Use of Resazurin To Rapidly Enumerate Bdellovibrio and Like Organisms and Evaluate Their Activities

A method to rapidly quantify predatory bacterial cell populations using resazurin reduction to resorufin and its resulting fluorescence kinetics (dF/dt) are described. The reliability of this method to measure the predatory populations was demonstrated with the type strain, Bdellovibrio bacteriovorus HD100, as well as B. bacteriovorus 109J and two natural isolates, Halobacteriovorax strains JA-1 and JA-3, with clear correlation when densities were between 107 and 109 PFU/ml. Resazurin was also used to evaluate how B. bacteriovorus HD100 and Halobacteriovorax strain JA-1 respond to harmful conditions, i.e., exposure to sodium dodecyl sulfate (SDS), with both the dF/dt and PFU/ml indicating Halobacteriovorax strain JA-1 is more sensitive to this surfactant. Tests were also performed using media of different osmolalities, with the dF/dt values matching the 24-h predatory activities reasonably well. Finally, this method was successfully applied in near real-time analyses of predator-prey dynamics and, when coupled with SDS, was capable of differentiating between the predatory and prey populations. All of these tests serve to prove this method is (i) very rapid, needing only 15 min from start to finish; (ii) very reliable with different predatory bacterial species; and (iii) very versatile as it can be easily adapted to measure predatory numbers and activities in a range of experiments

Staphylococcus aureus Sensitivity to Membrane Disrupting Antibacterials Is Increased under Microgravity

In a survey of the International Space Station (ISS), the most common pathogenic bacterium identified in samples from the air, water and surfaces was Staphylococcus aureus. While growth under microgravity is known to cause physiological changes in microbial pathogens, including shifts in antibacterial sensitivity, its impact on S. aureus is not well understood. Using high-aspect ratio vessels (HARVs) to generate simulated microgravity (SMG) conditions in the lab, we found S. aureus lipid profiles are altered significantly, with a higher presence of branch-chained fatty acids (BCFAs) (14.8% to 35.4%) with a concomitant reduction (41.3% to 31.4%) in straight-chain fatty acids (SCFAs) under SMG. This shift significantly increased the sensitivity of this pathogen to daptomycin, a membrane-acting antibiotic, leading to 12.1-fold better killing under SMG. Comparative assays with two additional compounds, i.e., SDS and violacein, confirmed S. aureus is more susceptible to membrane-disrupting agents, with 0.04% SDS and 0.6 mg/L violacein resulting in 22.9- and 12.8-fold better killing in SMG than normal gravity, respectively. As humankind seeks to establish permanent colonies in space, these results demonstrate the increased potency of membrane-active antibacterials to control the presence and spread of S. aureus, and potentially other pathogens

Loss of the lipopolysaccharide (LPS) inner core increases the electrocompetence of Escherichia coli

Mutations that shorten the lipopolysaccharide (LPS) inEscherichia coliwere found to significantly increase the number of transformants after electroporation. The loss of the LPS outer core increased the number of transformants with plasmid pAmCyan (3.3 kb) from 5.0 x 10(5)colony-forming units (CFU)/mu g in the wild-typeE. coliBW25113 to 3.3 x 10(7)CFU/mu g in a Delta waaGbackground, a 66.2-fold increase in efficiency. Truncation of the inner core improved this even further, with the Delta waaFmutant exhibiting the best transformation efficiencies obtained, i.e., a 454.7-fold increase in the number of colonies over the wild-type strain. Similar results were obtained when a larger plasmid (pDA1; 11.3 kb) was used, with the Delta waaFmutant once more giving the best transformation rates, i.e., a 73.7-fold increase. Subsequent tests proved that the enhanced transformabilities of these mutants were not due to a better survival or their surface charge properties, nor from preferential binding of these strains to the plasmid. Using N-phenyl-1-naphthylamine (NPN), we confirmed that the outer membranes of these mutant strains were more permeable. We also found that they leaked more ATP (3.4- and 2.0-fold higher for the Delta waaFand Delta waaGmutants, respectively, than wild-typeE. coliBW25113), suggesting that the inner membrane stability is also reduced, helping to explain how the DNA enters these cells more easily

Compounds affecting predation by and viability of predatory bacteria

Bdellovibrio-and-like organisms (BALOs) are a small group of bacteria that actively predate on other Gram-negative bacterial species. Although viewed mostly in a positive light, such as their potential use as living antibiotics to reduce pathogenic strain populations, several studies have also highlighted the need to control their activities, such as in the production of biodiesel. Consequently, this mini-review discusses research being conducted to characterize compounds and environmental settings that influence predation rates and the mechanisms by which they accomplish this, with a heavy emphasis on studies published within the last decade. Key points center dot This review discusses bacterial predators and factors impacting their activities. center dot Emphasis is on recent articles, particularly those discussing prey metabolites. center dot The implications on possible applications of bacterial predators are discussed

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 & PLUSMN; 0.9 %, 97.8 +/-& nbsp; 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)

Diffusible Signaling Factor, a Quorum-Sensing Molecule, Interferes with and Is Toxic Towards Bdellovibrio bacteriovorus 109J

Bdellovibrio bacteriovorus 109J is a predatory bacterium which lives by predating on other Gram-negative bacteria to obtain the nutrients it needs for replication and survival. Here, we evaluated the effects two classes of bacterial signaling molecules (acyl homoserine lactones (AHLs) and diffusible signaling factor (DSF)) have on B. bacteriovorus 109J behavior and viability. While AHLs had a non-significant impact on predation rates, DSF considerably delayed predation and bdelloplast lysis. Subsequent experiments showed that 50 ??M DSF also reduced the motility of attack-phase B. bacteriovorus 109J cells by 50% (38.2????????14.9 vs. 17????????8.9 ??m/s). Transcriptomic analyses found that DSF caused genome-wide changes in B. bacteriovorus 109J gene expression patterns during both the attack and intraperiplasmic phases, including the significant downregulation of the flagellum assembly genes and numerous serine protease genes. While the former accounts for the reduced speeds observed, the latter was confirmed experimentally with 50 ??M DSF completely blocking protease secretion from attack-phase cells. Additional experiments found that 30% of the total cellular ATP was released into the supernatant when B. bacteriovorus 109J was exposed to 200 ??M DSF, implying that this QS molecule negatively impacts membrane integrity. This is a preview of subscription content, log in to check access

A 1.74.12 mm3 Fully Integrated pH Sensor for Implantable Applications using Differential Sensing and Drift-Compensation

This paper presents a 1.7 ?? 4.1 ?? 2 mm3 pH sensor that is a fully integrated, stand-alone and implantable system. Instead of a bulky cm size Ag/AgCl electrode, we use a mm-size integrated platinum electrode, and differential sensing using ISFET and REFET pair to compensate for unstable fluid potential. We also propose a drift compensation technique in which the leakage from the source and drain through the gate oxide is canceled, reducing drift > 100 ?? . ?? 2019 JSAP

Sensitivity of predatory bacteria to different surfactants and their application to check bacterial predation

We evaluated the toxicity of surfactants against different predatory bacteria. Tests with Bdellovibrio bacteriovorus HD100 and SDS, an anionic surfactant, showed the predator was very sensitive; 0.02% SDS completely killed the predatory population (7-log loss; < 10 PFU/ml remaining) both when free-swimming or within the bdelloplast, i.e., intraperiplasmic. Similar results were also observed with B. bacteriovorus 109J and Peredibacter starrii. In contrast, none of the prey (E. coli, Klebsiella pneumoniae, Acinetobacter baumannii, or Pseudomonas sp. DSM 50906) viabilities were negatively affected by SDS. Triton X-100, a nonionic surfactant, was slightly less toxic towards B. bacteriovorus HD100 (viability loss of only 4-log), while two cationic surfactants, i.e., benzalkonium chloride (BZC) and cetyltrimethylammonium bromide (CTAB), were toxic towards both the predator and prey. Based on the above findings, we tested the potential use of SDS as a means to control predation. Addition of 0.02% SDS immediately halted predation based upon the prey bioluminescence, which leveled off and remained steady. This was confirmed using the predator viabilities; no predators were found in any of the samples where SDS was added. Consequently, low concentrations of SDS can be used as a simple means to control B. bacteriovorus HD100 activities