Xanthoferrin, the α‐hydroxycarboxylate‐type siderophore of Xanthomonas campestris pv. campestris, is required for optimum virulence and growth inside cabbage

Abstract

Xanthomonas campestris pv. campestris causes black rot, a serious disease of crucifers. Xanthomonads encode a siderophore biosynthesis and uptake gene cluster xss (Xanthomonas siderophore synthesis) involved in the production of a vibrioferrin-type siderophore. However, little is known about the role of the siderophore in the iron uptake and virulence of X. campestris pv. campestris. In this study, we show that X. campestris pv. campestris produces an α-hydroxycarboxylate-type siderophore (named xanthoferrin), which is required for growth under low-iron conditions and for optimum virulence. A mutation in the siderophore synthesis xssA gene causes deficiency in siderophore production and growth under low-iron conditions. In contrast, the siderophore utilization ΔxsuA mutant is able to produce siderophore, but exhibits a defect in the utilization of the siderophore-iron complex. Our radiolabelled iron uptake studies confirm that the ΔxssA and ΔxsuA mutants exhibit defects in ferric iron (Fe ) uptake. The ΔxssA mutant is able to utilize and transport the exogenous xanthoferrin-Fe complex; in contrast, the siderophore utilization or uptake mutant ΔxsuA exhibits defects in siderophore uptake. Expression analysis of the xss operon using a chromosomal gusA fusion indicates that the xss operon is expressed during in planta growth and under low-iron conditions. Furthermore, exogenous iron supplementation in cabbage leaves rescues the in planta growth deficiency of ΔxssA and ΔxsuA mutants. Our study reveals that the siderophore xanthoferrin is an important virulence factor of X. campestris pv. campestris which promotes in planta growth by the sequestration of Fe .

Co-regulation of Iron Metabolism and Virulence Associated Functions by Iron and XibR, a Novel Iron Binding Transcription Factor, in the Plant Pathogen Xanthomonas

“…Xanthomonas group of phytopathogens produces vibrioferrin type of siderophore (xanthoferrin) [12–14]. However, the contribution of vibrioferrin siderophore in virulence appears to be variable among xanthomonads.…”

“…Furthermore, in Xanthomonas campestris pv. campestris , it has been shown that genes involved in flagellar biogenesis, chemotaxis and iron uptake and metabolism (ferric iron uptake) are required for optimum virulence [14,50]. Interestingly, (XC_2234) flgB , XC_2298 ( motD ), XC_2241 ( flgI ), XC_2260 ( fliF ) genes, which are regulated by XibR (S3 Table; S9 Table), has been shown to be required for the virulence of Xcc [50].…”

“…Siderophores estimation of different Xcc strains was done by the HPLC quantification of the purified vibrioferrin from cell-free culture supernatant as described previously [13,14] with few modifications. Briefly, different Xcc strains were grown up to the density of 1.0 X 10 9 cells/ml in rich PS medium with respective antibiotics.…”

“…Total RNA of cells was isolated by Trizol (Invitrogen, CA, U.S.A.) method as per the manufacturer’s instructions. Real-time qRT-PCR was done in 7500 Real-time PCR system (Applied Biosystems) and analyzed by SDS relative quantification software (Applied Biosystems, USA) as described previously [34,14]. Primers used for the real-time qRT-PCR are listed in supporting S13 Table.…”

“…Xanthomonads encodes an xss ( X anthomonas s iderophore s ynthesis) operon which is required for the production of siderophore vibrioferrin under iron-restricted conditions, and iron metabolism plays a critical role in their virulence [12–14]. These phytopathogen use cell-cell signaling mediated by diffusible quorum sensing signal molecule to regulate the expression of iron uptake and metabolism functions contributing to virulence and growth within host [13,15].…”