Oomycetes and fungi:important parasites on marine algae

Considering that the field is largely unexplored and its importance to aquaculture,outline of oomycetes and fungi parasiting on marine algae was provided in this paper,including 15 species of oomycetes,six species of chytrids,31 Ascomycota species and one species of mitosporic fungi.In natrue,both the oomycetes and chytrids frequently occurred and induced prevalences of disease which could destroy the populations of host plants greatly.However,the parasites in Ascomycota on algae have never occurred as epidemics so far.Some issues relating to the field were discussed such as performing tests to satisfy Koch＇s postulates,investigations of host specificity,interactions between host and parasite and the potential effects of environmental factors on occurrence of a disease,which are urgent in need of further investigations.

Approaches to functional genomics in filamentous fungi

The study of gene function in filamentous fungi is a field of research that has made great advances in very recent years. A number of transformation and gene manipulation strategies have been developed and applied to a diverse and rapidly expanding list of economically important filamentous fungi and oomycetes. With the significant number of fungal genomes now sequenced or being sequenced, functional genomics promises to uncover a great deal of new information in coming years. This review discusses recent advances that have been made in examining gene function in filamentous fungi and describes the advantages and limitations of the different approaches.

A LAMP-assay-based specific microbiota analysis reveals community dynamics and potential interactions of 13 major soybean root pathogens

Soybean root diseases are associated with numerous fungal and oomycete pathogens;however,the community dynamics and interactions of these pathogens are largely unknown.We performed 13 loop-mediated isothermal amplification(LAMP)assays that targeted specific soybean root pathogens,and traditional isolation assays.A total of 159 samples were collected from three locations in the Huang-Huai-Hai region of China at three soybean growth stages(30,60,and 90 days after planting)in 2016.In LAMP results,we found that pathogen communities differed slightly among locations,but changed dramatically between soybean growth stages.Phytophthora sojae,Rhizoctonia solani,and Fusarium oxysporum were most frequently detected at the early stage,whereas Phomopsis longicolla,Fusarium equiseti,and Fusarium virguliforme were most common in the later stages.Most samples(86%)contained two to six pathogen species.Interestingly,the less detectable species tended to exist in the samples containing more detected species,and some pathogens preferentially co-occurred in diseased tissue,including P.sojae–R.solani–F.oxysporum and F.virguliforme–Calonectria ilicicola,implying potential interactions during infection.The LAMP detection results were confirmed by traditional isolation methods.The isolated strains exhibited different virulence to soybean,further implying a beneficial interaction among some pathogens.

GmDRR1,a dirigent protein resistant to Phytophthora sojae in Glycine max (L.) Merr.

Soil-borne pathogen Phytophthora sojae is an oomycete that causes devastating damage to soybean yield. To mine original resistant genes in soybean is an effective and environmentally-friend approach controlling the disease. In this study, soybean proteins were extracted from the first trifoliolates infected by predominant P. sojae race 1 and analyzed by twodimensional gel electrophoresis. Nineteen differently-expressed protein spots were detected, and 10 of them were further applied for Matrix-Assisted Laser Desorption/Ionization Time of Flight Mass Spectrometry Assay. One protein containing a dirigent （DIR） domain was identified and belonged to the DIR-b/d family. Therefore, it was named as GmDRR1 （Glycine max Disease Resistance Response 1）. Then, GmDRR1 gene was pathologically confirmed to be involved in the resistant to P. sojae in soybean. GmDRR1-GFP （green fluorescent protein） fusion proteins localized in the cell membrane. qRTPCR results showed GmDRR1 gene expressed differently in P. sojae resistant- and susceptible-soybean cultivars. By the promoter analysis, we found a haplotype H8 was existing in most resistant soybean varieties, while a haplotype H77 was existing in most susceptible soybean varieties. The H77 haplotype had seven SNPs （C to A, G to C, C to A, T to A, T to C, T to C, and T to A） and two single nucleotide insertions. The results supported that the expression difference of GmDRR1 genes between P. sojae resistant- and susceptible-soybean cultivars might depend on the GmDRR1 promoter SNPs. The results suggested that GmDRR1 was a dirigent protein involved in soybean resistant to P. sojae and paved a novel way for investigation of the molecular regulatory mechanism of the defense response to P. sojae in soybean.

Morphotaxonomic and phylogenetic analysis of Saprolegnia ferax from India

Saprolegnia ferax is isolated from polluted water sample collected from Mula River,Pune,Maharashtra,India.The isolated taxon is illustrated and compared with the morphotaxonomy based first record of Indian S.ferax isolated by Wani et al.(2017).The present study deals with the re-description of the isolated taxon S.ferax based on detail morphological features,sequence analysis and phylogeny of ITS and LSU regions of rDNA.

A Nicotiana benthamiana receptor-like kinase regulates Phytophthora resistance by coupling with BAK1 to enhance elicitin-triggered immunity

Cell-surface-localized leucine-rich-repeat receptorlike kinases(LRR-RLKs)are crucial for plant immunity.Most LRR-RLKs that act as receptors directly recognize ligands via a large extracellular domain(ECD),whereas LRR-RLK that serve as regulators are relatively small and contain fewer LRRs.Here,we identified LRR-RLK regulators using high-throughput tobacco rattle virus(TRV)-based gene silencing in the model plant Nicotiana benthamiana.We used the cell-death phenotype caused by INF1,an oomycete elicitin that induces pattern-triggered immunity,as an indicator.By screening 33 small LRR-RLKs(≤6 LRRs)of unknown function,we identified ELICITIN INSENSITIVE RLK 1(NbEIR1)as a positive regulator of INF1-induced immunity and oomycete resistance.Nicotiana benthamiana mutants of eir1 generated by CRISPR/Cas9-editing showed significantly compromised immune responses to INF1 and were more vulnerable to the oomycete pathogen Phytophthora capsici.NbEIR1 associates with BRI1-ASSOCIATED RECEPTOR KINASE 1(NbBAK1)and a downstream component,BRASSINOSTEROIDSIGNALING KINASE 1(NbBSK1).NbBSK1 also contributes to INF1-induced defense and P.capsici resistance.Upon INF1 treatment,NbEIR1 was released from NbBAK1 and NbBSK1 in vivo.Moreover,the silencing of NbBSK1 compromised the association of NbEIR1 with NbBAK1.We also showed that NbEIR1 regulates flg22-induced immunity and associates with its receptor,FLAGELLIN SENSING 2(NbFLS2).Collectively,our results suggest that NbEIR1 is a novel regulatory element for BAK1-dependent immunity.NbBSK1-NbEIR1 association is required for maintaining the NbEIR1/NbBAK1 complex in the resting state.

The genus Pythium in Taiwan,China(1)-a synoptic review

The genus Pythium,with slightly over 280 described species,has been classified traditionally with other filamentous,coenocytic,sporangia-producing fungi as“Phycomyetes”.However,with recent advances in chemical,ultrastructural and molecular studies,Pythium spp.are now considered as“fungus-like organisms”or“pseudo-fungi”and are placed in the Kingdom Chromista or Kingdom Straminopila,distinct from the true fungi of the Kingdom Fungi or Kingdom Mycota.They are widely distributed throughout the world as soil saprophytes or plant pathogens.Because of the warm moist maritime climate,Taiwan,China,is especially rich in Pythium species.To date,48 species of Pythium have been reported from Taiwan,China,with the dominant species being Py.vexans,Py.spinosum,Py.splendens,Py.aphanidermatum,Py.dissotocum and Py.acanthicum.There is no definite geographical distribution of Pythium spp.in Taiwan,China.Twenty nine species of Pythium have proven to be plant pathogens attacking a wide variety of woody and herbaceous plants primarily causing pre-and post-emergence seedling damping-off,root rot,stem rot and rotting of fruits,tubers and ginger rhizomes,resulting in serious economic losses.The most important plant pathogenic species include Py.aphanidermatum and Py.Myriotylum,which are most active during the hot and wet summer months;whereas Py.splendens,Py.spinosum,Py.ultimum and Py.irregulare cause the greatest damage in the cool winter.Most Pythium spp.are non-specific pathogens,infecting mainly juvenile or succulent tissues.This review attempts to assess the taxonomic position of the genus Pythium and provide details of the historical development of the study of Pythium as pathogens in Taiwan,China,causing diseases of sugarcane,trees,vegetables,fruits,specialty crops and flowering plants,as well as measures to control these diseases.Of special note is the introduction of the S-H mixture which,when used as soil amendment,effectively controls many soil-borne Pythium diseases during the early stages of plant growth.The diversity of Pythium species in Taiwan,China,is discussed in comparison with the situation in the mainland of China and suggestions are made to fully utilize Pythium spp.as agents for biological control,in industry and medicine.

A Host KH RNA-Binding Protein Is a Susceptibility Factor Targeted by an RXLR Effector to Promote Late Blight Disease

Plant pathogens deliver effector proteins that alter host processes to create an environment conducive to colonization. Attention has focused on identifying the targets of effectors and how their manipulation facil- itates disease. RXLR effector Pi04089 from the potato blight pathogen Phytophthora infestans accumu- lates in the host nucleus and enhances colonization when transiently expressed in planta. Its nuclear local- ization is required for enhanced P. infestans colonization. Pi04089 interacts in yeast and in planta with a putative potato K-homology （KH） RNA-binding protein, StKRBPI. Co-localization of Pi04089 and StKRBP1, and bimolecular fluorescence complementation between them, indicate they associate at nuclear speckles. StKRBP1 protein levels increased when it was co-expressed with Pi04089. Indeed, such accumu- lation of StKRBP1 was observed also on the first day of leaf colonization by the pathogen. Remarkably, overexpression of StKRBP1 significantly enhances P. infestans infection. Mutation of the nucleotide- binding motif GxxG to GDDG in all three KH domains of StKRBP1 abolishes its interaction with Pi04089, its localization to nuclear speckles, and its increased accumulation when co-expressed with the effector. Moreover, the mutant StKRBP1 protein no longer enhances leaf colonization by P. infestans, implying that nucleotide binding is likely required for this activity. We thus argue that StKRBP1 can be regarded as a sus- ceptibility factor, as its activity is beneficial to the pathogen.

All Roads Lead to Susceptibility: The Many Modes of Action of Fungal and Oomycete Intracellular Effectors

The ability to secrete effector proteins that can enter plant cells and manipulate host processes is a key determinant of what makes a successful plant pathogen.Here,we review intracellular effectors from filamentous(fungal and oomycete)phytopathogens and the host proteins and processes that are targeted to promote disease.We cover contrasting virulence strategies and effector modes of action.Filamentous pathogen effectors alter the fates of host proteins that they target,changing their stability,their activity,their location,and the protein partners with which they interact.Some effectors inhibit target activity,whereas others enhance or utilize it,and some target multiple host proteins.We discuss the emerging topic of effectors that target negative regulators of immunity or other plant proteins with activities that support susceptibility.We also highlight the commonly targeted host proteins that are manipulated by effectors from multiple pathogens,including those representing different kingdoms of life.

A G-type lectin receptor-like kinase regulates the perception of oomycete apoplastic expansin-like proteins

Phytophthora capsici is one of the most harmful pathogens in agriculture, which threatens the safe production of multiple crops and causes serious economic losses worldwide. Here, we identified a P. capsici expansin-like protein, Pc EXLX1, by liquid chromatography-tandem mass spectrometry from Nicotiana benthamiana apoplastic fluid infected with P. capsici. Clustered regularly interspaced short palindromic repeats/crispr associated protein9(CRISPR/Cas9)-mediated Pc EXLX1 knockout mutants exhibited significantly enhanced virulence,while the overexpression of Pc EXLX1 impaired the virulence. Prokaryotically expressed Pc EXLX1 activated multiple plant immune responses, which were BRI1-associated kinase 1(BAK1)-and suppressor of BIR1-1(SOBIR1)-dependent. Furthermore, overexpression of Pc EXLX1 homologs in N. benthamiana could also increase plant resistance to P. capsici. A G-type lectin receptor-like kinase from N. benthamiana, expansin-regulating kinase 1(ERK1), was shown to regulate the perception of Pc EXLX1 and positively mediate the plant resistance to P. capsici. These results reveal that the expansin-like protein, Pc EXLX1, is a novel apoplastic effector with plant immunity-inducing activity of oomycetes, perception of which is regulated by the receptor-like kinase, ERK1.

The Ubiquitin E3 Ligase PUB17 Positively Regulates Immunity by Targeting a Negative Regulator, KH17, for Degradation

Ubiquitination is a post-translational modification that regulates many processes in plants.Several ubiquitin E3 ligases act as either positive or negative regulators of immunity by promoting the degradation of different substrates.StPUB17 is an E3 ligase that has previously been shown to positively regulate immunity to bacteria,fungi and oomycetes,including the late blight pathogen Phytophthora infestans.Silencing of StPUB17 promotes pathogen colonization and attenuates Cf4/avr4 cell death.Using yeast-2-hybrid and co-immunoprecipitation we identified the putative K-homology(KH)RNA-binding protein(RBP),StKH17,as a candidate substrate for degradation by StPUB17.StKH17 acts as a negative regulator of immunity that promotes P.infestans infection and suppresses specific immune pathways.A KH RBP domain mutant of StKH17(StKH17GDDG)is no longer able to negatively regulate immunity,indicating that RNA binding is likely required for StKH17 function.As StPUB17 is a known target of the ubiquitin E3 ligase,StPOB1,we reveal an additional step in an E3 ligase regulatory cascade that controls plant defense.

The Role of Autophagy during Development of the Oomycete Pathogen Phytophthora infestans

Phytophthora infestans is a highly destructive plant path- ogen that causes late blight disease in plants such as potato and tomato （Fry and Goodwin, 1997）. As a typical oomycete, the primary infective propagule of P. infestans is the sporangium, which can either germinate directly or release biflagellate zoospores that encyst and germinate on the host subsequently （Grenville-Briggs et al., 2005）. Due to abundant formation of the sporangia and prolific sporulation, P. infestans can spread quickly in the host tissues and cause extensive necrosis of the infected plants （Kamoun and Smart, 2005）.

Molecular identification of oomycete species affecting aquaculture in Bangladesh

Fish mycotic disease outbreaks occur due to infections with oomycete pathogens such as Saprolegnia spp.and Aphanomyces invadans,and cause large-scale fish production losses.Despite its negative impact on aquaculture,little is known about the diversity of oomycete pathogens.The aim of this study was to identify the diversity of pathogenic oomycetes causing infections in major aquaculture zones of Mymensingh and Jessore regions in Bangladesh.A total of 449 water and infected fish samples were collected from 28 fish farms in both regions of which 29 samples were able to grow out with mycelia on the Potato Dextrose Agar(PDA)/Glucose Peptone Yeast Agar(GPYA)plates.Sequence database searches using the rRNA Internal Transcribed Spacer(ITS)region revealed that 15 belonged to Pythium spp.,12 were Aphanomyces invadans and two corresponded to Saprolegnia parasitica.Five isolates of Pythium spp.were identified to the species level:one was closely related to Pythium catenulatum,four to Pythium rhizo-oryzae,rest were identified up to genus.The Pythium spp.were only isolated from water samples whereas A.invadans and S.parasitica were found in fish lesions.Phylogenetic analysis revealed that a single A.invadans clone exists in the sampled area.The results obtained confirm the existence of pathogenic oomycetes in Bangladesh fish farms and this will pave future research on diversity,prevention and control measures.