Microbial community succession during lactate amendment and electron acceptor limitation reveals a predominance of metal-reducing Pelosinus spp

Abstract

The determination of the success of in situ bioremediation strategies is complex. By using controlled laboratory conditions, the influence of individual variables, such as U(VI), Cr(VI), and electron donors and acceptors on community structure, dynamics, and the metal-reducing potential can be studied. Triplicate anaerobic, continuous-flow reactors were inoculated with Cr(VI)-contaminated groundwater from the Hanford, WA, 100-H area, amended with lactate, and incubated for 95 days to obtain stable, enriched communities. The reactors were kept anaerobic with N(2) gas (9 ml/min) flushing the headspace and were fed a defined medium amended with 30 mM lactate and 0.05 mM sulfate with a 48-h generation time. The resultant diversity decreased from 63 genera within 12 phyla to 11 bacterial genera (from 3 phyla) and 2 archaeal genera (from 1 phylum). Final communities were dominated by Pelosinus spp. and to a lesser degree, Acetobacterium spp., with low levels of other organisms, including methanogens. Four new strains of Pelosinus were isolated, with 3 strains being capable of Cr(VI) reduction while one also reduced U(VI). Under limited sulfate, it appeared that the sulfate reducers, including Desulfovibrio spp., were outcompeted. These results suggest that during times of electron acceptor limitation in situ, organisms such as Pelosinus spp. may outcompete the more-well-studied organisms while maintaining overall metal reduction rates and extents. Finally, lab-scale simulations can test new strategies on a smaller scale while facilitating community member isolation, so that a deeper understanding of community metabolism can be revealed.

Fig 1

qPCR quantification data (A) and cell counts (B) for microbial consortia in triplicate anaerobic continuous flow reactors inoculated with groundwater from Hanford well H-100. Average values with standard error bars are presented.

Fig 2

Concentrations of actetate (closed symbols) and lactate (open symbols) (A), CO₂ (B), and CH₄ (C) from the triplicate anaerobic continuous flow reactors inoculated with groundwater from Hanford well H-100.

Fig 3

The original and temporal changes in the bacterial community composition according to pyrosequencing analysis from selected dates from triplicate continuous flow bioreactors of lactate-enriched Hanford well H-100 groundwater sample where Pelosinus (green) and Acetobacterium (orange) became dominant.

Fig 4

(A and B) Original archaeal groundwater community (A) and temporal changes in archaeal OTU abundance (B) from selected dates from triplicate continuous flow bioreactors of the lactate-enriched Hanford well H-100 groundwater sample. Methanosarcina OTUs are dotted, and Methanobacteria OTUs are solid. (C) Distance tree of sequence representatives from the individual archaeal clusters (97% level).

Fig 5

Triplot of RDA of the relative abundances of microbial genera determined by pyrosequencing analysis on selected dates from the triplicate continuous flow reactor experiment of the lactate-enriched Hanford well H-100 groundwater samples. Dashed arrows (blue) indicate genera associated with the variation in microbial community composition. Solid (black) arrows indicate metabolite data significantly associated with the variation.