THE DESTRUCTION OF DIDYMELLA LYCOPERSICI KLEB. IN TOMATO HAULM COMPOSTS

Experiments have been carried out to assess the possibility of the destruction by composting of mycelium of Didymella lycopersici in diseased tomato haulms. The results of laboratory trials with agar cultures and with infested tomato stems and fruits suggest that the fungus is destroyed if kept under moist conditions at 35° C. for from 3 to 6 days. Agar cultures buried for approximately this length of time in composting material at this or higher temperatures were dead when recovered. When composts made from haulms of tomatoes affected by Didymella stem rot were tested by placing them round the stem bases of tomato plants, or by growing tomato plants in them, few of the test plants became diseased. It was concluded that D. lycopersici present in tomato refuse could be destroyed by careful composting.     

The role of mineral nutrients in the increased growth response of tomato plants in solarized soil

Soil solarization is a non-chemical disinfestation technique that frequently promotes plant growth in the absence of known major pathogens, a phenomenon termed increased growth response (IGR). The effect of solarization on plant nutrients and their role in the IGR was studied with tomato plants grown in solarized or non-solarized (control) sandy soil, under controlled conditions. Solarization considerably increased the soil concentrations of water extractable N, K, Ca, Mg and Na at most sites, whereas Cl and DTPA extractable Mn, Zn, Fe and Cu were decreased by the treatment. Plant growth and specific leaf area were enhanced in solarized as well as in N-supplemented control soil. In tomato plants grown in solarized soil, concentrations of most nutrients in the xylem sap, including N, were increased compared to the control, whereas Cl and SO4 levels decreased. The most significant increase in leaf nutrient concentration caused by soil solarization was recorded for N. Furthermore, leaf N concentration was highly and positively correlated with shoot growth. The concentration of Cu increased in leaves from the solarization vs. the control treatment, whereas that of SO4 and Cl decreased, the latter presumably below the critical toxicity level. The correlation between shoot growth and leaf concentration was positive for Cu and inverse for Cl and SO4. In conclusion, we found that soil solarization significantly affects nutrient composition in tomato plants, and provided strong evidence that N, and eventually also Cl, play a major role in IGR.     

A highly conserved effector in Fusarium oxysporum is required for full virulence on Arabidopsis

Secreted-in-xylem (SIX) proteins of the vascular wilt pathogen Fusarium oxysporum f. sp. lycopersici are secreted during infection of tomato and function in virulence or avirulence. F. oxysporum formae speciales have specific host ranges but the roles of SIX proteins in diverse hosts are unknown. We identified homologs of F. oxysporum f. sp. lycopersici SIX1, SIX4, SIX8, and SIX9 in the genome of Arabidopsis infecting isolate Fo5176. A SIX4 homolog (termed Fo5176-SIX4) differed from that of F. oxysporum f. sp. lycopersici (Fol-SIX4) by only two amino acids, and its expression was induced during infection of Arabidopsis. Transgenic Arabidopsis plants constitutively expressing Fo5176-SIX4 had increased disease symptoms with Fo5176. Conversely, Fo5176-SIX4 gene knock-out mutants (Δsix4) had significantly reduced virulence on Arabidopsis, and this was associated with reduced fungal biomass and host jasmonate-mediated gene expression, the latter known to be essential for host symptom development. Full virulence was restored by complementation of Δsix4 mutants with either Fo5176-SIX4 or Fol-SIX4. Thus, Fo5176-SIX4 contributes quantitatively to virulence on Arabidopsis whereas, in tomato, Fol-SIX4 acts in host specificity as both an avirulence protein and a suppressor of other race-specific resistances. The strong sequence conservation for SIX4 in F. oxysporum f. sp. lycopersici and Fo5176 suggests a recent common origin.     

Cloning and disruption of pgx4 encoding an in planta expressed exopolygalacturonase from Fusarium oxysporum

Fusarium oxysporum f. sp. lycopersici, the causal agent of tomato vascular wilt, produces an array of pectinolytic enzymes, including at least two exo-alpha1,4-polygalacturonases (exoPGs). A gene encoding an exoPG, pgx4, was isolated with degenerate polymerase chain reaction primers derived from amino acid sequences conserved in two fungal exoPGs. pgx4 encodes a 454 amino acid polypeptide with nine potential N-glycosylation sites and a putative 21 amino acid N-terminal signal peptide. The deduced mature protein has a calculated molecular mass of 47.9 kDa, a pI of 8.0, and 51 and 49% identity with the exoPGs of Cochliobolus carbonum and Aspergillus tubingensis, respectively. The gene is present in a single copy in different formae speciales of F. oxysporum. Expression of pgx4 was detected during in vitro growth on pectin, polygalacturonic acid, and tomato vascular tissue and in roots and stems of tomato plants infected by F. oxysporum f. sp. lycopersici. Two mutants of F. oxysporum f. sp. lycopersici with a copy of pgx4 inactivated by gene replacement were as virulent on tomato plants as the wild-type strain.     

THE EFFECT OF CERTAIN SOIL TREATMENTS ON DIDYMELLA STEM ROT OF TOMATOES PART I. GLASSHOUSE EXPERIMENTS

When tomatoes were planted in steamed soil inoculated with spores of Didymella lycopersici Kleb., a large proportion became infected, but very few became infected in unsteamed soil similarly inoculated. In soil treated with formalin and subsequently inoculated the number of diseased plants was about half that in steamed soil. Reinfecting steamed soil with soil micro-organisms before inoculation with D. lycopersici reduced the amount of disease. Covering steamed soil with a layer of maiden loam filtered out a large proportion of D. lycopersici spores applied in water suspension and reduced the amount of disease. Addition of stable manure either before or after steaming did not affect the number of diseased plants. Steamed soil remained susceptible to infection by D. lycopersici for 4 months. D. lycopersici survived in a glasshouse soil over the winter, but the number of diseased plants was small. Cultivation may lessen the amount of disease by burying most of the infected surface layers of the soil. The peak of infection under glass occurred in May. The amount of disease and the yield were correlated, but with severe infection the yield varied with the time of infection and the distribution of diseased plants.     

Influence of Canopy Height and Fertilization Levels on the Resistance of Lycopersicon Hirsutum to Aculops Lycopersici (Acari: Eriophyidae)

The objective of this work was to study the effect of NK fertilization levels and canopy height on the resistance of Lycopersicon hirsutum and Lycopersicon esculentum to Aculops lycopersici (Acari: Eriophydae). The effects of NK fertilization levels and canopy height in the leaf size and density of trichomes and their effects on tridecan-2-one (2-TD) and undecan-2-one (2-UD) limiting the attack of A. lycopersici on tomato plants were assessed. Different NK fertilization levels had no effect on the resistance of L. hirsutum to A. lycopersici. No significant differences were found in attack rates of this mite on leaves of the top and median parts of L. hirsutum canopy. The type and density of trichomes were the main determining factor of A. lycopersici attack on tomato plants. High trichome densities and type VI glandular trichomes which produce tridecan-2-one are important resistance factors on tomato plants. L. hirsutum showed a high resistance level to A. lycopersici due to high densities of type VI glandular trichomes and consequently higher levels of tridecan-2-one in its leaves.     

THE EFFECT OF CERTAIN SOIL TREATMENTS ON DIDYMELLA STEM-ROT OF TOMATOES

About 93% of Didymella lycopersici spores were destroyed after 4 weeks incubation in unsterilized soil. A survey of the microflora of glasshouse soil receiving different treatments and inoculated with D. lycopersici showed no clear relation between numbers of any group of organisms and the incidence of stem rot. Sterilized soil was not made toxic to D. lycopersici by the growth of a number of soil microorganisms even after 9 months incubation, but addition of unsterilized soil or of a suspension of unsterilized soil quickly restored toxicity. Direct observations of spores in soil on slides showed that their fate varied with the treatment of the soil before inoculation. With fresh soil or air-dry soil moistened 2 or more days before inoculation, lysis of spores occurred. With air-dry soil moistened and inoculated simultaneously, some spores germinated but growth of germ tubes soon ceased. No direct connexion could be seen between the fate of the spores and soil microorganisms. Addition of glucose to unsterilized soil reduced its toxicity to D. lycopersici. Soils steamed for 1 min. or longer were not toxic to D. lycopersici , but soils steamed for very short periods were as toxic as unsterilized soils although the soil microflora was much reduced.     

Genome-wide dissection of<Emphasis Type="Italic"> Fusarium</Emphasis> resistance in tomato reveals multiple complex loci

Resistance to different pathogenic races of Fusarium oxysporum f. sp. lycopersici (F. o. lycopersici) was explored at two genomic levels in tomato. Six independent Fusarium resistance loci were identified by comparing the responses of a complete set of 53 lines carrying different introgressed regions of the Lycopersicon pennellii genome in a L. esculentum background. The loci confer varying degrees of resistance to different races of the pathogen. Corresponding map positions from different tomato species were aligned and in some cases revealed parallel resistance to F. o. lycopersici with qualitative changes in race specificities. One of the loci identified corresponds to the previously characterized complex resistance locus I2, which is involved in resistance to F. o. lycopersici race 2. A novel member of this locus, I2C-5, which belongs to the NBS-LRR family of resistance genes, was cloned and shown to confer partial resistance in transgenic plants. Thus, at a particular complex locus gene members can confer full or partial resistance to F. o. lycopersici race 2. The results of our whole-genome mapping analysis underline the robust independent origin of resistance to a particular disease and demonstrate the conservation of resistance features at syntenic loci, together with the rapid diversification of genes for innate resistance within loci.     

Inhibition of Some Rot Fungi Polygalacturonases by Allium cepa L. and Allium porrum L. Extracts

Extracts of Allium cepa and A. porrum - contain factors that inhibit to various extents polygalacturonases (PGs) produced in vitro by Sclerotinia sclerotiorum, Botrytis cinerea, Fusarium moniliforme, Phoma terrestris, Sclerotium cepivorum, Macrophomina phaseolina, Didymella bryoniae and Phoma lycopersici. The PG inhibition rank changed using leek or onion extract. The inhibition factors are possibly proteins, do not present particular specificity and act against PGs of fungi pathogens and non pathogens for these plant species.     

The presence of a virulence locus discriminates Fusarium oxysporum isolates causing tomato wilt from other isolates

Fusarium oxysporum is an asexual fungus that inhabits soils throughout the world. As a species, F. oxysporum can infect a very broad range of plants and cause wilt or root rot disease. Single isolates of F. oxysporum, however, usually infect one or a few plant species only. They have therefore been grouped into formae speciales (f.sp.) based on host specificity. Isolates able to cause tomato wilt (f.sp. lycopersici) do not have a single common ancestor within the F. oxysporum species complex. Here we show that, despite their polyphyletic origin, isolates belonging to f.sp. lycopersici all contain an identical genomic region of at least 8 kb that is absent in other formae speciales and non-pathogenic isolates, and comprises the genes SIX1, SIX2 and SHH1. In addition, SIX3, which lies elsewhere on the same chromosome, is also unique for f.sp. lycopersici. SIX1 encodes a virulence factor towards tomato, and the Six1, Six2 and Six3 proteins are secreted in xylem during colonization of tomato plants. We speculate that these genes may be part of a larger, dispensable region of the genome that confers the ability to cause tomato wilt and has spread among clonal lines of F. oxysporum through horizontal gene transfer. Our findings also have practical implications for the detection and identification of f.sp. lycopersici.     

Effector gene screening allows unambiguous identification of Fusarium oxysporum f. sp. lycopersici races and discrimination from other formae speciales

During infection of tomato, the fungus Fusarium oxysporum f. sp. lycopersici secretes several unique proteins, called 'secreted in xylem' (Six) proteins, into the xylem sap. At least some of these proteins promote virulence towards tomato and among them, all predicted avirulence proteins that can trigger disease resistance in tomato have been found. In this study, a large, worldwide collection of F. oxysporum isolates was screened for the presence of seven SIX genes ( SIX1 – SIX7 ). The results convincingly show that identification of F. oxysporum formae speciales and races based on host-specific virulence genes can be very robust. SIX1, SIX2, SIX3 and SIX5 can be used for unambiguous identification of the forma specialis lycopersici . In addition, SIX4 can be used for the identification of race 1 strains, while polymorphisms in SIX3 can be exploited to differentiate race 2 from race 3 strains. For SIX6 and SIX7 , close homologs were found in a few other formae speciales , suggesting that these genes may play a more general role in pathogenicity. Host specificity may be determined by the unique SIX genes, possibly in combination with the absence of genes that trigger resistance in the host.     

Tomatinase from Fusarium oxysporum f. sp. lycopersici defines a new class of saponinases.

Plants produce a variety of secondary metabolites, many of which have antifungal activity. Saponins are plant glycosides that may provide a preformed chemical barrier against phytopathogenic fungi. Fusarium oxysporum f. sp. lycopersici and other tomato pathogens produce extracellular enzymes known as tomatinases, which deglycosylate alpha-tomatine to yield less toxic derivatives. We have cloned and characterized the cDNA and genomic DNA encoding tomatinase from the vascular pathogen of tomato F. oxysporum f. sp. lycopersici. This gene encodes a protein (FoTom1) with no amino acid sequence homology to any previously described saponinase, including tomatinase from Septoria lycopersici. Although FoTom1 is related to family 10 glycosyl hydrolases, which include mainly xylanases, it has no detectable xylanase activity. We have overexpressed and purified the protein with a bacterial heterologous system. The purified enzyme is active and cleaves alpha-tomatine into the less toxic compounds tomatidine and lycotetraose. Tomatinase from F. oxysporum f. sp. lycopersici is encoded by a single gene whose expression is induced by alpha-tomatine. This expression is fully repressed in the presence of glucose, which is consistent with the presence of two putative CREA binding sites in the promoter region of the tomatinase gene. The tomatinase gene is expressed in planta in both roots and stems throughout the entire disease cycle of F. oxysporum f. sp. lycopersici.     

The arms race between tomato and Fusarium oxysporum

The interaction between tomato and Fusarium oxysporum f. sp. lycopersici has become a model system for the study of the molecular basis of disease resistance and susceptibility. Gene-for-gene interactions in this system have provided the basis for the development of tomato cultivars resistant to Fusarium wilt disease. Over the last 6 years, new insights into the molecular basis of these gene-for-gene interactions have been obtained. Highlights are the identification of three avirulence genes in F. oxysporum f. sp. lycopersici and the development of a molecular switch model for I-2, a nucleotide-binding and leucine-rich repeat-type resistance protein which mediates the recognition of the Avr2 protein. We summarize these findings here and present possible scenarios for the ongoing molecular arms race between tomato and F. oxysporum f. sp. lycopersici in both nature and agriculture.     

Most AAL toxin-sensitive Nicotiana species are resistant to the tomato fungal pathogen Alternaria alternata f. sp. lycopersici

The phytopathogenic fungus Alternaria alternata f. sp. lycopersici produces AAL toxins required to colonize susceptible tomato (Lycopersicon esculentum) plants. AAL toxins and fumonisins of the unrelated fungus Fusarium moniliforme are sphinganine-analog mycotoxins (SAMs), which are toxic for some plant species and mammalian cell lines. Insensitivity of tomato to SAMs is determined by the Alternaria stem canker gene 1 (Asc-1), and sensitivity is associated with a mutated Asc-1. We show that SAM-sensitive species occur at a low frequency in the Nicotiana genus and that candidate Asc-1 homologs are still present in those species. In Nicotiana spp., SAM-sensitivity and insensitivity also is mediated by a single codominant locus, suggesting that SAM-sensitive genotypes are host for A. alternata f. sp. lycopersici. Nicotiana umbratica plants homozygous for SAM-sensitivity are indeed susceptible to A. alternata f. sp. lycopersici. In contrast, SAM-sensitive genotypes of Nicotiana spegazzinii, Nicotiana acuminata var. acuminata, Nicotiana bonariensis, and Nicotiana langsdorffii are resistant to A. alternata f. sp. lycopersici infection concomitant with localized cell death. Additional (nonhost) resistance mechanisms to A. alternata f. sp. lycopersici that are not based on an insensitivity to SAMs are proposed to be present in Nicotiana species.     

Evaluation of Steam and Soil Solarization for Meloidogyne arenaria Control in Florida Floriculture Crops

Steam and soil solarization were investigated for control of the root-knot nematode Meloidogyne arenaria in 2 yr of field trials on a commercial flower farm in Florida. The objective was to determine if preplant steam treatments in combination with solarization, or solarization alone effectively controlled nematodes compared to methyl bromide (MeBr). Trials were conducted in a field with naturally occurring populations of M. arenaria. Treatments were solarization alone, steam treatment after solarization using standard 7.6-cm-diameter perforated plastic drain tile (steam 1), steam treatment following solarization using custom-drilled plastic drain tile with 1.6-mm holes spaced every 3.8 cm (steam 2), and MeBr applied at 392 kg/ha 80:20 MeBr:chloropicrin. Drain tiles were buried approximately 35 cm deep with four tiles per 1.8 by 30 m plot. Steam application followed a 4-wk solarization period concluding in mid-October. All steam was generated using a Sioux propane boiler system. Plots were steamed for sufficient time to reach the target temperature of 70°C for 20 min. Solarization plastic was retained on the plots during steaming and plots were covered with a single layer of carpet padding to provide additional insulation. The floriculture crops larkspur (Delphinium elatum and Delphinium × belladonna), snapdragon (Antirrhinum majus), and sunflower (Helianthus annuus) were produced according to standard commercial practices. One month after treatment in both years of the study, soil populations of M. arenaria were lower in both steam treatments and in MeBr compared to solarization alone. At the end of the season in both years, galling on larkspur, snapdragon, and sunflowers was lower in both steam treatments than in solarization. Both steam treatments also provided control of M. arenaria in soil at the end of the season comparable to, or exceeding that provided by MeBr. Both steam treatments also reduced M. arenaria in snapdragon roots comparable to, or exceeding control with MeBr. Meloidogyne arenaria in soil increased in solarization alone. Solarization alone also had higher gall ratings on larkspur, snapdragon, and sunflower than all other treatments. Steam provided excellent control of M. arenaria in this study.     

Field evaluation of plant growth-promoting Rhizobacteria amended transplant mixes and soil solarization for tomato and pepper production in Florida

Field trials were performed in Florida to evaluate tomato and pepper transplants amended with formulations of several plant growth-promoting rhizobacteria (PGPR) in a production system that included soil solarization. Transplants grown in five different formulations of PGPR were planted into plots treated by soil solarization, MeBr fumigation, or untreated soil. Treatments were assessed for incidence of several naturally occurring tomato and pepper pathogens including root-knot nematode (Meloidogyne incognita) and species of Pythium, Phytophthora, and Fusarium. Highly significant increases in tomato and pepper transplant growth occurred in response to most formulations of PGPR tested. Transplant vigor and survival in the field were improved by PGPR treatments in both tomato and pepper. Diseases of tomato caused by root-knot nematodes, Fusarium, Phytophthora, and Pythium were not affected by PGPR treatments. PGPR formulation LS261 reduced numbers of root-knot nematode galls on pepper while pepper root condition was improved with formulations LS213, LS256 and LS261. Individual PGPR strains affected the number of Pythium colonies isolated from pepper roots, but did not affect isolation of Pythium from tomato roots. Greater numbers of colonies of Pythium were isolated from pepper roots in the MeBr treatment and fewest in the solarization treatment. Numbers of colony forming units of Fusarium were significantly higher in the untreated soil than in MeBr fumigated or solarized soil with no effect of PGPR on isolation of Fusarium from either crop. Incidence of wilt symptoms on tomato was significantly lower in MeBr treated plots and highest in the untreated plots. Yield of extra large tomato fruit and total yield increased with PGPR formulation LS256. Yield of pepper was increased with formulations LS255 and LS256. Solarization combined with LS256 on pepper produced yields comparable to MeBr.     

QTLs for tomato powdery mildew resistance (Oidium lycopersici) in Lycopersicon parviflorum G1.1601 co-localize with two qualitative powdery mildew resistance genes

Tomato (Lycopersicon esculentum) is susceptible to the powdery mildew Oidium lycopersici, but several wild relatives such as Lycopersicon parviflorum G1.1601 are completely resistant. An F2 population from a cross of Lycopersicon esculentum cv. Moneymaker x Lycopersicon parviflorum G1.1601 was used to map the O. lycopersici resistance by using amplified fragment length polymorphism markers. The resistance was controlled by three quantitative trait loci (QTLs). Ol-qtl1 is on chromosome 6 in the same region as the Ol-1 locus, which is involved in a hypersensitive resistance response to O. lycopersici. Ol-qtl2 and Ol-qtl3 are located on chromosome 12, separated by 25 cM, in the vicinity of the Lv locus conferring resistance to another powdery mildew species, Leveillula taurica. The three QTLs, jointly explaining 68% of the phenotypic variation, were confirmed by testing F3 progenies. A set of polymerase chain reaction-based cleaved amplified polymorphic sequence and sequence characterized amplified region markers was generated for efficient monitoring of the target QTL genomic regions in marker assisted selection. The possible relationship between genes underlying major and partial resistance for tomato powdery mildew is discussed.