Neisseria meningitidis and Neisseria gonorrhoeae are differently adapted in the regulation of denitrification: single nucleotide polymorphisms that enable species-specific tuning of the aerobic–anaerobic switch

The closely related pathogenic Neisseria species N. meningitidis and N. gonorrhoeae are able to respire in the absence of oxygen, using nitrite as an alternative electron acceptor. aniA (copper-containing nitrite reductase) is tightly regulated by four transcriptional regulators: FNR (fumarate and nitrate reductase), NarP, FUR (Ferric uptake regulator) and NsrR. The four regulators control expression of aniA in N. meningitidis by binding to specific and distinct regions of the promoter. We show in the present study that FUR and NarP are both required for the induction of expression of aniA in N. meningitidis, and that they bind adjacent to one another in a non-co-operative manner. Activation via FUR/NarP is dependent on their topological arrangement relative to the RNA polymerase-binding site. Analysis of the sequence of the aniA promoters from multiple N. meningitidis and N. gonorrhoeae strains indicates that there are species-specific single nucleotide polymorphisms, in regions predicted to be important for regulator binding. These sequence differences alter both the in vitro DNA binding and the promoter activation in intact cells by key activators FNR (oxygen sensor) and NarP (which is activated by nitrite in N. meningitidis). The weak relative binding of FNR to the N. gonorrhoeae aniA promoter (compared to N. meningitidis) is compensated for by a higher affinity of the gonococcal aniA promoter for NarP. Despite containing nearly identical genes for catalysing and regulating denitrification, variations in the promoter for the aniA gene appear to have been selected to enable the two pathogens to tune differentially their responses to environmental variables during the aerobic–anaerobic switch.

Regulation of Denitrification Genes in Neisseria meningitidis by Nitric Oxide and the Repressor NsrR

The human pathogen Neisseria meningitidis is capable of growth using the denitrification of nitrite to nitrous oxide under microaerobic conditions. This process is catalyzed by two reductases: nitrite reductase (encoded by aniA) and nitric oxide (NO) reductase (encoded by norB). Here, we show that in N. meningitidis MC58 norB is regulated by nitric oxide via the product of gene NMB0437 which encodes NsrR. NsrR is a repressor in the absence of NO, but norB expression is derepressed by NO in an NsrR-dependent manner. nsrR-deficient mutants grow by denitrification more rapidly than wild-type N. meningitidis, and this is coincident with the upregulation of both NO reductase and nitrite reductase even under aerobic conditions in the absence of nitrite or NO. The NsrR-dependent repression of aniA (unlike that of norB) is not lifted in the presence of NO. The role of NsrR in the control of expression of aniA is linked to the function of the anaerobic activator protein FNR: analysis of nsrR and fnr single and nsrR fnr double mutants carrying an aniA promoter lacZ fusion indicates that the role of NsrR is to prevent FNR-dependent aniA expression under aerobic conditions, indicating that FNR in N. meningitidis retains considerable activity aerobically.

Denitrification measurements of sediments using cores and chambers

Denitrification is commonly measured using in situ benthic chambers or laboratory incubations of sediment cores. These techniques are similar in principle but differ considerably in cost and practicality. Despite widespread use of both techniques, it is uncertain whether they give comparable results. We compared cores and chambers for measuring fluxes (dissolved oxygen [DO], N 2, NH4+, NO3- and NO 2-) and denitrification efficiency at 2 sites in Port Phillip Bay, Victoria, Australia. Overall, denitrification efficiency was not significantly different between cores and chambers, but fluxes of DO, NO 3- and NO2- differed. Chambers demonstrated higher levels of oxygen consumption and net fluxes of NO 3- and NO2- out of the sediment, suggesting that denitrification and nitrification were closely coupled. In contrast, there was a greater relative importance for uncoupled denitrification in cores as indicated by reduced oxygen consumption and net fluxes of NO 3- into the sediment. We conclude that cores and chambers give different flux results and therefore are not comparable techniques for measuring denitrification. To ascertain the cause of this, we tested the hypothesis that cores failed to adequately incorporate the impacts of macrofauna on fluxes, due to the small size of cores relative to chambers. However, densities of macrofauna were not significantly different in cores and chambers. We then hypothesised that disturbance during core collection, transportation, and handling may account for differences, but cores deployed in situ and in the laboratory gave similar results. We suggest that compression of sediment during insertion of core cylinders into the sediment may account for differences between core and chamber fluxes.

Application of submerged membrane bioreactor for aquaculture effluent reuse

Discharging the nutrient rich aquaculture effluents into inland water bodies and oceans is becoming a serious concern due to the adverse effect that brings in the form of eutrophication and subsequent damages to those waters. A laboratory scale biological reactor consisting of a denitrifying compartment followed by a submerged membrane bioreactor (SMBR) compartment was used to treat 40 L d−1 of aquaculture effluent with an average concentration of 74 mg L−1 nitrate (NO3 − ). Sugar was added to the aquaculture effluent in order that to enter into the denitrifying compartment at a carbon: nitrogen ratio (C:N) of 2:1 and 4:1. A hollow fibre membrane with a pore size of 0.4 μm and a filtration area of 0.20 m2 was used in the SMBR and was operated at an average flux of 0.20 m3 m−2 d−1. An intermittent suction period of 12 min followed by a relaxation period of 3 min was maintained in the SMBR throughout the experiment. Different aeration rates of 1, 3, 5 and 10 Lpm were applied to the SMBR to determine the rate of membrane fouling and 5 Lpm aeration rate was found to be optimum with respect to the rate of fouling of membrane at a C:N ratio of 4:1. The average rate of fouling at 1, 3, 5 and 10 Lpm were 1.17, 0.70, 0.48 and 0.52 kPa d−1, respectively. The increase in the rate of fouling when the aeration was increased from 5 to 10 Lpm may be due to the breakage of suspended particles into finer particles which could have increased the fouling of membrane. It was also found that increasing the C:N ratio from 2:1 to 4:1 resulted in more cake being formed on the membrane surface as well as an increase in the reduction of NO3 − from 64% to 78%. Preliminary calculations show that 2.4 to 3.2 g of suspended solids could be accumulated per square meter of membrane surface before physical cleaning of membrane is required (at a transmembrane pressure of 20 kPa).

Performance of suspended and attached growth MBR systems in treating high strength synthetic wastewater

The performance of laboratory-scale attached growth (AG) and suspended growth (SG) membrane bioreactors (MBRs) was evaluated in treating synthetic wastewater simulating high strength domestic wastewater. This study investigated the influence of sponge suspended carriers in AG-MBR system, occupying 15% reactor volume, on the removal of chemical oxygen demand (COD), total nitrogen (TN) and total phosphorus (TP), and compared it to that of SG-MBR. Results showed that the removal efficiencies of COD, TN and TP in AG-MBR were 98%, 89% and 58%, respectively as compared to 98%, 74% and 38%, respectively in SG-MBR. Improved TN removal in AG-MBR systems was primarily based on simultaneous nitrification and denitrification (SND) process. These results infer that the presence of small bio-particles having higher microbial activity and the growth of complex biomass captured within the suspended sponge carriers resulted in improved TN and TP removal in AG-MBR.

Powdered activated carbon for fouling reduction of membrane in a pilot-scale recirculation aquaculture system

Recirculating aquaculture systems (RAS) are essential for the reduction in fresh water usage as well as the discharge of nutrients along with aquaculture effluents. A RAS consisting of an anoxic reactor, a membrane bioreactor (MBR) and a UV-disinfection unit was used to process 10,000 L/d of aquaculture effluent providing high-quality treated water for recirculation to a Barramundi fish culture. The system maintained low levels of nitrate (<20 mg/L), nitrite (<3 mg/L) and ammonia (<0.6 mg/L) in the fish tank. Permeate from the membrane that was recirculated to the fish tank contained <21 mg/L of nitrate, <2 mg/L of nitrite and 0 mg/L of ammonia. However, the rate of fouling of the membrane in the MBR was around 1.47 kPa/d, and the membrane in the MBR required cleaning due to fouling after 16 days. Cleaning of the membrane was initiated when the TMP reached around 25 to 30 kPa. In order to reduce the rate of fouling, 500 mg of powdered activated carbon (PAC) per litre of MBR volume was introduced, which decreased the rate of fouling to 0.90 kPa/d. Cleaning of membrane was needed only after 31 days of operation while maintaining the treated effluent quality. Thus the frequency of cleaning could be halved due to the introduction of PAC into the MBR.

The Nitric Oxide (NO)-Sensing Repressor NsrR of Neisseria meningitidis Has a Compact Regulon of Genes Involved in NO Synthesis and Detoxification

We have analyzed the extent of regulation by the nitric oxide (NO)-sensitive repressor NsrR from Neisseria meningitidis MC58, using microarray analysis. Target genes that appeared to be regulated by NsrR, based on a comparison between an nsrR mutant and a wild-type strain, were further investigated by quantitative real-time PCR, revealing a very compact set of genes, as follows: norB (encoding NO reductase), dnrN (encoding a protein putatively involved in the repair of nitrosative damage to iron-sulfur clusters), aniA (encoding nitrite reductase), nirV (a putative nitrite reductase assembly protein), and mobA (a gene associated with molybdenum metabolism in other species but with a frame shift in N. meningitidis). In all cases, NsrR acts as a repressor. The NO protection systems norB and dnrN are regulated by NO in an NsrR-dependent manner, whereas the NO protection system cytochrome c′ (encoded by cycP) is not controlled by NO or NsrR, indicating that N. meningitidis expresses both constitutive and inducible NO protection systems. In addition, we present evidence to show that the anaerobic response regulator FNR is also sensitive to NO but less so than NsrR, resulting in complex regulation of promoters such as aniA, which is controlled by both FNR and NsrR: aniA was found to be maximally induced by intermediate NO concentrations, consistent with a regulatory system that allows expression during denitrification (in which NO accumulates) but is down-regulated as NO approaches toxic concentrations.

Characterization of the extracellular polymeric substances and microbial community of aerobic granulation sludge exposed to cefalexin

This study characterizes the extracellular polymeric substances and bacterial community composition of aerobic granules exposed to cefalexin (CLX). The presence of CLX potentially decreases granular stabilities, resulting in a lowered granule diameter. Chemical oxygen demand and NH4+-N removal efficiencies were slightly decreased and the denitrification process was inhibited with CLX addition. Extracellular polymeric substance contents were significantly increased in aerobic granules exposed to CLX. The shifts of fluorescence intensities and peak locations in 3D-EEM fluorescence spectra indicated changes of EPS components. High-throughput sequencing analysis showed aerobic granules with CLX addition in synthetic wastewater had superior diversity of microbial species, and this was the reason that the level and components of EPS changed. The species richness for bacteria was increased from 341 to 352, which was revealed by Chao1. The Shannon index of diversity rose slightly from 3.59 to 3.73 with CLX addition. The abundance of Proteobacteria significantly decreased, while the abundance of Bacteroidetes and Chloroflexi underwent a highly significant increase in aerobic granules exposed to CLX.

Estimating hydrogen sulphide dissipation rate constant under the influence of different chemical dosing

Sewer odour and corrosion is caused by the reduction of sulphide ions and the release of hydrogen sulphide gas (H2S) into the sewer atmosphere. The reduction of sulphide is determined by its dissipation rate which depends on many processes such as emission, oxidation and precipitation that prevail in wastewater environments. Two factors that mainly affect the dissipation of sulphide are sewer hydraulics and wastewater characteristics; modification to the latter by dosing certain chemicals is known as one of the mitigation strategies to control the dissipation of sulphide. This study investigates the dissipation of sulphide in the presence of NaOH, Mg(OH)2, Ca(NO3)2 and FeCl3 and the dissipation rate is developed as a function of hydraulic parameters such as the slope of the sewer and the velocity gradient. Experiments were conducted in a 18m experimental sewer pipe with adjustable slope to which, firstly no chemical was added and secondly each of the above mentioned chemicals was supplemented in turn. A dissipation rate constant of 2×10^-6 for sulphide was obtained from experiments with no chemical addition. This value was then used to predict the sulphide concentration that was responsible for the emission of H2S gas in the presence of one of the above mentioned four chemicals. It was found that the performance of alkali substances (NaOH and Mg(OH)2) in suppressing the H2S gas emission was excellent while ferric chloride showed a moderate mitigating effect due to its slow reaction kinetics. Calcium nitrate was of little value since the wastewater used in this study experienced almost no biological growth. Thus the effectiveness of selected chemicals in suppressing H2S gas emission had the following order: NaOH ≥ Mg(OH)2 ≥ FeCl3 ≥ Ca(NO3)2.