Date Published: July 21, 2015
Publisher: Public Library of Science
Author(s): Ryan J. Blower, Stephanie M. Barksdale, Monique L. van Hoek, Robert A Harrison. http://doi.org/10.1371/journal.pntd.0003862
Abstract: Burkholderia thailandensis is a Gram-negative soil bacterium used as a model organism for B. pseudomallei, the causative agent of melioidosis and an organism classified category B priority pathogen and a Tier 1 select agent for its potential use as a biological weapon. Burkholderia species are reportedly “highly resistant” to antimicrobial agents, including cyclic peptide antibiotics, due to multiple resistance systems, a hypothesis we decided to test using antimicrobial (host defense) peptides. In this study, a number of cationic antimicrobial peptides (CAMPs) were tested in vitro against B. thailandensis for both antimicrobial activity and inhibition of biofilm formation. Here, we report that the Chinese cobra (Naja atra) cathelicidin NA-CATH was significantly antimicrobial against B. thailandensis. Additional cathelicidins, including the human cathelicidin LL-37, a sheep cathelicidin SMAP-29, and some smaller ATRA peptide derivatives of NA-CATH were also effective. The D-enantiomer of one small peptide (ATRA-1A) was found to be antimicrobial as well, with EC50 in the range of the L-enantiomer. Our results also demonstrate that human alpha-defensins (HNP-1 & -2) and a short beta-defensin-derived peptide (Peptide 4 of hBD-3) were not bactericidal against B. thailandensis. We also found that the cathelicidin peptides, including LL-37, NA-CATH, and SMAP-29, possessed significant ability to prevent biofilm formation of B. thailandensis. Additionally, we show that LL-37 and its D-enantiomer D-LL-37 can disperse pre-formed biofilms. These results demonstrate that although B. thailandensis is highly resistant to many antibiotics, cyclic peptide antibiotics such as polymyxin B, and defensing peptides, some antimicrobial peptides including the elapid snake cathelicidin NA-CATH exert significant antimicrobial and antibiofilm activity towards B. thailandensis.
Partial Text: Burkholderia pseudomallei is a Gram-negative soil bacterium which acts as a facultative intracellular pathogen that can infect both humans and animals, causing melioidosis. Melioidosis is endemic to Southeast Asia and Northern Australia, where the mortality rates are 50% and 19% respectively [1–3]. In addition, B. pseudomallei is of interest because it is considered a class B priority pathogen and a Tier 1 Select Agent and has potential for aerosol delivery. In this study, Burkholderia thailandensis is used as a model for B. pseudomallei . B. thailandensis is a BSL-2 organism closely related to B. pseudomallei with an LD50 in mice 1000-fold higher than that of B. pseudomallei, making it an easier and safer model organism with which to work . B. thailandensis has been successfully demonstrated to be a useful BSL-2 surrogate for B. pseudomallei [4,6–8] for both in vitro and in vivo experiments. Thus, B. thailandensis may be a good model in which to study the molecular actions of full length cathelicidins such as LL37 both as antibacterial and antibiofilm peptides against Burkholderia strains.
B. thailandensis and B. pseudomallei have a wide range of mechanisms for evading antibiotics and antimicrobial peptides. These mechanisms include, but are not limited to, having a more impermeable membrane, multi-drug-resistant efflux pumps, inactivation of host proteins, and modification of drug targets . In this study, we demonstrate that certain peptides can evade these mechanisms and exhibit antimicrobial activity against B. thailandensis, despite its reported extreme peptide resistance. We also demonstrate that D-amino acid peptides exhibit comparable antimicrobial activity . Finally, we describe the anti-biofilm activity of some of these peptides against B. thailandensis.