Activity of tick antimicrobial peptide from Ixodes persulcatus (persulcatusin) against cell membranes of drug-resistant Staphylococcus aureus

Abstract

Persulcatusin (IP), which is an antimicrobial peptide found in Ixodes persulcatus midgut, is active against Gram-positive bacteria such as Staphylococcus aureus. Multidrug-resistant bacteria in particular methicillin-resistant S. aureus (MRSA), vancomycinintermediate S. aureus (VISA) and vancomycin-resistant S. aureus (VRSA) are a worldwide clinical concern. In the present study, to explore the potential of IP as a new agent against multidrug-resistant S. aureus infections, we evaluated the antimicrobial activity of IP against multidrug-resistant S. aureus strains by MIC 90 , morphological observation with scanning electron microscope (SEM), and the calcein leakage assay of membrane integrity. Among the six antimicrobial peptides used in this study, IP exhibited the lowest MIC 90 values for both vancomycin-susceptible and -resistant S. aureus strains. The IP MIC 90 against a VISA strain was equivalent to vancomycin, while the MIC 90 against VRSA was relatively low. SEM observations indicated that bacterial cells exposed to IP were crumpled and showed prominent structural changes. Moreover, IP influenced the cell membranes of both MRSA and VRSA in a mere 5 min, leading to leakage of the preloaded calcein. Although a VISA strain was resistant to the action of IP on cell membrane, the MIC 90 of IP was lower than that of Nisin, suggesting that IP had another bactericidal mechanism in addition to cell membrane attack. Our results indicate that the synthetic tick antimicrobial peptide, IP exhibits strong antibacterial activity against multidrug-resistant S. aureus strains, including VRSA, via both cell membrane attack and another unknown mechanism. IP represents a promising candidate for a new anti-VRSA therapy. The appearance of antibiotic-resistant bacteria has resulted in bacterial diseases re-emerging as a threat after the many decades since the introduction of the first antibiotic penicillin. 1 The development of new antibacterial agents is urgently required, and antimicrobial peptides are viewed as ideal candidate agents. 5 Antimicrobial peptides are integral components of the innate immune system of all living organisms, including mammals, plants and insects. 6 Antimicrobial peptides result in strong natural defense in arthropod, particularly antimicrobial peptides of silk moth 7,8 and beetle, 9 which belong to the defensin family. Persulcatusin (IP), a tick antimicrobial peptide found in the midgut of Ixodes persulcatus, exhibits antimicrobial activity against Gram-positive bacteria such as S. aureus. 10-12 Furthermore, we have reported previously that S. aureus strains could not be isolated from I. persulcatus during feeding. 13 This is attributable to the antimicrobial activity of IP, which is highly expressed during blood feeding. 10,11 We hypothesized that IP could exhibit antimicrobial activity against VISA and VRSA, as well as methicillin-susceptible S. aureus and MRSA.In this study, we evaluated the antimicrobial activity of IP against multidrug-resistant S. aureus strains using MIC, scanning electron m...

Mechanisms of Action for Antimicrobial Peptides With Antibacterial and Antibiofilm Functions

“…In this review, a number of examples that make use of biophysical methods to determine MOAs that involve DNA/RNA targets (Park et al, 1998;Hsu et al, 2005;Nam et al, 2014), protein targets (Mardirossian et al, 2014(Mardirossian et al, , 2018Krizsan et al, 2015), stringent response inhibition (de la Fuente-Núñez et al, 2014;Pletzer et al, 2017b), and degradation of biofilms (Segev-Zarko et al, 2015;Ansari et al, 2017) were presented. A number of examples of how these biophysical approaches can be applied to bacteria (i.e., termed "biological" assays here) (de la Fuente-Núñez et al, 2014;Brannan et al, 2015;Miyoshi et al, 2017;Overall et al, 2019) serve to illustrate not only how the lines between these methods may become increasingly blurred in the future, but also how all the methods presented in this review, as well as direct visualization approaches [e.g., scanning electron microscopy (SEM) (Chileveru et al, 2015)] and other new approaches [e.g., small angle X-ray scattering (Von Gundlach et al, 2016, von Gundlach et al, 2019], will continue to be relevant.…”

“…HDPinduced membrane damage leads to calcein leakage, and results in an increase in fluorescence (Peisajovich and Shai, 2003;Fillion et al, 2014). This assay can also be adapted for use in bacterial cells to test whether an HDP has an effect on bacterial cytoplasmic membrane integrity (Miyoshi et al, 2017). Finally, smaller fluorophores, such as the Ca 2+ -dependent fluorophore Quin-2 (Tatulian and Kandel, 2019), can be used to probe pores of a smaller diameter (<1 nm).…”

Antimicrobial and anti-biofilm activity of hexadentated macrocyclic complex of copper (II) derived from thiosemicarbazide against Staphylococcus aureus

“…ATCC 29213 cells were grown till the exponential phase and resuspended at a concentration of 10 8 CFU/mL 31 . Cells were incubated with Cu Complex (50 µg/mL) for 60 min at 37 °C and cells without drug were used as a control.…”

“…Cells were incubated with Cu Complex (50 µg/mL) for 60 min at 37 °C and cells without drug were used as a control. Further, the cells were pelleted and fixed with 2% glutaraldehyde solution for 2 h at 4 °C as described earlier 31 . The cells were dehydrated in a graded series of alcohols.…”

Insect Antimicrobial Peptides, a Mini Review

“…Its similarity with other known tick AMPs is from 71% to 88% [ 121 , 122 ]. The structural integrity of persulcatusin is maintained by three S-S bonds, which are energetically important for the stability and the formation of the structure of α-helix and β-sheet [ 123 , 124 ]. This peptide can inhibit the growth of methicillin-sensitive S. aureus (MSSA) and methicillin-resistant S. aureus (MRSA) with the MIC of 0.156–1.25 μg/mL and 0.625–2.5 μg/mL, respectively [ 31 ].…”