Getting to the Root of Bacterial Hairs: What is “s”?

An atomic force microscope (AFM) was used to measure the steric forces of lipopolysaccharides (LPS) on the biofilm-forming bacteria, Pseudomonas aeruginosa. It is well known that LPS play a vital role in biofilm formation. These forces were characterized with a modified version of the Alexander and de Gennes (AdG) model for polymers, which is a function of equilibrium brush length, L, probe radius, R, temperature, T, separation distance, D, and an indefinite density variable, s. This last parameter was originally distinguished by de Gennes as the root spacing or mesh spacing depending upon the type of polymer adhesion; however since then it has been commonly thought of as the root spacing. This study aims to clarify the ambiguity of this parameter as a first step in characterizing biofilm formation. Varying the temperature and pH at which the steric forces of the LPS are measured and then analyzing the produced force curves with Matlab, should allow us to measure s. The Matlab program has been written to crop large numbers of force curves in accordance with the Alexander and de Gennes polymer model objectively and quickly. If s is the root spacing it should remain constant regardless of the changing polymer lengths, on the other hand if it is the mesh spacing it will be proportional to the temperature and pH. Preliminary data suggest that the LPS vary with temperature and pH. The data also suggest that s represents the mesh spacing. Once s has been described, further studies can be done to determine how environmental changes influence L, and s and consequently biofilm formation

LL37-Derived Fragments Improve the Antibacterial Potential of Penicillin G and Ampicillin against Methicillin-Resistant <i>Staphylococcus aureus</i>

Methicillin-resistant Staphylococcus aureus (MRSA) infections are a severe threat to public health. Antimicrobial peptides (AMPs) are novel and potential antimicrobials with specific antibacterial mechanisms. Our aim was to study the potential of LL37, FK16, and FK13 to enhance the anti-MRSA activity of antibiotics in vitro, particularly penicillin G and ampicillin. Our results showed that FK16 and FK13 have more synergistic inhibitory effects to MRSA strains when combined with penicillin G and ampicillin. In addition, AMPs exhibited strong membrane permeabilizing properties, and membrane permeabilizing effects can provide a possible explanation for the improved antibacterial effects of antibiotics, since permeabilizing AMPs have the potential to increase the access of antibiotics. To further study the electrostatic interactions among cationic AMPs with negatively charged bacteria, we measured the zeta potentials of three MRSA strains and also neutralized three MRSA strains with the addition of cationic AMPs. Further, we demonstrated the connection between membrane permeabilization and zeta potential neutralization. Finally, we treated MRSA strains with AMPs and characterized the MICs of penicillin G and ampicillin. FK16 was the most promising AMP among the three AMPs, since exposure to FK16 decreased the MICs of both penicillin G and ampicillin for all MRSA strains and also demonstrated more synergistic combinations when combined with antibiotics. AMP exposure and subsequent membrane permeabilization provide a possible pathway to re-sensitize drug-resistant bacteria to traditional antibiotics. Re-sensitization may help preserve the effectiveness of traditional antibiotics, thus providing a potential new strategy for fighting MRSA infections

Atomic Force Microscopy Study of the Effect of Lipopolysaccharides and Extracellular Polymers on Adhesion of Pseudomonas aeruginosa▿

The roles of lipopolysaccharides (LPS) and extracellular polymers (ECP) on the adhesion of Pseudomonas aeruginosa PAO1 (expresses the A-band and B-band of O antigen) and AK1401 (expresses the A-band but not the B-band) to silicon were investigated with atomic force microscopy (AFM) and related to biopolymer physical properties. Measurement of macroscopic properties showed that strain AK1401 is more negatively charged and slightly more hydrophobic than strain PAO1 is. Microscopic AFM investigations of individual bacteria showed differences in how the biopolymers interacted with silicon. PAO1 showed larger decay lengths in AFM approach cycles, suggesting that the longer polymers on PAO1 caused greater steric repulsion with the AFM tip. For both bacterial strains, the long-range interactions we observed (hundreds of nanometers) were inconsistent with the small sizes of LPS, suggesting that they were also influenced by ECP, especially polysaccharides. The AFM retraction profiles provide information on the adhesion strength of the biopolymers to silicon (Fadh). For AK1401, the adhesion forces were only slightly lower (Fadh = 0.51 nN compared to 0.56 nN for PAO1), but the adhesion events were concentrated over shorter distances. More than 90% of adhesion events for AK1401 were at distances of <600 nm, while >50% of adhesion events for PAO1 were at distances of >600 nm. The sizes of the observed molecules suggest that the adhesion of P. aeruginosa to silicon was controlled by ECP, in addition to LPS. Steric and electrostatic forces each contributed to the interfacial interactions between P. aeruginosa and the silicon surface