In order to clarify the molecular characteristics and genetic evolutionary relationships of the emerging soybean disease, cowpea mild mottle virus (CPMMV), the full-length genomic sequence of a CPMMV isolate CPMMV-NJ-JS (8 195 nt, GenBank accession number: OK625819) was cloned through reverse transcription polymerase reaction (RT-PCR) and rapid amplification of cDNA ends (RACE). The results of sequence alignment showed that the nucleotide identity of this isolate with Chinese isolates (Jiangsu isolate GenBank accession No.: MT366555.1, Hubei isolate GenBank accession No.: MW354939.1, etc.) reached 95.90%-97.02%, while the identity with foreign isolates (USA isolate GenBank accession No.: KC774020.1, Ghana isolate GenBank accession No.: Ghana NC_014730.1) was obviously reduced (83.00% and 63.78%). Phylogenetic analysis revealed that the clustering of CPMMV isolates are consistent with their geographical distributions, forming independent evolutionary branches. Chinese isolates form the Ib subgroup, which is obviously differentiated from the groups of foreign isolates (subgroups Ia and Ic). In addition, the amino acid sequences of the coat protein (CP) encoded by ORF5 are completely identical among Chinese isolates, showing a high degree of conservation, while other non-structural proteins (ORF1-ORF4, ORF6) exhibit a relatively high degree of variation. This study provides important data on the molecular characteristics, geographical distributions and evolutionary relationships of CPMMV, and offers a theoretical basis for the monitoring and prevention of soybean viral diseases.

A systematic survey was conducted to characterize viral pathogens affecting sweet cherry (Prunus avium L.) cultivars in Qinghai Province. During May—September 2024, 243 leaf samples showing suspected viral infection symptoms were collected from 10 towns across 5 counties/districts in Haidong, Xining and Hainan regions. The samples were tested using double antibody sandwich enzyme-linked immunosorbent assay (DAS-ELISA) targeting seven viruses: cherry green ring mottle virus (CGRMV), cherry virus A (CVA), little cherry virus 1 (LChV-1), little cherry virus 2 (LChV-2), prunus necrotic ringspot virus (PNRSV), cherry rasp leaf virus (CRLV) and prune dwarf virus (PDV). Six viruses were detected with a total positivity rate of 49.38% (120/243), excluding LChV-2. CVA was the most prevalent (33.33%, 81/243), followed by PNRSV (10.69%, 26/243). The detection rates for CRLV, CGRMV, PDV and LChV-1 were lower at 9.05%, 4.11%, 3.29% and 3.70%, respectively. Distinct symptoms were observed for single-virus infections, whereas mixed infections caused more severe and complex leaf deformities. Infection patterns revealed 29.63% (72/243) single infections and 19.75% (48/243) co-infections, with CVA+PNRSV being the most common combination (7.81%, 19/243). A small proportion of samples harbored more than three viruses. Distinct symptoms were observed in single-virus infections, whereas mixed infections resulted in more complex symptomatology. Among all samples, both the composition of virus species and infection incidence varied across different counties, and a positive correlation was observed between tree age and infection rate, which may be associated with the transmission of viruses through propagation materials. This study systematically detected and analyzed, for the first time, the diversity and distribution of viruses infecting sweet cherry in Qinghai Province, providing a scientific basis for the management of cherry viral diseases and the production of virus-free planting materials.

Potato late blight, caused by Phytophthora infestans, is a major disease that poses a severe threat to potato production. Cinnamoyl-CoA reductase (CCR), a key enzyme in lignin biosynthesis, enhances plant mechanical resistance by promoting lignin deposition and cell wall thickening. To investigate the role of StCCR in potato resistance to late blight, we cloned the StCCR gene from the resistant potato cultivar ‘Qingshu 9’. Bioinformatics analysis showed that the coding sequence (CDS) of StCCR is 999 bp in length, encoding a protein of 322 aa. Quantitative real-time PCR (qPCR) analysis revealed that StCCR expression was significantly higher in resistant cultivars than in susceptible ones (P<0.05), with the highest level observed in the stems, followed by the leaves and roots. An StCCR-overexpressing vector was constructed and introduced into tobacco via Agrobacterium-mediated transformation. The transgenic lines, which were generated using the "leaf disc" method, exhibited significantly higher lignin content than wild-type plants. Phloroglucinol staining further confirmed that the lignified cell walls in the stems of transgenic lines were markedly thicker. Following inoculation with P. infestans, StCCR overexpression led to increased H₂O₂ accumulation and elevated activities of antioxidant enzymes (SOD, POD, APX, and PAL); it also upregulated the expression of several defense-related genes, including NtPR1, NtPR5, NtPAL, and NtERF1, while suppressing the expression of NtJAZ1, a negative regulator in the jasmonic acid signaling pathway. Additionally, StCCR modulated the expression of lignin biosynthesis-related genes by upregulating NtCAD, NtCOMT and NtC4H while downregulating Nt4CL and NtC3H. These synergistic effects enhanced resistance to late blight. Collectively, our results demonstrate that StCCR positively regulates potato resistance to late blight by coordinating defense responses and lignin biosynthesis.

Fusarium sacchari is one of the primary pathogens responsible for Pokkah boeng disease in sugarcane. Kinesins play a crucial role in intracellular transport and are involved in various physiological processes, including embryonic development, axonal transport, and cell division, making them essential for cell morphogenesis, function maintenance, and survival. BimC (BimC kinesin motor domain protein) is a member of the kinesin-5 subfamily. In F. sacchari, the FsBimC gene is 3 492 bp in length and encodes a protein containing a BimC kinesin domain (KISc_BimC_Eg5) and a microtubule-binding superfamily domain (Mbs), which localizes to the nucleus. Quantitative real-time PCR analysis revealed that the expression level of FsBimC increases during the late stage of F. sacchari infection in sugarcane leaves. To investigate the role of this gene in the virulence of F. sacchari, an RNA interference (RNAi)-mediated silencing mutant, FsBimC-RNAi, was generated in this study. Compared to the wild-type strain, the FsBimC-RNAi mutant exhibited slower mycelial growth, reduced conidial production, a significantly decreased conidial germination rate, and an increased frequency of conidia producing two germ tubes. These results indicate that FsBimC positively regulates mycelial growth and conidiation, and is involved in the process of conidial germination. Furthermore, the virulence of the FsBimC-RNAi mutant towards both sugarcane leaves and whole plants was significantly attenuated. These findings provide a potential molecular target for the green control of Pokkah boeng disease in sugarcane.

Stem canker, caused by Fusarium zanthoxyli, is a devastating disease in prickly ash production. The limited understanding of pathogen-host interaction mechanisms has hindered the development of effective control strategies. Effector proteins, which are crucial in these interactions, offer key insights into pathogenic mechanisms. In this study, we focused on FzEP119, an effector protein gene previously identified to be significantly up-regulated during the early infection stage of F. zanthoxyli. We constructed a knockout mutant (ΔFzEP119) and a complemented strain (ΔFzEP119-C) to systematically analyze the function of this gene in the pathogenicity and biological characteristics of the fungus. The results showed that, compared to the wild-type strain (Fz-WT), the virulence of ΔFzEP119 was significantly reduced, with lesions only 37% the size of those caused by Fz-WT. Although ΔFzEP119 showed no significant differences in mycelial growth rate, sporulation, and spore germination rate compared to the wild-type, its germ tubes were significantly shorter. In terms of carbon source utilization, the mutant exhibited significantly faster colony growth than Fz-WT when maltose was used as the carbon source, but grew significantly slower on glucose. Regarding nitrogen source utilization, ΔFzEP119 grew significantly slower than the wild-type on yeast extract, beef extract, potassium nitrate, and sodium nitrate. Additionally, ΔFzEP119 exhibited significantly reduced tolerance to H₂O₂ stress, while its responses to Congo red and NaCl stresses were similar to those of the wild-type. The mycelial penetration ability of ΔFzEP119 was also markedly impaired. The complemented strain ΔFzEP119-C restored virulence and other related biological characteristics to wild-type levels. In conclusion, the effector protein FzEP119 is a positive regulator of virulence in F. zanthoxyli and is involved in regulating multiple physiological processes, including germ tube elongation, nutrient utilization, stress response, and hyphal penetration. This study lays a foundation for further elucidating the molecular function and mechanism of FzEP119.

Root-knot nematodes (Meloidogyne spp.) are globally important soil-borne pathogens that cause severe economic losses to agricultural crops annually. Planting resistant crops is a core strategy for managing this disease; however, virulent nematode populations possess the ability to overcome gene-based resistance, threa-tening the sustainable utilization of resistance genes. In this study, we comparatively analyzed the host adaptability, life cycle, and expression profiles of pathogenesis-related genes between the Me3-virulent Meloidogyne incognita population (virulent to the pepper Me3 gene) and an avirulent M. incognita population on resistant and susceptible pepper and tomato cultivars. Our results showed that the host adaptability of the Me3-virulent population was significantly higher on pepper than on tomato and was closely associated with the type of resistance gene: it partially overcame the resistance conferred by the pepper N gene but failed to break the resistance conferred by the tomato Mi-1 and Mi-9 genes. Histopathological observations revealed that the body size of the Me3-virulent population on resistant pepper was significantly smaller than that of the avirulent population (P<0.05), accompanied by a shortened life cycle. Gene expression analysis demonstrated that pathogenesis-related genes (e.g., pectate lyase and β-1,4-endoglucanase) were significantly upregulated in the Me3-virulent population (P<0.05), whereas genes associated with growth and development (e.g., heat shock protein and cuticle protein) were suppressed. This study provides a theoretical basis for elucidating the pathogenic mechanisms of virulent root-knot nematodes and for the rational deployment of resistance genes.

To investigate the response of the kiwifruit leaf microbiome to bacterial canker occurrence, this study analyzed the leaf tissues of ‘Donghong’ kiwifruit at different disease grades (healthy, moderately affected, and severely affected). Using high-throughput sequencing, we characterized microbial community composition and dynamics to assess how infection alters microbial diversity and community structure in kiwifruit leaves. The results demonstrated that infection by the canker pathogen Pseudomonas syringae pv. actinidiae (Psa) significantly altered the phyllosphere bacterial and fungal microbiota in kiwifruit leaves, with a more pronounced impact on bacterial community composition than on fungal communities. Correlation and network analyses demonstrated that Psa invasion significantly strengthened positive inter-kingdom correlations (bacteria-fungi) while exhibiting significant negative correlations with key beneficial bacterial genera (Bacillus, Flavobacterium, Sphingomonas). The inverse abundance patterns of these beneficial bacteria suggest a potential host resistance mechanism that may help to mitigate Psa infection. In summary, Psa distinctly restructured the leaf-associated bacterial and fungal communities, destabilizing native bacterial community structures to facilitate its own proliferation. Concurrently, the enrichment of putative probiotic bacteria—Bacillus, Flavobacterium, and Sphingomonas—may play a role in alleviating the severity of kiwifruit canker caused by Psa.

To investigate the effects of ampicillin (AMP) on the rhizosphere bacterial community of citrus infected with Huanglongbing (HLB), this study used diseased citrus (Citrus reticulata Blanco cv. Shatangju) plants as materials. Changes in the HLB pathogen (Candidatus Liberibacter asiaticus, CLas) load within the plants were monitored following periodic trunk injection of AMP, and alterations in the rhizosphere bacterial community were analyzed using 16S rDNA amplicon sequencing. The results demonstrated that HLB infection increased the richness of rhizosphere bacteria, while AMP treatment significantly reduced bacterial diversity to below that of the healthy control (sterile water-injected healthy plants). Furthermore, AMP significantly enriched bacterial genera with antimicrobial potential and plant immune-activating capabilities (such as Burkholderia-Caballeronia-Paraburkholderia), while reducing the relative abundance of potential pathogens, HLB-associated bacteria, and denitrifying bacteria. In addition, AMP decreased the abundance of some common bacterial genera and induced the enrichment of multiple pollutant-degrading genera, suggesting that plants may perceive AMP as an abiotic stress. This study reveals that ampicillin can enhance plant disease resistance by reshaping the rhizosphere microbiome, but its use may also pose potential ecological risks, necessitating further careful evaluation before field application.

Bacteria perceive external environmental changes via two-component regulatory systems and remodel their metabolic pathways to adapt to adverse conditions. In Pseudomonas bijieensis 2P24, the RetS-GacS/GacA signaling pathway fine-tunes the expression of non-coding small RNAs (sRNAs), such as RsmZ, to influence the synthesis of the antibiotic 2,4-diacetylphlorogulcinol (2,4-DAPG). This study employed techniques including gene knockout and complementation, as well as transcriptional/translational fusion analysis, to investigate the regulatory role of Lon protease on the protein level of the transcriptional regulator GacA within the GacS/GacA two-component system and on biocontrol-related traits. The results showed that, compared to the wild-type strain 2P24, the accumulation level of GacA protein and the expression level of rsmZ were significantly increased in the lon deletion mutant. Bacterial two-hybrid assay results indicated that Lon does not directly participate in the degradation process of GacA. High-performance liquid chromatography (HPLC) analysis further demonstrated that the lon deletion significantly enhanced 2,4-DAPG production in strain 2P24 and strengthened its biofilm formation capability. These findings suggest that in strain 2P24, Lon protease affects GacA protein stability through an indirect mechanism, thereby finely regulating the expression of the sRNA RsmZ and ultimately influencing biocontrol-related traits such as 2,4-DAPG synthesis and biofilm formation. This study provides a new perspective for an in-depth understanding of the role of the RetS-GacS/GacA signaling pathway in disease suppression by the biocontrol strain 2P24.

Phomopsis seed decay (PSD), caused by Diaporthe longicolla, is a severe threat to soybean production globally. To establish novel chemical control strategies and address pathogen resistance issues, the inhibitory effects of the triazole compound mefentrifluconazole on 93 D. longicolla isolates were evaluated in this study. The results revealed a significant dose-dependent inhibitory effect on mycelial growth based on in vitro toxicity assays, and the concentrations at which mycelial growth is inhibited 50% (EC₅₀) values ranged from 0.018 6 to 0.222 6 μg·mL^-1, with a mean value of (0.058 0 ± 0.014 6) μg·mL^-1, demonstrating an exceptional fungicidal activity. Microstructural analysis revealed that the normal morphology of mycelium was significantly disrupted after mefentrifluconazole treatment including increased hyphal branching and compromised vacuole integrity. Biochemical assays showed that cell membrane permeability was also altered after mefentrifluconazole treatment. At a treatment concentration of 6.100 μg·mL^-1, the content of ergosterol (a critical component of fungal cell membranes) decreased by 62.95% and the accumulation of malondialdehyde (MDA) increased 1.91 times, while the activities of peroxidase (POD) and catalase (CAT) were reduced by 51.65% and 39.34%, respectively. The in vivo leaf assays showed that a treatment concentration of 6.100 μg·mL^-1 of mefentrifluconazole exhibited 88.60% of protective efficacy and 71.89% of therapeutic efficacy against PSD. Field efficacy trials demonstrated that foliar application of mefentrifluconazole (180 g a.i.·hm^-2) significantly reduced the disease index of PSD, with a control efficacy of 68.61%.Collectively, these data demonstrated that mefentrifluconazole exhibits significant control efficacy against D. longicolla and can be used as a novel fungicide for management of PSD in soybean production systems.

Phytophthora root rot is a devastating disease in global soybean production and a key target for quarantine and control in China. To identify novel target genes for the specific detection of Phytophthora sojae and establish a corresponding quantitative detection system, this study conducted comparative genomic analyses of 13 soybean pathogens, eight other Phytophthora species, and P. sojae populations. A single-copy gene, PsRrp8 (Ribosomal RNA-processing 8), was identified, which exhibits high sequence polymorphism across species but is highly conserved within P. sojae. Among six candidate primer pairs designed, PsRrp8-qPCR-F1/R1 was selected to finally establish the qPCR detection system. The system demonstrated a specific melting peak temperature (Tm) of (84.0±0.5) ℃, a detection sensitivity of 0.332 pg·μL^-1, and a standard curve of y=-3.266x+33.44 (R²=0.9993). In 2024, the assay was successfully applied to detect P. sojae in 38 soybean fields with root rot symptoms, with a positive detection rate of 42.1%. A positive correlation was observed between disease severity and pathogen load. This study provides a new technical tool for the rapid detection, quarantine, and early warning of P. sojae.

Cymbidium mosaic virus (CymMV) and odontoglossum ringspot virus (ORSV) are the two most prevalent viruses infecting Phalaenopsis aphrodite plants. These two viruses often co-infect plants in the field, and severely threaten the healthy development of the P. aphrodite industry. In this study, a duplex reverse transcription polymerase chain reaction (RT-PCR) detection system that can simultaneously detect CymMV and ORSV was established based on the conserved sequences of the cp gene of CymMV and ORSV. The results demonstrated that the designed primers exhibited high specificity and the optimized reaction system was as follows: the primer volume for CymMV and ORSV detection were 0.2 μL and 0.4 μL (initial concentration 10 μmol·L^-1), respectively; the final concentrations of Taq DNA polymerase, dNTPs and Mg²⁺ were 3.5 U·μL^-1 , 0.5 mmol·L^-1 and 15 mmol·L^-1 , respectively; add ddH₂O to 15.0 μL. The optimal annealing temperature was 55 ℃ and the cycle number for PCR was 25. Using the established duplex RT-PCR detection system, both viruses can be simultaneously detected from the samples of diseased P. aphrodite plants at a 10⁶ fold dilution. A 77% co-infection rate of the two viruses was detected by the duplex RT-PCR detection system on P. aphrodite plants from market. The duplex RT-PCR detection system established in this study has high sensitivity and specificity, providing technical support for the precise detection of CymMV and ORSV.

Myrica rubra (Lour.) S. et Zucc. is an important fruit tree in China, particularly in Zhejiang Province. However, branch canker disease has been increasingly prevalent in major production areas in recent years, severely affecting the healthy development of the M. rubra industry. In this study, branches showing typical canker symptoms were collected, potential pathogenic fungi were isolated and identified, and multigene phylogenetic analysis based on five gene loci (act, ITS, LSU, rpb2, tub2), morphological characterization, and pathogenicity tests were conducted. The results confirmed that Cytospora myrtagena is the causal agent of branch canker disease in bayberry, representing the first report of C. myrtagena infecting M. rubra. These findings provide a theoretical basis for the prevention and control of branch canker disease and offer important practical guidance for field management.

Osmanthus fragrans Lour. is a widely cultivated ornamental tree species in China and it is also a well-known aromatic and medicinal plant. In the People's Park, Zijingshan Park and Henan Academy of Agricultural Sciences in Zhengzhou City, this plant was found to develop canker on branches with a disease incidence rate of 80%-100%. A fungal species was isolated using tissue isolation methods. Pathogenicity was confirmed by inoculating a representative isolate onto healthy branches of O. fragrans. The pathogenic fungus was identified as Nothophoma quercina by using morphological characteristics and multiple gene sequence analysis of ITS, β-tubulin, and RPB2, which is a closely related genus of Phoma. This fungal pathogen produced olive-green colony on PDA and formed spherical pycnidia with one-cell ellipsoidal conidia of (2.74-4.07) μm×(3.56-7.20) μm in size. This study is the first report of N. quercina causing branch canker of O. fragrans in China.

Pinus bungeana, a unique and endangered coniferous tree in China, possesses significant ecological and ornamental value. A needle segment spot disease observed in Hancheng, Shaanxi Province, severely threatens the growth of P. bungeana. The causal pathogen was isolated from infected needles using the tissue isolation method, and the pathogenicity was confirmed according to Koch's postulates. Based on morphological characteristics and phylogenetic analysis of concatenated ITS, TEF1-α, and β-tubulin sequences, the pathogen was identified as Fusarium acuminatum. In vitro toxicity assays revealed that all six tested fungicides inhibited the mycelial growth of F. acuminatum to varying degrees. Among them, three triazole fungicides (tebuconazole, difenoconazole, and hexaconazole) demonstrated strong inhibitory efficacy, with EC₅₀ values of 0.206, 0.324, and 0.677 μg·mL^-1, respectively. To our knowledge, this is the first report of F. acuminatum causing needle segment spot on P. bungeana. The results of this fungicide screening provide a basis for developing precise disease control strategies.

In May 2024, a novel leaf spot disease was observed on Cayratia japonica in planting areas of Jinhua City, Zhejiang Province, China. Initial symptoms appeared as small water-soaked lesions that developed into irregular brown necrotic spots with distinct yellow halos, ultimately causing leaf perforation. The disease incidence ranged from 35% to 60%. The causal agent was isolated using the tissue isolation method, and its pathogenicity was confirmed by satisfying Koch's postulates. Based on morphological characteristics, cultural traits, physiological and biochemical profiling, and molecular identification (including phylogenetic analysis of 16S rDNA, purH, and rpoB sequences), the pathogen was identified as Enterobacter mori. To the best of our knowledge, this is the first report of E. mori causing leaf spot on C. japonica worldwide. This study provides a foundation for early diagnosis and integrated management of this emerging disease.

The red celery planted in the experimental field of the Inner Mongolia Autonomous Region Agriculture and Animal Husbandry Technology Promotion Center exhibited leaf yellowing and mottling symptoms. Through high-throughput sequencing analysis of the infected samples, celery gammacytorhabdovirus 1 (CelGCRV1) was detected. The complete genome sequence of CelGCRV1 was obtained using RACE technology combined with RT-PCR and was named 23IM_RC1 (GenBank accession number: PQ552775). The sequence was blasted against the GenBank database of NCBI using the BLASTn program, and one published CelGCRV1 sequence (GenBank accession number: BK064347) was found. In this study, 160 previously published L protein sequences of Rhabdoviruses were collected. Then, their phylogenetic relationship with the L protein of the 23IM_RC1 virus was analyzed. The results showed that 23IM_RC1 belongs to the Gammacytorhabdovirus genus. Further analysis indicated that 23IM_RC1 has the closest genetic relationship with the isolate from Zhejiang, China. The complete genome nucleotide homology between 23IM_RC1 and 18 isolates of Gammacytorhabdovirus ranged from 43.1% to 99.6%. No recombination events were found in the 23IM_RC1 genome using the seven algorithms provided by RDP4. This paper is the first report of CelGCRV1 detection in China. The results of this study provide basis for the development of CelGCRV1 detection techniques and the study on its pathogenic mechanisms.

Ditylenchus destructor is a significant agricultural quarantine nematode in China, which is capable of causing substantial yield reductions in potatoes. As Inner Mongolia is a major potato-producing region in China, identifying the haplotype of D. destructor infecting potatoes is critical for the quarantine and control of this pathogen. Nematodes solated from dry rot potato tubers, collected in Saihan District, were characterized morphologically, identified molecularly and verified pathogenicity. The morphological characteristics of the nematode population in Saihan District closely resembled those of D. destructor. The pathogenicity experiment showed that potatoes inoculated with the Saihan population developed specific symptoms of stem nematode infection after 40 days. Amplification using D. destructor-specific primers produced a band of approximately 375 bp, which confirmed the identity of the nematodes as D. destructor. The phylogenetic tree constructed based on rDNA-ITS sequences demonstrated that the nematode population clustered with the A-type of D. destructor. Comparative analysis revealed that the Saihan population shared the highest sequence similarity (99.71%) with an A-type population from Chifeng City, Inner Mongolia, among other potato-infecting A-type populations. In conclusion, the pathogen causing potato dry rot in Saihan District was identified as A-type of D. destructor. These finding provided a theoretical basis for developing improved quarantine and control strategies against D. destructor in this region.