Mitigation of biofouling in membrane bioreactors by quorum-quenching bacteria during the treatment of metal-containing wastewater.

Quorum quenching (QQ) is an efficient way to mitigate membrane biofouling in a membrane bioreactor (MBR) during wastewater treatment. A QQ bacterium, Lysinibacillus sp. A4, was isolated and used to mitigate biofouling in an MBR during the treatment of wastewater containing metals. A QQ enzyme (named AilY) was cloned from A4 and identified as a metallo-ß-lactamase-like lactonase. The QQ activity of A4 and that of Escherichia coli BL21 (DE3) overexpressing AilY could be promoted by Fe2+, Mn2+, and Zn2+ while remaining unaffected by other metals tested. The two bacteria effectively mitigated biofouling by reducing the transmembrane pressure from around 30 to 20 kPa without negative influence on the COD, NH4+-N, or total phosphorus of the effluent. The relative abundance of Lysinibacillus sp. A4 increased greatly from 0.04 to 8.29% in the MBR with metal-containing wastewater, suggesting that Lysinibacillus sp. A4 could multiply quickly and adapt to this environment. Taken together, the findings suggested that A4 could tolerate metal to a certain degree, and this property could allow A4 to adapt well to metal-containing wastewater, making it a valuable strain for mitigating biofouling in MBR during the treatment of metal-containing wastewater.

Characterization of an N-acylhomoserine lactonase from Serratia sp. and its biofouling mitigation in a membrane bioreactor.

Membrane biofouling is a process that can impede the development of membrane bioreactor (MBR), which constitutes an important system of the wastewater treatment process. Membrane biofouling is governed by quorum sensing (QS), a communication system heavily dependent on the activities of signal molecules. Certain bacteria, known as quorum quenching (QQ) bacteria, can quench the QS process by destroying the signal molecules. These QQ bacteria are considered a sustainable and feasible way of mitigating membrane biofouling in MBR. In this study, a QQ enzyme (designated as AisZ) from a Serratia sp. was first identified and characterized. Escherichia coli BL21 expressing AisZ was able to degrade different QS signal molecules. Furthermore, these cells could also mitigate membrane biofouling in MBR during a 29-day operation by reducing the transmembrane pressure from 31 to 21 kPa. The metal ions Co2+ and Ni2+ were relatively important to AisZ in that they could significantly enhance the activity of AisZ and restore the EDTA-inactivated AisZ. Expression of the aisA gene was not influenced by Co2+, Ni2+ and QS signal molecules. AisZ might, therefore, extend the diversity of potential candidates for the mitigation of biofouling associated with membrane filtration technologies.

Biotreatment of Cr(VI) and pyrene combined water pollution by loofa-immobilized bacteria.

Hexavalent chromium (Cr(VI)) and pyrene are toxic pollutants that are difficult to remediate from soils and wastewater. Serratia sp. strains have been previously demonstrated to remove either Cr(VI) or pyrene and here a new isolate, called the Z6 strain, was demonstrated to remove both simultaneously. The removal occurs primarily by Cr(VI) reduction and pyrene biodegradation, and genome analysis suggests the removal mechanisms are the putative chromate reductase and two assumable pathways of pyrene degradation. The Z6 strain effectively removed most Cr(VI) (up to approximately 86%) and pyrene (up to approximately 57%) in seven different types of wastewater after 7 days of biotreatment. Additionally, the carrier loofa used for bacteria immobilization did not change the kinetics of Cr(VI) reduction or pyrene degradation. The carrier loofa was also effective for multiple uses, with removal capacity not being significantly affected over the first seven cycles with the same carrier loofa. These results provide data for developing practical biotreatment applications of Cr(VI) and pyrene contaminated sites.

Biotreatment of pyrene and Cr(VI) combined water pollution by mixed bacteria.

Pyrene and chromium (Cr(VI)) are persistent pollutants and cause serious environmental problems because they are toxic to organisms and difficult to remediate. The toxicity of pyrene and Cr(VI) to three crops (cotton, soybean and maize) was confirmed by the significant decrease in root and shoot biomass during growth in pyrene/Cr(VI) contaminated hydroponic solution. Two bacterial strains capable of simultaneous pyrene biodegradation and Cr(VI) reduction were isolated and identified as Serratia sp. and Arthrobacter sp. A mixture of the isolated strains at a ratio of 1:1 was more efficient for biotreatment of pyrene and Cr(VI) than either strain alone; the mixture effectively carried out bioremediation of contaminated water in a hydroponic system mainly through pyrene biodegradation and Cr(VI) reduction. Application of these isolates shows potential for practical microbial remediation of pyrene and Cr(VI) combined water pollution.

Remediation potential of immobilized bacterial consortium with biochar as carrier in pyrene-Cr(VI) co-contaminated soil.

Polycyclic aromatic hydrocarbons (PAHs) and potentially toxic trace elements (PTEs) soil contamination have become areas of concern. Bioaugmentation is regarded as an effective bioremediation method, however it is difficult to simultaneously degrade organic compounds and remove PTEs with individual microbial strains. Therefore, the objective of this study was to evaluate the feasibility of using immobilized microbial consortia, including two PAH-degrading bacterial strains (W1 and W2) and a Cr(VI)-reducing bacterium (Y2), for the remediation of pyrene-Cr(VI) co-contaminated soil. Three immobilization methods were investigated: (1) bacterial consortium adsorption onto biochar (BC), (2) bacterial consortium entrapment in alginate beads (AC), (3) bacterial consortium adsorption on biochar and sequential entrapment in alginate beads (BAC). In addition, a free bacterial consortium (FC) was also used for comparison. Ten treatments were designed to illustrate the bioremediation efficiency of the free and immobilized consortia. The results show that treatments AC and BAC resulted in more efficient Cr(VI) removal compared with BC and FC. Pyrene levels in AC and BAC microcosms were reduced from 42.33 ± 3.82 to 11.56 ± 1.37 and 7.48 ± 0.39 mg kg-1, respectively. Bioavailable Cr (VI) in AC and BAC was significantly lower than that in other microcosms after 28 days' incubation. Both AC and BAC microcosms exhibited a higher level of dehydrogenase and fluorescein diacetate hydrolysis activity. Furthermore, soil microbial diversity was higher in AC and BAC microcosms compared with the others. Thus, the entrapped consortia may be useful for bioremediation of pyrene and Cr (VI) without compromising soil ecology.

Simultaneous Cr(VI) reduction and Zn(II) biosorption by Stenotrophomonas sp. and constitutive expression of related genes.

OBJECTIVES: To investigate simultaneous Cr(VI) reduction and Zn(II) biosorption by a microorganism. RESULTS: A new isolate, Stenotrophomonas sp. TD3, simultaneously reduced Cr(VI) and achieved Zn(II) biosorption. The strain was resistant to other metals and salts, and its Zn biosorption could be stimulated greatly by CaCl2 and MgCl2, which has not been reported previously. Three putative Cr- or Zn-related genes, chrR, zupT and hmrE, were amplified and measured using quantitative real-time PCR to evaluate their mRNA expression levels in Stenotrophomonas sp. The three putative genes were not sensitive to Cr(6+), Zn(2+), Ca(2+), and Mg(2+), which suggested that they were constitutively expressed in Stenotrophomonas sp. TD3. CONCLUSIONS: These results improve our understanding of the mechanism and suggest that the strain is a potential candidate for facilitating remediation of Cr(VI) and Zn contaminated sites.

High-Quality Draft Genome Sequence of Leucobacter sp. Strain G161, a Distinct and Effective Chromium Reducer.

Here, we report the genome sequence for Leucobacter sp. strain G161 due to its distinct and effective hexavalent chromium reduction under aerobic growth conditions, followed by facultative anaerobic incubation. The draft genome sequence of Leucobacter sp. G161 comprises 3,554,188 bp, with an average G+C content of 65.3%, exhibiting 3,341 protein-coding genes and 55 predicted RNA genes.

Bioremediation of hexavalent chromate using permeabilized Brevibacterium sp. and Stenotrophomonas sp. cells.

Bioremediation has been found to be a useful method for removing hexavalent chromium (Cr(VI)), which is very toxic, from wastewater. Two strains of bacteria that were able to reduce Cr(VI) effectively were isolated from Cr(VI) contaminated soil samples and identified as Brevibacterium sp. K1 and Stenotrophomonas sp. D6, respectively, based on 16S rRNA gene sequence analyses. Brevibacterium sp. K1 and Stenotrophomonas sp. D6 could grow in Luria-Broth medium containing K2Cr2O7 at 1000 and 1600 mg/L, respectively, and they completely reduced the Cr(VI) in LB medium containing K2Cr2O7 at 200 mg/L within 72 h. Further analyses revealed that permeabilized K1 and D6 cells reduced Cr(VI) more effectively than did the resting cells. Triton X-100 was the best permeabilizing agent that was tested. The permeabilized cells of both strains could completely reduce Cr(VI) in industrial wastewater twice before needing to be replenished. The results suggested that these chromate-reducing bacteria are potential candidates for practical use biotreating industrial effluents containing Cr(VI) with Stenotrophomonas sp. D6 being the more effective bacterium.

Comparative evaluations on bio-treatment of hexavalent chromate by resting cells of Pseudochrobactrum sp. and Proteus sp. in wastewater.

Two marine bacterial strains, B5 and H24, were isolated from long-term Cr(VI) contaminated seawater and identified as Pseudochrobactrum and Proteus, respectively, based on 16S rRNA gene sequence analyses. Both strains were examined for their tolerance to Cr(VI) and other metal salts and their abilities to reduce Cr(VI) to trivalent chromium [Cr(III)]. Growing cells of Pseudochrobactrum sp. B5 and Proteus sp. H24 could tolerate Cr(VI) at a concentration of 2000 and 1500 mg/l and completely reduce 1000 mg/l Cr(VI) in LB medium within 96 and 144 h, respectively. Resting cells of the two strains were able to reduce 200mg/l Cr(VI) in Tris-HCl buffer within 16 and 24h, respectively. Furthermore, resting cells of both strains were able to reduce Cr(VI) in industrial wastewaters three times consecutively. Overall, this study provides evidence of the potential for application of chromate-reducing bacteria to direct Cr(VI) decontamination of industrial effluents.

Distinct and effective biotransformation of hexavalent chromium by a novel isolate under aerobic growth followed by facultative anaerobic incubation.

A bacterial isolate (G161) with high Cr(VI)-reducing capacity was isolated from Cr(VI)-contaminated soil and identified as Leucobacter sp. on the basis of 16S rRNA gene sequence analysis. The isolate was a Gram-positive, aerobic rod. The hexavalent chromate-reducing capability of the isolate was investigated under three conditions of oxygen stress. The isolate was found to reduce Cr(VI) under all conditions but performed most effectively during aerobic growth followed by facultative anaerobic incubation. Under these conditions, the isolate tolerated K(2)Cr(2)O(7) concentrations up to 1,000 mg/l and completely reduced 400 mg/l K(2)Cr(2)O(7) within 96 h. The strain reduced Cr(VI) over a wide range of pH (6.0-11.0) and temperatures (15-45 °C) with optimum performance at pH 8.0 and 35 °C. The presence of other metals, such as Ca(2+), Co(2+), Cu(2+), Mn(2+), Ni(2+), and Zn(2+), induced no effect or else played a stimulatory role on Cr(VI)-reduction activity of the strain. The strain was tested for Cr(VI) removal in wastewaters and proved capable of completely reducing the contained Cr(VI). This is the novel report of a bacterial growth and Cr(VI)-reduction process under sequential aerobic growth and facultative anaerobic conditions. The study suggested that the isolate possesses a distinct capability for Cr(VI) reduction which could be harnessed for the detoxification of chromate-contaminated wastewaters.

Expression and functional analysis of aminotransferase involved in indole-3-acetic acid biosynthesis in Azospirillum brasilense Yu62.

In this study, atrC (a novel gene from Azospirillum brasilense identified in our laboratory) was expressed in Escherichia coli, and SDS-PAGE analysis of the expressed AtrC revealed the apparent molecular weight of 45 kD. When analyzed under non-denaturing PAGE conditions and using L-tryptophan as a substrate, the purified AtrC protein exhibited aminotransferase activity, while crude protein extracts from A. brasilense Yu62 showed two activity bands with molecular masses estimated as 44 and 66 kD. Thus, we deduced that AtrC protein is identical to the 44 kD band of crude protein extracts. The optimal temperature and pH for the catalytic activity of the purified AtrC are 30 degrees C and pH 7.0, respectively.

Characterization of two trpE genes encoding anthranilate synthase alpha-subunit in Azospirillum brasilense.

The previous report from our laboratory has recently identified a new trpE gene (termed trpE2) which exists independently in Azospirillum brasilense Yu62. In this study, amplification of trpE(G) (termed trpE1(G) here) confirmed that there are two copies of trpE gene, one trpE being fused into trpG while the other trpE existed independently. This is the first report to suggest that two copies of the trpE gene exist in this bacterium. Comparison of the nucleotide sequence demonstrated that putative leader peptide, terminator, and anti-terminator were found upstream of trpE1(G) while these sequence features did not exist in front of trpE2. The beta-galactosidase activity of an A. brasilense strain carrying a trpE2-lacZ fusion remained constant at different tryptophan concentrations, but the beta-galactosidase activity of the same strain carrying a trpE1(G)-lacZ fusion decreased as the tryptophan concentration increased. These data suggest that the expression of trpE1(G) is regulated at the transcriptional level by attenuation while trpE2 is constantly expressed. The anthranilate synthase assays with trpE1(G)- and trpE2- mutants demonstrated that TrpE1(G) fusion protein is feedback inhibited by tryptophan while TrpE2 protein is not. We also found that both trpE1(G) and trpE2 gene products were involved in IAA synthesis.

Development of methods for determination of the residues of 15 pesticides in medicinal herbs Isatis indigotica Fort. by capillary gas chromatography with electron capture or flame photometric detection.

Two multiresidue methods were developed for the determination of 15 pesticides (organochlorines, organphosphorus compounds, pyrethroids, and fungicides) in medicinal herbs Isatis indigotica Fort. and its formulations. The analytical procedure is based on ultrasonic assisted extraction and liquid-liquid extraction (LLE). After solvents were added, the raw material or granule sample was sonicated in an ultrasonic water bath and then centrifuged, filtered, and cleaned up by LLE. The infusion sample was extracted with petroleum ether by LLE. The pesticide residues were determined by capillary gas chromatography with electron-capture or flame photometric detection. Recoveries with the method at concentrations between 0.4 microg/kg and 10 mg/kg ranged from 70.2 to 119.5% for raw material, 73.2 to 105.1% for granule formulation, and 72.8 to 113.3% for infusion formulation. The relative standard deviation values were <20% for all of the pesticides studied. The pesticide detection limits were within the ranges 0.3-0.5 microg/L for endosulfan, 3-7.5 microg/L for pyrethroids, 0.7-32.5 microg/L for organophosphorus pesticides, and 0.1-0.6 microg/L for the other pesticides. The proposed methods are simple and rapid and provide simultaneous determination of pesticide residues in Isatis indigotica Fort. with acceptable recoveries and repeatability and an adequate limit of determination.