Electrophoretic karyotype analysis of Crinipellis perniciosa, the causal agent of witches' broom disease of Theobroma cacao.
ABSTRACT Pulse-field gel electrophoresis (PFGE) was used to determine the genome size and characterize karyotypic differences in isolates of the cacao biotype of Crinipellis perniciosa (C-biotype). The karyotype analysis of four isolates from Brazil revealed that this biotype could be divided into two genotypes: one presenting six chromosomal bands and the other presenting eight. The size of the chromosomes ranged from 2.7 to 5.3 Mb. The different genotypes correlate with telomere-based PCR analysis. The isolates with six chromosomal bands had two that appeared to be doublets, as shown by densitometric analysis, indicating that the haploid chromosome number for this biotype is eight. The size of the haploid genomes was estimated at approximately 30 Mb by both PFGE and Feulgen-image analysis. DNA hybridization revealed that the rDNA sequences are clustered on a single chromosome and these sequences were located on different chromosomes in an isolate dependent manner. This is the first report of genome size and chromosomal polymorphism for the C-biotype of C. perniciosa.
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ABSTRACT: A nonautonomous element of 624 bp, called RetroCl1 (Retroelement Colletotrichum lindemuthianum 1), was identified in the plant pathogenic fungus Colletotrichum lindemuthianum. RetroCl1 contains terminal direct repeats (223 bp) that are surrounded by CTAGT sequences. It has a short internal domain of 178 bp and shows characteristics of terminal-repeat retrotransposon in miniature (TRIM) family. We used RetroCl1 sequence to develop molecular markers for the Colletotrichum genus. IRAP (Inter-Retrotransposon Amplified Polymorphism) and REMAP (Retrotransposon-Microsatellite Amplified Polymorphism) markers were used to analyze the genetic diversity of C. lindemuthianum. Fifty-four isolates belonging to different races were used. A total of 45 loci were amplified. The Nei index showed significant differences among the populations divided according to race, indicating that they are structured according to pathotype. No clear correlation between IRAP and REMAP markers with pathogenic characterization was found. C. lindemuthianum has high genetic diversity, and the analysis of molecular variance showed that 51% of variability is found among the populations of different races. The markers were also tested in different Colletotrichum species. In every case, multiple bands were amplified, indicating that these markers can be successfully used in different species belonging to the Colletotrichum genus.MIRCEN Journal of Applied Microbiology and Biotechnology 03/2012; 28(3):1087-95. · 1.08 Impact Factor
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ABSTRACT: Transposase sequence analysis is an important technique used to detect the presence of transposable elements in a genome. Putative transposase sequence was analyzed in the genome of the phytopathogenic fungus Moniliophthora perniciosa, the causal agent of witches' broom disease of cocoa. Sequence comparisons of the predicted transposase peptide indicate a close relationship with the transposases from the elements of the Tc1-Mariner superfamily. The analysis of the distribution of transposase sequence was done by means of PCR and Southern blot techniques in different isolates of the fungus belonging to C-, L-, and S-biotypes and collected from various geographical areas. The distribution profile of the putative transposase sequence suggests the presence of polymorphic copies among the isolates from C-biotypes. The total DNA hybridization profile of each isolate was used to calculate genetic distance and group by the UPGMA method. C-biotype isolates colleted from of the Bahia showed two hybridization profiles for the transposase sequence. Thus the two different fingerprinting profiles for transposase sequence reported here by Southern analysis could also be correlated to the presence of two different genotypes in Bahia, Brazil.Tropical Plant Pathology 10/2011; 36(5):276-286. · 0.51 Impact Factor
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ABSTRACT: Boto, a Class II transposable element, was characterized in the Moniliophthora perniciosa genome. The Boto transposase is highly similar to plant PIF-like transposases that belong to the newest Class II superfamily known as PIF/Harbinger. Although Boto shares characteristics with PIF-like elements, other characteristics, such as the transposase intron position, the position and direction of the second ORF, and the footprint indicate that Boto belongs to a new family of the PIF/Harbinger superfamily. Southern analyses detected 6 to 12 copies of Boto in C-biotype isolates and a ubiquitous presence among the C- and S-biotypes, as well as a separation in the C-biotype isolates from Bahia State in Brazil in at least two genotypic groups, and a new insertion in the genome of a C-biotype isolate maintained in the laboratory for 6 years. In addition to PCR amplification from a specific insertion site, changes in the Boto hybridization profile after the M. perniciosa sexual cycle and detection of Boto transcripts gave further evidence of Boto activity. As an active family in the genome of M. perniciosa, Boto elements may contribute to genetic variability in this homothallic fungus. This is the first report of a PIF/Harbinger transposon in the genome of a phytopathogenic fungus.Microbiology 10/2012; · 3.06 Impact Factor