Stationary phase Staphylococcus aureus, especially methicillin-resistant S. aureus (MRSA), has been widely associated with many persistent infections as well as biofilm-associated infections, which are challenging due to their increasing antibiotic resistance. α-Melanocyte stimulating hormone (α-MSH) is an antimicrobial peptide (AMP) with well-established potent activity against S. aureus, but little is known about its antimicrobial efficacy against the stationary phase of the bacteria. We investigated the in vitro activities of two palmitoylated analogues, Pal-α-MSH(6-13) and Pal-α-MSH(11-13), of the C-terminal fragments of α-MSH against biofilm-producing strains of methicillin-sensitive S. aureus (MSSA) and MRSA. While both the peptides demonstrated antistaphylococcal efficacy, Pal-α-MSH(11-13) emerged as the most effective AMP as palmitoylation led to a remarkable enhancement in its activity against stationary phase bacteria. Similar to α-MSH, both the designed analogues were membrane-active and exhibited improved bacterial membrane depolarization and permeabilization, as further confirmed via electron microscopy studies. Of the two peptides, Pal-α-MSH(11-13) was able to retain its activity in the presence of standard microbiological media, which otherwise is a major limiting factor toward the therapeutic use of α-MSH-based peptides. More importantly, Pal-α-MSH(11-13) was also highly effective in inhibiting the formation of biofilms. Furthermore, it did not lead to resistance development in MRSA cells even upon 18 serial passages at sub-MIC concentrations. These observations support the potential use of in the treatment of planktonic as well as sessile S. aureus infections.

Methicillin-resistant Staphylococcus aureus (MRSA), a biofilm-forming recalcitrant pathogen with a multidrug-resistant profile, poses a pandemic threat to human health and is the leading cause of severe infections in both healthcare and community settings. In this study, toward designing novel α-MSH-based peptides with enhanced activity and stability against MRSA, particularly its stationary phase and biofilm, we explored a design approach to augment the hydrophobicity of an 8-mer C-terminal α-MSH(6-13)-based peptide Ana-5 through the incorporation of a bulky unnatural amino acid. The designed Ana-peptides overcame the limitation of diminished activity in biological media and exhibited enhanced antistaphylococcal activity and cell selectivity. With membrane rupture as the primary mode of action, the peptides exhibited inhibitory potential against S. aureus biofilms. Importantly, the peptides did not exhibit any adverse effects in the in vivo toxicity studies and were also able to significantly alleviate bacterial infection in a systemic infection mice model study. Additionally, the peptides retained their activity in the presence of serum and displayed a low propensity toward resistance development in MRSA cells. Moreover, the observed synergistic potential of Ana-10 with conventional antibiotics could be vital in resurrecting discarded antibiotics. Thus, this study provides us with an exciting lead, Ana-10, for further development against biofilm-based chronic S. aureus infections.

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Aim: To analyse the microbial profile of canine persistent wound infections. Materials and Methods:The total wound samples (n=172) taken from both traumatic (140) and post-surgical (32) persistent wounds in canines were processed for routine microbial isolation and identification during a period of 15 months.Results: Staphylococcus intermedius was found to be the predominant isolate from all types of wounds under study. It was followed by Staphylococcus aureus, Pseudomonas aeruginosa, E. coli, Pasteurella spp., Corynaebacterium spp. and Bacillus spp. From different traumatic wounds of dogs, S. intermedius (92/140=65.7%) and from surgical wounds, P. aeruginosa (24/32=75%) were found to be the predominant isolates recovered whereas the most commonly isolated bacterial genus in both traumatic and surgical wounds of dogs was Staphylococcus spp. Conclusion:Canine wounds are polymicrobial in nature. Hence proper microbial laboratory diagnosis and presence of multiple organisms in a wound are to be taken into consideration for effective treatment of persistent wound infections in dogs.

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