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. 2020 Mar 13;9(3):215.
doi: 10.3390/pathogens9030215.

Isolation, Characterization, and Application of a Bacteriophage Infecting the Fish Pathogen Aeromonas hydrophila

Muhammad Akmal  1 , Aryan Rahimi-Midani  2 , Muhammad Hafeez-Ur-Rehman  1 , Ali Hussain  3 , Tae-Jin Choi  2
Affiliations

Isolation, Characterization, and Application of a Bacteriophage Infecting the Fish Pathogen Aeromonas hydrophila

Muhammad Akmal et al. Pathogens. .

Abstract

Bacteriophages are increasingly being used as biological control agents against pathogenic bacteria. In the present study, we isolate and characterize bacteriophage Akh-2 from Geoje Island, South Korea, to evaluate its utility in controlling motile Aeromonas septicemia. Akh-2 lysed four of the seven Aeromonas hydrophila strains tested. Transmission electron microscopy analysis showed that Akh-2 belongs to the Siphoviridae family, with head and tail sizes of 50 ± 5 and 170 ± 5 nm, respectively. One-step growth curve analysis revealed that the phage has a latent period of 50 ± 5 min and a burst size of 139 ± 5 plaque-forming units per infected cell. The phage appeared stable in a pH range of 6-8 and a temperature range of -80 to 46 °C. Based on next-generation sequencing analysis, its genome is 114,901 bp in size, with a 44.22% G + C content and 254 open reading frames. During an artificial induction of the disease, loach (Misgurnus anguillicaudatus) treated with Akh-2 showed an increased survival rate and time compared with the non-treated control. Our results suggest that Akh-2 is a potential biological agent for the treatment of Aeromonas infections in fish.

Keywords: Aeromonas hydrophila; aquaculture; bacteriophage; phage genomics; phage therapy.

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Conflict of interest statement

The authors declare no conflicts of interest.

Figures

Figure 1
Figure 1
Electron micrograph of bacteriophage Akh-2. The two arrows indicate the beginning and end of the 170-nm-long tail. Scale bar = 50 nm.
Figure 2
Figure 2
One-step growth curve of phage Akh-2. The results are the average of three replications with standard deviation as vertical lines.
Figure 3
Figure 3
Stability of phage Akh-2 at different temperatures. Phage Akh-2 (107 PFU/mL) was maintained at the indicated temperatures for three days, and then the titer was determined by plaque assay. The results are the average of three replications with standard deviation as vertical lines.
Figure 4
Figure 4
Stability of phage Akh-2 at different pH levels. Phage Akh-2 (107 PFU/mL) was maintained at the indicated pH levels for three days, and then the titer was determined by plaque assay. The results are the average of three replications with standard deviation as vertical lines.
Figure 5
Figure 5
Genomic comparison of phage Akh-2 and the reference A. hydrophila phage AhSzw-1 (GenBank No. MG676225.1), constructed using EasyFigure. Arrows represent ORFs. The level of identity is indicated by the gray shading.
Figure 6
Figure 6
Protective effect of Akh-2 against A. hydrophila in inoculated loach. Af-clipped loach were immersed in PBS (□), A. hydrophila (1 × 107 CFU/mL) (▨), or A. hydrophila (1 × 107 CFU/mL), followed by Akh-2 (1 × 108 PFU/mL) (■), and the survival rate was measured within 96 h. The results are the average of three replications with standard deviation as vertical lines (Table S2).
Figure 7
Figure 7
Protection of loach (M. anguillicaudatus) from A. hydrophila infection by bacteriophage Akh-2. (A) Non-infected negative control. (B) Loach inoculated with A. hydrophila showing hemorrhagic red spots. (C) Loach treated with phage Akh-2 after inoculation with A. hydrophila showing fewer and smaller hemorrhagic red spots (indicated by red boxes). The average fish size was 5 ± 2 cm.
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References

    1. The State of World Fisheries and Aquaculture 2016. FAO Fisheries and Aquaculture Department; Rome, Italy: 2016. Contributing to food security and nutrition for all.
    1. Luis A.I.S., Campos E.V.R., Oliveira J.L., Fraceto L.F. Trends in aquaculture sciences: From now to use of nanotechnology for disease control. Rev. Aquac. 2017;11:119–132. doi: 10.1111/raq.12229. - DOI
    1. Jun J.W., Kim J.H., Shin S.P., Han J.E., Chai J.Y., Park S.C. Protective effects of the Aeromonas phages pAh1-C and pAh6-C against mass mortality of the cyprinid loach (Misgurnus anguillicaudatus) caused by Aeromonas hydrophila. Aquaculture. 2013;416:289–295. doi: 10.1016/j.aquaculture.2013.09.045. - DOI
    1. Austin B., Austin D.A. Bacterial Fish Pathogens. Springer; Dordrecht, The Netherlands: 2012. Aeromonadaceae Representative (Aeromonas salmonicida) pp. 147–228.
    1. Lilley J., Hart D., Richards R.H., Roberts R.J., Cerenius L., Söderhäll K. Pan-Asian spread of single fungal clone results in large scale fish kills. Veter Rec. 1997;140:653–654. doi: 10.1136/vr.140.25.653. - DOI - PubMed

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