Increase In Number Of Cycles Research Articles

A Comparative Study to Assess the Efficacy of Two Different Estradiol Formulations during In Vitro Fertilization.

Improvements in stimulation protocols, introduction of vitrification, and changes in clinical practices have contributed to improved efficacy and safety of assisted reproductive technology (ART) procedures. This has also led to a concomitant increase in number of cycles requiring hormone replacement therapy (HRT) protocol for performing an embryo transfer. Successful implantation is dependent on endometrial thickness which in turn is regulated by temporal regulation of hormones. Careful control of estrogen levels determines uterine receptivity. One of the most used drugs for achieving appropriate endometrial lining of >7 mm in HRT is estradiol valerate. Although different estrogen formulations with varying physicochemical properties exist, there is not enough literature to support if the differences translate into a discernible clinical outcome in an in vitro fertilization (IVF) setting. Objective and Method. In this study, retrospective in nature, we compare the efficacy of oral estradiol hemihydrate with estradiol valerate in HRT cycles in 2,529 Indian women, undergoing treatment at a center in India between Jan 2017 and May 2019. Results. Our results primarily indicate that between the estradiol valerate and estradiol hemihydrate treatment groups, the implantation rate (IR) was 47.42% and 49.07%, respectively (P value 0.284), and the endometrial thickness (mean ± SEM in mm) that was achieved was 9.25 ± 0.038 mm and 9.57 ± 0.058 mm (P value < 0.001), respectively. There were no significant differences observed in the secondary outcome measures including clinical pregnancy rate, abortion rate, ectopic pregnancy, and live birth rate. Conclusions. Hence, this study concludes that oral estradiol hemihydrate and estradiol valerate are therapeutically equivalent and provide similar clinical outcomes in an IVF setting.

Stress sensitivity of formation during multi-cycle gas injection and production in an underground gas storage rebuilt from gas reservoirs

Permeability sensitivity to stress experiments were conducted on standard core samples taken from Wen 23 Gas Storage at multi-cycle injection and production conditions of the gas storage to study the change pattern of stress sensitivity of permeability. A method for calculating permeability under overburden pressure in the multi-cycle injection and production process was proposed, and the effect of stress sensitivity of reservoir permeability on gas well injectivity and productivity in UGS was analyzed. Retention rate of permeability decreased sharply first and then slowly with the increase of the UGS cycles. The stress sensitivity index of permeability decreased with the increase of cycle number of net stress variations in the increase process of net stress. The stress sensitivity index of permeability hardly changed with the increase of cycle number of net stress variations in the decrease process of net stress. With the increase of cycle number of net stress variation, the stress sensitivity index of permeability in the increase process of net stress approached that in the decrease process of net stress. The lower the reservoir permeability, the greater the irreversible permeability loss rate, the stronger the cyclic stress sensitivity, and the higher the stress sensitivity index of the reservoir, the stronger the reservoir stress sensitivity. The gas zones with permeability lower than 0.3×10−3 μm2 are not suitable as gas storage regions. Stress sensitivity of reservoir permeability has strong impact on gas well injectivity and productivity and mainly in the first few cycles.

Redox Stability of Tubular Solid Oxide Cell Using LaGaO3 Electrolyte Film Prepared by Dip-Coating

Reversible operation of SOFC is now important subjects from energy storage of renewable electric power from solar or wind power. At present, planer type cell design is widely used for this purpose, however, because of tight gas sealing, tubular type cell design is more suitable. In this study, tubular solid oxide cell using La0.9Sr0.1Ga0.8Mg0.2O3−δ (LSGM) thin electrolyte film was prepared by dip-coating and co-sintering process. NiO-YSZ porous substrate with 10 mm diameter and 30 mm length was used for preparation of the cell. For preventing reaction between LSGM and NiO, TiO2-Ce0.6La0.4O2/Ce0.6Mn0.3Fe0.1O2 was used for buffer layer. The cyclic operation of fuel cell and electrolysis mode was measured for investigating the stability of the prepared tubular cell at an intermediate temperature of 600℃ under the fuel of 20%steam-30%H2 in Ar. After 100 redox cycles under the applied current density of ±100 mA cm-2 , OCV was almost the same with the initial value before measurement. The terminal potential in SOFC operation became slightly lower and unstable with increasing cycle number, however, degradation in power density is not significantly observed. The cell also presents a relative stable performance in the electrolysis mode and it seems to be more active to SOEC than that to SOFC. From measurement of internal resistance, degradation may be related with the increased overpotential of anode. Therefore, aggregation and reoxidation of Ni in Ni-YSZ substrate is expected.In order to further increase performance of the cell, effects of Ce infiltration into NiO-YSZ contributes and it was found that the power density and electrolysis performance was much increased by the decreased internal resistance. The redox stability of NiO-YSZ-CeO2 cell was also measured and it also presents a stable redox performance after 100 times cycles by infiltration of CeO2.

Estimating Real-Time qPCR Amplification Efficiency from Single-Reaction Data.

Methods for estimating the qPCR amplification efficiency E from data for single reactions are tested on six multireplicate datasets, with emphasis on their performance as a function of the range of cycles n1–n2 included in the analysis. The two-parameter exponential growth (EG) model that has been relied upon almost exclusively does not allow for the decline of E(n) with increasing cycle number n through the growth region and accordingly gives low-biased estimates. Further, the standard procedure of “baselining”—separately estimating and subtracting a baseline before analysis—leads to reduced precision. The three-parameter logistic model (LRE) does allow for such decline and includes a parameter E0 that represents E through the baseline region. Several four-parameter extensions of this model that accommodate some asymmetry in the growth profiles but still retain the significance of E0 are tested against the LRE and EG models. The recursion method of Carr and Moore also describes a declining E(n) but tacitly assumes E0 = 2 in the baseline region. Two modifications that permit varying E0 are tested, as well as a recursion method that directly fits E(n) to a sigmoidal function. All but the last of these can give E0 estimates that agree fairly well with calibration-based estimates but perform best when the calculations are extended to only about one cycle below the first-derivative maximum (FDM). The LRE model performs as well as any of the four-parameter forms and is easier to use. Its proper implementation requires fitting to it plus a suitable baseline function, which typically requires four–six adjustable parameters in a nonlinear least-squares fit.

A new approach to thermo-mechanical treatment of steel T91 by multiple upsetting-extrusion in a ferritic range

The microstructure and mechanical properties of the ferritic-martensitic steel T91 after severe plastic deformation (SPD) and subsequent thermal treatment are investigated. SPD was realized by the method of multiple “upsetting-extrusion” (MUE) in temperature range of stability of ferritic phase (750-575 °C), thermal treatment of SPD specimens was carried out at temperatures 550–730 °C during 1–100 h. It is revealed that decrease of temperature of “upsetting-extrusion” and increase of cycle number to 5 induces the decrease of mean grain size to 145 nm and increase of degree of microstructure homogeneity. At the same time microhardness increases from 1800 MPa to 2850 MPa. Composition, shape, size and density of carbide precipitates in steel after SPD and thermal treatment are determined. Obtained submicron grain ferritic microstructure with high value of microhardness is stable at exposure temperature of 550 °C. It is shown that temperature variation of thermal treatment induces the considerable change in grain size and mechanical characteristics.

Experimental Investigation of the CO2 Huff and Puff Effect in Low-Permeability Sandstones with NMR.

For low-permeability sandstone reservoirs, CO2 huff and puff is an effective method for increasing oil recovery. Commonly, sandstone formations with low permeability have diverse pore and throat sizes and a complex pore-throat structure, which essentially affects the flow characteristics of CO2 and oil in the formation and further the CO2 huff and puff performance. It is necessary to understand the recovery degree of various microscale pore sizes under different operational parameters during CO2 huff and puff in tight sandstones. In this work, several experiments of cyclic CO2 injection are conducted with sandstone core samples with low permeability. Before and after the injection, the T2 spectra of the sandstone cores are compared using the NMR technique. We then discuss the micro residual oil distribution and recovery degree in different pores, such as micropores (<1 ms), small pores (1–10 ms), medium pores (10–100 ms), and macropores (>100 ms). It is found that the recovery degree in the different pores increases as the pore size increases. Oil can be recovered more easily from macropores and medium pores during the cyclic CO2 injection. The oil contained in micropores is relatively difficult to extract considering a high capillary force under immiscible conditions. It is found that the total recovery degree increases with the increase in soaking time. However, such a recovery degree increment in small pores is not as large as that achieved in medium and large pores. With the CO2 injection volume increase, the total recovery degree increases. When the CO2 injection volume is less than 1.5 PV, it is challenging to extract the oil from micropores and small pores. As the cycle number increases, the cyclic oil recovery decreases, and most of the oil is produced in the first cycle. This suggests that under the experimental conditions of this study, the cycle number of CO2 huff and puff shall not be more than 3. This work is important to further understand the CO2 huff and puff process for improving oil recovery in sandstone reservoirs with low permeability.

Ratcheting Analysis of Steel Plate under Cycling Loading using Dynamic Relaxation Method Experimentally Validated

The present study aimed to introduce a numerical method to study ratcheting strains of rectangular plates. A new numerical analysis was conducted by development of dynamic relaxation method combined with MATLAB software to evaluate the ratcheting behavior of the thin steel plate under mentioned loading condition. In order to verify the results, experimental tests were performed under stress-controlled conditions by a zwick/roell amsler HB100 machine and bending ratcheting of CK45 steel plate at room temperature was studied. Under stress-controlled conditions with non-zero mean stress, ratcheting behavior occurred on thin plate. Moreover, a finite element analysis was carried out by Abaqus using nonlinear isotropic/kinematic (combined) hardening model. The results showed that the rate of ratcheting strain decreased with an increase in cycle number. It was found that the hysteresis loops were wider in experimental method than those of other methods because of more energy dissipation. The numerical results are in a good agreement with the simulation and experimental data. Comparison of errors between these methods obviously demonstrate high accuracy of the new introduced method.

Amplicon-sequencing of raw milk microbiota: impact of DNA extraction and library-PCR

The highly complex raw milk matrix challenges the sample preparation for amplicon-sequencing due to low bacterial counts and high amounts of eukaryotic DNA originating from the cow. In this study, we optimized the extraction of bacterial DNA from raw milk for microbiome analysis and evaluated the impact of cycle numbers in the library-PCR. The selective lysis of eukaryotic cells by proteinase K and digestion of released DNA before bacterial lysis resulted in a high reduction of mostly eukaryotic DNA and increased the proportion of bacterial DNA. Comparative microbiome analysis showed that a combined enzymatic and mechanical lysis procedure using the DNeasy® PowerFood® Microbial Kit with a modified protocol was best suitable to achieve high DNA quantities after library-PCR and broad coverage of detected bacterial biodiversity. Increasing cycle numbers during library-PCR systematically altered results for species and beta-diversity with a tendency to overrepresentation or underrepresentation of particular taxa. To limit PCR bias, high cycle numbers should thus be avoided. An optimized DNA extraction yielding sufficient bacterial DNA and enabling higher PCR efficiency is fundamental for successful library preparation. We suggest that a protocol using ethylenediaminetetraacetic acid (EDTA) to resolve casein micelles, selective lysis of somatic cells, extraction of bacterial DNA with a combination of mechanical and enzymatic lysis, and restriction of PCR cycles for analysis of raw milk microbiomes is optimal even for samples with low bacterial numbers.Key points• Sample preparation for high-throughput 16S rRNA gene sequencing of raw milk microbiota.• Reduction of eukaryotic DNA by enzymatic digestion.• Shift of detected microbiome caused by high cycle numbers in library-PCR.

Deformation Mechanisms of CoCrFeMnNi High-Entropy Alloy Under Low-Cycle-Fatigue Loading

Plastic deformation during low-cycle fatigue (LCF) in equiatomic face-centered cubic (FCC) CoCrFeMnNi high-entropy alloy (HEA) is accumulated by dislocation substructure formation, which leads to crack initiation. Whilst these substructures have been reported before, little has been done to clarify their formation mechanisms and the effects of strain amplitude, cycle number and grain orientation. In this study, cyclic deformation behavior and microstructural evolution of CoCrFeMnNi were examined for two different grain sizes at room temperature. Microstructural investigations by transmission electron microscopy showed that, while the dislocation structures at low strain amplitude (0.3%) mainly consisted of planar slip bands, at higher strain amplitudes (0.5% and 0.7%), wavy-substructures including veins, wall, labyrinth and cells prevailed. Slip mode also changes from initially planar-slip to wavy-slip with increasing cycle numbers, which defines the cyclic stress response. Dislocations in veins, walls, labyrinth and cells are found to have different Burgers vectors, suggesting that apart from wavy-slip, multiple-slip also contributes to their formation. Moreover, distinct dislocation substructure in grains is dictated more by the constraints from neighboring grains rather than by their orientation. Additionally, the formation of various dislocation structures in a single grain is also linked to the constraint effects from the neighboring grains.

Fatigue study on AISI/SAE 1015 steel with shot peening under corrosive environments

This work evaluates alternative four-point bending fatigue stress in 2 < pH < 4 corrosive media of AISI/SAE 1015 steel subjected to shot peening surface treatment. A tri factorial experimental design with replications was used. The intervening factors were: i) load levels, 0.96, 0.85, 0.73, 0.65; ii) corrosive media; pH2, pH4, ambient; iii) shot-peening surface treatment; without any shot peening, S230 shot peening and CW-41 shot peening, for which a total of 108 specimens were tested with a reliability level of 95% and a probability of failure of 3%. Tests were performed on a universal machine in a three-point bending assembly with an electro-pneumatic system. The cycle counting and control were performed by a programmable logic controller. The results of the stress-life method in each one of the scenarios show that the best condition was shown in the probes with shot peening S230 and CW-41 treatment under atmospheric conditions, where the material tends to increase its fatigue behavior; the number of cycles reached 36990 and 37773 respectively. The corrosive effect of pH2 and pH4 reduced on probes without shot peening treatment with a maximum of 9862 and 8670 cycles respectively; however, for S230 and CW-41 shots, an increase in cycle number in pH2 and pH4 was evidenced, with the best result for S230 shot at pH4 with a maximum range of 21568. Likewise, the fractography analysis shows mixed fractures with fatigue failure mechanism; the characteristics found were beach marks, secondary cracks, localized oxidation (filiform, plates, pitting, stress, and compression corrosion), nucleation, microvoids, cleavage facets, intergranular and transgranular cracks.

Comparison of high pressure homogenization, selective thermal denaturation and glycosylation on textural properties of green soybean (Glycine max)tofu by TOPSIS analysis

Impacts of high pressure homogenization (HPH), selective thermal denaturation (STD) and glycosylation (GLY) on textural properties, water holding capacity (WHC) and yield of green soybean tofu were compared and optimal treatment was obtained by TOPSIS analysis. Firstly, the increase in HPH pressure and cycle numbers enhanced the quality of green soybean tofu. Hardness, WHC and yield of HPH-treated tofu (60 MPa, four cycles) were increased by 48.3%, 17.0%, 6.7%, respectively, compared with those untreated. Moreover, STD condition (85 °C, 10min) made hardness, WHC and yield of green soybean tofu increased by 109%, 15.4%, 21.6%, respectively. Accordingly, hardness and WHC of green soybean tofu increased and afterward declined (P < 0.05) with glucose from 0 to 2.5%, whereas it significantly increased with GLY temperature from 80 to 100 °C. Finally, through TOPSIS multi-index evaluation, STD treatment (85 °C, 10 min) was selected as the most effective way to improve textural characteristics, WHC and yield of green soybean tofu. Therefore, our research provides the optimum processing guidance for producing high-quality edible tofu, expanding the soybean products market.

Electrochemical Impedance Spectroscopy Characterization of a Bismuth Oxyiodide (BiOI) Electrochemical Cell in Terms of Various Morphologies

This investigation reports the electrochemical impedance spectroscopy (EIS) characterization of dip-successive ionic layer adsorption and reaction (SILAR)-prepared BiOI electrochemical cells in terms of various morphologies. Here, the photocathode BiOI has been prepared at various cycle numbers, which causes a change in the surface morphology that is observed by field emission scanning electron microscopy (FESEM). The increasing cycle number until 30 brings the change in the BiOI film's flaky structures, crystal size, and effective surface area. At 30-dip-SILAR cycle, the film's uniformity of flaky structure and effective surface area reach their maximum values, resulting in the lowest film resistance and lowest charge transfer resistance (at the BiOI–electrolyte interface) recorded via EIS analysis. Hence, at 30 cycles (thickness 500 nm), the maximum photovoltaic performance has been observed: short-circuit current density JSC = 0.6 mA/cm2, and open-circuit voltage VOC = 0.43 V.

Evaluating the durability, microstructure and mechanical properties of a clayey-sandy soil stabilized with copper slag-based geopolymer against wetting-drying cycles

This study was conducted on durability of the clayey-sand stabilized using the copper slag (CS)-based geopolymer against wetting-drying (W-D) cycles. In this investigation, the mix of 0, 10, and 15% CS and alkaline activator liquid was used. In this solution, the ratio of NaOH to Na2SiO3 is 70:30 and the weight ratio of CS to solution is 1:1. To evaluate the effects of CS content and NaOH concentration (8 M and 11 M) on the strength, durability, and microstructural variation, a series of experiments were conducted after different cycles. The tests carried out for this purpose include standard proctor compaction test, unconfined compressive strength (UCS), Atterberg limits, accumulative loss of mass, swell and shrinkage, ultrasonic P-wave velocity, pH, toxicity characteristic leaching procedure (TCLP), scanning electron microscopy (SEM), and energy-dispersive X-ray spectroscopy (EDS) tests. The UCS results demonstrated a remarkable reduction in cycle 2 due to W-D cycles, followed by a decrease in further cycles. As a result of Atterberg limits, lower PI leads to the lower potential to swell or shrinkage; thus, the lower loss of soil mass with a higher amount of CS content is obtained. The microstructure analysis showed that with the increase in cycle number, the abundance of microcracks and voids were increased. Moreover, dissolved geopolymerization products leached due to wet cycles and formed gel becomes weaker. The results showed that all the chosen mixtures were durable and endured all 12 W-D cycles. Furthermore, their mass loss was lower than allowable mass loss content of ASTM D559, suggesting that this stabilization method was appropriate for this soil.

Experimental study on repeatedly loaded foundation soil strengthened by wraparound geosynthetic reinforcement technique

In the recent past, the potential benefits of wraparound geosynthetic reinforcement technique for constructing the reinforced soil foundations have been reported. This paper presents the experimental study on the behaviour of model strip footing resting on sandy soil bed reinforced with geosynthetic in wraparound and planar forms under monotonic and repeated loadings. The geosynthetic layers were laid according to the reinforcement ratio to minimise the scale effect. It is found that for the same amount of reinforcement material, the wraparound reinforced model resulted in less settlement in comparison to planar reinforced models. The efficiency of wraparound reinforced model increased with the increase in load amplitude and the rate of total cumulative settlement substantially decreased with the increase in number of load cycles. The wraparound reinforced model has shown about 45% lower average total settlement in comparison to unreinforced model, while the double-layer reinforced model has about 41% lower average total settlement at the cost of approximately twice the material and 1.5 times the occupied land width ratio. Moreover, wraparound models have shown much greater stability in comparison to their counterpart models when subjected to incremental repeated loading.

Structural response of high strength S690 welded sections under cyclic loading conditions

High strength S690 steel offers an attractive solution for use in buildings and bridges, owing to the inherent high strength to self-weight ratio of the material which can lead to significant savings in terms of cost and time. However, take up in practice has been hampered by concerns related to the deterioration in mechanical properties of the S690 steel plates after welding as a result of changes in microstructure. Whilst recent studies have illustrated the high levels of ductility that can be provided by the S690 steel, it is essential to assess and quantify the inelastic cyclic response of welded sections made from the same material under earthquake attacks. This paper therefore presents an experimental investigation into the structural response of high strength S690 welded sections under various cyclic loading conditions. A detailed description of 32 cyclic tests, carried out under two different cyclic protocols with various combinations of target strains and loading frequencies, is provided. Particular focus is given to examining the strength degradation in the material, the number of effective cycles completed before fracture, and the energy dissipation performance. A direct comparsion is also provided between the cumulative cyclic response of the S690 welded sections and their unwelded S690 plate counterparts, based on which the deterioration in behaviour is readily quantified.It is shown that a significant deterioration in strength similarly occurs in both the unwelded and welded S690 sections with the increase in number of cycles, particularly under large target strains. However, in terms of ductility, the number of effective cycles to fracture of the S690 welded sections is found to be considerably lower than for the S690 steel plates, typically by a factor of 2 to 3. Moreover, the energy dissipation densities of the S690 welded sections are shown to be only 41% to 47% of those for the counterparts. Overall, in addition to highlighting the importance of conducting realistic cyclic tests for assessing ductility, the findings provide detailed insights into the structural response of the S690 welded sections in steel structures under earthquake attacks, including the influence of target strains and loading frequencies. The results also enable detailed quantification of the cyclic response for the purpose of numerical modelling as well as for determining reliable ductility criteria.

Effects of reuse on the properties of tantalum powders and tantalum parts additively manufactured by electron beam powder bed fusion

The cost of additively manufactured tantalum parts can be effectively lowered by reusing or recycling tantalum powder. To evaluate the effects of reuse of the tantalum powder on the properties of tantalum parts and scaffolds, this study investigated the characteristics of tantalum powder during cycling, including oxygen content, particle morphology, apparent density, tap density, and flowability. Besides, the influence of reuse time on the mechanical properties of electron beam powder bed fusion (EB-PBF) fabricated tantalum parts and scaffolds was studied with tantalum powder reused more than 30 cycles. The results indicated that particle size distribution of the tantalum powder was nearly unchanged with the increase of cycle number, accordingly apparent density, tap density, and fluidity. While the powder became less spherical with increasing reuse times and some particles showed noticeable distortion and rough surface after being reused 25 times. Moreover, the oxygen content of the tantalum powder increased progressively with increasing reuse times, leading to the decrease of plasticity of the dense tantalum after 15 reuse cycles, and some potential micro-defects appeared in the tantalum samples fabricated from EB-PBF process. However, the tensile strength of dense tantalum parts was not sensitive to the number of uses within the research range.

Parameter identification of fractional‐order model with transfer learning for aging lithium‐ion batteries

The aging of lithium-ion batteries (LiBs) is inevitable during their operation owing to their irreversible side reactions. It is practical to capture only the dominant physicochemical processes with a physics-based model for engineering applications, as the degradation mechanism of LiBs is complex and interconnected. Numerous factors dramatically affect the performance of LiBs; thus, it is necessary to use the real-time operational data to estimate the model parameters online. The combination of the degradation mechanism and real-time data can effectively reduce the complexity of the parameter identification, and offers practicality from the perspective of engineering applications. In this study, a transfer learning method based on a back propagation neural network (BPNN) is introduced as the data-driven parameter identification method for the physics-based fractional-order model (FOM) of the LiBs. Accelerated aging tests are designed for commercial LiBs to reduce the experimental time, and the reference performance tests under different aging states are implemented to capture the degradation modes with increasing cycle numbers. The initial key parameters of the fresh FOM are estimated based on the BPNN with two hidden layers. The model-based transfer learning is proposed to estimate the key parameters of the aging FOM. The parameter identification results at different aging states are validated under various working loads. The good fit with the experimental data indicated that the parameters can be identified accurately using the transfer learning with BPNN. The outstanding results show the effectiveness of the parameter identification method based on transfer learning.

Effect of deep cryogenic treatment parameters on martensite multi-level microstructures and properties in a lath martensite/ferrite dual-phase steel

Abstract The microstructures and properties of a low-carbon 20CrNi2Mo–Al steel with martensite/ferrite dual-phase after deep cryogenic treatment (DCT) with different parameters were investigated. It was found that, with the increase of DCT processing time (QCT-h process) and cycle number (QCT-t process), the ferrite size was refined and its distribution became more homogeneous; the hardness and impact toughness were simultaneously increased. Especially in the QCT-t processes, the microstructure refinement and the property improvement were highly significant. The impact toughness of samples treated through 3-cycle DCT were 29.4% higher than the samples without DCT. Meanwhile, the martensite multi-level microstructures under different DCT parameters were thoroughly investigated by optical microscopy (OM), scanning electron microscopy (SEM), transmission electron microscopy (TEM), electron backscattering diffraction (EBSD) and X-ray diffraction (XRD). Results showed that the size of martensite packet (dp), block (db) and lath (dl) decreased with increasing total DCT processing time. The Hall-Petch relationship was applied to discuss the relationship between the martensite multi-level microstructure and the impact toughness. According to the deflection degree of the crack propagation path at different crystallographic orientations, the effective control unit of the impact toughness was revealed to be block (db). The quantity of its boundaries increased most significantly among the three kinds of subunits, which had the strongest hindering effect on crack propagation.

Haplotype-level DNA metabarcoding from freshwater macroinvertebrate community samples

DNA metabarcoding is a robust method for environmental impact assessments of freshwater ecosystems that enables the simultaneous multi-species identification of complex mixed community samples from different origins using extracellular and total genomic DNA. The development and evaluation of DNA metabarcoding protocols for haplotype level resolution require attention, specifically for basic population genetic applications, i.e., analysis to allow genetic diversity estimations and dispersal abilities of the species present in the bulk community samples. Various literature has proposed using DNA metabarcoding for population genetics, and few studies have provided preliminary applications and proof of concepts that always refer to particular taxa. However, further exploration and assessment of the laboratory and bioinformatics strategies are required to unlock the potential of metabarcoding-based population-level ecological assessments. Here, we assessed the ability to infer haplotype information of freshwater macroinvertebrate species from DNA metabarcoding community sequence. Using mock samples with known Sanger-sequenced haplotypes, we also assayed the effects of PCR cycle for the detection and reduction of spurious haplotypes obtained from DNA metabarcoding. We tested our haplotyping strategy on a mock sample containing 20 specimens from four species with known haplotypes based on the 658-bp Folmer region of the mitochondrial cytochrome c oxidase (mtCOI) gene. The read processing and denoising-step resulted in 14 zero-radius operational taxonomic units (ZOTUs) of 421-bp length, with 12 ZOTUs having 100% match with 12 of the mock haplotype sequences. The remaining eight haplotypes that were not detected from the DNA metabarcoding dataset were all the A. decemseta samples (0.01, 0.05, 0.10 ng/μL DNA template concentrations), two E. bulba (0.01 and 0.05 ng/μL), E. latifolium (0.01 ng/μL), and two K. tibialis (0.01 and 0.10 ng/μL). Given that most of the undetected samples had low concentrations, we report the influence of initial DNA template concentration on the amplification from a mock community sample. Our observation is in accordance with previous studies that reported that samples or taxa with low DNA template concentrations have lower detection probability. Accordingly, abundant taxa or samples with high biomass tend to have higher detection probabilities than those rare, smaller or have low biomass from mixed-community samples. The difference in biomass affects haplotypes' detection since most of the large specimens would be retained after read processing. Hence, these factors need to be addressed when metabarcoding-based haplotyping is to be used to infer abundance-based analysis for population genetics applications. The phylogenetic-based analysis (Fig. 1) revealed that the two ZOTUs without taxonomic matches clustered with one of the species from the mock sample. This supports our observation that only the samples with low concentration were unrepresented from the DNA metabarcoding data. Although we still reported false positive detections because two of the 14 ZOTUs failed to have a 100% match with the mock reference sequences, we could at least identify them as A. decemseta sequences based on the phylogenetic approach. Quality passing reads relatively increased with increasing cycle number, and the relative abundance of each ZOTUs was consistent for each cycle number. This suggests that increasing the cycle number, from 24 to 64, did not affect the relative abundance of quality passing filter reads. Our study demonstrated that DNA metabarcoding data could be used to infer intraspecific variability, showing promise for possible applications in population-based genetic studies. As DNA metabarcoding becomes more established and laboratory protocols and bioinformatics pipelines are continuously being developed, our proof of concept study demonstrated that the method could be used to infer intraspecific variability, showing promise for possible applications on population-based genetic studies.

Dynamic current cycles effect on the degradation characteristic of a H2/O2 proton exchange membrane fuel cell

The durability and reliability of H2/O2 proton exchange membrane fuel cell (PEMFC) is a key factor that prevents its wide application in the civil field. PEMFC inevitably experience different dynamic loading cycles according to different power switching requirements during practical operation. To explore the degradation behavior under different dynamic cycles, a single H2/O2 PEMFC with 50 cm2 active area was operated under the circulating current density from 100 mAcm−2 to 600 mAcm−2, 100 mAcm−2 to 800 mAcm−2, and 100 mAcm−2 to 1000 mAcm−2, respectively. The change of polarization curve, performance degradation at different current density, Cyclic Voltammetry (CV), Electrochemical Impedance Spectroscopy (EIS) were characterized to investigate the performance degradation over dynamic current cycles. Besides, the Scanning Electron Microscopy (SEM) was used to evaluate the degradation of catalyst layer. The results indicated that the degradation rate of the fuel cell performance increased corresponding to the cycle number, at 1200 mA/cm2, it with a total performance degradation rate of 11.83% after 2000 dynamic loading cycles with the circulating current density from 100 mAcm−2 to 600 mAcm−2. The degradation of electrochemical performance such as CV and EIS was consistent with that of fuel cell performance. The degradation rate is accelerated with the increase of loading cycle number and load step amplitude. What’ more, EIS provides additional sensitivity to differentiate catalyst layer degradation within PEMFC. Moreover, the degradation of the catalyst layer became much more severe under a larger load step amplitude.