Influence of the ferric uptake regulator (Fur) protein on pathogenicity in Pectobacterium carotovorum subsp brasiliense

Show simple item record Tanui, Collins Kipngetich Shyntum, Divine Yufetar Priem, Stefan Louis Theron, Jacques Moleleki, Lucy N. 2017-08-11T06:29:18Z 2017-08-11T06:29:18Z 2017-05
dc.description S1 Fig. Schematic representation of how Pcb1692Δfur mutant strain was generated. A) Using specific set of primers, PCR amplifications of the fur upstream and downstream regions were generated as indicated in S1A Fig. Kanamycin cassette was amplified from pKD4 plasmid with primers Kan F and Kan R. B) Primers Fur F and R, were used in a PCR reaction consisting of, the fur upstream kanamycin and downstream PCR fragment to generate a PCR fusion product. C) The fusion product was electroporated into electrocompetent Pcb1692 to generate the Pcb1692Δfur mutant strain (S1D Fig). Both electrocompetent Pcb1692 and Pcb1692Δfur mutant strain were electroporated with empty pTrc99A. en_ZA
dc.description S2 Fig. PCR amplicons used to generate the fur mutant. Lane 1. DNA ladder, 2. fur downstream PCR fragment, 3. kanamycin cassette PCR product, 4. fur upstream PCR fragment, 5. Fusion product consisting of the downstream, kanamycin and upstream fragment. 6. The fragment used for complementation which contains the fur gene and its promoter region. 7. Control. en_ZA
dc.description S3 Fig. Schematic presentation of the putative promoter regions of selected genes in Pcb1692. Based on our qRT-PCR results, some of the genes under the Pcb1692 Fur regulon were aligned to the consensus fur box and the putative fur boxes for each gene is indicated by red boxes. en_ZA
dc.description.abstract Iron is an important nutrient for the survival and growth of many organisms. In order to survive, iron uptake from the environment must be strictly regulated and maintained to avoid iron toxicity. The ferric uptake regulator protein (Fur) regulates genes involved in iron homeostasis in many bacteria, including phytopathogens. However, to date, the role played by Fur in the biology of Pectobacterium carotovorum subsp. brasiliense (Pcb1692), an important pathogen of potatoes, has not yet been studied. To this end, we used the lambda recombineering method to generate a fur mutant strain of Pcb1692 and assessed the virulence and fitness of the mutant strain. The results showed that production of siderophores in Pcb1692Δfur increased compared to the Pcb1692 wild-type and the complemented strain Pcb1692Δfur-pfur. However, production of N-acyl homoserine lactone (AHLs), biofilm formation, exopolysaccharide (EPS) production, virulence on potato tubers and swimming motility, were all significantly decreased in Pcb1692Δfur compared to the wild-type and complemented Pcb1692Δfur-pfur strains. The Pcb1692Δfur mutant also demonstrated significant sensitivity to oxidative stress when exposed to H2O2. Consistent with phenotypic results, qRT-PCR results demonstrated that Fur down-regulates genes which encode proteins associated with: iron uptake (HasA-extracellular heme-binding protein and Ferrodoxin- AED-0004132), stress response (SodC-superoxide dismutase), plant cell wall degrading enzymes (PrtA and CelV) and motility (FlhC and MotA). We conclude that the ferric uptake regulator protein (Fur) of Pcb1692 regulates traits that are important to host-pathogens interactions. en_ZA
dc.description.department Forestry and Agricultural Biotechnology Institute (FABI) en_ZA
dc.description.department Microbiology and Plant Pathology en_ZA
dc.description.librarian am2017 en_ZA
dc.description.sponsorship The National Research Foundation (NRF), South Africa through Competitive Funding for Rated Researchers (CFRR) 98993; NRF Bioinformatics and Functional Genomics (BFG 93685) and NRF Research Technology and Transfer Fund (RTF) 98654. CKT was funded by University of Pretoria Bursary and SLP funded by the NRF Grant Holder Linked Bursary. SLP was also funded by Potatoes South Africa Transformation Bursary. en_ZA
dc.identifier.citation Tanui CK, Shyntum DY, Priem SL, Theron J, Moleleki LN (2017) Influence of the ferric uptake regulator (Fur) protein on pathogenicity in Pectobacterium carotovorum subsp. brasiliense. PLoS ONE 12(5): e0177647. 10.1371/journal.pone.0177647. en_ZA
dc.identifier.issn 1932-6203 (online)
dc.identifier.other 10.1371/journal.pone.0177647
dc.language.iso en en_ZA
dc.publisher Public Library of Science en_ZA
dc.rights © 2017 Tanui et al. This is an open access article distributed under the terms of the Creative Commons Attribution License. en_ZA
dc.subject Iron en_ZA
dc.subject Protein en_ZA
dc.subject Enzymes en_ZA
dc.subject Ferric uptake regulator (Fur) en_ZA
dc.subject Pathogenicity en_ZA
dc.subject Pectobacterium carotovorum subsp. brasiliense en_ZA
dc.title Influence of the ferric uptake regulator (Fur) protein on pathogenicity in Pectobacterium carotovorum subsp brasiliense en_ZA
dc.type Article en_ZA

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