Bacterial identification and classification are commonly performed based on the sequence of the 16S rRNA and several gold-standard marker genes, such as gyrB and recA. However, certain bacterial genera, including Xanthomonas and Pseudomonas, present challenges in clear classification at the species or subspecies level using these single-marker genes. Here, we explored the potential of the rfbA gene as a novel marker for bacterial classification. The rfbA gene, involved in lipopolysaccharide synthesis, is a conserved gene present in most gram-negative bacteria and is sufficiently short (approximately 850 bp). In this study, we identified that rfbA has undergone host-specific adaptive evolution, distinguishing it from conserved housekeeping genes. Therefore, we targeted Xanthomonas and Pseudomonas species to demonstrate the utility of rfbA in resolving taxonomic challenges at the pathovar level. These differences were strongly associated with the hosts of the respective strains, suggesting a potential evolutionary link between rfbA variation and host specificity.

Fire blight, caused by Erwinia amylovora, is an economically devastating disease affecting apple and pear orchards, and reliable detection is critical for effective management. However, field detection is challenging due to inhibitory compounds and the time-consuming nature of nucleic acid extraction, which limits the speed and accessibility of current diagnostic methods. Here, we present a CRISPR-Cas13a-based diagnostic platform designed for rapid, amplification-free, and extraction-free detection directly from plant material. In regions such as Korea where E. pyrifoliae is endemic, high genomic similarity between the two Erwinia species complicates accurate discrimination and poses a significant challenge for disease management. We identified E. amylovora-specific (EA-specific) single nucleotide polymorphisms and designed a panel of CRISPR RNAs (crRNAs) across multiple housekeeping genes and the 16S rRNA V3 region. Systematic screening with both synthetic RNA and mRNA revealed new crRNAs that maintained species specificity and sensitivity, achieving detection within minutes. To enable field-compatible sample processing, we developed and optimized a robust alkaline lysis workflow based on sequential NaOH lysis and HCl neutralization, which effectively released RNA from bacterial cells and remained compatible with crude Malus domestica leaf lysates. Under these extraction-free conditions, the assay achieved rapid, EA-specific detection of 1 × 106 CFUs/reaction within 15 minutes without nucleic acid purification or thermal cycling in the presence of plant material. This study establishes a practical framework for CRISPR-Cas13a diagnostics in plant pathology and provides a low-infrastructure strategy that can improve the speed and accuracy of fire blight surveillance and broader agricultural biosecurity efforts.

Plant pathogenic fungi have evolved molecular arsenals that enable them to successfully invade their host plants and ultimately achieve colonization. These mechanisms involve multifaceted and complex processes that require spatiotemporal regulation of various genes. The cosmopolitan genus Fusarium has been recognized worldwide as an important group of plant pathogens that exhibit diverse virulence mechanisms. This review seeks a broad overview of (a) how the virulence factors and their regulatory mechanisms are specifically utilized during the early invasion process in Fusarium species, (b) the gene regulatory mechanisms that govern this process, and (c) future directions for molecular genetics in disease control by directly targeting virulence factors in plant pathogenic fungi. By integrating current knowledge of key virulence factors and intrinsic mechanisms in Fusarium–plant systems, this work aims to provide a comprehensive understanding of Fusarium-mediated pathogenesis, with particular emphasis on the early stages of infection. Finally, we outline the shared and species-specific contributions of virulence factors, integrating findings from previous studies across individual Fusarium species.

Rice brown spot, caused by Bipolaris oryzae, typically occurs during the rice harvest season and can cause substantial yield losses. In 2023–2024, this disease emerged in rice cultivation areas of Korea. However, the population structure and genetic diversity of B. oryzae isolates remain unclear. Such information is necessary to effectively target and control rice brown spot. Thus, this study aimed to investigate the population structure of 50 B. oryzae isolates collected from the leaves, neck, and panicles of rice infected with brown spot through random amplified polymorphic DNA analysis. Among 140 primers tested, 30 were selected and applied, of which 5 exhibited significant polymorphisms among the isolates. The generated dendrogram revealed five clades with 92% similarity. Group A was the most predominant, comprising 84.0% of the total isolates (42/50), followed by Group C (8.0%, 4/50). Groups B, D, and E each contained one isolate (2%). These isolates were collected from the southern region of Korea and exhibited high genetic similarity (>95%). Two strains from Group A (F1305 and F1318) and one each from Groups B (F1248), C (F1253), D (F1317), and E (F1409) were selected and tested for their mycological characteristics and pathogenicity. Compared with the other strains, F1253, F1317, and F1409 exhibited higher conidial production and caused larger diseased leaf areas in the pathogenicity tests. These results suggest that the B. oryzae isolates that caused rice brown spot in 2023 are genetically homogeneous. This study may serve as a basis for developing targeted control strategies against brown spot.

Tree diseases associated with phytoplasma infections predominantly affecting nine host trees have serious impacts on tree growth and cause significant economic losses in South Korea. Loop-mediated isothermal amplification (LAMP)-based primers for early detection were developed to evaluate their accuracy. First, the 16S rRNA gene of phytoplasma was successfully amplified from the extracted DNA of various infected tree species using the polymerase chain reaction method. Two types diagnostic kits developed for phytoplasma detection were evaluated. The first kit detected phytoplasma infection within 30 min under isothermal conditions at 65°C, while the second kit did so within 40 min. Both kits could detect the nine different species of host trees infected with phytoplasma. When tested with 10 ng of the synthetic target gene, the FAM value became detectable at 10 min and remained consistent until 40 min. The lowest detection concentration was 0.01 pg/μL, and the limit of detection was 100 copies/μL. All of the phytoplasmas from nine diseased hosts were early detected. Furthermore, phytoplasma was not detected in healthy specimens, confirming the diagnostic kits’ accuracy in distinguishing between healthy and infected strains. The LAMP method confirmed rapid, accurate, and visually assessable detection of phytoplasma, suggesting it will enable early diagnosis of phytoplasma infections in South Korea.

Pratylenchus penetrans, an important soil pathogen, has been reported on various crops in the temperate regions of South Korea. In concern, there is an urgent need for a precise, species-specific quantitative polymerase chain reaction (qPCR) kit to detect and quantify root lesion nematodes for early pest management and to controls yield losses. The present study focuses on D2–D3 region, a known marker for molecular profiling of Pratylenchus sp. A primer set mined from the highly conserved D2–D3 region of P. penetrans was used in a SYBR green based qPCR assay. Initial examination identified P. penetrans from infested soil samples using morphological and phylogenetic analyses. The DPp7F12R primer set demonstrated significant specificity in identifying P. penetrants by both conventional polymerase chain reaction (PCR) and qPCR assays. Linear regression of serially diluted DNA from nematode and nematode inoculated soil revealed a limit of quantification of 2 picograms (r2 = 0.984), while also highlighting the impact of soil inhibitors. The qPCR using the DNA from varying densities of P. penetrans inoculated in soil demonstrated a robust correlation (r2 = 0.98), indicating the limit of detection down to single nematode. Primer specificity evaluation with field soil sample precisely detected only P. penetrants. Species-specific DPp7F12R facilitate the direct detection of P. penetrants from soil DNA in very shorter time. Reliability of PCR was confirmed using BLAST algorithm, which identified partial sequence of PCR amplicon (300 bp) as P. penetrants. Finally, PCR assay using DPp7F12R is crucial for early detection of P. penetrans infestations, helping improve the plant health.

Inoculation of Chinese cabbage leaves with high titer (107 cfu/ml) of the non-adapted bacteria Xanthomonas campestris pv. vesicatoria (Xcv) strain Bv5-4a.1 triggered rapid leaf tissue collapses and hypersensitive cell death (HCD) at 24 h. Electrolyte leakage and lipid peroxidation markedly increased in the Xcv-inoculated leaves. Defence-related gene expressions (BrPR1, BrPR4, BrChi1, BrGST1 and BrAPX1) were preferentially activated in the Xcv-inoculated leaves. The Xcv-triggered HCD was attenuated by continuous light but accelerated by a dark environment, and the prolonged high relative humidity also alleviated the HCD. Constant dark and increased relative humidity provided favorable conditions for the Xcv bacterial growth in the leaves. Pretreated fluridone (biosynthetic inhibitor of endogenous abscisic acid [ABA]) increased the HCD in the Xcv-inoculated leaves, but exogenous ABA attenuated the HCD. The pretreated ABA also reduced the Xcv bacterial growth in the leaves. These results highlight that the onset of HCD in Chinese cabbage leaves initiated by non-adapted pathogen Xcv Bv5-4a.1 and in planta bacterial growth was differently modulated by internal and external conditional changes.

The Colletotrichum gloeosporioides species complex includes many phytopathogenic species, causing anthracnose disease on a wide range of host plants and appearing to be globally distributed. Seventy-one Colletotrichum isolates in the complex from different plants and geographic regions in Korea were preserved in the Korean Agricultural Culture Collection (KACC). Most of them had been identified based on hosts and morphological features, this could lead to inaccurate species names. Therefore, the KACC isolates were re-identified using DNA sequence analyses of six loci, comprising internal transcribed spacer, gapdh, chs-1, his3, act, and tub2 in this study. Based on the combined phylogenetic analysis, KACC strains were assigned to 12 known species and three new species candidates. The detected species are C. siamense (n = 20), C. fructicola (n = 19), C. gloeosporioides (n = 9), C. aenigma (n = 5), C. camelliae (n = 3), C. temperatum (n = 3), C. musae (n = 2), C. theobromicola (n = 2), C. viniferum (n = 2), C. alatae (n = 1), C. jiangxiense (n = 1), and C. yulongense (n = 1). Of these, C. jiangxiense, C. temperatum, C. theobromicola and C. yulongense are unrecorded species in Korea. Host plant comparisons showed that 27 fungus-host associations are newly reported in the country. However, plant-fungus interactions need to be investigated by pathogenicity tests.

Cowpea mild mottle virus (CPMMV) is a global plant virus that poses a threat to the production and quality of legume crops. Early and accurate diagnosis is essential for effective managing CPMMV outbreaks. With the advancement in isothermal recombinase polymerase amplification and lateral flow strips technologies, more rapid and sensitive methods have become available for detecting this pathogen. In this study, we have developed a reverse transcription recombinase polymerase amplification combined with lateral flow strips (RT-RPA-LFS) method for the detection of CPMMV, specifically targeting the CPMMV coat protein (CP) gene. The RT-RPA-LFS assay only requires 20 min at 40°C and demonstrates high specificity. Its detection limit was 10 copies/μl, which is approximately up to 100 times more sensitive than RT-PCR on agarose gel electrophoresis. The developed RT-RPA-LFS method offers a rapid, convenient, and sensitive approach for field detection of CPMMV, which contribute to controlling the spread of the virus.

Physoderma fungal species cause faba bean gall (FBG) which devastates faba bean (Vicia faba L.) in the Ethiopian highlands. In three regions (Amahara, Oromia, and Tigray), the relative importance, distribution, intensity, and association with factors affecting FBG damage were assessed for the 2019 (283 fields) and 2020 (716 fields) main cropping seasons. A logistic regression model was used to associate biophysical factors with FBG incidence and severity. Amhara region has the highest prevalence of FBG (95.7%), followed by Tigray (83.3%), and the Oromia region (54%). Maximum FBG incidence (78.1%) and severity (32.8%) were recorded from Amhara and Tigray areas, respectively. The chocolate spot was most prevalent in West Shewa, Finfinne Special Zone, and North Shewa of the Oromia region. Ascochyta blight was found prevalent in North Shewa, West Shewa, Southwest Shewa of Oromia, and the South Gondar of Amhara. Faba bean rust was detected in all zones except for the South Gonder and North Shewa, and root rot disease was detected in all zones except South Gonder, South Wollo, and North Shewa of Amahara. Crop growth stage, cropping system, altitude, weed density, and fungicide, were all found to affect the incidence and severity of the FBG. Podding and maturity stage, mono-cropping, altitude (>2,400), high weed density, and non-fungicide were found associated with increased disease intensities. However, crop rotation, low weed infestation, and fungicide usage were identified as potential management options to reduce FBG disease.