scholarly journals Diverse Microhabitats Experienced by Halomonas variabilis on Salt-Secreting Leaves

2012 ◽  
Vol 79 (3) ◽  
pp. 845-852 ◽  
Author(s):  
Adrien Y. Burch ◽  
Omri M. Finkel ◽  
Juliana K. Cho ◽  
Shimshon Belkin ◽  
Steven E. Lindow
Keyword(s):  
Salt Concentration ◽  
Fluorescent Protein ◽  
Leaf Surface ◽  
Green Fluorescent Protein Fluorescence ◽  
Protein Fluorescence ◽  
Content Type ◽  
Osmotic Tolerance ◽  
Leaf Surfaces ◽  
High Concentrations ◽  
Green Fluorescent

ABSTRACTThe leaf surfaces of the salt-excreting treeTamarix aphyllaharbor a wide diversity of halophilic microorganisms, includingHalomonassp., but little is known of the factors that shape community composition in this extreme habitat. We isolated a strain ofHalomonas variabilisfrom the leaf surface ofT. aphyllaand used it to determine the heterogeneity of salt concentrations experienced by bacteria in this environment. This halophilic strain was transformed with aproU::gfpreporter gene fusion, the fluorescence of which was responsive to NaCl concentrations up to 200 g liter−1. These bioreporting cells were applied toT. aphyllaleaves and were subsequently recovered from dew droplets adhering to the leaf surface. Although cells from within a given dew droplet exhibited similar green fluorescent protein fluorescence, the fluorescence intensity varied between droplets and was correlated with the salt concentration measured in each drop. Growth ofH. variabiliswas observed in all droplets, regardless of the salt concentration. However, cells found in desiccated microniches between dew drops were low in abundance and generally dead. Other bacteria recovered fromT. aphylladisplayed higher desiccation tolerance thanH. variabilis, both in culture and on inoculated plants, despite having lower osmotic tolerance. Thus, theTamarixleaf surface can be described as a salty desert with occasional oases where water droplets form under humid conditions. While halotolerant bacteria such asHalomonasgrow in high concentrations of salt in such wet microniches, other organisms are better suited to survive desiccation in sites that are not wetted.

scholarly journals Effect of sequential C‐terminal tryptophans on green fluorescent protein fluorescence

FEBS Open Bio ◽  
2018 ◽  
Vol 8 (7) ◽  
pp. 1176-1183
Author(s):  
Shirou Tsuchida ◽  
Takumi Kanashiki ◽  
Shuhei Izumiya ◽  
Takuya Ichikawa ◽  
Ryusuke Kurosawa ◽  
...  
Keyword(s):  
Green Fluorescent Protein ◽  
Fluorescent Protein ◽  
Green Fluorescent Protein Fluorescence ◽  
Protein Fluorescence ◽  
Green Fluorescent

scholarly journals Microplate Bioassay for Nisin in Foods, Based on Nisin-Induced Green Fluorescent Protein Fluorescence

2003 ◽  
Vol 69 (7) ◽  
pp. 4214-4218 ◽  
Author(s):  
J. Reunanen ◽  
P. E. J. Saris
Keyword(s):  
Green Fluorescent Protein ◽  
Lactococcus Lactis ◽  
Culture Supernatant ◽  
Fluorescent Protein ◽  
Green Fluorescent Protein Fluorescence ◽  
Regulatory Proteins ◽  
Protein Fluorescence ◽  
Gene Encoding ◽  
Two Component ◽  
Green Fluorescent

ABSTRACT A plasmid coding for the nisin two-component regulatory proteins, NisK and NisR, was constructed; in this plasmid a gfp gene (encoding the green fluorescent protein) was placed under control of the nisin-inducible nisF promoter. The plasmid was transformed into non-nisin-producing Lactococcus lactis strain MG1614. The new strain could sense extracellular nisin and transduce it to green fluorescent protein fluorescence. The amount of fluorescence was dependent on the nisin concentration, and it could be measured easily. By using this strain, an assay for quantification of nisin was developed. With this method it was possible to measure as little as 2.5 ng of pure nisin per ml in culture supernatant, 45 ng of nisin per ml in milk, 0.9 μg of nisin in cheese, and 1 μg of nisin per ml in salad dressings.

scholarly journals Conditional Survival as a Selection Strategy To Identify Plant-Inducible Genes of Pseudomonas syringae

2003 ◽  
Vol 69 (10) ◽  
pp. 5793-5801 ◽  
Author(s):  
Maria L. Marco ◽  
Jennifer Legac ◽  
Steven E. Lindow
Keyword(s):  
Pseudomonas Syringae ◽  
Fluorescent Protein ◽  
Green Fluorescent Protein Fluorescence ◽  
Restriction Pattern ◽  
Selection Strategy ◽  
Gene Promoters ◽  
Protein Fluorescence ◽  
Novel Strategy ◽  
Green Fluorescent ◽  
Bean Leaves

ABSTRACT A novel strategy termed habitat-inducible rescue of survival (HIRS) was developed to identify genes of Pseudomonas syringae that are induced during growth on bean leaves. This strategy is based on the complementation of metXW, two cotranscribed genes that are necessary for methionine biosynthesis and required for survival of P. syringae on bean leaves exposed to conditions of low humidity. We constructed a promoter trap vector, pTrap, containing a promoterless version of the wild-type P. syringae metXW genes. Only with an active promoter fused to metXW on pTrap did this plasmid restore methionine prototrophy to the P. syringae metXW mutant B7MX89 and survival of this strain on bean leaves. To test this method, a partial library of P. syringae genomic DNA was constructed in pTrap and a total of 1,400 B7MX89 pTrap clones were subjected to HIRS selection on bean leaves. This resulted in the enrichment of five clones, each with a unique RsaI restriction pattern of their DNA insert. Sequence analysis of these clones revealed those P. syringae genes for which putative plant-inducible activity could be assigned. Promoter activity experiments with a gfp reporter gene revealed that these plant-inducible gene promoters had very low levels of expression in minimal medium. Based on green fluorescent protein fluorescence levels, it appears that many P. syringae genes have relatively low expression levels and that the metXW HIRS strategy is a sensitive method to detect weakly expressed P. syringae genes that are active on plants. Furthermore, we found that protected sites on the leaf surface provided a higher level of enrichment for P. syringae expressing metXW than exposed sites. Thus, the metXW HIRS strategy should lead to the identification of P. syringae genes that are expressed primarily in these areas on the leaf.

scholarly journals Green Fluorescent Protein-Marked Pseudomonas fluorescens: Localization, Viability, and Activity in the Natural Barley Rhizosphere

1999 ◽  
Vol 65 (10) ◽  
pp. 4646-4651 ◽  
Author(s):  
Bo Normander ◽  
Niels B. Hendriksen ◽  
Ole Nybroe
Keyword(s):  
Green Fluorescent Protein ◽  
Pseudomonas Fluorescens ◽  
Laser Scanning ◽  
Fluorescent Protein ◽  
Green Fluorescent Protein Fluorescence ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Protein Fluorescence ◽  
Dividing Cells ◽  
Green Fluorescent

ABSTRACT The gfp-tagged Pseudomonas fluorescensbiocontrol strain DR54-BN14 was introduced into the barley rhizosphere. Confocal laser scanning microscopy revealed that the rhizoplane populations of DR54-BN14 on 3- to 14-day-old roots were able to form microcolonies closely associated with the indigenous bacteria and that a majority of DR54-BN14 cells appeared small and almost coccoid. Information on the viability of the inoculant was provided by a microcolony assay, while measurements of cell volume, the intensity of green fluorescent protein fluorescence, and the ratio of dividing cells to total cells were used as indicators of cellular activity. At a soil moisture close to the water-holding capacity of the soil, the activity parameters suggested that the majority of DR54-BN14 cells were starving in the rhizosphere. Nevertheless, approximately 80% of the population was either culturable or viable but nonculturable during the 3-week incubation period. No impact of root decay on viability was observed, and differences in viability or activity among DR54-BN14 cells located in different regions of the root were not apparent. In dry soil, however, the nonviable state of DR54-BN14 was predominant, suggesting that desiccation is an important abiotic regulator of cell viability.

scholarly journals Biological Sensor for Sucrose Availability: Relative Sensitivities of Various Reporter Genes

2001 ◽  
Vol 67 (3) ◽  
pp. 1308-1317 ◽  
Author(s):  
William G. Miller ◽  
Maria T. Brandl ◽  
Beatriz Quiñones ◽  
Steven E. Lindow
Keyword(s):  
Reporter Gene ◽  
Fluorescence Intensity ◽  
Fluorescent Protein ◽  
Reporter Genes ◽  
Green Fluorescent Protein Fluorescence ◽  
Average Concentration ◽  
Epifluorescence Microscopy ◽  
Protein Fluorescence ◽  
Wide Range ◽  
Green Fluorescent

ABSTRACT A set of three sucrose-regulated transcriptional fusions was constructed. Fusions p61RYTIR, p61RYlac, and p61RYice contain thescrR sucrose repressor gene and the promoterlessgfp, lacZ, and inaZreporter genes, respectively, fused to the scrY promoter from Salmonella enterica serovar Typhimurium. Cells ofErwinia herbicola containing these fusions are induced only in media amended with sucrose, fructose, or sorbose. While a large variation in sucrose-dependent reporter gene activity was observed in cells harboring all gene fusions, fusions to theinaZ reporter gene yielded a much wider range of activity and were responsive to lower levels of sucrose than eitherlacZ or gfp. The lacZreporter gene was found to be more efficient than gfp, requiring approximately 300-fold fewer cells for a detectable response over all concentrations of sucrose. Similarly, inaZ was found to be more efficient than lacZ, requiring 30-fold fewer cells at 1.45 μM sucrose and 6,100-fold fewer cells at 29 mM sucrose for a quantifiable response. The fluorescence of individual cells containing p61RYTIR was quantified following epifluorescence microscopy in order to relate the fluorescence exhibited by populations of cells in batch cultures with that of individual cells in such cultures. While the mean fluorescence intensity of a population of individual cells increased with increasing concentrations of sucrose, a wide range of fluorescence intensity was seen among individual cells. For most cultures the distribution of fluorescence intensity among individual cells was log-normally distributed, but cells grown in intermediate concentrations of sucrose exhibited two distinct populations of cells, one having relatively low fluorescence and another with much higher fluorescence. When cells were inoculated onto bean leaves, whole-cell ice nucleation andgfp-based biological sensors for sucrose each indicated that the average concentration of sucrose on moist leaf surfaces was about 20 μM. Importantly, the variation in green fluorescent protein fluorescence of biosensor cells on leaves suggested that large spatial variations in sugar availability occur on leaves.

Sign in / Sign up

Export Citation Format

Share Document