Extracellular self-RNA: A danger elicitor in pepper induces immunity against bacterial and viral pathogens in the field.

Plants and animals serve as hosts for microbes. To protect themselves from microbe-induced damage, plants and animals need to differentiate self-molecules/signals from non-self, microbe-derived molecules. Damage-associated molecular patterns (DAMPs) are danger signals released from the damaged host tissue or present on the surface of stressed cells. Although a self-extracellular DNA has previously been shown to act as a DAMP in different plant species, the existence of a self-extracellular RNA (eRNA) as a danger signal in plants remains unknown. Here, we firstly evaluated the ability of a pepper self-eRNA to activate immunity against viral and bacterial pathogens under field conditions. Pepper leaves pre-infiltrated with self-eRNA exhibited reduced titer of the naturally occurring Tomato spotted wilt virus and diminished symptoms of Xanthomonas axonopodis pv. vesicatoria infection through eliciting defense priming of abscisic acid signaling. At the end of the growing season at 90 days after transplanting, pepper plants treated with self- and non-self-eRNAs showed no difference in fruit yield. Taken together, our discovery demonstrated that self-eRNA can successfully activate plant systemic immunity without any growth penalty, indicating its potential as a novel disease management agent against a broad range of pathogenic microbes.

Analysis of the Effect of Carbonation Rate on the Concrete Water Reservoir Structures According to Applied Waterproofing/Anticorrosive Methods.

This study seeks to analyze how the degree of carbonation and the application of waterproofing and anticorrosive materials affect carbonation in water reservoirs among the water treatment facilities managed by the Seoul Metropolitan Government. To guarantee similarity of the experimental group, 42 highly similar water reservoirs were selected from among the water supply reservoirs currently in operation in Seoul. On-site carbonation assessments were performed in order to derive the carbonation rate coefficients. In the water reservoirs with applied waterproofing and anticorrosive materials immediately after public service, the upper and lower limits were D = 1.13t and D = 0.29t, respectively, whereas those of the water reservoir applied with waterproofing and anticorrosive materials after 15 years of service life were D = 1.89t and D = 0.94t, respectively. The comparative analysis showed that the rate of reduction in the carbonation rate was about 10.4% to 16.8% in the water reservoirs applied with waterproofing and anticorrosive methods after 15 years of service life. However, reduction in the carbonation rate was about 46.4% to 74.3% in the water reservoirs applied with waterproofing and anticorrosive methods at the initial stage of service life. It was confirmed that the early application of waterproofing and anticorrosive materials is effective in suppressing carbonation of concrete water reservoir structures.

Elicitation of Innate Immunity by a Bacterial Volatile 2-Nonanone at Levels below Detection Limit in Tomato Rhizosphere.

Bacterial volatile compounds (BVCs) exert beneficial effects on plant protection both directly and indirectly. Although BVCs have been detected in vitro, their detection in situ remains challenging. The purpose of this study was to investigate the possibility of BVCs detection under in situ condition and estimate the potentials of in situ BVC to plants at below detection limit. We developed a method for detecting BVCs released by the soil bacteria Bacillus velezensis strain GB03 and Streptomyces griseus strain S4-7 in situ using solid-phase microextraction coupled with gas chromatography-mass spectrometry (SPME-GC-MS). Additionally, we evaluated the BVC detection limit in the rhizosphere and induction of systemic immune response in tomato plants grown in the greenhouse. Two signature BVCs, 2-nonanone and caryolan-1-ol, of GB03 and S4-7 respectively were successfully detected using the soil-vial system. However, these BVCs could not be detected in the rhizosphere pretreated with strains GB03 and S4-7. The detection limit of 2-nonanone in the tomato rhizosphere was 1 µM. Unexpectedly, drench application of 2-nonanone at 10 nM concentration, which is below its detection limit, protected tomato seedlings against Pseudomonas syringae pv. tomato. Our finding highlights that BVCs, including 2-nonanone, released by a soil bacterium are functional even when present at a concentration below the detection limit of SPME-GC-MS.

Genome Sequence of Ralstonia pseudosolanacearum SL1931, a Causal Phytopathogen of Bacterial Wilt Disease in Capsicum annuum and Nicotiana benthamiana.

Here, we report the genome sequence of Ralstonia pseudosolanacearum (R. solanacearum phylotype I) strain SL1931 (KACC10711), isolated from pepper (Capsicum annuum L.) stems; R. solanacearum is the causal pathogen of bacterial wilt. Strain SL1931 had a different type III effector profile than that of the reference genome strain GMI1000.

Crack-Bridging Property Evaluation of Synthetic Polymerized Rubber Gel (SPRG) through Yield Stress Parameter Identification.

Yield stress parameter derivation was conducted by stress-strain curve analysis on four types of grout injection leakage repair materials (GILRM); acrylic, epoxy, urethane and SPRG grouts. Comparative stress-strain curve analysis results showed that while the yield stress point was clearly distinguishable, the strain ratio of SPRG reached up to 664% (13 mm) before material cohesive failure. A secondary experimental result comprised of three different common component ratios of SPRG was conducted to derive and propose an averaged yield stress curve graph, and the results of the yield stress point (180% strain ratio) were set as the basis for repeated stress-strain curve analysis of SPRGs of up to 15 mm displacement conditions. Results showed that SPRG yield stress point remained constant despite repeated cohesive failure, and the modulus of toughness was calculated to be on average 53.1, 180.7, and 271.4 N/mm2, respectively, for the SPRG types. The experimental results of this study demonstrated that it is possible to determine the property limits of conventional GILRM (acrylic, epoxy and urethane grout injection materials) based on yield stress. The study concludes with a proposal on potential application of GILRM toughness by finite element analysis method whereby strain of the material can be derived by hydrostatic pressure. Comparative analysis showed that the toughness of SPRG materials tested in this study are all able to withstand hydrostatic pressure range common to underground structures (0.2 N/mm2). It is expected that the evaluation method and model proposed in this study will be beneficial in assessing other GILRM materials based on their toughness values.

Morpho-Anatomical Traits and Soluble Sugar Concentration Largely Explain the Responses of Three Deciduous Tree Species to Progressive Water Stress.

A better understanding of plant drought responses is essential to improve plant water use efficiency, productivity, and resilience to ever-changing climatic conditions. Here, we investigated the growth, morpho-anatomical, physiological, and biochemical responses of Quercus acutissima Carruth., Quercus serrata Murray, and Betula schmidtii Regel to progressive water-stress. Seedlings were subjected to well-watered (WW) and water-stressed (WS) conditions while regularly monitoring the soil volumetric water content, stem diameter (SD), height, biomass, stomatal conductance (gs), intercellular CO2 concentration (Ci), and leaf relative water content (RWC). We also investigated the variation in stomatal pore (SP) area, specific leaf area (SLA), root xylem vessel diameter (VD), and total soluble sugar (TSS) concentration between treatments. After 2 months, WS significantly suppressed SD growth of Q. acutissima and B. schmidtii but had no impact on Q. serrata. Total biomass significantly declined at WS-treated seedlings in all species. WS resulted in a smaller SLA than WW in all species. The SP of WS-treated seedlings of Q. acutissima and B. schmidtii significantly decreased, whereas it increased significantly with time in Q. serrata. Larger vessels (i.e., >100 to ≤ 130) were more frequent at WS for Q. acutissima and B. schmidtii, whereas smaller vessels (i.e., >40 to ≤ 90) were more frequent at WS than at WW for Q. serrata after 8 weeks. Tylosis was more frequent at WS than WW for Q. serrata and B. schmidtii at eighth week. WS seedlings showed lower gs, Ci, and RWC compared with WW-treated ones in Q. acutissima and B. schmidtii. TSS concentration was also higher at WS-treated seedlings in two Quercus species. Overall, principal component analysis (PCA) showed that SLA and SP are associated with WS seedlings of Q. serrata and B. schmidtii and the tylosis frequency, TSS, and VD are associated with WS seedlings of Q. acutissima. Therefore, water-stressed plants from all species responded positively to water stress with increasing experimental duration and stress intensity, and that is largely explained by morpho-anatomical traits and soluble sugar concentration. The present study should enhance our understanding of drought-induced tree growth and short-term tree-seedling responses to drought.

Anti-Seismic Performance Evaluation of Waterproofing Materials for Underground Pile Wall Structures.

This study introduces and demonstrates the application of an experimental regime for anti-seismic performance evaluation of waterproofing materials used for concrete pile walls. Concrete pile walls are subject to high degrees of seismic load, and the resultant stress can affect the waterproofing integrity of the structure, but there is currently no existing methodology or standard for evaluating this property of waterproofing materials. To propose and conduct this evaluation, a new testing apparatus was designed and manufactured to test an installed waterproofing material's seismic resistance performance. Under three different inclined angle conditions (0°, 10°, 20°), each with three different rotation speed conditions (10, 20 and 30 rotations per minute), three types of waterproofing materials were subjected to 30 s of increasing seismic stress and tested for their waterproofing performance. Waterproofing performance was determined by whether the specimen installed with the respective type of material was able to prevent leakage path formation during the seismic stress, and the performance was summarized and compared based on the average results for four specimens of each material type and the duration before leakage occurrence. Results of the demonstration testing yielded significantly different results for the tested material types, prompting the need to further investigate different types of waterproofing materials, products, and techniques for a comprehensive understanding of waterproofing material response properties against seismic stress. The demonstration process shown in this research was intended to serve as a proposal for the development of these performance evaluation criteria, methodologies, and equipment for possible future application.

First Report of Colletotrichum siamense Causing Anthracnose of Chili Pepper Fruit in Korea.

In October 2015, typical anthracnose symptoms were observed on approximately 15 to 20% of the chili fruits (cv. Manita) growing in Goesan County, Chungcheong Province, South Korea. Infection of fruits were characterized by the presence of circular, sunken lesions with concentric rings of orange conidial acervuli. Fresh samples were collected from the infected fruits and lesions from seven symptomatic fruits were cut into small pieces (5 mm2) and surface sterilized by soaking them in 1% sodium hypochlorite for 3 min, followed by rinsing thrice using sterilized water, and drying on sterilized filter paper. The tissue pieces were then placed on potato dextrose agar (PDA) and incubated at 25 ± 2°C with 12hrs photoperiod. After 2 to 3 days, single hyphal tips were transferred to fresh PDA and a total of seven isolates were selected from typical single hyphae. The upper surfaces of the colonies formed on PDA were white to gray in color with cottony mycelia, in which salmon-colored acervuli were clearly visible (Supplementary 1). Thirty conidia were examined; all were hyaline, smooth-walled, aseptate, straight, mainly cylindrical with round ends, 12 to 17 µm long, and 3 to 4.5 µm wide. Appressoria were oval to irregular inshape, dark brown in color, and range from 9.5 to 11.5 µm × 6.5 to 7.5 µm in sizes. Morphological characteristics of the seven isolates were identical and resembled those of C. siamense (Weir et al. 2012). To confirm the identification of the fungal isolates, DNA from seven isolates were extracted (Cenis et al. 1992) and the genes encoding glyceraldehyde-3-phosphate dehydrogenase (GAPDH), internal transcribed spacer (ITS) rDNA regions, and ß-Tublin-2 (TUB2) were partially amplified and sequenced. Sequences from all seven isolates were identical each other. Nucleotide sequences of ITS, GAPDH, and TUB2 from representative isolates CNU180002 and CNU180012 were deposited in GenBank under accession numbers MH085103, MH085105, and MH085107 for CNU180002 and MK033503, MK033504, and MK033505 for CNU180012, respectively. The sequences for all three genes exhibited 99 to 100% identity with C. siamense, GenBank accession nos. FJ972613 (ITS), FJ972575 (GAPDH), and FJ907438 (TUB2) for both isolates. A multi-locus phylogenetic tree with closely related reference sequences downloaded from the GenBank database demonstrated that these two isolates were aligned with C. siamense. Pathogenicity of isolates CNU180002 and CNU180012 was confirmed on healthy fruits (Manita) by using a pin-pricked wound/drop (1 mm depth) and non-wound/drop inoculation method (Oo et al. 2017) and control fruits were mock-inoculated with sterilized distilled water. Three fruits were inoculated for each isolate and pathogenicity test were repeated thrice. After inoculation, the fruits were placed on a sterilized paper tissue in moistened clean boxes with a relative humidity of approximately 90% and incubated for 7 days at 25°C in the dark. Disease symptoms were appeared 5 to 7 days after inoculation on wounded fruits whereas non-wounded fruits were observed after 10 days. The two isolates showed identical symptoms and control fruits remained symptomless. Both isolates were re-isolated from infected fruits and were identical to the original isolates in morphology characteristics as well as on molecular sequences of ITS, GAPDH and TUB2 genes. To our knowledge, this is the first report of anthracnose caused by C. siamense on chili pepper fruit in Korea.

Analysis on the Effects of External Temperature and Welding Speed on the Safety of EVA Waterproofing Sheet Joints by Hot Air Welding.

This study analyzes the optimal seasonal ambient temperature during welding and welding speed conditions for securing high tensile strength of ethylene vinyl acetate (EVA) waterproofing sheets bonded for roofing, installed by hot air welded joints (overlaps). Seven separate ambient temperature conditions (-10, -5, and 0 °C for winter conditions, 20 °C for the normal condition, and 25, 30, and 35 °C for summer conditions) were set for the test variable and seven speed conditions (3, 4, 5, 6, 7, 8, and 9 m/min) for hot air welding. Based on these conditions, EVA sheet joint specimens were prepared, and the tensile strength of the joint sections was tested and measured. Tensile strength results, compared to normal temperature conditions (20 °C) showed an increase in the summer temperature condition but a decrease during winter temperature conditions. The analysis on the effects of the welding speed showed that in summer temperature conditions (25, 30, and 35 °C), the optimum hot air welding speed is 4.3~9.0 m/min at 25 °C, 4.7~8.7 m/min at 30 °C and 5.2~8.6 m/min at 35 °C, whereas in winter (-10, -5, and 0 °C), the optimum hot air welding temperature is 3~4.1 m/min at -10 °C, 3~4.6 m/min at -5 °C and 3~4.9 m/min at 0 °C. Research results demonstrate that it is imperative to consider the welding speed in accordance to the respective seasonal temperature conditions to secure construction quality of the EVA joints for roofing.

Evaluation of Sheet-Coated Composite Waterproofing Joint Types by Analysis of Tensile Strength Change and Concrete Displacement Resistance Testing under Environmental Degradation.

Sheet-coated composite waterproofing (SCCW) have been developed to overcome the natural weakness of singly-ply coating or sheet waterproofing systems for roofing, but there are currently multiple types of SCCW joints. Conventional standard tensile strength testing results show that all SCCW joint types seem to pass the minimum requirement and current selection of SCCW type is dictated on the principle of 'higher tensile strength is better', but it has not been experimentally studied as to which type is the optimal to respond to environmental degradation while under the effect of zero-span tensile stress occurring during concrete joint displacement. In this study, five types of SCCW joints were tested: Overlap Bond (OB) type, Overlap Heated-Air Welding (OH) type, Butt Joint I Type (BI), Butt Joint T Type (BT), and Butt Joint Separation Movement Type (BS). These types of joint designs were subjected to Alkali, NaCl, and H2SO4 exposure, and temperature change (60 °C and -20 °C) for determining changes to tensile strength in the joint section. Tensile strength change results are compared to joint displacement resistance test results of specimens that were treated with chemical and temperature degradation. With the exception of chemical exposure conditioning, the Overlap type joints generally had higher tensile strength compared to the butt joint types, but joint displacement test results showed the opposite results, suggesting that complex joints found in SCCW require new evaluation method for quality assessment.

Evaluation Method of Relative Humidity Changes in Below-Grade Concrete Structure Space Depending on Different Waterproofing Material and Installation Method.

An evaluation method for assessing the difference in the relative humidity (RH) control performance of waterproofing material is proposed. For a demonstration of this evaluation method, two waterproofing materials (urethane coating and cementitious waterproofing material) installed with different methods (positive and negative side of concrete structure respectively) are exposed to temperature conditions representing three seasonal conditions: Summer (40 °C), spring/autumn (20 °C) and winter (4 °C). Condensation level changes on the inner side of the waterproofing material installed specimen is measured, and for derive criteria for comparison, three parameters based on the average RH, intercept RH (derived from a linear regression analysis of RH measurement), and maximum relative humidity are derived for each different waterproofing material installed specimen. Based on quality specification for underground concrete structures, the demonstration evaluation establishes provisional standard criteria of below 70% RH, and all three parameters are evaluated to determine whether the tested waterproofing material/method complies to the performance requirement. Additional analysis through linear regression and cumulative probability density graphs are derived to evaluate the RH consistency and range parameters. The evaluation regime demonstrates a quantitative RH analysis method and apparatus, and a newly designed evaluation criteria is used to compare the RH control performance of positive-side installed urethane waterproofing materials and negative-side installed cementitious waterproofing material.

Oil Leakage Evaluation for Selection of Emulsion-Based Non-Curable Synthetic Polymer Rubberized Gel (ENC-SPRG) as Waterproofing Material in Underground Structures.

A revised oil leakage evaluation regime is proposed in response to the oil leakage problems of emulsion-based non-curable synthetic polymer rubberized gel (ENC-SPRG) used as a waterproofing material in concrete slabs of residential underground structures. Oil leakage from ENC-SPRG can cause significant economic and environmental damage. As ENC-SPRG waterproofing material is relatively new in the global waterproofing market, a systematic quality control for ENC-SPRG products being manufactured and exported globally is currently non-existent. For the selection of optimal ENC-SPRG, six assessment parameters comprised of averaged and daily average oil leakage mass, averaged and daily average filler content settlement, oil leakage area, and oil leakage duration are proposed. Five ENC-SPRG product specimens are tested to obtain the property values of each parameter. The property values derived from the test results are compared between the tested ENC-SPRG product specimens. With the demonstration of this evaluation regime, a quantified method for a comparative assessment of ENC-SPRG type waterproofing materials is established.

Identification and Characterization of Colletotrichum Species Associated with Bitter Rot Disease of Apple in South Korea.

Bitter rot caused by Colletotrichum species is a common fruit rotting disease of apple and one of the economically important disease in worldwide. In 2015 and 2016, distinct symptoms of bitter rot disease were observed in apple orchards in five regions of South Korea. In the present study, infected apples from these regions were utilized to obtain eighteen isolates of Colletotrichum spp. These isolates were identified and characterized according to their morphological characteristics and nucleotide sequence data of internal transcribed spacer regions and glyceraldehyde-3-phosphate-dehydrogenase. Molecular analyses suggested that the isolates of Colletotrichum causing the bitter rot disease in South Korea belong to 4 species: C. siamense; C. fructicola; C. fioriniae and C. nymphaeae. C. siamense and C. fructicola belonged to Musae Clade of C. gloeosporioides complex species while C. fioriniae and C. nymphaeae belonged to the Clade 3 and Clade 2 of C. acutatum complex species, respectively. Additionally, we also found that the isolates of C. gloeosporioides species-complex were more aggressive than those in the C. acutatum species complex via pathogenicity tests. Taken together, our results suggest that accurate identification of Colletotrichum spp. within each species complex is required for management of bitter rot disease on apple fruit in South Korea.

Identification and Characterization of Alternaria iridiaustralis Causing Leaf Spot on Iris ensata in China.

In 2016, a severe leaf spot disease was found on Iris ensata Thumb. in Nanjing, China. The symptom was elliptical, fusiform, or irregularly necrotic lesion surrounded by a yellow halo, from which a small-spored Alternaria species was isolated. The fungus was identified as Alternaria iridiaustralis based on morphological characteristics. The pathogenicity tests revealed that the fungus was the causal pathogen of the disease. Phylogenic analyses using sequences of ITS, gpd, endoPG, and RPB2 genes confirmed the morphological identification. This study is the first report of A. iridiaustralis causing leaf spots on I. ensata in China.

Alternaria brassicifolii sp. nov. Isolated from Brassica rapa subsp. pekinensis in Korea.

A new species belonging to the genus Alternaria was isolated from the necrotic leaf spots of Brassica rapa subsp. pekinensis in Yuseong district, Daejeon, Korea. It is an occasional isolate, not an etiological agent, which is morphologically similar to A. broccoli-italicae, but differs in conidial size and conidiophore shape. Phylogenetic analysis using the sequence datasets of the internal transcribed spacer (ITS) region of the rDNA, glyceraldehyde-3-phosphate dehydrogenase (gpd), and plasma membrane ATPase genes showed that it is distantly related to A. broccoli-italicae and closely related to Alternaria species in the section Pseudoalternaria, which belonged to a clade basal to the section Infectoriae. Morphologically, the species is unique because it produces solitary conidia or conidial chains (two units), unlike the four members in the section Pseudoalternaria that produce conidia as short branched chains. It exhibits weak pathogenicity in the host plant. This report includes the description and illustration of A. brassicifolii as a new species.

Continuous cropping of endangered therapeutic plants via electron beam soil-treatment and neutron tomography.

Various medicinal plants are threatened with extinction owing to their over-exploitation and the prevalence of soil borne pathogens. In this study, soils infected with root-rot pathogens, which prevent continuous-cropping, were treated with an electron beam. The level of soil-borne fungus was reduced to ≤0.01% by soil electron beam treatment without appreciable effects on the levels of antagonistic microorganism or on the physicochemical properties of the soil. The survival rate of 4-year-old plant was higher in electron beam-treated soil (81.0%) than in fumigated (62.5%), virgin (78%), or untreated-replanting soil (0%). Additionally, under various soils conditions, neutron tomography permitted the monitoring of plant health and the detection of root pathological changes over a period of 4-6 years by quantitatively measuring root water content in situ. These methods allow continual cropping on the same soil without pesticide treatment. This is a major step toward the environmentally friendly production of endangered therapeutic herbs.

Characterization and Pathogenicity of New Record of Anthracnose on Various Chili Varieties Caused by Colletotrichum scovillei in Korea.

The anthracnose disease caused by Colletotrichum species is well-known as a major plant pathogen that primarily causes fruit rot in pepper and reduces its marketability. Thirty-five isolates representing species of Colletotrichum were obtained from chili fruits showing anthracnose disease symptoms in Chungcheongnam-do and Chungcheongbuk-do, South Korea. These 35 isolates were characterized according to morphological characteristics and nucleotide sequence data of internal transcribed spacer, glyceraldehyde-3-phosphate-dehydrogenase, and ß-tubulin. The combined dataset shows that all of these 35 isolates were identified as C. scovillei and morphological characteristics were directly correlated with the nucleotide sequence data. Notably, these isolates were recorded for the first time as the causes of anthracnose caused by C. scovillei on pepper in Korea. Forty cultivars were used to investigate the pathogenicity and to identify the possible source of resistance. The result reveals that all of chili cultivars used in this study are susceptible to C. scovillei.

Nicotiana benthamiana Matrix Metalloprotease 1 (NMMP1) gene confers disease resistance to Phytophthora infestans in tobacco and potato plants.

We previously isolated Nicotiana benthamiana matrix metalloprotease 1 (NMMP1) from tobacco leaves. The NMMP1 gene encodes a highly conserved, Zn-containing catalytic protease domain that functions as a factor in the plant's defense against bacterial pathogens. Expression of NMMP1 was strongly induced during interactions between tobacco and one of its pathogens, Phytophthora infestans. To elucidate the role of the NMMP1 in defense of N. benthamiana against fungal pathogens, we performed gain-of-function and loss-of-function studies. NMMP1-overexpressing plants had stronger resistance responses against P. infestans infections than control plants, while silencing of NMMP1 resulted in greater susceptibility of the plants to the pathogen. This greater susceptibility correlated with fewer NMMP1 transcripts than the non-silenced control. We also examined cell death as a measure of disease. The amount of cell death induced by the necrosis-inducing P. infestans protein 1, PiNPP1, was dependent on NMMP1 in N. benthamiana. Potato plants overexpressing NMMP1 also had enhanced disease resistance against P. infestans. RT-PCR analysis of these transgenic potato plants revealed constitutive up-regulation of the potato defense gene NbPR5. NMMP1-overexpressing potato plants were taller and produced heavier tubers than control plants. We suggest a role for NMMP1in pathogen defense and development.

Identification and Characterization of New Record of Grape Ripe Rot Disease Caused by Colletotrichum viniferum in Korea.

In 2016, grape fruits showing ripe rot symptom were found in fields of Korea. The fungus was isolated and identified as Colletotrichum viniferum based on morphological characteristics and nucleotide sequence data of the internal transcribed spacer, glyceraldehyde-3-phosphate dehydrogenase and ß-tubulin. To our knowledge, this is the first report of C. viniferum causing grape ripe rot disease of grape fruits in Korea.