Semrau, J. D. | bioRxiv Science

bioRxiv2026

Quantifying the Effect of Encapsulation on Microbial Mineralization Processes Using a Modified DNA Extraction Method

Microbially-mediated mineral precipitation is important in the natural world and to a number of emerging climate, energy, and water technologies. Central to many microbial mineralization processes is the preferential accumulation of precipitates on microbial surfaces. In some systems, including ureolytic microbially-induced carbonate precipitation (MICP), precipitates may entomb cells in a process termed "encapsulation." Encapsulation is known to greatly affect microbial systems, but understanding these relationships quantitatively has proven difficult, largely because nucleic acid extraction from encapsulated cells yields inconsistent results. Here, a demineralization protocol adapted from studies of natural microbialites is shown to recover genomic DNA from carbonate-encapsulated Sporosarcina pasteurii at yields indistinguishable from unmineralized cells and of sufficient quality for quantitative PCR. This method allows the degree of encapsulation to be expressed quantitatively as the amount of precipitate formed normalized to the estimated number of cells present, and allows growing populations to be tracked with increased precision. Simultaneous growth and mineralization experiments in mixed batch reactors suggest encapsulation-induced growth suppression at calcium concentrations of 250 mM, which are commonly employed in MICP under different physicochemical conditions. Variable rates of precipitate accumulation on cells, possibly stemming from local mass transport constraints, are proposed as a mechanism of calcium concentration-dependent growth suppression. Accordingly, mass transport principles are used to derive a reaction rate scaling function, allowing the observed growth and precipitation behavior to be replicated by a numerical model. These findings lay the groundwork for a more fundamental understanding of how encapsulation shapes microbial mineralization processes.

microbiology↗

bioRxiv2020

Enhancement of nitrous oxide emissions in soil microbial consortia via copper competition between proteobacterial methanotrophs and denitrifiers

Unique means of copper scavenging have been identified in proteobacterial methanotrophs, particularly the use of methanobactin, a novel ribosomally synthesized post-translationally modified polypeptide that binds copper with very high affinity. The possibility that copper sequestration strategies of methanotrophs may interfere with copper uptake of denitrifiers in situ and thereby enhance N2O emissions was examined using a suite of laboratory experiments performed with rice paddy microbial consortia. Addition of purified methanobactin from Methylosinus trichosporium OB3b to denitrifying rice paddy soil microbial consortia resulted in substantially increased N2O production, with more pronounced responses observed for soils with lower copper content. The N2O emission-enhancing effect of the soils native mbnA-expressing Methylocystaceae methanotrophs on the native denitrifiers was then experimentally verified with a Methylocystaceae-dominant chemostat culture prepared from a rice paddy microbial consortium as the inoculum. Lastly, with microcosms amended with varying cell numbers of methanobactin-producing Methylosinus trichosporium OB3b before CH4 enrichment, microbiomes with different ratios of methanobactin-producing Methylocystaceae to gammaproteobacterial methanotrophs incapable of methanobactin production were simulated. Significant enhancement of N2O production from denitrification was evident in both Methylocystaceae-dominant and Methylococcaceae-dominant enrichments, albeit to a greater extent in the former, signifying the comparative potency of methanobactin-mediated copper sequestration while implying the presence of alternative copper abstraction mechanisms for Methylococcaceae. These observations support that copper-mediated methanotrophic enhancement of N2O production from denitrification is plausible where methanotrophs and denitrifiers cohabit. ImportanceProteobacterial methanotrophs, groups of microorganisms that utilize methane as source of energy and carbon, have been known to utilize unique mechanisms to scavenge copper, namely utilization of methanobactin, a polypeptide that binds copper with high affinity and specificity. Previously the possibility that copper sequestration by methanotrophs may lead to alteration of cuproenzyme-mediated reactions in denitrifiers and consequently increase emission of potent greenhouse gas N2O has been suggested in axenic and co-culture experiments. Here, a suite of experiments with rice paddy soil slurry cultures with complex microbial compositions were performed to corroborate that such copper-mediated interplay may actually take place in environments co-habited by diverse methanotrophs and denitrifiers. As spatial and temporal heterogeneity allow for spatial coexistence of methanotrophy (aerobic) and denitrification (anaerobic) in soils, the results from this study suggest that this previously unidentified mechanism of N2O production may account for significant proportion of N2O efflux from agricultural soils.

microbiology↗