Broad-Spectrum Inhibitor of Bacterial Polyphosphate Homeostasis Attenuates Virulence Factors and Helps Reveal Novel Physiology of Klebsiella pneumoniae and Acinetobacter baumannii.

Nathan Roberge,Nolan Neville,Katya Douchant,Nadejda Boev,Zongchao Jia,Calvin Sjaarda,Prameet M Sheth,Curtis Noordhof

doi:10.3389/fmicb.2021.764733

Nathan Roberge, Nolan Neville + Show 6 more

Open Access

https://doi.org/10.3389/fmicb.2021.764733

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Abstract

Acinetobacter baumannii and Klebsiella pneumoniae currently rank amongst the most antibiotic-resistant pathogens, responsible for millions of infections each year. In the wake of this crisis, anti-virulence therapeutics targeting bacterial polyphosphate (polyP) homeostasis have been lauded as an attractive alternative to traditional antibiotics. In this work, we show that the small molecule gallein, a known G-protein βγ subunit modulator, also recently proven to have dual-specificity polyphosphate kinase (PPK) inhibition in Pseudomonas aeruginosa, in turn exhibits broad-spectrum PPK inhibition in other priority pathogens. Gallein treatment successfully attenuated virulence factors of K. pneumoniae and A. baumannii including biofilm formation, surface associated motility, and offered protection against A. baumannii challenge in a Caenorhabditis elegans model of infection. This was highlighted most importantly in the critically understudied A. baumannii, where gallein treatment phenocopied a ppk1 knockout strain of a previously uncharacterized PPK1. Subsequent analysis revealed a unique instance of two functionally and phenotypically distinct PPK1 isoforms encoded by a single bacterium. Finally, gallein was administered to a defined microbial community comprising over 30 commensal species of the human gut microbiome, demonstrating the non-disruptive properties characteristic of anti-virulence treatments as microbial biodiversity was not adversely influenced. Together, these results emphasize that gallein is a promising avenue for the development of broad-spectrum anti-virulence therapeutics.

Highlights

Encountering multi-drug resistant bacterial pathogens has become a common occurrence in modern medicine, and the frequency of identification is on the rise
Given that the small molecule gallein (Figure 1A) is a known polyP biosynthesis inhibitor in P. aeruginosa (Neville et al, 2021), we first attempted to identify whether gallein treatment could inhibit polyphosphate kinase 1 (PPK1) from other priority species
Gallein treatment demonstrated dosedependent inhibitory action against K. pneumoniae polyphosphate kinase 2 (PPK2) ATP synthesis (Supplementary Figure 2), reminiscent of the dualspecificity polyphosphate kinase (PPK) inhibition observed in P. aeruginosa

Summary

Introduction

Encountering multi-drug resistant bacterial pathogens has become a common occurrence in modern medicine, and the frequency of identification is on the rise. As antibiotic development has stagnated over the past several decades (Ventola, 2015; Andrei et al, 2019), many species have developed effective resistance strategies to circumvent conventional therapeutics (Rossolini et al, 2014; Dickey et al, 2017) To address this mounting threat, novel classes of drugs targeting an organism’s ability to infect, disseminate, and persist within a host, termed virulence factors, have been proposed as an alternative to antibiotics (Cross, 2008). This approach aims to attenuate virulence but avoid bactericidal activity, thereby reducing the selective pressures which are an inevitable consequence of traditional antibiotic use (Cegelski et al, 2008; Totsika, 2017). This can lead to re-infection, or increased susceptibility to other infectious agents including Clostridiodes difficile (Keeney et al, 2014), yet anti-virulence therapeutics should avoid these disastrous consequences (Totsika, 2016)

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