**4. Storage of phages**

Phage preparations for clinical use ought to be (i) endotoxin-free, (ii) phage must be intact with high titers [53–55]. (iii) Suitable storage and transport are crucial. (iv) protected from high temperature, extremely acidic, or alkaline conditions [56], and (v) phage stock should not be refrozen and rethawed [57]. The usefulness of preparation of phage lysate, modified treatment methods evolved and accepted for long-term storage of phage was elucidated. In a study that demonstrated the infectivity of the phages remained unaffected with chloroform and DMSO treatments and storage for 30 days to a year at 4°C 40°C [10, 58–60]. Infectivity of long-term stored bacteriophages at 80°C can be increased by adding 15%-25% glycerol to phage lysate preparations, and by rapid freezing and storage of phage infected bacteria at 70°C. Similarly, phage was shown to remain highly stable underneath normal storage conditions or also stable in NaCl and MgSO4 due to its stabilizing effect. Considerable numbers of viable phage have been described

to occur even after storage in distilled water. Phage isolates were found to remain stable upon storage at 4°C, or a rapid loss of phage infectivity was encountered with repeated freezing and thawing at 70°C. Phage infectivity could not be inhibited with trypsin, protease, ribonuclease treatments, or chloroform whilst the infectivity over the phage was inhibited together with lysozyme and SDS treatments [10, 59, 60]. The enzymatic treatments and inhibition of phage infectivity of several bacteriophages had been reported. Similarly, *Mycoplasma arthritidis* virulent 1 (MAV1) phage infectivity was reported to be unaffected by treatment with Triton X-100 and used to be resistant to non-ionic detergents [55, 61]. Phage survived a hundred percent at pH 7 and exhibited infectivity, whilst none of the phage survived at extreme pH conditions (pH 3 and pH 12) [10]. At a temperature below 37°C, phage JSF9 was shown to be stable whereas, at 50°C, the phage had been rapidly inactivated. Phage (VPP97) of *V. parahaemolyticus* have been shown to be stable up to 65°C and were totally inactivated at 70°C [10, 61–64]. Bacteriophages were detected to survive extremes over 95°C [52]. Bacteriophages such as T-ϕD0, TϕD2S, T-ϕHSIC, and T-ϕD1B exhibited a latent period ranging beyond 90°C [64]. The effects of temperature on the survival and infectivity of bacteriophages have clearly shown that the physicochemical parameters are very important for the survival and infectivity of phage [55, 58, 59]. Bacteriophages can be resilient to low/ high temperatures, salinity, pH, and ions. They can tolerate extreme environments. New data on these along with therapeutic phage survivability, methods of their preservation and transport shall be useful.
