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In vitro activity of Protegrin-1, alone and in combination with clinically useful antibiotics, against Acinetobacter baumannii strains isolated from surgical wounds

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Abstract

In the past few years the increasing incidence of hospital infections with Acinetobacter baumannii, especially in immunocompromised patients, and its proneness to develop multidrug resistance have been raising considerable concern. This study examines the antimicrobial and antibiofilm activity of protegrin 1 (PG-1), an antimicrobial peptide from porcine leukocytes, against A. baumannii strains isolated from surgical wounds. PG-1 was tested both alone and combined with the antibiotics commonly used in clinical settings. Its antimicrobial activity was evaluated by determination of minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC), checkerboard assays, and time-kill experiments. Its effects on biofilm inhibition/eradication were tested with crystal violet staining. The strains were grown in subinhibitory or increasing PG-1 concentrations to test the development of resistance. Mammalian cell toxicity was tested by XTT assays. PG-1 MICs and MBCs ranged from 2 to 8 µg/ml. PG-1 was most active and demonstrated a synergistic interaction with colistin, a last resort antibiotic. Interestingly, antagonism was never observed. In time-kill experiments, incubation with 2 × MIC for 30 min suppressed all viable cells. PG-1 did not select resistant strains and showed a limited effect on cell viability, but it did exert a strong activity against multidrug-resistant A. baumannii. In contrast, in our experimental conditions it had no effect on biofilm inhibition/eradication. PG-1 thus seems to be a promising antimicrobial agent against multidrug-resistant Gram-negative infections.

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Acknowledgements

We are grateful to Prof. Gian Maria Rossolini and Dr. Alberto Antonelli for providing the colistin-resistant A. baumannii strains.

Funding

This work was supported by “Progetto Strategico di Ateneo 2016 – UNIVPM: Study of new compounds and innovative strategies to control complicated bacterial skin infections”.

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Author notes

  1. Gianluca Morroni and Oriana Simonetti have contributed equally to this work.

Authors and Affiliations

  1. Infectious Diseases Clinic, Department of Biomedical Sciences and Public Health, Polytechnic University of Marche, Ancona, Italy

    Gianluca Morroni, Lucia Brescini, Andrea Giacometti & Oscar Cirioni

  2. Dermatological Clinic, Polytechnic University of Marche, Ancona, Italy

    Oriana Simonetti & Annamaria Offidani

  3. Microbiology Unit, Department of Biomedical Sciences and Public Health, Polytechnic University of Marche, Ancona, Italy

    Andrea Brenciani

  4. Faculty of Pharmacy, Medical University of Gdansk, Gdańsk, Poland

    Wojciech Kamysz & Damian Neubauer

  5. Faculty of Chemistry, University of Gdansk, Gdańsk, Poland

    Elzbieta Kamysz

  6. Histology, Department of Clinical and Molecular Sciences, Polytechnic University of Marche, Ancona, Italy

    Miriam Caffarini & Monia Orciani

  7. Microbiology Unit, Department of Life and Environmental Sciences, Polytechnic University of Marche, Ancona, Italy

    Eleonora Giovanetti

Authors
  1. Gianluca Morroni
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Corresponding author

Correspondence to Andrea Brenciani.

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Edited by: Volkhard A. J. Kempf.

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Figure S1

Checkerboard assays performed with serial dilutions of colistin (rows) and PG-1 (columns). Red squares: no growth; green squares: growth (TIFF 675 kb)

Figure S2

Quantification of the biofilms formed by the six representative A. baumannii strains. Panel A: biofilm formation assay. Panel B: biofilm reduction assay. Error bars represent ± SD. (TIFF 405 kb)

Figure S3

XTT assays involving HeLa ATCC CCL-2 cells exposed to different PG-1 concentrations. Red histograms, 48 h; blue histograms, 24 h. Concentrations (x-axis) are expressed in µg/ml; cell viability (y-axis) is expressed as percentage compared to control. Error bars represent ± SD. Asterisks indicated significant results vs the control group (p < 0.05) (TIFF 497 kb)

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Morroni, G., Simonetti, O., Brenciani, A. et al. In vitro activity of Protegrin-1, alone and in combination with clinically useful antibiotics, against Acinetobacter baumannii strains isolated from surgical wounds. Med Microbiol Immunol 208, 877–883 (2019). https://doi.org/10.1007/s00430-019-00624-7

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  • DOI: https://doi.org/10.1007/s00430-019-00624-7

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