Intense association of non-culturable endophytic bacteria with antibiotic-cleansed in vitro watermelon and their activation in degenerating cultures

The study was undertaken with a view to unravel the source of bacterial colony growth observed in a section of micropropagated triploid watermelon cultures that were supposedly cleansed of the associated endophytic bacteria through antibiotic treatment, and thereafter maintained under stringent ster...

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Bibliographic Details
Published in:Plant cell reports 2011-12, Vol.30 (12), p.2313-2325
Main Author: Thomas, Pious
Format: Article
Language:English
Subjects:
Anti-Bacterial Agents - pharmacology
Antibiotics
bacteria
Bacteria - drug effects
Bacteria - growth & development
Bacteria - isolation & purification
bacterial colonization
Bacteriological Techniques
Biological and medical sciences
Biomedical and Life Sciences
Biotechnology
Cell Biology
Cell culture
Citrullus - microbiology
Citrullus lanatus
Citrullus lanatus var. lanatus
Colonies
Colonization
Contaminants
Degeneration
DNA, Bacterial - genetics
Drug Resistance, Bacterial
Endophytes
Endophytes - drug effects
Endophytes - growth & development
Endophytes - isolation & purification
Fundamental and applied biological sciences. Psychology
hosts
Life Sciences
Media (culture)
microbial growth
micropropagation
Microscopy
Original Paper
Phylogeny
Plant Biochemistry
Plant Sciences
Polymerase Chain Reaction
Sterility
Supplementation
Tissue culture
Tissue Culture Techniques - methods
triploidy
watermelons
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Summary:The study was undertaken with a view to unravel the source of bacterial colony growth observed in a section of micropropagated triploid watermelon cultures that were supposedly cleansed of the associated endophytic bacteria through antibiotic treatment, and thereafter maintained under stringent sterility checks to prevent lateral intrusion of contaminants. Five different bacteria were retrieved from colony growth-displaying watermelon cultures that were previously treated with gentamycin and five isolates from cefazolin-treated stocks with the organisms showing tolerance to the respective antibiotic. These watermelon cultures were in degeneration phase (over 6 months after the previous sub-culturing), while the actively maintained counterpart stocks appeared healthy with no colony growth on different bacteriological media during tissue-screenings. The latter cultures, however, revealed abundant motile, tetrazolium-stained bacterial cells in microscopy, suggesting tissue colonization by non-culturable endophytes. PCR screening on healthy cultures endorsed tissue colonization by different bacterial phylogenic groups. A few organisms could be activated to cultivation from healthy watermelon stocks through host tissue extract supplementation, which also enhanced the growth of all the organisms. The study indicated that a fraction of antibiotic-tolerant bacteria survived intra-tissue in non-culturable form during the preceding cleansing activity, multiplied to substantial numbers thereafter, and turned cultivable in degenerating cultures contributed by tissue breakdown products. This study brings out the existence of a deep endophyte association in tissue cultures which is not easily dissociable. It also signifies the utility of in vitro system for investigations into plant–endophyte association and to bring normally non-culturable novel organisms to cultivation facilitating their future exploitation.
ISSN:0721-7714
1432-203X
DOI:10.1007/s00299-011-1158-z