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This research aimed to determine the optimal media and temperature conditions for the long-term storage of bacteriophages. In this study, the viability of Salmonella phages in 50% glycerol, 10% sodium chloride-magnesium sulfate (SM) buffer, and 5% dimethyl sulfoxide (DMSO) media at room temperature, 4oC, -20oC, and -80oC for 12 months was determined. In 50% glycerol, at the end of the experiment, no significant difference was found between four temperature conditions on phage density, ranging from 6.20-6.23 log10 PFU/mL (P>0.05). Under 10% SM medium, phage preservation at room temperature provided the optimum density at 6.31 log10 PFU/mL. In addition, phages preserved in a 5% DMSO medium were of similar density values across all temperature treatments. Still, their availability after 12 month-storage (88.0-88.5%) was significantly lower (P<0.05) than that of 50% glycerol and 10% SM. Moreover, for phage lysis capacity, low temperatures (4oC, -20oC, and -80oC) were superior to room temperature used for preservation. Considering the density, lysis capacity, and practical convenience, storing phages at 4°C in a 50% Glycerol medium is recommended.
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Ackermann, H.W., Tremblay, D. and Moineau, S., 2004. Long-term bacteriophage preservation. World Fed. Cult. Collect. Newslett. 38, 35-40.
Alvi, I.A., Asif, M., Tabassum, R., Abbas, Z. and Rehman, S., 2018. Storage of bacteriophages at 4°C leads to no loss in their titer after one year. Pakistan J. Zool. 50, 2395-2398.
Bonilla, N., Rojas, M.I., Netto Flores Cruz, G., Hung, S.H., Rohwer, F. and Barr, J.J., 2016. Phage on tap-a quick and efficient protocol for the preparation of bacteriophage laboratory stocks. PeerJ. 4, e2261.
Clark, W.A., 1962. Comparison of several methods for preserving bacteriophages. Appl. Environ. Microbiol. 10, 466-471.
Doss, J., Culbertson, K., Hahn, D., Camacho, J. and Barekzi, N., 2017. A review of phage therapy against bacterial pathogens of aquatic and terrestrial organisms. Viruses 9, 50.
Golec, P., Dąbrowski, K., Hejnowicz, M.S., Gozdek, A., Loś, J.M., Węgrzyn, G., Lobocka, M.B. and Loś, M., 2011. A reliable method for storage of tailed phages. J. Microbiol. Methods 84, 486-489.
González-Menéndez, E., Fernández, L., Gutiérrez, D., Rodríguez, A., Martínez, B. and García, P., 2018. Comparative analysis of different preservation techniques for the storage of Staphylococcus phages aimed for the industrial development of phage-based antimicrobial products. PloS One 13, e0205728.
Jepson, C. D. and March, J. B., 2004. Bacteriophage lambda is highly stable DNA vaccine delivery vehicle. Vaccine 22, 2413-2419.
Leung, S.S.Y., Parumasivam, T., Nguyen, A., Gengenbach, T., Carter, E.A., Carrigy, N.B., Wang, H., Vehring, R., Finlay, W.H., Morales, S, Britton, W.J., Kutter, E. and Chan, H.K., 2018. Effect of storage temperature on the stability of spray dried bacteriophage powders. Eur. J. Pharm. Biopharm. 127, 213-222.
Litt, P.K. and Jaroni, D., 2017. Isolation and physiomorphological characterization of Escherichia coli O157:H7-infecting bacteriophages recovered from beef cattle operations. Int. J. Microbiol. 2017(4), 1-12.
Malik, D. J., Sokolov, I. J., Vinner, G. K., Mancuso, F., Cinquerrui, S., Vladisavljevic, G. T., Clokie, M., Garton, N.J., Stapley, A. and Kirpichnikova, A., 2017. Formulation, stabilisation and encapsulation of bacteriophage for phage therapy. Adv. Colloid. Interface Sci. 249, 100-133.
Manohar, P., and Ramesh, N., 2019. Improved lyophilization conditions for long-term storage of bacteriophages. Sci. Rep. 9: 15242.
Mazur, P. and Kleinhans, F.W., 2008. Relationship between intracellular ice formation in oocytes of the mouse and Xenopus and the physical state of the external medium-a revisit. Cryobiology 56, 22-27.
Minitab, 2010. Minitab Reference Manual, Release 16.2.1 for Windows. Minitab Inc., United States.
Moghimian, P., Srot, V., Pichon, B.P., Facey, S.J. and van Aken, P.A., 2016. Stability of m13 phage in organic solvents. J. Biomater. Nanobiotechnol. 7, 72-77.
Nagao, Y., Ishiguro, H. and Nukina, N., 2000. DMSO and glycerol reduce bacterial death induced by expression of truncated N-terminal huntingtin with expanded polyglutamine tracts. Biochim. Biophys. Acta - Mol. Basis Dis., 1502(2), 247-256.
Novik, G., Sidarenka, A., Rakhuba, D. and Kolomiets, E., 2009. Cryopreservation of bifidobacteria and bacteriophages in belarusian collection of non-pathogenic microorganisms. J. Cult. Collect. 6, 76-84.
Olson, M. R., Axler, R.P. and Hicks, R. E., 2004. Effects of freezing and storage temperature on MS2 viability. J. Virol. Methods 122, 147-152.
Poxleitner, M., Pope, W., Jacobs-Sera, D., Silvanathan, V. and Hatfull, D., 2017. Phage Discovery Guide. Hevy Chase (US): Howard Hughes Medical Institute.
Puapermpoonsiri, U., Ford, S. J. & van der Walle, C. F., 2010. Stabilization of bacteriophage during freeze-drying. Int. J. Pharm. 389(1), 168-175.
Tahir, A., Asif, M., Abbas, Z. and Rehman, S.U., 2017. Three bacteriophages SA, SA2 and SNAF can control growth of milk isolated Staphylococcal species. Pakistan J. Zool. 49, 529-533.
Tovkach, F., Zhuminska, G. and Khushkina, A., 2012. Long-term preservation of unstable bacteriophages of enterobacteria. Mikrobiol. Z. 74, 60-66.
Vagenende, V., Yap, M.G.S. and Trout, B.L., 2009. Mechanisms of protein stabilization and prevention of protein aggregation by glycerol. Biochemistry 48, 11084-11096.
Wallis, C. and Melnick, J.L., 1968. Stabilization of enveloped viruses by dimethyl sulfoxide. J. Virol. 2(9), 953-954.