The recently described oomycete pathogen Phytophthora pisi causes root rot on pea and faba bean, while the closely related Phytophthora sojae is the causal agent of soybean root and stem rot. ...Differences in the pathogenicity factor repertoires that enable the two species to have distinct host specificity towards pea and soybean, were studied using tandem mass spectrometry in a global proteome study of hyphae and germinating cysts in P. pisi and P. sojae. In total 2775 proteins from P. pisi and 2891 proteins from P. sojae were identified. Fifty-eight orthologous proteins were more abundant in germinated cysts of both pathogens and thus identified as candidate proteins for the infective stage. Several of these proteins were associated with lipid transport and metabolism, and energy production. Twenty-three orthologous proteins were more abundant in hyphae of both pathogens and thus identified as candidate proteins for vegetative growth. Proteins uniquely present in germinating cysts of either P. pisi or P. sojae were considered as candidates for species-specific pathogenicity factors that may be involved in host specificity. Among these proteins were serine proteases, membrane transporters and a berberine-like protein. These results significantly expand the knowledge of the expressed proteome in P. pisi and P. sojae.
P. sojae and P. pisi are closely related species that specifically cause root rot on soybean and pea, respectively. The pathogenicity factors contributing to their host specificity remained unknown. We carried out a comparative large-scale proteome analysis of vegetative (hyphae) and infective (germinating cysts) life stages in P. pisi and P. sojae. This study provides knowledge of the common factors and mechanism involved in initiation of infection and species-specific proteins that may contribute to the host specificity of these pathogens. This knowledge will lead to a better understanding of the infection biology of these pathogens, allowing new possibilities towards developing alternative and effective plant protection measures.
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•Stored lipid compounds are utilized during cyst germination.•Protein candidates possibly involved in early infection process were identified.•Proteins specific to infective stage of each pathogen were identified.
Phytophthora niederhauserii, P. pisi, P. sojae and P. vignae are closely related species that are pathogenic to various legume plants. While P. sojae and P. vignae are reported to specifically infect ...soybean and cowpea, respectively, P. pisi is reported to attack pea and faba bean. Phytophthora niederhauserii is considered to have a broad host range. Zoospores of some Phytophthora species are chemotactically attracted to the isoflavones that are secreted by their host plants. The focus of the current study was to determine the chemotaxic behaviour of zoospores from closely related legume‐root infecting Phytophthora species and to investigate the correlation, if any, to host preference as determined by greenhouse pathogenicity tests. The results showed that P. sojae and P. vignae were attracted to the non‐soybean isoflavone prunetin as well as to the soybean isoflavones genistein and daidzein, which is in contrast with their host specificity on soybean and cowpea, respectively. On the other hand, P. pisi and P. niederhauserii were only attracted to prunetin, previously reported to be produced by pea, but not to the isoflavones associated with the non‐host soybean. The lack of responsiveness to genistein and daidzein in P. pisi may represent a recent adaptation to the host specialization towards pea. However, the affinity of P. niederhauserii to prunetin shows that this trait can also be present in taxa not specifically associated with legume hosts.
The recently reported red fluorescent protein DsRed from the reef coral
Discosoma sp. represents a new marker that has been codon-optimized for high expression in mammalian cells. To facilitate ...expression of DsRed in ascomycete fungi, we used the clone pDsRed-Express (Clontech) for constructing a plasmid vector, pPgpd-DsRed, containing the constitutive
Aspergillus nidulans glyceraldehyde 3-phosphate (
gpd) promoter. This vector was used for co-transformation of
Penicillium paxilli,
Trichoderma harzianum and
Trichoderma virens (syn.
Gliocladium virens) together with either pAN7-1 or gGFP, both containing a gene for hygromycin resistance for transformant selection. In addition, gGFP contains a green fluorescent protein (GFP) gene for expression in Ascomycetes. Expression of DsRed-Express was obtained in all three fungi, indicating that DsRed can be used as a highly effective vital marker in Ascomycetes. Dual marked transformants expressed both DsRed-Express and GFP in the same mycelium and were used for non-quantitative comparison of the intensity of the fluorescence using confocal laser scanning microscopy.
Trichoderma harzianum is an effective biocontrol agent against several fungal soilborne plant pathogens. However, possible adverse effects of this fungus on arbuscular mycorrhizal fungi might be a ...drawback in its use in plant protection. The objective of the present work was to examine the interaction between Glomus intraradices and T. harzianum in soil. The use of a compartmented growth system with root-free soil compartments enabled us to study fungal interactions without the interfering effects of roots. Growth of the fungi was monitored by measuring hyphal length and population densities, while specific fatty acid signatures were used as indicators of living fungal biomass. Hyphal 33P transport and beta-glucuronidase (GUS) activity were used to monitor activity of G. intraradices and a GUS-transformed strain of T. harzianum, respectively. As growth and metabolism of T. harzianum are requirements for antagonism, the impact of wheat bran, added as an organic nutrient source for T. harzianum, was investigated. The presence of T. harzianum in root-free soil reduced mat colonization by G. intraradices. The external hyphal length density of G. intraradices was reduced by the presence of T. harzianum in combination with wheat bran, but the living hyphal biomass, measured as the content of a membrane fatty acid, was not reduced. Hyphal 33P transport by G. intraradices also was not affected by T. harzianum. This suggests that T. harzianum exploited the dead mycelium but not the living biomass of G. intraradices. The presence of external mycelium of G. intraradices suppressed T. harzianum population development and GUS activity. Stimulation of the hyphal biomass of G. intraradices by organic amendment suggests that nutrient competition is a likely means of interaction. In conclusion, it seemed that growth of and phosphorus uptake by the external mycelium of G. intraradices were not affected by the antagonistic fungus T. harzianum; in contrast, T. harzianum was adversely affected by G. intraradices.
Different screening methods for selection of biological control agents (BCAs), for controlling soil and seed-borne diseases, are discussed. The shortcomings of laboratory methods focused on mechanism ...of action are discussed and we conclude that these methods should be used with caution if candidates with multifactorial or plant mediated mechanisms of control are to be obtained. In vitro screens may be useful for specific groups of microorganisms, thus, screens for antibiotics may be relevant for Streptomyces spp., and promising results have been obtained using soil plating or precolonized agar methods to screen for mycoparasitism and competitive saprophytic ability. Experience with screening in the Nordic programme 'Biological control of seed borne diseases in cereals' is summarized. Research in the four participating countries - Finland, Sweden, Norway and Denmark - followed the same paradigm: that of obtaining antagonists, well adapted to different Nordic environments, and developing them as effective BCAs. Potential antagonists were isolated from different sources and in planta screening methods were developed in order to optimize selection of antagonists effective against a range of seed borne pathogens. Screens in the laboratory or greenhouse were followed by screening in the field. The different screening procedures are compared and evaluated.PUBLICATION ABSTRACT
In six field experiments, seed treatment with Clonostachys rosea (IK726) significantly reduced disease caused by Fusarium culmorum. IK726 was active against the pathogen at average soil temperatures ...at sowing ranging from 6.2 to 12 °C. Both in the field experiments and in growth chamber experiments conducted in sand, dried and stored conidia of IK726 controlled F. culmorum as effectively as freshly harvested conidia. A high correlation was found between disease index ratings from field experiments and from corresponding growth chamber sand tests. Amendment with the stickers Pelgel or Sepiret did not influence control activity. The effective dosages of IK726 (cfu/seed) were estimated in bioassays and were very similar for freshly harvested conidia and for dried conidia. With a density of > 5×10^sup 3^ conidia per seed more than 80% disease control was repeatedly obtained with both types of conidia.PUBLICATION ABSTRACT
The influence of organic matter on the interactions between external mycelium of the arbuscular mycorrhizal (AM) fungus
Glomus intraradices, the bacterium
Burkholderia cepacia and other soil ...microorganisms was studied in a root-free sand environment. Organic matter amendment, in terms of ground barley leaves, markedly increased the growth of the external mycelium of
G. intraradices as estimated both with the fatty acid biomarker 16:1ω5 and hyphal length measurements. Mycelial proliferation of
G. intraradices in sand with organic matter was unaffected by both inoculation with
B. cepacia and a soil filtrate containing a mixed population of indigenous microorganisms. On the other hand, in the absence of organic matter, both inoculation with
B. cepacia and the soil filtrate reduced the growth of
G. intraradices, as estimated with measurements of 16:1ω5. In contrast,
B. cepacia inoculation increased hyphal length density of
G. intraradices in the absence of organic matter. Overall, the presence of external mycelium of
G. intraradices increased the bacterial biomass and counteracted a suppressive effect of
B. cepacia on the growth of saprotrophic fungi.
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