Biohydrogen production by a microbial consortium isolated from local hot spring

Biohydrogen production from microorganism is a form of renewable energy that could supplement the depletion of fossil fuels. In producing biohydrogen, microbial consortia are more feasible than pure cultures because of its operational ease and stability and it is more favourable energetically at elevated temperatures which enables thermophiles to reach higher biohydrogen production than mesophiles. The aim of this study was to isolate, enrich and screen microbial consortium from local hot spring for its potential in producing biohydrogen, to optimize the selected consortium for optimal biohydrogen production and to identify the microbial diversity community of the consortium. Sampling was conducted at Gadek, Cherana Putih, Gersik and Selayang hot spring and the samples were enriched in Mineral Salt Succinate medium. The enriched consortia were screened for biohydrogen production using Gas Chromatography-Thermal Conductivity Detector (GC-TCD) and the biohydrogen production of the selected consortium was optimized by one factor at a time (OFAT) method. The kinetic analysis of the growth and biohydrogen production of the consortium were analyzed using the modified Logistic growth equation and modified Gompertz equation respectively. The microbial diversity community of the consortia were observed using 16S rRNA polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE). To determine the microbial population dynamics of the consortia, 16S rRNA clone library were constructed for the consortia before and after optimization and sequencing data were analyzed using Mothur. Microbial consortium from Gadek hot spring (GDC) yielded the highest biohydrogen production compared to other consortia. The optimized condition (15% (v/v) inoculum size, 50°C, pH 7, 2 g/L sodium pyruvate and 0.5 g/L tryptone) showed a maximal biomass growth of 0.563 g dry cell weight/L and apparent specific growth rate of 0.959 h-1. Whilst the optimized hydrogen production potential was 86.2 mmol H2/L culture with the maximal production rate of 4.117 mmol/L h-1, biohydrogen yield obtained was 135.7 mmol H2/g biomass and the lag phase time was 5.1 hours. DGGE showed a slight microbial shift between the consortia before and after optimization. From the 16S rRNA clone library, 21 clones were obtained and a total of four operational taxonomic unit (OTU) were detected. Both consortia showed Firmicutes and Proteobacteria as the predominant phyla which have phylogeny affiliations to hydrogen producers. However, OTU_4 (Sporoacetegenium mesophilum) was only present in the consortium before optimization, OTU_1 (Thauera sp), OTU_2 (Paenibacillus barengoltzii) and OTU_3 (Sporomusaceae g. sp) were present in both consortia. Analysis showed the presence of OTU_2 and OTU_3 and the abundance of OTU_1 in the optimized consortium led to an increased in biohydrogen production of about 8 fold more from the consortium before optimization. In conclusion, this is the first study that reports a unique combination of Thauera sp., Paenibacillus barengoltzii and Sporomusaceae g. sp. which are able to produce a high amount of biohydrogen at the optimized condition

Biohydrogen production by a microbial consortium isolated from local hot spring

Biohydrogen production from microorganism is a form of renewable energy that could supplement the depletion of fossil fuels. In producing biohydrogen, microbial consortia are more feasible than pure cultures because of its operational ease and stability and it is more favourable energetically at elevated temperatures which enables thermophiles to reach higher biohydrogen production than mesophiles. The aim of this study was to isolate, enrich and screen microbial consortium from local hot spring for its potential in producing biohydrogen, to optimize the selected consortium for optimal biohydrogen production and to identify the microbial diversity community of the consortium. Sampling was conducted at Gadek, Cherana Putih, Gersik and Selayang hot spring and the samples were enriched in Mineral Salt Succinate medium. The enriched consortia were screened for biohydrogen production using Gas Chromatography-Thermal Conductivity Detector (GC-TCD) and the biohydrogen production of the selected consortium was optimized by one factor at a time (OFAT) method. The kinetic analysis of the growth and biohydrogen production of the consortium were analyzed using the modified Logistic growth equation and modified Gompertz equation respectively. The microbial diversity community of the consortia were observed using 16S rRNA polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE). To determine the microbial population dynamics of the consortia, 16S rRNA clone library were constructed for the consortia before and after optimization and sequencing data were analyzed using Mothur. Microbial consortium from Gadek hot spring (GDC) yielded the highest biohydrogen production compared to other consortia. The optimized condition (15% (v/v) inoculum size, 50°C, pH 7, 2 g/L sodium pyruvate and 0.5 g/L tryptone) showed a maximal biomass growth of 0.563 g dry cell weight/L and apparent specific growth rate of 0.959 h-1. Whilst the optimized hydrogen production potential was 86.2 mmol H2/L culture with the maximal production rate of 4.117 mmol/L h-1, biohydrogen yield obtained was 135.7 mmol H2/g biomass and the lag phase time was 5.1 hours. DGGE showed a slight microbial shift between the consortia before and after optimization. From the 16S rRNA clone library, 21 clones were obtained and a total of four operational taxonomic unit (OTU) were detected. Both consortia showed Firmicutes and Proteobacteria as the predominant phyla which have phylogeny affiliations to hydrogen producers. However, OTU_4 (Sporoacetegenium mesophilum) was only present in the consortium before optimization, OTU_1 (Thauera sp), OTU_2 (Paenibacillus barengoltzii) and OTU_3 (Sporomusaceae g. sp) were present in both consortia. Analysis showed the presence of OTU_2 and OTU_3 and the abundance of OTU_1 in the optimized consortium led to an increased in biohydrogen production of about 8 fold more from the consortium before optimization. In conclusion, this is the first study that reports a unique combination of Thauera sp., Paenibacillus barengoltzii and Sporomusaceae g. sp. which are able to produce a high amount of biohydrogen at the optimized condition

Strep-tag ii mutant maltose-binding protein for reagentless fluorescence sensing

Maltose-binding protein (MBP) is a periplasmic binding protein found in Gram negative bacteria. MBP is involved in maltose transport and bacterial chemotaxis; it binds to maltose and maltodextrins comprising α(1-4)-glucosidically linked linear glucose polymers and α(1-4)-glucosidically linked cyclodextrins. Upon ligand binding, MBP changes its conformation from an open to a closed form. This molecular recognition-transducing a ligand-binding event into a physical one-renders MBP an ideal candidate for biosensor development. Here, we describe the construction of a Strep-tag II mutant MBP for reagentless fluorescence sensing. malE, which encodes MBP, was amplified. A cysteine residue was introduced by site-directed mutagenesis to ensure a single label attachment at a specific site with a thiol-specific fluorescent probe. An environmentally sensitive fluorophore (IANBD amide) was covalently attached to the introduced thiol group and analysed by fluorescence sensing. The tagged mutant MBP (D95C) was purified (molecular size, ∼42 kDa). The fluorescence measurements of the IANBD-labelled Strep-tag II-D95C in the solution phase showed an appreciable change in fluorescence intensity (dissociation constant, 7.6±1.75 μM). Our mutant MBP retains maltose-binding activity and is suitable for reagentless fluorescence sensin

Construction of a strep-tag II mutant maltose binding protein for reagentless fluorescence sensing

Maltose binding protein (MBP) changes its conformational structure upon its ligand binding.This molecular recognition element that transduces a ligand-binding event into a physical one make MBP an ideal candidate for reagentless fluorescence sensing. MBP gene, (malE) was amplified from a pMaL-C4x plasmid vector and was fused to a Strep-Tag II pET-51b(+) vector. Strep-Tag II is a tag that will enable the MBP to be unidirectionally immobilized on solid supports. A cysteine mutant of the MBP was constructed by inverse PCR and the recombinant protein fusion was then purified by affinity purification using Strep-Tactin resin. To sense maltose binding, an environmentally sensitive fluorophore (IANBD amide) was covalently attached to the introduced thiol group. The tagged mutant MBP (D95C) was successfully generated and the protein was successfully purified with the expected molecular size of ~42 kDa observed on the SDS PAGE. The fluorescence measurements of the IANBD labeled of tagged mutant MBP (Strep-Tag II D95C) in the solution phase, showed an appreciable change in fluorescence intensity with dissociation constant, (Kd) of 7.6 ± 1.75 µM. Nonetheless, it could retain its ligand binding activity towards maltose. However, immobilization of Strep-Tag II D95C on solid surface suffered some limitation with the Strep-Tactin coated microwell plates because it did not give any dependable results to support the ligand binding activity of the site directed immobilized protein. Thus, this engineered mutant MBP (Strep-Tag II fused D95C) could be potentially developed for biosensor application with further improvement in protein immobilization method

Health effects of herbicides and its current removal strategies

ABSTRACTThe continually expanding global population has necessitated increased food supply production. Thus, agricultural intensification has been required to keep up with food supply demand, resulting in a sharp rise in pesticide use. The pesticide aids in the prevention of potential losses caused by pests, plant pathogens, and weeds, but excessive use over time has accumulated its occurrence in the environment and subsequently rendered it one of the emerging contaminants of concern. This review highlights the sources and classification of herbicides and their fate in the environment, with a special focus on the effects on human health and methods to remove herbicides. The human health impacts discussion was in relation to toxic effects, cell disruption, carcinogenic impacts, negative fertility effects, and neurological impacts. The removal treatments described herein include physicochemical, biological, and chemical treatment approaches, and advanced oxidation processes (AOPs). Also, alternative, green, and sustainable treatment options were discussed to shed insight into effective treatment technologies for herbicides. To conclude, this review serves as a stepping stone to a better environment with herbicides