Current Microbiology

, Volume 74, Issue 1, pp 103–113 | Cite as

Investigation of Endophytic Bacterial Community in Supposedly Axenic Cultures of Pineapple and Orchids with Evidence on Abundant Intracellular Bacteria

  • Natalia Pimentel Esposito-PolesiEmail author
  • Monita Fiori de Abreu-Tarazi
  • Cristina Vieira de Almeida
  • Siu Mui Tsai
  • Marcílio de Almeida


Asepsis, defined as the absence of microbial contamination, is one of the most important requirements of plant micropropagation. In long-term micropropagated cultures, there may occasionally occur scattered microorganism growth in the culture medium. These microorganisms are common plant components and are known as latent endophytes. Thus, the aim of this research was to investigate the presence of endophytic bacteria in asymptomatic pineapple and orchid microplants, which were cultivated in three laboratories for 1 year. Isolation and characterization of bacterial isolates, PCR–DGGE from total genomic DNA of microplants and ultrastructural analysis of leaves were performed. In the culture-dependent technique, it was only possible to obtain bacterial isolates from pineapple microplants. In this case, the bacteria genera identified in the isolation technique were Bacillus, Acinetobacter, and Methylobacterium. The scanning electron microscopy and transmission electron microscopy (SEM and TEM) analyses revealed the presence of endophytic bacteria in intracellular spaces in the leaves of pineapple and orchid microplants, independent of the laboratory or cultivation protocol. Our results strongly indicate that there are endophytic bacterial communities inhabiting the microplants before initiation of the in vitro culture and that some of these endophytes persist in their latent form and can also grow in the culture medium even after long-term micropropagation, thus discarding the concept of “truly axenic plants.”


Transmission Electron Microscopy Micrographs Bacterial Colonization Endophytic Bacterium Intracellular Space Band Profile 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.



The authors acknowledge the Fundação de Amparo a Pesquisa do Estado de São Paulo—FAPESP (Process No: 2008/07535-9) and CAPES for providing financial support. The authors also thank Elliot Kitajima and Francisco André Ossamu Tanaka (NAP/MEPA/ESALQ/USP) for use of the electronic microscope.


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Copyright information

© Springer Science+Business Media New York 2016

Authors and Affiliations

  • Natalia Pimentel Esposito-Polesi
    • 1
    Email author
  • Monita Fiori de Abreu-Tarazi
    • 1
  • Cristina Vieira de Almeida
    • 2
  • Siu Mui Tsai
    • 3
  • Marcílio de Almeida
    • 1
  1. 1.Biological Science Department, “Luiz de Queiroz” Superior College of AgricultureUniversity of São Paulo (ESALQ/USP)PiracicabaBrazil
  2. 2.In Vitro Palm Consultoria, Estudo e Desenvolvimento Biológico LtdaPiracicabaBrazil
  3. 3.Cell Biology and Molecular Laboratory, Nuclear Energy in Agriculture CenterUniversity of São PauloPiracicabaBrazil

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