Fungal Bioweathering of Mimetite and a General Geomycological Model for Lead Apatite Mineral Biotransformations

Fungi play important roles in biogeochemical processes such as organic matter decomposition, bioweathering of minerals and rocks, and metal transformations and therefore influence elemental cycles for essential and potentially toxic elements, e.g., P, S, Pb, and As. Arsenic is a potentially toxic metalloid for most organisms and naturally occurs in trace quantities in soil, rocks, water, air, and living organisms. Among more than 300 arsenic minerals occurring in nature, mimetite [Pb₅(AsO₄)₃Cl] is the most stable lead arsenate and holds considerable promise in metal stabilization for in situ and ex situ sequestration and remediation through precipitation, as do other insoluble lead apatites, such as pyromorphite [Pb₅(PO₄)₃Cl] and vanadinite [Pb₅(VO₄)₃Cl]. Despite the insolubility of mimetite, the organic acid-producing soil fungus Aspergillus niger was able to solubilize mimetite with simultaneous precipitation of lead oxalate as a new mycogenic biomineral. Since fungal biotransformation of both pyromorphite and vanadinite has been previously documented, a new biogeochemical model for the biogenic transformation of lead apatites (mimetite, pyromorphite, and vanadinite) by fungi is hypothesized in this study by application of geochemical modeling together with experimental data. The models closely agreed with experimental data and provided accurate simulation of As and Pb complexation and biomineral formation dependent on, e.g., pH, cation-anion composition, and concentration. A general pattern for fungal biotransformation of lead apatite minerals is proposed, proving new understanding of ecological implications of the biogeochemical cycling of component elements as well as industrial applications in metal stabilization, bioremediation, and biorecovery.     

Silver tolerance and accumulation in yeasts

Summary Debaryomyces hansenii (NCYC 459 and strain 75-21),Candida albicans (3153A),Saccharomyces cerevisiae (X2180-1B),Rhodotorula rubra (NCYC 797) andAureobasidium pullulans (IMI 45533 and ATCC 42371) were grown on solid medium supplemented with varying concentrations of AgNO₃. Although Ag⁺ is highly toxic towards yeasts, growth on solid media was still possible at Ag concentrations of 1–2 mM. Further subculture on higher Ag concentrations (up to 5 mM) resulted in elevated tolerance. The extent of Ag tolerance depended on whether Ag-containing plates were exposed to light prior to inoculation since light-mediated reduction of Ag⁺ to Ag⁰ resulted in the production of a less toxic silver species. Experimental organisms exhibited blackening of colonies and the surrounding agar during growth on AgNO₃-containing medium especially at the highest Ag concentrations tested. All organisms accumulated Ag from the medium; electron microscopy revealed that silver was deposited as electron-dense granules in and around cell walls and in the external medium. X-ray microprobe analysis indicated that these granules were metallic Ag⁰ although AgCl was also present in some organisms. Volatile and non-volatile reducing compounds were produced by several test organisms which presumably effected Ag⁺ reduction to Ag⁰.     

A Model Sheet Mineral System to Study Fungal Bioweathering of Mica

The general objective of this research was to examine fungal interactions with silicate minerals within the context of their roles in bioweathering. To achieve this, we used muscovite, a phyllosilicate mineral (KAl₂[(OH)₂|AlSi₃O₁₀]), in the form of a mineral sheet model system for ease of experimental manipulation and microscopic examination. It was found that test fungal species successfully colonized and degraded the surface of muscovite sheets in both laboratory and field experiments. After colonization by the common soil fungus Aspergillus niger, a network of hyphae covered the surface of the muscovite, and mineral dissolution or degradation was clearly evidenced by a network of fungal “footprints” that reflected coverage by the mycelium. For natural soil incubations, microorganisms associated with muscovite sheet material included biofilms of fungi and bacteria on the surface, while mineral encrustation or adhesion to microbial structures was also observed. Our results show that muscovite sheet is a good model mineral system for examination of microbial colonization and degradation, and this was demonstrated using laboratory and field systems, providing more evidence for the bioweathering significance of fungal activities in the context of silicate degradation and soil formation and development. The approach is also clearly applicable to other rock and mineral-based substrates and a variety of free-living and symbiotic microbial systems.     

Nodulation of Mimosa spp. by the beta-proteobacterium Ralstonia taiwanensis

Several beta-proteobacteria have been isolated from legume root nodules and some of these are thought to be capable of nodulating and fixing N2. However, in no case has there been detailed studies confirming that they are the active symbionts. Here, Ralstonia taiwanensis LMG19424, which was originally isolated from Mimosa pudica nodules, was transformed to carry the green fluorescent protein (gfp) reporter gene before being used to inoculate axenically-grown seedlings of M. pudica and M. diplotricha. Plants were harvested at various intervals for 56 days after inoculation, then examined for evidence of infection and nodule formation. Nodulation of both Mimosa spp. was abundant, and acetylene reduction assays confirmed that nodules had nitrogenase activity. Confocal laser scanning microscopy (CLSM) showed that fresh M. pudica nodules with nitrogenase activity had infected cells containing bacteroids expressing gfp. In parallel, fixed and embedded nodules from both Mimosa spp. were sectioned for light and electron microscopy, followed by immunogold labeling with antibodies raised against gfp and nitrogenase Fe (nifH) protein. Significant immunolabeling with these antibodies confirmed that R. taiwanensis LMG19424 is an effective N2-fixing symbiont of Mimosa spp. Both species were infected via root hairs and, in all respects, the nodule ontogeny and development was similar to that described for other mimosoid legumes. The nodules were indeterminate with a persistent meristem, an invasion zone containing host cells being invaded via prominent infection threads, and an N2-fixing zone with infected cells containing membrane-bound symbiosomes.     

Microemulsions are membrane-active, antimicrobial, self-preserving systems

Microemulsions are physically stable oil/water systems that have potential use as delivery systems for many pharmaceuticals which are normally of limited use due to their hydrophobicity, toxicity or inability to access the site of action. It has been suggested that microemulsions are self-preserving antimicrobials in their own right, although there is little evidence to support this. In this experiment, microemulsions of various compositions were formulated and tested for their stability and antimicrobial action. The physical stability of the different microemulsions was assessed by centrifugation at 4000g and by storage in a water bath at 37 degrees C for one month, during which no phase separation was observed. The antimicrobial activity of the microemulsions was tested using the compendial method, observation of the kinetics of killing, and transmission electron microscopy (TEM) of microemulsion-exposed cultures of Pseudomonas aeruginosa PA01. These latter experiments on Ps. aeruginosa indicated distinct signs of membrane disruption. The results indicated that the microemulsions are self-preserved, and that their killing of microbial cultures is very rapid and may be the result of membrane activity.     

The development and structure of root nodules on bambara groundnut [Voandzeia (Vigna)subterranea]

The infection of Vigna subterranea (formerly Voandzeia subterranea) by Bradyrhizobium strain MAO 113 (isolated from V. subterranea) was examined by light and transmission electron microscopy. Bacteria accumulated on the epidermis close to root hairs, and subsequently entered the latter via infection threads. Most of the steps involved in nodule formation were generally characteristic of determinate nodules, such as those which form on the closely related V. radiata. For example, nodule meristems were induced beneath the root epidermis adjacent to infected root hairs, but prior to infection of the meristem by rhizobia. Moreover, after the infection of some of the meristematic cells by the infection threads, and the release of the rhizobia into membrane-bound vesicles, the infection process ceased and dissemination of the rhizobia was by division of already-infected host cells. However, there were some aspects of this process in V. subterranea which have been more commonly described in indeterminate nodules. These include long infection threads entering a number of cells within the meristems simultaneously and a matrix within infection threads which was strongly labelled with immunogold monoclonal antibodies, MAC236 and MAC265, which recognize epitopes on an intercellular glycoprotein. The MAC236 and MAC265 antibodies also recognized material in the unwalled infection droplets surrounding bacteria which were newly-released from the infection threads. The amount of labelling shown was more characteristic of the long infection threads seen in indeterminate nodules such as pea (Pisum sativum) and Neptunia plena. The structure of mature V. subterranea nodules was similar to that described for other determinate nodules such as Glycine max, Vigna unguiculata and V.radiata, i.e. they were spherical and the infected zone consisted of both infected and uninfected cells. Surrounding the infected tissue was an inner cortex of uninfected cell layers containing the putative components of an oxygen diffusion barrier (including glycoprotein-occluded intercellular spaces), and an outer cortex with cells containing calcium oxalate crystals.