Effects of temperature and media on preservation and recovery of Mycobacterium tuberculosis strains

Abstract

were preserved directly to the respective temperature. In the second approach a systematic step method starting from RT to 4˚C, to -20˚C, and to -80˚C with one hour interval, was utilized. After each subsequent month for five months, strains were thawed by systematic approach starting from -80˚C to -20˚C to 4˚C to RT. For the second approach strains were directly thawed after five months preservation. The thawed 19 strains by both approaches were subcultured on Lowenstein Jensen, incubated at 37ºC for four weeks and -growth was graded according to the standard method. Cross-tabulation of grading of growth was performed to determine interrelation between temperature and media using 7H9 medium as the standard. Significant differences within and between the grading of each suspension media were determined by using the Kendall’s coefficient of concordance. A p<0.05 was regarded as significant. In the first approach at -80ºC OADC-7H9 had the highest yield while SDW had the lowest yield (p<0.05). At -20ºC SG had highest yield while SDW had the lowest yield (p<0.05). At 4ºC SG had highest yield while PBS had the lowest yield (p<0.05). At RT SM had the highest yield while TSB had the lowest grading (p<0.05). In the second approach, at -80ºC, 5% SG had the highest yield while SDW had the lowest yield p<0.05. At -20ºC, 5% SG had highest yield while SDW had the lowest yield p<0.05. At 4ºC, 5% SG had highest yield while PBS had the lowest yield (p<0.05). At RT SM had the highest yield while TSB had the lowest grading (p<0.05). When approaches one and two were compared for temperature and media, there was no statistically significant difference (p<0.05). This study showed that optimum recovery of MTB strains was mainly dependant on a combination of appropriate temperature and suitable media. The media that was consistent with high recovery was sodium glutamate while the best preservation temperature was -80ºC. More studies are required to determine the effect of 20 preservation using approach one and recovery using approach two and vice versa In addition extension of the preservation duration beyond five months should be further elucidated.