Biotransformation of phosphogypsum in media containing different forms of nitrogen.

Studies on the biotransformation of phosphogypsum (a waste product formed in the course of the production of phosphorous fertilizers) with the use of sulfate reducing bacteria (SRB) demonstrated that it is a good source of sulfates and biogenic elements for these bacteria, though the addition of organic carbon and nitrogen is necessary. The aim of this study was to investigate the form of nitrogen and C:N ratio in the medium on the growth of SRB community in cultures containing phosphogypsum. Batch community cultures of sulfate reducing bacteria were maintained in medium with phosphogypsum (5.0 g/l), different concentrations of sodium lactate (1.6 - 9.4 g/l) and different forms (NH4CI, CO(NH2)2, KNO3) and concentrations (0 - 250 mg/l) of nitrogen. The growth of SRB was studied in the C:N ratio of from 2:1 to 300:1. It was found that: 1 - the best source of nitrogen for SRB is urea, followed by ammonium, the worst were nitrates; 2 - the bacteria were also able to grow in medium without nitrogen but their activity was then by approximately 15% lower than in optimal growth conditions; 3 - in medium with KNO3 inhibition of sulfate reduction by approx. 50% was observed; 4 - the highest reduction of nitrates (removal of nitrate) in media with phosphogypsum and nitrates was at limiting concentrations of sodium lactate. This is probably caused by the selection under these conditions (low concentration of hydrogen sulfide) of denitrifying bacteria or sulfate reducing bacteria capable of using nitrates as an electron acceptor.

Biodegradation of phenol by bacterial strains from petroleum-refining wastewater purification plant.

The ability of four strains of bacteria derived from a biological petroleum-refining wastewater purification plant to carry out the biodegradation of phenol was studied. Two of the strains belonging to the genus Pseudomonas were found to be characterised by high effectiveness of the removal of phenol which was used as sole carbon and energy source (the strains were designated P1 and P2). In turn the effect of inoculum size, initial concentration of substrate (500 and 1,000 mg phenol/L) and temperature (10, 20 and 30 degrees C) on the rate of phenol degradation by strains P1, P2 and mixture of both was investigated. It was found that strain P1 which was identified as Pseudomonas fluorescens degraded phenol better than strain P2--Pseudomonas cepacia. The rate of phenol biodegradation was significantly affected by size of inoculum and temperature of incubation. Phenol was removed the fastest with the highest inoculum used. The optimal temperature was about 20 degrees C. At 10 and 30 degrees C the process of biodegradation was visibly inhibited. The rate of phenol utilisation was also found to decrease with increased concentration of substrate.

Phenol-degrading denitrifying bacteria in wastewater sediments.

An attempt was made to isolate denitrifying bacteria capable of degrading phenol from anoxic wastewater sediments and compost (vegetable soil). The greatest phenol-degrading activity was shown by autochthonous microflora from petroleum wastewater sediments (PWS) and by denitrifying microflora from nitrogenous wastewater reservoir sediment (NWRS). Eight strains of denitrifying bacteria able to degrade phenolic compounds were isolated, six of which were from the petroleum wastewater sediments. The highest activity of phenol degradation (23.5 and 23 mg/L/day) in denitrifying conditions was shown by strains 54/1 and 83/2, both from petroleum wastewater sediments, which were classified as Pseudomonas aeruginosa and Alcaligenes faecalis, respectively.

Adaptation of a phenol-degrading denitrifying bacteria to high concentration of phenol in the medium.

The growth and uptake of phenol by 8 strains isolated from wastewater sediments in stationary cultures in medium with increasing concentrations of phenol (from 100 to 600 mg/L) under denitrifying conditions were studied. All the strains grew in media containing 250 mg phenol/L and only strains 101/1, 83/2 and 21/1/ in consecutive passages visibly increased both specific growth rate (mu day-1) as well as phenol-degrading activity (mg/L x day). Consecutive passages of the culture in medium containing 400 mg phenol/L resulted in the elimination of 3 out of the 5 strains growing in the medium in the first passage. Only strain 101/1 demonstrated high specific growth rate and phenol-degrading activity in medium containing 600 mg phenol/L. In consecutive passages in medium containing 250, 400 and 600 mg phenol/L the specific growth (mu day-1) and phenol-degrading activity (mg/L x day) of P. aeruginosa 101/1 were 0.38 and 36; 0.12 and 19; 0.09 and 20, respectively.

An attempt to use selected strains of bacteria adapted to high concentrations of petroleum oil to increase the effective removal of petroleum products in excess activated sludge in laboratory conditions.

Forty two strains of bacteria were isolated from excess activated sludge from petroleum wastewater treatment plant. The strains were identified and classified to the following groups: Enterobacteriaceae (7 strains), Anitratum (3 strains), Pseudomonas (13 strains), Micrococcus (12 strains), Comamonas (2 strains), Xanthomonas (2 strains), Achromobacter (1 strain) and Vibrio-Aeromonas (1 strain). One of the isolates was found to be a yeast strain. Following preliminary selection ten strains, showing the best growth in medium with oil fraction as sole carbon source, were chosen for further studies. The selected strains belong to Pseudomonas (6 strains), Xanthomonas (2 strains), Micrococcus (1 strain) and Saccharomyces (1 strain). The strains were adapted to high oil concentration (500-2000 mg/L) and an attempt to use them to intensify removal of petroleum products from excess activated sludge was made. The sludge was inoculated with a mixture of the isolated strains. The experiment was carried out three times, each time with a fresh sample of the excess sludge. The obtained results show that the inoculation of activated sludge with the strains active against oil reduced the petroleum products content by 20% in 14 days. The greatest reduction of oil was observed in sludge with the lowest dry weight, that is with the greatest degree of hydration. The dry weight of the excess sludge did not significantly decrease during the course of the experiment, after having been inoculated with the mixture of strains.

Isolation and identification of bacteria from activated sludge purifying petroleum wastewaters.

Several strains growing well in minimal media with 500 and 1000 mg/l of oil or phenol as a sole carbon source were isolated from activated sludge purifying petroleum waste waters and identified. Five of the best growing strains classified as Arthrobacter, Pseudomonas, Xanthomonas and Enterobacter were selected and their capacity to remove petroleum components and phenol (in the oil fraction of petrochemical waste waters) was studied. The enzymatic activity of the strains, including respiration intensity and dehydrogenase activity was also determined. All the examined strains were found to use oils as the sole source of carbon (percent age of the oils reduction during cultivation of the individual strains ranged from 58 to 78). Phenol was completely reduced by only one strain. The rest of the strain removed only from 7 to 24% of this compound. The activity of dehydrogenases and the respiration intensity in the presence of the studied substrates -- oil and phenol was low for all the examined strains.

On the role of bacterial inoculum growth conditions on nitrate denitrification pathway.

Activities of nitrite and nitrate reductases as well as the efficiencies of denitrification carried out by Pseudomonas aeruginosa and Pseudomonas stutzeri under different culture conditions have been compared. The inoculum growth conditions had no influence on denitrification pathway in either strain. Activities of both reductases were higher in Pseudomonas stutzeri than in Pseudomonas aeruginosa cultures.

Denitrification of high concentrations of nitrites and nitrates in synthetic medium with different sources of organic carbon. III. Methanol.

The denitrification of nitrites and nitrates (1000 mg N/l) in medium containing methanol as a source of organic carbon was studied. Continuous cultures of mixed population of autochtonic microflora from bottom sludge of nitrogenous wastewater reservoir were set up in a chemostat-type column and packed bed reactor. The efficiency of denitrification of nitrates in packed bed reactor was 506.7 mg N/l/h whereas denitrification of nitrites was from 8.7 to 16.0 mg N/l/h depending on the granulation of the filing material. In the latter case 83% nitrogen was removed from the medium. One of the factors causing low efficiency of denitrification of nitrites is excessive alkalization of the medium in the bed. The use of a three-step bed with adjusted pH resulted in complete denitrification of nitrites with efficiency 60 mg N/l/h. The bacteria inside the bed were dominated by Paracoccus denitrificans and by Pseudomonas aeruginosa when nitrates were present. The sensitivity of P. denitrificans to high concentrations of nitrites seems to be the second factor contributing to low efficiency of denitrification with methanol as organic substrate.

Quantitative selection of denitrifying bacteria in continuous cultures and requirement for organic carbon. I. Starch.

A mixed population of bacteria from bottom sludge of nitrogen wastewater reservoir was incubated in continuous culture in medium containing 1000 mg nitrate nitrogen/l and starch. Maximal efficiency of denitrification was 5 mg N/l/h. Marked changes in participation of denitrifying bacteria (16-76%) among total number of bacteria was observed, this being dependent on the ratio of starch concentration (CS) to nitrogen concentration (CN) in the medium. The optimal CS/CN ratio ensuring highest participation of denitrifying bacteria was 3.2. The amount of starch required for the denitrification of a defined quantity of nitrogen is negatively correlated (r = -0.98) with the frequency of the occurrence of denitrifying bacteria (XD) and is: CS = (5.53-0.028XD) CN. The denitrifying bacteria in continuous culture were dominated, depending on CS/CN ratio and flow rate of medium, by Alcaligenes faecalis, Paracoccus denitrificans or Pseudomonas mendocina, that is species unable to hydrolyse starch.

Quantitative selection of denitrifying bacteria in continuous cultures and requirement for organic carbon. II. Maltose.

A mixed population of bacteria from industrial nitrogen wastewaters was incubated in continuous culture in medium containing 1000-1300 mg nitrate nitrogen per litre and maltose as a source of organic carbon. Maximal efficiency of denitrification was 8.6 mg N/1/h. The participation of denitrifying bacteria in the culture varied from 0% to about 80%, depending on the ratio of maltose concentration (CM) to nitrogen concentration (CN) in the medium. The optimal CM/CN ratio ensuring the greatest selection of denitrifying bacteria was 5.0, which calculated per organic carbon (CC/CN) gave the value of 2.1. The amount of maltose needed to denitrify a defined amount of nitrogens was negatively correlated (rxy = -0.95) with the frequency of denitrifying bacteria (XD) in the culture and was: CM = (4.20-0.026XD)CN. The denitrifying bacteria isolated from the studied continuous culture were dominated by Alcaligenes faecalis and Pseudomonas mendocina.

Bacterial microflora participating in the removal of nitrogen from industrial wastewaters by nitrification and denitrifaction.

Nitrification and denitrification were used to purify industrial nitrogenous-organic wastewaters containing 1,130 mg N/l (mainly in the form of ammonium and urea) and with mean COD 1,050 mg O2/l. The use of a five-step system consisting of two denitrifying beds and three nitrifying activated sludge reactors gave 99% removal of nitrogen and 54% reduction in COD during total retention time of the wastes of 81 hours. The participation of stage I nitrifying bacteria in the activated sludge was 4.4 to 5.8% and of stage II nitrifying bacteria 21.9 to 53.8%. Denitrifying bacteria in the denitrifying beds accounted for 90% of the isolated population, which was dominated by Alcaligenes faecalis and Paracoccus denitrificans.

Effect of microbiological hydrolysis of urea on the nitrification process.

The aim of this study was to determine the causes of previously observed inhibition of stage II of nitrification in nitrifying activated sludge treating industrial wastewaters containing ammonium, urea and nitrates. Experiments made with both industrial wastewaters and synthetic medium containing equivalent concentrations of nitrogen compounds demonstrated that the factor limiting the activity of nitrobacteria is microbiological hydrolysis of urea, the causative agent probably being free ammonia released in the course of this hydrolysis.

Denitrification of high concentrations of nitrites and nitrates in synthetic medium with different sources of organic carbon. II. Ethanol.

The course of denitrification of nitrites and nitrates (1000 mg N/l) in medium containing ethanol as a source of organic carbon was studied. Continuous cultures of a mixed population of autochtonous microflora isolated from the bottom sludge of a nitrogenous wastewater reservoir were set up in two types of culture units: chemostat-type column and anaerobic packed bed reactor. In medium with nitrites denitrification was found to be strongly inhibited by alkalization of the medium (efficiency of denitrification 17.7 mg N/l/h). When pH was adjusted the efficiency of nitrite denitrification was identical as for nitrates (166.3 mg N/l/h). The denitrification of 1 g of nitrate or nitrite nitrogen was found to require 2.00 and 1.28 g ethanol, respectively.

Nitrification of industrial waste-waters with high nitrogen concentration in aerated packed-bed reactors.

The use of a two-step aerated packed-bed reactor yielded 99% nitrification of inorganic nitrogenous wastes carrying 800 to 1,300 mg N/l (COD of the wastes 55 mg O2/1). The beds were aerated with air enriched in CO2 pH values between the units were routinely adjusted. The efficiency of nitrification was 33 mg N/l/h. A change of the composition of the wastes to nitrogenous-organic (COD 1,200 mg O2/l) caused a drop in the efficiency of nitrification to 24 mg N/l/h and the subsequent introduction of a three-step system. Concomitant COD reduction was 85% with 73% reduction occurring in the first bed. The number of auto- and heterotropic bacteria in the nitrogenous-organic wastes and effluents from the separate beds were determined.

Effect of various sources of organic carbon and high nitrite and nitrate concentrations on the selection of denitrifying bacteria. I. Stationary cultures.

The effect of methanol, ethanol, acetic acid and glucose together with NaNO2 or KNO3 (1,000 mg N/l) on the intensity of denitrification and selection of denitrifying bacteria from the bottom sludge of nitrogenous wastewater reservoir was examined. Denitrification was found to be the most efficient in medium with ethanol or acetic acid. The presence of glucose facilitated the selection of Alcaligenes faecalis whereas the other carbon sources enabled the selection of bacteria of the genus Pseudomonas: methanol -- P. fluorescens, ethanol -- P. mendocina. In medium with acetic acid species selection depends on the form of nitrogen: NaNO2 -- P. fluorescens, KNO3 -- P. aeruginosa.

Removal of nitrogen from industrial wastewaters by three-step nitrification and denitrification.

The removal of nitrogen from industrial wastewaters carrying about 1,000 mg NH4-N and urea-N/l was investigated on a laboratory scale. The use of a three-step nitryfying activated sludge with adjustment of pH from step to step resulted in 99% oxidation of both forms of nitrogen to nitrites. The efficiency of nitrification was 18 mg N/l/h. Total time of wastewater aeration depended on nitrogen concentration and was 33-54 hours. Complete dentrification of NO2-N was obtained in packed-bed reactor with the use of acetic acid as a carbon source. Efficiency of denitrification was 361 mg N/l/h.

Effect of high concentrations of nitrogen on mixed populations of nitrifying bacteria isolated from industrial nitrogenous wastewaters.

The effect of high concentrations of different forms of nitrogen (NH4+, NO2-, NO3- and urea N) on nitrification by mixed populations of nitrosobacters isolated from nitrogen fertilizer plant wastewaters and nitrobacters isolated from effluents from a biological bed treating these wastewaters was determined. The inhibitory activity (within the concentration values for industrial wastewaters) of only the reaction products was observed, i.e. nitrites for nitrification phase I and nitrates for nitrification phase II. A mixed population of phase II nitrifying bacteria is highly resistant to high concentrations of ammonia nitrogen (50% inhibition at 3,000 mg N/l).