Successive transitory distribution of Thaumarchaeota and partitioned distribution of Bathyarchaeota from the Pearl River estuary to the northern South China Sea.

Thaumarchaeota and Bathyarchaeota (formerly named Miscellaneous Crenarchaeotal Group, MCG) are globally occurring archaea playing potential roles in nitrogen and carbon cycling, especially in marine benthic biogeochemical cycle. Information on their distributional and compositional patterns could provide critical clues to further delineate their physiological and biochemical characteristics. Profiles of thaumarchaeotal and the total archaeal community in the northern South China Sea surface sediments revealed a successively transitional pattern of Thaumarchaeota composition using MiSeq sequencing. Shallow-sea sediment enriched phylotypes decreased gradually along the slope from estuarine and coastal marine region to the deep-sea, while deep-sea sediment enriched phylotypes showed a trend of increasing. Proportion of Thaumarchaeota within the total archaea increased with seawater depth. Phylotypes enriched in shallow- and deep-sea sediments were affiliated to OTUs originated from similar niches, suggesting that physiological adaption not geographical distance shaped the distribution of Thaumarchaeota lineages. Quantitative PCR also depicted a successive decrease of thaumarchaeotal 16S rRNA gene abundance from the highest at shallow-sea sites E708S and E709S (2.57 × 106 and 2.73 × 106 gene copies/g of dry sediment) to the lowest at deep-sea sites E525S and E407S (1.97 × 106 and 2.14 × 106 gene copies/g of dry sediment). Both of the abundance fractions of Bathyarchaeota subgroups (including subgroups 1, 6, 8, 10, 13, 15, 17, and ungrouped Bathyarchaeota) and the total Bathyarchaeota in the total archaea showed a negative distribution to seawater depth. Partitioned distribution of Bathyarchaeota fraction in the total archaea is documented for the first time in this study, and the shallow- and deep-sea Bathyarchaeota could account for 17.8 and 0.8%, respectively, on average. Subgroups 6 and 8, enriched subgroups in shallow-sea sediments, largely explained this partitioned distribution pattern according to seawater depth. Their prevalence in shallow-sea and suboxic estuarine sediments rather than deep-sea sediments hints that their metabolic properties of carbon metabolism are adapted to carbon substrates in these environments.

Distribution of tetracycline resistance genes and AmpC ß-lactamase genes in representative non-urban sewage plants and correlations with treatment processes and heavy metals.

The mixed development of livestock breeding and industry in non-urban zones is a very general phenomenon in China. Distribution of antibiotic resistance genes (ARGs) in non-urban sewage treatment systems has not been paid enough attentions. In this study, eleven tetracycline resistance genes (tetA, tetB, tetC, tetE, tetG, tetL, tetM, tetO, tetQ, tetS and tetX), four AmpC ß-lactamase genes (EBC, MOX, FOX and CIT) and four heavy metals (Cu, Zn, Cd and Pb) were detected and analyzed in four non-urban sewage plants with different sewage sources and different treatment processes in Guangzhou. The results showed that tetA and tetC were the most prevalent tetracycline resistance genes with the same detection frequency of 85% and EBC was the most prevalent AmpC ß-lactamase gene with a detection frequency of 75%. The relative abundance of tetracycline resistance genes was approximately 1.6 orders of magnitudes higher than that of AmpC ß-lactamase genes in all samples. A/O was the most effective process for the non-urban sewage plant receiving industrial or agricultural wastewater. Sedimentation was the most key process to eliminate ARGs from liquid phase. Most ARGs were carried in excess sludge rather than effluent. Significant correlation was found between the tet gene and Zn (r = 0.881, p < 0.01), followed by the AmpC gene and Cu (r = 0.847, p < 0.01), the tet gene and Cu (r = 0.714, p < 0.05). Therefore, the pollution of ARGs in the sewage treatment systems of non-urban zones co-polluted by heavy metals should be paid more attentions.

Antibiotic resistance genes (ARGs) in duck and fish production ponds with integrated or non-integrated mode.

Antibiotic resistance genes (ARGs) are emerging micropollutants with environmental persistence. Aquaculture environments are considered as potential reservoirs for ARGs pollution and horizontal gene transfer (HGT). This study analyzed water and sediment from eight culture ponds (integrated culture: duck-fish pond; monoculture: duck pond and fish pond) and a control pond (without any aquaculture activity) in Zhongshan, South China. Seventeen types of ARGs were detected in all ponds, which conferring resistance to four classes of antibiotics including tetracycline (tetA, tetB, tetC, tetE, tetG, tetL, tetA-P, tetM, tetO, tetS, tetW and tetX), AmpC beta-lactamase products (EBC and FOX), sulfonamide (sul1 and sul2) and erythromycin (ermA), with class 1 integron (intI1) as motility gene. The total concentrations of detected ARGs in culture pond water were much higher than control (about 1.6-4.0 times). Integrated culture showed lowest absolute abundance of ∑ARGs in water (3.686 × 107 copies mL-1) and the highest in sediment (4.574 × 108 copies g-1). Monoculture ponds showed higher relative abundance of ∑ARGs both in water (fish pond: 0.5149) and sediment (duck pond: 0.4919). As the main contributor to the ARGs abundance and significant correlations with ∑tet, ∑ARGs and intI1 (P < 0.01), tetA was suggested to be a potential indicator for the abundance of tetracycline resistance genes in these classes of aquaculture modes in the Pearl River Delta. This study provides a case for the ARGs abundance in aquaculture and as a reference for the upcoming health risk assessment in aquatic environment.

Occurrence of antibiotic resistance genes in landfill leachate treatment plant and its effluent-receiving soil and surface water.

The antibiotic resistance genes (ARGs) from urban waste may spread to the environment with the discharge of leachate. Fifteen types of ARGs, including tetracycline, sulfonamides, AmpC ß-lactamase and the class 1 integron gene were detected in the samples from the largest leachate treatment plant (LTP) in Guangzhou and its effluent receiving bodies (soil and surface water). The results showed that ARGs in leachates were in high levels and varied with seasons. The abundance of ARGs in the influent from high to low was in the turn of summer, winter, spring. About 2 to 4 orders of magnitude of ARGs were eliminated by the whole leachate treatment process. The predominant ARGs in the receiving soil were intI1, tetB, sul2, tetA and tetX, while those in the receiving surface water were sul2, intI1 and sul1, and the concentrations of ARGs in the receiving bodies were higher than those in the other natural bodies by 1 to 2 orders of magnitude. In addition, the results of bivariate correlation analysis showed that the abundances of ARGs (tetC, tetW, sul1, sul2, intI1 and FOX) were in significant correlation with the concentrations of heavy metals (Cu, Zn, Ni and Cr) (p < 0.05). LTPs are more likely to be sources of ARGs than wastewater treatment plant (WWTP) and need to be focused on.

Accurate Evaluation of Cone-Beam Computed Tomography to Volumetrically Assess Bone Grafting in Alveolar Cleft Patients.

OBJECTIVE: The aim of this study was to investigate the accuracy of cone-beam computed tomography (CBCT) to assess the volume of bone graft in alveolar cleft patients. MATERIALS AND METHODS: Twelve patients of unilateral alveolar cleft were included in this study. All patients were taken CBCT preoperative and 1 week postoperative. The digital imaging and communications in medicine (DICOM) files were imported into Simplant software and three-dimensional (3D) reconstruction of the alveolar defect was achieved. With 3D volumetric measurements module, the volume of alveolar cleft was calculated preoperatively. During operation, the syringe compression method was adopted to calculate the actual amount of bone graft. One week postoperative, CBCT scan was performed again to measure the bone volume grafted to the defect. The volumetric ratio of the syringe compression method to preoperative CBCT assessment and the volume difference between syringe compression method and postoperative CBCT assessment were analyzed to evaluate the accuracy of CBCT measurement. RESULTS: The 3D structure of the alveolar cleft and the boundary of bone graft was clear from CBCT images. The estimated volume of alveolar cleft by preoperative CBCT scans was 1.06 ±â€Š0.09 cm, and the actual amount of bone graft determined by the syringe compression method was 1.51 ±â€Š0.12 cm. The ratio between the latter to the former was 1.43 ±â€Š0.07. The calculated volume of bone graft by 1-week postoperative CBCT scans was 1.53 ±â€Š0.11 cm, with no significant difference compared with the actual amount of bone graft (P > 0.05). CONCLUSIONS: CBCT was an accurate measurement to calculate the volume of alveolar defect and bone graft in alveolar cleft patients. Preoperative scans could aid in quantitatively determining the bone amount needed to adequately fill the bone defect, and the postoperative scans give accurate follow-up evaluation after surgery.

Antioxidative responses of Pseudomonas fluorescens YZ2 to simultaneous exposure of Zn and Cefradine.

Binary pollution of both heavy metals and antibiotics has received increasing attentions for their joint effects of eco-toxicity and health hazards. To reveal the effects of mixtures of different pollutants on bacterial antioxidant response system, Pseudomonas fluorescens ZY2, a new strain isolated from swine wastewater, was chosen to determinate growth (bacterial density OD600), reactive oxygen species (ROS) concentration, protein concentration and superoxide dismutase (SOD) activity under exposure treatments of Zn, Cefradine or Zn + Cefradine. Bacterial densities of all the treatment groups increased significantly over the incubation time, but those containing pollutant addition were slightly lower than the control at different times of incubation. Both ROS concentration and SOD activity increased first and then decreased (p < 0.01) over time, which was opposite to the protein concentrations (p < 0.01), showing a much significant increase by Cefradine alone. With Zn concentration increasing from 40 to 160 mg/L, the intracellular SOD activity increased as a response to the improvement of ROS (p < 0.05), while the balance between ROS and SOD was broken down due to the disproportionate change of total SOD activity and ROS concentration, the bacterial densities therefore decreased for the weak resistance. With the combined treatment of Zn (200 mg/L) and Cefradine (1 mg/L), though the toxicity of Zn caused a much significant increase of ROS, the bacterial resistance was further improved showing a more significant increase of total SOD activity and the bacterial densities therefore increased bacterial growth. Zn concentration also affected the protein synthesis. Either single or binary stress induced the bacterial resistance by regulating SOD activity to eliminate ROS. All results of the bacterial oxidant stress, SOD response and protein synthesis in the combined treatment groups were more complicated than those in single treatment groups, which depended on the properties of the single treatment as well as the interaction between the two treatments upon bacterial activity. For P. fluorescens ZY2, the mediation of SOD activity to eliminate ROS in response to the combined exposure to Zn and Cefradine was first revealed as one of the co-resistance mechanisms, which is informative to further understanding the risk of antibiotics resistant bacteria to human and environmental health more accurately.

Endogenous nitric oxide in Pseudomonas fluorescens ZY2 as mediator against the combined exposure to zinc and cefradine.

A better understanding on the mechanism involved in bacterial resistance to combined exposure to antibiotics and heavy metals is helpful in implementing practices to mitigate their ecological risk and spread of resistance genes in microbial population. Pseudomonas fluorescens ZY2, a strain isolated from swine wastewater, was chosen to study its growth (bacterial density OD600), the formation of reactive oxygen species (ROS), nitric oxide (NO) and NO synthases (NOS) under Zn, cefradine or Zn + cefradine treatments. Using Zn and cefradine as representative heavy metal and antibiotic in this investigation, respectively, the resistance of P. fluorescens ZY2 to toxic chemical exposure was investigated. Bacterial densities of treatment groups significantly increased over the time of incubation, but less than the control. ROS, NO and NOS initially increased, but then decreased after the initial 8 h of culturing, and were positively related to Zn concentrations. Moreover, the formation of ROS, NOS, and NO was activated by cefradine at Zn of up to 160 mg/L, but inhibited at Zn of 200 mg/L whether cefradine was added or not. Zn concentration affected ROS and NO concentrations between treatments and also was closely related to the variation of the relative bacterial density. For P. fluorescens ZY2, the mediation of endogenous NO to overcome ROS in response to the combined exposure of Zn and cefradine was suggested as a co-resistance mechanism, which would be beneficial to evaluate the ecological risk of heavy metals and antibiotics.

Combined toxic effects of heavy metals and antibiotics on a Pseudomonas fluorescens strain ZY2 isolated from swine wastewater.

A Pseudomonas fluorescens strain ZY2, isolated from swine wastewater, was used to investigate the synergistic effects of five heavy metals (Pb, Cu, Zn, Cr(VI) and Hg) on bacterial resistance to antibiotics. Results indicate that the combined effects of antibiotic type, heavy metal type and concentration were significant (p < 0.01). Cross-resistance to Hg and antibiotics was the most noticeable. Moreover, the resistance to Hg and cefradine or amoxicillin, and Cr and amoxicillin were synergistic for low heavy metal concentrations, and turned antagonistic with increasing concentrations, while the resistances to Cr or Cu and cefradine, Pb or Cu and amoxicillin, Cu and norfloxacin showed reverse effects. In addition, resistance to Zn and amoxicillin were always synergetic, while resistance to Pb and cefradine or norfloxacin, Cr or Hg and norfloxacin as well as all the heavy metals and tetracycline were antagonistic. These results indicate that bacterial resistance to antibiotics can be affected by the type and concentration of co-exposed heavy metals and may further threaten people's health and ecological security severely via horizontal gene transfer.

[Analysis of hydrolytic enzyme activities on sludge aerobic/anoxic digestion after ultrasonic pretreatment].

In order to evaluate the function of sludge aerobic/anoxic digestibility by ultrasonic pretreatment. The SS, VSS and hydrolytic enzyme activities (amylase, glucosidase, protease, phosphatase) were measured before and after ultrasonic pretreatment (28 kHz, 0.15 kW x L(-1), 10 min). The results showed that the performances of aerobic/anoxic were greatly improved after ultrasonic pretreatment, the removal efficiency of VSS went to 44.3%, 7.8% better than of traditional aerobic/anoxic digestion. The variational trend of sludge hydrolytic enzyme activities increased firstly and then fell off during 13d digestion, the maximum of amylase activity and glucosidase activity in ultrasonic sludge, appeared in the 5 d, amylase activity was 0.104 micromol x g(-1) and glucosidase activity was 0.637 (micromol x g(-1). The maximum of intracellular protease activity and extracellular proteases activity in ultrasonic sludge, appeared in the 7 d, intracellular protease activity was 23.68 micromol x g(-1), higher than extracellular proteases activity, and it was playing a leading role in sludge digestion. The acid phosphatase activity of ultrasonic sludge was higher than the control sludge, and the alkaline phosphatase was sensitive to environment. So the alkaline phosphatase activity reduced when the internal properties of sludge was changed.

[Analysis of community structure on sludge aerobic/anoxic digestion after ultrasonic pretreatment].

The microbial community structure of sludge aerobic/anoxic digestion after ultrasonic pretreatment was studied by PCR amplification and DGGE based on 16S rDNA. The genomic DNA of sludge at different stages was extracted with SDS cell lysate method. After purification of DNA, the 16S rDNA genes (V3 region) were amplified by using the universal primers (F357GC and R518). The results of agarose gel (1.5%) electrophoresis showed that the PCR products were about 190 bp in length. The amplified DNA fragments were separated by paralleled DGGE with the denaturant (urea and acrylamide) from 30% to 60%. The sequences were used for homology analysis and phylogenetic trees were constructed. The DGGE profiles showed that the change of microbial diversity was in correspondence to different periods. Compared with 0 d, the diversities of microorganisms were 61.2%, 48.2%, 46.4%, 42.6% and 41.7%, respectively after 5 d, 10 d, 15 d, 20 d, 25 d. Shannon density index of bacteria experienced a process from a gradual reduction to stable state. This suggested that ultrasonic pretreatment had a significant impact on bacterial community structures. Cluster analysis of DGGE by UPGMA (unweighted air group method, arithmetic mean) divided all lanes into three clusters, which corresponded to different periods during the whole experiment. The sequences indicated that Firmicute, Genuscitrobacter, Bacilli, alpha-Proteobacteria, beta-Proteobacteria were the predominant microbial populations in the process of sludge aerobic/anoxic digestion after ultrasonic pretreatment.

Study on anaerobic treatment of wastewater containing hexavalent chromium.

A self-made anaerobic bio-filter bed which was inoculated with special sludge showed high efficiency in removing hexavalent chromium. When pump flow was 47 ml/min and COD(Cr) of wastewater was about 140 mg/L, it took 4 h to decrease the Cr6+ concentrations from about 60 mg/L to under 0.5 mg/L, compared with 14 h without carbon source addition. Cr6+ concentrations ranged from 64.66 mg/L to 75.53 mg/L, the system efficiency was excellent. When Cr6+ concentration reached 95.47 mg/L, the treatment time was prolonged to 7.5 h. Compared with the contrast system, the system with trace metals showed clear superiority in that the Cr6+ removal rate increased by 21.26%. Some analyses also showed that hexavalent chromium could probably be bio-reduced to trivalent chromium, and that as a result, the chrome hydroxide sediment was formed on the surface of microorganisms.