Genetic structure of <Emphasis Type="Italic">Pyrenophora teres</Emphasis> f. <Emphasis Type="Italic">teres</Emphasis> and <Emphasis Type="Italic">P. teres</Emphasis> f. <Emphasis Type="Italic">maculata</Emphasis> populations from western Canada

Infection by Pyrenophora teres f. teres (Ptt) or P. teres f. maculata (Ptm), the causal agents of the net and spot forms of net blotch of barley, respectively, can result in significant yield losses. The genetic structure of a collection of 128 Ptt and 92 Ptm isolates from the western Canadian provinces of Alberta (55 Ptt, 27 Ptm), Saskatchewan (58 Ptt, 46 Ptm) and Manitoba (15 Ptt, 19 Ptm) were analyzed by simple sequence repeat (SSR) marker analysis. Thirteen SSR loci were examined and found to be polymorphic within both Ptt and Ptm populations. In total, 110 distinct alleles were identified, with 19 of these shared between Ptt and Ptm, 75 specific to Ptt, and 16 specific to Ptm. Genotypic diversity was relatively high, with a clonal fraction of approximately 10 % within Ptt and Ptm populations. Significant genetic differentiation (PhiPT = 0.230, P = 0.001) was found among all populations; 77 % of genetic variation occurred within populations and 23 % between populations. Lower, but still significant genetic differentiation (PhiPT = 0.038, P = 0.001) was detected in Ptt, with 96 % of genetic variation occurring within populations. No significant genetic differentiation (PhiPT = 0.010, P = 0.177) was observed among Ptm populations. Isolates clustered in two distinct groups conforming to Ptt or Ptm, with no intermediate cluster. The high number of haplotypes observed, combined with an equal mating type ratio for both forms of the fungus, suggests that P. teres goes through regular cycles of sexual recombination in western Canada.     

Competence of <Emphasis Type="Italic">Frankliniella occidentalis</Emphasis> and <Emphasis Type="Italic">Frankliniella intonsa</Emphasis> strains as vectors for <Emphasis Type="Italic">Chrysanthemum stem necrosis virus</Emphasis>

The vector competence of Frankliniella occidentalis for Chrysanthemum stem necrosis virus (CSNV) was evaluated. Three vector strains with distinct competences for Tomato spotted wilt virus (TSWV) transmission were investigated, including an artificially selected strain (TsH) that has a particularly high competence (>90 %). Newly hatched larvae of F. occidentalis were given an acquisition access period of 5 days on CSNV-infected D. stramonium leaves, and reared to maturity. Their transmission efficiencies were examined using a leaf disk assay using Petunia x hybrida leaves. Following the leaf disk assay, the virus accumulation in the vectors was examined via a double antibody sandwich enzyme-linked immunosorbent assay (DAS-ELISA) of their bodies. The results showed that the CSNV acquisition and transmission efficiency of the TsH strain did not differ from those of the others, indicating that the competence of F. occidentalis as a vector for CSNV is not related to that for TSWV. The CSNV transmission and acquisition efficiencies of two F. intonsa strains (Hiroshima and Fukuoka) were also evaluated. In Hiroshima strain, 35 % of adults were viruliferous, but only two transmitters (3 %) were observed. In Fukuoka strain, 6 % were viruliferous, and no transmitters were observed. These results indicate that F. intonsa cannot be a major vector for CSNV. The accumulation of CSNV in the adults of F. occidentalis and F. intonsa evaluated using DAS-ELISA showed a significant difference in ELISA values among transmitter, viruliferous non-transmitter, and non-viruliferous individuals. These results clearly demonstrated that only transmitters that accumulated a threshold quantity of virus can transmit CSNV to plants.     

Quantitative detection of <Emphasis Type="Italic">Colletotrichum godetiae</Emphasis> and <Emphasis Type="Italic">C. acutatum sensu stricto</Emphasis> in the phyllosphere and carposphere of olive during four phenological phases

A duplex qPCR detection method was developed to detect and quantify Colletotrichum godetiae and C. acutatum sensu stricto (s.s.) in olive tissues. The method proved highly specific and sensitive with a detection limit of 10 pg for each pathogen. The analysis of green and senescent leaves, fertilized fruitlets with floral residues, green fruit and symptomatic and asymptomatic fruit collected in May, June, October and December revealed a high incidence of both C. godetiae and C. acutatum s.s. in Calabria, southern Italy. In comparison with previous reports, these results highlighted an ongoing population shift from C. godetiae to C. acutatum s.s. Interestingly, C. godetiae was slightly more abundant in terms of number of infected samples, yet the quantity of C. acutatum in infected samples was always higher, suggesting greater aggressiveness and/or sporulation ability of the latter pathogen. The populations of both C. godetiae and C. acutatum s.s. increased sharply in December even though both pathogens were detected widely in asymptomatic samples in May, June and October, confirming an important role of latent infections in the disease cycle. A large quantity of both C. godetiae (1.7 × 10⁸ cells/mg of tissue) and C. acutatum s.s. (7.5 × 10⁸ cells/mg of tissue) was estimated in symptomatic fruit, presenting an enormous inoculum potential for secondary infections. Two other important observations were a high incidence and quantity of both pathogens in senescent leaves and in fertilized fruitlets with floral residues as compared to green leaves.     

Long-term dynamics of causative agents of stem base diseases in winter wheat and reaction of Czech <Emphasis Type="Italic">Oculimacula</Emphasis> spp. and <Emphasis Type="Italic">Microdochium</Emphasis> spp<Emphasis Type="Italic">.</Emphasis> populations to prochloraz

Trichoderma aggressivum is an aggressive contaminant mould in the cultivation of Agaricus bisporus leading to severe reductions in mushroom yields. Production of fully colonised A. bisporus substrate in Europe is commonly carried out in large tunnels (Phase III), after which the substrate undergoes several bulk handling (mixing) operations before ending up on shelves in mushroom growing facilities. The work presented here studied the effect of Trichoderma aggressivum inoculum, substrate mixing and supplementation on Agaricus bisporus yields and evaluated four methods to detect T. aggressivum in bulk handled substrate. Inoculum dilution level was shown to correlate well with mushroom yield (P < 0.0001) with reductions of 2–6 % at the most dilute level (10^?4) and 60–100 % at the most concentrated level (10^?1), depending on the experiment. Supplementation, with or without T. aggressivum, had no significant effect on mushroom yield (P ≥ 0.85) but a high degree of substrate mixing was shown to significantly increase (P < 0.0001) T. aggressivum-associated crop losses. Four T. aggressivum detection methods were evaluated and a quantitative polymerase chain reaction (qPCR) method gave the most consistent and least variable results. Cycle threshold (CT) values ranged from 24 to 40, depending on the experiment and the inoculum dilution level, and false negatives (CT = 40) were reported on one occasion with the most dilute samples. The results indicate that Phase III mushroom substrate is vulnerable to infection by T. aggressivum when the fully colonised substrate is broken up and mixed during bulk handling operations, identifying a previously unidentified risk for Phase III substrate producers.     

Phytosynthesis of Silver Nanoparticles using Leaf Extracts from <Emphasis Type="Italic">Ocimum basilicum</Emphasis> and <Emphasis Type="Italic">Mangifira indica </Emphasis>and their Effect on some Biochemical Attributes of <Emphasis Type="Italic">Triticum aestivum</Emphasis>

In this investigation, leaf extracts of Ocimum basilicum and Mangifira indica were used as reducing agents for biosynthesis of silver nanoparticles (AgNPs). The biosynthesized AgNPs were authorized by UV-vis spectrophotometry and X?ray diffraction (XRD) analysis. AgNPs were obtained after 5?min of reaction at 80^oC. The formation of AgNPs was confirmed by the presence of absorption peaks at 439?nm using extract from O. basilicum and at 442.5?nm from M. indica. X?ray spectra showed strong peaks for the crystalline Ag. Shape and size of the biosynthesized AgNPs were studied using high resolution transmission electron microscope (HR-TEM). Size of the produced AgNPs was found to be 9–35?nm. Effect of the synthesized nanosilver was then investigated on some biochemical attributes of wheat plant (Triticum aestivum cultivar saka 92). The growth parameters such as shoot lengths, fresh and dry weight of shoot, chlorophyll, carbohydrate and protein contents in shoot of wheat plant were investigated. Application of AgNPs synthesized from Ocimum basilicum and from Mangifira indica at the concentrations of 20,40?ppm showed an increase in shoot length, fresh and dry weight of shoot, chlorophyll, total carbohydrate and protein content in shoot of wheat plants, beyond these concentrations an inhibitory effects were shown.     

Increased susceptibility of potato to <Emphasis Type="Italic">Rhizoctonia</Emphasis> diseases in <Emphasis Type="Italic">Potato leafroll virus</Emphasis>-infected plants

In Hokkaido potato fields, tubers produced from the plants with leaf curl symptoms caused by potato leaf roll virus (PLRV) were noted to be more densely covered with Rhizoctonia sclerotia. This observation led us to hypothesize that potato infected with PLRV would have an increased susceptibility to Rhizoctonia solani. To test this hypothesis, in a pot experiment, we inoculated PLRV-infected mother tubers with Rhizoctonia. As a result, PLRV-infected plants produced significantly fewer and smaller tubers than virus-free plants did, suggesting that PLRV-infected plants are more susceptible than virus-free plants to R. solani. Virus-free seed tubers should thus be used to reduce Rhizoctonia diseases.     

Characterization of <Emphasis Type="Italic">Dickeya</Emphasis> and <Emphasis Type="Italic">Pectobacterium</Emphasis> strains obtained from diseased potato plants in different climatic conditions of Norway and Poland

Soft rot and blackleg of potato caused by pectinolytic bacteria lead to severe economic losses in potato production worldwide. To investigate the species composition of bacteria causing soft rot and black leg of potato in Norway and Poland, bacteria were isolated from potato tubers and stems. Forty-one Norwegian strains and 42 Polish strains that formed cavities on pectate medium were selected for potato tuber maceration assays and sequencing of three housekeeping genes (dnaX, icdA and mdh) for species identification and phylogenetic analysis. The distribution of the species causing soft rot and blackleg in Norway and Poland differed: we have demonstrated that mainly P. atrosepticum and P. c. subsp. carotovorum are the causal agents of soft rot and blackleg of potatoes in Norway, while P. wasabiae was identified as one of the most important soft rot pathogens in Poland. In contrast to the other European countries, D. solani seem not to be a major pathogen of potato in Norway and Poland. The Norwegian and Polish P. c. subsp. carotovorum and P. wasabiae strains did not cluster with type strains of the respective species in the phylogenetic analysis, which underlines the taxonomic complexity of the genus Pectobacterium. No correlation between the country of origin and clustering of the strains was observed. All strains tested in this study were able to macerate potato tissue. The ability to macerate potato tissue was significantly greater for the P. c. subsp. carotovorum and Dickeya spp., compared to P. atrosepticum and P. wasabiae.     

Biochemical,physiological, and molecular characterization of <Emphasis Type="Italic">Dickeya dianthicola</Emphasis> (formerly named <Emphasis Type="Italic"> Erwinia chrysanthemi</Emphasis>) causing potato blackleg disease in Japan

The taxonomic assignment of Japanese potato blackleg isolates of Dickeya spp. has not been confirmed after the changes in their former name, Erwinia chrysanthemi. Therefore, we investigated and identified 23 representative isolates of Dickeya spp. from symptomatic stems of potatoes in Japan, with biochemical tests and phylogenetic sequence analysis using recA, dnaX, rpoD, gyrB, and 16S rDNA sequences. Results of our biochemical tests showed that all isolates can be assigned to phenon 5 and biovar 1, which are associated with D. dianthicola. Based on the recA, dnaX, rpoD, gyrB, and 16S rDNA sequences, all isolates are in the same clade with D. dianthicola and were clearly distinguished from D. chrysanthemi, D. dadantii, D. dadantii subsp. dieffenbachiae, D. solani, D. zeae, and D. paradisiaca. Therefore, we conclude that Dickeya spp. isolated from potatoes with blackleg symptoms in Japan are D. dianthicola.     

<Emphasis Type="Italic">Dittrichia viscosa</Emphasis> and<Emphasis Type="Italic">Rubus ulmifolius</Emphasis> as reservoirs of aphid parasitoids (Hymenoptera: Braconidae: Aphidiinae) and the role of certain Coccinellid species

The role of the self-sown shrubsDittrichia viscosa (L.) W. Greuter andRubus ulmifolius Schott as reservoirs of aphid parasitoids was investigated. In the field studies conducted,D. viscosa grew adjacent to crops of durum wheat and barley andR. ulmifolius grew adjacent to cotton. The relative abundance of the parasitoids of(a) Capitophorus inulae (Passerini) onD. viscosa, (b) Rhopalosiphum padi (Linnaeus) on durum wheat and barley,(c) Aphis ruborum (Börner) onR. ulmifolius, and(d) Aphis gossypii Glover on cotton in various parts of Greece, was assessed during the years 1996–2000. In 2000, the fluctuation of parasitization of the above four aphid species was recorded and the action of the aphidophagous predators of the family Coccinellidae was studied. It was observed thatAphidius matricariae Haliday predominated onC. inulae andR. padi in all sampling cases. In contrast,Lysiphlebus fabarum (Marshall) was the dominant species parasitizingA. ruborum onR. ulmifolius andA. gossypii on cotton in Thessaly (central Greece) and Macedonia (northern Greece), whereasLysiphlebus confusus Tremblay et Eady andBinodoxys acalephae (Marshall) were the dominant parasitoid species in Thrace (northern Greece).Coccinella septempunctata Linnaeus was the most abundant coccinellid species on durum wheat, whereasAdonia variegata (Goeze) predominated on cotton. However, coccinellid individuals were scarce on bothD. viscosa andR. ulmifolius. The present study indicated that these two shrubs can be regarded as useful reservoirs of aphid parasitoids.     

Histological observation of cucumber infected with <Emphasis Type="Italic">Corynespora cassiicola</Emphasis>

Erwinia amylovora is the causative agent of fire blight, which is a destructive bacterial disease of rosaceous plants. In Hungary Erwinia amylovora (Burrill) Winslow et al. was first detected in 1996. Since the appearance of fire blight, E. amylovora samples have been collected from different host plats from various geographic locations. A motif of eight nucleotides (ATTACAGA) is repeated 3–15 times in the PstI fragment of the pEa29 plasmid in Erwinia amylovora strains, and represents a valuable tool for strain classification. The number of short-sequence DNA repeats in plasmid pEa29 of 30 Hungarian isolates were determined by PCR assays and they ranged from five to ten. The SSR test is suitable for distinguishing the individual strains between the E. amylovora isolates. The examined isolates showed high pathogenicity on immature pear fruits. Several biochemical techniques, such as miniaturized API 20E, were applied on the samples. Differences were also revealed in microbiological assays like levan formation and colony morphology on semi-selective media. Examining the Hungarian Erwinia amylovora population by molecular analysis we can draw the conclusion that the population consists of different strains, which shows great diversity. E. amylovora is a widespread pathogen in Hungary, which is supported by the 30 strains isolated from various host plants from many parts of the country. The phenotypic diversity-evaluation of the E. amylovora strains showed, that they differ metabolically, like other plant pathogenic bacteria as reported by several authors. This is the first report on the diversity of E. amylovora strains isolated from Hungary.     

<Emphasis Type="Italic">Bradyrhizobium</Emphasis> isolated from huanglongbing (HLB) affected citrus trees reacts positively with primers for <Emphasis Type="Italic">Candidatus</Emphasis> Liberibacter asiaticus

Bradyrhizobium sp., a slow-growing nitrogen-fixing symbiotic bacterium of legumes and common root endophyte of other plants, is closely related to Candidatus Liberibacter asiaticus (Las), the uncultured putative pathogen associated with citrus huanglongbing (HLB). In attempts to isolate Las on a low-nutrient medium that had been used for the isolation of several uncultured bacteria of the alpha subclass of proteobacteria, slow-growing Bradyrhizobium spp. were isolated and identified by sequencing of 16S rDNA. The individual isolates tested weakly positive (Ct = 31.2–36.0) with the USDA primers commonly used in qPCR assays for Las in foliar tissues. Direct DNA extracts from roots of HLB symptomatic trees that contained sequences of Bradyrhizobium sp. had Ct values ranging from 31.2 to 36.5; sequences of Las were not present in those samples. Potential cross-reaction between DNA of members of the Rhizobiales and sequences amplified by the Las primers were tested in silico with the Primer-BLAST tool in NCBI. Similar to Las, Bradyrhizobium generated predicted 16S rDNA amplicon sizes of 78–79 bp with the qPCR primers and of 1167-1172 bp with the conventional PCR primers. Bradyrhizobium sequences of 16S rDNA had 1–7 mismatches and only 1 mismatch at the 3′ end of qPCR and conventional PCR primers confirming potential cross-reactivity. As Bradyrhizobium is usually not found in foliage, the USDA qPCR primers can be safely used to check leaves for the presence of Las, but a threshold value of 31.0 is recommended for Las detection in roots. Other primers should be tested for potential cross-reaction with members of the Rhizobiales.     

The chemotaxis regulator <Emphasis Type="Italic">pilG</Emphasis> of <Emphasis Type="Italic">Xylella fastidiosa</Emphasis> is required for virulence in <Emphasis Type="Italic">Vitis vinifera</Emphasis> grapevines

Type IV pili of X. fastidiosa are regulated by pilG, a response regulator protein putatively involved in chemotaxis-like operon sensing stimuli through signal transduction pathways. To elucidate the roles of pilG in pathogenicity of X. fastidiosa, the pilG-deletion mutant XfΔpilG and complemented strain XfΔpilG-C were generated. While all strains had similar growth curves in vitro, XfΔpliG showed significant reduction in cell-matrix adherence and biofilm production compared with wild-type X. fastidiosa and XfΔpilG-C. The genes pilE, pilU, pilT, and pilS were down-regulated in XfΔpliG when compared with its complemented strain and wild-type X. fastidiosa. Finally, no Pierce’s disease symptoms were observed in grapevines inoculated with XfΔpilG, whereas grapevines inoculated with the wild-type X. fastidiosa and complemented strain of XfΔpilG-C developed typical Pierce’s Disease (PD) symptoms. The results indicate that pilG has a role in X. fastidiosa virulence in grapevines.     

Infection of <Emphasis Type="Italic">Sorghum bicolor</Emphasis>, selected grass species and <Emphasis Type="Italic">Zea mays</Emphasis> by G<Emphasis Type="Italic">loeocercospora sorghi</Emphasis>, causal pathogen of zonate leaf spot

Zonate leaf spot (Gloeocercospora sorghi) is a common disease in Sorghum bicolor producing areas of the U.S., but little is known about its biology, virulence and severity on S. bicolor, Zea mays, and related crop grassweeds. Greenhouse studies were conducted to determine and compare the virulence and severity of G. sorghi on 10 commercially available sorghum hybrids, four Z. mays hybrids and selected grassweed species including Sorghum bicolor (grain sorghum and shattercane biotypes) and Sorghum halepense (Johnsongrass), two of the most problematic arable weeds. Plants from the respective species were inoculated with a local G. sorghi isolate and maintained in a dew-chamber at 24 °C for 24 h and then incubated under greenhouse conditions for 4 weeks. Plants were observed for lesion expression and rated using a modified Horsfall-Barrett scale (0–10). The first symptoms of infection were visible within 24 h following inoculation on shattercane and S. bicolor hybrids. Symptoms consisted of small, non-diagnostic purple lesions on the leaves. Results showed that S. bicolor, S. halepense and shattercane were susceptible to G. sorghi. All other species tested in this study were not infected. More particularly, disease severity, increased from a rating of 3 to 10 on sorghum and from 2 to 7 on S. halepense between 2 and 23 days after inoculation, respectively. However, disease severity on shattercane increased rapidly from 3.5 to 10 between 2 and 8 days after inoculation, respectively. Among the sorghum hybrids tested, FFR-322 appeared to be the most resistant to G. sorghi while Pioneer 83G66 appeared to be the most susceptible. Z. mays hybrids were not infected by the fungus used in this study. G. sorghi could be used effectively to manage shattercane and S. halepense infestations occurring in Z. mays and S. bicolor fields consisting of specific G. sorghi-resistant hybrids.     

Influence of exogenous cholesterol in Pythiaceae resistance to inhibition by <Emphasis Type="Italic">Trichoderma</Emphasis> antibiosis

The purpose of this study was to determine if exogenous cholesterol availability influenced Pythiaceae resistance to antibiosis. Characterisation of an isolate of Phytophthora erythroseptica and Pythium ultimum for tolerance to antibacterial compounds found that 0.05 g.l^?1 chloramphenicol inhibited mycelial growth by 96.6 % and 23.5 % respectively. However, the addition of cholesterol (0.01 g l^?1) to potato dextrose agar (PDA) containing 0.05 g l^?1 chloramphenicol was found to increase mycelial growth of P. erythroseptica, indicating a role for cholesterol in tolerance to inhibitory antibacterial compounds. To determine if this property extended to suppressive effects of a potential biocontrol agent, P. erythroseptica and P. ultimum were then tested against a cell-free filtrate of diffusible metabolites produced by a suppressive Trichoderma harzianum isolate in the presence and absence of cholesterol in PDA. In the absence of cholesterol, diffusible metabolites of the T. harzianum isolate were found to inhibit mycelial growth of P. erythroseptica and P. ultimum on PDA by 98 % and 63.6 % respectively (P?<?0.0001). However, the inhibitory effect of the metabolites was mitigated when 0.005 g l^?1 of cholesterol was present in PDA, with mycelial growth of P. ultimum and P. erythroseptica reduced by only 60.4 % and 41.8 %, respectively (P?<?0.0001), much less inhibition than was observed in the absence of cholesterol. These results demonstrated that access to exogenous cholesterol can influence the sensitivity of Pythiaceae species to antibiosis by positively influencing mycelial growth.     

Virulence profiling of <Emphasis Type="Italic">Xanthomonas oryzae</Emphasis> pv. <Emphasis Type="Italic">oryzae</Emphasis> isolates,causing bacterial blight of rice in India

Bacterial blight (BB) of rice caused by Xanthomonas oryzae pv. oryzae (Xoo), remains a major production constraint in rice cultivation especially in irrigated and rainfed lowland ecosystems in India. The pathogen is highly dynamic in nature and knowledge on pathotype composition among the Xoo population is imperative for designing a scientific resistance breeding program. In this study, four hundred isolates of Xoo collected from diverse rice growing regions of India were analyzed for their virulence and genetic composition. Virulence profiling was carried out on a set of differentials consisting of 22 near isogenic lines (NILs) of IR24 possessing different BB resistance genes and their combinations along with the checks. It was observed that different NILs possessing single BB resistance gene were susceptible to about 59–94% of the Xoo isolates except IRBB 13 (containing BB resistance gene xa13), which showed susceptibility to about 35% of the isolates. Based on the reaction of the Xoo isolates on the differentials, they were categorized into 22 pathotypes. Among the 22 pathotypes, IXoPt-1 and IXoPt-2 were least virulent and IXoPt # 18–22 were highly virulent. Pathotype IXoPt-19 which was virulent on all single BB resistance genes except xa13 constituted the major pathotype (22.5% isolates) and was widely distributed throughout India (16 states). This was followed by pathotype IXoPt-22 (17.25%) which was virulent on all the NILs possessing single BB resistance genes. Molecular analysis was carried out using two outwardly directed primers complementary to sequence of IS1112, a repetitive element of Xoo. A high level of genetic polymorphism was detected among these isolates and the isolates were grouped into 12 major clusters. The data indicated complex nature of evolution of the Xoo pathotypes and there was no strong correlation between pathotypes and genetic clusters as each genetic cluster was composed of Xoo isolates belonging to different pathotypes. The study indicated that none of the single BB resistance genes can provide broad spectrum resistance in India. However, two-gene combinations like xa5 + xa13 and different 3 or 4 genes combination like Xa4 + xa5 + xa13, Xa4 + xa13 + Xa21, xa5 + xa13 + Xa21 and Xa4 + xa5 + xa13 + Xa21 are broadly effective throughout India.     

Inhibitory effect of <Emphasis Type="Italic">Lycium europaeum</Emphasis> extracts on phytopathogenic soil-borne fungi and the reduction of late wilt in maize

The ability to control soil-borne pathogens in agriculture is highly conditioned by the restricted use of synthetic pesticides. Allelopathy, the antimicrobial activity of plant extracts, is a promising option against crop pathogens. Extracts from Lycium spp. such as L. barbarum, L. chinense and L. intricatum possess biological and therapeutic properties. Individual methanolic extracts from leaves and stems of the Mediterranean medicinal species L. europaeum collected in two locations of Tunisia were each evaluated in vitro against Verticillium dahliae (Vd), Sclerotinia sclerotiorum (Ss) and Harpophora maydis (Hm). The mycelial growth of the three fungi was significantly reduced by all the extracts at doses of 10 and 30 μl mL^?1 (equivalent to 1 and 3 mg plant tissue mL^?1). The sporulation of Hm was almost completely inhibited in all the amendments, but that of Vd was stimulated by one of the leaf extracts when 1 and 3 mg dried plant tissue mL^?1 were used. Sclerotia of Ss were formed in a smaller number, their total weight increasing at extract doses equivalent to 1 mg plant tissue mL^?1 and higher. In greenhouse, the pathogenicity of Hm was confirmed as early as 6 weeks after inoculation, since it caused significant decreases of weights in both roots and aboveground parts of maize. The detrimental effect of Hm on maize root weight in greenhouse was significantly counteracted by one of the leaf extracts added by watering. In total, 11 phenolic compounds were separated in the four extracts. The hydroxycinnamic acid family, including chlorogenic acid as a major compound, represented more than 50% of the total content in all the samples. Rutin was the most abundant flavonoid. The results of this work show the detrimental effect of L. europaeum extracts against the soil-borne pathogens Hm, Ss and Vd, and highlight their potential in crop protection if adequately developed into final products and used in combination with other tools.     

Accumulation of toxigenic <Emphasis Type="Italic">Fusarium</Emphasis> species and <Emphasis Type="Italic">Stenocarpella maydis</Emphasis> in maize grain grown under different cropping systems

Entomopathogenic nematodes in the genus Steinernema are associated with Xenorhabdus spp. bacteria. When steinernematid colonise an insect host the nematode-bacterium association overcomes the insect immune system and kills the host within 48 h. Xenorhabdus spp. produce secondary metabolites that are antifungal to protect nematode-infected cadavers from fungal colonization. The concentrated, or cell-free metabolites of X. szentirmaii exhibit high toxicity against various fungal plant pathogens and show potential as natural bio-fungicides. In the current study, we determined 1) thermo-stability, 2) dose-response, and 3) shelf-life of antifungal metabolites of X. szentirmaii against Monilinia fructicola (cause of brown rot of peach and other stone fruit) and Glomerella cingulata (cause of antharacnose). Thermo-stability was determined by autoclaving bacterial culture broths (121 °C and 15 psi for 15 min) and measuring fungal growth on in potato dextrose agar (PDA) containing 10% of the supernatants. Autoclaving had no impact on the antifungal activity of the secondary metabolites. Over a test period of 9 months, the activity of both extract types did not decline when stored at 4 or 20 °C. A dose-response study (10, 20, 40, 60, 80 and 100% supernatant-containing metabolite) using both phytopathogens demonstrated that a greater dose of supernatant increased antifungal activity. The antifungal-metabolite containing supernatant of X. szentirmaii has potential as a bio-fungicide. These results demonstrate the metabolite(s) are thermo-stable, they have a long shelf-life and require no stabilizing formulation, even at room temperature.     

Survival of <Emphasis Type="Italic">Curtobacterium flaccumfaciens</Emphasis> pv. <Emphasis Type="Italic">flaccumfaciens</Emphasis> in the soil under Brazilian conditions

Mustard clubroot, caused by Plasmodiophora brassicae, is a serious disease that affects Brassica juncea var. tumida Tsen, a mustard plant that is the raw material for a traditional fermented food manufactured in the Chongqing Municipality, People’s Republic of China. To find antagonistic bacteria for P. brassicae, 124 bacteria were obtained from the rhizosphere soil of B. juncea var. tumida grown in Fuling, Chongqing. Isolates were preliminarily chosen by evaluating the inhibition rate of the P. brassicae resting spore germination. The biocontrol effects of three antagonistic bacteria against clubroot on B. juncea var. tumida were evaluated in a greenhouse experiment. B18 showed the highest control efficiency, at 63.4% in the greenhouse test. In a field trial, B18 was also effective in controlling clubroot, but only at a 49.7% efficiency rate. According to 16S rDNA sequence analysis, strain B18 had a 100% sequence similarity with type strain Zhihengliuella aestuarii DY66^T (EU939716). Based on morphological, cultural, physiological and biochemical characteristics, the DNA G + C content, polar lipids, fatty acids, cell wall analysis, as well as DNA–DNA hybridization, strain B18 was identified as Z. aestuarii B18. Thus, the isolate B18 might have a potential biocontrol application for clubroot. We report for the first time that Z. aestuarii B18 can control clubroot.     

Characterization of <Emphasis Type="Italic">Bean common mosaic virus</Emphasis> and <Emphasis Type="Italic">Bean common mosaic necrosis virus</Emphasis> isolates in common bean growing areas in Turkey

Bean common mosaic virus (BCMV) and Bean common mosaic necrosis virus (BCMNV) are well-known legume-infecting potyviruses. The incidences of BCMV and BCMNV infections were determined by ELISA in 367 seed and leaf samples which were collected in 15 common bean-growing provinces of Turkey. Of the samples tested, 67 (18.2 %) occurred to be infected with BCMV, however only 5 (1.4 %) were infected with BCMNV. A total of 45 ELISA-positive samples were selected from single-virus infected ones to determine BCMV and BCMNV pathogenicity groups (PGs) by using a set of bean cultivars that contain different combinations of resistance genes. Some BCMV populations exhibiting unusual pathogenicity were identified. One of them, named TR-180, was found to overcome resistance conferred by bc-1, bc-1 ² , bc-2 and bc-2 ² recessive alleles in common bean and assigned to PG VII. This isolate shared high (99 %) sequence identity with previously identified BCMV RU-1 and RU-1-related strains (RU1-OR-B and RU1-OR-C) according to a BLAST analysis of the nucleotide sequences of RT-PCR amplified products comprising the complete coat protein and 3′ partial NIb regions. The isolates TR-203 and TR-256 produced a distinctive reaction pattern in the dominant I gene-bearing bean cultivars Amanda and Isabella at lower (<30 °C) temperatures and were classified into PG IVb. These isolates were found to be 99 % identical to US-1 strain based on 3′ terminal nucleotide sequences of the BCMV genome. A fourth isolate, TR-243, involved mixed BCMV populations, as confirmed by partial nucleotide sequence analysis; one was classified as belonging to PG VII being similar to TR-180, and another was assigned to PG IVb. In conclusion, on the basis of both the reactions of differential bean cultivars and ELISA results, most of BCMV isolates were assigned to pathogroup PG VII and BCMNV isolates to PG VIb. This study is the first to show that four recessive resistance alleles of common bean can be overcome by a single field isolate of BCMV, and that a wide range of BCMV pathogroups are present in Turkey.