Single Escherichia coli bacteria detection using a chemiluminescence digital microwell array chip.

Rapid and sensitive Escherichia coli (E. coli) detection is important in determining environmental contamination, food contamination, as well as bacterial infection. Conventional methods based on bacterial culture suffer from long testing time (24 h), whereas novel nucleic acid-based and immunolabelling approaches are hindered by complicated operation, the need of complex and costly equipment, and the lack of differentiation of live and dead bacteria. Herein, we propose a chemiluminescence digital microwell array chip based on the hydrolysis of 6-Chloro-4-methylumbelliferyl-ß-D-glucuronide by the ß-D-glucuronidase in E. coli to achieve fast single bacterial fluorescence detection. Taking the advantage of the picoliter microwells, single bacteria are digitally encapsulated in these microwells, thus the accurate quantification of E. coli can be realized by counting the number of positive microwells. We also show that the chemiluminescence digital microwell array chip is not affected by the turbidity of the test samples as well as the temperature. Most importantly, our method can differentiate live and dead bacteria through bacterial proliferation and enzyme expression, which is confirmed by detecting E. coli after pH and chlorination treatment. By comparing with the standard method of plate counting, our method has comparable performance but significantly reduces the testing time from over 24 h-2 h and 4 h for qualitative and quantitative analysis, respectively. In addition, the microfluidic chip is portable and easy to operate without external pump, which is promising as a rapid and on-site platform for single E. coli analysis in water and food monitoring, as well as infection diagnosis.

Trapping and Detection of Single Viruses in an Optofluidic Chip.

Accurate single virus detection is critical for disease diagnosis and early prevention, especially in view of current pandemics. Numerous detection methods have been proposed with the single virus sensitivity, including the optical approaches and immunoassays. However, few of them hitherto have the capability of both trapping and detection of single viruses in the microchannel. Here, we report an optofluidic potential well array to trap nanoparticles stably in the flow stream. The nanoparticle is bound with single viruses and fluorescence quantum dots through an immunolabeling protocol. Single viruses can be swiftly captured in the microchannel by optical forces and imaged by a camera. The number of viruses in solution and on each particle can be quantified via image processing. Our method can trap and detect single viruses in the 1 mL serum or water in 2 h, paving an avenue for the advanced, fast, and accurate clinical diagnosis, as well as the study of virus infectivity, mutation, drug inhibition, etc.

Appling hydrolysis acidification-anoxic-oxic process in the treatment of petrochemical wastewater: From bench scale reactor to full scale wastewater treatment plant.

A hydrolysis acidification (HA)-anoxic-oxic (A/O) process was adopted to treat a petrochemical wastewater. The operation optimization was carried out firstly by a bench scale experimental reactor. Then a full scale petrochemical wastewater treatment plant (PCWWTP, 6500 m(3) h(-1)) was operated with the same parameters. The results showed that the BOD5/COD of the wastewater increased from 0.30 to 0.43 by HA. The effluent COD was 54.4 mg L(-1) for bench scale reactor and 60.9 mg L(-1) for PCWWTP when the influent COD was about 480 mg L(-1) on optimized conditions. The organics measured by gas chromatography-mass spectrometry (GC-MS) reduced obviously and the total concentration of the 5 organics (1,3-dioxolane, 2-pentanone, ethylbenzene, 2-chloromethyl-1,3-dioxolane and indene) detected in the effluent was only 0.24 mg L(-1). There was no obvious toxicity of the effluent. However, low acute toxicity of the effluent could be detected by the luminescent bacteria assay, indicating the advanced treatment is needed. The clone library profiling analysis showed that the dominant bacteria in the system were Acidobacteria, Proteobacteria and Bacteriodetes. HA-A/O process is suitable for the petrochemical wastewater treatment.

[Difference of Species Sensitivities for Aquatic Life Criteria in China and the USA].

Recently, China has commenced water quality criteria (WQC) research using the species sensitivity distribution (SSD) method. However, it is difficult to obtain sufficient toxicity data on native species for thousands of contaminants. In this study, the feasibility of deriving aquatic life criteria using toxicity data from non-native species was analyzed (taking the USA as an example). Firstly, 10 pollutants [As(Ⅲ), Cr(Ⅵ), Hg, Cu, Pb, Zn, nitrobenzene, parathion, chlorpyrifos, tributyltin] with plenty of toxicity data (at least 3 Phyla and 8 Families) both in China and the USA were selected based on the recommended aquatic life criteria of 58 pollutants by the US EPA. Secondly, the two-sample Kolmogorov-Smirnov (K-S) test and comparison method of important points of HC5 and HC50 (hazardous concentrations for 5% of the species and 50% of the species) were used to determine the difference of species sensitivity between China and the USA. The result of two-sample K-S test showed that the difference of species sensitivity to Cu, chlorpyrifos and tributyltin between the two countries was significant (P<0.05). Additionally, there were significant differences for HC5 values [Cr(Ⅵ), chlorpyrifos] between the two countries, as well as the HC50 (chlorpyrifos). As a result, there were significant differences for the species sensitivity to some pollutants [Cr(Ⅵ), chlorpyrifos, etc.] between China and the USA. Therefore, in this situation, "over-protection" or "non-protection" may happen when using the criteria values or the toxicity data of American species directly to protect the Chinese native species.

[Evaluation of the acute toxicity of pharmaceutical wastewater to luminescent bacteria].

Acute toxicity of wastewater from 5 nodes of technological process in the pharmaceutical factory sewage treatment station was studied by luminescent bacteria tests. The EC50, TUa and LID of the wastewater in underground regulating tanks was 3.44%, 29 and 625, respectively, indicating the water was extremely/highly toxic; for the wastewater in surface regulating tanks, the EC50, TUa and LID was 2.46%, 41 and 244, respectively, also extremely/highly toxic; for the wastewater in middle sediment tanks, the EC50 > 100% and LID was 10, which was moderately toxic; for the wastewater in secondary sediment tanks and the final effluents, the EC50 was above 100% and LID was 1, with no observed toxicity. The results indicated that the existing treatment process effectively reduced the acute toxicity of the pharmaceutical wastewater to luminescent bacteria, the effluents showed no observed toxicity to luminescent bacteria, which was lower than the relative effluent limits of pharmaceutical wastewater. The wastewater in lower concentration did not inhibit the luminosity, but enhanced the luminosity.

Progress of environmental management and risk assessment of industrial chemicals in China.

With China's rapid economic growth, chemical-related environmental issues have become increasingly prominent, and the environmental management of chemicals has garnered increased attention from the government. This review focuses on the current situation and the application of risk assessment in China's environmental management of industrial chemicals. The related challenges and research needs of the country are also discussed. The Chinese government promulgated regulations for the import and export of toxic chemicals in 1994. Regulations for new chemical substances came into force in 2003, and were revised in 2010 based on the concept of risk management. In order to support the implementation of new regulations, Guidance for Risk Assessment of Chemicals is under development in an attempt to provide the concepts and techniques of risk assessment. With increasing concern and financial support from Chinese government, China is embarking on the fast track of research and development in environmental management of industrial chemicals.

[Conjugation and immunogenic evaluation of complete immunogen for the small molecular environmental pollutants 2,4-D].

Conjugation of complete immunogen for 2,4-dichlorophenoxyacetic acid (2,4-D) was studied. 2,4-D was cross-linked to bovine serum albumin (BSA, carrier) by 1-ethyl-3-(3-dimethyl- aminopropyl) carbodiimide hydrochloride (EDC). The conjugation reaction was found to be more effective at 4 degrees C and incubated for 18 hour. 2,4-D was dissolved in 0.05 mol/L phosphate buffer between 10.0 - 12.0 mg/mL concentration, pH was adjusted to be 5.4 - 6.1. When the weight of added EDC was below 12 mg, the more EDC was added the higher substitution degree complete immunogen was synthesized. Complete immunogen of various substitution degree (2,4-D: protein) were applied to immunize balb/c mice. The conjugates of 2,4-D and poly-L-lysine was applied as coating antigen. It was experimentally found that complete immunogen of substitution degree 12 and 18 are more immunogenic than that of substitution degree 6 and 25. None-specific adsorption between antiserum that was produced by complete immunogen of substitution degree 18 and coating antigen was very weak, and the antiserum contained more 2,4-D specific antibody. It could be used as the immunogen for the preparation monoclonal antibody.