Role of bacterial pyrroloquinoline quinone in phosphate solubilizing ability and in plant growth promotion on strain <Emphasis Type="Italic">Serratia</Emphasis> sp. S119

The aim of this study was to analyze if cofactor pyrroquinoline quinone from Serratia sp. S119 is involved in the inorganic phosphate solubilization mechanism and in its ability to promote the plant growth. Site directed mutagenesis was performed to obtain a pqqE- minus mutant of strain Serratia sp. S119. The phosphate solubilization ability, gluconate and PQQ production of the mutant Serratia sp. RSL (pqqE-) was analyzed. Mutant RSL (pqqE-) showed significant decrease in P soluble and gluconic acid levels produced and undetectable levels of PQQ cofactor compared with wild-type strain. Complementation with synthetic PQQ cofactor restored P solubilization and gluconate production reaching the levels produced by wild-type strain. PqqE gene sequence indicated that it is highly conserved within Serratia strains and its product shows conserved motifs found in other PqqE proteins of several bacteria. The effect of the inoculation of the PQQ- mutant on peanut and maize plants was evaluated in pot assays. Plants growth parameters showed no differences among the different treatments indicating that PQQ from Serratia sp. S119 is not involved in the growth promotion of these plants. PQQ cofactor is essential for phosphate solubilization ability of Serratia sp. S119 but is not required for growth promotion of peanut and maize plants.     

Expression and characteristics of rice xylanase inhibitor OsXIP,a member of a new class of antifungal proteins

It has been hypothesized that xylanase inhibitors play important roles in plant defense against microbial pathogens. Currently, there is little information available about xylanase inhibitor OsXIP in rice and its gene expression. We cloned a xylanase inhibitor gene OsXIP from rice (Oryza sativa L. cv. Nipponbare) genomic DNA. To determine the function of OsXIP, we generated OsXIP-overexpressing transgenic rice plants. The transgenic plants had significantly higher OsXIP expression and showed enhanced defense response to Magnaporthe oryzae compared to the wild-type plants. The results also showed that the increased OsXIP expression was accompanied by the up-regulation of pathogenesisrelated genes. To clarify the OsXIP expression pattern, a ProOsXIP::GUS vector was constructed and transgenic plants were obtained. GUS staining results revealed that OsXIP showed organ-specific expressions in rice plants. OsXIP was primarily expressed in the roots and in the veins, but it was weakly expressed in the leaves. Analyses of the OsXIP expression in response to biotic and abiotic stresses indicated that it was drastically induced by biotic stresses and methyl jasmonate treatment. OsXIP, a member of a new class of antifungal proteins, may function as a barrier that prevents the cell wall degradation by xylanases excreted by fungal pathogens. The OsXIP was found to be a stressresponsive gene and it could take part in plant defense via a JA-mediated signaling pathway.     

Quantitative proteomic analyses reveal that RBBI3.3, a trypsin inhibitor protein,plays an important role in <Emphasis Type="Italic">Magnaporthe oryzae</Emphasis> infection in rice

Magnaporthe oryzae (M. oryzae) is the causative agent of rice blast, the most destructive rice disease in China. This study was designed to ascertain the molecular mechanisms of the response of rice to M. oryzae infection to facilitate the breeding of new high-quality and disease-resistant rice varieties using isobaric tags for relative and absolute quantification (iTRAQ) combined with a high-throughput mass spectrometry identification platform. M. oryzae infection models were constructed with the resistant rice cultivar Gumei2 and the non-resistant cultivar Lijiangxintuanheigu (LTH). The results showed that total of 1541 proteins were identified, among which 843 proteins were overlapping between the two biological replicates analyses. Seventy-one proteins were classified as fungi-responsive. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses revealed the important roles of these proteins in metabolic processes. Detailed phenotypic analyses revealed that the trypsin inhibitor RBBI3.3 was effective in inhibiting the initial formation of appressoria. Our quantitative proteomic study provides insights into the molecular mechanism underlying M. oryzae resistance in the incompatible rice Gumei2. The identification of RBBI3.3 as a key defense regulator highlights a new possibility for disease control.     

The Receptor-Like Cytoplasmic Kinase OsRLCK102 Regulates XA21-Mediated Immunity and Plant Development in Rice

Receptor-like cytoplasmic kinases (RLCKs) belong to a large subgroup of kinases that play pivotal roles in plant development and in protecting plants from various stresses. Here, we report the isolation and characterization of rice OsRLCK102, from the OsRLCK VII subgroup. Silencing of OsRLCK102 compromised receptor kinase XA21-mediated resistance to Xanthomonas oryzae pv. oryzae (Xoo) but did not affect plant basal resistance to Xoo or Magnaporthe oryzae (M. oryzae). Plants with silenced OsRLCK102 exhibit architecture alterations, including reduced plant height, enlarged angle of the lamina joint, decreased rates of seed setting and enhanced sensitivity to hormone brassinolide (BR). Collectively, our study reveals that OsRLCK102 positively regulates XA21-mediated immunity and negatively regulates rice development through BR signaling in rice.     

<Emphasis Type="Italic">Lsi1</Emphasis>-regulated Cd uptake and phytohormones accumulation in rice seedlings in presence of Si

Silicon (Si) is known for its role in regulating the response of plants to imposed abiotic stresses. Since the stresses generally hinder production of a crop, such as rice, the exploration of the biochemistry and plant physiology relating to the function is of interest. Indeed, recently, there were reports on the function of Lsi1 in regulating the tolerance of rice to cadmium (Cd) stress. This study compared the kinetics of the Cd uptakes in Lemont wild type rice and its transgenic lines exposed to Cd with or without exogenous Si supply. At the same time, changes on the endogenous phytohormones and growth of the rice seedlings were monitored. Genetically, Lsi1 overexpression was found to downregulate K_m and V_max of Cd uptake kinetics in the plants under Cd stress, especially in the presence of Si. On the other hand, Lsi1 RNAi upregulated K_m and V_max regardless whether Si was present or not. It implied that Lsi1 could be capable of regulating Si as well as Cd transports. Under Cd stress, addition of Si reduced the Cd uptake of the rice lines in the order of Lsi1-overexpression line?>?Lemont?>?Lsi1-RNAi line. In addition, it also affected the chlorophyll biosynthesis and dry mass accumulation of the rice plants under Cd stress. Analyses on phytohormones including IAA, GA₃, JA, SA and ABA, as well as physiological functions, of the seedlings further verified the active involvement of Lsi1 in the complex defense system of the plants against Cd stress.     

Isolation,characterization and virulence of bacteria from <Emphasis Type="Italic">Ostrinia nubilalis</Emphasis> (Lepidoptera: Pyralidae)

Isolation, characterization and virulence of the culturable bacteria from entire tissues of larval Ostrinia nubilalis (Hübner) (Lepidoptera: Pyralidae) were studied to obtain new microbes for biological control. A total of 16 bacteria were isolated from living and dead larvae collected from different maize fields in the Eastern Black Sea Region of Turkey. The bacterial microbiota of O. nubilalis were identified as Pseudomonas aeruginosa (On1), Brevundimonas aurantiaca (On2), Chryseobacterium formosense (On3), Acinetobacter sp. (On4), Microbacterium thalassium (On5), Bacillus megaterium (On6), Serratia sp. (On7), Ochrobactrum sp. (On8), Variovorax paradoxus (On9), Corynebacterium glutamicum (On10), Paenibacillus sp. (On11), Alcaligenes faecalis (On12), Microbacterium testaceum (On13), Leucobacter sp. (On14), Leucobacter sp. (On15) and Serratia marcescens (On16) based on their morphological and biochemical characteristics. A partial sequence of the 16S rRNA gene was also determined to confirm strain identification. The highest insecticidal activities were obtained from P. aeruginosa On1 (80%), Serratia sp. On7 (60%), V. paradoxus On9 (50%) and S. marcescens On16 (50%) against larvae 14 days after treatment (p < 0.05). Also, the highest activity from previously isolated Bacillus species was observed from Bacillus thuringiensis subsp. tenebrionis Xd3 with 80% mortality within the same period (p < 0.05). Our results indicate that P. aeruginosa On1, Serratia sp. On7, V. paradoxus On9, S. marcescens On16 and B. thuringiensis subsp. tenebrionis Xd3 show potential for biocontrol of O. nubilalis.     

Improvement of seedling and panicle blast resistance in <Emphasis Type="Italic">Xian</Emphasis> rice varieties following <Emphasis Type="Italic">Pish</Emphasis> introgression

Rice blast is a serious disease caused by the filamentous ascomycetous fungus Magnaporthe oryzae. Incorporating disease resistance genes in rice varieties and characterizing the distribution of M. oryzae isolates form the foundation for enhancing rice blast resistance. In this study, the blast resistance gene Pish was observed to be differentially distributed in the genomes of rice sub-species. Specifically, Pish was present in 80.5% of Geng varieties, but in only 2.3% of Xian varieties. Moreover, Pish conferred resistance against only 23.5% of the M. oryzae isolates from the Geng-planting regions, but against up to 63.2% of the isolates from the Xian-planting regions. Thus, Pish may be an elite resistance gene for improving rice blast resistance in Xian varieties. Therefore, near-isogenic lines (NILs) with Pish and the polygene pyramid lines (PPLs) PPL^Pish/Pi1, PPL^Pish/Pi54, and PPL^Pish/Pi33 in the Xian background Yangdao 6 were generated using a molecular marker-assisted selection method. The results suggested that (1) Pish significantly improved rice blast resistance in Xian varieties, which exhibited considerably improved seedling and panicle blast resistance after Pish was introduced; (2) PPLs with Pish were more effective than the NILs with Pish regarding seedling and panicle blast resistance; (3) the PPL seedling and panicle blast resistance was improved by the complementary and overlapping effects of different resistance genes; and (4) the stability of NIL and PPL resistance varied under different environmental conditions, with only PPL^Pish/Pi54 exhibiting highly stable resistance in three natural disease nurseries (Jianyang, Jinggangshan, and Huangshan). This study provides new blast resistance germplasm resources and describes a novel molecular strategy for enhancing rice blast resistance.     

Characterization of harpin<Subscript>Xoo</Subscript> induced hypersensitive responses in non host plant,tobacco

Harpin proteins encoded by hrp genes are bacterial protein elicitors that can stimulate hypersensitive response (HR) in non-host plants. HR-related pathogen resistance involves a complex form of programmed cell death (PCD). It is increasingly viewed as a key component of the hypersensitive disease response of plants. Currently, the evidence of harpin proteins-induced PCD is deficient though it exhibits phenotypic parallels with HR, and the mechanism of harpin proteins-induced PCD is not well understood. In this study, we demonstrate that harpin_Xoo protein from Xanthomonas oryzae pv. oryzae of rice bacterial blight expressed and isolated from bacterial cells acted as an agent to induce PCD in infiltrated tobacco plants. Treatment of tobacco leaves with harpin_Xoo induced typical PCD-related morphological and biochemical changes including cell shrinkage and nuclear DNA degradation. We further analyzed the expression of several genes in signal transduction pathway of PCD in tobacco plants by real-time qRT-PCR analysis using EF-1α as an endogenous control. Our results showed that the expression of NtDAD1 was down-regulated and the expression of BI-1, tpa1 and aox1 was up-regulated following the infiltration of harpin_Xoo into tobacco leaves. Our data suggest that harpin_Xoo can induce PCD with the coordination of PCD-related genes in infiltrated tobacco leaves, providing evidence to further investigate the signal transduction pathways of HR and PCD.     

Influence of Different Rice Cultivars on <Emphasis Type="Italic">Schizotetranychus oryzae</Emphasis> Development

Schizotetranychus oryzae Rossi de Simons (Acari: Tetranychidae) is considered one of the most important phytophagous mite in rice cultivation in the Americas South, Central, and North. This study aimed to examine some biological aspects of S. oryzae developing on leaves of three different cultivars of rice [Oryza sativa (L.)—Poaceae] produced in the state of Rio Grande do Sul, Brasil. The plants were kept in a room at 25?±?1°C, with natural light (photophase of approximately 14 h) and 70?±?5% relative humidity. During the immature stages, observations were carried out daily at 7 am, 1 pm, and 7 pm. The results showed that the mean duration of the egg–adult period in days were similar between cultivars evaluated (Irga 424, 11.27?±?0.13; Taim, 11.21?±?0.14 and Sinuelo, 11.13?±?0.15). Egg–adult viability on Irga 424, Sinuelo, and Taim was 61.9, 85.71, and 90.48%, respectively, being lowest on Irga 424 (χ2?=?28.62, p?<?0.0001). The duration of the immature stages was not affected by cultivar, but on Irga 424, egg–adult viability and female longevity were lower. The results of this study can help select O. sativa cultivar resistant to S. oryzae. However, historically, the IRGA 424 has lower populations of S. oryzae in field conditions.     

Effectiveness of cyanobacteria and green algae in enhancing the photosynthetic performance and growth of willow (<Emphasis Type="Italic">Salix viminalis</Emphasis> L.) plants under limited synthetic fertilizers application

The physiological response of plants to triple foliar biofertilization with cyanobacteria and green algae under the conditions of limited use of chemical fertilizers was investigated. Triple foliar biofertilization with intact cells of Microcystis aeruginosa MKR 0105, Anabaena sp. PCC 7120, and Chlorella sp. significantly enhanced physiological performance and growth of plants fertilized with a synthetic fertilizer YaraMila Complex (1.0, 0.5, and 0.0 g per plant). This biofertilization increased the stability of cytomembranes, chlorophyll content, intensity of net photosynthesis, transpiration, stomatal conductance, and decreased intercellular CO2 concentration. Applied monocultures augmented the quantity of N, P, K in plants, the activity of enzymes, such as dehydrogenases, RNase, acid or alkaline phosphatase and nitrate reductase. They also improved the growth of willow plants. This study revealed that the applied nontoxic cyanobacteria and green algae monocultures have a very useful potential to increase production of willow, and needed doses of chemical fertilizers can be reduced.     

Endogenous Salicylic Acid Levels and Signaling Positively Regulate <Emphasis Type="Italic">Arabidopsis</Emphasis> Response to Polyethylene Glycol-Simulated Drought Stress

Salicylic acid (SA) functions in the plant response to drought stress were assessed using SA-altering Arabidopsis mutants, including snc1 (with constitutively high levels of SA) and its nahG-transformed plants (named as snc1/nahG, with a comparable SA level to the wild type), sid2 and transgenic line nahG (both with SA deficiency), and npr1-1 (with SA signaling blockage). The drought stress was simulated by polyethylene glycol (PEG)-6000 treatment. Compared with wild-type (wt) plants, the snc1 plants displayed obvious easing of PEG-induced growth inhibition, leaf water loss, and photosynthesis-related impairment, whereas in nahG, sid2, and npr1-1 mutants the effect was more severe. PEG stress reduced stomatal conductance, to a higher extent in the snc1 line, whereas it was lower in nahG, sid2, and npr1-1 lines as compared with the wt. The snc1 plants accumulated higher levels of H₂O₂ than the other genotypes tested. PEG stress increased activities of superoxide dismutase and peroxidase, but decreased activities of catalase in all lines tested, to a greater extent in snc1 and less in sid2, nahG, and npr1-1 relative to wt. Proline was significantly increased, especially in snc1 line at 6 % and higher PEG stress. Noticeably, the performance of snc1 under PEG stress was dependent on SA levels, as the expression of nahG in snc1 plants did not only significantly reduce SA levels, but largely reversed the above-mentioned parameters, as well as eliminated the drought tolerance. Based on these data, it was concluded that endogenous SA levels and signaling provided a protective role in the Arabidopsis response to PEG-simulated drought.     

Elevation of Defense Network in Chilli Against <Emphasis Type="Italic">Colletotrichum capsici</Emphasis> by Phyllospheric <Emphasis Type="Italic">Trichoderma</Emphasis> Strain

Biocontrol strategies have been mainly focused on proposing the use of biocontrol agents (BCAs) isolated from the rhizospheric region of the plant for protection against phytopathogens. The present study evaluates the effectiveness of phyllospheric Trichoderma isolates in elevating the defense responses in chilli against Colletotrichum capsici infection and comparing its efficiency to the conventionally recommended rhizospheric Trichoderma strains. The elicitation of the defense network in the plants was analyzed using biochemical assays for important enzymes, that is, PAL, PO, PPO, TPC, SOD along with the total protein level in challenged plants over untreated and unchallenged control plants. The results recorded 2.1, 5.18, 3, 0.67, and 0.5-fold increases in TPC, PAL, PO, PPO, and total protein content in BHUF4 (phyllopsheric Trichoderma isolate)-treated plants when compared to control plants under C. capsici challenge. This was at par with the increment recorded in T16A (rhizospheric Trichoderma isolate)-treated chilli plants. The increment in growth parameters was also recorded after treatment with the isolated Trichoderma strains. Interestingly, the phyllospheric isolate (BHUF4) treatment recorded comparable growth promotion in chilli plants recording 36, 62, and 60 % increases in one of the major parameters of plant growth, that is, root length, no. of leaves, and dry weight, respectively. This study proposes the use of combined application of both rhizospheric as well as phyllospheric Trichoderma isolates for better and all around protection of plants against foliar as well as soil phytopathogens. This would be a novel approach in biological control strategy for better management of anthracnose disease of chilli.     

Salicylic acid promotes plant growth and salt-related gene expression in <Emphasis Type="Italic">Dianthus superbus</Emphasis> L. (Caryophyllaceae) grown under different salt stress conditions

Salt stress is a critical factor that affects the growth and development of plants. Salicylic acid (SA) is an important signal molecule that mitigates the negative effects of salt stress on plants. To elucidate salt tolerance in large pink Dianthus superbus L. (Caryophyllaceae) and the regulatory mechanism of exogenous SA on D. superbus under different salt stresses, we conducted a pot experiment to evaluate leaf biomass, leaf anatomy, soluble protein and sugar content, and the relative expression of salt-induced genes in D. superbus under 0.3, 0.6, and 0.9% NaCl conditions with and without 0.5 mM SA. The result showed that exposure of D. superbus to salt stress lead to a decrease in leaf growth, soluble protein and sugar content, and mesophyll thickness, together with an increase in the expression of MYB and P5CS genes. Foliar application of SA effectively increased leaf biomass, soluble protein and sugar content, and upregulated the expression of MYB and P5CS in the D. superbus, which facilitated in the acclimation of D. superbus to moderate salt stress. However, when the plants were grown under severe salt stress (0.9% NaCl), no significant difference in plant physiological responses and relevant gene expression between plants with and without SA was observed. The findings of this study suggest that exogenous SA can effectively counteract the adverse effects of moderate salt stress on D. superbus growth and development.     

Efficient harvesting of <Emphasis Type="Italic">Synechocystis</Emphasis> sp. PCC 6803 with filamentous fungal pellets

Cyanobacteria play a major role as direct producers of biofuels, such as ethanol and butanol, with the aid of genetic engineering. However, development of a new harvesting-technology is essential to achieve economic viability of biofuel production from cyanobacteria. In this study, we demonstrated the feasibility of harvesting the unicellular cyanobacterium Synechocystis sp. PCC 6803 using pre-made filamentous fungal pellets and investigated key factors affecting efficiency of harvest, including fungal strain, pellet quantity (number of pellets), initial pH, and organic carbon source. Synechocystis sp. PCC 6803 cells attached to Aspergillus oryzae pellets, indicating that this fungal pellet had a desirable harvesting effect, while Rhizopus oryzae pellets had no effect on harvesting. Increasing pellet quantity and adding organic carbon sources, such as glucose and xylose, improved the harvesting efficiency of Aspergillus oryzae pellet; efficiency was not affected by the initial pH.     

Evaluation of insecticidal activity of a bacterial strain, <Emphasis Type="Italic">Serratia</Emphasis> sp. EML-SE1 against diamondback moth

To identify novel bioinsecticidal agents, a bacterial strain, Serratia sp. EML-SE1, was isolated from a dead larva of the lepidopteran diamondback moth (Plutella xylostella) collected from a cabbage field in Korea. In this study, the insecticidal activity of liquid cultures in Luria-Bertani broth (LBB) and nutrient broth (NB) of a bacterial strain, Serratia sp. EML-SE1 against thirty 3rd and 4th instar larvae of the diamondback moth was investigated on a Chinese cabbage leaf housed in a round plastic cage (Ø 10×6 cm). 72 h after spraying the cabbage leaf with LBB and NB cultures containing the bacterial strain, the mortalities of the larvae were determined to be 91.7% and 88.3%, respectively. In addition, the insecticidal activity on potted cabbage containing 14 leaves in a growth cage (165×83×124 cm) was found to be similar to that of the plastic cage experiment. The results of this study provided valuable information on the insecticidal activity of the liquid culture of a Serratia species against the diamondback moth.