Siderophore receptor PupA as a marker to monitor wild-type Pseudomonas putida WCS358 in natural environments.
For application of genetically engineered fluorescent Pseudomonas spp., specific markers are required for monitoring of wild-type Pseudomonas strains and their genetically modified derivatives in natural environments. In this study, the specific siderophore receptor PupA of plant growth-promoting Pseudomonas putida WCS358 was used as a marker to monitor wild-type strain WCS358. After introduction into natural soil and rhizosphere environments, strain WCS358 could be recovered efficiently on a medium amended with 300 microM pseudobactin 358. Although low population densisties of indigenous pseudomonads (less than or equal to 10(3)/g of soil or root) were recovered on the pseudobactin 358-amended medium, subsequent agglutination assays with a WCS358-specific polyclonal antiserum enabled accurate monitoring of populations of wild-type strain WCS358 over a range of approximately 10(3) to 10(7) CFU/g of soil or root. Genetic analysis of the background population by PCR and Southern hybridization revealed that natural occurrence of the pupA gene was limited to a very small number of indigenous Pseudomonas spp. which are very closely related to P. putida WCS358. The PupA marker system enabled the study of differences in rhizosphere colonization among wild-type strain WCS358, rifampin-resistant derivative WCS358rr, and Tn5 mutant WCS358::xylE. Chromosomally mediated rifampin resistance did not affect the colonizing ability of P. putida WCS358. However, Tn5 mutant WCS358::xylE colonized the radish rhizosphere significantly less than did its parental strain.
Department of Plant Ecology and Evolutionary Biology, Utrecht University, The Netherlands.
J M
JM
Raaijmakers
W
W
Bitter
H L
HL
Punte
P A
PA
Bakker
P J
PJ
Weisbeek
B
B
Schippers
D001425:Bacterial Outer Membrane Proteins; D001426:Bacterial Proteins / Q000032:analysis* / Q000235:genetics; D001483:Base Sequence; D015415:Biomarkers; D015139:Blotting, Southern; D004269:DNA, Bacterial / Q000032:analysis; D004352:Drug Resistance, Microbial; D004784:Environmental Monitoring / Q000379:methods*; D008969:Molecular Sequence Data; D010944:Plants / Q000382:microbiology*; D016133:Polymerase Chain Reaction; D016958:Pseudomonas putida / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification*; D011956:Receptors, Cell Surface / Q000032:analysis* / Q000235:genetics; D012293:Rifampin / Q000494:pharmacology; D012988:Soil Microbiology; D011198:Solanum tuberosum / Q000382:microbiology
D001425:Bacterial Outer Membrane Proteins; D001426:Bacterial Proteins; D015415:Biomarkers; D004269:DNA, Bacterial; D011956:Receptors, Cell Surface; C047631:pseudobactin receptor, Pseudomonas; C035902:siderophore receptors; D012293:Rifampin