Acinetobacter baumannii is one of many bacteria that have evolved to live socially in communities called “biofilms.” Like a small town, biofilms make it easier for bacteria to survive, providing strength in numbers against antibodies and other microbes, protecting them against environmental hazards such as drying out, detergents, and antibiotics, and even making it easier for them to metabolize food. A biofilm is made out of extracellular polysaccharides, which are the building materials that hold the cell clusters together and grant the colonies the benefits of increased size and better surface adhesion.
Because of the bacteria's ability to form biofilms, which allow the pathogen to persist in any environment, be it a catheter or the human body, A. baumannii was named an urgent threat by the CDC in 2019. Biofilm infections are particularly common in tissues such as the mucous membranes of the respiratory system and urinary tract, though A. baumannii can also cause infections in the bloodstream, which can lead to a fatal condition known as sepsis.
Cartoon of a catheter where A. baumannii happily resides source
A. baumannii relies on contaminated hospital equipment as a major source of infection in medical settings. This bacteria primarily affects people with medical supports such as ventilators or catheters, as the bacteria can easily survive on such surfaces, though it can also take root in wound sites during prolonged hospital stays. Its virulence (severity or harmfulness of a pathogen) is due in part to its ability to survive on these surfaces for long periods of time, and though good hygiene practices prevent the person to person transmission, immunocompromised patients can also contract it by touching a contaminated surface. Treatment for infection of this pathogen is often personalized due to its capacity to develop resistance to antibiotics by modifying its outer membrane as well as by quickly acquiring new genes from its environment.
One of the features of A. baumannii that allows it to persist on surfaces and in infections is the unique structure of its lipid A. Lipid A is a common membrane feature in many bacteria and is extremely important in regards to virulence and the identification and regulation of the body’s immune response to infection. Unlike other gram-negative bacteria, it has a hepta-acylated lipid A as opposed to being hexa-acylated, which is more common. The hepta-acylated lipid A that A. baumannii has protects the outer membrane and reduces any host immune recognition. It also helps with bacterial resistance to CAMPs (strong immune system response regulators).
The goal of a study done by Joseph Boll et al. was to study exactly how the lipid A structure of A. baumannii functions to protect the bacteria from both desiccation (drying out) and the human immune system, and to investigate whether they could control the sensitivity of the microbe to stress both from the environment and the human immune system by altering its structure.
In their study, Boll et al. investigated A. baumannii’s lipid A synthesis with the hypothesis that if lipid A biosynthesis was inhibited by targeting an acyltransferase, LpxMab(A. baumannii LpxM), then A. baumannii fitness would be decreased, thus providing a novel target by which to combat the pathogen. In other words, the primary goal of this study was to see if they could manipulate the synthesis of lipid A in order to make the pathogen more susceptible to the human immune system and to desiccation. A study of this nature is important given the growing concern in the medical community about drug-resistant infections. By finding a way to target lipid A in the cell membrane, the pathogen could be killed in a way that bypasses the difficulty of its resistance to antibiotics and possibly introduces new research for treating other drug-resistant diseases.
Boll et al. studied two acyltransferases, LpxM and LpxL, which were believed to be involved in modifying Lipid A. Unlike other pathogens, A. baumannii evolved a PagP-independent mechanism (it does not use the acetyltransferase PagP that most do to synthesize its hepta-acylated Lipid A) to synthesize its protective hepta-acylated Lipid A, using instead the dual acyltransferases LpxM and LpxL. Boll et al. went through eight different experiments to look at the two acyltransferases LpxM and LpxL, as well as other factors that lead to modified lipid A in A. baumannii. One of the experiments looked at how A. baumannii LpxM contributes to vertebrate and polymyxin (antibiotic) cationic antimicrobial peptides (CAMP) resistance in A. baumannii by survival assays (which determines the reproductive ability of a cell or group of cells). CAMPs typically serve as therapeutic alternatives since they target the conserved lipid A component of gram-negative outer membranes to lyse the bacterial cell. In order to get around this, A. baumannii has fortified its outer membrane with hepta-acylated lipid A to protect against CAMP-dependent cell lysis.
A. baumannii was incubated with antibiotics C18, polymyxin A, or polymyxin E for two hours. After which they were serially diluted and plated to recover viable bacteria. The results showed that A. baumannii LpxM dependent acylation promotes resistance to vertebrate CAMPs and medically relevant polymyxin antibiotics. The next experiment used a virulence model to show the survival dependency on A. baumannii LpxM in Galleria mellonella, a species of moth.
Desiccation survival assay of A. baumannii in LB broth and human serum
In this study, aside from protecting the microbe from the human immune system, A. baumannii LpxM-dependent acylation is essential for the microbe to survive environment stressors, specifically desiccation which is shown in the eighth experiment (fig. 8). When talking about desiccation, we are referring to the drying out of living organisms or the state of extreme dryness which causes microorganisms to not be able to grow and divide. Figure 8 is both interesting to look at and important in understanding how A. baumannii is able to survive in harsh environments such as a hospital. This figure shows the A. baumannii desiccation survival assay that was performed on the two wild-types and complemented strains. Figure 8A shows the pre and post desiccation survival of each A. baumannii strain where they used LB broth, which is a typical bacterial growth medium to suspend the cells. Figure 8B shows the pre and post desiccation survival of each A. baumannii strain where they used a human serum to suspend the cells. Following desiccation, all groups were found to still have cells and were spotted again. For both trials, the strains were spotted on agar in a 10-fold dilution series before and after desiccation on polystyrene. Their results showed that A. baumannii is able to survive before and after desiccation, when suspended in human serum. This leads to the question of what it depends on to survive desiccation? The experimenters found that the results from the desiccation assay suggest that the lipid components of the outer membrane contribute to A. baumannii desiccation survival. A. baumannii’s unique ability to survive under desiccation gives the result of increased infection and transmission rates with wild-type cells persisting under desiccative conditions for weeks or months, finding treatment options are imperative.
Different mutants of A. baumannii, washed in either a generic growth medium suspension of LB broth or a human serum in a ten-fold dilution series, were placed on a polystyrene surface to study how each mutant would survive drying out. Each plate was taken from the desiccated samples to see if any of the cells were able to survive, with the solid gray plates showing multi-cell survival and the single bright spots representing the survival of only a few individual cells. This experiment not only shows that wild-type A. baumannii is able to survive desiccation in a human-cell growth medium but also that its survival is related to the structure of its outer membrane. The microbe’s ability to survive dryness is part of how it persists on hospital surfaces, including ventilators and catheters. Since its survival was impacted in the mutant strains the experiment indicates that targeting the structure of the outer membrane could be a strategy to combat the pathogen.
One of the most urgent concerns for human health is the increasing number of multidrug-resistant infections as a result of dependence on antibiotics. There are many examples of pathogens that develop immunity to medication, such as MRSA and tuberculosis. Modern medicine is already trying to come up with non-antibiotic solutions to this microscopic arms race, and this study on A. baumannii introduces an intriguing route of study into how to sensitize resistant microbes to treatments. In their paper, Boll et al. show that inhibiting LpxMAb is an effective way to cause cell membrane of A. baumannii to become sensitized to a wide range of host and polymyxin CAMPs, potentially introducing a novel mode of enhanced drug delivery as a synergistic antimicrobial agent, as well as making it harder for the pathogen to survive drying out on surfaces. By making the pathogens sensitive to CAMPs there is an added benefit in that CAMPs could be an alternative to widely-prescribed antibiotics. For example, CAMPs such as magainin, which works by disordering outer membrane acyl chains, and cecropin, which disrupts lipid elements in biological membranes, along with other molecules could be used with an LpxMAb inhibitor to lyse (kill) multidrug-resistant A. baumannii infections.
Studies such as this one will become increasingly necessary if we are to combat ever-adapting pathogens since it is on the whole harder for microbes to develop immunity to agents that destroy their membranes structurally.
Acinetobacter baumannii viewed from under a microscope and on a agar plate
Besides informing future methods of drug delivery and treatment, studies such as Boll et al.’s desiccation experiment have broad implications in creating new and better antiseptics, since the best treatment for any infection is prevention. By having a better understanding of how and why tough to kill microbes work as they do, doctors and scientists will be better equipped to stop infections even before they start.
Charlotte Landeryou '20 is an English major and biology minor. While at Mount Holyoke, her main focus was creative writing but also learned her way around a microscope. Her passions outside of class are writing and reproductive justice, and following graduation she plans to spend a gap year focusing on both. Her long term plan is to become a midwife. Her favorite hobbies are snuggling cats and searching for the perfect coffee.
Others coming soon!
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