Understanding the molecular survival strategies of Acinetobacter baumannii and developing strategies to disable them.

The discovery of penicillin over 70 years ago, and its subsequent uptake by healthcare systems around the world, revolutionised the treatment of bacterial infections. It marked the beginning of a golden age in antibiotic discovery with new classes of antibiotics being routinely discovered, saving millions of lives globally. However, towards the end of the last century the rate of discovery slowed to a near standstill. This lack of discovery has been compounded by the rapid emergence and spread of bacterial pathogens that exhibit resistance to multiple antibiotic treatments. A 2018 report from the World Health Organisation placed Acinetobacter baumannii at the top of a global priority list of bacteria in urgent need of novel treatment strategies. A. baumannii is an opportunistic bacteria that can infect individuals who are already sick leading to a variety of life threatening clinical complications and death. This creates a problem particularly in hospitals where most A. baumannii outbreaks occur. Prior to the 2000s, A. baumannii infections were relatively infrequent and typically susceptible to most front line antibiotics. However, there has been a rapid increase in the number of these infections, such that this pathogen now accounts for 20% of all infections seen in Intensive Care Units (ICUs) worldwide. These infections are also becoming increasingly difficult to treat, with up to 70% of A. baumannii isolated from patients being multidrug resistant. Research into new strategies to prevent and treat A. baumannii infections is now a matter of global priority in order to maintain sustainable access to effective treatments. One key strategy that these bacteria use to stop antibiotics working properly is by forming a community of cells called a biofilm. By coming together in these communities, bacteria are protected from antibiotics, with up to 1,000 times more antibiotic being needed to kill bacteria in a biofilm community compared to bacteria on their own. Another strategy used by A. baumannii is the ability to survive on surfaces like handrails, desks, hospital beds and ventilators, without food or water for months at a time. This survival ability allows this bacteria to survive in hospitals long after any infected patients have left, only to remerge when a sick individual comes in contact with an infected surface. Despite the role that these two survival mechanisms play in the spread and difficulty in treating this pathogen, very little is known about the genes that control these survival strategies. This proposal aims to build on considerable preliminary data by characterising key genes and pathways that regulate the ability of A. baumannii to survive on dry surfaces and to form biofilms. We also aim to identify new drugs that will disrupt these survival stratagies and could potentially be the next generation of antibiotics needed to prevent a post-antibiotic era. A. baumannii is a particular problem for patients with wounds from trauma, surgery or burns. In fact, it has been known to cause outbreaks in specialist wound treatment centres such as Burn ICUs. We have also developed a highly innovative invertebrate assay that will be used to study wound colonisation and biofilm formation. We will explore new ways to deliver drugs to wounds infected with A. baumannii by developing new wound dressings that contain our next generation antibiotics. The work outlined in the proposal has the potential to rapidly advance our understanding of this pathogen at a genetic level, giving novel insights into the key survival mechanisms that have been central to its emergence over the last 20 years. This proposal also has the potential to lead to the development of novel compounds that disable the ability of this pathogen to survive antibiotic treatment in patients and/or survive on hospital surfaces for long periods of time.

Biofilm formation and desiccation tolerance are central factors in the pathogenic success of Acinetobacter baumannii. Pilot data supporting this proposal has implicated two novel regulators in the regulation of these survival phenotypes. This proposal takes a highly interdisciplinary and disruptive approach to characterising these regulators and preventing biofilm formation and desiccation tolerance by this pathogen. This includes the use of cutting-edge technologies such as high through-put programmable robotics and hydrogel printing.
For Aim 1, this proposal will use multi-omics tools to define the molecular mechanisms underpinning the regulation of biofilm formation and desiccation tolerance in A. baummanii. Specifically, this will involves exploring the transcriptomic and lipidomic impact of the newly identified survival regulators. Interactomic analysis will use a custom A. baumannii Bacterial Two Hybrid Library to identify key protein-protein interactions involved in regulating biofilm formation and desiccation tolerance. For Aim 2, this proposal will use workflows optimised for phytochemical identification to identify anti-biofilm and anti-desiccation compounds from lead bioactive extracts. The bioactivity of these compounds will be tested in hydrogels. The mechanisms of action of anti-biofilm and anti-desiccation compounds will be investigated by using robotic screening of a mutant library. For Aim 3, a highly innovative invertebrate model of infection will be used to explore the role of these survival regulators in vivo. This will give high resolution insights into biofilm formation dynamics in vivo and ex vivo desiccation kinetics at different stages of infection. This model will also be used to determine at what stages of wound colonisation these survival regulators are expressed. This part of the proposal will also explore the clinical potential of anti-biofilm and anti-desiccation compounds identified in Aim 2 by testing their in vivo efficacy.