Host plants exercise a selective pressure on their endophytic bacteria. Host plant changes and repeated passages on the new species may modify gene frequencies and generate aplotypes distinct from the population on the original host plant. The virulent strain Erwinia amylovora (Ea) OMP-BO 1077/7 was inoculated into shoots of two year-old pear, apple and hawthorn plants. After 21 days at 25°C, four isolates were randomly purified and identified from cankers on each host species. One reisolate, randomly chosen for each species, was reinoculated into shoots of the same host. Six passages were made on each host species for a total of 72 pure cultures from which the genomic DNA was analyzed by AFPL. The genetic distance between the Ea subpopulations was determined by the Dice similarity coefficient which pointed out that the isolates from host plants other than pear were undistinguishable between themselves. However, the AFLP technique compares profiles obtained by the PCR amplification of DNA fragments selected on the entire genome. In this research work we hypothesized that the adaptation to other hosts may modify the genetic and/or genotypic frequencies in the Ea subpopulations, with possible effects even on the virulence. The dspEF locus next to the hrp gene cluster of Ea is essential for pathogenicity, but not for elicitation of the hypersensitive reaction. Moreover DspE is a homolog of AvrE of Pseudomonas syringae. In this work we have started a comparative analysis of the sequence of the dspE gene which can be one of the likely target of selective pressure in different hosts. For this purpose oligonucleotides based on the dspE gene of Ea321 strain were used as primers in a PCR with the total DNA of Ea 1077/7 as template. An amplicone was similar in size to dspE gene ( 5.6 kb) and its restriction fragment analysis was carried out using the restriction endonucleases: EcoRI, AluI, HaeII, MspI, HhaI, MseI, RsaI. As a result of the comparative analysis of the amplicone, obtained from several Ea strains isolated from different host plants, the variability related to the adaptation to the host plant was studied.
Genetic diversity of Erwinia amylovora subpopulations using AFLP and molecular analysis of the dspE gene: role of different host plants / Minardi P.; Mucini S.; Mazzucchi U.. - STAMPA. - (2007), pp. 229-229. (Intervento presentato al convegno 13th International Congress on Molecular Plant-Microbe Interactions tenutosi a Sorrento (Italy) nel 21-27 luglio 2007).
Genetic diversity of Erwinia amylovora subpopulations using AFLP and molecular analysis of the dspE gene: role of different host plants
MINARDI, PAOLA;MUCINI, SARA;MAZZUCCHI, UMBERTO
2007
Abstract
Host plants exercise a selective pressure on their endophytic bacteria. Host plant changes and repeated passages on the new species may modify gene frequencies and generate aplotypes distinct from the population on the original host plant. The virulent strain Erwinia amylovora (Ea) OMP-BO 1077/7 was inoculated into shoots of two year-old pear, apple and hawthorn plants. After 21 days at 25°C, four isolates were randomly purified and identified from cankers on each host species. One reisolate, randomly chosen for each species, was reinoculated into shoots of the same host. Six passages were made on each host species for a total of 72 pure cultures from which the genomic DNA was analyzed by AFPL. The genetic distance between the Ea subpopulations was determined by the Dice similarity coefficient which pointed out that the isolates from host plants other than pear were undistinguishable between themselves. However, the AFLP technique compares profiles obtained by the PCR amplification of DNA fragments selected on the entire genome. In this research work we hypothesized that the adaptation to other hosts may modify the genetic and/or genotypic frequencies in the Ea subpopulations, with possible effects even on the virulence. The dspEF locus next to the hrp gene cluster of Ea is essential for pathogenicity, but not for elicitation of the hypersensitive reaction. Moreover DspE is a homolog of AvrE of Pseudomonas syringae. In this work we have started a comparative analysis of the sequence of the dspE gene which can be one of the likely target of selective pressure in different hosts. For this purpose oligonucleotides based on the dspE gene of Ea321 strain were used as primers in a PCR with the total DNA of Ea 1077/7 as template. An amplicone was similar in size to dspE gene ( 5.6 kb) and its restriction fragment analysis was carried out using the restriction endonucleases: EcoRI, AluI, HaeII, MspI, HhaI, MseI, RsaI. As a result of the comparative analysis of the amplicone, obtained from several Ea strains isolated from different host plants, the variability related to the adaptation to the host plant was studied.I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
