Distinguishing Bacteriolytic from Bacteriostatic Activity of an Antibiotic Agent by Real-Time Quantitative PCR

Abstract:

A simple and rapid real-time quantitative PCR assay was devised to discriminate bacteriolytic from bacteriostatic activity for a given antibacterial agent. Bacteria suspension was incubated with the compound solution, the mixture was centrifuged and supernatant was removed completely, the obtained pellet was then used as the DNA template for PCR. Then the bacteriostatic and bacteriolytic activity can be inferred by the quantity of PCR product. Moreover, the parameters that influence the assay sensitivity was discussed.

• Periodical: Advanced Materials Research (Volumes 343-344)
• Pages: 357-360
• DOI: https://doi.org/10.4028/www.scientific.net/AMR.343-344.357
• Online since: September 2011
• Authors: Zhi Ping Wang
• Keywords: Bacteriolytic, Bacteriostatic, Real Time Quantitative PCR

References

[1] G. Mardones, and A. Venegas, Chromogenic plate assay distinguishing bacteriolytic from bacteriostatic activity of an antibiotic agent, J. Microbio. Metho, vol. 40, pp.199-206, (2000).

DOI: 10.1016/s0167-7012(00)00125-1

[2] DeGrado W.F., Kezdy, F.J., and Kaiser, E.T., Design, synthesis and characterization of a cytotoxic peptide with melittin-like activity, J. Am. Chem. Soc, vol. 103, pp.679-681, (1981).

DOI: 10.1021/ja00393a035

[3] Kaiser, E.T., and Kezdy, F.J., Secondary structures of proteins and peptides in amphiphilic environments, Proc. Natl. Acad. Sci. USA, vol. 80, pp.1137-1143, (1983).

DOI: 10.1073/pnas.80.4.1137

[4] Ganz, T., Selsted, M.E., and Lehrer, R.I., Defensins, Eur. J. Haematol, vol. 44, pp.1-8, (1990).

[5] Habermann, E., Bee and wasp venoms, Science, vol. 177, p.314–322, (1972).

[6] Steiner, H., Hultmark, D., Engstro¨m, A., Bennich, H., and Boman H.G., Sequence and specificity of two antibacterial proteins involved in insect immunity, , Nature, vol. 292, pp.246-248, (1981).

DOI: 10.1038/292246a0

[7] Okada, M., and Natori, S., Primary structure of sarcotoxin I, an antibacterial protein induced in the hemolymph of Sarcophaga peregrina (Flesh Fly) larvae, J. Biol. Chem., vol. 260, pp.7174-7177, (1985).

DOI: 10.1016/s0021-9258(17)39590-x

[8] Zasloff, M., and Magainins, a class of antimicrobial peptides from Xenopus skin: isolation, characterization of two active forms, and partial cDNA sequence of a precursor, Proc. Natl. Acad. Sci. USA, vol. 84, pp.5449-5454, (1987).

DOI: 10.1073/pnas.84.15.5449

[9] Westerhoff, H.V., Juretic, D., Hendler, R.W., and Zasloff, M., Magainins and the disruption of membrane-linked free-energy transduction, Proc. Natl. Acad. Sci. USA, vol. 86, pp.6597-6601, (1989).

DOI: 10.1073/pnas.86.17.6597

[10] Christensen, B., Fink, J., and Merrifield, R.B., Mauzerall, D., Channel-forming properties of cecropins and related model compounds incorporated into planar lipid membranes, Proc. Natl. Acad. Sci. USA, vol. 85, pp.5072-5076., (1988).

DOI: 10.1073/pnas.85.14.5072

[11] Cruciani, R.A., Stanley, E.F., Zasloff, M., Lewis, D.L., and Barker, J.L., The antibiotic magainin II from the African clawed frog forms an anion permeable ionophore in artificial membranes, J. Biophys, vol. 53, pp. 9a, (1988).

[12] Kagan, B.L., Selsted, M.E., Ganz, T., Lehrer, and R.I., Antimicrobial defensin peptides form voltage-dependent ion-permeable channels in planar lipid bilayer membranes. Proc, Natl Acad. Sci. USA, vol. 87, pp.210-214, (1990).

DOI: 10.1073/pnas.87.1.210

[13] Bessalle, R., Kapitkovsky, A., Gorea, A., Shalit, I., Fridkin, and M., All-D-magainin: chirality, antimicrobial activity and proteolytic resistance, FEBS Lett, vol. 274, pp.151-155. (1990).

DOI: 10.1016/0014-5793(90)81351-n

[14] Flouret, G., and du Vigneaud, V., The synthesis of D-oxytocin, the enantiomer of the posterior pituitary hormone, oxytocin, J. Am. Chem. Soc. vol. 87, pp.3775-3776, (1965).

DOI: 10.1021/ja01094a045

[15] Morley, J.S., Tracy, H.J., and Gregory, R.A., Structure–function relationships in the active C-terminal tetrapeptide sequence of gastrin, Nature, vol. 207, pp.1356-1359, (1965).

DOI: 10.1038/2071356a0

[16] Stewart, J.M., and Woolley, D.W., All-D-bradykinin and the problem of peptide antimetabolites, Nature, vol. 206, pp.619-620, (1965).

DOI: 10.1038/206619b0

[17] Casteels, P., and Tempst, P., Apidaecin-type peptide antibiotics function through a non-poreforming mechanism involving stereospecificity. Biochem. Biophys, Res. Commun, vol. 199, pp.339-345, (1994).

DOI: 10.1006/bbrc.1994.1234

[18] Z Wang, and S Zhang. The role of lysozyme and complement in the antibacterial activity of zebrafish (Danio rerio) egg cytosol. Fish & shellfish immunology, vol. 29, pp.773-777, (2010).

DOI: 10.1016/j.fsi.2010.07.002