Solutes tend to concentrate on surfaces, and thus cells might respond to the higher osmolarity or concentration of particular ions at surfaces. Additionally, bacteria that are situated in biofilms on a surface experience lower oxygen and higher cell density conditions, and might interpret these conditions as location cues. Other modes of surface sensing include responses to physical perturbation of the membrane upon adherence, such as the Cpx two-component system in E. coli , or responding to the increased torque that appendages such as flagella might encounter upon their interaction with surfaces. Thus, it appears that bacteria have a variety of mechanisms with which they can sense surfaces. Different conditions might trigger biosurfactant production in different bacteria, depending on the function of the surfactant to a given species and habitat. However, are there limitations to the tasks a given surfactant can be used for? Although biosurfactant production has been noted for decades, the significance of their different chemical structures is only starting to be appreciated. For instance, it has been found that small changes in peptide components of Bacillus surfactants result in large changes of their anti-fungal and antimicrobial properties. However, as of yet there are no good guidelines on what surfactant structures are appropriate for a given type of bacterial function. This is in contrast to synthetic surfactants, where manufacturers have developed many tools for choosing appropriate surfactant formulations from thousands of synthetic surfactants. One goal of this research is to identify biosurfactants with different physical properties, and determine how these properties affect the biological roles they play to the producing organism. A particularly important property that was focused on in this study is the water solubility of biosurfactants, a proxy for their hydrophilic lipophilic balance HLB. HLB values are a scalar factor that reflects the degree to which a surfactant is hydrophilic or lipophilic, with a value of zero reflecting a completely lipophilic molecule,maceta cuadrada 25 x 25 a value of 10 corresponding to a compound with equivalent hydrophobic and hydrophilic groups, and values over 10 descriptive of predominantly hydrophilic molecules. 

This value is of great significance commercially since it is used to determine appropriate functions of surfactants. For example, common surfactants such as SDS and Tween 20 have high HLB values and are therefore best suited for emulsifying a hydrophobic substance into the water phase. On the other hand, surfactants such as Silwet® L-77 with HLB values near 10 are more suited for wetting, or spreading of a water phase over surfaces such as leaves. At the other end of the spectrum, lipophilic surfactants are best at forming inverseemulsions of water into oil. Although biosurfactants were originally proposed to be used by bacteria to solubilize hydrophobic nutrient sources , by the HLB classification alone it is obvious that only a small subset of biosurfactants would be optimal for this purpose. Biosurfactant producers are common in the environment, with around 10% of culturable bacteria in a given environment readily exhibiting this trait. Given their prevalence, the general field of microbiology will benefit from a better understanding of biosurfactant production. Additionally, in order for humans to best utilize biosurfactants, it should be informative to discover their natural functions which, in turn, might reveal novel applications for these molecules. Biosurfactants have been implicated in a large variety of functions beyond hydrocarbon emulsification. In aqueous environments, bacteria might use surfactants to coat themselves and/or surfaces to alter adherence or deherence. On the other hand, terrestrial surfaces often only harbor thin films of water; bacteria in such habitats often experience water stress and suffer from low diffusional nutrient fluxes. In this circumstance, biosurfactants might prevent evaporation or act as osmotic agents, thus maintaining thicker water films, relieving water stress and increasing microbial access to nutrients. Their ability to lower the surface tension of water has been implicated in promoting aerial hyphal growth , while their emulsification properties might enable delivery of antagonistic compounds. Because biosurfactants are amphiphilic, they can insert into membranes, and some surfactants have thus been noted for their potent membrane disrupting and resultant antimicrobial properties. 

Biosurfactants appear essential for biofilm formation in some bacteria , while they appear to prevent biofilm formation in others. Indeed, the anti-adhesive properties of some biosurfactants make them excellent candidates for coating medical devices. Additionally, some biosurfactants are proposed to act as auto inducers to signal cellular differentiation. Obviously all these traits do not apply to a given biosurfactant, but is inclusive of a rather broad spectrum of diverse molecules. Biosurfactant research would greatly benefit from further categorizations of biosurfactants based on their physical properties and demonstration of functions in which they participate.A classic function of biosurfactant activity is its enhancement of bacterial motility across soft agar plates. This motility, termed swarming motility, is an active form of translocation and is generally reliant on flagellar motility and biosurfactant production. Although biosurfactants are necessary for swarming motility in many bacteria, their production provides no benefit to swimming motility, and it is difficult to imagine a natural environment that would support the large local population sizes necessary for swarming motility. Nonetheless, it is widely assumed that biosurfactant production supports bacterial movement in vivo. How exactly might biosurfactants be beneficial to motility, and under what natural conditions do they aid motility? This question is addressed in chapter 6. Biosurfactant production has been noted in many bacterial species, but few bacterial habitats allow for as easy observation and manipulation of surfactant production as do leaves. Thus, the phyllosphere is an excellent setting in which to test the biological roles of biosurfactant production. Epiphytic bacteria not only survive, but readily flourish on leaves despite the high UV exposure, cycles of desiccation and hydration, rapid temperature fluctuations,macetas de plastico 25 litros and low and heterogeneous nutrient availability found on most leaves. It has been shown that growth of surfactant-producing bacteria on a plant can change the wettability of the leaf. It has previously been postulated that such biosurfactant production might be beneficial to the epiphytic life of bacteria and it is widely assumed that the plant environment is especially enriched with biosurfactant producers for this reason. 

It is already known that once inside the leaf, surfactant production by bacteria such as P. syringae is important for the development of disease symptoms, most likely through the induction of plant cell leakage. However, it remains unclear how biosurfactants specifically aid epiphytic growth of bacteria. Continuous water films may not normally form on such waxy surfaces; by decreasing the interfacial tension between the leaf surface and dispersed water droplets, biosurfactants could increase the wetted surface area of the leaf. Such enlarged water films might increase the distribution of locally abundant nutrients that might be separated by waxy regions of the leaf which would not otherwise be wetted by water. During periods of abundant leaf surface water, it is hypothesized that epiphytes will leave cellular aggregates in which they survive and explore the leaf surface, moving between dispersed nutrient-rich sites ; surfactant-mediated enlarged wetted areas might enable increased regions over which such motility could occur. Furthermore, surfactants might have lubricating properties, and increase bacterial motility on leaves by decreasing potential attractive forces that could immobilize bacteria on surfaces. Besides increasing growth through redistribution of nutrients and bacteria, surfactants might also increase nutrient or water availability in those sites already colonized by bacteria through their plasticizing effect on the cuticle. A number of plant-associated organisms have been studied for biosurfactant production, but few have been directly tested for the role of these compounds in planta. When surfactant-deficient mutants have been tested in planta, the focus is usually on the contributions of the biosurfactants to virulence or to the membrane-disruptive, phytotoxic properties of these molecules. A few studies have attempted to include movement in their assessment of biosurfactant roles, but the results are generally mixed; it is difficult to pinpoint the exact cause of a deficiency of colonization of plant surfaces by a mutant. Thus, although it has been speculated that the decreased fitness of biosurfactant mutants is due to their decreased motility and/or access to nutrients, neither of these factors have been directly proven on plants. Although there is a paucity of research on the role of different types of biosurfactants in the phyllosphere, the widespread use of synthetic surfactants in agriculture has provided a large source of information that might be applied to biosurfactants. Surfactants are capable of solubilizing plant epicuticular wax, thus diminishing the barrier of nutrient diffusion from the leaf onto the surface, although solubilization will only occur at concentrations above the critical micelle concentration. Biosurfactant production could potentially reach high enough local concentrations in bacterial aggregates to solubilize and strip away adjacent waxes if the biosurfactant is suited for solubilizing hydrophobic substances into water.

At lower concentrations, surfactants will have different effects on the cuticle depending on their structures. Hydrophilic surfactants, when adsorbed into the cuticle, will increase the hydration of the cuticle and therefore increase the movement of not only water but also water-soluble molecules. Alternatively, although hydrophobic surfactants readily adsorb into the cuticle, they do not increase the hydration but rather the fluidity of cuticular waxes that, in turn, increases the rate of diffusion of hydrophobic compounds across the cuticle. Additionally, movement of water and bacteria into the apoplast is normally prevented by the high surface tension of water, but can occur spontaneously when the surface tension of the liquid is reduced such as in Zebrina purpusii when the surface tension of liquid is less than 30 dyn/cm. Similarly, during plant invasion, pathogens could be employing a surfactant with high surface tension lowering abilities to facilitate water entry into stomata and other openings. Biosurfactants have been implicated in a wide variety of roles, and all of these roles might prove true in specific situations. However, it is important to start defining what types of surfactants are good at achieving a given result. The goal of this dissertation is to examine biosurfactant production in the phyllosphere with an emphasis on the plant-associated Pseudomonas syringae, in which several surfactants that it produces will be characterized and studed for their specific roles in the phyllosphere, based on clues from their genetic regulation. Biosurfactant-producing organisms have classically been identified by their ability to emulsify and utilize hydrocarbons as a nutrient source. It has only been recently appreciated that biosurfactants are produced by bacteria for many reasons other than access to hydrophobic nutrient sources. Among the numerous functions identified, are their use for swarming motility , biofilm structure and maintenance, and delivery of insoluble signals. Biosurfactants have been identified that can either promote biofilms or disperse them on root and abiotic surfaces. Additionally, some biosurfactants have been noted for their membrane-disrupting and thus zoosporicidal or antimicrobial activity. An unexplored arena where biosurfactants may prove particularly important is the colonization of waxy leaf surfaces. In order to survive on leaf surfaces, epiphytes must be able to access limited and spatially heterogeneous nutrient supplies and endure daily fluctuations in moisture availability in forms such as dew and rainfall. Continuous water films may not normally form on such waxy surfaces, and surfactants might thus aid in diffusion of compounds across the plant. If the bacteria have a pathogenic life phase, they must first have a method to enter plant tissue after which they create a favorable apoplastic environment for growth. It is already known that once inside the leaf, bacteria such as P. syringae use surfactants to cause plant cell leakage and disease symptoms. However, some studies have also implicated biosurfactants in the pre-pathogenic stages of plant-associated bacteria. Pseudomonas syringae pv. syringae B728a, a sequenced model organism with a prominent epiphytic lifestyle, produces biosurfactants. A study of the genetic regulation of biosurfactant production should provide insight into its function in this species. The identification of mutants altered in surfactant production would be an important first step in this process. However, an effective method of identifying such mutants needed to be found. Many studies have compared various screening methods to identify biosurfactant producers from limited collections of environmental isolates. Some of the most commonly used methods for analyzing biosurfactant production are drop-collapse, emulsification, and tensiometric evaluation. However, when many strains need to be assessed for surfactant production, the drop-collapse assay has been the method of choice.