Donor Strains of the Soft-Rot Bacterium Erwinia chrysanthemi and Conjugational Transfer of the Pectolytic Capacity

Abstract

Donor strains of Erwinia chrysanthemi ICPB EC16, a member of the soft-rot (pectolytic) section of the enterobacterial genus Erwinia , were obtained by chromosomal integration of an F′ lac + plasmid originating from Escherichia coli . These stable donor strains, selected from an unstable F′ lac + heterogenote by repeated platings of single Lac + colonies on lactose minimal agar, do not segregate (as does the parent F′ lac + heterogenote) into Lac − or F − clones, in either the presence or absence of acridine orange. One representative donor strain (from the 12 that have been selected) has been examined in more detail; it can transfer ade + , gal + , gtu + (utilization of galacturonate), his + , lac + , leu + , lys + , mcu + (multiple carbohydrate utilization), pat + (production of polygalacturonic acid trans -eliminase), thr + , and trp + in a polarized manner to appropriate recipient strains of E. chrysanthemi ; the frequencies of ade + , leu + , and thr + transfer were higher than those of the other markers tested to date. This donor strain transfers lac + genes during a 6-h mating on membranes; most of the Lac + recombinants are donors of chromosomal markers. The kinetics of entry as well as the frequencies of transfer of chromosomal markers indicate that thr + and leu + enter the recipient as proximal markers and that lac + enters as a distal marker. Analysis of the recombinants demonstrates close linkage between thr and leu, ade and thr, his and pat , and his and trp loci. The results suggest that the integration of F′ lac + into the chromosome of E. chrysanthemi has occurred at a region adjacent to the leu-thr loci, and that the chromosome is transferred in the following sequence: origin---- leu--thr--ade--lys--mcu--pat--his--trp--gal--gtu--lac --F. Plant-tissue maceration occurs in Pat + recombinants and not in Pat − recombinants, even though both form another pectolytic enzyme, hydrolytic polygalacturonase. This genetic evidence supports the idea that the E. chrysanthemi polygalacturonic acid trans -eliminase plays an essential role in bringing about plant-tissue maceration.