Surface adhesion strategies are widely employed by bacterial pathogens during establishment and systemic spread in their host. A variety of cell-surface appendages such as pili, fimbriae, and afimbrial adhesins are involved in these processes. The phytopathogen Xylella fastidiosa employs several of these structures for efficient colonization of its insect and plant hosts. Among the adhesins encoded in the X. fastidiosa genome, three afimbrial adhesins, XadA1, Hsf/XadA2, and XadA3, are predicted to be trimeric autotransporters with a C-terminal YadA-anchor membrane domain. We analyzed the individual contributions of XadA1, XadA2, and XadA3 to various cellular behaviors both in vitro and in vivo. Using isogenic X. fastidiosa mutants, we found that cell-cell aggregation and biofilm formation were severely impaired in the absence of XadA3. No significant reduction of cell-surface attachment was found with any mutant under flow conditions. Acquisition by insect vectors and transmission to grapevines were reduced in the XadA3 deletion mutant. While the XadA3 mutant was hypervirulent in grapevines, XadA1 or XadA2 deletion mutants conferred lower disease severity than the wild-type strain. This insight of the importance of these adhesive proteins and their individual contributions to different aspects of X. fastidiosa biology should guide new approaches to reduce pathogen transmission and disease development. [Formula: see text] Copyright © 2022 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Xylella fastidiosa colonizes the xylem network of host plant species as well as the foregut of its required insect vectors to ensure efficient propagation. Disease management strategies remain inefficient due to a limited comprehension of the mechanisms governing both insect and plant colonization. It was previously shown that X. fastidiosa has a functional chitinase (ChiA), and that chitin likely serves as a carbon source for this bacterium. We expand on that research, showing that a chiA mutant strain is unable to grow on chitin as the sole carbon source. Quantitative PCR assays allowed us to detect bacterial cells in the foregut of vectors after pathogen acquisition; populations of the wild-type and complemented mutant strain were both significantly larger than the chiA mutant strain 10 days, but not 3 days, post acquisition. These results indicate that adhesion of the chiA mutant strain to vectors may not be impaired, but that cell multiplication is limited. The mutant was also affected in its transmission by vectors to plants. In addition, the chiA mutant strain was unable to colonize host plants, suggesting that the enzyme has other substrates associated with plant colonization. Lastly, ChiA requires other X. fastidiosa protein(s) for its in vitro chitinolytic activity. The observation that the chiA mutant strain is not able to colonize plants warrants future attention to be paid to the substrates for this enzyme.

ABSTRACT Cell density-dependent regulation of gene expression in Xylella fastidiosa that is crucial to its switching between plant hosts and insect vectors is dependent on RpfF and its production of 2-enoic acids known as diffusible signal factor (DSF). We show that X. fastidiosa produces a particularly large variety of similar, relatively long-chain-length 2-enoic acids that are active in modulating gene expression. Both X. fastidiosa itself and a Pantoea agglomerans surrogate host harboring X. fastidiosa RpfF (XfRpfF) is capable of producing a variety of both saturated and unsaturated free fatty acids. However, only 2-cis unsaturated acids were found to be biologically active in X. fastidiosa. X. fastidiosa produces, and is particularly responsive to, a novel DSF species, 2-cis-hexadecanoic acid that we term XfDSF2. It is also responsive to other, even longer 2-enoic acids to which other taxa such as Xanthomonas campestris are unresponsive. The 2-enoic acids that are produced by X. fastidiosa are strongly affected by the cellular growth environment, with XfDSF2 not detected in culture media in which 2-tetradecenoic acid (XfDSF1) had previously been found. X. fastidiosa is responsive to much lower concentrations of XfDSF2 than XfDSF1. Apparently competitive interactions can occur between various saturated and unsaturated fatty acids that block the function of those agonistic 2-enoic fatty acids. By altering the particular 2-enoic acids produced and the relative balance of free enoic and saturated fatty acids, X. fastidiosa might modulate the extent of DSF-mediated quorum sensing. IMPORTANCE X. fastidiosa, having a complicated lifestyle in which it moves and multiplies within plants but also must be vectored by insects, utilizes DSF-based quorum sensing to partition the expression of traits needed for these two processes within different cells in this population based on local cellular density. The finding that it can produce a variety of DSF species in a strongly environmentally context-dependent manner provides insight into how it coordinates the many genes under the control of DSF signaling to successfully associate with its two hosts. Since the new DSF variant XfDSF2 described here is much more active than the previously recognized DSF species, it should contribute to plant disease control, given that the susceptibility of plants can be greatly reduced by artificially elevating the levels of DSF in plants, creating “pathogen confusion,” resulting in lower virulence.

The rpfF gene from Xylella fastidiosa, encoding the synthase for diffusible signal factor (DSF), was expressed in ‘Freedom’ grape to reduce the pathogen's growth and mobility within the plant. Symptoms in such plants were restricted to near the point of inoculation and incidence of disease was two- to fivefold lower than in the parental line. Both the longitudinal and lateral movement of X. fastidiosa in the xylem was also much lower. DSF was detected in both leaves and xylem sap of RpfF-expressing plants using biological sensors, and both 2-Z-tetradecenoic acid, previously identified as a component of X. fastidiosa DSF, and cis-11-methyl-2-dodecenoic acid were detected in xylem sap using electrospray ionization mass spectrometry. A higher proportion of X. fastidiosa cells adhered to xylem vessels of the RpfF-expressing line than parental ‘Freedom’ plants, reflecting a higher adhesiveness of the pathogen in the presence of DSF. Disease incidence in RpfF-expressing plants in field trials in which plants were either mechanically inoculated with X. fastidiosa or subjected to natural inoculation by sharpshooter vectors was two- to fourfold lower in than that of the parental line. The number of symptomatic leaves on infected shoots was reduced proportionally more than the incidence of infection, reflecting a decreased ability of X. fastidiosa to move within DSF-producing plants.

ABSTRACT Temperature, among other environmental factors, influences the incidence and severity of many plant diseases. Likewise, numerous traits, including the expression of virulence factors, are regulated by temperature. Little is known about the underlying genetic determinants of thermoregulation in plant-pathogenic bacteria. Previously, we showed that the expression of both fliC (encoding flagellin) and syfA (encoding a nonribosomal polypeptide synthetase) was suppressed at high temperatures in Pseudomonas syringae . In this work, we used a high-throughput screen to identify mutations that conferred overexpression of syfA at elevated temperatures (28°C compared to 20°C). Two genes, Psyr_2474, encoding an acyl-coenzyme A (CoA) dehydrogenase, and Psyr_4843, encoding an ortholog of RppH, which in Escherichia coli mediates RNA turnover, contribute to thermoregulation of syfA . To assess the global role of rppH in thermoregulation in P. syringae , RNA sequencing was used to compare the transcriptomes of an rppH deletion mutant and the wild-type strain incubated at 20°C and 30°C. The disruption of rppH had a large effect on the temperature-dependent transcriptome of P. syringae , affecting the expression of 569 genes at either 20°C or 30°C but not at both temperatures. Intriguingly, RppH is involved in the thermoregulation of ribosome-associated proteins, as well as of RNase E, suggesting a prominent role of rppH on the proteome in addition to its effect on the transcriptome.

Xylella fastidiosa and Xanthomonas citri subsp. citri, that cause citrus variegated chlorosis (CVC) and citrus canker diseases, respectively, utilize diffusible signal factor (DSF) for quorum sensing. DSF, produced by RpfF, are similar fatty acids in both organisms, although a different set of genes is regulated by DSF in each species. Because of this similarity, Xylella fastidiosa DSF might be recognized and affect the biology of Xanthomonas citri. Therefore, transgenic Citrus sinensis and Carrizo citrange plants overexpressing the Xylella fastidiosa rpfF were inoculated with Xanthomonas citri and changes in symptoms of citrus canker were observed. X. citri biofilms formed only at wound sites on transgenic leaves and were thicker; however, bacteria were unable to break through the tissue and form pustules elsewhere. Although abundant growth of X. citri occurred at wound sites on inoculated transgenic leaves, little growth was observed on unwounded tissue. Genes in the DFS-responsive core in X. citri were downregulated in bacteria isolated from transgenic leaves. DSF-dependent expression of engA was suppressed in cells exposed to xylem sap from transgenic plants. Thus, altered symptom development appears to be due to reduced expression of virulence genes because of the presence of antagonists of DSF signaling in X. citri in rpfF-expressing plants.

Significance Release of outer membrane vesicles (OMVs) is a general feature of Gram-negative bacteria. Most studies have addressed the mechanisms of their formation or the cargo they can carry, but other roles remain to be explored further. Here we provide evidence for a novel role for OMVs in Xylella fastidiosa , a bacterial pathogen that colonizes the xylem of important crop plants. OMVs, whose production is suppressed by a quorum-sensing system, serve as an autoinhibitor of cell adhesion to surfaces, thereby blocking attachment-driven biofilm formation that would restrict movement within the xylem and thus colonization of plants. The ubiquity of OMV formation in the bacterial world suggests that these extracellular products may have alternative roles that might modulate movement and biofilm formation.

The hypothesis that a wild-type strain of Xylella fastidiosa would restore the ability of rpfF mutants blocked in diffusible signal factor production to be transmitted to new grape plants by the sharpshooter vector Graphocephala atropunctata was tested. While the rpfF mutant was very poorly transmitted by vectors irrespective of whether they had also fed on plants infected with the wild-type strain, wild-type strains were not efficiently transmitted if vectors had fed on plants infected with the rpfF mutant. About 100-fewer cells of a wild-type strain attached to wings of a vector when suspended in xylem sap from plants infected with an rpfF mutant than in sap from uninfected grapes. The frequency of transmission of cells suspended in sap from plants that were infected by the rpfF mutant was also reduced over threefold. Wild-type cells suspended in a culture supernatant of an rpfF mutant also exhibited 10-fold less adherence to wings than when suspended in uninoculated culture media. A factor released into the xylem by rpfF mutants, and to a lesser extent by the wild-type strain, thus inhibits their attachment to, and thus transmission by, sharpshooter vectors and may also enable them to move more readily through host plants.

ABSTRACT Cell-cell signaling in Xylella fastidiosa has been implicated in the coordination of traits enabling colonization in plant hosts as well as insect vectors. This cell density-dependent signaling has been attributed to a diffusible signaling factor (DSF) produced by the DSF synthase RpfF. DSF produced by related bacterial species are unsaturated fatty acids, but that of X. fastidiosa was thought to be different from those of other taxa. We describe here the isolation and characterization of an X. fastidiosa DSF (XfDSF) as 2(Z)-tetradecenoic acid. This compound was isolated both from recombinant Erwinia herbicola expressing X. fastidiosa rpfF and from an X. fastidiosa rpfC deletion mutant that overproduces DSF. Since an rpfF mutant is impaired in biofilm formation and underexpresses the hemagglutinin-like protein-encoding genes hxfA and hxfB, we demonstrate that these traits can be restored by ca. 0.5 µM XfDSF but not by myristic acid, the fully saturated tetradecenoic acid. A phoA-based X. fastidiosa biosensor that assesses DSF-dependent expression of hxfA or hxfB revealed a high level of molecular specificity of DSF signaling. IMPORTANCE X. fastidiosa causes diseases in many important plants, including grape, where it incites Pierce’s disease. Virulence of X. fastidiosa for grape is coordinated by cell-cell signaling molecules, designated DSF (Diffusible Signaling Factor). Mutants blocked in DSF production are hypervirulent for grape, suggesting that virulence is suppressed upon DSF accumulation and that disease could be controlled by artificial elevation of the DSF level in plants. In this work, we describe the isolation of the DSF produced by X. fastidiosa and the verification of its biological activity as an antivirulence factor. We also have developed X. fastidiosa DSF biosensors to evaluate the specificity of cell-cell signaling to be investigated.

ABSTRACT Eal is an EAL domain protein in Xylella fastidiosa homologous to one involved in resistance to tobramycin in Pseudomonas aeruginosa . EAL and HD-GYP domain proteins are implicated in the hydrolysis of the secondary messenger bis-(3′-5′)-cyclic dimeric GMP (cyclic di-GMP). Cell density-dependent communication mediated by a Diffusible Signal Factor (DSF) also modulates cyclic di-GMP levels in X. fastidiosa , thereby controlling the expression of virulence genes and genes involved in insect transmission. The possible linkage of Eal to both extrinsic factors such as antibiotics and intrinsic factors such as quorum sensing, and whether both affect virulence, was thus addressed. Expression of eal was induced by subinhibitory concentrations of tobramycin, and an eal deletion mutant was more susceptible to this antibiotic than the wild-type strain and exhibited phenotypes similar to those of an rpfF deletion mutant blocked in DSF production, such as hypermotility, reduced biofilm formation, and hypervirulence to grape. Consistent with that, the rpfF mutant was more susceptible than the wild-type strain to tobramycin. Therefore, we propose that cell-cell communication and antibiotic stress can apparently lead to similar modulations of cyclic di-GMP in X. fastidiosa , resulting in similar phenotypes. However, the effect of cell density is dominant compared to that of antibiotic stress, since eal is suppressed by RpfF, which may prevent inappropriate behavioral changes in response to antibiotic stress when DSF accumulates.

Xylella fastidiosa, like related Xanthomonas species, employs an Rpf cell-cell communication system consisting of a diffusible signal factor (DSF) synthase, RpfF, and a DSF sensor, RpfC, to coordinate expression of virulence genes. While phenotypes of a ΔrpfF strain in Xanthomonas campestris could be complemented by its own DSF, the DSF produced by X. fastidiosa (XfDSF) did not restore expression of the XfDSF-dependent genes hxfA and hxfB to a ΔrpfF strain of X. fastidiosa, suggesting that RpfF is involved in XfDSF sensing or XfDSF-dependent signaling. To test this conjecture, rpfC and rpfF of X. campestris were replaced by those of X. fastidiosa, and the contribution of each gene to the induction of a X. campestris DSF-dependent gene was assessed. As in X. fastidiosa, XfDSF-dependent signaling required both X. fastidiosa proteins RpfF and RpfC. RpfF repressed RpfC signaling activity, which in turn was derepressed by XfDSF. A mutated X. fastidiosa RpfF protein with two substitutions of glutamate to alanine in its active site was incapable of XfDSF production yet enabled a response to XfDSF, indicating that XfDSF production and the response to XfDSF are two separate functions in which RpfF is involved. This mutant was also hypervirulent to grape, demonstrating the antivirulence effects of XfDSF itself in X. fastidiosa. The Rpf system of X. fastidiosa is thus a novel example of a quorum-sensing signal synthase that is also involved in the response to the signal molecule that it synthesizes.

In Xylella fastidiosa the fatty acid signal molecule diffusible signaling factor (DSF) is produced and sensed by components of the regulation of pathogenicity factors (rpf) cluster; lack of DSF production in RpfF mutants results in a non-vector-transmissible phenotype yet cells are hypervirulent to grape. rpfB has not been characterized in Xylella fastidiosa, although its homolog has been suggested to be required for DSF synthesis in Xanthomonas campestris pv. campestris. We show that RpfB is involved in DSF processing in both Xylella fastidiosa and Xanthomonas campestris, affecting the profile of DSF-like fatty acids observed in thin-layer chromatography. Although three fatty acids whose production is dependent on RpfF were detected in Xylella fastidiosa and Xanthomonas campestris wild-type strains, their respective rpfB mutants accumulated primarily one chemical species. Although no quantifiable effect of rpfB on plant colonization by Xylella fastidiosa was found, insect colonization and transmission was reduced. Thus, RpfB apparently is involved in DSF processing, and like Xanthomonas campestris, Xylella fastidiosa also produces multiple DSF molecules. It is possible that Xylella fastidiosa coordinates host vector and plant colonization by varying the proportions of different forms of DSF signals via RpfB.

A novel luxR homolog, termed XagR, in Xanthomonas axonopodis pv. glycines, the cause of soybean pustule, controls expression of pip, yapH, and at least 77 other genes. Although XagR and Pip are required for full virulence of X. axonopodis pv. glycines to soybean, constitutive overproduction of XagR suppresses infection. The xagR-dependent induction of pip occurs in planta only 2 days or more after inoculation. Although the transcription of xagR appears constitutive, XagR accumulates only in cells that have colonized soybean plants for more than 2 days suggesting that some components produced during the infection process mediate post-transcriptional control, likely by protecting XagR from proteolytic degradation. XagR modulates the adhesiveness of the pathogen during the infection process by suppressing the adhesin YapH. Although yapH mutants incite more infections of soybean leaves than the wild-type strain when topically applied under dry conditions, the mutant causes fewer infections when leaves are subject to simulated rain events after inoculation. Likewise, yapH mutants and cells in which XagR was overexpressed exhibited much more egress from infected leaves than the wild-type strain. Thus, XagR differentially modulates expression of a variety of genes during the infection process in response to feedback from plant molecules elaborated during infection to coordinate processes such as invasion, infection, and cell egress needed to complete the disease cycle.

The benefits of bioluminescence for nonsymbiotic marine bacteria have not been elucidated fully. One of the most commonly cited explanations, proposed more than 30 y ago, is that bioluminescence augments the propagation and dispersal of bacteria by attracting fish to consume the luminous material. This hypothesis, based mostly on the prevalence of luminous bacteria in fish guts, has not been tested experimentally. Here we show that zooplankton that contacts and feeds on the luminescent bacterium Photobacterium leiognathi starts to glow, and demonstrate by video recordings that glowing individuals are highly vulnerable to predation by nocturnal fish. Glowing bacteria thereby are transferred to the nutritious guts of fish and zooplankton, where they survive digestion and gain effective means for growth and dispersal. Using bioluminescence as bait appears to be highly beneficial for marine bacteria, especially in food-deprived environments of the deep sea.

The Escherichia coli yjbEFGH operon, encoding genes involved in exopolysaccharide production, has previously been shown to be induced by osmotic stress and to be negatively regulated by σ38. Promoter analysis suggested that like most E. coli genes, its transcription is driven by the housekeeping sigma factor σ70. Indeed, manipulation of any of the other five alternative sigma factors did not affect its induction by osmotic stress. Surprisingly, when assayed in a strain expressing low levels of σ70, yjbEFGH induction in response to osmotic stress was higher than in a strain expressing normal levels of σ70. Similar phenomena were observed in the σ70-driven promoters of sulA, uvrA, recA, fecI, entC and lacZ, the transcription of which is directly controlled by a repressor protein (LexA, Fur and LacI), but not in promoters of the housekeeping genes ftsA and ftsY, or in σ38-driven treA promoter. Since transcription factors are generally present in the cell in low numbers, we hypothesize that a decrease in σ70, that drives the expression of lexA, fur and lacI as well, further diminishes their number in the cell and thus de-represses the induction of genes which are subjected to their repression.

Xylella fastidiosa (Xf) produces an unsaturated fatty acid signal molecule called diffusible signal factor (DSF) that modulates gene expression in cells as they reach high numbers in plants. By increasing the expression of a variety of afimbrial adhesins while decreasing the expression of pili involved in twitching motility as well as extracellular enzyme involved in degrading pit membranes, DSF accumulation suppresses virulence of Xf in grape. We thus are exploring different ways to elevate DSF levels in plants to achieve “pathogen confusion.” Plants expressing rpfF from Xf produce low levels of DSF and are highly resistant to Pierce’s disease (PD). Chloroplast targeting of RpfF substantially increased DSF production in grape; these plants are currently being tested for disease resistance. Xf moved much less rapidly in rpfF-transformed grape, colonized many fewer xylem vessels, and achieved a much lower population size indicating that elevated DSF levels suppressed movement within the plant. Topical applications of DSF-containing bacterial extracts also reduced the severity of PD suggesting that DSF is relatively mobile within the plant and that exogenous sources of DSF might also be applied in various ways to achieve suppression of pathogen mobility and hence virulence. Some reduction in disease severity was observed in grape scions grafted to DSF-producing rootstocks suggesting that DSF produced by rootstocks can somewhat move to scions and confer disease control’ the control of disease was substantially less than that seen in transformed scions however. While certain bacteria such as Rhizobium etli can produce DFS when transformed with rpfF from Xf, and to colonize grape slowly after inoculation, modest levels of disease control are conferred by pre-treatment of grape suggesting that a more rapidly multiplying and internally mobile endophytic bacterium might be superior for disease control as a surrogate DSF producer. As studies of pathogen confusion will be greatly facilitated by having an improved bioassay for the DSF produced by Xf, we have been developing several immunological and biochemical means to assay for the presence of DSF using Xf itself as a bioindicator. LAYPERSON SUMMARY: Xylella fastidiosa (Xf) produces an unsaturated fatty acid signal molecule called diffusible signal factor (DSF) that changes its gene expression in cells as they reach high numbers in plants. Accumulation of DSF in Xf cells, which presumably normally occurs as cells become numerous within xylem vessels, causes a change in many genes in Xf, but the overall effect is to suppress the virulence of Xf in plants. We have investigated DSF-mediated cell-cell signaling in Xf with the aim of developing cell-cell signaling disruption (pathogen confusion) as a means of controlling Pierce’s disease (PD). We have investigated both the role of DFS-production by Xf on its behavior within plants, the manner in which other bacterial strains affect such cell signaling, the extent to which other endophytes could modulate density-dependent behaviors and virulence in Xf by interfering with cell-cell signaling, performed genetic transformation of grape to express DSF, and explored other means to alter DSF abundance in plants to achieve PD control. Elevating DSF levels in plants should reduce movement of Xf in the plant. We have produced bacterial endophyte strains that can produce large amounts of DSF and, by moving within plants apparently they can alter the abundance of DSF sufficiently to reduce the virulence of Xf. Given that DSF overabundance appears to mediate an attenuation of virulence in Xf we have transformed grape with the rpfF gene of Xf to enable DSF production in plants; such grape plants produce at least some DSF and are much less susceptible to disease. Higher levels of expression of DSF have been obtained in plants by targeting the biosynthetic enzymes to the chloroplast. Some reduction in disease severity was observed in grape scions grafted to DSF-producing rootstocks suggesting that DSF produced by rootstocks can somewhat move to scions and confer disease control. INTRODUCTION We have found that the virulence of Xylella fastidiosa (Xf) is strongly regulated in a cell density-dependent fashion by accumulation of a signal molecule called diffusible signal factor (DSF) encoded by rpfF and involving signal transduction that requires other rpf genes. We now have shown that Xf makes a DSF molecule that is recognized by Xanthomonas campestris pv. campestris (Xcc) but slightly different than the DSF of Xcc (Figure 1). In striking contrast to that of Xcc, rpfFmutants of Xf blocked in production of DSF, exhibit dramatically increased virulence to plants, however, they are unable to be spread from plant to plant by their insect vectors since they do not form a biofilm within the insect. These Figure 1

The movement of Xylella fastidiosa (Xf) in plants as well as insect transmission is controlled by a small diffusible signal factor (DSF) that accumulates when cells are at high cell densities. Pathogen behavior can be changed and disease reduced by altering the abundance of DSF in plants in a form of "pathogen confusion." To enable new strategies of pathogen confusion we have chemically characterized the most abundant DSF produced by grape strains of Xf under the control of the rpfF gene as 2-Z-tetradecenoic acid (hereafter called C14-cis). Additional related chemicals forms of DSF with biological activity may exist, and we are currently determining their relative activity and if such molecules cooperate in regulating gene expression in Xf. The DSF of Xf is structurally related to, but distinct from, the DSF made by Xanthomonas campestris pv. campestris (Xcc). While an Xcc eng:gfp based biosensor for DSF can detect as little as about 1 uM of DSF produced by Xcc, more than about 100 uM of C14-cis is required for detection. Biological assays for the presence of C14-cis are being developed in Xf. As the expression of genes conferring type IV pili and thus twitching are suppressed while those involved in EPS production and production of various cell adhesins are induced in the presence of DSF in Xf, we are developing Xf-based bioassays for C14-cis using an rpfF mutant of Xf that cannot produce DSF but which can respond to exogenous C14-cis. Twitching motility of the rpfF mutant was suppressed in the presence of as little as 1 uM exogenous C14 cis while cell-cell adhesiveness and cell-surface adhesiveness was enhanced. Further bioassays based on immunological detection of cell surface adhesins or EPS as well as by quantifying mRNA associated with these genes in Xf are being developed. Initial results suggest that the responsiveness of Xf to C14-cis is dependent on the physiological state of cells; young, actively- growing cells appear to respond less than older cells. C14-cis, as well as the Sodium salt of this fatty acid which is highly water soluble, have been produced and have been used as topical and injected treatments of grape that have subsequently been challenge inoculated with Xf for tests of disease control; modest decreases in disease control with these treatments revealed a need to better assess the efficiency with which DSF was introduced into plants. Substantial decreases in disease severity and incidence of vessel colonization with cells of Xf were obtained after application of DSF-containing bacterial extracts, revealing the importance of efficient methods of introduction of this signal molecule for disease control. We have also designed and synthesized some DSF-analogs and will soon test them for their ability to alter pathogen gene expression and behavior in culture as well as control disease.