Lysin LysMK34 ofAcinetobacter baumanniiBacteriophage PMK34 Has a Turgor Pressure-Dependent Intrinsic Antibacterial Activity and Reverts Colistin Resistance

Abstract

The prevalence of extensively and pandrug-resistant Acinetobacter baumannii leaves little or no therapeutic options for treatment for this bacterial pathogen. Bacteriophages and their lysins represent attractive alternative antibacterial strategies in this regard. We used the extensively drug-resistant A. baumannii strain MK34 to isolate the bacteriophage PMK34 (vB_AbaP_PMK34). This phage shows fast adsorption and lacks virulence genes, nonetheless its narrow host spectrum based on capsule recognition limits broad application. PMK34 is a Fri1virus member of the Autographiviridae and has a 41.8 kb genome (50 ORFs), encoding an endolysin (LysMK34) with potent muralytic activity (1499.9 ± 131 U/μM), a typical mesophilic thermal stability up to 55 °C and a broad pH activity range (4-10). LysMK34 has an intrinsic antibacterial activity up to 4.8 and 2.4 log units for A. baumannii and Pseudomonas aeruginosa strains, respectively, but only when a high turgor pressure is present. Addition of 0.5 mM EDTA or application of an osmotic shock after treatment can compensate for the lack of a high turgor pressure. The combination of LysMK34 and colistin results in up to 32-fold reduction of the MIC of colistin, and colistin-resistant strains are re-sensitized in both Mueller-Hinton broth and 50 % human serum. As such, LysMK34 may be used to safeguard the applicability of colistin as a last-resort antibiotic. Importance A. baumannii is one of the most challenging pathogens for which development of new and effective antimicrobials is urgently needed. Colistin is a last resort antibiotic and even colistin-resistant A. baumannii strains exist. Here, we present a lysin that sensitizes A. baumannii for colistin and can revert colistin resistance to colistin susceptibility. The lysin also shows a strong, turgor pressure dependent intrinsic antibacterial activity, providing new insights in the mode-of-action of lysins with intrinsic activity against Gram-negative bacteria.

Discovering the Potentials of Four Phage Endolysins to Combat Gram-Negative Infections

“…In the case of native, unmodified enzymes, their combination with antibiotics seems to be much more promising. For example, the synergistic effects of endolysins and antibiotics (colistin) in vitro and in vivo therapies were shown ( Schirmeier et al, 2018 ; Abdelkader et al, 2020 ; Blasco et al, 2020 ). If developers succeed in defining an appropriate dosing schedule (beyond a single dose) as a basis for successful clinical studies for patients with confirmed infection due to drug-resistant pathogens or who experience recurrent or relapse infections, endolysins may provide an adjunctive therapy option.…”

LysSAP26, a New Recombinant Phage Endolysin with a Broad Spectrum Antibacterial Activity

“…This is similar to the mechanism of action of LysSS lysin, while the lysozyme-like Ablysin perforates the cell wall, followed by total cell shrinkage [ 4 , 10 ]. Abdelkader et al reported recently that LysMK34 lysin has turgor pressure-dependent antibacterial activity against A. baumannii [ 23 ]. We hypothesized that LysSAP26 would penetrate into the outer membrane and degrade peptidoglycan partly, and the damaged bacteria eventually would be lysed by force of turgor pressure.…”

Lysins breaking down the walls of Gram-negative bacteria, no longer a no-go

“…Whereas some studies report moderate antibacterial activities (<1-3 log reductions), other studies report a high bactericidal in vitro effect (>5 log reductions) [10,18,19 ]. However, assessing and comparing the potential of these lysins is complicated by the lack of standardized protocols and the variety of conditions used.…”

“…For example, dilute suspension buffers result in cells with a high intracellular turgor pressure, which are more prone to osmotic lysis by such lysins than when the turgor pressure is reduced in more physiological buffers. Also the diluent used for plating may cause osmotic effects and further affects the observed intrinsic antibacterial activity [19 ]. In addition, some studies make use of Tris as a buffer component in antibacterial assays.…”