Abstract
Larval therapy (LT) is an alternative treatment for healing chronic wounds; its action is based on debridement, the removal of bacteria, and stimulating granulation tissue. The most important mechanism when using LT for combating infection depends on larval excretions and secretions (ES). Larvae are protected against infection by a spectrum of antimicrobial peptides (AMPs); special interest in AMPs has also risen regarding understanding their role in wound healing since they degrade necrotic tissue and kill different bacteria during LT. Sarconesiopsis magellanica (Diptera: Calliphoridae) is a promising medically-important necrophagous fly. This article reports a small AMP being isolated from S. magellanica ES products for the first time; these products were obtained from third-instar larvae taken from a previously-established colony. ES were fractionated by RP-HPLC using C18 columns for the first analysis; the products were then lyophilised and their antimicrobial activity was characterized by incubation with different bacterial strains. These fractions’ primary sequences were determined by mass spectrometry and de novo sequencing; five AMPs were obtained, the Sarconesin fraction was characterized and antibacterial activity was tested in different concentrations with minimum inhibitory concentrations starting at 1.2 μM. Potent inhibitory activity was shown against Gram-negative (Escherichia coli D31, E. coli DH5α, Salmonella enterica ATCC 13314, Pseudomonas aeruginosa 27853) and Gram-positive (Staphylococcus aureus ATCC 29213, S. epidermidis ATCC 12228, Micrococcus luteus A270) bacteria. Sarconesin has a significant similarity with Rho-family GTPases which are important in organelle development, cytoskeletal dynamics, cell movement, and wound repair. The data reported here indicated that Sarconesin could be an alternative candidate for use in therapeutics against Gram-negative and Gram-positive bacterial infections. Our study describes one peptide responsible for antibacterial activity when LT is being used. The results shown here support carrying out further experiments aimed at validating S. magellanica AMPs as novel resources for combating antibacterial resistance.
Highlights
IntroductionLarval therapy (LT) involves applying sterile larvae (usually Diptera from the Calliphoridae family) to an infected nonhealing wound (Raposio et al, 2017); its action is based on four mechanisms: removing necrotic tissue (debridement), disinfecting microorganisms, including methicillin-resistant Staphylococcus aureus (MRSA) (Robinson, 1935; Mumcuoglu, 2001; Bexfield et al, 2004; Nigam et al, 2010), inhibiting and eradicating biofilms (van der Plas et al, 2008; Cazander et al, 2009; Gottrup and Jorgensen, 2011) and stimulating granulation tissue for enhancing healing (Church, 1996; Thomas A.M. et al, 1999; Sherman et al, 2000; Mumcuoglu, 2001; Wolff and Hansson, 2005; Spilsbury et al, 2008).It has been proposed that larvae release antimicrobial ingredients into wounds in response to infection; some of these ingredients are low molecular weight bacteriostatic compounds, such as p-hydroxybenzoic acid, p-hydroxyphenylacetic acid, dioxopiperazine proline (Huberman et al, 2007) and an enigmatic compound (C10H16N6O9) known as seraticin (Nigam et al, 2010)
The present work led to finding a new sequence from S. magellanica; its antibacterial activity was screened and its biochemical and structural properties were elucidated by sequence homology
One antimicrobial peptides (AMPs) responsible for the antibacterial activity previously reported in S. magellanica was found (Diaz-Roa et al, 2014)
Summary
Larval therapy (LT) involves applying sterile larvae (usually Diptera from the Calliphoridae family) to an infected nonhealing wound (Raposio et al, 2017); its action is based on four mechanisms: removing necrotic tissue (debridement), disinfecting microorganisms, including methicillin-resistant Staphylococcus aureus (MRSA) (Robinson, 1935; Mumcuoglu, 2001; Bexfield et al, 2004; Nigam et al, 2010), inhibiting and eradicating biofilms (van der Plas et al, 2008; Cazander et al, 2009; Gottrup and Jorgensen, 2011) and stimulating granulation tissue for enhancing healing (Church, 1996; Thomas A.M. et al, 1999; Sherman et al, 2000; Mumcuoglu, 2001; Wolff and Hansson, 2005; Spilsbury et al, 2008).It has been proposed that larvae release antimicrobial ingredients into wounds in response to infection; some of these ingredients are low molecular weight bacteriostatic compounds, such as p-hydroxybenzoic acid, p-hydroxyphenylacetic acid, dioxopiperazine proline (Huberman et al, 2007) and an enigmatic compound (C10H16N6O9) known as seraticin (Nigam et al, 2010). Other compounds are antimicrobial peptides (AMPs) originating from the immune system that, when applied into wounds, contribute to their healing (Thomas A.M. et al, 1999; Bexfield et al, 2004). Lucifensin is one of the well characterized AMPs; it is derived from Lucilia sericata larvae and has been found as a constituent of larval excretions and secretions (ES) (Cerovsky et al, 2010). This molecule was originally isolated from Lucilia sericata larval intestine, later being detected in the salivary glands, fat body and haemolymph. It has been shown that it is the larval immune system which induces the production of these substances in the fat body when activated in response to an infectious environment (Valachova et al, 2013) for its rapid release into the haemolymph
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