This study focuses on the evaluation of the performance of Chlorella sp. in removing nutrient in aquaculture wastewater and its correlation with the kinetic growth of Chlorella sp. The treatment was applied with various Chlorella sp. inoculation dosage ranging from 0% to 60% (v/v) of wastewater. The optimum inoculation dosage was recorded at 30% (v/v) with effluent concentration of ammonia and orthophosphate recording at 0.012mgL(-1) and 0.647mgL(-1), respectively on Day 11. The optimum dosage for bio-flocculation process was obtained at 30mgL(-1) of Aspergillus niger with a harvesting efficiency of 97%. This type of development of phytoremediation with continuous bio-harvesting could promote the use of sustainable green technology for effective wastewater treatment.
The marine dinoflagellate Alexandrium minutum is known to produce saxitoxins that cause paralytic shellfish poisoning in human worldwide through consumption of the contaminated shellfish mollusks. Despite numerous studies on the growth physiology and saxitoxin production of this species, the knowledge on the molecular basis of nutrient uptakes in relation to toxin production in this species is limited. In this study, relative expressions of the high-affinity transporter genes of nitrate, ammonium, and phosphate (AmNrt2, AmAmt1 and AmPiPT1) and the assimilation genes, nitrate reductase (AmNas), glutamine synthase (AmGSIII) and carbamoyl phosphate synthase (AmCPSII) from A. minutum were studied in batch clonal culture condition with two nitrogen sources (nitrate: NO3- or ammonium: NH4+) under different N:P ratios (high-P: N:P of 14 and 16, and low-P: N:P of 155). The expression of AmAmt1 was suppressed in excess NH4+-grown condition but was not observed in AmNrt2 and AmNas. Expressions of AmAmt1, AmNrt2, AmNas, AmGSIII, AmCPSII, and AmPiPT1 were high in P-deficient condition, showing that A. minutum is likely to take up nutrients for growth under P-stress condition. Conversely, relative expression of AmCPSII was incongruent with cell growth, but was well correlated with toxin quota, suggesting that the gene might involve in arginine metabolism and related toxin production pathway. The expression of AmGSIII is found coincided with higher toxin production and is believed to involve in mechanism to detoxify the cells from excess ammonium stress. The gene regulation observed in this study has provided better insights into the ecophysiology of A. minutum in relation to its adaptive strategies in unfavorable environments.
The initial rate of NH(3)/NH(4)(+) accumulation in a medium containing L(3) Teladorsagia circumcincta was 0.18-0.6 pmol h(-1) larva(-1), which increased linearly with larval density. However it appeared that the larva-generated external concentration of NH(3)/NH(4)(+) did not exceed about 130 μM. The rate of NH(3)/NH(4)(+) accumulation increased with temperature between 4 °C and 37 °C, declined with increasing pH or increasing external NH(3)/NH(4)(+) concentration and was not significantly affected by the concentration of the phosphate buffer or by exsheathing the larvae. We infer from these data that the efflux of NH(3)/NH(4)(+) is a diffusive process and that the secreted or excreted NH(3)/NH(4)(+) is generated enzymatically rather than dissociating from the surface of the nematode. The enzymatic source of the NH(3)/NH(4)(+) is yet to be identified. Since the concentration of NH(3)/NH(4)(+) in the rumen and abomasum is higher than 130 μM, it is unlikely that T. circumcincta contributes to it, but NH(3)/NH(4)(+) may be accumulated from the rumen fluid by the nematode.
An appropriate tillage method must be implemented by maize growers to improve phosphorus dynamics in the soil in order to increase phosphorus uptake by plant. The objective of this study was to investigate the effects of tillage systems on phosphorus and its fractions in rhizosphere and non-rhizosphere soils under maize. An experimental field was established, with phosphate fertilizers applied to four treatment plots: continuous rotary tillage (CR), continuous no-tillage (CN), plowing-rotary tillage (PR), and plowing-no tillage (PN). Under the different tillage methods, the available P was increased in the non-rhizosphere region. However, the concentration of available P was reduced in the rhizosphere soil region. The soil available P decreased with the age of the crop until the maize reached physiological maturity. The non-rhizosphere region had 132.9%, 82.5%, 259.8%, and 148.4% more available P than the rhizosphere region under the CR, PR, CN, and PN treatments, respectively. The continuous no-tillage method (CN) improved the uptake of soil phosphate by maize. The concentrations of Ca2-P, Ca8-P, Fe-P, Al-P and O-P at the maturity stage were significantly lower than other seedling stages. However, there was no significant relationship between total P and the P fractions. Therefore, a continuous no-tillage method (CN) can be used by farmers to improve phosphorus availability for spring maize. Soil management practices minimizing soil disturbance can be used to impove phosphorus availability for maize roots, increase alkaline phosphatase activity in the rhizosphere soil and increase the abundance of different phosphorus fractions.
Primers corresponding to conserved bacterial repetitive of BOX elements were used to show that BOX-DNA sequences are widely distributed in phosphate solubilizing Pseudomonas strains. Phosphate solubilizing Pseudomonas was isolated from oil palm fields (tropical soil) in Malaysia. BOX elements were used to generate genomic fingerprints of a variety of Pseudomonas isolates to identify strains that were not distinguishable by other classification methods. BOX-PCR, that derived genomic fingerprints, was generated from whole purified genomic DNA by liquid culture of phosphate solubilizing Pseudomonas. BOX-PCR generated the phosphate solubilizing Pseudomonas specific fingerprints to identify the relationship between these strains. This suggests that distribution of BOX elements' sequences in phosphate solubilizing Pseudomonas strains is the mirror image of their genomic structure. Therefore, this method appears to be a rapid, simple, and reproducible method to identify and classify phosphate solubilizing Pseudomonas strains and it may be useful tool for fast identification of potential biofertilizer strains.
Use of phosphate-solubilizing bacteria (PSB) as inoculants has concurrently increased phosphorous uptake in plants and improved yields in several crop species. The ability of PSB to improve growth of aerobic rice (Oryza sativa L.) through enhanced phosphorus (P) uptake from Christmas island rock phosphate (RP) was studied in glasshouse experiments. Two isolated PSB strains; Bacillus spp. PSB9 and PSB16, were evaluated with RP treatments at 0, 30 and 60 kg ha(-1). Surface sterilized seeds of aerobic rice were planted in plastic pots containing 3 kg soil and the effect of treatments incorporated at planting were observed over 60 days of growth. The isolated PSB strains (PSB9 and PSB16) solubilized significantly high amounts of P (20.05-24.08 mg kg(-1)) compared to non-inoculated (19-23.10 mg kg(-1)) treatments. Significantly higher P solubilization (24.08 mg kg(-1)) and plant P uptake (5.31 mg plant(-1)) was observed with the PSB16 strain at the highest P level of 60 kg ha(-1). The higher amounts of soluble P in the soil solution increased P uptake in plants and resulted in higher plant biomass (21.48 g plant(-1)). PSB strains also increased plant height (80 cm) and improved root morphology in aerobic rice. The results showed that inoculation of aerobic rice with PSB improved phosphate solubilizing activity of incorporated RP.
Extensive use of metals in various industrial applications has caused substantial environmental pollution. Molybdenum-reducing bacteria isolated from soils can be used to remove molybdenum from contaminated environments. In this work we have isolated a local bacterium with the capability to reduce soluble molybdate to the insoluble molybdenum blue. We studied several factors that would optimize molybdate reduction. Electron donor sources such as glucose, sucrose, lactose, maltose and fructose (in decreasing efficiency) supported molybdate reduction after 24 h of incubation with optimum glucose concentration for molybdate reduction at 1.5% (w/v). The optimum pH, phosphate and molybdate concentrations, and temperature for molybdate reduction were pH 6.5, 5.0, 25 to 50 mM and 37 degrees C, respectively. The Mo-blue produced by cellular reduction exhibited a unique absorption spectrum with a maximum peak at 865 nm and a shoulder at 700 nm. Metal ions such as chromium, cadmium, copper, silver and mercury caused approximately 73, 71, 81, 77 and 78% inhibition of the molybdenum-reducing activity, respectively. All of the respiratory inhibitors tested namely rotenone, azide, cyanide and antimycin A did not show any inhibition to the molybdenum-reducing activity suggesting components of the electron transport system are not responsible for the reducing activity. The isolate was tentatively identified as Enterobacter sp. strain Dr.Y13 based on carbon utilization profiles using Biolog GN plates and partial 16S rDNA molecular phylogeny.
The need to isolate efficient heavy metal reducers for cost effective bioremediation strategy have resulted in the isolation of a potent molybdenum-reducing bacterium. The isolate was tentatively identified as Serratia sp. strain DRY5 based on the Biolog GN carbon utilization profiles and partial 16S rDNA molecular phylogeny. Strain DRY5 produced 2.3 times the amount of Mo-blue than S. marcescens strain Dr.Y6, 23 times more than E. coli K12 and 7 times more than E. cloacae strain 48. Strain DRY5 required 37 degrees C and pH 7.0 for optimum molybdenum reduction. Carbon sources such as sucrose, maltose, glucose and glycerol, supported cellular growth and molybdate reduction after 24 hr of static incubation. The most optimum carbon source that supported reduction was sucrose at 1.0% (w/v). Ammonium sulphate, ammonium chloride, glutamic acid, cysteine, and valine supported growth and molybdate reduction with ammonium sulphate as the optimum nitrogen source at 0. 2% (w/v). Molybdate reduction was optimally supported by 30 mM molybdate. The optimum concentration of phosphate for molybdate reduction was 5 mM when molybdate concentration was fixed at 30 mM and molybdate reduction was totally inhibited at 100 mM phosphate. Mo-blue produced by this strain shows a unique characteristic absorption profile with a maximum peak at 865 nm and a shoulder at 700 nm, Dialysis tubing experiment showed that 95.42% of Mo-blue was found in the dialysis tubing suggesting that the molybdate reduction seen in this bacterium was catalyzed by enzyme(s). The characteristics of isolate DRY5 suggest that it would be useful in the bioremediation ofmolybdenum-containing waste.
Black pepper production in Malaysia was restricted by various diseases. Hazardous chemical products appear to be the best solution to control diseases in black pepper cultivation. However, persistence of chemical residues in peppercorns could affect the quality of exports and consumptions. Application of fertilizers is crucial to sustain pepper growth and high yield. But, continuous use of chemical fertilizers could affect the soil ecosystem and eventually restrict nutrient uptake by pepper roots. Therefore, we propose biological approaches as an alternative solution instead of chemical products to sustain pepper cultivation in Malaysia. In this study, we have isolated a total of seven indigenous rhizobacteria antagonistic to soil-borne Fusarium solani, the causal fungus of slow decline, the most serious debilitating disease of black pepper in Malaysia. The isolated bacteria were identified as Bacillus subtilis, Bacillus siamensis, Brevibacillus gelatini, Pseudomonas geniculata, Pseudomonas beteli, Burkholderia ubonensis and Burkholderia territorii. These bacteria were effective in production of antifungal siderophore with the amount of 53.4 %-73.5 % per 0.5 mL of cell-free supernatants. The bacteria also produced appreciable amount of chitinase with chitinolytic index was ranged from 1.19 to 1.76. The bacteria have shown phosphate solubilizing index within 1.61 to 2.01. They were also efficient in ACC deaminase (0.52 mM-0.62 mM) and ammonia (60.3 mM-75.3 mM) production. The isolated antagonists were efficacious in stimulation of black pepper plant growth and root development through IAA (10.5 μg/mL-42.6 μg/mL) secretion. In conclusion, the isolated rhizobacteria are potent to be developed not only as biocontrol agents to minimize the utilization of hazardous chemicals in black pepper disease management, but also developed as bio-fertilizers to improve black pepper plant growth due to their capabilities in plant growth-promotion.
Phosphate solubilizing bacteria (PSB) can convert insoluble form of phosphorous to an available form. Applications of PSB as inoculants increase the phosphorus uptake by plant in the field. In this study, isolation and precise identification of PSB were carried out in Malaysian (Serdang) oil palm field (University Putra Malaysia). Identification and phylogenetic analysis of 8 better isolates were carried out by 16S rRNA gene sequencing in which as a result five isolates belong to the Beta subdivision of Proteobacteria, one isolate was related to the Gama subdivision of Proteobacteria, and two isolates were related to the Firmicutes. Bacterial isolates of 6upmr, 2upmr, 19upmnr, 10upmr, and 24upmr were identified as Alcaligenes faecalis. Also, bacterial isolates of 20upmnr and 17upmnr were identified as Bacillus cereus and Vagococcus carniphilus, respectively, and bacterial isolates of 31upmr were identified as Serratia plymuthica. Molecular identification and characterization of oil palm strains as the specific phosphate solubilizer can reduce the time and cost of producing effective inoculate (biofertilizer) in an oil palm field.
Phosphorus is one of the most abundant minerals in the human body. It is essential for almost all biochemical activities through ATP formation, intracellular signal transduction, cell membrane formation, bone mineralization, DNA and RNA synthesis, and inflammation modulation through various inflammatory cytokines. Phosphorus levels must be optimally regulated, as any deviations may lead to substantial derangements in glucose homeostasis. Clinical studies have reported that hyperphosphatemia can increase an individual's risk of developing metabolic syndrome. High phosphate burden has been shown to impair glucose metabolism by impairing pancreatic insulin secretion and increasing the risk of cardiometabolic disorders. Phosphate toxicity deserves more attention as metabolic syndrome is being seen more frequently worldwide and should be investigated further to determine the underlying mechanism of how phosphate burden may increase the cardiometabolic risk in the general population.
In this work we report on the isolation of a local molybdenum-reducing bacterium. The bacterium reduced molybdate or Mo(6+) to molybdenum blue (oxidation states between 5+ to 6+). Electron donors that supported cellular growth were sucrose, maltose, mannitol, fructose, glucose and starch (in decreasing order) with sucrose supporting formation of the highest amount of molybdenum blue at 10 g/l after 24 hours of static incubation. The optimum molybdate and phosphate concentrations that supported molybdate reduction were 20 and 5 mM, respectively. Molybdate reduction was optimal at 37 degrees C. The molybdenum blue produced from cellular reduction exhibited a unique absorption spectrum with a maximum peak at 865 nm and a shoulder at 700 nm. The isolate was tentatively identified as S. marcescens strain Dr.Y9 based on carbon utilization profiles using Biolog GN plates and partial 16S rDNA molecular phylogeny. No inhibition of molybdenum-reducing activity was seen using electron transport system (ETS) inhibitors such as antimycin A, 1HQNO (Hydroxyquinoline-N-Oxide), sodium azide and cyanide suggesting that the ETS of this bacterium is not the site of molybdate reduction.
The cytosolic mevalonate (MVA) pathway in Hevea brasiliensis latex is the conventionally accepted pathway which provides isopentenyl diphosphate (IPP) for cis-polyisoprene (rubber) biosynthesis. However, the plastidic 2-C-methyl-D-erythritol 4-phosphate (MEP) pathway may be an alternative source of IPP since its more recent discovery in plants. Quantitative RT-PCR (qRT-PCR) expression profiles of genes from both pathways in latex showed that subcellular compartmentalization of IPP for cis-polyisoprene synthesis is related to the degree of plastidic carotenoid synthesis. From this, the occurrence of two schemes of IPP partitioning and utilization within one species is proposed whereby the supply of IPP for cis-polyisoprene from the MEP pathway is related to carotenoid production in latex. Subsequently, a set of latex unique gene transcripts was sequenced and assembled and they were then mapped to IPP-requiring pathways. Up to eight such pathways, including cis-polyisoprene biosynthesis, were identified. Our findings on pre- and post-IPP metabolic routes form an important aspect of a pathway knowledge-driven approach to enhancing cis-polyisoprene biosynthesis in transgenic rubber trees.
Linalool and nerolidol are terpene alcohols that occur naturally in many aromatic plants and are commonly used in food and cosmetic industries as flavors and fragrances. In plants, linalool and nerolidol are biosynthesized as a result of respective linalool synthase and nerolidol synthase, or a single linalool/nerolidol synthase. In our previous work, we have isolated a linalool/nerolidol synthase (designated as PamTps1) from a local herbal plant, Plectranthus amboinicus, and successfully demonstrated the production of linalool and nerolidol in an Escherichia coli system. In this work, the biochemical properties of PamTps1 were analyzed, and its 3D homology model with the docking positions of its substrates, geranyl pyrophosphate (C10) and farnesyl pyrophosphate (C15) in the active site were constructed. PamTps1 exhibited the highest enzymatic activity at an optimal pH and temperature of 6.5 and 30 °C, respectively, and in the presence of 20 mM magnesium as a cofactor. The Michaelis-Menten constant (Km) and catalytic efficiency (kcat/Km) values of 16.72 ± 1.32 µM and 9.57 × 10-3 µM-1 s-1, respectively, showed that PamTps1 had a higher binding affinity and specificity for GPP instead of FPP as expected for a monoterpene synthase. The PamTps1 exhibits feature of a class I terpene synthase fold that made up of α-helices architecture with N-terminal domain and catalytic C-terminal domain. Nine aromatic residues (W268, Y272, Y299, F371, Y378, Y379, F447, Y517 and Y523) outlined the hydrophobic walls of the active site cavity, whilst residues from the RRx8W motif, RxR motif, H-α1 and J-K loops formed the active site lid that shielded the highly reactive carbocationic intermediates from the solvents. The dual substrates use by PamTps1 was hypothesized to be possible due to the architecture and residues lining the catalytic site that can accommodate larger substrate (FPP) as demonstrated by the protein modelling and docking analysis. This model serves as a first glimpse into the structural insights of the PamTps1 catalytic active site as a multi-substrate linalool/nerolidol synthase.
Growth and productivity of rice are negatively affected by soil salinity. However, some salt-tolerant rhizosphere-inhabiting bacteria can improve salt resistance of plants, thereby augmenting plant growth and production. Here, we isolated a total of 53 plant-growth-promoting rhizobacteria (PGPR) from saline and non-saline areas in Bangladesh where electrical conductivity was measured as >7.45 and <1.80 dS/m, respectively. Bacteria isolated from saline areas were able to grow in a salt concentration of up to 2.60 mol/L, contrary to the isolates collected from non-saline areas that did not survive beyond 854 mmol/L. Among the salt-tolerant isolates, Bacillus aryabhattai, Achromobacter denitrificans, and Ochrobactrum intermedium, identified by comparing respective sequences of 16S rRNA using the NCBI GenBank, exhibited a higher amount of atmospheric nitrogen fixation, phosphate solubilization, and indoleacetic acid production at 200 mmol/L salt stress. Salt-tolerant isolates exhibited greater resistance to heavy metals and antibiotics, which could be due to the production of an exopolysaccharide layer outside the cell surface. Oryza sativa L. fertilized with B. aryabhattai MS3 and grown under 200 mmol/L salt stress was found to be favoured by enhanced expression of a set of at least four salt-responsive plant genes: BZ8, SOS1, GIG, and NHX1. Fertilization of rice with osmoprotectant-producing PGPR, therefore, could be a climate-change-preparedness strategy for coastal agriculture.
The present study aims to investigate the influence of Staphylococcus aureus, Escherichia coli and Enterococcus faecalis in public market wastewater on the removal of nutrients in terms of ammonium (NH4-) and orthophosphate (PO43) using Scenedesmus sp. The removal rates of NH4- and orthophosphate PO43- and batch kinetic coefficient of Scenedesmus sp. were investigated. The phycoremediation process was carried out at ambient temperature for 6 days. The results revealed that the pathogenic bacteria exhibited survival potential in the presence of microalgae but they were reduced by 3-4 log at the end of the treatment process. The specific removal rates of NH4- and PO43- have a strong relationship with initial concentration in the public market wastewater (R2 = 0.86 and 0.80, respectively). The kinetic coefficient of NH4- removal by Scenedesmus sp. was determined as k = 4.28 mg NH4- 1 log10 cell mL-1 d-1 and km = 52.01 mg L-1 (R2 = 0.94) while the coefficient of PO43- removal was noted as k = 1.09 mg NH4- 1 log10 cell mL-1 d-1 and km = 85.56 mg L-1 (R2 = 0.92). It can be concluded that Scenedesmus sp. has high competition from indigenous bacteria in the public market wastewater to remove nutrients, with a higher coefficient of removal of NH4- than PO43.
The effect of temperature on the efficiency of organics and nutrients removal during the cultivation of aerobic granular sludge (AGS) in biological treatment of synthetic wastewater was studied. With this aim, three 3 L sequencing batch reactors (SBRs) with influent loading rate of 1.6 COD g (L d)(-1) were operated at different high temperatures (30, 40 and 50 °C) for simultaneous COD, phosphate and ammonia removal at a complete cycle time of 3 h. The systems were successfully started up and progressed to steady state at different cultivation periods. The statistical comparison of COD, phosphate and ammonia for effluent from the three SBRs revealed that there was a significant difference between groups of all the working temperatures of the bioreactors. The AGS cultivated at different high temperatures also positively correlated with the accumulation of elements including carbon, oxygen, phosphorus, silicon, iron, aluminium, calcium and magnesium that played important roles in the granulation process.
Bacteria with the ability to tolerate, remove, and/or degrade several xenobiotics simultaneously are urgently needed for remediation of polluted sites. A previously isolated bacterium with sodium dodecyl sulfate- (SDS-) degrading capacity was found to be able to reduce molybdenum to the nontoxic molybdenum blue. The optimal pH, carbon source, molybdate concentration, and temperature supporting molybdate reduction were pH 7.0, glucose at 1.5% (w/v), between 25 and 30 mM, and 25°C, respectively. The optimum phosphate concentration for molybdate reduction was 5 mM. The Mo-blue produced exhibits an absorption spectrum with a maximum peak at 865 nm and a shoulder at 700 nm. None of the respiratory inhibitors tested showed any inhibition to the molybdenum-reducing activity suggesting that the electron transport system of this bacterium is not the site of molybdenum reduction. Chromium, cadmium, silver, copper, mercury, and lead caused approximately 77, 65, 77, 89, 80, and 80% inhibition of the molybdenum-reducing activity, respectively. Ferrous and stannous ions markedly increased the activity of molybdenum-reducing activity in this bacterium. The maximum tolerable concentration of SDS as a cocontaminant was 3 g/L. The characteristics of this bacterium make it a suitable candidate for molybdenum bioremediation of sites cocontaminated with detergent pollutant.
The wide use of whole-genome sequencing approach in the modern genomic era has opened a great opportunity to reveal the prospective applications of halophilic bacteria. Robertkochia marina CC-AMO-30DT is one of the halophilic bacteria that was previously taxonomically identified without any inspection on its biotechnological potential from a genomic aspect. In this study, we present the whole-genome sequence of R. marina and demonstrated the ability of this bacterium in solubilizing phosphate by producing phosphatase. The genome of R. marina has 3.57 Mbp and contains 3107 predicted genes, from which 3044 are protein coding, 52 are non-coding RNAs, and 11 are pseudogenes. Several phosphatases such as alkaline phosphatases and pyrophosphatases were mined from the genome. Further genomic study (phylogenetics, sequence analysis, and functional mechanism) and experimental data suggested that the alkaline phosphatase produced by R. marina could potentially be utilized in promoting plant growth, particularly for plants on saline-based agricultural land.