Microbial diversity and anaerobic hydrocarbon degradation potential in an oil-contaminated mangrove sediment

ABSTRACT: BACKGROUND: Mangrove forests are coastal wetlands that provide vital ecosystem services and serve as barriers against natural disasters like tsunamis, hurricanes and tropical storms. Mangroves harbour a large diversity of organisms, including microorganisms with important roles in nutrient cycling and availability. Due to tidal influence, mangroves are sites where crude oil from spills farther away can accumulate. The relationship between mangrove bacterial diversity and oil degradation in mangrove sediments remains poorly understood. RESULTS: Mangrove sediment was sampled from 0--5, 15--20 and 35--40 cm depth intervals from the Surui River mangrove (Rio de Janeiro, Brazil), which has a history of oil contamination. DGGE fingerprinting for bamA, dsr and 16S rRNA encoding fragment genes, and qPCR analysis using dsr and 16S rRNA gene fragment revealed differences with sediment depth. CONCLUSIONS: Analysis of bacterial 16S rRNA gene diversity revealed changes with depth. DGGE for bamA and dsr genes shows that the anaerobic hydrocarbon-degrading community profile also changed between 5 and 15 cm depth, and is similar in the two deeper sediments, indicating that below 15 cm the anaerobic hydrocarbon-degrading community appears to be well established and homogeneous in this mangrove sediment. qPCR analysis revealed differences with sediment depth, with general bacterial abundance in the top layer (0--5 cm) being greater than in both deeper sediment layers (15--20 and 35--40 cm), which were similar to each other.     

Changes in bacterial communities from anaerobic digesters during petroleum hydrocarbon degradation

Anaerobic biodegradation of petroleum hydrocarbons (PHC) to methane has been recognized to occur in oil reservoirs and contaminated surface sites alike. This process could be employed efficiently for the treatment of contaminated materials, including petrochemical wastes and PHC-contaminated soil, since no external electron acceptor is required. Moreover, the controlled production of methane in digestion plants, similarly to the anaerobic digestion (AD) of energy crops or organic residues, would enable for energy recovery from these wastes. At present, little is known about the bacterial communities involved in and responsible for hydrocarbon fermentation, the initial step in PHC conversion to methane. In the present study, the fate of two different methanogenic communities derived from the AD of wastewater (WWT) and of biowaste, mixed with PHC-contaminated soil (SWT), was monitored during incubation with PHC using denaturing gradient gel electrophoresis (DGGE) of 16S rDNA genes amplified with Bacteria-specific primers. During 11 months of incubation, slight but significant degradation of PHC occurred in both sludges and distinct bacterial communities were developing. In both sludges, Bacteroidetes were found. In addition, in WWT, the bacterial community was found to be dominated by Synergistetes and Proteobacteria, while Firmicutes and unidentified members were abundant in SWT. These results indicate that bacterial communities from anaerobic digesters can adapt to and degrade petroleum hydrocarbons. The decontamination of PHC-containing waste via fermentative treatment appears possible.     

Positive FNR-like control of anaerobic arginine degradation and nitrate respiration in Pseudomonas aeruginosa.

A mutant of Pseudomonas aeruginosa was characterized which could not grow anaerobically with nitrate as the terminal electron acceptor or with arginine as the sole energy source. In this anr mutant, nitrate reductase and arginine deiminase were not induced by oxygen limitation. The anr mutation was mapped in the 60-min region of the P. aeruginosa chromosome. A 1.3-kb chromosomal fragment from P. aeruginosa complemented the anr mutation and also restored anaerobic growth of an Escherichia coli fnr deletion mutant on nitrate medium, indicating that the 1.3-kb fragment specifies an FNR-like regulatory protein. The arcDABC operon, which encodes the arginine deiminase pathway enzymes of P. aeruginosa, was rendered virtually noninducible by a deletion or an insertion in the -40 region of the arc promoter. This -40 sequence (TTGAC....ATCAG) strongly resembled the consensus FNR-binding site (TTGAT....ATCAA) of E. coli. The cloned arc operon was expressed at low levels in E. coli; nevertheless, some FNR-dependent anaerobic induction could be observed. An FNR-dependent E. coli promoter containing the consensus FNR-binding site was expressed well in P. aeruginosa and was regulated by oxygen limitation. These findings suggest that P. aeruginosa and E. coli have similar mechanisms of anaerobic control.     

Adding handles to unhandy substrates: anaerobic hydrocarbon activation mechanisms

In spite of their chemical inertness, hydrocarbons are degraded by microorganisms in the complete absence of oxygen. As all known aerobic hydrocarbon degradation pathways start with oxygen-dependent reactions, hydrocarbon catabolism in anaerobes must be initiated by novel biochemical reactions. In recent years, the enzymes catalyzing oxygen-independent activation of several hydrocarbons have been identified. Surprisingly, a variety of reactions seems to be employed to overcome the activation barrier of different hydrocarbons. This review presents the current understanding on some of these reactions and the associated degradation pathways: oxygen-independent hydroxylation as employed in ethylbenzene metabolism, fumarate addition to methyl or methylene carbons in toluene or alkane degradation, and only recently discovered reactions such as methylation of naphthalene or anaerobic methane oxidation via reverse methanogenesis.     

水体沉积物中微生物厌氧呼吸耦合多环芳烃降解的研究进展

多环芳烃(Polycyclic aromatic hydrocarbons,PAHs)是一种具有致癌、致畸、致突变的持久性有机污染物。本文在分析国内外主要水体沉积物中PAHs污染状况的基础上,综述了近几年有关厌氧水体沉积物中微生物以硝酸盐、Fe(III)以及硫酸盐为电子受体进行呼吸耦合PAHs降解的研究概况。此外,还总结了基于微生物的PAHs降解基因组、蛋白质组、代谢组以及菌群水平上互作网络的研究进展,以期为进一步加速PAHs污染水体沉积物原位生物修复提供科学理论参考。     

Sulfonates: novel electron acceptors in anaerobic respiration

The enrichment and isolation in pure culture of a bacterium, identified as a strain of Desulfovibrio, able to release and reduce the sulfur of isethionate (2-hydroxyethanesulfonate) and other sulfonates to support anaerobic respiratory growth, is described. The sulfonate moiety was the source of sulfur that served as the terminal electron acceptor, while the carbon skeleton of isethionate functioned as an accessory electron donor for the reduction of sulfite. Cysteate (alanine-3-sulfonate) and sulfoacetaldehyde (acetaldehyde-2-sulfonate) could also be used for anaerobic respiration, but many other sulfonates could not. A survey of known sulfate-reducing bacteria revealed that some, but not all, strains tested could utilize the sulfur of some sulfonates as terminal electron acceptor. Isethionate-grown cells of Desulfovibrio strain IC1 reduced sulfonate-sulfur in preference to that of sulfate; however, sulfate-grown cells reduced sulfate-sulfur in preference to that of sulfonate. Received: 2 May 1996 / Accepted: 8 June 1996     

Mixed-valence cytoplasmic iron granules are linked to anaerobic respiration

Intracellular granules containing ferric and ferrous iron formed in Shewanella putrefaciens CN32 during dissimilatory reduction of solid-phase ferric iron. It is the first in situ detection at high resolution (150 nm) of a mixed-valence metal particle residing within a prokaryotic cell. The relationship of the internal particles to Fe(III) reduction may indicate a respiratory role.     

Pseudomonas aeruginosa anaerobic respiration in biofilms: relationships to cystic fibrosis pathogenesis

Recent data indicate that cystic fibrosis (CF) airway mucus is anaerobic. This suggests that Pseudomonas aeruginosa infection in CF reflects biofilm formation and persistence in an anaerobic environment. P. aeruginosa formed robust anaerobic biofilms, the viability of which requires rhl quorum sensing and nitric oxide (NO) reductase to modulate or prevent accumulation of toxic NO, a byproduct of anaerobic respiration. Proteomic analyses identified an outer membrane protein, OprF, that was upregulated approximately 40-fold under anaerobic versus aerobic conditions. Further, OprF exists in CF mucus, and CF patients raise antisera to OprF. An oprF mutant formed poor anaerobic biofilms, due, in part, to defects in anaerobic respiration. Thus, future investigations of CF pathogenesis and therapy should include a better understanding of anaerobic metabolism and biofilm development by P. aeruginosa.     

Bacterial anaerobic respiration and electron transfer relevant to the biotransformation of pollutants

Bacterial anaerobic respiration is one of the most ancient and essential metabolism processes, possessing the characteristics of both flexibility and high diversity, and a very close relationship with the physiological function in the ecological environment. Under anaerobic conditions, bacteria and anthropogenic substances can form coupling process facilitating terminal electron transfer. Several forms of bacterial anaerobic respiration and electron transfer related to the biotransformation of pollutants, including respiration with humics, sulfonates, halogenated chemicals, azo compounds, TNTs, metallic and non-metallic elements, are reviewed in this paper. These respirations and electron transfers on diverse electron acceptors in the environment have important biotechnological implications because these biochemical reactions have their roles on the transformation/degradation of toxic substances and the cycling of organic carbon as well as many inorganic elements. Furthermore, remediation of sites contaminated with toxic pollutants based on bacterial anaerobic respirations is being recognized widely.     

A new hydrocarbon degradation fungus: Verticillium ecanii

Summary Verticillium lecanii was isolated from tar lumps resulting from a marine oil spill. It has not previously been reported to utilize hydrocarbons.     

Aerobic and anaerobic respiration in the intact spinach chloroplast

Aerobic and anaerobic chloroplastic respiration was monitored by measuring ¹⁴CO₂ evolution from [¹⁴C]glucose in the darkened spinach (Spinacia oleracea) chloroplast and by estimating the conversion of fructose 1,6-bisphosphate to glycerate 3-phosphate in the darkened spinach chloroplast in air with O₂ or in N₂ with nitrite or oxaloacetate as electron acceptors. The pathway of ¹⁴CO₂ evolution from labeled glucose in the absence and presence of the inhibitors iodoacetamide and glycolate 2-phosphate under air or N₂ were those expected from the oxidative pentose phosphate cycle and glycolysis. Of the electron acceptors, O₂ was the best (2.4 nanomoles CO₂ per milligram chlorophyll per hour), followed by nitrite and oxaloacetate. With respect to glycerate 3-phosphate formation from fructose 1,6-bisphosphate, methylene blue increased the aerobic rate from 3.7 to 5.4 micromoles per milligram chlorophyll per hour. A rate of 4.8 micromoles per milligram chlorophyll per hour was observed under N₂ with nitrite and oxaloacetate.     

Aerobic and anaerobic bacterial respiration monitored by electrodes.

A technique is described by which both oxygen and nitrate (or nitrate or chlorate) levels were continuously monitored during bacterial respiration. Paracoccus (Micrococcus) denitrificans and Escherichia coli oxidizing succinate rapidly ceased to reduce nitrate when oxygen was available, and equally rapidly commenced nitrate reduction when all the oxygen had been consumed. By contrast, membrane vesicles isolated from P. denitrificans reduced oxygen and nitrate simultaneously. The respiratory nitrate reductase in intact cells of P. denitrificans appeared to be inacessible to chlorate present in the reaction medium, and it is suggested that the nitrate reductase is orientated on the plasma membrane so that nitrate gains access from the inner (cytosolic) face.     

The anaerobic oyster heart: Coupling of glucose and aspartate fermentation

Summary The interrelationships of carbohydrate and amino acid metabolism during anaerobiosis were investigated in the ventricle of the intertidal oyster,Crassostrea gigas. While the ventricle accumulates alanine and succinate in a 21 ratio during anoxia, these end products appear to arise from different precursors. Thus glucose-¹⁴C is metabolized mainly to alanine-¹⁴C (55% of glucose carbon appears in alanineversus 3% in succinate) by the anoxic ventriclein vitro while succinate-¹⁴C is the principle end product of aspartate-¹⁴C catabolism. Glutamate-¹⁴C is poorly metabolized by the anoxic ventricle, and correspondingly, while ventricular aspartate concentrations drop during anoxia, those of other amino acids do not. A metabolic scheme coupling glucose and aspartate catabolism in this facultative anaerobe is proposed. The detection of a third, as yet incompletely identified, anaerobic end product produced by the ventricle is reported.     

Microbial petroleum degradation: use of mixed hydrocarbon substrates

Methods of examining hydrocarbons to estimate the microbial degradation of petroleum are compared. Gas-liquid chromatography with a mixed hydrocarbon substrate has been shown to be useful in evaluating microbial potential for degradation of a number of hydrocarbons.     

Fermentation and anaerobic respiration by Rhodospirillum rubrum and Rhodopseudomonas capsulata

Rhodospirillum rubrum and Rhodopseudomonas capsulata were able to grow anaerobically in the dark either by a strict mixed-acid fermentation of sugars or, in the presence of an appropriate electron acceptor, by an energy-linked anaerobic respiration. Both species fermented fructose without the addition of accessory oxidants, but required the initial presence of bicarbonate before fermentative growth could begin. Major products of R. rubrum fermentation were succinate, acetate, propionate, formate, hydrogen, and carbon dioxide; R. capsulata produced major amounts of lactate, acetate, succinate, hydrogen, and carbon dioxide. R. rubrum and R. capsulata were also capable of growing strictly through anaerobic, respiratory mechanisms. Nonfermentable substrates, such as succinate, malate, or acetate, supported growth only in the presence of an electron acceptor such as dimethyl sulfoxide or trimethylamine oxide. Carbon dioxide and dimethyl sulfide were produced during growth of R. rubrum and R. capsulata on succinate plus dimethyl sulfoxide. Molar growth yields from cultures grown anaerobically in the dark on fructose plus dimethyl sulfoxide were 3.8 to 4.6 times higher than values obtained from growth on fructose alone and were 56 to 60% of the values obtained from aerobic, respiratory growth with fructose. Likewise, molar growth yields from anaerobic, respiratory growth conditions with succinate plus dimethyl sulfoxide were 51 to 54% of the values obtained from aerobic, respiratory growth with succinate. The data indicate that dimethyl sulfoxide or trimethylamine oxide as a terminal oxidant is approximately 33 to 41% as efficient as O₂ in conserving energy through electron transport-linked respiration.