Characterization of Tn904 insertions in octopine Ti plasmid mutants of Agrobacterium tumefaciens.

Seven Tn904 insertion mutants of pTi Ach5 affecting Agrobacterium tumefaciens virulence were studied. The mutant character was shown to be plasmid borne. Four of these mutants were avirulent and carried an insertion in restriction endonuclease HpaI fragment 12, a 3.3-megadalton fragment, which therefore appears to be a Ti plasmid region essential for virulence. Two mutants were attenuated in virulence. The inserts mapped close to HpaI fragment 12. One mutant giving rise to small tumors with excessive adventitious root formation on Kalanchoe daigremontiana carried an insertion in the right side of the common sequence in the deoxyribonucleic acid of the Ti plasmid detected in crown gall tumors. The insertion behavior of Tn904 was studied by analyzing 11 independently isolated and randomly chosen mutants. The Tn904 inserts did not affect oncogenicity, tumor morphology, bacterial transfer functions, octopine catabolism functions, or vital parts of the Ti plasmid, such as the origin of replication. Most of the Tn904 inserts were concentrated in a small part of the map. The size of additional deoxyribonucleic acid as a result of Tn904 inserts varied between 5 and 15 megadaltons. In two cases a Ti plasmid was found with two Tn904 insertions at different positions.     

The structure and function of fen lakes in relation to water table management in The Netherlands

Most relatively large (> 250 ha) and shallow (< 2 m), Dutch lakes originate from large-scale peat mining. Originally, their water tables were regulated by natural processes such as evapotransporation, precipitation, drainage and seepage (up and downwards). These lakes used to be oligo to mesotrophic, and the phytoplankton consisted mainly of diatoms and green algae. However, 30 years ago filamentous cyanobacteria were also present. Owing to the intensification of agriculture in the surroundings of the lakes, their management developed more and more towards a man-made reservoir function. This management resulted in inflow of eutrophic water from the agricultural areas in winter. In summer the lakes were increasingly used as a water supply. This supply is compensated by water imported mainly from the River Rhine. The impacts of this reservoir function on the limnology of the Dutch fen lakes are discussed with the Tjeukemeer, the Loosdrechtse Plassen and the Reeuwijkse Plassen as examples. All these lakes have become typical eutrophic waters with high turbidity, dominance of filamentous cyanobacteria and bream and complete loss of submerged macrophytes. Finally, methods and results of manipulation procedures to restore these waters are discussed.     

Isolation of a recombination deficient Agrobacterium tumefaciens mutant.

Summary The isolation of a recombination deficient (Rec^-) strain of Agrobacterium tumefaciens is described. Strain LBA 4011 was mutagenized with nitrosoguanidine and after segregation 18,000 colonies were replica plated and UV irradiated. Twentytwo UV sensitive strains were isolated and tested for methylmethanesulphonate (MMS) sensitivity. Six of these strains were more MMS-sensitive than LBA 4011. A Ti plasmid that was genetically marked with Tn 1 (Cb^R) was introduced in these strains and the rescue of the Cb^R marker during superinfection with an incompatible cointegrate plasmid Ti::R 702 was determined. One strain exhibited a large reduction in rescue frequency. It is concluded that the latter strain was recombination deficient. This property did not influence the induction of plant tumours.This paper forms part of a Ph.D. Thesis submitted at Leiden University by the first author     

Spatial ecology of host–parasitoid interactions: a threatened butterfly and its specialised parasitoid

Habitat conservation for threatened temperate insect species is often guided by one of two paradigms: a metapopulation approach focusing on patch area, isolation and number; or a habitat approach focusing on maintaining high quality habitat for the focal species. Recent research has identified the additive and interacting importance of both approaches for maintaining populations of threatened butterflies. For specialised host-parasitoid interactions, understanding the consequences of habitat characteristics for the interacting species is important, because (1) specialised parasitoids are particularly vulnerable to the consequences of fragmentation, and (2) altered interaction frequencies resulting from changes to habitat management or the spatial configuration of habitat are likely to have consequences for host dynamics. The spatial ecology of Cotesia bignellii, a specialist parasitoid of the threatened butterfly Euphydryas aurinia, was investigated at two spatial scales: within habitat patches (at the scale of individual aggregations of larvae, or ‘webs’) and among habitat patches (the scale of local populations). Parasitism rates were investigated in relation to larval web size, vegetation sward height and host density. Within patches, the probability of a larval webs being parasitized increased significantly with increasing number of larvae in the web, and parasitism rates increased significantly with increasing web isolation. The proportion of webs parasitized was significantly and negatively correlated with cluster density. Among habitat patches the proportion of parasitized webs decreased as cluster density increased. Clusters with a high proportion of larval webs parasitized tended to have lower parasitism rates per larval web. These results support the call for relatively large and continuous habitat patches to maintain stable parasitoid and host populations. Conservation efforts directed towards maintenance of high host plant density could allow E. aurinia to reduce parasitism risk, while providing C. bignellii with sufficient larval webs to allow population persistence.     

NO removal in continuous BioDeNOx reactors: Fe(II)EDTA2- regeneration, biomass growth, and EDTA degradation

BioDeNOx is a novel technique for NOx removal from industrial flue gases. In principle, BioDeNOx is based on NO absorption into an aqueous Fe(II)EDTA2- solution combined with biological regeneration of that scrubber liquor in a bioreactor. The technical and economical feasibility of the BioDeNOx concept is strongly determined by high rate biological regeneration of the aqueous Fe(II)EDTA2- scrubber liquor and by EDTA degradation. This investigation deals with the Fe(II)EDTA2- regeneration capacity and EDTA degradation in a lab-scale BioDeNOx reactor (10-20 mM Fe(II)EDTA2-, pH 7.2 +/- 0.2, 55 degrees C), treating an artificial flue gas (1.5 m3/h) containing 60-155 ppm NO and 3.5-3.9% O2. The results obtained show a contradiction between the optimal redox state of the aqueous FeEDTA solution for NO absorption and the biological regeneration. A low redox potential (below -150 mV vs. Ag/AgCl) is needed to obtain a maximal NO removal efficiency from the gas phase via Fe(II)EDTA2- absorption. Fe(III)EDTA- reduction was found to be too slow to keep all FeEDTA in the reduced state. Stimulation of Fe(III)EDTA- reduction via periodical sulfide additions (2 mM spikes twice a week for the conditions applied in this study) was found to be necessary to regenerate the Fe(II)EDTA2- scrubber liquor and to achieve stable operation at redox potentials below -150 mV (pH 7.2 +/- 0.2). However, redox potentials of below -200 mV should be avoided since sulfide accumulation is unwanted because it is toxic for NO reduction. Very low values for biomass growth rate and yield, respectively, 0.043/d and 0.009 mg protein per mg ethanol, were observed. This might be due to substrate limitations, that is the electron acceptors NO and presumably polysulfide, or to physiological stress conditions induced by the EDTA rich medium or by radicals formed in the scrubber upon the oxidation of Fe(II)EDTA2- by oxygen present in the flue gas. Radicals possibly also induce EDTA degradation, which occurs at a substantial rate: 2.1 (+/-0.1) mM/d under the conditions investigated.     

Effect of forest stand type on host plant quality and direct and indirect effects on pine sawfly performance

1. Observed lower levels of herbivory in mixed compared with monoculture stands have been hypothesized to depend on top-down forces, through higher predation pressure by natural enemies or through bottom-up mechanisms through plant quality effects on herbivore performance.
2. In this study, we compared the performance measured as host plant induced mortality, cocoon weight, and predation mortality of the European pine sawfly Neodiprion sertifer (Geoffroy) (Hymenoptera, Diprionidae) in mixed and monoculture forest stands.
3. We did not observe a difference in host plant induced mortality, cocoon weight, or predation mortality between mixed and monoculture forest stands. We did find an effect of local conditions around each experimental tree on pine sawfly performance. For example, the nitrogen content of pine needles is negatively affected by the proportion of pine around the experimental tree, which in turn increases the survival of sawfly larvae.
4. The results suggest that local conditions around individual trees are more important for the performance of the European pine sawfly than stand type, i.e. mixed or monoculture plant stands.
5. We conclude that the ongoing trend for diversification within commercial forestry calls for more research where the effects of both bottom-up and top-down effects are studied at several spatial scales.

     

Comparision of historical and recent data on hydrochemistry and phytoplankton in the Rijnland area (The Netherlands)

In two canals and two lakes in the western part of the Netherlands a comparision is made between data on water chemistry and phytoplankton from 1941/1942 with recent data. Orthophosphate in particular, but also inorganic nitrogen, has increased tremendously, especially in the Gouwe canal, where Rhinewater enters the area. The inorganic N/P mass ratio decreased in the last 45 years, indicating that the limiting nutrient has changed from phosphate in 1941 to nitrogen in 1987.The average seston volume, measured by filtering 1001 water through a plankton net (50 µm), has doubled. In the early 1940s the blue-green alga Microcystis aeruginosa regularly formed waterblooms, as it does now. The plankton composition seems to have become impoverished in the last decennia, since several taxa have disappeared. Others are strongly reduced in number. The saprobic index has not changed.Based on the relations between chlorophyll-a and biological oxygen demand (BOD) and between transparency and seston volume, BOD, dry weight and ash free dry weight in 1987 chlorophyll-a and transparency in 1941 are estimated. The average chlorophyll-a concentration in the lakes has doubled or tripled in the last 45 years and the mean transparency in the Gouwe canal declined from 75 to 50 cm. Submerged higher plants at some sites have disappeared in the last decennia. The results can be used to develop ecological objectives for combatting eutrophication in canals and lakes and possibly in the river Rhine.