Defense response of a pepper cultivar cv. Sy-2 is induced at temperatures below 24°C

Temperature is one of the most important environmental factors that influence plant growth and development. Recent studies imply that plants show various responses to non-extreme ambient temperatures. Previously, we have found that a pepper cultivar cv. Sy-2 (Capsicum chinense) shows developmental defects at temperatures below 24°C. In this study, to gain new insights into the temperature sensitivity of cv. Sy-2, temperature-sensitive genes were screened using microarray techniques. At restrictive temperature of 20°C, almost one-fourth of the 411 up-regulated genes were defense related or predicted to be defense related. Further expression analyses of several defense-related genes showed that defense-related genes in cv. Sy-2 were constitutively expressed at temperatures below 24°C. Moreover, accumulation of high level of salicylic acid (SA) in cv. Sy-2 grown at 20°C suggests that the defense response is activated in the absence of pathogens. To confirm that the defense response is induced in cv. Sy-2 below 24°C, we evaluated the resistance to biotrophic bacterial pathogen Xanthomonas campestris pv. vesicatoria and necrotrophic fungal pathogen Cercospora capsici. Cv. Sy-2 showed enhanced resistance to X. campestris pv. vesicatoria, but not to C. capsici.     

Overexpression of the <Emphasis Type="Italic">Activated Disease Resistance 1-like1</Emphasis> (<Emphasis Type="Italic">ADR1-L1</Emphasis>) Gene Results in a Dwarf Phenotype and Activation of Defense-Related Gene Expression in <Emphasis Type="Italic">Arabidopsis thaliana</Emphasis>

The protein encoded by the activated disease resistance 1-like1 (ADR1-L1) gene (locus name, At4g33300) belongs to the activated disease resistance 1 (ADR1) family of coiled-coil nucleotide-binding site leucine-rich repeat-type disease resistance proteins. This family contains four proteins and they have specific features in their amino acid sequences. It has been reported that ADR1 protein belongs to the ADR1 family, which is related to not only defense response but also drought tolerance. We found that transgenic plants overexpressing the ADR1-L1 gene showed a dwarf phenotype and morphological change in leaves. The expression levels of defense-related genes and the resistance to Pseudomonas syringae pv. tomato DC3000 were increased in transgenic plants. However, enhancement of drought tolerance and activation of abiotic response genes were not observed. When the growth temperature was changed from 22°C to 28°C, the expression of defense-related genes and the enhancement of resistance to a bacterial pathogen were suppressed and the dwarf phenotype and morphological change of leaves recovered.     

The Effect of Temperature on Symptom Expression and Colonization in Resistant and Susceptible Carnation Cultivars Infected with Fusarium oxysporum f. sp. dianthi

Six commercial carnation cultivars were inoculated with Fusarium oxysporum f. sp. dianthi race 2, and grown under three different temperature regimes. Colonization by the pathogen and development of wilt symptoms were assessed at intervals up to 40 days. No symptoms and very little colonization were seen in any of the cultivars at 14–15°C. At a temperature of 22°C, the cultivars were clearly differentiated into three groups: those with resistance, partial resistance or susceptibility to the pathogen depending on the severity of symptoms and the extent of fungal colonization. Symptom severity was associated with the extent of colonization. This differentiation was not seen at 26°C, when all cultivars except the most resistant, cv.‘Carrier 929′, rapidly became diseased and died by 23 days after inoculation. ‘Carrier 929’ also showed some wilt symptoms at this temperature and was colonized throughout the height of the stem after 40 days. The pathogen caused disease at 26°C by a combination of vascular wilting and stem base and root rotting. Fungal colonization was assayed at 22°C by the dilution plate/homogenization method and by estimation of fungal chitin in a highly resistant (‘Carrier 929′) and in a highly susceptible (‘Red Baron’) cultivar. Both methods of assay gave similar results. In ‘Red Baron’, colonization increased slowly up to 20 days after inoculation then progressed rapidly, closely following the development of severe wilt symptoms. In ‘Carrier 929’, colonization remained very low. The low level of fungal biomass in ‘Carrier 929’ compared with ‘Red Baron’ indicated that the former cultivar showed true resistance as opposed to tolerance to the disease.     

Studies on the physiological effects of viruses on sweet potato yield in Kenya

Much of the economic value of soybean (Glycine max) is based on the amount of protein and oil produced in the seeds. To examine the influence of temperature on seed oil and protein concentration, immature soybean seeds (cv. Williams 82) were grown in vitro at temperatures of 17°C, 21°C, 25°C, 29°C and 33°C. Dry growth rate (DGR) was calculated to be maximal at 23.7°C. Oil and protein concentration and seed growth rate did not show statistical difference (P > 0.05) within the temperature range from 21–29°C. Across all temperatures, however, a quadratic regression on oil concentration (R2 = 0.66) showed a minimum at 24.1°C and a quadratic regression on protein concentration (R2 = 0.59) showed a minimum at 24.3°C. Dilution by increased dry matter accumulation in the seed accounted for much of the variation in oil and protein concentration and the two concentrations were equally affected across temperatures. Consequently, oil and protein concentrations were positively related over the tested range of temperature. It was concluded that under these conditions the rate of dry matter accumulation by soybean seeds was critical in influencing seed oil and protein concentrations.     

Priming,signaling, and protein production associated with induced resistance by <Emphasis Type="Italic">Bacillus amyloliquefaciens</Emphasis> KPS46

Bacillus amyloliquefaciens KPS46 is a rhizobacterium that induces systemic protection in soybean (Glycine max L.) against several diseases and enhances plant growth. In this study, treatment of soybean seed with KPS46 provided protection to leaves from bacterial pustule, caused by Xanthomonas axonopodis pv. glycines (Xag). KPS46 treatment also increased phenolic content and β-1,3-glucanase and peroxidase activity levels in leaves over non-treated plants. Differential expression of these traits was more rapid and pronounced when KPS46 treated plants were infected with Xag, this pattern indicating priming. Also associated with induced resistance by KPS46 was increased production of salicylic acid (SA) and jasmonic acid (JA) in soybean leaves, suggesting both SA- and JA-dependent signaling pathways are systemically triggered by KPS46 seed treatment. When KPS46 was applied to Arabidopsis roots, however, resistance against Pseudomonas syringae pv. tomato (Pst) was induced only in host genotypes with intact jasmonate, ethylene, and auxin sensitivity. Thus, induced resistance against Pst by KPS46 was SA independent and JA/ethylene dependent. Proteins induced in soybean leaves by KPS46 seed treatment and by the seed treatment in combination with pathogen inoculation were determined by proteomic analysis. Among 20 proteins upregulated in KPS46-treated plants, compared with non-treated plants, only three were defense related. In plants that received both KPS46 treatment and inoculation with Xag, nine of the 20 upregulated protein, as compared with proteins produced Xag inoculated plants having no KPS46 treatment, were defense related. This pattern of increased induction of defense-related proteins following pathogen infection of KPS46 treated plants supported priming by KPS46. Aside from proteins with defense-related function, most of the proteins induced by KPS46 were involved in metabolism and energy conversion, reflecting the strong direct positive effect that KPS46 has on soybean growth.     

Wheat defense genes in fungal (Puccinia striiformis) infection

Wheat stripe rust, caused by Puccinia striiformis f. sp. tritici, is one of the most important diseases of wheat worldwide. To isolate defense-related genes against the pathogen, a suppression subtractive hybridization library was constructed for an incompatible interaction. From the library, 652 sequences were determined to be unigenes, of which 31 were determined as genes involved in signal transduction and 77 were predicted to encode defense-related proteins. Expression patterns of 12 selected signal transduction and defense-related genes were determined using quantitative real-time polymerase chain reaction. Signal transduction genes started increasing their expression at 12 h post inoculation (hpi), and expressions of the most of the transport and resistance-related genes were induced at 18 hpi. The gene expression results indicate specific molecular and cellular activities during the incompatible interaction between wheat and the stripe rust pathogen. In general, the expression increase of wheat signal transduction genes soon after inoculation with the pathogen inducing various defense-related genes, including reactive oxygen species, ATP-binding cassette (ABC) transporters, pathogenesis-related proteins, and genes involved in the phenylpropanoid pathway. The activities of these defense genes work in a sequential and concerted manner to result in a hypersensitive response.     

A cotyledon double inoculation technique for evaluating resistance to anthracnose (Colletotrichum orbiculare) and scab (Cladosporium cucumerinum) in cucumber

A technique for simultaneous inoculation of cucumber cotyledons with Colletotrichum orbiculare race 1 and Cladosporium cucumerinum has been developed. The procedure permitted both resistant and susceptible plants to be recovered. Seedlings were grown at 20°C and inoculated 24 h after emergence with Colletotrichum orbiculare (200 spores in 2 μ1 of water) and Cladosporium cucumerinum (1000 spores in 5 μ1 of water) followed by 48 h of incubation in the dark at 20°C and 100% r.h., and 48 h in a 20°C lighted growth chamber. Seedlings were then moved to a growth chamber at 21°C at night and at 26°C during the day for 4 days and plants were rated as resistant or susceptible 8 days after inoculation. No interference in the expression of resistance or susceptibility of cultivars to either pathogen was detected in simultaneous inoculations.     

Role of hydrogen peroxide and ascorbate-glutathione pathway in host resistance to bacterial wilt of eggplant

Ralstonia solanacearum, a soil-borne bacterium causes bacterial wilt, is a lethal disease of eggplant (Solanum melongena L.). However, the first line of defense mechanism of R. solanacearum infection remains unclear. The present study focused on the role of induced H₂O₂, defense-related enzymes of ascorbate-glutathione pathway variations in resistant and susceptible cultivars of eggplant under biotic stress. Fifteen cultivars of eggplant were screened for bacterial wilt resistance, and the concentration of antioxidant enzymes were estimated upon infection with R. solanacearum. A quantitative real-time PCR was also carried out to study the expression of defense genes. The concentration of H₂O₂ in the pathogen inoculated seedlings was two folds higher at 12 h after pathogen inoculation compared to control. Antioxidant enzymes of ascorbate-glutathione pathway were rapidly increased in resistant cultivars followed by susceptible and highly susceptible cultivars upon pathogen inoculation. The enzyme activity of ascorbate-glutathione pathway correlates by amplification of their defense genes along with pathogenesis-related protein-1a (PR-1a). The expressions of defense genes increased 2.5?3.5 folds in resistant eggplant cultivars after pathogen inoculation. The biochemical and molecular markers provided an insight to understand the first line of defense responses in eggplant cultivars upon inoculation with the pathogen.     

Effects of changing temperature regimes on resistance to races of Melampsora medusae in a cultivar of poplar

Leaf discs of Populus deltoides cv. W-79/307 inoculated with race 4–C of Melampsora medusae, give a compatible reaction when incubated at 16°C (LT), but an incompatible reaction at 26°C (HT). When, over a 12–day period, sets of inoculated leaf discs were reciprocally transferred between the temperature regimes (LT to HT, or HT to LT), incubation for as short as 15 h at HT resulted in incompatibility which was not reversed by subsequent incubation at LT. In contrast, incubation of the inoculated discs at LT for at least 4 days was necessary for the development of a compatible reaction following transfer to HT. Further, incompatibility induced in discs by inoculation with race 4–C and incubation at HT is epistatic to expected compatibility following subsequent inoculation with race 4–M, a temperature non-sensitive biotype. The rapidity, irreversibility and epistatic nature of the temperature-induced incompatibility suggests that initial recognition in this pathosystem may be for incompatibility. The significance of these results in this host/pathogen system is discussed.     

Seed germination of the weed Rumex obtusifolius after on-farm conventional, biodynamic and vermicomposting of cattle manure

This study investigated whether composting methods differ in their impact on seed germination of Rumex obtusifolius (broad‐leaved dock). Weed seeds were buried in windrows of cattle farmyard manure, removed at monthly intervals and germinated during the course of 7 months. Composting methods differed in the maximum temperatures reached (63°C for conventional and biodynamic composting and 35°C for vermicomposting), the addition of 1000 m^?2 earthworms (Eisenia fetida) for vermicomposting and the inoculation of biodynamic preparations for biodynamic composting. After 1 month in windrows, germination rate of Rumex seeds was significantly higher in vermicompost (48%) than in conventional (28%) or biodynamic compost (18%). After 2 months in windrows, 26% of the seeds germinated in vermicomposting windrows, while those inserted in conventional and biodynamic windrows showed a negligible germination (0% and 2%, respectively). After 3 and 4 months, only seeds under vermicomposting germinated (22% and 3%, respectively). No germination was determined when seeds were inserted for longer than 4 months in any of the treatments. Seeds stored at room temperature germinated at 89% over the course of the experiment. Results suggest that the maximum temperature reached in windrows is not the single main factor reducing weed seed germination during composting.     

Induction of some defense-related genes and oxidative burst is required for the establishment of systemic acquired resistance in <Emphasis Type="Italic">Capsicum annuum</Emphasis>

The inoculation of primary pepper leaves with an avirulent strain of Xanthomonas campestris pv. vesicatoria induced systemic acquired resistance (SAR) in the non-inoculated, secondary leaves. This SAR response was accompanied by the systemic expression of the defense-related genes, a systemic microoxidative burst generating H₂O₂, and the systemic induction of both ion-leakage and callose deposition in the non-inoculated, secondary leaves. Some defense-related genes including those encoding PR-1, chitinase, osmotin, peroxidase, PR10, thionin, and SAR8.2 were markedly induced in the systemic leaves. The conspicuous systemic accumulation of H₂O₂ and the strong increase in peroxidase activity in the pepper leaves was suggested to play a role in the cell death process in the systemic micro-hypersensitive responses (HR), leading to the induction of the SAR. Treatment of the primary leaves with diphenylene iodinium (DPI), an inhibitor of oxidative burst, substantially reduced the induction of some of the defense-related genes, and lowered the activation of the oxidative bursts in the systemic leaves distant from the site of the avirulent pathogen inoculation and subsequently SAR. Overall, these results suggest that the induction of some defense-related genes as well as a rapid increase in oxidative burst is essential for establishing SAR in pepper plants.     

Alpha-momorcharin, a RIP produced by bitter melon, enhances defense response in tobacco plants against diverse plant viruses and shows antifungal activity in vitro

Alpha-momorcharin (α-MMC) is type-1 ribosome inactivating proteins (RIPs) with molecular weight of 29 kDa and has lots of biological activity. Our recent study indicated that the α-MMC purified from seeds of Momordica charantia exhibited distinct antiviral and antifungal activity. Tobacco plants pre-treated with 0.5 mg/mL α-MMC 3 days before inoculation with various viruses showed less-severe symptom and less reactive oxygen species (ROS) accumulation compared to that inoculated with viruses only. Quantitative real-time PCR analysis revealed that the replication levels of viruses were lower in the plants treated with the α-MMC than control plants at 15 days post inoculation. Moreover, the coat protein expression of viruses was almost completely inhibited in plants which were treated with the α-MMC compared with control plants. Furthermore, the SA-responsive defense-related genes including non-expressor of pathogenesis-related genes 1 (NPR1), PR1, PR2 were up-regulated and activities of some antioxidant enzymes including superoxide dismutase (SOD), catalase (CAT), peroxidase (POD) were increased after the α-MMC treatment. In addition, the α-MMC (500 μg/mL) revealed remarkable antifungal effect against phytopathogenic fungi, in the growth inhibition range 50.35–67.21 %, along with their MIC values ranging from 100 to 500 μg/mL. The α-MMC had also a strong detrimental effect on spore germination of all the tested plant pathogens along with concentration as well as time-dependent kinetic inhibition of Sclerotinia sclerotiorum. The α-MMC showed a remarkable antiviral and antifungal effect and hence could possibly be exploited in crop protection for controlling certain important plant diseases.     

Physiological dormancy in seeds of tropical montane woody species in Hawai`i

Worldwide, there is relatively little information on seed dormancy and germination of tropical montane species. Our aim was to help fill this knowledge gap by conducting seed dormancy/germination studies on woody species from this vegetation zone in Hawai`i. All species had water-permeable seeds with a fully developed embryo. Seeds of 29 species (23 genera) were incubated in light/dark at 15/6, 20/10 and 25/15°C and germination monitored at 2-week intervals for 16–128 weeks. Seeds of Chenopodium oahuense, Dubautia menziesii and Silene lanceolata were non-dormant (ND) and those of 26 other species had physiological dormancy (PD); 10 of the 26 species had conditional PD. The optimum germination temperature regime(s) was (were) 25/15°C, 17 species; 25/10 and 20/10°C, 2; 20/10°C, 6; 20/10 and 15/6°C, 2; and 15/6°C, 2. Worldwide, PD in the woody genera included in our study is more common than ND. In addition to its contribution to the world biogeography of seed dormancy/germination, this study will be useful to conservation biologists who need to germinate seeds of tropical montane species.     

Effects of temperature and light on germination and early seedling development of the pine pink orchid (Bletia purpurea)

Orchid seed physiology is a poorly understood phenomenon owing to an emphasis on production and the challenges associated with propagating orchids from minute seed. We investigated the role of simulated south Florida temperatures and illumination (dark and 12 h photoperiod) in regulating germination and seedling development using asymbiotic seed germination assays of Bletia purpurea. Our objectives were to determine whether in situ germination is limited by seasonal temperatures and to determine whether temperature alters responses to illumination. Bletia purpurea seeds were able to germinate to > 90% under all treatments. The greatest germination after 3 weeks was observed at 29/19°C under continual darkness and at 25°C under dark and illuminated conditions. The slowest germination was observed at simulated winter temperatures (22/11°C). Illumination initially inhibited germination and development, but resulted in equal or greater development by week six. Germination under 22/11°C was strongly inhibited by illumination, indicating an interaction between temperature and light sensing systems.     

Effect of Temperature Regimes on Germination of Dimorphic Seeds of Atriplex prostrata

Dimorphic seeds of Atriplex prostrata were removed from cold dry storage monthly over a one year period to test for fluctuations in seed dormancy and germination rate. For each seed type, four replicates of 25 seeds were exposed to four alternating night/day temperature regimes mimicking seasonal fluctuations in Ohio: 5/15 °C; 5/25 °C; 15/25 °C and 20/35 °C with a corresponding 12-h photoperiod (20 μmol m⁻² s⁻¹; 400 – 700 nm). We found a significant three-way interaction of seed size, temperature and month for both percent germination and the rate of germination. Large seeds showed the greatest germination at the 20/35 °C and 5/25 °C temperature regimes and small seeds at the 5/25 °C regime. Large seeds had greater germination at all temperatures as compared to small seeds. Large seeds had the fastest germination rates at 20/35 °C followed by 5/25 °C whereas small seeds had the fastest rates at 5/25 °C followed by 20/35 °C. This revised version was published online in July 2006 with corrections to the Cover Date.     

Identification of rice genes induced in a rice blast-resistant mutant

To clarify mechanisms of rice blast resistance in rice plants we used suppression subtractive hybridization (SSH) to isolate genes induced upon rice blast inoculation in a rice blast-resistant mutant. A total of 26 rice cDNAs were isolated and found to have elevated expression upon rice blast infection in a rice blast-resistant derivative, SHM-11, of the rice cultivar, Sanghaehyanghyella. Sequencing of the cDNAs revealed that many of the proteins they encoded had been previously described as involved in plant responses against pathogen attack. Two interesting groups of the defense-related proteins consisted of three different PR5 homologues and four different protease inhibitors, all highly expressed in the rice blast mutant. Genes encoding proteins involved in signal transduction and regulation were also identified, including translation initiation factor eIF5A, C2 domain DNA binding protein, putative rice EDS and putative receptor like kinase. Most of the identified cDNAs were highly expressed 24 h after blast inoculation. Our results suggest that a pathway regulating defense gene expression may be altered in the mutant, resulting in early induction of the defense genes upon fungal infection.