homodispersity, sphericity and particle size range: a method is described for the synthesis of homodisperse haematite (α-Fe 2 O 3 ) sols containing particles whose sizes vary from 35 nm up to 700 nm. This method is principally based on the gradual growth of haematite seeds in supersaturated FeCl 3 solutions (heterogeneous nucleation) up to the desired paticle size. As slight deviations in the composition of the growth medium have drastic effects on the shape of the final colloid, emphasize is given to the description of optimal synthesis conditions for spherically shaped particles. Kinetic experiments, performed to unravel the principles of particle growth revealed that the precipitation process is governed by diffusion.The coagulation experiments described in chapter 5 are monitored by turbidity measurements. Therefore the characterisation of the optical properties of the sols is a prerequisite for further studies. The sols under investigation are homodiperse and contain spherically shaped particles. This makes them particularly suited for such an evaluation, as their scattering behaviour can be interpreted in terms of the Mie theory. Such a comparsion, leading to values for the refractive index (n) and the absorption coefficient (K) in the wavelength range from 400 to 800 nm is made in chapter 3 . The agreement between calculations and experiments is good for any wave length in the visible range, and for any particle size studied, although the particles are monocrystalline and no perfect spheres.Any study dealing with electrostatic stabilisation demands some knowledge of the electrochemical behaviour of the system under study. Chapter 4 pays attention to the electrochemical characterisation of the haematite surface by comparing haematite samples from different origines. In these studies, potentiometric titrations, streaming potential measurements on haematite-coated capillaries and micro electrophoresis were used as the experimental tools to get access to the surface properties. Instead of providing unambiguous data, being valid for all iron oxides occuring in the (α-Fe 2 O 3 ) modification, the reported experiments emphasize that the crystal structure of the bulk phase is not the exclusive parameter in determining the electrochemical behaviour of an oxide. The purification procedure, or a heat treatment of the sol (aqueous or dried) plays an important part as well. There is some evidence that the crystal habit of the haematite surface is pH-dependent and that in some cases precipitated amorphous oxide may share in determining the surface properties. Though not going too much into details, the study gives evidence of the fact that the charging mechanisms of haematite are more complicated than expected on the grounds of purely crystallographic considerations.Inevitably this finding has its impact on the question of colloid stability, which is the dominating item of chapter 5 . Considering the outcome of the electrochemical study, is it still justified to assume that the haematite particles meet the demands of the stability theory regarding the sharpness of the boundary between bulk material and surrounding liquid? How could a diffuse surface layer be accounted for in existing stability models? Within the restrictions set by such questions, chapter 5 deals with the item of colloid stability with special reference to the influence of particle size on stability: though the value of the critical coagulation concentration depends on particle size and shows a minimum (!), the slopes of the log W - log C plots are virtually size independent. Such trends can be accounted for by the concepts of the DUO theory, if coagulation reversibility and shear effects are incorporated in the analysis. Deviations from sphericity, which are definitely observed for the systems under investigation. might explain some of the observed effects as is shown by some simple double layer calculations dealing with orientational effects in the interaction of a cubic particle with a half space.Finally, chapter 6 reflects on the preceding Items and pays attention to its limitations. Furthermore, it points to subjects which deserve further elaboration and mentions the means to make them experimentally accessible.

This thesis is concerned with the purification and properties of hydrogenase from the obligate anaerobic rumen bacterium Megasphaera elsdenii. In chapter 1 the motives underlying this thesis, the physiological role of hydrogenase in some heterotrophs, including Megasphaera elsdenii, as well as a comparison of physico-chemical and kinetic properties of purified hydrogenase preparations from microorganisms, showing great variations in physiology and taxonomy, are given. The physiological role and the physico-chemical and kinetic properties of the hydrogenases from Megasphaera elsdenii and Clostridium pasteurianum show great similarities.In chapter 2 the procedure for the anaerobic purification of hydrogenase from Megasphaera elsdenii is given. Two activity bands are separated on DEAE-cellulose chromatography, of which one (fraction I; see Table 1 of this chapter) represents a different hydrogenase or a complex of the enzyme in fraction II. Kinetics with the purified enzyme of the hydrogen production activity at pH 8, with the electron donors flavodoxin hydroquinone, reduced ferredoxin and methyl viologen semiquinone show non-linear double reciprocal plots of the activity versus the electron donor concentration. Two kinetic models were developed, with an identical general rate equation for the hydrogen production activity, which describe a random mechanism for the reaction of the oxidized enzyme with a proton and a reduced electron donor. These models also indicate that the hydrogenase accepts the two electrons necessary for hydrogen production in two separate, independent steps. Hydrogen oxidation with methyl and benzyl viologen shows, in contrast to hydrogen production, Michaelis Menten kinetics. In chapter 3 effectors of the hydrogenase activity, such as salts (which have hydrophylic properties) Me 2 SO and ethylene glycol (which have hydrophobic properties), and oxidants (oxygen, ferricyanide, Cl 2 Ind, (bi)sulphite) are described. The data show that the more chaotropic the anion of the salt the greater the increase in activity, per mole salt used. Both Me 2 SO and ethylene glycol inhibit the hydrogen evolution activity, however, Me 2 SO stimulates the hydrogen oxidation activity, while ethylene glycol does not affect this activity. Careful oxidation of the reduced enzyme with (bi)sulphite or Cl 2 Ind irreversibly increases the activity of the enzyme by about 60%; in contrast, oxygen and ferricyanide inactivate the enzyme. This irriversibly oxidized enzyme, which shows identical kinetic properties to the reduced enzyme, is more resistant to the effects of oxygen, Me 2 SO and storage, than the reduced enzyme. Difficulties encountered in ascribing redox potentials to the observed EPR spectra of hydrogenase at several redox states, such as the effects of temperature on the Nernst equation, on the apparent pH and on the midpoint potentials of the redox species, are described.In chapter 4 the theoretical aspects are described to determine the hydrogen production activity spectrophotometrically in a series coupled redox reactions as function of the pH and redox potential, as well as the limitations of this method. The hydrogen production activity is determined from the decrease in concentration of the reduced electron donor, or increase in the concentration of the oxidized electron donor. The changes in concentration of the reduced or oxidized electron donor do not equal the amount of hydrogen produced but only represent a measure of the actual amount of hydrogen produced.In chapter 5 the method as described in chapter 4 is used to study the effects of pH and redox potential on the hydrogen production activity. The hydrogen production activity is strongly pH- and redox potential-dependent; the activity patterns observed as function of pH and redox potential are independent of the nature of the electron donor used and thus represent a property of the enzyme. If at a given pH the activities are considered to express the degree of reduction of the enzyme the dependence of the activity on the redox potential represents a n=2 type redox titration curve with an 'apparent midpoint potential, which corresponds with the potential of the hydrogen electrode at that pH. To explain the effects of redox potential, proton and electron donor concentration on the hydrogenase activity Model I of chapter 2 was slightly adapted without changing its general rate equation for the hydrogen production activity. This model also explains the 'loss' of electrons of the reduced enzyme in EPR spectroscopy as well as the formation of HD together with DD during H 2 -D 2 O exchange experiments.

Data concerning reproduction, survival and dispersal of the whirligig water beetle Gyrinus marinus Gyll . were collected between 1974 and 1983 by observations and experiments in the laboratory and in a field area with about 10 populations distributed over 15 pools.The purpose of the study was to assess the importance of the dispersal activities for the population dynamics of the species.The main results can be summarised as follows: In a year two new generations emerge, one in summer and another in autumn (which subsequently hibernates).The mean survival chance is rather low as is the variation in survival chance. (Variation in survival during hibernation is, however. high.)Although there is much variation in egg production between females, the mean egg production per population varies little between different populations or years. The variation in survival during larval and pupal development is greater than that of egg production. Thus variation in egg production can have only a minor influence on the variation in numbers of emerging beetles.Exchange between populations by flying is limited to a few per cent of the specimens. It only takes place during favourable weather conditions.Intensive exchange of individuals occurs by swimming between (sub)populations that are connected by water. Exchange by swimming seems to be a result of every day swimming actvity.Key factor analyses indicate that survival during hibernation is the most important factor in determining fluctuation in population size. Larval development is a distant second. According to these analyses, all other processes, activities and developmental stages seem of minor importance for fluctuations in population size. This means that changes in the numbers of beetles is mainly dependent on factors to which the beetles cannot react directly. However, simulation experiments with stochastic models show that stability in population size is mainly achieved by the exchange of Individuals between populations Even the low rate of exchange by flight appears significantly to decrease the chance of extinction.The importance of exchange for the survival chance of such populations has implications for the exploitations (e.g. fishing) and control (e.g. pests) of natural populations, and the preservation of natural habitats.

Bij de ontwikkeling van een sedimenteermethode voor de bepaling van het botgehalte in separatorvlees wordt gebruik gemaakt van een vloeistof met een hoge relatieve dichtheid. Bij dit onderzoek wordt nagegaan welke vloeistof hiervoor het meest in aanmerking komt.

Data concerning reproduction, survival and dispersal of the whirligig water beetle Gyrinus marinus Gyll . were collected between 1974 and 1983 by observations and experiments in the laboratory and in a field area with about 10 populations distributed over 15 pools.The purpose of the study was to assess the importance of the dispersal activities for the population dynamics of the species.The main results can be summarised as follows: In a year two new generations emerge, one in summer and another in autumn (which subsequently hibernates).The mean survival chance is rather low as is the variation in survival chance. (Variation in survival during hibernation is, however. high.)Although there is much variation in egg production between females, the mean egg production per population varies little between different populations or years. The variation in survival during larval and pupal development is greater than that of egg production. Thus variation in egg production can have only a minor influence on the variation in numbers of emerging beetles.Exchange between populations by flying is limited to a few per cent of the specimens. It only takes place during favourable weather conditions.Intensive exchange of individuals occurs by swimming between (sub)populations that are connected by water. Exchange by swimming seems to be a result of every day swimming actvity.Key factor analyses indicate that survival during hibernation is the most important factor in determining fluctuation in population size. Larval development is a distant second. According to these analyses, all other processes, activities and developmental stages seem of minor importance for fluctuations in population size. This means that changes in the numbers of beetles is mainly dependent on factors to which the beetles cannot react directly. However, simulation experiments with stochastic models show that stability in population size is mainly achieved by the exchange of Individuals between populations Even the low rate of exchange by flight appears significantly to decrease the chance of extinction.The importance of exchange for the survival chance of such populations has implications for the exploitations (e.g. fishing) and control (e.g. pests) of natural populations, and the preservation of natural habitats.

Since the foundation of colloid chemistry as a branch of science, much attention has been paid to the subject of colloid stability, i.e. the stability of colloid systems against aggregation. Gradually, our knowledge of the mechanisms involved has improved and models were developed, comprised in the DUO theory, which form the basis of a quantitative description of the stability of a colloidal system. There is plenty of experimental evidence which substantiate the correctness of the principles of the DUO theory, and hence, this theory is regarded as one of the fundaments of colloid chemistry. However, in one respect the theory is not confirmed by experiments: calculations predict pronounced size effects, but in practice stability seems to be little affected by particle size.It was the purpose of this study to gain insight in this contradictory matter. Chapter 1 offers a more extended introduction to the problem as well as the outline of this study.Chapter 2 focusses attention on a model system which meets the outlined requirements regarding surface charge, homodispersity, sphericity and particle size range: a method is described for the synthesis of homodisperse haematite (α-Fe 2 O 3 ) sols containing particles whose sizes vary from 35 nm up to 700 nm. This method is principally based on the gradual growth of haematite seeds in supersaturated FeCl 3 solutions (heterogeneous nucleation) up to the desired paticle size. As slight deviations in the composition of the growth medium have drastic effects on the shape of the final colloid, emphasize is given to the description of optimal synthesis conditions for spherically shaped particles. Kinetic experiments, performed to unravel the principles of particle growth revealed that the precipitation process is governed by diffusion.The coagulation experiments described in chapter 5 are monitored by turbidity measurements. Therefore the characterisation of the optical properties of the sols is a prerequisite for further studies. The sols under investigation are homodiperse and contain spherically shaped particles. This makes them particularly suited for such an evaluation, as their scattering behaviour can be interpreted in terms of the Mie theory. Such a comparsion, leading to values for the refractive index (n) and the absorption coefficient (K) in the wavelength range from 400 to 800 nm is made in chapter 3 . The agreement between calculations and experiments is good for any wave length in the visible range, and for any particle size studied, although the particles are monocrystalline and no perfect spheres.Any study dealing with electrostatic stabilisation demands some knowledge of the electrochemical behaviour of the system under study. Chapter 4 pays attention to the electrochemical characterisation of the haematite surface by comparing haematite samples from different origines. In these studies, potentiometric titrations, streaming potential measurements on haematite-coated capillaries and micro electrophoresis were used as the experimental tools to get access to the surface properties. Instead of providing unambiguous data, being valid for all iron oxides occuring in the (α-Fe 2 O 3 ) modification, the reported experiments emphasize that the crystal structure of the bulk phase is not the exclusive parameter in determining the electrochemical behaviour of an oxide. The purification procedure, or a heat treatment of the sol (aqueous or dried) plays an important part as well. There is some evidence that the crystal habit of the haematite surface is pH-dependent and that in some cases precipitated amorphous oxide may share in determining the surface properties. Though not going too much into details, the study gives evidence of the fact that the charging mechanisms of haematite are more complicated than expected on the grounds of purely crystallographic considerations.Inevitably this finding has its impact on the question of colloid stability, which is the dominating item of chapter 5 . Considering the outcome of the electrochemical study, is it still justified to assume that the haematite particles meet the demands of the stability theory regarding the sharpness of the boundary between bulk material and surrounding liquid? How could a diffuse surface layer be accounted for in existing stability models? Within the restrictions set by such questions, chapter 5 deals with the item of colloid stability with special reference to the influence of particle size on stability: though the value of the critical coagulation concentration depends on particle size and shows a minimum (!), the slopes of the log W - log C plots are virtually size independent. Such trends can be accounted for by the concepts of the DUO theory, if coagulation reversibility and shear effects are incorporated in the analysis. Deviations from sphericity, which are definitely observed for the systems under investigation. might explain some of the observed effects as is shown by some simple double layer calculations dealing with orientational effects in the interaction of a cubic particle with a half space.Finally, chapter 6 reflects on the preceding Items and pays attention to its limitations. Furthermore, it points to subjects which deserve further elaboration and mentions the means to make them experimentally accessible.

This thesis is concerned with the purification and properties of hydrogenase from the obligate anaerobic rumen bacterium Megasphaera elsdenii. In chapter 1 the motives underlying this thesis, the physiological role of hydrogenase in some heterotrophs, including Megasphaera elsdenii, as well as a comparison of physico-chemical and kinetic properties of purified hydrogenase preparations from microorganisms, showing great variations in physiology and taxonomy, are given. The physiological role and the physico-chemical and kinetic properties of the hydrogenases from Megasphaera elsdenii and Clostridium pasteurianum show great similarities.In chapter 2 the procedure for the anaerobic purification of hydrogenase from Megasphaera elsdenii is given. Two activity bands are separated on DEAE-cellulose chromatography, of which one (fraction I; see Table 1 of this chapter) represents a different hydrogenase or a complex of the enzyme in fraction II. Kinetics with the purified enzyme of the hydrogen production activity at pH 8, with the electron donors flavodoxin hydroquinone, reduced ferredoxin and methyl viologen semiquinone show non-linear double reciprocal plots of the activity versus the electron donor concentration. Two kinetic models were developed, with an identical general rate equation for the hydrogen production activity, which describe a random mechanism for the reaction of the oxidized enzyme with a proton and a reduced electron donor. These models also indicate that the hydrogenase accepts the two electrons necessary for hydrogen production in two separate, independent steps. Hydrogen oxidation with methyl and benzyl viologen shows, in contrast to hydrogen production, Michaelis Menten kinetics. In chapter 3 effectors of the hydrogenase activity, such as salts (which have hydrophylic properties) Me 2 SO and ethylene glycol (which have hydrophobic properties), and oxidants (oxygen, ferricyanide, Cl 2 Ind, (bi)sulphite) are described. The data show that the more chaotropic the anion of the salt the greater the increase in activity, per mole salt used. Both Me 2 SO and ethylene glycol inhibit the hydrogen evolution activity, however, Me 2 SO stimulates the hydrogen oxidation activity, while ethylene glycol does not affect this activity. Careful oxidation of the reduced enzyme with (bi)sulphite or Cl 2 Ind irreversibly increases the activity of the enzyme by about 60%; in contrast, oxygen and ferricyanide inactivate the enzyme. This irriversibly oxidized enzyme, which shows identical kinetic properties to the reduced enzyme, is more resistant to the effects of oxygen, Me 2 SO and storage, than the reduced enzyme. Difficulties encountered in ascribing redox potentials to the observed EPR spectra of hydrogenase at several redox states, such as the effects of temperature on the Nernst equation, on the apparent pH and on the midpoint potentials of the redox species, are described.In chapter 4 the theoretical aspects are described to determine the hydrogen production activity spectrophotometrically in a series coupled redox reactions as function of the pH and redox potential, as well as the limitations of this method. The hydrogen production activity is determined from the decrease in concentration of the reduced electron donor, or increase in the concentration of the oxidized electron donor. The changes in concentration of the reduced or oxidized electron donor do not equal the amount of hydrogen produced but only represent a measure of the actual amount of hydrogen produced.In chapter 5 the method as described in chapter 4 is used to study the effects of pH and redox potential on the hydrogen production activity. The hydrogen production activity is strongly pH- and redox potential-dependent; the activity patterns observed as function of pH and redox potential are independent of the nature of the electron donor used and thus represent a property of the enzyme. If at a given pH the activities are considered to express the degree of reduction of the enzyme the dependence of the activity on the redox potential represents a n=2 type redox titration curve with an 'apparent midpoint potential, which corresponds with the potential of the hydrogen electrode at that pH. To explain the effects of redox potential, proton and electron donor concentration on the hydrogenase activity Model I of chapter 2 was slightly adapted without changing its general rate equation for the hydrogen production activity. This model also explains the 'loss' of electrons of the reduced enzyme in EPR spectroscopy as well as the formation of HD together with DD during H 2 -D 2 O exchange experiments.