An Antifungal Polycyclic Tetramate Macrolactam, Heat-Stable Antifungal Factor (HSAF), Is a Novel Oxidative Stress Modulator in Lysobacter enzymogenes.

Polycyclic tetramate macrolactams (PoTeMs) are a fast-growing family of antibiotic natural products found in phylogenetically diverse microorganisms. Surprisingly, none of the PoTeMs have been investigated for potential physiological functions in their producers. Here, we used heat-stable antifungal factor (HSAF), an antifungal PoTeM from Lysobacter enzymogenes, as a model to show that PoTeMs form complexes with iron ions, with an association constant (Ka ) of 2.71 × 106 M-1 The in vivo and in vitro data showed formation of 2:1 and 3:1 complexes between HSAF and iron ions, which were confirmed by molecular mechanical and quantum mechanical calculations. HSAF protected DNA from degradation in high concentrations of iron and H2O2 or under UV radiation. HSAF mutants of L. enzymogenes barely survived under oxidative stress and exhibited markedly increased production of reactive oxygen species (ROS). Exogenous addition of HSAF into the mutants significantly prevented ROS production and restored normal growth in the mutants under the oxidative stress. The results reveal that the function of HSAF is to protect the producer microorganism from oxidative damage rather than as an iron-acquisition siderophore. The characteristic structure of PoTeMs, a 2,4-pyrrolidinedione-embedded macrolactam, may represent a new iron-chelating scaffold of microbial metabolites. The study demonstrated a previously unrecognized strategy for microorganisms to modulate oxidative damage to the cells.IMPORTANCE PoTeMs are a family of structurally distinct metabolites that have been found in a large number of bacteria. Although PoTeMs exhibit diverse therapeutic properties, the physiological function of PoTeMs in the producer microorganisms had not been investigated. HSAF from Lysobacter enzymogenes is an antifungal PoTeM that has been subjected to extensive studies for mechanisms of biosynthesis, regulation, and antifungal activity. Using HSAF as a model system, we here showed that the characteristic structure of PoTeMs, a 2,4-pyrrolidinedione-embedded macrolactam, may represent a new iron-chelating scaffold of microbial metabolites. In L. enzymogenes, HSAF functions as a small-molecule modulator for oxidative damage caused by iron, H2O2, and UV light. Together, the study demonstrated a previously unrecognized strategy for microorganisms to modulate oxidative damage to the cells. HSAF represents the first member of the fast-growing PoTeM family of microbial metabolites whose potential biological function has been studied.

Stepwise flow diagram for the development of formulations of non spore-forming bacteria against foliar pathogens: The case of Lysobacter capsici AZ78.

The formulation is a significant step in biopesticide development and is an efficient way to obtain consistency in terms of biological control under field conditions. Nonetheless, there is still a lack of information regarding the processes needed to achieve efficient formulation of non spore-forming bacterial biological control agents. In response to this, we propose a flow diagram made up of six steps including selection of growth parameters, checking of minimum shelf life, selection of protective additives, checking that the additives have no adverse effects, validation of the additive mix under field conditions and choosing whether to use additives as co-formulants or tank mix additives. This diagram is intended to provide guidance and decision-making criteria for the formulation of non spore-forming bacterial biological control agents against foliar pathogens. The diagram was then validated by designing an efficient formulation for a Gram-negative bacterium, Lysobacter capsici AZ78, to control grapevine downy mildew caused by Plasmopara viticola. A harvest of 10(10)L. capsici AZ78cellsml(-1) was obtained in a bench top fermenter. The viability of cells decreased by only one order of magnitude after one year of storage at 4°C. The use of a combination of corn steep liquor, lignosulfonate, and polyethyleneglycol in the formulation improved the survival of L. capsici AZ78 cells living on grapevine leaves under field conditions by one order of magnitude. Furthermore, the use of these additives also guaranteed a reduction of 71% in P. viticola attacks. In conclusion, this work presents a straightforward stepwise flow diagram to help researchers develop formulations for biological control agents that are easy to prepare, stable, not phytotoxic and able to protect the microorganims under field conditions.

Characterization of water-soluble dark-brown pigment from Antarctic bacterium, Lysobacter oligotrophicus.

Lysobacter oligotrophicus strain 107-E2(T) isolated from Antarctica produces dark-brown colored water-soluble pigment, in addition to hydrolases and lytic enzymes. The production of pigment is a common characteristic among members of the genus Lysobacter, but the identity of the pigments has been unknown. In this study, we identified the pigment from L. oligotrophicus as melanin pigment (Lo-melanin) by chemical and spectroscopic analyses. Although melanin is generally insoluble in both aqueous and organic solvents, the results in this study revealed that Lo-melanin shows water-solubility by means of the added polysaccharide chain. Lo-melanin production of L. oligotrophicus was increased by ultraviolet (UV) exposure, and survival rate of Escherichia coli under UV-irradiated condition was increased by the addition of Lo-melanin to the medium.

Lysobacter xinjiangensis sp. nov., a moderately thermotolerant and alkalitolerant bacterium isolated from a gamma-irradiated sand soil sample.

A yellow-pigmented bacterial strain, designated RCML-52(T), was isolated from an abandoned gold mine in the desert in Xinjiang, China. Strain RCML-52(T) was Gram-negative, aerobic and non-motile. Phylogenetic analysis based on 16S rRNA gene sequences showed that strain RCML-52(T) was affiliated with the genus Lysobacter. Strain RCML-52(T) exhibited <95.6 % 16S rRNA gene sequence similarity to the type strains of all species of the genus Lysobacter. The major fatty acids were iso-C(16 : 0) (27.6 %), iso-C(15 : 0) (19.1 %), iso-C(17 : 1)ω9c (16.4 %), iso-C(11 : 0) 3-OH (6.5 %) and iso-C(11 : 0) (5.3 %). The DNA G+C content was 69.7 mol%. The major isoprenoid quinone was Q-8. On the basis of phylogenetic, phenotypic and chemotaxonomic analysis, strain RCML-52(T) should be assigned to a novel species of the genus Lysobacter, for which the name Lysobacter xinjiangensis sp. nov. is proposed. The type strain is RCML-52(T) (=CCTCC AB 208194(T) =KCTC 22558(T)).