The study was aimed primarily at cleansing the in vitro-decline displaying long-term micropropagated triploid watermelon 'Arka Manik' cultures from covert bacteria and their further field testing. Disinfectant treated shoots showed endophytic survival but to a lesser extent in shoot-tips. Culturing the NaOCl (5 min) treated shoot-tips on filter paper bridges in liquid watermelon medium containing single antibiotic (gentamycin (Gm), amoxycillin (Ax) or cefazolin (Cz) at 50, 250, 500 or 1000 mg l )1 ) for 1 month followed by repeated indexing of medium and tissue for two-four subculture passages facilitated the cleansing of cultures with 12.5% recovery as monitored for 2 years. Partial bleaching damage by NaOCl, phytotoxicity to higher levels of antibiotics, poor growth response in the initial sucrose-free medium and rampant hyperhydricity came in the way of a higher recovery. The effectiveness of the above approach was ascertained after back inoculating clean cultures with a mixture of Gram-positive and Gram-negative bacteria yielding 30, 45 and 35% clean cultures in Gm, Ax and Cz (250 mg l )1 for 1 month) treatments, respectively in modified medium compared with 10% recovery after mere HgCl 2 surface sterilization. The results indicated that antibiotic treatment was essential but not its choice, and extended culture-indexing subsequent to disinfection or antibiotic treatment was crucial in identifying clean stocks. Cleansed cultures, showed restoration of growth but a drop in rooting. Most of the in vitro cultures appeared normal and trueto-type during the 7-10 year period in vitro but a small proportion of bacteria-harboring stocks displayed 'epigenetic variations'. Acclimatized plants and those in the field also appeared true-to-type but for a minor proportion derived from bacteria-harboring stocks. Field-plants which originated from bacteria-freed stocks after 9 years of continuous culturing were normal and fertile validating the possibility of keeping treasured cultures in vitro for long periods if covert contaminants are checked.Abbreviations: Ax -amoxycillin; BIM -Bacteriological indexing medium; CoBa+ -covert bacteria harboring culture; CoBa--culture devoid of bacteria in covert form; Cz -cefazolin; Gm -gentamycin; PP bag -polypropylene bag; PR -propagation rate; WBNV -watermelon bud necrosis virus; WMM -watermelon multiplication medium; WRM -watermelon rooting medium; 2x -diploid; 3x -triploid
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