Biocontrol mechanism of Myxococcus fulvus B25-I-3 against Phytophthora infestans and its control efficiency on potato late blight.

Phytophthora infestans is the pathogen of potato late blight, which is one of the most serious diseases of the potato. Myxobacteria, especially Myxococcus, become a valuable biological control resource due to their preponderant abilities to produce various secondary metabolites with novel structure and remarkable biological activity. In a previous study, Myxococcus fulvus B25-I-3 with antagonistic activity against P. infestans was isolated from an environmental sample by rabbit fecal induction method. The biocontrol mechanism of M. fulvus B25-I-3 against P. infestans and its control efficiency on potato late blight were studied. The results showed that the active substances produced by strain B25-I-3 had strong inhibitory effect on the asexual reproduction and sexual reproduction of P. infestans. In addition, the active substances could reduce the content of soluble proteins and the activity of the protective enzymes (polyphenol oxidase, peroxidase, phenylalanine ammonia lyase, superoxide dismutase) in P. infestans and increase the oxidative damage and permeability of cell membrane. And the active substances could inhibit the infection of the detached potato leaves by P. infestans significantly. In conclusion, M. fulvus B25-I-3 can produce active substances against P. infestans and has potential value to develop into biological pesticides for the control of potato late blight. The completion of this work may provide basic data for the isolation and identification of active substances and the development of pesticides against potato late blight.

A Post-segregational Killing Mechanism for Maintaining Plasmid PMF1 in Its Myxococcus fulvus Host.

Although plasmids provide additional functions for cellular adaptation to the environment, they also create a metabolic burden, which causes the host cells to be less competitive with their siblings. Low-copy-number plasmids have thus evolved several mechanisms for their long-term maintenance in host cells. pMF1, discovered in Myxococcus fulvus 124B02, is the only endogenous autonomously replicated plasmid yet found in myxobacteria. Here we report that a post-segregational killing system, encoded by a co-transcriptional gene pair of pMF1.19 and pMF1.20, is involved in maintaining the pMF1 plasmid in its host cells. We demonstrate that the protein encoded by pMF1.20 is a new kind of nuclease, which is able to cleave DNA in vitro. The nuclease activity can be neutralized by the protein encoded by pMF1.19 through protein-protein interaction, suggesting that the protein is an immune protein for nuclease cleavage. We propose that the post-segregational killing mechanism of the nuclease toxin and immune protein pair encoded by pMF1.20 and pMF1.19 is helpful for the stable maintenance of pMF1 in M. fulvus cells.

Nonadaptive processes can create the appearance of facultative cheating in microbes.

Adaptations to social life may take the form of facultative cheating, in which organisms cooperate with genetically similar individuals but exploit others. Consistent with this possibility, many strains of social microbes like Myxococcus bacteria and Dictyostelium amoebae have equal fitness in single-genotype social groups but outcompete other strains in mixed-genotype groups. Here we show that these observations are also consistent with an alternative, nonadaptive scenario: kin selection-mutation balance under local competition. Using simple mathematical models, we show that deleterious mutations that reduce competitiveness within social groups (growth rate, e.g.) without affecting group productivity can create fitness effects that are only expressed in the presence of other strains. In Myxococcus, mutations that delay sporulation may strongly reduce developmental competitiveness. Deleterious mutations are expected to accumulate when high levels of kin selection relatedness relax selection within groups. Interestingly, local resource competition can create nonzero "cost" and "benefit" terms in Hamilton's rule even in the absence of any cooperative trait. Our results show how deleterious mutations can play a significant role even in organisms with large populations and highlight the need to test evolutionary causes of social competition among microbes.

Comparative analysis of myxococcus predation on soil bacteria.

Predator-prey relationships among prokaryotes have received little attention but are likely to be important determinants of the composition, structure, and dynamics of microbial communities. Many species of the soil-dwelling myxobacteria are predators of other microbes, but their predation range is poorly characterized. To better understand the predatory capabilities of myxobacteria in nature, we analyzed the predation performance of numerous Myxococcus isolates across 12 diverse species of bacteria. All predator isolates could utilize most potential prey species to effectively fuel colony expansion, although one species hindered predator swarming relative to a control treatment with no growth substrate. Predator strains varied significantly in their relative performance across prey types, but most variation in predatory performance was determined by prey type, with Gram-negative prey species supporting more Myxococcus growth than Gram-positive species. There was evidence for specialized predator performance in some predator-prey combinations. Such specialization may reduce resource competition among sympatric strains in natural habitats. The broad prey range of the Myxococcus genus coupled with its ubiquity in the soil suggests that myxobacteria are likely to have very important ecological and evolutionary effects on many species of soil prokaryotes.

Hdsp, a horizontally transferred gene required for social behavior and halotolerance in salt-tolerant Myxococcus fulvus HW-1.

Myxococcus fulvus HW-1, a salt-tolerant bacterial strain, which was isolated from a coastal environment, changes its behavior with different salinities. To study the relationship between behavioral shifts and the adaption to oceanic conditions, the HW-1 strain was randomly mutagenized using transposon insertion, producing a dispersed-growing mutant, designated YLH0401. The mutant did not develop fruiting bodies and myxospores, was deficient in S-motility, produced less extracellular matrix and was less salt tolerant. The YLH0401 strain was determined to be mutated by a single insertion in a large gene of unknown function (7011 bp in size), which is located in a horizontally transferred DNA fragment. The gene is expressed during the vegetative growth stage, as well as highly and stably expressed during the development stage. This horizontally transferred gene may allow Myxococcus to adapt to oceanic conditions.

Myxococcus-from single-cell polarity to complex multicellular patterns.

Myxococcus xanthus creates complex and dynamic multicellular patterns as it swarms. The cells have two polar gliding engines: pulling type IV pili at their leading pole and pushing slime secretory nozzles at their lagging pole. Evidence is presented that slime secretion is vital for cell survival and that the peptidoglycan/cytoskeleton serves as a template to keep both engines oriented in the same direction. Swarming requires that each cell periodically reverse its gliding direction. For the leading pole to become the trailing pole, old engines are inactivated at both ends while new engines are being created at both ends. Reversal is initiated by a small G-protein reversal switch; a pulse of frzE approximately P from a reversal clock triggers MglA to bind GTP. Mgl.GTP then recognizes the engines that are currently in use and inactivates both of them. Meanwhile, new engines appear as instructed by the template, and the cell starts to glide in the opposite direction.

Discovery of the autonomously replicating plasmid pMF1 from Myxococcus fulvus and development of a gene cloning system in Myxococcus xanthus.

Myxobacteria are very important due to their unique characteristics, such as multicellular social behavior and the production of diverse and novel bioactive secondary metabolites. However, the lack of autonomously replicating plasmids has hindered genetic manipulation of myxobacteria for decades. To determine whether indigenous plasmids are present, we screened about 150 myxobacterial strains, and a circular plasmid designated pMF1 was isolated from Myxococcus fulvus 124B02. Sequence analysis showed that this plasmid was 18,634 bp long and had a G+C content of 68.7%. Twenty-three open reading frames were found in the plasmid, and 14 of them were not homologous to any known sequence. Plasmids containing the gene designated pMF1.14, which encodes a large unknown protein, were shown to transform Myxococcus xanthus DZ1 and DK1622 at high frequencies ( approximately 10(5) CFU/microg DNA), suggesting that the locus is responsible for the autonomous replication of pMF1. Shuttle vectors were constructed for both M. xanthus and Escherichia coli. The pilA gene, which is essential for pilus formation and social motility in M. xanthus, was cloned into the shuttle vectors and introduced into the pilA-deficient mutant DK10410. The transformants subsequently exhibited the ability to form pili and social motility. Autonomously replicating plasmid pMF1 provides a new tool for genetic manipulation in Myxococcus.

The Bacillus and Myxococcus developmental networks and their transcriptional regulators.

Studies of endospore formation by Bacillus subtilis and fruiting body development of Myxococcus xanthus have revealed key features of regulatory networks that govern temporal and spatial gene expression in bacteria. In B. subtilis, sigma factor cascades, modulated by other types of transcription factors, regulate genes in two cell types that form and communicate with each other during starvation-induced sporulation. In M. xanthus, starving cells also send signals that alter gene expression, but the cascade to emerge so far involves transcription factors other than sigma factors. A hundred thousand cells coordinate their movements to build a fruiting body in which spores form. The two regulatory networks are compared, and questions that remain are identified.

New locus important for Myxococcus social motility and development.

The mts locus in salt-tolerant Myxococcus fulvus HW-1 was found to be critical for gliding motility, fruiting-body formation, and sporulation. The homologous genes in Myxococcus xanthus are also important for social motility and fruiting-body development. The mts genes were determined to be involved in cell-cell cohesion in both myxobacterial species.

Gliding motility and polarized slime secretion.

Myxococcus leaves a trail of slime on agar as it moves. A filament of slime can be seen attached to the end of a cell, but it is seen only at one end at any particular moment. To identify genes essential for A motility, transposon insertion mutations with defective A motility were studied. Fifteen of the 33 mutants had totally lost A motility. All these mutant cells had filaments of slime emerging from both ends, indicating that bipolar secretion prevents A motility. The remaining 18 A motility mutants, also produced by gene knockout, secreted slime only from one pole, but they swarmed at a lower rate than A(+) and are called 'partial' gliding mutants, or pgl. For each pgl mutant, the reduction in swarm expansion rate was directly proportional to the reduction in the coefficient of elasticotaxis. The pgl mutants have a normal reversal frequency and normal gliding speed when they move. But their probability of movement per unit time is lower than pgl(+) cells. Many of the pgl mutants are produced by transposon insertions in glycosyltransferase genes. It is proposed that these glycosyltransferases carry out the synthesis of a repeat unit polysaccharide that constitutes the slime.

Adaptation of salt-tolerant Myxococcus strains and their motility systems to the ocean conditions.

More and more studies have indicated that myxobacteria are able to live in seawater conditions, which, however, can decrease the fruiting body formation ability and also the adventurous (A) and social (S) motility systems of the myxobacteria. To learn the adaptation mechanism of the salt-tolerant myxobacteria to marine conditions, we analyzed 10 salt-tolerant Myxococcus strains of their fruiting body formation and motility. The isolates were from marine samples and possessed different levels of salt tolerance. They had the dual motility system and formed fruiting bodies in the presence of suitable seawater concentrations. Some high salt-tolerant strains even lost their fruiting abilities in the absence of seawater. In response to the presence of seawater, the S-motility was found to be increased in the high salt-tolerants but decreased in the low salt-tolerants. The A-motility, on the other hand, was observed in all the salt-tolerant Myxococcus strains, but increased or decreased in response to the presence of seawater. Perceived shifts of fruiting body formation abilities and motilities discovered in the salt-tolerant Myxococcus strains suggested an ecological adaptation of myxobacterial social behaviors to the marine environments.

Social interactions in myxobacterial swarming.

Swarming, a collective motion of many thousands of cells, produces colonies that rapidly spread over surfaces. In this paper, we introduce a cell-based model to study how interactions between neighboring cells facilitate swarming. We chose to study Myxococcus xanthus, a species of myxobacteria, because it swarms rapidly and has well-defined cell-cell interactions mediated by type IV pili and by slime trails. The aim of this paper is to test whether the cell contact interactions, which are inherent in pili-based S motility and slime-based A motility, are sufficient to explain the observed expansion of wild-type swarms. The simulations yield a constant rate of swarm expansion, which has been observed experimentally. Also, the model is able to quantify the contributions of S motility and A motility to swarming. Some pathogenic bacteria spread over infected tissue by swarming. The model described here may shed some light on their colonization process.

Evolution of an obligate social cheater to a superior cooperator.

Obligate relationships have evolved many times and can be parasitic or mutualistic. Obligate organisms rely on others to survive and thus coevolve with their host or partner. An important but little explored question is whether obligate status is an evolutionarily terminal condition or whether obligate lineages can evolve back to an autonomous lifestyle. The bacterium Myxococcus xanthus survives starvation by the social development of spore-bearing fruiting bodies. Some M. xanthus genotypes defective at fruiting body development in isolation can nonetheless exploit proficient genotypes in chimaeric groups. Here we report an evolutionary transition from obligate dependence on an altruistic host to an autonomous mode of social cooperation. This restoration of social independence was caused by a single mutation of large effect that confers fitness superiority over both ancestral genotypes, including immunity from exploitation by the ancestral cheater. Thus, a temporary state of obligate cheating served as an evolutionary stepping-stone to a novel state of autonomous social dominance.

How myxobacteria glide.

BACKGROUND: Many microorganisms, including myxobacteria, cyanobacteria, and flexibacteria, move by gliding. Although gliding always describes a slow surface-associated translocation in the direction of the cell's long axis, it can result from two very different propulsion mechanisms: social (S) motility and adventurous (A) motility. The force for S motility is generated by retraction of type 4 pili. A motility may be associated with the extrusion of slime, but evidence has been lacking, and how force might be generated has remained an enigma. Recently, nozzle-like structures were discovered in cyanobacteria from which slime emanated at the same rate at which the bacteria moved. This strongly implicates slime extrusion as a propulsion mechanism for gliding. RESULTS: Here we show that similar but smaller nozzle-like structures are found in Myxococcus xanthus and that they are clustered at both cell poles, where one might expect propulsive organelles. Furthermore, light and electron microscopical observations show that slime is secreted in ribbons from the ends of cells. To test whether the slime propulsion hypothesis is physically reasonable, we construct a mathematical model of the slime nozzle to see if it can generate a force sufficient to propel M. xanthus at the observed velocities. The model assumes that the hydration of slime, a cationic polyelectrolyte, is the force-generating mechanism. CONCLUSIONS: The discovery of nozzle-like organelles in various gliding bacteria suggests their role in prokaryotic gliding. Our calculations and our observations of slime trails demonstrate that slime extrusion from such nozzles can account for most of the observed properties of A motile gliding.

Bacterial gliding motility: multiple mechanisms for cell movement over surfaces.

The mechanisms responsible for bacterial gliding motility have been a mystery for almost 200 years. Gliding bacteria move actively over surfaces by a process that does not involve flagella. Gliding bacteria are phylogenetically diverse and are abundant in many environments. Recent results indicate that more than one mechanism is needed to explain all forms of bacterial gliding motility. Myxococcus xanthus "social gliding motility" and Synechocystis gliding are similar to bacterial "twitching motility" and rely on type IV pilus extension and retraction for cell movement. In contrast, gliding of filamentous cyanobacteria, mycoplasmas, members of the Cytophaga-Flavobacterium group, and "adventurous gliding" of M. xanthus do not appear to involve pili. The mechanisms of movement employed by these bacteria are still a matter of speculation. Genetic, biochemical, ultrastructural, and behavioral studies are providing insight into the machineries employed by these diverse bacteria that enable them to glide over surfaces.

Myxococcus xanthus displays Frz-dependent chemokinetic behavior during vegetative swarming.

Myxococcus xanthus has been shown to utilize both directed (tactic) and undirected (kinetic) movements during different stages of its complex life cycle. We have used time-lapse video microscopic analysis to separate tactic and kinetic behaviors associated specifically with vegetatively swarming cells. Isolated individual cells separated by a thin agar barrier from mature swarms showed significant increases in gliding velocity compared to that of similar cells some distance from the swarm. This orthokinetic behavior was independent of the frequency of reversals of gliding direction (klinokinesis) but did require both the Frz signal transduction system and S-motility. We propose that M. xanthus uses Frz-dependent, auto-orthokinetic behavior to facilitate the dispersal of cells under conditions where both cell density and nutrient levels are high.

Further characterization and in situ localization of chain-like aggregates of the gliding bacteria Myxococcus fulvus and Myxococcus xanthus.

For the first time, chain-like aggregates, called "strands," have been enriched from crude cell wall preparations of liquid-grown vegetative cells of two strains of Myxococcus xanthus. These strands are highly isomorphic to macromolecular structures, previously described for Myxococcus fulvus (Lünsdorf and Reichenbach, J. Gen. Microbiol. 135:1633-1641, 1989). The strands are morphologically composed of ring elements, consisting of six or more peripheral protein masses and possibly three small central masses. The ring elements are linked by two parallel strings of filamentous proteins, called elongated elements, which keep the ring elements at a constant distance. The overall dimensions of the ring elements are 16.6 +/- 1.0 nm (n = 55) for M. xanthus Mx x48 and 16.4 +/- 1.5 nm (n = 37) for M. xanthus DK 1622. The distance between the ring elements, as a measure of the length of the elongated elements, is 16.6 +/- 1.1 nm (n = 59) for strain Mx x48 and 15.5 +/- 0.6 nm (n = 41) for strain DK 1622. Characteristically, the strands and oligomeric forms thereof show a strict association with the outer membrane. In situ studies of freeze-fractured cells of M. fulvus showed ring elements, isomorphic to those described for M. xanthus, within the periplasm; they appeared in parallel rows just below the outer membrane but not in direct contact with the cytoplasmic membrane. A three-dimensional model summarizes the morphological data. It is hypothesized that the chain-like strands, as building blocks of a more complex belt-like continuum, represent the peripheral part of the gliding machinery, which transforms membrane potential energy into mechanical work.