Changes in element availability induced by sterilization in heavy metal contaminated substrates: A comprehensive study.

Microbiome analyses of soils and microcosm experiments depend on conditions that include sterilization in order to perform experimental manipulation of microbial communities. Still, they should represent conditions close to nature. When using metal contaminated soils, sterilization methods might alter metal availability. Here, four typical metal contaminated substrates were analyzed, representing different contamination histories and soil types. They included two very poor substrates, as they are often found at metal contaminated sites. The low contents in organic carbon and nitrogen as well as two substrates with slightly higher nutrient availability were used to perform a comprehesive study for element availability changes induced by sterilization. Autoclaving, dry heat or gamma raγ sterilization were applied and compared to a non-treated control. The sterile substrates were analyzed using sequential extraction to account for different associations of the elements. Metals forming specific (hydro)oxide layers were specifically analyzed since they in turn may also impact other metals or ions. In addition, (heavy) metals and (micro)nutrients were analyzed for changes in speciation. The effects of autoclaving (wet heat) was found acceptable, while γ-ray irradiation did show unexpected changes in metal associations, especially for one substrate. Dry heat changed metal availability to the highest degree.

Siderophore production by streptomycetes-stability and alteration of ferrihydroxamates in heavy metal-contaminated soil.

Heavy metal-contaminated soil derived from a former uranium mining site in Ronneburg, Germany, was used for sterile mesocosms inoculated with the extremely metal-resistant Streptomyces mirabilis P16B-1 or the sensitive control strain Streptomyces lividans TK24. The production and fate of bacterial hydroxamate siderophores in soil was analyzed, and the presence of ferrioxamines E, B, D, and G was shown. While total ferrioxamine concentrations decreased in water-treated controls after 30 days of incubation, the sustained production by the bacteria was seen. For the individual molecules, alteration between neutral and cationic forms and linearization of hydroxamates was observed for the first time. Mesocosms inoculated with biomass of either strain showed changes of siderophore contents compared with the non-treated control indicating for auto-alteration and consumption, respectively, depending on the vital bacteria present. Heat stability and structural consistency of siderophores obtained from sterile culture filtrate were shown. In addition, low recovery (32 %) from soil was shown, indicating adsorption to soil particles or soil organic matter. Fate and behavior of hydroxamate siderophores in metal-contaminated soils may affect soil properties as well as conditions for its inhabiting (micro)organisms.

Growth of streptomycetes in soil and their impact on bioremediation.

The impact of the extremely heavy metal resistant actinomycete Streptomyces mirabilis P16B-1 on heavy metal mobilization/stabilization, phytoremediation and stress level of plants was analyzed in the presence and absence of Sorghum bicolor in sterile microcosms containing highly metal contaminated or control soil. For control, a metal sensitive S. lividans TK24 was used. The metal contents with respect to the mobile and specifically adsorbed fractions of the contaminated soil were considerably decreased by addition of both, living and dead biomass of the strains, with the heavy metal resistant S. mirabilis P16B-1 showing considerably higher impact. Both strains could grow in control soil, while only S. mirabilis P16B-1 formed new tip growth in the metal contaminated soil. A plant growth promoting effect was visible for S. mirabilis P16B-1 in contaminated soil enhancing the dry weight of inoculated Sorghum plants. Thus, metal resistant strains like S. mirabilis P16B-1 are able to enhance phytoremediation of heavy metal contaminated soils.

GC content-independent amino acid patterns in bacteria and archaea.

Every organism can be characterized by the amino acid composition of its proteome. So far it was assumed that these compositions are determined by the GC content of the DNA or, in some cases, by extreme lifestyles, like thermophily or halophily. Here, we focussed our analysis on eight amino acids, each of which is encoded by both, GC and AT rich codons, to identify finer amino acid patterns beyond the GC dominance. We investigated the conceptually translated proteomes of 1029 bacterial and archaeal strains with sequenced genomes for amino acid composition. Using correspondence analysis, we found that phylogenetic groups within bacteria and archaea generally can be discriminated from other groups due to their amino acid composition. In some cases, single organisms, e.g. Treponema pallidum strains or Mycoplasma penetrans, are characterized by extreme amino acid compositions. We assume that our data could provide a basis for a new approach to analyze evolution of bacterial and archaeal groups. Furthermore, for single organisms, the detailed knowledge of the amino acid composition of the entire proteome encoded in the genome could lead to a better understanding, important for pharmaceutical or biotechnological applications. We recommend that information about amino acid compositions should be provided in databases, comparable to the GC content of genomes.

In silico prediction of potential metallothioneins and metallohistins in actinobacteria.

Metallothioneins and metallohistins are short peptides with a high cysteine and/or histidine content able to coordinate metals intracellularly, thereby increasing the tolerance against elevated concentrations of metals. Because of their features, they can be detected by in silico prediction from proteomes annotated from sequenced genomes. Here, we analyzed 73 sequenced actinobacterial genomes for peptides (≤ 100 amino acids) with a high content of cysteine and histidine (≥ 15%) and identified 103 putative metallothioneins and metallohistins. For 45 of these peptides, we found similarities to metal binding protein domains, including zinc fingers, heavy metal transporters or eukaryotic metallothioneins, which can serve as proof-of-principle in underscoring a potential function as metal binding peptides. An evolutionary origin from metal containing domains of enzymes is discussed and metallohistins not containing cysteine are described for the first time for bacteria.

Directed synthesis of a heterobimetallic complex based on a novel unsymmetric double-Schiff-base ligand: preparation, characterization, reactivity and structures of hetero- and homobimetallic nickel(II) and zinc(II) complexes.

A series of bimetallic zinc(II) and nickel(II) complexes based on the novel dinucleating unsymmetric double-Schiff-base ligand benzoic acid [1-(3-{[2-(bispyridin-2-ylmethylamino)ethylimino]methyl}-2-hydroxy-5-methylphenyl)methylidene]hydrazide (H(2)bpampbh) has been synthesized and structurally characterized. The metal centers reside in two entirely different binding pockets provided by the ligand H(2)bpampbh, a planar tridentate [ONO] and a pentadentate [ON(4)] compartment. The utilized ligand H(2)bpampbh has been synthesized by condensation of the single-Schiff-base proligand Hbpahmb with benzoic acid hydrazide. The reaction of H(2)bpampbh with two equivalents of either zinc(II) or nickel(II) acetate yields the homobimetallic complexes [Zn(2)(bpampbh)(mu,eta(1)-OAc)(eta(1)-OAc)] (ZnZn) and [Ni(2)(bpampbh)(mu-H(2)O)(eta(1)-OAc)(H(2)O)](OAc) (NiNi), respectively. Simultaneous presence of one equivalent zinc(II) and one equivalent nickel(II) acetate results in the directed formation of the heterobimetallic complex [NiZn(bpampbh)(mu,eta(1)-OAc)(eta(1)-OAc)] (NiZn) with a selective binding of the nickel ions in the pentadentate ligand compartment. In addition, two homobimetallic azide-bridged complexes [Ni(2)(bpampbh)(mu,eta(1)-N(3))]ClO(4) (NiNi(N(3))) and [Ni(2)(bpampbh)(mu,eta(1)-N(3))(MeOH)(2)](ClO(4))(0.5)(N(3))(0.5) (NiNi(N(3))(MeOH)(2)) were synthesized. In all complexes, the metal ions residing in the pentadentate compartment adopt a distorted octahedral coordination geometry, whereas the metal centers placed in the tridentate compartment vary in coordination number and geometry from square-planar (NiNi(N(3))) and square-pyramidal (ZnZn and NiZn), to octahedral (NiNi and NiNi(N(3))(MeOH)(2)). In the case of complex NiNi(N(3)) this leads to a mixed-spin homodinuclear nickel(II) complex. All compounds have been characterized by means of mass spectrometry as well as IR and UV/Vis spectroscopies. Magnetic susceptibility measurements show significant zero-field splitting for the nickel-containing complexes (D=2.9 for NiZn, 2.2 for NiNi(N(3)), and 0.8 cm(-1) for NiNi) and additionally a weak antiferromagnetic coupling (J=-1.4 cm(-1)) in case of NiNi. Electrochemical measurements and photometric titrations reveal a strong Lewis acidity of the metal center placed in the tridentate binding compartment towards external donor molecules. A significant superoxide dismutase reactivity against superoxide radicals was found for complex NiNi.