Banana Fusarium wilt, caused by Fusarium oxysporum f. sp. cubense tropical race 4 (Foc TR4), poses a severe threat to the sustainable development of the global banana industry. Glycosylphosphatidylinositol (GPI)-anchored proteins play essential roles in maintaining fungal cell wall integrity and mediating host interactions; however, their functions in Foc TR4 remain largely unknown. In this study, we identified and characterized a GPI-anchored protein gene, FocGPI1, in Foc TR4. Bioinformatic analyses showed that FocGPI1 is highly conserved among plant pathogenic fungi and contains a functional signal peptide and a predicted GPI-anchoring site. We further generated knockout (ΔFocGPI1), complementation (FocGPI1-C), and overexpression (FocGPI1-OE) mutant strains via homologous recombination. Pathogenicity assays demonstrated that deletion of FocGPI1 significantly reduced the virulence of Foc TR4 on banana. ΔFocGPI1 also exhibited markedly impaired hyphal growth, conidiation and hyphal penetration. In addition, ΔFocGPI1 showed increased sensitivity to oxidative, and cell wall integrity stresses and altered utilization of cellulose, pectin, and chitin. Transcriptome analysis revealed that, during early infection, ΔFocGPI1 significantly induced host reactive oxygen species bursts and callose deposition, accompanied by upregulation of multiple plant defense-related genes. Subcellular localization analysis indicated that FocGPI1 exhibits a special dual localization at the plasma membrane and nucleus in plant cells. Collectively, FocGPI1 is a critical effector required for virulence in Foc TR4. It facilitates successful infection by modulating fungal growth, stress tolerance, and suppression of host basal immune responses. This study provides new insights into the pathogenic mechanisms of Foc TR4 and suggests that FocGPI1 could serve as a potential molecular target for effective control of banana wilt.

Continuous cropping obstacles in greenhouse muskmelon cultivation pose a significant threat to sustainable production. While leguminous green manures are known to mitigate soil degradation in other crops, their efficacy and micro-ecological mechanisms in muskmelon systems remain unexplored. Here, we demonstrate for the first time that winter planting of two leguminous green manures, common vetch (Vicia sativa L.) and smooth vetch (Vicia villosa Roth var. glabresens Koch), during fallow periods alleviates continuous cropping obstacles by reshaping soil micro-ecology. Field trials revealed that both green manures significantly increased muskmelon yield (13.71% and 10.68%, respectively), elevated soil pH and organic matter, and reduced salinity (EC by 51.1% and total salt by 35.7%). High-throughput sequencing uncovered enriched microbial diversity, with beneficial taxa (Pirellula, Gemmata, Myceliophthora, Talaromyces) positively correlated with yield, while suppressing pathogenic fungi (Fusarium). Redundancy analysis highlighted soil pH and organic matter as key drivers of beneficial microbial recruitment, whereas salinity promoted harmful taxa. This study establishes a green manure-driven micro-ecological remediation framework, providing a cost-effective strategy for sustainable muskmelon cultivation in southern China.

Melon (Cucumis melo L.) is an important economic crop in China, with a planting area of about 500,000 hectares, ranking first in the world. In May 2024, anthracnose symptoms were found on melon plants, particularly severe on the mature fruits, in Binyang County, Guangxi, China. Disease incidence was between 30% to 60% in four surveyed planting areas. The symptoms on fruits initially appeared as water-soaked lesions, gradually turning into dark brown sunken lesions, sometimes with cracks. Additionally brown necrotic lesions with yellowish edges appeared on the leaves. For pathogen isolation, lesion edge tissues (3×3 mm) of fruits were surface-sterilized in 75% ethanol (30 s) and 1% sodium hypochlorite (1 min), rinsed in sterile distilled water, and plated on potato dextrose agar (PDA) amended with streptomycin sulphate (30 mg/l) for 4 days at 28°C in the dark. Ten pure isolates with similar morphology were obtained by transferring hyphal tips to new PDA plates. Colonies were round with smooth margins. Mycelium was sparse, initially pale gray, then changed to dark gray with numerous black microsclerotia after 14 days and generated a small amount of orange conidial masses afer 30 days of cultivation. Conidia were single-celled, hyaline, slightly curved, tapered tip and truncate base, with an oil globule at center, and 18.9 to 22.2 × 3.2 to 4.7 μm (n = 50). Setae initiated from an acervuli, were dark brown, septate, straight, pointed, and measuring 85.5 to 146.3 × 4.2 to 5.5 μm. Appressoria were light brown, elliptic to claviform or slightly lobed. Morphological characters were similar to Colletotrichum truncatum (Damm et al. 2009). Two representative isolates M1 and M3 were used for molecular identification. The partial internal transcribed spacer (ITS) region, actin (ACT), β-tubulin (TUB2), chitin synthase (CHS-1), and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) genes were amplified with ITS1/ITS4, ACT512F/ACT783R, BT2A/BT2B, CHS-79F/CHS-345R, and GDF1/GDR1 primers, respectively. Sequences were deposited in GenBank (ITS: PQ549938, PQ549939; Actin: PQ562860, PQ562861; TUB2: PQ562866, PQ562867; CHS-1: PQ562862, PQ562863; GAPDH: PQ562864, PQ562865) and showed 97% to 100% similarity with C. truncatum strains. A maximum likelihood phylogenetic tree based on the concatenated these five loci in MEGA-X showed the clustering of the isolates M1 and M3 in the C. truncatum clade. Pathogenicity tests were performed twice in a greenhouse at 25 to 30°C with 90% relative humidity. The healthy living fruits were slightly wounded by sterilized needle. Then spore suspension (106 conidia/ml) of isolates M1 and M3 were inoculated onto the wounds (10 μl/wound). For each isolate, five fruits were inoculated. Control fruits were treated with sterile water. After 7 days, all the inoculated fruits showed brown lesions resembling natural symptoms, whereas no symptoms appeared on the negative controls. The same fungus was re-isolated from the symptomatic fruits, thus completing Koch’s postulates. Based on morphological and molecular characteristics and a pathogenicity test, the pathogen was identified as C. truncatum. Previously, C. truncatum was reported to cause melon anthracnose in Brazil (Assunção et al. 2024) and watermenlon anthracnose in China (Guo et al. 2022). To our knowledge, this is the first report of C. truncatum causing anthracnose on melon in China. Knowing the causal agent is important to control this disease effectively.

To screen out the bio-control soil microorganisms for preventing melon wilt, soil microbial compositions in rhizospheres between wilt-resistant and susceptible melon varieties were analyzed. The results showed that the soil fungal richness in rhizospheres of wilt-resistant melon varieties (MT) was significantly higher than that of wilt-susceptible melon varieties (MS). Additionally, in comparison with MS, soil bacterial compositions, such as Proteobacteria, Bacteroidota, Acidibacter, Streptomyces, etc., and the soil fungal compositions, such as Penicillium, Derxomyces, Aspergillus, and Talaromyces, enriched; also, Trichoderma, Gibellulopsis, and Pseudallescheria decreased in rhizospheres of wilt-resistant melon varieties (MT). Moreover, Mycothermus, Zopfiella, and Cladorrhinum were the unique soil-dominant fungal genera in rhizospheres of MT. All the above results suggested that the soil bacterial communities, such as Proteobacteria, Bacteroidota, Acidibacter, Streptomyces, etc., and the soil fungal communities, such as Penicillium, Derxomyces, Aspergillus, Talaromyces Mycothermus, Zopfiella, and Cladorrhinum, could be speculated as the potential soil bio-control microorganisms for preventing melon wilt.

Pear (Pyrus pyrifolia) is an economically important fruit and widely planted in China. In September 2023, soft rot disease was observed on more than 20% of pear fruits (cv. Jinghua) in a plantation of approximately 1.0 ha of Shilin County, Kunming City, Yunnan Province, China. The initial symptoms manifested as water-soaked lesions. Then, they rapidly expanded and covered the whole fruit, which was soft and necrotic. At severe stage, infected fruits turned brown and dropped prematurely. To isolate and identify the causal pathogen, two diseased fruits were collected and pieces of rotted fruit tissue were cut into small fragments (5×5 mm), disinfested in 75% ethanol (30 s) and 2% sodium hypochlorite (60 s), and rinsed three times with sterile distilled water. The sterilized sections were soaked in 2 ml of sterile water and shaken for 3 min in a vortex oscillator. The suspension was streaked on Luria-Bertani (LB) agar medium. After incubation at 28°C for 36 h, single colonies were restreaked three times to obtain purified isolations. Thirteen bacterial strains with similar morphology were isolated from different fruits (seven strains in one fruit and six in the other), and their colonies were yellowish white, round, and convex with smooth surfaces on the LB agar plates. Representative strains YNPA1 and YNPA2, isolated from different fruits, were selected for further analyses. The two strains were Gram-negative, tested negative for citrate and catalase, but positive for sucrose and glucose. The 16S rRNA gene of these strains (GenBank accession nos. PP917741, PP917742) was amplified using primer pair 27F/1492R and sequenced. BlastN searches revealed that the obtained sequences shared >99% identity with Pectobacterium aroidearum type strain (GenBank accession no. NR_159926). A multilocus sequence analysis based on concatenated sequences of five housekeeping genes was conducted, of which gyrA (product sizes: 987bp, GenBank accession no. PP928290, PP928291), icdA (589bp, PP928292, PP928293), mdh (547bp, PP928288, PP928289), proA (734bp, PP928294, PP928295), and rpoS genes (892bp, PP928296, PP928297) was amplified, using primer pair gyrA1/gyrA4, icdA400F/icdA977R, mdh86F/mdh628R, proAF1/proAR1, rpoS1/rpoS2, respectively (Ma et al. 2007; Waleron et al. 2008). The reconstructed maximum likelihood tree (Tamura-Nei model; with 1,000 bootstrap replicates) showed that strains YNPA1 and YNPA2 clustered with strains of P. aroidearum. Pathogenicity tests were performed on three healthy pear fruits by injecting 10 μl of bacterial suspensions of P. aroidearum strains YNPA1 and YNPA2 (108 CFU/ml), respectively; another three healthy control pear fruits were injected with 10 μl of sterile water. All tested fruits were covered with plastic bags and incubated at 28°C and 80% humidity in a growth chamber. After 3 days, all inoculated pear fruits showed soft rot symptoms resembling those observed in the plantation, while control fruits remained symptom-free. Bacteria were successfully reisolated from the symptomatic tissues and identified as P. aroidearum by PCR and sequencing of the 16S rRNA gene as described above. P. aroidearum has been reported as a pathogen of Syngonium podophyllum (Xu et al. 2020) and Prunus persica (Liang et al. 2022) in China. The current study expands the known host range of P. aroidearum and helps raise attention for controlling pathogen's spread.

Melon is an important horticultural crop. It has a long history of cultivation in China, which is the country with the largest production and consumption of melon. In March 2023, at the Modern Agricultural Experiment Station in Guangxi, China (23°15'26" N, 108°27'47" E), we observed bacterial fruit blotch symptoms on about 10% of 'Zhongtian No. 9' melon plants in an 8 ha field. Symptoms were mainly on two-week-old cotyledons. Leaves were collected from five sites. Infected tissues were sterilized with 75% ethanol, cut into 4 × 4 mm pieces, and ground in ddH₂O. The suspension was spread on KB agar plates and incubated at 28℃ for 2 days. Thirty single colonies were selected and streaked on new KB agar plates. The colonies were milky white, round, smooth, raised, opaque, and 1-2 mm in diameter. Three single colonies were isolated and sequenced for 16S rRNA gene. BLAST analysis showed >99% nucleotide sequence homology with Paracidovorax citrulli ICMP 7500 (NR_041758) and representative strain M6 (CP029373). One strain, named GXM1, had its 16S rRNA gene sequence deposited in NCBI with accession number PQ461200. Specific primers of P. citrulli BFB/BFB1 and BFB/BFB2 (Yang et al. 2019) were used for further identification, the result showed that the strain belonged to P. citrull group I. The whole genome of GXM1 was sequenced and assembled and submitted to NCBI under accession number GCA_042610265.1, with 99.3% genome-wide similarity to the P. citrulli model strain ICMP7500 (NR_041758.1) (based on dDDH analysis; Meier-Kolthoff et al. 2013) and 99.69% (based on ANI analysis; Richter et al. 2015). In the Biolog Microbial Identification System, Version 4.2 (Biolog Inc., Hayward, CA), GXM1 were similar with a match probability of 77.5% to P. citrulli. P.citrulli can trigger hypersensitive response in Nicotiana tabacum (Zhang et al., 2018). The GXM1 bacterial suspension (3×108 CFU·mL-1) was injection into 4-week-old N. tabacum leaves. The results showed typical HR after 24 h post injection. GXM1 was inoculated into nine watermelon and nine melon plants (10⁶ CFU·mL⁻¹). A 10 mM MgCl₂ solution served as the negative control, and the strain AAC00-1 (NC_008752.1) as the positive control, each inoculated into nine plants. After inoculation, plants were maintained at 75% relative humidity and 28℃ for 2 days. Plants inoculated with GXM1 and AAC00-1 exhibited typical bacterial fruit blotch (BFB) symptoms: irregular brown necrotic spots with faint chlorotic halos. The negative control plants remained symptomless and healthy. The pathogenicity tests were repeated three times with the similar results. Bacteria isolated from diseased melon leaves formed creamy white, circular, smooth, convex, opaque colonies on KB agar, consistent with the original isolate. Sequencing of the 16S rRNA genes of three colonies and BLAST analysis showed nucleotide sequence homology exceeding 99% with the type strain P. citrulli ICMP 7500. Based on pathogenic, biochemical, and molecular characteristics, the isolates from melon plants in Guangxi province (China) were identified as Paracidovorax citrulli. This is the first report of Paracidovorax citrulli infecting melon in Guangxi. Considering favorable environmental conditions, the pathogen can cause significant loses in melon production in Guangxi province (China). It is necessary to take phytosanitary measures to prevent spread of the pathogen.

Syzygium grijsii is an evergreen shrub belonging to the family Myrtaceae, and widely cultivated in southern China as an ornamental medicinal plant. In May 2022, anthracnose symptoms were observed on leaves of S. grijsii planted in a nursery (N22°55'46″, E108°22'11″) in Nanning, Guangxi Province, China. More than 30% of leaves were infected. Initially, irregular brown spots (1 to 2 mm in diameter) formed on the leaves, with a slight depression in the center, then expanded into large, dark-brown lesions. In severe infections, lesions coalesced and covered the entire leaf, causing wilt and fall off the plant. To identify the pathogen, 30 diseased leaves were collected from five plants. Leaf tissues (5 × 5 mm) were cut from the infected margins, surface sterilized (75% ethanol 10 s, 2% NaClO 5 min, rinsed three times with sterile water), then placed on potato dextrose agar (PDA), and incubated at 28℃ in darkness. After 5 days, 16 fungal isolates with similar morphology were obtained from 30 plated tissues. Colonies on PDA were abundant with grayish-white fluffy mycelia, and yellowish-white on the back. Conidia were one-celled, hyaline, smooth-walled, cylindrical with narrowing at the center, blunt at the ends, and ranged from 11.35 to 22.14 × 4.88 to 7.67 μm (n=100). Morphological characteristics of the isolates were similar to the descriptions of Colletotrichum sp. (Prihastuti et al. 2009). Five representative isolates (Cs34, Cs31, Cs32, Cs33 and Cs35), which were preserved in the Guangxi Key Laboratory of Biology for Crop Diseases and Insect Pests, were selected for molecular identification. The ITS (Nos. OQ618199, OR539576 to OR539579), TUB2 (Nos. OQ630972, OR545076 to OR545079), ACT (Nos. OQ685919, OR545060 to OR545063), CHS-1 (Nos. OQ685917, OR545068 to OR545071), GAPDH (Nos. OQ685916, OR545072 to OR545075), and CAL (Nos. OQ685918, OR545064 to OR545067) sequences showed >99% identity to those of Colletotrichum siamense ex-type culture ICPM 18578 (Nos. JX010171, JX009924, JX009714 and JX009518) and strain C1315.2 (Nos. JX009865 and JX010404) in GenBank. Multigene phylogenetic analyses (ITS, TUB, ACT, CHS-1, GAPDH, and CAL) using the Maximum likelihood method indicated that the 5 isolates were clustered with C. siamense. To perform pathogenicity tests, three one-year-old healthy S. grijsii plants were inoculated with conidial suspension (1 × 106 conidia/ml) of isolate Cs34 by brushing gently with a soft paintbrush, each plant was inoculated with 3 leaves. The same number of plants were inoculated with sterile water as control, and pathogenicity tests were performed three times. All plants were kept in an artificial climatic box at 28℃, with a 90% humidity and a 12 h light/dark cycle. Similar symptoms to those of the field were observed on all inoculated leaves after 5 days, whereas controls remained symptomless. Reisolated fungi from the diseased leaves were confirmed to be C. siamense by morphology and molecular characterization, confirming Koch's postulates. C. siamense has been reported causing anthracnose on Crinum asiaticum (Khoo et al. 2022) in Malaysia, and Erythrina crista-galli in China (Li et al. 2021). To our knowledge, this is the first report of C. siamense causing anthracnose on S. grijsii in China. The results of pathogen identification provide crucial information for control strategies of the disease.

Studies have shown that plant endophytic microbial communities are ubiquitous and closely related to plant growth and health. To clarify the mechanism of the melon varieties with high resistant to wilt, the endophytic microbial compositions and metabolites in roots of melon varieties with high resistant ability to wilt were analyzed. The results showed that the abundances of Firmicutes, Ascomycota, Bacillus, Bradyrhizobium, Amycolatopsis, Actinospica, and Catenulispora all increased in roots of wilt high resistant melon varieties (MT) which compared to wilt susceptible melon varieties (MS). Meanwhile, Ochrobactrum, Bordetella, Roseateles, Staphylococcus, Acidovorax, Amycolatopsis, Catenulispora, Promicromonospora, and Gymnopilus were the unique endophytic microbes in roots of MT. Moreover, in comparison with the MS varieties, the functions of Defense mechanisms, Secondary metabolites biosynthesis, transport and catabolism, Nucleotide transport and metabolism, Signal transduction mechanisms, Coenzyme transport and metabolism, Carbohydrate transport and metabolism and Amino acid transport and metabolism all increased in roots of MT varieties. Additionally, the nucleotide metabolism and biosynthesis of cofactors metabolic pathways were also significantly increased in roots of MT varieties. On the other hand, the untargeted metabolome results showed that Biosynthesis of various plant secondary metabolites, Nucleotide metabolism and Biosynthesis of cofactors metabolic pathways were significantly increased in the expression of MT varieties; and the content of metabolic compounds such as flavonoids, Cinnamic acid compounds, Organic acid compounds, and Nucleotides were increased. In addition, the correlation between microbiome and metabolome indicates a significant correlation between the two. All above results suggested that higher abundant antagonistic microbes and metabolic functions of endophytes in roots of wilt high resistant melon varieties (MT) were the important mechanisms for their high resistance to wilt.

Cordyline fruticosa is a shrub plant, commonly used in landscape, and distributed in the tropical regions of southern China. In September 2022, anthracnose symptoms were found on this species in Nanning, Guangxi, China. The disease incidence was between 30% to 80% and disease severity was 10% to 30% in five surveyed planting areas. The symptoms initially appeared as small, round, brown spots on leaves. As the disease developed, the lesions turned gray-white with brown borders and yellow halos. Some spots coalesced into larger irregular shapes and even leading to leaf blight. Small segments of the diseased tissues (3×3 mm) were cut from the leaves, surface-sterilized by dipping in a 1% sodium hypochlorite solution for 1 min, rinsed three times with sterile distilled water, and plated on potato dextrose agar (PDA). These plates were incubated at 28°C in the dark for 5 days. Ten fungal isolates with similar morphology were consistently isolated from these diseased tissues. The colonies on PDA were initially white with sparse aerial mycelia and turned pale orange with abundant orange conidial masses on the center after 8 days of culture. The reverse color was pale orange. No sclerotia or setae were found in culture. Conidia were single-celled, hyaline, straight, cylindrical with round ends, and 12.2 to 17.8 µm long (mean 14.9 µm) and 3.9 to 7.3 µm wide (mean 4.8 µm, n=50). The morphological characteristics of these isolates were similar to the Colletotrichum cordylinicola (Sharma et al., 2014). Genomic DNA of two isolates Z3 and Z4 generated from monospore culture was extracted using a fungal DNA extraction kit (Solarbio, Beijing, China). Partial sequences of internal transcribed spacer (ITS), partial actin (ACT), chitin synthase (CHS-1), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), and beta-tubulin (TUB2) were amplified using the primer pairs ITS1/ITS4, ACT-512F/ACT-783R, CHS-79F/CHS-345R, GDF1/GDR1, and BT2A/BT2B (Lin et al., 2022), respectively. All the sequences (GenBank accession nos. OQ509909, OQ509910, OQ658690, OQ658691, and OK649310 to OK649314) showed 99% to 100% identity with those of C. cordylinicola in GenBank database. A phylogenetic tree based on concatenated sequences of ITS, ACT, CHS-1, TUB, and GAPDH using maximum likelihood analysis by MEGA X software revealed that Z3 and Z4 clade with reference strains of C. cordylinicola (OJX010226 and MK935473). Based on morphological observation and multi-gene sequence analysis, the isolates were identified as C. cordylinicola (Phoulivong et al., 2010). To assess their pathogenicity, conidial suspensions (106 conidia/ml) of C. cordylinicola were inoculated onto 10 healthy living leaves wounded by slight puncturing (10 μl/wounded spot). Control leaves were treated with sterile water. All inoculated and control plants were maintained under high relative humidity (~90%) and 28℃ in a climate chamber. After 8 days, all the inoculated leaves showed brown lesions resembling natural symptoms, whereas the control group remained symptom-free. The same fungus was re-isolated from the symptomatic leaves, thus completing Koch's postulates. C. cordylinicola is a species of the C. gloeosporioides complex (Weir et al., 2012). It has been reported to cause anthracnose on C. fruticosa in USA and Thailand (Phoulivong et al., 2010; Sharma et al., 2014). To our knowledge, this is the first report of C. cordylinicola causing anthracnose on C. fruticosa in China. Knowing the causal agent is essential to control the serious disease effectively.

Banana is an important fruit and food crop in tropical and subtropical regions worldwide. Banana production is seriously threatened by Fusarium wilt of banana (FWB), a disease caused by Fusarium oxysporum f. sp. cubense, and biological control is an important means of curbing this soil-borne disease. To reveal the effects of biocontrol agents on inhibiting FWB and altering the soil bacterial community under natural ecosystems, we conducted experiments at a banana plantation. The control efficiency of a compound microbial agent (CM), Paenibacillus polymyxa (PP), Trichoderma harzianum (TH), and carbendazim (CA) on this disease were compared in the field. Meanwhile, the alterations in structure and function of the rooted soil bacterial community in different treatments during the vigorous growth and fruit development stages of banana were analyzed by microbiomics method. The results confirmed that the different biocontrol agents could effectively control FWB. In particular, CM significantly reduced the incidence of the disease and showed a field control efficiency of 60.53%. In terms of bacterial community, there were no significant differences in the richness and diversity of banana rooted soil bacteria among the different treatments at either growth stage, but their relative abundances differed substantially. CM treatment significantly increased the ratios of Bacillus, Bryobacter, Pseudomonas, Jatrophihabitans, Hathewaya, and Chujaibacter in the vigorous growth stage and Jatrophihabitans, Occallatibacter, Cupriavidus, and 1921-3 in the fruit development stage. Furthermore, bacterial community function in the banana rooted soil was affected differently by the various biocontrol agents. CM application increased the relative abundance of multiple soil bacterial functions, including carbohydrate metabolism, xenobiotic biodegradation and metabolism, terpenoid and polyketide metabolism, lipid metabolism, and metabolism of other amino acids. In summary, our results suggest that the tested biocontrol agents can effectively inhibit the occurrence of banana Fusarium wilt and alter the soil bacterial community in the field. They mainly modified the relative abundance of bacterial taxa and the metabolic functions rather than the richness and diversity. These findings provide a scientific basis for the use of biocontrol agents to control banana Fusarium wilt under field conditions, which serves as a reference for the study of the soil microbiological mechanisms of other biocontrol agents.

Banana (Musa spp.) is an economically important fruit and food crop globally as well as in China. In March 2023, a bulb rot disease was observed on more than 20% of cultivated dwarf bananas in a plantation in Wuming County of Guangxi Province, a major hub of banana production in China. Infected plants showed crackles at the basal part of stem and were relatively dwarf, while yellowing of the leaves was not observed. When the rhizomes were cut open, water-soaked lesions with a yellow or black margin can be seen in the bulb. In severe infections, the internal tissue became dry or wet rot, and there was typical dark-brown cavity formation in the bulb. The rot was limited to the bulb. To isolate the causal agent, dissected diseased tissues (5×5 mm) were surface sterilized with 75% ethanol (30 s) and 2% NaClO (3 min), followed by three rinses with sterile water. The sterilized sections were soaked in 2 mL of sterile water and shaken for 5 min in a vortex oscillator. The suspension was streaked on Luria-Bertani (LB) agar medium, and incubated at 28℃ for 24 h. Single colonies were re-streaked three times to obtain purified isolation. Twelve pure bacterial cultures with similar morphology were isolated from three plants taken from the field. The bacterial colonies were yellowish white, mucoid, round, and raised with translucent surfaces on the LB agar plate. Three strains Gxkv1, Gxkv2 and Gxkv3 were selected for further analyses. The 16S rDNA gene (GenBank Accession OR461756, PP094726 and PP109349) were amplified using primer pair 27F/1492R (Frank et al. 2008). Comparing 16S sequences against GenBank showed 99.86%-100% sequence identity to Klebsiella variicola strain (MZ475068) for the three isolates Gxkv1 (1,398/1,398 bp), Gxkv2 (1,398/1,396 bp) and Gxkv3 (1,398/1,398 bp). A multilocus phylogenetic analysis was conducted by neighbor-joining method (1,000 bootstrap values) based on three housekeeping gene sequences of gyrA (GenBank Accession No. OR515493, PP105747, PP105748), rpoB (OR515494, PP105751, PP105752 ) and infB (OR515495, PP105749, PP105750) genes which were amplified by gyrA-A/gyrA-C, CM31b/CM7 and infB867F/infB1819R primer sets, respectively (Rosenblueth et al. 2004). The results of phylogenetic analysis showed the three strains belong to the K. variicola clade. A pathogenicity test was conducted on six healthy 3-month-old dwarf banana plants by spraying 10 mL of bacterial suspensions of Gxkv1 (108 CFU/mL) into the rhizome which wounded with a sterilized needle; another six healthy control plants were sprayed with 10 mL of sterile water. Following inoculation, the plants were placed in a greenhouse at 28-32°C. After 30 days, all inoculated plants showed symptoms similar to those observed in the field, while the control plants remained healthy. Bacteria were successfully reisolated from the symptomatic tissues and identified to be K. variicola by PCR mentioned above. K. variicola has been reported to cause rhizome rot of banana in India (Loganathan et al. 2021), and to cause plantain soft rot in Haiti (Fulton et al. 2021). Besides, previous reports from China only showed K. variicola causing banana sheath rot (Fan et al. 2015, Sun et al. 2023). To our knowledge, this is the first report of bulb rot disease of banana caused by K. variicola in Guangxi Province, China. This finding will provide important information for studying the epidemiology and management of this pathogen.

Sweet persimmon (Diospyros kaki L.) is a fruit of significant nutritional and commercial value in Asia. In summer 2023, leaf spots were observed affecting 20 to 30% of sweet persimmon trees in a commercial orchard located in Gongcheng City, Guangxi, China. Initially, the infected leaves exhibited sparse light brown spots on their upper surface, which subsequently evolved into brown circular to irregular lesions encircled by a yellow halo. Eventually, these lesions became densely distributed across the leaves leading to insufficient nutrient accumulation in the fruit. To isolate the pathogen, diseased leaves were cut into small pieces (5×5 mm), disinfected with 75% ethanol for 15 seconds, followed by 1% NaClO for 1minute, rinsed three times with sterile water, and then transferred onto potato dextrose agar (PDA) plates. The plates were then incubated in darkness for 3 days at 25°C. Pure cultures were obtained using the hyphal-tip method and single-spore isolation. On PDA, the colonies initially appeared fluffy and white after 24 hours, turning yellowish or red after 3 days. Macroconidia (average length of 26.1 μm in length × 4.3 μm in width, n = 50) exhibited dorsiventral curvature and were hyaline, with 3 to 5 septa. Microconidia (average length of 9.45 μm in length × 3.4 μm in width, n = 50) were hyaline, aseptate, and oval. Two representative isolates, Gxfky1 and Gxfky2, were selected for further molecular analyses. Their internal transcribed spacer (ITS) region rDNA gene were amplified via PCR and sanger sequenced (GenBank Accession Nos. PP506475, PP506593) using the primer pair ITS1/ITS4 (White et al. 1990), showing more than 99% sequence identity with Fusarium kyushuense type-material strain NRRL3509 (NR_152943) according to BLASTn analysis in NCBI. To further confirm the identity of the isolates, four gene sequences were amplified: RPB1 (PP532864, PP532865), RPB2 (PP532866, PP532867), TEF1 (PP580505, PP580506), and TUB2 (PP532862, PP532863), using the F5/G2R, 5f2/11ar, EF1/EF2, and T1/T2 primer sets, respectively (O'Donnell et al., 1997; O'Donnell et al., 2010). A multi-locus maximum likelihood phylogenetic analysis revealed that Gxfky1 and Gxfky2 clustered with strains F. kyushuense with 100% bootstrap support. Pathogenicity tests using Gxfky1 and Gxfky2 were conducted on leaves of two-year-old sweet persimmon plants using non-wound inoculation. Specifically, 5-mm mycelial plugs and sterile agar plugs were placed on six leaves and secured with cling film, with six plugs each for the inoculation treatment and negative control, respectively. They were then incubated in a greenhouse at room temperature (25 ± 2°C) with a relative humidity of 70 to 80%. After 5 days, the same symptoms on naturally infected plants were observed on leaves inoculated with mycelium, while no symptoms were observed on the controls. The same fungus were reisolated from the inoculated leaves and identified based on morphology and the TEF1 gene sequence, thus fulfilling Koch's postulates. Fusarium kyushuense has previously been reported to cause diseases in various plant species, including maize (Cao et al., 2021), rice (Wang et al., 2024), and tobacco (Wang et al., 2013). To our knowledge, this is the first report of F. kyushuense causing leaf spot on sweet persimmon in China, which expands the known host range of this pathogen.

[目的]从不同作物的根围土壤中分离筛选香蕉枯萎病菌拮抗菌,对拮抗菌进行鉴定并测定其防病效果,以期为香蕉枯萎病的生物防治提供新的菌种资源.[方法]以尖孢镰刀菌古巴专化型4号生理小种(Fusarium oxysporum f.sp.cubense 4,Foc4)Foc 1402菌株为对象,采用稀释涂布法对不同作物根围土壤中的细菌进行分离,并使用平板对峙法和牛津杯法筛选其中的香蕉枯萎病拮抗菌.根据细菌菌落形态、生理生化特征结合16S rDNA序列分析对拮抗菌进行分类鉴定.使用威廉斯B6香蕉杯苗测定拮抗菌对香蕉枯萎病的盆栽防治效果.[结果]从不同作物根围分离获得345株细菌,从中筛选获得7株对香蕉枯萎病菌具有较强拮抗活性的菌株,抑菌率最高可达72.65％.依据形态学观察、生理生化反应和16S rDNA序列分析结果,菌株Ba02、Ba310、Ba48、BBa62和Ba63被鉴定为解淀粉芽孢杆菌(Bacillus amyloliquefaciens),菌株Be11被鉴定为洋葱伯克霍尔德氏菌(Burkholderia cepacia),菌株Pt05被鉴定为土地类芽孢杆菌(Paenibacillus terrae).盆栽试验结果显示,7株拮抗细菌对香蕉枯萎病的防治效果在20.00％～68.89％,其中以菌株Be11的防治效果最好,为68.89％.[结论]从不同作物的根围土壤中筛选获得7株对Foc具有较强拮抗活性的细菌,其中,土地类芽孢杆菌为香蕉枯萎病生防菌家族的新成员.筛选获得的不同种类拮抗菌可作为研制香蕉枯萎病生防菌剂的候选菌株资源,在后续开发抗病复合菌剂方面具有良好的应用前景.

Acidovorax citrulli is a seed-borne bacterium that causes bacterial fruit blotch of watermelon and other cucurbit plants worldwide. It uses a type III secretion system to inject type III effectors (T3Es) into plant cells, which affect the host immune responses and facilitate pathogen colonization. However, the current understanding of the specific molecular mechanisms and targets of these effectors in A. citrulli is limited. In this study, we characterized a novel T3E called AopU in A. citrulli group II strain Aac5, which shares homology with XopU in Xanthomonas oryzae. The Agrobacterium-mediated gene transient expression system was used to study the effect of AopU on host immunity. The results showed that AopU localized on the cell membrane and nucleus of Nicotiana benthamiana, inhibited reactive oxygen species burst induced by flg22 and the expression of marker genes associated with pathogen-associated molecular pattern-triggered immunity, but activated salicylic acid and jasmonic acid signal pathways. Further investigations revealed that AopU interacts with E3 ubiquitin ligase ClE3R in watermelon, both in vitro and in vivo. Interestingly, the deletion of aopU did not affect the virulence of A. citrulli, suggesting that AopU may have functional redundancy with other effectors in terms of its role in virulence. Collectively, these findings provide new insights into the mechanism of plant immune responses regulated by A. citrulli T3Es.

Pomegranate (Punica granatum L.) is a deciduous shrub or small tree that is native to Iran and Afghanistan. It is also a commercially important fruit tree in China and worldwide. In the summer of 2022, a serious root rot disease occurred in some pomegranate orchards in Xichuan County(32º42´ N, 111º48´ E), Henan Province, China, with an incidence of ~30%. Symptoms included leaf yellowing and wilting, root browning and rotting, and stem-base cracking, eventually leading to defoliation and death. To isolate the causal agent, small pieces (5×5 mm) of diseased root from six trees were surface-sterilized by dipping in 2% NaClO for 8 min followed by 70% ethanol for 15 s, rinsed five times with sterile water, and plated on potato dextrose agar (PDA), then incubated at 28°C in the dark for 5 days. Fifteen pure fungal isolates with the same morphological characteristics were obtained from 24 pieces of roots. All isolates produced white fluffy mycelia. Microconidia were hyaline, oval or reniform, with zero to one septa and dimensions of 7.1 to 19.9 (average 14.5 )× 3.8 to 8.0 (average 5.6) μm (n = 100). Macroconidia were sickle-shaped, one to four septate, and 20.1 to 40.8 (average 26.5) × 4.8 to 8.6 (average 6.5) μm (n = 100). Chlamydospores were spherical, single, in pairs or chains, and 5.6 to 9.8 (average 6.8) µm in diameter (n = 100). Based on the above characteristics, the pathogens were identified as Fusarium sp. (Leslie and Summerell 2006). Genomic DNA was extracted from mycelia of two representative isolates Fs1 and Fs3. The internal transcribed spacer (ITS), translation elongation factor 1-alpha (TEF-1α) and RNA polymerase II second largest subunit (RPB2) sequences were PCR amplified using primer pairs of ITS1/ITS4, EF1/EF2, and RPB2-5f2/RPB2-7cr, RPB2-7cf/RPB2-11ar (O'Donnell et al., 2022), respectively. BLAST analysis showed that the ITS, TEF-1α and RPB2 sequences of isolates Fs1(GenBank accession nos. OK001765, OQ921726 and OQ928396) and Fs3 (GenBank accession nos. OK001771, OQ921727 and OQ928397) showed 99%-100% identity with multiple GenBank sequences of Fusarium falciforme (KY617066, MN064683, KF255514, OQ933361, KY556711 and ON331935). A phylogenetic tree based on concatenated sequences of ITS, TEF-1α and RPB2 using maximum-likelihood analysis revealed that both isolates Fs1 and Fs3 were in the same clade with F. falciforme strains. Based on the morphological and molecular characteristics, the isolates were identified as members of F. falciforme. For pathogenicity testing, conidial suspensions (1×108 spores /mL) of isolates Fs1 and Fs3 were poured onto the roots of healthy pomegranate that had been planted in pots two months previously. Ten plants were inoculated for each isolate. Control plants were drenched with sterile water. After 3 months, inoculated plants developed leaf yellowing and wilting accompanied by root browning and rotting, much like symptoms observed in field plants. The same fungi re-isolated from the experimental plants were confirmed to be F. falciforme by morphology and sequence analysis. This is the first report of F. falciforme causing root rot on pomegranate. F. falciforme is a ubiquitous soil-borne pathogen that causes root rot on multiple plants around the world (Xu F., et al. 2022; Qiu R., et al. 2023). The results of pathogen identification are essential precursors to development of effective control of the disease.

Rosemary (Rosmarinus officinalis L.) is an aromatic, evergreen, medicinally important shrub and widely used for cooking, tea, cosmetics as well as medicinal materials. It is grown in many countries including China that had more than 9300 hm2 of commercial cultivation area in 2021. In March 2020, a leaf spot disease sporadic occurred in field rosemarry plants in Nanyang City (32º51´ N, 111º36´ E), Henan Province, China. The disease outbreaked in September with a disease incidence of 57-83%. Symptoms initially appeared as small brown leaf spots that gradually expanded into dark blackbrown irregular lesions. Most of the spots started from the leaf tip or leaf margin, and gradually spread to the leaf base, resulting in heavy defoliation especially on rainy days. Diseased leaf segments (1×3 mm) were surface-sterilized by dipping in 1% sodium hypochlorite for 1 min, rinsed three times with sterile distilled water, and plated on potato dextrose agar, then incubated at 28°C in the dark for 5 days. Twelve fungal isolates with the same morphological characteristics were obtained from nine affected leaves. The fungal colonies were initially white and turned gray brown with flocculent aerial mycelia and a whorled back. Conidia were frequently born in a long chain, with a short beak, brown or light-brown, 13.2 to 48. 7 (average 26.1) × 4.0 to 13.1 (average 8.0) μm in size (n=148) with 0 to 8 transverse and 0 to 3 longitudinal/oblique septa. Phenotypic features of the isolates agreed with those of Alternaria alternata (Simmons et al. 2007). Two isolates Aa1 and Aa2 were randomly selected for molecular and pathogenicity tests. DNA was extracted from mycelia. Partial sequences of internal transcribed spacer (ITS) and translation elongation factor 1-alpha (TEF1-α) were amplified using the primer pairs ITS1/ITS4 and EFI-728F/EFI-986R (Wei et al. 2022), respectively. The GenBank accession nos. were OK036714 and OK036715 for ITS, and ON951980 and ON951981 for TEF1-α of Aa1 and Aa2, respectively, with a maximal identity of greater than 99% to multiple A. alternata strains. In the neighbour joining phylogenetic tree of the amplified ITS and TEF1-α sequences both Aa1 and Aa2 clustered with A. alternata strains, clearly separating them from other Alternaria spp. For pathogenicity test, conidial suspensions (1×106 spores /mL) of Aa1 and Aa2 were separately sprayed on healthy one-year-old rosemary plants (n=3) with their leaves slightly wounded with a sterilized needle. Control plants (n=3) were sprayed with sterile water. Both inoculated and control plants were incubated at 90% RH, 28 °C. After 14 days, all the inoculated leaves showed black brown lesions similar to those on naturally affected field plants, whereas controls remained symptomless. Fungal cultures with the same phenotypic features as the inocula were constantly re-isolated from the infected leaves. A. alternata was reported as pathogen causing foliar necrosis on rosemary in Italy (Perello et al.1995) and leaf spot (or leaf blight) on multiple plant species such as Actaea dahurica (Hai et al. 2022), and Ligustrum japonicum (Wei et al. 2022) in China. This is the first report of A. alternata causing leaf black spot on rosemary in China.

甜瓜是我国重要的蔬菜作物,海南是我国重要的甜瓜产区,而病毒病的发生严重影响了海南甜瓜的生产.综述了近几年在海南发生的较为严重的几种病毒病害,并在此基础上分析了病毒病的流行原因,并提出了相应的防控措施.及时了解病毒病害发生的种类、有效使用化学药剂防控田间传播介体、培育抗病品种是当前防控病毒病害的重要举措,旨在为甜瓜的安全生产、病毒病害防治提供科学依据和参考.

为探明海南地区冬春作甜瓜病毒病的主要病原种类和危害情况,2022—2023年冬春季从海南甜瓜主产区采集34份疑似感染病毒病的甜瓜样品,利用高通量测序(high-throughput sequencing)和聚合酶链式反应(polymerase chain reaction,PCR)技术对甜瓜病毒病的病原种类进行鉴定和分析.结果表明:甜瓜黄斑病毒(melon yellow spot virus,MYSV)、西瓜银斑驳病毒(watermelon silver mottle virus,WSMoV)和中国南瓜曲叶病毒(squash leaf curl China virus,SLCCNV)引起的甜瓜病毒病在海南多个产区普遍发生.其中,MYSV检出率最高,为73.53％;其次为WSMoV,检出率61.76％;且呈现MYSV和WSMoV复合侵染现象,复合侵染率为47.06％.

Fusarium wilt of banana is a devastating disease caused by Fusarium oxysporum f. sp. cubense (Foc). It has restricted the development of the banana industry worldwide and is particularly serious in China because of the large planting areas and special planting patterns. However, there is no rapid and accurate approach to detect the Foc strains that specifically occur in China because of the rich genetic diversity observed in this pathosystem. In this study, we evaluated the performance of 10 previously published PCR primer pairs on 103 representative Foc strains in China and neighboring countries and screened out a set of primers (Foc-specific primer pair SIX9-Foc-F/R, Foc R1-specific primer pair SIX6b-210-F/R, Foc R4-specific primer pair Foc-1/2, and Foc TR4-specific primer pair W2987F/R) suitable for the detection of Foc strains in China and the surrounding Southeast Asian countries. Moreover, we developed a molecular detection system to accurately identify the different physiological races of Foc. The findings of this study provide technical support for preventing and controlling the spread of Fusarium wilt of banana in the field in China.