Microbiology & Immunology - Theses

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Author: Alquethamy, Saleh Fahad S × Type: PhD thesis ×

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    Investigating Metal Ion Homeostasis in Acinetobacter baumannii
    Alquethamy, Saleh Fahad S ( 2022)
    Acinetobacter baumannii is a Gram-negative nosocomial pathogen associated with significant disease in immunocompromised individuals. Due to the prevalence of carbapenem-resistant A. baumannii clinical isolates, the WHO has called for novel antimicrobial therapies to combat this pathogen. Mammalian manipulation of metal ion abundance at the host-pathogen interface contributes to control of bacterial infection. Metals, such as copper and zinc, can be removed via sequestration strategies to starve the pathogen of essential micronutrients, or increased to exploit the antimicrobial properties of these metals. Bacterial pathogens can subvert host-mediated metal stress using highly efficacious uptake and efflux pathways to tightly regulate cellular metal homeostasis. Bioinformatic studies have shown that A. baumannii harbours an extensive repertoire of metal ion homeostasis mechanisms, but these have remained poorly characterised. Here, we have characterised the pathways associated with zinc and copper tolerance in A. baumannii. By combining molecular microbiological analyses with inductively coupled plasma-mass spectrometry and transcriptional assays, this work revealed a role for the resistance nodulation cell-division transporter CzcCBA, and the cation diffusion facilitator transporter CzcD, in zinc resistance, and the P-type ATPase CopA, in copper resistance. Despite the potential toxicity of zinc in excess, A. baumannii requires zinc for numerous cellular functions. Accordingly, zinc recruitment in A. baumannii is also a crucial virulence factor. The high-affinity periplasmic solute-binding protein ZnuA is a component of the primary zinc uptake pathway. ZnuA role in A. baumannii Zn uptake has been phenotypically characterised, but the molecular mechanism in which ZnuA acquires Zn remains unknown. Our work characterised ZnuA structural and biochemical properties. This revealed the first structure of A. baumannii ZnuA and shows that the zinc-binding site is comprised by three strictly conserved histidine residues that contribute to metal-recruitment, albeit to different extents. Collectively, these analyses provide new insights into the metal homeostasis mechanisms of A. baumannii and establish a foundation for future antimicrobial therapeutic development to target these pathways.