Identification of pathogenic fungi and preliminary screening for resistance in <Emphasis Type="Italic">Jatropha curcas</Emphasis> L. germplasm

Sexual reproduction in fungi is controlled by mating type genes, which are located at the MAT locus. In this study, we investigated the structure of this locus in the phytopathogenic fungus Phyllosticta citricarpa, the causal agent of citrus black spot disease. Despite intensive study, its sexual state has never been observed in single-spore culture. Through analysis of the genome sequences of two individual P. citricarpa isolates, the sequence of the DNA lyase gene was identified and, as previously reported in the literature, the mating type genes were located in the 3′ flanking region of this gene. The results suggested that P. citricarpa is heterothallic, owing to the exclusive presence of the MAT1–1 or MAT1–2 gene in individual strains. In order to characterize the MAT locus, we designed primers to amplify this region. P. citricarpa was found to have complete and apparently functional copies of MAT genes, containing α-1 and HMG domains, present in different isolates. In addition to MAT1–2-1 and MAT1–1-1 genes, the MAT1–1-4 gene was located in the 5′ flanking region of the MAT1–1-1 gene and the MAT1–2-5 gene was located in 5′ flanking region of the MAT1–2-1 gene. A multiplex PCR protocol was also developed to differentiate P. citricarpa idiomorphs, which can be used in distribution and incidence studies of mating type strains, in order to determine the occurrence of sexual reproduction and to facility crossing studies. Furthermore, in Brazil, the two idiomorphs occur in a 1:1 ratio, which is expected in sexually reproducing populations.     

Pathogenicity,Morpho-Species and Mating Types of <Emphasis Type="Italic">Alternaria</Emphasis> spp. causing Alternaria blight in <Emphasis Type="Italic">Pistacia</Emphasis> spp. in Turkey

Alternaria genus includes many plant pathogens on numerous hosts, causing leaf spots, rots and blights. Alternaria blight has been observed as one of the important fungal diseases of pistachio (Pistacia vera L.) as well as its wild relatives (P. terebinthus, P. lentiscus, P. khinjuk, P. atlantica, P. mutica) in Turkey. Alternaria species were sampled from Pistacia spp. hosts from different geographic regions in Turkey during field trips in late spring to early fall of 2013. Alternaria blight symptoms were observed mainly on fruits and rarely on leaves. Four hundred and twenty two of the isolates were morphologically defined as A. alternata, A. tenuissima, A. arborescens and also intermediate morpho-species between A. alternata/A. arborescens. Pathogenicity of the isolates was confirmed with host inoculations on detached fruits. Mating types of 270 isolates of Alternaria spp. from the collection were identified using a PCR-based mating type assay that amplifies either a MAT1-1 or a MAT1-2 fragment from the mating locus. Although a strongly clonal population structure was expected due to the putative asexual reproduction of these fungi, both idiomorphs were detected at equal frequencies at several different spatial scales. The distribution of mating types within each geographic region, within host species as well as in overall collection was not significantly different from 1:1. Amplified fragments of partial idiomorph sequences were obtained for representative isolates. Parsimony trees were depicted based on sequence data of mating type genes for these representative isolates as well as some other Alternaria species obtained by Genebank. Several point mutations presented a few clusters which are supported by high bootsrapped values. The Alternaria blight disease agents both from cultivated and wild hosts were pathogenic on pistachio which may cause difficulties to control the disease because of extensity of pathogen sources. Besides, equal mating type distribution of the pathogen at both geographic and host species levels suggests a potential for sexual reproduction of Alternaria spp. in Turkey.     

Fungicide resistance profile and genetic structure of <Emphasis Type="Italic">Botrytis cinerea</Emphasis> from greenhouse crops in Cyprus

Botrytis cinerea is a complex species prone to fungicide resistance and characterized by enormous genetic diversity. During 2013, 220 B. cinerea isolates causing gray mold were collected from greenhouse-grown crops in the regions of Ammochostos, Larnaca, and Limassol (Cyprus). Sensitivities of the sampled populations to seven botryticides with different modes of action were screened in vitro. The results of this in vitro screening highlighted the widespread phenomenon of fungicide resistance in greenhouses, since only 8.6 % of the isolates were sensitive to all botryticides. Resistance to thiophanate-methyl was the most prevalent, with frequencies ranging from 53.8 % to 80 %. Similarly, high resistance frequencies were observed for pyraclostrobin (27.1 to 78.9 %) and boscalid (28.2 to 66.2 %). Multiple fungicide resistant phenotypes were predominant, covering 67.3 % of the population, with frequencies of 80.0, 37.5, 53.8, 83.1, and 60.2 % in cucumber, eggplant, green bean, strawberry, and tomato, respectively. No fludioxonil-resistant isolates were observed. Botrytis cinerea and Botrytis group S genotypes comprised the gray mold population. B. cinerea was predominant within cucumber, eggplant and strawberry, whereas both genotypes were in equilibrium in green bean and tomato. However, Botrytis group S was found in all hosts. B. cinerea was the most prevalent in the majority of fungicide resistance phenotypes from strawberry, while genotype distributions within tomato were generally more balanced. B. pseudocinerea was not detected in the sampled population. Overall, frequency of the mating type allele MAT1–1 was higher to MAT1–2, underlying their unequal distribution in the population. However, cases of 1:1 distribution were apparent within particular subpopulations, suggesting that mating in the field cannot be excluded.     

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.     

Distribution and characterization of Dothistroma needle blight pathogens on <Emphasis Type="Italic">Pinus mugo</Emphasis> in Slovakia

The occurrence and distribution of Dothistroma needle blight (DNB) on Pinus mugo was studied in 2014–2015 around the Slovakia. In total, 42 localities were investigated both native and planted ones. Symptoms of DNB were observed on 35 localities only on planted shrubs. All these 35 localities are new P. mugo DNB stands. No DNB symptoms were observed in natural and naturally regenerated plantations. DNA was extracted from a total of 236 isolates and eight needle samples. Based on the ITS-rDNA comparisons and using species specific primers, both pathogenic Dothistroma species were detected: D. septosporum and D. pini. Isolates of D. septosporum had ITS sequences identical to D. septosporum from Europe and both mating types were identified with slight predominance of MAT2. The ratio of D. septosporum mating types varies significantly between sites, ranging from an equal proportion of each mating type to single mating type populations. D. pini ITS sequence grouped with D. pini from Ukraine, Russia and Switzerland and only MAT2 was found.     

Characterization of <Emphasis Type="Italic">Myoviridae</Emphasis> and <Emphasis Type="Italic">Podoviridae</Emphasis> family bacteriophages of <Emphasis Type="Italic">Erwinia amylovora</Emphasis> from Hungary - potential of application in biological control of fire blight

The virulence spectrum of 300 isolates of Xanthomonas oryzae pv. oryzae (Xoo), representing 17 districts of Punjab, Pakistan was elucidated through inoculation on a set of six rice IRRI-differentials. The virulence level was assessed by using principal component and cluster analysis. Among six principal components (PCs), PC-1 exhibited 59.3 % of the total variance. The highly virulent isolates clusters on the positive side of the ordination away from the point of intersection of PC1 and PC2 and classifies the Xoo isolates from slow disease to the highest disease causing entities. The 300 isolates were categorized into 29 pathotypes (Pt1-29) wherein the highly virulent pathotype (Pt-1), comprises of 39 Xoo isolates were widespread in 12 districts. The majority of Xoo isolates were moderately to least virulent (21.7–43 %) and average disease progress curves confirmed the field reactions of these pathotype clusters for an efficient recognition of Xoo isolates. Interaction of the pathogen with differentials harboring different resistant genes was well investigated in the current study for future management approaches for which the surveillance of the new Xoo pathotypes may expedite the disease resistant rice breeding programme in the country.     

Development of specific primers based on the genomes of <Emphasis Type="Italic">Penicillium</Emphasis> spp. for rapid detection of <Emphasis Type="Italic">Penicillium digitatum</Emphasis> among fungal isolates in citrus

Specific primers targeting Penicillium digitatum were developed based on fungal genes RPB1 and cmd, which are conserved among the genomes of Penicillium spp. The specific primers were designed based on the mutational sites in the homologous regions of the conserved genes. The results indicated that primer pairs RPB1–1 and cmd-3 were specific enough to distinguish Penicillium digitatum (N1) from Penicillium chrysogenum (Q), Penicillium italicum (A10) and Penicillium expansum (L) when the DNA samples were diluted 100-fold. To further verify the effectiveness and specificity of the two primer pairs RPB1–1 and cmd-3, 38 strains of fungal isolates from sources related to citrus were detected using both primer pairs, and 14 candidate P. digitatum strains were identified. Then, the fourteen candidate P. digitatum strains were further identified as P. digitatum by morphological and molecular methods, which confirmed the detection accuracy and reliability of the specific primer pairs RPB1–1 and cmd-3 as molecular markers of P. digitatum. This work may significantly facilitate the rapid identification of P. digitatum in the citrus industry.     

Identification and characterization of <Emphasis Type="Italic">Colletotrichum</Emphasis> spp. associated with chili anthracnose in peninsular Malaysia

Anthracnose fruit rot caused by Colletotrichum spp. is a serious post-harvest disease of chili fruits (Capsicum spp.). One hundred-thirty isolates of Colletotrichum spp. were isolated from anthracnose of green and red cayenne pepper (Capsicum annuum) and bird’s eye chili (Capsicum frutescens). The isolates were morphologically identified as Colletotrichum acutatum sensu lato (62 isolates), Colletotrichum truncatum (54 isolates), and Colletotrichum gloeosporioides sensu lato (14 isolates). Molecular identification and phylogenetic analyses were based on internal transcribed spacer regions, β-tubulin, actin, and glyceraldehyde-3-phosphate dehydrogenase genes, and the isolates were re-identified as C. scovillei (58 isolates), C. truncatum (54 isolates), C. siamense (11 isolates), C. fioriniae (four isolates), and C. fructicola (3 isolates). Maximum likelihood trees using combined sequences showed that isolates of the same species grouped in the same main clade with the isolates used for comparison. Pathogenicity testing showed that the tested isolates from each species were pathogenic towards green and red Capsicum annuum and Capsicum frutescens upon treatment of wounded fruit, using both the mycelial plug and conidial suspension methods. Only five isolates of C. truncatum and seven isolates of C. scovillei were found to be pathogenic upon treatment of unwounded fruit. The occurrence of five Colletotrichum spp. (C. siamense, C. fructicola, C. scovillei, C. fioriniae, and C. truncatum) associated with chili anthracnose in Peninsular Malaysia indicates that correct species identification is important to formulate not only effective disease management, but also effective quarantine policy.     

Root-specific expression of defensin in transgenic tobacco results in enhanced resistance against <Emphasis Type="Italic">Phytophthora parasitica</Emphasis> var. <Emphasis Type="Italic">nicotianae</Emphasis>

Phytophthora species are soil-borne pathogens that damage plants in both agro- and natural ecosystems. To suppress the devastating pathogen, we generated a root-specific expression system using a specific promoter (pPRP3) conferring elevated expression of the target gene in roots that are very susceptible to soil-borne pathogens. To verify root-specific expression, we compared β-glucuronidase (GUS) expression driven by a constitutive or root-specific promoters in shoots and roots. In histochemical and fluorometric assays, GUS activity was detected in whole tobacco plants when GUS expression was driven by p35S, but was detected only in the roots by pPRP3. We then expressed a pepper defensin (J1–1) gene in tobacco to elucidate its effect on plant resistance. The accumulation of J1–1 was also tissue-specific in transgenic tobacco plants. Finally, transgenic plants carrying GUS or J1–1 genes in combination with p35S or pPRP3 were inoculated with Phytophthora parasitica var. nicotianae and Pythium aphanidermatum. Disease symptoms were significantly suppressed in transgenic plants that accumulated J1–1, regardless of the promoter used. Furthermore, the expression of PR genes was induced in J1–1 transgenic plants, exhibiting much higher levels in p35S-driven J1–1 plants than in pPRP3::J1–1 plants. These results demonstrated that J1–1 transgenic plants were primed for enhanced expression of PR genes, which provided synergistic effects with the defensin for disease resistance.     

Comparison among Japanese isolates of <Emphasis Type="Italic">Pseudomonas savastanoi</Emphasis> pv. <Emphasis Type="Italic">savastanoi</Emphasis>, causal agent of olive knot disease

Olive knot disease in Japan was first reported in Shizuoka Prefecture in 2014, and the causal agent was identified as Pseudomonas savastanoi pv. savastanoi. Subsequently, olive trees having knots were also found in Aichi and Kanagawa Prefectures in 2015, and the isolates from knots were also suspected to be P. savastanoi pv. savastanoi through preliminary examinations. Therefore, the Aichi and Kanagawa isolates were identified through comparison of isolates from three prefectures. Phylogenic analysis based on 16S rDNA and housekeeping genes (gyrB, rpoD, gltA and gap1) revealed that the isolates belonged to the same cluster as the pathotype strain, ICMP4352^PT. The iaaM, H and L genes, which are involved in promotion of symptoms, and the ina gene coding the ice nucleation protein, were detected by PCR from all the isolates. In rep-PCR (ERIC and REP) analyses, the isolates yielded DNA fragment-banding patterns that were nearly identical to that of ICMP4352^PT, but slight variations in banding patterns were observed among them. In a pathogenicity test, the isolates formed distinct knots on olive and pink jasmine. Phenotypic properties of the isolates were almost identical to those of ICMP4352^PT, with the exception of d-sorbitol utilization. Consequently, Aichi and Kanagawa isolates from olive were identified as P. savastanoi pv. savastanoi, and several genetic diversities in terms of rep-PCR were found in the Japanese population of P. savastanoi pv. savastanoi, indicating their heterogeneity.     

A climatic risk analysis of the threat posed by the South American leaf blight (SALB) pathogen <Emphasis Type="Italic">Microcyclus ulei</Emphasis> to major rubber producing countries

In 2010, symptoms of cobweb disease were observed on cultivated Pleurotus eryngii crops in Spain. Based on morphological and genetic analyses, the causal agent of cobweb was identified as Cladobotryum mycophilum. Pathogenicity tests on fruit bodies were performed using conidial suspensions of three C. mycophilum isolates. The causal agent was re-isolated in 80–85 % of the fruit bodies inoculated internally and 15–40 % of those fruit bodies inoculated on the cap surface. The results pointed to a certain resistance of the P. eryngii cap surface to the mycelium of C. mycophilum. Two cropping trials inoculated with C. mycophilum were set up to evaluate the pathogenicity of the causal agent of cobweb in two casings. At the end of the growth cycle, 50–60 % of the inoculated blocks cased with mineral soil, and 20–33 % of the inoculated blocks cased with black peat showed cobweb symptoms. This difference in the appearance of the disease and its aggressiveness may be partly explained by different electrical conductivity values of the casing materials used. In vitro sensitivity of the C. mycophilum isolates and P. eryngii strains against four fungicides (chlorothalonil, prochloraz-Mn, thiabendazole and thiophanate-methyl) was assessed in radial growth experiments on fungicide-amended media. The most effective fungicides for inhibiting the in vitro growth of C. mycophilum were prochloraz-Mn and chlorothalonil, while prochloraz-Mn was also the most selective fungicide between P. eryngii and C. mycophilum, and chlorothalonil was the most toxic fungicide against the P. eryngii mycelium.     

Characterisation of <Emphasis Type="Italic">Aureobasidium pullulans</Emphasis> isolates from <Emphasis Type="Italic">Vitis vinifera</Emphasis> and potential biocontrol activity for the management of bitter rot of grapes

Aureobasidium isolated from Vitis vinifera (cv Chardonnay) grapevine tissues were characterised using morphological and molecular techniques. Species level identification of 29 isolates was accomplished by partial amplification and sequencing of the ITS region (ITS1–5.8S–ITS2) using universal primers ITS1 and ITS4. A comparison of nucleotide sequences using BLAST followed by phylogenetic analysis revealed that all isolates examined were Aureobasidium pullulans. Strain level discrimination of a total of 100 epiphytic Aureobasidium isolates including three reference strains was successfully carried out using two inter simple sequence repeat (ISSR) primers, (AAC)5 and (GTG)5 and the Intron Splice Junction R1 (ISJ-R1) primer in which 24, 24 and 15 scorable bands were produced for each primer, respectively. The high level of genetic variation recorded among the isolates further highlighted the high levels of strain diversity among A. pullulans residing on grapevines. Thirty-two epiphytic Aureobasidium isolates were examined for their ability to inhibit the growth of Greeneria uvicola, responsible for bitter rot of grapes. Using an in-vitro dual-culture antagonism assay, all isolates inhibited the growth of G. uvicola (Isolates DAR 77272 and DAR 77273) with inhibition ranging from 15 to 85%. Three Aureobasidium isolates were then examined for their ability to inhibit G. uvicola when co-inoculated onto detached berries, leaves and grape bunches growing on potted vines in a glass house. All isolates reduced the severity of bitter rot infection. The results indicate that A. pullulans has the potential to suppress bitter rot of grapes.     

Pathogenicity and Fungicide Sensitivity of <Emphasis Type="Italic">Rhizoctonia solani</Emphasis> and <Emphasis Type="Italic">R. cerealis</Emphasis> Isolates

The Rhizoctonia solani species consists of multinucleate isolates that belong to anastomosis groups AG1–AG3 and differ in virulence and host affinity. R. cerealis is a binucleate species of anastomosis group AG-D which causes sharp eyespot, a common plant disease in Poland. Rhizoctonia spp. is a ubiquitous soil pathogen that poses a significant threat for global crop production due to the absence of effective crop protection products. The aim of this study was to determine the virulence of R. solani and R. cerealis isolates towards Beta vulgaris, Zea mays, Triticum spelta and T. aestivum seedlings, to confirm the presence of endopolygalacturonase genes pg1 and pg5 in the genomes of the tested isolates and to evaluate the tested isolates’ sensitivity to triazole, strobilurin, imidazole and carboxamide fungicides. All tested isolates infected B. vulgaris seedlings. but none of them were virulent against Z. mays plants. R. solani isolates AG4 PL and AG2-2IIIB PL were characterized by the highest virulence (average infestation score of 2.37 and 2.53 points on a scale of 0–3 points) against sugar beet seedlings. The prevalence of infections caused by most of the analysed isolates (in particular R. solani AG4 J—11.8, and R. cerealis RC2—0.78) was higher in spelt than in bread wheat. The virulence of the analysed isolates was not correlated with the presence of pg1 and pg5 genes. The efficacy of the tested fungicides in controlling Rhizoctonia spp. infections was estimated at 100% (propiconazole + cyproconazole), 98.8% (penthiopyrad), 95.4% (tebuconazole) and 78.3% (azoxystrobin).     

Morphological and molecular characterization of <Emphasis Type="Italic">Diaporthe (</Emphasis>anamorph <Emphasis Type="Italic">Phomopsis)</Emphasis> complex and pathogenicity of <Emphasis Type="Italic">Diaporthe aspalathi</Emphasis> isolates causing stem canker in soybean

The complex of Diaporthe (asexual morph) species occurring on soybean constitutes an important pathogenic group associated with diseases such as pod and stem blight, seed decay and stem canker. Stem canker, caused by Diaporthe aspalathi, has been reported as the most aggressive form of canker and its occurrence has limited soybean crop productivity in the southern United States. The main form of pathogen control is the use of stem canker resistant soybean varieties. In this study, strains of Diaporthe and Phomopsis were isolated from stem and seeds of soybean in different locations in South America during the years 1989–2014. Genomic DNA from 26 isolates were analyzed by PCR-restriction fragment length polymorphism (RFLP) and Amplified Fragment Length Polymorphism (AFLP) techniques, and sequencing of internal transcribed spacer (ITS) regions of ribosomal DNA. The molecular analysis of ITS sequences by alignment with those of ex-type strains deposited in GenBank and morphological characteristics allowed the identification of Phomopsis longicolla, D. phaseolorum var. sojae, D. caulivora and D. aspalathi. An analysis of the pathogenicity of 13 isolates of D. aspalathi inoculated in soybean genotypes carrying different resistance genes to stem canker (Rdm1, Rdm2, Rdm3, Rdm4, Rdm5 and Rdm?) enabled us to identify the occurrence of at least three races of D. aspalathi occurring in Brazil. Among the isolates identified as D. aspalathi, both molecular and phenotypic analyses showed clustering depending on the date of collection and pathogenicity, which revealed the existence of variability of the pathogen.     

Outcome of sexual reproduction in the <Emphasis Type="Italic">Phytophthora infestans</Emphasis> population in Estonian potato fields

In this study, the Estonian population of Phytophthora infestans was characterized with mating type, sensitivity to metalaxyl, virulence on 11 potato R-gene differentials and 12 SSR markers to show the outcome of potential sexual reproduction in the population. During the three years 2010–2012, 141 P. infestans isolates, collected from 23 potato fields, showed quite a high and stable frequency of the A2 mating type, 48% of the total population. In 87% of all sampled potato fields, both mating types were recorded, suggesting continuous sexual reproduction of P. infestans and possible oospore production. Metalaxyl-sensitive isolates prevailed in all three years (68 out of 99 isolates). Amongst the 95 isolates tested, 51 virulence races were found. The race structure was diverse, and most pathotypes were unique, appearing only once; the two most common pathotypes, 1.2.3.4.6.7.10.11 and 1.2.3.4.7.10.11, comprised 35% of the population. The P. infestans population was genetically highly diverse and most of the multilocus genotypes (MLGs) appeared only once. Furthermore, all of the MLGs appeared in only one of the three sampling years. Our results confirm that the high diversity in the Estonian P. infestans population is most likely the result of frequent sexual reproduction, which benefits the survival, adaptability and diversity of the pathogen in the climate of North-Eastern Europe.     

Characterization of <Emphasis Type="Italic">Trichoderma</Emphasis> species isolated in Ecuador and their antagonistic activities against phytopathogenic fungi from Ecuador and Japan

Native Trichoderma spp. were isolated from agricultural fields in several regions of Ecuador. These isolates were characterized via morphological observation as well as molecular phylogenetic analysis based on DNA sequences of the rDNA internal transcribed spacer region, elongation factor-1α gene and RNA polymerase subunit II gene. Fifteen native Trichoderma spp. were identified as T. harzianum, T. asperellum, T. virens and T. reesei. Some of these strains showed strong antagonistic activities against several important pathogens in Ecuador, such as Fusarium oxysporum f. sp. cubense (Panama disease) and Mycosphaerella fijiensis (black Sigatoka) on banana, as well as Moniliophthora roreri (frosty pod rot) and Moniliophthora perniciosa (witches’ broom disease) on cacao. The isolates also showed inhibitory effects on in vitro colony growth tests against Japanese isolates of Fusarium oxysporum f. sp. lycopersici, Alternaria alternata and Rosellinia necatrix. The native Trichoderma strains characterized here are potential biocontrol agents against important pathogens of banana and cacao in Ecuador.     

Efficacy of some rhizospheric and endophytic bacteria <Emphasis Type="Italic">in vitro</Emphasis> and as seed coating for the control of <Emphasis Type="Italic">Fusarium culmorum</Emphasis> infecting durum wheat in Tunisia

Sixty two rhizospheric and endophytic bacterial strains were evaluated for their biocontrol effect on two aggressive Fusarium culmorum isolates (Fc2 and Fc3). We observed that 35 % and 23 % of the tested strains inhibited the in vitro growth of Fc2 and Fc3 respectively. The observed antagonism was due to inhibition by contact (13–19 % of the strains) or at distance (10–16 % of the strains) for both fungal isolates. Some of the antagonistic bacteria showed the ability to produce diffuse and/or volatile compounds that inhibit the growth, the sporulation and macroconidia germination of F. culmorum. None of the tested antagonistic bacteria showed chitinase activity on synthetic medium. The sequencing of the 16S rDNA genes of some antagonistic bacteria showed that they belong to the genera Bacillus, Pseudomonas and Microbacterium. The double inoculation of durum wheat seeds by the antagonistic bacterial strains (B13, B18, BSE1, BSE3 and B16E) and the two F. culmorum isolates showed that germination and seedling vigor were generally improved in vitro. The percentage of infected seeds was also reduced. In greenhouse trials, the biocontrol effectiveness of F. culmorum was dependant from the virulence of the fungal strain and the specificity of the antagonistic interaction between bacterial and fungal strains. The bacterial strains B18 and B16E reduced F. culmorum infection on durum wheat plants probably due to their antagonistic and plant growth promoting activities and they may be used in a mixture as seed biopriming inoculum for plant growth bio-promoting and Fusarium wheat diseases biocontrol.     

Identification,virulence factors characterization,pathogenicity and aggressiveness analysis of <Emphasis Type="Italic">Fusarium</Emphasis> spp., causing wheat head blight in Iran

Fusarium head blight (FHB), mainly caused by Fusarium graminearum species complex (FGSC) and also by other species of this genus, is one of the most destructive cereal diseases with high yield losses and mycotoxin contamination worldwide. The aim of this study was to identify Fusarium species, characterize their virulence factors such as trichothecene genotypes and cell wall degrading enzymes (CWDEs), and also investigate virulence of the isolates obtained from wheat plants with FHB symptoms in Golestan province of Iran. Among 41 isolates tested, 24 were F. graminearum sensu stricto (s.s.), six were F. proliferatum, four were F. culmorum, three isolates belonged to each of F. subglutinans and F. meridionale species and one isolate of F. asiaticum was identified. Among Fusarium isolates, the nivalenol (NIV) genotype could be found more frequently, followed by 3-acetyl deoxynivalenol (3-ADON) and 15-acetyl deoxynivalenol (15-ADON) genotypes. Production of trichothecenes in autoclaved rice cultures was analyzed by gas chromatography (GC) and confirmed by GC–MS. The mean levels of NIV, 3-ADON and 15-ADON produced by Fusarium spp. were 824, 665 and 622 μg kg^?1, respectively. All Fusarium isolates were capable of producing CWDEs, mainly cellulase and xylanase. Lipase and pectinase activities appeared later and at less quantities. In overall, the isolates FH1 of F. graminearum and FH8 of F. proliferatum showed the maximum activity of CWDEs, which was correlated with high level of their virulence and aggressiveness on wheat. On the other hand, correlation was observed between the level and type of trichothecene produced by each isolate and its virulence on wheat. Virulence of trichothecene producing isolates was higher than that of non-trichothecene producing isolates. Our results suggested that CWDEs and trichothecenes, as virulence factors, have considerable roles on virulence and aggressiveness of the pathogen. This is the first report on the effect of trichothecenes and CWDEs on virulence and aggressiveness of Fusarium spp. associated with FHB disease in wheat growing regions of Iran.     

Genome-wide identification of mildew resistance locus O (<Emphasis Type="Italic">MLO</Emphasis>) genes in tree model poplar (<Emphasis Type="Italic">Populus trichocarpa</Emphasis>): powdery mildew management in woody plants

Poplars are economically important fast growing trees. They are exposed to broad range of fungal diseases like powdery mildew (PM). MLOs (mildew resistance locus O), as plant susceptibility genes, act as negative regulators and whose loss-of-functions confer complete resistance to PM disease. Herein, work identified the MLO gene family members in poplar, a woody model species. A total of 26 identified MLOs (annotated as PtMLO1–26) were distributed on 14 poplar chromosomes either individually or in groups of two to four. PtMLO genes encoded a polypeptide of 341–593 residues with a characteristic MLO domain structure. One tandem and eight segmental duplications were revealed in PtMLO genes. PtMLO proteins anchored at plasma membrane and had putative 5–9 TMDs with extracellular/cytosolic N- and C-terminuses. They were rich in leucine (9.1–12.9%), which is reported to play roles in defense response signaling. The C-terminal calmodulin-binding domain (CaMBD), reported to modulate the signaling mechanism in the defense response, was completely preserved in all PtMLOs, except PtMLO6. This domain was partially absent in PtMLO6 which is inferred to be a different MLO type or a pseudogene with a lost/impaired function in PM response. Besides, second and third cytoplasmic loops that are critical for PM-susceptibility were identified in PtMLOs. Particularly, PtMLO17, 18, 19, and 24 genes, inferred from Arabidopsis-poplar comparative phylogeny, were identified as potential candidates that may be involved in poplar-PM resistance. Notably, inductions of 14 PtMLO genes were detected in probes of microarray data such as GSE56865, GSE16417, and GSE23726 under different fungal infections indicating their involvements in plant defense. Overall, this work provided a basis for woody plant genomics for the effective and better management of poplar-PM disease.     

Population features of <Emphasis Type="Italic">Xanthomonas arboricola</Emphasis> pv. <Emphasis Type="Italic">pruni</Emphasis> from <Emphasis Type="Italic">Prunus spp.</Emphasis> orchards in northern Italy

Bacterial leaf/fruit spot and canker of stone fruits, caused by Xanthomonas arboricola pv. pruni, is a recurrent disease in Italy. A set of 23 strains has been isolated in peach and plum orchards in an intensively stone fruit cultivated area located in north-eastern Italy. They were all identified as X. arboricola pv. pruni by means of phytopathological and serological features: hypersensitive reaction on bean pods, pathogenicity test on immature peach or plum fruitlets, identification by immunofluorescence assay and conventional PCR. Phylogenetic analysis based on sequencing of the gyrB housekeeping gene of the isolates showed that they formed a unique clade, well characterised and separated from other xanthomonads. An insight into the genetic population features was attempted by rep-PCR analysis, using the ERIC, REP and BOX primers. The combined rep-PCR fingerprints showed a slight intra-pathovar variation within our isolates, which grouped in five close clusters. Copper resistance has been assessed in vitro for our whole X. arboricola pv. pruni collection, highlighting that two isolates show a level of resistance in vitro up to 200 ppm of copper. Nonetheless, the copLAB gene cluster, present in many other species of Xanthomonads, was not detected in any isolate, confirming the presence of a still unknown mechanism of copper detoxification in our Xanthomonads arboricola pv. pruni tolerant/resistant strains.