Halophilic Martelella sp. AD-3 enhanced phenanthrene degradation in a bioaugmented activated sludge system through syntrophic interaction.

Polycyclic aromatic hydrocarbons (PAHs) are a group of common recalcitrant pollutant in industrial saline wastewater that raised significant concerns, whereas traditional activated sludge (AS) has limited tolerance to high salinity and PAHs toxicity, restricting its capacity to degrade PAHs. It is therefore urgent to develop a bioaugmented sludge (BS) system to aid in the effective degradation of these types of compounds under saline condition. In this study, a novel bioaugmentation strategy was developed by using halophilic Martelella sp. AD-3 for effectively augmented phenanthrene (PHE) degradation under 3% salinity. It was found that a 0.5∼1.5% (w/w) ratio of strain AD-3 to activated sludge was optimal for achieving high PHE degradation activity of the BS system with degradation rates reaching 2.2 mg⋅gVSS-1⋅h-1, nearly 25 times that of the AS system. Although 1-hydroxy-2-naphthoic acid (1H2N) was accumulated obviously, the mineralization of PHE was more complete in the BS system. Reads-based metagenomic coupled metatranscriptomic analysis revealed that the expression values of ndoB, encoding a dioxygenase associated with PHE ring-cleavage, was 5600-fold higher in the BS system than in the AS system. Metagenome assembly showed the members of the Corynebacterium and Alcaligenes genera were abundant in the strain AD-3 bioaugmented BS system with expression of 10.3±1.8% and 1.9±0.26%, respectively. Moreover, phdI and nahG accused for metabolism of 1H2N have been annotated in both above two genera. Degradation assays of intermediates of PHE confirmed that the activated sludge actually possessed considerable degradation capacity for downstream intermediates of PHE including 1H2N. The degradation capacity ratio of 1H2N to PHE was 87% in BS system, while it was 26% in strain AD-3. These results indicated that strain AD-3 contributed mainly in transforming PHE to 1H2N in BS system, while species in activated sludge utilized 1H2N as substrate to grow, thus establishing a syntrophic interaction with strain AD-3 and achieving the complete mineralization of PHE. Long-term continuous experiment confirmed a stable PHE removal efficiency of 93% and few 1H2N accumulation in BS SBR system. This study demonstrated an effective bioaugmented strategy for the bioremediation of saline wastewater containing PAHs.

Effect of rhamnolipids on degradation of anthracene by two newly isolated strains, Sphingomonas sp. 12A and Pseudomonas sp. 12B.

Anthracene is a PAH that is not readily degraded, plus its degradation mechanism is still not clear. Thus, two strains of bacteria-degrading bacteria were isolated from longterm petroleum-polluted soil and identified as Sphingomonas sp. 12A and Pseudomonas sp. 12B by a 16S rRNA sequence analysis. To further enhance the anthracene-degrading ability of the two strains, the biosurfactants produced by Pseudomonas aeruginosa W3 were used, which were characterized as rhamnolipids. It was found that these rhamnolipids dramatically increased the solubility of anthracene, and a reverse-phase HPLC assay showed that the anthracene degradation percentage after 18 days with Pseudomonas sp. 12B was significantly enhanced from 34% to 52%. Interestingly, their effect on the degradation by Sphingomonas sp. 12A was much less, from 35% to 39%. Further study revealed that Sphingomonas sp. 12A also degraded the rhamnolipids, which may have hampered the effect of the rhamnolipids on the anthracene degradation.

[Distribution Characteristics of Sulfonamide Antibiotic Resistance Genes in a Drinking Water Source in East China].

Antibiotic resistance genes (ARGs) in drinking water sources have attracted widespread attention due to the threat they pose to water security and human health. This study mainly focused on the distribution of sulfonamide ARGs (sul 1, sul 2) and one integrase gene (intI 1) in water and sediment using qualitative and fluorescent quantitative PCR, based on previous work on the characteristics of 13 kinds of sulfonamides in a drinking water source in East China. Results showed that the three target genes were all detected in water and sediment. The sul 1 gene was the sulfonamide ARG with highest concentration, with 1.5×104-6.4×105 copies·mL-1 in source water and maximum concentration of 1.6×108 copies·g-1 in sediment. Concentration of sul 1 was 0.6-2.2, 0.5-1.9 order of magnitudes higher than sul 2 and intI 1 genes, respectively. There was no significant difference between the absolute concentrations of sul 1, sul 2, and intI 1 in inflow and outflow. However, in the case of sediment, absolute abundances of sul 1, sul 2, and intI 1 in outflow were higher than those in inflow. The maximum concentration of sul 1 was detected in outflow in summer (6.4×105 copies·mL-1). The concentration of intI 1 was higher in winter compared to other seasons. There was a positive correlation between sul 1 and 13 sulfonamides (r=0.69, P<0.05), and the relative concentration of sul 1 and amount of sulfamethoxazole were significantly positively related (r=0.79, P<0.01). There were also positive correlations between the relative concentrations of intI 1 and sul 1, sul 2 (r:0.80 and 0.73, P<0.05), respectively, suggesting that intI 1 played an important role in horizontal gene transfer of sulfonamide ARGs in this drinking water source. This study provides basic data for monitoring pollution of ARGs, as well as a basis for controlling ARG pollution in the drinking water environment and making management decisions.

[Distribution Characteristics and Health Risk Assessment of Thirteen Sulfonamides Antibiotics in a Drinking Water Source in East China].

Trace levels of residual antibiotics in drinking water sources may threaten public health and becomes a serious issue in modern society. Occurrence and distribution of 13 sulfonamides antibiotics in a drinking water source in East China were investigated using solid phase extraction-high performance liquid chromatography-tandem mass spectrometry. The results revealed that all 13 sulfonamides were detected with the total concentrations ranging from 10.5 ng·L-1 to 238.5 ng·L-1. Sulfamethoxazole (SMX) and sulfonamide (SAM) presented the 100% detected frequency, and the maximum concentrations reached 107.0 ng·L-1 and 43.1 ng·L-1, respectively. Sulfonamides contamination levels in the inlet were relatively higher than those in the outlet. The concentration of SMX was substantially unchanged in different positions of drinking water source. Seasonal variation of sulfonamide concentration showed that the concentrations were higher in winter and spring compared with those in summer and autumn. The range of total sulfonamides concentration detected in winter and spring was from 110.8 ng·L-1 to 117.9 ng·L-1, which was 3.6-3.8 times higher than those in summer and autumn. Human health risk characterization of residual sulfonamides in drinking water source was based on the assessment of risk quotients (RQs) for which different life stages were taken into account. The results showed that no sulfonamides had a RQs higher than 1. Trimethoprim (TMP) was the most risky antibiotic to the 0-3 months old infants with RQs reaching 0.001. Although the sulfonamides detected in the drinking water source do not pose any risk to the peoples' health, the implication of long-time exposure and comprehensive risk still deserve attention.

[Isolation, charcaterization of an anthracene degrading bacterium Martelella sp. AD-3 and cloning of dioxygenase gene].

Anthracene, among the 16 US EPA polycyclic aromatic hydrocarbons (PAHs), is a typical low molecular weight environmental contaminant, which gains concern on its biodegradation under hypersaline condition. In this study, an anthracene-degrading bacterial strain was isolated from highly saline petroleum-contaminated soil. Based on its physiological, biochemical characteristics and 16S rDNA sequence analysis, the bacteria was preliminary identified and named as Martelella sp. AD-3. The strain was able to utilize anthracene as sole carbon source for growth and the degradation occurred under broad salinities (0.1% to 10%) and varying pHs (6.0 to 10.0). The optimized degradation conditions were initial concentration 25 mg x L(-1), culture temperature 30 degrees C, pH 9.0 and salinity 3%. And 94.6% of anthracene was degraded by strain AD-3 under the optimal conditions within 6 days. Degenerate primers design was performed with a reported dioxygenase alpha subunit homologous gene. A length of 307 bp fragment of the partial dioxygenase gene sequences (GenBank accession: JF823991.1) was amplified by nested PCR. The clones amino acid sequence from strain AD-3 showed 95% identity to that of the partial naphthalene dioxygenase large-subunit from Marinobacter sp. NCE312 (AF295033). The results lay a foundation for the further study of molecular mechanism involved in the PAHs biodegradation by strain AD-3.

Characterization of the Haloarcula hispanica amyH gene promoter, an archaeal promoter that confers promoter activity in Escherichia coli.

Archaea form a third domain of life that is distinct from Bacteria and Eukarya. According to the current knowledge, the basal transcription machinery of Archaea (including the core promoter architecture, the RNA polymerase, and the basal transcription factors) closely resembles that of Eukarya in structure and function, while differing considerably from the bacterial paradigm. In the present study, the promoter region of the halophilic archaeon Haloarcula hispanica's amyH gene was isolated and characterized, and it was surprisingly revealed that the amyH gene promoter could confer promoter activity (i.e., drive transcription) in haloarchaea (Archaea) as well as in Escherichia coli (Bacteria), where the transcriptions driven are initiated at the same adenine base. Further investigation revealed that the core structure of the amyH gene promoter possesses a combination of the typical structural characteristics of archaeal promoter, which are eukaryotic-like, and those of bacterial promoter. Our results indicate that the core promoter structures of some archaeal genes may possess a combination of eukaryotic- and bacterial-like features, and moreover, suggest a possible evolutionary relationship between basal transcription signals and transcription mechanisms of Archaea and the other two domains of life.