Mutations That Disrupt Either thepqqor thegdhGene ofRahnella aquatilisAbolish the Production of an Antibacterial Substance and Result in Reduced Biological Control of Grapevine Crown Gall

Abstract

Rahnella aquatilis HX2, a biocontrol agent for grapevine crown gall caused by Agrobacterium vitis, produces an antibacterial substance that inhibits the growth of A. vitis in vitro. In this study, we show that MH15 and MH16, two Tn5-induced mutants of HX2, have lost their abilities to inhibit A. vitis and have reduced biocontrol activities; they grow in logarithmic phase at a rate similar to that of the wild type and have single Tn5 insertions. They are also impaired in producing pyrroloquinoline quinone (PQQ) or glucose dehydrogenase (GDH). Complementation of MH15 and MH16 with cosmid clones of CP465 and CP104 from an HX2 DNA library restored the antibiosis, biocontrol, and PQQ or GDH production phenotypes. A 6.7-kb BamHI fragment from CP465 that fully restored the MH15-affected phenotypes was cloned and sequenced. Sequence analysis of the mutated DNA region resulted in the identification of seven open reading frames (ORFs), six of which share significant homology with PQQ-synthesizing genes in other bacteria, designated pqqA through pqqF. Meanwhile, A 5.5-kb PstI fragment from CP104 fully complemented the MH16 mutant and contained a single ORF highly similar to that of genes coding for GDHs. An in-frame gdh deletion mutant has the same phenotypes as the Tn5 mutant of MH16. Complementation of both deletion and Tn5 gdh mutants restored the affected phenotypes to wild-type levels. Our results suggest that an antibacterial substance plays a role in biocontrol of A. vitis by HX2.

Distribution and Properties of the Genes Encoding the Biosynthesis of the Bacterial Cofactor, Pyrroloquinoline Quinone

“…Five bacteria with demonstrated PQQ biosynthetic capacity were selected for this study: Klebsiella pneumoniae , Methylobacterium extorquens AM1 , Gluconobacter oxydans 621H , Rahnella aquatilis and Streptomyces rochei (19–23). Each of the genes in the pqq operons in these five bacteria (shown in Table 1) was used as starting points for bioinformatics analyses.…”

Characterization of the Biocontrol Activity of Pseudomonas fluorescens Strain X Reveals Novel Genes Regulated by Glucose

“…The oxidation of glucose to gluconic acid is catalysed by membrane-bound quinoprotein glucose dehydrogenases (Gcd) that are involved either in biocontrol of plant pathogens [10], [13] or in pathogenicity of bacteria in mammals [14]. Among various quinoprotein dehydrogenase enzymes in bacteria, Gcd uses pyrroloquinoline quinone (PQQ) as an essential cofactor [15].…”

Pyrroloquinoline Quinone Biosynthesis GenepqqC, a Novel Molecular Marker for Studying the Phylogeny and Diversity of Phosphate-Solubilizing Pseudomonads

“…Several studies with bacterial mutants unable to produce PQQ and gluconic acid have demonstrated the intimate relation of the cofactor to phosphate solubilization processes (7,17). Besides its relevant role in P solubilization, PQQ is reported to be a potent growth-promoting factor for bacteria and plants, has antioxidant properties (5), and is directly related to the production of antimicrobial substances (7,14,36) as well as to the induction of systemic plant defenses (17). Hence, the cofactor PQQ has multiple plant beneficial effects.…”

“…The genes responsible for PQQ production have been cloned and sequenced in several bacterial genera, including Pseudomonas, Methylobacterium, Acinetobacter, Klebsiella, Enterobacter, and Rahnella (5,14,17,36,42). In P. fluorescens B16, the PQQ operon is formed by 11 …”