A Virulent Salmonella enterica Serovar Enteritidis Phage SE2 with a Strong Bacteriolytic Activity of Planktonic and Biofilmed Cells

Birendra R. Tiwari; Shukho Kim; Jungmin Kim

doi:10.4167/jbv.2013.43.3.186

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Tiwari, Kim, and Kim: A Virulent Salmonella enterica Serovar Enteritidis Phage SE2 with a Strong Bacteriolytic Activity of Planktonic and Biofilmed Cells

Original Article

J Bacteriol Virol 2013;43(3):186-194.

Published online: 9 February 2013

DOI: https://doi.org/10.4167/jbv.2013.43.3.186

A Virulent Salmonella enterica Serovar Enteritidis Phage SE2 with a Strong Bacteriolytic Activity of Planktonic and Biofilmed Cells

Birendra R. Tiwari, Shukho Kim¹, Jungmin Kim^*

Department of Microbiology, Kyungpook National University, School of Medicine, Daegu, Korea

^*Corresponding author: Jungmin Kim, M.D., Ph.D. Department of Microbiology, School of Medicine, Kyungpook National University, Daegu 700-422, Korea. Phone: +82-53-420-4845, Fax: +82-53-427-5664, e-mail: minkim@knu.ac.kr

^** This research was supported by Kyungpook National University Research Fund, 2012.

¹ Present address: Hazardous Substances Analysis Division. Daegu Regional Food & Drug Administration, Daegu.

Received 7 June 201324 June 2013 Accepted 31 June 2013

This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/3.0) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract

Salmonella enterica serovar Enteritidis is one of the major food borne pathogens. Utilizing lytic bacteriophages against this pathogen can be a new and effective approach for the prevention of food-contamination and food-borne infection. In this study, we isolated and characterized a Salmonella Enteritidis specific lytic bacteriophage (phage SE2). The bacteriolytic activity of planktonic and biofilmed cells against an antibiotic resistant strain of Salmonella Enteritidis was also evaluated. Phage SE2 revealed an efficient bacteriolytic effect with biofilm dispersing ability and could maintain its virulence even at extreme pH and temperature. It can be a potential biotherapeutic agent against Salmonella Enteritidis.

Keywords: Virulent, Salmonella Enteritidis, Bacteriophage, Drug resistance, Biofilm

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Figure 1.

Adsorption rate and burst size of Phage SE2. (A) Adsorption rate of Phage SE2. (B) Burst size of Phage SE2. L, Latent time (10 minutes); b, average burst size (155 PFU/host cells at 35 minutes).

Figure 2.

Ability of phage SE2 to kill bacteria and produce progenies at different temperature. (A) Ability of phage SE2 to kill bacteria at 37°C, ambient temperature and 4°C. (B) Ability of phage to produce progenies during infection at 37°C, ambient temperature (AT) and 4°C.

Figure 3.

Bacteriolytic activity of the phage SE2 to biofilmed cells. (A) Biofilmed cells of Salmonella Enteritidis JB-201 were treated with tryptic soy broth (control), phage SE2 (10¹¹ PFU/ml) or ciprofloxacin (0.5 μg/ml) for 4 hours at 37°C. Mean bacterial CFU was calculated from the triplicates. (B) Field emission scanning electron microscopic (FE-SEM) images showing biofilm dispersion activity of phage SE2. FE-SEM images showed that almost all the biofilmed cells were lysed with the dispersion of biofilm's matrix by phage SE2, very few cells, cell debris and ghost like bacteria can be seen on phage treated biofilms images (lower) compared that of phage untreated control (upper). At least three independent experiments were performed.

Table 1.

Host range of phage SE2 against Salmonella enterica serovars Enteritidis and Gallinarum.

Salmonella Serovars	Year of isolation	Plaques¹	Salmonella Serovars	Year of isolation	Plaques¹
S. Enteritidis 012	2000	++	S. Gallinarum I	?	++
S. Enteritidis 013	2000	++	S. Gallinarum II	?	++
S. Enteritidis 014	2000	++	S. Gallinarum 08vs-38	2008	++
S. Enteritidis 015	2002	++	S. Gallinarum 08vs-178	2008	++
S. Enteritidis 018	2002	++	S. Gallinarum 08vs-3	2008	++
S. Enteritidis 09	2005	++	S. Gallinarum 08vs-4	2008	++
S. Enteritidis 010	2005	++	S. Gallinarum 08vs-5	2008	++
S. Enteritidis 011	2005	++	S. Gallinarum 08vs-6	2008	++
S. Enteritidis 015	2006	++	S. Gallinarum 08vs-8	2008	++
S. Enteritidis 016	2006	++	S. Gallinarum 08vs-10	2008	++
S. Enteritidis 017	2006	++	S. Gallinarum 08vs-18	2008	++
S. Enteritidis 01	2009	++	S. Gallinarum 08vs-19	2008	++
S. Enteritidis 09	2009	++	S. Gallinarum 08vs-20	2008	++
S. Enteritidis PT-4	?	++	S. Gallinarum 08vs-21	2008	++
S. Enteritidis Jb-10-201	2010	++	S. Gallinarum 08vs-23	2008	++
S. Enteritidis 5-17-6	2010	++	S. Gallinarum 08vs-25	2008	++
S. Enteritidis 5-6-5	2010	+	S. Gallinarum 08vs-27	2008	++
S. Enteritidis 5-6-2	2010	+	S. Gallinarum 08vs-33	2008	++
			S. Gallinarum 08vs-34	2008	++
			S. Gallinarum 08vs-36	2008	+
			S. Gallinarum 08vs-38	2008	+
			S. Gallinarum 08vs-42	2008	+
			S. Gallinarum 08vs-43	2008	++
			S. Gallinarum 08vs-45	2008	++
			S. Gallinarum 08vs-51	2008	++
			S. Gallinarum 08vs-4	2008	++
			S. Gallinarum 08vs-52	2008	++
			S. Gallinarum 08vs-53	2008	++
			S. Gallinarum 08vs-56	2008	++
			S. Gallinarum 08vs-57	2008	++
			S. Gallinarum 08vs-165	2008	++
			S. Gallinarum 08vs-168	2008	++
			S. Gallinarum 08vs-173	2008	++
			S. Gallinarum 08vs-178	2008	++
			S. Gallinarum 08vs-188	2008	++
			S. Gallinarum 08vs-190	2008	++
			S. Gallinarum 08vs-191	2008	++
			S. Gallinarum 08vs-200	2008	++
			S. Gallinarum 08vs-202	2008	++
			S. Gallinarum 08vs-236	2008	++
			S. Gallinarum 08vs-240	2008	++
			S. Gallinarum 08vs-251	2008	++
			S. Gallinarum 09vs-6	2009	+
			S. Gallinarum 09vs-7	2009	+
			S. Gallinarum 09vs-8	2009	+
			S. Gallinarum 09vs-9	2009	++

¹ ++, large clear lysis, +, small clear lysis

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