MlrA, an Essential Enzyme for Microcystins and Nodularin on First Step Biodegradation in Microcystin-Degrading Bacteria.

Microcystin-degrading bacteria first degrade microcystins by microcystinase A (MlrA) to cleave the cyclic structure of microcystins at the Adda-Arg site of microcystin-LR, microcystin-RR, and microcystin-YR, but the cleavage of the other microcystins was not clear. In our study, the microcystin-degrading bacterium Sphingopyxis sp. C-1 as wild type and that of mlrA-disrupting mutant, Sphingopyxis sp. CMS01 were used for microcystins biodegradation. The results showed MlrA degraded microcystin-LA, microcystin-LW, microcystin-LY, microcystin-LF, and nodularin. MlrA could cleave the Adda-L-amino acid site.

Dynamics of the prokaryotic and eukaryotic microbial community during a cyanobacterial bloom.

Toxic cyanobacterial blooms frequently develop in eutrophic freshwater bodies worldwide. Microcystis species produce microcystins (MCs) as a cyanotoxin. Certain bacteria that harbor the mlr gene cluster, especially mlrA, are capable of degrading MCs. However, MC-degrading bacteria may possess or lack mlr genes (mlr+ and mlr- genotypes, respectively). In this study, we investigated the genotype that predominantly contributes to biodegradation and cyanobacterial predator community structure with change in total MC concentration in an aquatic environment. The 2 genotypes coexisted but mlr+ predominated, as indicated by the negative correlation between mlrA gene copy abundance and total MC concentration. At the highest MC concentrations, predation pressure by Phyllopoda, Copepoda, and Monogononta (rotifers) was reduced; thus, MCs may be toxic to cyanobacterial predators. The results suggest that cooperation between MC-degrading bacteria and predators may reduce Microcystis abundance and MC concentration.

Recombinant protein expression of Moringa oleifera lectin in methylotrophic yeast as active coagulant for sustainable high turbid water treatment.

The natural coagulant Moringa oleifera lectin (MoL) as cationic protein is a promising candidate in coagulation process of water treatment plant. Introducing the gene encoding MoL into a host, Pichia pastoris, to secrete soluble recombinant protein is assessed in this study. Initial screening using PCR confirmed the insertion of MoL gene, and SDS-PAGE analysis detected the MoL protein at 8 kDa. Cultured optimization showed the highest MoL protein at 520 mg/L was observed at 28 °C for 144 h of culturing by induction in 1% methanol. Approximately, 0.40 mg/mL of recombinant MoL protein showed 95 ± 2% turbidity removal of 1% kaolin suspension. In 0.1% kaolin suspension, the concentration of MoL at 10 µg/mL exhibits the highest turbidity reduction at 68 ± 1%. Thus, recombinant MoL protein from P. pastoris is an effective coagulant for water treatment.

Phenotypic and genetic characterization of multidrug-resistant Staphylococcus aureus in the tropics of Southeast Asia.

Antibiotic resistance has become a major public health problem throughout the world. The presence of antibiotic-resistant bacteria such as Staphylococcus aureus and antibiotic resistance genes (ARGs) in hospital wastewater is a cause for great concern today. In this study, 276 Staph. aureus isolates were recovered from hospital wastewater samples in Malaysia. All of the isolates were screened for susceptibility to nine different classes of antibiotics: ampicillin, ciprofloxacin, gentamicin, kanamycin, erythromycin, vancomycin, trimethoprim and sulfamethoxazole, chloramphenicol, tetracycline and nalidixic acid. Screening tests showed that 100 % of Staph.aureus isolates exhibited resistance against kanamycin, vancomycin, trimethoprim and sulfamethoxazole and nalidixic acid. Additionally, 91, 87, 50, 43, 11 and 8.7 % of isolates showed resistance against erythromycin, gentamicin, ciprofloxacin, ampicillin, chloramphenicol and tetracycline, respectively. Based on these results, 100 % of isolates demonstrated multidrug-resistant (MDR) characteristics, displaying resistance against more than three classes of antibiotics. Of 276 isolates, nine exhibited resistance to more than nine classes of tested antibiotics; these were selected for antibiotic susceptibility testing and examined for the presence of conserved ARGs. Interestingly, a high percentage of the selected MDR Staph.aureus isolates did not contain conserved ARGs. These results indicate that non-conserved MDR gene elements may have already spread into the environment in the tropics of Southeast Asia, and unique resistance mechanisms against several antibiotics may have evolved due to stable, moderate temperatures that support growth of bacteria throughout the year.

Photocatalytic removal of microcystin-LR by advanced WO3-based nanoparticles under simulated solar light.

A series of advanced WO3-based photocatalysts including CuO/WO3, Pd/WO3, and Pt/WO3 were synthesized for the photocatalytic removal of microcystin-LR (MC-LR) under simulated solar light. In the present study, Pt/WO3 exhibited the best performance for the photocatalytic degradation of MC-LR. The MC-LR degradation can be described by pseudo-first-order kinetic model. Chloride ion (Cl-) with proper concentration could enhance the MC-LR degradation. The presence of metal cations (Cu2+ and Fe3+) improved the photocatalytic degradation of MC-LR. This study suggests that Pt/WO3 photocatalytic oxidation under solar light is a promising option for the purification of water containing MC-LR.

Enhanced removal of contaminant using the biological film, anoxic-anaerobic-aerobic and electro-coagulation process applied to high-load sewage treatment.

In order to explore a new treatment process applying to decentralized domestic sewage treatment, and enhance removal of total nitrogen (TN), total phosphorus (TP) and chemical oxygen demand (COD), a novel system integrating anoxic-anaerobic-aerobic (reversed A2O) and electro-coagulation (EC) process was studied, and complex biological media (CMB) was used as suspended carrier for biofilm development. In this work, TN, TP and COD removal performance were investigated with consideration of three major factors, hydraulic retention time (HRT), organic loading rate (OLR) and sludge recycle ratio (SRR). Results showed that (1) The optimum HRT was between 8 and 12 h. The removal efficiencies of TN, TP and COD were about 68%, 95% and 95%, respectively. (2) With the increase of OLR, the removal efficiency of TN increased slowly. But it increased first and then declined for COD and TP removal. Their maximum were attained when OLR was 1.8 g(COD)/(L d), and they were 96% and 93%, respectively. (3) The optimum SRR was 75%. The COD, TN and TP removal efficiencies were about 95%, 72% and 98%, respectively. In this system, the maximum TN and COD removal were achieved in anoxic tank, but it was achieved in aerobic tank for TP removal. The EC bed enhanced the effluent quality, especially the efficiency in advanced P removal. From these results, it was concluded that the new process could be a reliable option for providing excellent effluent quality.

Compensation of blue phase I by blue phase II in optoeletronic device.

Compensation effect of blue phase I (BP I) with blue phase II (BP II) liquid crystal was demonstrated. BP I and BP II were co-exist in the optoeletronic device by polymer stabilization. Consequently, disadvantages of BP I and BP II were greatly improved by compensation effect and resulted in high contrast ratio, low hysteresis and fast falling time. Mechanism of compensation effect was explained by relaxation ability of lattice structure under electrical field and compensation structure was well confirmed by Bragg's reflectance spectrum and Commission International de l'Éclairage chromaticity diagram.

Degradation of microcystins by an electrochemical oxidative electrode cell.

Microcystins (MCs), which are produced by cyanobacteria, are one of the most serious problems that threaten quality of drinking water and public health. In this study, an electrolysis cell with no electrolyte is demonstrated to degrade MCs (MC-RR, MC-YR and MC-LR) in both high and low concentrations. In addition, degradation of MCs was studied under different current densities. The results revealed that the electrolysis cell could degrade MCs successfully. It was observed that degradation of a single MC was faster than mixed types and statistical analysis revealed that the degradation rate of all the three MCs did not show much difference in mixed degradation. Analysis of hydroxyl radical concentration suggested a possible role of the hydroxyl radical in degradation of MCs. We propose that the electrolysis cell could be a promising treatment for effective removal of MCs in situ, especially in water purification plants where low amounts of salts (electrolytes) are present.

Microcystin-LR incorporated into colonic cells through probenecid-sensitive transporters leads to upregulated MCP-1 expression induced by JNK activation.

Harmful algae that inhabit eutrophic lakes produce cyanotoxic microcystins. Therefore, the relationship between chronic exposure to microcystins via drinking water and organ disorders has been investigated. The present study aimed to determine whether representative microcystin-LR is involved in increased monocyte chemoattractant protein-1 (MCP-1) expression in rat colonic mucosa and enterocyte-like differentiated Caco-2 cells. The mRNA expression of MCP-1 was increased in the colons of rats administered with microcystin-LR, compared with controls. Furthermore, mRNA levels of MCP-1 expression significantly and positively correlated with those of Adhesion G Protein-Coupled Receptor E1 (ADGRE1; EMR1; F4/80), an indicator of macrophage infiltration, suggesting that increased MCP-1 expression induced by microcystin-LR promotes macrophage infiltration into the colon. Microcystin-LR increased MCP-1 expression in enterocyte-like differentiated Caco-2 cells, by activating c-Jun N-terminal kinase (JNK), but not extracellular signal-regulated kinase (ERK) or p38. The findings of transporter inhibitors indicated that microcystin-LR is incorporated into cells via ATP Binding Cassette (ABC) or solute carrier (SLC) transporters other than the organic anion transporting polypeptides (OATPs)1B1, 1B3, 2B1, and 1A2, which this leads to increased MCP-1 expression in the colon through activating JNK. Thus, increased MCP-1 expression induced by microcystin-LR might be a trigger for initiating tumorigenesis with inflammation in the colon because increased MCP-1 expression induces inflammation associated with macrophage infiltration into the colon, and chronic inflammation is associated with the initiation of tumorigenesis.

Enhancement of astaxanthin accumulation using black light in Coelastrum and Monoraphidium isolated from Malaysia.

Microalgae are important microorganisms which produce potentially valuable compounds. Astaxanthin, a group of xanthophyll carotenoids, is one of the most powerful antioxidants mainly found in microalgae, yeasts, and crustaceans. Environmental stresses such as intense light, drought, high salinity, nutrient depletion, and high temperature can induce the accumulation of astaxanthin. Thus, this research aims to investigate the effect of black light, also known as long-wave ultraviolet radiation or UV-A, as a stressor on the accumulation of astaxanthin as well as to screen the antioxidant property in two tropical green algal strains isolated from Malaysia, Coelastrum sp. and Monoraphidium sp. SP03. Monoraphidium sp. SP03 showed a higher growth rate (0.66 day-1) compared to that of Coelastrum sp. (0.22 day-1). Coelastrum sp. showed significantly higher accumulation of astaxanthin in black light (0.999 g mL culture-1) compared to that in control condition (0.185 g mL-1). Similarly, Monoraphidium sp. SP03 showed higher astaxanthin content in black light (0.476 g mL culture-1) compared to that in control condition (0.363 g mL culture-1). Coelastrum sp. showed higher scavenging activity (30.19%) when cultured in black light condition, indicating a correlation between the antioxidant activity and accumulation of astaxanthin. In this study, black light was shown to possess great potential to enhance the production of astaxanthin in microalgae.

Draft Genome Sequence of the Prazosin-Degrading Bacillus sp. Strain PR5, Isolated from a River Receiving Hospital and Urban Wastewater in Malaysia.

We report the complete genome sequence of Bacillus sp. strain PR5, isolated from a river receiving hospital and urban wastewater in Malaysia, which demonstrated a high capability for degrading prazosin. This genome sequence of 4,525,264 bp exhibited 41.5% GC content, 4,402 coding sequences, and 32 RNAs.

Limitation of nutrients stimulates musty odor production by Streptomyces sp. isolated from a tropical environment.

Musty odor production by actinomycetes is usually related to the presence of geosmin and 2-methylisoborneol (2-MIB), which are synthesized by enzymes encoded by the geoA and tpc genes, respectively. Streptomyces spp. strain S10, which was isolated from a water reservoir in Malaysia, has the ability to produce geosmin when cultivated in a basal salt (BS) solid medium, but no 2-MIB production occurred during growth in BS medium. Strain S10 could produce higher levels of geosmin when the phosphate concentration was limited to 0.05 mg/L, with a yield of 17.53 ± 3.12 ✕ 105 ng/L, compared with growth in BS medium. Interestingly, 2-MIB production was suddenly detected when the nitrate concentration was limited to 1.0 mg/L, with a yield of 1.4 ± 0.11 ✕ 105 ng/L. Therefore, it was concluded that phosphate- and nitrate-limiting conditions could induce the initial production of geosmin and 2-MIB by strain S10. Furthermore, a positive amplicon of geoA was detected in strain S10, but no tpc amplicon was detected by PCR analysis. Draft genome sequence analysis showed that one open reading frame (ORF) contained a conserved motif of geosmin synthase with 95% identity with geoA in Streptomyces coelicolor A3 (2). In the case of the tpc genes, it was found that one ORF showed 23% identity to the known tpc gene in S. coelicolor A3(2), but strain S10 lacked one motif in the N-terminus.

Removal of fluoride from aqueous solution by adsorption onto Kanuma mud.

Kanuma mud, a geomaterial, is used as an adsorbent for the removal of fluoride from water. The influences of contact time, solution pH, adsorbent dosage, initial fluoride concentration and co-existing ions were investigated by batch equilibration studies. The rate of adsorption was rapid with equilibrium being attained after about 2 h, and the maximum removal of fluoride was obtained at pH 5.0-8.0. The Freundlich isotherm model was found to represent the measured adsorption data well. The negative value of the thermodynamic parameter ΔG suggests the adsorption of fluoride by Kanuma mud was spontaneous, the endothermic nature of adsorption was confirmed by the positive ΔH value. The negative ΔS value for adsorbent denoted decreased randomness at the solid/liquid interface. The adsorption process using Kanuma mud followed the pseudo-second-order kinetic model. Fluoride uptake by the Kanuma mud was a complex process and intra-particle diffusion played a major role in the adsorption process. It was found that adsorbed fluoride could be easily desorbed by washing the adsorbent with a solution of pH 12. This indicates the material could be easily recycled.

Draft Genome Sequence of the Microcystin-Degrading Bacterium Novosphingobium sp. Strain MD-1.

This report describes the whole-genome sequence of a microcystin-degrading bacterium, Novosphingobium sp. strain MD-1, isolated from a lake in Japan. The Novosphingobium sp. strain MD-1 genome had a total length of 4,617,766 bp. Moreover, strain MD-1 showed a conserved microcystin-degrading gene cluster (mlrA to mlrF), similar to Sphingopyxis sp. strain C-1.

Elucidation of prazosin biodegradation by isolated Bacillus spp. from the tropical environment.

Prazosin (PRZ), a drug used to treat hypertensive patients, is an emergent contaminant in water systems. PRZ is an alpha-adrenergic receptor blocker used to treat anxiety, and is believed to reach the environment through human excretion, irresponsible disposal of unused medicine, and waste products from manufacturing plants. The purpose of this research was to isolate and characterize potential microbes for PRZ biodegradation and to identify the degradation pathway. After screening, isolated strain STP3 showed a capability for PRZ degradation and was chosen for further analysis. Resting cell assays with PRZ were conducted to identify the intermediate metabolites formed from biodegradation by Fourier transform ion cyclotron resonance mass spectrometer (FT-ICR MS) analysis. Two metabolites degraded from PRZ by STP3 were successfully found, and as these metabolites are derived from the main structure of PRZ, their presence proved PRZ degradation. Draft genome sequencing analysis of STP3 was performed to identify potential enzymes for PRZ biodegradation based on the metabolites found.

Influence of metal addition on ethanol production with Pichia stipitis ATCC 58784.

Trace metals always act as cofactors or coenzymes in many cellular processes. Deficiency or excess of some metals will affect the fermentation of lignocellulosic hydrolysate. In order to make sure the deficient or excessive states of metals in culture medium, metal contents analysis was conducted in Pichia stipitis ATCC 58784 cells, synthetic medium, and diluted acid hydrolysate of rice straw. The results showed that Cu, Ni, and Co were deficient, and Al was a little excessive. So the influences of Cu(2+), Al(3+), Ni(2+), and Co(2+) additions on the growth and ethanol production of ATCC 58784 were further researched. Low concentration additions of Cu(2+) and Al(3+) (<0.24 mM and <0.23 mM, respectively) improved biomass growth of ATCC 58784 by 34 and 13%, respectively; however, higher concentrations decreased biomass growth. On the other hand, addition of Cu(2+) (0.39 mM) did not affect volumetric ethanol production significantly (P = 0.05) and addition of Al(3+) (0.38 mM) showed no influence on volumetric ethanol production (P = 0.68). Addition of 0.074 mM Co(2+) inhibited biomass growth of ATCC 58784 by 13% and volumetric ethanol production by 10%. The biomass growth and volumetric ethanol production of ATCC 58784 was arrested by the addition of 0.33 mM of Ni(2+) by 53 and 65%, respectively.

Electrochemical degradation of phenol using electrodes of Ti/RuO(2)-Pt and Ti/IrO(2)-Pt.

Electrochemical degradation of phenol was evaluated at two typical anodes, Ti/RuO(2)-Pt and Ti/IrO(2)-Pt, for being a treatment method in toxic aromatic compounds. The influences of current density, dosage of NaCl, initial phenol concentration on electrochemical phenol degradation were investigated. It was found that Ti/RuO(2)-Pt anode had a higher oxygen evolution potential than Ti/IrO(2)-Pt anode, which will increase the current efficiency for electrochemical degradation, and the instantaneous current efficiency (ICE) was relatively higher at the initial time during phenol electrolysis. HOCl formed during electrolysis would play an important role on the oxidation of phenol. For the Ti/RuO(2)-Pt anode, phenol concentration decreased from around 8mg/L to zero after 30min of electrolysis with 0.3g/L NaCl as supporting electrolyte at the current density of 10mA/cm(2). Although phenol could be completely electrochemical degraded at both Ti/RuO(2)-Pt and Ti/IrO(2)-Pt anodes, phenol degradation was slower at the Ti/IrO(2)-Pt anode than at the Ti/RuO(2)-Pt anode due to the fact that passivation was to be found at the Ti/IrO(2)-Pt anode.

Response of microcystin biosynthesis and its biosynthesis gene cluster transcription in Microcystis aeruginosa on electrochemical oxidation.

Microcystin-LR (MC-LR), which is one of the most commonly found microcystins (MCs) in fresh water, has been proved to be a potential tumour promoter and classified as 2B by the International Agency for Research on Cancer. MC-LR decomposition and inhibition of MC-LR production in Microcystis aeruginosa were investigated under electrolysis condition using an electrolysis cell consisting of Ti/Pt electrodes and Nafion membrane. The relationship between the decrease in MC-LR concentration and transcription of MC-LR synthesis gene clusters was determined by performing real-time reverse transcription polymerase chain reaction (RT-qPCR) to monitor changes in the levels of transcription encoding mcyB and mcyD (cDNA to DNA) in M. aeruginosa NIES 1086 under electrolysis condition and three different conditions (i.e. oxygenated, air aerated and unaerated) as controls. Cell density decreased from day 2 under electrolysis than under the three controls. Intracellular MC-LR concentration was approximately 33 fg cell-1 under electrolysis from days 4 to 8, while those in the other conditions ranged in 40-50 fg cell-1. The mcyB transcription continuously decreased from day 2 to nondetectable level in day 6 under electrolysis, while this transcription was stabilised under the three controls. This result suggested that oxidative stress, such as hydroxyl radicals, played an important role in the down-regulation of mcyB and mcyD gene transcription level and the MC-LR concentration and cell density of M. aeruginosa.

Molecular characterization of multi-drug resistant Escherichia coli isolates from tropical environments in Southeast Asia.

The emergence of antibiotic resistance among multidrug-resistant (MDR) microbes is of growing concern, and threatens public health globally. A total of 129 Escherichia coli isolates were recovered from lowland aqueous environments near hospitals and medical service centers in the vicinity of Kuala Lumpur, Malaysia. Among the eleven antibacterial agents tested, the isolates were highly resistant to trimethoprim-sulfamethoxazole (83.7%) and nalidixic acid (71.3%) and moderately resistant to ampicillin and chloramphenicol (66.7%), tetracycline (65.1%), fosfomycin (57.4%), cefotaxime (57.4%), and ciprofloxacin (57.4%), while low resistance levels were found with aminoglycosides (kanamycin, 22.5%; gentamicin, 21.7%). The presence of relevant resistance determinants was evaluated, and the genotypic resistance determinants were as follows: sulfonamides (sulI, sulII, and sulIII), trimethoprim (dfrA1 and dfrA5), quinolones (qnrS), ß-lactams (ampC and blaCTX-M), chloramphenicol (cmlA1 and cat2), tetracycline (tetA and tetM), fosfomycin (fosA and fosA3), and aminoglycosides (aphA1 and aacC2). Our data suggest that multidrug-resistant E. coli strains are ubiquitous in the aquatic systems of tropical countries and indicate that hospital wastewater may contribute to this phenomenon.

Microbial Diversity in Decaying Oil Palm Empty Fruit Bunches (OPEFB) and Isolation of Lignin-degrading Bacteria from a Tropical Environment.

Oil palm empty fruit bunches (OPEFB) are the most abundant, inexpensive, and environmentally friendly lignocellulosic biomass in Malaysia. Investigations on the microbial diversity of decaying OPEFB may reveal microbes with complex enzymes that have the potential to enhance the conversion of lignocellulose into second-generation biofuels as well as the production of other value-added products. In the present study, fungal and bacterial diversities in decaying OPEFB were identified using Illumina MiSeq sequencing of the V3 region of the 16S rRNA gene and V4 region of the 18S rRNA gene. Fungal diversity in decaying OPEFB was dominated by the phylum Ascomycota (14.43%), while most of the bacterial sequences retrieved belonged to Proteobacteria (76.71%). Three bacterial strains isolated from decaying OPEFB, designated as S18, S20, and S36, appeared to grow with extracted OPEFB-lignin and Kraft lignin (KL) as the sole carbon source. 16S rRNA gene sequencing identified the 3 isolates as Paenibacillus sp.. The molecular weight distribution of KL before and after degradation showed significant depolymerization when treated with bacterial strains S18, S20, and S36. The presence of low-molecular-weight lignin-related compounds, such as vanillin and 2-methoxyphenol derivatives, which were detected by a GC-MS analysis, confirmed the KL-degrading activities of isolated Paenibacillus strains.