Exploring the capacity for anaerobic biodegradation of polycyclic aromatic hydrocarbons and naphthenic acids by microbes from oil-sands-process-affected waters

Both polycyclic aromatic hydrocarbons (PAHs) and naphthenic acids (NAs) are natural components of fossil fuels, but they are also widespread toxic and environmentally persistent pollutants. They are the major cause of environmental toxicity in oil-sands-process waters (OSPW). This study aimed to investigate the anaerobic biodegradation of the PAHs pyrene and 2-methylnaphthalene, and the NAs adamantane-1-carboxylic acid and a "natural" NA mixture (i.e., acid-extractable NAs from OSPW) under sulfate-reducing and methanogenic conditions by a microbial community derived from an oil sands tailings pond. Using gas-chromatography mass spectrometry (GC-MS), the rate of biodegradation was measured in relation to changes in bacterial community composition. Only 2-methylnaphthalene was significantly degraded after 260 days, with significantly more degradation under sulfate-reducing (40%) than methanogenic conditions (25%). During 2-methylnaphthalene biodegradation, a major metabolite was produced and tentatively identified as 2-naphthoic acid. Denaturing gradient gel electrophoresis (DGGE) demonstrated an increase in intensity of bands during the anaerobic biodegradation of 2-methylnaphalene, which derived from species of the genera Fusibacter, Alkaliphilus, Desulfobacterium, Variovorax, Thaurea, and Hydrogenophaga. Despite the biodegradation of 2-methylnaphthalene, this study demonstrates that, under anaerobic conditions, NAs and high-molecular-weight PAHs are the predominant molecules likely to persist in OSPW. Therefore alternative remediation strategies are required

A stable genetic polymorphism underpinning microbial syntrophy

Syntrophies are metabolic cooperations, whereby two organisms co-metabolize a substrate in an interdependent manner. Many of the observed natural syntrophic interactions are mandatory in the absence of strong electron acceptors, such that one species in the syntrophy has to assume the role of electron sink for the other. While this presents an ecological setting for syntrophy to be beneficial, the potential genetic drivers of syntrophy remain unknown to date. Here, we show that the syntrophic sulfate-reducing species Desulfovibrio vulgaris displays a stable genetic polymorphism, where only a specific genotype is able to engage in syntrophy with the hydrogenotrophic methanogen Methanococcus maripaludis. This 'syntrophic' genotype is characterized by two genetic alterations, one of which is an in-frame deletion in the gene encoding for the ion-translocating subunit cooK of the membrane-bound COO hydrogenase. We show that this genotype presents a specific physiology, in which reshaping of energy conservation in the lactate oxidation pathway enables it to produce sufficient intermediate hydrogen for sustained M. maripaludis growth and thus, syntrophy. To our knowledge, these findings provide for the first time a genetic basis for syntrophy in nature and bring us closer to the rational engineering of syntrophy in synthetic microbial communities

Co-existence of physiologically similar sulfate-reducing bacteria in a full-scale sulfidogenic bioreactor fed with a single organic electron donor

A combination of culture-dependent and independent methods was used to study the co-existence of different sulfate-reducing bacteria (SRB) in an upflow anaerobic sludge bed reactor treating sulfate-rich wastewater. The wastewater was fed with ethanol as an external electron donor. Twenty six strains of SRB were randomly picked and isolated from the highest serial dilution that showed growth (i.e. 108). Repetitive enterobacterial palindromic polymerase chain reaction and whole cell protein profiling revealed a low genetic diversity, with only two genotypes among the 26 strains obtained in the pure culture. The low genetic diversity suggests the absence of micro-niches within the reactor, which might be due to a low spatial and temporal micro-heterogeneity. The total 16S rDNA sequencing of two representative strains L3 and L7 indicated a close relatedness to the genus Desulfovibrio. The two strains differed in as many as five physiological traits, which might allow them to occupy distinct niches and thus co-exist within the same habitat. Whole cell hybridisation with fluorescently labeled oligonucleotide probes was performed to characterise the SRB community in the reactor. The isolated strains Desulfovibrio L3 and Desulfovibrio L7 were the most dominant SRB, representing 30–35% and 25–35%, respectively, of the total SRB community. Desulfobulbus-like bacteria contributed for 20–25%, and the Desulfobacca acetoxidans-specific probe targeted approximately 15–20% of the total SRB. The whole cell hybridisation results thus revealed a consortium of four different species of SRB that can be enriched and maintained on a single energy source in a full-scale sulfidogenic reactor

Anaerobic degradability of wool scouring effluent

The anaerobic degradability of wool scouring effluent was investigated in batch cultures. The results were compared to the degradation of cellulose and sterile activated sludge. Wool scouring effluent was clearly more difficult to degrade anaerobically than cellulose or the biomass of activated sludge. The maximum biogas production rate from wool scouring effluent was about 46% and 31 % of the maximal rates obtained from activated sludge and cellulose powder, respectively. The slow conversion rate and also the low percentage of the organics degraded showed that wool scouring effluent is particularly difficult to degrade anaerobically. Our results suggest that very long residence times (> 30 days) are required to successfully convert most of the organics in wool scouring effluent into biogas. The large digester size required questions the economics of such a treatment. However, in contrast to waste streams containing communal wastes, biomass waste or carbohydrates, wool scouring effluent as feed material is unlikely to cause digester failure by acidification, which would make its anaerobic digestion more stable and more easily controlable

Concurrent lactic and volatile fatty acid analysis of microbial fermentation samples by gas chromatography with heat pre-treatment

Organic acid analysis of fermentation samples can be readily achieved by gas chromatography (GC), which detects volatile organic acids. However, lactic acid, a key fermentation acid is non-volatile and can hence not be quantified by regular GC analysis. However the addition of periodic acid to organic acid samples has been shown to enable lactic acid analysis by GC, as periodic acid oxidizes lactic acid to the volatile acetaldehyde. Direct GC injection of lactic acid standards and periodic acid generated inconsistent and irreproducible peaks, possibly due to incomplete lactic acid oxidation to acetaldehyde. The described method is developed to improve lactic acid analysis by GC by using a heat treated derivatization pre-treatment, such that it becomes independent of the retention time and temperature selection of the GC injector. Samples containing lactic acid were amended by periodic acid and heated in a sealed test tube at 100°C for at least 45 min before injecting it to the GC. Reproducible and consistent peaks of acetaldehyde were obtained. Simultaneous determination of lactic acid, acetone, ethanol, butanol, volatile fatty acids could also be accomplished by applying this GC method, enabling precise and convenient organic acid analysis of biological samples such as anaerobic digestion and fermentation processes

Cyanocobalamin enables activated sludge bacteria to dechlorinate hexachloro-1,3-butadiene to nonchlorinated gases

The ability of activated sludge obtained from a local wastewater treatment plant to dechlorinate hexachloro-1,3-butadiene (HCBD) in the presence of either acetate or lactate and cyanocobalamin was investigated. Results from headspace analysis indicated complete dechlorination of HCBD by the accumulation of fully dechlorinated C4 gases (1-buten-3-yne, 1,3-butadiene, and 1,3-butadiyne). Dechlorination products were not observed in the control cultures without cyanocobalamin. Examination of control cultures revealed that the disappearance of HCBD from the headspace was partly due to adsorption into the biomass. However, the key for dechlorination was the shuttle (cyanocobalamin) rather than specific microbial enzymatic activity. The hypothesis that the bacteria reduced cyanocobalamin, which in turn reductively dechlorinated HCBD, was supported by the finding that cyanocobalamin reduced by zero-valent zinc resulted in complete dechlorination. The significance of the findings is that, in contrast to prior work where specific anaerobic bacteria (enrichments or pure cultures) were believed to be necessary for dechlorination resulting in only partly dechlorinated products, the currect data show that nonspecific aerobic activated sludge bacteria can be employed for complete HCBD dechlorination at rates sufficiently high to be considered for bioremediation projects