First report of Fusarium meridionale causing Fusarium head blight of wheat in Henan Province, China.

Fusarium graminearum and F. asiaticum have been found as a major cause of Fusarium head blight (FHB) of wheat (Triticum aestivum L.), especially in Henan Province of China (Zhang et al. 2014; Xu et al. 2021). In May 2021, a survey to determine the composition of Fusarium species infecting wheat heads was conducted in commercial fields in Henan. A total of 395 diseased spikes with premature whitening symptom were collected from 31 commercial fields in Henan. Symptomatic spikelets were excised, surface-sterilized for 10 s in 70% ethanol followed by 1 min in 3% sodium hypochlorite, rinsed three times with autoclaved distilled water, and then plated onto potato dextrose agar (PDA) medium. Isolated colonies that resembled Fusarium species were transferred to fresh PDA plates and purified using a single spore method. Species were identified based on sequence analysis of the translation elongation factor-1α (TEF) and trichothecene 3-Oacetyltransferase (Tri 101) gene (Proctor et al. 2009). The results indicated that F. graminearum (43.3%), F. asiaticum (47.8%), F. pseudograminearum (6.6%) were the main causal agents of FHB in Henan. However, nine isolates (2.3%) were found to be identical to F. meridionale by sequence comparison in GenBank, and eight isolates of which came from three fields with 1% to 2% diseased spikes near Reservoir Luhun (34.1255° N, 112.1111° E, altitude: 388 m above sea level), Songxian County of Henan. The isolates of F. meridionale were transferred onto carnation leaf agar (CLA) and incubated at 20℃ under black light blue illumination. Macroconidia were abundant, relatively slender, curved to almost straight, commonly six- to seven-septate, and 27.0 to 61.0 (average 44.0) µm × 3.2 to 6.8 (average 5.3) µm. Microconidia were not observed. The TEF sequences (Accession nos. OM460748 to OM460756) and the Tri 101 sequences (OM460759 to OM460767) of the nine isolates showed 99 to 100% similarity with the TEF and Tri 101 sequences of F. meridionale NRRL 28436 and NRRL 28723 (AF212435 and AF212436 (TEF); AF212582 and AF212683 (Tri 101)). To complete Koch's postulates, the pathogenicity of the fungus was tested by using the single floret inoculation method by injecting 20-µl conidial suspension (5 × 105 conidia per milliliter) into healthy inflorescences of wheat cultivar Bainong 207 at anthesis in the field. Another 30 healthy inflorescences were injected with sterile distilled water. The heads were covered with polyethylene bags that were removed after 2 days. Twenty days after inoculation, while control inflorescences were asymptomatic, the F. meridionale-inoculated inflorescences showed 12% bleached spikelets per spike. By using the methodology described above, the fungus was re-isolated from infected spikelets of inoculated wheat heads but not from the controls. Although F. meridionale has frequently been reported in association with Fusarium ear rot (FER) of maize in Chongqing City and Gansu Province (Zhang et al. 2014; Zhou et al. 2018), and rice FER in Sichuan Province (Dong et al. 2020), to our knowledge, this is the first report of F. meridionale from diseased wheat heads in Henan, China. Further investigation is needed to gain a better understanding of this species by collecting isolates from different cropping system in Henan, which maize-wheat and rice-wheat rotation fields have coexisted in the region.

Candidate powdery mildew resistance gene in wheat landrace cultivar Hongyoumai discovered using SLAF and BSR-seq.

BACKGROUND: Wheat powdery mildew, caused by Blumeria graminis f. sp. tritici (Bgt), is an important disease affecting wheat production. Planting resistant cultivars is an effective, safe, and economical method to control the disease. Map construction using next-generation sequencing facilitates gene cloning based on genetic maps and high-throughput gene expression studies. In this study, specific-locus amplified fragment sequencing (SLAF) was used to analyze Huixianhong (female parent), Hongyoumai (male parent) and two bulks (50 homozygous resistant and 50 susceptible F2:3 segregating population derived from Huixianhong × Hongyoumai to determine a candidate gene region for resistance to powdery mildew on the long arm of chromosome 7B in wheat landrace Hongyoumai. Gene expressions of candidate regions were obtained using bulked segregant RNA-seq in 10 homozygous resistant and 10 susceptible progeny inoculated by Bgt.. Candidate genes were obtained using homology-based cloning in two parents. RESULTS: A 12.95 Mb long candidate region in chromosome 7BL was identified, and five blocks in SLAF matched the scaffold of the existing co-segregation marker Xmp1207. In the candidate region, 39 differentially expressed genes were identified using RNA-seq, including RGA4 (Wheat_Chr_Trans_newGene_16173)-a disease resistance protein whose expression was upregulated in the resistant pool at 16 h post inoculation with Bgt. Quantitative reverse transcription (qRT)-PCR was used to further verify the expression patterns in Wheat_Chr_Trans_newGene_16173 that were significantly different in the two parents Hongyoumai and Huixianhong. Two RGA4 genes were cloned based on the sequence of Wheat_Chr_Trans_newGene_16173, respectively from two parent and there was one amino acid mutation: S to G in Huixianhong on 510 loci. CONCLUSION: The combination of SLAF and BSR-seq methods identified a candidate region of pmHYM in the chromosome 7BL of wheat landrace cultivar Hongyoumai. Comparative analysis between the scaffold of co-segregating marker Xmp1207 and SLAF-seq showed five matching blocks. qRT-PCR showed that only the resistant gene Wheat_Chr_Trans_newGene_16173 was significantly upregulated in the resistant parent Hongyoumai after inoculation with Bgt, and gene cloning revealed a difference in one amino acid between the two parent genes, indicating it was involved in the resistance response and may be the candidate resistance gene pmHYM.

First report of Fusarium falciforme causing root rot of soybean (Glycine max L.) in Henan, China.

Soybean (Glycine max L.) is an important crop in China owing to its high oil and protein content, with approximately 9.88 million ha of production in 2020. In September 2021, soybean plants showing wilting, root necrosis, and brown discoloration were observed, with an average incidence of approximately 36% in seven fields in Yongcheng City and Shangqiu City, Henan, China. Fungi were isolated from small pieces of symptomatic root tissues after being surface-sterilized (70% ethanol for 50 s followed by 3% NaClO for 1.0 min), rinsed three times in sterile distilled water, and then placed on PDA and incubated at 25℃ for 5 days in the dark. Single-spore cultures of twenty isolates were obtained by dilution plating (Leslie and Summerell 2006), and then were cultured on carnation leaf agar at 25℃ for 14 days. Macroconidia were mostly 3-septate, hyaline, falcate, with slightly curved apexes, with well-developed foot cells and blunt apical cells, and measured 29.3 to 45.0 (average 34.7) µm × 4.6 to 8.0 (average 6.0) µm. Microconidia were one to two celled, hyaline, and measured 11.9 to 29.0 (average 20.1) µm × 3.9 to 7.6 (average 5.7) µm. These morphological characteristics were consistent with previous descriptions of the Fusarium solani species complex (FSSC) (Leslie and Summerell 2006; Summerell et al. 2003). Partial sequences of translation elongation factor-1α (TEF) and RNA polymerase II subunit (RPB2) gene were PCR amplified using region specific primers as described by O'Donnell et al. (2008). The nucleotide sequences obtained from twenty isolates were deposited in GenBank with accession numbers of ON375405-ON375423, ON697187 (TEF) and ON331917-ON331936 (RPB2). Phylogenetic analysis revealed the isolates were nested within F. falciforme based on the DNA sequences of the above two genes (Chitrampalam and Nelson 2016). Pathogenicity tests of two representative isolates (21BeanYC3-3 and 21BeanYC7-5) were performed on two-week-old healthy soybean seedlings (cv. Shengdou 101) by injecting and cutting root method with a conidial suspension (1×106 conidia per mL) of F. falciforme (2 mL to one seedling). Control seedlings were inoculated with 2 mL distilled water. After 40 days under 25℃, 16h light/8h dark, the root system of all inoculated soybean plants exhibited dark brown lesions over the entire taproot, while control plants remained healthy. The fungus was reisolated from inoculated plants and identified as F. falciforme based on morphological characteristics and molecular methods described above. To our knowledge, this is the first report of root rot in soybean (Glycine max L.) caused by F. falciforme in Henan, China. The results are important for soybean production and breeding programs.

Fusarium pseudograminearum biomass and toxin accumulation in wheat tissues with and without Fusarium crown rot symptoms.

Fusarium crown rot (FCR) is an important and devastating disease of wheat (Triticum aestivum) caused by the fungus Fusarium pseudograminearum and related pathogens. Using two distinct susceptible cultivars, we investigated the isolation frequencies of F. pseudograminearum and quantified its biomass accumulation and the levels of the associated toxins deoxynivalenol (DON) and DON-3-glucoside (D3G) in inoculated field-grown wheat plants. We detected F. pseudograminearum in stem, peduncle, rachis, and husk tissues, but not in grains, whereas DON and D3G accumulated in stem, rachis, husk, and grain tissues. Disease severity was positively correlated with the frequency of pathogen isolation, F. pseudograminearum biomass, and mycotoxin levels. The amount of F. pseudograminearum biomass and mycotoxin contents in asymptomatic tissue of diseased plants were associated with the distance of the tissue from the diseased internode and the disease severity of the plant. Thus, apparently healthy tissue may harbor F. pseudograminearum and contain associated mycotoxins. This research helps clarify the relationship between F. pseudograminearum occurrence, F. pseudograminearum biomass, and mycotoxin accumulation in tissues of susceptible wheat cultivars with or without disease symptoms, providing information that can lead to more effective control measures.

Biological control of Fusarium crown rot of wheat with Chaetomium globosum 12XP1-2-3 and its effects on rhizosphere microorganisms.

Chaetomium globosum is a common plant endophytic fungi that exhibits great biocontrol potential in plant disease. Fusarium crown rot (FCR) is an important disease in wheat that seriously threatens wheat production worldwide. The control effect of C. globosum against wheat FCR remains unclear. In this study, we introduced an identified C. globosum 12XP1-2-3 and tested its biological control potential against wheat FCR. The hypha and fermentation broth exhibited an antagonistic effect against Fusarium pseudograminearum. Results from indoor experiments showed that C. globosum 12XP1-2-3 might delay the onset of symptoms of brown stem base and significantly reduced the disease index (37.3%). Field trials showed that wheat seeds coated with a spore suspension of 12XP1-2-3 grew better than the control seeds, had control effects of 25.9-73.1% on FCR disease, and increased wheat yield by 3.2-11.9%. Analysis of rhizosphere microorganisms revealed that seeds coated with C. globosum ('Cg' treatment) had a greater effect on fungal rather than on bacterial alpha diversity and may improve the health state of rhizosphere microorganisms, as reflected by the significantly increased fungal Shannon index at Feekes 11 and the increased complexity of the bacterial co-occurrence network but decreased complexity of the fungal network. Moreover, the accumulation of beneficial bacteria such as Bacillus and Rhizobium at Feekes 3, and Sphingomonas at Feekes 7 in the 'Cg' treatment may be the important contributions to healthier wheat growth state, significantly reduced relative abundance of Fusarium at Feekes 11, and reduced occurrence of FCR disease. These results provide a basis for further research on the mechanism of action of C. globosum and its application in the biological control of FCR in the field.

Spatial Distribution of Root and Crown Rot Fungi Associated With Winter Wheat in the North China Plain and Its Relationship With Climate Variables.

The distribution frequency of pathogenic fungi associated with root and crown rot of winter wheat (Triticum aestivum) from 104 fields in the North China Plain was determined during the period from 2013 to 2016. The four most important species identified were Bipolaris sorokiniana (24.0% from roots; 33.7% from stems), Fusarium pseudograminearum (14.9% from roots; 27.8% from stems), Rhizoctonia cerealis (1.7% from roots; 4.4% from stems), and Gaeumannomyces graminis var. tritici (9.8% from roots; 4.4% from stems). We observed that the recovered species varied with the agronomic zone. Fusarium pseudograminearum was predominant in regions 1 and 3, whereas F. graminearum, F. acuminatum, and R. cerealis were predominant in regions 2 and 4. The incidence of F. pseudograminearum and R. cerealis was significantly different between regions 1 and 4, while no significant association was found in the distribution of the other species and the agronomic zones. A negative correlation between the frequency of occurrence of F. pseudograminearum and mean annual precipitation during 2013-2016 (r = -0.71; P < 0.01) in the North China Plain and a positive correlation between the mean annual precipitation during 2013-2016 and the frequency of occurrence of F. asiaticum (r = 0.74; P < 0.01) were observed. Several Fusarium species were also found with low frequencies of ~2.1%-3.4 % (F. graminearum, F. acuminatum, and F. sinensis) and ~0.1%-1.3% (F. equiseti, F. oxysporum, F. proliferatum, F. culmorum, F. avenaceum, and F. asiaticum). In more than 93% of the fields, from the root and crown tissues of wheat, two or more root and crown rot species were isolated. The coexistence of Fusarium spp. and B. sorokiniana in one field (65.4%) or in individual plants (11.6%) was more common than for the other species combinations. Moreover, this is the first report on the association between F. sinensis and root and crown rot of wheat. Our results would be useful in the framing guidelines for the management of root and crown rot fungi in wheat in different agronomic zones of the North China Plain.