Papers by Hung Kim Doan
Fusarium oxysporum f. sp. vasinfectum (FOV) race 4 is a seedborne pathogen of cotton. A series of... more Fusarium oxysporum f. sp. vasinfectum (FOV) race 4 is a seedborne pathogen of cotton. A series of seed treatments using hot water at various temperatures (55 to 90°C) for various lengths of time (105 seconds to 20 min) was tested on seed of both Pima (Gossypium barbadense) and Upland (G. hirsutum) cultivars to determine the optimum combination to eliminate the pathogen from seed without reducing seed germination or vigor. In 2012, the incidence of FOV race 4 in an Upland seed lot was reduced by ~56% without reducing seed germination and vigor when FOV race 4-infected cotton seed was immersed in a hot water bath for 20 min at 60°C. In 2013, infected Upland seed was pretreated at 23°C and agitated on a shaker at 100 rpm for 1 hour in sterile deionized water or sterile 30% potato dextrose broth (PDB). The seed was then immersed for 20 min at 60°C in one of the following: water, 10% solution of household bleach (0.8% NaOCl, w/v), 0.5% sodium lauryl sulfate, azoxystrobin (0.24 g a.i.), fludioxonil (0.50 g a.i.), thiabendazole (0.42 g a.i.), thiophanate-methyl (0.70 g a.i.), or a fungicide bath containing all four fungicides. A pretreatment immersion in PDB or water followed by a 20 minute immersion in a 60°C fungicide slurry containing all four fungicides or thiophanate-methyl alone were the most effective pretreatment-treatment combinations in reducing FOV (by ~85%) in seed and avoiding loss of seed germination and vigor based on recovery of the fungus on petri plates and greenhouse grow-out assays. FOV was completely eliminated from infected seed when the seed was immersed in water at 23°C for 1 hour at 100 rpm followed by a 20 minute immersion in a solution of thiophanate-methyl heated to 70°C, but seed germination was reduced by 36% and vigor was reduced by 38%.
In the summer of 2012, an outbreak of a newly discovered root and basal stalk rot of wild rice (Z... more In the summer of 2012, an outbreak of a newly discovered root and basal stalk rot of wild rice (Zizania palustris L.) cv. Franklin was observed in a 16-ha field in Big Valley, Lassen County, California (GPS coordinates 41°08′41.93″ N 121°10′07.49″ W). Infected plants exhibiting rot and dieback of roots and stalks were in various stages of decline, including death. Symptomatic stem and root tissues from affected plants were surface disinfected in 1% NaOCl for 90 s and placed on PARP agar plates, which were then incubated at 25°C in the dark for 1 week. Hyphal tips were used to start and maintain the organism in pure cultures. Isolates were transferred into petri plates with water and sterilized blades of turfgrass for the production of hyphae and reproductive structures. Isolates had coenocytic hyphae and produced zoospores 20 to 30 μm in diameter outside of sporangia (75 to 160 × 46 to 110 μm) from a naked mass of protoplasm, unlike from a vesicle, which is characteristic of Pythium spp. (2). Based on these morphological features, the isolate was tentatively identified as a Pythiogeton sp. Total genomic DNA was extracted from mycelia using the DNeasy Plant Mini Kit (Qiagen Inc., Valencia, CA). The internal transcribed spacers (ITS) 1 and 2 flanking the 5.8S rRNA regions were amplified by PCR and sequenced using universal ITS5 and ITS4 primers. A BLAST search of the 855-bp sequences revealed 98% similarity with a sequence of P. ramosum isolate Pg-164 (GenBank Accession No. JQ610190.1). The 21 nucleotide differences suggest that the isolate from wild rice may be an unreported species. The sequences were submitted to GenBank (KF719169). To fulfill Koch's postulates and confirm pathogenicity, 100 wild rice seeds were surface disinfected in 1% NaOCl for 90 s and placed in a 500 ml sterile pot with 250 g of autoclaved sand. Three 5 mm-diameter disks from the margin of a 7-day-old culture growing on PARP were placed in each of five pots. As a control, three 5 mm-diameter disks from a non-inoculated PARP plate were placed in five different pots, and five pots with autoclaved sand were not inoculated. All pots were kept in a randomized complete block design at 25°C for 14 days under a 14-h photoperiod. The pathogenicity test was repeated three times. After 14 days, the inoculated plants in all tests developed root and basal stalk rot, consistent with the symptoms observed on diseased wild rice in the field. The Pythiogeton sp. was consistently re-isolated on PARP from symptomatic plants but not from control plants. The non-inoculated wild rice plants remained asymptomatic. DNA sequences of the ITS region of the re-isolated Pythiogeton sp. revealed 100% identity with the isolate from the field. There have been reports of P. zeae on corn in Korea and P. zizaniae on water bamboo in Taiwan (1,2,3). This is the first report of a Pythiogeton sp. on wild rice.
Fusarium wilt of cotton, caused by the soilborne
fungus Fusarium oxysporum f. sp. vasinfectum (F... more Fusarium wilt of cotton, caused by the soilborne
fungus Fusarium oxysporum f. sp. vasinfectum (FOV),
is a widespread and devastating disease occurring
in most cotton-growing regions of the world. The
most successful strategy to manage Fusarium wilt is
the use of resistant cultivars. Recently, two sources
of germplasm, MD25-26ne and MD25-27, which
have superior yield, fiber quality, and resistance
against Fusarium wilt and root-knot nematodes
(Meloidogyne spp.), were released. The purpose
of this study was to evaluate the response of eight
Upland cotton (including MD25-26ne and MD25-
27) and two Pima cultivars (susceptible and resistant
controls) against seven genotypes of FOV (races
1, 2, 3, 4, and 8 and genotypes LA108 and LA140)
in a greenhouse trial. To inoculate plants, roots of
2-wk-old seedlings of each cultivar were dipped for
4 min in a conidial suspension (106 conidia/mL) of
each FOV genotype. Based on dry shoot weights
and extent of vascular discoloration; Phytogen 800
followed by MD25-26ne and MD25-27 were the
most resistant cultivars against multiple genotypes
of FOV. FOV race 4 caused mild symptoms on PHY
800, PHY 98M-2983, MD25-26ne, and MD25-27 but
caused severe symptoms on DP 744 and Rowden. In
addition, Rowden was highly susceptible to FOV
races 1, 2, and 8 and LA108 and LA140. Phytogen
elite lines PHY x1, PHY x2, and PHY x3 were
moderately susceptible to multiple genotypes of
FOV. These data indicate that PHY 800, MD25-26ne,
and MD25-27 can be recommended for planting in
fields with previous histories of Fusarium wilt and
can be used in breeding programs as sources of
resistance to Fusarium wilt.
Phytopathology, Jan 20, 2015
The study of microorganisms that reside on plant leaf surfaces, or phyllosphere microbiology, gre... more The study of microorganisms that reside on plant leaf surfaces, or phyllosphere microbiology, greatly benefits from the availability of artificial surfaces that mimic in one or more ways the complexity of foliage as a microbial habitat. These leaf surface proxies range from very simple, such as nutrient agars that can reveal the metabolic versatility or antagonistic properties of leaf-associated microorganisms, to the very complex, such as silicon-based casts that replicate leaf surface topography down to nanometer resolution. In this review, we summarize the various uses of artificial surfaces in experimental phyllosphere microbiology and discuss how these have advanced our understanding of the biology of leaf-associated microorganisms and the habitat they live in. We also provide an outlook into future uses of artificial leaf surfaces, foretelling a greater role for microfluidics to introduce biological and chemical gradients into artificial leaf environments, stressing the import...
A rapid and reliable molecular diagnostic assay, AmplifyRP Acceler8, was developed for the direct... more A rapid and reliable molecular diagnostic assay, AmplifyRP Acceler8, was developed for the direct detection of Fusarium oxysporum f. sp. vasinfectum (FOV) race 4, a virulent genotype of the Fusarium wilt pathogen of cotton (Gossypium spp.), in soil and cotton tissue. Unlike traditional polymerase chain reaction (PCR) assays, the recombinase polymerase amplification based-assay described here utilizes an advanced isothermal technology where the amplification is carried out at a single constant temperature, 39°C, without the need of a thermal cycler. The AmplifyRP Acceler8 diagnostic assay consistently detected FOV race 4 from all infected tissue samples. The test is rapid, simple, and more sensitive than conventional PCR. The AmplifyRP Acceler8 diagnostic assay detected DNA from FOV race 4 at concentrations of 1 ng/μL and above. In addition, it did not amplify DNA from otherknown FOV races (races 1, 2, 3, 6, and 8). The whole process from sample preparation to reading the results can be completed in as little as 30 min. The test can detect FOV race 4 from cotton taproots, petioles, and stems.
Conference Presentations by Hung Kim Doan
The porosity of the outer poultry eggs shell allows pathogens to arrive inside egg by penetrating... more The porosity of the outer poultry eggs shell allows pathogens to arrive inside egg by penetrating the eggshell. Under
humid ambient storing conditions in refrigerators, eggs can be colonized by fungi and bacteria. Volatile organic
compounds (VOCs) are produced by bacteria and fungi as they proliferate, and these chemicals are emitted back
through the eggshell into the environment. Some of these compounds have low molecular weight and high volatility,
and almost 200 VOCs have been previously identified from contaminated eggs. In this work, we present the study of
metabolites from fungi inside hen eggs and their correlation with the fungi species-specific VOCs. Fresh-eggs(n=2)
and 2months old-eggs(n=4) were stored under normal ambient humid conditions, and they were analyzed using noninvasive
techniques: Solid Phase Microextraction Headspace Gas Chromatography Mass Spectrometry(SPME-HSGC/
MS) and Headspace Gas Chromatography Differential Mobility Spectrometry(HS-GC/DMS). Eggs were placed
inside vials (neck-size-70mm,PTFE-cap,capacity-240ml) that were previously cleaned in a vacuum-oven(160ºC,3h),
and samples were taken after flushing the vials with dryUHP air for 2min. Upon experimetns completion, all eggs
were opened to check fungus presence and identification. DNA was extracted from mycelia of fungal cultures on
APDA, and bacterial colonies that were grown on KB medium. PCR products were sequenced and the fungi and
bacteria were identified using NCBI BLAST results. Petri dishes with grown fungi from eggs were analyzed with
SPME-GC/MS for metabolite confirmation. Almost 100 VOCs have been identified with GC/MS and are classified in
5-groups: fungi (present only in the eggs containing fungi); old (present only in the old eggs that may or may not
contain the fungi); healthy (present only in the eggs not containing the fungi); fresh (present only in the fresh eggs)
and all eggs (present in all eggs). Specific compounds that have been previously attributed to the identified fungi
(Botrytis cinerea and Cladosporium macrocarpum) have been matched with the identified compounds from the fungiinfected
eggs as well as from the analyzed colonies that were grown in vitro in petri-dishes. GC/MS results allow for
clear differentiation between non-infected eggs and eggs infected with fungi. Similarly to GC/MS results,
approximately 100 VOCs that are present in the eggs and absent in the vials with dry air blanks have been identified
in positive ion mode with GC/DMS. Additionally, 4 VOCs have been identified in the negative ion mode. The
advantage of GC/DMS is that it can detect both positive and negative ions in a single measurement, while only one
ion mode could be used at a time in some mass spectrometers . Although chemical identification with GC/DMS
sensor may be not straightforward or not possible, the sensor can be useful as a VOC monitoring tool in production
processes. Visual inspection of tested eggs revealed differences between non-infected eggs and eggs infected with
fungi. Non-invasive sampling of eggs pathogens. GC/DMS chemical signatures to be potentially used in production
process.
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Papers by Hung Kim Doan
fungus Fusarium oxysporum f. sp. vasinfectum (FOV),
is a widespread and devastating disease occurring
in most cotton-growing regions of the world. The
most successful strategy to manage Fusarium wilt is
the use of resistant cultivars. Recently, two sources
of germplasm, MD25-26ne and MD25-27, which
have superior yield, fiber quality, and resistance
against Fusarium wilt and root-knot nematodes
(Meloidogyne spp.), were released. The purpose
of this study was to evaluate the response of eight
Upland cotton (including MD25-26ne and MD25-
27) and two Pima cultivars (susceptible and resistant
controls) against seven genotypes of FOV (races
1, 2, 3, 4, and 8 and genotypes LA108 and LA140)
in a greenhouse trial. To inoculate plants, roots of
2-wk-old seedlings of each cultivar were dipped for
4 min in a conidial suspension (106 conidia/mL) of
each FOV genotype. Based on dry shoot weights
and extent of vascular discoloration; Phytogen 800
followed by MD25-26ne and MD25-27 were the
most resistant cultivars against multiple genotypes
of FOV. FOV race 4 caused mild symptoms on PHY
800, PHY 98M-2983, MD25-26ne, and MD25-27 but
caused severe symptoms on DP 744 and Rowden. In
addition, Rowden was highly susceptible to FOV
races 1, 2, and 8 and LA108 and LA140. Phytogen
elite lines PHY x1, PHY x2, and PHY x3 were
moderately susceptible to multiple genotypes of
FOV. These data indicate that PHY 800, MD25-26ne,
and MD25-27 can be recommended for planting in
fields with previous histories of Fusarium wilt and
can be used in breeding programs as sources of
resistance to Fusarium wilt.
Conference Presentations by Hung Kim Doan
humid ambient storing conditions in refrigerators, eggs can be colonized by fungi and bacteria. Volatile organic
compounds (VOCs) are produced by bacteria and fungi as they proliferate, and these chemicals are emitted back
through the eggshell into the environment. Some of these compounds have low molecular weight and high volatility,
and almost 200 VOCs have been previously identified from contaminated eggs. In this work, we present the study of
metabolites from fungi inside hen eggs and their correlation with the fungi species-specific VOCs. Fresh-eggs(n=2)
and 2months old-eggs(n=4) were stored under normal ambient humid conditions, and they were analyzed using noninvasive
techniques: Solid Phase Microextraction Headspace Gas Chromatography Mass Spectrometry(SPME-HSGC/
MS) and Headspace Gas Chromatography Differential Mobility Spectrometry(HS-GC/DMS). Eggs were placed
inside vials (neck-size-70mm,PTFE-cap,capacity-240ml) that were previously cleaned in a vacuum-oven(160ºC,3h),
and samples were taken after flushing the vials with dryUHP air for 2min. Upon experimetns completion, all eggs
were opened to check fungus presence and identification. DNA was extracted from mycelia of fungal cultures on
APDA, and bacterial colonies that were grown on KB medium. PCR products were sequenced and the fungi and
bacteria were identified using NCBI BLAST results. Petri dishes with grown fungi from eggs were analyzed with
SPME-GC/MS for metabolite confirmation. Almost 100 VOCs have been identified with GC/MS and are classified in
5-groups: fungi (present only in the eggs containing fungi); old (present only in the old eggs that may or may not
contain the fungi); healthy (present only in the eggs not containing the fungi); fresh (present only in the fresh eggs)
and all eggs (present in all eggs). Specific compounds that have been previously attributed to the identified fungi
(Botrytis cinerea and Cladosporium macrocarpum) have been matched with the identified compounds from the fungiinfected
eggs as well as from the analyzed colonies that were grown in vitro in petri-dishes. GC/MS results allow for
clear differentiation between non-infected eggs and eggs infected with fungi. Similarly to GC/MS results,
approximately 100 VOCs that are present in the eggs and absent in the vials with dry air blanks have been identified
in positive ion mode with GC/DMS. Additionally, 4 VOCs have been identified in the negative ion mode. The
advantage of GC/DMS is that it can detect both positive and negative ions in a single measurement, while only one
ion mode could be used at a time in some mass spectrometers . Although chemical identification with GC/DMS
sensor may be not straightforward or not possible, the sensor can be useful as a VOC monitoring tool in production
processes. Visual inspection of tested eggs revealed differences between non-infected eggs and eggs infected with
fungi. Non-invasive sampling of eggs pathogens. GC/DMS chemical signatures to be potentially used in production
process.
fungus Fusarium oxysporum f. sp. vasinfectum (FOV),
is a widespread and devastating disease occurring
in most cotton-growing regions of the world. The
most successful strategy to manage Fusarium wilt is
the use of resistant cultivars. Recently, two sources
of germplasm, MD25-26ne and MD25-27, which
have superior yield, fiber quality, and resistance
against Fusarium wilt and root-knot nematodes
(Meloidogyne spp.), were released. The purpose
of this study was to evaluate the response of eight
Upland cotton (including MD25-26ne and MD25-
27) and two Pima cultivars (susceptible and resistant
controls) against seven genotypes of FOV (races
1, 2, 3, 4, and 8 and genotypes LA108 and LA140)
in a greenhouse trial. To inoculate plants, roots of
2-wk-old seedlings of each cultivar were dipped for
4 min in a conidial suspension (106 conidia/mL) of
each FOV genotype. Based on dry shoot weights
and extent of vascular discoloration; Phytogen 800
followed by MD25-26ne and MD25-27 were the
most resistant cultivars against multiple genotypes
of FOV. FOV race 4 caused mild symptoms on PHY
800, PHY 98M-2983, MD25-26ne, and MD25-27 but
caused severe symptoms on DP 744 and Rowden. In
addition, Rowden was highly susceptible to FOV
races 1, 2, and 8 and LA108 and LA140. Phytogen
elite lines PHY x1, PHY x2, and PHY x3 were
moderately susceptible to multiple genotypes of
FOV. These data indicate that PHY 800, MD25-26ne,
and MD25-27 can be recommended for planting in
fields with previous histories of Fusarium wilt and
can be used in breeding programs as sources of
resistance to Fusarium wilt.
humid ambient storing conditions in refrigerators, eggs can be colonized by fungi and bacteria. Volatile organic
compounds (VOCs) are produced by bacteria and fungi as they proliferate, and these chemicals are emitted back
through the eggshell into the environment. Some of these compounds have low molecular weight and high volatility,
and almost 200 VOCs have been previously identified from contaminated eggs. In this work, we present the study of
metabolites from fungi inside hen eggs and their correlation with the fungi species-specific VOCs. Fresh-eggs(n=2)
and 2months old-eggs(n=4) were stored under normal ambient humid conditions, and they were analyzed using noninvasive
techniques: Solid Phase Microextraction Headspace Gas Chromatography Mass Spectrometry(SPME-HSGC/
MS) and Headspace Gas Chromatography Differential Mobility Spectrometry(HS-GC/DMS). Eggs were placed
inside vials (neck-size-70mm,PTFE-cap,capacity-240ml) that were previously cleaned in a vacuum-oven(160ºC,3h),
and samples were taken after flushing the vials with dryUHP air for 2min. Upon experimetns completion, all eggs
were opened to check fungus presence and identification. DNA was extracted from mycelia of fungal cultures on
APDA, and bacterial colonies that were grown on KB medium. PCR products were sequenced and the fungi and
bacteria were identified using NCBI BLAST results. Petri dishes with grown fungi from eggs were analyzed with
SPME-GC/MS for metabolite confirmation. Almost 100 VOCs have been identified with GC/MS and are classified in
5-groups: fungi (present only in the eggs containing fungi); old (present only in the old eggs that may or may not
contain the fungi); healthy (present only in the eggs not containing the fungi); fresh (present only in the fresh eggs)
and all eggs (present in all eggs). Specific compounds that have been previously attributed to the identified fungi
(Botrytis cinerea and Cladosporium macrocarpum) have been matched with the identified compounds from the fungiinfected
eggs as well as from the analyzed colonies that were grown in vitro in petri-dishes. GC/MS results allow for
clear differentiation between non-infected eggs and eggs infected with fungi. Similarly to GC/MS results,
approximately 100 VOCs that are present in the eggs and absent in the vials with dry air blanks have been identified
in positive ion mode with GC/DMS. Additionally, 4 VOCs have been identified in the negative ion mode. The
advantage of GC/DMS is that it can detect both positive and negative ions in a single measurement, while only one
ion mode could be used at a time in some mass spectrometers . Although chemical identification with GC/DMS
sensor may be not straightforward or not possible, the sensor can be useful as a VOC monitoring tool in production
processes. Visual inspection of tested eggs revealed differences between non-infected eggs and eggs infected with
fungi. Non-invasive sampling of eggs pathogens. GC/DMS chemical signatures to be potentially used in production
process.