Genetic and Pathogenic Analyses of Colletotrichum gloeosporioides Isolates from Strawberry and Noncultivated Hosts

Colletotrichum crown rot of strawberry in Florida is caused primarily by Colletotrichum gloeosporioides. To determine potential inoculum sources, isolates of Colletotrichum spp. from strawberry and various noncultivated plants growing in the areas adjacent to strawberry fields were collected from different sites. Species-specific internal transcribed spacer primers for C. gloeosporioides and C. acutatum were used to identify isolates to species. Random amplified polymorphic DNA (RAPD) markers were used to determine genetic relationships among isolates recovered from noncultivated hosts and diseased strawberry plants. Selected isolates also were tested for pathogenicity on strawberry plants in the greenhouse. In all, 39 C. gloeosporioides and 3 C. acutatum isolates were recovered from diseased strawberry crowns, and 52 C. gloeosporioides and 1 C. acutatum isolate were recovered from noncultivated hosts. In crown inoculation tests, 18 of the 52 C. gloeosporioides isolates recovered from noncultivated hosts were pathogenic to strawberry. Phylogenetic analysis using RAPD marker data divided isolates of C. gloeosporioides from noncultivated hosts into two separate clusters. One cluster contained 50 of the 52 isolates and a second cluster contained 2 isolates that were homothallic in culture. Isolates from strawberry were interspersed within the cluster containing the 50 isolates that were recovered from noncultivated hosts. The results are not inconsistent with the hypothesis that C. gloeosporioides isolates obtained from strawberry and noncultivated hosts adjacent to strawberry fields are from the same population and that noncultivated hosts can serve as potential inoculum sources for Colletotrichum crown rot of strawberry.

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Genetic-Relationships as Assessed by Molecular Markers and Cross-Infection Among Strains of Colletotrichum gloeosporioides

Australian Journal of Botany ◽

10.1071/bt9940009 ◽

1994 ◽

Vol 42 (1) ◽

pp. 9 ◽

Cited By ~ 19

Author(s):

HL Hayden ◽

KG Pegg ◽

EAB Aitken ◽

JAG Irwin

Keyword(s):

Rapd Markers ◽

Colletotrichum Gloeosporioides ◽

Rapd Analysis ◽

Genetic Relationships ◽

Random Amplified Polymorphic Dna ◽

Colletotrichum Acutatum ◽

Fruit Species ◽

Pathogenicity Tests ◽

Colletotrichum Musae ◽

Highly Virulent

Morphological characterisation allows isolates of Colletotrichum gloeosporioides, Colletotrichum musae and Colletotrichum acutatum to be identified only to species level. Pathogenicity tests and random amplified polymorphic DNA (RAPD) markers distinguished a mango biotype of C. gloeosporioides from eight other isolates of C gloeosporioides obtained from five different fruit species. Using these procedures, it was also possible to distinguish C. acutatum and C. musae both from each other, and from the C. gloeosporioides isolates. In cross-infectivity studies, isolates of C. gloeosporioides displayed a wide host range with the exception of isolates from mango, which were highly virulent on mango only. Teleomorphic isolates of C. gloeosporioides were clustered together by RAPD analysis. This work has demonstrated the existence of a biotype of C. gloeosporioides which shows specialisation to mango.

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Identification and characterization of RAPD markers inferring genetic relationships among Pine species

Genome ◽

10.1139/g01-121 ◽

2002 ◽

Vol 45 (1) ◽

pp. 51-58 ◽

Cited By ~ 30

Author(s):

K K Nkongolo ◽

P Michael ◽

W S Gratton

Keyword(s):

Pinus Banksiana ◽

Rapd Markers ◽

Genetic Relationships ◽

Random Amplified Polymorphic Dna ◽

Morphological Characteristics ◽

Rflp Analysis ◽

Pinus Monticola ◽

Species Specific ◽

Specific Sequences ◽

Pine Species

Total genomic DNAs were extracted from several populations of pine species and amplified using oligonucleotides of random sequences. Polymorphism in random amplified polymorphic DNA (RAPD) markers was high and sufficient in distinguishing each of the species. Genetic relationships among eight pine species (Pinus sylvestris, Pinus strobus, Pinus rigida, Pinus resinosa, Pinus nigra, Pinus contorta, Pinus monticola, and Pinus banksiana) from different provenances were analyzed. The degree of band sharing was used to evaluate genetic distance between species and to construct a phylogenetic tree. In general, the dendrogram corroborated the description of relationships based on morphological characteristics and crossability, but also provided new insights into pine taxonomy. RAPD markers specific to some pine species were cloned and sequenced. PCR amplifications using pairs of designed specific primers revealed that all the cloned sequences were likely genus specific because they were not found in spruce or larch. True species-specific sequences were identified using designed primers flanking cloned RAPD fragments. The analysis of RAPD fragment sequences confirmed the genetic relationships among species. A 2281-bp RAPD band called PI-Mt-Stb-23 from P. strobus was used as a probe in restriction fragment length polymorphism (RFLP) analysis and produced distinct banding patterns for each species examined, consistent with the highly polymorphic character of DNA-fingerprinting probes.Key words: Pine, RAPD, RFLP, cloning, species-specific sequences.

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GENETIC FINGERPRINTING OF SNAP BEAN CULTIVARS WITH RAPD MARKERS

HortScience ◽

10.21273/hortsci.27.6.573g ◽

1992 ◽

Vol 27 (6) ◽

pp. 573g-574

Author(s):

Paul Skroch ◽

Jim Nienhuis

Keyword(s):

Rapd Markers ◽

Genetic Relationships ◽

Rapd Marker ◽

Pedigree Information ◽

Banding Patterns ◽

Snap Bean ◽

Marker Data ◽

Varietal Identification ◽

The One ◽

And Control

The genetic variation in a population of one hundred Snap Bean varieties, including processing and garden types, was studied using RAPD markers. All one hundred genotypes were distinguished by unique combinations of banding patterns. These unique “fingerprints” were tested for repeatability. Certain bands were very reliable and can be used for varietal identification. The RAPD marker data was also used to estimate genetic relationships among a subset of the one hundred lines. The results of the analysis agreed with known pedigree information. These markers will allow more precise monitering and control of germplasm by those who are involved with the breeding and production of superior seed.

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Selection for Pathogenicity to Strawberry in Populations of Colletotrichum gloeosporioides from Native Plants

Phytopathology ◽

10.1094/phyto-97-9-1130 ◽

2007 ◽

Vol 97 (9) ◽

pp. 1130-1140 ◽

Cited By ~ 22

Author(s):

S. J. MacKenzie ◽

T. E. Seijo ◽

D. E. Legard ◽

L. W. Timmer ◽

N. A. Peres

Keyword(s):

Restriction Enzyme ◽

Colletotrichum Gloeosporioides ◽

Native Plants ◽

Random Amplified Polymorphic Dna ◽

Rapd Marker ◽

Crown Rot ◽

Enzyme Analysis ◽

Restriction Enzyme Analysis ◽

Selection For ◽

Native Host

Colletotrichum gloeosporioides causes a serious crown rot of strawberry and some isolates from native plants are pathogenic to strawberry. C. gloeosporioides from lesions on wild grape and oak were sampled at two sites adjacent to commercial strawberry fields in Florida and two distant sites. Random amplified polymorphic DNA (RAPD) marker data and restriction enzyme digests of amplified rDNA were used to determine whether isolates were from the same C. gloeosporioides subgroup that infects strawberry. There were 17 to 24 native host isolates from each site that clustered with a group of strawberry crown isolates based on RAPD markers. Among strawberry isolates, there were two rDNA genotypes identified by restriction enzyme analysis. Both genotypes were present among native host isolates sampled from all four sites. There was some evidence that the different rDNA genotypes differentiated two closely related subpopulations, although the proportion of pathogenic isolates from native hosts among the two different genotypes was not different. The incidence of isolates pathogenic to strawberry was greater at sites close to strawberry fields relative to sites distant from strawberry fields for isolates with a BstUI(−)/MspI(+) rDNA genotype (44 versus 13%), a BstUI(+)/MspI(−) genotype (57 versus 16%), or when both genotypes were analyzed together (46 versus 15%). Based on these results, it appears that the C. gloeosporioides subgroup that causes crown rot on strawberry is widely distributed in Florida and that selection for pathogenicity on strawberry occurs in the area where this host is grown in abundance.

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Oversummer Survival of Inoculum for Colletotrichum Crown Rot in Buried Strawberry Crown Tissue

Plant Disease ◽

10.1094/pdis.2001.85.7.750 ◽

2001 ◽

Vol 85 (7) ◽

pp. 750-754 ◽

Cited By ~ 29

Author(s):

A. R. Ureña-Padilla ◽

D. J. Mitchell ◽

D. E. Legard

Keyword(s):

Colletotrichum Gloeosporioides ◽

Crown Rot ◽

Colletotrichum Acutatum ◽

Initial Increase ◽

Spore Morphology ◽

Plant Debris ◽

Crown Tissue ◽

Colletotrichum Spp ◽

Species Specific ◽

Dna Primers

The oversummer survival of Colletotrichum gloeosporioides in strawberry crown tissue under field conditions was investigated in 1998 and 1999. Strawberry crowns infected naturally with C. gloeosporioides were placed inside cloth bags containing field soil, buried in the field at 5 or 13 cm, then recovered over 6 months of each year. The recovered crowns were plated onto a Colletotrichum spp. semiselective medium and speciated by colony, spore morphology, and molecular markers with species-specific DNA primers. Pathogenicity of selected isolates was confirmed by greenhouse bioassays on strawberry. Of the 428 isolates of Colletotrichum spp. recovered from buried crowns, 96% were C. gloeosporioides and 4% Colletotrichum acutatum. Following an initial increase in the detection of the fungus, survival of C. gloeosporioides was stable for 2 to 3 weeks, then declined. No Colletotrichum spp. were detected after burial for 56 days in 1998 and 98 days in 1999. Because the time between crop seasons is typically more than 170 days, these data support the hypothesis that inoculum of C. gloeosporioides does not survive in buried plant debris between seasons in Florida and, therefore, oversummering crop debris does not contribute inoculum for epidemics of Colletotrichum crown rot in Florida.

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Segregation for Resistance to Eastern Filbert Blight in Progeny of `Zimmerman' Hazelnut

Journal of the American Society for Horticultural Science ◽

10.21273/jashs.131.6.731 ◽

2006 ◽

Vol 131 (6) ◽

pp. 731-737 ◽

Cited By ~ 8

Author(s):

China F. Lunde ◽

Shawn A. Mehlenbacher ◽

David C. Smith

Keyword(s):

Rapd Markers ◽

Chromosome Region ◽

Random Amplified Polymorphic Dna ◽

Rapd Marker ◽

Segregation Ratio ◽

Enzyme Linked Immunosorbent Assay ◽

Resistance Locus ◽

Resistance Allele ◽

Reciprocal Translocations ◽

Eastern Filbert Blight

Eastern filbert blight (EFB), caused by the fungus Anisogramma anomala (Peck) E. Müller, is an important disease of european hazelnut (Corylus avellana L.) in the Pacific northwestern United States. In 1989, a chance seedling free of EFB was discovered adjacent to a severely diseased orchard near Troutdale, Ore. This selection, subsequently named `Zimmerman', was crossed with three susceptible selections. Based on morphological characters and incompatibility alleles, we speculated that `Zimmerman' (S1 S3) was a hybrid between `Barcelona' (S1 S2) and `Gasaway' (S3 S26). The three seedling populations were inoculated with spores of the pathogen in a greenhouse test and assayed by indirect enzyme-linked immunosorbent assay (ELISA) and by observation of canker incidence. The observed segregation fit a 3 resistant : 1 susceptible ratio in all three progenies, in contrast to the 1 : 1 ratio found when the resistant pollinizer `Gasaway' was crossed to susceptible genotypes. Random amplified polymorphic DNA (RAPD) marker UBC 152800 linked to the resistance gene in `Gasaway' co-segregated with the resistant phenotype in all three populations with 2%, 4%, and 6% recombination, respectively. Seed germination and transplanting records did not provide evidence of selection in favor of resistant seedlings. Pollen germination was 71% in `Gasaway', 29% in `Zimmerman', and 18% in `Barcelona', indicating possible selection at the gametophytic level. Subsequently 16 resistant seedlings of `Zimmerman' were crossed with the highly susceptible selection OSU 313.078. Segregation fit a 3 : 1 ratio in 14 of the 16 progenies, and showed a surplus of resistant seedlings in the other two. None showed a 1 : 1 segregation. Resistance co-segregated with two RAPD markers that flank the `Gasaway' resistance allele. To test allelism of resistance from `Gasaway' and `Zimmerman', VR 6-28 with resistance from `Gasaway' was crossed with `Zimmerman'. Eight resistant selections from this progeny were crossed with OSU 313.078. Five of the eight progenies segregated 3 : 1, two progenies segregated 1 : 1, and OSU 313.078 × OSU 720.056 gave only resistant offspring. The ratios indicate that OSU 720.056 is homozygous resistant and that `Zimmerman' and `Gasaway' share a common resistance allele. Reciprocal translocations have been reported in hazelnut cultivars, including `Barcelona', the leading cultivar in Oregon. `Zimmerman' appears to be a hybrid of `Barcelona' and `Gasaway', but because of cytogenetic abnormalities, `Zimmerman' may have inherited two copies of the chromosome region that contain the resistance locus and flanking RAPD markers. If the region containing the resistance were attached to two independent centromeres, a 3 : 1 segregation ratio for disease response and flanking markers would be expected, and we propose this as the most likely explanation. Resistance from `Gasaway' and `Zimmerman' has been called “immunity” or “complete resistance.” However, we noted a few seedlings with small cankers, nearly all of which lacked sporulating stromata. Flanking RAPD markers indicate that the resistance allele is present in these seedlings. Although not “immune” or “completely resistant,” `Gasaway' and `Zimmerman' transmit a very high level of resistance.

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Segregation of random amplified polymorphic DNA markers and strategies for molecular mapping in tetraploid alfalfa

Genome ◽

10.1139/g93-112 ◽

1993 ◽

Vol 36 (5) ◽

pp. 844-851 ◽

Cited By ~ 23

Author(s):

K. F. Yu ◽

K. P. Pauls

Keyword(s):

Chromosome Segregation ◽

Dna Markers ◽

Rapd Markers ◽

Molecular Mapping ◽

Random Amplified Polymorphic Dna ◽

Rapd Marker ◽

Female Parent ◽

Linkage Groups ◽

Double Reduction ◽

Chromatid Segregation

An F1 population was used to analyze the inheritance of random amplified polymorphic DNA (RAPD) markers in tetraploid alfalfa. Of the 32 RAPD markers that were used for a segregation analysis in this study, 27 gave ratios that are consistent with random chromosome and random chromatid segregation at meiosis. However, among all of the RAPD markers (121) that were screened in this study, only one example of a double reduction, that is typical of chromatid segregation, was observed. These results indicate that random chromosome segregation is likely the predominant but not the exclusive mode of inheritance for tetraploid alfalfa. χ2 analyses of cosegregation for RAPD marker pairs derived from the female parent revealed nine linkages that fell into four linkage groups. The recombination fractions among linked marker pairs ranged from 1 to 37%. These are the first molecular linkage groups reported in tetraploid alfalfa. In addition, various strategies for molecular mapping in the tetraploid alfalfa genome are proposed that should be of interest to plant breeders who are planning to use molecular markers for alfalfa or other tetraploid species.Key words: RAPD markers, tetraploid alfalfa, segregation, linkage groups.

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Identification and Characterization of Benomyl-Resistant and -Sensitive Populations of Colletotrichum gloeosporioides from Statice (Limonium spp.)

Phytopathology ◽

10.1094/phyto-96-0542 ◽

2006 ◽

Vol 96 (5) ◽

pp. 542-548 ◽

Cited By ~ 40

Author(s):

Marcel Maymon ◽

Aida Zveibil ◽

Shimon Pivonia ◽

Dror Minz ◽

Stanley Freeman

Keyword(s):

Colletotrichum Gloeosporioides ◽

Sequence Analyses ◽

Specific Primers ◽

Tubulin Genes ◽

Colletotrichum Spp ◽

Polymerase Chain ◽

Species Specific ◽

Identification And Characterization ◽

Propagation Material

Sixty-four isolates of Colletotrichum gloeosporioides were isolated from infected Limonium spp. cultivated in 12 different locations in Israel. All isolates were identified as belonging to the C. gloeosporioides complex by species-specific primers. Of these isolates, 46 were resistant to benomyl at 10 μg/ml and 18 were sensitive to this concentration of fungicide. Based on arbitrarily primed polymerase chain reaction of all isolates and internal transcribed spacer-1 sequence analyses of 12 selected isolates, the benomyl-resistant and -sensitive populations belong to two distinct genotypes. Sequence analyses of the β-tubulin genes, TUB1 and TUB2, of five sensitive and five resistant representative isolates of C. gloeosporioides from Limonium spp. revealed that the benomyl-resistant isolates had an alanine substitute instead of a glutamic acid at position 198 in TUB2. All data suggest that the resistant and sensitive genotypes are two independent and separate populations. Because all Limonium plant propagation material is imported from various geographic regions worldwide, and benomyl is not applied to this crop or for the control of Colletotrichum spp. in Israel, it is presumed that plants are bearing quiescent infections from the points of origin prior to arrival.

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Phenotype Instability in Botrytis cinerea in the Absence of Benzimidazole and Dicarboximide Fungicides

Phytopathology ◽

10.1094/phyto.2001.91.3.307 ◽

2001 ◽

Vol 91 (3) ◽

pp. 307-315 ◽

Cited By ~ 22

Author(s):

L. F. Yourman ◽

S. N. Jeffers ◽

R. A. Dean

Keyword(s):

Botrytis Cinerea ◽

High Frequency ◽

Rapd Markers ◽

Random Amplified Polymorphic Dna ◽

Rapd Marker ◽

Potato Dextrose Agar ◽

Multiple Generations ◽

Rapd Fingerprints ◽

Growth Room ◽

Conidium Germination

Stability of phenotypes of isolates of Botrytis cinerea that were sensitive or resistant to benzimidazole and dicarboximide fungicides was examined in the absence of fungicides in laboratory and growth room experiments. Twelve greenhouse isolates of B. cinerea were subcultured on potato dextrose agar (PDA) for 20 generations and on geranium seedlings for 15 generations. Three isolates of each of the following four phenotypes were used: sensitive to the fungicides thiophanate-methy1 (a benzimidazole) and vinclozolin (a dicarboximide) (STSV), resistant to both fungicides (RTRV), resistant to thiophanate-methy1 and sensitive to vinclozolin (RTSV), and sensitive to thiophanate-methy1 and resistant to vinclozolin (STRV). In three trials on PDA, 36 populations were subcultured; 8 populations changed phenotypes by the end of 20 generations, as determined by conidium germination on fungicide-amended medium. Five of the eight initially were STRV; the resulting phenotypes were STSV, RTSV, and RTRV. Populations from eight other isolates exhibited temporary changes in phenotype during intermediate generations on PDA but reverted to initial phenotypes by the twentieth generation; five of these populations changed to phenotype RTRV. In two geranium seedling trials, each of the 12 greenhouse isolates was inoculated onto a set of three seedlings for each generation, and diseased tissue that developed was used to initiate the next generation. Therefore, a total of 72 populations of B. cinerea were subcultured in the two trials; 5 of these populations changed phenotype at the end of 15 generations. Three of the five initially were STRV; these changed to phenotypes STSV or RTRV. In each of the two trials on geranium seedlings, a population subcultured from one STSV isolate changed phenotype one to phenotype RTRV and one to phenotype RTSV. In all trials, no population resistant to thiophanate-methy1 changed to a thiophanate-methy1-sensitive phenotype, and no population changed to phenotype STRV. Random amplified polymorphic DNA (RAPD) fingerprints were generated with the 12 initial isolates and 49 isolates subcultured on PDA or geranium seedlings. Cluster analyses of RAPD markers showed that subcultured isolates exhibiting the same phenotype clustered together and that subcultured isolates derived from a common greenhouse isolate but with different phenotypes were in different clusters. Some populations that did not change phenotype exhibited considerable differences in RAPD marker patterns. The results of this study indicate that, in the absence of fungicides, sensitive populations of B. cinerea can develop resistance to thiophanate-methy1 and vinclozolin, and this resistance can be maintained in populations through multiple generations. Populations resistant only to vinclozolin (STRV) exhibited a high frequency of phenotype change, and populations resistant to both fungicides (RTRV) were stable.

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Isolation of Genotype- and Chromosome-specific DNA Markers in a Biotype of Colletotrichum gloeosporioides Using Random Amplified Polymorphic DNA Analysis

Australian Journal of Botany ◽

10.1071/bt96131 ◽

1998 ◽

Vol 46 (1) ◽

pp. 143

Author(s):

Agnieszka M. Poplawski ◽

John A. G. Irwin ◽

John M. Manners

Keyword(s):

Dna Sequences ◽

Fungal Pathogen ◽

Rapd Markers ◽

Dna Analysis ◽

Colletotrichum Gloeosporioides ◽

Rapd Analysis ◽

Random Amplified Polymorphic Dna ◽

Dna Probes ◽

Race 2 ◽

Biotype B

Genetic markers that distinguish fungal genotypes are important tools for genetic analysis of heterokaryosis and parasexual recombination in fungi. Random amplified polymorphic DNA (RAPD) markers that distinguish two races of biotype B of Colletotrichum gloeosporioides infecting the legume Stylosanthes guianensis were sought. Eighty-five arbitrary oligonucleotide primers were used to generate 895 RAPD bands but only two bands were found to be specifically amplified from DNA of the race 3 isolate. These two RAPD bands were used as DNA probes and hybridised only to DNA of the race 3 isolate. Both RAPD bands hybridised to a dispensable 1.2 Mb chromosome of the race 3 isolate. No other genotype-specific chromosomes or DNA sequences were identified in either the race 2 or race 3 isolates. The RAPD markers hybridised to a 2 Mb chromosome in all races of the genetically distinct biotype A pathogen which infects other species of Stylosanthes as well as S. guianensis. The experiments indicate that RAPD analysis is a potentially useful tool for obtaining genotype- and chromosome-specific DNA probes in closely related isolates of one biotype of this fungal pathogen.

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