Antimicrobial Agents and Chemotherapy–June’11, Part 1

The content of the June issue of Antimicrobial Agents and Chemotherapy is now available. Its the 6th issue of volume 55. There are quite some research articles published in relation to antimicrobials against biofilms.

Antimicrobial Peptoids Are Effective against Pseudomonas aeruginosa Biofilms (doi:10.1128/AAC.01516-10)

Bacteria in biofilms are known to be more resistant towards antimicrobials than their planktonic (free living) counterparts.

A group of scientists from Stanford University, California and Northwestern University, Evanston have investigated the effects of peptoids, Antimicrobial peptides (AMPs) and conventional antibiotics against the formation and removal of Pseudomonas aeruginosa biofilms. AMPs are short aminoacid sequences are known for their nonspecific mechanism of bactericidal activity.
It is hypothesized that, because they act non specifically, the bacteria might not be able to develop resistance towards them. But these natural peptides are susceptible to many proteases in the cell and in most cases, their bioavailability is low. Peptoids in the other hand are synthetically prepared peptides which also has antimicrobial property, in short they are non-natural mimics of AMPs.

In this study, the authors have demonstrated the ability of peptoids to reduce the viability of P. aeruginosa biofilms at their minimum inhibitory concentrations (MIC). Whereas the antibiotics showed comparable results only at 8 to 30 times higher concentrations than their MICs. Therefore, the authors propose the peptoids as a novel class of antibiotics to overcome antibacterial resistance in biofilms that makes the conventional antibiotics less effective. 

Dynamics of the Action of Biocides in Pseudomonas aeruginosa Biofilms (doi:10.1128/AAC.01760-10)

Another group of scientists from France (INRA, AgroParisTech and STERIS S.A.) and Switzerland (University Hospital of Lausanne) jointly published their results from their investigation of the biocidal activity of Peracetric acid (PAA) and benzalkonium chloride (BAC) against P. aeruginosa biofilms. PAA is an oxidizing agent and BAC is a quaternary ammonium compound.

Biofilms are generally recognized as heterogeneous structures, having subpopulations within – with different physiological states and resistance phenotypes. The group has exploited the time-lapse confocal laser scanning microscopy (CLSM) to study the biocidal activity of grown P. aeruginosa biofilms loosing  the intracellular fluorescence triggered by the cleavage of fluorogenic esterase substrate,  when the cell membrane of the bacteria in the biofilm is compromised due to the biocide activity.

This is not a very new technique, the authors quote, but P.aeruginosa particularly is not a good candidate for calcein-AM (fluorescent labeling molecule) protocol as reported earlier, due to their intense efflux pump activity which does not allow the fluorescent residues to be retained in the cells. For this reason, the authors used Chemchrome V6 marker/Chemsol B16 staining buffer kit which can block the efflux pump activity and so the intracellular fluorescence is maintained until there is a membrane damage caused by the agents under study.  

As expected, the results showed that the biofilms are more resistant to the biocide treatments than the planktonic cells. The biofilms required 15-20 and 100 fold higher concentrations for PAA and BAC respectively. Interestingly, there was a non-uniform pattern of bactericidal activity observed for the BAC treatment. The cells in the exterior were immediately susceptible, while the cells in the interior of the biofilms were active for a significant amount of time before their fluorescence is reduced. The above observation suggests that there is a transport-limitation of BAC to the interior of the biofilm. Moreover, there were still some cells which retained the fluorescence even at the end of the experimental time, 25 minutes. This pattern was not observed in the case of PAA.

The authors have also demonstrated that the cells when removed from the biofilm, devoid of polysaccharides and proteins (from the biofilms) were equally susceptible as their planktonic counter parts which suggests that the resistance or the transport-boundary is caused manly by the biofilm components like proteins and carbohydrates. The measure of the biofilm components also support the above. Needless to say, the observation of the intact cells still fluorescing until the end of the experimental time span can be explained by the local physiological status of the cells/subpopulations which might express highly resistant phenotypes.

Quorum Sensing Inhibitors Increase the Susceptibility of Bacterial Biofilms to Antibiotics In Vitro and In Vivo (doi:10.1128/AAC.00045-11)

Another interesting study was reported by group of Belgium scientists from Ghent university and University of Antwerp on the use of quorum sensing inhibitors to increase the susceptibility of biofilms to antibiotics.

Quorum sensing (QS), the chemical communication system for the bacteria is an interesting filed of research. QS is an important part of biofilm formation and it has been shown that blocking the QS signals could stop the bacteria from forming the biofilm. I have a post written about how bacteria talk using the QS system here. The authors of this study have evaluated several QS inhibitors on the susceptibility of bacterial biofilms to antibiotics both in vitro (in cells in test tubes) and in vivo (in animal models).

The authors selected Tobramycin, vancomycin and Clindamycin as the test antibiotics, together with baicalin hydrate, cinnamaldehyde and hamamelitannin as the QS inhibitors. Combinations of the QS inhibitors and antibiotics were tested against biofilms formed by different strains each of Burkholderia cenocepacia, B. multivorans, P. aeruginosa and Staphylococcus aureus. For the in vivo studies, they examined the infections in Galleria mollonella larvae, Baenorhabditis elegans and BALB/c mice.

Together with the MIC and animal survival assays the authors have demonstrated that by combining QS inhibitors with conventional antibiotics, the treatment efficiency can be improved against biofilm related infections.

Some more continued in Part-2