In Vitro Activities of Nisin and Nisin Derivatives Alone and In Combination with Antibiotics against Staphylococcus Biofilms.

Des Field,R Paul Ross,Paul D Cotter,Colin Hill,Rory O’ Connor

doi:10.3389/fmicb.2016.00508

Abstract

The development and spread of pathogenic bacteria that are resistant to the existing catalog of antibiotics is a major public health threat. Biofilms are complex, sessile communities of bacteria embedded in an organic polymer matrix which serve to further enhance antimicrobial resistance. Consequently, novel compounds and innovative methods are urgently required to arrest the proliferation of drug-resistant infections in both nosocomial and community environments. Accordingly, it has been suggested that antimicrobial peptides could be used as novel natural inhibitors that can be used in formulations with synergistically acting antibiotics. Nisin is a member of the lantibiotic family of antimicrobial peptides that exhibit potent antibacterial activity against many Gram-positive bacteria. Recently we have used bioengineering strategies to enhance the activity of nisin against several high profile targets, including multi-drug resistant clinical pathogens such as methicillin-resistant Staphylococcus aureus, vancomycin-resistant enterococci, staphylococci, and streptococci associated with bovine mastitis. We have also identified nisin derivatives with an enhanced ability to impair biofilm formation and to reduce the density of established biofilms of methicillin resistant S. pseudintermedius. The present study was aimed at evaluating the potential of nisin and nisin derivatives to increase the efficacy of conventional antibiotics and to assess the possibility of killing and/or eradicating biofilm-associated cells of a variety of staphylococcal targets. Growth curve-based comparisons established that combinations of derivatives nisin V + penicillin or nisin I4V + chloramphenicol had an enhanced inhibitory effect against S. aureus SA113 and S. pseudintermedius DSM21284, respectively, compared to the equivalent nisin A + antibiotic combinations or when each antimicrobial was administered alone. Furthermore, the metabolic activity of established biofilms treated with nisin V + chloramphenicol and nisin I4V + chloramphenicol combinations revealed a significant decrease in S. aureus SA113 and S. pseudintermedius DSM21284 biofilm viability, respectively, compared to the nisin A + antibiotic combinations as determined by the rapid colorimetric XTT assay. The results indicate that the activities of the nisin derivative and antibiotic combinations represent a significant improvement over that of the wild-type nisin and antibiotic combination and merit further investigation with a view to their use as anti-biofilm agents.

Highlights

Staphylococcus aureus and S. pseudintermedius are major human and/or animal pathogens
The dramatic rise in antibiotic-resistance has stimulated renewed efforts to identify, develop or redesign existing antimicrobial agents active against these multi-resistant bacteria. Due to their many unique properties, the lantibiotics have become the focus of many biomedical and pharmaceutical research groups due to their demonstrable high potency in vitro, multiple modes of action and ability to destroy target cells rapidly (Cotter et al, 2005a; Cavera et al, 2015a). Because lantibiotics such as nisin are produced as gene-encoded pre-peptides, they are infinitely more suited than classical antibiotics to bioengineering which could lead to the generation of a new source of potent antimicrobials
We set out to examine for the first time, the ability of nisin and two enhanced bioengineered nisin derivatives in conjunction with a selection of conventional antibiotics to control a range of Staphylococcus sp, including human and veterinary pathogens, with the ultimate aim of identifying superior anti-biofilm combinations

Summary

Introduction

Staphylococcus aureus and S. pseudintermedius are major human and/or animal pathogens. The development of anti-biofilm therapeutics has generally focused on interfering with quorum sensing, inhibition of adhesion, enhancement of dispersion, or bacteriophage-based treatments (Mataraci and Dosler, 2012) Another potential strategy to reduce biofilm-associated resistance is through synergistic effects of antimicrobial agents in combination, which can enhance antibiofilm activities and help to prevent or delay the emergence of resistance. Nisin is used as a food preservative in over 50 countries and has been approved in the EU and by the US Food and Drug Administration (FDA; Delves-Broughton et al, 1996) Both nisin A (and its natural variant nisin Z) are effective against the Gram-positive pathogens responsible for bovine mastitis and have been incorporated into a number of products devoted to restricting or treating such infections (Sears et al, 1992; Wu et al, 2007). The bioengineering of nisin has generated variants that exhibit improved antimicrobial activity against strains of clinical relevance (methicillin resistant S. aureus (MRSA), vancomycinresistant enterococci (VRE), vancomycin-intermediate S. aureus (VISA), methicillin-resistant S. pseudintermedius (MRSP), and C. difficile) but has brought about the widening of its antimicrobial spectrum (Field et al, 2008, 2012; Molloy et al, 2013)

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Conclusion