Relative importance of bacteriocin-like genes in antagonism of Xanthomonas perforans tomato race 3 to Xanthomonas euvesicatoria tomato race 1 strains.

Bacterial spot of tomatoes and peppers is caused by Xanthomonas campestris pv. vesicatoria. Pohronezny and Volin (34) estimated that as high as 50% loss of marketable fruit was due to bacterial spot on tomatoes. Control of bacterial spot of tomato is difficult when high temperatures and high moisture exist. Bactericides, such as fixed coppers and streptomycin, have provided the primary means of control (30, 45, 46); however, streptomycin-resistant mutants and copper-tolerant strains became prevalent on treated plants (46). Marco and Stall (30) showed that many Xanthomonas euvesicatoria strains were tolerant of copper and addition of mancozeb to copper sprays improved control of the copper-tolerant strains (6, 30). However, they also showed that this treatment is insufficient when conditions favorable for disease development exist. On tomato, three races, designated tomato race 1 (T1), T2, and T3, were identified based on their reaction on three tomato genotypes (25, 26, 44). Recent reclassification of the xanthomonads associated with bacterial spot of tomato has changed the species classification of these races. Currently, T1 strains are classified as X. euvesicatoria, T2 strains are classified as X. vesicatoria, and T3 strains are now classified as Xanthomonas perforans (24). T1 and T2 were first identified in 1990 (55) and are distributed worldwide (3), although only T1 strains were present in Florida until the early 1990s. In 1991, T3 appeared in Florida fields (26) and eventually predominated (22). This new race had antagonistic activity toward T1 strains (8) and was determined to be a new group designated group C (23). Genetically, T3 strains are most closely related to T1 strains based on DNA similarity values (23). In a previous study, wild type (WT) T3 strains were shown to be antagonistic toward WT T1 strains (8). Tudor-Nelson et al. (52) identified three cosmid clones, designated BCN-A, BCN-B, and BCN-C, which were found to confer the ability to inhibit a sensitive T1 strain in plate assays. These compounds were determined to have narrow inhibition spectra and fit the definition of a bacteriocin described by Reeves (36) based on the following criteria: (i) the presence of a biologically active protein moiety, (ii) inducibility with mitomycin C, and (iii) non-self inhibition (52). The three clones isolated by Tudor-Nelson et al. (52) were unique in activity and specificity based on differential inhibition of selected sensitive T1 and T2 strains (52). It has been speculated that the bacteriocins may confer in part or completely this competitive advantage. Of the bacteriocins which have been well characterized in the field of bacteriology, the most studied of the gram-negative bacteria bacteriocins is the colicin family found in Escherichia coli. The genes encoding the bacteriocins of E. coli are found on plasmids. The modes of action of these colicins range from pore formation in the cell membrane to nuclease activity against DNA, rRNA, and tRNA (54). Most of the bacteriocins produced by gram-negative bacteria are large proteins and, in the case of the colicins, range in size from 449 to 629 amino acids for the pore-forming bacteriocins and 178 to 777 amino acids for the nuclease bacteriocins (54). The extensive research on the colicin family of bacteriocins has offered many helpful insights into the evolution of bacteriocins and the relationships between the producing and sensitive strains. Theories have developed from studying this family related to the evolution of bacteriocins. It was hypothesized that new bacteriocin-producing strains arise through recombination (4, 47), as well as mutations (37, 38). It is likely that many closely related bacteria acquire bacteriocins by horizontal transfer. For instance, homology was detected between a bcnA-specific probe and genomic DNA of X. campestris pv. glycines strain 1717 (51). In addition, a subclone from X. perforans T3 strain 91-118 that conferred immunity to BCN-A also conferred immunity to X. campestris pv. glycines (16, 52). There has been conjecture by Riley and Wertz (39) that bacteriocins may serve as anticompetitors enabling the strain to invade an established microbial community. This appears to suggest that bacteriocins may play a defensive role in limiting competition among bacteria, essentially giving the producing cells a competitive advantage over the nonproducing sensitive strains. The ability of bacteriocin-producing strains to reduce sensitive strains when grown together has been shown previously (13, 18, 33, 42, 49, 53). In this study, we show the relative importance of bacteriocins of T3 strains of X. perforans for their antagonistic activity toward T1 strains in greenhouse and field studies and present information that one bacteriocin-like compound may be a negative factor in competition between strains.