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S E Lindow

Publications and source records attributed to S E Lindow.

At least 37 records · Page 2Linked to original sources
PubMed1996

Utility of microcosm studies for predicting phylloplane bacterium population sizes in the field.

Population sizes of two ice nucleation-active strains of Pseudomonas syringae were compared on leaves in controlled environments and in the field to determine the ability of microcosm studies to predict plant habitat preferences in the field. The P. syringae strains investigated were the parental strains of recombinant deletion mutant strains deficient in ice nucleation activity that had been field tested for their ability to control plant frost injury. The population size of the P. syringae strains was measured after inoculation at three field locations on up to 40 of the same plant species that were studied in the growth chamber. There was seldom a significant relationship between the mean population size of a given P. syringae strain incubated under either wet or dry conditions in microcosms and the mean population size which could be recovered from the same species when inoculated in the field. Specifically, on some plant species, the population size recovered from leaves in the field was substantially greater than from that species in a controlled environment, while for other plant species field populations were significantly smaller than those observed under controlled conditions. Population sizes of inoculated P. syringae strains, however, were frequently highly positively correlated with the indigenous bacterial population size on the same plant species in the field, suggesting that the ability of a particular plant species to support introduced bacterial strains is correlated with its ability to support large bacterial populations or that indigenous bacteria enhance the survival of introduced strains. Microcosm studies therefore seem most effective at assessing possible differences between parental and recombinant strains under a given environmental regime but are limited in their ability to predict the specific population sizes or plant habitat preferences of bacteria on leaves under field conditions.

Journal Article↗
PubMed1995

Differential effects of growth temperature on ice nuclei active at different temperatures that are produced by cells of Pseudomonas syringae.

The temperature at which ice-nucleating bacteria are grown causes differences of 100- to 10,000-fold in the fraction of cells that nucleate ice at a given temperature (ice nucleation frequency). Ice nucleation frequencies of cells of Pseudomonas syringae grown at temperatures that ranged from 9 to 33 degrees C were examined in order to more accurately characterize physiological effects on ice nuclei active at temperatures of from about -2 to -10 degrees C, the temperature range for this phenotype. Large differences in ice nucleation frequency occurred at all but the lowest assay temperatures in cells of P. syringae grown in the temperature range of 15 to 33 degrees C. These differences in ice nucleation frequency may be attributed, at least in part, to post-translational factors. Because other studies have indicated that ice nuclei active at the lowest assay temperatures may reflect the amount of ice nucleation protein produced, while higher nucleation temperatures reflect aggregates of this ice nucleation protein, data was normalized to the frequency of ice nuclei active at the lowest ice nucleation temperatures (which also correspond to the most abundant nuclei). This was done in order to develop a baseline of comparison for cells grown at different temperatures that more clearly shows possible post-translational effects such as aggregation of the nucleation protein. After this normalization was performed, and in contrast to the results noted above, the number of ice nuclei in cells grown at 9, 15, and 20 degrees C that were active at different assay temperatures was very similar. Differences in ice nucleation frequency that occurred over all assay temperatures in cells grown between 9 and 20 degrees C may be attributed to differences in the total number of nuclei present in the population of cells. The large effects of growth temperature on nucleation frequency have important implications for estimating numbers of ice nucleating bacteria in environmental samples by determining the number of bacterial ice nuclei in such samples.

Bacterial Outer Membrane Proteins↗
PubMed1995

Membrane fluidity as a factor in production and stability of bacterial ice nuclei active at high subfreezing temperatures.

Detailed measurements were made of the rate of appearance of bacterial ice nuclei upon cooling of suspensions of Pseudomonas syringae cells and the disappearance of ice nuclei upon warming of the cells before assay for ice nucleation activity. While no substantial change in numbers of ice nuclei active at either -5 or at -9 degrees C was observed in cells that were grown at temperatures lower than 24 degrees C and cooled to 21 degrees C before assay, large increases in -5 but not -9 degrees C ice nuclei were observed in cells grown at temperatures greater than 24 degrees C. Ice nucleation activity of cells subjected to a decrease in temperature before assay increased immediately upon temperature shift, but 8 to 12 min was required before maximum rates of increase in numbers of ice nuclei were observed. The rate of appearance of ice nuclei in cell suspensions incubated at relatively cold temperatures prior to assay was substantially less than those incubated at temperatures approaching 24 degrees C. Cells rapidly lost ice nucleation activity when warmed to above 27 degrees C before assay; the rate of loss of ice nuclei in cells grown at a given temperature increased rapidly as the temperature to which they were warmed before assay increased. Ice nuclei disappeared most rapidly when cells grown at low temperatures were warmed before assay, suggesting that ice nucleus stability was lower in highly fluid membranes. The logarithm of the half-life of ice nuclei in cells was directly related to the concentration of the membrane fluidizing agent, 2-phenethyl alcohol, in which they were suspended.

Cell Membrane↗
PubMed1995

Enhanced Epiphytic Coexistence of Near-Isogenic Salicylate-Catabolizing and Non-Salicylate-Catabolizing Pseudomonas putida Strains after Exogenous Salicylate Application.

The hypothesis that epiphytic bacterial populations can coexist through nutritional resource partitioning was tested with the near-isogenic bacterial strain pair Pseudomonas putida R20 and R20(pNAH7). The plasmid pNAH7 conferred upon R20 the ability to catabolize salicylate as a sole carbon source in vitro. P. putida R20(pNAH7) also catabolized exogenously applied salicylate in planta and reached a significantly larger epiphytic population size than the near-isogenic parental strain R20 under the same conditions. This supports previous observations that epiphytic populations on plants grown under nitrogen-sufficient conditions are limited by carbon availability. In the absence of exogenous salicylate, R20 and R20(pNAH7) competed for and partitioned endogenous carbon according to their inoculum proportion in replacement series experiments, exhibiting a low level of coexistence. In the presence of exogenous salicylate, however, R20(pNAH7) was solely able to catabolize the additional carbon and achieved a higher level of coexistence with R20 than was possible in the absence of exogenous carbon.

Journal Article↗
PubMed1995

Altered Epiphytic Colonization of Mannityl Opine-Producing Transgenic Tobacco Plants by a Mannityl Opine-Catabolizing Strain of Pseudomonas syringae.

The plasmid pYDH208, which confers the ability to catabolize the mannityl opines mannopine and agropine, was mobilized into the nonpathogenic Pseudomonas syringae strain Cit7. The growth of the mannityl opine-catabolizing strain Cit7(pYDH208) was compared with that of the near-isogenic non-opine-catabolizing strain Cit7xylE on leaves of wild-type tobacco (Nicotiana tabacum cv. Xanthi) and transgenic mannityl opine-producing tobacco plants (N. tabacum cv. Xanthi, line 2-26). The population size of Cit7(pYDH208) was significantly greater on the lower leaves of transgenic plants than on middle or upper leaves of those plants. The population size of Cit7(pYDH208) on lower leaves of transgenic plants was also significantly greater than the population size of Cit7xylE on similar leaves of wild-type plants. High-voltage paper electrophoresis demonstrated higher levels of mannityl opines in washings from lower- and mid-level leaves than in washings from upper-level leaves. The ability of Cit7(pYDH208) to catabolize mannityl opines in the carbon-limited phyllosphere increased the carrying capacity of the lower leaves of transgenic plants for Cit7(pYDH208). In coinoculations, the increase in the ratio of population sizes of Cit7(pYDH208) to Cit7xylE on transgenic plants was apparently due to a subtle difference in the growth rates of the two strains and to the difference in final population sizes. An ability to utilize additional carbon sources on the transgenic plants also enabled Cit7(pYDH208) to achieve a higher degree of coexistence with Cit7xylE on transgenic plants than on wild-type plants. This supports the hypothesis that the level of coexistence between epiphytic bacterial populations can be altered through nutritional resource partitioning.

Journal Article↗
PubMed1994

Survival, Growth, and Localization of Epiphytic Fitness Mutants of Pseudomonas syringae on Leaves.

Among 82 epiphytic fitness mutants of a Pseudomonas syringae pv. syringae strain that were characterized in a previous study, 4 mutants were particularly intolerant of the stresses associated with dry leaf surfaces. These four mutants each exhibited distinctive behaviors when inoculated onto and into plant leaves. For example, while none showed measurable growth on dry potato leaf surfaces, they grew to different population sizes in the intercellular spaces of bean leaves and on dry bean leaf surfaces, and one mutant appeared incapable of growth in both environments although it grew well on moist bean leaves. The presence of the parental strain did not influence the survival of the mutants immediately following exposure of leaves to dry, high-light incubation conditions, suggesting that the reduced survival of the mutants did not result from an inability to produce extracellular factors in planta. On moist bean leaves that were colonized by either a mutant or the wild type, the proportion of the total epiphytic population that was located in sites protected from a surface sterilant was smaller for the mutants than for the wild type, indicating that the mutants were reduced in their ability to locate, multiply in, and/or survive in such protected sites. This reduced ability was only one of possibly several factors contributing to the reduced epiphytic fitness of each mutant. Their reduced fitness was not specific to the host plant bean, since they also exhibited reduced fitness on the nonhost plant potato; the functions altered in these strains are thus of interest for their contribution to the general fitness of bacterial epiphytes.

Journal Article↗
PubMed1994

Comparison of the Behavior of Epiphytic Fitness Mutants of Pseudomonas syringae under Controlled and Field Conditions.

The epiphytic fitness of four Tn5 mutants of Pseudomonas syringae that exhibited reduced epiphytic fitness in the laboratory was evaluated under field conditions. The mutants differed more from the parental strain under field conditions than under laboratory conditions in their survival immediately following inoculation onto bean leaves and in the size of the epiphytic populations that they established, demonstrating that their fitness was reduced more under field conditions than in the laboratory. Under both conditions, the four mutants exhibited distinctive behaviors. One mutant exhibited particularly large population decreases and short half-lives following inoculation but grew epiphytically at near-wild-type rates, while the others exhibited reduced survival only in the warmest, driest conditions tested and grew epiphytically at reduced rates or, in the case of one mutant, not at all. The presence of the parental strain, B728a, did not influence the survival or growth of three of the mutants under field conditions; however, one mutant, an auxotroph, established larger populations in the presence of B728a than in its absence, possibly because of cross-feeding by B728a in planta. Experiments with B728a demonstrated that established epiphytic populations survived exposure of leaves to dry conditions better than newly inoculated cells did and that epiphytic survival was not dependent on the cell density in the inoculum. Three of the mutants behaved similarly to two nonpathogenic strains of P. syringae, suggesting that the mutants may be altered in traits that are missing or poorly expressed in naturally occurring nonpathogenic epiphytes.

Journal Article↗
PubMed1994

Coexistence among Epiphytic Bacterial Populations Mediated through Nutritional Resource Partitioning.

The levels of coexistence between Pseudomonas syringae and various nonpathogenic epiphytic species in the phyllosphere of beans (Phaseolus vulgaris) were assessed by using replacement series. The epiphytic species Pseudomonas fluorescens, Pantoea agglomerans, Stenotrophomonas maltophilia, and Methylobacterium organophilum were all capable of exhibiting higher levels of coexistence with P. syringae than was observed with a near-isogenic P. syringae strain pair. The ecological similarity of the epiphytes was estimated with niche overlap indices derived from in vitro carbon source utilization profiles. The level of coexistence of the epiphytes was inversely correlated with the ecological similarity of the strains. Hence, the level of coexistence between the epiphytes was proportional to the degree of niche differentiation, defined as the ability to utilize carbon sources not utilized by a competing strain. Comparisons of utilization profiles for groups of carbon sources (amino acids, organic acids, and carbohydrates) indicated the types of carbon sources for which the strains likely competed in the bean phyllosphere. P. fluorescens and P. syringae strains probably competed for most carbon sources. S. maltophilia and M. organophilum strains probably competed with P. syringae for most organic acids but few amino acids or carbohydrates. P. agglomerans strains probably competed with P. syringae for most amino acids and organic acids but few carbohydrates. A variable level of coexistence observed between P. agglomerans and P. syringae probably reflected the variability in abundance in the bean phyllosphere of the carbohydrates that P. agglomerans utilized exclusively.

Journal Article↗
PubMed1994

A biological sensor for iron available to bacteria in their habitats on plant surfaces.

A sensor responsive to iron was constructed by fusing a promoterless ice nucleation activity gene (inaZ) to an iron-regulated promoter of a genomic region involved in pyoverdine (fluorescent siderophore) (pvd) production in Pseudomonas syringae. Cells of Pseudomonas fluorescens and P. syringae that contained the pvd-inaZ fusion expressed iron-responsive ice nucleation activity in the bean rhizosphere and phyllosphere, respectively, and in culture. Addition of Fe(III) to leaves or soil reduced the apparent transcription of the pvd-inaZ reporter gene, as shown by a reduction in the number of ice nuclei produced, indicating that Fe(III) was primarily responsible for mediating transcription of the pvd-inaZ gene even in natural environments. A Pseudomonas sp. strain having an intact iceC gene, which conferred Fe-insensitive expression of ice nucleation activity, was included in all studies to account for small strain- or environment-dependent differences in the ability of bacterial cells to produce ice nuclei. Thus, a comparison of the ice nucleation activity conferred by pvd-inaZ with the activity conferred by iceC revealed the bioavailability of iron in culture or natural habitats. The relative ice nucleation activities expressed by strains containing iceC or pvd-inaZ indicated that, while not abundant, Fe(III) is not present at extremely low concentrations at all microsites colonized by bacteria on plant surfaces. Biological sensors that are constructed by fusing inaZ to chemically responsive promoters provide a novel way to characterize chemical constituents of microbial habitats.

Journal Article↗
PubMed1994

Inoculum Density-Dependent Mortality and Colonization of the Phyllosphere by Pseudomonas syringae.

Pseudomonas syringae inocula containing cell concentrations ranging from 10 to 10 cells per ml were applied to the primary leaves of bean plants. The plants were incubated under conditions of high temperature and illumination and low relative humidity. Bacterial mortality rates and the proportional population decline of the inoculum were lowest at the highest inoculum concentrations. Addition of a high concentration of heat-killed cells to the inoculum containing a low concentration of viable cells significantly reduced both the mortality rate and the proportional population decline of the viable cells. The mechanisms underlying this density-dependent mortality may include cooperative protective effects of extracellular factors, such as bacterial extracellular polysaccharides, and physical protection by neighboring cells. Although epiphytic populations derived from inoculum concentrations of 10 or 10 cells per ml tended toward 10 CFU/g, the presumed carrying capacity of the leaf, populations derived from lower inoculum concentrations never achieved this carrying capacity. Assuming that epiphytic populations of P. syringae reside in discrete protected sites, our results suggest that at low inoculum concentrations, following a period of environmental stress, the number of viable cells may have dropped to zero in some sites; hence, the carrying capacity of the leaf could not be achieved.

Journal Article↗
PubMed1994

Ecological Similarity and Coexistence of Epiphytic Ice-Nucleating (Ice) Pseudomonas syringae Strains and a Non-Ice-Nucleating (Ice) Biological Control Agent.

De Wit replacement series were used to study competitive interactions between epiphytic IcePseudomonas syringae strains and the biological frost control agents IceP. syringae TLP2del1 and Pseudomonas fluorescens A506. Mixtures containing two strains in different proportions but at a constant total population size were inoculated onto potato leaves. The population sizes of each strain and the total population size were determined when the community had reached equilibrium. A near-isogenic P. syringae strain pair exhibited an interaction similar to that expected for strains competing equally for limiting environmental resources. Replacement series with nonisogenic Ice and IceP. syringae strain pairs suggested that these strains competed for limiting resources according to their relative competitive abilities. There was no evidence of any niche differentiation between the IceP. syringae strains and the IceP. syringae strain. The growth responses of epiphytes following addition of nutrients to the phyllosphere indicated that the epiphytic P. syringae populations were nutrient limited and that, under growth chamber conditions, the populations were more limited by the availability of carbon than by the availability of nitrogen. Determination of in vitro carbon source utilization profiles provided further evidence for the lack of niche differentiation between the Ice and the IceP. syringae strains. Niche overlap indices calculated for the IceP. syringae strains with respect to IceP. syringae TLP2del1 were uniformly high, indicating ecological similarity, and were consistent with the observed low level of coexistence. The biological frost control agent P. fluorescens A506 replaced P. syringae. This was correlated with a high degree of niche overlap between these species.

Journal Article↗
PubMed1993

A model of the three-dimensional structure of ice nucleation proteins.

Bacterial ice-nucleation proteins are among the most active natural ice nucleants and can reduce the supercooling point of water in plants, thereby reducing the ability of sensitive plants to avoid damaging ice formation. We describe a structural model for bacterial ice-nucleation proteins based on molecular modelling. This model predicts a largely planar extended molecule, with one side serving as a template for orienting water into an ice lattice and the other side interacting with the membrane. The model also predicts that single molecules can form aggregates of unlimited size by interdigitation.

Amino Acid Sequence↗
PubMed1993

Bacterial ice nucleation: significance and molecular basis.

Several bacterial species are able to catalyze ice formation at temperatures as warm as -2 degrees C. These microorganisms efficiently catalyze ice formation at temperatures much higher than most organic or inorganic substances. Because of their ubiquity on the surfaces of frost-sensitive plants, they are responsible for initiating ice formation, which results in frost injury. The high temperature of ice catalysis conferred by bacterial ice nuclei makes them useful in ice nucleation-limited processes such as artificial snow production, the freezing of some food products, and possibly in future whether modification schemes. The rarity of other ice nuclei active at high subfreezing temperature, and the ease and sensitivity with which ice nuclei can be quantified, have made the use of a promoterless bacterial ice nucleation gene valuable as a reporter of transcription. Target genes to which this promoter is fused can be used in cells in natural habitats. Warm-temperature ice nucleation sites have also been extensively studied at a molecular level. Nucleation sites active at high temperatures (above -5 degrees C) are probably composed of bacterial ice nucleation protein molecules that form functionally aligned aggregates. Models of ice nucleation proteins predict that they form a planar array of hydrogen binding groups that closely complement that of an ice crystal face. Moreover, interdigitation of these molecules may produce a large contiguous template for ice formation.

Bacteria↗
PubMed1993

Isolation and characterization of hydroxylamine-induced mutations in the Erwinia herbicola ice nucleation gene that selectively reduce warm temperature ice nucleation activity.

Cells of ice nucleation active bacterial species catalyse ice formation over the temperature range of -2 to -12 degrees C. Current models of ice nucleus structure associate the size of ice nucleation protein aggregates with the temperature at which they catalyse ice formation. To better define the structural features of ice nucleation proteins responsible for the functional heterogeneity of ice nuclei within a genetically homogeneous collection of cells we used in vitro chemical mutagenesis to isolate mutants with reduced ability to nucleate ice at warm assay temperatures but which retain normal or near normal nucleation activity at cold temperatures (WIND, i.e. warm ice nucleus-deficient mutants). Nearly half of the mutants obtained after hydroxylamine mutagenesis of the iceE gene from Erwinia herbicola had this phenotype. The phenotypes and location of lesions on the genetic map of iceE were determined for a number of mutants. All WIND mutations were restricted to the portion of iceE encoding the repetitive region of the polypeptide. DNA sequencing of two WIND mutants revealed single nucleotide substitutions changing a conserved serine or glycine residue to phenylalanine and serine, respectively. The implications of these findings in structure/function models for the ice nucleation protein are discussed.

Amino Acid Sequence↗
PubMed1993

Invasion and Exclusion among Coexisting Pseudomonas syringae Strains on Leaves.

The invasion and exclusion abilities of coexisting Pseudomonas syringae strains were quantified on leaves. Twenty-nine P. syringae strains were inoculated onto plants in 107 pairwise combinations. All pairs were duplicated so that each strain was inoculated both first as an antagonist strain (day 0) and second as a challenge strain (day 3). The population size of each strain in a mixture was quantified on day 6 following incubation under moist conditions. For P. syringae strains, the presence of an established population often significantly reduced the growth of subsequently arriving challenge strains on the leaf surface. Invasion and exclusion abilities, quantified by contrasting population sizes of challenge strains in the presence and in the absence of another strain, varied significantly among P. syringae strains and were partly a function of the particular strain pair. The population size of a strain when present alone on a leaf was not predictive of invasion or exclusion ability. Successful invaders were significantly less likely to exclude challenge populations than were nonsuccessful invaders. Population sizes of successful excluders were negatively correlated with population sizes of coexisting challenge strains, while population sizes of successful invaders were positively correlated with those of coexisting antagonist strains. The patterns of interaction among coexisting strains suggest mechanisms for successful invasion and exclusion among P. syringae strains on leaves.

Journal Article↗
PubMed1993

Effect of phenotypic plasticity on epiphytic survival and colonization by Pseudomonas syringae.

The bacterial epiphyte Pseudomonas syringae MF714R was cultured on agar or in broth or collected from colonized leaves; it was then inoculated onto greenhouse-grown bean plants incubated in a growth chamber at low relative humidity or in the field or onto field-grown bean plants. Cells cultured in liquid medium survived the least well after inoculation of leaf surfaces under all conditions. Cells cultured in solid medium exhibited the highest percent survival and desiccation tolerance in the growth chamber but generally survived less well in the field than did cells harvested from plants. Cells harvested from plants and inoculated onto plants in the field usually exhibited the highest percent survival, started to increase in population earlier, and reached a higher number than did cells cultured in vitro. Differences in field survival were apparently not attributable to differential UV tolerance. The observed effects of phenotypic plasticity on epiphytic survival and colonization should be considered in risk assessment studies, in studies of bacterial epidemiology, and in the use of microbial antagonists for biological pest control.

Bacteriological Techniques↗
PubMed1993

Novel method for identifying bacterial mutants with reduced epiphytic fitness.

In order to identify novel traits involved in epiphytic colonization, a technique for the rapid identification of bacterial mutants with quantitatively different population sizes in a natural habitat based on measurements of ice nucleation activity was developed. The threshold freezing temperatures of leaves harboring different numbers of cells of ice nucleation-active Pseudomonas syringae B728a differed substantially. While few leaves containing less than about 10 cells per g (fresh weight) froze at assay temperatures of -2.75 degrees C or higher, nearly all leaves froze at these temperatures when population sizes of this strain increased to about 10 cells per g (fresh weight). Presumptive epiphytic fitness mutants could readily be identified as strains which initiated freezing in fewer leaves than did other strains within a given experiment. Most Tn5-induced mutants of strain B728a which conferred a low frequency of ice nucleation on inoculated bean leaves generally had a smaller population size than the parental strain at the time of the leaf freezing assay. The leaf freezing assay was capable of differentiating samples which varied by approximately three- to fivefold in mean bacterial population size.

Journal Article↗