Fungal Trunk Diseases Research Articles

Fungal Trunk Diseases Causing Decline of Apricot and Plum Trees in the Czech Republic.

Fungal trunk diseases (FTDs) have been a significant threat to the global stone fruit industry. FTDs are caused by a consortium of wood-decaying fungi. These fungi colonize woody tissues, causing cankers, dieback, and other decline-related symptoms in host plants. In this study, a detailed screening of the fungal microbiota associated with the decline of stone fruit trees in the Czech Republic was performed. The wood fragments of plum and apricot trees showing symptoms of FTDs were subjected to fungal isolation. The partial internal transcribed spacer region, partial beta-tubulin, and translation elongation factor 1-α genes were amplified from genomic DNA extracted from fungal cultures. All isolates were classified, and the taxonomic placement of pathogenic strains was illustrated in phylogenetic trees. The most abundant pathogenic genus was Dactylonectria (31%), followed by Biscogniauxia (13%), Thelonectria (10%), Eutypa (9%), Dothiorella (7%), Diplodia (6%), and Diaporthe (6%). The most frequent endophytic genus was Aposphaeria (17%). The pathogenicity of six fungal species (Cadophora daguensis, Collophorina africana, Cytospora sorbicola, Dothiorella sarmentorum, Eutypa lata, and E. petrakii var. petrakii) to four Prunus spp. was evaluated, and Koch's postulates were fulfilled. All tested isolates caused lesions on at least one Prunus sp. The most aggressive species was E. lata, which caused the largest lesions on all four tested Prunus spp., followed by E. petrakii var. petrakii and D. sarmentorum. Japanese plum (Prunus salicina) and almond (P. amygdalus) were the most susceptible hosts, while apricot (P. armeniaca) was the least susceptible host in the pathogenicity trial.

Characterization and pathogenicity of fungal species associated with hazelnut trunk diseases in North-western Italy

AbstractItaly is the second largest hazelnut producer worldwide and Piedmont is one of the most productive regions in the country. The changing climatic condition and fungal trunk diseases (FTD) can have a severe impact on this crop. Particularly, the considerable spread of Cytospora cankers (‘Mal dello stacco’) and dieback represent a serious concern for producers. Thus, considering the limited studies on the causal agents, different surveys were conducted in seven hazelnut orchards during 2021 and 2022. Eight fungal species were identified: Anthostoma decipiens, Botryosphaeria dothidea, Diaporthe eres, Dia. rudis, Diplodia seriata, Dip. subglobosa, Dothiorella parva and Nothophoma brennandiae. Species identification was achieved through multilocus phylogeny and morphology assessment. All the fungal species were pathogenic on healthy hazelnut plants (cultivar Tonda Gentile) and A. decipiens and Dia. eres were the most aggressive. The present study is the first report of B. dothidea and Dia. eres as causal agents of FTD on hazelnut in Italy and of Dia. rudis, Dip. subglobosa and N. brennandiae worldwide. Moreover, the study provides clarification of the fungal pathogens associated with FTD on this crop in Piedmont, thus laying the base for further studies on epidemiology, ecology and management strategies.

Molecular Identification and Pathogenicity of Diaporthe eres and D. hongkongensis (Diaporthales, Ascomycota) Associated with Cherry Trunk Diseases in China.

This study aimed to identify fungal species associated with trunk diseases of sweet cherries (Prunus avium) in several commercial cherry orchards in Beijing, Guizhou and Shandong provinces, China. In total, eighteen fungal strains that fitted well into the species concept of Diaporthe were isolated. Based on both morphological and multi-locus phylogenetic analyses of internal transcribed spacer region (ITS), beta-tubulin (tub-2), calmodulin (Cal) and translation elongation factor 1-α (tef1-α) sequencing data, fourteen isolates were identified as Diaporthe eres, while four isolates were classified as D. hongkongensis. Here, we report D. hongkongensis causing sweet cherry branch dieback disease and, further, we confirmed the host association of D. eres with sweet cherries in China. A pathogenicity assay revealed the ability of both D. eres and D. hongkongensis to cause shoot necrosis and stem lesions on Prunus avium cv. 'Brooks' (mean lesion lengths of 1.86 cm and 1.56 cm, respectively). The optimal temperature for the growth of both Diaporthe species was tested. The optimal growth temperature for D. hongkongensis was 30 °C, and the 25-28 °C temperatures were the most favorable for the growth of D. eres strains. This research advances the understanding of fungal trunk diseases in fruit crops, particularly gummosis and branch dieback disease in Chinese cherry orchards, and will aid growers in making decisions about cultural practices and disease management.

Assessing the relationship between cordon strangulation, dieback, and fungal trunk disease symptom expression in grapevine

Assessing the relationship between cordon strangulation, dieback, and fungal trunk disease symptom expression in grapevine

Fungal trunk diseases of fruit trees in Europe: pathogens, spread and future directions

Production from crops of pome, stone fruit, nut, berry fruit, citrus, grapevine, and olive is increasingly threatened by fungal trunk diseases (FTD). These diseases and the consequent production losses are major problems. Many fungi (including Botryosphaeriaceae, Calosphaeriaceae, Diaporthaceae, Diatrypaceae, Nectriaceae, Phaeomoniellaceae, Pleosporaceae, Togniniaceae, Valsaceae) infect host wood, mainly through wounds and subsequent colonization of woody tissues, causing symptoms such as cankers, gummosis, wood rotting, blight and dieback. Propagative plant material, seedlings and fruit play a significant role in pathogen spread. Several abiotic factors (e.g. shifts in cultural practices and climate change) are involved in the disease development. This paper reviews recent literature on FTD of fruit crops, particularly focusing on the European status of pathogen occurrence. Case studies are described related to diseases of apple, citrus, grapevine, berry, nut and stone fruit, and olive trees. Aspects related to epidemiology and the increase in disease incidence along with the future perspectives on the FTD research are also discussed.

Grapevine nursery propagation material as source of fungal trunk disease pathogens in Uruguay.

Grapevine fungal trunk diseases (GTDs) have become a serious problem for grapevines worldwide. Nursery vines infected during the propagation process are considered one of the main ways of dissemination of GTD pathogens. In this study, we examined the status of GTDs in grapevine planting material, from rootstocks and scion mother cuttings to grafted rooted vines ready to plant, according to the local nursery propagation process. During 2018-2019, internal symptoms of GTDs were examined in 2400 propagation materials and fungal isolations were carried out from a subsample of 1026 selected materials. Our results revealed that nursery grapevine plants produced in Uruguay have a high incidence of GTDs, regardless of the scion/rootstock combination. Typical brown to black streaks and sectorial wood necrosis were observed in materials on all propagation stages, with a markedly increasing incidence throughout the nursery process, reaching almost 100% in grafted rooted vines ready to plant. Botryosphaeria dieback, Petri disease and black-foot disease were the main GTDs found. The results showed that Botryosphaeria dieback and Petri disease pathogens infect materials from the early stages of the process, with a marked increase towards the end of the plant production process, whereas black-foot disease pathogens were found exclusively in vines ready to plant. Diaporthe dieback pathogens were also detected in materials in all stages but in a low proportion (less than 10% of infected material). Based on single locus analysis, the 180 isolates selected were placed into eight genera and 89% identified within 22 fungal species associated with GTDs, with Phaeoacremonium oleae and Diaporthe terebinthifolii as new records on grapevine worldwide. Our results have concluded that locally produced vines are one of the main ways of dissemination of GTD pathogens and showed that a nursery sanitation programme is required to reduce the incidence of these diseases.

Silverleaf (Chondrostereum purpureum) Effects on Japanese Plum (Prunus salicina).

Silverleaf is an important fungal trunk disease of fruit crops, such as Japanese plum (Prunus salicina). It is known that infection by Chondrostereum purpureum results in discolored wood, “silvered” foliage, and tree decline. However, effects on fruit yield and quality have not been assessed. Therefore, the objectives of this study were to determine C. purpureum pathogenicity on P. salicina and the effects on physiology, fruit yield, and quality, in Chile, in 2019 and 2020. Wood samples from affected plum trees were collected in the Chilean plum productive area. Fungi were isolated by plating wood sections from the necrosis margin on culture media. Morphological and molecular characteristics of the isolates corresponded to C. purpureum (98%). Representative isolates were inoculated from healthy plum plants and after 65-d incubation, wood necrotic lesions and silver leaves were visible. Fungi were reisolated, fulfilling Koch’s postulates. To determine Silverleaf effects, xylem water potential and fruit yield and quality were measured in healthy and Silverleaf-diseased plum trees ‘Angeleno’. Water potential was altered in diseased trees, and fruit yield was reduced by 51% (2019) and by 41% (2020) compared to fruit from healthy trees. Moreover, cover-colour, equatorial-diameter, and weight were reduced, and fruit were softer, failing to meet the criteria to be properly commercialized and exported to demanding markets.

Comparison of different inoculation methods in the investigation of fungal trunk diseases in grapevines

Amaç: Asma gövde hastalıkları, birçok fungal bitki patojeninin neden olduğu kompleks hastalıklardır. Bu çalışmada dört farklı hastalığı (Petri ve Siyah Bacak Hastalığı, Botryosphaeria ve Phomopsis Geriye Ölüm Hastalıkları) temsil eden bazı fungal patojenler için en uygun inokulasyon yönteminin saptanması amaçlanmıştır.Yöntem ve Bulgular: Phaeomoniella chlamydospora, Dactylonectria macrodidyma, Neofusicoccum parvum ve Diaporthe ampelina türü funguslar dormant asma kalemlerine veya yeşil sürgünlere farklı yöntemlerle inokule edilmişlerdir. Dormant kalemlere uygulanan yöntemler; a) kabuk altına miselyal agar disk inokulasyonu, b) tepeden agar disk inokulasyonu, c) ksileme enjeksiyon yöntemi ve d) yeşil sürgünlere agar disk inokulasyonudur. Patojenlerin kalemler ve yeşil sürgünlere inokulasyonunu takiben 2 ay süreyle gelişmelerine izin verilmiş ve sonra dokularda oluşan lezyon uzunlukları ölçülmüştür. Sonuçlara göre tüm patojenler tüm inokulasyon yöntemleriyle, hem dormant kalemler hem de yeşil sürgünler üzerinde lezyonlar meydana getirmiş ancak ölçülen lezyon uzunlukları kullanılan yöntemlere göre farklılık göstermiştir. En uzun lezyonların oluştuğu patojen ve inokulasyon metodu eşleşmesi şu şekilde gerçekleşmiştir; Diaporthe ampelina için yeşil sürgüne miseliyal agar disk inokulasyonu (72.2 mm), D. macrodidyma ve P. chlamydospora için ksileme enjeksiyon (sırasıyla 23.0 ve 66.1 mm) ve Neofusicoccum parvum için kabul altına miseliyal agar disk inokulasyonu (51.5 mm).Genel Yorum: Asmalarda fungal gövde hastalıklarının araştırılmasında birçok araştırmacı tarafından farklı inokulasyon yöntemleri kullanılsa da, patojenisite testlerinin bu çalışmada denenen yöntemlerden herhangi biri ile yapılabileceği ortaya koyulmuştur. Çok sayıda spor üreten türlerle yapılan çalışmalarda, ksileme enjeksiyon yönteminin daha makul sonuçlar vereceği görülmüştür. Miselyal gelişimi güçlü ve piknit üreten türler için ise kabuk altına ya da tepeden agar disk inokulasyon yöntemlerinin daha uygun olduğu anlaşılmıştır.Çalışmanın Önemi ve Etkisi: Bir araştırmada, inokulasyon yönteminin seçimi o araştırmanın sonuçlarını doğrudan etkilemektedir. Bu çalışmanın sonuçlarının, asma gövde hastalıkları ile yapılacak araştırmalarda uygun bir inokulasyon yönteminin seçimine yardımcı olabileceği düşünülmektedir.

Grapevine trunk disease in European and Mediterranean vineyards: occurrence, distribution and associated disease-affecting cultural factors

A survey was carried out in 22 European and some non-European Mediterranean countries in 2015 and 2016, to gain insights into the main fungal grapevine trunk diseases (GTDs) in Europe. Information was obtained from 105 viticulture regions, representing approx. four million ha of vineyards. Vintage and table wines are the main industry products, followed by nursery plants and table grapes. Diverse scion cultivars are grown, with 13 cultivars providing 50% of survey respondent production, and Riesling Italico, Chardonnay, Cabernet Sauvignon and Merlot predominating. The five most used rootstocks are SO4, 110R, K5BB, 41B and 1103P, together representing about 50% of survey respondent production. Despite the diversity of grapevine training, Guyot and cordon remain the most widespread methods, and grapevines are mainly hand-pruned. The use of pneumatic shears and mechanical pruning is becoming more common in some countries. The survey questionnaire proposed a simplified set of symptoms or diseases occurring in mature vineyards [apoplexy, esca complex leaf symptoms (including Grapevine Leaf Stripe Disease, GLSD), dead cordon, Eutypa related dieback, Botryosphaeria or unidentified agent dieback, Phomopsis cane and leaf spot]. Apoplexy and esca/GLSD, defining the esca complex, were the most frequent and increasing syndromes in almost all countries, except Israel and the United Kingdom, where these diseases are not present or not recorded in the survey. Dead cordon and Phomopsis cane and leaf spot were mentioned as occurring in a large number of regions but not frequently. Within individual countries, the profile of GTDs varied according to the region, and to pedo-climatic and production conditions. GTDs on young vines were uncommon, but remain a subject of concern in some of European countries with large nursery production. This overview on spread and relevance of GTDs in Europe and related aspects will be a useful starting point for policy makers and for collaborative research on factors contributing to the increasing disease incidence of GTDs.

Genetic Variation in Resistance to Valsa canker is Related to Arbutin and Gallic Acid Content in Pyrus bretschneideri

Pear Valsa canker is a fungal trunk disease caused by Valsa pyri. Phenolic compounds are ubiquitous in plants and usually contribute to plant resistance against biotic stress. To investigate the association between phenolic compounds and level of resistance to V. pyri, we quantified the contents of individual phenolic compounds in the cortex and phloem of stems from 8 cultivars of Pyrus bretschneideri. Significant variation in the levels of all compounds was found among the cultivars. Correlation analysis revealed an inverse correlation between levels of arbutin and gallic acid with the degree of canker resistance. This suggested that these phenolic compounds are beneficial to V. pyri infection. These data could be valuable for breeding cultivars of P. bretschneideri with greater resistance to V. pyri.

High-throughput amplicon sequencing-based analysis of active fungal communities inhabiting grapevine after hot-water treatments reveals unexpectedly high fungal diversity

Abstract The ecology of total fungal communities in grapevine is so far largely derived from studies on culture-dependent methods or cultivation-independent rDNA approaches. Sequencing the ribosomal RNA transcripts (rRNA) would rather reveal the functionally and metabolically active important taxa of the fungal community and provide insights into its activity in the wood. The present study investigated changes in the potentially active fungal communities of internal grapevine wood after Hot-Water Treatment (HWT) in planting material from Czech Republic and Spain at two different times during the propagation process and from two plant zones. We examined fungal communities using both traditional isolation into culture and high-throughput amplicon sequencing (HTAS) of the internal transcribed spacer 2 (ITS2) region in extracted total RNA. HTAS from metatranscriptomic RNA increased the resolution of the fungal community analysis and revealed a highly diverse mycobiota of grapevine wood compared to the traditional method. Fungal diversity differed between grapevine genotypes and showed a temporal variation over the vegetative period. Grapevine planting materials exhibited high fungal diversity after HWT, which demonstrates that the HWT process does not sterilize the internal wood of grapevine. HWT reduced the infection caused by fungal trunk disease pathogens but was not completely effective in eliminating their growth. This study provides important and practically useful insights into the dynamics of active fungal communities in hot-water treated plants, and represents the first study of active fungal communities on grapevine grafted plants by comparing traditional and next-generation sequencing methods.

Managing Grapevine Trunk Diseases With Respect to Etiology and Epidemiology: Current Strategies and Future Prospects.

Fungal trunk diseases are some of the most destructive diseases of grapevine in all grape growing areas of the world. Management of GTDs has been intensively studied for decades with some great advances made in our understanding of the causal pathogens, their epidemiology, impact, and control. However, due to the breadth and complexity of the problem, no single effective control measure has been developed. Management of GTD must be holistic and integrated, with an interdisciplinary approach conducted in both nurseries and vineyards that integrates plant pathology, agronomy, viticulture, microbiology, epidemiology, biochemistry, physiology, and genetics. In this review, we identify a number of areas of future prospect for effective management of GTDs worldwide, which, if addressed, will provide a positive outlook on the longevity of vineyards in the future.

Fungal trunk diseases: a problem beyond grapevines?

Grapevine trunk diseases (GTDs) are caused by a broad range of taxonomically unrelated fungi that occur wherever grapes are grown and are the main biotic factor limiting vineyard productivity and longevity (Bertsch et al., 2013). GTDs cause untenable economic losses to the grapevine industry worldwide. For example, they are considered a ‘national crisis’ in France, where it has been estimated that 12% of the vineyards are currently economically nonviable due to these maladies causing losses of about €1 billion (Lorch, 2014). Because GTDs are chronic and there is currently no option to eradicate the infections, unproductive vineyards must be replanted, at a worldwide annual cost estimate of €1·132 billion (Hofstetter et al., 2012). The establishment, progress, severity and spread of GTDs have been associated with (i) drought, (ii) limited availability of effective fungicides, (iii) pressure to increase yields, (iv) lack of pruning wound protection due to cost of labour, and/or (v) poor low-quality propagation material. Growers are demanding solutions. Many vineyards with high GTD incidence were planted in the mid-to-late 1990s, during which wine-grape prices were increasing (Volpe et al., 2008). These vineyards were being replanted in the wake of phylloxera, and nurseries could barely meet the demand for grape plants. In 2009 researchers declared the worldwide impact of GTDs and prioritized research goals (Bertsch et al., 2009). Despite scientific advances, there are still unanswered questions, for example about how the pathogens spread and which climate conditions favour infection. This limits the efficacy of GTD management practices. Other crops, such as almond, pistachio and walnut, are now experiencing planting booms (INC, 2015). The significance of these and other tree crops is that they host some of the same GTD pathogens. While researchers fine-tune strategies for managing GTDs in grape, the pathogens may alternate to tree crops, thus endangering the productivity and longevity of orchards and vineyards alike. Recent reports of high disease incidence in pome and stone fruits, nut crops, small fruits and olive in California (Úrbez-Torres et al., 2013), Iran (Mohammadi et al., 2015), Italy (Carlucci et al., 2015), South Africa (Cloete et al., 2011) and Spain (Gramaje et al., 2012) highlight the need for a focus on this new set of hosts. The production systems are different; not all practices for GTD management in grape are feasible in tree crops, especially with dwindling numbers of skilled farm labourers. Tree crop nurseries are struggling to meet demands for plant material and, from our experience with GTDs, a lack of first-quality plants means that low-quality plants are sold, too. Are these plantings doomed to a lifetime of poor productivity? Is the sustainability of the fruit and tree nut industries at risk? Yours sincerely, David Gramaje et al.

Proteomic insights into changes in grapevine wood in response to esca proper and apoplexy

Among fungal grapevine trunk diseases, esca proper poses a significant threat for viticulture. Apoplexy, mainly occurring on grapevines affected by esca proper, is also a threat. To verify if different responses are activated in the woody tissues of apoplectic (A) and esca proper-affected (E) vines, two-dimensional gel electrophoresis coupled to mass spectrometry analysis was used to examine changes in the trunk wood of E and A field-grown plants. Asymptomatic and black streaked trunk (BST) wood from A and E plants were compared to asymptomatic and BST wood of control plants. Twenty-seven differentially expressed protein spots were identified. For eleven targeted proteins, expression of the relative transcripts was also monitored by qRT-PCR. Hierarchical tree clustering revealed differences in the distribution of spots containing carbohydrate metabolism proteins and heat shock proteins between asymptomatic- and BST-wood of grapevine, irrespective of the type of plant examined (control or diseased grapevines). Asymptomatic wood was mainly characterized by down-expression of proteins involved in cell growth and defense responses. The proteome of BST wood, characterized by extensive presence of grapevine trunk disease agents, revealed over-expression of proteins involved in defense. There was no evidence of strong different response in the trunk wood of apoplectic and esca proper affected plants. This could mean that, despite the different feature of these external crown symptoms, grapevine responses at trunk wood level is very similar in both cases. This plant response might therefore either simply be due to the fact that plants can react in the same way to different stresses, or that apoplexy represents a different effect provoked by the same cause or causal agent(s).

Fungi associated to grapevine trunk diseases in young plants in Asturias (Northern Spain)

Dark discolorations of the vascular vessels of 3-year-old potted plants of Asturian grapevine cultivar (Northern Spain), were observed during pruning. These symptoms can be associated to fungal trunk diseases that, in the last decades, are affecting young vineyards all over the world. Cross sections from root, trunk and canes of 19 young grapevine plants were analyzed for the presence of pathogenic fungi associated with these diseases. Non-pathogenic fungi were isolated from both asymptomatic and symptomatic samples, showing that dark discolorations, in some cases, were a consequence of abiotic causes. Regarding pathogenic fungi, Cylindrocarpon spp. colonies were the most frequent, isolated mainly from roots and from asymptomatic tissue. Botryosphaeria spp. colonies were mostly isolated from trunk and from sections with dark discolorations generated by pruning. Phaeoacremonium spp. was isolated from all the organs (roots, trunk and canes). Only one colony of Libertella spp. was isolated. These results suggest the need of a standard protocol, combining treatments and management activities, to be performed in nurseries to limit the spread of these diseases.    

Efficacy of paste and liquid fungicide formulations to protect pruning wounds against pathogens associated with grapevine trunk diseases in Chile

Trunk diseases are important on grapevines (Vitis vinifera L.) in most countries where grapevines are grown. In Chile, Diplodia seriata, Inocutis sp. and Phaeomoniella (Pa.) chlamydospora are the most prevalent pathogens associated with this syndrome. It has been demonstrated that fresh pruning wounds are the main infection route for fungal trunk disease pathogens. The objective of this study was to evaluate the efficacy of paste and spray fungicide applications in protecting pruning wounds against D. seriata, Inocutis sp. and Pa. chlamydospora on ‘Cabernet Sauvignon’ grapevines in two important Chilean grapevine production areas. The fungicides used in this study were benomyl, pyraclostrobin, tebuconazole, and thiophanate-methyl, which were applied as paste and liquid fungicide on fresh pruning wounds pre- and post-inoculation with mycelium plugs of the different fungal pathogens used in this study. The efficacy of the fungicides was assessed by measuring the length of the discoloration of the vascular system and percent re-isolation obtained on fresh pruning wounds. The results obtained demonstrate that infections caused by D. seriata, Inocutis sp. and Pa. chlamydospora can be significantly reduced using both paste and spray applications of benomyl, pyraclostrobin, tebuconazole and thiophanate-methyl and that the paste formulations provided a better control than spray applications. Regardless of the fungicide and application method used, the pre-inoculation treatments (performed 24 h before inoculation) resulted in better control than the 24 h post-inoculation treatments. The current study showed that Inocutis sp. can induce vascular discoloration on grapevine spurs. This study supports a broader use of benzimidazole fungicides for the control of the devastating trunk diseases in all grape productions worldwide.

Contribution for a better understanding of grapevine fungal trunk diseases in the Portuguese Dão wine region

Summary. Esca and Petri disease, two of the most important fungal trunk diseases of grapevine, are responsible for significant losses by causing premature decline and dieback in vineyards worldwide. The Portuguese Dao wine region is no exception. Local winegrowers’ knowledge on Grapevine Trunk Diseases (GTD) in general and of esca and Petri disease in particular is incomplete. The real scope of those problems has been based largely on individual perceptions rather than on a methodical evaluation of the situation. In order to get a full picture of the diseases impact, a leaflet with color pictures was produced and issued to winegrowers, accompanied by a simple questionnaire. The results of this survey represent a first indication of the extent of grapevine trunk diseases in the Dao wine region, specifically its economic impact and relevance to the local wine industry. In conjunction with the survey, several samples of wood collected from esca and Petri disease symptomatic vines, identified during the survey throughout the entire region, were processed and a collection of isolates of Phaeomoniella chlamydospora obtained. To determine the intra-specific variability among these isolates, morphological, cultural and molecular characteristics were evaluated. A protocol to study the pathogenicity with P. chlamydospora was conducted, consisting on the inoculation of cv. Touriga Nacional’s spurs with a previously studied P. chlamydospora isolate.

Transformation of Fungal Grapevine Trunk Disease Pathogens with the Green Fluorescent Protein Gene

Eutypa lata and Phaeomoniella chlamydospora are important fungal grapevine trunk disease pathogens. Studies of their epidemiology and disease progression are hampered by the slow growth rates of the fungi and a lack of molecular tools. A protoplast-based transformation system was developed using hygromycin resistance as a positive selectable marker. The system was used to introduce a green fluorescent protein reporter gene that was expressed in the hyphae of both species. The reporter strains will be useful tools for further epidemiological studies. Furthermore the transformation system will allow for genetic dissection of candidate pathogenicity or virulence genes by targeted replacement or disruption, in order to discern the role of these genes in the disease process.