Simulated Leaching of Foliar Applied Copper Bactericides on the Soil Microbiome Utilizing Various Beta Diversity Resemblance Measurements.

Copper bactericides are routinely used to control Xanthomonas perforans (XP), causal agent of bacterial spot of tomato. Given the widespread tolerance to copper in XP strains in FL, USA, nanotechnology-based elemental composites have gained interest for their potential applications in agriculture in part due to their enhanced antimicrobial properties and toxicity to copper-tolerant strains. However, little is known about the potential impact of conventional copper bactericides as well as nano-based elemental composites on soil microbial communities, as determined by high-throughput sequencing of the 16S rDNA. We compared the effects of 2 and 200 µg/mL of core-shell (CS), a metallic copper composite, and a conventional copper bactericide + mancozeb (Cu+Man) on the soil microbiome. These treatments were compared to three controls, the microbial profile of the soil prior to application of copper products, a water application, and spiking the soil with a soilborne phytobacterium, Ralstonia solanacearum (RS). The RS treatment was included to determine if downstream analysis could detect the artificial inoculation. Utilizing multiple ß diversity measurements, each emphasizing various tenets of ecology, provided a greater perspective of the effects the treatments had on the microbiome. Analysis of HTS data revealed that the two treatments containing field applied rates of metallic copper, CS 200 and Cu+Man, had the largest impact on the soil microbiome at seven-days posttreatment compared to water. However, we simulated field applied rates of CS 200 entering the soil by treating soil with CS 2 and determined this concentration had a negligible effect on the soil microbiome. IMPORTANCE Nanotechnology-based elemental composites have gained popularity for their potential applications in plant disease management due to their enhanced antimicrobial properties. However, little is known about their potential impact on the environment. Foliar applications of nano metallic composites upon leaching into the soil have the potential to impact soil microbial populations that in turn influence soil health. Utilizing multiple ß diversity measurements, high-throughput sequencing analysis revealed that field applied rates of metallic copper (200 µg/mL) from an advanced copper composite (core-shell [CS]) and a conventional copper bactericide in combination with mancozeb had the largest impact on the soil microbiome compared to water and nontreated control. To simulate leaching from the leaf surface, a lower concentration (2 µg/mL) of CS was also applied to the soil and had a negligible effect on the soil microbiome. Thus, field applied rates of CS may have a minimal effect on soil microbial communities.

Improved deferred antagonism technique for detecting antibiosis.

The deferred antagonism technique has been utilized for several decades for detecting antibiosis activity. Most protocols require the elimination of antibiotic-producing cells by exposing them to chloroform vapour, UV radiation or filter sterilizing the filtrate steps that require additional time and expense to complete. We provide a modified approach to current soft agar overlay practices, which involves addition of antibiotics to the soft agar overlay to inhibit growth of the producer but not the indicator strain. This technique can be used to reproducibly and efficiently screen for antibiotic production with ease. We demonstrate the effectiveness of this technique with three bacterial systems: inhibition of the bacterial spot of tomato pathogen, Xanthomonas euvesicatoria, by its pathogenic competitor Xanthomonas perforans; and inhibition of the fire blight pathogen, Erwinia amylovora, by Pantoea vagans C9-1 or Pseudomonas fluorescens A506. SIGNIFICANCE AND IMPACT OF THE STUDY: Deferred antagonism assays are used commonly to observe antibiotic production by micro-organisms. Killing or removing the producer cells prior to introduction of the indicator strain is a standard practice but requires additional time and special handling procedures. We evaluated a modification of the assay, where the overlay medium is amended with an antibiotic to which the indicator strain is resistant and the producer strain is sensitive. This modification obviates extra steps to kill the producer strain prior to overlaying with the indicator strain and provides a rapid, consistent and cost-effective method to detect antibiosis.

Fungicide Application Timing Essential for the Management of Leaf Spot and Fruit Rot on Pomegranate (Punica granatum L.) in Florida.

Pomegranate (Punica granatum L.) has emerged as an alternative fruit crop for growers dealing with devastating threats to citrus and avocado in the southeastern United States. However, foliar and fruit diseases are major constraints to commercial pomegranate production. Replicated field trials were performed in 2015 at three separate sites in Florida (Dover, Plant City, and Parrish) to evaluate Merivon (pyraclostrobin and fluxapyroxad), Luna Experience (fluopyram + tebuconazole), and a rotational program consisting of these two fungicides as well as Penncozeb 75DF (mancozeb) for the management of leaf spot and fruit rot of pomegranate. The fungicide programs were evaluated on the varieties Vietnam in Dover, FL; Angel Red in Plant City, FL; as well as Christina, Azadi, Vikusnyi, Alsirinnar, Sakerdze, and Wonderful in Parrish, FL. Mean leaf spot severity varied across sites at the end of each trial in August, with values ranging from 4.5 to 62.5% in Plant City, 7.5 to 45.8% in Dover, and 4.5 to 54.2% in Parrish. Based on area under the disease progress curve, all treatments that included at least one at bloom application significantly reduced disease levels compared with postbloom treatments and the nontreated control across all trial sites. Based on fruit rot incidence, Luna Experience and Merivon, when applied twice at bloom in the beginning of the season, reduced disease by 66.6 and 88.4%, respectively, in comparison with the nontreated control in Plant City. A rotational program further reduced disease by at least 97% at the end of the season in Parrish. The proper timing of fungicide application and rotation with different modes of action provided a feasible disease management option for pomegranate growers in the Southeast United States.

Particle-size dependent bactericidal activity of magnesium oxide against Xanthomonas perforans and bacterial spot of tomato.

Bacterial spot, caused by Xanthomonas spp., is a highly destructive disease of tomatoes worldwide. Copper (Cu) bactericides are often ineffective due to the presence of Cu-tolerant strains. Magnesium oxide (MgO) is an effective alternative to Cu bactericides against Xanthomonas spp. However, the effects of particle size on bactericidal activity and fruit elemental levels are unknown. In this study, nano (20 nm) and micron (0.3 and 0.6 µm) size MgO particles were compared for efficacy. Nano MgO had significantly greater in vitro bactericidal activity against Cu-tolerant X. perforans than micron MgO at 25-50 µg/ml. In field experiments nano and micron MgO applied at 200 and 1,000 µg/ml were evaluated for disease control. Nano MgO at 200 µg/ml was the only treatment that consistently reduced disease severity compared to the untreated control. Inductively Coupled Plasma Optical Emission Spectroscopy revealed that nano MgO applications did not significantly alter Mg, Cu, Ca, K, Mn, P and S accumulation compared to fruits from the untreated plots. We demonstrated that although both nano MgO and micron MgO had bactericidal activity against Cu-tolerant strains in vitro, only nano MgO was effective in bacterial spot disease management under field conditions.

Dwiroopa punicae sp. nov. (Dwiroopaceae fam. nov., Diaporthales), associated with leaf spot and fruit rot of pomegranate (Punica granatum).

During a survey of diseases affecting pomegranate in the southeastern USA we identified a unique species of Diaporthales causing leaf spotting and fruit rot. Objectives of this study were to provide a morphological description of the putative new species, use DNA sequence data of three gene loci (LSU, ITS and rpb2) to accurately place the fungus within the Diaporthales, and to prove Koch's postulates. Morphological and phylogenetic comparisons confirmed the fungus to represent a new species and family, for which the names Dwiroopa punicae sp. nov. and Dwiroopaceae fam. nov. are proposed. This is the first report of a species of Dwiroopa being pathogenic to Punica granatum.

Nano-Magnesium Oxide: A Novel Bactericide Against Copper-Tolerant Xanthomonas perforans Causing Tomato Bacterial Spot.

Bacterial spot caused by Xanthomonas perforans causes significant damage on tomato in Florida. Due to the presence of copper (Cu)-tolerant X. perforans strains, Cu bactericides are not effective in disease management. Hence, there is a critical need to find alternatives for Cu. Antibacterial activity of magnesium oxide (Nano-MgO), and other metal oxide nanoparticles, were evaluated against a Cu-tolerant and -sensitive X. perforans strain. In vitro experiments demonstrated high antibacterial activity of Nano-MgO against both strains compared with the commercial Cu. The minimum inhibitory concentration of Nano-MgO is 25 µg/ml and the minimum bactericidal concentration is 100 µg/ml against a Cu-tolerant X. perforans strain after 4 h of exposure. Structural changes in the bacterial membrane following exposure to Nano-MgO treatments compared with the controls were observed using transmission electron microscopy. In two greenhouse experiments with a Cu-tolerant strain, bacterial spot severity was significantly reduced by Nano-MgO at 200 µg/ml compared with Cu-ethylene bis-dithiocarbamate (grower standard), and the untreated control (P = 0.05). In three field experiments, Nano-MgO at 200 µg/ml significantly reduced disease severity with no negative impact on yield compared with the untreated control. Inductively coupled plasma mass spectrometric analysis of the fruit confirmed that Nano-MgO application did not lead to the accumulation of Mg, Cu, Ca, K, Mn, P, and S. This study is the first to demonstrate the potential of Nano-MgO against bacterial spot of tomato.

Reclassification of Xanthomonas gardneri (ex Sutic 1957) Jones et al. 2006 as a later heterotypic synonym of Xanthomonas cynarae Trébaol et al. 2000 and description of X. cynarae pv. cynarae and X. cynarae pv. gardneri based on whole genome analyses.

Multilocus sequence analysis of Xanthomonas species revealed a very close relationship between Xanthomonas cynarae, an artichoke pathogen and Xanthomonas gardneri, a tomato and pepper pathogen. Results of whole genome sequence comparisons using average nucleotide identity between representative strains of X. gardneri and X. cynarae were well above the threshold of 95-96 %. Inoculations of X. gardneri strains in artichoke leaves caused mild disease symptoms, but only weak symptoms were observed in the bracts. Both X. cynarae and X. gardneri grew equally and caused typical bacterial spot symptoms in pepper after artificial inoculation. However, X. cynarae induced a hypersensitive reaction in tomato, while X. gardneri strains were virulent. Pathogenicity-associated gene clusters, including the protein secretion systems, type III effector profiles, and lipopolysaccharide cluster were nearly identical between the two species. Based on our results from whole genome sequence comparison, X. gardneri and X. cynarae belong to the same species. The name X. cynarae has priority and X. gardneri should be considered as a later heterotypic synonym. An emended description of X. cynarae (type strain=CFBP 4188T, =DSM 16794T) is given. However, due to the host specificity in artichoke and tomato, two pathovars, X. cynarae pv. cynarae and X. cynarae pv. gardneri, are proposed.

Advanced Copper Composites Against Copper-Tolerant Xanthomonas perforans and Tomato Bacterial Spot.

Bacterial spot, caused by Xanthomonas spp., is a widespread and damaging bacterial disease of tomato (Solanum lycopersicum). For disease management, growers rely on copper bactericides, which are often ineffective due to the presence of copper-tolerant Xanthomonas strains. This study evaluated the antibacterial activity of the new copper composites core-shell copper (CS-Cu), multivalent copper (MV-Cu), and fixed quaternary ammonium copper (FQ-Cu) as potential alternatives to commercially available micron-sized copper bactericides for controlling copper-tolerant Xanthomonas perforans. In vitro, metallic copper from CS-Cu and FQ-Cu at 100 µg/ml killed the copper-tolerant X. perforans strain within 1 h of exposure. In contrast, none of the micron-sized copper rates (100 to 1,000 µg/ml) from Kocide 3000 significantly reduced copper-tolerant X. perforans populations after 48 h of exposure compared with the water control (P < 0.05). All copper-based treatments killed the copper-sensitive X. perforans strain within 1 h. Greenhouse studies demonstrated that all copper composites significantly reduced bacterial spot disease severity when compared with copper-mancozeb and water controls (P < 0.05). Although there was no significant impact on yield, copper composites significantly reduced disease severity when compared with water controls, using 80% less metallic copper in comparison with copper-mancozeb in field studies (P < 0.05). This study highlights the discovery that copper composites have the potential to manage copper-tolerant X. perforans and tomato bacterial spot.

A Multiplex Real-Time PCR Assay Differentiates Four Xanthomonas Species Associated with Bacterial Spot of Tomato.

Bacterial spot of tomato, a major problem in many tomato production areas, is caused by Xanthomonas euvesicatoria, X. vesicatoria, X. perforans, and X. gardneri. In order to detect and identify the bacterial spot pathogens, we evaluated a region of hrpB operon as a source for primers and probes for real-time polymerase chain reaction (PCR). A 420-bp fragment of the hrpB7 gene was amplified by PCR from 75 strains representing the four species. The PCR products were sequenced and phylogenetic analysis revealed that hrpB7 is highly conserved within each species, with a single-nucleotide polymorphism (SNP) among the X. vesicatoria strains. X. euvesicatoria and X. perforans varied by two SNP. Four probes and two primer sets were designed to target the four bacterial spot pathogens based on their hrpB7 gene sequences. In order to simultaneously detect the four bacterial spot pathogens, the four probes and two primer sets were optimized for a multiplex real-time TaqMan PCR assay. The optimized multiplex assay was determined to be highly specific to the four bacterial spot pathogens. Because the optimized multiplex assay facilitated the identification of each bacterial spot pathogen from pure cultures and infected plant tissue, it holds great potential as a diagnostic tool.

Multilocus Sequence Analysis Reveals Genetic Diversity in Xanthomonads Associated With Poinsettia Production.

Xanthomonas axonopodis pv. poinsettiicola is traditionally identified as the primary causal agent of bacterial leaf spot on poinsettia (family Euphorbiaceae). Sixty-seven strains of xanthomonads isolated from lesions associated with several species within the family Euphorbiaceae were collected over a 64-year period. The pathogenicity of these strains was compared on several potential hosts and they were analyzed by multilocus sequence analysis (MLSA) using six housekeeping genes. The 67 Xanthomonas strains associated with poinsettia production were separated into three distinct clades based on MLSA. The first clade identified contained the X. axonopodis pv. poinsettiicola reference strain (LMG849PT). A second clade was more closely related to X. hortorum pv. pelargonii (LMG7314PT) and the third clade contained the X. codiaei type strain (LMG8678T). This analysis indicated that there may also be other closely related pathovars or species of Xanthomonas that can infect poinsettia. Strains from the three clades could not be distinguished by symptoms or virulence on poinsettia plants. Strains capable of infecting geranium were found in all three clades, although the extent of leaf spot formation and number of systemic infections were significantly less than those produced by X. hortorum pv. pelargonii strains, typically the main causal agent of bacterial leaf spot on geranium. Clade III also contained strains isolated from zebra plant (Aphelandra squarrosa, family Acanthaceae), which is a newly recognized host for X. codiaei and X. axonopodis pv. poinsettiicola. Xanthomonas leaf spot is a serious threat to poinsettia production that can be caused by several Xanthomonas spp. that can infect different ornamental plant hosts. It is imperative that growers maintain a strict sanitation program because reservoirs of inoculum can occur on a number of ornamental hosts.

First Report of Atypical Xanthomonas euvesicatoria Strains Causing Bacterial Spot of Tomato in Nigeria.

Bacterial spot (BS) is an important disease of tomato in Nigeria (2). Although a xanthomonad was isolated from tomato in Nigeria and characterized using phenotypic and pathogenicity tests, the bacterium was not characterized genetically to confirm the species. To determine the species associated with BS, leaves were collected in fields in northwestern Nigeria from tomato plants showing typical BS symptoms, which consisted of dark, irregular-shaped brown leaf spots that coalesced, resulting in a blighted appearance. Isolations from individual lesions were made on nutrient agar (NA). Yellow, mucoid colonies typical of Xanthomonas were isolated from 14 lesions and all were determined to be amylolytic (3). To determine the races of these strains, bacterial suspensions of the tomato strains, derived from 24-h cultures grown on NA at 28°C, were adjusted to 108 CFU/ml and infiltrated into leaves of tomato and pepper differential genotypes (5). The tomato strains elicited hypersensitive reactions (HRs) on the four pepper differential lines and an HR on the tomato genotype FL 216, which contains the R gene Xv3, but elicited susceptible reactions on the tomato genotypes Hawaii 7998 and Bonny Best. These reactions are typical of X. perforans tomato race 3 strains (5). Multilocus sequence analysis (MLSA) of six housekeeping genes (fusA, lacF, gyrB, gltA, gapA, and lepA) was used to further analyze four representative strains (1) (GenBank Accession Nos. KJ938581 to KJ938584, KJ938588 to KJ938591, KJ938595 to KJ938598, KJ938602 to KJ938605, KJ938629 to KJ938632, and KJ938636 to KJ938639, respectively). A partial sequence of hrpB2 was also made since the four Xanthomonas species associated with BS can be differentiated based on sequence divergence of this gene (3) (KJ938609 to KJ938621 and KJ938628). The housekeeping gene sequences were aligned along with other Xanthomonas sequences imported from the National Center for Biotechnology Information (NCBI) database ( www.ncbi.nlm.nih.gov ) using the MUSCLE tool from MEGA software, 5.2.2. Maximum likelihood phylogenetic trees constructed for the six housekeeping gene sequences individually and in concatenation revealed that the strains grouped most closely with the X. euvesicatoria reference strain 85-10 but more distantly to X. perforans. The hrpB2 sequence, which is highly conserved for each Xanthomonas species pathogenic on tomato (4), was sequenced from the tomato strains. These sequences were identical to the hrpB2 sequence from X. perforans strains but different from X. euvesicatoria. Although BS is common in Nigeria, to our knowledge, this represents a unique group of X. euvesicatoria strains from tomato that are identical to X. perforans based on pathogenic reactions on tomato and pepper and hrpB2 sequence identity but are more closely related to X. euvesicatoria based on the six housekeeping gene sequences. References: (1) N. F. Almeida et al. Phytopathology 100:208, 2010. (2) E. U. Opara and F. J. Odibo. J. Mol. Genet. 1:35, 2009. (3) J. B. Jones et al. Syst. Appl. Microbiol. 27:755, 2004. (4) A. Obradovic et al. Eur. J. Plant Pathol. 88:736, 2004. (5) R. E. Stall et al. Annu. Rev. Phytopathol. 47:265, 2009.

First Report of Xanthomonas euvesicatoria Causing Bacterial Spot Disease in Pepper in Northwestern Nigeria.

Bacterial spot (BS) has been reported as an important disease on pepper in Nigeria (4). Xanthomonas campestris pv. vesicatoria was identified as the causal agent using phenotypic and pathogenicity tests; however, X. campestris pv. vesicatoria is a synonym for two genetically distinct groups that have been elevated to the species X. euvesicatoria and X. vesicatoria (2). Furthermore, the latter two species and X. gardneri cause similar diseases on pepper (2). In order to determine the species associated with BS on pepper, leaves with irregular, dark brown lesions were collected from pepper plants in fields from northwestern Nigeria, and isolations were made on nutrient agar (NA). Yellow, mucoid colonies typical of Xanthomonas were isolated. Six strains isolated from pepper were determined to be non-amylolytic. For race determinations, bacterial suspensions of the pepper strains, derived from 24-h cultures grown on NA at 28°C, were adjusted to 108 CFU/ml and infiltrated into leaves of tomato and pepper differential genotypes (5). The six pepper strains elicited HRs on the tomato differential genotypes. The strains produced a susceptible reaction on all pepper differentials and were designated as pepper race 6 (5). Multilocus sequence analysis (MLSA) using six housekeeping genes (fusA, lacF, gyrB, gltA, gapA, and lepA) was used to further analyze the strains (1) (GenBank Accession Nos. KJ938585 to KJ938587, KJ938592 to KJ938594, KJ938599 to KJ938601, KJ938606 to KJ938608, KJ938633 to KJ938635, and KJ938640 to KJ938642). A partial sequence of hrpB2 was also sequenced since the four Xanthomonas species associated with BS can be differentiated based on sequence divergence (3) (KJ938622 to KJ938627). The housekeeping gene sequences were aligned along with other Xanthomonas sequences imported from the NCBI database using muscle tool from MEGA software, 5.2.2. Maximum likelihood phylogenetic trees constructed for the six housekeeping gene sequences individually and in concatenation revealed that the Nigerian pepper strains were identical to the X. euvesicatoria reference strain 85-10. Although BS is common in Nigeria, to our knowledge, this represents the first report for this pepper pathogen in Nigeria. References: (1) N. F. Almeida et al. Phytopathology 100:208, 2010. (3) J. B. Jones et al. System Appl. Microbiol. 27:755, 2004. (4) A. Obradovic et al. Eur. J. Plant Pathol. 88:736, 2004. (2) E. U. Opara and F. J. Odibo. J. Mol. Gen. 1:35, 2009. (5) R. E. Stall et al. Ann. Rev. Phytopathol. 47:265, 2009.

Phenological and phytochemical changes correlate with differential interactions of Verticillium dahliae with broccoli and cauliflower.

Cauliflower (Brassica oleracea var. botrytis subvar. cauliflora) is susceptible to wilt caused by Verticillium dahliae but broccoli (B. oleracea var. italica subvar. cyamosa) is not. Infection of broccoli and cauliflower by a green fluorescent protein-expressing isolate of V. dahliae was examined using epifluorescence and confocal laser-scanning microscopy to follow infection and colonization in relation to plant phenology. Plant glucosinolate, phenolic, and lignin contents were also assayed at 0, 4, 14, and 28 days postinoculation. V. dahliae consistently infected and colonized the vascular tissues of all cauliflower plants regardless of age at inoculation, with the pathogen ultimately appearing in the developing seed; however, colonization decreased with plant age. In broccoli, V. dahliae infected and colonized root and stem xylem tissues of plants inoculated at 1, 2, or 3 weeks postemergence. However, V. dahliae colonized only the root xylem and the epidermal and cortical tissues of broccoli plants inoculated at 4, 5, and 6 weeks postemergence. The frequency of reisolation of V. dahliae from the stems (4 to 22%) and roots (10 to 40%) of mature broccoli plants was lower than for cauliflower stems (25 to 64%) and roots (31 to 71%). The mean level of aliphatic glucosinolates in broccoli roots was 6.18 times higher than in the shoots and did not vary with age, whereas it was 3.65 times higher in cauliflower shoots than in the roots and there was a proportional increase with age. Indole glucosinolate content was identical in both cauliflower and broccoli, and both indole and aromatic glucosinolates did not vary with plant age in either crop. Qualitative differences in characterized glucosinolates were observed between broccoli and cauliflower but no differences were observed between inoculated and noninoculated plants for either broccoli or cauliflower. However, the phenolic and lignin contents were significantly higher in broccoli following inoculation than in noninoculated broccoli or inoculated cauliflower plants. The increased resistance of broccoli to V. dahliae infection was related to the increase in phenolic and lignin contents. Significant differential accumulation of glucosinolates associated with plant phenology may also contribute to the resistant and susceptible reactions of broccoli and cauliflower, respectively, against V. dahliae.

Molecular variation among isolates of Verticillium dahliae and polymerase chain reaction-based differentiation of races.

Verticillium dahliae is a soilborne fungal pathogen that causes vascular wilt in a variety of economically important crops worldwide. There are two races of V. dahliae that infect tomato and lettuce. Although race-1-specific resistance has been identified in both tomato and lettuce, no resistant sources are available for race 2. Molecular analyses were employed to characterize the genetic variability and race structure of 101 isolates of V. dahliae from a variety of hosts, mainly from central and coastal California, and 10 isolates exotic to this area. Analyses of the 16 simple sequence repeat (SSR) markers illustrated that tomato subpopulations from central California were distinct relative to the marigold subpopulations. In contrast, cotton and olive isolates showed admixture with tomato isolates. Analyses of both the ribosomal DNA intergenic spacer regions and SSR markers revealed high genetic variability among isolates but were unable to delineate races of V. dahliae. However, a polymerase chain reaction (PCR) assay was applied to amplify a race-1-specific amplicon from the isolates in many hosts from different geographic areas, and was coupled with virulence assays for validation of the data. Results of the PCR assay showed 100% concordance with the virulence assay to differentiate race 1 from race 2 of 48 isolates from tomato. The results indicate that the PCR assay can be applied to differentiate the two races to support our related aim of breeding host resistance, and further reveal insights into the distribution of races in tomato and lettuce cropping systems in California.

Colonization of resistant and susceptible lettuce cultivars by a green fluorescent protein-tagged isolate of Verticillium dahliae.

Interactions between lettuce and a green fluorescent protein (GFP)-expressing, race 1 isolate of Verticillium dahliae, were studied to determine infection and colonization of lettuce cultivars resistant and susceptible to Verticillium wilt. The roots of lettuce seedlings were inoculated with a conidial suspension of the GFP-expressing isolate. Colonization was studied with the aid of laser scanning confocal and epi-fluorescence microscopes. Few differences in the initial infection and colonization of lateral roots were observed between resistant and susceptible cultivars. Hyphal colonies formed on root tips and within the root elongation zones by 5 days, leading to the colonization of cortical tissues and penetration of vascular elements regardless of the lettuce cultivar by 2 weeks. By 8 to 10 weeks after inoculation, vascular discoloration developed within the taproot and crown regions of susceptible cultivars well in advance of V. dahliae colonization. Actual foliar wilt coincided with the colonization of the taproot and crown areas and the eruption of mycelia into surrounding cortical tissues. Advance colonization of stems, pedicels, and inflorescence, including developing capitula and mature achenes was observed. Seedborne infection was limited to the maternal tissues of the achene, including the pappus, pericarp, integument, and endosperm; but the embryo was never compromised. Resistant lettuce cultivars remained free of disease symptoms. Furthermore, V. dahliae colonization never progressed beyond infected lateral roots of resistant cultivars. Results indicated that resistance in lettuce may lie with the plant's ability to shed infected lateral roots or to inhibit the systemic progress of the fungus through vascular tissues into the taproot.

Effect of Organic Amendments on Soilborne and Foliar Diseases in Field-Grown Snap Bean and Cucumber.

Several paper mills in Wisconsin have programs for spreading paper mill residuals (PMR) on land. A growing number of vegetable farmers recognize the agronomic benefits of PMR applications, but there have been no investigations on the use of PMR for control of vegetable crop diseases. Our objective was to determine the effect of PMR amendments on soilborne and foliar diseases of cucumber and snap bean grown on a sandy soil. Raw PMR, PMR composted without bulking agent (PMRC), or PMR composted with bark (PMRBC) were applied annually in a 3-year rotation of potato, snap bean, and pickling cucumber. Several naturally occurring diseases were evaluated in the field, along with in situ field bioassays. All amendments suppressed cucumber damping-off and Pythium blight and foliar brown spot of snap bean. Both composts reduced the incidence of angular leaf spot in cucumber. In a separate field experiment planted with snap bean for two consecutive years, all amendments reduced common root rot severity in the second year. In a greenhouse experiment, the high rate of PMRBC suppressed anthracnose of snap bean. These results suggest that the application of raw and composted PMR to sandy soils has the potential to control several soilborne and foliar diseases.