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Alcaligenaceae and Chromatiaceae were previously reported as the specific pollution bioindicators in the receiving river water contaminated by palm oil mill effluent (POME) final discharge. Considering the inevitable sensitivity of bacteria under environmental stresses, it is crucial to assess the survivability of both bacteria in the fluctuated environmental factors, proving their credibility as POME pollution bioindicators in the environment. In this study, the survivability of Alcaligenaceae and Chromatiaceae from facultative pond, algae (aerobic) pond and final discharge were evaluated under varying sets of temperature (25-40°C), pH (pH 7-9) and low/high total suspended solid (TSS) contents of POME collected during low/high crop seasons of oil palm, respectively. Following treatment, the viability status and compositions of the bacterial community were assessed using flow cytometry-based assay and high-throughput Illumina MiSeq, respectively, in correlation with the changes of physicochemical properties. The changes in temperature, pH and TSS indeed changed the physicochemical properties of POME. The functionality of bacterial cells was also shifted where the viable cells and high nucleic acid contents reduced at elevated levels of temperature and pH but increased at high TSS content. Interestingly, the Alcaligenaceae and Chromatiaceae continuously detected in the samples which accounted for more than 0.5% of relative abundance, with a positive correlation with biological oxygen demand (BOD5) concentration. Therefore, either Alcaligenaceae or Chromatiaceae or both could be regarded as the reliable and specific bacterial indicators to indicate the pollution in river water due to POME final discharge despite the fluctuations in temperature, pH and TSS.

Excessive fertilizer applications in oil palm plantations are conventionally done to increase the oil yield, but they result in high production cost and environmental pollution. There have been only separate reports on the effects of fertilizer application on soil physical, chemical characteristics, and microbial biodiversity. Therefore, this study was conducted to determine the correlation between soil characteristics and soil microbial biodiversity in oil palm plantation after long-term frequent chemical fertilizer application compared with secondary soil, using molecular methods of polymerase chain reaction denaturing gradient gel electrophoresis (PCR-DGGE) and MiSeq. Secondary forest soil was chosen as the control. The results showed that after 25 years of fertilizer application, the total nitrogen and organic carbon contents decreased from low to very low scale, indicating soil infertility condition. Reduction of Firmicutes was related to suppression of soil borne diseases, and Bacteroidetes which is an indicator of soil health were both almost eliminated after 25 years of fertilizer application. In conclusion, long-term inorganic fertilizer application reduced the soil nitrogen, and organic carbon, altered beneficial microbes in the soil.

Biochar has proven to be a feasible additive for mitigating nitrogen loss during the composting process. This study aims to evaluate the influence of biochar addition on bacterial community and physicochemical properties changes, including ammonium (NH4+), nitrite (NO2−) and nitrate (NO3−) contents during the composting of poultry manure. The composting was carried out by adding 20% (w/w) of biochar into the mixture of poultry manure and rice straw with a ratio of 2:1, and the same treatment without biochar was prepared as a control. The finished product of control compost recorded the high contents of NO2− and NO3− (366 mg/kg and 600 mg/kg) with reduced the total NH4+ content to 10 mg/kg. Meanwhile, biochar compost recorded a higher amount of total NH4+ content (110 mg/kg) with low NO2− and NO3− (161 mg/kg and 137 mg/kg) content in the final composting material. The principal component analysis showed that the dynamics of dominant genera related to Halomonas, Pusillimonas, and Pseudofulvimonas, all of which were known as nitrifying and denitrifying bacteria, was significantly correlated with the dynamic of NO2− and NO3− content throughout the composting process. The genera related to Pusillimonas, and Pseudofulvimonas appeared as the dominant communities as the NO2− and NO3− increased. In contrast, as the NO2− and NO3− concentration decreased, the Halomonas genus were notably enriched in biochar compost. This study revealed the bacterial community shifts corresponded with the change of physicochemical properties, which provides essential information for a better understanding of monitoring and improving the composting process.

Zero-emission technology for palm oil mill effluent (POME) has led to a breakthrough in the palm oil industry in relation to the goal of sustainable development. However, there are limited resources on how this technology has affected the bacterial community in the receiving river that has previously been polluted with POME final discharge. Thus, the current study assessed the recoverability of the unexplored bacterial community in the receiving water of a constructed river water system post-zero emission of POME final discharge. An artificial river water system was constructed in this study, where the viability status and the composition of the bacterial community were assessed for 15 days using a flow cytometry-based assay and high-throughput sequencing by Illumina MiSeq, respectively. The zero-emission of POME final discharge reduced not only the physicochemical properties and nutrient contents of the receiving water, but also the bacterial cells’ viability from 40.3% to 24.5% and shifted the high nucleic acid (HNA) to low nucleic acid (LNA) content (38.7% to 34.5%). The proposed POME bacterial indicators, Alcaligenaceae and Chromatiaceae were not detectable in the rainwater (control) but were detected in the artificial river water system after the introduction of POME final discharge at the compositions of 1.0–1.3% and 2.2–5.1%, respectively. The implementation of a zero-emission system decreased the composition of Chromatiaceae from 2.2% on day 8 until it was undetectable on day 15, while Alcaligenaceae was continuously reduced from 1.2% to 0.9% within that similar time frame. As indicated by principal coordinate (PCO) analysis, the reductions in biological oxygen demand (BOD5) would further diminish the compositions of these bioindicators. The zero-emission of POME final discharge has demonstrated its efficacy, not only in reducing the polluting properties, but also in the bacterial biodiversity rebound in the affected water system.

Escherichia coli has approximately 4300 open reading frames and about 178 of them are annotated as pseudogenes. The existence of the pseudogenes in E. coli had raised the question of whether they contribute to any cell function. Recently, several pseudogenes have been studied; however, these genes are still considered rare and tend to be ignored as so-called junk. Our recent study has shown several pseudogenes in E. coli are essential in hydrogen metabolism. ydfW is one of the pseudogenes associated with hydrogen metabolism. Here, we found carbon dioxide and nitrate in the fermentation medium of mutant ydfW suggesting that this strain may induce nitrate pathways. The Fdh-N complex is comprised of FdnG, FdnH, and FdnI subunits which correspond to the nitrate-nitrite pathway. This finding is supported by transcription analysis that shows formate dehydrogenase (fdnG) and nitrate reductase (narG) are significantly induced (25- and 18-fold, respectively). FdnG and NarG play a significant role in the reduction of nitrate into nitrite; hence, their genes are induced by the build-up of nitrate. Based on in silico docking analysis, we predict that YdfW is covalently bound to the Fdh-N complex in the cytoplasm. Hence, deletion of ydfW is expected has influenced the nitrate-nitrite pathway. Subsequently, the situation will affect the hydrogen production in E. coli.

The main aim of our study was to explore the genome sequence of the inclusion body hepatitis associated Fowl adenovirus serotype 8b (FAdV-8b) UPM04217 and to study its genomic organisation. The nucleotide sequence of the whole genome of FAdV-8b UPM04217 was determined by using the 454 Pyrosequencing platform and the Sanger sequencing method. The complete genome was found to be 44,059 bp long with 57.9% G + C content and shared 97.5% genome identity with the reference FAdV-E genome (HG isolate). Interestingly, the genome analysis using ORF Finder, Glimmer3 and FGENESV predicted a total of 39 open reading frames (ORFs) compared to the FAdV-E HG that possessed 46 ORFs. Fourteen ORFs located within the central genomic region and 16 ORFs located within the left and right ends of the genome were assigned as being the high protein-coding regions. The fusion of the small ORFs at the right end terminal specifically in ORF22 and ORF33 could be the result of gene truncation in the FAdV-E HG. The frame shift mutation in ORF25 and other mutations in ORF13 and ORF17 might have lead to the emergence of genes that could have different functions. Besides, one of the minor capsid components, pVI, in FAdV-8b UPM04217 shared the highest similarity of 93% with that of FAdV-D, while only 47% similarity was found with FAdV-E. From the gene arrangement layout of the FAdV genome, FAdV-8b UPM04217 showed intermediate evolution between the FAdV-E HG and the FAdV-D although it was apparently more similar to the FAdV-E HG. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (10.1007/s13337-019-00530-9) contains supplementary material, which is available to authorized users.

Chitosan nanofiber has a highly uniform structure of 20–50 nm in diameter and shows high dispersibility in water due to its submicron size and high surface-to-volume ratio. The stacked nanofibers film is useful for breathability because it has a gap with a size of several tens of nm or more. However, the chemical bonds between the nanofibers cannot be broken during use. In this study, the thin films were obtained by filtration of chitosan nanofibers and 3-glycidoxypropyltrimethoxysilane (GPTMS) mixture. The addition of GPTMS changed the wettability, mechanical property and stability in water of the thin films. Bacitracin zinc salt (BZ) has been used for the localized dermatological medicines and loaded in the films. BZ interacted electrostatically with the thin films matrix and the release of BZ was controlled by the amount of GPTMS. A higher released amount of BZ showed higher antibacterial effects toward S. aureus. The film was also tested their toxicity by L929 fibroblasts. The release of less than 11.9 μg of BZ showed antibacterial effects, but were not toxic for fibroblast cells.

A novel deep-branching archaeal lineage was discovered at high-temperature hot springs around Oyasukyo Gorge in Akita Prefecture, Japan. Actively boiling hot spring water contained >1×104 microbes mL–1. The microbial community composition assessed by analyzing 16S rRNA gene amplicons revealed that the dominant bacterial phyla were Proteobacteria and Aquificae (>50% of the microbial composition) in samples collected in 2016 and 2019, respectively. Approximately 10% of the reads obtained in both years were not assigned to any taxonomy. The more detailed phylogenetic positions of the unassigned sequences identified using a clone library and phylogenetic tree showed that they formed a clade that was independent, distantly related to known phyla, and had low similarity (

The role of bacterial interaction is vital to control bacterial functions; however, it has not been fully understood in microbial consortia (including anaerobic digestion). In this study, fluorouracil (FU), which is an anticancer agent and a quorum sensing (QS) inhibitor to some of the Gram-negative bacteria was found to inhibit methane production from sewage sludge under anaerobic conditions, as shown in a result where methane production in the presence of FU was eight times lower than the control (sewage sludge without FU). Whereas FU did not influence the hydrolysis process, in the acidogenesis/acetogenesis process, butyrate, and acetate accumulated in samples with FU. Also, in the methanogenesis process, FU remarkably inhibited methane production by acetoclastic methanogens rather than by the hydrogenotrophic ones. This result agreed with the result that growth and methane production of Methanosarcina acetivorans C2A was inhibited in the presence of FU. However, the inhibitory effect of FU was high in the condition that both bacteria and archaea were active. It indicates that FU influences methanogens and bacteria in the process of methane fermentation. The analyses of microbial communities (bacteria and archaea) showed that the abundance ratio of the Firmicutes phyla is high, and hydrogenotrophic methanogens become dominant in the presence of FU. Conversely, the abundance of Spirochaetes significantly decreased under FU. The inhibition of methane production by FU was due to the growth inhibition of methanogenic archaea and the changes in the composition of the bacterial population.

Despite efforts to address the composition of the microbial community during the anaerobic treatment of palm oil mill effluent (POME), its composition in relation to biodegradation in the full-scale treatment system has not yet been extensively examined. Therefore, a thorough analysis of bacterial and archaeal communities was performed in the present study using MiSeq sequencing at the different stages of the POME treatment, which comprised anaerobic as well as facultative anaerobic and aerobic processes, including the mixed raw effluent (MRE), mixing pond, holding tank, and final discharge phases. Based on the results obtained, the following biodegradation processes were suggested to occur at the different treatment stages: (1) Lactobacillaceae (35.9%) dominated the first stage, which contributed to high lactic acid production; (2) the higher population of Clostridiaceae in the mixing pond (47.7%) and Prevotellaceae in the holding tank (49.7%) promoted acetic acid production; (3) the aceticlastic methanogen Methanosaetaceae (0.6-0.8%) played a role in acetic acid degradation in the open digester and closed reactor for methane generation; (4) Syntrophomonas (21.5-29.2%) appeared to be involved in the degradation of fatty acids and acetic acid by syntrophic cooperation with the hydrogenotrophic methanogen, Methanobacteriaceae (0.6-1.3%); and (5) the phenols and alcohols detected in the early phases, but not in the final discharge phase, indicated the successful degradation of lignocellulosic materials. The present results contribute to a better understanding of the biodegradation mechanisms involved in the different stages of the full-scale treatment of POME.

This study was undertaken to confirm the reliability of the proposed potential bioindicators, Alcaligenaceae and Chromatiaceae to specifically indicate contamination in river water by palm oil mill effluent (POME) final discharge. The use of bioindicators could assist in determining the specific cause of contamination in water bodies. Alcaligenaceae and Chromatiaceae were shown by a previous study to be present in river water contaminated with the final discharge from the treatment of POME. In the present study, the compositions of the bacterial communities in the POME final discharge obtained from four different palm oil mills which used different biotreatments of POME were elucidated using high-throughput MiSeq. The four POME final discharges studied showed different species richness and evenness among them. However, the bacterial community compositions in the different final discharges exhibited almost similar patterns in that the phylum Proteobacteria was dominant in all the samples. The proposed bioindicators, the Alcaligenaceae and Chromatiaceae families, were found to be present in all the four final discharges despite the different characteristics of the mills and the different biotreatment processes used by them. These bioindicators were also strongly and positively correlated with the biochemical oxygen demand (BOD5) concentration. This makes them reliable bacterial indicators to detect the presence of POME final discharge in river water.

A thorough outlook on the effect of palm oil mill effluent (POME) final discharge towards bacterial community dynamics in the receiving river is provided in this study by using a high-throughput MiSeq. The shift of bacterial composition could be used to determine the potential bacterial indicators to indicate contamination caused by POME. This study showed that the POME final discharge did not only alter the natural physicochemical properties of the river water but also caused the reduction of bacterial diversity in the receiving river. The Chromatiaceae and Alcaligenaceae which were not detected in the upstream but were detected in the downstream part of the river are proposed as the indicator bacteria to indicate the river water contamination caused by POME final discharge. The emergence of either one or both bacteria in the downstream part of the river were shown to be carried over by the effluent. Therefore, an accurate pollution monitoring approach using bacterial indicator is expected to complement the conventional POME pollution assessment method which is currently dependent on the physicochemical properties of the final discharge. This is the first study that reported on the potential indicator bacteria for the assessment of river water contamination caused by POME final discharge.

The potential utilization of phospholine gum, a by-product of the palm oil industry was evaluated using waste sewage sludge (WSS) as a substrate as well as a microbial source to mimic methane production by enteric fermentation. Ruminant animals release enteric methane through their digestive process. The enteric methane is one of the greenhouse gases that can contribute to global warming and should be prevented. In this study, methane production was remarkably inhibited by adding phospholine gum to WSS, even at a low concentration. Phospholine gum reduced the activity of methanogens and Lactobacillus sp. and Megasphaera sp. which are known as important ruminal microorganisms were detected as bacterial species induced by the addition of phospholine gum to WSS. Also, the addition of phospholine gum triggered an increase in protein concentrations as well as protease activities and stimulated to produce protease and cellulase by which phospholine gum may be degraded. Furthermore, a significant amount of propionate was produced in the presence of phospholine gum. Thus, phospholine gum inhibits methane production without inhibiting the stages of hydrolysis and acidogenesis/acetogenesis. Finally, methane fermentation using the rumen derived from a goat was also inhibited by phospholine gum. Therefore, these results indicate that the phospholine gum has great potential to inhibit methane production as a feed additive for ruminant animals.

A thermophilic Thermobifida fusca strain UPMC 901, harboring highly thermostable cellulolytic activity, was successfully isolated from oil palm empty fruit bunch compost. Its endoglucanase had the highest activity at 24 hours of incubation in carboxymethyl-cellulose (CMC) and filter paper. A maximum endoglucanase activity of 0.9 U/mL was achieved at pH 5 and 60 °C using CMC as a carbon source. The endoglucanase properties were further characterized using crude enzyme preparations from the culture supernatant. Thermal stability indicated that the endoglucanase activity was highly stable at 70 °C for 24 hours. Furthermore, the activity was found to be completely maintained without any loss at 50 °C and 60 °C for 144 hours, making it the most stable than other endoglucanases reported in the literature. The high stability of the endoglucanase at an elevated temperature for a prolonged period of time makes it a suitable candidate for the biorefinery application.

This study aims to elucidate the mechanism of carbon partitioning towards starch and triacylglycerol (TAG) under nitrogen stress. Whole-transcriptome analysis was performed using RNA sequencing before being validated with qRT-PCR. The strain, Chlorella vulgaris UPSI-JRM01, exhibited a two-stage response to nitrogen stress: i) carbohydrate accumulation, plastid protein degradation and amino acid biosynthesis; and ii) lipid accumulation, carbohydrate degradation and DNA damage. In the first stage, the genes responsible for plastid protein degradation (CD4A), amino acid biosynthesis (arg, GLSN, AHCY, win1), carbon fixation (RuBisCO, GapC, fba, TK) and starch synthesis (SS, SBE2) were upregulated. In the second stage, the upregulation of genes responsible for starch degradation (SP), glycolysis (PFO, PK, ACS), fatty acid biosynthesis (FabH, accD) and TAG accumulation (DGAT2) were observed. By contrast, the components of photosystem I (PsbD, PsbC, PsbB), photosystem II (PsaA), cytochrome b6/f (PetA, PetG) and F-type ATPase (beta, alpha) were downregulated. The results suggested that nitrogen stress triggered high starch accumulation before the carbon was partitioned towards triacylglycerol (TAG) for long-term energy storage by two different pathways: chloroplastic TAG synthesis and glycerolipid metabolism. The starch build-up functions served as a rapid response to nitrogen stress, whereas subsequent lipid accumulation benefited the long-term storage of energy. This research provides in-depth understanding of the metabolic changes triggered by nitrogen stress.

Palm oil mill effluent (POME) contains a substantial amount of nutrients and a significant amount of indigenous microorganisms that consume the nutrients for their growth. In this study, we demonstrated that cultivation of microalgae in POME was able to reduce its carbon and nutrient contents. Three native microalgae isolates, namely, Coelastrella sp. UKM4, Chlamydomonas sp. UKM6 and Scenedesmus sp. UKM9, were cultured in sterilized and unsterilized anaerobic POME to evaluate the microalgae-bacteria interactions for phycoremediation of POME. The results showed higher COD removal in unsterilized POME than in sterilized POME due to microalgae-bacteria consortia interaction. Of the three microalgae isolates, the interaction of Scenedesmus sp. UKM9 with bacteria showed the highest COD and PO43− removal percentages. For NH4+ removal, all three isolates showed removal of more than 80 % in both sterilized and unsterilized POME. Quantitative real-time polymerase chain reaction analysis showed that the quantities of bacteria were constant after the treatment, while the quantities of microalgae changed. Metagenomics analysis showed that the community of bacteria shifted after microalgae cultivation in POME, with Actinobacteria, Bacteroidetes, Planctomycetes and Proteobacteria being the major phyla.

Microorganisms are progressively adopted as reliable and specific bioindicators of pollution, with Alcaligenaceae and Chromatiaceae being recently proposed to indicate contamination in river water due to palm oil mill effluent (POME) final discharge. This study was conducted to evaluate the reliability of these bacteria to act as specific bioindicators by assessing their detections in correlation with environmental factors in rivers polluted by POME final discharge when compared to by other pollution sources. By using the Illumina MiSeq high throughput sequencing platform, bacterial diversities and compositions were compared among unpolluted (upstream) river, polluted rivers due to POME final discharge and other rivers polluted by mining, chemical and automotive industries. To correlate between bacterial compositions and metabolisms in rivers subjected to different pollutants sources, the viability and the ratio of the high and low nucleic acids (HNA/LNA) bacterial cells were also compared by using a double staining assay based on flow cytometry. Interestingly, Alcaligenaceae and Chromatiaceae were found only in POME final discharge polluted rivers (>0.15%) but were not detected in rivers polluted by other pollutants. Higher bacterial cell viability was detected in rivers polluted by POME final discharge (86–91%) as compared to the other polluted rivers (15–80%). From the viable fractions of the bacterial cells, higher HNA cells (91–93%) were quantified in the POME final discharge receiving rivers but with only 5–18% in the other rivers. These trends indicated that the nutrients carried by the POME final discharge enhanced the growth of bacteria in the receiving river water that might include Alcaligenaceae and Chromatiaceae. These results suggested that either Alcaligenaceae or Chromatiaceae or both were unique in the POME final discharge polluted rivers but not in the others. They could function as specific and reliable bacterial bioindicators relevant for the establishment of a complementary monitoring system for POME pollutant discharge.

Quorum sensing (QS) plays a key role in activating bacterial functions through small molecules called autoinducers. In this study, the QS of Gram-negative bacteria in waste sewage sludge (WSS) was downregulated by adding the quorum quenching enzyme, AiiM lactonase, which cleaved the acyl-homoserine lactone (AHL) autoinducer signals from Gram-negative bacteria, and subsequently methane production was inhibited by over 400%. The pH was lowered after 2 days in the anaerobic fermentation whereas protease activity at the hydrolysis step was almost the same with or without AiiM. The production of acetic acid significantly increased during the fermentation in the presence of AiiM. The bacterial community at day 2 indicated that the population of Gram-positive bacteria increased in the presence of AiiM, and the percentage of Gram-negative bacteria decreased in the WSS containing AiiM. The change in the bacterial community in the presence of AiiM may be due to the different antimicrobial agents produced in the WSS because some of the Gram-positive bacteria were killed by adding the solid-phase extraction (SPE) fraction from the WSS without AiiM. In contrast, the SPE fraction with AiiM had reduced bactericidal activity against Gram-negative bacteria. Thus, bacterial signaling between Gram-negative bacteria is critical for methane production by the microbial consortia.

Efficient approaches for the utilization of waste sewage sludge have been widely studied. One of them is to use it for the bioenergy production, specifically methane gas which is well-known to be driven by complex bacterial interactions during the anaerobic digestion process. Therefore, it is important to understand not only microorganisms for producing methane but also those for controlling or regulating the process. In this study, azithromycin analogs belonging to macrolide, ketolide, and lincosamide groups were applied to investigate the mechanisms and dynamics of bacterial community in waste sewage sludge for methane production. The stages of anaerobic digestion process were evaluated by measuring the production of intermediate substrates, such as protease activity, organic acids, the quantification of bacteria and archaea, and its community dynamics. All azithromycin analogs used in this study achieved a high methane production compared to the control sample without any antibiotic due to the efficient hydrolysis process and the presence of important fermentative bacteria and archaea responsible in the methanogenesis stage. The key microorganisms contributing to the methane production may be Clostridia, Cladilinea, Planctomycetes, and Alphaproteobacteria as an accelerator whereas Nitrosomonadaceae and Nitrospiraceae may be suppressors for methane production. In conclusion, the utilization of antibiotic analogs of macrolide, ketolide, and lincosamide groups has a promising ability in finding the essential microorganisms and improving the methane production using waste sewage sludge.

Anaerobic digestion is an effective method for reducing the by-product of waste-activated sludge (WAS) from wastewater treatment plants and for producing bioenergy from WAS. However, only a limited number of studies have attempted to improve anaerobic digestion by targeting the microbial interactions in WAS. In this study, we examined whether different antibiotics positively, negatively, or neutrally influence methane fermentation by evaluating changes in the microbial community and functions in WAS. Addition of azithromycin promoted the microbial communities related to the acidogenic and acetogenic stages, and a high concentration of soluble proteins and a high activity of methanogens were detected. Chloramphenicol inhibited methane production but did not affect the bacteria that contribute to the hydrolysis, acidogenesis, and acetogenesis digestion stages. The addition of kanamycin, which exhibits the same methane productivity as a control (antibiotic-free WAS), did not affect all of the microbial communities during anaerobic digestion. This study demonstrates the simultaneous functions and interactions of diverse bacteria and methanogenic Archaea in different stages of the anaerobic digestion of WAS. The ratio of Caldilinea, Methanosarcina, and Clostridium may correspond closely to the trend of methane production in each antibiotic. The changes in microbial activities and function by antibiotics facilitate a better understanding of bioenergy production.

九州工業大学博士学位論文（要旨）学位記番号：生工博甲第275号 学位授与年月日：平成28年12月27日

1 Introduction and literature review||2 General materials and methods||3 Bacterial soldiers in waste activated sludge – understanding of the bacterial natural selection for the future successful bioremediation –||4 Impact of different antibiotics on methane production using waste-activated sludge: mechanisms and microbial community dynamics||5 Effect of macrolides and lincosamides on methane production using waste activated sludge: mechanisms and microbial community dynamics||6 Inhibition of a mimic enteric methane fermentation by a palm oil industrial waste, phospholine gum – understanding of microbial community dynamics –||7 Concluding remarks and suggestions for further research, Waste activated sludge （WAS） is a by-product that has been abundantly produced from wastewater treatment plant after secondary biological treatment. The disposal of WAS through landfill or incineration has created more vulnerable problems in term of cost and environmental issues. Many strategies have adapted bioremediation and bioenergy production in order to completely utilize this WAS for sustainable environment and biodiversity conservation. These processes are driven by a complex microorganisms in WAS. Therefore, the successful achievement of both strategies; bioremediation and bioenergy production can be clarified based on the microbial community profile through next-generation sequencing analysis, MiSeq. Bioremediation strategies utilize specific and powerful exogenous microorganisms to assist in the degradation of various harmful pollutants in WAS. However, this inoculant is unable to retain in WAS for a long period of time to carry out their specific functions in degradation. The interaction between this inoculant and indigenous microbes presence in WAS is suspected to be one of the main reasons. Therefore, through the microbial community analysis in this study, some indigenous bacteria have been identified as bacterial soldiers that are responsible in killing the inoculant bacteria. The enhanced green fluorescence protein （EGFP） expressing E. coli was used as the inoculant with the fluorescence marker and some antibiotic-resistant genes. The indigenous bacteria identified as bacterial soldiers were mainly from Comamonadaceae, Myxococcales and Sphingobacteriales communities as later proved by co-culture interaction with inoculant. Bioenergy production through methane fermentation is another approach that has been taken to utilize WAS. Many studies have been done in laboratory scale to find an efficient methane production approach including application of antibiotic, azithromycin. The effect of antibiotics on microbial diversity during anaerobic digestion stages is important to understand their mechanisms and functions in methane production. Therefore, in this study, different antibiotics that produced different methane profile were used to clarify the microbial interactions and regulatory systems in each stages of anaerobic digestion process. During anaerobic digestion, hydrolysis and fermentation stages were efficiently occurred due to the activation of hydrolytic and fermentative bacterial communities by all antibiotics used in this study. However, some antibiotics have shaped the unfavorable conditions for methanogens in methanogenesis stages. This unbalanced of microbial communities has affected on methane production. Thus, a balanced of microbial communities in all stages of anaerobic digestion is important for efficient methane production as shown by some antibiotics, including azithromycin. The importance of microbial communities was shown in bioremediation and methane fermentation processes. Another application using by-product taken from refining process of palm oil industry, phospholine gum was done for methane fermentation in WAS. It was found that phospholine gum has inhibited methane production but not affected other anaerobic digestion stages. Therefore, based on these phenomena, phospholine gum has potential to be applied as feed additive for ruminants in mitigating the enteric methane emission which contribute to greenhouse gases. In conclusion, various reflections and dynamics have been shown by microbial diversity due to the different effects and conditions in the biological processes. This has provided a better and clear understanding on their functions and mechanisms in ensuring a successful bioremediation process and bioenergy production as the excellent approaches for sustainable environment and biodiversity conservation., 九州工業大学博士学位論文 学位記番号：生工博甲第275号 学位授与年月日：平成28年12月27日, 平成28年度