Selection and characterization of aerobic bacteria capable of degrading commercial mixtures of low-ethoxylated nonylphenols

Aims:  Isolation and characterization of new bacterial strains capable of degrading nonylphenol ethoxylates (NP n EO) with a low ethoxylation degree, which are particularly recalcitrant to biodegradation.
Methods and Results:  Seven aerobic bacterial strains were isolated from activated sludges derived from an Italian plant receiving NP n EO-contaminated wastewaters after enrichment with a low-ethoxylated NP n EO mixture. On the basis of 16S rDNA sequence, the strains were positioned into five genera: Ochrobactrum , Castellaniella , Variovorax , Pseudomonas and Psychrobacter . Their degradation capabilities have been evaluated on two commercial mixtures, i.e. Igepal CO-210 and Igepal CO-520, the former rich in low ethoxylated congeners and the latter containing a broader spectrum of NP n EO, and on 4- n -nonylphenol (NP). The strains degraded Igepal CO-210, Igepal CO-520 and 4- n -NP all applied at the initial concentration of 100 mg l⁻¹, by 35–75%, 35–90% and 15–25%, respectively, after 25 days of incubation.
Conclusions:  Some of the isolated strains, in particular the Pseudomonas strains BCb12/1 and BCb12/3, showed interesting degradation capabilities towards low ethoxylated NP n EO congeners maintaining high cell vitality.
Significance and Impact of the Study:  Increased knowledge of bacteria involved in NP n EO degradation and the possibility of using the isolated strains in tailored process for a tertiary biological treatment of effluents of wastewater treatment plants.     

Role of heterotrophic aerobic denitrifying bacteria in nitrate removal from wastewater

With the increase in industrial and agricultural activities, a large amount of nitrogenous compounds are released into the environment, leading to nitrate pollution. The perilous effects of nitrate present in the environment pose a major threat to human and animal health. Bioremediation provides a cost-effective and environmental friendly method to deal with this problem. The process of aerobic denitrification can reduce nitrate compounds to harmless dinitrogen gas. This review provides a brief view of the exhaustive role played by aerobic denitrifiers for tackling nitrate pollution under different ecological niches and their dependency on various environmental parameters. It also provides an understanding of the enzymes involved in aerobic denitrification. The role of aerobic denitrification to solve the issues faced by the conventional method (aerobic nitrification–anaerobic denitrification) in treating nitrogen-polluted wastewaters is elaborated.     

Biochemistry of nitrate respiration in Pseudomonas stutzeri. I. Aerobic and nitrate respiration routes of carbohydrate catabolism

Spangler, W. J. (Oregon State University, Corvallis), and C. M. Gilmour. Biochemistry of nitrate respiration in Pseudomonas stutzeri. I. Aerobic and nitrate respiration routes of carbohydrate catabolism. J. Bacteriol. 91:245-250. 1966.-The metabolic pathways of glucose catabolism were studied in Pseudomonas stutzeri under aerobic conditions and under conditions of nitrate respiration. Studies on both glucose and gluconate catabolism, by the radiorespirometric method, indicated that these substrates are degraded in the same manner, i.e., the Entner-Doudoroff and pentose phosphate pathways. There appeared to be no major shift in primary metabolic pathways when nitrate was used as the terminal hydrogen acceptor in nitrate respiration as opposed to aerobic respiration with free molecular oxygen. It was shown that glucose is not degraded to any appreciable extent under anaerobic conditions in the absence of nitrate. Tentative evidence suggests that the tricarboxylic acid cycle functions under both conditions of oxygen relationships and that the rate of carbon oxidation via the tricarboxylic acid cycle is slower with nitrate respiration than under aerobic conditions.     

Oxygen and nitrate respiration in a reed swamp sediment from a eutrophic lake

Seasonal measurements of the oxygen and nitrate uptake by a reed swamp sediment were carried out in a shallow, eutrophic Danish lake, Arreskov Sø. The oxidation of organic carbon in the sediment by aerobic and nitrate respiration was 290 and 188 g C m⁻² yr⁻¹ respectively. During winter, nitrate respiration amounted to 94% of the total carbon oxidation, whereas it was zero during summer. On an annual basis nitrate respiration constituted 39% of total respiration. Sediment nitrate uptake was correlated to nitrate concentration. In consequence of this the nitrate uptake rates varied during the year from zero in summer to 55 mg N m⁻² d⁻¹ in spring.
Oxygen uptake rates varied from 30 to 250 mg O₂ m⁻² h⁻¹ during the year, with a maximum uptake in August. The oxygen uptake per year was calculated to 860 g O₂ m⁻². The oxygen uptake rate was correlated to lake temperature and Kjeldahl nitrogen content of the sediment. The oxygen uptake rate, however, showed no correlation with loss on ignition of the sediment. A Q₁₀-value of 2.2 was found for lake measurements in the temperature interval of 5–15°C. The corresponding O₁₀-value in the laboratory was 2.6. A high microbial biomass indicated by the maximum content of Kjeldahl nitrogen and the lowest ratio of loss on ignition on Kjeldahl nitrogen appeared in late August, when the maximum oxygen uptake occurred. The oxygen uptake rate increased during the time interval from sampling to the start of the experiments.     

虾池中具有降解硝酸盐或亚硝酸盐能力的细菌多样性

水产养殖过程中,氮素积累日益严重,而其中亚硝酸盐由于转化速度低,毒性强,对养殖的危害更加突出。对从对虾养殖池中通过反硝化条件选择性富集培养得到的具有去除硝酸盐及亚硝酸盐能力的细菌进行筛选,结果分离到27株能够还原硝酸盐的异养细菌,其中24株在7d内能有效地降低硝酸盐和亚硝酸盐浓度;特别是LZX22、LZX27、LZX23、LZX21等4株使硝酸盐氮由起始的422．25mg／L降至4．00mg／L以下,亚硝酸盐浓度也降至0．40mg／L以下。对这些菌株进行16S rDNA系统发育分析,结果显示：27株菌分属于5个不同的类群,α-Proteobacteria（1）,γ-Proteobacteria（10）,Actinobacteria（12）,Firmicutes（3）,和Bacteroides（1）;它们在系统发育上分别与11个属相近,分别是Pseudomonas,Halomonas,Acinetobacter,Paracoccus,Arthrobacter,Microbacterium,Cellulosimicrobium,Bacillus,Stenotrophom,和Sphingobacterium。表明所分析的虾池中具有去除硝酸盐和亚硝酸盐能力的细菌具有较高的多样性,特别是多株细菌为首次报道具有去除硝酸盐和亚硝酸盐的能力,为下一步筛选亚硝酸盐高效去除细菌提供了丰富的菌种资源。     

Nitrogenase activity and nitrate respiration inAzospirillum spp.

The interaction between nitrate respiration and nitrogen fixation inAzospirillum lipoferum andA. brasilense was studied. All strains examined were capable of nitrogen fixation (acetylene reduction) under conditions of severe oxygen limitation in the presence of nitrate. A lag phase of about 1 h was observed for both nitrate reduction and nitrogenase activity corresponding to the period of induction of the dissimilatory nitrate reductase. Nitrogenase activity ceased when nitrate was exhausted suggesting that the reduction of nitrate to nitrite, rather than denitrification (the further reduction of nitrite to gas) is coupled to nitrogen fixation. The addition of nitrate to nitrate reductase negative mutants (nr^-) ofAzospirillum did not stimulate nitrogenase activity. Under oxygen-limited conditionsA. brasilense andA. lipoferum were also shown to reduce nitrate to ammonia, which accumulated in the medium. Both species, including strains ofA. brasilense which do not possess a dissimilatory nitrite reductase (nir^-) were also capable of reducing nitrous oxide to N₂.     

Cryptic niche differentiation of novel sediment ecotypes of Ruegeria pomeroyi correlates with nitrate respiration

Marine intertidal sediments fluctuate in redox conditions and nutrient availability, and they are also known as an important sink of nitrogen mainly through denitrification, yet how denitrifying bacteria adapt to this dynamic habitat remains largely untapped. Here, we investigated novel intertidal benthic ecotypes of the model pelagic marine bacterium Ruegeria pomeroyi DSS-3 with a population genomic approach. While differing by only 1.3% at the 16S rRNA gene level, members of the intertidal benthic ecotypes are complete denitrifiers whereas the pelagic ecotype representative (DSS-3) is a partial denitrifier lacking a nitrate reductase. The intertidal benthic ecotypes are further differentiated by using non-homologous nitrate reductases and a different set of genes that allow alleviating oxidative stress and acquiring organic substrates. In the presence of nitrate, the two ecotypes showed contrasting growth patterns under initial oxygen concentrations at 1 vol% versus 7 vol% and supplemented with different carbon sources abundant in intertidal sediments. Collectively, this combination of evidence indicates that there are cryptic niches in coastal intertidal sediments that support divergent evolution of denitrifying bacteria. This knowledge will in turn help understand how these benthic environments operate to effectively remove nitrogen.     

Distribution of heterotrophic aerobic marine bacteria in sediment in Eastern Harbour of Alexandria

In Eastern Harbour (EH) of Alexandria (Egypt), where an under-water museum is planned to be built, the distribution of some heterotrophic bacteria and their relationships to physical parameters and biochemical composition of the sedimentary organ ic matter were investigated. The Eastern Harbour is a relatively shallow semi-closed basin and is sheltered from the sea by an art ificial breakwater leaving two openings, El-Boughaz and El-Silsila. Seven stations covering the area of the EH were selected and surface sediments were collected seasonally from spring to winter 2002. The near bottom temperature varied seasonally with a minimum value in winter and a maximum in summer. In contrast to the temperature values, dissolved oxygen exhibited maximum values in cold seasons. The seasonal average of the total organic carbon ranged from 0.48 ± 0.16 to 4.42 ± 2.46%, while the total organic nitroge n ranged from 0.07±0.08 to 0.42±0.38%. The total carbohydrate had minimum and maximum values of 273 μg g^?1 and 6539 μg g^?1. The combined amino acids represented the dominant biochemical class of organic matter in the EH sediments with an average of 365 ± 1911 μg g^?1. The total bacterial count ranged from 1.4 × 10⁴ to 1.4 × 10⁷ colony forming unit (CFU) g^?1 sediment dry weight. Amylolytic bacterial group was recorded in almost all sites and seasons, while proteolytic bacteria were dominant in spring and au tumn. The variation in the abundance of amylolytic and proteolytic bacterial groups was found to be parallel to the variation in soluble carbohydrates and free amino acids. High percentage of H₂S-producing bacteria was reported during summer at some stations confirming the low oxygen content of the sediment at these sites. Agar-degrading bacteria were found only in warm seasons. The count of co liform bacteria in the EH sediments was very low (<10 CFU g^?1) during all monitored seasons indicating that the EH marine environment was almost free of domestic waste discharge during this period.     

Nitrate respiration,denitrification, and utilization of nitrogen sources by aerobic carbon monoxide-oxidizing bacteria

We describe the ability of carboxydotrophic bacteria for nitrate respiration or denitrification. Four out of fourteen strains examined could denitrify heterotrophically forming N₂ (Pseudomonas carboxydoflava) or N₂O (Pseudomonas carboxydohydrogena, Pseudomonas compransoris, and Pseudomonas gazotropha). Three carried out a heterotrophic nitrate respiration (Arthrobacter 11/x, Azomonas B1, and Azomonas C2). P. carboxydohydrogena could use H₂ as electron donor for nitrate respiration under chemolithoautotrophic growth conditions. CO did not support denitrification or nitrate respiration of carboxydotrophic bacteria, although the free energy changes of the reactions would be sufficiently negative to allow growth. CO at 50 kPa was a weak inhibitor of N₂O-reduction in carboxydotrophic and non-carboxydotrophic bacteria and decelerated denitrifying growth. Carboxydotrophic bacteria could utilize a wide range of N-sources. Results obtained with a plasmid-cured mutant of Pseudomonas carboxydovorans OM5 showed, that genes involved in nitrogen assimilation entirely reside on the chromosome. In the presence of an suitable electron donor, most carboxydotrophic bacteria could carry out a reduction of nitrate to nitrite that did not support growth and did not lead to the formation of ammonia.This article is dedicated to Professor Hans G. Schlegel on the occasion of his 65th birthday and in admiration for his élan and eternal idealism     

Isolation and characterization of aerobic bacteria capable of the degradation of synthetic and natural melanoidins from distillery effluent

Melanoidins, complex biopolymer of amino-carbonyl compounds are the major coloring and polluting constituents of distillery wastewaters. In this study, three aerobic melanoidin-degrading bacteria (RNBS1, RNBS3 and RNBS4) were isolated from soil contaminated with distillery effluent and characterized as Bacillus licheniformis (RNBS1), Bacillus sp. (RNBS3) and Alcaligenes sp. (RNBS4) by biochemical tests and 16S rRNA gene sequence analysis. The degradation of synthetic and natural melanoidins was studied by using the axenic and mixed bacterial consortium. Results have revealed that the mixed consortium was more effective compared to axenic culture decolorizing 73.79 and 69.83% synthetic and natural melanoidins whereas axenic cultures RNBS1, RNBS3 and RNBS4 decolorized 65.88, 62.56 and 66.10% synthetic and 52.69, 48.92 and 59.64% natural melanoidins, respectively. The HPLC analysis of degraded samples has shown reduction in peak areas compared to controls, suggesting that decrease in color intensity might be largely attributed to the degradation of melanoidins by isolated bacteria.     

Importance of unattached bacteria and bacteria attached to sediment in determining potentials for degradation of xenobiotic organic contaminants in an aerobic aquifer.

The bacterial abundance, distribution, and degradation potential (in terms of degradation versus lack of degradation) for four xenobiotic compounds in an aerobic aquifer sediment have been examined in laboratory and field experiments. The xenobiotic compounds studied were benzene, toluene, o-xylene, and naphthalene (all at concentrations of approximately 120 micrograms/liter). The aerobic degradation experiments ran for approximately 90 days at 10 degrees C, which corresponded to the groundwater temperature. At the end of the experiment, the major part of the microbial biomass, quantified as acridine orange direct counts, was attached to the groundwater sediment (18 x 10(6) to 25 x 10(6) cells per g [dry weight], and only a minor part was unattached in the groundwater (0.6 x 10(6) to 5.5 x 10(6) cells per ml). Experiments involving aquifer sediment suspensions showed identical degradation potentials in the laboratory and in the field. However, laboratory experiments involving only groundwater (excluding aquifer sediment) showed less degradation potential than in situ experiments involving only groundwater, indicating that the manipulation or approach of the laboratory experiments could affect the determination of the degradation potentials. No differences were observed between the groundwater-only and the sediment compartments in the in situ experiments in the ability to degrade the compounds, but the maximum degradation rates were substantially lower in the groundwater-only compartment. Preparations used in laboratory experiments for studying the degradation potential for xenobiotic organic contaminants should contain sediment to obtain the highest numbers of bacteria as well as the broadest and most stable degradation.(ABSTRACT TRUNCATED AT 250 WORDS)     

Importance of unattached bacteria and bacteria attached to sediment in determining potentials for degradation of xenobiotic organic contaminants in an aerobic aquifer.

The bacterial abundance, distribution, and degradation potential (in terms of degradation versus lack of degradation) for four xenobiotic compounds in an aerobic aquifer sediment have been examined in laboratory and field experiments. The xenobiotic compounds studied were benzene, toluene, o-xylene, and naphthalene (all at concentrations of approximately 120 micrograms/liter). The aerobic degradation experiments ran for approximately 90 days at 10 degrees C, which corresponded to the groundwater temperature. At the end of the experiment, the major part of the microbial biomass, quantified as acridine orange direct counts, was attached to the groundwater sediment (18 x 10(6) to 25 x 10(6) cells per g [dry weight], and only a minor part was unattached in the groundwater (0.6 x 10(6) to 5.5 x 10(6) cells per ml). Experiments involving aquifer sediment suspensions showed identical degradation potentials in the laboratory and in the field. However, laboratory experiments involving only groundwater (excluding aquifer sediment) showed less degradation potential than in situ experiments involving only groundwater, indicating that the manipulation or approach of the laboratory experiments could affect the determination of the degradation potentials. No differences were observed between the groundwater-only and the sediment compartments in the in situ experiments in the ability to degrade the compounds, but the maximum degradation rates were substantially lower in the groundwater-only compartment. Preparations used in laboratory experiments for studying the degradation potential for xenobiotic organic contaminants should contain sediment to obtain the highest numbers of bacteria as well as the broadest and most stable degradation.(ABSTRACT TRUNCATED AT 250 WORDS)     

The influence of nitrate concentration upon the end-products of nitrate dissimilation by bacteria in anaerobic salt marsh sediment

Abstract Experiments were carried out with slurries of saltmarsh sediment to which varying concentrations of nitrate were added. The acetylene blocking technique was used to measure denitrification by accumulation of nitrous oxide, while reduction of nitrate to nitrite and ammonium was also measured. There was good recovery of reduced nitrate and at the smallest concentration of nitrate used (250 μM) there was approximately equal reduction to either ammonium or nitrous oxide (denitrification). Nitrite was only a minor end-product of nitrate reduction. As the nitrate concentration was increased the proportion of the nitrate which was denitrified to nitrous oxide increased, to 83% at the greatest nitrate concentration used (2 mM), while reduction to ammonium correspondingly decreased. This change was attributed either to a greater competitiveness by the denitrifiers for nitrate as the ratio of electron donor to electron acceptor decreased; or to the increased production of nitrite rather than ammonium by fermentative bacteria under high nitrate, the nitrite then being reduced to nitrous oxide by denitrifying bacteria.     

Regulators of aerobic and anaerobic respiration in Bacillus subtilis.

Two Bacillus subtilis genes, designated resD and resE, encode proteins that are similar to those of two-component signal transduction systems and play a regulatory role in respiration. The overlapping resD-resE genes are transcribed during vegetative growth from a very weak promoter directly upstream of resD. They are also part of a larger operon that includes three upstream genes, resABC (formerly orfX14, -15, and -16), the expression of which is strongly induced postexponentially. ResD is required for the expression of the following genes: resA, ctaA (required for heme A synthesis), and the petCBD operon (encoding subunits of the cytochrome bf complex). The resABC genes are essential genes which encode products with similarity to cytochrome c biogenesis proteins. resD null mutations are more deleterious to the cell than those of resE. resD mutant phenotypes, directly related to respiratory function, include streptomycin resistance, lack of production of aa3 or caa3 terminal oxidases, acid accumulation when grown with glucose as a carbon source, and loss of ability to grow anaerobically on a medium containing nitrate. A resD mutation also affected sporulation, carbon source utilization, and Pho regulon regulation. The data presented here support an activation role for ResD, and to a lesser extent ResE, in global regulation of aerobic and anaerobic respiration i B.subtilis.     

Isolation of microorganisms capable of degrading isoquinoline under aerobic conditions.

Isoquinoline-degrading microbial cultures were isolated from oil- and creosote-contaminated soils. The establishment of initial enrichment cultures required the use of emulsified isoquinoline. Once growth on isoquinoline was established, isoquinoline emulsification was no longer required for utilization of isoquinoline as the sole source of carbon and nitrogen by these cultures. An isoquinoline-degrading Acinetobacter strain was isolated from one of the enrichment cultures. The degradation of isoquinoline was accompanied by the accumulation of a red cell-associated pigment and of 1-hydroxyisoquinoline, which was further degraded to unknown intermediary ring-cleavage products and carbon dioxide.     

Phosphorus composition and release in sediment bacteria of the genus Pseudomonas during aerobic and anaerobic conditions

A substantial amount of sediment phosphorus can be bound in bacterial biomass. In this study the fractional composition of phosphorus in the bacteria Pseudomonas was determined by sequential extraction with ammonium chloride, sodium hydroxide and hydrochloric acid according to the scheme of Hieltjes & Lijklema (1980). Both non-labelled and ³²P-labelled bacteria were used for fractionation. Up to 80% of the bacterial phosphorus was found in the NaOH-nRP fraction, which is in agreement with the results of Hupfer & Uhlman (1992) for Acinetobacter and activated sludge obtained with the sequential extraction scheme of Psenner et al. (1985). A significant correlation was found between bacterial biomass and the amount of phosphorus retained in the NaOH-nRP fraction when sediments were fractionated. Additional experiments with ³²P-labelled Pseudomonas in sediment-water systems were performed in order to follow bacterial release of phosphorus under aerobic and anaerobic conditions. These studies did not sustain the hypothesis that anaerobic conditions lead to rapid release of phosphorus from bacterial cells.     

Enrichment of mixed cultures capable of aerobic degradation of 1,2-dibromoethane.

1,2-dibromoethane (DBE) is a common environmental contaminant; it is potentially carcinogenic and has been detected in soil and groundwater supplies. Most of the biodegradation studies to date have been performed under anaerobic conditions or in the context of soil remediation, where the pollutant concentration was in the parts per billion range. In this work a mixed bacterial culture capable of complete aerobic mineralization of concentrations of DBE up to 1 g liter(-1) under well-controlled laboratory conditions was enriched. In order to verify biodegradation, formation of biodegradation products as well as the disappearance of DBE from the biological medium were measured. Complete mineralization was verified by measuring stoichiometric release of the biodegradation products. This mixed culture was found to be capable of degrading other halogenated compounds, including bromoethanol, the degradation of which has not been reported previously.