Comparative analysis of antibiotic resistance, immunofluorescent colony staining, and a transgenic marker (bioluminescence) for monitoring the environmental fate of rhizobacterium

Abstract

Field releases of the wild-type plant growth-promoting rhizobacterium Pseudomonas fluorescens 89B-27, its bioluminescent derivative GEM-8 (89B-27::Tn4431), and a spontaneous rifampin-resistant variant (R34) were established on cucumber in 1994 and 1995 to examine the efficiency of these marker systems for estimating the wild-type population. Seed and root samples were taken 0, 7, 14, 21 or 28, 35 or 42, and 70 days after planting in each year and processed for enumeration by spiral plating or immunofluorescent colony staining (IFC). In both years, the populations of 89B-27, R34, and GEM-8, as measured by IFC, were not significantly different (P > 0.05) from each other at each sampling time. However, the populations of R34 and GEM-8, as measured by spiral plating and differentiation based on their respective phenotypes, were significantly lower (P < 0.05) than the wild-type populations and their IFC-determined populations. These data indicate that traditional marker systems may underestimate populations and hence the survival and colonization of genetically marked bacteria.

Crude oil degradation efficiency of a recombinant Acinetobacter baumannii strain and its survival in crude oil-contaminated soil microcosm

“…strain B13 [30]. There are certain shortcomings in using PCR-based fingerprinting techniques with repeat element primers to monitor survivability of the introduced strains [3,8]. The use of reporter genes for efficient monitoring of these biological agents circumvents the mentioned shortcomings of conventional PCR-based DNA fingerprinting techniques [3,6,29].…”

“…There are certain shortcomings in using PCR-based fingerprinting techniques with repeat element primers to monitor survivability of the introduced strains [3,8]. The use of reporter genes for efficient monitoring of these biological agents circumvents the mentioned shortcomings of conventional PCR-based DNA fingerprinting techniques [3,6,29]. Thus, the aim of the present investigation was to evaluate the stability of a reporter gene in wild strain A. baumannii S30 and observe the survival and effect of the foreign insert on the metabolic capability of the wild-type strain.…”

“…The bioaugmentation of microbes with high efficiency for such degradation is thus a proven technique for efficient bioremediation. Genetically modified organisms (GMOs) with increased capacity to degrade and tolerate toxic compounds present in crude oil can be an alternative choice [3]. However, use of GMOs is an environmental concern and subjected to widespread controversy.…”

Applicability of the 16S–23S rDNA internal spacer for PCR detection of the phytostimulatory PGPR inoculant Azospirillum lipoferum CRT1 in field soil

“…These plant growth-promoting rhizobacteria (PGPR) have been extensively used over the past 30 years in field inoculation to improve crop yield and quality (Okon and Labandera-Gonzalez 1994). Evaluating survival and fate of PGPR inoculants is part of this process (Tsushima et al 1995;Mahaffee et al 1997;El Zemrany et al 2006).…”

Development of a strain-specific genomic marker for monitoring a Bacillus subtilis biocontrol strain in the rhizosphere of tomato

“…For detection of wild-type microorganisms released in the environment, molecular monitoring methods based on PCR techniques using natural genome polymorphism have largely facilitated the discrimination at strain level, thus minimizing the drawbacks of introduced markers (Mahaffee et al, 1997). Natural polymorphism can be detected by use of the random amplified polymorphic DNA (RAPD) procedure (Williams et al, 1990), in which very small amounts of total DNA are submitted to PCR using a single synthetic oligonucleotide of a random sequence as a primer.…”