Simultaneous amplification of DNA in a multiplex circular array shaped continuous flow PCR microfluidic chip for on-site detection of bacterial.

Based on time to place conversion, continuous flow polymerase chain reaction (CF-PCR) can realize a rapid amplification of DNA by running the PCR reagent in a serpentine microchannel but a larger space is required for each sample, which greatly reduces the efficiency of the CF-PCR. Herein, we propose a multiplex circular array shaped CF-PCR microfluidic chip for on-site detection of bacteria. There were 12 serpentine microchannels which were distributed on the disc in an annular form, and each microchannel consisted of an inlet for sample injection, and an outlet for the detection of the PCR products based on fluorescence. Samples could be simultaneously driven into each inlet by a one-to-twelve diverter through a syringe. Moreover, the method of adding fluorescent dyes at the end of the microchannel can solve the inhibition effect of excessive fluorescent dyes on the PCR reaction. The process finished with simultaneous amplification of 12 different target genes from Porphyromonas gingivalis, Treponema denticola, Tannerella forsythia, and Escherichia coli, and on-site detection of their corresponding positives within 23 min. The fastest detectable PCR reaction time was 5.38 ± 0.2 min at a flow rate of 1 mL h-1. For E. coli, the minimum detectable concentration was 2.5 × 10-3 ng µL-1 in this microfluidic system. Such a system can increase the throughput of CF-PCR for point-of-care testing of pathogens.

Lower fluidic resistance of double-layer droplet continuous flow PCR microfluidic chip for rapid detection of bacteria.

BACKGROUND: Rapid diagnosis of harmful microorganisms demonstrated its great importance for social health. Continuous flow PCR (CF-PCR) can realize rapid amplification of target genes by placing the microfluidic chip on heaters with different temperature. However, bubbles and evaporation always arise from heating, which makes the amplification not stable. Water-in-oil droplets running in CF-PCR microfluidic chip with uniform height takes long time because of the high resistance induced by long meandering microchannel. To overcome those drawbacks, we proposed a double-layer droplet CF-PCR microfluidic chip to reduce the fluidic resistance, and meanwhile nanoliter droplets were generated to minimize the bubbles and evaporation. RESULTS: Experiments showed that (1) fluidic resistance could be reduced with the increase of the height of the serpentine microchannel if the height of the T-junction part was certain. (2) Running speed, the size and the number of generated droplets were positively correlated with the cross-sectional area of the T-junction and water pressure. (3) Droplet fusion happened at higher water pressure if other experimental conditions were the same. (4) 0.032 nL droplet was created if the cross-sectional area of T-junction and water pressure were 1600 µm2 (40 × 40 µm) and 7 kPa, respectively. Finally, we successfully amplified the target genes of Porphyromonas gingivalis within 11'16″ and observed the fluorescence from droplets. SIGNIFICANCE AND NOVELTY: Such a microfluidic chip can effectively reduce the high resistance induced by long meandering microchannel, and greatly save time required for droplets CF-PCR. It offers a new way for the rapid detection of bacterial.

A Tropical Cyclone Center Location Method Based on Satellite Image.

Accurately detecting and locating the center of the tropical cyclone is critical for the trajectory forecasting. This study proposed an automatic method for centers' location of the tropical cyclones based on the visible or the infrared satellite images. The morphological structure of the tropical cyclone is modeled using the circular pattern. The tropical cyclone center is located based on regional pixels instead of skeleton points. All pixels in a segmented cloud cluster vote for a 2-dimensional accumulator. The center of the cloud cluster is computed by the mean voting distances, which are calculated by fitting quadratic functions in every column of the two-dimensional (2D) accumulator. Then, a linear function is fitted according to the functional relationship between the mean voting distance and voting angle. The fitted coefficients of the linear function are the center coordinates of the tropical cyclone. The proposed method for centers location of the tropical cyclones is tested using visible and infrared satellite images. The results of center location are compared with the best track provided in JMA datasets.

The Mitigation Effect of Park Landscape on Thermal Environment in Shanghai City Based on Remote Sensing Retrieval Method.

The mitigation effects of park green space on Urban Heat Island (UHI) have been extensively documented. However, the relative effects of the configuration of park components on land surface temperature (LST) inside the park and indicators (i.e., park cooling intensity and distance) surrounding the park is largely unknown. Therefore, the main objective of this study is to explore the quantitative impacts of configuration and morphology features under different urban park scales on the cooling effect. In this study, based on Landsat-8 OLI/TIRS images on 3 August 2015 and 16 August 2020 during summer daytime, the LSTs of Shanghai City were retrieved by atmospheric correction method. Then, the relationships of park landscape features with LSTs in the park and typical indicators representing cooling efficiency of 24 parks on different grades were analyzed. The results showed that the average temperature in urban parks was, respectively, 1.46 °C and 1.66 °C lower than that in the main city of Shanghai in 2015 and 2020, suggesting that urban parks form cold islands in the city. The landscape metrics of park area (PA), park perimeter (PP), green area (GA) and water area (WA), were key characteristics that strong negatively affect the internal park LSTs. However, the park perimeter-to-area ratio (PPAR) had a significant positive power correlation with the park LSTs. Buffer zone analysis showed that LST cools down by about 0.67 °C when the distance from the park increases by 100 m. The Maximum Cooling Distance (MCD) for 2015 and 2020 had a significant correlation with PA, PC, PPAR, GA and WA, and increased sharply within the park area of 20 ha. However, the medium park group had the largest Maximum Cooling Intensity (MCI) in both periods, followed by the small park group. There could be a trade-off relationship between the MCD and MCI in urban parks, which is worth pondering to research. This study could be of great significance for planning and constructing park landscapes, alleviating Urban Heat Island effect and improving urban livability.

A continuous flow PCR array microfluidic chip applied for simultaneous amplification of target genes of periodontal pathogens.

The concept of time to place conversion makes using a continuous flow polymerase chain reaction (CF-PCR) microfluidic chip an ideal way to reduce the time required for amplification of target genes; however, it also brings about low throughput amplicons. Although multiplex PCR can simultaneously amplify more than one target gene in the chip, it may easily induce false positives because of cross-reactions. To circumvent this problem, we herein fabricated a microfluidic system based on a CF-PCR array microfluidic chip. By dividing the chip into three parts, we successfully amplified target genes of Porphyromonas gingivalis (P.g), Tannerella forsythia (T.f) and Treponema denticola (T.d). The results demonstrated that the minimum amplification time required for P.g, T.d and T.f was 2'07'', 2'51'' and 5'32'', respectively. The target genes of P.g, T.d and T.f can be simultaneously amplified in less than 8'05''. Such a work may provide a clue to the development of a high throughput CF-PCR microfluidic system, which is crucial for point of care testing for simultaneous detection of various pathogens.

An insight into the retrogradation behaviors and molecular structures of lotus seed starch-hydrocolloid blends.

The retrogradation behaviors and molecular structures of lotus seed starch (LS) combined with different hydrocolloids, chitosan (CS), guar gum (GG) and xanthan (XN), were investigated. Following an autoclave treatment, the storage modulus (G') value of LS-CS increased more rapidly than LS alone, indicating an increase in starch retrogradation. This might result from intermolecular interactions, increased ordered structure, decreased weight-average molecular weight (Mw) and greater leached amylose content in LS-CS system. The LS-GG and particularly LS-XN blends showed the opposite trend. The much greater Mw of LS-XN was mainly attributed to the lower retrogradation rate. The enwrapping effect of GG or XN on LS, as observed by confocal laser scanning microscopy, might retard their retrogradation by limiting the granules' swelling and the amylose leaching. Overall, the changes in the interaction force, ordered structure, Mw, leached amylose and microstructure were related to retrogradation behaviors of LS-hydrocolloid blends following an autoclave treatment.

Polysaccharide fractions from Fortunella margarita affect proliferation of Bifidobacterium adolescentis ATCC 15703 and undergo structural changes following fermentation.

In this study, the relationships between the proliferation effect of polysaccharide fractions from the citrus shrub Fortunella margarita on Bifidobacterium adolescentis ATCC 15703 and their resulting structural changes were investigated. Four polysaccharide fractions, FP20, FP40, FP60, and FP80, were obtained by graded precipitation at ethanol concentrations of 20%, 40%, 60% and 80%, respectively. The results showed that polysaccharide fractions, especially FP20, FP40, and FP60, enhanced the proliferation of B. adolescentis ATCC 15703 and their effects were better than those of FP80, inulin or glucose. Moreover, acetic acid was mainly produced during fermentation. After fermentation, the molecular weight of polysaccharide fractions decreased and the tightly structural chain conformations of FP20, FP40, and FP60 changed to highly branched structures. The prebiotic effect of these polysaccharide fractions might be related to their molecular weight and chain conformation. Thus, these polysaccharides could be used as potential prebiotics.

Effect of guar gum on the physicochemical properties and in vitro digestibility of lotus seed starch.

The effect of guar gum (GG) on the physicochemical properties and digestibility of lotus seed starch (LS) subjected to autoclaving-cooling were investigated. Compared to LS alone, LS with 0.03% or 0.15% GG displayed higher crystallinity and short-range order, resulting in the formation of a denser crystal structure. It might contribute to the reduced swelling power, final viscosity, and setback viscosity for those complexes. Coating effect and hydrophilicity were more apparent in LS containing 0.30%, 0.60%, or 0.90% GG, which weakened their crystal structure and increased swelling power, final viscosity, and setback viscosity. The addition of GG delayed starch digestion in vitro, decreased the content of rapidly digestible starch, and increased the content of resistant starch, especially when 0.30% GG was added to 30% LS. The changes in crystal structure and viscosity partially explained the reduced digestibility.