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The plant cell wall is a chemically complex network composed mainly of polysaccharides. Cell wall polysaccharides surround and protect plant cells and are responsible for the stability and rigidity of plant tissue. Pectin is a major component of primary cell wall and the middle lamella of plants. However, pectin biosynthesis in planta and the mechanisms underlying the influence of structural differences arising from a modified biosynthesis machinery on functional properties remain poorly understood. In our research, the changes in the chemical structures of cell wall polysaccharides after transgenic modification of potato tuber polysaccharides were examined. The cell wall material from potato wild-type varieties, from known and from new potato transgenic lines targeting changes of the homogalacturonan or rhamnogalacturonan I backbone were isolated and characterized. The modified cell wall polysaccharides were examined by determining their individual monosaccharide levels on fresh weight base and their cell wall characteristic parameters, and levels of acetylation and methyl esterification of cell wall pectin. Data for both targeted and non-targeted structures of cell wall polysaccharides from wild-type and transgenic potatoes were obtained. A shorter galactan side chain was found from the buffer soluble pectin and calcium bound pectin of β-galactosidase (β-Gal) transgenic lines. All pectin fractions from rhamnogalacturonan lyase (RGL) transgenic lines had less galactan chains attached to their rhamnogalacturonan I backbones. Two uridine diphosphate-glucose 4-epimerase (UGE) transgenic lines, UGE 45 and UGE 51, had diverse effects on length of the galactan side chain. The xyloglucans from the RGL and UGE transgenic lines retained its XXGG building blocks but differed in the proportion of repeating units compared to the respective wild-type varieties. In contrast, the β-Gal transgenic lines predominantly consisted of XXXG-type xyloglucan in the 4 M alkali extract, but showed XXGG-type building blocks in 1 M alkali extract. In addition, a quick-screening method was validated and used to analyze 31 transgenic lines and their respective wild-type potato varieties. An overall comparison of pectin backbone, pectin side chains, acetylation and methyl-esterification of pectin, pectin content and (hemi)cellulose content of cell wall polysaccharides from these transgenic lines provided a better insight in the frequency, level and combination of both targeted and non-targeted structural changes compared to that of their respective wild-type varieties. The same evaluation method was used to correlate cell wall composition in wild-type and selected transgenic lines and their established gene expression with the texture of corresponding cooked potato cubes. Changed physical properties for the genetically modified tubers could be connected to specific cell wall characteristics. Tubers from transgenic lines containing cell wall pectin with short galactan side chains were less firm after cold processing compared to wild-type tubers. The enhanced understanding of transgenic modifications of potato tubers resulting into significant targeted and non-targeted modifications in cell wall polysaccharides will lead to a better selection of potato lines with tailored cell wall characteristics and desired properties of the tubers during processing. Potato cell walls are composed of pectin, hemicellulose and cellulose. Cell wall polysaccharides are responsible for the stability, rigidity and flexibility of plant tissue. Pectin, a major component of primary plant cell walls, primarily consists of homogalacturonan (HG) and rhamnogalacturonan I (RG-I). To understand the structure–function relationships of potato cell wall pectin, this study aimed to identify the characteristics of both pectin and other polysaccharides as present in cell wall material (CWM) and of individual polysaccharide populations from wild-type potato varieties and their respective transgenic potato lines. Chapter 1 gives a general introduction to the fine chemical structures of potato cell wall polysaccharides, the main models of cell wall architecture and the cell wall-degrading enzymes, which include pectinases, hemicellulases and cellulases. In addition, transgenic modification of the cell wall through the heterologous expression of various enzymes from fungal or plant origin that could modify potato cell wall polysaccharides in planta is addressed. Transgenic modifications of potato cell wall polysaccharides that targeted pectin structures and cellulose levels are summarised. However, due to unsuccessful starch removal during CWM isolation and incomplete analysis of CWM yield and composition, characteristics regarding the different cell wall polysaccharides from previously-studied transgenic potato lines are hardly available. CWMs were extracted from the Karnico (wild-type) potato and its transgenic lines that expressed either β-galactosidase or rhamnogalacturonan lyase (Chapter 2). Improved starch removal procedures proved to be successful. Pectic polysaccharides were fractionated from CWMs of wild-type potato and its transgenic lines β-Gal-14 and RGL-18. Most β-Gal-14 pectin populations had less galactose (Gal) than wild-type, indicating that the transgenic line had shorter galactan side chains, although the side chain length differed for individual pectin populations. The ratio of HG:RG-I was introduced to evaluate the pectin backbone structure. High HG:RG-I ratios were consistently found in RGL-18 pectic polysaccharide populations. A low level of RG-I segments in combination with lower Gal contents indicated the removal of the galactan-rich RG-I segments in all pectin populations of RGL transgenic lines. In addition, RGL-18 transgenic modification increased the methyl-esterification and lowered the acetylation of pectins present in hot buffer extracts, when compared to wild-type. No effect on pectin esterification was found for β-Gal transgenic lines. Side effects of the mutation generated unexpected changes in the various pectin populations. The xyloglucan structure was extensively modified after transgenic modification of the pectin structure. Two xyloglucan extracts were obtained from the Karnico and its β-Gal-14 and RGL-18 transgenic lines (Chapter 3). The extracts of the Karnico and RGL-18 lines were mainly comprised of the XXGG-type xyloglucan as represented by XXGG and XSGG as predominant repeating units. In contrast, the XXXG-type xyloglucan was primarily present in the β-Gal-14 4 M alkali extract built up by LLUG repeats, although XXGG type of xyloglucan was present in the 1M alkali extract. Both the RGL and β-Gal transgenic lines had different proportions of xyloglucan building blocks (XSGG/XXGG ratios) than wild-type. After transgenic modification of pectin backbone or pectin side chains, the xyloglucan structures has been biosynthetically modified by plant itself. Uridine diphosphate (UDP)-glucose 4-epimerase (UGE) catalyses the conversion of UDP-glucose into UDP-galactose, which hypothetically should lead to more galactose being built into the cell wall polysaccharides. CWMs from the Kardal (wild-type) potato and its UGE45-1 and UGE51-16 transgenic lines were isolated, fractionated and characterised (Chapter 4). Both the UGE45 and UGE51 genes encoded for UGE enzymes, but the corresponding transgenic lines exhibited different modifications of the galactan side chains and of other cell wall structures. The Gal content of CWM from the UGE45-1 transgenic line was 38% higher than that of the wild-type and resulted in longer pectin side chains. The Gal content present in CWM from UGE51-16 was 17% lower than that of wild-type, which resulted in a slightly shorter galactan side chains for most pectin populations. Both UGE transgenic lines showed a decreased acetylation and an increased methyl-esterification of the cell wall pectin. Side effects were found in the xyloglucan structures of the transgenes as reflected by different proportions of XSGG/XXGG repeating units in comparison to wild-type. Pectin side chain biosynthesis had not only a varying level of galactan side chain modification, but also influenced the structure and possibly the interaction of other cell wall polysaccharides. In Chapter 5, a new screening strategy is introduced to evaluate higher numbers of transgenic potato tubers via CWM yield and sugar composition. A total of four wild-type potato varieties and 31 transgenic lines were evaluated to determine the effects on targeted structures including RG-I or HG pectin backbone elements, galactan or arabinogalactan side chains, acetyl groups of pectin and cellulose levels. Modification of the pectin backbone or pectin side chains in the transgenic lines has either a simultaneous increase or simultaneous decrease of HG:RG-I ratio, side chain length and methyl-esterification of pectin. The pectin esterification transgenic line exhibited only limited side effects. The cellulose level targeted lines had also high HG:RG-I ratios, longer galactan chains and similar pectin content compared to the wild-type, indicative for a less branched pectin backbone with longer side chains. From the monosaccharide composition data, various pectin and cell wall characteristics parameters are suggested as powerful indicators of cell wall polysaccharide structure. In Chapter 6, the achievements of this research are summarised and discussed in the context of potato cell wall architecture. The strategy and outcome of a quick screening method for multiple transgenic lines and an in-depth analysis of individual pectin and xyloglucan populations for the evaluation of potato CWMs is discussed. Furthermore, the texture of steam-cooked potatoes and the stability of potato cubes after freeze-thaw cycles are correlated with gene expression and cell wall composition in wild-type and selected transgenetically modified potato tubers. CWMs from transgenetically modified potatoes showed different physical properties during processing. In isolated CWMs, acetylation of cell wall pectin, molar Gal levels and starch content were the main parameters that could be related to the texture or firmness of tubers. Tubers from transgenic lines that resulted in shorter pectin side chains felt apart more easily after several freeze-thaw cycles than wild-type tubers and tubers with an increased length of pectin side chains. The modification of both targeted as well as non-targeted structures have now been shown to occur in many different potato transgenic lines, but precise mechanisms and consequences for the cell wall architecture remain unclear. Research performed so far, as well as research needed for getting a better understanding of plant cell wall architecture, is discussed.

In 1990 is in Nederland een werkgroep opgericht om jaarlijks gevallen van massale bijensterfte te inventariseren die volgens getroffen imkers veroorzaakt zijn door blootstelling aan gewasbeschermingsmiddelen (Oomen, 1992). De werkgroep jubileert dit jaar en heeft daarom haar bevindingen met spuitschade in de afgelopen 25 jaar samengevat

The effects of a pesticide mixture (asulam, fluazinam, lambda-cyhalothrin, and metamitron) on aquatic ecosystems were investigated in 20 outdoor aquatic microcosms. Ten of the microcosms simulated mesotrophic aquatic ecosystems dominated by submerged macrophytes (Elodea). The others simulated eutrophic ecosystems with a high Lemna surface coverage (Lemna). This paper describes the fate of the chemicals as well as their effects on the growth of Myriophyllum spicatum and the periphytic algal community. In the Elodea-dominated microcosms significant increase in the biomass and alterations of species composition of the periphytic algae were observed, but no effect on M. spicatum growth could be recorded in response to the treatment. The opposite was found in the Lemna-dominated microcosms, in which decreased growth of M. spicatum was observed but no alterations could be found in the periphytic community. In the Elodea-dominated microcosms the species composition of the periphytic algae diverged from that of the control following treatment with 0.5% spray drift emission of the label-recommended rate (5% for lambda-cyhalothrin), while reduced growth of M. spicatum in the Lemna-dominated microcosms was recorded at 2% drift (20% for lambda-cyhalothrin). This study shows that the structure of the ecosystem influences the final effect of pesticide exposure.

Deze studie heeft als doel om de effecten van klimaatverandering op de land- en tuinbouw in kaart te brengen, in de huidige situatie en voor de (midden)lange termijn (2050). Op basis daarvan worden de risico’s en kansen voor de sector geïdentificeerd en de adaptatiemaatregelen waarmee risico’s kunnen worden opvangen en kansen worden benut. De doelstelling van deze studie is het actualiseren van kwetsbaarheden, risico’s en kansen voor de Nederlandse land- en tuinbouw gerelateerd aan klimaatverandering. De beoogde uitkomsten zijn: Een concrete aanduiding van de aard en omvang kwetsbaarheid en klimaatrisico’s, Een lijst van mogelijke indicatoren voor kwetsbaarheid.

Veldproeven zijn een belangrijk onderdeel van de milieurisicobeoordeling bij de toelatingsprocedure voor teelt van genetisch gemodificeerde gewassen , met name om eventuele nadelige effecten op zogenaamde niet-doelwitorganismen te onderkennen.

Deze studie heeft als doel om de effecten van klimaatverandering op de land- en tuinbouw in kaart te brengen, in de huidige situatie en voor de (midden)lange termijn (2050). Op basis daarvan worden de risico’s en kansen voor de sector geïdentificeerd en de adaptatiemaatregelen waarmee risico’s kunnen worden opvangen en kansen worden benut. De doelstelling van deze studie is het actualiseren van kwetsbaarheden, risico’s en kansen voor de Nederlandse land- en tuinbouw gerelateerd aan klimaatverandering. De beoogde uitkomsten zijn: Een concrete aanduiding van de aard en omvang kwetsbaarheid en klimaatrisico’s, Een lijst van mogelijke indicatoren voor kwetsbaarheid.

Veldproeven zijn een belangrijk onderdeel van de milieurisicobeoordeling bij de toelatingsprocedure voor teelt van genetisch gemodificeerde gewassen , met name om eventuele nadelige effecten op zogenaamde niet-doelwitorganismen te onderkennen.

De rol van pesticiden in een moderne bedrijfsvoering

In 1990 is in Nederland een werkgroep opgericht om jaarlijks gevallen van massale bijensterfte te inventariseren die volgens getroffen imkers veroorzaakt zijn door blootstelling aan gewasbeschermingsmiddelen (Oomen, 1992). De werkgroep jubileert dit jaar en heeft daarom haar bevindingen met spuitschade in de afgelopen 25 jaar samengevat

Because of the political, social and environmental problems associated with dependency on fossil fuels, there is considerable interest in alternative energy sources. Hydrogen is regarded as a promising option, particularly as a fuel for road vehicles. The Dutch Energy Research Centre (ECN) recently published a vision of the future, in which it suggested that by 2050 more than half of all cars in the Netherlands could be running on hydrogen. A switch to using hydrogen as the primary energy source for road vehicles would have far-reaching social consequences. As with all technological developments, opportunities would be created, but drawbacks would inevitably be encountered as well. Some of the disadvantages associated with hydrogen are already known, and are to some degree manageable. It is likely, however, that other drawbacks would come to light only once hydrogen-powered cars were actually in use. With that thought in mind, and in view of the social significance of a possible transition to hydrogen, it was decided that the Health Council should assess the positive and negative effects that hydrogen use could have on public health. It is particularly important to make such an assessment at the present early stage in the development of hydrogen technologies, so that gaps in existing scientific knowledge may be identified and appropriate strategies may be developed for addressing such gaps. This report has been produced by the Health and Environment Surveillance Committee, which has special responsibility for the identification of important correlations between environmental factors and public health.

Because of the political, social and environmental problems associated with dependency on fossil fuels, there is considerable interest in alternative energy sources. Hydrogen is regarded as a promising option, particularly as a fuel for road vehicles. The Dutch Energy Research Centre (ECN) recently published a vision of the future, in which it suggested that by 2050 more than half of all cars in the Netherlands could be running on hydrogen. A switch to using hydrogen as the primary energy source for road vehicles would have far-reaching social consequences. As with all technological developments, opportunities would be created, but drawbacks would inevitably be encountered as well. Some of the disadvantages associated with hydrogen are already known, and are to some degree manageable. It is likely, however, that other drawbacks would come to light only once hydrogen-powered cars were actually in use. With that thought in mind, and in view of the social significance of a possible transition to hydrogen, it was decided that the Health Council should assess the positive and negative effects that hydrogen use could have on public health. It is particularly important to make such an assessment at the present early stage in the development of hydrogen technologies, so that gaps in existing scientific knowledge may be identified and appropriate strategies may be developed for addressing such gaps. This report has been produced by the Health and Environment Surveillance Committee, which has special responsibility for the identification of important correlations between environmental factors and public health.

This report describes the evaluation of the 2006 proposal for the risk assessment of persistence of plant protection products in soil. The proposal considered three protection goals and proposed tiered assessment and decision schemes for each protection goal. The three schemes appeared to be consistent, both internally and with each other. It was found that both pore water concentrations and total content have to be considered in the soil risk assessment. The evaluation has been performed for five substances with all available information from both registration dossiers and open literature. Nevertheless, insufficient information was available to evaluate all aspects of the proposal. In practice this means that pesticide industry has to provide additional information for many dossiers. Furthermore, it was found that existing information often needs to be re-interpreted and a need for standardisation of evaluation of terrestrial (semi-)field experiments was observed. The proposal would require specific expertise and investments of evaluating authorities as well as stakeholders. To better understand fate and effects of persistent substances, it is recommended to investigate the behaviour of substances in the field over longer periods, to perform exposure concentration measurements while performing ecotoxicological tests, to develop protocols for testing effects on fungi, and to gain the necessary experience on the conduct and interpretation of (semi-)field studies with respect to the relation between exposure and effects of plant protection products.

This report describes the evaluation of the 2006 proposal for the risk assessment of persistence of plant protection products in soil. The proposal considered three protection goals and proposed tiered assessment and decision schemes for each protection goal. The three schemes appeared to be consistent, both internally and with each other. It was found that both pore water concentrations and total content have to be considered in the soil risk assessment. The evaluation has been performed for five substances with all available information from both registration dossiers and open literature. Nevertheless, insufficient information was available to evaluate all aspects of the proposal. In practice this means that pesticide industry has to provide additional information for many dossiers. Furthermore, it was found that existing information often needs to be re-interpreted and a need for standardisation of evaluation of terrestrial (semi-)field experiments was observed. The proposal would require specific expertise and investments of evaluating authorities as well as stakeholders. To better understand fate and effects of persistent substances, it is recommended to investigate the behaviour of substances in the field over longer periods, to perform exposure concentration measurements while performing ecotoxicological tests, to develop protocols for testing effects on fungi, and to gain the necessary experience on the conduct and interpretation of (semi-)field studies with respect to the relation between exposure and effects of plant protection products.

In Nederland zijn zijn voor de akkerbouw en de vollegrondsgroententeelt slechts vier biologische (niet-synthetische) middelen toegestaan: Bacillus thuringiensis, pyretrine + piperonylbutoxide, koperoxychloride en zwavel. Een opsomming van de tot nu toe bekende eigenschappen (doelorganismen; toegelaten gewassen; effectiviteit; giftigheid voor niet-doelorganismen; milieueffecten)

De rol van pesticiden in een moderne bedrijfsvoering

In this thesis the exposure to pesticides of fruit growers in The Netherlands was studied as well as its relation to reproductive health effects. The most commonly used fungicide, captan, was used as a marker for exposure. Several exposure studies were carried out during application of captan, and work in the orchards. As exposure per unit of time was in the same order of magnitude for Very different tasks such as application and re-entry, individual time spent on different tasks is crucial for estimating total exposure. As day-to-day variability in exposure was high, within as well as between workers, repeated measurements are necessary to estimate individual exposure accurately. For the application, respiratory exposure was predominantly related to the preparation of tank mixtures. Cabin use on the tractor was the most prominent determinant of dermal exposure during spraying. During re-entry dislodgeable foliar residue was the most prominent determinant of exposure for both respiratory and dermal exposure. Uptake of captan into the human body was measured with biological monitoring of tetrahydrophtalimide in urine of exposed farmers after application. Urine levels were related to dermal exposure of specific skin areas. The consequences of these findings for the way a 'skin notation' is set, is discussed. Another study was carried out to evaluate methods for subjective assessment of pesticide exposure in fruit growing by experts. Experts seem to recognize the most important determinants of exposure. In an epidemiological study time to pregnancy was used to study effects of pesticide exposure. In addition, the relationship with offspring sex ratio was explored. The findings of the study on time to pregnancy indicate that an adverse effect of exposure to pesticides on fecundability is likely. As very few epidemiological studies have been carried out on the effects of pesticides on reproduction, no firm conclusions can be drawn about specific pesticides or other related factors responsible for the observed effect on time to pregnancy or on offspring sex ratio. The indication, however, justifies more attention for possible effects of pesticide exposure on reproduction. In this light, reduction of exposure to pesticides as far as achievable is needed. The validity of the observed effects on reproduction and the underlying mechanisms have to be elucidated.

In this thesis the exposure to pesticides of fruit growers in The Netherlands was studied as well as its relation to reproductive health effects. The most commonly used fungicide, captan, was used as a marker for exposure. Several exposure studies were carried out during application of captan, and work in the orchards. As exposure per unit of time was in the same order of magnitude for Very different tasks such as application and re-entry, individual time spent on different tasks is crucial for estimating total exposure. As day-to-day variability in exposure was high, within as well as between workers, repeated measurements are necessary to estimate individual exposure accurately. For the application, respiratory exposure was predominantly related to the preparation of tank mixtures. Cabin use on the tractor was the most prominent determinant of dermal exposure during spraying. During re-entry dislodgeable foliar residue was the most prominent determinant of exposure for both respiratory and dermal exposure. Uptake of captan into the human body was measured with biological monitoring of tetrahydrophtalimide in urine of exposed farmers after application. Urine levels were related to dermal exposure of specific skin areas. The consequences of these findings for the way a 'skin notation' is set, is discussed. Another study was carried out to evaluate methods for subjective assessment of pesticide exposure in fruit growing by experts. Experts seem to recognize the most important determinants of exposure. In an epidemiological study time to pregnancy was used to study effects of pesticide exposure. In addition, the relationship with offspring sex ratio was explored. The findings of the study on time to pregnancy indicate that an adverse effect of exposure to pesticides on fecundability is likely. As very few epidemiological studies have been carried out on the effects of pesticides on reproduction, no firm conclusions can be drawn about specific pesticides or other related factors responsible for the observed effect on time to pregnancy or on offspring sex ratio. The indication, however, justifies more attention for possible effects of pesticide exposure on reproduction. In this light, reduction of exposure to pesticides as far as achievable is needed. The validity of the observed effects on reproduction and the underlying mechanisms have to be elucidated.