Abstract

It is known that many pathogens produce high-affinity iron uptake systems like siderophores and/or proteins for utilizing iron bound to heme-containing molecules, which facilitate iron-acquisition inside a host. In mutualistic digestive-tract associations, iron uptake systems have not been as well studied. We investigated the importance of two iron utilization systems within the beneficial digestive-tract association Aeromonas veronii and the medicinal leech, Hirudo verbana. Siderophores were detected in A. veronii using chrome azurol S. Using a mini Tn5, a transposon insertion in viuB generated a mutant unable to utilize iron using siderophores. The A. veronii genome was then searched for genes potentially involved in iron utilization bound to heme-containing molecules. A putative outer membrane heme receptor (hgpB) was identified with a transcriptional activator, termed hgpR, downstream. The hgpB gene was interrupted with an antibiotic resistance cassette in both the parent strain and the viuB mutant, yielding an hgpB mutant and a mutant with both iron uptake systems inactivated. In vitro assays indicated that hgpB is involved in utilizing iron bound to heme and that both iron utilization systems are important for A. veronii to grow in blood. In vivo colonization assays revealed that the ability to acquire iron from heme-containing molecules is critical for A. veronii to colonize the leech gut. Since iron and specifically heme utilization is important in this mutualistic relationship and has a potential role in virulence factor of other organisms, genomes from different Aeromonas strains (both clinical and environmental) were queried with iron utilization genes of A. veronii. This analysis revealed that in contrast to the siderophore utilization genes heme utilization genes are widely distributed among aeromonads. The importance of heme utilization in the colonization of the leech further confirms that symbiotic and pathogenic relationships possess similar mechanisms for interacting with animal hosts.

Highlights

  • Bacteria-host interactions can manifest many different outcomes

  • Bacterial Strains and Growth Conditions The A. veronii strains were cultured at 30◦C and Escherichia coli at 37◦C in Luria broth (LB) (Sambrook and Russell, 2001), or low iron medium (LIM) (Cox, 1994) or LB agar plates (15 g/l of BactoAgar)

  • A. veronii (HM21) Produces a Siderophore and Utilizes Heme-containing Molecules as an Iron Source Our goal was to determine if the A. veronii leech-isolate, HM21, requires an iron utilization system for successfully colonizing the leech gut

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Summary

Introduction

Bacteria-host interactions can manifest many different outcomes. These outcomes can range from the commonly investigated pathogenic ones, in which microbes have a negative effect on a host, to mutualistic interactions, in which both microbe and host benefit. The symbionts require many molecular tools for successful colonization and persistence within this environment (Graf, 2006; Silver et al, 2007a; Nelson et al, 2012). These molecular tools can be critical during the first 24 h of incubation, especially the ability of A. veronii to lyse erythrocytes (Maltz and Graf, 2011). A. veronii, a Gram-negative, facultative anaerobe, is both a symbiont of H. verbana and a human pathogen. The simplicity of the leech gut microbiome and the ability to genetically manipulate A. veronii allows one to study factors important for microbemicrobe interactions and microbe-host interactions in a naturally occurring mutualistic symbioses (Graf, 2006)

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