Fom Race Research Articles

Development and Evaluation of a New Source of Tolerance to Fusarium Wilt Race 1.2 in Melon

Melon Fusarium wilt, caused by Fusarium oxysporum f. sp. melonis (Fom), is one of the most prevalent fungal diseases affecting melon (Cucumis melo L.). We aimed at finding an effective resource for breeding cultivars tolerant to Fom race 1.2, which includes pathotypes that cause yellowing (race 1.2y) and wilting (race 1.2w). We screened 294 melon accessions that originated mainly in Africa and Asia for tolerance to race 1.2y. The highest level of tolerance was observed in the Indian accession PI124550; one of the five PI124550 plants tested showed no disease symptoms. From this plant, a tolerant inbred line, YR01, was developed after seven rounds of selfing combined with selection for tolerance to race 1.2y. YR01 had a high level of tolerance not only to race 1.2y, but also to race 1.2w. It was more tolerant to race 1.2y than the tolerant reference control ‘Isabelle’ and was equally tolerant to race 1.2w. Analysis of F1 hybrids and an F2 population developed by crossing YR01 with susceptible ‘Earl’s Favorite Harukei 3’ suggested that YR01 had multiple recessive and dominant (or codominant) genes for tolerance to race 1.2y and one dominant gene for tolerance to race 1.2w. These results indicate that PI124550 and its derivative YR01 are a promising breeding material with novel genes conferring practically useful tolerance to both pathotypes of race 1.2.

Pathotype diversity of Fusarium oxysporum f. sp. mungcola causing wilt on mungbean (Vigna radiata)

Fusarium wilt, caused by Fusarium oxysporum f. sp. mungcola (Fom), is an increasingly serious disease of mungbean (Vigna radiata (L.) R.Wilczek) in China. Pathogenic variability has been observed among Fom isolates; however, there are no reports describing Fom races or pathotypes. Thus, this study was conducted with the aim of developing a set of pathotype differentials to reveal Fom pathotype diversity by assessing virulence variability of Fom isolates. First, 105 mungbean cultivars were screened against a standard virulent Fom isolate (F08). Eleven of the 105 cultivars were selected as candidate differentials of Fom according to resistance phenotype and genetic background. Second, the resistance of the 11 candidate differential cultivars was tested against 30 Fom isolates from different geographical origins in China. Highly significant differences were observed among isolate × cultivar interaction patterns, indicating that pathotype differentiation exists in Fom isolates. Based on the different reaction patterns combining with genetic background, seven of the 11 cultivars were selected to constitute a set of differential hosts of Fom pathotype, used to distinguish pathotypes of 84 Fom isolates from different geographical regions by evaluating the virulence reaction pattern. The results showed that the 84 Fom isolates were defined as 12 pathotypes. Finally, we tried to confirm whether the 12 Fom pathotypes could be distinguished by a PCR-based diagnostic method based on the two genes (SIX6 and SIX11) reported to be associated with Fom pathogenicity. However, the Fom pathotype could not be distinguished by variation of the PCR products or their resulting sequences of the two genes. This is the first study to develop a set of Fom pathotype differential hosts and identify 12 Fom pathotypes, which provides important information for resistance breeding and disease control.

Construction of a genome-anchored, high-density genetic map for melon (Cucumis melo L.) and identification of Fusarium oxysporum f. sp. melonis race 1 resistance QTL.

Four QTLs and an epistatic interaction were associated with disease severity in response to inoculation with Fusarium oxysporum f. sp. melonis race 1 in a recombinant inbred line population of melon. The USDA Cucumis melo inbred line, MR-1, harbors a wealth of alleles associated with resistance to several major diseases of melon, including powdery mildew, downy mildew, Alternaria leaf blight, and Fusarium wilt. MR-1 was crossed to an Israeli cultivar, Ananas Yok'neam, which is susceptible to all of these diseases, to generate a recombinant inbred line (RIL) population of 172 lines. In this study, the RIL population was genotyped to construct an ultra-dense genetic linkage map with 5663 binned SNPs anchored to the C. melo genome and exhibits the overall high quality of the assembly. The utility of the densely genotyped population was demonstrated through QTL mapping of a well-studied trait, resistance to Fusarium wilt caused by Fusarium oxysporum f. sp. melonis (Fom) race 1. A major QTL co-located with the previously validated resistance gene Fom-2. In addition, three minor QTLs and an epistatic interaction contributing to Fom race 1 resistance were identified. The MR-1×AY RIL population provides a valuable resource for future QTL mapping studies and marker-assisted selection of disease resistance in melon.

Plant-pathogen interactions controlled by theFom-1/PrvandFom-2loci of melon

We cloned the genomic locus for melon resistance towards Fusarium oxysporum f.sp. melonis (FOM) races 0 and 1, and the Papaya ring spot virus (PRSV). The two adjacent genes encode proteins of the NBL (nucleotide binding site-leucine rich repeat) family; Prv carries an extra NB domain, and both have alternative, differentially expressed splice variants. The genes' expression patterns were studied using a promoter-reporter system in transgenic melon roots: both genes were expressed around the vascular tissue and in root tips. In our quest for possible fungal proteins that could interact with Fom-2, another resistance gene, we isolated FOM homologs of secreted in xylem (six) genes that, in Fusarium oxysporum f.sp. lycopersici, encode Avr factors. Two such homologs were found, but their distribution among FOM races did not fit the “gene for gene” model. We performed a proteomic analysis of the xylem sap of melon plants that were infected with FOM-0. A total of 513 melon protein and 9 fungal proteins could be annotated against the melon and FOM genomes, respectively. Many of the melon proteins were up-regulated by inoculation.

Two genes for resistance to Fusarium oxysporum f. sp. radicis-cucumerinum in melon (Cucumis melo, Cucurbitaceae)

Fusarium oxysporum f. sp. radicis-cucumerinum (fusarium crown rot, Forc) is an emerging pathogen of melons, Cucumis melo. Forc incites rotting of the plant crown, causing the plants to die prematurely. Most melon accessions are susceptible to this pathogen but the cantaloupe ‘Hemed’ is highly resistant. ‘Hemed’ was crossed with the susceptible muskmelon ‘Dulce’ to ascertain the mode of inheritance of resistance to Forc. Parental-, filial-, and backcross-generation plants were grown under controlled conditions and inoculated with Forc by using a root-dip methodology. Plants were scored as resistant or susceptible at 18 days post inoculation (dpi), when all plants of the susceptible parent ‘Dulce’ were dead or showed severe disease symptoms. At 18 dpi, plants of ‘Hemed’, the F1, and the backcross to ‘Hemed’ were resistant. The F2 population segregated in accordance with a 13:3 ratio, deviating significantly from a 3:1 ratio of resistant to susceptible. The backcross to ‘Dulce’ segregated in accordance with a 1:1 ratio of resistant to susceptible. These results indicate that ‘Hemed’ carries two genes for resistantance to Forc, one dominant and one recessive, which are herein designated, respectively, as Forc-1 and forc-2. Resistance to Forc is not co-inherited with resistance to F. oxysporum f. sp. melonis (fusarium wilt, Fom) races 0, 1, and 2, as both ‘Hemed’ and ‘Dulce’ are resistant to Fom races 0 and 2 and susceptible to Fom race 1.

Development of a Fusarium oxysporum f. sp. melonis functional GFP fluorescence tool to assist melon resistance breeding programmes

Fusarium wilt, caused by Fusarium oxysporum f. sp. melonis (Fom), is one of the most widespread and devastating melon diseases. This vascular disease is caused by the colonization of melon xylem vessels by any of the four Fom races reported (r0, r1, r2 and r1,2, subdivided into r1,2w and r1,2y). The macroscopic evaluation of disease symptoms (disease rating, DR) at several days post‐inoculation (dpi) with Fom spores has been the traditional method to determine the resistance of melon accessions to this fungal pathogen. In this study, one isolate from each Fom race was transformed by Agrobacterium tumefaciens to constitutively express the green fluorescent protein (GFP). Fom‐GFP transformants, as virulent as the corresponding wildtype races, were selected to develop an inoculation assay based on the non‐invasive evaluation of the fluorescence emitted by Fom‐GFP. It was determined that melon root neck was the appropriate area to follow Fom‐GFP and a fluorescence signal rating (FSR) was established in parallel to DR determination. This method allowed the evaluation of GFP signal in the root neck of inoculated melon seedlings at 11–15 dpi. The GFP signal was scored in 62 melon accessions/breeding lines inoculated with different Fom‐GFP, followed by evaluation of the macroscopic DR in the aerial part of melon seedlings at 20–28 dpi. Correlation analysis demonstrated a direct and significant relationship between FSR and DR. This method has shown to be an effective and reliable tool that can assist Fom resistance breeding programmes in melon.

Molecular characterization of Fom-1 gene and development of functional markers for molecular breeding of resistance to Fusarium race 2 in melon

Melon Fusarium wilt disease caused by the soil-borne pathogen Fusarium oxysporum f. sp. melonis (Fom) is one of the most devastating melon diseases worldwide. Recently, the Fom-1 gene responsible for resistance against Fom races 0 and 2 was cloned. In this study we amplified, cloned and sequenced full genomic DNA and cDNA of Fom-1 from several melon resistant and susceptible accessions using three pairs of primers designed within this gene. Sequence analyses showed that this gene contains four exons interrupted by three introns. The comparative sequence analysis of the cloned cDNA amplicons from resistant and susceptible genotypes revealed eight nucleotide substitutions, within Fom-1 coding regions, among which four were non-synonymous. RT-PCR revealed that the Fom-1 expression is induced by Fom race 2 inoculation. The Fom-1 predicted protein (FOM-1) exhibits a tripartite modular structure composed of an N-terminal TIR domain, a central NB–ARC domain and a C-terminal LRR domain. FOM-1 from resistant melon accessions share four amino acid differences relative to the FOM-1 protein in susceptible ones. Two amino acid substitutions N56K and R103H were located at the FOM-1 TIR domain and the substitution E385K in the NB–ARC domain. Based on single nucleotide polymorphisms within the coding region of the Fom-1 locus, we have generated two CAPS markers, Fom-1R and Fom-1S. Results from screening various melon accessions clearly demonstrated the usefulness of both functional CAPS markers in the marker-assisted selection for melon breeding programs.

Genetic Diversity and Pathogenicity of Fusarium oxysporum f.sp. melonis Races from Different Areas of Italy

AbstractFusarium wilt caused by Fusarium oxysporum f.sp. melonis (FOM) is a devastating disease of melon worldwide. Pathogenicity tests performed with F. oxysporum isolates obtained from Italian melon‐growing areas allowed to identify thirty‐four FOM isolates and the presence of all four races. The aims of this work were to examine genetic relatedness among FOM isolates by race determination and to perform phylogenetic analyses of identified FOM races including also other formae speciales of F. oxysporum of cucurbits. Results showed that FOM race 1,2 was the most numerous with a total of eighteen isolates, while six and nine isolates were identified as race 0 and 1, respectively, and just one isolate was assigned to race 2. Phylogenetic analysis was performed by random amplified polymorphic DNA (RAPD) profiling and by translation elongation factor‐1α (TEF‐1α) sequencing. The analysis of RAPD profiles separated FOM races into two distinct clades. Clade 1, which included races 0, 1 and 1,2, was further divided into ‘subclade a’ which grouped almost all race 1,2 isolates, and into ‘subclade b’ which included race 0 and 1 isolates. Clade 2 comprised only race 2 isolates. The phylogenetic analysis based on TEF‐1α separated FOM from the other formae speciales of F. oxysporum. Also with TEF‐1α analysis, FOM races 0, 1 and 1,2 isolates grouped in one single clade clearly separated from FOM race 2 isolates which grouped closer to F. oxysporum f.sp. cucumerinum. RAPD technique was more effective than TEF‐1α in differentiating FOM race 1,2 isolates from those belonging to the closely related races 0 and 1. Both phylogenetic analyses supported the close relationship between the three different FOM races which might imply the derivation from one another and the different origin of FOM race 2.

Physiological races of Fusarium oxysporum f.sp. melonis in Tunisia

Fusarium wilt of melon, caused by Fusarium oxysporum f.sp. melonis (Fom), is an important disease; races of the pathogen were identified by inoculating differential standard host cultivars. A total of ten isolates that were obtained from 23 fields located in four different geographical regions were identified as pathogenic. Results indicate that all four known Fom races, namely, 0, 1, 2 and 1.2, were found in north and middle Tunisia. Race 1.2 was the most prevalent.

Breeding melon for resistance to Fusarium wilt: recent developments

AbstractMelon Fusarium wilt (MFW), caused byFusarium oxysporumf. sp.melonis(Fom), is one of the most destructive diseases of melon (Cucumis meloL.). The development and deployment of resistant cultivars is generally considered to be the best approach to control MFW. Based on the host resistance genes associated with variants of this pathogen,Fomisolates were classified into four physiological races designated 0, 1, 2, and 1,2. Two dominant resistance genes,Fom-1andFom-2, control resistance to races 0 and 2, and 0 and 1, respectively.Fomisolates classified as race 1,2 are able to induce disease in melon lines carrying the above resistance genes. Many sources of resistance toFomraces 0, 1, and 2 have been reported. Partial resistance to race 1,2 controlled by polygenic recessive genes was only detected in a few Far Eastern melon accessions, except for the breeding line BIZ where complete resistance was described. Identification of DNA markers tightly linked to genes conferring resistance toFomhas immediate application in MFW resistance breeding programs. TheFom-2gene has been cloned, and it encodes a protein with a nucleotide binding site (NBS) and leucine-rich repeats domain (LRR). Based on the sequence of this domain, some molecular markers linked to this gene were developed. Several DNA markers linked toFom-1have also been described. However, the usefulness of these markers was variety-dependent. Therefore, their combined use would be very useful in marker assisted selection for introducing resistance toFomraces 0 and 2 in melon. Recently, these markers were used for the positional cloning of this gene, which encoded a protein with a NBS–LRR domains that shows similarity to the toll and interleukin-1 receptores (TIR). RegardingFomrace 1,2, nine QTL were detected on five linkage groups by composite interval mapping. In this paper we review the current knowledge of MFW disease, and focus on genetic resistance toFomand marker-assisted selection for resistance.

Differential response of Cucumis melo to Fusarium oxysporum f. sp. melonis race 1.2 isolates

Fusarium wilt incited by Fusarium oxysporum f. sp. melonis (Fom) causes severe losses in melon crops. Four physiological races of Fom have been identified: 0, 1, 2 and 1.2. In most cases, resistance to race 1.2 has been described as recessive, polygenic and not race specific. However, some evidence of race-specific effects within melon resistance to race 1.2 has been reported. In this work, we study these effects and assess whether they are due to race-specific resistance. Seeds were obtained from 14 melon accessions that exhibit some level of resistance to race 1.2, and from the lines ‘Charentais-Fom1’ (resistant to races 0 and 2), ‘Charentais-Fom-2’ (resistant to races 0 and 1), and ‘Dinero F1’ (with partial resistance to Fom race 1.2). Melon seedlings were artificially inoculated using two different procedures: ‘continuous shaking’ and ‘tray immersion’. Symptom severity was assessed on leaves using a rating scale from 0 (no symptoms) to 4 (death of the plant). Symptoms were recorded weekly over the four-week period following the first appearance of symptoms and the area under the disease progress curve (AUDPC) was calculated. Six Fom isolates (3 from pathotype Y and 3 from pathotype W) were used in the inoculation. The less aggressive ‘tray immersion’ procedure seems to be more appropriate for detecting the typically small resistance factors of this type of polygenic partial resistance. ‘Kogane Nashi Makuwa’, ‘BG-5384’, ‘Shiro Uri Okayama’, ‘C-211’ and the control, ‘Dinero F1’, showed a high level of resistance to all Fom isolates. However, some genotype × isolate effects were also detected. ‘Baza’, when inoculated with isolate Fom 9302, and ‘Korça, when inoculated with Fom 37mls.1.2W, showed resistance levels similar to that of ‘Dinero F1’; this effect was not observed when ‘Baza’ and ‘Korça’ were inoculated with other isolates. These results are characteristic of race-specific resistance and offer evidence for the presence of this type of resistance to Fom race 1.2 in melons.

Inheritance of race 1.2 Fusarium wilt resistance in four melon cultivars

The resistance to Fusarium oxysporum f.sp. melonis (Fom) race 1.2 has been studied in melons, such as the Portuguese accession ‘BG-5384’ and in the Japanese ‘Shiro Uri Okayama’, ‘Kogane Nashi Makuwa’, and ‘C-211’, since a good characterization of the resistance is necessary before its introgression into commercial varieties. These four melon accessions showed a high level of resistance to races 0, 1, and 2 of Fom, indicating that the partial resistance to the race 1.2 previously detected may not have been race specific. To determine the mode of inheritance of the resistance to Fom race 1.2, the F1, F2, BCPR, and BCPS generations from the crosses between the four resistant accessions above and ‘Piel de Sapo’, a Fom race 1.2 susceptible melon, were developed. They were subsequently inoculated with two Fom isolates, one from the pathotype 1.2Y and the other from the pathotype 1.2W. The area under the disease progress curve was determined for each inoculated plant, and the data were analyzed. We show that the resistance seen in these accessions is polygenically inherited with a complex genetic control because many epistatic interactions were detected. The three epistatic effects; additivity × additivity, dominance × dominance, and dominance × additivity are present and significant, with differing magnitudes from one cross to the next. The relatively low heritabilities, and these epistatic effects make difficult the improvement of the resistance, from these sources, through a standard selection procedure.

Distinct colonization patterns and cDNA-AFLP transcriptome profiles in compatible and incompatible interactions between melon and different races of Fusarium oxysporum f. sp. melonis.

BackgroundFusarium oxysporum f. sp. melonis Snyd. & Hans. (FOM) causes Fusarium wilt, the most important infectious disease of melon (Cucumis melo L.). The four known races of this pathogen can be distinguished only by infection on appropriate cultivars. No molecular tools are available that can discriminate among the races, and the molecular basis of compatibility and disease progression are poorly understood. Resistance to races 1 and 2 is controlled by a single dominant gene, whereas only partial polygenic resistance to race 1,2 has been described. We carried out a large-scale cDNA-AFLP analysis to identify host genes potentially related to resistance and susceptibility as well as fungal genes associated with the infection process. At the same time, a systematic reisolation procedure on infected stems allowed us to monitor fungal colonization in compatible and incompatible host-pathogen combinations.ResultsMelon plants (cv. Charentais Fom-2), which are susceptible to race 1,2 and resistant to race 1, were artificially infected with a race 1 strain of FOM or one of two race 1,2 w strains. Host colonization of stems was assessed at 1, 2, 4, 8, 14, 16, 18 and 21 days post inoculation (dpi), and the fungus was reisolated from infected plants. Markedly different colonization patterns were observed in compatible and incompatible host-pathogen combinations. Five time points from the symptomless early stage (2 dpi) to obvious wilting symptoms (21 dpi) were considered for cDNA-AFLP analysis. After successful sequencing of 627 transcript-derived fragments (TDFs) differentially expressed in infected plants, homology searching retrieved 305 melon transcripts, 195 FOM transcripts expressed in planta and 127 orphan TDFs. RNA samples from FOM colonies of the three strains grown in vitro were also included in the analysis to facilitate the detection of in planta-specific transcripts and to identify TDFs differentially expressed among races/strains.ConclusionOur data suggest that resistance against FOM in melon involves only limited transcriptional changes, and that wilting symptoms could derive, at least partially, from an active plant response.We discuss the pathogen-derived transcripts expressed in planta during the infection process and potentially related to virulence functions, as well as transcripts that are differentially expressed between the two FOM races grown in vitro. These transcripts provide candidate sequences that can be further tested for their ability to distinguish between races.Sequence data from this article have been deposited in GenBank, Accession Numbers: HO867279-HO867981.

Race Distribution, Vegetative Compatibility and Pathogenicity of Fusarium oxysporum f. sp. melonis Isolates in Greece

AbstractRaces and vegetative compatibility groups (VCGs) in Greek isolates of Fusarium oxysporum f. sp. melonis(Fom) were characterized. Three races (0, 2 and 1–2) among 12 isolates tested and two VCGs among 19 isolates tested, were identified. Race 1–2 was the most common and race 1 was not detected. One widespread VCG corresponded to a VCG previously reported from Israel (coded 0138), and included seven isolates of races 0 and 1–2. The other VCG, which was unclassified, included four isolates of races 0, 2 and 1–2. The latter VCG was detected only in a specific melon‐growing location of Evros. The remaining eight isolates tested for VCG did not show positive reactions with other isolates, with each other or with the testers of VCGs 0135 or 0138, although they produced complementary mutants. Using two inoculation methods, the local cv. ‘Golden Head’ was found susceptible to all known Fom races, and especially to race 1–2. These results show the presence of more than one VCG and the widespread distribution of the race 1–2, in Greece.

Atomic integral containing three odd powersof interelectronic separation coordinates

Fusarium wilt incited by Fusarium oxysporum f. sp. melonis (Fom) causes severe losses in melon crops. Four physiological races of Fom have been identified: 0, 1, 2 and 1.2. In most cases, resistance to race 1.2 has been described as recessive, polygenic and not race specific. However, some evidence of race-specific effects within melon resistance to race 1.2 has been reported. In this work, we study these effects and assess whether they are due to race-specific resistance. Seeds were obtained from 14 melon accessions that exhibit some level of resistance to race 1.2, and from the lines ‘Charentais-Fom1’ (resistant to races 0 and 2), ‘Charentais-Fom-2’ (resistant to races 0 and 1), and ‘Dinero F1’ (with partial resistance to Fom race 1.2). Melon seedlings were artificially inoculated using two different procedures: ‘continuous shaking’ and ‘tray immersion’. Symptom severity was assessed on leaves using a rating scale from 0 (no symptoms) to 4 (death of the plant). Symptoms were recorded weekly over the four-week period following the first appearance of symptoms and the area under the disease progress curve (AUDPC) was calculated. Six Fom isolates (3 from pathotype Y and 3 from pathotype W) were used in the inoculation. The less aggressive ‘tray immersion’ procedure seems to be more appropriate for detecting the typically small resistance factors of this type of polygenic partial resistance. ‘Kogane Nashi Makuwa’, ‘BG-5384’, ‘Shiro Uri Okayama’, ‘C-211’ and the control, ‘Dinero F1’, showed a high level of resistance to all Fom isolates. However, some genotype × isolate effects were also detected. ‘Baza’, when inoculated with isolate Fom 9302, and ‘Korça, when inoculated with Fom 37mls.1.2W, showed resistance levels similar to that of ‘Dinero F1’; this effect was not observed when ‘Baza’ and ‘Korça’ were inoculated with other isolates. These results are characteristic of race-specific resistance and offer evidence for the presence of this type of resistance to Fom race 1.2 in melons.