Characterization of the fludioxonil and phenamacril dual resistant mutants of Fusarium graminearum.

Fusarium graminearum is an important fungal pathogen causing Fusarium head blight (FHB) in wheat and other cereal crops worldwide. Due to lack of resistant wheat cultivars, FHB control mainly relies on application of chemical fungicides. Both fludioxonil (a phenylpyrrole compound) and phenamacril (a cyanoacrylate fungicide) have been registered for controlling FHB in China, however, fludioxonil-resistant isolates of F. graminearum have been detected in field. To evaluate the potential risk of dual resistance of F. graminearum to both compounds, fludioxonil and phenamacril dual resistant (DR) mutants of F. graminearum were obtained via fungicide domestication in laboratory. Result showed that resistance of the DR mutants to both fludioxonil and phenamacril were genetically stable after sub-cultured for ten generations or stored at 4 °C for 30 days on fungicide-free PDA. Cross-resistance assay showed that the DR mutants remain sensitive to other groups of fungicides, including carbendazim, tebuconazole, pydiflumetofen, and fluazinam. In addition, the DR mutants exhibited defects in mycelia growth, conidiation, mycotoxin deoxynivalenol (DON) production, and virulence Moreover, the DR mutants displayed increased sensitivity to osmotic stress. Sequencing results showed that amino acid point mutations S217L/T in the myosin I protein is responsible for phenamacril resistance in the DR mutants. Our results indicate that mutations leading to fludioxonil and phenamacril dual resistance could result in fitness cost for F. graminearum. Our results also suggest that the potential risk of F. graminearum developing resistance to both fludioxonil and phenamacril in field could be rather low, which provides scientific guidance in controlling FHB with fludioxonil and phenamacril.

Overproduction of mycotoxin biosynthetic enzymes triggers Fusarium toxisome-shaped structure formation via endoplasmic reticulum remodeling.

Mycotoxin deoxynivalenol (DON) produced by the Fusarium graminearum complex is highly toxic to animal and human health. During DON synthesis, the endoplasmic reticulum (ER) of F. graminearum is intensively reorganized, from thin reticular structure to thickened spherical and crescent structure, which was referred to as "DON toxisome". However, the underlying mechanism of how the ER is reorganized into toxisome remains unknown. In this study, we discovered that overproduction of ER-localized DON biosynthetic enzyme Tri4 or Tri1, or intrinsic ER-resident membrane proteins FgHmr1 and FgCnx was sufficient to induce toxisome-shaped structure (TSS) formation under non-toxin-inducing conditions. Moreover, heterologous overexpression of Tri1 and Tri4 proteins in non-DON-producing fungi F. oxysporum f. sp. lycopersici and F. fujikuroi also led to TSS formation. In addition, we found that the high osmolarity glycerol (HOG), but not the unfolded protein response (UPR) signaling pathway was involved in the assembly of ER into TSS. By using toxisome as a biomarker, we screened and identified a novel chemical which exhibited high inhibitory activity against toxisome formation and DON biosynthesis, and inhibited Fusarium growth species-specifically. Taken together, this study demonstrated that the essence of ER remodeling into toxisome structure is a response to the overproduction of ER-localized DON biosynthetic enzymes, providing a novel pathway for management of mycotoxin contamination.

The transcription factor FgStuA regulates virulence and mycotoxin biosynthesis via recruiting the SAGA complex in Fusarium graminearum.

The conserved Spt-Ada-Gcn5-Acetyltransferase (SAGA) complex controls eukaryotic transcription by modifying acetylation of histones. However, the mechanisms for this complex in regulating the transcription of target-specific genes remain largely unknown in phytopathogenic fungi. A filamentous fungal-specific transcription factor FgStuA was identified to interact with the SAGA complex physically. The coordinative mechanisms of FgStuA with the SAGA complex in regulating secondary metabolism and virulence were investigated in Fusarium graminearum with genetic, biochemical and molecular techniques. The transcription factor FgStuA binds to a 7-bp cis-element (BVTGCAK) of its target gene promoter. Under mycotoxin deoxynivalenol (DON) induction conditions, FgStuA recruits the SAGA complex into the promoter of TRI6, a core regulator of the DON biosynthesis gene cluster, leading to enhanced transcription of TRI6. During this process, we found that FgStuA is subject to acetylation by the SAGA complex, and acetylation of FgStuA plays a critical role for its enrichment in the TRI6 promoter. In addition, FgStuA together with the SAGA complex modulates fungal virulence. This study uncovers a novel regulatory mechanism of a transcription factor, which recruits and interacts with the SAGA complex to activate specific gene expression in pathogenic fungi.

The posttranscriptional regulator CsrA affects multidrug resistance and biocontrol activity in Lysobacter enzymogenes.

AIMS: The posttranscriptional regulator CsrA regulates many cellular processes, including stress responses in diverse bacteria. However, the role of CsrA in multidrug resistance (MDR) and biocontrol activity in Lysobacter enzymogenes strain C3 (LeC3) remains unknown. METHODS AND RESULTS: In this study, we demonstrated that deletion of the csrA gene resulted in the initial slow growth of LeC3 and reduced its resistance to multiple antibiotics, including nalidixic acid (NAL), rifampicin (RIF), kanamycin (Km), and nitrofurantoin (NIT). Loss of the csrA gene also reduced its ability in inhibiting hypha growth of Sclerotium sclerotiorum and influenced its extracellular cellulase and protease activities. Two putative small noncoding regulatory RNAs (sRNAs), referred to as csrB and csrC, were also revealed in the genome of LeC3. Double deletion of csrB and csrC in LeC3 led to increased resistance to NAL, RIF, Km, and NIT. However, no difference was observed between LeC3 and the csrB/csrC double mutant in their suppression of S. sclerotiorum hypha growth and production of extracellular enzymes. CONCLUSION: These results suggest that CsrA in LeC3 not only conferred its intrinsic MDR, but also contributed to its biocontrol activity.

Examination of Large Chromosomal Inversions in the Genome of Erwinia amylovora Strains Reveals Worldwide Distribution and North America-Specific Types.

Erwinia amylovora is a relatively homogeneous species with low genetic diversity at the nucleotide level. However, phenotypic differences and genomic structural variations among E. amylovora strains have been documented. In this study, we identified 10 large chromosomal inversion (LCI) types in the Spiraeoideae-infecting (SI) E. amylovora strains by combining whole genome sequencing and PCR-based molecular markers. It was found that LCIs were mainly caused by homologous recombination events among seven rRNA operons (rrns) in SI E. amylovora strains. Although ribotyping results identified inter- and intra-variations in the internal transcribed spacer (ITS1 and ITS2) regions among rrns, LCIs tend to occur between rrns transcribed in the opposite directions and with the same tRNA content (tRNA-Glu or tRNA-Ile/Ala) in ITS1. Based on the LCI types, physical/estimated replichore imbalance (PRI/ERI) was examined and calculated. Among the 117 SI strains evaluated, the LCI types of Ea1189, CFBP1430, and Ea273 were the most common, with ERI values at 1.31, 7.87, and 4.47°, respectively. These three LCI types had worldwide distribution, whereas the remaining seven LCI types were restricted to North America (or certain regions of the United States). Our results indicated ongoing chromosomal recombination events in the SI E. amylovora population and showed that LCI events are mostly symmetrical, keeping the ERI less than 15°. These findings provide initial evidence about the prevalence of certain LCI types in E. amylovora strains, how LCI occurs, and its potential evolutionary advantage and history, which might help track the movement of the pathogen.

The sphinganine C4-hydroxylase FgSur2 regulates sensitivity to azole antifungal agents and virulence of Fusarium graminearum.

Lipid rafts consisting of ergosterol and sphingolipids in the lipid membrane of cells play important roles in various cellular processes. However, the functions of sphingolipids and their synthetic genes in phytopathogenic fungi have not been well understood yet. In this study, we conducted genome-wide searches and carried out systematic gene deletion analysis of the sphingolipid synthesis pathway in Fusarium graminearum, a causal agent of Fusarium head blight of wheat and other cereal crops worldwide. Mycelial growth assays showed that deletion of FgBAR1, FgLAC1, FgSUR2 or FgSCS7 resulted in markedly reduced hyphal growth. Fungicide sensitivity tests showed that the sphinganine C4-hydroxylase gene FgSUR2 deletion mutant (ΔFgSUR2) exhibited significantly increased susceptibility to azole fungicides. In addition, this mutant displayed a remarkable increase in cell membrane permeability. Importantly, ΔFgSUR2 was defective in deoxynivalenol (DON) toxisome formation, leading to dramatically decreased DON biosynthesis. Moreover, the deletion of FgSUR2 resulted in dramatically decreased virulence of the pathogen on host plants. Taken together, these results indicate that FgSUR2 plays an important role in regulating the susceptibility to azoles and virulence of F. graminearum.

The calcium-calcineurin and high-osmolarity glycerol pathways co-regulate tebuconazole sensitivity and pathogenicity in Fusarium graminearum.

The calcium-calcineurin and high-osmolarity glycerol (HOG) pathways play crucial roles in fungal development, pathogenicity, and in responses to various environmental stresses. However, interaction of these pathways in regulating fungicide sensitivity remains largely unknown in phytopathogenic fungi. In this study, we investigated the function of the calcium-calcineurin signalling pathway in Fusarium graminearum, the causal agent of Fusarium head blight. Inhibitors of Ca2+ and calcineurin enhanced antifungal activity of tebuconazole (an azole fungicide) against F. graminearum. Deletion of the putative downstream transcription factor FgCrz1 resulted in significantly increased sensitivity of F. graminearum to tebuconazole. FgCrz1-GFP was translocated to the nucleus upon tebuconazole treatment in a calcineurin-dependent manner. In addition, deletion of FgCrz1 increased the phosphorylation of FgHog1 in response to tebuconazole. Moreover, the calcium-calcineurin and HOG signalling pathways exhibited synergistic effect in regulating pathogenicity and sensitivity of F. graminearum to tebuconazole and multiple other stresses. RNA-seq data revealed that FgCrz1 regulated expression of a set of non-CYP51 genes that are associated with tebuconazole sensitivity, including multidrug transporters, membrane lipid biosynthesis and metabolism, and cell wall organization. Our findings demonstrate that the calcium-calcineurin and HOG pathways act coordinately to orchestrate tebuconazole sensitivity and pathogenicity in F. graminearum, which may provide novel insights in management of Fusarium disease.

Fungicide Resistance: Progress in Understanding Mechanism, Monitoring, and Management.

Fungicide treatments are often essential for maintaining healthy crops and to achieve reliable and high-quality yields. However, continued use of fungicides with the same modes of action can lead to development of fungicide resistance, which has emerged in various plant pathogens and is a serious threat to effective crop protection. Exploration of resistance mechanisms is critical for resistance monitoring and management. This brief review summarizes advances during the past five decades in understanding the molecular resistance mechanisms of plant pathogenic fungi and oomycetes to major classes of fungicides, including benzimidazoles, myosin inhibitors, sterol demethylation inhibitors, quinone outside inhibitors, succinate dehydrogenase inhibitors, anilinopyrimidines, carboxylic acid amides, and oxysterol-binding protein homolog inhibitors. Based on known resistance mechanisms, PCR- and loop-mediated isothermal amplification-based approaches have been developed to allow high-throughput monitoring and early/rapid detection of emerging resistance. Classical principles in fungicide resistance management are also summarized, including using different modes of action of fungicides, limiting the number of applications of the chemicals with site-specific modes of action, and avoidance of their eradicant use. Future studies on novel strategies of disease management, including development of epigenetics- and RNA-based fungicides, will provide valuable knowledge for management of fungicide resistance.

RpoN Regulon in Erwinia amylovora Revealed by Transcriptional Profiling and In Silico Binding Site Analysis.

Erwinia amylovora causes a devastating fire blight disease in apples and pears. One of the main virulence determinants in E. amylovora is the hypersensitive response (HR) and pathogenicity (hrp)-type III secretion system (T3SS), which is activated by the RpoN-HrpL sigma factor cascade. However, the RpoN regulon in E. amylovora has not been investigated. In this study, we determined the RpoN regulon in E. amylovora by combining RNA-seq transcriptomic analysis with in silico binding site analysis. RNA-seq revealed that 262 genes, approximately 7.5% genes in the genome of E. amylovora, were differentially transcribed in the rpoN mutant as compared with the wild type. Specifically, genes associated with virulence, motility, nitrogen assimilation, the PspF system, stress response, and arginine biosynthesis are positively regulated by RpoN, whereas genes associated with biosynthesis of amino acids and sorbitol transport are negatively regulated by RpoN. In silico binding site analysis identified 46 potential target genes with a putative RpoN binding site, and the upstream sequences of six, three, and three genes also contain putative GlnG, PspF, and YfhA binding sites, respectively. Overall, RpoN directly regulates genes associated with virulence, nitrogen assimilation, the PspF system, motility and the YfhA/YfhK two-component regulatory system.

Spectinomycin resistance in Lysobacter enzymogenes is due to its rRNA target but also relies on cell-wall recycling and purine biosynthesis.

Resistance to spectinomycin emerged after widely used for treatment of gonorrhea. Previous studies revealed that Lysobacter enzymogenes strain C3 (LeC3) exhibited elevated level of intrinsic resistance to spectinomycin. In this study, we screened a Tn5 transposon mutant library of LeC3 to elucidate the underlying molecular mechanisms of spectinomycin resistance. Insertion sites in 15 out of 19 mutants recovered with decreased spectinomycin resistance were located on two ribosomal RNA operons at different loci, indicating the pivotal role of ribosomal RNAs in conferring spectinomycin resistance in L. enzymogenes. The other mutants harbored mutations in the tuf, rpoD, mltB, and purB genes. Among them, the tuf and rpoD genes, respectively, encode a translation elongation factor Tu and an RNA polymerase primary sigma factor. They both contribute to protein biosynthesis, where ribosomal RNAs play essential roles. The mltB gene, whose product is involved in cell-wall recycling, was not only associated with resistance against spectinomycin, but also conferred resistance to osmotic stress and ampicillin. In addition, mutation of the purB gene, for which its product is involved in the biosynthesis of inosine and adenosine monophosphates, led to decreased spectinomycin resistance. Addition of exogenous adenine at lower concentration in medium restored the growth deficiency in the purB mutant and increased bacterial resistance to spectinomycin. These results suggest that while cell-wall recycling and purine biosynthesis might contribute to spectinomycin resistance, target rRNAs play critical role in spectinomycin resistance in L. enzymogenes.

Thiamine Is Required for Virulence and Survival of Pseudomonas syringae pv. tomato DC3000 on Tomatoes.

Pseudomonas syringae pv. tomato DC3000 (PstDC3000) is an important plant pathogen that infects tomatoes and Arabidopsis. Thiamine and its derivative thiamine pyrophosphate (TPP) are cofactors that play an important role in the growth and survival of many bacterial microorganisms. However, the role of thiamine-related genes has not been determined in PstDC3000. Hence, to investigate the role of TPP in growth, resistance to stresses, and virulence of PstDC3000, double and quadruple mutants of thiamine biosynthesis-related genes (thiD/E, thiS/G, and thiD/E/S/G deletion mutants) as well as a single mutant of a lipoprotein-related gene (apbE) were constructed. Our results showed that growth of the thiD/E, thiS/G, and thiD/E/S/G mutants in the mannitol-glutamate (MG) medium was significantly lower than that of the wild type (WT) and their growth could be restored to the WT level with the addition of exogenous thiamine, whereas mutation of the apbE gene did not affect its growth in vitro. While tolerance to acid, osmotic, and oxidative stresses for the double mutants was similar to the WT, tolerance to stresses for the apbE mutant was reduced as compared to the WT. In addition, all four mutants exhibited reduced virulence and growth in tomatoes. However, when the double and quadruple mutants were inoculated with exogenous thiamine, the virulence and growth rate of these mutants were restored to the WT level. These results indicated that the thiD/E, thiS/G, and thiD/E/S/G mutants exhibiting growth deficiency in planta are probably due to a lack of thiamine biosynthesis, thus reducing colonization in tomatoes. On the other hand, it is possible that the apbE mutant exhibited reduced stress tolerances, thus resulting in reduced colonization. Overall, our findings suggest that the thiamine biosynthetic (TBS) pathway plays an important role in the colonization and infection of PstDC3000. Therefore, the thiamine biosynthetic pathway could be used as the target to develop new control measures for a bacterial spot in tomatoes.

Fusarium graminearum FgSdhC1 point mutation A78V confers resistance to the succinate dehydrogenase inhibitor pydiflumetofen.

BACKGROUND: Fusarium head blight (FHB) caused by Fusarium graminearum complex (Fg) is a devastating disease of cereal crops worldwide. The succinate dehydrogenase inhibitor, pydiflumetofen, was registered for management of FHB in China in 2019. Previously, laboratory-induced pydiflumetofen-resistant (PyR) mutants of Fg have been characterized. However, resistance situation of Fg to pydiflumetofen in the field remains largely unknown. RESULTS: After screening 6468 isolates of Fg from various regions of China, six PyR isolates were identified. All six resistant isolates exhibited no fitness penalties based on mycelial growth, conidiation and virulence. However, no cross-resistance between pydiflumetofen and azoxystrobin, tebuconazole or fludioxonil in Fg was detected. Genome-sequencing revealed that all six PyR isolates contained a point mutation A78V in FgSdhC1 (FgSdhC1A78V ). Genetic replacement assay further confirmed that FgSdhC1A78V conferred resistance of Fg to pydiflumetofen. Based on this, a mismatch allele-specific polymerase chain reaction was developed for rapidly detecting the PyR isolates containing the FgSdhC1A78V mutation in Fg. CONCLUSION: This is the first time that resistance of Fg to pydiflumetofen in the field was reported and point mutation FgSdhC1A78V conferring resistance of Fg to pydiflumetofen was confirmed. This study provides critical information for monitoring and managing pydiflumetofen resistance in Fg.

Fusarium BP1 is a reader of H3K27 methylation.

Histone H3 lysine 27 methylation catalyzed by polycomb repressive complex 2 (PRC2) is conserved from fungi to humans and represses gene transcription. However, the mechanism for recognition of methylated H3K27 remains unclear, especially in fungi. Here, we found that the bromo-adjacent homology (BAH)-plant homeodomain (PHD) domain containing protein BAH-PHD protein 1 (BP1) is a reader of H3K27 methylation in the cereal fungal pathogen Fusarium graminearum. BP1 interacts with the core PRC2 component Suz12 and directly binds methylated H3K27. BP1 is distributed in a subset of genomic regions marked by H3K27me3 and co-represses gene transcription. The BP1 deletion mutant shows identical phenotypes on mycelial growth and virulence, as well as similar expression profiles of secondary metabolite genes to the strain lacking the H3K27 methyltransferase Kmt6. More importantly, BP1 can directly bind DNA through its PHD finger, which might increase nucleosome residence and subsequently reinforce transcriptional repression in H3K27me3-marked target regions. A phylogenetic analysis showed that BP1 orthologs are mainly conserved in fungi. Overall, our findings provide novel insights into the mechanism by which PRC2 mediates gene repression in fungi, which is distinct from the PRC1-PRC2 system in plants and mammals.

The NDR kinase-MOB complex FgCot1-Mob2 regulates polarity and lipid metabolism in Fusarium graminearum.

Members of the NDR (nuclear Dbf2-related) protein-kinase family are essential for cell differentiation and polarized morphogenesis. However, their functions in plant pathogenic fungi are not well understood. Here, we characterized the NDR kinase FgCot1 and its activator FgMob2 in Fusarium graminearum, a major pathogen causing Fusarium head blight (FHB) in wheat. FgCot1 and FgMob2 formed a NDR kinase-MOB protein complex. Localization assays using FgCot1-GFP or FgMob2-RFP constructs showed diverse subcellular localizations, including cytoplasm, septum, nucleus and hyphal tip. ΔFgcot1 and ΔFgmob2 exhibited serious defects in hyphal growth, polarity, fungal development and cell wall integrity as well as reduced virulence in planta. In contrast, lipid droplet accumulation was significantly increased in these two mutants. Phosphorylation of FgCot1 at two highly conserved residues (S462 and T630) as well as five new sites synergistically contributed its role in various cellular processes. In addition, non-synonymous mutations in two MAPK (mitogen-activated protein kinase) proteins, FgSte11 and FgGpmk1, partially rescued the growth defect of ΔFgmob2, indicating a functional link between the FgCot1-Mob2 complex and the FgGpmk1 signalling pathway in regulating filamentous fungal growth. These results indicated that the FgCot1-Mob2 complex is critical for polarity, fungal development, cell wall organization, lipid metabolism and virulence in F. graminearum.

Protein acetylation and deacetylation in plant-pathogen interactions.

Protein acetylation and deacetylation catalysed by lysine acetyltransferases (KATs) and deacetylases (KDACs), respectively, are major mechanisms regulating various cellular processes. During the fight between microbial pathogens and host plants, both apply a set of measures, including acetylation interference, to strengthen themselves while suppressing the other. In this review, we first summarize KATs and KDACs in plants and their pathogens. Next, we introduce diverse acetylation and deacetylation mechanisms affecting protein functions, including the regulation of enzyme activity and specificity, protein-protein or protein-DNA interactions, subcellular localization and protein stability. We then focus on the current understanding of acetylation and deacetylation in plant-pathogen interactions. Additionally, we also discuss potential acetylation-related approaches for controlling plant diseases.

Interplay of two transcription factors for recruitment of the chromatin remodeling complex modulates fungal nitrosative stress response.

Nitric oxide (NO) is a diffusible signaling molecule that modulates animal and plant immune responses. In addition, reactive nitrogen species derived from NO can display antimicrobial activities by reacting with microbial cellular components, leading to nitrosative stress (NS) in pathogens. Here, we identify FgAreB as a regulator of the NS response in Fusarium graminearum, a fungal pathogen of cereal crops. FgAreB serves as a pioneer transcription factor for recruitment of the chromatin-remodeling complex SWI/SNF at the promoters of genes involved in the NS response, thus promoting their transcription. FgAreB plays important roles in fungal infection and growth. Furthermore, we show that a transcription repressor (FgIxr1) competes with the SWI/SNF complex for FgAreB binding, and negatively regulates the NS response. NS, in turn, promotes the degradation of FgIxr1, thus enhancing the recruitment of the SWI/SNF complex by FgAreB.

The RsmA RNA-Binding Proteins in Pseudomonas syringae Exhibit Distinct and Overlapping Roles in Modulating Virulence and Survival Under Different Nutritional Conditions.

The post-transcriptional regulator RsmA globally controls gene expression in bacteria. Previous studies showed that RsmA2 and RsmA3 played critical roles in regulating type III secretion system (T3SS), motility, syringafactin, and alginate productions in Pseudomonas syringae pv. tomato strain DC3000 (PstDC3000). In this study, we investigated global gene expression profiles of the wild-type PstDC3000, the rsmA3 mutant, and the rsmA2/A3 double mutant in the hrp-inducing minimum medium (HMM) and King's B (KB) medium. By comparing the rsmA2/A3 and rsmA3 mutants to PstDC3000, a total of 1358 and 1074 differentially expressed genes (DEGs) in HMM, and 870 and 1463 DEGs in KB were uncovered, respectively. When comparing the rsmA2/A3 mutant with the rsmA3 mutant, 277 and 741 DEGs in HMM and KB, respectively, were revealed. Transcriptomic analysis revealed that the rsmY, rsmZ, and rsmX1-5 non-coding small RNAs (ncsRNAs) were positively affected by RsmA2 and RsmA3, while RsmA3 positively regulates the expression of the rsmA2 gene and negatively regulates both rsmA1 and rsmA5 gene expression. Comparative transcriptomic analysis showed that RsmA2 and RsmA3 synergistically influenced the expression of genes involved in T3SS and alginate biosynthesis in HMM and chemotaxis in KB. RsmA2 and RsmA3 inversely affected genes involved in syringafactin production in HMM and ribosomal protein biosynthesis in KB. In addition, RsmA2 played a major role in influencing genes involved in sarcosine and thiamine biosynthesis in HMM and in mannitol and phosphate metabolism in KB. On the other hand, genes involved in fatty acid metabolism, cellulose biosynthesis, signal transduction, and stress responses were mainly impacted by RsmA3 in both HMM and KB; whereas RsmA3 played a major role in controlling genes involved in c-di-GMP, phosphate metabolism, chemotaxis, and capsular polysaccharide in HMM. Furthermore, regulation of syringafactin production and oxidative stress by RsmA2 and RsmA3 was experimentally verified. Our results suggested the potential interplay among the RsmA proteins, which exhibit distinct and overlapping roles in modulating virulence and survival in P. syringae under different nutritional conditions.

Advances in Understanding Fungicide Resistance in Botrytis cinerea in China.

Gray mold, caused by Botrytis cinerea, is a devastating disease that causes significant yield losses in various economically important plants. Fungicide application is one of the main strategies for management of gray mold; however, B. cinerea has developed resistance to various groups of fungicide. In China, benzimidazole-, dicarboximide-, and quinone outside inhibitor-resistant populations of B. cinerea have become dominant. Substitute mutations in fungicide target genes are responsible for resistance in B. cinerea. Based on known resistance mechanisms, molecular methods including loop-mediated isothermal amplification have been developed for rapid detection of resistant isolates of B. cinerea. Because B. cinerea is able to quickly develop resistance to various fungicides, various integrated strategies have been implemented in the last decade, including biological and agricultural practices, to manage fungicide resistance in B. cinerea.

Complete Genome Sequence of the Fire Blight Pathogen Strain Erwinia amylovora Ea1189.

Erwinia amylovora causes fire blight, the most devastating bacterial disease of apples and pears in the United States and worldwide. The model strain E. amylovora Ea1189 has been extensively used to understand bacterial pathogenesis and molecular mechanisms of bacterial-plant interactions. In this work, we sequenced and assembled the de novo genome of Ea1189, using a combination of long Oxford Nanopore Technologies and short Illumina sequence reads. A complete gapless genome assembly of Ea1189 consists of a 3,797,741-bp circular chromosome and a 28,259-bp plasmid with 3,472 predicted genes, including 78 transfer RNAs, 22 ribosomal RNAs, and 20 noncoding RNAs. A comparison of the Ea1189 genome to previously sequenced E. amylovora complete genomes showed 99.94 to 99.97% sequence similarity with 314 to 946 single nucleotide polymorphisms. We believe that the availability of the complete genome sequence of strain Ea1189 will further support studies to understand evolution, diversity and structural variations of Erwinia strains, as well as the molecular basis of E. amylovora pathogenesis and its interactions with host plants, thus facilitating the development of effective management strategies for this important disease.

The RasGEF FgCdc25 regulates fungal development and virulence in Fusarium graminearum via cAMP and MAPK signalling pathways.

Ras GTPases act as molecular switches to control various cellular processes by coupling integrated signals in eukaryotes. Activities of Ras GTPases are triggered by Ras GTPase guanine nucleotide exchange factors (RasGEFs) in general, whereas the role of RasGEF in plant pathogenic fungi is largely unknown. In this study, we characterized the only RasGEF protein in Fusarium graminearum, FgCdc25, by combining genetic, cytological and phenotypic strategies. FgCdc25 directly interacted with RasGTPase FgRas2, but not FgRas1, to regulate growth and sexual reproduction. Mutation of the FgCDC25 gene resulted in decreased toxisome formation and deoxynivalenol (DON) production, which was largely depended on cAMP signalling. In addition, FgCdc25 indirectly interacted with FgSte11 in FgSte11-Ste7-Gpmk1 cascade, and the ΔFgcdc25 strain totally abolished the formation of infection structures and was nonpathogenic in planta, which was partially recovered by addition of exogenous cAMP. In contrast, FgCdc25 directly interplayed with FgBck1 in FgBck1-MKK1-Mgv1 cascade to negatively control cell wall integrity. Collectively, these results suggest that FgCdc25 modulates cAMP and MAPK signalling pathways and further regulates fungal development, DON production and plant infection in F. graminearum.