Study on the inhibitory activity and mechanism of Mentha haplocalyx essential oil nanoemulsion against Fusarium oxysporum growth.

Mentha haplocalyx essential oil (MEO) has demonstrated inhibitory effects on Fusarium oxysporum. Despite its environmentally friendly properties as a natural product, the limited water solubility of MEO restricts its practical application in the field. The use of nanoemulsion can improve bioavailability and provide an eco-friendly approach to prevent and control Panax notoginseng root rot. In this study, Tween 80 and anhydrous ethanol (at a mass ratio of 3) were selected as carriers, and the ultrasonic method was utilized to produce a nanoemulsion of MEO (MNEO) with an average particle size of 26.07 nm. Compared to MTEO (MEO dissolved in an aqueous solution of 2% DMSO and 0.1% Tween 80), MNEO exhibited superior inhibition against F. oxysporum in terms of spore germination and hyphal growth. Transcriptomics and metabolomics results revealed that after MNEO treatment, the expression levels of certain genes related to glycolysis/gluconeogenesis, starch and sucrose metabolism were significantly suppressed along with the accumulation of metabolites, leading to energy metabolism disorder and growth stagnation in F. oxysporum. In contrast, the inhibitory effect from MTEO treatment was less pronounced. Furthermore, MNEO also demonstrated inhibition on meiosis, ribosome function, and ribosome biogenesis in F. oxysporum growth process. These findings suggest that MNEO possesses enhanced stability and antifungal activity, which effectively hinders F. oxysporum through inducing energy metabolism disorder, meiotic stagnation, as well as ribosome dysfunction, thus indicating its potential for development as a green pesticide for prevention and control P. notoginseng root rot caused by F.oxyosporum.

First report of sweet cherry root rot caused by Fusarium solani in China.

Sweet cherry (Prunus avium L.) has become an economically important fruit in China. And its cultivation area has significantly expanded over the last three decades (Wang et al. 2020; Zhao et al. 2023). In July 2023, wilting of cherry trees was observed in a cherry plantation in Wenchuan County (31°51'N, 103°56'E, altitude: 1,510 m) in Sichuan Province and approximately 27% of the trees showed symptoms of root rot including soft roots, dark brown to black lesions, yellowing and wilted leaves, and a distinct yellow-brown core discoloration of the inner root core when cut in cross-section. To isolate the causal pathogens, six infected sweet cherry plants with rootstock 'Daqingye' from Cerasus pseudocerasus were randomly selected from the orchard and then the intertwined diseased and healthy roots (5mm× 5mm × 2mm) were washed with sterile water to remove surface soil. The root samples were surface sterilized with 75% ethanol for 30 seconds and NaClO for 30 seconds and washed three times with distilled water. The disinfected tissues were placed on potato dextrose agar (PDA) and incubated at 27°C in darkness for 5 days (Zhao et al. 2024). A total of nine fungal isolates with similar morphological characteristics were obtained. The colony obtained through single-spore purification displays a red reverse side and a concentric ring pattern on the front, with a sparse surface. Macroconidia were relatively slender with a curve, like sickle shape, 0 to 3 septate measuring (25.8 to 46.1) µm× (4.2 to 7.5) µm, respectively (n=20). The morphological characteristics were consistent with the description of Fusarium spp. (Li et al. 2021). Among these isolates, only HB5 was selected for additional molecular identification. Three target genes, including the internal transcribed spacer (ITS), partial translation elongation factor 1-alpha (TEF), and RNA polymerase second largest subunit (RPB2) were amplified using the primers ITS1/ITS4, TEF1-728/FTEF1-re, and fRPB2-5F/fRPB2-7r, respectively (Groenewald et al. 2013; Carbone and Kohn 1999; Reeb et al. 2004). Sequences of HB5 was deposited in GenBank (ITS, PP388208; TEF, PP580036; RPB2, PP580035). A BLAST search revealed high similarity to those of F. solani sequences with 99%, 100% and 100% respectively (MN013858.1, JF740846.1, OR371902.1), and a multilocus phylogenetic tree was generated to represent the molecular identification results. Pathogenicity studies were conducted on the rootstocks from 'Daqingye' of Cerasus pseudocerasus in 1 liter plastic flowerpots. The seedlings were incubated in a constant temperature incubator at 25°C with a humidity level of 65% for two weeks. Following the growth of green leaves, 200ml (1x106 spores/ml) of spore suspensions were poured into pots. After 4 weeks of inoculation, the same symptoms of the inoculated plants were observed consistent with those shown in the field , while control plants were inoculated with distill water with asymptomatic. The inoculated pathogen was confirmed both morphologically and molecularly as described earlier, thereby fulfilling Koch's postulates. It has been reported that Fusarium solani has been reported to cause root rot in various plants in China, including Actinidia sppt, Zanthoxylum bungeanum, Fragaria×ananassa Duch (Song et al.2022; Li et al. 2023; Zhao et al. 2024). To our knowledge, this is the first report of Fusarium solani causing root rot in sweet cherry (Prunus avium). We here also report the severity and outbreak of this disease, which has been found in other regions in recent years and may become prevalent. Further research on disease management strategies is urgently needed to protect sweet cherry production.

White root rot of Bletilla striata: the pathogen, biological characterization, and fungicide screening.

Bletilla striata is an endangered traditional medicinal herb in China. In May 2020, the emergence of white root rot severely impacted the quality and yield of B. striata, affecting about 5% of the plants at plant nurseries of the Chengdu Academy of Agricultural and Forestry Sciences. Through a series of experiments and evaluations, the pathogen was identified as Fusarium solani. This is the first report of B. striata white root rot caused by F. solani in Sichuan, China. To better understand this disease and provide data support for its control, a combination of morphological, molecular characterisation and pathogenicity determination was used in this study for assessment. Meanwhile, the effects of different carbon and nitrogen sources, culture medium, temperature, photoperiod and pH on mycelial growth and spore production of F. solani were investigated. In addition, effective fungicides were screened and the concentration ratios of fungicides were optimized using response surface methodology (RSM). The experimental results showed that sucrose was the optimum carbon source for the pathogen, and the optimum temperature and pH were 25°C and pH 7, respectively, while light did no significant effect. Effective fungicides were screened, among which difenoconazole showed the strongest inhibition with EC50 of 142.773 µg/mL. The optimum fungicide concentration scheme (difenoconazole, pyraclostrobin, and thiophanate-methyl at 395.42, 781.03, and 561.11 µg/mL, respectively) was obtained using response surface methodology (RSM) to improve the inhibition rate of 92.24 ± 0.34%. This study provides basic data for the pathogen characterization of B. striata white root rot and its potential fungicides in Sichuan, China. In addition, the optimal fungicide concentration ratios were obtained through response surface methodology (RSM) optimization, which significantly enhanced the fungicidal effect and provided a scientific basis for the future control of B. striata white root rot.

Diversity and Pathogenicity of Fusarium Root Rot Fungi from Canola (Brassica napus) in Alberta, Canada.

Root rot disease poses a significant threat to canola (Brassica napus), underscoring the need for a comprehensive understanding of its causal agents for more effective disease mitigation. The composition and diversity of fungal pathogens associated with root rot of canola in Alberta, Canada, were evaluated from plant tissue samples collected in 2021 and 2022. The study revealed Fusarium spp. as the predominant pathogens found in almost all surveyed fields. Fusarium avenaceum, F. redolens, and F. solani were among the most frequently recovered species. Greenhouse trials confirmed their pathogenicity, with F. avenaceum and F. sporotrichioides found to be particularly aggressive. Additionally, F. sporotrichioides and F. commune were identified for the first time as canola root rot pathogens. Inoculation with isolates of most species resulted in significant reductions in seedling emergence, plant height, and shoot and root dry weights. Analysis of translation elongation factor 1-α (TEF-1α) and internal transcribed spacer (ITS) sequences confirmed the identity of the Fusarium spp., while concatenating the ITS and TEF-1α sequences enabled improved species differentiation. Geographic and year effects did not influence fungal diversity or aggressiveness, as determined by principal component analysis. This study emphasized the high diversity and impact of Fusarium spp. in causing canola root rot.

First Report of Root Rot on Rhizome of Polygonatum kingianum Caused by Aspergillus awamori.

Polygonatum kingianum Coll. et Hemsl (Huangjing), which belongs to the family Asparagaceae, is a perennial traditional Chinese herb with homologous medicinal and edible value (Liu et al., 2021). Huangjing is known to promote blood circulation; it has anti-inflammatory properties, increases immunity, and provides hypoglycemic treatments (Ma et al., 2019). Root rot-infected P. kingianum exhibited withering yellow leaves and stems, rhizome rot, slowed growth, and plant death. In recent years, with an average incidence of up to 45%, the spread of HJ root rot (rhizome and stem bases) has resulted in a significant reduction in the quality and up to 63% reduction in the yields of Sichuan Junlian (104.5°E, 28.2°N) and Guizhou Zhunyi (107.0°E, 27.7°N). After collecting the diseased samples, we used the tissue isolation method to isolate the pathogenic fungi (Wu et al., 2020). Four fungal isolates associated with root rot were obtained: HJ-G2 (two strains), HJ-G3 (one strain), HJ-G4 (one strain), and HJ-G6 (two strains), of which HJ-G2 and HJ-G6 were the dominant species. To determine pathogenicity of each strain, tests were conducted by wounding rhizomes wth an inoculation needle and the pathogen strain was inoculated onto the wound and symptoms observed. The results reveal that HJ-G6 exhibited the strongest pathogenicity against P. kingianum (Figure 1). The HJ-G6 colonies were black, grew rapidly, and produced a large number of spores (Figure 1). A spherical apical sac (conidial head) is formed at the top with two palisades of cells, metulae and phialides, which are shaped radially and produce a large number of spores with 2-5 um in diameter (Figure 2). Morphological observations revealed that the isolate was consistent with Aspergillus awamori (Naher et al., 2021). To further confirm the fungal species, the ribosomal internal transcribed spacer (ITS), ß-tubulin (TUB), and elongation factor 1-alpha (EF-1a) gene regions were amplified with ITS1/ITS4, Bt2a/ Bt2b, and EF1/EF2. Primer and PCR amplification were performed as previously described (Paul et al., 2017). The sequences were compared with those obtained from GenBank. The ITS sequences (GenBank accession number OR682143) of the isolates (HJ-G6) were 100% identical to those of the strain PANCOM10 (GenBank accession number MT007535.1) of Aspergillus awamori. The EF-1a sequences (GenBank accession OR752352) of the isolates (HJ-G6) were 98% identical with strain ITEM 4777 (GenBank accession FN665402.1) of Aspergillus awamori. The TUB sequences (GenBank accession number OR752351) of the isolate (HJ-G6) were 100% identical with strain AF158 (GenBank accession MH781275.1) of Aspergillus awamori. Three maximum likelihood trees were constructed using MEGA v5.0 (Kumar et al., 2018) based on the sequences (ITS, TUB, and EF-1a) of the HJ-G6 strain and that of Aspergillus spp. previously deposited in GenBank (Paul et al., 2017). Phylogenetic analysis showed that HJ-G6 belonged to the Aspergillus awamori clade (Figure 3). Combined with morphological analysis and DNA sequencing, HJ-G6 was identified as Aspergillus awamori. To verify pathogenicity, P. kingianum roots were inoculated with the colonized agar discs of the isolates. P. kingianum plants inoculated with uncolonized agar discs were used as controls. After inoculation, P. kingianum roots were moved to the inoculation chamber under high humidity at 28 °C for 1 d and then transferred to a greenhouse. Previous studies have reported that Fusarium sp. are root rot pathogens in the rhizomes of medicinal plants (Pang et al., 2022; Song et al., 2023). In this study, HJ-G2, HJ-G3, and HJ-G4 were used as the positive controls. Typical symptoms of root rot appeared 3 days after inoculation and were similar to those observed in the field, whereas the control plants remained symptomless. According to the results of the inoculation experiment, the pathogenicity of Aspergillus awamori to P. kingianum root rot was significantly stronger than that of Fusarium (Figure 1). The pathogen was isolated from the rotting root of P. kingianum and the ITS region was sequenced again. Alignment analysis of the ITS sequences revealed that the causal agents were consistent with those of the original isolates. These studies fulfill Koch's postulates. As far as we know, this is the first report of Aspergillus awamori causing root rot in P. kingianum.

Genome sequence of Fusarium oxysporum strain ByF01, the causal agent of root rot of Knoxia roxburghii in China.

OBJECTIVES: Knoxia roxburghii is a member of the madder (Rubiaceae) family. This plant is cultivated in different areas of China and recognized for its medicinal properties, which leads to its use in traditional Chinese medicine. The incidence of root rot was 10-15%. In June 2023, the causal agent of root rot on K. roxburghii was identified as Fusarium oxysporum. To the best of our knowledge, this is the first report of the complete genome of F. oxysporum strain ByF01 that is the causal agent of root rot of K. roxburghii in China. The results will provide effective resources for pathogenesis on K. roxburghii and the prevention and control of root rot on this host in the future. DATA DESCRIPTION: To understand the molecular mechanisms used by F. oxysporum to cause root rot on K. roxburghii, strain ByF01 was isolated from diseased roots and identified by morphological and molecular methods. The complete genome of strain ByF01 was then sequenced using a combination of the PacBio Sequel IIe and Illumina sequencing platforms. We obtained 54,431,725 bp of nucleotides, 47.46% GC content, and 16,705 coding sequences.

First report of Cigar Tobacco Root Rot Caused by Setophoma terrestris in China.

Cigar tobacco (Nicotiana tabacum L.) is widely planted in Yunnan, which is becoming an important economic crop in China. In March 2023, root rot of cigar tobacco (cv. Yunxue 38) was observed in Baoshan (98°51'E, 24°58'N), and in July 2022 root rot of tobacco (cv. Yunyan 87) was observed in Dali (99°54'E, 26°30'N), Yunnan Province, China. The average disease incidences surveyed in the fields reached 10%. At the early stage, the bottom leaves showed wilting and turned yellow, and the roots became brown. Following the disease development, the color of roots turned to dark brown and ultimately necrosis. To isolate the causal agent, small pieces (5×5 mm) of diseased root from 6 symptomatic plant samples (three samples of cv. Yunxue 38 and three samples of cv. Yunyan 87) were cut. Pieces were surface-sterilized by dipping in 75% ethanol for 30 s, rinsed three times with sterile distilled water, then transferred to potato dextrose agar (PDA) medium and incubated at 28°C in the dark. Six fungal isolates cultured for 14 days were obtained. They were morphologically similar, so a representative isolate was selected for the following experiment. The colonies grew slowly on PDA, and their color were light pink initially, then changed to amaranth. Hyphae were hyaline and septate. Microconidia were hardly produced on PDA plates. After 14 days of culture on V8 juice agar, the colonies showed white aerial mycelia, and ellipsoidal and transparent conidia were observed, which measured 6.5 to 8.3 × 3.4 to 5.0 µm (n=20). Also, the pycnidia were measured 150 to 220 µm, that were subglobose in dark brown with brown setae. These morphological characteristics of 22DL91 were identical to S. terrestris (Boerema et al. 2004). For molecular identification, DNA was extracted and the PCR products of ITS region and polymerase II second largest subunit (RPB2), amplified with the primers ITS1/ITS4 and RPB2-5F/RPB2-7cR, were sequenced. By BLASTn analysis, the obtained ITS sequences showed 100% homology and the RPB2 sequences showed 95% homology with S. terrestris strains in GenBank (accession ON006851 and OM417590). The sequences were deposited in NCBI with accession numbers OR539491 (ITS) and OR554276 (RPB2), respectively. Based on the morphology and phylogenetic analysis, the isolate was 22DL91 identified as S. terrestris. Pathogenicity was evaluated on 50-day-old cigar tobacco seedlings (cv. Yunxue 38) and tobacco seedlings (cv. Yunyan 87). Ten plants were inoculated with 20 mL of conidial suspension of 105 conidia/mL poured onto the roots and ten control seedlings dipped in sterile water as controls (Luo et al. 2023). After 14 days, all inoculated seedlings showed the symptoms with leaves yellowing and root rot, whereas the control seedlings had no symptoms. Moreover, the fungus S. terrestris was reisolated from the infected roots, fulfilling Koch's postulates. This fungus was previously known to cause pink root on garlic in China (Zhang et al. 2019). To our knowledge, this is the first report of S. terrestris causing root rot of Nicotiana tabacum in China. Therefore, this finding will provide valuable information for prevention and management of root rot on tobacco.

First Report of Root Rot of Cathaya argyrophylla caused by Fusarium oxysporum in China.

Cathaya argyrophylla [Chun & Kuang.] is an ancient relict plant and its embryonic development is similar to that of Pinus species. This has important scientific value for studying the phylogeny of Pinaceae (Wu et al. 2023). In July 2022, root rot was detected in the seedling cultivation base of C. argyrophylla in Daozhen County, Guizhou Province, China (28.89 °N, 107.6 °E). The incidence of the disease was 30% (n = 100); the susceptible plants wilted, leaves withered, and roots showed brown-to-black lesions and rot. Ten root tissues were randomly collected from the edges of the lesions of six symptomatic susceptible plants. The tissues were sterilized with 75% alcohol for 30 seconds, followed by 2-minute immersion in 3% sodium hypochlorite. After washing with sterile water, the tissues were incubated on potato dextrose agar (PDA; BoWei, Shanghai) at 28 ℃ for five days. Four single-spore cultures were obtained using a single-spore isolation method (Gong et al., 2010). Single-spore cultures grew rapidly on PDA. After five days of incubation, the colonies were white and pink, indicating a large amount of aerial mycelia. Microconidia were ovate or ellipsoid, measuring 5.0-10.0 × 1.5-3.0 µm (n = 50); Macroconidia were falcate, slightly curved or straight, measuring 19.5-28.5 × 2.0-6.0 µm (n = 50). Based on morphological features, the pathogen was considered to be Fusarium spp. (Leslie and Summerell 2006). Three representative strains, GF5, GF6, and GF7, were selected for molecular identification, and genomic DNA was extracted to confirm morphological diagnosis. The internal transcribed spacer (ITS) (White et al. 1990) was amplified using primers ITS1/ITS4, and the ß-tubulin gene (Varga et al. 2011) was amplified using primers Bt2a/Bt2b. The ITS and ß-tubulin sequences were aligned with GenBank, and amplification of the genes from the three isolates was consistent. The ITS (OP482273) and ß-tubulin (OR825353) sequences of GF5 were stored in GenBank, and their homology with Fusarium oxysporum HC131(accession numbers MW600442 and MW670451) was 99 to 100%. Maximum likelihood analysis using MEGA 11.0 showed that isolate GF5 belongs to F. oxysporum. The reconstructed phylogenetic tree confirmed the phylogenetic position of the isolate GF5. The pathogenicity test was carried out using GF5 and GF6 isolates. The taproots of ten 3-year-old C. argyrophylla plants were washed, and then the roots were immersed in a 2 × 106/mL conidial suspension for one hour. Ten plants with sterile water were used as controls. After planting in pots (30 × 25 cm) with sterilized forest soil, the plants were cultured in a greenhouse (25 ℃ and 12-hour photoperiod). Thirty days after inoculation, all plants inoculated with the isolated pathogen showed wilting symptoms, and the roots showed typical root rot symptoms, whereas the control group showed no symptoms. The pathogens re-isolated from all inoculated plants were morphologically identical and had ITS sequences identical to F. oxysporum, validating Koch's hypothesis. The pathogenicity test was repeated twice and similar results were obtained. Although this fungus has been previously reported to cause root diseases in hosts, such as Musa nana Lour. and Pinus massoniana Lamb. (He et al. 2010; Luo et al. 2020), to our knowledge, this is the first report of F. oxysporum causing root rot in C. argyrophylla. These findings provide a basis for the development of management strategies for C. argyrophylla infection.

Characterization and Pathogenicity of Soil-Borne Pathogens in Gloriosa superba: Effects of Single and Multiple-Pathogen Co-Infection on Disease Responses.

Glory lily (Gloriosa superba), an ornamental climbing plant, contains the bioactive compound colchicine, attracting attention from the pharmaceutical industry. However, soil-borne pathogens have emerged as a serious threat to the cultivation of glory lily, leading to substantial economic losses in the southern parts of India. Among these, the three major pathogens are Macrophomina phaseolina, Fusarium oxysporum, and Agroathelia rolfsii, causing dry root rot (also referred to as charcoal rot), wilt, and stem rot, respectively. Here, we characterised these pathogens using morphological characteristics and phylogenetic analysis of DNA sequences related to the internal transcribed spacer (ITS) of ribosomal DNA, calmodulin (CAL) and translation elongation factor (TEF)-1α. Further, in the pathogenicity tests, the inoculation of M. phaseolina alone resulted in lesions measuring 7.54±0.01 mm on tubers and 90% seedling mortality. This severity was comparable to the simultaneous inoculation of all three pathogens, indicating the prominence of dry root rot among soil-borne diseases. This study marks the first detailed investigation of soil-borne pathogens combined infection in G. superba, contributing to the understanding of fungal disease complexity in medicinal plants.

Whole Genome Sequencing Reveals Novel Insights about the Biocontrol Potential of Burkholderia ambifaria CF3 on Atractylodes lancea.

Root rot caused by Fusarium spp. is the most destructive disease on Atractylodes lancea, one of the large bulks and most common traditional herbal plants in China. In this study, we isolated a bacterial strain, CF3, from the rhizosphere soil of A. lancea and determined its inhibitory effects on F. oxysporum in both in vitro and in vivo conditions. To deeply explore the biocontrol potential of CF3, we sequenced the whole genome and investigated the key pathways for the biosynthesis of many antibiotic compounds. The results revealed that CF3 is a member of Burkholderia ambifaria, harboring two chromosomes and one plasmid as other strains in this species. Five antibiotic compounds were found that could be synthesized due to the existence of the bio-synthesis pathways in the genome. Furthermore, the synthesis of antibiotic compounds should be confirmed by in vitro experiments and novel compounds should be purified and characterized in further studies.

Virulent Fusarium isolates with diverse morphologies show similar invasion and colonization strategies in alfalfa.

Root rot is a major disease that causes decline of alfalfa production, and Fusarium is a major pathogen associated with root rot. In this study, 13 Fusarium isolates were obtained from alfalfa with root rot in Gansu Province, the major alfalfa production region in China. The isolates were characterized by molecular genotyping (ITS, TEF 1-α and RPB2 fragments) and identified as six species, which included the F. acuminatum, F. incarnatum, F. oxysporum, F. proliferatum, F. redolens, and F. solani. We found that their morphology varied significantly at both the macro- and micro-levels, even for those from the same species. We developed a low cost and fast pathogenicity test and revealed that all isolates were pathogenic to alfalfa with typical root rot symptoms such as leaf yellowing and brown lesions on the root and stem. However, the virulence of the isolates differed. We also found that the conidia of all isolates germinated as early as 24 hours post inoculation (hpi), while hyphae colonized the root extensively and invaded the xylem vessel by 48 hpi. Together our results reveal that different virulent Fusarium isolates use a similar invasion strategy in alfalfa. This natural plant-fungus pathosystem is intriguing and warrants further examination, particularly with regard to efforts aimed at mitigating the impact of multiple similar vascular pathogens on infected alfalfa plants.

First Report of Strawberry Root Rot Caused by Fusarium falciforme in China.

Root rot caused by Fusarium spp. is a destructive disease affecting agricultural regions worldwide. Strawberries (Fragaria × ananassa Duch.) are an economically important crop in China. In March 2023, root rot was observed in strawberries grown in Jinan, Shandong Province, China. Symptoms included leaf wilt, necrotic roots, and plant death (Figure 1). Four strawberry samples (two symptomatic and two asymptomatic) were collected from ~2-acre fields where the disease incidence rate ranged from 2 to 3%. Tissue pieces (5 mm × 5 mm × 5 mm) from two healthy and two diseased strawberry root tissues were surface-disinfected with 75% ethanol for 3 min, treated with 10% sodium hypochlorite for 5 min, and washed three times with sterile water. These pieces were cultured for 5 days at 28°C on potato dextrose agar (PDA) containing 200 mg/L timentin. Typical Fusarium spp. like growth was observed on plates with the two symptomatic samples. Two representative fungal isolates (CM1 and CM2) with similar morphological characteristics were purified using the single-spore method (Figure 1). CM1 showed an average growth rate of 5 mm/d in PDA and comprised of several white-to-cream aerial mycelia after 5 d. After cultivation in carnation leaf agar medium for 7 d, falciform macroconidia, with blunt apical cells and slightly hooked basal cells comprising 3 to 4 septa of varying sizes (20 to 39)×(3.6 to 6.7 µm) were observed (n=50) (Figure 1). The chlamydospores were spherical, terminal or intercalary, solitary or chain-forming, and 3.1-10.5 µm in diameter (Figure 1). The microconidia on PDA were (5.8 to 13.6)× (2.5 to 3.3) µm in size (n=50). These morphological characteristics are consistent with previous descriptions of the Fusarium solani species complex (FSSC). DNA was extracted using the CTAB method (Stenglein and Balatti 2006). The internal transcribed spacer (ITS), translation elongation factor 1-α gene (tef1), RNA polymerase II largest subunit (rpb1), and RNA polymerase II second largest subunit (rpb2) were amplified and sequenced using specific primers (O'Donnell et al. 2010). The ITS (OR526528, OR526529), tef1 (OR536947, OR536948), rpb1 (OR536949, OR536950), and rpb2 (OR536951, OR536952) sequences of the CM1 and CM2 isolates were uploaded to the NCBI database. BLASTn analysis revealed that the ITS, tef1, rpb1, and rpb2 sequences were 99.1-100% identical to those of the Fusarium falciforme reference strains NRRL 54989 and NRRL 54978. A phylogenetic tree based on the ITS, tef1, rpb1, and rpb2 sequences was generated using MEGA v.11 via the maximum-likelihood method (Tamura et al. 2021). CM1 clustered with the Fusarium falciforme reference strains NRRL 54989 and NRRL 54978 and belonged to the FSSC based on its morphological and molecular characteristics (Figure 2). To test for pathogenicity, the roots of nine 3-month-old healthy strawberry (cv. Akihime) plants were exposed to conidial suspensions (1×108 spores/mL) of the CM1 isolate. Another nine root samples were treated with sterile water and used as controls. All strawberry plants were maintained in a growth chamber under a 12/12 h light/dark cycle at 28°C and 90% relative humidity and the experiment was repeated three times. After one month, the inoculated plants had withered and died, and the pith became dark red (similar to field plants) (Figure 1). The fungi isolated from the experimental plants were confirmed as F. falciforme using morphological and sequence analyses. F. falciforme causes root rot in several species including Nicotiana tabacum (Qiu et al. 2023) and Weigela florida (Shen et al. 2020); however, this study is the first to report root rot caused by F. falciforme in strawberries in China. Overall, F. falciforme infection poses a threat to strawberry production and breeding.

Bio-synthesis and characterization of silver nanoparticles from Trichoderma species against cassava root rot disease.

Cassava root rot disease caused by the fungal pathogens Fusarium solani and Lasiodiplodia theobromae produces severe damages on cassava production. This research was conducted to produce and assess silver nanoparticles (AgNPs) synthesized by Trichoderma harzianum for reducing root rot disease. The results revealed that using the supernatants of T. harzianum on a silver nitrate solution changed it to reddish color at 48 h, indicating the formation of AgNPs. Further characterization was identified using dynamic light scattering (DLS) and scanning electron microscope (SEM). DLS supported that the Z-average size is at 39.79 nm and the mean zeta potential is at - 36.5 mV. SEM revealed the formation of monodispersed spherical shape with a diameter between 60-75 nm. The antibacterial action of AgNPs as an antifungal agent was demonstrated by an observed decrease in the size of the fungal colonies using an increasing concentration of AgNPs until the complete inhibition growth of L. theobromae and F. solani at > 58 µg mL-1 and at ≥ 50 µg mL-1, respectively. At in vitro conditions, the applied AgNPs caused a decrease in the percentage of healthy aerial hyphae of L. theobromae (32.5%) and of F. solani (70.0%) compared to control (100%). The SR-FTIR spectra showed the highest peaks in the first region (3000-2800 cm-1) associated with lipids and fatty acids located at 2962, 2927, and 2854 cm-1 in the AgNPs treated samples. The second region (1700-1450 cm-1) consisting of proteins and peptides revealed the highest peaks at 1658, 1641, and 1548 cm-1 in the AgNPs treated samples. The third region (1300-900 cm-1), which involves nucleic acid, phospholipids, polysaccharides, and carbohydrates, revealed the highest peaks at 1155, 1079, and 1027 cm-1 in the readings from the untreated samples. Finally, the observed root rot severity on cassava roots treated with AgNPs (1.75 ± 0.50) was significantly lower than the control samples (5.00 ± 0.00).

First Report of Phytopythium litorale Causing Root Rot on Kousa Dogwood in Tennessee and the United States.

Kousa dogwood (Cornus kousa) is an economically important woody ornamental crop that exhibits creamy, white, pointed bracts in late spring, and reddish to pink drupe fruits in late summer and fall. It bears shiny dark green leaves that become reddish-purple to scarlet in the fall. In August of 2023, 3-year-old container grown C. kousa var. chinensis plants in a commercial nursery in Warren Co., Tennessee, exhibited severe yellowing, dieback and root rot symptoms (Fig. 1a and 1b). Dark brown to black lesions were observed in the root and crown region of the plants. Disease severity was 40% to 60% of root area affected, and disease incidence was approximately 40% of 1,000 plants. Surface-sterilized (10% NaOCl: 1 min) symptomatic root tissues were plated on V8-PARPH and incubated at 25°C. Sparse aerial mycelium, showing a distinct rosette or faint radiate to chrysanthemum colony pattern, was observed within four days of incubation (Fig. 2). All isolates produced ovoid or subglose, papillate, and proliferating sporangia in grass blade water cultures (Dervis et al. 2020). Sporangia measured as 19.18 to 24.80 µm X 18.08 to 22.16 µm (n = 50) with a length/width ratio of 1.06 to 1.11. Zoospores observed were between 7.07 to 9.98 µm in diameter (n = 50). Oogonia and oospores were not produced. The ribosomal internal transcribed spacer (ITS) and large subunit (LSU), as well as mitochondrial cytochrome oxidase subunit II (COX-II) genetic markers were amplified and sequenced using primer pairs ITS1/ITS4 (White et al. 1990), NL1/NL4 (Baten et al. 2014), and cox2-F/cox2-RC4 (Choi et al. 2015), respectively. The ITS, LSU, and COX-II sequences of isolates FBG6343, FBG6344 (ITS: PP458373 and PP461387; LSU: PP461390 and PP461391; COXII: PP477112 and PP477113) were 100% identical to those of MN306118, HQ643386, and MN206732, respectively. Based on the morphology (Nechwatal and Mendgen 2006) and sequence data, the isolates were identified as Phytopythium litorale (Nechw.) Abad, De Cock, Bala, Robideau, Lodhi & Lévesque. The pathogenicity test was performed on 3-year-old C. kousa var. chinensis plants grown in a 3-gal container to fulfill Koch's postulates. Kousa dogwood plants were drench inoculated (800 ml/plant) with a pathogen slurry (two plates of 7-day-old culture/liter) of isolates FBG6343 and FBG6364 (five plants per isolate). Control plants were drenched with agar slurry without the pathogen. The study was conducted in a greenhouse maintained at 21 to 23°C and 70% relative humidity with a 16-h photoperiod and irrigated twice a day for 2 min using an overhead irrigation system. Forty-five days after inoculation, plants showed dieback symptoms, and dark brown lesions developed in the roots of all inoculated plants. Isolates with morphology and sequences identical to those of FBG6343 and FBG6364 were recovered from root tissues of all inoculated plants. All control plants remained symptom-free, and P. litorale was not isolated from the root tissue. Previously, P. litorale was reported to cause disease on apple, kiwi, planatus, and rhododendron (Dervis et al. 2020; Li et al. 2021; Mert et al. 2020; Polat et al. 2023). To our knowledge, this is the first report of P. litorale causing root rot of kousa dogwood in Tennessee and the United States. Identification of this pathogen as the causal agent is crucial to developing timely management practices.

First report of Stagonosporopsis pogostemonis causing root rot on Strawberry in China.

In Henan, strawberry cultivation occurs on approximately 10,000 hectares, with annual production approaching 230,000 tons. In April 2022, a root rot disease with a 10% incidence rate was observed on the strawberry cultivars 'Ningyu' and 'Sweet Charlie' grown in plastic greenhouses (0.7 ha) located in Xingyang (113.39°E, 34.79°N), Henan, China. Disease symptoms included yellowing of the outer mature leaves, stunted growth, and subsequent wilting of the entire plant. The roots developed dark brown spots, which gradually turned necrotic (Figures 1a, 1b). To determine the causal agent, four symptomatic plants (two plants per cultivar) were collected. Twelve symptomatic root tissues (three root tissue samples per plant) were surface sterilized with 75% ethanol and 0.1% mercuric chloride, washed thrice in sterile water, air dried, and then placed on PDA at 25°C for 3 days. Eight pure isolates were obtained by hyphal-tip isolation (Fang 2007). Each colony had a dark olivaceous green to brown, cottony appearance with a round margin, and the reverse side was grey-black near the center (Figure 1c). Conidia were ellipsoidal, aseptate, with rounded ends, and 3.1 to 4.8 µm × 1.0 to 2.5 µm in size (Figure 1d). Chlamydospores were ellipsoidal, pale brown, and 7.9 to 11.9 µm × 7.6 to 10.7 µm in size (Figure 1e). A representative fungus isolate, designated as Z5, was selected for further molecular identification. Genomic DNA was extracted from the mycelia of isolate Z5, and four gene partial regions (ITS, TUB2, RPB2, and LSU) were amplified using the primer pairs ITS1/ITS4, Bt-2a/Bt-2b, RPB2-5F/RPB2-7CR and LROR/LR5, respectively (White et al.1990, O'Donnell et al.1997, Reeb et al. 2004, Rehner and Samuels 1994). PCR products were sequenced and submitted to GenBank with the following accession numbers OQ130480 (ITS), OQ190093 (TUB2), OQ190092 (RPB2), and OQ255570 (LSU). BLASTn search revealed that the ITS, TUB2, RPB2, and LSU gene sequences of isolate Z5 showed 99.42% (513/516 bp), 99.69% (320/321 bp), 100% (1071/1071 bp), and 100% (857/857 bp) identity with those of ex-type S. pogostemonis stain ZHKUCC 21-0001 (Dong et al. 2021), respectively. A phylogenetic tree was constructed showing that Z5 clustered with S. pogostemonis (Figure 2). The isolates in this study were identified as S. pogostemonis based on morphological and molecular evidence. To confirm pathogenicity, five one-month-old 'Ningyu' cultivar strawberry seedlings were planted in sterilized nursery soil mixed with wheat grains (0.5% w/w) coated with Z5 mycelia (Fang 2007). An equal number of strawberry seedlings were placed in pots filled with non-infected potting mix to serve as controls. The seedlings were kept in a greenhouse under a 12 h light/dark photoperiod at 25°C. After two weeks, the inoculated seedlings displayed symptoms such as leaf wilting and root necrosis, similar to those observed in the greenhouses, while the control seedlings showed no symptoms (Figures 1f, 1g). The experiment was performed thrice. The identical fungus was re-isolated from the symptomatic roots and identified as S. pogostemonis based on morphological characteristics and molecular analysis, thus fulfilling Koch's postulates. This is the first report of S. pogostemonis causing root rot on strawberries worldwide. Our findings will contribute to a more comprehensive study on investigating and managing this disease.

A new Schizophyllum commune strain as a potential biocontrol agent against blueberry root rot.

In recent years, blueberry root rot has been caused mainly by Fusarium commune, and there is an urgent need for a green and efficient method to control this disease. To date, research on Schizophyllum commune has focused on antioxidant mechanisms, reactive dye degradation, etc., but the mechanism underlying the inhibition of pathogenic microorganisms is still unclear. Here, the control effects of S. commune on F. commune and blueberry root rot were studied using adversarial culture, tissue culture, and greenhouse pot experiments. The results showed that S. commune can dissolve insoluble phosphorus and secrete various extracellular hydrolases. The results of hyphal confrontation and fermentation broth antagonism experiments showed that S. commune had a significant inhibitory effect on F. commune, with inhibition rates of 70.30% and 22.86%, respectively. Microscopy results showed distortion of F. commune hyphae, indicating that S. commune is strongly parasitic. S. commune had a significant growth-promoting effect on blueberry tissue-cultured seedlings. After inoculation with S. commune, inoculation with the pathogenic fungus, or inoculation at a later time, the strain significantly reduced the root rot disease index in the potted blueberry seedlings, with relative control effects of 79.14% and 62.57%, respectively. In addition, S. commune G18 significantly increased the antioxidant enzyme contents in the aboveground and underground parts of potted blueberry seedlings. We can conclude that S. commune is a potential biocontrol agent that can be used to effectively control blueberry root rot caused by F. commune in the field.

Pathogenicity, Host Resistance, and Genetic Diversity of Fusarium Species under Controlled Conditions from Soybean in Canada.

Fusarium spp. are commonly associated with the root rot complex of soybean (Glycine max). Previous surveys identified six common Fusarium species from Manitoba, including F. oxysporum, F. redolens, F. graminearum, F. solani, F. avenaceum, and F. acuminatum. This study aimed to determine their pathogenicity, assess host resistance, and evaluate the genetic diversity of Fusarium spp. isolated from Canada. The pathogenicity of these species was tested on two soybean cultivars, 'Akras' (moderately resistant) and 'B150Y1' (susceptible), under greenhouse conditions. The aggressiveness of the fungal isolates varied, with root rot severities ranging from 1.5 to 3.3 on a 0-4 scale. Subsequently, the six species were used to screen a panel of 20 Canadian soybean cultivars for resistance in a greenhouse. Cluster and principal component analyses were conducted based on the same traits used in the pathogenicity study. Two cultivars, 'P15T46R2' and 'B150Y1', were consistently found to be tolerant to F. oxysporum, F. redolens, F. graminearum, and F. solani. To investigate the incidence and prevalence of Fusarium spp. in Canada, fungi were isolated from 106 soybean fields surveyed across Manitoba, Saskatchewan, Ontario, and Quebec. Eighty-three Fusarium isolates were evaluated based on morphology and with multiple PCR primers, and phylogenetic analyses indicated their diversity across the major soybean production regions of Canada. Overall, this study contributes valuable insights into host resistance and the pathogenicity and genetic diversity of Fusarium spp. in Canadian soybean fields.

Allochthonous Trichoderma Isolates Boost Atractylodes lancea Herb Quality at the Cost of Rhizome Growth.

Atractylodes lancea is a perennial herb whose rhizome (AR) is a valuable traditional Chinese medicine with immense market demand. The cultivation of Atractylodes lancea faces outbreaks of root rot and deterioration in herb quality due to complex causes. Here, we investigated the effects of Trichoderma spp., well-known biocontrol agents and plant-growth-promoters, on ARs. We isolated Trichoderma strains from healthy ARs collected in different habitats and selected three T. harzianum strains (Th2, Th3 and Th4) with the strongest antagonizing effects on root rot pathogens (Fusarium spp.). We inoculated geo-authentic A. lancea plantlets with Th2, Th3 and Th4 and measured the biomass and quality of 70-day-old ARs. Th2 and Th3 promoted root rot resistance of A. lancea. Th2, Th3 and Th4 all boosted AR quality: the concentration of the four major medicinal compounds in ARs (atractylon, atractylodin, hinesol and ß-eudesmol) each increased 1.6- to 18.2-fold. Meanwhile, however, the yield of ARs decreased by 0.58- to 0.27-fold. Overall, Th3 dramatically increased the quality of ARs at a relatively low cost, namely lower yield, showing great potential for practical application. Our results showed selectivity between A. lancea and allochthonous Trichoderma isolates, indicating the importance of selecting specific microbial patches for herb cultivation.

The Host Range of Fusarium proliferatum in Western Canada.

Fusarium proliferatum is associated with the root rot of many plant species, but knowledge of its impact on western Canadian field crops is limited. This study assessed the host range of this fungus and its effect on plant emergence, plant height, and shoot and root dry weights in repeated greenhouse experiments with wheat, barley, faba beans, peas, lentils, canola, lupine, and soybeans. Infection was confirmed via PCR, and principal component analysis determined the utility of different parameters in assessing host responses. All crops were at least partly susceptible, developing mild to severe disease at the seedling and adult stages, and showing significant reductions in growth. In general, the barley and wheat demonstrated higher tolerances to infection, followed by the faba bean and the pea. The soybean, canola, lupine, and lentil were most susceptible. The canola and the soybean were particularly vulnerable to F. proliferatum at the pre-emergence stage, while infection greatly reduced the lentil's biomass. Reductions in the barley's emergence and other growth parameters, however, occurred only under a high inoculum concentration. Variability in root rot severity among cultivars of the same crop indicated some diversity in host reactions within species. Nonetheless, the absence of fully-resistant crops may pose challenges in managing F. proliferatum in western Canadian cropping systems.

A MYB-related transcription factor regulates effector gene expression in an oomycete pathogen.

Phytophthora pathogens possess hundreds of effector genes that exhibit diverse expression patterns during infection, yet how the expression of effector genes is precisely regulated remains largely elusive. Previous studies have identified a few potential conserved transcription factor binding sites (TFBSs) in the promoters of Phytophthora effector genes. Here, we report a MYB-related protein, PsMyb37, in Phytophthora sojae, the major causal agent of root and stem rot in soybean. Yeast one-hybrid and electrophoretic mobility shift assays showed that PsMyb37 binds to the TACATGTA motif, the most prevalent TFBS in effector gene promoters. The knockout mutant of PsMyb37 exhibited significantly reduced virulence on soybean and was more sensitive to oxidative stress. Consistently, transcriptome analysis showed that numerous effector genes associated with suppressing plant immunity or scavenging reactive oxygen species were down-regulated in the PsMyb37 knockout mutant during infection compared to the wild-type P. sojae. Several promoters of effector genes were confirmed to drive the expression of luciferase in a reporter assay. These results demonstrate that a MYB-related transcription factor contributes to the expression of effector genes in P. sojae.