CFU Density Research Articles

Survival of Beauveria brongniartii in the soil after application as a biocontrol agent against the European cockchafer Melolontha melolontha

The survival of the entomopathogenic fungusBeauveria brongniartii and its efficacyagainst the European cockchafer, Melolontha melolontha, was examined over 16months after application as a biologicalcontrol agent in different soil types inSwitzerland. In the absence of M. melolontha,the reduction of the number of the CFU in thesoil was nearly 90%. In soils with highorganic content and increased catalase activitythe decline in the CFU density of B.brongniartii was more pronounced. At siteswhere M. melolontha was present, thesurvival was significantly longer. Theapplication of B. brongniartii resultedin a maximal infection rate of 75% in the M. melolontha population. The number of CFU ofB. brongniartii in the soil started todecrease only when the M. melolonthapopulation was reduced during the epizootic(reduction of 46% on average). The rapiddecrease of the fungus in the absence of thehost gives further evidence of the highspecificity of the fungus and that asaprophytic multiplication without the host isunlikely. The fungus was isolated on aselective medium as well as by baiting withGalleria mellonella. The latter methoddid not reveal the significant differencesmentioned above, which leads to the assumptionthat the two methods may select for differentfungal strains.

Colonization pattern of primary tomato roots by Pseudomonas fluorescens A6RI characterized by dilution plating, flow cytometry, fluorescence, confocal and scanning electron microscopy

Early colonization of primary tomato roots, grown in vitro, by Pseudomonas fluorescens A6RI, introduced by seed bacterization, was monitored for 7 days in three different root zones (zone A, apex + elongation + young hairy zone; zone B, hairy zone; zone C, old hairy zone + collar). Bacterial quantification was assessed by enumeration of (i) colony forming units (cfu) after dilution plating and of (ii) total bacterial cells by flow cytometry. Bacterial distribution and organization in the root zones were analyzed by fluorescence, confocal and scanning electron microscopy. For all sampling dates and zones, the densities of total bacterial cells were significantly higher than those of the cfu. The kinetics of cfu densities varied according to the root zone. Their density decreased with time in zone A, while no variation with time was recorded in zones B and C. Densities of total bacterial cells did not show any significant temporal variation for any of the root zones. Microscopic analyses allowed the characterization of the distribution and organizational patterns of the bacterial cells according to time and space. In 3-day-old plants, bacteria were mostly present as single cells and were evenly distributed in the two root zones analyzed (A and B). In 5- and 7-day-old plants, distribution and organization differed according to the root zone. In zone A, only few single cells were observed, whereas zones B and C were mostly covered by cells localized between epidermal root cells and organized in pairs and strings, respectively.

Survival of Conidia of Clonostachys rosea on Stored Barley Seeds and Their Biocontrol Efficacy Against Seed-borne Bipolaris sorokiniana

Survival and biocontrol activity of Clonostachys rosea (isolate IK726) conidia during storage on barley seeds were investigated. The initial density of colony forming conidia on seed was 4 2 10 3 to 9 2 10 4 colony forming units (cfu)/seed. After 5 months storage at 4°C, the density decreased by less than one order of magnitude and the biocontrol efficacy against seedling blight caused by seed-borne Bipolaris sorokiniana was maintained at a significantly high level ( > 80% disease reduction) for > 5 months. Conidial survival on seeds stored at 20°C declined more rapidly than at 4°C, and biocontrol efficacy was significantly reduced after 3-5 months. However, conidia produced on solid media over 20 days survived better than conidia produced in liquid culture and conidia from solid media produced over 12 days. In contrast, when seeds treated with conidia were packed with silica gel and stored at 20°C, the cfu density decreased by less than one order of magnitude after 5 months and the biocontrol efficacy was still high after 6 months. A dose-response curve revealed that 103 cfu/seed were needed for 80% control of seedling blight. Similar control was obtained in storage experiments when approximately 103 cfu/seed were recovered from seed, indicating that conidia which survived also retained a high ability to control disease.

Staphylococcal Colonization in Atopic Dermatitis Treatment with Topical Tacrolimus (Fk506)

Staphylococcal Colonization in Atopic Dermatitis Treatment with Topical Tacrolimus (Fk506)

Studies on the Growth of Escherichia coli O157:H7 Strains at 45.5°C

The objectives of the present report were to examine the ability of 18 strains of Escherichia coli O157:H7 to grow in EC broth at 42.4, 43.5, 44.5, and 45.5°C, and to document the incidence of phenotypic variants present in low numbers that are capable of growth at 45.5°C in EC broth. Among the 18 strains of E. coli O157:H7 studied, only 3 were capable of producing turbid growth with gas formation in EC broth at 45.5°C with 1 × 102 initial CFU/ml. Higher initial densities of CFU resulted in turbid growth and gas formation in EC broth at 45.5°C with all strains. The presence of bile salts #3 in EC broth was found to be inhibitory at 45.5° C. All 18 strains were found to be capable of growth at 45.5°C in nonselective media. The ability of at least one sensitive strain to grow in EC broth at 45.5°C was found to be dependent on the initial number of CFU/ml. Prior growth of cells of a sensitive strain in EC broth at 45.5°C from a cell density of 2.0 × 107 to 8.0 × 107 CFU/ml followed by removal of cells and reinoculation at a cell density of 2.0 × 106 CFU/ml resulted in growth at 45.5°C that did not occur without such conditioning of the inhibitory medium. These results indicate that the ability of most strains of E. coli O157:H7 to grow in EC broth at 45.5°C is dependent on the initial density of CFU and that at low densities of CFU the ability to initiate growth is dependent on either low numbers of phenotypic variants tolerant to the presence of bile salts #3 in EC broth at 45.5°C or to conditioning of the medium with prior elevated numbers of cells.

Regulation of transformation frequency by exogenous and endogenous growth factors in rat tracheal epithelial cells.

The purpose of our studies was to re-evaluate the rat tracheal epithelial (RTE) transformation system and to identify critical variables that affect the development of enhanced growth variants (EGV). The enhanced growth variant colony, which is a preneoplastic cell variant, is the quantifiable transformation endpoint in RTE cultures. Using a standard protocol the frequency of EGV colony formation was shown to be inversely related to the number of clonogenic cells (CFU) seeded per dish in control cultures as well as in cultures treated with the transforming agent 6-nitrochrysene (6-NC). Experiments showed that the major mechanisms that underlie the CFU density-dependent inhibition of EGV colony formation are depletion of growth factors from and accumulation of autocrine TGF-beta in the media. Thus the cells themselves are creating the selection environment, which allows only the EGVs to survive. The effects of agents such as 6-NC, which increase the frequency of EGV colony formation, are to induce a cellular phenotype that is less susceptible to the selection environment. We showed that TGF-beta-neutralizing antibodies added to the selection media significantly increased EGV colony formation in control cultures but not in 6-NC-exposed cultures. In addition we demonstrated that the development of EGV colonies is much less susceptible to inhibition by (exogenous) TGF-beta in 6-NC-exposed than in control cultures. Thus spontaneous and 6-NC EGV colony formation are distinguishable based on TGF-beta sensitivity. To conduct quantitative cell transformation experiments with RTE cells it is essential that the number of surviving CFU per dish is the same in control and treated cultures. Under the conditions used in the studies described here, 350-500 CFU per culture was found to be the optimum CFU density. Besides 6-NC, agents that have been shown to increase EGV colony frequency under conditions similar to those described here are nitrosamines, NNK, nickel compounds and X-rays.

Residue Placement and Fungicide Effects on Fungal Communities in Conventional and No-Tillage Soils

Residue Placement and Fungicide Effects on Fungal Communities in Conventional and No-Tillage Soils

Potential application of genetic transformants of Fusarium oxysporum f. sp. raphani for assessing fungal autecology

To pursue the behavior of a soil-borne plant pathogen, Fusarium oxysporum f. sp. raphani, in soils, a genetically-transformed form of the pathogen was used as a model fungus. Strain SS1 of the pathogen was transformed to resistance to hygromycin B (Hyg) by introducing the vector plasmid, pDH25, into the fungal chromosome. Transformant PEG-4 among the transformants was chosen as a model inoculum, because it possessed the same abilities of vegetative growth and conidiation as the wild-type strain, as well as considerable resistance to Hyg and mitotic stability of the marker phenotype. Population dynamics of the transformant was periodically monitored after inoculation into a soil. By using Komada's selective medium containing Hyg, PEG-4 could be specifically detected even at a low density of several CFU g −1 soil. Within 3 days after inoculation, the number of transformant propagules increased rapidly within soil aggregatesand then the population remained stable in this soil fraction during 90 days of incubation. Hyg-resistant fusaria recovered from the soil were confirmed to be from propagules of transformant PEG-4 by Southern blot analysis of their genomic DNA. These results suggest that the use of genetic transformants of this Fusarium pathogen makes it possible to monitor sensitively the behavior of the fungus in soils and to assess fungal autecology.