Inhibition of Nitrate Reduction by Chromium(VI) in Anaerobic Soil Microcosms

Chromium is often found as a cocontaminant at sites polluted with organic compounds. For nitrate-respiring microbes, Cr(VI) may be not only directly toxic but may also specifically interfere with N reduction. In soil microcosms amended with organic electron donors, Cr(VI), and nitrate, bacteria oxidized added carbon, but relatively low doses of Cr(VI) caused a lag and then lower rates of CO₂ accumulation. Cr(VI) strongly inhibited nitrate reduction; it occurred only after soluble Cr(VI) could not be detected. However, Cr(VI) additions did not eliminate Cr-sensitive populations; after a second dose of Cr(VI), bacterial activity was strongly inhibited. Differences in microbial community composition (assayed by PCR-denaturing gradient gel electrophoresis) driven by different organic substrates (glucose and protein) were smaller than when other electron acceptors had been used. However, the selection of bacterial phylotypes was modified by Cr(VI). Nine isolated clades of facultatively anaerobic Cr(VI)-resistant bacteria were closely related to cultivated members of the phylum Actinobacteria or Firmicutes. In Bacillus cereus GNCR-4, the nature of the electron donor (fermentable or nonfermentable) affected Cr(VI) resistance level and anaerobic nitrate metabolism. Our results indicate that carbon utilization and nitrate reduction in these soils were contingent upon the reduction of added Cr(VI). The amount of Cr(VI) required to inhibit nitrate reduction was 10-fold less than for aerobic catabolism of the same organic substrate. We speculate that the resistance level of a microbial process is directly related to the diversity of microbes capable of conducting it.Chromium(VI) is a toxic metal that can negatively affect bioremediation of organic compounds in sites where chromium and organic pollutants cooccur (36). Under oxygen-limited conditions, chromium(VI) can be reduced (biologically or chemically) to insoluble and relatively nontoxic Cr(III) (22). Despite the potential interactions between biotic and chemical components, the responses of anaerobic microbial activities to Cr(VI) have not been well studied (6, 7, 42, 43).Under anaerobic conditions, an important factor in the catabolism of organic carbon is the availability of electron acceptors. Nitrate is of special interest because it is often found as a copollutant in contaminated soils (18). Nitrate-reducing bacteria are facultative anaerobes commonly found in environmental samples and can couple the reduction of nitrate to the oxidation of diverse organic substrates (10, 13). The effect of Cr(VI) on natural denitrifying communities or pure cultures of denitrifying bacteria is not well characterized (8, 29). The environmental effects of Cr(VI) on denitrification are of particular interest because in addition to acute toxicity to the cell, Cr(VI) may compete with nitrate as an electron acceptor (15, 30). However, in other denitrifying bacteria (for example, Staphylococcus spp.), no competitive interactions were reported (45).The purpose of this study was to extend our work on the effects of Cr(VI) upon microbes in soil that mediate discrete chemoheterotrophic processes such as the use of O₂ (30) or Fe⁺³ (26) as terminal electron acceptors. We examined denitrification to determine whether the putative direct impact of Cr(VI) on the biochemistry of nitrate reduction would alter community dynamics from what had been observed with other terminal electron acceptors. In addition, we can add this data set to previous work to analyze the range of sensitivities to Cr(VI) that were found across a broad array of chemoheterotrophic processes.