4,356 research outputs found
Titanium Dioxide Nanoparticle Humidity Microsensors Integrated with Circuitry on-a-Chip
A humidity microsensor integrated with a readout circuit on-a-chip fabricated using the commercial 0.18 ÎĽm CMOS (complementary metal oxide semiconductor) process was presented. The integrated sensor chip consists of a humidity sensor and a readout circuit. The humidity sensor is composed of a sensitive film and interdigitated electrodes. The sensitive film is titanium dioxide prepared by the sol-gel method. The titanium dioxide is coated on the interdigitated electrodes. The humidity sensor requires a post-process to remove the sacrificial layer and to coat the titanium dioxide. The resistance of the sensor changes as the sensitive film absorbs or desorbs vapor. The readout circuit is employed to convert the resistance variation of the sensor into the output voltage. The experimental results show that the integrated humidity sensor has a sensitivity of 4.5 mV/RH% (relative humidity) at room temperature
Rapid identification and application of Lactobacillus plantarum, Lactobacillus paracasei and Lactobacillus pentosus using multiplex polymerase chain reaction and species-specific primers, targeting 16S ribosomal RNA and recA genes
Background and Objective: Various products on the market contain probiotics such as lactic acid bacteria, which are promoted with a wide range of benefits. Functionality of these products is linked to the specific strains, bacterial species and viable cell counts. This study aimed to assess conformity of targeted lactic acid bacterial species and viable cell counts in commercially available probiotic products with their labeling, ensuring efficacy of the products.
Material and Methods: Multiplex polymerase chain reaction technique was developed using specific primers to effectively differentiate lactic acid bacteria in probiotic products. Therefore, strains used in the products were targeted and relevant nucleotide sequence data were searched to select two sets of polymerase chain reaction primer pairs of L. pla-F/R and L. para-F/R, targeting 16S ribosomal RNA genes, and L .pen-F/R, targeting recA genes.
Results and Conclusion: The individual primer sets produced the expected target products that matched the labeling for the tested strains of Lactobacillus plantarum, Lactobacillus paracasei and Lactobacillus pentosus. Then, specificity assessing was carried out using multiplex primer sets for single strains, pairwise combinations and triple combinations of lactic acid bacteria. After verifying specificity for all the three strains under similar polymerase chain reaction conditions, sensitivity of the multiplex polymerase chain reaction was investigated by assessing various dilutions of the three lactic acid bacterial strains and commercially available probiotic products. These findings demonstrated potential uses of multiplex polymerase chain reaction in lactic acid bacterial detection techniques. In conclusion, specific primer sets can be used in multiplex polymerase chain reaction to rapidly and effectively detect lactic acid bacterial strains in commercial products.
Conflict of interest: The authors declare no conflict of interest.
Lactic acid bacteria (LAB) are addressed for their beneficial effects on the human gastrointestinal tract, enhan-cing overall immune health. There are recent interests in development of functional LAB products. Demands for the probiotic functional foods are rapidly increasing due to the increased consumer awareness of food effects on health [1]. Researchers have assessed potential uses of probiotics in dairy and nondairy products and their viabilities during storage [2]. They have concluded that the final product should contain a minimum of 106-107 viable cells per serving to benefit consumer health [2]. Their studies have shown that mislabeling of probiotic species is common in commercial products [3]. Lack of appropriate identification of the strains and false efficacy claims have led to confusion. Probiotic products available in the market often use mixed strains. Thus, there are needs to monitor conformities of the labeled bacterial species and viable cell counts.
Polymerase chain reaction (PCR) technique has success-fully been used to detect and differentiate viruses and bacteria in various foods [4]. Rapid and reliable nature of PCR provides a valuable tool for distinguishing closely related species within two groups of lactobacilli [5]. This technique has been used to rapidly identify Lactobacillus plantarum in kimchi [6]. Oligonucleotide primers have been developed from sequences between the 16S and 23S rRNA genes, enabling identification of various lactobacilli strains in dairy products and probiotics using PCR [7]. Further-more, strain-specific PCR can be used for the rapid identi-fication of lactobacilli isolated from food samples [8] and specific identification of ten common lactobacilli and bifi-dobacteria strains in fermented milks [9].
In this study, multiplex PCR method was developed to investigate applicability of molecular detection techniques for LAB using three sets of specific primers sourced from the literature. Moreover, 16S rRNA gene sequences were targeted to explore molecular LAB detection. By amplifying 16S rRNA and recA gene sequences through PCR, this study simultaneously detected three LAB strains as well as mixed LAB strains in the products. Sensitivity of detection was assessed to establish a simple, reliable rapid method appropriate for the effective identification of LAB strains in probiotic products.
Materials and Methods
2.1 Bacterial strains and culture conditions
The LAB strains were stored in a -80 °C freezer. Before the experiments, strains were activated twice using lactobacilli MRS broth (Difco, Detroit, MI, USA) supplemented with 0.05% (w/w) L-cysteine (Merck, Taipei, Taiwan). Strains were cultured under optimal growth conditions at 37 °C for 24 h. Reference strains were provided by the Bioresources Collection and Research Center (BCRC, Hsin-Chu, Tai-wan). The L. pentosus BCRC 17972 and 17973, L. plantar-um F7-1 and L. paracasei BCRC 12193, 12188, 12248 and 17002 were used in this study. Seven strains were used in the current study as well. The commercial probiotic product was purchased from Li-Fong, Tainan, Taiwan, for PCR detection. Each sachet of the bacterial powder contained a high level of viable probiotic cells with 5.0 × 1010 CFU.g-1. Specific strains included in the product were L. plantarum LP112, L. paracasei LPC188 and L. pentosus LPE588. ThIS study was carried out at Testing and Analysis Center for Food and Cosmetics, HungKuang University, Taichung City, Taiwan.
2.2 Genomic DNA preparation and polymerase chain reaction primers
Total chromosomal DNA of the LAB cells was extracted using Blood and Tissue Genomic DNA Extraction Miniprep System (Viogene, Taipei, Taiwan) based on the manuf-acturer’s instructions. Specific primer sequences for the LAB detection are shown in Table 1. Experiments were repeated thrice [10-12].
2.3 Polymerase chain reaction amplification
Method was carried out according to [13]. For each PCR cycle, denaturation, annealing and extension were carried out at 94 °C for 60 s, 57 °C for 60 s and 72 °C for 120 s, respectively. Final extension was carried out at 72 °C for 5 min.
2.4 Sensitivity of the polymerase chain reaction assay
A 24-h culture of the LAB strain was serially diluted 10-fold with sterile water. Purification of DNA was carried out as described in Section 2.2 [14].
2.5 Polymerase chain reaction detection in the commercial probiotic product
The probiotic product was purchased from Li-Fong, Tainan, Taiwan. After diluting the product to 108–106, 1 ml of the diluted sample was collected and DNA extraction was carried out. Then, 2 μl of the extracted DNA was used for multiplex PCR. Experiments were repeated thrice.
Results and Discussion
3.1 Multiplex Polymerase chain reaction
Figure 1 shows gel electrophoresis results of the multiplex PCR for DNA detection of individual LAB strains. Results demonstrated specificity of the three primer sets for their respective target genes in each strain. Small interferences were seen for L. plantarum F7-4 with no effects on amplification of other strains. Figure 1 shows gel electrophoresis results of multiplex PCR for DNA detection of two LAB strains. Results demonstrated that L.pla-F/R, L.pen-F/R and L.para-F/R primer sets could accurately amplify DNA from combination of two LAB strains. No nonspecific products were observed. Thus, 57°C was determined as the optimal annealing temperature for successful primer binding and DNA polymerase activity.
Gel electrophoresis results validated effectiveness and specificity of the multiplex PCR for identifying and discriminating various LAB strains. These findings demonstrated applicability of the developed primer sets and verified their suitability for use in multiplex PCR. The optimized annealing temperature ensured robust amplifica-tion without nonspecific amplification products.
3.2 Sensitivity assessment of the lactic acid bacterial strains using multiplex polymerase chain reaction
In Figure 2A, DNA extracted directly from the mixed cultures of three LAB strains (109, 108 and 107 cfu ml-1) are shown. At a concentration of 106 cfu ml-1, only L. paracasei (BCRC 12188) demonstrated amplifications in multiplex PCR, indicating that detection sensitivity of L. plantarum and L. pentosus was limited to 107 cfu ml-1. Figure 2B shows mixed cultures of the three LAB strains at similar concen-trations after preculture in MRS broth (37 °C, 24 h). Multi-plex PCR detected all the three strains at a sensitivity of 106 cfu ml-1, suggesting that L. plantarum and L. pentosus proliferated and could be detected. Figure 2 shows detection limits and proliferation capabilities of the three LAB strains using multiplex PCR. Results indicated that preculture of the mixed cultures in MRS broth improves detection sensitivity, enabling accurate identification of L. plantarum and L. pentosus strains at lower concentrations.
3.3 Preculture and mixed various concentrations of lactic acid bacterial strains using multiplex polymerase chain reaction
Figure 3A shows gel electrophoresis results of the multiplex PCR carried out on DNA samples extracted from a mixture of L. plantarum (109 cfu ml-1) with two other LAB strains after preculture for 24 h. In Lanes 5–8, amplification products of L. pentosus and L. paracasei diluted to 106 cfu ml-1 were less expressed. The PCR amplification products of 106 cfu ml-1 DNA could be observed. The L. pentosus BCRC 17973 demonstrated a bacterial count of ~109 cfu ml-1 after 24 h of cultivation, whereas L. paracasei BCRC 12188 showed increased bacterial count, suggesting that preculture could enhance detection rate of low-concen-tration bacterial strains. Figure 3B shows gel electrophoresis results of the multiplex PCR on DNA extracted from a mixture of L. plantarum (108 cfu ml-1) with two other LAB strains after preculture for 24 h. The PCR products in Lanes 1–16 indicated that all the three sets of species-specific primer pairs yielded the expected PCR products for various concentrations of the bacterial suspensions after 24 h of preculture. Figure 3 shows effectiveness of the multiplex PCR in detecting L. plantarum at various concentrations in a mixed culture with other LAB strains. Results highlighted effects of preculture on enhancing assay detection rate and sensitivity.
3.4 Polymerase chain reaction detection of the commercial probiotic product
Figure 4 demonstrates results of the multiplex PCR. Whether directly detected or precultured, the three LAB strains provided DNA amplification products from 107 to 109 cfu ml-1. It was suggested that 107 cfu ml-1 was the detection limit of the product (not detected when diluted to 106 cfu ml-1).
The PCR-based species identification is a critical highly valuable tool for detecting and identifying bacteria. It offers advantages, including time efficiency and reliability in microbial identification. Compared to traditional methods such as culture-based techniques, PCR can provide results in a relatively short time. It eliminates the need of time-consuming cultivation of bacteria, allowing for further rapid identification and subsequent decision-making processes. In industrial uses, species identification is critical in selecting and developing bacterial strains appropriate for specific purposes.
Whether in food, agriculture or biotechnology industries, PCR-based species identification enables researchers to screen and select the most appropriate bacterial species for desired characteristics and functions.
The 16S rRNA gene in the ribosomal RNA of prokaryotes is the best molecular marker for bacterial evolutionary analysis. This is due to several gene characteristics, including its presence across various species, abundance, sufficient sequence length and presence of conserved and variable loci. The 16S rRNA gene is widely used in identifying lactobacilli and a commonly rapid technique for bacterial classification and identification in dairy products. Caro et al. reported that partial sequencing of 16S rRNA genes is often used for lactobacilli identification [15]. The RecA protein is a DNA recombinase that plays critical roles in DNA repair and recombination processes of bacteria. It is encoded by the recA gene in the genomes of various prokaryotic microorganisms. The recA gene and its corresponding protein have extensively been studied and used in various research, including evolutionary analysis, phylogenetic studies and identification of bacterial strains. Conserved nature and functional importance of the RecA protein make it a valuable molecular marker for understanding genetic relatedness and evolutionary relationships within bacteria. By comparing the recA gene sequences of various bacterial strains, researchers can have insights into their genetic diversity, evolutionary history and phylogenetic classification.
Lu et al. developed a multiplex PCR method that could effectively be used in key vaginal microbiota evaluation in women with bacterial vaginosis [16]. You et al. used multiplex PCR to detect six species of L. acidophilus group [17]. Petri et al. used multiplex PCR for the rapid identification of wine-associated LAB [18]. Settanni et al. introduced a method and reported its state-of-the-art uses for microbial identification in foods and beverages [19]. Sciancalepore et al. described use of a simple, low-cost, rapid sensitive method based on droplet-based multiplex PCR directly on food matrices for the simultaneous detection of bacterial genes involved in biogenic amine synthesis [20]. Specific primers were designed based on similar sequences and multiplex PCR was optimized for the simultaneous identification of L. plantarum, L. pentosus and L. paraplantarum [21]. Sul et al. developed a multiplex PCR to detect Lactobacillus and Bifidobacterium spp. in commercial probiotic products [13].
Previously, Gram-negative broth enrichment was used in studies, followed by immunomagnetic separation multiplex PCR to enable the simultaneous detection of Salmonella spp. and enterohemorrhagic Escherichia coli in food samples, irrespective of their significant discrepancy in cell counts [22]. A PCR primer set derived from the seque-nce in 16S to 23S internal transcribed spacer (ITS) region was also developed for the specific detection of B. adoles-centis in probiotics. This primer set included potentials for inspecting dairy food and environmental samples [14]. To ensure food safety during direct vat inoculation of Paocai, a propidium monoazide-based quantitative PCR method was developed to quantify L. plantarum NCU116 fermentation starter, as well as Saccharomyces spp. and potentially present pathogenic bacteria [23]. Plate counting was carried out and demonstrated similar results to quantitative PCR analysis, indicating appropriateness and effectiveness of absolute quantitative PCR for rapidly detecting microbial composition in the Paocai system [23]. In a recent study, surveillance of Lactobacillus bacteremia was carried out using biochemical and conventional-PCR assays. However, these methods could not provide target quantification and might lead to false-positive results [24]. To address this limitation, a L. rhamnosus-specific quantitative PCR assay was developed. This assay delivers accurate and reproduci-ble results, leveraging specificity of a TaqMan probe, targ-eting unique 16S rDNA sequences of L. rhamnosus [24].
Conclusion
In summary, the three sets of PCR primer combinations, targeting various LAB strains, demonstrated specificity and generated the expected amplicons during PCR amplifica-tion. Use of multiplex PCR in LAB genomic detection showed potentials and was appropriate for detecting various species in food products. Multiplex PCR decreased experimental cost and time, eliminating the need of time-consuming sequencing processes. Therefore, this method is expected to contribute to the reliability of probiotic labeling systems by facilitating strain identification. This molecular technique offers a valuable tool for quality control, product development and microbial monitoring of probiotic strains in various fields.
Conflict of Interest
The authors report no conflict of interest.
Authors Contributions
Conceptualization, TCC; methodology, LZY; data curation, LZY; writing, TCC.
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Developing Mobile BIM/2D Barcode-Based Automated Facility Management System
Facility management (FM) has become an important topic in research on the operation and maintenance phase. Managing the work of FM effectively is extremely difficult owing to the variety of environments. One of the difficulties is the performance of two-dimensional (2D) graphics when depicting facilities. Building information modeling (BIM) uses precise geometry and relevant data to support the facilities depicted in three-dimensional (3D) object-oriented computer-aided design (CAD). This paper proposes a new and practical methodology with application to FM that uses an integrated 2D barcode and the BIM approach. Using 2D barcode and BIM technologies, this study proposes a mobile automated BIM-based facility management (BIMFM) system for FM staff in the operation and maintenance phase. The mobile automated BIMFM system is then applied in a selected case study of a commercial building project in Taiwan to verify the proposed methodology and demonstrate its effectiveness in FM practice. The combined results demonstrate that a BIMFM-like system can be an effective mobile automated FM tool. The advantage of the mobile automated BIMFM system lies not only in improving FM work efficiency for the FM staff but also in facilitating FM updates and transfers in the BIM environment
COMPARISON OF PLAYER’S CENTER OF MASS MOVEMENT BETWEEN HIGH AND LOW IMPACT POSITIONS IN TENNIS FOREHAND STROKE
During the tennis forehand stroke, the displacement of body center of mass (COM) changes with the body movement. The COM movement influences the recovery from one stroke to the next. Therefore, the purpose of this study is to investigate the differences of COM movement and joint kinematics between high and low-impact positions on different skilled players. This study adopted a 3-D motion analysis system for recording and tracing the advanced (n = 5; level 3-4) and intermediate (n = 7; level 5-6) athletes’ motion of whole body during high and low-impact positions in tennis forehand stroke. The results showed that significant difference was not found between both impact positions and level groups in ball velocity. Advanced group showed greater anterior/posterior displacement than the intermediate group in low-impact position that increased the kinetic energy
An interactively recurrent functional neural fuzzy network with fuzzy differential evolution and its applications
In this paper, an interactively recurrent functional neural fuzzy network (IRFNFN) with fuzzy differential evolution (FDE) learning method was proposed for solving the control and the prediction problems. The traditional differential evolution (DE) method easily gets trapped in a local optimum during the learning process, but the proposed fuzzy differential evolution algorithm can overcome this shortcoming. Through the information sharing of nodes in the interactive layer, the proposed IRFNFN can effectively reduce the number of required rule nodes and improve the overall performance of the network. Finally, the IRFNFN model and associated FDE learning algorithm were applied to the control system of the water bath temperature and the forecast of the sunspot number. The experimental results demonstrate the effectiveness of the proposed method
Associations of parental bonding and adolescent internet addiction symptoms with depression and anxiety in parents of adolescents with attention deficit/hyperactivity disorder
Objectives: The aim of the present study was to evaluate the associations of parental bonding and adolescents’ Internet addiction symptoms with depression and anxiety in parents of adolescents with attention deficit/hyperactivity disorder (ADHD). Methods: Parental depression and anxiety symptoms, parental bonding, and adolescents’ Internet addiction symptoms were assessed in 46 parent-child dyads using the Center for Epidemiological Studies Depression Scale, State-Trait Anxiety Inventory, Parental Bonding Instrument (PBI), and Chen Internet Addiction Scale, respectively. Forward stepwise multiple regression analysis was used to examine the associations of parental bonding and adolescents’ Internet addiction symptoms with parental depression and anxiety. Results: Low care/affection on the PBI was significantly associated with parental depression, and overprotection on the PBI and adolescents’ Internet addiction were significantly associated with parental anxiety. Discussion: Parental bonding and adolescents’ Internet addiction are related to depression and anxiety in parents of adolescents with ADHD
Plasmonic Circular Nanostructure for Enhanced Light Absorption in Organic Solar Cells
This study attempts to enhance broadband absorption in advanced plasmonic circular nanostructures (PCN). Experimental results indicate that the concentric circular metallic gratings can enhance broadband optical absorption, due to the structure geometry and the excitation of surface plasmon mode. The interaction between plasmonic enhancement and the absorption characteristics of the organic materials (P3HT:PCBM and PEDOT:PSS) are also examined. According to those results, the organic material's overall optical absorption can be significantly enhanced by up to ~51% over that of a planar device. Additionally, organic materials are enhanced to a maximum of 65% for PCN grating pitch = 800 nm. As a result of the PCN's enhancement in optical absorption, incorporation of the PCN into P3HT:PCBM-based organic solar cells (OSCs) significantly improved the performance of the solar cells: short-circuit current increased from 10.125 to 12.249 and power conversion efficiency from 3.2% to 4.99%. Furthermore, optimizing the OSCs architectures further improves the performance of the absorption and PCE enhancement
Developing Mobile- and BIM-Based Integrated Visual Facility Maintenance Management System
Facility maintenance management (FMM) has become an important topic for research on the operation phase of the construction life cycle. Managing FMM effectively is extremely difficult owing to various factors and environments. One of the difficulties is the performance of 2D graphics when depicting maintenance service. Building information modeling (BIM) uses precise geometry and relevant data to support the maintenance service of facilities depicted in 3D object-oriented CAD. This paper proposes a new and practical methodology with application to FMM using BIM technology. Using BIM technology, this study proposes a BIM-based facility maintenance management (BIMFMM) system for maintenance staff in the operation and maintenance phase. The BIMFMM system is then applied in selected case study of a commercial building project in Taiwan to verify the proposed methodology and demonstrate its effectiveness in FMM practice. Using the BIMFMM system, maintenance staff can access and review 3D BIM models for updating related maintenance records in a digital format. Moreover, this study presents a generic system architecture and its implementation. The combined results demonstrate that a BIMFMM-like system can be an effective visual FMM tool
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