The ascomycete fungus Colletotrichum gloeosporioides infects a wide range of plant hosts and causes enormous economic losses in the world. The transcription factors (TFs) play an important role in development and pathogenicity of many organisms. In this study, we found that the C₂H₂ TF CgCrzA is localized in both cytoplasm and nucleus under standard condition, and it translocated from cytoplasm to nucleus in a calcineurin-dependent manner. Moreover, the ΔCgCrzA was hypersensitive to cell wall perturbing agents and showed severe cell wall integrity defects. Deletion of the CgCRZA inhibited the development of invasive structures and lost pathogenicity to plant hosts. Our results suggested that calcineurin-responsive TF CgCrzA was not only involved in regulating cell wall integrity, but also in morphogenesis and virulence in C. gloeosporioides.

Nutrient manipulation is a promising strategy for controlling plant diseases in sustainable agriculture. Although many studies have investigated the relationships between certain elements and plant diseases, few have comprehensively explored how differing mineral nutrition levels might affect plant-fungal pathogen interactions, namely plant susceptibility and resistance. Here, we systematically explored the effects of the seven mineral elements that plants require in the greatest amounts for normal development on the susceptibility of soybean plants (Glycine max) to Fusarium oxysporum infection in controlled greenhouse conditions. Nitrogen (N) negligibly affected plant susceptibility to infection in the range 4 to 24 mM for both tested soybean cultivars. At relatively high concentrations, phosphorus (P) increased plant susceptibility to infection, which led to severely reduced shoot and root dry weights. Potassium (K), calcium (Ca), magnesium (Mg), sulfur (S), and iron (Fe) induced plant resistance to infection as their concentrations were increased. For K and Ca, moderate concentrations had a positive effect on plant resistance to the pathogen, whereas relatively high doses of either element adversely affected plant growth and promoted disease symptoms. Further experiments were conducted, assessing disease suppression by selected combinations of macro-elements and Fe at screened concentrations, i.e., K (9 mM) plus Fe (0.2 mM), and S (4 mM) plus Fe (0.2 mM). The disease index was significantly reduced by the combination of K plus Fe. In conclusion, this systematic investigation of soybean plant responses to F. oxysporum infection provides a solid basis for future environmentally-friendly choices for application in soybean disease control programs.

Early warning services for crop diseases are valuable when they provide timely forecasts that farmers can utilize to inform their disease management decisions. In South Korea, collaborative disease controls that utilize unmanned aerial vehicles are commonly performed for most rice paddies. However, such controls could benefit from seasonal disease early warnings with a lead time of a few months. As a first step to establish a seasonal disease early warning service using seasonal climate forecasts, we developed the EPIRICE Daily Risk Model for rice blast by extracting and modifying the core infection algorithms of the EPIRICE model. The daily risk scores generated by the EPIRICE Daily Risk Model were successfully converted into a realistic and measurable disease value through statistical analyses with 13 rice blast incidence datasets, and subsequently validated using the data from another rice blast experiment conducted in Icheon, South Korea, from 1974 to 2000. The sensitivity of the model to air temperature, relative humidity, and precipitation input variables was examined, and the relative humidity resulted in the most sensitive response from the model. Overall, our results indicate that the EPIRICE Daily Risk Model can be used to produce potential disease risk predictions for the seasonal disease early warning service.

Xanthomonas campestris pv. campestris (Xcc), the pathogen of black rot which is the most destructive disease of Brassica vegetables throughout the world. Here, we reported two novel sequence-characterized amplified region (SCAR) markers (i.e., XccR6-60 and XccR6-67) for the detection of Xcc race 6 via re-alignment of the complete genome sequences of Xcc races/strains/pathovars. The specificity of SCAR primer sets was verified by mean of PCR amplification using the genomic DNA template of Xcc races/strains/pathovars and two other plant infecting bacterial strains. The PCR result revealed that the XccR6-60 and XccR6-67 primer sets amplified 692-bp and 917-bp DNA fragments, respectively, specifically from race 6, while no visible amplification was detected in other samples. In addition, the SCAR primers were highly sensitive and can detect from a very low concentration of genomic DNA of Xcc race 6. However, the complete genome sequence of Xcc race 6 is not yet publicly available. Therefore, the cloning and sequencing of XccR6-60 and XccR6-67 fragments from race 6 provide more evidence of the specificity of these markers. These results indicated that the newly developed SCAR markers can successfully, effectively and rapidly detect Xcc race 6 from other Xcc races/strains/pathovars as well as other plant pathogenic bacteria. This is the first report for race-specific molecular markers for Xcc race 6.

Erwinia pyrifoliae is a Gram-negative bacterial plant pathogen that causes black shoot blight in apple and pear. Although earlier studies reported the genome comparison of Erwinia species, E. pyrifoliae strains for such analysis were isolated in 1996. In 2014, the strain E. pyrifoliae EpK1/15 was newly isolated in the apple tree showing black shoot blight in South Korea. This study aimed to better understand the similarities and differences caused by natural variations at the genomic level between newly isolated E. pyrifoliae EpK1/15 and the strain Ep1/96, which were isolated almost 20 years apart. Several comparative genomic analyses were conducted, and Clusters of Orthologous Groups of proteins (COG) database was used to classify functional annotation for each strain. E. pyrifoliae EpK1/15 had similarities with the Ep1/96 strain in stress-related genes, Tn3 transposase of insertion sequences, type III secretion systems, and small RNAs. The most remarkable difference to emerge from this comparison was that although the draft genome of E. pyrifoliae EpK1/15 was almost conserved, Epk1/15 strain had at least three sorts of structural variations in functional annotation according to COG database; chromosome inversion, translocation, and duplication. These results indicate that E. pyrifoliae species has gone natural variations within almost 20 years at the genomic level, and we can trace their similarities and differences with comparative genomic analysis.

The purpose of this study was to analyze the genetic and pathogenic characteristics of Ralstonia pseudosolanacearum in roses in Korea, and to examine the similarities and differences between Korean isolates and the first-reported European strains. Between 2017 and 2019, seventeen isolates from rose plants were identified as R. pseudosolanacearum using Ralstonia-specific primers. All 17 isolates were identified as race 1 using race-specific primers, and were confirmed as biovar 3 due to their ability to utilize carbon sources. Multiplex PCR using phylotype discriminating specific primers identified the 17 isolates as phylotype I. Sequevar comparison with reference sequevars using the sequences of the egl, mutS, and fliC genes, and only the egl gene, revealed that the strains evaluated in this study corresponded to sequevar I-33. The pathogenicity in roses differed depending on the rose cultivars. The different methods used for the genetic characterization of R. pseudosolanacearum indicate that the 17 rose bacterial wilt isolates had the same genetic characteristics. The lack of genetic variation in these isolates indicates their recent introduction from other countries (likely European countries). Therefore, appropriate quarantine and control measures should be taken in order to avoid further increases in the pathogenicity and/or secondary host range of R. pseudosolanacearum through genetic mutation.

Pepper and tomato plants infected with two Clavibacter species, C. capsici and C. michiganensis have shown different patterns of disease development depending on their virulence. Here, we investigated how pepper and tomato plants respond to infection by the high-virulent or low-virulent Clavibacter strains. For this, we chose two strains of each Clavibacter species to show different virulence level in the host plants. Although low-virulent strains showed less disease symptoms, they grew almost the same level as the high-virulent strains in both plants. To further examine the response of host plants to Clavibacter infection, we analyzed the expression patterns of plant defense-related genes in the leaves inoculated with different strains of C. capsici and C. michiganensis. Pepper plants infected with high-virulent C. capsici strain highly induced the expression of CaPR1, CaDEF, CaPR4b, CaPR10, and CaLOX1 at 5 days after inoculation (dai), but their expression was much less in low-virulent Clavibacter infection. Expression of CaSAR8.2 was induced at 2 dai, regardless of virulence level. Expression of GluA, Pin2, and PR2 in tomato plants infected with high-virulent C. michiganensis were much higher at 5 dai, compared with mock or low-virulent strain. Expression of PR1a, Osmotin-like, Chitinase, and Chitinase class 2 was increased, regardless of virulence level. Expression of LoxA gene was not affected by Clavibacter inoculation. These results suggested that Clavibacter infection promotes induction of certain defense-related genes in host plants and that differential expression of those genes by low-virulent Clavibacter infection might be affected by their endophytic lifestyle in plants.

Areca palm yellow leaf (AYL) disease caused by the 16SrI group phytoplasma is a serious threat to the development of the Areca palm industry in China. The 16S rRNA gene sequence was utilized to establish a rapid and efficient detection system efficient for the 16SrI-B subgroup AYL phytoplasma in China by loop-mediated isothermal amplification (LAMP). The results showed that two sets of LAMP detection primers, 16SrDNA-2 and 16SrDNA-3, were efficient for 16SrI-B subgroup AYL phytoplasma in China, with positive results appearing under reaction conditions of 64℃ for 40 min. The lowest detection limit for the two LAMP detection assays was the same at 200 ag/μl, namely approximately 53 copies/μl of the target fragments. Phytoplasma was detected in all AYL disease samples from Baoting, Tunchang, and Wanning counties in Hainan province using the two sets of LAMP primers 16SrDNA-2 and 16SrDNA-3, whereas no phytoplasma was detected in the negative control. The LAMP method established in this study with comparatively high sensitivity and stability, provides reliable results that could be visually detected, making it suitable for application and research in rapid diagnosis of AYL disease, detection of seedlings with the pathogen and breeding of disease-resistant Areca palm varieties.

Malva vein clearing virus (MVCV) is a member of the Potyvirus species, and has a negative impact on the aesthetic development of Alcea rosea. It was first reported in Germany in 1957, but its complete genome sequence data are still scarce. In the present work, A. rosea leaves with vein-clearing and mosaic symptoms were sampled and analyzed with small RNA deep sequencing. By denovo assembly the raw sequences of virus-derived small interfering RNAs (vsiRs) and whole genome amplification of malva vein cleaning virus SX strain (MVCV-SX) by specific primers targeting identified contig gaps, the full-length genome sequences (9,645 nucleotides) of MVCV-SX were characterized, constituting of an open reading frame that is long enough to encode 3,096 amino acids. Phylogenetic analysis showed that MVCV-SX was clustered with euphorbia ringspot virus and yam mosaic virus. Further analyses of the vsiR profiles revealed that the most abundant MVCV-vsiRs were between 21 and 22 nucleotides in length and a strong bias was found for "A" and "U" at the 5′-terminal residue. The results of polarity assessment indicated that the amount of sense strand was almost equal to that of the antisense strand in MVCV-vsiRs, and the main hot-spot region in MVCV-SX genome was found at cylindrical inclusion. In conclusion, our findings could provide new insights into the RNA silencing-mediated host defence mechanism in A. rosea infected with MVCV-SX, and offer a basis for the prevention and treatment of this virus disease.

The parasitic weed, Orobanche crenata, is one of the most devastating constraint for faba bean production in Mediterranean regions. Plant host defense induction was reported as one of the most appropriate control methods in many crops. The aim of this study was to elucidate the effect of salicylic acid (SA) and indole acetic acid (IAA) on the induction of faba bean resistance to O. crenata under the field and controlled experimental conditions. Both hormones were tested on two contrasting faba bean genotypes: Giza 843 (partially resistant to O. crenata) and Lobab (susceptible) at three different application methods (seed soaking, foliar spray, and the combination of both seed soaking and foliar spray). Soaking seeds in SA or IAA provided the highest protection levels reaching ~75% compared to the untreated control plants. Both elicitors limited the chlorophyll content decrease caused by O. crenata infestation and increased phenolic compound production in host plants. Phenylalanine ammonia lyase, peroxidase, and polyphenol oxidase activities were stimulated in the host plant roots especially in the susceptible genotype Lobab. The magnitude of induction was more obvious in infested than in non-infested plants. Histological study revealed that both SA and IAA decreased the number of attached O. crenata spikes which could be related to specific defense responses in the host plant roots.

An understanding of the contribution of secondary metabolites (SMs) to the antagonistic and biocontrol activities of bacterial biocontrol agents serves to improve biocontrol potential of the strain. In this study, to evaluate the contribution of each SM produced by Pseudomonas fluorescens NBC275 (Pf275) to its antifungal and biocontrol activity, we combined in silico analysis of the genome with our previous study of transposon (Tn) mutants. Thirteen Tn mutants, which belonged to 6 biosynthetic gene clusters (BGCs) of a total 14 BGCs predicted by the antiSMASH tool were identified by the reduction of antifungal activity. The biocontrol performance of Pf275 was significantly dependent on 2,4-diacetylphloroglucinol and pyoverdine. The clusters that encode for arylpolyene and an unidentified small linear lipopeptide influenced antifungal and biocontrol activities. To our knowledge, our study identified the contribution of SMs, such as a small linear lipopeptide and arylpolyene, to biocontrol efficacy for the first time.

Barley yellow dwarf virus (BYDV) is an economically important plant pathogen that causes stunted growth, delayed heading, leaf yellowing, and purple leaf tip, thereby reducing the yields of cereal crops worldwide. In the present study, a reverse transcription recombinase polymerase amplification (RT-RPA) assay was developed for the detection of BYDV in oat leaf samples. The RT-RPA assay involved incubation at an isothermal temperature (42℃) and could be performed rapidly in 5 min. In addition, no cross-reactivity was observed to occur with other cereal-infecting viruses, and the method was 100 times more sensitive than conventional reverse transcription polymerase chain reaction. Furthermore, the assay was validated for the detection of BYDV in both field-collected oat leaves and viruliferous aphids. Thus, the RT-RPA assay developed in the present study represents a simple, rapid, sensitive, and reliable method for detecting BYDV in oats.

The potential transmission of plant pathogenic viruses through processed foods could be a source of concern for global crop production; however, there is a lack of supporting evidence. The present study was conducted to investigate the presence of plant pathogenic viruses in five samples of gochujang (fermented red pepper paste) manufactured in Korea. Several viruses infecting pepper were detected by reverse transcription-polymerase chain reaction, among which the pepper mild mottle virus (PMMoV) was detected in all five samples, at concentrations ranging from 2.8 to 7.0 (log₁₀ copies/ml). In addition, PMMoV was observed by transmission electron microscopy in all five samples. The samples exhibited viral pathogenicity to Nicotiana benthamiana plants, indicating that global trade of processed products could be a possible source of the transmission of plant viruses.

Satsuma dwarf virus (SDV) seriously damages citrus production by reducing the quality and yield of fruit. To avoid contamination with SDV, mother trees are checked to be SDV-free in advance of nursery tree distribution. In this study, we compared an immunochromatographic assay (ICA) kit with double-antibody sandwich enzyme-linked immunosorbent assay (DAS-ELISA) for large-scale diagnosis of SDV in orchardgrown trees in Shizuoka Prefecture, Japan. The two methods gave conflicting results for 11 of 1,705 samples, all of which were negative by DAS-ELISA but positive by ICA. The samples scored as positive by either DAS-ELISA or ICA were analyzed by reverse transcription polymerase chain reaction and all were confirmed to be positive. These results validate the use of ICA as a screening method for large-scale diagnosis. Strain discrimination revealed that 16 of 22 isolates belonged to SDV, while citrus mosaic virus (CiMV) infection only and co-infection (SDV and CiMV) were in a minority.