Sec4/Rab8 is one of the well-studied members of the Rab GTPase family, previous studies have shown that Sec4/Rab8 crucially promotes the pathogenesis of phytopathogens, but the upstream regulators of Rab8 are still unknown. Here, we have identified two Sec2 homologues FgSec2A and FgSec2B in devastating fungal pathogen Fusarium graminearum and investigated their functions and interactions with FgRab8 by live-cell imaging, genetic and functional analyses. Yeast two-hybrid assay shows that FgSec2A specifically interacts with FgRab8DN(N123I) and itself. Importantly, FgSec2A is required for growth, conidiation, DON production and virulence of F. graminearum. Live-cell imaging shows that FgSec2A and FgSec2B are both localized to the tip region of hyphae and conidia. Both N-terminal region and Sec2 domain of FgSec2A are essential for its function, but not for localization, whereas the C-terminal region is important for its polarized localization. Furthermore, constitutively active FgRab8CA(Q69L) partially rescues the defects of ΔFgsec2A. Consistently, FgSec2A is required for the polarized localization of FgRab8. Finally, FgSec2A and FgSec2B show partial functions, but FgSec2A does not interact and co-localize with FgSec2B. Taken together, these results indicate that FgSec2A acts as a FgRab8 guanine nucleotide exchange factor and is necessary for polarized growth, DON production and pathogenicity in F. graminearum.

Summary In filamentous fungi, hyphal growth depends on the continuous delivery of vesicles to the growing tips. It is unclear how fast‐growing hyphae coordinate simultaneous cell extension and expansion in the tip cells. We have functionally characterized 12 TBC (Tre‐2/Bub2/Cdc16) domain‐containing proteins in Fusarium graminearum. Among them, FgMsb3 is found to regulate hyphal tip expansion and to be required for pathogenicity. The regulatory mechanism of FgMsb3 has been further investigated by genetic, high‐resolution microscopy and high‐throughput co‐immunoprecipitation strategies. The FgMsb3 protein localizes at the polarisome and the hyphal apical dome (HAD) where it acts as a GTPase‐activating protein for FgRab8 which is required for apical secretion‐mediated growth and pathogenicity. Deletion of FgMSB3 causes excessive polarized trafficking but blocks the fusion of FgSnc1‐associated vesicles to the plasma membrane. Moreover, we establish that FgSpa2 interacts with FgMsb3, enabling FgMsb3 tethering to the polarisome. Loss of FgSpa2 or other polarisome components (FgBud6 and FgPea2) causes complete shifting of FgMsb3 to the HAD and this affects the polarized growth and pathogenicity of the fungus. In summary, we conclude that FgSpa2 regulates FgMsb3–FgRab8 cascade and this is crucial for creating a steady‐state equilibrium that maintains continuous polarized growth and contributes to the pathogenicity of F. graminearum.

Fusarium graminearum is a fungal pathogen that causes Fusarium head blight (FHB) in wheat and barley. Autophagy is a highly conserved vacuolar degradation pathway essential for cellular homeostasis in which Atg9 serves as a multispanning membrane protein important for generating membranes for the formation of phagophore assembly site. However, the mechanism of autophagy or autophagosome formation in phytopathogens awaits further clarifications. In this study, we identified and characterized the Atg9 homolog (FgAtg9) in F. graminearum by live cell imaging, biochemical and genetic analyses. We find that GFP-FgAtg9 localizes to late endosomes and trans-Golgi network under both nutrient-rich and nitrogen starvation conditions and also show its dynamic actin-dependent trafficking in the cell. Further targeted gene deletion of FgATG9 demonstrates that it is important for growth, aerial hyphae development, and pathogenicity in F. graminearum. Furthermore, the deletion mutant (ΔFgatg9) shows severe defects in autophagy and lipid metabolism in response to carbon starvation. Interestingly, small GTPase FgRab7 is found to be required for the dynamic trafficking of FgAtg9, and co-immunoprecipitation (Co-IP) assays show that FgAtg9 associates with FgRab7 in vivo. Finally, heterologous complementation assay shows that Atg9 is functionally conserved in F. graminearum and Magnaporthe oryzae. Taken together, we conclude that FgAtg9 is essential for autophagy-dependent development and pathogenicity of F. graminearum, which may be regulated by the small GTPase FgRab7.

Objective To identify the pathogens responsible for neonatal sepsis in a high-volume women and children's hospital in Southwest China. Methods We retrospectively studied 133 neonates who were admitted to the West China Women and Children's Hospital between 2008 and 2012 for sepsis. The clinical characteristics of the patients were recorded, and the antibiotic sensitivities of the isolated bacteria were determined. Results All of the included patients had clinical symptoms of sepsis, and subsequent blood cultures confirmed the infection. Almost 80% of patients were infected with coagulase-negative staphylococci (52.8%), Escherichia coli (23.6%), Klebsiella pneumoniae (16.0%) or Staphylococcus aureus (7.5%). Neonates who were infected with gram-negative bacteria, particularly K. pneumoniae, had lower birth weights and were admitted to hospital within 24 hours of birth. Additionally, 87.5% of the isolated K. pneumoniae strains were resistant to third generation cephalosporins. Conclusion Coagulase-negative staphylococci were the most common pathogens found in neonatal sepsis. Moreover, neonatal sepsis caused by gram-negative bacteria was more often observed in newborns of low birth weight. The isolated strains of gram-negative bacteria were highly resistant to cephalosporins. This observation highlights the issue of antibiotic-resistant pathogens in the clinical setting, which poses an added risk to infants presenting with sepsis.

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