Characterising four Sarconesiopsis magellanica (Diptera: Calliphoridae) larval fat body-derived antimicrobial peptides.

BACKGROUND: The inappropriate use of antibiotics has led to the accelerated growth of resistance to antibiotics. The search for new therapeutic strategies (i.e., antimicrobial peptides-AMPs) has thus become a pressing need. OBJECTIVE: Characterising and evaluating Sarconesiopsis magellanica larval fat body-derived AMPs. METHODS: Fat body extracts were analysed by reversed-phase high-performance liquid chromatography (RP-HPLC); mass spectrometry was used for characterising the primary structure of the AMPs so found. ProtParam (Expasy) was used for analysing the AMPs' physico-chemical properties. Synthetic AMPs' antibacterial activity was evaluated. FINDINGS: Four new AMPs were obtained and called sarconesin III, IV, V and VI. Sarconesin III had an α-helix structure and sarconesins IV, V and VI had linear formations. Oligomer prediction highlighted peptide-peptide interactions, suggesting that sarconesins III, V and VI could form self-aggregations when in contact with the microbial membrane. AMPs synthesised from their native molecules' sequences had potent activity against Gram-positive bacteria and, to a lesser extent, against Gram-negative and drug-resistant bacteria. Sarconesin VI was the most efficient AMP. None of the four synthetic AMPs had a cytotoxic effect. MAIN CONCLUSIONS: S. magellanica larval fat body-derived antimicrobial peptides are an important source of AMPs and could be used in different antimicrobial therapies and overcoming bacterial resistance.

Rondonin: antimicrobial properties and mechanism of action.

Infectious diseases are among the major causes of death in the human population. A wide variety of organisms produce antimicrobial peptides (AMPs) as part of their first line of defense. A peptide from Acanthoscurria rondoniae plasma, rondonin-with antifungal activity, a molecular mass of 1236 Da and primary sequence IIIQYEGHKH-was previously studied (UniProt accession number B3EWP8). It showed identity with the C terminus of subunit 'D' of the hemocyanin of the Aphonopelma hentzi spider. This result led us to propose a new pathway of the immune system of arachnids that suggests a new function to hemocyanin: production of antimicrobial peptides. Rondonin does not interact with model membranes and was able to bind to yeast nucleic acids but not bacteria. It was not cytotoxic against mammalian cells. The antifungal activity of rondonin is pH-dependent and peaks at pH ˜ 4-5. The peptide presents synergism with gomesin (spider hemocyte antimicrobial peptide-UniProtKB-P82358) against human yeast pathogens, suggesting a new potential alternative treatment option. Antiviral activity was detected against RNA viruses, measles, H1N1, and encephalomyocarditis. This is the first report of an arthropod hemocyanin fragment with activity against human viruses. Currently, it is vital to invest in the search for natural and synthetic antimicrobial compounds that, above all, present alternative mechanisms of action to first-choice antimicrobials.

Characterising four Sarconesiopsis magellanica (Diptera: Calliphoridae) larval fat body-derived antimicrobial peptides

BACKGROUND The inappropriate use of antibiotics has led to the accelerated growth of resistance to antibiotics. The search for new therapeutic strategies (i.e., antimicrobial peptides-AMPs) has thus become a pressing need. OBJECTIVE Characterising and evaluating Sarconesiopsis magellanica larval fat body-derived AMPs. METHODS Fat body extracts were analysed by reversed-phase high-performance liquid chromatography (RP-HPLC); mass spectrometry was used for characterising the primary structure of the AMPs so found. ProtParam (Expasy) was used for analysing the AMPs' physico-chemical properties. Synthetic AMPs' antibacterial activity was evaluated. FINDINGS Four new AMPs were obtained and called sarconesin III, IV, V and VI. Sarconesin III had an α-helix structure and sarconesins IV, V and VI had linear formations. Oligomer prediction highlighted peptide-peptide interactions, suggesting that sarconesins III, V and VI could form self-aggregations when in contact with the microbial membrane. AMPs synthesised from their native molecules' sequences had potent activity against Gram-positive bacteria and, to a lesser extent, against Gram-negative and drug-resistant bacteria. Sarconesin VI was the most efficient AMP. None of the four synthetic AMPs had a cytotoxic effect. MAIN CONCLUSIONS S. magellanica larval fat body-derived antimicrobial peptides are an important source of AMPs and could be used in different antimicrobial therapies and overcoming bacterial resistance.

Effects of the Natural Peptide Crotamine from a South American Rattlesnake on Candida auris, an Emergent Multidrug Antifungal Resistant Human Pathogen.

Invasive Candida infections are an important growing medical concern and treatment options are limited to a few antifungal drug classes, with limited efficacies depending on the infecting organism. In this scenario, invasive infections caused by multiresistant Candida auris are emerging in several places around the world as important healthcare-associated infections. As antimicrobial peptides (AMPs) exert their activities primarily through mechanisms involving membrane disruption, they have a lower chance of inducing drug resistance than general chemical antimicrobials. Interestingly, we previously described the potent candicidal effect of a rattlesnake AMP, crotamine, against standard and treatment-resistant clinical isolates, with no hemolytic activity. We evaluated the antifungal susceptibility of several Candida spp. strains cultured from different patients by using the Clinical and Laboratory Standards Institute (CLSI) microdilution assay, and the antifungal activity of native crotamine was evaluated by a microbial growth inhibition microdilution assay. Although all Candida isolates evaluated here showed resistance to amphotericin B and fluconazole, crotamine (40-80 µM) exhibited in vitro activity against most isolates tested. We suggest that this native polypeptide from the South American rattlesnake Crotalus durissus terrificus has potential as a structural model for the generation of a new class of antimicrobial compounds with the power to fight against multiresistant Candida spp.

Juruin: an antifungal peptide from the venom of the Amazonian Pink Toe spider, Avicularia juruensis, which contains the inhibitory cystine knot motif.

The aim of this study was to screen the venom of the theraposid spider Avicularia juruensis for the identification of antimicrobial peptides (AMPs) which could be further used as prototypes for drug development. Eleven AMPs, named juruentoxins, with molecular weight ranging from 3.5 to 4.5 kDa, were identified by mass spectrometry after the soluble venom was separated by high performance liquid chromatography. Juruentoxins have a putative inhibitory cystine knot (ICK) motif, generally found in neurotoxins, which are also resistant to proteolysis. One juruentoxin that has 38 amino acid residues and three disulfide bonds were characterized, to which we proposed the name Juruin. Based on liquid growth inhibition assays, it has potent antifungal activity in the micromolar range. Importantly, Juruin lacks haemolytic activity on human erythrocytes at the antimicrobial concentrations. Based on the amino acid sequence, it is highly identical to the insecticidal peptides from the theraposid spiders Selenocosmia huwena, Chilobrachys jingzhao, and Haplopelma schmidti from China, indicating they belong to a group of conserved toxins which are likely to inhibit voltage-gated ion channels. Juruin is a cationic AMP, and Lys22 and Lys23 show maximum positive charge localization that might be important for receptor recognition. Although it shows marked sequence similarity to neurotoxic peptides, Juruin is a novel exciting molecule with potent antifungal activity, which could be used as a novel template for development of drugs against clinical resistant fungi strains.