Identification of culturable and originally non-culturable endophytic bacteria isolated from shoot tip cultures of banana cv. Grand Naine

Abstract

In this article we describe the identification of endophytic bacteria belonging to three groups isolated from shoot tip cultures of banana cv. Grand Naine in a recent study (Thomas et al. 2008) based on partial 16S rRNA gene sequence homology analysis. The first group included banana stocks that displayed obvious colony growth on MS based tissue culture medium during the first in vitro passage. The second group constituted stocks that were tissue indexnegative for cultivable bacteria initially but turned index-positive after a few to several (4-8) in vitro passages while the third group formed one sub-stock that turned index-positive after about 18 passages. The organisms belonged to about 20 different genera comprising of a, b, c-proteobacteria, Gram-positive firmicutes and actinobacteria. Visibly expressing easily cultured organisms during the first in vitro passage included Enterobacter, Klebsiella, Ochrobactrum, Pantoea, Staphylococcus and Bacillus spp. Organisms of second group that were not detected or nonculturable originally constituted Brevundimonas, Methylobacterium, Alcaligenes, Ralstonia, Pseudomonas, Corynebacterium, Microbacterium, Staphylococcus, Oceanobacillus and Bacillus spp. while the third group that turned cultivable after extended in vitro culturing included mostly non-filamentous actinobacteria (Brachybacterium, Brevibacterium, Kocuria and Tetrasphaera spp.). The identification results suggested that the endophytes of second and third groups were not strictly obligate or fastidious microbes but those surviving in viable but-non-culturable (VBNC) state and displaying gradual activation to cultivable form during continuous tissue culturing. Several of the organisms isolated are known as beneficial ones in agriculture while some organisms have possible implications in human health. The use of tissue cultures for isolating uncommon endophytes is discussed.Supply of live bacterial cultures or genetic material for research purpose is subject to their revival from glycerol stocks (as some of the organisms showed poor tolerance) and the requestor obtaining written permission from the Director General,

A New Approach to Analyzing Endophytic Actinobacterial Population in the Roots of Banana Plants (Musa sp., AAA)

“…The actinobacterial taxa were not consistent with previous results. Streptomyces was the most frequently isolated genus from surface-sterilized banana roots [7] , nevertheless, it can not be isolated or detected in previous studies [1,6,22] . In the study, total 11 reads belonged to Streptomyces or Streptomycetaceae were detected by Illumina-based analysis.…”

Técnicas para el aislamiento, identificación y caracterización molecular de cepas de Ralstonia solanacearum relacionadas a Moko del plátano

“…However, even in this medium, isolating this bacteria is difficult, due to the growth of other microorganisms (Champoiseau et al, 2009;Kalpage y De Costa, 2015), particularly bacteria of the genera Klebsiella, Erwinia, and Pseudomonas. These three gram-negative bacteria, like other species of Ralstonia, such as Ralstonia mannitolilytica, are majority endophytic in banana plants (Thomas et al, 2008;Ganen et al, 2009;Souza et al, 2013). These bacteria are common contaminants in R. solanacearum plantations, since they present a faster growth and similar microscopic and colonial morphologies (French et al, 1995;Nasim, 2011).…”

“…This report describes a strategy that makes the isolation and genetic characterization of R. solanacearum easier, from banana diseased with Moko. (Thomas et al, 2008;Ganen et al, 2009;Souza et al, 2013). Estas bacterias son contaminantes comunes en los cultivos de R. solanacearum debido a que presentan mayor capacidad de crecimiento y muestran morfologías microscópicas y coloniales similares (French et al, 1995;Nasim, 2011).…”

“…Ramesh et al, (2009) point out that genus Pseudomonas is opportunist during the colonization of the host by R. solanacearum. Along with Pseudomonas, the bacteria Klebsiella, Erwinia, and other species of the genus Ralstonia, such as Ralstonia mannitolilytica, are mostly endophytic of the banana (Thomas et al, 2008;Ganen et al, 2009;Souza et al, 2013). The bacteria recovered from the 3 samples that produced faint lines on the ImmunoStrips presented fluorescence on the B-King medium and did not amplify in any of the PCR diagnosis reactions for R. solanacearum.…”

“…The identity of the R. solanacearum strains was confirmed by sequencing the amplified fragment (Thomas et al, 2008;Ganen et al, 2009;Souza et al, 2013). Las bacterias recuperadas de las 3 muestras que dieron las líneas tenues en las inmunotiras presentaron fluorescencia sobre el medio B-King y no amplificaron en ninguna de las reacciones de PCR de diagnóstico para R. solanacearum.…”