Controlling Fusarium Head Blight in oat

Oats (Avena sativa) is a versatile crop grown worldwide for animal feed and human consumption. Humanoat consumption has recently risen due to its various health benefits. However, oats are susceptible toFusarium head blight (FHB) caused by various Fusarium fungi. FHB reduces yield and leads to mycotoxinaccumulation. The most commonly reported mycotoxins in oat are trichothecenes deoxynivalenol (DON)and T-2/HT-2 toxins. Trichothecenes inhibit eukaryotic protein biosynthesis and cause acute and chronictoxicoses in human and animals. Effective control of FHB is important for ensuring safety and quality ofoats. This thesis examines various aspects of FHB in oats, relevant to the development of better FHBcontrol strategies.Accurate FHB symptom identification is crucial for breeding resistant oats, but the symptoms of FHB arecryptic, causing errors in scoring the disease during trials. This work presents an affordable method forassessing FHB symptoms in oats by de-hulling mature seeds. Symptoms of blackening and discolorationof the oat kernels significantly correlate with Fusarium DNA and mycotoxin accumulation and thus canbe used as quantitative disease indicators.To enhance pathogen resistance, identifying and characterizing plant resistance genes is key. In thiswork two oat genes coding for DON-detoxifying UDP-glucosyltransferases (UGTs) were identified andcharacterised. Transcripts of two oat UGTs were highly upregulated in response to DON treatment andF.graminearum infection. The genes conferred resistance to several trichothecenes when expressed inyeast. Both UGTs, recombinantly expressed in E.coli were confirmed for their ability to detoxify DON.These genes could potentially be used for developing genetic markers for FHB resistance in oat.Further in this thesis, biocontrol possibilities for FHB in oats are investigated. The fungal BCAClonostachys rosea's potential against FHB is examined. Treating oat spikelets with C. rosea reducedFusarium DNA and DON content in mature kernels. C.rosea enhanced both rate of DON detoxificationand expression of DON-detoxifying UGTs. Furthermore, there was significant upregulation of markers ofinduced resistance, including PR proteins and the WRKY23 transcription factor, indicating that thebiocontrol effect of C. rosea is attributed to the induction of plant defences.Additionally, oats' own endophytes were explored for FHB biocontrol. Fungal endophytes from oatspikelets were isolated and tested for reducing FHB in greenhouse trials. The most successful isolatePseudozyma flocculosa significantly reduced FHB symptoms, F. graminearum biomass, and DONaccumulation in oat. Treatment of oat with P. flocculosa induced expression of genes encoding for PRproteins, known to be involved in FHB resistance

28th Fungal Genetics Conference

Full abstracts from the 28th Fungal Genetics Conference Asilomar, March 17-22, 2015

벼 도열병균의 긴 비암호화 리보핵산 분석 및 짧은 비암호화 리보핵산과의 상호작용

학위논문(박사) -- 서울대학교대학원 : 농업생명과학대학 협동과정 농생명유전체학전공, 2023. 2. 이용환.단백질을 암호화하는 구역 및 암호화하는 서열이 없는 구역에서도 전사는 일어난다. 비암호화 리보핵산은 단백질을 만드는 정보가 없지만 유전자를 조절함으로써 전사 과정, 전사 후 과정, 번역 과정, 번역 후 과정에서 일어나는 조절 과정에 관여한다. 비암호화 리보핵산은 200개의 염기보다 긴 경우 긴 비암호화 리보핵산(lncRNA)으로 간주된다. 시퀀싱(sequencing) 분석 기술이 발전하면서 비암호화 리보핵산 전사체가 축적되고 기능 분석이 수행되고 있다. 긴 비암호화 리보핵산은 발달 과정, 비생물적 자극에 대한 반응, 기주와 미생물의 상호작용에 참여한다고 보고되었다. 그러나 제한된 종에서의 연구로 인해 식물병원성 곰팡이에서는 긴 비암호화 리보핵산에 대한 역할에 대한 이해가 부족하다. 해당 연구는 기주에 대한 반응에서 긴 비암호화 리보핵산의 역할을 이해하기 위해 병이 발생하는 동안 벼 도열병균(Magnaporthe oryzae)에서 프로파일링(profiling)을 수행했다. 긴 비암호화 리보핵산을 확인 후 기능과 관련이 있을 수 있는 유전체 서열 특징과 발현 경향을 분석했다. 추가적으로, 기능을 할 가능성이 큰 긴 비암호화 리보핵산을 조사하기 위해서 감염 단계에 특이적으로 발현한 경우의 대상 유전자를 탐색했다. 유전자 분석 결과는 긴 비암호화 리보핵산은 세포벽 분해와 기주의 면역체계 회피 같은 역할을 수행하여 병원성에 관여한다고 제시해 준다. 긴 비암호화 리보핵산은 단독으로 또는 짧은 비암호화 리보핵산(sRNA)와 협력해서 기능한다. 상호작용 방식은 일반적으로 세 가지가 있다. 전자가 후자의 전구체가 되는 경우, 후자가 전자를 조절하는 경우, 전자가 후자의 활동을 조절하는 경우로 구분할 수 있다. 곰팡이에서는 이들의 상호작용에 대한 이해가 부족한 상황이다. 벼 도열병균에서 상호작용을 밝히기 위해 짧은 비암호화 리보핵산의 생합성 유전자가 없는 상황에서 두 비암호화 리보핵산의 프로파일링을 수행했다. 그 과정에서 짧은 비암호화 리보핵산 중 잔해를 배제하기 위해서 리보핵산 간섭 도구에 의해 처리되는 것들을 선별했다. 대상 유전자의 분석 결과 상호작용의 종류에 따라 다른 생물학적 과정과 연관되어 있음을 밝혔다. 해당 연구는 비암호화 리보핵산의 레퍼토리를 제공하여 생물학적 기능을 알아보기 위한 기능적 연구의 기반을 제공한다. 또한 종합적인 연구를 통해 두 종류의 비암호화 리보핵산의 상호작용에 대한 이해를 돕고, 병원성을 포함하는 생물학적 과정에서 이들이 핵심 요소라는 점을 제안한다. 따라서 본 연구는 식물 병원성 곰팡이에서 복잡한 조절망에 대한 연구 방향을 제시한다.Transcription occurs in the protein-coding regions as well as the regions where any protein-coding sequence is absent. Although these non-coding RNAs lack coding potential, they play roles in transcriptional, post-transcriptional, translational, and post-translational regulation by controlling protein-coding genes. Non-coding RNAs, which are longer than 200 nucleotides, are considered as long non-coding RNAs (lncRNAs). As the sequencing technology has advanced, a repertoire of lncRNA transcriptomes has been accumulated and the functional characterization of each lncRNA has been performed. LncRNAs have been reported to participate in the development, responses to abiotic stresses, and host-microbe interaction. However, their role in plant fungal pathogens was poorly understood due to the limited range of studied species. In this study, we profiled lncRNAs of the rice blast fungus, Magnaporthe oryzae, during disease development to decipher the role of lncRNAs in response to the host. We identified lncRNAs and analyzed their genomic feature and expression pattern to understand their properties, which could be related to their functions. Moreover, specifically expressed lncRNAs in infection stages and their target genes were identified to investigate functional lncRNAs. The analysis of target gene functions suggests that these lncRNAs play roles in pathogenesis such as cell wall degradation and evasion of host immunity. LncRNAs could function solely or in cooperation with small RNAs (sRNAs). LncRNAs generally interact with sRNAs in three ways. LncRNAs could be precursors of sRNAs, be regulated by sRNAs, and regulate sRNA activity. However, their interaction is not well understood in fungi. We profiled lncRNAs and sRNAs in the defect of sRNA biogenesis machinery genes to unravel their interaction in M. oryzae. We selected sRNAs processed by RNA interference machinery to filter out the debris. The analysis of genes targeted by non-coding RNAs suggests that two classes of non-coding RNAs be involved in different biological processes depending on the type of interaction. This study provides a repertoire of non-coding RNAs and a foundation for functional studies to elucidate their biological roles. This comprehensive study helps to understand the crosstalk between two classes of non-coding RNAs and suggests that non-coding RNAs can be key regulators in biological processes including pathogenesis. Taken together, this work shed light on the complex regulatory network in plant pathogenic fungi.CHAPTER I. Long non-coding RNA in fungi 1 ABSTRACT 2 INTRODUCTION 3 I. LncRNA profiling in fungi 5 II. Biological roles of lncRNAs in fungi 9 PERSPECTIVE 14 LITERATURE CITED 15 CHAPTER II. Genome-wide profiling of long non-coding RNA of the rice blast fungus Magnaporthe oryzae during infection 26 ABSTRACT 27 INTRODUCTION 28 MATERIALS AND METHODS I. RNA extraction and strand‐specific sequencing 31 II. Collection of in planta RNA-seq data 31 III. Transcriptome assembly 32 IV. LncRNA identification 32 V. LncRNA conservation analysis 35 VI. Assessment of stage specificity and prediction of stage-specific lncRNAs 35 VII. Target gene prediction 36 VIII. Validation of lncRNA transcript production 37 RESULTS 39 I. Genome-wide identification of lncRNAs in M. oryzae 39 II. Genomic features of M. oryzae lncRNAs 43 III. Expression of lncRNA transcripts during infection 46 IV. Prediction of stage-specifically expressed lncRNA 50 V. Verification of lncRNA production 57 DISCUSSION 60 LITERATURE CITED 63 CHAPTER III. Comprehensive genome-wide analysis of non-coding RNAs reveals functions of lncRNA-sRNA crosstalk in the rice blast fungus Magnaporthe oryzae 71 ABSTRACT 72 INTRODUCTION 73 MATERIALS AND METHODS I. Collection of RNA-seq and sRNA-seq data 76 II. RNA-seq data analysis 76 III. sRNA-seq data analysis 77 IV. Target gene prediction and analysis 78 RESULTS 79 I. Identification of lncRNAs and Dicer-dependent sRNAs 79 II. Identification of small RNAs originating from lncRNAs 84 III. Identification of sRNAs regulating lncRNA expression 89 IV. Construction of a lncRNA-sRNA-mRNA network 92 DISCUSSION 94 LITERATURE CITED 97 ABSTRACT (in Korean) 104박

A Review on Disease Detection, Pathogen Identification and Population Genetics in Fungi

Based on knowing the fungi how much they are important in causing holistic impacts on human welfare, the present review initiated with reviewing the different disease detection and pathogen identification methods of plant pathogenic fungi and over viewing the population genetics of fungi. Currently, more and more diagnostic laboratories and inspection agencies are using molecular methods for detection and identification of diseases caused by plant pathogenic fungi. Better understanding of fungal-plant interactions, pathogenicity factors, rapid and accurate detection of fungal pathogens to species or strain level are a crucial prerequisite for disease surveillance and development of novel disease control strategies. Molecular technology increases understanding of the biology and population structures of plant pathogens, provides quick and accurate answers to epidemiological questions about plant diseases, and supports disease management decisions. Both domestic and international plant quarantine stations must be facilitated with high throughput, rapid and sensitive methods for detection of quarantine pathogens. The methods of population genetics offer powerful tools to elucidate the life histories of important plant pathogens and address fundamental questions about the biology of these organisms. Increased knowledge of the population biology of pathogens is likely to lead to better management of disease in agricultural ecosystems. Eventhough every pathogen detection methods starting from conventional up-to the most sophisticated real-time PCR, they have their own advantages and drawbacks, but selecting of the most appropriate one based on its multi-dimensional importance is crucial at this time. In Ethiopia, most of the detection methods are based on the conventional one, but these methods are not as effective as molecular detection methods. So developing and utilizing these more effective, time sever and easy assays of fungal disease detection and pathogen identification methods is important. Keywords: Agricultural Ecosystems, Fungal-Plant Interactions, PCR, Pathogenicity Factors, Real-Time PC

Computational and molecular study of terpene synthase genes in Trichoderma

[ES] Trichoderma comprende un amplio número de especies, de gran interés en el manejo de enfermedades de las plantas de cultivo y la industria. El amplio rango de estilos de vida de estas especies está respaldado por su diversidad de Metabolitos Secundarios (MSs). En este trabajo, se ha utilizado una combinación de enfoques de minería genómica y genómica comparativa generando una extensa visión sobre el potencial de biosíntesis de MSs en Trichoderma

Program and abstracts from the 24th Fungal Genetics Conference

Abstracts of the plenary and poster sessions from the 24th Fungal Genetics Conference, March 20-25, 2007, Pacific Grove, CA

27th Fungal Genetics Conference

Program and abstracts from the 27th Fungal Genetics Conference Asilomar, March 12-17, 2013

Proteomics of Plant Pathogenic Fungi

Plant pathogenic fungi cause important yield losses in crops. In order to develop efficient and environmental friendly crop protection strategies, molecular studies of the fungal biological cycle, virulence factors, and interaction with its host are necessary. For that reason, several approaches have been performed using both classical genetic, cell biology, and biochemistry and the modern, holistic, and high-throughput, omic techniques. This work briefly overviews the tools available for studying Plant Pathogenic Fungi and is amply focused on MS-based Proteomics analysis, based on original papers published up to December 2009. At a methodological level, different steps in a proteomic workflow experiment are discussed. Separate sections are devoted to fungal descriptive (intracellular, subcellular, extracellular) and differential expression proteomics and interactomics. From the work published we can conclude that Proteomics, in combination with other techniques, constitutes a powerful tool for providing important information about pathogenicity and virulence factors, thus opening up new possibilities for crop disease diagnosis and crop protection

XXIII Fungal Genetics Conference

Program and abstracts from the 23rd Fungal Genetics Conference and Poster Abstracts at Asilomar, March 15-20, 200