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Antifungal Activity and Physicochemical Properties of a Novel Antimicrobial Protein AMP-17 from Musca domestica

YANG, LONG-BING

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03 set 2019

Antifungal Activity and Physicochemical Properties of a Novel Antimicrobial Protein AMP-17 from Musca domestica's Cover Image

Polish Journal of Microbiology

Volume 68 (2019): Numero 3 (Settembre 2019)

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Categoria dell'articolo: original-paper

Pubblicato online: 03 set 2019

Pagine: 383 - 390

Ricevuto: 15 giu 2019

Accettato: 04 ago 2019

DOI: https://doi.org/10.33073/pjm-2019-041

Parole chiave
antimicrobial peptides, AMP-17, antifungal activity, stability, hemolytic activity

© 2019 Anton Pyskun et al., published by Sciendo

This work is licensed under the Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International License.

Time- kill curves of the AMP-17 against the fungal species tested. The growth of five fungi was monitored in the presence of 100 μg/ml AMP-17. Growth of C. albicans (A), C. tropicalis (B), C. krusei (C), C. parapsilosis (D), and C. neoformans with AMP-17 were monitored by reading the optical density (OD562) of the cultures. 100-μg/ml fluconazole was used as a positive control for fungi. Sabouraud’s dextrose broth (0 μg/ml AMP-17) was used as a negative control. The OD562 values of the remaining viable cells were monitored in 96-well plates at different times.

Influences of various NaCl or MgCl2 concentrations on the antifungal activity of AMP-17. The MIC value of AMP-17 against C. albicans was determined under the influence of different concentrations of NaCl or MgCl2 according to the conditions recommended by CLSI (2008).

Influences of trypsin/pepsin/proteinase K on AMP-17 when used against C. albicans. Fifty μl of the proteases (at a concentration of 1 mg/ml) and 50 μl AMP-17 (at a concentration of 37.5 μg/ml) were mixed in a 96-well plate and placed in a 37°C water bath for 2, 20, 40, 60, 80, 100, and 120 min. Then, it was subjected to CFU enumeration after being treated with 100 μl of the fungal suspension for 18 hours. PBS was used as a negative control. Compared with PBS control, *P < 0.05; **P < 0.01.

Hemolytic activities of AMP-17 in vitro. The diluted human red blood cells (100 μl) were incubated with AMP-17 at 37°C for 1 h and spun at 225×g for 10 minutes. The supernatants (100 μl) were transferred to a 96-well plate for measurement of the absorbance of the supernatants at 450 nm by the ultra-microplate spectrophotometer. 0.1% Triton-X 100 and 0.01M PBS were used as a positive control and a negative control, respectively. Fig. 4A is based on Fig. 4B calculated by the hemolysis rate formula. Fig. 4B was the original experimental diagram.

MIC values of AMP-17 against the fungi species tested_

Peptide	MIC (μg/ml)
Peptide	C. albicansATCC10231	C. kruseiIFM56881	C. tropicalisIFM57016	C. parapsilosisATCC22019	C. neoformansIFM51426
AMP-17	18.75	18.75	18.75	18.75	9.375

MFC values of AMP-17 against the fungi species tested_

Peptide	MIC (μg/ml)
Peptide	C. albicansATCC10231	C. kruseiIFM56881	C. tropicalisIFM57016	C. parapsilosisATCC22019	C. neoformansIFM51426
AMP-17	37.5	37.5	37.5	37.5	18.75

Influences of temperature and the number of freeze-thaw times on the antifungal activity of AMP-17_

Temperature (98°C)(min)	Colony count( $\bar{x}$ ± SD)	Temperature (−80°C)(Times)	Colony count( $\bar{x}$ ± SD)
5	69.50 ± 4.95^**	1	(240.50 ± 3.54^**) × 10²
20	6.00 ± 2.83^**	2	(273.50±3.54^**) × 10²
30	4.50 ± 2.12^**	4	(342.00 ± 8.49^**) × 10²
60	6.50 ± 0.71^**	6	(357.00 ± 1.41^**) × 10²
90	9.50 ± 3.54^**	8	(351.50 ± 6.36^**) × 10²
120	3.50 ± 0.71^**	10	(370.50 ± 3.54^**) × 10²
Negative control (ddH₂O)	565900.00 ± 17112.00	Negative control (ddH₂O)	(5507.00 ± 147.08) × 10²

Lingua:: Inglese

Frequenza di pubblicazione:: 4 volte all'anno
Argomenti della rivista:: Scienze biologiche, Microbiologia e virologia

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