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Rice is responsible for the stable crop of 3 billion people worldwide, about half of Asian depends on it, and rice is grown in more than 100 countries. Rice diseases can lead to devastating economic loss by decreasing yield production, disturbing a stable food supply and demand chain. The most commonly used method to control rice disease is chemical control. However, misuse of chemical control can cause environmental pollution, residual toxicity, and the emergence of chemical-resistant pathogens, the deterioration of soil quality, and the destruction of biodiversity. In order to control rice diseases, research on alternative biocontrol is actively pursued including microorganism-oriented biocontrol agents. Microbial agents control plant disease through competition with and antibiotic effects and parasitism against plant pathogens. Microorganisms isolated from the rice rhizosphere are studied comprehensively as biocontrol agents against rice pathogens. Bacillus sp., Pseudomonas sp., and Trichoderma sp. were reported to control rice diseases, such as blast, sheath blight, bacterial leaf blight, brown spot, and bakanae diseases. Here we reviewed the microorganisms that are studied as biocontrol agents against rice diseases.

Heavy reliance on synthetic pesticides for crop protection has become increasingly unsustainable, calling for robust alternative strategies that do not degrade the environment and vital ecosystem services. There are numerous reports of successful disease control by various microbes used in small-scale trials. However, inconsistent efficacy has hampered their large-scale application. A better understanding of how beneficial microbes interact with plants, other microbes, and the environment and which factors affect disease control efficacy is crucial to deploy microbial agents as effective and reliable pesticide alternatives. Diverse metabolites produced by plants and microbes participate in pathogenesis and defense, regulate the growth and development of themselves and neighboring organisms, help maintain cellular homeostasis under various environmental conditions, and affect the assembly and activity of plant and soil microbiomes. However, research on the metabolites associated with plant health-related processes, except antibiotics, has not received adequate attention. This review highlights several classes of metabolites known or suspected to affect plant health, focusing on those associated with biocontrol and belowground plant–microbe and microbe–microbe interactions. The review also describes how new insights from systematic explorations of the diversity and mechanism of action of bioactive metabolites can be harnessed to develop novel crop protection strategies.

The genome of the rice blast fungus Magnaporthe oryzae contains several types of transposable elements (TEs), and some TEs cause genetic variation that allows M. oryzae to evade host detection. We studied how five abundant TEs in rice pathogens, Pot3, Pot2, MAGGY, Line-like element (MGL) and Mg-SINE, are expressed under diverse conditions related to growth, development, and stress. Expression of Pot3 and Pot2 was activated in germinated conidia and mycelia treated with tricyclazole. Retrotransposon MAGGY was highly expressed in appressoria and tricyclazole-treated mycelia. MAGGY and Pot2 were also activated during the early and late stages of perithecia development. MGL was up-regulated in conidia and during conidial germination but not during appressorium formation. No noticeable expression of Mg-SINE was observed under most conditions. Our results should help investigate if and how condition-specific expressions of some TEs contribute to the biology and evolution of M. oryzae.

An endolichenic fungus, Xylaria grammica strain EL000614, showed strong nematicidal effects against plant pathogenic nematode, Meloidogyne incognita by producing grammicin. We report genome assembly of X. grammica EL000614 comprised of 25 scaffolds with a total length of 54.73 Mb, N50 of 4.60 Mb, and 99.8% of BUSCO completeness. GC contents of this genome were 44.02%. Gene families associated with biosynthesis of secondary metabolites or regulatory proteins were identified out of 13,730 gene models predicted.

Microscopic observation of root disease onset and progression is typically performed by harvesting different plants at multiple time points. This approach prevents the monitoring of individual encounter sites over time, often mechanically damages roots, and exposes roots to unnatural conditions during observation. Here, we describe a method developed to avoid these problems and its application to study Fusarium oxysporum-Arabidopsis thaliana interactions. This method enabled three-dimensional, time-lapse imaging of both A. thaliana and F. oxysporum as they interact via the use of confocal and multi-photon microscopy and facilitated inquiries about the genetic mechanism underpinning Fusarium wilt.

The rice blast fungus, Magnaporthe oryzae, is one of the most economically important crop diseases. In addition, rice-M. oryzae interaction is a classical gene-for-gene host-pathogen system. Race variation in pathogen groups was proposed as the main mechanism for rapid break-down of resistance in newly introduced rice cultivars. These new pathogen race variations may be caused by changes in an avirulence gene, such as point mutations, insertion of transposons, and frame shifts. The avirulence gene AVR-Pita1 is representative avirulence gene in which all of these mutations are reported. In this review, we present a useful information for avirulence gene AVR-Pita1 and its homologous genes AVR-Pita2 and AVR-Pita3. We also review examples that cause mutations in these evolutionarily significant genes.

Pathogens utilize multiple types of effectors to modulate plant immunity. Although many apoplastic and cytoplasmic effectors have been reported, nuclear effectors have not been well characterized in fungal pathogens. Here, we characterize two nuclear effectors of the rice blast pathogen Magnaporthe oryzae. Both nuclear effectors are secreted via the biotrophic interfacial complex, translocated into the nuclei of initially penetrated and surrounding cells, and reprogram the expression of immunity-associated genes by binding on effector binding elements in rice. Their expression in transgenic rice causes ambivalent immunity: increased susceptibility to M. oryzae and Xanthomonas oryzae pv. oryzae, hemibiotrophic pathogens, but enhanced resistance to Cochliobolus miyabeanus, a necrotrophic pathogen. Our findings help remedy a significant knowledge deficiency in the mechanism of M. oryzae–rice interactions and underscore how effector-mediated manipulation of plant immunity by one pathogen may also affect the disease severity by other pathogens., Plant pathogens secrete various effectors to manipulate host immunity. Here, Kim et al. describe two Magnaporthe oryzae effectors that translocate into the nuclei of infected rice cells and reprogram expression of immunity-associated genes, increasing susceptibility to hemibiotrophic pathogens.

The fungus Aspergillus oryzae strain BP2-1 was isolated from the traditional malted starter culture nuruk. We report here the draft whole-genome sequence of A. oryzae BP2-1, which is comprised of 14 scaffolds with a total length of 39,455,382 bp and a GC content of 47.13%.

Acetylation of histone H3 lysine 56 (H3K56) by the fungal-specific histone acetyltransferase Rtt109 plays important roles in maintaining genome integrity and surviving DNA damage. Here, we investigated the implications of Rtt109-mediated response to DNA damage on development and pathogenesis of the rice blast fungus Magnaporthe oryzae (anamorph: Pyricularia oryzae). The ortholog of Rtt109 in M. oryzae (MoRtt109) was found via sequence homology and its functionality was confirmed by phenotypic complementation of the Saccharomyces cerevisiae Rtt109 deletion strain. Targeted deletion of MoRtt109 resulted in a significant reduction in acetylation of H3K56 and rendered the fungus defective in hyphal growth and asexual reproduction. Furthermore, the deletion mutant displayed hypersensitivity to genotoxic agents, confirming the conserved importance of Rtt109 in genome integrity maintenance and genotoxic stress tolerance. Elevated expression of DNA repair genes and the results of the comet assay were consistent with constitutive endogenous DNA damage. Although the conidia produced from the mutant were not impaired in germination and appressorium morphogenesis, the mutant was significantly less pathogenic on rice leaves. Transcriptomic analysis provided insight into the factors underlying phenotypic defects that are associated with deficiency of H3K56 acetylation. Overall, our results indicate that MoRtt109 is a conserved histone acetyltransferase that affects proliferation and asexual fecundity of M. oryzae through maintenance of genome integrity and response to DNA damage.

The red flour beetle, Tribolium castaneum, is one of the most common and economically important pests of stored cereal products worldwide. Furthermore, these beetles can act as vectors for several fungal post-harvest diseases. In this study, we collected T. castaneum from 49 rice processing complexes (RPCs) nationwide during 2016-2017 and identified contaminating fungal species on the surface of the beetles. Five beetles from each region were placed on potato dextrose agar media or Fusarium selection media after wet processing with 100% relative humidity at 27°C for one week. A total of 142 fungal isolates were thus collected. By sequence analysis of the internal transcribed spacer region, 23 fungal genera including one unidentified taxon were found to be associated with T. castaneum. The genus Aspergillus spp. (28.9%) was the most frequently present, followed by Cladosporium spp. (12.0%), Hyphopichia burtonii (9.2%), Penicillium spp. (8.5%), Mucor spp. (6.3%), Rhizopus spp. (5.6%), Cephaliophora spp. (3.5%), Alternaria alternata (2.8%) and Monascus sp. (2.8%). Less commonly identified were genera Fusarium, Nigrospora, Beauveria, Chaetomium, Coprinellus, Irpex, Lichtheimia, Trichoderma, Byssochlamys, Cochliobolus, Cunninghamella, Mortierella, Polyporales, Rhizomucor and Talaromyces. Among the isolates, two known mycotoxin-producing fungi, Aspergillus flavus and Fusarium spp. were also identified. This result is consistent with previous studies that surveyed fungal and mycotoxin contamination in rice from RPCs. Our study indicates that the storage pest, T. castaneum, would play an important role in spreading fungal contaminants and consequently increasing mycotoxin contamination in stored rice.

Background: Histamine in food is known to cause food poisoning and allergic reactions. We usually ingest histamine in cooked food, but there are few studies about the influence of cooking method on the histamine level. Objective: The purpose of this study was to determine the influence of cooking methods on the concentration of histamine in foods. Methods: The foods chosen were those kinds consumed frequently and cooked by grilling, boiling, and frying. The histamine level of the food was measured using enzyme-linked immunosorbent assay. Results: Grilled seafood had higher histamine levels than raw or boiled seafood. For meat, grilling increased the histamine level, whereas boiling decreased it. For eggs, there was not much difference in histamine level according to cooking method. Fried vegetables had higher histamine levels than raw vegetables. And fermented foods didn’t show much difference in histamine level after being boiled. Conclusion: The histamine level in food has changed according to the cooking method used to prepare it. Frying and grilling increased histamine level in foods, whereas boiling had little influence or even decreased it. The boiling method might be helpful to control the effect of histamine in histamine-sensitive or susceptible patients, compared with frying and grilling. (Ann Dermatol 29(6) 706∼714, 2017)

Incidence rates of diseases in kiwiberry orchards were investigated monthly from late June to late September in Gwangyang and Boseong in 2015 and 2016. The impact of postharvest fruit rot was investigated during ripening after harvest. Bacterial canker was only observed on one single tree in 2015, but black rot, powdery mildew, leaf spot and blight, and postharvest fruit rot diseases were problematic throughout the study period in both 2015 and 2016. Incidence rates of the diseases varied with kiwiberry cultivar, region and sampling time. Incidence rates of powdery mildew, leaf spot and blight diseases increased significantly during the late growing stages near fruit harvest, while black rot peaked in late August. Incidence rate of postharvest fruit rot on fruit without fruit stalks was less than half of fruit with fruit stalks, regardless of kiwiberry cultivars. Among the four cultivars, Mansu was relatively resistant to black rot and postharvest fruit rot diseases. In our knowledge, this is the first report of various potential pathogens of kiwiberry in Korea.

The fungal genus Cochliobolus describes necrotrophic pathogens that give rise to significant losses on rice, wheat, and maize. Revealing plant mechanisms of non-host resistance (NHR) against Cochliobolus will help to uncover strategies that can be exploited in engineered cereals. Therefore, we developed a heterogeneous pathosystem and studied the ability of Cochliobolus to infect dicotyledons. We report here that C. miyabeanus and C. heterostrophus infect Arabidopsis accessions and produce functional conidia, thereby demonstrating the ability to accept Brassica spp. as host plants. Some ecotypes exhibited a high susceptibility, whereas others hindered the necrotrophic disease progression of the Cochliobolus strains. Natural variation in NHR among the tested Arabidopsis accessions can advance the identification of genetic loci that prime the plant's defence repertoire. We found that applied phytotoxin-containing conidial fluid extracts of C. miyabeanus caused necrotic lesions on rice leaves but provoked only minor irritations on Arabidopsis. This result implies that C. miyabeanus phytotoxins are insufficiently adapted to promote dicot colonization, which corresponds to a retarded infection progression. Previous studies on rice demonstrated that ethylene (ET) promotes C. miyabeanus infection, whereas salicylic acid (SA) and jasmonic acid (JA) exert a minor function. However, in Arabidopsis, we revealed that the genetic disruption of the ET and JA signalling pathways compromises basal resistance against Cochliobolus, whereas SA biosynthesis mutants showed a reduced susceptibility. Our results refer to the synergistic action of ET/JA and indicate distinct defence systems between Arabidopsis and rice to confine Cochliobolus propagation. Moreover, this heterogeneous pathosystem may help to reveal mechanisms of NHR and associated defensive genes against Cochliobolus infection.

The Magnaporthe oryzae snodprot1 homolog (MSP1), secreted by M. oryzae, is a cerato-platanin family protein. msp1-knockout mutants have reduced virulence on barley leaves, indicating that MSP1 is required for the pathogenicity of rice blast fungus. To investigate the functional roles of MSP1 and its downstream signaling in rice, recombinant MSP1 was produced in Escherichia coli and was assayed for its functionality. Application of MSP1 triggered cell death and elicited defense responses in rice. MSP1 also induced H2O2 production and autophagic cell death in both suspension-cultured cells and rice leaves. One or more protein kinases triggered cell death, jasmonic acid and abscisic acid enhanced cell death, while salicylic acid suppressed it. We demonstrated that the secretion of MSP1 into the apoplast is a prerequisite for triggering cell death and activating defense-related gene expression. Furthermore, pretreatment of rice with a sublethal MSP1 concentration potentiated resistance to the pathogen. Taken together, our results showed that MSP1 induces a high degree of cell death in plants, which might be essential for its virulence. Moreover, rice can recognize MSP1, resulting in the induction of pathogen-associated molecular pattern-triggered immunity.

Isolates of the rice blast fungus show a range of tissue-specificities infecting leaves, nodes, neck and panicles. Although neck and panicle blast cause significantly greater yield losses than the leaf blast, virulence tests of the blast isolates have been performed only rice leaves instead of neck and panicles. In this study, we have developed a virulence test method for neck and panicle blast. We selected three representative isolates from each of leaf, neck, and panicle blast. We observed that severe disease lesions developed on the neck and the panicles when the infected rice plants were incubated in a dew chamber for 48 h instead of 24 h when tested on leaves. Unlike the leaf blast, a typical lesion on the neck and panicles appeared after 14 days post-infection as opposed to 7 days with leaf blast. This method will be applied to examine tissue-specificity of the rice blast fungus isolates.

Hot pepper (Capsicum annum L.) anthracnose has been a destructive disease in commercial pepper production fields (Harp et al., 2008; Lewis-Ivey et al., 2004; Park and Kim, 1992), and in Korea, the disease is estimated to cause a loss of more than US$100 million annually (Kim and Park, 1998). Furthermore, since the pepper anthracnose outbreak in 2002, this disease has received high priority (Kim et al., 2008). Several Colletotrichum spp., such as C. acutatum, C. coccodes, C. dematium, and C. gloeosporioides, thought to be causal agents of anthracnose (Park and Kim, 1992). However, more recently, C. acutatum was identified as the primary Colletotrichum species for anthracnose in pepper (Kim et al., 2008). Similar outbreaks of anthracnose on peppers have occurred in Ohio (Lewis-Ivey et al., 2004) and Florida, USA (Harp et al., 2008), and C. acutatum was identified as the primary Colletotricum species for anthracnose in pepper (Lewis-Ivey et al., 2004). C. acutatum (teleomorph: Glomerella acutata) is an important anthracnose pathogen on a wide range of host plants, causing significant economic loss in various crops, including apple, almond, citrus, strawberry, tomato, and hot pepper (Sutton, 1992; Freeman et al., 1998; Peres et al., 2005). Various fungicides have been identified to control C. acutatum, including copper compounds (e.g., copper hydroxide), the quinone outside inhibitors (azoxystrobin, trifloxystrobin, or pyraclostrobin), triazoles, dithiocarbamates, and benzimidazole compounds (Harp et al., 2014; Wedge et al., 2007). Notably, C. acutatum is tolerant to benomyl and other benzimidazole fungicides (Adaskaveg and Hartin, 1997; Peres et al., 2002; Talhinhas et al., 2002; Talhinhas et al., 2005). The azole fungicides are the most effective in inhibiting in vitro growth of C. acutatum (Paredes and Munoz, 2002), but the rapid development of fungicide resistant strains has limited their use. An understanding of the fungicide-resistant mechanisms will help to enhance control of anthracnose in pepper. Phytopathogenic fungi have developed various biological mechanisms that provide resistance to fungicides or abiotic stresses. Genes responsible for this resistance include ATP-binding cassette (ABC) transporters. For example, a gene deletion mutant of an ABC transporter, ABC1in Magnaporthe oryzae, showed hypersensitivity to several drugs (Urban et al., 1999). Moreover, ABC1 (Urban et al., 1999), ABC3 (Schneider and Hunke, 1998), ABC4 (Gupta and Chattoo, 2008), and ABC5 (Kim et al., 2013) from M. oryzae, GpABC1 from Gibberella pulicaris (Fleissner et al., 2002), Mgatr4 from Mycosphaerella graminicola (Stergiopoulos et al., 2003) and BcatrB from Botrytis cinerea (Schoonbeek et al., 2001) are required for pathogenicity. ABC-transporter proteins utilize energy derived from the hydrolysis of ATP to “pump” the substrate across a membrane, thus effectively reducing intracellular concentration to less toxic levels. The proteins are defined by the presence of amino acid sequences such as the ABC-ATPase domain, ABC domain, or nucleotide-binding domain. This domain contains the two peptide motifs Walker A (p-loop) and a hydrophobic Walker B motif (Walker et al., 1982). Both motifs are involved in ATP-binding proteins and identified as ATP signatures (Hyde et al., 1990). In addition, transmembrane domains are embedded in cell membranes that consist of at least six transmembranes. Until now, no ABC transporter genes have been isolated and characterized in C. acutatum. To begin defining the functional significance of the ABC transporter gene in C. acutatum, we are the first to identify a partial cDNA that encoded an ABC transporter, CaABC1, in C. acutatum. We also present the corresponding full-length gene structure of CaABC1 with the motifs. C. acutatum CaABC1 is most closely related to the ABC transporter {"type":"entrez-protein","attrs":{"text":"XP_007590216","term_id":"615436115","term_text":"XP_007590216"}}XP_007590216 of C. fioriniae. CaABC1 also shares a high degree of homology with the other Colletotrichum spp., including C. higginsianum, C. sublineola, C. graminicola, C. orbiculare, and C. gloeosporioides. CaABC1 was up-regulated in conidiation, abiotic stresses, and multiple fungicides. To our knowledge, this is the first structural and functional analysis of an ABC transporter gene in C. acutatum. Our results will provide the basis for further study on the function of ABC transporter genes in fungicide resistance and pathogenicity in C. acutatum.

Fusarium head blight (FHB) caused by the filamentous fungus Fusarium graminearum is one of the most severe diseases threatening the production of small grains. Infected grains are often contaminated with mycotoxins such as zearalenone and trichothecences. During survey of contamination by FHB in rice grains, we found a bacterial isolate, designated as BN1, antagonistic to F. graminearum. The strain BN1 had branching vegetative hyphae and spores, and its aerial hyphae often had long, straight filaments bearing spores. The 16S rRNA gene of BN1 had 100% sequence identity with those found in several Streptomyces species. Phylogenetic analysis of ITS regions showed that BN1 grouped with S. sampsonii with 77% bootstrap value, suggesting that BN1 was not a known Streptomyces species. In addition, the efficacy of the BN1 strain against F. graminearum strains was tested both in vitro and in vivo. Wheat seedling length was significantly decreased by F. graminearum infection. However, this effect was mitigated when wheat seeds were treated with BN1 spore suspension prior to F. graminearum infection. BN1 also significantly decreased FHB severity when it was sprayed onto wheat heads, whereas BN1 was not effective when wheat heads were point inoculated. These results suggest that spraying of BN1 spores onto wheat heads during the wheat flowering season can be efficient for plant protection. Mechanistic studies on the antagonistic effect of BN1 against F. graminearum remain to be analyzed.

Rice blast, caused by the pathogen Magnaporthe oryzae, is a serious hindrance to rice production and has emerged as an important model for the characterization of molecular mechanisms relevant to pathogenic development in plants. Similar to other pathogenic fungi, conidiation plays a central role in initiation of M. oryzae infection and spread over a large area. However, relatively little is known regarding the molecular mechanisms that underlie conidiation in M. oryzae. To better characterize these mechanisms, we identified a conidiation-defective mutant, ATMT0225B6 (MoCDC15T-DNA), in which a T-DNA insertion disrupted a gene that encodes a homolog of fission yeast cdc15, and generated a second strain containing a disruption in the same allele (ΔMoCDC15T-DNA). The cdc15 gene has been shown to act as a coordinator of the cell cycle in yeast. Functional analysis of the MoCDC15T-DNA and ΔMoCDC15T-DNA mutants revealed that MoCDC15 is required for conidiation, preinfection development and pathogenicity in M. oryzae. Conidia from these mutants were viable, but failed to adhere to hydrophobic surface, a crucial step required for subsequent pathogenic development. All phenotypic defects observed in mutants were rescued in a strain complemented with wild type MoCDC15. Together, these data indicate that MoCDC15 functions as a coordinator of several biological processes important for pathogenic development in M. oryzae.

The soilborne fungal pathogen Fusarium oxysporum causes vascular wilt and root rot diseases in many plant species. We investigated the role of cyclic AMP-dependent protein kinase A of F. oxysporum (FoCPKA) in growth, morphology, and root attachment, penetration, and pathogenesis in Arabidopsis thaliana. Affinity of spore attachment to root surfaces of A. thaliana, observed microscopically and measured by atomic force microscopy, was reduced by a loss-of-function mutation in the gene encoding the catalytic subunit of FoCPKA. The resulting mutants also failed to penetrate into the vascular system of A. thaliana roots and lost virulence. Even when the mutants managed to enter the vascular system via physically wounded roots, the degree of vascular colonization was significantly lower than that of the corresponding wild-type strain O-685 and no noticeable disease symptoms were observed. The mutants also had reduced vegetative growth and spore production, and their hyphal growth patterns were distinct from those of O-685. Coinoculation of O-685 with an focpkA mutant or a strain nonpathogenic to A. thaliana significantly reduced disease severity and the degree of root colonization by O-685. Several experimental tools useful for studying mechanisms of fungal root pathogenesis are also introduced.

Cauliflower (Brassica oleracea var. botrytis subvar. cauliflora) is susceptible to wilt caused by Verticillium dahliae but broccoli (B. oleracea var. italica subvar. cyamosa) is not. Infection of broccoli and cauliflower by a green fluorescent protein-expressing isolate of V. dahliae was examined using epifluorescence and confocal laser-scanning microscopy to follow infection and colonization in relation to plant phenology. Plant glucosinolate, phenolic, and lignin contents were also assayed at 0, 4, 14, and 28 days postinoculation. V. dahliae consistently infected and colonized the vascular tissues of all cauliflower plants regardless of age at inoculation, with the pathogen ultimately appearing in the developing seed; however, colonization decreased with plant age. In broccoli, V. dahliae infected and colonized root and stem xylem tissues of plants inoculated at 1, 2, or 3 weeks postemergence. However, V. dahliae colonized only the root xylem and the epidermal and cortical tissues of broccoli plants inoculated at 4, 5, and 6 weeks postemergence. The frequency of reisolation of V. dahliae from the stems (4 to 22%) and roots (10 to 40%) of mature broccoli plants was lower than for cauliflower stems (25 to 64%) and roots (31 to 71%). The mean level of aliphatic glucosinolates in broccoli roots was 6.18 times higher than in the shoots and did not vary with age, whereas it was 3.65 times higher in cauliflower shoots than in the roots and there was a proportional increase with age. Indole glucosinolate content was identical in both cauliflower and broccoli, and both indole and aromatic glucosinolates did not vary with plant age in either crop. Qualitative differences in characterized glucosinolates were observed between broccoli and cauliflower but no differences were observed between inoculated and noninoculated plants for either broccoli or cauliflower. However, the phenolic and lignin contents were significantly higher in broccoli following inoculation than in noninoculated broccoli or inoculated cauliflower plants. The increased resistance of broccoli to V. dahliae infection was related to the increase in phenolic and lignin contents. Significant differential accumulation of glucosinolates associated with plant phenology may also contribute to the resistant and susceptible reactions of broccoli and cauliflower, respectively, against V. dahliae.

Antioxidant and antibacterial activities of Phlomis umbrosa Turcz. (PU) ethanol extracts were investigated in vitro. Total polyphenols, flavonoids and FRAP values of PU extracts were /mg, /mg and 0.26 mmol /g, respectively. The PU extracts also showed strong DPPH scavenging activity and antibacterial effects against Staphylococcus aureus. In addition, the DPPH scavenging effect of the seasoning sauces was increased by the addition of PU extracts. This study was also conducted to evaluate the quality characteristics of seasoned pork treated with PU extracts during storage at for 21 days. Seasoned pork was produced containing 0, 0.5, and 1% PU extracts and 0.1% BHA. The total bacteria and TBARS values of seasoned pork added with PU were decreased compared to the control, whereas coliform bacteria were not detected after longer storage periods. With regards to sensory evaluation, seasoned pork treated with 0.5% PU had higher juiciness, flavor and overall acceptability compared to seasoned fork. These results suggest the possibility of manufacturing seasoned pork with PU extracts for the improvement of shelf-life and quality.

Department of Plant Pathology, The Pennsylvania State University, University Park 16802, USA(Received on September 27, 2010; Accepted on November 2, 2010)Genetic instability of the rice blast fungus Magnaportheoryzae has been suggested as a major factor underlyingthe rapid breakdown of host resistance in the field.However, little information is available on the mech-anism of genetic instability. In this study, we assessedthe stability of repetitive DNA elements and several keyphenotypic traits important for pathogenesis afterserially transferring two isolates though rice plants andan artificial medium. Using isolate 70-15, we obtained atotal of 176 single-spore isolates from 10 successiverounds of culturing on artificial medium. Another 20isolates were obtained from germ tubes formed at thebasal and apical cells of 10 three-celled conidia. Addi-tionally, 60 isolates were obtained from isolate KJ201after serial transfers through rice plants and an arti-ficial medium. No apparent differences in phenotypes,including mycelial growth, conidial morphologies, coni-diation, conidial germination, appressorium formation,and virulence, or in DNA fingerprints using MGR586,MAGGY, Pot2, LINE, MG-SINE and PWL2 as probeswere observed among isolates from the same parent iso-late. Southern hybridization and sequence analysis oftwo avirulence genes, AVR-Pita1 and AVR-Pikm, show-ed that both genes were also maintained stably during10 successive generations on medium and plants. How-ever, one reversible loss of restriction fragments wasfound in the telomere-linked helicase gene ( TLH1) family,suggesting some telomere regions may be more unstablethan the rest of the genome. Taken together, our resultssuggest that phenotype and genotype of M. oryzae iso-lates do not noticeably change, at least up to 10 successivegenerations on a cultural medium and in host plants.Keywords :genetic stability, genotypes, Magnaporthe grisea,phenotypes, repeat sequencesMagnaporthe oryzae is a heterothallic ascomycete fungusthat causes rice blast, one of the most devastating diseasesin rice throughout the world. Since there is no evidence forsexual reproduction in the field in most parts of the world,asexual spores (conidia) are the most important source ofinoculum. Through the production of conidia, M. oryzaenormally undergoes 8-11 disease cycles in rice plants underfield conditions during one growing season (Han et al.,1997). Although asexual reproduction is expected to pro-duce genetically identical clones, the high rate of appear-ance of new races with the ability to infect previouslyresistant varieties of rice, suggests that M. oryzae may havehigh mutation rates in avirulence genes, which determinerace (also called pathotype in M. oryzae) (Kang et al., 2001;Kang and Lee, 2000; Zhou et al., 2007).Although the most effective control strategy for rice blastis cultivating resistant varieties, it has often achieved onlyshort-term success due to frequent breakdown of newlyintroduced resistance. This resistance breakdown has beenattributed to genetic variability in M. oryzae (Bonman et al.,1986; Correa-Victoria and Zeigler, 1993; Correa-Victoriaand Zeigler, 1993). Various mechanisms have been sug-gested to explain the frequent generation of race variants,including heterokaryosis, parasexuality, and mutations(Genovesi and Magill, 1976; Wu and Magill, 1995; Xia andCorrell, 1995; Zeigler et al., 1997). Parasexual recombina-tion of genetic markers, resulting new genetic variants, hasbeen demonstrated in the laboratory for M. oryzae, sug-gesting the potential for parasexual recombination togenerate new races (Noguchi et al., 2006; Zeigler et al.,1997). However, no evidence for parasexuality has beenfound in nature for M. oryzae. The activity of transposableDNA elements is another possible mechanism that has beensuggested (Kang et al., 2001; Nishimura et al., 2000; Zhouet al., 2007). For example, mobilization of Pot2, Pot3, andLINE elements has been shown to cause variation by insert-ing into promoter region or coding sequences of avirulence

A Gram-negative rhizobacterial isolate (LSW25) antagonistic to Pseudomonas corrugata, a vein necrosis pathogen of tomato, and promotes the growth of tomato seedlings was isolated from surface-sterilised tomato roots. A spontaneous rifampicin-resistant mutant (LSW25R) was selected to facilitate its tracking, and identified as Pseudomonas sp. and named as Pseudomonas sp. LSW25R (LSW25R), based on its sequences of the internal transcribed spacer (ITS) region and 16 S rRNA gene. LSW25R inhibited mycelial growth of 12 other plant fungal pathogens such as Botrytis cinerea on V8 agar plates. By using a scanning electron microscope, LSW25R colonised not only the root surface around the natural aperture of tomato radicles but also under epidermal cells like endophytic bacteria. LSW25R successfully colonised the roots of tomato, eggplant and pepper seedlings, significantly promoted the fresh weight, height and dry matter of tomato plants at 108 cfu·ml−1, and increased the plant growth of eggplants and peppers at 104 cfu·ml−1, suggesting that the optimal population density of LSW25R for growth promotion varies from species to species. Moreover, densities of LSW25R inside roots and the lowest leaf of tomato plants were > 9.3 × 103 cfu·g−1. Although the growth promotion of tomato by LSW25R was observed under N- or Ca-deficient conditions as well as a standard nutrient condition, the uptake of calcium was increased only under the standard nutrient condition. In a hydroponic system, LSW25R not only successfully colonised the rhizosphere during cultivation due to its broad spectrum of antifungal activity and endophytic colonisation, but also reduced blossom-end rot of tomato fruits presumably through increasing calcium uptake.

In planta secretion of fungal pathogen proteins, including effectors destined for the plant cell cytoplasm, is critical for disease progression. However, little is known about the endoplasmic reticulum (ER) secretion mechanisms used by these pathogens. To determine if normal ER function is crucial for fungal pathogenicity, Magnaporthe oryzae genes encoding proteins homologous to yeast Lhs1p and Kar2p, members of the heat shock protein 70 family in Saccharomyces cerevisiae, were cloned and characterized. Like their yeast counterparts, both LHS1 and KAR2 proteins localized in the ER and functioned in an unfolded protein response (UPR) similar to the yeast UPR. Mutants produced by disruption of LHS1 were viable but showed a defect in the translocation of proteins across the ER membrane and reduced activities of extracellular enzymes. The Δlhs1 mutant was severely impaired not only in conidiation, but also in both penetration and biotrophic invasion in susceptible rice (Oryza sativa) plants. This mutant also had defects in the induction of the Pi-ta resistance gene–mediated hypersensitive response and in the accumulation of fluorescently-labeled secreted effector proteins in biotrophic interfacial complexes. Our results suggest that proper processing of secreted proteins, including effectors, by chaperones in the ER is requisite for successful disease development and for determining host-pathogen compatibility via the gene-for-gene interaction.

The avirulence (AVR) gene AVR-Pita in Magnaporthe oryzae prevents the fungus from infecting rice cultivars containing the resistance gene Pi-ta. A survey of isolates of the M. grisea species complex from diverse hosts showed that AVR-Pita is a member of a gene family, which led us to rename it to AVR-Pita1. Avirulence function, distribution, and genomic context of two other members, named AVR-Pita2 and AVR-Pita3, were characterized. AVR-Pita2, but not AVR-Pita3, was functional as an AVR gene corresponding to Pi-ta. The AVR-Pita1 and AVR-Pita2 genes were present in isolates of both M. oryzae and M. grisea, whereas the AVR-Pita3 gene was present only in isolates of M. oryzae. Orthologues of members of the AVR-Pita family could not be found in any fungal species sequenced to date, suggesting that the gene family may be unique to the M. grisea species complex. The genomic context of its members was analyzed in eight strains. The AVR-Pita1 and AVR-Pita2 genes in some isolates appeared to be located near telomeres and flanked by diverse repetitive DNA elements, suggesting that frequent deletion or amplification of these genes within the M. grisea species complex might have resulted from recombination mediated by repetitive DNA elements.

A previous study of the diversity and population structure of the rice blast fungus, Magnaporthe oryzae, over a 20-year period in Korea, found novel fingerprint haplotypes each year, and the authors hypothesized that populations might experience annual bottlenecks. Based on this model, we predicted that M. oryzae populations would have little or no genetic differentiation among geographic regions because rice blast is commonly found throughout Korea each year and M. oryzae would have to disperse from small populations surviving annually between rice crops. To test this hypothesis, we sampled M. oryzae from rice fields in eight provinces in Korea in a single year (1999). In four provinces, we sampled from a set of rice cultivars commonly grown in commercial fields (group I); because of low disease incidence in four other provinces, we could not sample from commercial fields and instead sampled from scouting plots of different cultivars set up for detecting new pathotypes of M. oryzae (group II). All isolates were genotyped with DNA fingerprint probes MGR586 and MAGGY, a telomere-linked gene family member TLH1, the PWL2 host specificity gene and mating type. Fingerprint haplotypes clustered into two distinct lineages corresponding to the two sets of cultivars (groups I and II), with haplotype similarities of 71% between lineages and >76% within lineages. Isolates from the same cultivar within group I were genetically differentiated among locations, and isolates within the same location were differentiated among cultivars. Differentiation for TLH1 and PWL2 was significant (P < 0.03), but not as strong as for fingerprint markers. Similar analyses were not possible among group II isolates because too few isolates were available from any one cultivar. All isolates were in the same mating type, Mat1-1, ruling out sexual reproduction as a source of novel haplotypes. When the 1999 samples were compared with the historical samples from the previous study, haplotypes of group I formed a separate cluster, while those of group II clustered with haplotypes from the historical sample. Altogether, geographic subdivision, monomorphism of mating type, and correlation of haplotypes to sets of cultivars are not consistent with the hypothesis of repeated turnover of haplotypes. Instead, the previous correlations of haplotypes to year might have been caused by inadequate sampling of haplotypes each year, highlighting the need for studies of population genetics to be conducted with systematic samples collected to address specific questions.

Phytophthora species are devastating plant pathogens in both agricultural and natural environments. Due to their significant economic and environmental impact, there has been increasing interest in Phytophthora genetics and genomics, culminating in the recent release of three complete genome sequences (P. ramorum, P. sojae, and P. infestans). In this study, genome and other large sequence databases were used to identify over 225 potential genetic markers for phylogenetic analyses. Here, we present a genus-wide phylogeny for 82 Phytophthora species using seven of the most informative loci (approximately 8700 nucleotide sites). Our results support the division of the genus into 10 well-supported clades. The relationships among these clades were rigorously evaluated using a number of phylogenetic methods. This is the most comprehensive study of Phytophthora relationships to date, and many newly discovered species have been included. A more resolved phylogeny of Phytophthora species will allow for better interpretations of the overall evolutionary history of the genus.

An insertion sequence identified as a solo long terminal repeat (LTR) of a new rice copia-like retrotransposon was detected in the ORE of the Pi-b gene from the rice cv. Nipponbare, and was designated as Osr1. Osr1 consists of a 6386 bp nucleotide sequence including 965 bp LTRs on both ends with an 82％ nucleotide sequence identity to the wheat Tarl retrotransposon on reverse transcriptase. Nucleotide divergence was noted among the individual LTRs, as well as the coding region of Osr1. Various restriction fragment length polymorphism (RFLP) of LTR were detected in indica cultivars, whereas, only a few could be detected in the japonica cultivars. The population of Osr1 is lower in the wild-type rice compared with that in the domesticated cultivars. The insertion of LTR sequence in the h-b gene in the susceptible cultivar suggested that retro-tyansposon-mediated insertional mutation might play an important role in the resistance breakdown, as well as in the evolution of resistance genes in rice.

Sequence analysis of a 13-kb telomeric region in O-137, a rice pathogenic isolate of Magnaporthe grisea, uncovered a novel gene, designated TLH1 (telomere-linked helicase 1). The TLH1 gene is a member of a gene family, and the sequences flanking this gene family have also been amplified. Genetic mapping showed that most members of the TLH gene family are tightly linked to the telomeres. A physical mapping technique, termed RecA-mediated Achilles’ heel cleavage, and cloning and sequencing of two additional telomeres of O-137 associated with the TLH gene family confirmed that most members of the TLH gene family are located within 10 kb from the telomeric repeat. A survey of M. grisea strains from diverse hosts revealed that the gene family is ubiquitously present among rice pathogens, but is absent from almost all isolates of hosts other than rice. The gene family appears to be highly dynamic, undergoing frequent deletion/amplification events. Given the presence of similar helicase gene families in chromosome ends of Saccharomyces cerevisiae and Ustilago maydis, the initial association of helicase genes with fungal telomeres might date back to very early stages of the fungal evolution.

Rice blast disease caused by Magnaporthe oryzae is one of the most devastating fungal diseases of rice and results in a huge loss of rice productivity worldwide. During the infection process, M. oryzae secretes a large number of glycosyl hydrolase proteins into the host apoplast to digest the cell wall and facilitate fungal ingression into host tissues. In this study, we identified a novel arabinofuranosidase-B (MoAbfB) protein that is secreted by M. oryzae during fungal infection. Deletion of MoAbfB from M. oryzae resulted in reduced disease severity in rice. Biochemical assays revealed that the MoAbfB protein exhibited arabinofuranosidase activity and caused degradation of rice cell wall components. Interestingly, pre-treatment of rice with the MoAbfB protein inhibited fungal infection by priming defence gene expression. Our findings suggest that MoAbfB secretion affects M. oryzae pathogenicity by breaking down the host cell wall, releasing oligosaccharides that may be recognized by the host to trigger innate immune responses.

Outbreaks of rice blast have been a threat to the global production of rice. Members of the Magnaporthe grisea species complex cause blast disease on a wide range of gramineous hosts, including cultivated rice and other grass species. Recently, based on phylogenetic analyses and mating tests, isolates from crabgrass were separated from the species complex and named M. grisea. Then other isolates from grasses including rice were named as M. oryzae. Here, we collected 103 isolates from 11 different species of grasses in Korea and analyzed their phylogenetic relationships and pathogenicity. Phylogenetic analyses of multilocus sequences and DNA fingerprinting revealed that the haplotypes of most isolates were associated with their hosts. However, six isolates had different haplotypes from the expectation, suggesting potential host shift in nature. Results of pathogenicity tests demonstrated that 42 isolates from crabgrass and 19 isolates from rice and other grasses showed cross-infectivity on rice and crabgrass, respectively. Interestingly, we also found that the isolates from rice had a distinct deletion in the calmodulin that can be used as a probe.

Previously, we showed that AMP-activated protein kinase (AMPK) negatively regulates receptor activator of nuclear factor-κB ligand-induced osteoclast formation in vitro. The present study investigated the effect of (-)-epigallocathechin-3-gallate (EGCG), an AMPK activator, on ovariectomy (OVX)-induced bone loss in mice. Female mice subjected to OVX were administered EGCG for 8 weeks. We measured total-body bone mineral density (BMD) before and after the operation at an interval of 4 weeks. We performed micro-computed tomography (micro-CT) of the tibia and bone histomorphometric examination of the femur. Western blot analysis was additionally performed, to detect levels of the phosphorylated and total forms of AMPK-α in calvarial extracts. EGCG prevented OVX-induced body weight gain. The OVX control did not show a significant increase in BMD values at baseline and after treatment, unlike the sham control. EGCG attenuated OVX-induced bone loss. Micro-CT experiments revealed that EGCG induced a significant increase in trabecular bone volume and trabecular number and a decrease in trabecular spacing compared to the OVX control. Histomorphometric analyses further showed that EGCG suppressed osteoclast surface and number. Phosphorylated AMPK expression was significantly elevated in bone following EGCG treatment. Our findings collectively indicate that EGCG decreases OVX-induced bone loss via inhibition of osteoclasts.

Knowledge remains limited about how fungal pathogens that colonize living plant cells translocate effector proteins inside host cells to regulate cellular processes and neutralize defense responses. To cause the globally important rice blast disease, specialized invasive hyphae (IH) invade successive living rice (Oryza sativa) cells while enclosed in host-derived extrainvasive hyphal membrane. Using live-cell imaging, we identified a highly localized structure, the biotrophic interfacial complex (BIC), which accumulates fluorescently labeled effectors secreted by IH. In each newly entered rice cell, effectors were first secreted into BICs at the tips of the initially filamentous hyphae in the cell. These tip BICs were left behind beside the first-differentiated bulbous IH cells as the fungus continued to colonize the host cell. Fluorescence recovery after photobleaching experiments showed that the effector protein PWL2 (for prevents pathogenicity toward weeping lovegrass [Eragrostis curvula]) continued to accumulate in BICs after IH were growing elsewhere. PWL2 and BAS1 (for biotrophy-associated secreted protein 1), BIC-localized secreted proteins, were translocated into the rice cytoplasm. By contrast, BAS4, which uniformly outlines the IH, was not translocated into the host cytoplasm. Fluorescent PWL2 and BAS1 proteins that reached the rice cytoplasm moved into uninvaded neighbors, presumably preparing host cells before invasion. We report robust assays for elucidating the molecular mechanisms that underpin effector secretion into BICs, translocation to the rice cytoplasm, and cell-to-cell movement in rice.

International audience; Fusarium species are among the most important phytopathogenic and toxigenic fungi. To understand the molecular underpinnings of pathogenicity in the genus Fusarium, we compared the genomes of three phenotypically diverse species: Fusarium graminearum, Fusarium verticillioides and Fusarium oxysporum f. sp. lycopersici. Our analysis revealed lineage-specific (LS) genomic regions in F. oxysporum that include four entire chromosomes and account for more than one-quarter of the genome. LS regions are rich in transposons and genes with distinct evolutionary profiles but related to pathogenicity, indicative of horizontal acquisition. Experimentally, we demonstrate the transfer of two LS chromosomes between strains of F. oxysporum, converting a non-pathogenic strain into a pathogen. Transfer of LS chromosomes between otherwise genetically isolated strains explains the polyphyletic origin of host specificity and the emergence of new pathogenic lineages in F. oxysporum. These findings put the evolution of fungal pathogenicity into a new perspective.

Using isolates collected over 2 decades, we determined the population structure and dynamics of the rice blast fungus, Magnaporthe grisea, in Korea at both the genotypic and phenotypic levels. Pathotype analysis on 6,315 isolates collected from 328 rice cultivars from 1981 to 2000 revealed the presence of a total of 91 pathotypes. Among these 91 patho-types, nine dominated, comprising 76.5% of the isolates. The expected number of pathotypes (corrected for sample size) increased significantly during the course of this study. On average, six (ranging from 0 to 20) new commercial cultivars were introduced annually between 1981 and 1998. However, the overall cultivar diversity, estimated using the Shannon index, was low. Most of the new cultivars were not planted to a large area because the seven most common cultivars each year occupied over 70% of the rice-cultivated area. The frequencies of the nine dominant patho-types from these seven cultivars were highly correlated with those from the entire set of cultivars. To understand genetic diversity within and between pathotypes, 176 isolates collected from 1984 to 1999 were randomly sampled and analyzed by DNA fingerprinting. High similarities were observed among isolates; overall similarities were greater than 63% in combined MGR586 and MAGGY DNA fingerprints. Unlike most other populations of M. grisea, DNA fingerprints showed no clear lineage structure. No groups were supported by bootstrap values greater than 10%. Furthermore, there was no significant correlation between DNA fingerprint similarities and pathotypes. Genetic similarity was significantly greater (P < 0.001) within years than between years, although the difference was small. Our data suggest that M. grisea populations in Korea have been mostly dominated by a single clonal lineage. We cannot conclude from these data that selection by the host population has been a major force in the evolution of M. grisea in Korea.

Fusarium oxysporum, a major soil-borne fungal pathogen, causes vascular wilt, damping-off, and root rot diseases on over 100 cultivated plant species. Mechanisms of root colonization by F. oxysporum in Arabidopsis thaliana were studied through in planta 3-dimensional time-lapse documentation using confocal and multi-photon microscopy. Data from individual encounter sites were acquired repeatedly over a several day period without physical manipulation or retrieval from the growth chamber. In vivo observations were facilitated by transformation of F. oxysporum for constitutive cytoplasmic expression of the fluorescent protein ZsGreen, and host responses were monitored using autofluorescence or GFP-tagged endoplasmic reticulum. Penetration into the vascular system occurred primarily in the meristematic region of primary and lateral roots. Fungal hyphae may release phytotoxin(s) that compromise host cells not directly in contact with hyphae. This novel approach was essential for visualizing the dynamic interactions between F. oxysporum and A. thaliana from both the host and pathogen sides.

Mitogen-activated protein kinase (MAPK) cascade(s) is important for plant defense/stress responses. Though MAPKs have been identified and characterized in rice (Oryza sativa L.), a monocot cereal crop research model, the first upstream component of the kinase cascade, namely MAPK kinase kinase (MAPKKK) has not yet been identified. Here we report the cloning of a novel rice gene encoding a MAPKKK, OsEDR1, designated based on its homology with the Arabidopsis MAPKKK, AtEDR1. OsEDR1, a single copy gene in the genome of rice, encodes a predicted protein with molecular mass of 113046.13 and a pI of 9.03. Using our established two-week-old rice seedling in vitro model system, we show that OsEDR1 has a constitutive expression in seedling leaves and is further up-regulated within 15 min upon wounding by cut, treatment with the global signals jasmonic acid (JA), salicylic acid (SA), ethylene (ethephon, ET), abscisic acid, and hydrogen peroxide. In addition, protein phosphatase inhibitors, fungal elicitor chitosan, drought, high salt and sugar, and heavy metals also dramatically induce its expression. Moreover, OsEDR1 expression was altered by co-application of JA, SA, and ET, and required de novo synthesized protein factor(s) in its transient regulation. Furthermore, using an in vivo system we also show that OsEDR1 responds to changes in temperature and environmental pollutants-ozone and sulfur dioxide. Finally, OsEDR1 expression varied significantly in vegetative and reproductive tissues. These results suggest a role for OsEDR1 in defense/stress signalling pathways and development.

Recent investigation of bacteria isolated from samples submitted to the Plant Disease Diagnostic Laboratory, Pennsylvania Department of Agriculture indicated that in 1995, Xanthomonas gardneri (ex Sutic 1957) (2) caused a leaf spot on tomato plants (Lycopersicon esculentum Mill.). In 1995, we examined 185 tomato and 36 pepper samples (13 field, 2 garden center, 38 greenhouse, 4 residence, 16 field-grown transplant, and 148 greenhouse-grown transplant samples). A processing company representative collected samples showing symptoms of bacterial spot of tomato on a hybrid, whole pack processing tomato, from a 16-ha field in Northumberland County, PA exhibiting almost 50% crop infection. Symptoms consisted of circular- to irregularly shaped, dark brown spots, 8 CFU/ml. Lower leaf surfaces of tomato (cvs. Bonnie Best and Walter) and pepper (cvs. California Wonder and Early Niagara) plants were gently rubbed with sterile cheesecloth that was moistened with the inoculum. Strain PDA80951-95 caused leaf spots, with chlorotic haloes and occasional coalescence on both tomato and pepper, within 2 weeks at 15 s of mist per 20 min at 20 to 35°C in a secured greenhouse chamber. X. gardneri was only reisolated from symptomatic plants and its identity was confirmed by rep-PCR and absence of amylolytic and pectolytic activities. Negative controls consisting of X. campestris pv. campestris and sterile tap water did not show symptoms. A known type strain of X. gardneri was not included as a positive control for pathogenicity studies because this species is not known to occur in the United States (2). To our knowledge, this is the first report of bacterial spot on tomato plants caused by X. gardneri in Pennsylvania and the United States. Since the first occurrence in 1995, bacterial spot caused by X. gardneri reoccurred in Pennsylvania tomato fields in 2001 and consecutively from 2003 to 2009. Reference: (1) D. A. Cuppels et al. Plant Dis. 90:451, 2006. (2) J. B. Jones et al. Syst. Appl. Microbiol. 27:755, 2004. (3) A. M. Quezado-Duval et al. Plant Dis. 88:15, 2004. (4) D. J. Versalovic et al. Methods Mol. Cell Biol. 5:25, 1994.

Black pod or Phytophthora pod rot is the most economically important and widespread disease of cacao, Theobroma cacao L. Total losses due to Phytophthora exceed $400 million worldwide (1), and several species are known to attack cacao with P. palmivora (E.J. Butler) E. J. Butler as the most common. All plant parts are infected, but pod infections are particularly damaging. Symptoms resembling those of black pod disease were observed at the National Plant Germplasm Collection System of cacao at the USDA-ARS Tropical Agriculture Research Station (TARS) in Mayaguez, Puerto Rico for a number of years. During May of 2005, to determine the etiology of the disease, small, surface disinfested sections of pod lesions were placed on water agar and incubated for 4 days. The formation of papillate, deciduous, ellipsoidal to ovoid sporangia produced on sympodial sporangiophores on fruits, fit the description of P. palmivora and the identification was confirmed on cultures on water agar (2). Chlamydospores were readily observed in diseased pods and observed in pure cultures on V8 agar (2). Eight, single hyphal tips were transferred to potato dextrose agar (PDA) (Sigma-Aldrich, St. Louis, MO) and maintained as stock cultures. For pathogenicity tests, healthy mature pods were surface disinfested and placed in a humidity chamber lined with moist paper towels. Eight isolates were tested on four fruits per isolate and the pathogenicity test was repeated once. Inoculum was prepared by growing each isolate on PDA for 5 days with irradiation at 24°C, adding approximately 3.0 ml of water to each plate, dislodging the sporangia with a glass rod, mixing the suspension, estimating spore numbers with a hemacytometer, and adjusting to 104 sporangia per ml. A small, sterile scalpel was used to make an approximately 20.0 mm cut on the fruit epidermis, and approximately 0.2 ml of inoculum was placed on the wound. Pods were evaluated daily for 2 weeks. For molecular analysis, each of the eight cultures were grown in 50% potato dextrose broth to produce mycelia for DNA extraction using the FastDNA kit (Q-Biogen1, Irvine, CA). The internal transcribed spacer (ITS) region of the ribosomal RNA gene cluster was amplified, purified, and sequenced for all eight isolates. The ITS sequences of GenBank Accession Nos. DQ987915 to DQ987922 were identical and exhibited strong similarity (>99% identity) to that of three previously described isolates of P. palmivora from cacao (GenBank Accession Nos. AF 228097, AF467093, and AF467089). P. palmivora has been reported on citrus, coconut, black pepper, and Arracacia xanthorrhiza in Puerto Rico (2,3) and inoculum may have originated from these host or imported on cacao planted into the cacao collection before 2000. USDA-ARS-TARS is the official site for the cacao germplasm collection, thus, a detailed integrated pest management plan that includes the evaluation for resistance, sanitation measures, and use of fungicides to reduce disease levels has been implemented. Decreasing incidence and severity of this disease is a top priority. To our knowledge, this is the first report of P. palmivora on cacao in Puerto Rico. References: (1) M. C. T. Braga et al. Agrotropica 1:108, 1989. (2) D. Erwin and O. K. Ribeiro. Phytophthora Diseases Worldwide. The American Phytopathological Society. St. Paul, MN, 1996. (3) E. Rosa-Marquez. J. Agric. Univ. P. R. 84:53, 2000.

Regulation of gene expression by transcription factors (TFs) helps plant pathogens to interact with the host plants and to sustain a pathogenic lifestyle in the environmental changes. Elucidating novel functions of TFs is, therefore, crucial for understanding pathogenesis mechanisms of plant pathogens. Magnaporthe oryzae, the rice blast pathogen, undergoes a series of developmental morphogenesis to complete its infection cycle. To understand TF genes implicated in pathogenic development of this fungus, two Zn(II)2Cys6 TF genes, MoCOD1 and MoCOD2, whose expression was notably induced during conidiation, were functionally characterized. Targeted deletion of MoCOD1 resulted in defects in conidiation and pathogenicity due to defects in appressorium formation and invasive growth within the host cells. MoCOD2 was also a critical regulator in conidiation and pathogenicity, but not in conidial germination and appressorium formation. When rice plants were inoculated with conidia of the ΔMocod2 mutant, rapid accumulation of dark brown granules was observed around the infection sites in the plant cells and no visible disease symptom was incited. Taken together, both MoCOD1 and MoCOD2 play important roles in conidiation and pathogenicity of the rice blast fungus.