A note on the physiological state of epiphytic bacteria compared with planktonic bacteria in two organically-enriched watercourses

Epiphytic bacteria were compared with planktonic bacteria in two watercourses enriched with sewage works effluent. Colony-forming units as a percentage of acridine orange direct counts were higher for epiphytic bacteria in one watercourse. The percentage of cells capable of intracellular reduction of 2-( p -iodophenyl)-3-( p -nitrophenyl)-5-phenyl tetrazolium chloride (INT) to INT-formazan was higher for epiphytic bacteria in both watercourses. The greater viability of epiphytic over planktonic bacteria was less evident, however, than in previously studied non-enriched gravel pit ponds. This suggests that the greater availability of dissolved organic nutrients in the enriched watercourses reduced stress on planktonic bacteria.     

Neustonic versus epiphytic bacteria of eutrophic lake and their biodegradation ability on deltamethrin

This study evaluated biodegradation of the insecticide deltamethrin (1 μg l⁻¹) by pure cultures of neustonic (n = 25) and epiphytic (n = 25) bacteria and by mixed cultures (n = 1), which consisted of a mixture of 25 bacterial strains isolated from the surface microlayer (SM ≈ 250 μm) and epidermis of the Common Reed (Phragmites australis, (Cav.) Trin. ex Steud.) growing in the littoral zone of eutrophic lake Chełmżyńskie. Results indicate that neustonic and epiphytic bacteria are characterized by a similar average capacity to degrade deltamethrin. After a 15-day incubation, bacteria isolated from the surface microlayer reduced the initial concentration of deltamethrin by 60%, while the average effectiveness of the bacteria found on the Common Reed equaled 47%.     

Relations between suspended bacteria, epiphytic bacteria and submerged vegetation over the spring growing season in a calcareous headstream

SUMMARY. 1. Quantitative observations on suspended bacteria, epiphytic bacteria and submerged vegetation, in a calcareous headstream in North Humberside, were made at regular intervals from January to June 1984.
2. Downstream increase in concentration of suspended bacteria was observed along a 135 m richly-vegetated section but not over a 150 m vegetation-free section, hence there was measurable drift loss of bacteria only from within the vegetated section.
3. The total number of epiphytic bacteria within the vegetated section was estimated from the density of epiphytic bacteria per unit dry weight of submerged vegetation and the total dry weight of submerged vegetation.
4. The total number of epiphytic baeteria within the vegetated section decreased frtim 2.5 × 10¹⁵ in January to 0.8 × l0¹⁵ March before increasing to 4.7 × 10¹⁵ by June. This pattern was related to deerease and subsequent increase in density of epiphytic bacteria per unit dry weight of vegetation. The total dry weight of submerged vegetation inereased throughout the study period.
5. The daily drift loss of bacteria from the vegetated section averaged 53% (range 9–132%) of the total epiphytic bacteria, a loss rate which might reasonably be supported by the epiphyte population.
6. The dimensions and volume of suspended bacteria increased, between the source and the downstream limit of the vegetated section, to resemble those of epiphytic bacteria.
7. The results suggested that released epiphytic bacteria might largely be the source of suspended bacteria in this headstream.     

Plant-microbe interactions: identification of epiphytic bacteria and their ability to alter leaf surface permeability

Bacteria were either isolated from leaf surfaces of Hedera helix or obtained from a culture collection in order to analyse their effect on barrier properties of isolated Hedera and Prunus laurocerasus cuticles. On the basis of the 16S rDNA sequences the genera of the six bacterial isolates from Hedera were identified as Pseudomonas sp., Stenotrophomonas sp. and Achromobacter. Water permeability of cuticles isolated from H. helix was measured before and after inoculation with the six bacterial strains. In addition water permeability of cuticles isolated from P. laurocerasus was measured before and after inoculation with the three bacterial strains Pseudomonas aeruginosa, Xanthomonas campestris and Corynebacterium fascians. Rates of water diffusing across isolated cuticles of both species significantly increased by up to 50% after inoculation with all bacterial strains. Obtained results show that epiphytic bacteria have the ability of increasing water permeability of Hedera and Prunus cuticles, which in turn should increase the availability of water and dissolved compounds in the phyllopshere. Consequently, living conditions in the habitat phyllosphere are improved. It can be concluded that the ability to change leaf surface properties will improve epiphytic fitness of leaf surface bacteria.     

Effect of sampling scale on the assessment of epiphytic bacterial populations

Bacterial populations on above-ground plant surfaces were estimated at three different biological scales, including leaflet disks, entire leaflets, and whole plants. The influence of sample scale on the estimation of mean bacterial population size per unit and per gram and on the variability among sampling units was quantified at each scale. Populations were highly variable among sampling units at every scale examined, suggesting that there is no optimal scale at which sample variance is reduced. The distribution of population sizes among sample units was sometimes, but not consistently, described by the lognormal. Regardless of the sampling scale, expression of population sizes on a per gram basis may not reduce variance, because population size was not generally a function of sample unit weight within any single sampling scale. In addition, the data show that scaling populations on a per gram basis does not provide a useful means of comparing population estimates from samples taken at different scales. The implications of these results for designing sampling strategies to address specific issues in microbial ecology are discussed. Correspondence to: L.L. Kinkel     

Seasonal variation in the abundance and heterotrophic activity of suspended bacteria in two lowland rivers

SUMMARY. 1. Water samples were collected over two years from the Yorkshire Ouse and Yorkshire Derwent and the following were measured: (i) concentration of directly-counted bacteria (free-living and particle-bound), (ii) concentration of colony-forming units, (iii) bacterial heterotrophic activity (turnover rate for glucose assimilation), (iv) specific activity (turnover rate per bacterium), (v) a range of environmental variables.
2. The abundance and activity of suspended bacteria showed similar seasonal periodicities in both rivers.
3. Free-living bacteria were usually more numerous than particle-bound bacteria; low concentration of free-living bacteria and maxima of particle-bound bacteria usually occurred in winter.
4. Concentration of colony-forming units varied irregularly, but lowest levels were found in summer.
5. Turnover rate and turnover rate per bacterium showed distinct summer maxima.
6. Multiple-regression analysis was used to relate bacterial variables to subsets (chosen by factor analysis) of environmental variables; up to 89% of variation in bacterial variables was related to the combined effects of variation by variables in the chosen subsets.     

Metabolic footprint of epiphytic bacteria on Arabidopsis thaliana leaves

The phyllosphere, which is defined as the parts of terrestrial plants above the ground, is a large habitat for different microorganisms that show a high extent of adaption to their environment. A number of hypotheses were generated by culture-independent functional genomics studies to explain the competitiveness of specialized bacteria in the phyllosphere. In contrast, in situ data at the metabolome level as a function of bacterial colonization are lacking. Here, we aimed to obtain new insights into the metabolic interplay between host and epiphytes upon colonization of Arabidopsis thaliana leaves in a controlled laboratory setting using environmental metabolomics approaches. Quantitative nuclear magnetic resonance (NMR) and imaging high-resolution mass spectrometry (IMS) methods were used to identify Arabidopsis leaf surface compounds and their possible involvement in the epiphytic lifestyle by relative changes in compound pools. The dominant carbohydrates on the leaf surfaces were sucrose, fructose and glucose. These sugars were significantly and specifically altered after epiphytic leaf colonization by the organoheterotroph Sphingomonas melonis or the phytopathogen Pseudomonas syringae pv. tomato, but only to a minor extent by the methylotroph Methylobacterium extorquens. In addition to carbohydrates, IMS revealed surprising alterations in arginine metabolism and phytoalexin biosynthesis that were dependent on the presence of bacteria, which might reflect the consequences of bacterial activity and the recognition of not only pathogens but also commensals by the plant. These results highlight the power of environmental metabolomics to aid in elucidating the molecular basis underlying plant–epiphyte interactions in situ.     

Supercooling ability in two populations of the land snail Helix pomatia (Gastropoda: Helicidae) and ice-nucleating activity of gut bacteria

The land snail Helix pomatia (Gastropoda: Helicidae) is widely distributed in Northern and Central Europe where it may experience subzero temperatures during winter months. Its supercooling ability was studied in two populations of H. pomatia. One population originated from Southern Sweden (Gotaland) and the other from Central France (Auvergne). In the experimental design, they were acclimated, over 2 weeks, to artificial winter conditions (hibernation, T=5 degrees C). The Swedish snails showed a rather limited supercooling ability (temperature of crystallization, T(c)=-6.4+/-0.8 degrees C), significantly greater, however, than the supercooling capacity of the population from France (T(c)=-4.6+/-1.4 degrees C). In artificial spring conditions (3 months of hibernation followed by a progressive acclimation, over 2 weeks, to activity at T=20 degrees C), both populations exhibited a similar high T(c) (-2.0+/-1.0 degrees C). The lower T(c) of hibernating Swedish snails could be due to a greater loss of body water, accompanied by a higher concentration of solutes in the hemolymph. In both populations, the variation in hemolymph osmolality measured between hibernating (250-270 mOsm kg(-1)) and active (165-215 mOsm kg(-1)) snails may be explained by the variation in body water mass and did not suggest the production of colligative cryoprotectants. Moreover, the three bacterial strains, Buttiauxella sp., Kluyvera sp., and Tatumella sp. (Enterobacteriaceae) which were isolated from fed snails, but absent in starved snails, did not show any ice-nucleating activity at temperatures higher than -9 degrees C. Only the strain Kluyvera sp. initiated nucleation at -9 degrees C. This strain, therefore, is a weak, also termed a Type III or Class C ice-nucleating active bacterium, but with no influence on the supercooling ability of individual snails. In summary, fluctuations in body water mass of hibernating snail populations, triggering changes in osmolyte concentration, rather than the presence of endogenous ice-nucleating-active bacteria, accounts for fluctuations in their T(c).     

Rapid and dissimilar response of ammonia oxidizing archaea and bacteria to nitrogen and water amendment in two temperate forest soils

Biochemical processes relevant to soil nitrogen (N) cycling are performed by soil microorganisms affiliated with diverse phylogenetic groups. For example, the oxidation of ammonia, representing the first step of nitrification, can be performed by ammonia oxidizing bacteria (AOB) and, as recently reported, also by ammonia oxidizing archaea (AOA). However, the contribution to ammonia oxidation of the phylogenetically separated AOA versus AOB and their respective responsiveness to environmental factors are still poorly understood. The present study aims at comparing the capacity of AOA and AOB to momentarily respond to N input and increased soil moisture in two contrasting forest soils. Soils from the pristine Rothwald forest and the managed Schottenwald forest were amended with either NH(4)(+)-N or NO(3)(-)-N and were incubated at 40% and 70% water-filled pore space (WFPS) for four days. Nitrification rates were measured and AOA and AOB abundance and community composition were determined via quantitative PCR (qPCR) and terminal restriction length fragment polymorphism (T-RFLP) analysis of bacterial and archaeal amoA genes. Our study reports rapid and distinct changes in AOA and AOB abundances in the two forest soils in response to N input and increased soil moisture but no significant effects on net nitrification rates. Functional microbial communities differed significantly in the two soils and responded specifically to the treatments during the short-term incubation. In the Rothwald soil the abundance and community composition of AOA were affected by the water content, whereas AOB communities responded to N amendment. In the Schottenwald soil, by contrast, AOA responded to N addition. These results suggest that AOA and AOB may be selectively influenced by soil and management factors.     

Physicochemical and in situ observations on the adhesion of gliding bacteria to surfaces

The ability of Flexibacter BH3 to adhere to solid surfaces and to overcome the horizontal drag involved in gliding across the surfaces was considered in terms of the Stefan adhesion principle. The extracellular slime produced by Flexibacter BH3 was suitable as a Stefan adhesive because it exhibited viscous properties characteristic of a linear colloid, increasing the adhesiveness of the bacterium but allowing translational motion across the surface. The water-soluble slime was a glycoprotein, containing glucose, fucose, galactose and some uronic acid. Vesicles and tubules on the outer surface of Flexibacter BH3 possessed trilaminar membranes, contained 2-keto-3-deoxyoctonate (KDO), and showed identity with phenol-extracted lipopolysaccharide (LPS) in gel-diffusion tests.Sections of Flexibacter BH3 gliding on a gold film overlaying an agar medium reveraled a highly convuluted cell envelope outer membrane, portions of which closely conformed to the microcontours of the gold surface. Possible mechanisms of gliding are discussed in relation to this close association with solid surface features, to the finding that flexibility and spiral motion are not essential for gliding, and to evidence revealing the extrusion of slime in advance of pathfinder bacteria.Abbreviations used KDO 2-keto-3-deoxyoctonate - LPS lipopolysaccharide     

A regression technique for the estimation of epiphytic invertebrate populations*

SUMMARY. 1. A regression method is proposed for the estimation of populations of epiphytic invertebrates. Small samples of macrophytes and attached animals are taken by gentle enclosure. Regression analysis is used to relate the number of animals collected to the macrophyte species composition and ibiomass in these small samples. These relationships estimate the number of organisms of each taxon per unit mass of each macrophyte species. Areal population density is estimated by multiplication of macrophyte mass-specific invertebrate density by standing macrophyte biomass. 2. The regression method yields population density estimates several times greater than the best of current methods for several fauna. Differences are most pronounced for active organisms such as water mites, amphipods, cladocerans, copepods, lepidopterans, ostracods, and trichopterans. 3. Precision levels obtained using the regression method are comparable to other techniques. The regression technique automatically provides estimates of macrophyte species-specific colonization density and the abundance of organisms swimming among macrophytes in littoral areas.     

Quantitative assessment of hemolymph metabolites in two physiological states and two populations of the land snail Helix pomatia

Hemolymph metabolite composition in ectothermic species is mainly constrained by trophic and climatic habitat conditions. In temperate regions, ectothermic species have to face subzero temperatures in winter, to which they typically respond with a state of inactivity. With use of ultra-performance liquid chromatography and gas chromatography-mass spectrometry techniques, we investigated the hemolymph metabolite composition of the land snail Helix pomatia with respect to physiological states (activity and hibernation) in a mountain population (800 m above sea level) and a valley population (150 m above sea level) in Germany. The dry masses of active snails as well as the saccharide and amino acid concentrations in active snails were higher in the mountain population than in the valley population. These differences between populations might reflect differences in microhabitat conditions, such as climate and vegetal food, and consequent differences in metabolic activity. Galactose was the most abundant component in hemolymph besides glucose. Both saccharides might indicate glycolytic activity, which could provide energy for locomotion and foraging. In hibernation, glutamate, α-alanine, glycine, aspartate, serine, homoserine, hydroxyproline, glycerol, and triglycerides were accumulated in both populations. The concentrations were correlated with a decrease in body supercooling point. Therefore, these metabolites might have a role in the cold hardiness of H. pomatia that should be further investigated in a functional study.     

Physiological state of epiphytic bacteria on submerged stems of the reed Phragmites australis compared with planktonic bacteria in gravel-pit ponds

Suspensions of epiphytic bacteria from submerged stems of Phragmites australis , collected from a gravel-pit pond, were prepared by treatment in a stomacher and by brushing. The two procedures were equally successful in dislodging bacteria. These epiphytic bacteria were compared with planktonic bacteria in water samples from within the reed bed. Colony-forming units (cfu) as a percentage of acridine-orange direct counts (AODCs), percentage of cells capable of intracellular reduction of 2-( p -iodophenyl)-3-( p -nitrophenyl)-5-phenyl tetrazolium chloride (INT) to INT-formazan, percentage of cells able to form microcolonies, and cell length were all greater for epiphytic bacteria. Epiphytic bacteria of P. australis from six further gravel-pit ponds were also compared with bacterioplankton; cfu as a percentage of AODCs, and percentage of cells capable of INT reduction were again greater for epiphytic bacteria. Because the epiphytic bacteria in these non-organically-enriched, gravel-pit ponds were physiologically different from the planktonic bacteria it is suggested that there was not continual casual exchange, by largely identical bacterial cells, between the epiphytic and planktonic mode. Instead, epiphytic and planktonic populations were independent of each other and/or if there was exchange then bacteria were more successful while in the attached mode, perhaps because of greater organic-nutrient availability at the stem surface.     

Temporal variation in density of epiphytic bacteria on submerged vegetation in a calcareous stream

The density of epiphytic bacteria on (1) leaves, (2) stems and petioles, and (3) whole shoots of submerged Nasturtium officinale, Apium nodiflorum and Glyceria fluitans was determined from January to June 1984 in a calcareous stream in North Humberside. The density on leaves was less than on stems and petioles; reasons for this are suggested. The density on all three species decreased from late January to February-March and then increased to June. This temporal change was related to environmental variables (e. g. negative correlation with discharge and positive correlation with temperature), a situation which contrasts with Baker & Orr (1986) who found no such relationships in a Southern England chalk stream.     

Multiple regions of internalin B contribute to its ability to turn on the Ras-mitogen-activated protein kinase pathway

Internalin B (InlB) is a protein present on the surface of Listeria monocytogenes that mediates bacterial entry into mammalian cells. It is thought that InlB acts by binding directly to the hepatocyte growth factor (HGF) receptor, present on the surface of host cells. Binding of InlB to the HGF receptor results in mitogen-activated protein (MAP) kinase and phosphoinositide 3-kinase activation, followed by changes in the organization of the actin cytoskeleton. Here we have compared signaling by HGF and InlB. Whereas stimulation with equivalent concentrations of HGF and InlB elicits similar activation of the HGF receptor, we observed striking differences in downstream activation of MAP kinase. InlB leads to a greater activation of the Ras-MAP kinase pathway than does HGF. The leucine-rich repeat region, which was previously shown to be sufficient for binding and activation of the HGF receptor, lacks the ability to super-activate the Ras-MAP kinase pathway. Analysis of a series of deletion mutants suggests that it is the B repeat region between the leucine-rich repeat and GW domains that endows InlB with an increased ability to turn on the Ras-MAP kinase pathway. These unexpected observations suggest that HGF and InlB use alternative mechanisms to turn on cellular signaling pathways.     

Comparative studies on metabolic activity of planktonic,benthic and epiphytic bacteria

Summary Differences have been noted in metabolic activity of bacteria isolated from water, mud and from Canadian pondweed (Elodea canadensis).The epiphytic bacteria showed the greatest metabolic activity in every instance, benthic bacteria were least active and the water isolates were intermediate.Non-chromogenic bacteria were metabolically more active than the chromogens.Casamino acids proved to be the most readily oxidized substrate among the compounds studied. Glucose, succinate, fumarate and gluconate were readily used by most epiphytic and planktonic bacteria. Acetate was intermediate and fructose was the least suitable substrate for these organisms. Casamino acids proved to be the best substrate for all three groups, although the benthic bacteria were less active with this substrate than the former two groups of bacteria. Gluconate, succinate, pyruvate and acetate were intermediate, and glucose and fructose were least suitable for most of the benthic bacteria.     

The effect of drought on photosynthesis in two epiphytic and two terrestrial tropical fern species

Water-withholding for 5 to 7 weeks and subsequent re-watering were made on potted plants of two epiphytic (E) and two terrestrial (T) fern species, which were collected from a seasonal tropical rainforest and had been grown in a screenhouse with 5 % irradiance for 4 months. During the water stress, the two E species completely closed stomata when frond relative water content (RWC) reached about 70 % with fairly constant maximum photochemistry efficiency (F_v/F_m), while the two T species kept partial stomata opening until RWC reached 45 % and reduction in F_v/F_m at the late stage. Also, chlorophyll content as indicated by a spectral reflectance index was gradually reduced in three species. Physiological recovery was completed after 3-d re-watering for the E species, which was more rapid than for the T species. The gas exchange measurements and regression analyses indicated higher photosynthetic water use efficiency in the E species than in the T species.