Chitin elicitor receptor kinase 1 (CERK1) is required for the non-host defense response of <Emphasis Type="Italic">Arabidopsis</Emphasis> to <Emphasis Type="Italic">Fusarium oxysporum</Emphasis> f. Sp. <Emphasis Type="Italic">cubense</Emphasis>

Banana wilt disease is a typical vascular disease caused by the fungal pathogen Fusarium oxysporum f. sp. cubense 4 (Foc 4). Pattern recognition receptors in the plant cell membrane can recognize pathogen-associated molecular patterns (PAMPs) to activate multi-layer defense responses, including defense gene expression, stomatal closure, reactive oxygen species (ROS) burst and callose deposition, to limit pathogen growth. In the present study, we found that chitin elicitor receptor kinase 1 (CERK1) was required for the non-host resistance of Arabidopsis thaliana to Foc B2 (a strain of Foc 4). The cerk1 mutant had weaker defense responses after Foc B2 treatment, including lower expression of PAMP- and salicylic acid-responsive genes, no stomatal closure, lower ROS level and less callose deposition, than that of the wild-type plant. Consistent with this, the cerk1 mutant plants exhibited higher susceptibility to non-host pathogen Foc B2. These results suggest the crucial importance of CERK1 in Foc B2-triggered non-host resistance.     

Rapid screening of <Emphasis Type="Italic">Musa</Emphasis> species for resistance to Fusarium wilt in an <Emphasis Type="Italic">in vitro</Emphasis> bioassay

In order to accelerate breeding and selection for disease resistance to Fusarium wilt, it is important to develop bioassays which can differentiate between resistant and susceptible cultivars efficiently. Currently, the most commonly used early bioassay for screening Musa genotypes against Fusarium oxysporum f. sp. cubense (Foc) is a pot system, followed by a hydroponic system. This paper investigated the utility of in vitro inoculation of rooted banana plantlets grown on modified medium as a reliable and rapid bioassay for resistance to Foc. Using a scale of 0 to 6 for disease severity measurement, the mean final disease severities of cultivars expressing different levels of disease reaction were significantly different (P ≤ 0.05). Twenty-four days after inoculation with Foc tropical race 4 at 10⁶ conidia ml⁻¹, the plantlets of two susceptible cultivars had higher final disease severities than that of four resistant cultivars. Compared with ‘Guangfen No.1’, ‘Brazil Xiangjiao’ is highly susceptible to tropical race 4 and its mean final disease severity was the highest (5.27). The plantlets of moderately resistant cultivar ‘Formosana’ had a mean final disease severity (3.53) lower than that of ‘Guangfen No.1’ (4.33) but higher than that of resistant cultivars: ‘Nongke No.1’, GCTCV-119, and ‘Dongguan Dajiao’ (1.87, 1.73, and1.53, respectively). Promising resistant clones acquired through non-conventional breeding techniques such as in vitro selection, genetic transformation, and protoplast fusion could be screened by the in vitro bioassay directly. Since there is no acclimatization stage for plantlets used in the bioassay, it helps to improve banana breeding efficiency.     

Baseline sensitivity of <Emphasis Type="Italic">Phytophthora capsici</Emphasis> to the strobilurin fungicide benzothiostrobin and the efficacy of this fungicide

Fusarium oxysporum f. sp. cubense tropical race 4 (Foc TR4) has seriously threatened global banana production. Recently, identification of Foc TR4 in the Caribbean and Mozambique has aroused widespread concern because it may be directly related to food security for millions of people in Africa and Latin America. To identify a resistant or highly resistant germplasm, 129 accessions were evaluated for Foc TR4 resistance in greenhouses and 100 of them were also assessed in the field. In total, 10 accessions were highly resistant (HR) to Foc TR4; these include four from the AA group, two from the BB group, one of the East African highland bananas (EAHBs), two plantains, and one from wild relatives of banana. Of these, Pahang (AA), Calcutta 4 (AA), and Musa itinerans exhibited the highest degree of resistance with an index of disease (ID2) less than 10, and Foc TR4 could not infect any of them in the field. Furthermore, 31 accessions from AA, AB, AAB, AAAB, and ABB groups and wild relatives were identified as a resistant (R) germplasms. All tested EAHBs and plantains exhibited resistance or high resistance to Foc TR4 infection. Our results provide valuable genetic resources for banana breeding and for studying the mechanisms underlying Fusarium wilt resistance. Furthermore, evaluation of EAHBs and plantains provided a rational for local banana producers and researchers to grow EAHBS and plantains.     

Impact of carrot resistance on development of the <Emphasis Type="Italic">Alternaria</Emphasis> leaf blight pathogen (<Emphasis Type="Italic">Alternaria dauci</Emphasis>)

The interaction between Alternaria dauci and two carrot cultivars differing in their resistance to leaf blight was investigated by microscopy. The fungal development between 1 and 15 days post-inoculation was quite similar in the susceptible cv. Presto and the partially resistant cv. Texto: After conidial germination, leaf adhesion of the pathogen was achieved with mucilaginous filaments; hyphae penetrated the leaves directly with/without the formation of appressoria-like structures or via stomata; the fungus spread by epiphytic hyphae with hyphopodia and subcuticular mycelia. Intense necrotic plant cell reactions occurred under the fungal structures. At 21 days post-inoculation, typical features of fungal development were noted for each cultivar: growing hyphae emerged from stomata in cv. Presto, whereas conidiophores without conidia were observed in cv. Texto. Leaf tissues of both cultivars were strongly damaged and vesicle-like structures (assumed to be plant phenolics) were abundantly present between mesophyll cells. A real-time PCR method was developed for in planta quantification of A. dauci. Between 1 and 15 days post-inoculation, the fungal biomass was equivalent in the two cultivars and was about fourfold higher in cv. Presto than cv. Texto at 21 and 25 days post-inoculation. Taken together, our results indicated that A. dauci was able to colonize both cultivars in a similar manner during the first steps of the interaction, then fungal development in the partially resistant cultivar was restricted due to putative plant defence reactions. The results of this study enhance the overall understanding of infection processes in the A. dauci-carrot pathosystem.     

A study on the susceptibility of the model legume plant <Emphasis Type="Italic">Medicago truncatula</Emphasis> to the soil-borne pathogen <Emphasis Type="Italic">Fusarium oxysporum</Emphasis>

Fusarium wilt is a soil-borne disease caused by formae specialis of Fusarium oxysporum on a large number of cultivated and wild plants. The susceptibility of the model legume plant Medicago truncatula to Fusarium oxysporum was studied by root-inoculating young plants in a miniaturised hydroponic culture. Among eight tested M. truncatula lines, all were susceptible to F. oxysporum f.sp. medicaginis, the causal agent of Fusarium wilt in alfalfa. However, a tolerant line, F83005.5, and a susceptible line, A17, could be distinguished by scoring the disease index. The fungus was transformed with the GFP marker gene and colonisation of the plant roots was analysed by epifluorescence and confocal microscopy. A slightly atypical pattern of root colonisation was observed, with massive fungal growth in the cortex. Although colonisation was not significantly different between susceptible and tolerant plants, the expression of some defence-related genes showed discrimination between both lines. A study with 10 strains from various host-plants indicated that M. truncatula was a permissive host to F. oxysporum.     

Seed transmission of<Emphasis Type="Italic">Fusarium oxysporum</Emphasis> f.sp.<Emphasis Type="Italic">lactucae</Emphasis>

Twenty-seven seed samples belonging to the lettuce cultivars most frequently grown in Lombardy (northwestern Italy), in an area severely affected by Fusarium wilt of lettuce, were assayed for the presence ofFusarium oxysporum on a Fusarium-selective medium. Isolations were carried out on subsamples of seeds (500 to 1500) belonging to the same seed lots used for sowing, and either unwashed or disinfected in 1% sodium hypochloride. The pathogenicity of the isolates ofF. oxysporum obtained was tested in four trials carried out on lettuce cultivars of the butterhead type, very susceptible to Fusarium wilt. Nine of the 27 samples of seeds obtained from commercial seed lots used for sowing in fields affected by Fusarium wilt were contaminated byF. oxysporum. Among the 16 isolates ofF. oxysporum obtained, only one was isolated from disinfected seeds. Three of the isolates were pathogenic on the tested cultivars of lettuce, exhibiting a level of pathogenicity similar to that of the isolates ofF. oxysporum f.sp.lactucae obtained from infected wilted plants in Italy, USA and Taiwan, used as comparison. The results obtained indicate that lettuce seeds are a potential source of inoculum for Fusarium wilt of lettuce. The possibility of isolatingF. oxysporum f.sp.lactucae, although from a low percent of seeds, supports the hypothesis that the rapid spread of Fusarium wilt of lettuce observed recently in Italy is due to the use of infected propagation material. Measures for prevention and control of the disease are discussed. http://www.phytoparasitica.org posting Dec. 16, 2003.     

Microbial community responses associated with the development of <Emphasis Type="Italic">Fusarium oxysporum</Emphasis> f. sp. <Emphasis Type="Italic">cucumerinum</Emphasis> after 24-epibrassinolide applications to shoots and roots in cucumber

Fusarium wilt, caused by Fusarium oxysporum f. sp. cucumerinum (FO), is one of the major diseases in cucumber (Cucumis sativus) production. Root and foliar applications of 24-epibrassinolide (EBL), an immobile phytohormone with antistress activity, were evaluated for their effects on the incidence of Fusarium wilt and changes in the microbial population and community in roots of cucumber plants. EBL pre-treatment to either roots or shoots significantly reduced disease severity followed by an improved plant growth regardless of the treatment methods applied. EBL applications decreased the Fusarium population on root surfaces and in nutrient solution, but increased the population of fungi and actinobacteria on root surfaces. PCR-DGGE analysis showed that FO-inoculation had significant effects on the bacterial community on root surfaces as expressed by a decreased diversity index and evenness index, but EBL applications alleviated these changes. Moreover, several kinds of decomposing bacteria and growth-promoting bacteria were identified from root surfaces of FO-inoculated plants and EBL-pre-treated plants, respectively. Overall, these results show that the microbial community on root surfaces was affected by a complex interaction between phytohormone-induced resistance and plant pathogens.     

Interaction of root colonizing biocontrol agents demonstrates the antagonistic effect against <Emphasis Type="Italic">Fusarium oxysporum</Emphasis> f. sp. <Emphasis Type="Italic">lycopersici</Emphasis> on tomato

Plant growth promoting Bacillus subtilis MSS9 and Bacillus licheniformis MSS14 were isolated from the tomato rhizosphere. These isolates were capable of inhibiting the fungal pathogen, Fusarium oxysporum f. sp. lycopersici causing fusarium wilt in tomato, tested by dual culture method and by mycolytic enzyme production. The isolates have the capacity to form biofilm on the microtitre plate. Scanning electron microscopy revealed good colonization capacity of Bacillus licheniformis MSS14 on tomato plant root as compared to Bacillus subtilis MSS9, pot experiments were also analyzed to study the effects of both rhizobacterial cultures on pathogen development and plant growth. It was observed that MSS14 reduces the incidence of Fusarium oxysporum f. sp. lycopersici in tomato and there was significant increase in vegetative parameters like root length, shoot length, plant wet weight, dry weight and chlorophyll content after which indicates that the root colonization property of the culture MSS14 helps in enhancing the biocontrol capacity against pathogen than that of MSS9.     

Impaired purine biosynthesis affects pathogenicity of <Emphasis Type="Italic">Fusarium oxysporum</Emphasis> f. sp. <Emphasis Type="Italic">melonis</Emphasis>

The vascular wilt pathogen Fusarium oxysporum f. sp. melonis causes worldwide yield losses of muskmelon. In this study, we characterized a UV-induced non-pathogenic mutant (strain 4/4) of F. oxysporum f. sp. melonis, previously identified as a potential biological control agent. During comparative analysis of vegetative growth parameters using different carbon sources, mutant strain 4/4 showed a delay in development and secretion of extracellular enzymes, compared to the wild type strain. Amendments of the growth medium with yeast extract, adenine or hypoxanthine, but not guanine, complemented the growth defect of strain 4/4, as well as secretion and partial activity of cellulases and endopolygalacturonases, indicating that the strain is an adenine auxotroph. Incubation of strain 4/4 conidia in adenine solution, prior to inoculation of muskmelon plants, partially restored pathogenicity to the mutant strain.     

<Emphasis Type="Italic">Lupinus luteus</Emphasis>, a new host of <Emphasis Type="Italic">Phytophthora cinnamomi</Emphasis> in Spanish oak-rangeland ecosystems

Phytophthora cinnamomi is an aggressive pathogen on Lupinus luteus (yellow lupin), causing root rot, wilting and death of this crop, common in oak-rangeland ecosystems ('dehesas') in south-western Spain. The oomycete, the main cause of Quercus decline in the region, was isolated from roots of wilted lupins in the field. Artificial inoculations on four cultivars of L. luteus reproduced the symptoms of the disease, both in pre- and post-emergence stages, recovering the pathogen from necrotic roots. These results suggest the potential of yellow lupin as inoculum reservoir for the infection of Quercus roots. This is the first report of P. cinnamomi as root pathogen of L. luteus.     

Infection and colonization of strawberry by <Emphasis Type="Italic">Gnomonia fragariae</Emphasis> strain expressing green fluorescent protein

Gnomonia fragariae is a poorly studied ascomycete, which was recently demonstrated to be a cause of severe root rot and petiole blight of strawberry. The pathogen was genetically transformed with the GFP as a vital marker and hygromycin resistance gene. Several stable transformants were obtained, which did not differ in their phenotype from the wild type isolate. Using one of the GFP-tagged isolates the infection process and colonization of roots and petioles of host plant by the pathogen were studied. Fluorescence microscopy examinations of the inoculated plants at different time points showed that plant infection occurs 24 h after inoculation and intensively continues during first 3 days. The specific penetration sites on epidermal cells and preferences in colonization for certain root and petiole tissues were observed. The pathogen intensively colonized and destroyed cortex of roots and petioles and spread rapidly longitudinally within intercellular spaces. The petioles were colonized by the hyphae, which grew mostly in the intracellular spaces of the cortical cells while in the roots the intracellular growth of hyphae occurred only in the later stages of infection. The fungus was also capable to infect the vascular tissues of petioles although these were not the primary tissues colonized by the pathogen. The mature ascomata were formed on the infected petiole bases several weeks after the inoculation. This study presents a genetic transformation method for Gnomonia fragariae and it demonstrates details on infection process and colonization of root, crown and petiole tissues of strawberry by the pathogen.     

Screening of cultivated and wild adzuki bean for resistance to race 3 of <Emphasis Type="Italic">Cadophora gregata</Emphasis> f. sp. <Emphasis Type="Italic">adzukicola</Emphasis>, cause of brown stem rot

Two diseases of adzuki bean, brown stem rot (BSR, caused by Cadophora gregata f. sp. adzukicola) and adzuki bean Fusarium wilt (AFW, caused by Fusarium oxysporum f. sp. adzukicola), are serious problems in Hokkaido and have been controlled using cultivars with multiple resistance. However, because a new race of BSR, designated race 3, was identified, sources of parental adzuki bean for resistance to race 3 were needed. Therefore, we examined 67 cultivars and lines of cultivated and wild adzuki bean maintained at the Tokachi Agricultural Experiment Station using a root-dip inoculation method. Consequently, nine adzuki bean cultivars, one wild adzuki bean accession and 30 lines (including two lines resistant to all the three races of BSR and AFW) were confirmed to be resistant or tolerant to race 3 of BSR, and we found a cultivar Akamame as well as a wild adzuki bean Acc2515 to be a new source for a resistance gene to the race 3. This cultivar also holds promise as a source of resistance against other races of BSR and AFW.     

Modifications in the potato rhizosphere during infestations of <Emphasis Type="Italic">Globodera rostochiensis</Emphasis> and subsequent effects on the growth of <Emphasis Type="Italic">Rhizoctonia solani</Emphasis>

Two controlled environment experiments were conducted to explore the hypothesis that invasion and damage caused to potato roots by the potato cyst nematode Globodera rostochiensis might result in quantitative or qualitative changes in the release of root exudates to subsequently affect the growth of Rhizoctonia solani (AG3) in the potato rhizosphere. The growth of five R. solani isolates was compared on media amended either with root exudates from G. rostochiensis-infested or uninfested potato (cv. Désirée) plants at different time intervals after the introduction of the nematodes. In Experiment 1, the growth of R. solani was higher on medium amended with potato root exudates from G. rostochiensis-infested compared to uninfested plants, collected 4, 6, 8 and 12 days after the G. rostochiensis treatments were administered. Similarly, in Experiment 2, R. solani isolates grew faster on medium amended with potato root exudates from G. rostochiensis-infested than uninfested plants. This trend was particularly pronounced at the 12-day collection. At this time, 49% of the G. rostochiensis juveniles in roots were found to belong to the juvenile moults J2 and J3, indicating that root exudates were modified during the earlier stages of juvenile invasion. Carbohydrate analysis of root exudates indicated significantly higher levels of sucrose in root exudates from G. rostochiensis-infested than uninfested plants, whereas no significant differences were found in total nitrogen content. The results are discussed to help elucidate the mechanism behind the disease complex found between G. rostochiensis and R. solani in previous field research.     

Pathogenicity of indole-3-acetic acid producing fungus <Emphasis Type="Italic">Fusarium delphinoides</Emphasis> strain GPK towards chickpea and pigeon pea

An indole-3-acetic acid (IAA) producing fungal strain was isolated from chickpea grown rhizospheric soil samples. Based on morphological and Internal Transcribed Spacer (ITS) region sequence analysis the new isolate was identified as Fusarium delphinoides. The Fusarium delphinoides strain produces and secretes IAA in-vitro as identified by HPLC and Mass spectrometry. The IAA production is dependent on tryptophan (Trp) as a nitrogen source in the medium. The IAA production is influenced by growth conditions such as pH of the medium, concentration of Trp and the nature of the carbon source. Additional nitrogen sources repress Trp dependent IAA production. Glucose and Trp served as the best carbon and nitrogen sources respectively. Pathogenicity of Fusarium delphinoides towards the plants was tested by electrolyte, nutrient leakage analysis and also by scoring the disease symptoms. Two cultivars of chickpea (ICCV-10 and L-550) and two cultivars of pigeon pea (Maruti and PT-221) were assessed for the pathogenicity by inoculating with spores of Fusarium delphinoides. The inoculation induced symptoms of Fusarium wilt as in the case of Fusarium oxysporum f. sp. ciceris (FOC), a known pathogen causing Fusarium wilt in chickpea. Electrolyte and nutrient leakage from the infected plants were used to assess the resistance, tolerance (moderately resistance) and susceptibility of the plants to the infection. Based on the results, both the pigeon pea cultivars (Maruti and PT-221) were rated as resistant, and ICCV-10 was rated as a tolerant cultivar of chickpea. However, chickpea cultivar L −550 was found to be a susceptible host for infection by Fusarium delphinoides. These results suggest that Fusarium delphinoides, which belongs to the Fusarium dimerum species group, is an IAA producing plant pathogen and causes wilt in chickpea. Further, along with pathogenicity tests, electrolyte and nutrient leakage analysis can be used to assess the pathogenicity of pathogenic fungi.     

Characterization of <Emphasis Type="Italic">Phytophthora clandestina</Emphasis> races on <Emphasis Type="Italic">Trifolium subterraneum</Emphasis> in Western Australia

Phytophthora clandestina is a causal agent of root rot disease of subterranean clover in Western Australia (W.A). As a significant number of isolates of P. clandestina from W.A. could not previously be designated using existing differentials, a comprehensive set of subterranean clover (Trifolium subterraneum) cultivars was used as differentials to delineate a broader range of races of the pathogen. One hundred and one isolates of the pathogen collected from W.A. were screened on nine subterranean clover cultivars, of which seven were found to be useful as host differentials. A total of 10 races (in contrast to the five recognized previously) were defined and differentiated using octal nomenclature, presenting a clearer picture of the racial distribution of P. clandestina among W.A. isolates. Differences were found in the race populations between Australian states and are therefore important to the selection/breeding of cultivars for specific regions of Australia to counter the predominant race populations and for enforcing quarantine measures in relation to seed movements within and outside Australia. The octal nomenclature used provides a sound basis for follow-up studies and future race designations. Races 173 and 177 in this study were widely distributed and were the most common races in W.A., and together constitute 80% of the isolates characterized. While six of the seven host differentials were resistant to isolates belonging to race 001 and all were resistant to 000, it is of concern that only one differential was resistant to 157 and 173 and that none of the host differentials were resistant to 177. Our approach to P. clandestina race delineation is clearly conservative and is different from previous studies. The octal nomenclature we applied in this study is not only scientifically sound but also will facilitate rapid recognition and characterization of the races.     

Cloning of the pathogenicity-related gene <Emphasis Type="Italic">FPD1</Emphasis> in <Emphasis Type="Italic">Fusarium oxysporum</Emphasis> f. sp. <Emphasis Type="Italic">lycopersici</Emphasis>

We selected a reduced-pathogenicity mutant of Fusarium oxysporum f. sp. lycopersici, a tomato wilt pathogen, from the transformants generated by restriction enzyme-mediated integration (REMI) transformation. The gene tagged with the plasmid in the mutant was predicted to encode a protein of 321 amino acids and was designated FPD1. Homology search showed its partial similarity to a chloride conductance regulatory protein of Xenopus, suggesting that FPD1 is a transmembrane protein. Although the function of FPD1 has not been identified, it does participate in the pathogenicity of F. oxysporum f. sp. lycopersici because FPD1-deficient mutants reproduced the reduced pathogenicity on tomato.The nucleotide sequence data reported are available in the DDBJ/EMBL/GenBank databases under the accession number AB110097     

Cell interactions between a nonpathogenic <Emphasis Type="Italic">Fusarium oxysporum</Emphasis> strain and root tissues of <Emphasis Type="Italic">Eucalyptus viminalis</Emphasis>

Nonpathogenic isolates of Fusarium oxysporum can be successful antagonists of pathogenic forms of the same fungal species that commonly attacks crop plants. The characteristics that distinguish nonpathogenic from pathogenic forms are not well understood. In this study, the mode of root colonization of Eucalyptus viminalis seedlings by a nonpathogenic F. oxysporum strain is described at the ultrastructural level. Root systems of E. viminalis plants were inoculated with nonpathogenic F. oxysporum strain Fo47 in an in vitro model system. Changes in the occurrence of nonesterified and methyl-esterified pectins in colonized E. viminalis roots were evaluated by in situ immunolabeling using two monoclonal antibodies, JIM 5 and JIM 7. Modes of penetration and root colonization patterns in E. viminalis seedlings by the nonpathogenic fungus were similar to those described for pathogenic forms of F. oxysporum. However, root interactions differed in that the nonpathogenic fungus did not induce host tissue damage. No papilla-like appositions were observed in host cells in response to invading hyphae, which did not disrupt the host plasma membrane in many cases, suggesting that a biotrophic relationship was established. Root colonization by the nonpathogenic strain did not induce alteration in JIM 7 labeling of methyl-esterified pectin in E. viminalis cell walls, whereas nonesterified pectin was detected to a significantly greater extent in cell walls of roots colonized by the fungus. Pectin components decreased slightly only at points of hyphal contact with host cells. Because nonpathogenic strains utilize pectin in pure culture, host control over enzyme activity or production by the fungi may at least partly explain their compatible interactions with host tissues.     

<Emphasis Type="Italic">In vitro</Emphasis> bacterization of banana (<Emphasis Type="Italic">Musa</Emphasis> spp.) with native endophytic and rhizospheric bacterial isolates: Novel ways to combat <Emphasis Type="Italic">Fusarium</Emphasis> wilt

Compared to conventional planting material, micropropagated plantlets are highly susceptible to Fusarium wilt because they are free from beneficial root inhabitants. We aimed to introduce mixtures of beneficial microbes in the plantlets in the rooting medium under in vitro conditions rather than by field applications. Endophytes and rhizobacteria from different banana cultivars and plantation areas were screened and characterized. Under in vitro conditions, banana tissue culture plantlets were bacterized with the prospective endophytes, Bacillus subtilis strain EPB56 and EPB10 and the rhizobacteria, Pseudomonas fluorescens strain Pf1 and effects of in vitro bacterization were investigated against Fusarium oxysporum f. sp. cubense race 1 under glasshouse and field conditions. Inoculation of bananas during micropropagation allowed for the omission of minerals and salts as well as vitamins from the growing media while resulting in plantlets close to double size compared to the controls with full strength media. All endophyte and rhizobacteria strains tested resulted in significant reductions in Fusarium infection in the glasshouse and field and in significantly better plant growth. The three-way combination of bacteria resulted in 78% disease reduction and more than doubled the yields compared to the untreated controls across two field experiments. Three-way inoculation led to yields of 23 and 24 kg/ bunch compared to chemical disease control (13; 15 kg/bunch) and untreated controls (10; 13 kg/bunch) in the two field experiments. Under glasshouse conditions, activity of defence enzymes was significantly increased by all inoculation treatments. Inoculation in vitro led to the establishment of the microorganisms in the plant system before delivering to the farming community. Micropropagation combined with the establishment of a beneficial microbial consortium should complement the micropropagated plants for easier adaptation under field conditions.     

Combining sub-lethal heating and on-farm wastes: effects on <Emphasis Type="BoldItalic">Fusarium oxysporum</Emphasis> f.sp. <Emphasis Type="BoldItalic">cumini</Emphasis> causing wilt of cumin

In a 2-year field study, the effects of four heating regimes established by varying temperature or duration of heating on efficiency of on-farm wastes as soil amendments in controlling Fusarium oxysporum f.sp. cumini (Foc) causing wilt on cumin (Cuminum cyminum L.), were ascertained. Significant improvement in reduction of Foc propagules was achieved with the increase in duration and amount of heat. In 2000, mild heating under shade (heat level 4), 31.8–65.9% reduction in Foc propagules was estimated in all the amendments at 0–30 cm soil depth, which improved by 75.7–86.5% in the treatment where Foc-infested soil brought from the laboratory was exposed to direct heat (heat level 3) and amendments and irrigation were applied. Foc propagules were reduced by 76.6–88.3% when infested soil was exposed continuously to dry heat for 56 days (heat level 2), improving efficiency of amendments by 0.9–13.5% compared with heat level 3. After 56 days of exposure to dry heat, elevating the temperature by mulching amended soil with a transparent polyethylene film (50 μm thick) for 20 days (heat level 1) augmented reduction by 80.2–95.5%. In the second season, combining 0.04% of onion, Verbisina encelioides or mustard oil-cake with mustard residues (0.18%) improved the reduction in Foc propagules at all heat levels even at the lower soil depth. Maximum reduction (94.9–100%) in Foc propagules at heat levels 1 to 3 was achieved when residues of Verbisina were supplemented with those of mustard. These results suggest a new approach to improve the control of Foc by combining prolonged heating with on-farm wastes such as Verbisina residues and one summer irrigation.