Predatory Bacteria: A Potential Ally against Multidrug-Resistant Gram-Negative Pathogens

Multidrug-resistant (MDR) Gram-negative bacteria have emerged as a serious threat to human and animal health. Bdellovibrio spp. and Micavibrio spp. are Gram-negative bacteria that prey on other Gram-negative bacteria. In this study, the ability of Bdellovibrio bacteriovorus and Micavibrio aeruginosavorus to prey on MDR Gram-negative clinical strains was examined. Although the potential use of predatory bacteria to attack MDR pathogens has been suggested, the data supporting these claims is lacking. By conducting predation experiments we have established that predatory bacteria have the capacity to attack clinical strains of a variety of ß-lactamase-producing, MDR Gram-negative bacteria. Our observations indicate that predatory bacteria maintained their ability to prey on MDR bacteria regardless of their antimicrobial resistance, hence, might be used as therapeutic agents where other antimicrobial drugs fail. © 2013 Kadouri et al

Evolutionary diversification of the RomR protein of the invasive deltaproteobacterium, Bdellovibrio bacteriovorus

This is the final version. Available on open access from Nature Research via the DOI in this recordBdellovibrio bacteriovorus is a predatory deltaproteobacterium that encounters individual Gram-negative prey bacteria with gliding or swimming motility, and then is able to invade such prey cells via type IVa pilus-dependent mechanisms. Movement control (pili or gliding) in other deltaproteobacteria, such as the pack hunting Myxococcus xanthus, uses a response regulator protein, RomRMx (which dynamically relocalises between the cell poles) and a GTPase, MglAMx, previously postulated as an interface between the FrzMx chemosensory system and gliding or pilus-motility apparatus, to produce regulated bidirectional motility. In contrast, B. bacteriovorus predation is a more singular encounter between a lone predator and prey; contact is always via the piliated, non-flagellar pole of the predator, involving MglABd, but no Frz system. In this new study, tracking fluorescent RomRBd microscopically during predatory growth shows that it does not dynamically relocalise, in contrast to the M. xanthus protein; instead having possible roles in growth events. Furthermore, transcriptional start analysis, site-directed mutagenesis and bacterial two-hybrid interaction studies, indicate an evolutionary loss of RomRBd activation (via receiver domain phosphorylation) in this lone hunting bacterium, demonstrating divergence from its bipolar role in motility in pack-hunting M. xanthus and further evolution that may differentiate lone from pack predators.Leverhulme TrustMedical Research Council (MRC)Biotechnology and Biological Sciences Research Council (BBSRC

Ankyrin-mediated self-protection during cell invasion by the bacterial predator Bdellovibrio bacteriovorus

Predatory Bdellovibrio bacteriovorus are natural antimicrobial organisms, killing other bacteria by whole-cell invasion. Self-protection against prey-metabolizing enzymes is important for the evolution of predation. Initial prey entry involves the predator’s peptidoglycan DD-endopeptidases, which decrosslink cell walls and prevent wasteful entry by a second predator. Here we identify and characterize a self-protection protein from B. bacteriovorus, Bd3460, which displays an ankyrin-based fold common to intracellular pathogens of eukaryotes. Co-crystal structures reveal Bd3460 complexation of dual targets, binding a conserved epitope of each of the Bd3459 and Bd0816 endopeptidases. Complexation inhibits endopeptidase activity and cell wall decrosslinking in vitro. Self-protection is vital — DBd3460 Bdellovibrio deleteriously decrosslink self-peptidoglycan upon invasion, adopt a round morpholog, and lose predatory capacity and cellular integrity. Our analysis provides the first mechanistic examination of self-protection in Bdellovibrio, documents protection-multiplicity for products of two different genomic loci, and reveals an important evolutionary adaptation to an invasive predatory bacterial lifestyle

Feedback control architecture and the bacterial chemotaxis network.

PMCID: PMC3088647This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.Bacteria move towards favourable and away from toxic environments by changing their swimming pattern. This response is regulated by the chemotaxis signalling pathway, which has an important feature: it uses feedback to 'reset' (adapt) the bacterial sensing ability, which allows the bacteria to sense a range of background environmental changes. The role of this feedback has been studied extensively in the simple chemotaxis pathway of Escherichia coli. However it has been recently found that the majority of bacteria have multiple chemotaxis homologues of the E. coli proteins, resulting in more complex pathways. In this paper we investigate the configuration and role of feedback in Rhodobacter sphaeroides, a bacterium containing multiple homologues of the chemotaxis proteins found in E. coli. Multiple proteins could produce different possible feedback configurations, each having different chemotactic performance qualities and levels of robustness to variations and uncertainties in biological parameters and to intracellular noise. We develop four models corresponding to different feedback configurations. Using a series of carefully designed experiments we discriminate between these models and invalidate three of them. When these models are examined in terms of robustness to noise and parametric uncertainties, we find that the non-invalidated model is superior to the others. Moreover, it has a 'cascade control' feedback architecture which is used extensively in engineering to improve system performance, including robustness. Given that the majority of bacteria are known to have multiple chemotaxis pathways, in this paper we show that some feedback architectures allow them to have better performance than others. In particular, cascade control may be an important feature in achieving robust functionality in more complex signalling pathways and in improving their performance

Engulfment, persistence and fate of Bdellovibrio bacteriovorus predators inside human phagocytic cells informs their future therapeutic potential

In assessing the potential of predatory bacteria, such as Bdellovibrio bacteriovorus, to become live therapeutic agents against bacterial infections, it is crucial to understand and quantify Bdellovibrio host cell interactions at a molecular level. Here, we quantify the interactions of live B. bacteriovorus with human phagocytic cells, determining the uptake mechanisms, persistence, associated cytokine responses and intracellular trafficking of the non-growing B. bacteriovorus in PMA-differentiated U937 cells. B. bacteriovorus are engulfed by U937 cells and persist for 24h without affecting host cell viability and can be observed microscopically and recovered and cultured post-uptake. The uptake of predators is passive and depends on the dynamics of the host cell cytoskeleton; the engulfed predators are eventually trafficked through the phagolysosomal pathway of degradation. We have also studied the prevalence of B. bacteriovorus specifc antibodies in the general human population. Together, these results quantify a period of viable persistence and the ultimate fate of B. bacteriovorus inside phagocytic cells. They provide new knowledge on predator availability inside hosts, plus potential longevity and therefore potential efficacy as a treatment in humans and open up future fields of work testing if predators can prey on host-engulfed pathogenic bacteria

A Transcriptional “Scream” Early Response of E. coli Prey to Predatory Invasion by Bdellovibrio

We have transcriptionally profiled the genes differentially expressed in E. coli prey cells when predatorily attacked by Bdellovibrio bacteriovorus just prior to prey cell killing. This is a brief, approximately 20–25 min period when the prey cell is still alive but contains a Bdellovibrio cell in its periplasm or attached to and penetrating its outer membrane. Total RNA was harvested and labelled 15 min after initiating a semi-synchronous infection with an excess of Bdellovibrio preying upon E. coli and hybridised to a macroarray spotted with all predicted ORFs of E. coli. SAM analysis and t-tests were performed on the resulting data and 126 E. coli genes were found to be significantly differentially regulated by the prey upon attack by Bdellovibrio. The results were confirmed by QRT-PCR. Amongst the prey genes upregulated were a variety of general stress response genes, potentially “selfish” genes within or near prophages and transposable elements, and genes responding to damage in the periplasm and osmotic stress. Essentially, the presence of the invading Bdellovibrio and the resulting damage to the prey cell elicited a small “transcriptional scream”, but seemingly no specific defensive mechanism with which to counter the Bdellovibrio attack. This supports other studies which do not find Bdellovibrio resistance responses in prey, and bodes well for its use as a “living antibiotic”

Measuring and modelling the response of Klebsiella pneumoniae KPC prey to Bdellovibrio bacteriovorus predation, in human serum and defined buffer

In worldwide conditions of increasingly antibiotic-resistant hospital infections, it is important to research alternative therapies. Bdellovibrio bacteriovorus bacteria naturally prey on Gram-negative pathogens, including antibiotic-resistant strains and so B. bacteriovorus have been proposed as "living antibiotics" to combat antimicrobially-resistant pathogens. Predator-prey interactions are complex and can be altered by environmental components. To be effective B. bacteriovorus predation needs to work in human body fluids such as serum where predation dynamics may differ to that studied in laboratory media. Here we combine mathematical modelling and lab experimentation to investigate the predation of an important carbapenem-resistant human pathogen, Klebsiella pneumoniae, by B. bacteriovorus in human serum versus buffer. We show experimentally that B. bacteriovorus is able to reduce prey numbers in each environment, on different timescales. Our mathematical model captures the underlying dynamics of the experimentation, including an initial predation-delay at the predator-prey-serum interface. Our research shows differences between predation in buffer and serum and highlights both the potential and limitations of B. bacteriovorus acting therapeutically against K. pneumoniae in serum, informing future research into the medicinal behaviours and dosing of this living antibacterial

Fluorescent D-amino-acids reveal bi-cellular cell wall modifications important for Bdellovibrio bacteriovorous predation

Modification of essential bacterial peptidoglycan (PG) containing cell walls can lead to antibiotic resistance, for example β-lactam resistance by L,D-transpeptidase activities. Predatory Bdellovibrio bacteriovorus are naturally antibacterial and combat infections by traversing, modifying and finally destroying walls of Gram-negative prey bacteria, modifying their own PG as they grow inside prey. Historically, these multi-enzymatic processes on two similar PG walls have proved challenging to elucidate. Here, with a PG labelling approach utilizing timed pulses of multiple fluorescent D-amino acids (FDAAs), we illuminate dynamic changes that predator and prey walls go through during the different phases of bacteria:bacteria invasion. We show formation of a reinforced circular port-hole in the prey wall; L,D-transpeptidaseBd mediated D-amino acid modifications strengthening prey PG during Bdellovibrio invasion and a zonal mode of predator-elongation. This process is followed by unconventional, multi-point and synchronous septation of the intracellular Bdellovibrio, accommodating odd- and even-numbered progeny formation by non-binary division

Investigating the Responses of Human Epithelial Cells to Predatory Bacteria

One beguiling alternative to antibiotics for treating multi-drug resistant infections are Bdellovibrio-and-like-organisms (BALOs), predatory bacteria known to attack human pathogens. Consequently, in this study, the responses from four cell lines (three human and one mouse) were characterized during an exposure to different predatory bacteria, Bdellovibrio bacteriovorus HD100, Bacteriovorus BY1 and Bacteriovorax stolpii EB1. TNF-alpha levels were induced in Raw 264.7 mouse macrophage cultures with each predator, but paled in comparison to those obtained with E. coli. This was true even though the latter strain was added at an 11.1-fold lower concentration (p < 0.01). Likewise, E. coli led to a significant (54%) loss in the Raw 264.7 murine macrophage viability while the predatory strains had no impact. Tests with various epithelial cells, including NuLi-1 airway, Caco2, HT29 and T84 colorectal cells, gave similar results, with E. coli inducing IL-8 production. The viabilities of the NuLi-1 and Caco-2 cells were slightly reduced (8%) when exposed to the predators, while T84 viability remained steady. In no cases did the predatory bacteria induce actin rearrangement. These results clearly demonstrate the gentle natures of predatory bacteria and their impacts on human cells.ope

Bacterial Chemotaxis in an Optical Trap

An optical trapping technique is implemented to investigate the chemotactic behavior of a marine bacterial strain Vibrio alginolyticus. The technique takes the advantage that the bacterium has only a single polar flagellum, which can rotate either in the counter-clock-wise or clock-wise direction. The two rotation states of the motor can be readily and instantaneously resolved in the optical trap, allowing the flagellar motor switching rate to be measured under different chemical stimulations. In this paper the focus will be on the bacterial response to an impulsive change of chemoattractant serine. Despite different propulsion apparati and motility patterns, cells of V. alginolyticus apparently use a similar response as Escherichia coli to regulate their chemotactic behavior. Specifically, we found that the switching rate of the bacterial motor exhibits a biphasic behavior, showing a fast initial response followed by a slow relaxation to the steady-state switching rate . The measured can be mimicked by a model that has been recently proposed for chemotaxis in E. coli. The similarity in the response to the brief chemical stimulation in these two different bacteria is striking, suggesting that the biphasic response may be evolutionarily conserved. This study also demonstrated that optical tweezers can be a useful tool for chemotaxis studies and should be applicable to other polarly flagellated bacteria