Isolation and identification of a bacteriocin with antibacterial and antibiofilm activity from Citrobacter freundii.

Multi- and pan-antibiotic-resistant bacteria area major health challenge in hospital settings. Furthermore,when susceptible bacteria establish surface-attached biofilm populations, they become recalcitrant to antimicrobial therapy. Therefore, there is a need for novel antimicrobials that are effective against multi-drug-resistant and surface-attached bacteria. A screen to identify prokaryote-derived antimicrobials from a panel of over 100 bacterial strains was performed. One compound isolated from Citrobacter freundii exhibited antimicrobial activity against a wide range of Gram-negative bacteria and was effective against biofilms. Random transposon mutagenesis was performed to find mutants unable to produce the antimicrobial compound.Transposons mapped to a bacteriocin gene located on a small plasmid capable of replication in Escherichia coli. The plasmid was sequenced and found to be highly similar to a previously described colicinogenic plasmid.Expression of the predicted bacteriocin immunity gene conferred bacteriocin immunity to E. coli. The predicted bacteriocin gene, colA-43864, expressed in E. coli was sufficient to generate anti-microbial activity, and purified recombinant ColA-43864 was highly effective in killing E. coli, Citrobacter species, and Klebsiella pneumoniae cells in a planktonic and biofilm state. This study suggests that bacteriocins can be an effective way to control surface-attached pathogenic bacteria.

Inhibition of predation by Bdellovibrio bacteriovorus and Micavibrio aeruginosavorus via host cell metabolic activity in the presence of carbohydrates.

Bdellovibrio bacteriovorus and Micavibrio aeruginosavorus are highly motile Gram-negative predatory bacteria with the potential of being used as biocontrol agents or living antibiotics. It was suggested previously that sugar-binding proteins play a role in M. aeruginosavorus and B. bacteriovorus host specificity and predator-prey interactions. The effect of carbohydrates on predation was reexamined in this study. It was demonstrated that the presence of carbohydrates could indeed block predation. However, further investigation demonstrated that inhibition of predation was due to medium acidification by the metabolic activity of the host and not to a blocking of a putative sugar-binding protein. The data presented here might be of value when storing, growing, and cultivating predatory bacteria, as well as when considering environmental conditions that might influence predation in the field.

The Use of Commercially Available Alpha-Amylase Compounds to Inhibit and Remove Staphylococcus aureus Biofilms.

Staphylococcus aureus, a versatile human pathogen, is commonly associated with medical device infections. Its capacity to establish and maintain these infections is thought to be related to its ability to form adherent biofilms. In this study, commercially available α-amylase compounds from various biological sources were evaluated for their ability to reduce and prevent biofilm formation of several S. aureus isolates. Our data demonstrates that α-amylase compounds can rapidly detach biofilms of S. aureus, as well as inhibit biofilm formation. Our data also demonstrates that α-amylase compounds have an ability to reduce and disassociate S. aureus cell-aggregates grown in liquid suspension. These findings suggest that commercially available α-amylase compounds could be used in the future to control S. aureus biofilm-related infections.

A new method for isolating host-independent variants of Bdellovibrio bacteriovorus using E. coli auxotrophs.

Bdellovibrios are Gram-negative bacteria that are characterized by predatory behavior. Although Bdellovibrios exhibit an obligatory parasitic life cycle, it is possible to isolate Bdellovibrio variants that no longer require host cells for their growth. In this study, a new method for isolating Bdellovibrio bacteriovorus host-independent (HI) variants was developed. Filtered B. bacteriovorus prey cells were cultured with E. coli diaminopimelic acid (DAP) auxotrophs as host cells. Thereafter, the lysate was plated on DAP minus media, allowing only HI colonies to develop. Using this method, we have isolated numerous HI variants and demonstrated that the emergence of HI variants may be occurring at a higher frequency than was previously suggested.