Molecular characterization of a stable antisense chalcone synthase phenotype in strawberry (Fragaria x ananassa)

An octaploid (Fragaria x ananassa cv. Calypso) genotype of strawberry was transformed with an antisense chalcone synthase (CHS) gene construct using a ripening related CHS cDNA from Fragaria x ananassa cv. Elsanta under the control of the constitutive CaMV 35S promoter via Agrobacterium tumefaciens. Out of 25 transgenic lines, nine lines showed a reduction in CHS mRNA accumulation of more than 50% as compared to the untransformed cv. Calypso control. The antisense CHS construct was found to be integrated into the genome, with a copy number ranging from one to four. The pigmentation of the fruit was only affected when less than 5% of the control CHS expression level was detected. A stable antisense phenotype over a period of 4 years was obtained in the primary transgenic lines at a rate of 1:20. As a consequence of the reduced activity of CHS, the levels of anthocyanins, flavonols, and proanthocyanidins were downregulated and precursors of the flavonoid pathway were shunted to the phenylpropanoid pathway leading to highly increased levels of cinnamoyl glucose (520% of control), caffeoyl glucose (816% of control), and feruloyl glucose (1092% of control) as well as p-coumaryl alcohol (363% of control) and p-coumaryl-1-acetate (1079% of control), which occur only as trace components in untransformed control fruits. These results demonstrate that the introduction of an antisense CHS construct in strawberry results in an unpredictable biochemical phenotype, thereby confirming that CHS function is an important regulatory point of substrate flow between the flavonoid and the phenylpropanoid pathways.     

Native Grass Facilitates Mycorrhizal Colonisation and P Uptake of Tree Seedlings in Two Anthropogenic Substrates

Two microcosm experiments were conducted to study the role of extraradical mycelium (ERM) of arbuscular mycorrhizal fungi (AMF) in establishment and growth of tree species used for revegetation in anthropogenic substrates. Inoculated or non-inoculated Acer pseudoplatanus, Alnus glutinosa or Salix purpurea seedlings were grown with Calamagrostis epigejos (a grass spontaneously colonising degraded ecosystems) in two substrates (fly ash and coal mine spoil) either in direct root contact or in rhizoboxes with interaction only via ERM network. In both experiments, inoculation with AMF mostly had a positive effect on the growth of trees and increased the aggregation of fly ash. When plants grew in direct root-to-root contact, grass presence negatively affected tree growth, but it significantly improved mycorrhizal development (colonization of tree seedlings, spore number and ERM length). When grass and tree roots interacted via the ERM network, tree seedlings were successfully colonised by the ERM spreading from the C. epigejos roots. Mechanical disturbance of the ERM links between plants reduced AMF development and tree height in both substrates, but tree shoot biomass was not affected. In fly ash, inoculated, non-disturbed treatments showed significant transfer of ³²P from the grass to the tree seedlings. It can be concluded that roots of A. pseudoplatanus, A. glutinosa or S. purpurea seedlings can be colonised from the ERM network radiating from quickly growing grasses, which can act as important agents for AMF distribution and facilitate mycorrhization of planted trees. In particular for willow, grass seems to be an essential nurse plant to achieve successful root colonisation.     

Interaction of arbuscular mycorrhizal fungi and rhizobia: Effects on flax yield in spoil‐bank clay

This study focused on the application of native strains of arbuscular mycorrhizal fungi (AMF) and Sinorhizobium in effective crop production during reclamation of coal‐mine spoil banks. Two greenhouse experiments were conducted in spoil‐bank clay with a low dose of organic amendment to determine whether the microbial inoculation improves growth and utility qualities of two cultivars of Linum usitatissimum L. (oil and fiber flax). Inoculation with two native AMF isolates (Glomus mosseae, G. intraradices, and their mixture) significantly increased growth and shoot phosphorus (P) concentration of both flax cultivars. Inoculated fiber flax plants produced fivefold more fibers than the uninoculated ones. In oil flax, mycorrhizal inoculation significantly but quantitatively to a minor degree decreased the concentration of nonsaturated fatty acids in the seed oil. A mixture of five native Sinorhizobium sp. strains supported growth and P uptake of oil flax only in the absence of AMF. However, these beneficial effects of the bacteria were significantly lower as compared to AMF. No synergic action of Sinorhizobium strains and AMF was observed, and their interactions were often even antagonistic. Inoculation with AMF significantly decreased population density of Sinorhizobium in the soil. These results suggest that a careful selection of suitable bacterial strains is necessary to provide effective AMF combinations and maximize flax‐growth support.     

Immunoblotting, ELISA and culture evidence for Chlamydiaceae in sows on 258 Belgian farms

The prevalence of Chlamydiaceae infections on 258 closed pig breeding farms in Belgium was examined. For this purpose, 258 farms were randomly selected in the provinces West-Vlaanderen (44%), Oost-Vlaanderen (20%), Antwerpen (10%) and Vlaams-Brabant (6%). Of all farms examined, 96.5% were positive for Chlamydia-specific antibodies in ELISA and most were moderately to strongly positive. ELISA results revealed only 9 (3.5%) sero-negative farms. None of the ELISA negative sera reacted in immunoblotting. Only 212 of 249 ELISA positive sera reacted positive in immunoblotting. Additionally, 23 autopsy samples were examined by isolation in Vero cells. The major outer membrane sequence of the one isolate obtained showed 98.6% amino acid homology to the one of Chlamydophila psittaci strain CP3, formerly isolated from a pigeon. Present observations indicate that chlamydial infections are nearly endemic in the Belgian pig population and that Belgian pigs can become infected with C. psittaci. Nevertheless, the role and significance of Chlamydiaceae as pathogens in pigs remain unsolved and require further investigation.     

Declining plant species richness of grassland ditch banks—a problem of colonisation or extinction?

Small-scale landscape elements, such as ditch banks are an important remaining source of biodiversity in many agricultural landscapes, including the Western Peat District in the Netherlands. Unfortunately, plant species richness is declining even in these habitats. To understand the factors threatening biodiversity, we studied demographic traits (occupancy, trend, colonisation and extinction) for a large number of plant species, in a 25-year long data set. We developed a method to investigate the relative importance of colonisation and extinction for species increase and decrease in multi-species assemblages. We show that colonisation has been more important for determining species trends than extinction. Decreasing species were small and characterised by low nutrient tolerance and high light requirements, indicating that competitive ability influences species trends. The mechanism by which high nutrient levels reduce plant diversity appears to be closely related to colonisation (germination and seedling establishment). Local management should therefore continue to focus on nutrient reduction and the creation of regeneration sites. Yet, these measures will be insufficient for restoring species richness since isolation also hampers species increase. Therefore, to maximise the effects of local management, additional regional management solutions are required to improve seed dispersal for example, from nature reserves.     

A new method for bovine embryo production: a potential alternative to superovulation

Eight cows were used to study the feasibility of transvaginal ultrasound-guided puncture of follicles as a method for the collection of immature oocytes for embryo production in vitro. In six trials at intervals of seven days, 104 oocytes were collected. After in vitro maturation and fertilisation the 104 oocytes were transferred to the oviducts of sheep. Six days later, 75 oocytes were recovered by flushing the oviducts. Twenty-four per cent of the recovered oocytes/embryos had developed into transferable and viable morulae and, or, blastocysts. The data show that this non-surgical and repeated collection of immature oocytes can be used successfully for the in vitro production of bovine embryos. The procedure may produce yields of embryos comparable to those obtainable by conventional superovulation procedures.     

Foot-and-mouth disease and the African buffalo (Syncerus caffer). 1. Carriers as a source of infection for cattle

Ten pregnant buffalo cows, six of which were subsequently shown to be carriers of SAT 1, 2 and 3 viruses, were captured in the Kruger National Park (KNP) and allowed to calve in captivity. The buffalo cows and calves were separated by a fence from 6 FMD susceptible cattle but the buffalo and cattle were obliged to use common drinking troughs and hay racks. Over a period of 15 months, during which the buffalo calves lost their maternally-derived immunity, neither the buffalo calves nor the susceptible cattle became infected with FMD virus. By the end of the observation period, however, only 1 buffalo cow still had detectable virus in its oesophageal/pharyngeal specimens.     

Short-term interval baiting to combat the re-emergence of fox rabies in Rhineland Palatinate (Germany) in 2005

In 2005, the final phase of terrestrial rabies eradication in Germany was put at risk by a severe setback due to re-introduction of the disease in Rhineland-Palatinate from neighbouring Hesse after seven years of absence. The rapid westward spread of the disease prompted veterinary authorities to react swiftly and apply a new yet unproven vaccination strategy to rapidly increase herd immunity in an almost unprotected fox population to stop the epidemic. The cornerstones of this emergency oral rabies vaccination strategy, i. e. vaccination intervals, identification of high risk spots, real time epidemiological assessment, capable to eliminate rabies within 13 months after incursion are described here. This strategy may be used as a template to tackle similar emergency situations in Europe in the future.