Distribution of Archaeal Communities along the Coast of the Gulf of Finland and Their Response to Oil Contamination.

The Baltic Sea is vulnerable to environmental changes. With the increasing shipping activities, the risk of oil spills remains high. Archaea are widely distributed in many environments. However, the distribution and the response of archaeal communities to oil contamination have rarely been investigated in brackish habitats. Hence, we conducted a survey to investigate the distribution, diversity, composition, and species interactions of indigenous archaeal communities at oil-contaminated sites along the coast of the Gulf of Finland (GoF) using high-throughput sequencing. Surface water and littoral sediment samples were collected at presumably oil-contaminated (oil distribution facilities) and clean sites along the coastline of the GoF in the winter 2015 and the summer 2016. Another three samples of open sea surface water were taken as offshore references. Of Archaea, Euryarchaeota dominated in the surface water and the littoral sediment of the coast of the GoF, followed by Crenarchaeota (including Thaumarchaeota, Thermoprotei, and Korarchaeota based on the Greengenes database used). The unclassified sequences accounted for 5.62% of the total archaeal sequences. Our study revealed a strong dependence of the archaeal community composition on environmental variables (e.g., salinity, pH, oil concentration, TOM, electrical conductivity, and total DNA concentration) in both littoral sediment and coastal water in the GoF. The composition of archaeal communities was season and ecosystem dependent. Archaea was highly diverse in the three ecosystems (littoral sediment, coastal water, and open sea water). Littoral sediment harbored the highest diversity of archaea. Oil was often detected in the littoral sediment but rarely detected in water at those presumably contaminated sites. Although the composition of archaeal community in the littoral sediment was sensitive to low-input oil contamination, the unchanged putative functional profiles and increased interconnectivity of the archaeal core species network plausibly revealed resilience and the potential for oil degradation. Halobacteriaceae and putative cytochrome P450 pathways were significantly enriched in the oil-contaminated littoral sediment. The archaeal taxa formed highly interconnected and interactive networks, in which Halobacteriaceae, Thermococcus, and methanogens were the main components, implying a potential relevant trophic connection between hydrocarbon degradation, methanogenesis, sulfate reduction, and/or fermentative growth.

Contribution of increased mutagenesis to the evolution of pollutants-degrading indigenous bacteria.

Bacteria can rapidly evolve mechanisms allowing them to use toxic environmental pollutants as a carbon source. In the current study we examined whether the survival and evolution of indigenous bacteria with the capacity to degrade organic pollutants could be connected with increased mutation frequency. The presence of constitutive and transient mutators was monitored among 53 pollutants-degrading indigenous bacterial strains. Only two strains expressed a moderate mutator phenotype and six were hypomutators, which implies that constitutively increased mutability has not been prevalent in the evolution of pollutants degrading bacteria. At the same time, a large proportion of the studied indigenous strains exhibited UV-irradiation-induced mutagenesis, indicating that these strains possess error-prone DNA polymerases which could elevate mutation frequency transiently under the conditions of DNA damage. A closer inspection of two Pseudomonas fluorescens strains PC20 and PC24 revealed that they harbour genes for ImuC (DnaE2) and more than one copy of genes for Pol V. Our results also revealed that availability of other nutrients in addition to aromatic pollutants in the growth environment of bacteria affects mutagenic effects of aromatic compounds. These results also implied that mutagenicity might be affected by a factor of how long bacteria have evolved to use a particular pollutant as a carbon source.

Strategy of Pseudomonas pseudoalcaligenes C70 for effective degradation of phenol and salicylate.

Phenol- and naphthalene-degrading indigenous Pseudomonas pseudoalcaligenes strain C70 has great potential for the bioremediation of polluted areas. It harbours two chromosomally located catechol meta pathways, one of which is structurally and phylogenetically very similar to the Pseudomonas sp. CF600 dmp operon and the other to the P. stutzeri AN10 nah lower operon. The key enzymes of the catechol meta pathway, catechol 2,3-dioxygenase (C23O) from strain C70, PheB and NahH, have an amino acid identity of 85%. The metabolic and regulatory phenotypes of the wild-type and the mutant strain C70ΔpheB lacking pheB were evaluated. qRT-PCR data showed that in C70, the expression of pheB- and nahH-encoded C23O was induced by phenol and salicylate, respectively. We demonstrate that strain C70 is more effective in the degradation of phenol and salicylate, especially at higher substrate concentrations, when these compounds are present as a mixture; i.e., when both pathways are expressed. Moreover, NahH is able to substitute for the deleted PheB in phenol degradation when salicylate is also present in the growth medium. The appearance of a yellow intermediate 2-hydroxymuconic semialdehyde was followed by the accumulation of catechol in salicylate-containing growth medium, and lower expression levels and specific activities of the C23O of the sal operon were detected. However, the excretion of the toxic intermediate catechol to the growth medium was avoided when the growth medium was supplemented with phenol, seemingly due to the contribution of the second meta pathway encoded by the phe genes.

Functional redundancy in phenol and toluene degradation in Pseudomonas stutzeri strains isolated from the Baltic Sea.

In the present study we describe functional redundancy of bacterial multicomponent monooxygenases (toluene monooxygenase (TMO) and toluene/xylene monooxygenase (XylAM) of TOL pathway) and cooperative genetic regulation at the expression of the respective catabolic operons by touR and xylR encoded regulatory circuits in five phenol- and toluene-degrading Pseudomonas stutzeri strains. In these strains both toluene degradation pathways (TMO and Xyl) are active and induced by toluene and phenol. The whole genome sequence of the representative strain 2A20 revealed the presence of complete TMO- and Xyl-upper pathway operons together with two sets of lower catechol meta pathway operons, as well as phenol-degrading operon in a single 292,430bp contig. The much lower GC content and analysis of the predicted ORFs refer to the plasmid origin of the approximately 130kb region of this contig, containing the xyl, phe and tou genes. The deduced amino acid sequences of the TMO, XylA and the large subunit of phenol monooxygenase (LmPH) show 98-100% identity with the respective gene products of the strain Pseudomonas sp. OX1. In both strains 2A20 and OX1 the meta-cleavage pathways for catechol degradation are coded by two redundant operons (phe and xyl). We show that in the strain 2A20 TouR and XylR are activated by different effector molecules, phenol and toluene, respectively, and they both control transcription of the xyl upper, tou (TMO) and phe catabolic operons. Although the growth parameters of redundant strains did not show advantage at toluene biodegradation, the functional redundancy could provide better flexibility to the bacteria in environmental conditions.