Madilyn Fletcher

Analysis of genomic sequences by Chaos Game Representation

MOTIVATION Chaos Game Representation (CGR) is an iterative mapping technique that processes sequences of units, such as nucleotides in a DNA sequence or amino acids in a protein, in order to find the coordinates for their position in a continuous space. This distribution of positions has two properties: it is unique, and the source sequence can be recovered from the coordinates such that distance between positions measures similarity between the corresponding sequences. The possibility of using the latter property to identify succession schemes have been entirely overlooked in previous studies which raises the possibility that CGR may be upgraded from a mere representation technique to a sequence modeling tool. RESULTS The distribution of positions in the CGR plane were shown to be a generalization of Markov chain probability tables that accommodates non-integer orders. Therefore, Markov models are particular cases of CGR models rather than the reverse, as currently accepted. In addition, the CGR generalization has both practical (computational efficiency) and fundamental (scale independence) advantages. These results are illustrated by using Escherichia coli K-12 as a test data-set, in particular, the genes thrA, thrB and thrC of the threonine operon.

Effect of Growth Conditions and Surface Characteristics of Aquatic Bacteria on Their Attachment to Solid Surfaces

Summary: The physico-chemical basis for the effects of nutrient conditions on the attachment of four freshwater isolates, Pseudomonas fluorescens, Enterobacter cloacae, Chromobacterium sp. and Flexibacter sp., to hydrophobic (PD) and relatively hydrophilic (TCD) polystyrene surfaces was investigated. Different nutrient conditions and growth rates resulted in changes in the physico-chemistry of the bacterial surfaces, measured either by liquid contact angles on lawns of cells or by hydrophobic and electrostatic interaction chromatography of cells, and in different levels of attachment to the substrata. The phenotypic changes in cell surfaces and the levels of attachment were different for each species. Levels of bacterial adhesion differed for the two substrata, indicating different adhesion interactions with PD and TCD surfaces. Treatment of attached cells with chloramphenicol did not cause detachment of any of the bacteria from PD or TCD, whereas periodate and protease treatment removed some attached cells, the degree of detachment depending on the species. The presence of complex organic molecules, in the liquid phase and conditioning the solid surface, influenced the extent of bacterial attachment, the effect depending on the substratum, organic concentration and bacterial species. The results suggest that changes in nutrient conditions in natural aquatic habitats will affect the attachment of individual bacterial species differently, thus influencing the population structure of developing biofilms.

Simultaneous Transport of Two Bacterial Strains in Intact Cores from Oyster, Virginia: Biological Effects and Numerical Modeling

ABSTRACT The transport characteristics of two adhesion-deficient, indigenous groundwater strains, Comamonas sp. strain DA001 and Erwinia herbicola OYS2-A, were studied by using intact sediment cores (7 by 50 cm) from Oyster, Va. Both strains are gram-negative rods (1.10 by 0.56 and 1.56 by 0.46 μm, respectively) with strongly hydrophilic membranes and a slightly negative surface charge. The two strains exhibited markedly different behaviors when they were transported through granular porous sediment. To eliminate any effects of physical and chemical heterogeneity on bacterial transport and thus isolate the biological effect, the two strains were simultaneously injected into the same core. DA001 cells were metabolically labeled with 35S and tagged with a vital fluorescent stain, while OYS2-A cells were metabolically labeled with 14C. The fast decay of 35S allowed deconvolution of the two isotopes (and therefore the two strains). Dramatic differences in the transport behaviors were observed. The breakthrough of DA001 and the breakthrough of OYS2-A both occurred before the breakthrough of a conservative tracer (termed differential advection), with effluent recoveries of 55 and 30%, respectively. The retained bacterial concentration of OYS2-A in the sediment was twofold higher than that of DA001. Among the cell properties analyzed, the statistically significant differences between the two strains were cell length and diameter. The shorter, larger-diameter DA001 cells displayed a higher effluent recovery than the longer, smaller-diameter OYS2-A cells. CXTFIT modeling results indicated that compared to the DA001 cells, the OYS2-A cells experienced lower pore velocity, higher porosity, a higher attachment rate, and a lower detachment rate. All these factors may contribute to the observed differences in transport.

Adsorption of Bacterial Surface Polymers to Attachment Substrata

SUMMARY: Extracellular polysaccharide (EP) and lipopolysaccharide (LPS) were isolated from three strains of Pseudomonas fluorescens (a wild-type and two mutants) and the adsorption isotherms (relationship between amount of polymer adsorbed and bulk liquid concentration of polymer in solution) of these polymers to hydrophobic, tissue culture treated and sulphonated polystyrene surfaces were measured. The adsorption properties of the polymers were then related to the ability of the three bacterial strains to attach to the polystyrene surfaces in an attempt to elucidate the attachment mechanisms. A Langmuir adsorption isotherm equation was applied to the data, and the mathematical constants thus derived indicated if and at what concentration each surface became polymer-saturated and whether multilayer adsorption occurred. EP isolated from a crenated mutant (strain with the greatest attachment ability) adsorbed at higher concentrations than EP from wild-type and mucoid strains, and the isotherm indicated multilayer adsorption. EP from the mucoid strain (strain with little attachment ability) showed comparatively little adsorption. The isotherm of wild-type LPS was very similar to that of EP from the mucoid strain. Polymer adsorption to the three surface types was different and was generally consistent with the different degrees of bacterial attachment to the surfaces.

8 Methods for Studying Adhesion and Attachment to Surfaces

Publisher Summary The attachment of bacteria to solid surfaces has significant and often serious implications in a number of industrial, clinical, and ecological areas. Examples are the initial fouling of manmade structures, such as pipelines, heat exchangers, and ship hulls, of biomaterials used for medical implants or prosthetic devices and of particulates or larger surfaces in natural aquatic and soil environments. However, most attachment studies fall into a second category—that is, those that are laboratory based and designed to measure some specific aspect of the adhesion process. They are generally designed to measure (a) the adhesion ability of particular organisms, (b) the effect of particular environmental conditions on adhesion, or (c) the degree to which bacteria attach to a particular surface.

Bacterial Metabolism in Biofilms

Bacteria that are attached to surfaces frequently appear to differ metabolically from their free-living counterparts. There are two probable reasons for such physiological differences: (1) the physicochemical conditions at the solid surface-liquid interface are not the same as those in the bulk phase, and thus influence bacterial metabolism through environmental effects, and (2) the bacteria in biofilms are often situated in close proximity to other bacteria, microorganisms, and often macroorganisms, and are influenced by synergistic, mutualistic, competitive, or antagonistic interactions among biofilm members.

Rapid screening method for detection of bacterial mutants with altered adhesion abilities

A method has been developed to rapidly screen large numbers of bacterial mutants and identify those with altered adhesion to polystyrene surfaces. The wells in 96-well polystyrene microtiter plates were used as the attachment surfaces; both tissue culture-treated and nontreated polystyrene plates were used to allow comparison of adhesion ability to surfaces with different water-wettabilities. Numbers of attached bacteria were measured by staining them with Congo Red, eluting the stain with ethanol, and measuring the absorbance of the eluted stain in the wells with a microtiter plate reader. Assays with Pseudomonas fluoresences strain H2 and two spontaneous mutants with altered adhesion ability demonstrated the reproducibility of the technique, and assays on a series of mutants obtained by transposon mutagenesis demonstrated the ability of the method to allow identification of strains with altered adhesion abilities.

Adhesion of anaerobic microorganisms to solid surfaces and the effect of sequential attachment on adhesion characteristics

The attachment of three anaerobic microorganisms, Desulfomonile tiedjei, Syntrophomonas wolfei, and Desulfovibrio sp. strain G11, was investigated to determine if the presence of one species could influence the adhesion of another species to glass surfaces. The results indicated that the numbers and distribution of attached cells of one species could be influenced considerably by the presence of another species and the order in which the test species were exposed to the surface. D. tiedjei was found to detach readily from surfaces when it was not the primary colonizer. The attachment of Desulfovibrio G11 as the primary colonizer appeared to be stabilized by exposure to another test species. Under certain experimental conditions the test organisms formed close associations with each other on the surfaces. These findings demonstrate that the characteristics of anaerobic community biofilms can be determined by both the adhesion characteristics of the individual species and the interactions among those microorganisms.

Attachment of a Pseudomonas-like bacterium and Bacillus coagulans to solid surfaces and adsorption of their S-layer proteins

SUMMARY: The role of S-layer proteins in bacterial adhesion to solid surfaces was investigated by determining whether there was a relationship between the adsorption of S-layer protein and the attachment of the source bacteria to a series of substrata exhibiting a range of water-wettabilities. Polystyrene substrata, prepared by treatment with H2SO4, provided advancing water contact angles ranging from 76° to 46°. The test bacteria were a Pseudomonas-like strain, designated EU2, and the thermophilic bacterium Bacillus coagulans. In two out of four cases, S-layer adsorption paralleled cell attachment. Numbers of attached EU2 and amount of S-layer adsorption in phosphate buffer both increased with increasing substratum hydrophobicity. Numbers of attached B. coagulans and S-layer adsorption in distilled deionized water both decreased with increasing substratum hydrophobicity. The inconsistencies in attachment and S-layer adsorption observed in the remaining experiments were possibly due to the fact that S-layer proteins were free to adsorb by both inner and outer faces, whereas S-layer on the cell could adsorb only by the outer face. The results indicated that S-layers may play a role in bacterial adhesion to surfaces, but that the adhesiveness of S-layers depends upon their specific chemistry and environmental conditions such as medium composition and temperature.

The Measurement of Bacterial Attachment to Surfaces in Static Systems

There have been many laboratory studies that have attempted to measure the adhesive properties of bacteria or their attachment to solid surfaces in laboratory systems. Many of these investigations have used “static” systems, which were still or gently agitated and where there was no directional flow. (Flow cell systems for evaluating adhesion are dealt with elsewhere in this volume.) These static system studies have employed a variety of designs, using numerous types of bacteria, substrata, and environmental conditions. They have usually been carried out in order to evaluate (a) the adhesiveness of specific bacteria, (b) the suitability of particular materials as attachment substrata, or (c) the influence of environmental factors on the attachment process. Some laboratory procedures can also be applied to studies in natural environments (e.g. water, soil, water delivery systems, cooling towers), but they must first be validated in the laboratory. Such methods include microcosopy, analysis of biochemical markers, and nucleic acid hybridization.