Luminometric analyses of plant root colonization by bioluminescent pseudomonads

Abstract

Bioluminescence was used to study root colonization in nonsterile soil by Tn5-luxAB mutants of four Pseudomonas sp. strains. These bioluminescent strains colonized cucumber, cotton, maize, and soybean roots following seed treatment. In general, there were significant interactions between host and strain. Soybean and cucumber supported the best bacterial colonization, followed by maize and cotton. Across all crops, the best colonizing strains were GR7.4L and GR20.5L, with mean populations of about log 6.0 (colony-forming units + 1)/root system, followed by strains 61.9A.3L and 86.139LR with mean populations of about log 5.0 and 4.5 (colony-forming units + 1)/root system, respectively. The Lux+ strains colonized abundantly the upper root system, while their populations decreased progressively down the root system, and their root-tip populations were independent of mean populations at the crown and middle-root segments. Bioluminescence was observed visually from colonies growing on selective media and photometrically on roots with a luminometer and a charge-coupled device camera. Embedding roots, in situ root luminometer readings, and root-imaging techniques were less sensitive for detecting bacteria on roots than dilution-plating and broth enrichment techniques using selective media. The number of bacteria and competition with other rhizosphere microorganisms affected the sensitivity of the technique used. Selective broth enrichment was less time consuming than the dilution-plating technique. Bioluminescence is a simple and fast method to evaluate colonization of roots by bacteria.Key words: genetic marker, bioluminescence, lux genes, root colonization, Pseudomonas sp.