Stephan Declerck

Effects of Glomus intraradices on the reproduction of the burrowing nematode (Radopholus similis) in dixenic culture

Abstract  The effects of Glomus intraradices on the reproduction of the burrowing nematode Radopholus similis were studied under dixenic culture conditions. The life cycles of both the arbuscular mycorrhizal fungus (AMF) and the nematode were completed in presence of each other and a transformed carrot root as host. The AMF suppressed the R. similis population by almost 50% and thus increased protection of the root against the nematode. This reduction was significant for both females and males within roots. There was no correlation between nematode population density and either AMF internal root colonization, external hyphal development or spore production. These results demonstrate that the dixenic system, although artificial, is a valuable tool for studying AMF–nematode interactions, complementing the classical experimental approaches.

Vitality and genetic fidelity of white-rot fungi mycelia following different methods of preservation.

Basidiomycetes present specific problems with regard to their preservation, because most of them do not form resistant propagules in culture but exist only as mycelium. Usually these fungi can only be preserved by serial transfer on agar (labour-intensive procedures that can increase the danger of variation or loss of physiological or morphological features), or cryopreserved in liquid nitrogen (expensive). Cryopreservation at -80 degrees C and lyophilisation could be good alternatives. In this work we set up and tested six protocols of cryopreservation at -80 degrees C, and 12 protocols of lyophilisation on 15 isolates of white-rot fungi (WRF) belonging to 10 species. The tested protocols were mainly characterized by the use of different growth media, protectants, time and number of perfusion with protectants and finally by the typology and origin of the samples to be cryopreserved (mycelium/agar plug, whole colony) or to lyophilise (mycelium/agar plug, mycelium fragment, whole colony). Cryopreservation and lyophilisation outcomes were checked, at morphological (macro- and microscopic features), physiological (growth rate and laccase, Mn-independent and Mn-dependent peroxidases activities) and genetic level (Amplified Fragment Length Polymorphisms analysis - AFLP). Vitality of all fungi was successfully preserved by all cryopreservation protocols at -80 degrees C, and by two lyophilisation methods. Our results showed that cryopreservation at -80 degrees C did not produce morphological changes in any isolate, while two isolates were affected by lyophilisation. None of the physiological features were lost, even though growth rate and enzyme activities were somehow influenced by all preservation methods. AFLP analysis showed that only the two isolates that varied in their morphology after lyophilisation produced a different DNA fingerprint pattern in comparison with that obtained before lyophilisation. These findings provide evidence that cryopreservation at -80 degrees C and lyophilisation are suitable alternatives to liquid nitrogen cryopreservation for preservation of some WRF strains.

Effects of two sterol biosynthesis inhibitor fungicides (fenpropimorph and fenhexamid) on the development of an arbuscular mycorrhizal fungus.

The effects of different concentrations (0.2, 2, 20, 200mgl(-1)) of two sterol biosynthesis inhibitor (SBI) fungicides, i.e. fenpropimorph and fenhexamid, were evaluated on the spore germination, germ tube elongation, sporulation, and root colonization of Glomus intraradices grown monoxenically in association with transformed carrot roots. The percentage of germinated spores incubated on the SBI fungicides and the length of the germ tubes decreased with increasing concentrations of both fungicides. However, for spore germination this impact was fungistatic rather than fungicidal. Extraradical mycelium architecture and spore production in contact with the SBI fungicides were also strongly impacted at high concentration (20mgl(-1)). Conversely, the colonization of roots developing in the fungicide-free compartment, but interconnected with the extraradical mycelium developing on the SBI fungicides, appeared unaffected. Our results demonstrated that the monoxenic culture system could be used as a standardized, reproducible technique to compare the impacts of different molecules on arbuscular mycorrhizal fungi, and for the initial screening of new candidate molecules before registration.

Characterization of an endophytic whorl-forming Streptomyces from Catharanthus roseus stems producing polyene macrolide antibiotic

An endophytic whorl-forming Streptomyces sp. designated as TS3RO having antifungal activity against a large number of fungal pathogens, including Sclerotinia sclerotiorum, Rhizoctonia solani, Colletotrichum gloeosporioides, Cryphonectria parasitica, Fusarium oxysporum, Pyrenophora tritici-repentis, Epidermophyton floccosum, and Trichophyton rubrum, was isolated from surface-sterilized Catharanthus roseus stems. Preliminary identification showed that Streptomyces cinnamoneus subsp. sparsus was its closest related species. However, strain TS3RO could readily be distinguished from this species using a combination of phenotypic properties, 16S rDNA sequence similarity, and phylogenetic analyses. Thus, the whorl-forming Streptomyces sp. strain TS3RO is likely a new subspecies within the Streptomyces cinnamoneus group. Direct bioautography on a thin-layer chromatography plate with Cladosporium cucumerinum was conducted throughout the purification steps for bioassay-guided isolation of the active antifungal compounds from the crude extract. Structural elucidation of the isolated bioactive compound was obtained via LC-MS spectrometry, UV-visible spectra, and nuclear magnetic resonance data. It revealed that fungichromin, a known methylpentaene macrolide antibiotic, was the main antifungal component of TS3RO strain, as shown by thin-layer chromatography bioautography. This is the first report of an endophytic whorl-forming Streptomyces isolated from the medically important plant Catharanthus roseus.

Uptake and translocation of uranium by arbuscular mycorrhizal fungi under monoxenic culture conditions

ABSTRACT Fundamental information on the transport of uranium by mycelium of arbuscular mycorrhizal (AM) fungi is reported and discussed in this chapter. The monoxenic cultivation system was shown to be convenient to investigate the processes of uptake and translocation of U by AM fungi, as it was already reported for some other elements. The evidence that extraradical AM hyphae can take up U and translocate it towards the host roots was clearly shown. Further research to understand the mechanisms involved and future prospects were also suggested.

Report on Workshop on “Biological Control in Potato Production

Thirty-three scientists from 22 countries, mostly not only from Europe (Austria, Belgium, France, Finland, Germany, Ireland, The Netherlands, Slovenia, Switzerland, the UK) but also from Peru and Kenya, actively participated as experts in the EAPR workshop on biological control in potato production. Each participant rated both his own expertise in potato and in biological control, on a scale from 1 (very low) to 5 (very high). The mean rate of expertise of the panel was 3.60 (medium to high) for potato expertise and 2.89 (medium) for biological control. Additionally, the experts were asked to (i) identify the major research areas in biological control, (ii) choose major targets to focus on plant pathogenic fungi, bacteria, and viruses as well as nematode and insect pests, (iii) specify the major gaps and needs in potato biological control, and finally (iv) provide indications on possible technological locks for innovation in biological control.