Bioassay and baiting methods for determining occurrence of races of Phytophthora clandestina in soil

1996 ◽  
Vol 36 (7) ◽  
pp. 815 ◽  
Author(s):  
A Purwantara ◽  
SP Flett ◽  
PJ Keane
Keyword(s):  
Disease Severity ◽  
Root Rot ◽  
Subterranean Clover ◽  
Soil Samples ◽  
Field Soil ◽  
Flooded Soil ◽  
Aphanomyces Euteiches ◽  
Differential Cultivars ◽  
Pure Cultures ◽  
Race 1

The method currently used for determining races of Phytophthora clandestina requires isolation of pure cultures of the pathogen and testing of their pathogenicity on a range of differential cultivars. To date, the pathogen has not been isolated directly from soil and isolation of the pathogen from naturally infected seedlings is laborious. A bioassay involving the planting of differential cultivars in soil samples in small planting trays was developed to identify races of P. clandestina in soil. The specific races of the pathogen in the soil were determined by assessing the disease severity and the extent of sporulation of the pathogen on the roots of the differential cultivars. A more rapid baiting method using cotyledons of differential cultivars in flooded soil samples was also developed to determine the presence of different races in the samples. Both bioassays were used to confirm the presence of race 0 and race 1 in separate paddocks at Rutherglen, northern Victoria, over 4 seasons. The presence in field soil of another root rot pathogen of subterranean clover, Aphanomyces euteiches, was also detected using these techniques.

Occurrence of Phytophthora clandestina in Trifolium subterraneum paddocks in Australia

2001 ◽  
Vol 41 (2) ◽  
pp. 187 ◽  
Author(s):  
R. Aldaoud ◽  
W. Guppy ◽  
L. Callinan ◽  
S. F. Flett ◽  
K. A. Wratten ◽  
...  
Keyword(s):  
Western Australia ◽  
Root Rot ◽  
New South ◽  
New South Wales ◽  
South Australia ◽  
Subterranean Clover ◽  
Trifolium Subterraneum ◽  
Soil Samples ◽  
South Wales ◽  
Differential Cultivars

In 1995–96, a survey of soil samples from subterranean clover (Trifolium subterraneum L.) paddocks was conducted across Victoria, South Australia, New South Wales and Western Australia, to determine the distribution and the prevalence of races of Phytophthora clandestina (as determined by the development of root rot on differential cultivars), and the association of its occurrence with paddock variables. In all states, there was a weak but significant association between P. clandestina detected in soil samples and subsequent root rot susceptibility of differential cultivars grown in these soil samples. Phytophthora clandestina was found in 38% of the sampled sites, with a significantly lower prevalence in South Australia (27%). There were significant positive associations between P. clandestina detection and increased soil salinity (Western Australia), early growth stages of subterranean clover (Victoria), mature subterranean clover (South Australia), recently sown subterranean clover (South Australia), paddocks with higher subterranean clover content (Victoria), where herbicides were not applied (South Australia), irrigation (New South Wales and Victoria), cattle grazing (South Australia and Victoria), early sampling dates (Victoria and New South Wales), sampling shortly after the autumn break or first irrigation (Victoria), shorter soil storage time (Victoria) and farmer’s perception of root rot being present (Victoria and New South Wales). Only 29% of P. clandestina isolates could be classified under the 5 known races. Some of the unknown races were virulent on cv. Seaton Park LF (most resistant) and others were avirulent on cv. Woogenellup (most susceptible). Race 1 was significantly less prevalent in South Australia than Victoria and race 0 was significantly less prevalent in New South Wales than in South Australia and Western Australia. This study revealed extremely wide variation in the virulence of P. clandestina. The potential importance of the results on programs to breed for resistance to root rot are discussed. in South Australia.

scholarly journals A Rapid Method Using PCR-Based SCAR Markers for the Detection and Identification of Phoma sclerotioides: The Cause of Brown Root Rot Disease of Alfalfa

Plant Disease ◽  
2002 ◽  
Vol 86 (9) ◽  
pp. 928-932 ◽  
Author(s):  
R. C. Larsen ◽  
C. R. Hollingsworth ◽  
G. J. Vandemark ◽  
M. A. Gritsenko ◽  
F. A. Gray
Keyword(s):  
Root Rot ◽  
Random Amplified Polymorphic Dna ◽  
Dna Amplification ◽  
Pythium Ultimum ◽  
Soil Samples ◽  
Root Tissue ◽  
Aphanomyces Euteiches ◽  
Amplification Product ◽  
Phoma Sclerotioides ◽  
Brown Root Rot

A rapid technique for identification and detection of Phoma sclerotioides, the causal agent of brown root rot of alfalfa, has been developed using polymerase chain reaction (PCR). Amplification products obtained from random amplified polymorphic DNA (RAPD) reactions were cloned and sequenced, and two extended primer sets were designed from the resulting data that were used to detect sequence-characterized DNA markers. A single 499-bp DNA amplification product was consistently obtained from primers PSB12499 that was specific for 19 isolates of P.sclerotioides but was not produced from Phoma medicaginis or Phoma betae, or from other soilborne pathogens including Aphanomyces euteiches, Rhizoctonia solani, Fusarium oxysporum, Pythium ultimum, or Phytophthora infestans. A 499-bp amplification product was also produced from root tissue known to be infected with the fungus as verified by microscopic examination. A similar PCR product was obtained from soil samples collected from fields with an established infection of P. sclerotioides on alfalfa. This PCR-based assay enables detection of P. sclerotioides from alfalfa root tissue and in soil samples in a single day, including extraction of DNA, compared with standard methods that require up to 100 days for identification using agar media.

scholarly journals Quantifying Aphanomyces euteiches in Alfalfa with a Fluorescent Polymerase Chain Reaction Assay

2002 ◽  
Vol 92 (3) ◽  
pp. 265-272 ◽  
Author(s):  
G. J. Vandemark ◽  
B. M. Barker ◽  
M. A. Gritsenko
Keyword(s):  
Polymerase Chain Reaction ◽  
Disease Severity ◽  
Pcr Analysis ◽  
Aphanomyces Euteiches ◽  
Chain Reaction ◽  
Plant Samples ◽  
Race 1 ◽  
Polymerase Chain ◽  
Race 2 ◽  
Pathogen Dna

A polymerase chain reaction (PCR) assay using a set of specific primers and a dual-labeled probe (TaqMan) was developed to quantify the amount of Aphanomyces euteiches DNA in alfalfa plants exhibiting varying levels of disease severity. The study included isolates of race 1 and race 2 of A. euteiches. The assay also discriminated between alfalfa populations for resistance based on analysis of DNA extracted from bulked plant samples. Analysis of individual plants and bulked plant samples of standard check populations with both pathogen isolates resulted in Spearman rank correlations between pathogen DNA content and disease severity index ratings that were greater than 0.75 and highly significant (P < 0.0005). In experiments with a race 1 isolate, the amount of pathogen DNA present in the resistant check WAPH-1 was significantly less than in the susceptible check Saranac. In experiments with a race 2 isolate, the amount of pathogen DNA in the resistant check WAPH-5 was significantly less than in either of the susceptible checks, Saranac and WAPH-1. Discrimination between commercial cultivars based on quantitative PCR analysis of bulked plant samples was similar to classification based on visual assessment of disease severity.

scholarly journals Inoculation with Nonpathogenic Fusarium solani Increases Severity of Pea Root Rot Caused by Aphanomyces euteiches

Plant Disease ◽  
2002 ◽  
Vol 86 (4) ◽  
pp. 411-414 ◽  
Author(s):  
R. D. Peters ◽  
C. R. Grau
Keyword(s):  
Disease Severity ◽  
Fusarium Solani ◽  
Root Rot ◽  
Aphanomyces Euteiches ◽  
Disease Symptoms ◽  
Pea Root ◽  
Pea Seedlings ◽  
Pea Root Rot

Aphanomyces euteiches is an important root-rotting pathogen of pea. When recovering isolates of A. euteiches from infested soils in Wisconsin using pea as a bait host, isolates of Fusarium solani often were recovered. Experiments were established to compare disease symptoms of pea seedlings inoculated with isolates of A. euteiches and F. solani alone or in combination. Inoculation of pea seedlings with either of two isolates of A. euteiches produced typical root rot symptoms. However, inoculation of pea seedlings with an isolate of F. solani resulted in no disease symptoms, indicating that the isolate was nonpathogenic to pea. Co-inoculation of pea seedlings with A. euteiches and the nonpathogenic isolate of F. solani resulted in significantly (P = 0.05) greater disease severity than inoculation with A. euteiches alone. Both A. euteiches and F. solani could be reisolated, individually or together, from pea seedlings following individual or co-inoculations, respectively. Although the mechanisms of interaction between these two species are unknown, the synergism documented in this study indicates that the interactions of pathogens with nonpathogens may affect development of disease symptoms.

scholarly journals Characteristics and Frequency of Aphanomyces euteiches Races 1 and 2 Associated with Alfalfa in the Midwestern United States

Plant Disease ◽  
2001 ◽  
Vol 85 (7) ◽  
pp. 740-744 ◽  
Author(s):  
D. K. Malvick ◽  
C. R. Grau
Keyword(s):  
Root Rot ◽  
High Frequency ◽  
Rapd Analysis ◽  
Random Amplified Polymorphic Dna ◽  
Aphanomyces Euteiches ◽  
History Of ◽  
Race 1 ◽  
Race 2 ◽  
Genotypic Characteristics ◽  
Distribution Frequency

Aphanomyces root rot of alfalfa, caused by Aphanomyces euteiches, kills seedlings and causes decline of established plants in slowly drained soils. Two races of A. euteiches that are pathogenic to alfalfa have been identified. Despite the contribution of race 1 resistance to establishment and yield of alfalfa, race 1-resistant alfalfa cultivars perform poorly in some fields infested with A. euteiches. Many isolates of A. euteiches obtained from the soils of problematic fields are of a race 2 phenotype. The purpose of this study was to determine distribution, frequency, and pathogenic and genotypic characteristics of race 1 (R1) and race 2 (R2) isolates from 21 fields: 13 in Wisconsin, 7 in Minnesota, and 1 in Kentucky. A. euteiches was successfully isolated from the soil of 16 of the 21 fields; 405 isolates were obtained from Wisconsin, 4 from Minnesota, and 48 from Kentucky. Pathogenicity and race phenotype of isolates were characterized on Saranac (susceptible to R1 and R2 isolates) and WAPH-1 (resistant to R1 and susceptible to R2 isolates) alfalfa populations. One Wisconsin field with no recent history of alfalfa production had a high frequency (51%) of R2 isolates, and 43% of all isolates were R2 from fields with a history of alfalfa production. In a location that was planted continuously to pea for 30 years, 27% of the isolates were R2. Random amplified polymorphic DNA (RAPD) analysis of three R1 and three R2 isolates with eight primers generated 43 total polymorphic bands; however, none of the bands were uniquely associated with race phenotype. Cluster analysis based on RAPD bands revealed no consistent genotypic distinctions between R1 and R2 isolates of A. euteiches. Evaluation of eight commercial alfalfa cultivars for resistance to two R1 and two R2 isolates demonstrated that most are susceptible to R2 isolates; however, those selected for R2 resistance express resistance to R2 isolates. The results suggest that R2 isolates represent a widespread risk to alfalfa cultivars having resistance only to R1 isolates in fields with varied cropping histories.

scholarly journals Detection of Fusarium oxysporum f. sp. melonis Race 1 in Soil in Colima State, Mexico

Plant Disease ◽  
2004 ◽  
Vol 88 (12) ◽  
pp. 1383-1383 ◽  
Author(s):  
M. de Cara ◽  
E. J. Fernández ◽  
R. Blanco ◽  
J. C. Tello Marquina ◽  
F. J. Estrada ◽  
...  
Keyword(s):  
Fusarium Species ◽  
Selective Medium ◽  
Soil Samples ◽  
Potato Dextrose Agar ◽  
Conidial Suspension ◽  
State University ◽  
Yield Losses ◽  
Pennsylvania State University ◽  
Differential Cultivars ◽  
Race 1

During the winters of 2002 and 2003, a wilt occurred in melons cultivated on 1,500 ha in Colima State, Mexico. Yield losses reached 25% of final production, despite soil disinfestation with 60% methyl bromide and 40% chloropicrin. On the basis of the observation of plants with necrotic xylem, yellowing, and wilting of leaves, this disease was identified provisionally as Fusarium wilt. During February 2003, four soil samples from affected fields were plated onto a Fusarium-selective medium (1), which resulted in the detection of 2,260 ± 357, 179 ± 76, 668 ± 357, and 1,391 ± 256 CFU/g of F. oxysporum (3). Thirty-one randomly chosen isolates were used to inoculate differential cultivars of melon as described by Risser et al. (4). The cultivars were Amarillo Canario (susceptible to all races), Diana (resistant to races 0 and 2), Tango (resistant to races 0 and 1), and Vulcano (resistant to races 0, 1, and 2) (2). Ten plants of each cultivar, grown on sterilized vermiculite, were inoculated at the first true-leaf stage by drenching with 200 ml of a conidial suspension (1 × 105 CFU/ml) of each isolate. Noninoculated plants of each cultivar served as controls. Plants were maintained in a growth chamber with a 16-h photoperiod (18 × 103 lux) and temperatures at 23 to 25°C. Yellowing, wilt, and vascular discoloration symptoms developed on cvs. Amarillo Canario and Diana following inoculation with each of the 31 isolates, while noninoculated plants remained symptomless. F. oxysporum was consistently reisolated on potato dextrose agar from the affected plants. On the basis of the combination of affected cultivars, all isolates were identified as F. oxysporum f. sp. melonis race 1. To our knowledge, this is the first report of F. oxysporum f. sp. melonis race 1 in Colima State, Mexico. References: (1) H. Komada. Rev. Plant Prot. Res. 8:114, 1975. (2) J. Marín Rodríquez. Portagrano 2004. Vadmecum de Variedades Hortícolas. Agrobook, Spain. 2004. (3) P. E. Nelson et al. Fusarium Species: An Illustrated Manual for Identification. Pennsylvania State University Press, University Park, 1983. (4) G. Risser et al. Phytopathology 66:1105, 1976.

scholarly journals Aphanomyces euteiches Race 2 in Central Illinois Alfalfa Fields

Plant Disease ◽  
2002 ◽  
Vol 86 (5) ◽  
pp. 560-560 ◽  
Author(s):  
D. Malvick
Keyword(s):  
Root Rot ◽  
Soil Samples ◽  
Disease Index ◽  
Growth Chamber ◽  
Aphanomyces Euteiches ◽  
First Report ◽  
Dead Plant ◽  
Race 2 ◽  
Central Illinois

Approximately 260,000 ha of alfalfa is grown in Illinois. Two soil samples were collected randomly from slowly drained thin patches in each of four established alfalfa fields near Urbana in 2001. Plants in the thin patches were asymptomatic. Aphanomyces euteiches Drechs. was baited from the soil with cv. Saranac alfalfa seedlings and was isolated from 3- to 4-week-old infected seedlings using a medium containing metalaxyl and benomyl (1,2). It is difficult to isolate A. euteiches from field-grown roots. One to seven isolates were obtained per field, and all were identified as A. euteiches based on morphology (1,2). A. euteiches (races R1 and R2) causes root rot of alfalfa in slowly drained fields in Iowa, Kentucky, and Wisconsin (1,2). The race of 13 isolates was determined in tests repeated once with alfalfa populations Saranac (susceptible to R1 and R2), WAPH-1 (resistant only to R1), and WAPH-5 (resistant to R1 and R2) (1). Twelve 7-day-old seedlings in each of three pots per population were inoculated with 103 zoospores per seedling in a growth chamber (25°C). A disease index (DI) was determined 12 days later by scoring plants on a 1 to 5 scale, where 5 is a dead plant (1). Race was based on DI, R1: DI ≥3 for Saranac and <3 for WAPH-1, and R2: DI > 3 for Saranac and WAPH-1. The DI was 1.0 for noninoculated plants. All isolates were R2; the DI was >3.0 for inoculated Saranac and WAPH-1 and <3.0 for WAPH-5. To our knowledge, this is the first report of A. euteiches races in Illinois, and this pathogen was reported previously only from northwest Illinois. Control of Aphanomyces root rot is based on resistance; however, few alfalfa cultivars are resistant to R2. References: (1) D. Malvick and C. Grau. Plant Dis. 85:740, 2001. (2) G. Munkvold and W. Carlton. Plant Dis. 79:1251,1995.

Suppression of Root Diseases and Weeds in Peas(Pisum sativum)Treated with Dinitrophenol and Dinitroaniline Herbicides

Weed Science ◽  
1978 ◽  
Vol 26 (6) ◽  
pp. 589-593
Author(s):  
R. F. Sacher ◽  
H. J. Hopen ◽  
B. J. Jacobsen
Keyword(s):  
Pisum Sativum ◽  
Weed Control ◽  
Disease Severity ◽  
Root Rot ◽  
Fungal Pathogens ◽  
Severity Index ◽  
Disease Severity Index ◽  
Aphanomyces Euteiches ◽  
Root Diseases ◽  
Dinitroaniline Herbicides

Dinoseb (2-sec-butyl-4,6-dinitrophenol) and certain dinitroaniline herbicides suppressed root rot caused byAphanomyces euteichesDrechs. and other fungal pathogens in peas(Pisum sativumL.). Dinoseb was the most effective. At 6.72 kg/ha pre-plant incorporated (PPI) it reduced the disease severity index (DSI) in glasshouse bioassays to the level considered safe for general use by commercial growers. In fields tests disease severity was reduced and yield of peas was increased with dinoseb. Dinitroaniline herbicides reduced DSI values to the level considered safe for early crop peas. A simplified DSI method of measuring root rot based on mid-season root symptoms was found to be effective. Weed control was superior with the dinitroanilines and propachlor (2-chloro-N-isoproylacetanilide). Dinoseb significantly reduced weed populations relative to untreated plots. Several dinitroaniline treatments caused stunting and stand reduction.

Aphanomyces Euteiches, a Cause of Root Rot of Subterranean Clover in Victoria.

1985 ◽  
Vol 14 (2) ◽  
pp. 34 ◽  
Author(s):  
FC Greenhalgh ◽  
PR Merriman ◽  
PJ Keane
Keyword(s):  
Root Rot ◽  
Subterranean Clover ◽  
Aphanomyces Euteiches
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