Public attitudes and risk perception toward land application of biosolids within the south-eastern United States.

A descriptive-correlational study of biosolids recycling was conducted in the south-eastern United States to assess current knowledge, attitudes and risk perceptions of participants in two communities that land apply biosolids as part of their waste management programs. One community, Amelia County VA, has been outspoken against biosolids recycling in the past, whereas the second community, Knoxville, TN region, has voiced few concerns about biosolids recycling. Additionally, gender differences within the entire study population were assessed. A 45-question telephone survey, utilizing a 4-point Likert scale, was developed and administered to 311 randomly selected adults in the two regions. Commonalities identified during the study revealed key risk perceptions by the public regarding biosolids regulations, treatment, and application. Given current perceptions and knowledge, respondents felt that the benefits derived from biosolids recycling do not offset the perceived health and safety risks. However, as distance between application and personal property increased, a decrease in opposition of biosolids reuse became evident for all respondents. Survey participants were dissatisfied with the level of stakeholder involvement in research and decision-making processes concerning biosolids. The outspoken Amelia County residents perceived greater health risks due to inadequate treatment of biosolids and odorous emissions during the application process than the less engaged Knox Metro respondents. Significant gender differences were observed with sampled females perceiving greater risks to health and safety from biosolids recycling than males. There was also indication that decisions and risks were not sufficiently communicated to the public, leading to respondents being inadequately informed about biosolids land application in both communities. Community-specific outreach programs must address these public risk perceptions and the differences in perception caused by gender and issue awareness to assist solid waste managers in developing and implementing successful biosolids land application systems that are acceptable to the public.

Transcription levels (amoA mRNA-based) and population dominance (amoA gene-based) of ammonia-oxidizing bacteria.

A population shift of ammonia-oxidizing bacteria (AOB) was described within a bench-scale activated sludge process treating an industrial wastewater in a previous report (Kuo et al. in Environ Eng Sci 23:507-520, 2006). In this investigation, transcriptional levels (amoA mRNA-based) of the three AOB groups (i.e., RI-27, B2-3, and Nitrosomonas nitrosa) identified in the treatment process were determined by quantitative real-time reverse transcription (RT-PCR) assays to circuitously evaluate AOB ammonia-oxidizing activity and to assess the presumed correlation between cellular activity and the dominant (greatest number) AOB population. Results demonstrated that the AOB group with higher amoA mRNA levels dominated the overall AOB population in the wastewater treatment process. Although AOB population dominance did not correlate well with transcripts at a normalized cellular level (amoA mRNA/DNA ratio), overall amoA mRNA levels did reflect the activity of distinct AOB groups under different N-loading conditions. Thus, an additional molecular parameter (amoA mRNA) was successfully utilized to assess timely shifts in AOB population structure that may impact nitrification treatment performance.

Response of Nitrobacter spp. ribosomal gene and transcript abundance following nitrite starvation and exposure to mechanistically distinct inhibitors.

The Nitrobacter spp. rRNA gene (rDNA) and relative rRNA transcript abundance (rRNAt/rDNA ratio) were evaluated in response to sudden changes in the nitrite oxidation rate. The rDNA abundance poorly indicated sudden transitions in the rate, whereas the relative rRNAt abundance usually varied quickly and significantly. In response to changes in nitrite concentration, 8 h were required for the rRNAt/rDNA ratio to transition from a minimum value at nitrite starvation (approximately 0.07) to a maximum value with excess nitrite present (approximately 4), and 5 h were required for this metric to return to the minimum value after nitrite starvation re-ensued. Generally, the relative rRNAt abundance dropped significantly after 4.5 h of exposure to three different inhibitors. A sharp decline in the rRNAt/rDNA ratio occurred during exposure to 3,5-DCP (from 4 down to 0.2) even as the fractional inhibition level remained low (< 0.10); the minimum ratio value was observed when nitrite oxidation was completely inhibited. The ratio decreased significantly during exposure to azide (from 4 to 0.5) and H+ (from 2 to 0.2), but only when the fractional inhibition levels were high (> 0.8). Interestingly, when the pH was suddenly changed to 4.5, inhibiting nitrite oxidation completely, the rRNAt/rDNA metric did not decline suggesting that rRNAt processing was inhibited. This effect was not observed during severe inhibition with 3,5-DCP and azide. Overall, the findings indicate the relative rRNAt abundance can be used to closely track in situ Nitrobacter spp. activity and in most instances will reveal inhibition events with the potential to impact treatment performance in reactors where Nitrobacter spp. are dominant.

Concurrent nitrite oxidation and aerobic denitrification in activated sludge exposed to volatile fatty acids.

The goal of this research was to investigate the simultaneous occurrence of nitrification and denitrification by activated sludge exposed to volatile fatty acids (VFAs) during aerobic wastewater treatment using a single-stage reactor. A mixture of VFAs was spiked directly into a continuous-stirred tank reactor (CSTR) to assess subsequent impacts on nitrite removal, nitrate formation, CO(2) fixation, total bacterial density, and dominant nitrite oxidizing bacteria (NOB) concentration (i.e., Nitrospira). The activity of the periplasmic nitrate reductase (NAP) enzyme and the presence of nap gene were also measured. A rapid decrease in the nitrate formation rate (>70% reduction) was measured for activated sludge exposed to VFAs; however, the nitrite removal rate was not reduced. The total bacterial density and Nitrospira concentration remained essentially constant; therefore, the reduction in nitrate formation rate was likely not due to heterotrophic uptake of nitrogen or to a decrease in the dominant NOB population. Additionally, VFA exposure did not impact microbial CO(2) fixation efficiency. The activity of NAP enzyme increased in the presence of VFAs suggesting that nitrate produced as a consequence of nitrite oxidation was likely further reduced to gaseous denitrification products via catalysis by NAP. Little, if any, nitrogen was discharged in the aqueous effluent of the CSTR after exposure to VFAs demonstrating that activated sludge treatment yielded compounds other than those typically produced solely by nitrification.

Volatile fatty acid impacts on nitrite oxidation and carbon dioxide fixation in activated sludge.

Batch test were performed to assess nitrite removal, nitrate formation, CO2 fixation, gaseous nitrogen production and microbial density in activated sludge exposed to volatile fatty acid (VFA) mixtures. Nitrite removal and nitrate formation were both affected by the presence of VFAs, but to different degrees. Nitrate formation rates were reduced to a greater extent (79%) than nitrite removal rates (36%) resulting in an apparent unbalanced nitrite oxidation reaction. Since the total bacterial density and the nitrite oxidizing bacteria (NOB, Nitrospira) concentration remained essentially constant under all test conditions, the reduction in rates was not due to heterotrophic uptake of nitrogen or to a decrease in the NOB population. In contrast to the nitrogen results, VFAs were not found to impact CO2 fixation efficiency. It appeared that nitrite oxidation occurred when VFAs were present since the oxidation of nitrite provides energy for CO2 fixation. However, nitrate produced from the oxidation of nitrite was reduced to gaseous nitrogen products. N2O gas was detected in the presence of VFAs which was a clear indication that VFAs stimulated an alternative pathway, such as aerobic denitrification, during biotransformation of nitrogen in activated sludge.

Emergence of competitive dominant ammonia-oxidizing bacterial populations in a full-scale industrial wastewater treatment plant.

Ammonia-oxidizing bacterial populations in an industrial wastewater treatment plant were investigated with amoA and 16S rRNA gene real-time PCR assays. Nitrosomonas nitrosa initially dominated, but over time RI-27-type ammonia oxidizers, also within the Nitrosomonas communis lineage, increased from below detection to codominance. This shift occurred even though nitrification remained constant.

Power analysis for real-time PCR quantification of genes in activated sludge and analysis of the variability introduced by DNA extraction.

The aims of this study were to determine the power of discrimination of the real-time PCR assay for monitoring fluctuations in microbial populations within activated sludge and to identify sample processing points where methodological changes are needed to minimize the variability in target quantification. DNA was extracted using a commercially available kit from mixed liquor samples taken from the aeration tank of four bench-scale activated-sludge reactors operating at 2-, 5-, 10-, and 20-day solid retention times, with mixed-liquor volatile suspended solid (MLVSS) values ranging from 260 to 2,610 mg/liter. Real-time PCR assays for bacterial and Nitrospira 16S rRNA genes were chosen because they represent, respectively, a highly abundant and a less-abundant bacterial target subject to clustering within the activated sludge matrix. The mean coefficient of variation in DNA yields (measured as microgram of DNA per milligram of MLVSS) in triplicate extractions of 12 different samples was 12.2%. Based on power analyses, the variability associated with DNA extraction had a small impact on the overall variability of the real-time PCR assay. Instead, a larger variability was associated with the PCR assay. The less-abundant target (Nitrospira 16S rRNA gene) had more variability than the highly abundant target (bacterial 16S rRNA gene), and samples from the lower-biomass reactors had more variability than samples from the higher-biomass reactors. Power analysis of real-time PCR assays indicated that three to five samples were necessary to detect a twofold increase in bacterial 16S rRNA genes, whereas three to five samples were required to detect a fivefold increase in Nitrospira 16S rRNA genes.

Real-time PCR quantification of nitrifying bacteria in a municipal wastewater treatment plant.

Real-time PCR assays using TaqMan or Molecular Beacon probes were developed and optimized for the quantification of total bacteria, the nitrite-oxidizing bacteria Nitrospira, and Nitrosomonas oligotropha-like ammonia oxidizing bacteria (AOB) in mixed liquor suspended solids (MLSS) from a municipal wastewater treatment plant (WWTP) using a single-sludge nitrification process. The targets for the real-time PCR assays were the 16S rRNA genes (16S rDNA) for bacteria and Nitrospira spp. and the amoA gene for N. oligotropha. A previously reported assay for AOB 16S rDNA was also tested for its application to activated sludge. The Nitrospira 16S rDNA, AOB 16S rDNA, and N. oligotropha-like amoA assays were log-linear over 6 orders of magnitude and the bacterial 16S rDNA real-time PCR assay was log-linear over 4 orders of magnitude with DNA standards. When these real-time PCR assays were applied to DNA extracted from MLSS, dilution of the DNA extracts was necessary to prevent PCR inhibition. The optimal DNA dilution range was broad for the bacterial 16S rDNA (1000-fold) and Nitrospira 16S rDNA assays (2500-fold) but narrow for the AOB 16S rDNA assay (10-fold) and N. oligotropha-like amoA real-time PCR assay (5-fold). In twelve MLSS samples collected over one year, mean cell per L values were 4.3 +/- 2.0 x 10(11) for bacteria, 3.7 +/- 3.2 x 10(10) for Nitrospira, 1.2 +/- 0.9 x 10(10) for all AOB, and 7.5 +/- 6.0 x 10(9) for N. oligotropha-like AOB. The percent of the nitrifying population was 1.7% N. oligotropha-like AOB based on the N. oligotropha amoA assay, 2.9% total AOB based on the AOB 16S rDNA assay, and 8.6% nitrite-oxidizing bacteria based on the Nitrospira 16S rDNA assay. Ammonia-oxidizing bacteria in the wastewater treatment plant were estimated to oxidize 7.7 +/- 6.8 fmol/hr/cell based on the AOB 16S rDNA assay and 12.4 +/- 7.3 fmol/hr/cell based on the N. oligotropha amoA assay.

Quantification of Nitrosomonas oligotropha-like ammonia-oxidizing bacteria and Nitrospira spp. from full-scale wastewater treatment plants by competitive PCR.

Utilizing the principle of competitive PCR, we developed two assays to enumerate Nitrosomonas oligotropha-like ammonia-oxidizing bacteria and nitrite-oxidizing bacteria belonging to the genus NITROSPIRA: The specificities of two primer sets, which were designed for two target regions, the amoA gene and Nitrospira 16S ribosomal DNA (rDNA), were verified by DNA sequencing. Both assays were optimized and applied to full-scale, activated sludge wastewater treatment plant (WWTP) samples. If it was assumed that there was an average of 3.6 copies of 16S rDNA per cell in the total population and two copies of the amoA gene per ammonia-oxidizing bacterial cell, the ammonia oxidizers examined represented 0.0033% +/- 0.0022% of the total bacterial population in a municipal WWTP. N. oligotropha-like ammonia-oxidizing bacteria were not detected in an industrial WWTP. If it was assumed that there was one copy of the 16S rDNA gene per nitrite-oxidizing bacterial cell, Nitrospira spp. represented 0.39% +/- 0.28% of the biosludge population in the municipal WWTP and 0.37% +/- 0.23% of the population in the industrial WWTP. The number of Nitrospira sp. cells in the municipal WWTP was more than 62 times greater than the number of N. oligotropha-like cells, based on a competitive PCR analysis. The results of this study extended our knowledge of the comparative compositions of nitrifying bacterial populations in wastewater treatment systems. Importantly, they also demonstrated that we were able to quantify these populations, which ultimately will be required for accurate prediction of process performance and stability for cost-effective design and operation of WWTPs.

Coronary Angioplasty of an Anomalous Coronary Artery in the Elderly: Case Report and Review of the Literature.

Newer developments in biotechnology and increasing operator experience have broadened the clinical applicability of percutaneous transluminal coronary angioplasty. These advances have allowed the treatment of patients with variant and anomalous coronary arteries and the treatment of an increasingly elderly population. A case of successful percutaneous transluminal coronary angioplasty in a 90-year-old man with unstable angina and an anomalous origin of the left main coronary artery is reported.