Methylmercury formation in biofilms of Geobacter sulfurreducens

Abstract

IntroductionMercury (Hg) is a major environmental pollutant that accumulates in biota predominantly in the form of methylmercury (MeHg). Surface-associated microbial communities (biofilms) represent an important source of MeHg in natural aquatic systems. In this work, we report MeHg formation in biofilms of the iron-reducing bacterium Geobacter sulfurreducens.MethodsBiofilms were prepared in media with varied nutrient load for 3, 5, or 7 days, and their structural properties were characterized using confocal laser scanning microscopy, cryo-scanning electron microscopy and Fourier-transform infrared spectroscopy.ResultsBiofilms cultivated for 3 days with vitamins in the medium had the highest surface coverage, and they also contained abundant extracellular matrix. Using 3 and 7-days-old biofilms, we demonstrate that G. sulfurreducens biofilms prepared in media with various nutrient load produce MeHg, of which a significant portion is released to the surrounding medium. The Hg methylation rate constant determined in 6-h assays in a low-nutrient assay medium with 3-days-old biofilms was 3.9 ± 2.0 ∙ 10−14 L ∙ cell−1 ∙ h−1, which is three to five times lower than the rates found in assays with planktonic cultures of G. sulfurreducens in this and previous studies. The fraction of MeHg of total Hg within the biofilms was, however, remarkably high (close to 50%), and medium/biofilm partitioning of inorganic Hg (Hg(II)) indicated low accumulation of Hg(II) in biofilms.DiscussionThese findings suggest a high Hg(II) methylation capacity of G. sulfurreducens biofilms and that Hg(II) transfer to the biofilm is the rate-limiting step for MeHg formation in this systems.

Biochemical characterization and mercury methylation capacity of Geobacter sulfurreducens biofilms grown in media containing iron hydroxide or fumarate

“…Biofilms prepared on glass substrates were examined using Nikon A1R confocal microscope and the volume was estimated using BiofilmQ software, as previously described [ 23 , 29 ]. Briefly, biofilms were stained with Syto9 dye (Invitrogen) in dark for 15 min and imaged every 1.74 μm along the biofilm height using a DIC 20× objective.…”

“…The detailed protocol of Hg(II) methylation assay with G. sulfurreducens biofilms is described elsewhere [ 23 ]. That protocol was used here with some minor adjustments, as follows.…”

“…This assay buffer allows supporting cell metabolic activity without substantial cell growth, and it has been previously used in studies reporting Hg(II) methylation assays with G . sulfurreducens [ 12 , 16 , 17 , 23 ]. MeHg formed in the assay was quantified by isotope dilution analysis using Me 200 Hg (synthesized in house according to Snell et al [ 30 ]).…”

“…Geobacter species is one important group of Hg(II) methylators in the environment including in natural periphytic biofilms [ 21 ], and G. sulfurreducens has been used as a model in studies of environmental and biological factors controlling Hg(II) methylation [ [11] , [12] , [13] , [14] , [15] ]. However, there are no studies of MeHg formation by G. sulfurreducens biofilms cultivated on Fe(III) hydroxides despite the environmental relevance of these systems, and overall, the research on Hg(II) methylation in biofilms is still scarce [ [22] , [23] , [24] , [25] ].…”

The Extracellular Vesicles Containing Inorganic Polyphosphate of Candida Yeast upon Growth on Hexadecane

“…Phylogenetic analyses showed the close evolutionary relationships of C. maltosa with C. tropicaiis , C. viswanathii , C. aibicans , C. parapsiiosis and C. guiiiiermondii , forming a subgroup within this genus [ 42 ]. Biofilms are also used as a structure-forming matrix for the food industry and sorption of heavy metals in bioremediation [ 43 , 44 , 45 , 46 ]. These materials have good stability and biocompatibility [ 47 ] and have significant potential for various biomedical and biotechnological applications [ 48 , 49 ].…”