High nucleotide sequence variation of avirulent gene, AVR-Pita1, in Thai rice blast fungus population.

Rice blast disease, caused by Magnaporthe oryzae, is one of the most importance diseases of rice production worldwide. The keyrole of defense mechanism to combat this fungus in rice follows the gene-for-gene concept, which a plant resistant (R) gene product recognizes a fungal avirulent (AVR) effector and triggers the hypersensitive response. However, the AVR genes have been shown to be rapidly evolving resulting in high level of genetic diversity. The aims of this study were to examine the nucleotide sequence variation of AVR-Pita1 gene in Thai rice blast isolates and to identify the severity of blast disease using isogenic line of Pita gene. Seventy-six rice blast isolates collected from different parts of Thailand were used. Gene specific primers for AVR-Pita1 gene coding sequence were designed and used for identifying the genetic diversity of AVR-Pita1 gene by PCR amplification and sequencing. The obtained sequences were analysed for genetic variation and genetic relationship. Our results revealed the association between the sequence variations of AVR-Pita1 and selective forces from Pita gene. This phenomenon demonstrated the coevolution between rice blast resistant gene in rice and avirulent gene in blast fungus. The information about variation and evolutionary mechanisms of AVR gene obtained from this study can be used in rice blast resistant breeding programme.

Antimicrobial compounds from endophytic Streptomyces sp. BCC72023 isolated from rice (Oryza sativa L.).

An endophytic actinomycete strain BCC72023 was isolated from rice (Oryza sativa L.) and identified as the genus Streptomyces, based on phenotypic, chemotaxonomic and 16S rRNA gene sequence analyses. The strain showed 99.80% similarity compared with Streptomyces samsunensis M1463(T). Chemical investigation led to the isolation of three macrolides, efomycins M (1), G (2) and oxohygrolidin (3), along with two polyethers, abierixin (4) and 29-O-methylabierixin (5). To our knowledge, this is the first report of efomycin M being isolated from a natural source. The compounds were identified using spectroscopic techniques and comparison with previously published data. All compounds exhibited antimalarial activity against the Plasmodium falciparum, K-1 strain, a multidrug-resistant strain, with IC50 values in a range of 1.40-5.23 µg/ml. In addition, these compounds were evaluated for biological activity against Mycobacterium tuberculosis, Bacillus cereus, Colletotrichum gloeosporioides and Colletotrichum capsici, as well as cytotoxicity against both cancerous (MCF-7, KB, NCI-H187) and non-cancerous (Vero) cells.

Micromonospora soli sp. nov., isolated from rice rhizosphere soil.

An actinomycete strain SL3-70(T) was isolated from a rice field and characterised using a polyphasic approach. The morphological and chemotaxonomical characteristics of strain SL3-70(T) indicate that it belongs to the genus Micromonospora. The phylogenetic analysis of the nearly complete 16S rRNA gene sequence revealed that strain SL3-70(T) is a member of the genus Micromonospora, and is closely related to Micromonospora echinaurantica DSM 43904(T) (99.1 % 16S rRNA gene sequence similarity) and Micromonospora kangleipakensis MBRL 34(T) (98.8 %). DNA-DNA relatedness between strain SL3-70(T) and its relatives ranged from 21.2 % ± 0.6 to 38.7 % ± 0.4. The results obtained from our study indicate that strain SL3-70(T) represents a novel species of the genus Micromonospora, for which the name Micromonospora soli sp. nov. is proposed. The type strain is SL3-70(T) (=BCC 67268(T); =NBRC 110009(T)).

Micromonospora oryzae sp. nov., isolated from roots of upland rice.

An actinomycete strain, designated CP2R9-1T, was isolated from root internal tissues of upland rice (Oryza sativa). Based on a polyphasic approach, strain CP2R9-1T was characterized as a member of the genus Micromonospora. meso-Diaminopimelic acid and 3-OH-diaminopimelic acid were present in the cell-wall peptidoglycan. The polar lipids were diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylglycerol, phosphatidylinositol, phosphatidylinositol mannosides, two unidentified phospholipids and four unidentified polar lipids. Predominant menaquinones were MK-9(H4), MK-9(H6) and MK-10(H4). Whole-cell sugars consisted of ribose, xylose, arabinose and glucose. Phylogenetic analysis of the nearly complete 16S rRNA gene sequence suggested that strain CP2R9-1T was closely related to Micromonospora haikouensis 232617T (99.32 % similarity), Micromonospora carbonacea DSM 43168T (99.18 %) and Micromonospora krabiensis MA-2T (99.16 %). Strain CP2R9-1T was distinct from its closest relatives based on low levels of DNA-DNA relatedness (21.3 ± 0.1-41.7 ± 0.7 %) and phenotypic differences. The results presented in this study showed that strain CP2R9-1T represents a novel species of the genus Micromonospora, for which the name Micromonospora oryzae sp. nov. is proposed. The type strain is CP2R9-1T ( = BCC 67266T = NBRC 110007T).

Localization and expression of EDS5H a homologue of the SA transporter EDS5.

BACKGROUND: An important signal transduction pathway in plant defence depends on the accumulation of salicylic acid (SA). SA is produced in chloroplasts and the multidrug and toxin extrusion transporter ENHANCED DISEASE SUSCEPTIBILITY5 (EDS5; At4g39030) is necessary for the accumulation of SA after pathogen and abiotic stress. EDS5 is localized at the chloroplast and functions in transporting SA from the chloroplast to the cytoplasm. EDS5 has a homologue called EDS5H (EDS5 HOMOLOGUE; At2g21340) but its relationship to EDS5 has not been described and its function is not known. RESULTS: EDS5H exhibits about 72% similarity and 59% identity to EDS5. In contrast to EDS5 that is induced after pathogen inoculation, EDS5H was constitutively expressed in all green tissues, independently of pathogen infection. Both transporters are located at the envelope of the chloroplast, the compartment of SA biosynthesis. EDS5H is not involved with the accumulation of SA after inoculation with a pathogen or exposure to UV stress. A phylogenetic analysis supports the hypothesis that EDS5H may be an H(+)/organic acid antiporter like EDS5. CONCLUSIONS: The data based on genetic and molecular studies indicate that EDS5H despite its homology to EDS5 does not contribute to pathogen-induced SA accumulation like EDS5. EDS5H most likely transports related substances such as for example phenolic acids, but unlikely SA.

Micromonospora endophytica sp. nov., an endophytic actinobacteria of Thai upland rice (Oryza sativa).

An actinobacterial strain, DCWR9-8-2(T), was isolated from a leaf of Thai upland rice (Oryza sativa) collected in Chumporn province, Thailand. Strain DCWR9-8-2(T) is Gram-stain-positive aerobic bacteria that produce single spores directly on the vegetative hypha. Cell wall peptidoglycan of this strain exhibits meso-diaminopimelic acid and glycine, the reducing sugars of whole-cell hydrolysate are arabinose, glucose, ribose, xylose and small amount of mannose. The phospholipid profiles in the membrane are comprised of phosphatidylethanolamine, diphosphatidylglycerol, phosphatidylglycerol, phosphatidylinositol, phosphatidylinositol mannosides. The major menaquinones are MK-9(H4) and MK-10(H6). The diagnostic cellular fatty acids are iso-C16:0 and iso-C15:0. The G+C content of the genomic DNA is 72.5 mol%. The result of 16S rRNA sequence analysis of the strain revealed that this strain was closely related to Micromonospora auratinigra TT1-11(T) (99.25%). On the other hand, the result of gyrB gene sequence analysis revealed that this strain was closed to M. eburnea JCM 12345(T) (96.30%). In addition, a combination of DNA-DNA hybridization results and some phenotypic properties supported that this strain should be judged as a novel species of the genus Micromonospora, for which the name M. endophytica sp. nov. is proposed. The type strain is DCWR9-8-2(T) (=BCC 67267(T)=NBRC 110008(T)).

EDS5, an essential component of salicylic acid-dependent signaling for disease resistance in Arabidopsis, is a member of the MATE transporter family.

The eds5 mutant of Arabidopsis (earlier named sid1) was shown previously to accumulate very little salicylic acid and PR-1 transcript after pathogen inoculation and to be hypersusceptible to pathogens. We have isolated EDS5 by positional cloning and show that it encodes a protein with a predicted series of nine to 11 membrane-spanning domains and a coil domain at the N terminus. EDS5 is homologous with members of the MATE (multidrug and toxin extrusion) transporter family. EDS5 expression is very low in unstressed plants and strongly induced by pathogens and UV-C light. The transcript starts to accumulate 2 hr after inoculation of Arabidopsis with an avirulent strain of Pseudomonas syringae or UV-C light exposure, and it stays induced for approximately 2 days. EDS5 also is expressed after treatments with salicylic acid, indicating a possible positive feedback regulation. EDS5 expression after infection by certain pathogens as well as after UV-C light exposure depends on the pathogen response proteins EDS1, PAD4, and NDR1, indicating that the signal transduction pathways after UV-C light exposure and pathogen inoculation share common elements.