Abstract: We investigated the genetic diversity, extent of recombination, natural selection, and population divergence of Ralstonia solanacearum samples obtained from sources worldwide. This plant pathogen causes bacterial wilt in many crops and constitutes a serious threat to agricultural production due to its very wide host range and aggressiveness. Five housekeeping genes, dispersed around the chromosome, and three virulence-related genes, located on the megaplasmid, were sequenced from 58 strains belonging to the four major phylogenetic clusters (phylotypes). Whereas genetic variation is high and consistent for all housekeeping loci studied, virulence-related gene sequences are more diverse. Phylogenetic and statistical analyses suggest that this organism is a highly diverse bacterial species containing four major, deeply separated evolutionary lineages (phylotypes I to IV) and a weaker subdivision of phylotype II into two subgroups. Analysis of molecular variations showed that the geographic isolation and spatial distance have been the significant determinants of genetic variation between phylotypes. R. solanacearum displays high clonality for housekeeping genes in all phylotypes (except phylotype III) and significant levels of recombination for the virulence-related egl and hrpB genes, which are limited mainly to phylotype strains III and IV. Finally, genes essential for species survival are under purifying selection, and those directly involved in pathogenesis might be under diversifying selection.
Abstract: The genetic diversity among a worldwide collection of 120 strains of Ralstonia solanacearum was assessed by restriction fragment length polymorphism (RFLP) analysis of amplified fragments from the hrp gene region. Five amplified fragments appeared to be specific to R. solanacearum. Fifteen different profiles were identified among the 120 bacterial strains, and a hierarchical cluster analysis distributed them into eight clusters. Each cluster included strains belonging to a single biovar, except for strains of biovars 3 and 4, which could not be separated. However, the biovar 1 strains showed rather extensive diversity since they were distributed into five clusters whereas the biovar 2 and the biovar 3 and 4 strains were gathered into one and two clusters, respectively. PCR-RFLP analysis of the hrp gene region confirmed the results of previous studies which split the species into an "Americanum" division including biovar 1 and 2 strains and an "Asiaticum" division including biovar 3 and 4 strains. However, the present study showed that most of the biovar 1 strains, originating from African countries (Reunion Island, Madagascar, Zimbabwe, and Angola) and being included in a separate cluster, belong to the "Asiaticum" rather than to the "Americanum" division. These African strains could thus have evolved separately from other biovar 1 strains originating from the Americas.
Abstract: A survey of bacterial wilt in China collected 286 strains of Ralstonia solanacearum from 17 plant species in 13 Chinese provinces to investigate genetic diversity using the biovar (bv.) and phylotype classification schemes. A phylotype-specific multiplex-PCR showed that 198 isolates belonged to phylotype I (bv. 3, 4 and 5) and 68 to phylotype II (bv. 2 and bv. 1). A phylogenetic analysis examined the partial sequence of the egl and hrpB gene of all strains and the genetic diversity of 95 representatives was reported, demonstrating that Chinese strains are partitioned into phylotype I (Asia) and II (Americas). Phylotype I strains (historically typed bv. 3, 4 and 5), had considerable phylogenetic diversity, including 10 different sequevars: seven previously described sequevars 12 to 18 and three new sequevars: 34, 44 and 48. Chinese strains Z1, Z2, Z3, Z7, Pe74 and Tm82 were not genetically distinguishable from the edible ginger reference strain ACH92 (r4-bv. 4) for sequevar 16. This is believed to be the first report of this ginger group in China. All Chinese bv. 2 strains falling into the genetically and phenotypically diverse phylotype II were placed into phylotype IIB sequevar 1 (historically the Andean race3-bv. 2 potato brown rot agent). In both the egl and hrpB sequence-based trees, strains isolated from mulberry were present in two distinct branches found in sequevars 12 and 48 (reference strains R292 and M2, respectively).
Abstract: The genetic diversity among strains in a worldwide collection of Ralstonia solanacearum, causal agent of bacterial wilt, was assessed by using three different molecular methods. PCR-RFLP analysis of the hrp gene region was extended from previous studies to include additional strains and showed that five amplicons were produced not only with all R. solanacearum strains but also with strains of the closely related bacteria Pseudomonas syzygii and the blood disease bacterium (BDB). However, the three bacterial taxa could be discriminated by specific restriction profiles. The PCR-RFLP clustering, which agreed with the biovar classification and the geographical origin of strains, was confirmed by AFLP. Moreover, AFLP permitted very fine discrimination between different isolates and was able to differentiate strains that were not distinguishable by PCR-RFLP. AFLP and PCR-RFLP analyses confirmed the results of previous investigations which split the species into two divisions, but revealed a further subdivision. This observation was further supported by 16S rRNA sequence data, which grouped biovar 1 strains originating from the southern part of Africa.
Abstract: Two extant nomenclature systems were reconciled to relate six mitochondrial DNA (mtDNA) haplotypes of Phytophthora infestans, the oomycete pathogen causing late blight disease on potato and tomato. Carter's haplotypes I-a and I-b were included in Goodwin's haplotype A, while Carter's haplotypes II-a and II-b were included in Goodwin's haplotype B. In addition, haplotypes E and F were included in Carter's haplotype I-b. The mutational differences separating the various haplotypes were determined, and we propose that either haplotype I-b(A) or haplotype I-a(A) is the putative ancestral mtDNA of P. infestans, because either can center all the other haplotypes in a logical stepwise network of mutational changes. The occurrence of the six haplotypes in 548 isolates worldwide was determined. Haplotypes I-a and II-a were associated with diverse genotypes worldwide. As previously suggested, haplotype I-b was found only in the US-1 clonal lineage and its variants (n = 99 isolates from 16 countries on 5 continents), and haplotype II-b was limited to the US-6 clonal lineage and its derivatives (n = 36). In a confirmation of a previous suggestion, the randomly mating population in the Toluca Valley of central Mexico (n = 78) was monomorphic for mtDNA haplotype I-a(A). We hypothesize that selection there may be driving the dominance of that single mtDNA haplotype.
Abstract: Morphology has often been used as an indicator of variability within species. The present study investigated morphological and physiological characteristics of isolates of Phytophthora cinnamomi collected from diseased vegetation communities at Anglesea, Victoria, and isolates collected from other regions in the State. Characteristics studied included growth rate on potato-dextrose agar (PDA), corn-meal agar and V8-juice agar at 24°C, growth rate on V8 agar at 15°C, colony morphology on PDA, sporangial and gametangial morphology, sporangial production and mating type. Phenotypic variation was demonstrated in radial growth rate, colony morphology and sporangial dimensions. Sporangial and oogonial dimensions and sporangial production were not significantly different between isolates from different geographical regions. All isolates were found to be of the A2 mating type suggesting variation was derived asexually. Paragynal associations, in an organism characteristically defined as amphigynal, were observed following crossing with A1 isolates. This is the first such study undertaken in southern Victoria. The findings highlight the importance of appropriate management of an area of such high conservation value as the Anglesea Heath to contain the current infection and to prevent introduction of new isolates into the area.
Abstract: AFLP fingerprinting analysis of Fusarium circinatum, Nirenberg and O’Donnell and relative species was carried out. Ten primer-pairs that could generate abundant polymorphism fragments were screened. A total of 298 nucleotide acid fragments were amplified with the primers from the template of the 17 strains of Fusarium spp., among which 283 fragments were polymorphic. Percentage of polymorphic loci produced by each pair of AFLP primer-pair was 94.97% in average and varied from 89.29% to 100%. All these data indicated that considerable genetic variation existed among F. circinatum and relative species at DNA level. Molecular genetic distances among Fusarium spp. were calculated, and the relationship among them was described quantitatively. Compared with biological species, the result of cluster analysis was basically similar to the phenotypic classification of species. Genetic diversity of E-AT/M-CAA AFLP fingerprinting of Fusarium spp. was analyzed, and specific and difference bands for each species and all Fusarium section Liseola tested were identified based on the E-AT/M-CAA AFLP fingerprinting.
Abstract: The Fusarium wilt caused by Fusarium oxyspoum f. sp. cubense (Foc) is a major biotic constraint for banana production. The characteristics of F. oxyspoum f. sp. cubense isolates were investigated using electrophoretic studies of isozyme and whole-cell protein. The morphological characteristics of the isolates were very similar to each other. All the Foc isolates were pathogenic to banana cultivar ‘Nanjangud Rasabale’ but they did not induce any disease symptoms on cultivar ‘Cavendish’. F. oxyspoum (Isolate 6) did not induce wilt symptoms on either ‘Nanjangud’ or ‘Cavendish’ cultivar. Isozyme banding patterns showed 46 scoreable markers and cluster analysis with UPGMA using genetic distance showed that the isolates belonged to three main groups. Group 1 contained isolates 1, 2, 4, 5, 7 and isolate 3 and 6 were placed in group 2 and 3. Results indicated that the estimated intra-specific variation may be more pronounced with isozyme analysis than with protein markers. The level of isozyme variability detected within F. oxysporum f.sp. cubense suggested that it is reliable, efficient and effective in determining genetic relationships among Foc isolates.
Abstract: Isozyme variation was studied in 94 isolates of Phytophthora colocasiae originating from Indonesia, Papua New Guinea, the Philippines, Thailand and Vietnam. Eight polymorphic enzyme systems (HK, PGM, PGI, Gluco, MDH, ICD, 6PDH, ME) revealed 52 isozyme patterns (zymotypes), each uniquely characterized by the presence or absence of 60 electromorphs. A core sample of 20 isolates was subsequently analysed with RAPD markers. Seven primers were used successfully and all profiles were reproducible. Clear bands were revealed and, in some cases, allowed differentiation between isolates exhibiting identical zymotypes. Results indicate that throughout this vast geographic region, taro leaf blight is caused by numerous and distinct strains that are genetically variable. Variation occurs within and between countries. The geographical distribution of zymotypes shows that none is common to two different countries. Although the differences in pathogenicity are not yet established, different P. colocasiae genotypes are likely to recombine and evolve rapidly as this species is heterothallic. From these results, a long-term breeding strategy is recommended for taro (Colocasia esculenta) based on recurrent selection using a wide genetic base composed of carefully selected parents from diverse geographic origins to maximize multigenic resistance in progenies.
Abstract:Phytophthora infestans was isolated from potato leaves and tubers collected from different parts of Finland in 1990–96 and Norway in 1993–96. Isolates were assessed for mating type, resistance to metalaxyl and virulence phenotype. In Finland 15% of 200 isolates tested and in Norway 25% of 642 tested were A2 mating type. Differences in the A1/A2 ratio were evident among regions and A2 was not found in northern areas. In Finland the frequency of metalaxyl-resistant isolates, among 1834 tested, decreased from 59% in 1990–95 to 2% in 1996. In Norway 59% of 491 isolates tested were resistant to metalaxyl in 1996. Among 269 Finnish and 105 Norwegian isolates, all known virulence genes were found in both countries and race 1.3.4.7.10.11 was the most common (resistance gene R9 was not included in the differential set). Oospores were observed in potato leaves from three locations in the southern part of Norway. The implications of the 'new' populations in the Nordic countries are discussed.
Abstract