When Xcd-lux was coinoculated with the mixture of five guttation bacteria, the size of the Xcd-lux population declined progressively during incubation; the sizes of the populations of Xcd-lux coinoculated with the bacterial mixture 3, 7, and 10 days after inoculation were significantly different (P = 0.01) from the sizes of the corresponding populations when Xcd-lux was inoculated alone (Fig.1A and G). 5). The sizes of populations of Xcd-lux in sterile distilled water and phosphate buffer determined 15 days after inoculation were 6.41 and 5.91 log CFU/ml, respectively. The initial densities of Xcd-lux and total bacteria were 6.34 ± 0.06 and 6.71 ± 0.04 log CFU/ml (means of four replicates), respectively. Below these five bacterial strains are referred to guttation bacteria. Thus, it may be possible to improve the efficacy of a mixture by identifying the trivial strains in the mixture and replacing them with beneficial species. Anthurium ‘New Era’, UH1402, ushers in a new era of bacterial blight–resistant cut flowers suitable for screenhouse production. Effects of guttation bacteria on growth and survival of Xcd-lux in filter-sterilized guttation fluids. This experiment was conducted twice. Such a balanced and self-sustaining bacterial community is ideal for biological control if the same phenomenon can be reproduced in planta. (A) First test with nonwounded leaves. A mixture containing the five guttation bacteria was sprayed onto foliage of cultivar Marian Seefurth plants. Growth and survival of Xcd-lux in guttation fluids from various anthurium cultivars. It was confirmed in this and previous studies that treatment-examiner interactions were not significant when disease severity data were assessed by three examiners (data not shown). It was reported previously that the inhibition of Erwinia amylovora by antibiotics produced by strains of E. herbicola was reduced in the presence of various amino acids (31). Effects of mineral nutrients (concentration, 100 μM) added to guttation fluid on the inhibition of Xcd-lux by guttation bacteria. Anthurium and Onion bacterial blight; Back to the list. For comparison (as a second control), guttation fluid inoculated with only Xcd-lux (plus 15 μl of sterile distilled water and 15 μl of phosphate buffer) was prepared in order to examine the survival of Xcd-lux when guttation bacteria and nutrients were not added. Effects of guttation bacteria on suppression of foliar infection by Xcd-lux.Pretreatment of anthurium leaves with mixtures of guttation bacteria significantly reduced infection by Xcd-lux of both intact (nonwounded) and wounded (notched) leaves (Fig.8). Individual guttation fluids typically contained five to eight predominant bacterial species, as judged by colony types and morphology observed on TZC medium. Do not add … The populations of the individual strains remained near the initial inoculum levels for at least 14 days. More importantly, the observation that a complex nutrient source (peptone) was more efficacious than a single carbon source (glucose) in reversing the inhibition indicated that survival of the pathogen in the presence of guttation bacteria may be affected by the number and kinds of nutrient sources present in the guttation fluid. 3 p. (Commodity Fact Sheet; CFS-AN-4A). The average values calculated from the data collected by the three examiners (percentage data) were transformed by the arcsine transformation and then analyzed by analysis of variance. Data points represent means of four replicates. Samples obtained from the same leaf were pooled in a sterile glass tube and stored at 5°C until the amount of guttation fluid exceeded 4 ml for all plants. There are few publications that report biocontrol studies on the ability of antagonistic bacteria strains to inhibit the pathogens of anthurium blight. Watering your anthurium plants by placing six ice cubes on the soil and allowing them to melt once per week keeps the leaves from getting wet. More studies on the effects of carbon and nitrogen sources on disease suppression by guttation bacteria should provide key information which can be used for biological control of anthurium blight with mixtures of bacterial species. In July 2007, symptoms of bacterial blight were observed on leaves of anthurium plants growing in a commercial greenhouse in central Poland. Moreover, only the pathogen was eliminated from a mixture containing the pathogen and the five guttation bacteria, and the populations of the five guttation bacteria were sustained for 14 days in the guttation fluid. Mixture A consisted of the five guttation bacteria (GUT3, GUT4, GUT5, GUT6, and GUT9). Negative images of bioluminescence emission from infected leaves were scanned with a computer and converted to positive images by using Adobe Photoshop (Adobe Systems Inc., Mountain View, Calif.). dieffenbachiae [27]), is an important disease in Hawaii, as well as other tropical and subtropical regions. Disease incidence was approximately 10% at the time of inspection. The results of two repeated experiments indicated that nonfiltered guttation fluids from cultivar Marian Seefurth were more inhibitory than nonfiltered guttation fluids from cultivar ARCS, Kalapana, or Tropic Mist. Growing plants under plastic or glass houses coupled with drip irrigation rather than overhead or sprinkler irrigation reduced the spread of the bacteria through aerosols and water splash and significantly reduced the incidence of blight in anthurium seedling culture (29). The inhibitory effects of mixture C (containing five other strains obtained from cultivar Marian Seefurth) and mixture F (containing five strains obtained from cultivars Ellison Onizuka and Nitta) were similar to the inhibitory effects of mixture A. Then 15 μl of an Xcd-lux cell suspension and 15 μl of a cell suspension containing the guttation bacteria were inoculated into the guttation fluid in order to determine the survival of Xcd-lux in the guttation fluid in the presence of each mineral nutrient (final concentration, 100 μM). This indicated that there would be no bacterial blight re-infection even if old pots were used immediately even without … The tubes were incubated and the cell densities of Xcd-lux and the guttation bacteria were determined 0, 1, 3, 7, and 14 days after inoculation as described above. ASM journals are the most prominent publications in the field, delivering up-to-date and authoritative coverage of both basic and clinical microbiology. Inhibitory effects of various bacterial mixtures on growth of Xcd-lux in filter-sterilized guttation fluid. Survival of Xcd-lux in guttation fluids of anthurium plants and isolation of inhibitory bacterial strains.Guttation fluids were collected from cultivar ARCS, Marian Seefurth, and UH1060 plants (eight plants per cultivar). The bars represent the means of 10 or 12 observations. Survival of Xcd-lux in guttation fluids of anthurium plants and isolation of inhibitory bacterial strains. The same principle may apply for the enhanced survival of Xcd-lux in guttation fluid containing peptone. Cell suspensions of Xcd-lux and a mixture containing the five guttation bacteria were inoculated into two tubes containing filter-sterilized guttation fluid from cultivar Marian Seefurth. Resident bacterial communities in the guttation fluids of various anthurium cultivars were highly inhibitory to the anthurium blight pathogen, X. campestris pv. The plants were removed from the bags at night and placed in a glasshouse to allow slow drying of the leaves. bacterial leaf blight of anthurium Supawadee Kumsingkaew* and Angsana Akarapisan Department of Entomology and Plant Pathology, Faculty of Agriculture, Chiang Mai University, Chiang Mai 50200, Thailand Supawadee Kumsingkaew and Angsana Akarapisan (2014) Efficiency of Bacillus subtilis EPB14 as biocontrol to control bacterial leaf blight of anthurium. Bacterial Blight. An outbreak of bacterial blight in the 1980s had a severe impact on Hawaii’s local anthurium industry (21, 22). Values marked by asterisks were significantly different (P = 0.01) from the corresponding values for Xcd-lux inoculated alone, as determined by the SNK test. Progression of foliar infection by Xcd-lux in bacterium-treated and nontreated anthurium leaves, as monitored by bioluminescence. When guttation fluids were filter sterilized, the sizes of the populations of the pathogen were not significantly reduced for at least 14 days. The daily minimum and maximum temperatures in the glasshouse were 18 to 22 and 26 to 30°C, respectively. The first is changing how they are watered. On the next day, leaves were wounded by notching them (arrowheads), and the same bacterial mixture was placed on the wounds. The tubes were incubated at 28°C as described above. Mixture F consisted of two strains isolated from cultivar Ellison Onizuka and three strains isolated from cultivar Nitta. The nonwounded plants were treated in the same way, as described above. Xanthomonas blight on anthuriums is caused by Xanthomonas campestris pv. campestris and X. campestris pv. Disease incidence was approximately 10% at the time of inspection. Cultivar Marian Seefurth is highly susceptible to foliar infection, and the other three cultivars are resistant (5). Treatments which were included in this test on A. andraeanum were water treated controls (inoculated and noninoculated), two rates of fosetyl aluminum in two formulations (Aliette 80WP and Aliette … and Ochrobactrum sp. Alternative methods of disease control are needed to ensure protection of the crop from future disease outbreaks. For each day, bars marked by the same letter are not significantly different (P = 0.01), as determined by the SNK test. In this test, the cell densities of the five guttation bacteria were determined individually on the basis of the different colony morphologies of the bacteria on TZC medium containing 100 μg of cycloheximide per ml. Peptone and yeast extract significantly (P = 0.01) increased the number of total bacteria. Bars marked by the same letter were not significantly different (P = 0.01), as determined by the SNK test. An equivalent amount of the nonfiltered guttation fluid was placed in a second tube. (F) Xcd-lux inoculated with GUT9. The experiment was repeated by using six cultivar Marian Seefurth plants per treatment. Effects of guttation bacteria on survival of Xcd-lux in the filter-sterilized guttation fluid.The size of the Xcd-lux population in the filter-sterilized guttation fluids remained close to the initial population size in the absence of guttation bacteria (Fig.1A). At 7 days after inoculation, the size of the population of Xcd-lux in the guttation fluid containing peptone (in the presence of guttation bacteria) was significantly greater (P = 0.01) than the size of the population in the absence of guttation bacteria (in the absence of additional nutrients). This suggests that there are key component strains (species) in a bacterial community that are responsible for inhibition and that a lack of the key organisms in bacterial mixtures eliminates the inhibitory effects on the pathogen. The five guttation bacteria found in this study appear to be common bacterial species indigenous to anthurium leaves. Filtration also removes other microorganisms, such as fungi, algae, and protozoans, from guttation fluids. The differences in the initial sizes of the populations of all bacteria were not significant for cultivars, as determined by the SNK test. analysis, and a Biolog MicroPlate system (Biolog, Inc., Hayward, Calif.) analysis. The two other strains were identified as nonfluorescent pseudomonads. means you agree to our use of cookies. Images of bioluminescence emission from the leaves recorded on X-ray film revealed that infection was initiated at the wound sites and advanced rapidly into the vascular tissues in nontreated leaves (Fig.9). It is, however, impossible to treat the disease. 7). Guttation fluids were collected from leaves that produced more than 500 μl overnight (six, six, and two cultivar ARCS, Marian Seefurth, and UH1060 samples, respectively), and 500 μl of each fluid was placed in a sterile test tube (100 by 13 mm) and used to determine the effect of the fluid on the growth of Xcd-lux. The pathogen, X. campestris pv. The images represent the leaves analyzed in the first trial, which had the disease severity indices closest to the average values. The effects of the filtered guttation fluids on Xcd-lux were examined by determining the number of CFU per milliliter after 0, 1, 3, and 7 days of incubation at 28°C. For comparison, two tubes containing filtered guttation fluid which had not been inoculated previously with any bacteria were incubated with Xcd-lux. Novelanthurium hybrids produced by tissue culture will be indexed for disease and nematode … All of the plants were later inoculated with Xcd-lux. The tubes were incubated as described above. In July 2007, symptoms of bacterial blight were observed on leaves of anthurium plants growing in a commercial greenhouse in central Poland. Populations of beneficial bacteria decline after foliar application on anthurium plants. A recent report that bacterial blight occurs in The Netherlands and that the pathogen was isolated from propagative materials en route from The Netherlands to India (19) indicates that the disease is not restricted to tropical and subtropical regions. Various biological factors may have affected bacterial strains on the leaf surface; these factors include survival, mobility, and subsequent colonization of the hydathodes. The estimated size of the initial inoculum of Xcd-lux was 6.41 ± 0.09 log CFU/ml (mean of eight observations). In: Proceedings of 6th international conference on plant pathogenic bacteria. The missing datum point was estimated by using a general linear model. Four ecofriendly materials viz., turmeric powder impregnated in sodium bicarbonate (0.15%), neem oil (2%), Pseudomonas fluorescens (a proprietary product at 1.5%) and cow dung extract (7.5%) were compared with streptocycline (100 µg ml-1) and Captan (0.3%) for their efficacy in controlling bacterial blight of anthurium. Generally, using cultural controls is not as effective for bacterial leaf spot diseases as for some other diseases like Botrytis blight and downy mildew. The effect of guttation bacteria on disease suppression was more evident in notched leaves than in intact leaves. There is no actual treatment for bacterial blight. The white background illumination is bioluminescence from Xcd-lux recorded on X-ray film. This research was supported by the U. S. Department of Agriculture Special Grants Program for Tropical and Subtropical Agricultural Research (agreement no. This cultivar is known to be highly susceptible to bacterial blight (5). When Xcd-lux was inoculated into filter-sterilized guttation fluids from cultivar Marian Seefurth in which the mixture of five strains or individual indigenous strains had been grown for 14 days before the preparation was filter sterilized, the size of the Xcd-lux population dropped from the initial level (7.09 ± 0.05 log CFU/ml) only to 6.73 ± 0.20 log CFU/ml (for the mixture) or 7.04 ± 0.07 log CFU/ml (for GUT5) after 7 days of incubation. The estimated size of the initial inoculum of Xcd-lux was 6.69 ± 0.08 log CFU/ml (mean of seven observations). 1985. Bars marked by the same letter were not significantly different (P = 0.01), as determined by the SNK test. Effects of guttation bacteria on growth and survival of Xcd-lux in filter-sterilized guttation fluids.Guttation fluids were collected from plants of four different cultivars (cultivars Marian Seefurth, ARCS, Kalapana, and Nitta); the fluids collected from each cultivar were pooled and then filter sterilized. Spraying guttation bacteria onto intact leaves reduced the disease severity index to approximately two-thirds the value obtained for nontreated leaves by day 41 (Fig. 3). dieffenbachiae which also causes leaf spot and blight diseases of many other aroids. While much is known about biochemical and physiological events in host-bacterium interactions, biotic factors in guttation fluids have been inadequately studied. Twenty-eight bacterial isolates from rhizospheric regions were identified as different Bacillus spp. Statistical analysis.The data from the in vitro tests performed to determine the inhibition of Xcd-lux growth in the guttation fluids (and the data for the total bacterial population) were analyzed by analysis of variance. The disinfested leaves were each covered with a clean plastic bag in the evening, and the plants were watered. Hara AH, Tsang MMC, Jacobsen CM, Yogi-Chun JAT, Hata TY, Niino-DuPonte RY (2004) Pest management strategies for anthuriums. This phenomenon was observed more frequently with some cultivars (e.g., cultivars ARCS and UH1060) than with others. No mixture or pair of other leaf-inhabiting xanthomonads (X. campestris pv. Then, the cell suspensions were mixed at different ratios to prepare four replicates (1:2:1:2:1, 2:1:2:1:2, 1:2:2:1:2, and 2:1:1:2:1 for mixtures consisting of five strains; 1:2:1:2, 2:1:2:1, 1:2:2:1, and 2:1:1:2 for mixture E consisting of only four strains), and 15 μl of each mixture was inoculated into 1.47 ml of filter-sterilized guttation fluid from cultivar Marian Seefurth. dieffenbachiae []), is an important disease in Hawaii, as well as other tropical and subtropical regions.An outbreak of bacterial blight in the 1980s had a severe impact on Hawaii’s local anthurium … Use of the bioluminescent strain has also allowed accurate evaluation of cultivar susceptibility in the foliar infection phase without dependence on symptom expression (5). Strains GUT3, GUT4, GUT5, GUT6, and GUT9 were grown on YDC medium plates for 2 days, and cells of each strain were suspended in sterile distilled water and adjusted to an optical density at 600 nm of 0.1 (cell densities, ∼1.0 × 108 CFU/ml). Honolulu (HI): University of Hawaii. Management is the only avenue. The contribution of Keoki N. Nunies to this project is acknowledged. After 2 weeks, all of the guttation fluid samples were individually filter sterilized, and 1.5 ml of each filtered sample was inoculated with 15 μl of a suspension of Xcd-lux cells. As a control, sterile distilled water was added to the guttation fluid. Effects of guttation bacteria on survival of Xcd-lux in the filter-sterilized guttation fluid. Symbols: ●, Xcd-lux; ○, GUT3; ▵, GUT4; ×, GUT5; □, GUT6; ▴, GUT9. The initial inoculum size was 7.00 log CFU/ml, and the size of the population progressively declined to 4.15 ± 1.16 log CFU/ml for cultivar Marian Seefurth (six samples), to 4.81 and 6.46 log CFU/ml for cultivar UH1060 (two samples), and to 5.94 ± 0.44 log CFU/ml for cultivar ARCS (six samples) after 7 days of incubation. In bacterium-treated leaves, in contrast, there was no evidence that infections advanced from the wound sites, but infection through hydathodes at the leaf margins was evident (Fig. Inhibitory effects of various bacterial mixtures on growth of Xcd-lux in filter-sterilized guttation fluid.All six bacterial mixtures that were added to filter-sterilized guttation fluids significantly (P = 0.01) reduced the sizes of the populations of Xcd-lux during 8 days of incubation in filter-sterilized guttation fluid. dieffenbachiae in guttation fluids (xylem sap exuded from leaf margins) of anthuriums were suppressed by several bacterial strains indigenous to leaves of various anthurium cultivars. Inhibition of growth was not observed in filter-sterilized guttation fluids and was restored to original levels only by reintroducing specific mixtures of bacteria into filter-sterilized guttation fluids. Individual guttation bacteria had no effect on the growth or survival of Xcd-lux when they were coinoculated into the filter-sterilized guttation fluids (Fig. Bars = 5 cm. How to Treat Bacterial Blight. Enter multiple addresses on separate lines or separate them with commas. This devastating disease has limited anthurium production not only in Hawaii, but throughout the world where anthuriums are produced. based on standard bacteriological tests (9, 23), a fatty acid analysis, an API-NFT system (bioMérieux Vitek, Inc., Hazelwood, Mo.) Then, 2 ml of each subsample was sterilized by filtration, and 1.485 ml was placed in a sterile test tube. The proposed research isdesigned to bring all components of an integrated pest management program together. Four replicate samples (one for each cultivar) were used for each strain and for the mixture. using 16S rRNA gene sequencing. Survival of Xcd-lux in filter-sterilized and nonsterile guttation fluids from various anthurium cultivars.Guttation fluids were collected from cultivar Alii, ARCS, Ellison Onizuka, Kalapana, Marian Seefurth, Nitta, Tropic Mist, and UH1060 plants (four plants per cultivar). The initial population of total bacteria in each guttation fluid was determined by dilution plate counting on TZC medium containing 100 μg of cycloheximide per ml. The initial densities of Xcd-lux and total bacteria were 6.35 ± 0.04 and 6.72 ± 0.05 log CFU/ml (means of four replicates), respectively. For comparison, 15-μl portions of the suspension were inoculated into equivalent amounts of sterile distilled water and phosphate buffer (two tubes each). The anthurium industry is important to the State of Hawaii but production of anthurium cut flowers isreduced by bacterial blight disease and damage by burrowing nematodes. 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