Comparing denitrification estimates for a Texas estuary by using acetylene inhibition and membrane inlet mass spectrometry

Abstract

Characterizing denitrification rates in aquatic ecosystems is essential to understanding how systems may respond to increased nutrient loading. Thus, it is important to ensure the precision and accuracy of the methods employed for measuring denitrification rates. The acetylene (C2H2) inhibition method is a simple technique for estimating denitrification. However, potential problems, such as inhibition of nitrification and incomplete inhibition of nitrous oxide reduction, may influence rate estimates. Recently, membrane inlet mass spectrometry (MIMS) has been used to measure denitrification in aquatic systems. Comparable results were obtained with MIMS and C2H2 inhibition methods when chloramphenicol was added to C2H2 inhibition assay mixtures to inhibit new synthesis of denitrifying enzymes. Dissolved-oxygen profiles indicated that surface layers of sediment cores subjected to the MIMS flowthrough incubation remained oxic whereas cores incubated using the C2H2 inhibition methods did not. Analysis of the microbial assemblages before and after incubations indicated significant changes in the sediment surface populations during the long flowthrough incubation for MIMS analysis but not during the shorter incubation used for the C2H2 inhibition method. However, bacterial community changes were also small in MIMS cores at the oxygen transition zone where denitrification occurs. The C2H2 inhibition method with chloramphenicol addition, conducted over short incubation intervals, provides a cost-effective method for estimating denitrification, and rate estimates are comparable to those obtained by the MIMS method.

FIG. 1.

Selected profiles of dissolved-oxygen (O₂) concentration with depth in sediment cores subjected to MIMS flowthrough incubation and C₂H₂ inhibition incubation. Concentrations were measured post-incubation periods. Different lines and symbols represent different replicate cores, with both sites represented.

FIG. 2.

Comparison of depths of O₂ penetration into the sediment cores pre- and postincubation under different incubation techniques for all samples collected at both sites. Amendment cores were incubated with He headspace and carbon and nitrogen additions; no-amendment cores were incubated without additions; MIMS cores were incubated with a flowthrough system. **, significant difference within treatment group (α = 0.05; P < 0.001 [ANOVA]). Bars indicate standard deviations (SD).

FIG. 3.

Average denitrification estimates obtained by MIMS flowthrough treatment and acetylene inhibition with the addition of chloramphenicol for Corpus Christi Bay (CCB) and the Nueces River (NR). Bars indicate SD.

FIG. 4.

Average denitrification rates estimated using the C₂H₂ inhibition method with sediment cores and slurries from the Corpus Christi Bay (CCB) and the Nueces River (NR). Amendment cores and slurries were incubated with a helium headspace, and additions of nitrogen and carbon were made prior to incubation. **, significant difference within treatment group (α = 0.05; P < 0.001 [ANOVA]). Bars indicate SD.

FIG. 5.

DGGE profiles of 200-bp 16S rRNA gene fragments. Panel I represents the 0- to 1-cm-depth horizon, and panel II represents the 3- to 4-cm-depth horizon. A, B, and C indicate, respectively, before treatment, after C₂H₂ inhibition, and after flowthrough incubation.