Characterization of Pectobacterium brasiliense strains from potato and vegetables in Israel

Pectobacterium brasiliense (Pbr) infects a wide range of crops worldwide, causing potato blackleg and soft rot and vegetable soft rots. This study aimed to characterize the genetic diversity and virulence variability among 68 Pbr strains isolated from either symptomless potato progeny tubers, diseased potato plants, ware potatoes wash water, or vegetables grown in Israel, as well as strains isolated from symptomless seed tubers grown in Europe, or diseased potato plants grown in France. The collection was typed using PCR and TaqMan real-time PCR analyses, dnaX sequence analysis, pulsed-field gel electrophoresis (PFGE), and pectolytic activity. dnaX phylogeny grouped almost all strains in a common genetic clade related to Pbr, which was distinct from the other Pectobacterium species. PFGE analysis identified two main clusters, including one major group of 47 strains with 95%–100% similarity. Maceration assays on two potato cultivars showed significant differences between strains but with no correlations with the source of the strains nor the status of the host (with/without symptoms). Molecular (dnaX sequences and PFGE profiles) and phenotypic analyses (tuber maceration tests) showed that the tested Pbr strains are not a homogeneous group. Analysis of the tested Pbr strains isolated from potato and vegetables grown in fields with a history of potato cultivation suggests that seed tubers imported from Europe may be the main source for Pbr in Israel. To the best of our knowledge, this is the first study that describes biodiversity and population structure of P. brasiliense isolated from potato and vegetables under hot climate conditions.     

Detection of Erwinia herbicola pv. gypsophilae in gypsophila plants by PCR

Three PCR primer pairs, based on the cytokinins (etz) or IAA biosynthetic genes, were used for detecting Erwinia herbicola pv. gypsophilae in Gypsophila paniculata plants. The primers were specific to all gall-forming E. herbicola strains and distinguished them from saprophytic strains associated with gypsophila plants or from other gall-forming bacteria. In pure culture of the pathogen, less than one bacterial cell was detected with nested PCR using the etz primers - an increase of 100-fold in sensitivity as compared with single-round PCR. In the presence of plant extract a reduction of tenfold in sensitivity was observed by nested PCR. When cells were grown on a semi-selective medium prior to PCR (Bio-PCR), five cells from pure culture of the pathogen were detected. The bacteria could be detected by nested-PCR or Bio-PCR in symptomless gypsophila cuttings after 7 days. The Bio-PCR procedure described in this study can be used to establish disease-free nuclear stock of mother plants of gypsophila.     

Identification of Fungal rDNA Associated with Soil Suppressiveness Against Heterodera schachtii Using Oligonucleotide Fingerprinting

ABSTRACT To understand the nature of a soil with suppressiveness against Heterodera schachtii, an rDNA analysis was used to identify fungi associated with H. schachtii cysts obtained from soils possessing various levels of suppressiveness. Because H. schachtii cysts isolated from these suppressive soils can transfer this beneficial property to nonsuppressive soils, analysis of the microorganisms associated with the cysts should lead to the identification of the causal organisms. Five soil treatments, generated by mixing different amounts of suppressive and fumigation-induced nonsuppressive soils, were infested with second-stage juveniles of H. schachtii and cropped with mustard-greens. Fungi were identified through an rDNA analysis termed oligonucleotide fingerprinting of ribosomal RNA genes (OFRG). Cysts obtained from soil mixtures consisting of 10 and 100% suppressive soil predominantly contained fungal rDNA with high sequence identity to Dactylella oviparasitica. The dominant fungal rDNA in the cysts isolated from the soil mixtures composed of 0.1 and 1% suppressive soil had high sequence identity to Fusarium oxysporum. Polymerase chain reaction (PCR) amplifications performed with sequence-selective primers corroborated the treatment-specific distribution of rDNA clones obtained by the OFRG analysis. When these sequence-selective PCR primers were used to examine H. schachtii cysts from biocidal soil treatments that produced various levels of suppressiveness, only the D. oviparasitica-like rDNA was consistently identified in the highly suppressive soils.