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Wen-Tso Liu

Publications and source records attributed to Wen-Tso Liu.

At least 19 recordsLinked to original sources
PubMed2006

Effects of target length on the hybridization efficiency and specificity of rRNA-based oligonucleotide microarrays.

The effect of target size on microarray hybridization efficiencies and specificity was investigated using a set of 166 oligonucleotide probes targeting the 16S rRNA gene of Escherichia coli. The targets included unfragmented native rRNA, fragmented rRNA ( approximately 20 to 100 bp), PCR amplicons (93 to 1,480 bp), and three synthetic single-stranded DNA oligonucleotides (45 to 56 bp). Fluorescence intensities of probes hybridized with targets were categorized into classes I (81 to 100% relative to the control probe), II (61 to 80%), III (41 to 60%), IV (21 to 40%), V (6 to 20%), and VI (0 to 5%). Good hybridization efficiency was defined for those probes conferring intensities in classes I to IV; those in classes V and VI were regarded as weak and false-negative signals, respectively. Using unfragmented native rRNA, 13.9% of the probes had fluorescence intensities in classes I to IV, whereas the majority (57.8%) exhibited false-negative signals. Similar trends were observed for the 1,480-bp PCR amplicon (6.6% of the probes were in classes I to IV). In contrast, after hybridization of fragmented rRNA, the percentage of probes in classes I to IV rose to 83.1%. Likewise, when DNA target sizes were reduced from 1,480 bp to 45 bp, this percentage increased approximately 14-fold. Overall, microarray hybridization efficiencies and specificity were improved with fragmented rRNA (20 to 100 bp), short PCR amplicons (<150 bp), and synthetic targets (45 to 56 bp). Such an understanding is important to the application of DNA microarray technology in microbial community studies.

DNA, Bacterial↗
PubMed2006

Miniaturized platforms for the detection of single-nucleotide polymorphisms.

Conventional methods for detecting single-nucleotide polymorphisms (SNPs), the most common form of genetic variation in human beings, are mostly limited by their analysis time and throughputs. In contrast, advances in microfabrication technology have led to the development of miniaturized platforms that can potentially provide rapid high-throughput analysis at small sample volumes. This review highlights some of the recent developments in the miniaturization of SNP detection platforms, including microarray-based, bead-based microfluidic and microelectrophoresis-based platforms. Particular attention is paid to their ease of fabrication, analysis time, and level of throughput.

Electrophoresis↗
PubMed2006

Technology development to explore the relationship between oral health and the oral microbial community.

The human oral cavity contains a complex microbial community that, until recently, has not been well characterized. Studies using molecular tools have begun to enumerate and quantify the species residing in various niches of the oral cavity; yet, virtually every study has revealed additional new species, and little is known about the structural dynamics of the oral microbial community or how it changes with disease. Current estimates of bacterial diversity in the oral cavity range up to 700 species, although in any single individual this number is much lower. Oral microbes are responsible for common chronic diseases and are suggested to be sentinels of systemic human diseases. Microarrays are now being used to study oral microbiota in a systematic and robust manner. Although this technology is still relatively young, improvements have been made in all aspects of the technology, including advances that provide better discrimination between perfect-match hybridizations from non-specific (and closely-related) hybridizations. This review addresses a core technology using gel-based microarrays and the initial integration of this technology into a single device needed for system-wide studies of complex microbial community structure and for the development of oral diagnostic devices.

Journal Article↗
PubMed2006

Biological filtration limits carbon availability and affects downstream biofilm formation and community structure.

Carbon removal strategies have gained popularity in the mitigation of biofouling in water reuse processes, but current biofilm-monitoring practices based on organic-carbon concentrations may not provide an accurate representation of the in situ biofilm problem. This study evaluated a submerged microtiter plate assay for direct and rapid monitoring of biofilm formation by subjecting the plates to a continuous flow of either secondary effluent (SE) or biofilter-treated secondary effluent (BF). This method was very robust, based on a high correlation (R(2) = 0.92) between the biomass (given by the A(600) in the microtiter plate assay) and the biovolume (determined from independent biofilms developed on glass slides under identical conditions) measurements, and revealed that the biomasses in BF biofilms were consistently lower than those in SE biofilms. The influence of the organic-carbon content on the biofilm community composition and succession was further evaluated using molecular tools. Terminal restriction fragment length polymorphism analysis of 16S rRNA genes revealed a group of pioneer colonizers, possibly represented by Sphingomonadaceae and Caulobacter organisms, to be common in both SE and BF biofilms. However, differences in organic-carbon availabilities in the two water samples eventually led to the selection of distinct biofilm communities. Alphaproteobacterial populations were confirmed by fluorescence in situ hybridization to be enriched in SE biofilms, while Betaproteobacteria were dominant in BF biofilms. Cloning analyses further demonstrated that microorganisms adapted for survival under low-substrate conditions (e.g., Aquabacterium, Caulobacter, and Legionella) were preferentially selected in the BF biofilm, suggesting that carbon limitation strategies may not achieve adequate biofouling control in the long run.

Bacteria↗
PubMed2006

Nanoparticles and their biological and environmental applications.

Nanoparticles exhibit unique physical properties (such as particle aggregation and photoemission, and electrical and heat conductivities) and chemical properties (such as catalytic activity), and hence have received much attention from scientists and researchers in different areas of biological sciences. In this review, we briefly summarize the major types of nanoparticle that have been used so far and discuss the possible applications of these nanoparticles in biological and environmental research, and the potential environmental and health impacts associated with the use of these nanoparticles.

Biosensing Techniques↗
PubMed2005

Microfluidic device as a new platform for immunofluorescent detection of viruses.

A bead-based microfluidic device was developed and demonstrated to achieve rapid and sensitive enzyme-linked immunosorbent assay (ELISA) with quantum dots as the labeling fluorophore for virus detection. In comparison to standard ELISA performed on the same virus, the minimal detectable concentration of the target virus was improved from 360 to 22 ng mL-1, the detection time was shortened from >3.25 h to <30 min, and the amount of antibody consumed was reduced by a factor of 14.3.

Animals↗
PubMed2005

Biofilm formation characteristics of bacterial isolates retrieved from a reverse osmosis membrane.

High-quality water purification systems using reverse osmosis (RO) membrane separation have faced a major challenge related to biofilm formation on the membrane surface, or biofouling. To understand this issue, the biofilm formation characteristics of four bacterial isolates previously retrieved from an RO membrane treating potable water were investigated. Biofilm formation of all four isolates occurred to different extents in microtiter plates and could be related to one or more cell properties (hydrophobicity, surface charge, and motility). For Dermacoccus sp. strain RO12 and Microbacterium sp. strain RO18, bacterial adhesion was facilitated by cell surface hydrophobicity, and for Rhodopseudomonas sp. strain RO3, adhesion was assisted by its low surface charge. Sphingomonas sp. strain RO2 possessed both twitching and swarming motilities, which could be important in mediating surface colonization. Further, strains RO2, RO3, and RO12 did not exhibit swimming motility, suggesting that they could be transported to RO membrane surfaces by other mechanisms such as convective permeate flow. The biofilm formation of RO2 was further tested on different RO membranes made of cellulose acetate, polyamide, and thin film composite in continuous flow cell systems. The resultant RO2 biofilms were independent of membrane surface properties and this was probably related to the ex-opolysaccharides secreted bythe biofilm cells. These results suggested that RO2 could colonize RO membranes effectively and could be a potential fouling organism in RO membranes for freshwater purification.

Bacterial Physiological Phenomena↗
PubMed2005

Environmental microbiology-on-a-chip and its future impacts.

Rapid advances in microfabrication, DNA and protein microarray and microfluidic technologies have enabled the development of fully-integrated, miniaturized systems. These so called 'laboratory-on-a-chip' (LOC) devices perform sample preparation (i.e. concentration, separation and purification) together with biochemical reactions and detection steps in a simple and automated manner. We believe LOC technology for environmental microbiology studies will have immediate impacts on microbial monitoring by achieving detection and identification within minutes at the single-cell level, and on microbial ecology by deepening the understanding of microbial community structure and diversity and correlating these with niche-specific functions within a micro space. In the long run, significant impacts are anticipated on environmental metagenomics and proteomics.

Environmental Microbiology↗
PubMed2005

In situ identification and characterization of the microbial community structure of full-scale enhanced biological phosphorous removal plants in Japan.

Fluorescent in situ hybridization (FISH) and polyphosphate (polyP) staining methods were used to characterize the microbial community structure of 13 activated sludge samples taken from nine different Japanese wastewater treatment plants with and without enhanced biological phosphorous removal (EBPR) activities. FISH with published rRNA-targeted oligonucleotide probes for important bacterial groups involving in the EBPR process revealed that Rhodocyclus-related polyphosphate accumulating organisms (PAOs) and glycogen accumulating organisms from a gammaproteobacterial lineage GB were the predominant populations detected, representing 4-18% and 10-31% of EUBmix-stained cells, respectively, in those samples. However, a considerable proportion of Rhodocyclus-related PAO cells were observed with no polyP granules accumulated based on polyP staining. This was further supported by a poor correlation between Rhodocyclus-related PAO population and sludge total phosphorous (TP) contents. In contrast, high correlations between polyP-stained cells and sludge TP contents were observed. In particular, among those polyP-stained cells in samples Ariake_A2O and Nakano_AO, more than 85% of them could not be targeted by probe PAOmix. These non-Rhodocyclus-related PAOs included populations from other bacterial divisions and members of the Betaproteobacteria other than those in Rhodocyclus-related group.

Bacteria↗
PubMed2005

Emission characteristics of fluorescent labels with respect to temperature changes and subsequent effects on DNA microchip studies.

The effects of temperature, salt concentration, and formamide concentration on the emission characteristics of commonly used fluorescent labels were evaluated on DNA microchips. The emission intensities of different fluorophores without hybridization were observed to vary, each to a different extent, to mainly temperature changes. Rhodamine red, TAMRA (tetramethylrhodamine), and dyes from the carbocyanide group exhibited the largest variations, and Texas Red and Oregon Green exhibited the smallest variations. This temperature dependency was shown to affect results obtained during melting curve analysis in DNA microarray studies. To minimize the bias associated with the temperature-dependent emission of different fluorescent labels, a normalization step was proposed.

Fluorescein-5-isothiocyanate↗
PubMed2004

LabArray: real-time imaging and analytical tool for microarrays.

UNLABELLED: Microarrays have been used to perform high-throughput genetic analyses such as single-nucleotide polymorphisms detection and microbial genome analysis. Some of these analyses require real-time monitoring of the hybridization signals with respect to a varying experimental condition, such as temperature. However, current microarray imaging and analysis packages typically do not possess such real-time capabilities. Therefore, microarray image analyses are often time-consuming and labour-intensive. LabArray was developed to expedite such processes by enabling real-time monitoring of microarray signals. AVAILABILITY: LabArray is available at http://www.eng.nus.edu.sg/civil/Labarray/labarray.htm CONTACT: cveliuwt@nus.edu.sg SUPPLEMENTARY INFORMATION: Screenshots and instructions for use are available at the above website.

Algorithms↗
PubMed2004

Filter-based microfluidic device as a platform for immunofluorescent assay of microbial cells.

A filter-based microfluidic device was combined with immunofluorescent labeling as a platform to rapidly detect microbial cells. The coin-sized device consisted of micro-chambers, micro-channels and filter weirs (gap = 1-2 microm), and was demonstrated to effectively trap and concentrate microbial cells (i.e., Cryptosporidium parvum and Giardia lamblia), which were larger in size than the weir gap. After sample injection, a staining solution containing fluorescently-labeled antibodies was continuously provided into the device (flow rate = 20 microl min(-1)) to flush the microbial cells toward the weirs and to accelerate the fluorescent labeling reaction. Using a staining solution that was 10 to 100 times more dilute than the recommended concentration used in a conventional glass method, those target cells with a fluorescent signal-to-noise ratio of 12 could be microscopically observed at single-cell level within 2 to 5 min prior to secondary washing.

Animals↗
PubMed2004

Microbial community structure in a thermophilic anaerobic hybrid reactor degrading terephthalate.

A thermophilic terephthalate-degrading methanogenic consortium was successfully enriched for 272 days in an anaerobic hybrid reactor, and the microbial structure was characterized using terminal RFLPs, clone libraries and fluorescence in-situ hybridization with rRNA-targeted oligonucleotide probes. All the results suggested that Methanothrix thermophila-related methanogens, Desulfotomaculum-related bacterial populations in the Gram-positive low-G + C group, and OP5-related populations were the key members responsible for terephthalate degradation under thermophilic methanogenic conditions except during periods when the reactor experienced heat shock and pump failure. These perturbations caused a significant shift in bacterial population structure in sludge samples taken from the sludge bed but not from the surface of the packing materials. After system recovery, many other bacterial populations emerged, which belonged mainly to the Gram-positive low-G + C group and Cytophaga-Flexibacter-Bacteroides, as well as beta-Proteobacteria, Planctomycetes and Nitrospira. These newly emerged populations were probably also capable of degrading terephthalate in the hybrid system, but were out-competed by those bacterial populations before perturbations.

Anaerobiosis↗
PubMed2004

Identification and occurrence of tetrad-forming Alphaproteobacteria in anaerobic-aerobic activated sludge processes.

In an acetate-fed anaerobic-aerobic membrane bioreactor, a deteriorated enhanced biological phosphorus removal (EBPR) community was developed (as determined based on the chemical profiles of organic substrate, soluble phosphate, and intracellular carbohydrate and polyhydroxyalkanote (PHA) concentrations). Microscopic observations revealed the dominance of tetrad-forming organisms (TFOs), of which the majority stained positively for PHA under anaerobic conditions. Fluorescence in situ hybridization (FISH) confirmed that the Alphaproteobacteria (85.0+/-7.0% of total cells) were the most dominant group. A 16S rRNA gene clone library specific for the Alphaproteobacteria indicated that most 16S rRNA gene clones (61% of total clones) were closely affiliated with 'Defluvicoccus vanus', forming a cluster within subgroup 1 of the Alphaproteobacteria. Combined PHA staining and FISH with specific probes designed for the members of the 'Defluvicoccus' cluster suggested diversity within this TFO cluster, and that these TFOs were newly identified glycogen-accumulating organisms in EBPR systems. However, these 'Defluvicoccus'-related TFOs were only seen in low abundance in 12 different EBPR and non-EBPR systems, suggesting that they were not the key populations responsible for the deterioration of full-scale EBPR processes.

Aerobiosis↗
PubMed2004

Evaluating single-base-pair discriminating capability of planar oligonucleotide microchips using a non-equilibrium dissociation approach.

The capability of planar rRNA-based oligonucleotide microarrays for single-base-pair discrimination was evaluated using an approach that compares the non-equilibrium dissociation profiles and dissociation temperatures (Tds) of all probe-target duplexes simultaneously. Three sets of 16S rRNA gene specific probes at different levels of specificity were used along with their counter probes for individual sets having either one or two mismatches (MM) to their targets at specific external (next to terminus) and various internal positions. Criteria based on the Td approach and a discrimination index (DI) were proven to be competent in discriminating PM from internal MM duplexes, but not always for external MM duplexes. Maximal DI for separating PM duplexes from ones with two and one internal MM usually occurred at temperatures approximately 5-10 degrees C and 10-15 degrees C, respectively, higher than the Tds of the PM duplexes. Washing buffer type and salt concentration, and MM number and position were shown statistically to affect dissociation profiles, Td, and single-base-pair discriminating capability. The reusability potential of the planar microchip was further demonstrated.

Base Pair Mismatch↗
PubMed2002

Characterization of microbial community in granular sludge treating brewery wastewater.

The diversity and distribution of microbes within brewery-degrading anaerobic sludge granules were studied using various molecular techniques. Molecular cloning of small-subunit rRNA gene sequences indicated that all archaeal clones were affiliated with Methanosaeta concillii (>99% sequence similarity), and the bacterial clones were mostly affiliated with a not-yet-cultured Clostridium cluster (48 out of 99 clones) in the low G + C gram-positive group, Xanthomonas spp. in the gamma-subclass of Proteobacteria (30 clones), and Desulfovibrio spp. (16 clones) in the delta-subclass of Proteobacteria. Slot-blot hybridization indicated that archaeal cells from the Methanomicrobiales (58.4% of total rRNA), Methanobacterials (3.3%) and Methanococcales (1.0%) accounted for 62.4% of the total community rRNA. The rest of the microbial populations were the clostridial cluster (27.3% of total rRNA) and Desulfovibrio spp. (9.4%). Fluorescence in-situ hybridization with domain and group-specific oligonucleotide probes further revealed a multi-layer granular architecture. On the surface layer, the hydrolytic clostridial species and hydrogenotrophic Methanobacteriales were the predominant. In the middle layer, mostly H2-producing acetogens from the delta-Proteobacteria (i.e., Desulfovibrio spp.), hydrogenotrophic Methanobacteriales and aceticlastic Methanosaeta were observed to presumably form a syntrophic association. Finally, the center core consisted of microcolonies of Methanosaeta cells.

Archaea↗
PubMed2002

Microbial community dynamics during start-up of acidogenic anaerobic reactors.

Start-up of two acidogenic reactors under mesophilic (37 degrees C) and thermophilic (55 degrees C) conditions was carried out with methanogenic granular sludge as an inoculum and dairy wastewater as feed. During these 71 days of the start-up period, microbial community dynamics in these two acidogenic reactors, as monitored by denaturing gradient gel electrophoresis (DGGE) and dot-blot hybridization with group-specific oligonucleotide probes, was correlated to reactor performance. Due to pH drop to 5.5, DGGE community fingerprints for domains Bacteria and Archaea populations showed significant shifts after 13 days of operation, and this change was accompanied with an increase in volatile fatty acid production, a decrease in methane formation, and rapid sludge disintegration. Dot-blot hybridization results further indicated that the decrease in methane production was related to the decrease in Archaea population in particular with methanogens from 34.1% of total 16S-rRNA in the seed sludge to 8% within the first 13 days, and to 2-5% at day 71. Among the methanogens monitored, the class Methanomicrobiales was the most abundant followed up by Methanobacteriales and Methanococcales. Due to an elevated temperature, the microbial community change was more significant and rapid in the thermophilic reactor than in the mesophilic reactor. Significant microbial population changes took place at the first 13 days for both reactors, but a longer period up to 71 days was required to establish a microbial community with a stable metabolic activity.

Animals↗