Bdellovibrio Research Articles

Strain-specific predation of Bdellovibrio bacteriovorus on Pseudomonas aeruginosa with a higher range for cystic fibrosis than for bacteremia isolates

This work aimed to evaluate the predatory activity of Bdellovibrio bacteriovorus 109J on clinical isolates of Pseudomonas aeruginosa selected from well-characterized collections of cystic fibrosis (CF) lung colonization (n = 30) and bloodstream infections (BSI) (n = 48) including strains selected by genetic lineage (frequent and rare sequence types), antibiotic resistance phenotype (susceptible and multidrug-resistant isolates), and colony phenotype (mucoid and non-mucoid isolates). The intraspecies predation range (I-PR) was defined as the proportion of susceptible strains within the entire collection. In contrast, the predation efficiency (PE) is the ratio of viable prey cells remaining after predation compared to the initial inoculum. I-PR was significantly higher for CF (67%) than for BSI P. aeruginosa isolates (35%) probably related to an environmental origin of CF strains whereas invasive strains are more adapted to humans. I-PR correlation with bacterial features such as mucoid morphotype, genetic background, or antibiotic susceptibility profile was not detected. To test the possibility of increasing I-PR of BSI isolates, a polyhydroxyalkanoate depolymerase deficient B. bacteriovorus bd2637 mutant was used. Global median I-PR and PE values remained constant for both predators, but 31.2% of 109J-resistant isolates were susceptible to the mutant, and 22.9% of 109J-susceptible isolates showed resistance to predation by the mutant, pointing to a predator–prey specificity process. The potential use of predators in the clinical setting should be based on the determination of the I-PR for each species, and the PE of each particular target strain.

Production of 3',3'-cGAMP by a Bdellovibrio bacteriovorus promiscuous GGDEF enzyme, Bd0367, regulates exit from prey by gliding motility.

Bacterial second messengers are important for regulating diverse bacterial lifestyles. Cyclic di-GMP (c-di-GMP) is produced by diguanylate cyclase enzymes, named GGDEF proteins, which are widespread across bacteria. Recently, hybrid promiscuous (Hypr) GGDEF proteins have been described in some bacteria, which produce both c-di-GMP and a more recently identified bacterial second messenger, 3′,3′-cyclic-GMP-AMP (cGAMP). One of these proteins was found in the predatory Bdellovibrio bacteriovorus, Bd0367. The bd0367 GGDEF gene deletion strain was found to enter prey cells, but was incapable of leaving exhausted prey remnants via gliding motility on a solid surface once predator cell division was complete. However, it was unclear which signal regulated this process. We show that cGAMP signalling is active within B. bacteriovorus and that, in addition to producing c-di-GMP and some c-di-AMP, Bd0367 is a primary producer of cGAMP in vivo. Site-directed mutagenesis of serine 214 to an aspartate rendered Bd0367 into primarily a c-di-GMP synthase. B. bacteriovorus strain bd0367S214D phenocopies the bd0367 deletion strain by being unable to glide on a solid surface, leading to an inability of new progeny to exit from prey cells post-replication. Thus, this process is regulated by cGAMP. Deletion of bd0367 was also found to be incompatible with wild-type flagellar biogenesis, as a result of an acquired mutation in flagellin chaperone gene homologue fliS, implicating c-di-GMP in regulation of swimming motility. Thus the single Bd0367 enzyme produces two secondary messengers by action of the same GGDEF domain, the first reported example of a synthase that regulates multiple second messengers in vivo. Unlike roles of these signalling molecules in other bacteria, these signal to two separate motility systems, gliding and flagellar, which are essential for completion of the bacterial predation cycle and prey exit by B. bacteriovorus.

Integration of Bdellovibrio spp. with SODIS and Moringa oleifera flocculation to target multi-drug resistant Klebsiella pneumoniae and Pseudomonas aeruginosa

Combination treatments using Bdellovibrio bacteriovorus PF13 and/or TWPF pre-treatment; solar disinfection (SODIS); and Moringa oleifera flocculation, were investigated for the reduction of antibiotic resistant Klebsiella pneumoniae and Pseudomonas aeruginosa. After the various stages of treatment, culture- and molecular-based methods were employed to quantify the respective bacteria. Results indicated that pre-treatment with the B. bacteriovorus strains did not reduce the P. aeruginosa cell concentration. The subsequent exposure of the pre-treated and non-pre-treated samples to SODIS (6 h) then significantly reduced the P. aeruginosa cell counts and gene copies (EMA-qPCR) by up to 7.50 logs, and 4.37 logs, respectively. In contrast, pre-treatment using both B. bacteriovorus strains (PF13 and TWPF) in combination with SODIS significantly reduced the K. pneumoniae cell counts (8.46 logs) and gene copies (4.40 logs) to below detection limit. While significant log reductions were obtained for the K. pneumoniae samples pre-treated with B. bacteriovorus PF13 or TWPF, and SODIS only (no pre-treatment), K. pneumoniae still persisted. Furthermore, flocculation treatment following SODIS did not significantly reduce the concentration of K. pneumoniae or P. aeruginosa. The antibiogram of the target organisms was compared before and after each treatment stage using the VITEK® 2 Compact System. No difference in the organisms’ susceptibility to the tested antibiotics was observed, with both K. pneumoniae and P. aeruginosa maintaining their MDR and XDR status, respectively. It is thus recommended that the interaction and predation kinetics of employing multiple predatory bacteria as a pre-treatment to SODIS is explored, as these treatments eradicated the MDR K. pneumoniae.

Biological Control of Acinetobacter baumannii: In Vitro and In Vivo Activity, Limitations, and Combination Therapies.

The survival, proliferation, and epidemic spread of Acinetobacter baumannii (A. baumannii) in hospital settings is associated with several characteristics, including resistance to many commercially available antibiotics as well as the expression of multiple virulence mechanisms. This severely limits therapeutic options, with increased mortality and morbidity rates recorded worldwide. The World Health Organisation, thus, recognises A. baumannii as one of the critical pathogens that need to be prioritised for the development of new antibiotics or treatment. The current review will thus provide a brief overview of the antibiotic resistance and virulence mechanisms associated with A. baumannii’s “persist and resist strategy”. Thereafter, the potential of biological control agents including secondary metabolites such as biosurfactants [lipopeptides (surfactin and serrawettin) and glycolipids (rhamnolipid)] as well as predatory bacteria (Bdellovibrio bacteriovorus) and bacteriophages to directly target A. baumannii, will be discussed in terms of their in vitro and in vivo activity. In addition, limitations and corresponding mitigations strategies will be outlined, including curtailing resistance development using combination therapies, product stabilisation, and large-scale (up-scaling) production.

Asymmetric peptidoglycan editing generates cell curvature in Bdellovibrio predatory bacteria

Peptidoglycan hydrolases contribute to the generation of helical cell shape in Campylobacter and Helicobacter bacteria, while cytoskeletal or periskeletal proteins determine the curved, vibrioid cell shape of Caulobacter and Vibrio. Here, we identify a peptidoglycan hydrolase in the vibrioid-shaped predatory bacterium Bdellovibrio bacteriovorus which invades and replicates within the periplasm of Gram-negative prey bacteria. The protein, Bd1075, generates cell curvature in B. bacteriovorus by exerting LD-carboxypeptidase activity upon the predator cell wall as it grows inside spherical prey. Bd1075 localizes to the outer convex face of B. bacteriovorus; this asymmetric localization requires a nuclear transport factor 2-like (NTF2) domain at the protein C-terminus. We solve the crystal structure of Bd1075, which is monomeric with key differences to other LD-carboxypeptidases. Rod-shaped Δbd1075 mutants invade prey more slowly than curved wild-type predators and stretch invaded prey from within. We therefore propose that the vibrioid shape of B. bacteriovorus contributes to predatory fitness.

Validating Flow Cytometry as a Method for Quantifying Bdellovibrio Predatory Bacteria and Its Prey for Microbial Ecology.

ABSTRACTBdellovibrio bacteriovorus is a predatory, Gram-negative bacteria that feeds on many pathogenic bacteria and has been investigated as a possible solution for mitigating biofilms in different fields. The application depends on more fundamental ecological studies into the dynamics between Bdellovibrio and their prey. To do so requires an accurate, reliable, and, preferably rapid, way of enumerating the cells. Flow cytometry (FCM) is potentially a rapid, accurate, and inexpensive tool for this, but it has yet to be validated in the enumeration of Bdellovibrio. In this study, we developed a protocol to measure the number of Bdellovibrio in samples of various densities using FCM and compared the results with those of other methods: optical density (OD), PFU assay (PFU), and quantitative PCR (qPCR). We observed a strong correlation between values obtained using FCM and PFU (ρ = 0.923) and FCM and qPCR (ρ = 0.987). Compared to optical density there was a much weaker correlation (ρ = 0.784), which was to be expected given the well-documented uncertainty in converting optical density (OD) to cell numbers. The FCM protocol was further validated by demonstrating its ability to distinguish and count mixed populations of Bdellovibrio and the prey Pseudomonas. Thus, the accuracy of FCM as well as its speed and reproducibility make it a suitable alternative for measuring Bdellovibrio cell numbers, especially where many samples are required to capture the dynamics of predator-prey interactions.IMPORTANCE The rise of antibiotic resistance and the unwanted growth of bacteria is a universally growing problem. Predatory bacteria can be used as a biological alternative to antibiotics because they grow by feeding on other bacteria. To apply this effectively requires further study and a deeper understanding of the forces that drive a prey population to elimination. Initially, such studies require more reliable methods to count these cells. Flow cytometry (FCM) is potentially a rapid, accurate, and inexpensive tool for this, but it has yet to be validated for predatory bacteria. This study develops a protocol to count the predatory bacteria Bdellovibrio bacteriovorus and its Pseudomonas prey using FCM and compare the results with those of other methods, demonstrating its ability for studies into B. bacteriovorus predation dynamics. This could lead to the use of B. bacteriovorus for killing bacterial biofilms in fields, such as drinking water and agriculture.

Bacterial predation in extreme environments

Predatory bacterium are being investigated as a living anti-biotic for multiple real-world settings. One such predatory bacterium, Bdellovibrio bacteriovorus (Bb), has been investigated for the past 50 years yet the strategies in which Bb locate prey is still unknown. Previously, we hypothesized that Bb exploit passive hydrodynamic forces to assist in prey location. Due to the energy-taxing nature of this strategy, we investigated how their dynamics may change in extreme environments where location of prey must be balanced with energy expenditure.

Death by a thousand cuts: investigating pore formation by magainin 2 in the predatory bacterium Bdellovibrio bacteriovorus

Bdellovibrio bacteriovorus is a predatory bacterium that has been proposed as a living antibiotic, potentially capable of clearing or preventing bacterial infections in vivo. However, bdellovibrio has never been successfully established in an animal model, possibly because the conditions under which bdellovibrio thrives in the lab (30 °C, ambient oxygen) are quite different than physiological conditions. Thus, we are interested in how the viability of bdellovibrio might change under physiological conditions, particularly with respect to the activity of antimicrobial peptides (AMPs).

Engineered Bdellovibrio bacteriovorus: A countermeasure for biofilm-induced periodontitis

Periodontitis is caused by subgingival plaque biofilm mainly composed of Gram-negative bacteria, affecting human populations at a high prevalence. Antibiotics therapy is the traditional method for periodontitis, but its efficiency in removing plaque biofilm is limited. With the aim to find an effective therapeutic method to treat periodontitis, we reported a material-assistant micro-organism (MAMO) strategy to eradicate Gram-negative bacteria biofilm using ZnO nanorods engineered Bdellovibrio (ZnO@Bdello). Bdellovibrio bacteriovorus is a natural Gram-negative bacteria predator with strong motility. ZnO nanorods are a kind of common piezoelectric material that senses mechanical pressure and produces reactive oxygen species (ROS). During the predation process of Bdellovibrio, the collision of Bdellovibrio triggered ZnO nanorods to produce ROS to enhance the removal of plaque biofilm. Both in vitro and in vivo studies showed ZnO@Bdello could remove biofilm effectively. In addition, ZnO@Bdello possessed superior competence in alleviating inflammation and inhibiting bone resorption in rats and rabbit periodontitis models. Importantly, this strategy does not induce detectable pathogen resistance and side effects.

Predation of colistin- and carbapenem-resistant bacterial pathogenic populations and their antibiotic resistance genes in simulated microgravity

As mankind evaluates moving toward permanently inhabiting outer space and other planetary bodies, alternatives to antibiotic that can effectively control drug-resistant pathogens are needed. The activity of one such alternative, Bdellovibrio bacteriovorus HD100, was explored here, and was found to be as active or better in simulated microgravity (SMG) conditions as in flask and normal gravity (NG) cultures, with the prey viabilities decreasing by 3- to 7-log CFU/mL in 24 h. The activity of B. bacteriovorus HD100 under SMG was also appraised with three different carbapenem- and colistin-resistant pathogenic bacterial strains. In addition to being more efficient at killing two of these pathogens under SMG conditions (with losses of 5- to 6-log CFU/mL), we also explored the ability of B. bacteriovorus HD100 to hydrolyze the carbapenem- and colistin-resistant gene pools, i.e., mcr-1, blaKPC-2 and blaOXA-51, present in these clinical isolates. We found removal efficiencies of 97.4 ± 0.9 %, 97.8 ± 0.4 % and 99.3 ± 0.1 %, respectively, in SMG cultures, while similar reductions were also seen in the flask and NG cultures. These results illustrate the potential applicability of B. bacteriovorus HD100 as an antibiotic to combat the ever-growing threat of multidrug-resistant (MDR) pathogens during spaceflight, such as in the International Space Station (ISS).

Predation Strategies of the Bacterium Bdellovibrio bacteriovorus Result in Overexploitation and Bottlenecks.

With increasing antimicrobial resistance, alternatives for treating infections or removing resistant bacteria are urgently needed, such as the bacterial predator Bdellovibrio bacteriovorus or bacteriophage. Therefore, we need to better understand microbial predator-prey dynamics. We developed mass-action mathematical models of predation for chemostats, which capture the low substrate concentration and slow growth typical for intended application areas of the predators such as wastewater treatment, aquaculture, or the gut. Our model predicted that predator survival required a minimal prey cell size, explaining why Bdellovibrio is much smaller than its prey. A predator considered to be "too good" (attack rate too high, mortality too low) overexploited its prey, leading to extinction (tragedy of the commons). Surprisingly, a predator taking longer to produce more offspring outcompeted a predator producing fewer offspring more rapidly (rate versus yield trade-off). Predation was only efficient in a narrow region around optimal parameters. Moreover, extreme oscillations under a wide range of conditions led to severe bottlenecks. These could be avoided when two prey species became available in alternating seasons. A bacteriophage outcompeted Bdellovibrio due to its higher burst size and faster life cycle. Together, results suggest that Bdellovibrio would struggle to survive on a single prey, explaining why it must be a generalist predator and suggesting it is better suited than phage to environments with multiple prey. IMPORTANCE The discovery of antibiotics led to a dramatic drop in deaths due to infectious disease. Increasing levels of antimicrobial resistance, however, threaten to reverse this progress. There is thus a need for alternatives, such as therapies based on phage and predatory bacteria that kill bacteria regardless of whether they are pathogens or resistant to antibiotics. To best exploit them, we need to better understand what determines their effectiveness. By using a mathematical model to study bacterial predation in realistic slow growth conditions, we found that the generalist predator Bdellovibrio is most effective within a narrow range of conditions for each prey. For example, a minimum prey cell size is required, and the predator should not be "too good," as this would result in overexploitation risking extinction. Together these findings give insights into the ecology of microbial predation and help explain why Bdellovibrio needs to be a generalist predator.

Quinolone Signals Related to Pseudomonas Quinolone Signal-Quorum Sensing Inhibits the Predatory Activity of Bdellovibrio bacteriovorus.

Bdellovibrio bacteriovorus is one of the predatory bacteria; therefore, it can act as a novel “living antibiotic,” unlike the current antibiotics. Here the predation of Escherichia coli by B. bacteriovorus was inhibited in the presence of Pseudomonas aeruginosa. This study investigated whether P. aeruginosa-induced predation inhibition is associated with bacterial quorum sensing (QS). Each las, rhl, or pqs QS mutant in P. aeruginosa was used to check the predatory activity of E. coli cells using B. bacteriovorus. As a result, the predatory activity of B. bacteriovorus increased in a mutant pqs QS system, whereas wild-type PA14 inhibited the predatory activity. Moreover, the addition of 4-hydroxy-2-heptylquinoline (HHQ) or the analog triggered the low predatory activity of B. bacteriovorus and killed B. bacteriovorus cells. Therefore, a defensive action of P. aeruginosa against B. bacteriovorus is activated by the pqs QS system, which produces some quinolone compounds such as HHQ.

Ocular Delivery of Predatory Bacteria with Cryomicroneedles Against Eye Infection.

The development of potent antibiotic alternatives with rapid bactericidal properties is of great importance in addressing the current antibiotic crisis. One representative example is the topical delivery of predatory bacteria to treat ocular bacterial infections. However, there is a lack of suitable methods for the delivery of predatory bacteria into ocular tissue. This work introduces cryomicroneedles (cryoMN) for the ocular delivery of predatory Bdellovibrio bacteriovorus (B. bacteriovorus) bacteria. The cryoMN patches are prepared by freezing B. bacteriovorus containing a cryoprotectant medium in a microneedle template. The viability of B. bacteriovorus in cryoMNs remains above 80% as found in long‐term storage studies, and they successfully impede the growth of gram‐negative bacteria in vitro or in a rodent eye infection model. The infection is significantly relieved by nearly six times through 2.5 days of treatment without substantial effects on the cornea thickness and morphology. This approach represents the safe and efficient delivery of new class of antimicrobial armamentarium to otherwise impermeable ocular surface and opens up new avenues for the treatment of ocular surface disorders.

Secretion systems play a critical role in resistance to predation by Bdellovibrio bacteriovorus

Bdellovibrio bacteriovorus, a Gram-negative predatory bacterium belonging to the Bdellovibrio and like organisms (BALOs), predates on Gram-negative bacteria. BALO strains differ in prey range but so far, the genetic basis of resistance against BALO predation is hardly understood. We developed a loss-of-function approach to screen for sensitive mutants in a library of strain M6, a predation-resistant strain of the plant pathogen Acidovorax citrulli. The screen is based on tracking the growth of a B. bacteriovorus strain expressing the fluorescent reporter Tdtomato in mutant pools to reveal predation-sensitive variants. Two independent loci were identified in mutant strains exhibiting significant levels of susceptibility to the predator. Genes in the two loci were analysed using both protein sequence homology and protein structure modeling. Both were secretion-related proteins and thus associated to the bacterial cell wall. Successful complementation of gspK, a gene encoding for a minor pseudopilin protein confirmed the involvement of the type II secretion system in A. citrulli M6 resistance. This proof of concept study shows that our approach can identify key elements of the BALO-prey interaction, and it validates the hypothesis that mutational changes in a single gene can drastically impact prey resistance to BALO predation.

Bacterial chromosome segregation: New insights into non-binary replication and division

Bdellovibrio bacteriovorus divides in a non-binary manner resulting in an even or odd number of progeny. A new study tracks the spatiotemporal dynamics of chromosome segregation in this species and shows that the process is dependent on the conserved ParA-ParB-parS system.

Bdellovibrio bacteriovorus phosphoglucose isomerase structures reveal novel rigidity in the active site of a selected subset of enzymes upon substrate binding.

Glycolysis and gluconeogenesis are central pathways of metabolism across all domains of life. A prominent enzyme in these pathways is phosphoglucose isomerase (PGI), which mediates the interconversion of glucose-6-phosphate and fructose-6-phosphate. The predatory bacterium Bdellovibrio bacteriovorus leads a complex life cycle, switching between intraperiplasmic replicative and extracellular 'hunter' attack-phase stages. Passage through this complex life cycle involves different metabolic states. Here we present the unliganded and substrate-bound structures of the B. bacteriovorus PGI, solved to 1.74 Å and 1.67 Å, respectively. These structures reveal that an induced-fit conformational change within the active site is not a prerequisite for the binding of substrates in some PGIs. Crucially, we suggest a phenylalanine residue, conserved across most PGI enzymes but substituted for glycine in B. bacteriovorus and other select organisms, is central to the induced-fit mode of substrate recognition for PGIs. This enzyme also represents the smallest conventional PGI characterized to date and probably represents the minimal requirements for a functional PGI.

Chromosome choreography during the non-binary cell cycle of a predatory bacterium

SummaryIn bacteria, the dynamics of chromosome replication and segregation are tightly coordinated with cell-cycle progression and largely rely on specific spatiotemporal arrangement of the chromosome. Whereas these key processes are mostly investigated in species that divide by binary fission, they remain mysterious in bacteria producing larger number of descendants. Here, we establish the predatory bacterium Bdellovibrio bacteriovorus as a model to investigate the non-binary processing of a circular chromosome. We found that its single chromosome is highly compacted in a polarized nucleoid that excludes freely diffusing proteins during the non-proliferative stage of the cell cycle. A binary-like cycle of DNA replication and asymmetric segregation is followed by multiple asynchronous rounds of replication and progressive ParABS-dependent partitioning, uncoupled from cell division. Finally, we provide the first evidence for an on-off behavior of the ParB protein, which localizes at the centromere in a cell-cycle-regulated manner. Altogether, our findings support a model of complex chromosome choreography leading to the generation of variable, odd, or even numbers of offspring and highlight the adaptation of conserved mechanisms to achieve non-binary reproduction.

Clearance of Gram-Negative Bacterial Pathogens from the Ocular Surface by Predatory Bacteria

It was previously demonstrated that predatory bacteria are able to efficiently eliminate Gram-negative pathogens including antibiotic-resistant and biofilm-associated bacteria. In this proof-of-concept study we evaluated whether two species of predatory bacteria, Bdellovibrio bacteriovorus and Micavibrio aeruginosavorus, were able to alter the survival of Gram-negative pathogens on the ocular surface. Clinical keratitis isolates of Pseudomonas aeruginosa (strain PAC) and Serratia marcescens (strain K904) were applied to the ocular surface of NZW rabbits followed by application of predatory bacteria. At time intervals, surviving pathogenic bacteria were enumerated. In addition, B. bacteriovorus and S. marcescens were applied to porcine organ culture corneas under contact lenses, and the ocular surface was examined by scanning electron microscopy. The ocular surface epithelial layer of porcine corneas exposed to S. marcescens, but not B. bacteriovorus was damaged. Using this model, neither pathogen could survive on the rabbit ocular surface for longer than 24 h. M. aeruginosavorus correlated with a more rapid clearance of P. aeruginosa but not S. marcescens from rabbit eyes. This study supports previous evidence that predatory bacteria are well tolerated by the cornea, but suggest that predatory bacteria do not considerably change the ability of the ocular surface to clear the tested Gram-negative bacterial pathogens from the ocular surface.

Modeling the Life Cycle of the Intramitochondrial Bacterium "Candidatus Midichloria mitochondrii" Using Electron Microscopy Data.

ABSTRACT“Candidatus Midichloria mitochondrii” is a Gram-negative bacterium that lives in strict intracellular symbiosis with the hard tick Ixodes ricinus, forming one of the most intriguing endosymbiosis described to date. The bacterium is capable of durably colonizing the host mitochondria, a peculiar tropism that makes “Ca. Midichloria mitochondrii” a very interesting tool to study the physiology of these cellular organelles. The interaction between the symbiont and the organelle has, however, been difficult to characterize. A parallelism with the predatory bacterium Bdellovibrio bacteriovorus has been drawn, suggesting the hypothesis that “Ca. Midichloria mitochondrii” could prey on mitochondria and consume them to multiply. We studied the life cycle of the bacterium within the host oocytes using a multidisciplinary approach, including electron microscopy, molecular biology, statistics, and systems biology. Our results were not coherent with a predatory-like behavior by “Ca. Midichloria mitochondrii” leading us to propose a novel hypothesis for its life cycle. Based on our results, we here present a novel model called the “mitochondrion-to-mitochondrion hypothesis.” Under this model, the bacterium would be able to move from mitochondrion to mitochondrion, possibly within a mitochondrial network. We show that this model presents a good fit with quantitative electron microscopy data.

Active Biopolymeric Films Inoculated with Bdellovibrio bacteriovorus, a Predatory Bacterium

The objective of the present work was to evaluate novel active films made with biopolymeric matrices as carriers of a living Bdellovibrio bacteriovorus HD100 strain, a predatory bacterium with antimicrobial potentials against pathogens. Biopolymer films were prepared by a casting method using the following mixtures: collagen/sodium alginate/sorbitol (CA-S), collagen/sodium alginate/glycerol (CA-G), and tapioca starch/sodium alginate/glycerol (StA-G). The effects of the film formulations on the viability of the B. bacteriovorus was investigated by using Fourier Transform Infrared (FTIR) spectroscopy, Differential Scanning Calorimetry (DSC), and Scanning Electron Microscopy (SEM). SEM showed that Bdellovibrio bacteriovorus morphology was not altered in the polymeric films. FTIR spectroscopy provided information about the structural composition of the films. CA-S showed less reduction in the viability of B. bacteriovorus after its entrapment; thus, CA-S proved to be a better agent for the immobilization and preservation of B. bacteriovorus to enhance its predatory activities during application against Escherichia coli.