Cucurbit chlorotic yellows virus (CCYV) can infect a variety of melons, mainly affecting the leaves and photosynthesis which ultimately results in reduced fruit weight. CCYV was detected by siRNA sequencing of suspected viral disease samples from Hunan province. Further analysis by RT-PCR confirmed that CCYV can naturally infect pumpkins with a detection rate of 5.78%. This is the first time that CCYV is being reported to naturally infect pumpkins in mainland China and it is found that the occurrence of pumpkins-infecting CCYV in Hunan province is increasing each year. The CCYV CP gene was cloned and analyzed for phylogeny, genetic differences, selection pressure and genetic drift. It was found that geographical distribution of the CCYV population was significantly correlated. Natural selection and genetic drift are the main drivers of CCYV evolution, but there is a positive selection effect in Chinese populations. Since CCYV is undergoing adaptive changes.CCYV population is expanding.Keeping in view the current spread of this virus, it might emerge as serious pathogen and cause more crop losses in the future. Thus, the results of our research are of significant importance for the early detection and prevention of CCYV infecting pumpkins.

Some VQ motif-containing proteins interact with WRKY transcription factors, and play essential regulatory roles in disease defense response. In the present study, quantitative real-time PCR was performed to analyze the expression profiles of rice VQ gene family in response to three defense-related hormones and sheath blight fungus Rhizoctonia solani. The results showed that more than one-third of rice VQ gene members were responsive to at least one treatment. OsVQ2,-11 and -35 were significantly upregulated by salicylic acid (SA), jasmonic acid (JA) and ethylene (ET). Among them, OsVQ2 was also obviously induced by R. solani. Specially, OsVQ2, -35, -34 and -37 were the most markedly induced genes under the treatments of SA, JA, ET and R. solani, respectively. Prediction of cis-elements revealed that mesophyll expression modules and pathogen-responsive elements containing W-box core sequences were over-represented within the promoters of R. solani-responsive VQ genes, in basically consistent with the gene expression patterns and the tissue identity of pathogen infection. These findings suggest that some rice VQ genes may be involved in the defense response against sheath blight possibly through SA, JA or ET signaling pathway, and provide a solid foundation for gene function study in the future.

Sexual reproduction plays an important role in the life cycle and evolution of the fungi. Mating type genes are the master regulators of sexual reproduction. The MAT1-2 strain of Villosiclava virens contains two mating type genes, MAT1-2-1 and MAT1-2-8. However, how these two genes regulate sexual reproduction of V. virens remains largely unknown. In this study, we demonstrated the expression pattern of MAT1-2-1 and MAT1-2-8 at different stages of infection and development. We also analyzed the structural characteristics of these two proteins. The results showed that the expression level of MAT1-2-1 was reduced during infection stages. MAT1-2-8 was up-regulated at the early infection stage (5 dpi), and then decreased during late infection stage. Compared with the vegetative mycelium stage, the expression levels of both MAT1-2-1 and MAT1-2-8 were reduced in the four stages of sexual development sclerotia formation, sclerotia germination, stromata primordium formation and mature stromata, and the expression level was the lowest in the sclerotia formation stage. Bioinformatics analysis showed that MAT1-2-1 and MAT1-2-8 have phosphorylation sites, but no secretion peptides and no obvious transmembrane domains. Homologous alignment analysis showed that MAT1-2-1 shared high homology with its homolog in Epichloë typhina, and MAT1-2-8 shared high homology with its homolog in Metarhizium. Further study showed that MAT1-2-1 interacted with MAT1-2-8, and they were mainly localized in the nucleus and the cytoplasm, respectively. In addition, we identified several candidate proteins that might interact with MAT1-2-1 via mass spectrometry, including the putative Ran exchange factor Prp20/Pim1 (KDB12229.1), the putative rRNA processing protein (Ebp2) (KDB12923.1), and histone H1 (KDB12711.1). Collectively, these results provide fundamental understanding biological functions of the mating type genes MAT1-2-1 and MAT1-2-8 on sexual reproduction.

Cyclic diguanylate monophosphate (c-di-GMP) is a second messenger in many phytopathogenic bacteria and regulates a variety of biological functions including pathogenicity. However, no functional study on c-di-GMP in Acidovorax citrulli has ever been reported. The gene Aave_2620 in A. citrulli contains domains related to c-di-GMP metabolism. We generated the ΔAave_2620 mutant in A. citrulli strain Aac5, as well as its complementation strain ΔAave_2620comp. The virulence, biofilm formation and bacterial growth were significantly reduced in the mutant ΔAave_2620 compared to its wild-type Aac5. The mutant ΔAave_2620 was also decreased in the ability triggering hypersensitive reaction on non-host tobacco, and decreased in swimming motility. The complemented strain ΔAave_2620comp recovered all aforementioned biological traits. The expression of genes related to Type III secretion system, virulence, flagellum assembly and chemotaxis were significantly affected in mutant ΔAave_2620, indicating the importance of gene Aave_2620 in the virulence of A. citrulli.

Potato virus Y is one of the major viruses infecting tobacco plant and different strains induce different symptom in tobacco. Some PVY isolates induce veinal necrosis in Nicotiana tabacum and severely affect the yield and quality of tobacco leaves. The PVY isolate A12 belongs to the NTN-NW strain, and induces symptom of mosaic, instead of veinal necrosis in Nicotiana tabacum cv. Xanthi. The other NTN-NW isolates that induce veinal necrosis in Nicotiana tabacum have the amino acid residue lysine (K) at both positions 182 and 245, while A12 has the residue arginine (R) at both positions. The corresponding amino acid residues at positions 182 and 245 are also K in the HC-Pro of PVY necrosis isolate N605. Mutation was introduced to infectious clone PVY^N605-GFP via site-directed mutagenesis. The inoculation results showed that the mutant with substitution of R for K at position 182 in HC-Pro could not induce veinal necrosis in N. tabacum cv. Xanthi. Western blot results showed that there was no significant difference in the CP accumulation levels between the wild type and mutant PVY. The RNA silencing suppression assay showed that there was no significant change between wild type and mutant HC-Pro. Therefore, we concluded that the residue K at position 182 of HC-Pro was an critical amino acid responsible for tobacco veinal necrosis, and presumed that the residue R at position 182 of HC-Pro was accountable for the failure of A12 to induce tobacco veinal necrosis.

Squash blight caused by Phytophthora capsici is one of the primary diseases of seed-used pumpkin in Heilongjiang, China. It leaded to serious rotten and dead seedlings and the planting area decreased year by year. This study investigated and analysed the planting situation and the blight occurrence of seed-used pumpkin in Heilongjiang from 2017 to 2019. Further, 106 strains of pathogens obtained from the areas with high incidence of squash blight were separated and purified with the methods of tissues isolation. The identification of pathogens was done by the morphological and molecular biology. The pathogenicity of some representative isolated strains was determined on the leaves in vitro, and the pathogenicity of the tested isolates was stronger than that of the control standard isolates. Using the combination methods of root inoculation and leaf inoculation, 112 preserved germplasm resources of seed-used pumpkin materials were identified and evaluated for the resistance of squash blight. Two materials showed high resistance, 3 materials showed resistance, 9 materials showed moderate resis-tance and the rest of the materials were susceptible. This study provided a theoretical basis for occurring and harming of squash blight on seed-used pumpkin, analysis of pathogenicity in Heilongjiang, also provided the source of the materials for the resistant breeding, the new resistant source screening and the resistant gene mining of seed-used pumpkin.

Soilborne disease Fusarium wilt, caused by Fusarium oxysporum and F. solani threatens chrysanthemum production seriously. However, there was no report on the difference of physiological and biochemical responses between the resistant and susceptible varieties. To establish the physiological and biochemical indexes for the selection of resistant varieties, two Fusarium strains with strong pathogenicity were inoculated into the seedlings of five Chrysanthemum morifolium varieties, and the morphological, physiological and biochemical responses of different varieties infected with the pathogen were measured in this study.    The results showed that the disease index of C.morifolium was negaticely correlated with disease resistance after infection . With the development of the disease, the content of leaf chlorophyll (Chl) content increased slightly and then decreased, and the content of Chl in resistant variety ‘Qiaofenge' was higher than the other varieties tested. The content of malondialdehyde (MDA) increased after infection. The MDA content was significantly higher in susceptible variety ‘Linglong' than those in resistant varieties, and it was negatively correlated with disease resistance of varieties. The proline content increased rapidly in resistant varieties, showing that the higher the content of peoline, the stronger the disease resistance of varieties, The content of soluble protein (SPC) increased firstly and then decreased during infection and it was higher in resistant varieties than that in susceptible ones. The activities of leaf superoxide dismutase (SOD) and peroxidase (POD) increased first and then decreased following infection. SOD and POD activities in resistant varieties increased rapidly and they were higher than susceptible ones.In summary, through evaluation of resistance and measurement of various physiological indicators and disease index after inoculation,the resistance of five Ground cover chrysanthemum varieties to F. oxysporum from high to low was ‘Qiaofenge' > ‘Ruhe'>‘ Huoyan '> ‘Xianhong' > ‘Lingnong', while the disease resistance to F. solani from high to low was ‘Qiaofenge'> ‘Ruhe' > ‘Xianhong'> ‘ Huoyan' > ‘Lingnong'. This provides basis for deployment of chrysanthemum cultivars in production and molecular bree-ding of disease resistant .

Puccinia triticina, the cause of wheat leaf rust, is a heteroecious parasite. Its alternate hosts include mainly Thalictrum spp., and several species in genera Isopyrum, Anchusa, Clematis, and Echium. So far, only two Thalictrum species, T. minus L. and T. petaloideum L., have been reported as alternate hosts for the rust pathogen in China. In the present study, two new species of Thalictrum, T. minus var. hypoleucum and T. baicalense, collected from Shaanxi and Gansu provinces were tested their susceptibility to P. triticina through artificially inoculation to determine as alternate hosts for the rust fungus under controlled conditions. Simultaneously, we collected aecial samples from naturally infected tissues of T. baicalense plants and tested them to determine whether they were P. triticina by sequence alignment of ITS regions and sexual cycle of the rust fungus could occur in field. The results showed that T. minus var. hypoleucum and T. baicalense can infect the two Thalictrum species to complete their pycnial and aecial stages, and that aeciospores can infect susceptible wheat cultivar to produce uredinia, indicating T. minus var. hypoleucum and T. baicalense serve as alternate hosts for P. triticina. Twenty two aecial samples collected from naturally infected leaves of T. baicalense were identified as P. triticina as they had 95%-96% sequence homologies with the ITS region sequences of P. triticina submitted in the NCBI database. This hinted P. triticina could infect susceptible Thalictrum spp. to complete sexual cycle under natural conditions in China.

To clarify the development status of the resistance of Phytophthora infestans to fluopicolide in one-cropping areas, the sensitivities of 520 P. infestans strains collected from main potato production areas of Hebei Province, Inner Mongolia Autonomous Region, Liaoning province, Jilin province and Heilongjiang Pro-vince during 2012 and 2016 to fluopicolide were determined by the mycelial growth inhibition test. Also the control efficacies of 9 conventional fungicides to potato late blight were assayed in the field through spraying stems and leaves with fungicide. The results showed that the population of P. infestans tended to be less sensitive to fluopicolide and developed low resistance to fluopicolide widely all over the main potato production area of Hebei province, Inner Mongolia Autonomous Region, Liaoning Province, Jilin Province and Heilongjiang Province, with the average resistance factor and index of 3.45 and 0.48, respectively. The resistance frequency was 90.80% among all tested strains where the strains with low resistance accounted for 90.35%. Three moderate resistant strains were firstly detected in 2016. The trials of control efficacy of fungicides were performed in field in 2016 and 2017 with fungicide sprayed four times under the recommended doses. The result showed that the control efficacy of fluopicolide·propamocarb hydrochloride 687.5 g·L^-1 SC was significantly higher than mefenoxam·mancozeb 68% WG, mancozeb 80% WP, azoxystrobin 250 g·L^-1 SC and fluazinam 500 g·L^-1 SC, but the control efficacy of fluopicolide·propamocarb hydrochloride 687.5 g·L^-1 SC in 2017 (86%) was slightly lower than that in 2016 (89.5%). Above all results, the mixing formulation of flupicolide and propamocarb hydrochloride fluopicolide·propamocarb hydrochloride 687.5 g·L^-1 SC still had a high efficacy in controlling late blight currently, but it was necessary to monitor the dynamics of the resistance of P. infestans to fluopicolide, and to carry out the management strategies of resistance, including limiting the application times of fluopicolide·propamocarb hydrochloride 687.5 g·L^-1 SC within a growth season and applying fluopicolide·propamocarb hydrochloride 687.5 g·L^-1 SC in turn with other fungicides with different mechanism of action to control late blight.

To determine the sensitivity of Magnaporthe oryzae to trifloxystrobin, 220 M. oryzae isolates, obtained from 11 rice producing regions in Liaoning Province during 2018 and 2019, were tested by mycelium growth rate method. Then the resistance level of the tested isolates was analyzed. The results showed that the EC₅₀ values of M. oryzae to trifloxystrobin ranged from 0.011 1 to 0.498 3μg·mL^-1. The difference between the highest EC₅₀ value and the lowest EC₅₀ value was 44.89 times. The sensitivity distribution frequency of M. oryzae to trifloxystrobin did not conform to the normal distribution, suggesting that the population with decreased sensitivity to trifloxystrobin was emerged. According to the theory that the sensitivity of wild sensitive pathogens to fungicides was normally distributed, the average EC₅₀ value of 91.82% strains ((0.043 0±0.017 9) μg·mL^-1) which were conform to normal distribution, was used as the baseline sensitivity of M. oryzae to trifloxystrobin. The range of the resistance ratio of M. oryzae to trifloxystrobin in Liaoning Province was from 0.256 9 to 11.493 8 with the average value at 0.27, and the frequency of resistant isolates was 6.82%. At present, most isolates were sensitive to trifloxystrobin and the fungicide can be used as the main fungicide to control rice blast. However, the resistant strains have been found in some regions, which indicated that the alternate fungicides with different mechanisms should be used in turn with trifloxystrobin.

Bacterial Fruit Blotch (BFB), caused by Acidovorax citrulli, is a typical seed-borne bacterial di-sease on cucurbitaceae crops, including watermelon and melon. It has been reported that A. citrulli can enter into the viable but non-culturable (VBNC) state induced by copper and resuscitate under appropriate conditions, which acted as a potential primary inoculum source in the field. In this study, bacterial cells of A. citrulli strain AAC00-1 was treated at 50℃, 55℃ and 4℃, respectively, which artificially stimulated the temperatures of hot water treatment and seed storage in production. The viability and culturability of A. citrulli cells were determined by flow cytometry (FCM) and agar plating, respectively. It showed that AAC00-1 can enter into VBNC state by unfavorable temperatures. All viable cells of AAC00-1 entered into the VBNC state after treated at 55℃ for 20 min and 30 min. The treated bacterial suspensions of AAC00-1 were injected into the cotyledon of watermelon seedlings for pathogenicity test. The inoculated plants showed no symptoms of BFB, and no culturable cells can be isolated from the inoculated seedlings. This indicates that VBNC cells of AAC00-1 can not be resuscitated in vivo in watermelon seedling. This study analyzed the ability of A. citrulli to enter into the VBNC state under unfavorable temperature conditions, and provided a theoretical basis for primary inoculum of BFB in the field, which has important significance for controlling of BFB in cucurbits production.

The colonization ability of Bacillus subtilis NCD-2 in the rhizosphere of ‘Jimian 11' is associated with L-proline in its root exudates. The effect of exogenous L-proline on the growth and biofilm formation of strain NCD-2 was determined in this study. The result showed that addition of L-proline to the MSgg medium did not affect on the growth of strain NCD-2, while the yield of biofilm was increased significantly and the degree of wrinkling and stereo-structure of biofilm were more obvious. The quantitative determination of biofilm formation was performed by crystal violet staining. The results showed that a higher concentration of L-proline at 1.250 to 10.000 mg·mL^-1 (but not 0.625 mg·mL^-1) could increase biofilm formation of strain NCD-2 compared to the mock control. The expression levels of tasA and epsA genes, two biofilm formation-related markers were analyzed by RT-qPCR after treatment with L-proline. Expression of the tasA gene was significantly increased by 2.53 to 9.98 times at different concentrations of L-proline. However, expression of the epsA gene was only increased in 0.625 mg·mL^-1 L-proline, and the expression level of the epsA gene was lower than that of tasA gene. These results suggest that supply of L-proline enhances biofilm formation of B. subtilis NCD-2 through boosting the production of extracellular protein TasA rather than increasing bacterial growth.

Blackleg and stem canker of oilseed rape are two diseases caused by two closely-related ascomycetous fungi, namely Leptosphaeria biglobosa (blackleg) and L. maculans (stem canker). In China, only L. biglobosa has been found in oilseed rape-producing areas, whereas L. maculans has not been reported in this crop. Therefore, L. maculans is officially treated as a quarantine plant pathogenic fungus in China. Both fungi formed pycnidia and conidia with similar morphology, and produced similar symptoms, which can be observed until maturity. It is difficult to identify both diseases quickly and accurately in the field. Therefore, it is imperative to develop a rapid, sensitive and high throughput method for detection and identification of the two fungi. This study was done to establish an easy-to-use detection system based on the loop-mediated isothermal amplification (LAMP) principle. Five specific primers for L. biglobosa and six specific primers for L. maculans were designed based on the ITS-rDNA sequences of the two fungi. Results showed that the established LAMP methods for detecting L. biglobosa and L. maculans are optimally performed at 65°C for 40 min and 50 min, respectively, with high detection specificity. Results of the sensitivity test indicated that threshold of the template DNA for both fungi reached down to the fg level, which was 100 and 1 000 times lower than that in the conventional PCR detection of L. biglobosa and L. maculans, respectively. Results also indicated that the LAMP for L. biglobosa had a positive detection with DNA extracted from diseased stems of oilseed rape showing typical blackleg symptoms, whereas the LAMP for L. maculans had a negative detection from these diseased stem samples. This result suggests that the blackleg disease on oilseed rape was caused by L. biglobosa, but not by L. maculans, and identification of the pathogen (L. biglobosa) in the diseased stems was confirmed by the convenient PCR with L. biglobosa- and L. maculans-specific primers. Therefore, the LAMP techniques developed in this study have a promising potential for detection of both pathogens in diseased tissues of oilseed rape.

The selective medium was based on WA medium, and added streptomycin sulfate, doxodine and carbendazim. The appropriate concentrations of three agents were 100 μg·mL^-1, 1 μg·mL^-1and 50 μg·mL^-1, respectively. The selective culture medium for isolation of Rhizoctonia cerealis from the stem of infected wheat has the characteristics of time saving, labor saving and high success rate of isolation, and can be operated directly in non-sterile conditions; selective medium can be used to detect the sclerotia in soil, and provide some reference for predicting the occurrence and prevalence of wheat sharp eyespot.

Xanthomonas campestris pv. campestris (Xcc) is an important pathogen that infects cruciferous plants. Extracellular cellulases are important for full virulence of Xcc. In Xcc 8004, nine genes were annotated to encode extracellular cellulases and one of them (XC0639) was the major cellulase gene. This work studied the effect of the bioinformatically predicted cellulase binding domain (CBD) of XC0639 on enzyme activity by constructing XC0639 CBD deletion mutant named D0639(CBD) and a complementary strain of D0639(CBD) named C0639(CBD). Interestingly, there was no significant difference among the cellulase activities produced by D0639(CBD), C0639(CBD), and the wild-type strain 8004, suggesting that the predicted CBD is not involved in the enzyme activity of XC0639. This contention was supported by the results showing that the cellulase activities exhibited by a mutant (D9) with deletion of all 9 annotated cellulase genes, D9 carrying an entire XC0639 in trans, D9 carrying an XC0639 lacking its CBD in trans, and the wild type strain 8004 were not significantly different. This finding provides a base for further study of the catalytic mechanism of the major cellulase XC0639 in Xcc.

This article reports the new taxonomy system (2019) approved by the International Committee on Taxonomy of Viruses (ICTV) in March 2020, which fully adopted a 15-rank taxonomic structure, namely: realm, subrealm, kingdom, subkingdom, phylum, subphylum, class, subclass, order, suborder, family, subfamily, genus, subgenus, species. Viruses whose hosts are plants include plant viruses and subviral infection factors (satellite virus, satellite and viroid). There are 1 608 species of plant viruses, belonging to 2 realms, 3 kingdoms, 8 phyla, 13 classes, 16 orders, 31 families, 8 subfamilies, 132 genera and 3 subgenera. Subviral infection factors include 33 species of viroids belonging to 2 families and 8 genera, 6 species of satellite viruses belonging to 4 genera, and 142 species of satellites belonging to 2 families, 2 subfamilies and 13 genera.

A new soil-borne disease of Salvia miltiorrhiza occurred in Anguo county of Hebei province in recent years. These pathogens were isolated and identified, and these pathogenicities of the isolates on S. miltiorrhiza were evaluated Fifteen fungal isolates were obtained from S. miltiorrhiza exhibiting typical symptoms of soil-borne disease, and these isolates were confirmed as the pathogens of S. miltiorrhiza by testing Koch's postulates. Under microscope, all isolates showed wheel branched conidiophores and could produce black radial microsclerotia on the media, thus, it was speculated that these isolates belong to the genus Verticillium. These isolates showed the highest internal transcribed spacer (ITS) sequence similarity with the species of V. dahliae, and were further identified as V. dahliae by PCR amplification with V. dahliae specific primers. Among them, 13 isolates were defoliated strains and 2 isolates were non-defoliated strains identified by PCR amplification with defoliated and non-defoliated specific primers. Taken together, the new soil-borne disease of S. miltiorrhiza was identified as verticillium wilt caused by V. dahliae. The artificial inoculation experiment revealed the obvious differences both in pathogenicities among V. dahliae isolates and in host resistances among the varieties of S. miltiorrhiza. To our knowledge, this is the first report of verticillium wilt of S. miltiorrhiza in China.

In order to determine whether the crumpling symptoms of Manihot esculenta Crantz were caused by the infection of begomoviruses. Leaf samples of M. esculenta plants with suspected viral symptoms were collec-ted from the fields in Honghe, Yunnan. Four complete DNA-A and six betasatellite genome sequences were obtained by PCR, cloned, and sequenced. Comparisons performed with the complete nucleotide sequence revealed the presence of two begomoviruses, tobacco curly shoot virus (TbCSV) and ageratum yellow vein China virus (AYVCNV) in the M. esculenta samples. The TbCSV isolate from M. esculenta was most closely related to TbCSV-YN2247 (KX290925) isolated from Yunnan (with 96.67% sequence identity). The AYVCNV isolates from M. esculenta were most closely related to AYVCNV-YN4326 (KU601622) (with 95.80% sequence identity). Two betasatellites, malvastrum yellow vein betasatellite (MaYVB) and ageratum yellow vein China virus betasatellite (AYVCNVB) were detected in the M. esculenta samples. The MaYVB-YN6332-12 isolates were most closely related to MaYVB-Y216 (KX290925) (with 96.4% sequence identity) , whereas, the AYVCNB-YN6338-17 isolates were most closely related to AYVCNB-Hn9 (KU601622) (with 90.4% sequence identity). This study represents M. esculenta plants as a new host of TbCSV and AYVCNV, also the first report of mono-partite begomoviruses and associated DNA betasatellites complex infecting M. esculenta.

To clarify the relationship between BcPDR1 gene and PKA coding genes of the cAMP signaling pathway in Botrytis cinerea. Real-time PCR was used to analyze the expression levels of BcPKA1, BcPKA2 and BcPKAR in BcPDR1 gene mutants and the expression levels of BcPDR1 in mutants of BcPKA1, BcPKA2 and BcPKAR. It was found that the expression levels of PKA coding genes BcPKA1, BcPKA2 and BcPKAR in BcPDR1 gene mutations were significantly higher than those in wild-type BC22 and the BcPDR1 complementing strain. The expression levels of BcPDR1 gene in RNAi mutations of BcPKA1, BcPKA2 and BcPKAR genes were significantly lower than those in wild-type BC22. We constructed the over-expression isolates of BcPKA1, BcPKA2 and BcPKAR in ΔBcpdr1 background, respectively. The colony morphology, sclerotium forming, mycelial morphology, growth rate, conidiation, and pathogenicity of the over-expression isolates significantly different from ΔBcpdr1, and closer to the wild-type BC22. These results suggest that the expression of BcPDR1 was closely related to BcPKA1, BcPKA2 and BcPKAR in B. cinerea. The BcPDR1 gene negatively regulates the expression of BcPKA1, BcPKA2 and BcPKAR, while BcPKA1, BcPKA2 and BcPKAR genes positively regulate the expression of the BcPDR1. In a summary, our findings would increase understanding the molecular mechanism of the BcPDR1 gene in growth, development and pathogenicity in B. cinerea.

In eukaryotes, gene expression can be regulated at transcription and post transcription levels to adapt to the biological processes of growth and development under different time and space environments. The PUF RNA-binding proteins belong to a family of posttranscriptional regulator with a conserved Pumilio homology domain (PUM-HD), and controls stability and translation of mRNA through specific binding with their target. In this study, CRISPR/CAS9-mediated genome editing technology was used to knockout PsM90, which encodes a PUF RNA-binding protein in Phytophthora sojae, and analyzed its biological functions. We found that the PsM90-knockout mutants produced significantly less oospores (only 32% of the wild-type P. sojae strain), the oospore wall was thinner and organelles in oospores failed to differentiate normally. The pathogenicity of zoospores to hypocotyls of soybean etiolated seedlings was also slightly decreased. In addition, the mutants did not show any significant differences in the growth of vegetative hyphae, sporangium development, zoospore release, and cyst germination. These results revealed PsM90 as an important gene involved in the sexual development of P. sojae, and provided insights for further studies on the molecular regulation mechanism of the oomycete-specific oospore development process.

Mitogen-activated Protein Kinase (MAPK)signaling pathway plays important roles in the regulation of physiological properties and virulence of Fusarium oxysporum f. sp. cubense race 4 (Foc4). To reveal the transcription mechanisms of transcription factor FoSwi6, the transcriptome profiles of Foc4 wild type strain and ΔFoSwi6 mutant were described using RNA sequencing technology. The results showed that there were 5 728 differentially expressed genes (DEGs) in ΔFoSwi6 mutant, of which 2 682 were up-regulated and 3 046 were down-regulated. The Gene Ontology (GO) classification showed that the DEGs were mainly involved in metabolic process, cellular process and single-organism process of biological process group, and catalytic activity and binding of molecular function group. The KEGG pathway analysis showed that the DEGs were mainly involved in ribosome, glyoxylate and dicarboxylate metabolism, biosynthesis of amino acids and 2-oxocarboxylic acid metabolism pathway. In silico analysis of the DEGs suggested that the 12 DEGs were related to conidiation, growth, oxidative stress, cell wall biosynthesis, sterol biosynthesis, nitrogen assimilation, toxin biosynthesis and virulence of Foc4. These data would provide theoretical basis for description of virulence mechanisms of Foc.

Grapevine berry inner necrosis virus (GINV) can cause obvious chlorotic mottling symptoms on grape leaves. Currently, the molecular mechanism of its pathogenesis, especially the interaction between GINV and host protein has not been reported. In this study, the cDNA library of GINV-infected Beta grapevine leaves was constructed, and GINV coat protein (CP) was used as the bait to screen the host factors in the library. The cDNA library constructed in this study contains 1.6×10⁷ clones with average size of inserted fragments being more than 1.0 kb, and the quality meets the requirements of cDNA library screening. A total of 55 candidate genes interacted with GINV CP in yeast. Sequencing and verification of positive clones showed that these genes encoded a total of 17 host proteins, including chloroplast photosynthesis-related proteins, ubiquitination-related proteins and defense-related proteins, and so on. This study provides a basis for further research on GINV pathogenicity and mechanism of interaction

Chinese wheat mosaic virus (CWMV) is one of the most economically important virus causing wheat soil-borne mosaic virus disease in China. The RNA 1 and RNA 2 fragments of CWMV were ligated to pCB301-Rz vector, producing recombinant plasmids pCWMV-RNA1 and pCWMV-RNA2, which could induce mosaic and deformation on Nicotiana benthamiana plants and yellow mosaic on wheat leaves. The genes for green fluorescent protein (GFP) was cloned to the 5'- end of the cysteine-rich gene of pCWMV-RNA2 and could produce green fluorescence in N. benthamiana plants. The codons for Asp (D₁₀₀₁) of RdRp in RNA1 and Glu (E₁₂₇) of CP in RNA2 were mutated to that for Ala. Compared with wild-type CWMV, the resulting mutants, CWMV-RdRp-D1001A, CWMV-CP-E127A and CWMV-RdRp-D1001A/CP-E127A, showed significantly lower virulence and less CP accumulation in N. benthamiana plants.

Black point disease, caused by the dominant pathogen Bipolaris sorokiniana, is a severe wheat grain disease. In order to analyze the genetic characters and detect the resistance loci for black point, the resistant genotype Shannong4143 and susceptible genotype Wanyuanbai1 and their recombinant inbred line (RIL) population at F₇ were planted at three experimental locations in 2018-2019 wheat growing season. Resistance identification was carried out using the method of spraying spore suspension of B. sorokiniana and bagging for moisture. The genetic analysis was carried out by mixed inheritance model of major genes plus polygenes. The extreme resistant and susceptible inbred lines were selected to build a mixed pool. Combining 660K SNP arrays, the resistant loci were detected by bulked segregate analysis (BSA). The results showed that: the resistance of wheat to black point caused by B. sorokiniana was in accordance with the "additive - epistatic inheritance model of 4 major genes", and heritability of the major gene was 0.88-0.95. Thirty-six loci were detected by BSA, which were located on 14 chromosomes of 1A, 2B (6), 2D (2), 3A, 3B (2), 3D, 4A (6), 4D, 5A (4), 5B (6), 5D (3), 7A, 7B and 7D, respectively. A total of 13, 15, and 8 loci were from the A, B, and D genomes, respectively. Among the 36 loci detected, 18 loci were new loci for resistance to black point. The results of this study laid a foundation for the fine mapping of resistance loci to black point and development of molecular markers in the breeding for resistant cultivars to black point.

Rice and Magnaporthe oryzae (M. oryzae) constitutes an ideal pathosystem for studying host-pathogen interaction in cereals crops. Luoping County, southwest China's Yunnan Province, is a main rice-producing area and one of the major hotspots of rice blast disease. In the field, the composition of M. oryzae population is complex and the information flow is strong. The isolation of single spore strains and the study of avirulence genes are the important basis for revealing the mechanism of virulence variation of M. oryzae and the comprehensive control of rice blast disease. In this study, we isolated 120 single spore strains from the field samples collec-ted from Luoping in 2017. The morphology, sporulation capacity, presence/absence (P/A) polymorphisms of seven avirulence (Avr) genes, and the relationship between avirulence gene variation and pathogenicity of these strains were studied systematically. Our results showed that the culture characteristics and sporulation ability were diversity among strains. The P/A frequency of seven Avr genes in 120 field strains was different, in which ACE1, Pwl2, and Avr-Pizt had the highest frequency of 100%, Avr-Pia had the lowest frequency of 5%, and Avr-Pita1, Avr-Pik, Avr-Pii were 99%, 99%, and 89%, respectively. Importantly, thirty-three M. oryzae isolates containing three Avr-Pik allele copies were identified for the first time. The rice monogenic lines were susceptible to the strains with the corresponding Avr genes complete deletion or functional variation. This result indicated that pathogen could shed off or modify its Avr gene to dodge past the resistance mechanism of the host plant thus making it susceptible. Our findings not only enrich the genetic resources of M. oryzae, lay an important foundation for studying the mechanisms of rice-M. oryzae interaction, but also provide valuable information for controlling rice blast disease.

Resistance detecting and monitoring are critical to manage the fungicides and pathogen populations. We collected 133 isolates of Septoria piricola Desm from pear from 11 counties in Hebei Province. The sensitivities to tebuconazole, iminoctadine trialbesilate, pyraclostrobin and cyproconazole were determined by the mycelial growth method in the laboratory. The correlation between tebuconazole and iminoctadine trialbesilate, pyraclostrobin and cyproconazole were further evaluated. The results showed that EC₅₀ values of tebuconazole for all isolates ranged from 0.0392 to 2.0522 μg·mL^-1 with the average at 0.5486±0.0012 μg·mL^-1. The ratio of lowest and highest EC₅₀ values of tebuconazole for the isolates from the different regions ranged from 2.7 to 7.4. All the isolates did not exhibit the significant difference on the sensitivity to tebuconazole with the average EC₅₀ values from 0.1639 to 0.9124 μg·mL^-1. In compare with the baseline sensitivities for mycelia growth of S. piricola to iminoctadine trialbesilate and pyraclostrobin with the average values of 0.2621±0.0014 μg·mL^-1 and 0.2750±0.0017 μg·mL^-1, respectively, there was no significant correlation of sensitivity between tebuconazole and iminoctadine trialbesilate, pyraclostrobin, while correlation to cyproconazole. Therefore, tebuconazole and iminoctadine trialbesilate, tebuconazole and pyraclostrobin can be mixed, and the best mix ratio is 1∶6 and 4∶1, respectively. The control efficacy, compared with the single fungicide application for pear brown spot caused by S. piricola, was significant differences between tebuconazole + iminoctadine trialbesilate (30+180 μg·mL^-1) and tebuconazole + pyraclostrobin (100+25 μg·mL^-1) that was 95.49%, 93.48% in 10 days after spraying application just once, and 86.51%, 87.40% or 69.02%, 70.33% in 10 or 50 days after the third applications, respectively. The combination of the fungicides displayed a significantly better effectiveness and duration of protection than with their use only.

Coniothyrium minitans is a mycoparasite of the plant pathogenic fungus Sclerotinia sclerotiorum. It can parasitize sclerotia of S. sclerotiorum, reducing apothecial production and disease incidence. However, in the natural soils, the effect of sclerosphere microbes on infection of S. sclerotiorum sclerotia by C. minitans is poorly understood. This study was focused on the ecology of the sclerosphere microorganism community of S. sclerotiorum sclerotia, using isolation and molecular techniques. We analyzed the sclerosphere microorganism community of S. sclerotiorum in different treatments, including sampling time and burial depth of sclerotia. Results showed that the populations of bacteria and fungi in sclerosphere soil were significantly higher than those in sclerotia-free soils. Two hundred and fifty-three bacteria and one hundred and eighty fungi were isolated from sclerosphere soil. Base on bacterial 16S rDNA and fungal internal transcribed spacer of ribosomal DNA sequences, we identified Pseudomonas and Bacillus as the dominant bacterial populations in the sclerosphere. Whereas, Penicillium was the dominant fungus population in the sclerosphere. In the dual culture in vitro assays, there were 25 and 22 sclerosphere bacterial strains with antagonistic activity against S. sclerotiorum and C. minitans, respectively. Furthermore, sclerotial infection assay showed that seven bacterial strains significantly suppressed infection of sclerotia by C. minitans in vitro. These results suggest that in the field, sclerosphere bacteria might play an important role in suppression of the infection of sclerotia by C. minitans.

In this study, bioinformatics methods were used to conduct phylogenetic, conservative domain, tissue-specific, and expression regularity analysis of BTB family genes during resistance to biological and abiotic stresses in Zea mays. It was found that all 70 BTB family proteins in Z. mays contain a conserved BTB domain except AC206223.3, which can be divided into 11 subfamilies. The BTB family genes of maize had obvious tissue expression characteristics, and the expression levels show obvious differences under high temperature, low temperature, ultraviolet, high salt, drought stress, and in the process of Fusarium verticillioide infection. The expression levels of BTB-TAZ subfamily genes were significantly changed after SA, JA and ET treatment. These results showed that BTB family genes play an important role in resisting biological and abiotic stresses. It would lay the foundation for elucidating the function and mechanism of BTB family genes in Z. mays.

In order to identify the pathogenic fungi of Atractylodes macrocephala root rot in Baoding, Hebei Province, eighty strains of Fusarium species were isolated from 120 A. macrocephala plants with root rot symptoms with the isolation frequency at 66.7%. Fusarium species were identified by molecular techniques with specific primers, TEF-1α gene sequencing together with morphological characteristics. Thirty-nine F. solani strains were identified with the highest percentage at 48.8%, and 32 F. oxysporum strains were isolated with the percentage at 40.0%. Seven F. equiseti and two F. brachygibbosum strains accounted for 8.8% and 2.5%, respectively. The pathogenicity test on A. macrocephala showed that F. solani and F. oxysporum caused the typical symptoms of root rot. Koch's postulates were fulfilled following re-isolation and identification of the isolates of F. solani and F. oxysporum. Taken together, the results showed that F. solani and F. oxysporum were the pathogens of A. macrocephala root rot in Baoding, Hebei Province.

In 2016-2019, sixty new varieties and 31 main cultivated varieties were evaluated for their resistance to sugarcane brown stripe disease under field condition. The results showed that 32 (53.33%) of 60 new sugarcane varieties were highly to moderately resistant, and 28 (46.67%) were susceptible to highly susceptible. Twenty-one (67.74%) of 31 main cultivated sugarcane varieties were highly to moderately resistant, and ten (32.26%) were susceptible to highly susceptible. This work may facilitate brown stripe resistance breeding and provide new elite resistant varieties for effectively control of sugarcane brown stripe.

Potato black scurf caused by the soil-borne pathogenic fungus Rhizoctonia solani is one of the main causes of potato continuous cropping obstacles. A biocontrol bacterium Paenibacillus polymyxa ZF129 had signi-ficant inhibitory effect on Rhizoctonia solani that the inhibition rate on mycelial growth was 55.29% and control efficacy in greenhouse was 84.76%. Meanwhile, ZF129 exhibited significant and broad inhibitory spectrum against six pathogenic bacteria and eight pathogenic fungi.

Early studies have revealed the uneven distribution of Candidatus Liberibacter asiaticus (CLas) in Huanglongbing (HLB)-affected citrus trees. Bacteria of CLas were found to be enriched in the fruit, especially in the pith. In this study, we further investigated the dynamics of CLas population in excised citrus fruits. The CLas-infected Shatangju fruits were collected and kept for 4 weeks under constant temperature and moisture. The concentrations of CLas in the pith of fruits were detected weekly, and the fruits on trees were detected at the same time as the control. Our results showed that the total DNA, citrus DNA and CLas DNA from the excised fruits all decreased during the storage, but the CLas concentration in the pith of excised fruit increased firstly, and then decreased with time. The CLas concentrations in the pith of diseased fruit at 14 d and 28 d after picking were higher than that of the first day and reached the highest at 14 d. Comparing with the fruits on the tree, CLas concentrations in the pith of excised fruits were also significantly higher at the 14 d and 28 d. With the degrading of total DNA in fruit pith, the proportion of CLas DNA was increased, which could be served as a suitable material for the DNA extraction and CLas genome sequencing.

Rice bakanae disease is an important disease responsible for serious rice production loss in China. The current study aimed to reveal the species of seedborne rice bakanae pathogens in main rice planting regions. A total of 111 strains of putative Fusarium spp. were isolated from the 66 rice seed samples via the washing method and plate culture method. Based on morphological features, 24 representative strains were selected and further analyzed based on the translation elongation factor 1α gene (TEF-1α) sequences and morphological characteristics. Among the 111 isolates, 45.94% and 52.25% were proved to be Fusarium fujikuroi and F. proliferatum, respectively. There were 22 of the 66 seed samples carrying bakanae pathogens. F. fujikuroi was the most frequent species (18.18%), followed by F. proliferatum (16.67%) and F. andiyazi (1.52%). Hydroponic method was used to nurse the 22 rice seed samples, and the bakanae disease was measured in the seedlings. The pathogens both from interior and exterior of rice seeds were found to be able to cause bakanae disease. The pathogens in the interior, exterior, or both interior and exterior of rice seeds induced diseases at the frequency of 3.73%, 6.5% and 15.56%, respectively, at the seedling stage. In addition, F. fujikuroi-infected seeds induced diseases in the seedlings at higher frequency than F. proliferatum did.

In 2018, a new disease with necrotic symptoms affecting the leaves of Ziziphus jujuba Mill. cv. Dongzao was observed in Qingyuan city, Guangdong province, China. The disease incidence reached 17% in fields. In order to identify the pathogen of Ziziphus jujuba Mill. cv. Dongzao leaf necrotic disease in Qingyuan city, Guangdong province, two isolates of the pathogen were obtained by diseased tissue isolation method. Two isolates were identified by pathogenicity test, physiological and biochemical characteristics and molecular biolo-gical identification. The results of pathogenicity test showed that Ziziphus jujuba Mill. cv. Dongzao plants inoculated with the isolates of the bacterium could infect Ziziphus jujuba Mill. cv. Dongzao and caused the same symptoms as the diseased Ziziphus jujuba Mill. cv. Dongzao plants in the fields. The physiological and biochemical characteristics of QYZ1 and QYZ2 were same to that of Pantoea agglomerans ATCC27155 except for melibiose utilization. The results of phylogenetic analysis showed that 16S rDNA gene sequences of two isolates share high identify of 99% with P. agglomerans TH81 (CP031649), P. agglomerans C410P1 (CP016889), P. vagans FDAARGOS_160 (CP014129) and P. vagans C9-1 (CP002206). The results of phylogenetic multilocus sequence analysis showed that QYZ1 and QYZ2 were clustered into a group with P. agglomerans TH81. The pathogen of Ziziphus jujuba Mill cv. Dongzao leaf necrotic disease was identified as P. agglomerans based on the results of physiological and biochemical characteristics and molecular biological identification. This is the first recorded report of necrotic disease caused by P. agglomerans on Ziziphus jujuba Mill. cv. Dongzao in Guangdong province, China.

Located in the center of the Yangtze River in the southeastern coast of China, the South Jiangsu area has fertile soil and plentiful rainfall, and vegetable production is an important industry in this region. Virus diseases on the vegetables in this area are frequent and serious. However, little has been done on the systemic investigation of species and trends of vegetable viruses in the South Jiangsu area. This study adopts the method of viral nucleic acid detection. From May 2018 to December 2019, 232 suspected virus-infected samples were collected from five main vegetable crops in Nanjing, Wuxi, Zhenjiang, Suzhou and Changzhou. Among a total of 12 detected virus species, seven were detected in Nanjing, four in Wuxi, two in Zhenjiang, six in Suzhou, and seven in Changzhou. Eleven viruses were found in Solanaceae vegetables, nine in Cucurbitaceae vegetables and seven in Leguminous vegetables. Among them, pepper mild mottle virus (PMMoV), tomato chlorosis virus (ToCV), tomato mosaic virus (ToMV), cucurbit chlorotic yellows virus (CCYV), tomato yellow leaf curl virus (TYLCV) and pepper vein yellow virus (PeVYV) were detected most frequently. In addition, the Southern tomato virus (STV), tobacco vein distorting virus (TVDV), tobacco mild green mosaic virus (TMGMV) and PeVYV identified in this study were the first time reported in Jiangsu province. At the same time, multiple virus infections were prevalent in vegetable crops, and up to 5 virus infections have been found in the tested samples, among which PMMoV and other viruses have a high frequency and a variety of mixed infections. The findings of this study provide an important basis for understanding the types, distribution and occurrence trends of major vegetable viral diseases in southern Jiangsu and for the development of effective surveillance and control measures.

Downy mildew (Peronospora farinosa f. sp. chenopodii) is one of the common diseases affecting the growth of quinoa. In order to explore the effect of downy mildew-infection on the metabolism of quinoa leaves, the metabolite profiles of quinoa leaves in the control group and the treatment group were analyzed by using the metabonomics method. A total of 126 metabolites were identified, including 43 organic acids, 17 amino acids, 27 sugars and 39 other substances. The ten major metabolites causing the significant variations between the treatment group and control group were found by using PCA and T-test analyses. Among them, N-heptadecane, succinic semialdehyde, isoleucine, and serine increased significantly in the treatment group (P<0.01), while malic acid, citric acid, sucrose, tyramine, stigmasterol and aspartic acid decreased significantly (P<0.01). The infection of downy mildew caused the abnormal changes of metabolites in several metabolic pathways, including glycoside metabolism, nucleic acid metabolism, amino acid metabolism, tricarboxylic acid cycle, and ɤ- aminobutyric acid.

SPX-domain-containing proteins (SPXs) play an important role in inorganic phosphate (Pi) sen-sing, signaling, and transport in eukaryotes. Related studies on SPXs of maize have not been reported. In this study, 15 SPXs were obtained by searching the database of Pfam (http://pfam.sanger.ac.uk/) using bioinformatics methods, and were divided into SPX-EXS, SPX, SPX-Zinc finger, and SPX-MFS subfamilies by phylogenetic analysis and conserved domain analysis. The tissue expression specificity analysis of ZmSPXs found that different genes had significantly different expression levels in different developmental stages and different tissues, showing obvious tissue specificity. ZmSPXs family genes have specific high and low expression genes under heat, cold, salt, UV, and drought abiotic stress. The gene expression levels of ZmSPXs family genes in Fusarium verticillioides infection in different periods are obviously different. RT-qPCR was used to analyze the expression levels of ZmSPXs under phosphorus stress, and the results showed that most of the genes were highly expressed under phosphorus stress. This study clarified the number of genes, phylogenetic relationships, conserved domains, tissue expression characteristics and expression patterns under biological and abiotic stresses of SPXs in maize, which could provide basic data for clarifying the function and mechanism of SPXs.

Cantaloupe is one of the important cash crops in Xinjiang province. Cucurbit aphid-borne yellows virus (CABYV) is an important worldwide cucurbit xanthosis virus, which is widely distributed in the main growing areas of cantaloupe of Xinjiang. CABYV is serious threaten to the quality and yield of cantaloupe. In this study, a total of 120 leaf samples were randomly selected from Akesu city of Xinjiang province, among which 38 samples were positive to CABYV by RT-PCR test. A new CABYV isolate was isolated, sequenced and cloned from these positive samples. The identity and phylogenetic were analyzed based on the CP gene nucleotide sequences and amino acid sequences of 23 isolates. The results showed that the 23 CABYV isolates were divided into 2 distinct evolutionary populations according to regional differences. Further analysis of genetic diversity in different CABYV populations revealed there are significant genetic differences between CABYV host and geographic populations. Selection pressure analysis and neutrality test indicated that CABYV population may undergone population expansion or background selection recently, and negative selection may be one of the reasons for genetic variation of CABYV. The knowledge on occurrence, damage, genetic structure and evolutionary mechanism of CABYV presented in this study may be useful in the design of sustainable management strategies for controlling CABYV.

Pine wilt disease is a worldwide quarantine disease that seriously threatens forest resources such as pine forests and ecological security. The Feminization (fem) gene family is involved in sex determination and differentiation in the model Caenorhabditis elegans. In this research, the pine wood nematode fem-1 gene was cloned and its expression characteristics and functions were studied. The full-length cDNA of fem-1 is 856 bp long,containing a 681 bp open reading frame and encodes a 226-amino-acids protein, which shares same clade with that of C. elegans in phylogenetic analysis. The results of real-time quantitative PCR(RT-qPCR)and in situ hybridization showed that fem-1 gene was expressed in all developmental stages of pine wood nematodes, with the highest expression level in the second juvenile stage. The fem-1 gene was expressed in the rear of the body from the egg stage to the third juvenile stage. In the later stage of development, it was mostly expressed in subcutaneous cells and rarely occurred in the intestinal tract, gonadal and other parts. After the fem-1 gene was silenced by RNA interference technology, the female-male ratio of the nematode population decreased, indicating that the fem-1 gene may be involved in the sex differentiation and determination of pine wood nematodes. This study is helpful for understanding the expression characteristics and function of fem-1 gene in B. xylophilus, and provides a theoretical basis for further research on the reproductive development of pine wood nematodes.