Behavior ofPseudomonas fluorescens within the hydrodynamic boundary layers of surface microenvironments

Phase, darkfield, and computer-enhanced microscopy were used to observe the surface microenvironment of flow cells during bacterial colonization. Microbial behavior was consistent with the assumptions used previously to derive surface colonization kinetics and to calculate surface growth and attachment rates from cell number and distribution. Surface microcolonies consisted of closely packed cells. Each colony contained 2ⁿ cells, where n is the number of cell divisions following attachment. Initially, cells were freely motile while attached, performing circular looping movements within the plane of the solid-liquid interface. Subsequently, cells attached apically, maintained a fixed position on the surface, and rotated. This type of attachment was reversible and did not necessarily lead to the formation of microcolonies. Cells became irreversibly attached by progressing from apical to longitudinal attachment. Longitudinally attached cells increased in length, then divided, separated, moved apart laterally, and slid next to one another. This resulted in tight cell packing and permitted simultaneous growth and adherence. After approximately 4 generations, individual cells emigrated from developing microcolonies to recolonize the surface at new locations. Surface colonization byPseudomonas fluorescens can thus be subdivided into the following sequential colonization phases: motile attachment phase, reversible attachment phase, irreversible attachment phase, growth phase, and recolonization phase.     

Effect of laminar flow velocity on the kinetics of surface recolonization by Mot+ and Mot− Pseudomonas fluorescens

Computer-enhanced microscopy (CEM) was used to monitor bacteria colonizing the inner surfaces of a 1×3 mm glass flow cell. Image analysis provided a rapid and reliable means of measuring microcolony count, microcolony area, and cell motility. The kinetics of motile and nonmotilePseudomonas fluorescens surface colonization were compared at flow velocities above (120m sec^–1) and below (8m sec^–1) the strain's maximum motility rate (85m sec^–1). A direct attachment assay confirmed that flagellated cells undergo initial attachment more rapidly than nonflagellated cells at high and low flow. During continuous-flow slide culture, neither the rate of growth nor the timing of recolonization (cell redistribution within surface microenvironments) were influenced by flow rate or motility. However, the amount of reattachment of recolonizing cells was both flow and motility dependent. At 8m sec^–1 flow, motility increased reattachment sixfold, whereas at 120m sec^–1 flow, motility increased reattachment fourfold. The spatial distribution of recolonizing cells was also influenced by motility. Motile cells dispersed over surfaces more uniformly (mean distance to the nearest neighbor was 47.0m) than nonmotile cells (mean distance was 14.2m) allowing uniform biofilm development through more effective redistribution of cells over the surface during recolonization. In addition, motile cell backgrowth (where cells colonize against laminar flow) occurred four times more rapidly than nonmotile cell backgrowth at low flow (where rate of motility exceeded flow), and twice as rapidly at high flow (where flow exceeded the rate of motility). The observed backgrowth of Mot⁺ cells against high flow could only have occurred as the result of motile attachment behavior. These results confirm the importance of motility as a behavioral mechanism in colonization and provides an explanation for enhanced colonization by motile cells in environments lacking concentration gradients necessary for chemotactic behavior.     

Flagellar Motility Confers Epiphytic Fitness Advantages upon Pseudomonas syringae

The role of flagellar motility in determining the epiphytic fitness of an ice-nucleation-active strain of Pseudomonas syringae was examined. The loss of flagellar motility reduced the epiphytic fitness of a normally motile P. syringae strain as measured by its growth, survival, and competitive ability on bean leaf surfaces. Equal population sizes of motile parental or nonmotile mutant P. syringae strains were maintained on bean plants for at least 5 days following the inoculation of fully expanded primary leaves. However, when bean seedlings were inoculated before the primary leaves had expanded and bacterial populations on these leaves were quantified at full expansion, the population size of the nonmotile derivative strain reached only 0.9% that of either the motile parental or revertant strain. When fully expanded bean primary leaves were coinoculated with equal numbers of motile and nonmotile cells, the population size of a nonmotile derivative strain was one-third of that of the motile parental or revertant strain after 8 days. Motile and nonmotile cells were exposed in vitro and on plants to UV radiation and desiccating conditions. The motile and nonmotile strains exhibited equal resistance to both stresses in vitro. However, the population size of a nonmotile strain on leaves was less than 20% that of a motile revertant strain when sampled immediately after UV irradiation. Epiphytic populations of both motile and nonmotile P. syringae declined under desiccating conditions on plants, and after 8 days, the population size of a nonmotile strain was less than one-third that of the motile parental or revertant strain.     

Motility as an intestinal colonization factor for Campylobacter jejuni

The colonization of the intestinal tract of suckling mice by Campylobacter jejuni was examined by orally challenging the mice with a wild-type strain and several nonmotile mutant strains which were isolated after treating the wild-type strain with mutagens. The wild-type strain had colonized the lower portion of the small intestine, the caecum and the colon 2 d after inoculation. Two nonmotile strains, one of which (M8) had lost all the flagellar structure including the filament, the hook and the basal structure, and the other (M1) which had lost only the filament region, were both cleared from the intestinal tract 2 d after challenge. Another nonmotile strain (M14), which had a complete flagellar structure like that of the wild-type strain, did not colonize and was cleared from the intestinal tract like the other nonmotile and nonflagellated strains. One atypically motile strain (M5), which had a shorter flagellar filament than that of the wild-type strain, colonized the intestinal tract only when mice were challenged with a large inoculum. None of the mice challenged with either the wild-type or any of the mutant strains showed signs of illness. We concluded that motility is an important factor in the colonization of the intestinal tract of suckling mice by C. jejuni.     

Characterizing the adhesion of motile and nonmotile Escherichia coli to a glass surface using a parallel-plate flow chamber

A parallel-plate flow chamber was used to measure the attachment and detachment rates of Escherichia coli to a glass surface at various fluid velocities. The effect of flagella on adhesion was investigated by performing experiments with several E. coli strains: AW405 (motile); HCB136 (nonmotile mutant with paralyzed flagella); and HCB137 (nonmotile mutant without flagella). We compared the total attachment rates and the fraction of bacteria retained on the surface to determine how the presence and movement of the flagella influence transport to the surface and adhesion strength in this dynamic system. At the lower fluid velocities, there was no significant difference in the total attachment rates for the three bacterial strains; nonmotile strains settled at a rate that was of the same order of magnitude as the diffusion rate of the motile strain. At the highest fluid velocity, the effect of settling was minimized to better illustrate the importance of motility, and the attachment rates of both nonmotile strains were approximately five times slower than that of the motile bacteria. Thus, different processes controlled the attachment rate depending on the parameter regime in which the experiment was performed. The fractions of motile bacteria retained on the glass surface increased with increasing velocity, whereas the opposite trend was found for the nonmotile strains. This suggests that the rotation of the flagella enables cells to detach from the surface (at the lower fluid velocities) and strengthens adhesion (at higher fluid velocities), whereas nonmotile cells detach as a result of shear. There was no significant difference in the initial attachment rates of the two nonmotile species, which suggests that merely the presence of flagella was not important in this stage of biofilm development.     

The role of motility in the in vitro attachment of Pseudomonas putida PaW8 to wheat roots

The attachment of motile and non-motile strains of Pseudomonas putida PaW8 to sterile wheat roots was assessed in both non-competitive and intra-specific competitive assays. The motile strain showed significantly greater attachment to wheat roots than non-motile strains in phosphate buffer. Overall, the motile strain attached better than the non-motile strain at 10(6), 10(7) and 10(8) cfu ml(-1) in competitive assays and at 10(6) and 10(7) cfu ml(-1) in non-competitive assays. When attachment was studied in Luria broth no significant difference between motile and non-motile strains was detected. P. putida PaW8 cells marked with the luxAB genes were used to compare direct detection of attached cells by luminometry with indirect detection by dilution plate counts following extraction from root material. Although direct detection permitted a rapid assessment (60 s) of attachment to surfaces, dilution plate counts provided a more sensitive method for quantification of bacteria. The detection limits were approximately 10 cfu root(-1) using dilution plate counts compared with 1000 cfu root(-1) using luminometry. All results highlighted the importance of motility for the attachment of P. putida to plant roots in simple model systems. To take this work further, studies to assess the role of motility using complex non-sterile systems are needed.     

Competitive Advantage Provided by Bacterial Motility in the Formation of Nodules by Rhizobium meliloti

The effect of motility on the competitive success of Rhizobium meliloti in nodule production was investigated. A motile strain formed more nodules than expected when mixed at various unfavorable ratios with either flagellated or nonflagellated nonmotile derivatives. We conclude that motility confers a selective advantage on rhizobia when competing with nonmotile strains.     

Rhizosphere Selection of Highly Motile Phenotypic Variants of Pseudomonas fluorescens with Enhanced Competitive Colonization Ability

Phenotypic variants of Pseudomonas fluorescens F113 showing a translucent and diffuse colony morphology show enhanced colonization of the alfalfa rhizosphere. We have previously shown that in the biocontrol agent P. fluorescens F113, phenotypic variation is mediated by the activity of two site-specific recombinases, Sss and XerD. By overexpressing the genes encoding either of the recombinases, we have now generated a large number of variants (mutants) after selection either by prolonged laboratory cultivation or by rhizosphere passage. All the isolated variants were more motile than the wild-type strain and appear to contain mutations in the gacA and/or gacS gene. By disrupting these genes and complementation analysis, we have observed that the Gac system regulates swimming motility by a repression pathway. Variants isolated after selection by prolonged cultivation formed a single population with a swimming motility that was equal to the motility of gac mutants, being 150% more motile than the wild type. The motility phenotype of these variants was complemented by the cloned gac genes. Variants isolated after rhizosphere selection belonged to two different populations: one identical to the population isolated after prolonged cultivation and the other comprising variants that besides a gac mutation harbored additional mutations conferring higher motility. Our results show that gac mutations are selected both in the stationary phase and during rhizosphere colonization. The enhanced motility phenotype is in turn selected during rhizosphere colonization. Several of these highly motile variants were more competitive than the wild-type strain, displacing it from the root tip within 2 weeks.     

Redox Interactions between Saccharomyces cerevisiae and Saccharomyces uvarum in Mixed Culture under Enological Conditions

Wine yeast starters that contain a mixture of different industrial yeasts with various properties may soon be introduced to the market. The mechanisms underlying the interactions between the different strains in the starter during alcoholic fermentation have never been investigated. We identified and investigated some of these interactions in a mixed culture containing two yeast strains grown under enological conditions. The inoculum contained the same amount (each) of a strain of Saccharomyces cerevisiae and a natural hybrid strain of S. cerevisiae and Saccharomyces uvarum. We identified interactions that affected biomass, by-product formation, and fermentation kinetics, and compared the redox ratios of monocultures of each strain with that of the mixed culture. The redox status of the mixed culture differed from that of the two monocultures, showing that the interactions between the yeast strains involved the diffusion of metabolite(s) within the mixed culture. Since acetaldehyde is a potential effector of fermentation, we investigated the kinetics of acetaldehyde production by the different cultures. The S. cerevisiae-S. uvarum hybrid strain produced large amounts of acetaldehyde for which the S. cerevisiae strain acted as a receiving strain in the mixed culture. Since yeast response to acetaldehyde involves the same mechanisms that participate in the response to other forms of stress, the acetaldehyde exchange between the two strains could play an important role in inhibiting some yeast strains and allowing the growth of others. Such interactions could be of particular importance in understanding the ecology of the colonization of complex fermentation media by S. cerevisiae.     

phlF− mutant of Pseudomonas fluorescens J2 improved 2,4-DAPG biosynthesis and biocontrol efficacy against tomato bacterial wilt

Pseudomonas fluorescens J2 can produce 2,4-diacetylphloroglucinol (2,4-DAPG) as the main antibiotic compound and effectively inhibits the wilt pathogens Ralstonia solanacearum and Fusarium oxysporum. The phlF which negatively regulates the 2,4-DAPG synthesis in strain J2 was disrupted by homologous recombination to construct a mutant strain J2-phlF⁻. The mutant J2-phlF⁻ produced much more 2,4-DAPG and showed higher inhibitory effect on R. solanacearum than the wild type strain J2 in vitro. The mutant J2-phlF⁻ also showed more colonization of tomato roots and higher inhibition to R. solanacearum in soil than wild type strain J2. The biocontrol efficiency of mutant J2-phlF⁻ was higher against tomato bacterial wilt than wild type strain J2, but the differences were not significant. However, the application of both strains with organic fertilizer improved the colonization and biocontrol efficiency against tomato bacterial wilt and mutant strain J2-phlF⁻ showed higher biocontrol efficiency against tomato bacterial wilt than wild type strain J2. Both strains, J2 and J2-phlF⁻, could also promote the growth of tomato plants.     

Effect of transposon-induced motility mutations on colonization of the host light organ by Vibrio fischeri.

Vibrio fischeri is found both as a free-living bacterium in seawater and as the specific, mutualistic light organ symbiont of several fish and squid species. To identify those characteristics of symbiosis-competent strains that are required for successful colonization of the nascent light organ of juvenile Euprymna scolopes squids, we generated a mutant pool by using the transposon Mu dI 1681 and screened this pool for strains that were no longer motile. Eighteen independently isolated nonmotile mutants that were either flagellated or nonflagellated were obtained. In contrast to the parent strain, none of these nonmotile mutants was able to colonize the juvenile squid light organ. The flagellated nonmotile mutant strain NM200 possessed a bundle of sheathed polar flagella indistinguishable from that of the wild-type strain, indicating that the presence of flagella alone is not sufficient for colonization and that it is motility itself that is required for successful light organ colonization. This study identifies motility as the first required symbiotic phenotype of V. fischeri.     

Root-to-Root Travel of the Beneficial Bacterium Azospirillum brasilense

The root-to-root travel of the beneficial bacterium Azospirillum brasilense on wheat and soybean roots in agar, sand, and light-textured soil was monitored. We used a motile wild-type (Mot⁺) strain and a motility-deficient (Mot^-) strain which was derived from the wild-type strain. The colonization levels of inoculated roots were similar for the two strains. Mot⁺ cells moved from inoculated roots (either natural or artificial roots in agar, sand, or light-textured soil) to noninoculated roots, where they formed a band-type colonization composed of bacterial aggregates encircling a limited part of the root, regardless of the plant species. The Mot^- strain did not move toward noninoculated roots of either plant species and usually stayed at the inoculation site and root tips. The effect of attractants and repellents was the primary factor governing the motility of Mot⁺ cells in the presence of adequate water. We propose that interroot travel of A. brasilense is an essential preliminary step in the root-bacterium recognition mechanism. Bacterial motility might have a general role in getting Azospirillum cells to the site where firmer attachment favors colonization of the root system. Azospirillum travel toward plants is a nonspecific active process which is not directly dependent on nutrient deficiency but is a consequence of a nonspecific bacterial chemotaxis, influenced by the balance between attractants and possibly repellents leaked by the root.     

Quantification of 2,4-Diacetylphloroglucinol-Producing Pseudomonas fluorescens Strains in the Plant Rhizosphere by Real-Time PCR

A real-time PCR SYBR green assay was developed to quantify populations of 2,4-diacetylphloroglucinol (2,4-DAPG)-producing (phlD⁺) strains of Pseudomonas fluorescens in soil and the rhizosphere. Primers were designed and PCR conditions were optimized to specifically amplify the phlD gene from four different genotypes of phlD⁺ P. fluorescens. Using purified genomic DNA and genomic DNA extracted from washes of wheat roots spiked with bacteria, standard curves relating the threshold cycles (C_Ts) and copies of the phlD gene were generated for P. fluorescens strains belonging to genotypes A (Pf-5), B (Q2-87), D (Q8r1-96 and FTAD1R34), and I (FTAD1R36). The detection limits of the optimized real-time PCR assay were 60 to 600 fg (8 to 80 CFU) for genomic DNA isolated from pure cultures of P. fluorescens and 600 fg to 6.0 pg (80 to 800 CFU, corresponding to log 4 to 5 phlD⁺ strain CFU/rhizosphere) for bacterial DNA extracted from plant root washes. The real-time PCR assay was utilized to quantify phlD⁺ pseudomonads in the wheat rhizosphere. Regression analysis of population densities detected by real-time PCR and by a previously described phlD-specific PCR-based dilution endpoint assay indicated a significant linear relationship (P = 0.0016, r² = 0.2). Validation of real-time PCR assays with environmental samples was performed with two different soils and demonstrated the detection of more than one genotype in Quincy take-all decline soil. The greatest advantage of the developed real-time PCR is culture independence, which allows determination of population densities and the genotype composition of 2,4-DAPG producers directly from the plant rhizospheres and soil.     

Campylobacter jejuni Strains Compete for Colonization in Broiler Chicks

Campylobacter jejuni isolates possess multiple adhesive proteins termed adhesins, which promote the organism's attachment to epithelial cells. Based on the proposal that one or more adhesins are shared among C. jejuni isolates, we hypothesized that C. jejuni strains would compete for intestinal and cecal colonization in broiler chicks. To test this hypothesis, we selected two C. jejuni strains with unique SmaI pulsed-field gel electrophoresis macrorestriction profiles and generated one nalidixic acid-resistant strain (the F38011 Nal^r strain) and one streptomycin-resistant strain (the 02-833L Str^r strain). In vitro binding assays revealed that the C. jejuni F38011 Nal^r and 02-833L Str^r strains adhered to LMH chicken hepatocellular carcinoma epithelial cells and that neither strain influenced the binding potential of the other strain at low inoculation doses. However, an increase in the dose of the C. jejuni 02-833L Str^r strain relative to that of the C. jejuni F38011 Nal^r strain competitively inhibited the binding of the C. jejuni F38011 Nal^r strain to LMH cells in a dose-dependent fashion. Similarly, the C. jejuni 02-833L Str^r strain was found to significantly reduce the efficiency of intestinal and cecal colonization by the C. jejuni F38011 Nal^r strain in broiler chickens. Based on the number of bacteria recovered from the ceca, the maximum number of bacteria that can colonize the digestive tracts of chickens may be limited by host constraints. Collectively, these data support the hypothesis that C. jejuni strains compete for colonization in chicks and suggest that it may be possible to design novel intervention strategies for reducing the level at which C. jejuni colonizes the cecum.     

Mechanisms and Rates of Bacterial Colonization of Sinking Aggregates

Quantifying the rate at which bacteria colonize aggregates is a key to understanding microbial turnover of aggregates. We used encounter models based on random walk and advection-diffusion considerations to predict colonization rates from the bacteria's motility patterns (swimming speed, tumbling frequency, and turn angles) and the hydrodynamic environment (stationary versus sinking aggregates). We then experimentally tested the models with 10 strains of bacteria isolated from marine particles: two strains were nonmotile; the rest were swimming at 20 to 60 μm s⁻¹ with different tumbling frequency (0 to 2 s⁻¹). The rates at which these bacteria colonized artificial aggregates (stationary and sinking) largely agreed with model predictions. We report several findings. (i) Motile bacteria rapidly colonize aggregates, whereas nonmotile bacteria do not. (ii) Flow enhances colonization rates. (iii) Tumbling strains colonize aggregates enriched with organic substrates faster than unenriched aggregates, while a nontumbling strain did not. (iv) Once on the aggregates, the bacteria may detach and typical residence time is about 3 h. Thus, there is a rapid exchange between attached and free bacteria. (v) With the motility patterns observed, freely swimming bacteria will encounter an aggregate in <1 day at typical upper-ocean aggregate concentrations. This is faster than even starving bacteria burn up their reserves, and bacteria may therefore rely solely on aggregates for food. (vi) The net result of colonization and detachment leads to a predicted equilibrium abundance of attached bacteria as a function of aggregate size, which is markedly different from field observations. This discrepancy suggests that inter- and intraspecific interactions among bacteria and between bacteria and their predators may be more important than colonization in governing the population dynamics of bacteria on natural aggregates.     

Attachment of Agrobacteria to Grape Cells

The presence of the Ti plasmid favorably influences the attachment of agrobacteria to grape callus cells, especially during the early stages of a 2-h incubation. Agrobacterium strains attached to a similar extent to both the crown gall-resistant cultivar (Catawba), Vitis labruscana, and the crown gall-susceptible cultivar (Chancellor), Vitis sp. Attachment of the virulent strain to grape callus cells is blocked by the avirulent strain HLB-2 in both the tissue culture cell suspension and the seedling root systems.     

Rhizoremediation of Trichloroethylene by a Recombinant, Root-Colonizing Pseudomonas fluorescens Strain Expressing Toluene ortho-Monooxygenase Constitutively

Trichloroethylene (TCE) was removed from soils by using a wheat rhizosphere established by coating seeds with a recombinant, TCE-degrading Pseudomonas fluorescens strain that expresses the tomA⁺ (toluene o-monooxygenase) genes from Burkholderia cepacia PR1₂₃(TOM_23C). A transposon integration vector was used to insert tomA⁺ into the chromosome of P. fluorescens 2-79, producing a stable strain that expressed constitutively the monooxygenase at a level of 1.1 nmol/min · mg of protein (initial TCE concentration, 10 μM, assuming that all of the TCE was in the liquid) for more than 280 cell generations (36 days). We also constructed a salicylate-inducible P. fluorescens strain that degraded TCE at an initial rate of 2.6 nmol/min · mg of protein in the presence of 10 μM TCE [cf. B. cepacia G4 PR1₂₃(TOM_23C), which degraded TCE at an initial rate of 2.5 nmol/min · mg of protein]. A constitutive strain, P. fluorescens 2-79TOM, grew (maximum specific growth rate, 0.78 h⁻¹) and colonized wheat (3 × 10⁶ CFU/cm of root) as well as wild-type P. fluorescens 2-79 (maximum specific growth rate, 0.77 h⁻¹; level of colonization, 4 × 10⁶ CFU/cm of root). Rhizoremediation of TCE was demonstrated by using microcosms containing the constitutive monooxygenase-expressing microorganism, soil, and wheat. These closed microcosms degraded an average of 63% of the initial TCE in 4 days (20.6 nmol of TCE/day · plant), compared to the 9% of the initial TCE removed by negative controls consisting of microcosms containing wild-type P. fluorescens 2-79-inoculated wheat, uninoculated wheat, or sterile soil.     

The role of bacterial motility in the survival and spread of Pseudomonas fluorescens in soil and in the attachment and colonisation of wheat roots

Motile and non-motile strains of Pseudomonas fluorescens SBW25 were constructed using different combinations of the lacZY, xylE and aph marker genes which allowed their detection and differentiation in soil, root and seed samples. The survival of motile and non-motile strains was investigated in both non-competitive and competitive assays in water and non-sterile soil. Although there was no difference between strains in water, the motile strain survived in significantly greater numbers than the non-motile strain after 21 days in soil. There was no significant difference between competitive assays, where motile and non-motile cells were co-inoculated into soil, and non-competitive assays where strains were inoculated separately. Bacterial survival decreased as matric potential increased from -224 to -17 kPa but matric potential had no significant effect on motile compared to non-motile strains. Vertical spread of both motile and non-motile strains was detected 6.4 mm from the inoculum zone after 14 days in the absence of percolating water. There was no significant difference, for either strain, in distance moved from the inoculum zone after 14, 26 or 40 days. The motile strain had a significant advantage in attachment to sterile wheat roots in both non-competitive and competitive studies. When the spatial colonisation of wheat root systems was assessed in non-sterile soil, there was no significant difference between the motile and non-motile strain from either seed or soil inoculum. However, when the whole root system was assessed as one sample unit, differences could be detected. Bacterial motility could contribute to survival in soil and the initial phase of colonisation, where attachment and movement onto the root surface are important.     

Swimming Marine Synechococcus Strains with Widely Different Photosynthetic Pigment Ratios Form a Monophyletic Group

Unicellular marine cyanobacteria are ubiquitous in both coastal and oligotrophic regimes. The contribution of these organisms to primary production and nutrient cycling is substantial on a global scale. Natural populations of marine Synechococcus strains include multiple genetic lineages, but the link, if any, between unique phenotypic traits and specific genetic groups is still not understood. We studied the genetic diversity (as determined by the DNA-dependent RNA polymerase rpoC1 gene sequence) of a set of marine Synechococcus isolates that are able to swim. Our results show that these isolates form a monophyletic group. This finding represents the first example of correspondence between a physiological trait and a phylogenetic group in marine Synechococcus. In contrast, the phycourobilin (PUB)/phycoerythrobilin (PEB) pigment ratios of members of the motile clade varied considerably. An isolate obtained from the California Current (strain CC9703) displayed a pigment signature identical to that of nonmotile strain WH7803, which is considered a model for low-PUB/PEB-ratio strains, whereas several motile strains had higher PUB/PEB ratios than strain WH8103, which is considered a model for high-PUB/PEB-ratio strains. These findings indicate that the PUB/PEB pigment ratio is not a useful characteristic for defining phylogenetic groups of marine Synechococcus strains.     

Behavior of bacterial stream populations within the hydrodynamic boundary layers of surface microenvironments

Phase and computer-enhanced microscopy were used to observe the surface microenvironment of continuous-flow slide cultures during microbial colonization and to document the diversity of bacterial colonization maneuvers among natural stream populations. Surface colonization involved 4 discrete types of cell movement, which were designated as packing, spreading, shedding, and rolling maneuvers. Each maneuver appeared to be associated with a specific species population within the community. The packing maneuver resulted in the formation of a monolayer of contiguous cells, while spreading maneuvers resulted in a monolayer of adjacent cells. During the shedding maneuver, cells attached perpendicular to the surface and the daughter cells were released. The rate of growth of new daughter cells gradually decreased as the attached mother cell aged. During the rolling maneuver, cells were loosely attached and continuously somersaulted across the surface as they grew and divided. Only those populations with a packing maneuver conformed fully to the assumptions of kinetics used previously to calculate growth and attachment rates from cell number and distribution. Consequently, these kinetics are not applicable to stream communities unless fluorescent antisera are used to study specific species populations within natural communities. Virtually all of the cells that attached to the surface were viable and underwent cell division. The abundance of unicells on surfaces incubated in situ was thus primarily the consequence of bacterial colonization behavior (shedding and spreading maneuvers) rather than the adhesion of dead or moribund cells.