Shenghua Wei

Ardisia crenata Sims, belonging to the family Myrsinaceae, has high medicinal and economic value. In July 2021, root rot disease was observed in plantations located in Ziyun (106°08'45″ E, 25°75'15″ N) and Xiuwen (106°46'53″ E, 26°54'01″ N) Counties, Guizhou Province, China, with an incidence rate of approximately 30% at fruit drop stage. The disease manifested as root softening, blackening, and phloem rot, while the aerial parts showed progressive yellowing, curling, and withering of leaves. Ultimately, the plant died. Nine symptomatic root segments were collected from fifteen infected plants, surface sterilized in 5% NaClO and 75% ethanol for 1 minute each, washed three times with sterile water, and incubated on potato dextrose agar (containing 10μg/mL of chloramphenicol) for five days at 28°C. The hyphal tip technique (Senanayake et al. 2020) was used to obtain pure cultures. Sixteen strains exhibiting similar morphological characteristics were isolated from the infected tissues. The colonies of these isolated cultures appeared white and turned light grey after 4 days. Five isolates were selected and grown on 2% water agar for 7 days for morphological characterization. Conidia were single-celled, hyaline, spindle-shaped to oval, and measured 12.04 to 19.47 µm long and 4.63 to 7.41 µm wide (n = 50). Based on these morphological features, the isolates were suspected to be Neofusicoccum parvum (Pavlic et al. 2009). For molecular identification, strain-7 was randomly selected as a representative individual to extract DNA. The internal transcribed spacer (ITS) region was amplified using primers ITS1/ITS4 (White et al. 1990). Additionally, the translation elongation factor 1-alpha (TEF1) gene was amplified using primers EF1-728F/EF1-986R (Rehner et al. 2005), and the β-tubulin (TUB2) gene was amplified using primers BT2a/BT2b (Glass et al. 1995). The sequences of strain-7 (accession number OR789487[ITS]; PV209690[TEF1]; PV209691[TUB2]) were deposited in GenBank, demonstrating a sequence homology of 99% to 100% (569/570, 265/265, 448/448) with N. parvum YBF5-1 (accession numbers PQ222752, PQ227810 and PQ227811). Based on morphological features and Neighbor-Joining method analyses of combined ITS, TEF1, and TUB2 gene sequences, strain-7 was identified as N. parvum. A pathogenicity test was performed using strain-7 by inoculating roots of 12-month-old A. crenata seedlings. The test was repeated seven times. Healthy seedlings were cut vertically with a sterile knife at 2 cm from the edge of one stem to create root damage. Spore suspensions (150 ml, 1 × 106 conidia/ml) of N. parvum were applied on the cut side, while the control group was watered with the same volume of sterile water. All plants were kept in the same glasshouse under natural conditions. After 7 days, some yellowish spots appeared on the leaf surface of some plants, with the edges turning dry and curled. The branches and leaves turned completely yellow, and the roots rotted extensively on the 14th day, whereas the control group remained asymptomatic. To satisfy Koch's postulates, N. parvum was reisolated from the inoculated plants but not from the control. N. parvum has been reported to cause leaf spot disease on Macadamia integrifolia and Vitis heyneanan in China (Li et al. 2023; Wu et al. 2015). This is the first report of N. parvum causing root rot in A. crenata in China. These findings provide a basis for the early detection of A. crenata root rot and the formulation of targeted control measures.

Epimedium sagittatum (Berberidaceae), a medicinal plant with a documented history of over 2,000 years in China for the treatment of kidney yang deficiency and rheumatism, has been cultivated in Tongren City, Guizhou Province (108.79°E, 27.54°N). During the period of 2023-2024, rhizome rot disease was detected in 1- to 2-year-old E. sagittatum plants within local plantations. The initial symptoms manifested as darkening of the stem base and scales, eventually leading to complete wilting and rhizome rot. A total of twelve symptomatic plants were collected for pathogen isolation. The diseased rhizome tissues underwent surface sterilization using 5% sodium hypochlorite for 30 seconds, followed by 75% ethanol for 30 seconds, and were then rinsed three times with sterile water before being cultured on potato dextrose agar (PDA) at 25°C. After 3-5 days, hyphal tips were transferred to fresh PDA for purification. This process resulted in the isolation of two distinct Fusarium species. Pathogenicity tests on 10-month-old cultivated E. sagittatum confirmed that the isolate JFB-2 induced rhizome rot symptoms identical to those observed in the field. Following a five-day incubation period at 25°C under dark conditions, the isolate JFB-2 developed dense, cottony mycelia, with colonies exhibiting a coloration ranging from white to pale purple or light pink. The macroconidia were hyaline and falcate (sickle-shaped), possessing 3-5 septa, and measured 36.5 (29.5-42.5) × 3.6 (3-4.5) μm (n=50). For molecular identification, the internal transcribed spacer (ITS) region, translation elongation factor 1-alpha (TEF), and RNA polymerase II subunit B2 (RPB2) genes of JFB-2 were amplified using the primers ITS1/ITS4, EF1-R6/F6, and RPB2-5F/7cR, as described by Zhang et al. (2024). The sequences obtained for ITS (GenBank accession PV546364), TEF (PV550274), and RPB2 (PV550273) exhibited greater than 98.9% identity and at least 98% query coverage when compared to F. oxysporum reference sequences (MN856306.1 for ITS, PP737057.1 for TEF, and ON316723.1 for RPB2). Phylogenetic analysis, employing Bayesian methods on concatenated ITS and TEF sequences (Zhang et al., 2020), positioned JFB-2 within the F. oxysporum clade with a posterior probability of 100%. The pathogenicity of the JFB-2 strain was confirmed in healthy E. sagittatum plants. The roots of three replicate plants were wounded and inoculated with 150 mL of a conidial suspension at a concentration of 1×10⁶ conidia/mL. These plants were then maintained in a humid chamber at a temperature of 25±1°C and 85% relative humidity. Control plants were treated with sterile double-distilled water. After a period of 15 days, the inoculated plants exhibited symptoms consistent with those observed in the field, and F. oxysporum was successfully reisolated from the diseased tissues. The internal transcribed spacer (ITS) sequence of the reisolated pathogen (accession number PQ656171.1) was identical to that of the JFB-2 strain, thereby satisfying Koch's postulates. F. oxysporum has previously been identified as a pathogen responsible for rhizome rot in various plants, including onions (Wesoly et al., 2024), Polygonatum cyrtonema (An et al., 2024), and Polygonatum odoratum (Xu et al., 2022). To the best of our knowledge, this study represents the first documented instance of F. oxysporum infecting E. sagittatum in China, offering valuable insights for the management of diseases in medicinal plant cultivation.

Ardisia crispa(Myrsinaceae) is an ethnomedicine with horticultural and important medicinal values. Its morphology is complex, and its identification is difficult. We analyse the chloroplast genome characteristics and phylogenetic position of A. crispa to provide basic research data for the identification of A. crispa species and resource conservation. This study assemble and annotate the chloroplast genome of A. crispa and to compare it with the chloroplast genome within Ardisia. The A. crispa chloroplast genome is 156,785 bp in length, with a typical quadripartite structure containing 131 genes, including 86 protein-coding genes, 37 tRNA genes, and 8 rRNA genes; a total of 59 SSRs sites were identified, and the codon preference of this chloroplast genome is greater in A/U than in G/C, and leucine is the amino acid with the highest frequency of use. The chloroplast genomes of the nine Ardisia species are conserved in gene content and number, with more stable boundaries and less variation. In the phylogenetic tree, A. crispa is clustered on a branch with A. crispa var dielsii, and is closely related to A. mamillata and A. pedalis. In this study, we constructed and analyzed the chloroplast genome structure of A. crispa, and conducted phylogenetic analysis using the whole chloroplast genome sequence data of Ardisia plants, which is of great significance in understanding the genetic basis of A. crispa and adaptive evolution in Ardisia plants, and this will lay the foundation for the future research on A. crispa resource conservation and species identification.

The genus of Epimedium belongs to Berberidaceae family, which is famous for their medicinal and ornamental value. In recent years, Epimedium has attracted increasing attention due to their medicinal and nutritive value. In this research, we reported the complete chloroplast (cp) genome of Epimedium enshiense. The complete chloroplast of this species is 157,076 bp in length, including a pair of invert repeat regions (IRS) (25,833 bp) that is divided by a large single copy area (LSC) (88,340 bp) and a small single copy area (SSC) (17,070 bp). The circular chloroplast genome of E. enshiense contains 112 unique genes, composing of 78 protein-coding genes, 30 tRNA, and four rRNA genes. Phylogenetic analysis indicates that E. enshiense has a closer relationship with E. dolichostmon.

Epimedium simplicifolium Ying is a rare and endangered species endemic to Guizhou, China. In this study, the complete chloroplast (cp) genome of E. simplicifolium was sequenced. The total cp genomes size was 158,745 bp in length, with 38.7% of GC content, including four distinct regions: the large single-copy region (LSC, 88,413 bp), small single-copy region (SSC, 15,386 bp), and a pair of inverted repeat regions (IRs, each for 27,473 bp). The whole cp genome of E. simplicifolium encoded 131 unique genes, including 86 protein-coding genes, 37 tRNA genes, and 8 rRNA genes. Phylogenetic analysis with the previously reported cp genomes showed that all the samples of Epimedium are grouped into one group, which can be easily separated from the other genera of the Berberidaceae, and the E. simplicifolium is a separate species from the rest of the Epimedium. This plastid genome is the first report for the E. simplicifolium and will be useful data for developing markers for further studies on resolving the relationship within the Berberidaceae.

Epimedium shuichengense S. Z. He is a rare and endangered species endemic to Guizhou, China. In this study, the complete chloroplast (cp) genome of E. shuichengense was sequenced. The total cp genomes size was 157,046 bp in length, with 38.8% of GC content, including four distinct regions: the large single-copy region (LSC, 88,394 bp), small single-copy region (SSC, 17,048 bp), and a pair of inverted repeat regions (IRs, each for 20,820 bp). The whole cp genome of E. shuichengense encoded 130 unique genes, including 85 protein-coding genes, 37 tRNA genes, and 8 rRNA genes. Phylogenetic analysis with the previously reported cp genomes showed that all the samples of Epimedium are grouped into one group, which can be easily separated from out groups, and E. shuichengense with E. borealiguizhouense into a supported branch.

Epimedium dewuense S. Z. He is a rare and endangered endemic to China. The complete chloroplast (cp) genome of E. dewuense was reported in this study. The total cp genomes size was 157,056 bp in length, with 38.8% of GC content, and a typical quadripartite structure, the large single-copy region (LSC, 88,346 bp), small single-copy region (SSC, 26,768 bp) that separate by a pair of inverted repeat regions (IRs, each for 20,820 bp). The whole cp genome of E. dewuense contained 130 unique genes, including 37 tRNA, 7 rRNA, and 86 protein-coding genes. This plastid genome is the first report for the E. dewuense and will be useful data for developing markers for further studies on resolving the relationship within the Berberidaceae.