confocal laser scanning microscope

  • 文章类型: Journal Article
    水生生态系统正面临越来越多的污染物暴露,对水生物种的稳定和健康构成潜在威胁。这项研究的重点是评估单次和联合暴露于80nm聚苯乙烯纳米塑料(PS-NP,0.1、1、10、20mg/L)和十溴二苯醚(BDE-209,300ng/L)生物蓄积性14天,增长,自由漂浮蕨类植物的光合作用和氧化应激。PS-NP主要积累在S.natans的表皮和毛状体中。同时,超氧化物歧化酶(SOD)和丙二醛(MDA)含量显著升高,而过氧化物酶(POD),过氧化氢酶(CAT),总抗氧化能力(T-AOC),相对增长率(RGR)下降。此外,沉水叶中的叶绿素含量下降,而在PS-NP浓度为0.1和1mg/L时,漂浮叶片中的含量增加。然而,随着PS-NP浓度的增加,淹没叶和漂浮叶的叶绿素含量均呈下降趋势。在PS-NP和BDE-209共同暴露下,MDA含量显著升高,而CAT,POD,SOD,T-AOC和RGR显著降低(p<0.05)。我们的结果显示,与单一曝光相比,在共同暴露于PS-NP和BDE-209的S.natans中观察到更明显的生态毒性效应。这些发现为淡水生态系统中BDE-209和PS-NPs可能存在的环境风险提供了宝贵的观点,有助于制定保护水生生物和生态系统的有效管理战略。这项研究强调了迫切需要了解新兴污染物对不同水生生物的毒性作用,强调保护和保存水生生态系统的重要性。
    Aquatic ecosystems are facing increasing exposure to pollutants, posing potential threats to the stability and wellness of aquatic species. This study focused on evaluating the impacts of single and combined exposure to 80 nm polystyrene nanoplastics (PS-NPs, 0.1, 1, 10, 20 mg/L) and decabromodiphenyl ether (BDE-209, 300 ng/L) for 14 days on the bioaccumulation, growth, photosynthesis and oxidative stress in the free-floating fern Salvinia natans. PS-NPs primarily accumulated in the epidermis and trichomes of S. natans. Meanwhile, the levels of superoxide dismutase (SOD) and malondialdehyde (MDA) were significantly increased, while those for peroxidase (POD), catalase (CAT), total antioxidant capacity (T-AOC), and relative growth rate (RGR) decreased. Furthermore, the chlorophyll contents in submerged leaves were decreased, while those in floating leaves were increased at PS-NPs concentrations of 0.1 and 1 mg/L. However, the chlorophyll contents in both submerged and floating leaves displayed a decreasing trend with increasing concentrations of PS-NPs. Under the co-exposure of PS-NPs and BDE-209, the contents of MDA were significantly elevated, whereas CAT, POD, SOD, T-AOC and RGR were significantly decreased (p < 0.05). Our results revealed that, compared to single exposure, more pronounced ecotoxic effects are observed in S. natans under co-exposure to PS-NPs and BDE-209. These findings offer valuable perspectives into the possible environmental risks of BDE-209 and PS-NPs in freshwater ecosystems, contributing to the development of effective management strategies for protecting aquatic organisms and ecosystems. This research highlights the urgent need to understand the toxic effects of emerging contaminants on different aquatic organisms, emphasizing the importance of protecting and preserving aquatic ecosystems.
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  • 文章类型: Journal Article
    利用高温共聚焦激光扫描显微镜(HT-CLSM)研究了Y对低冷却速率条件(10℃/min)下7Mo超级奥氏体不锈钢(7MoSASS)凝固过程的影响。原位观察结果表明,Y样品促进了奥氏体成核密度的增加。成核10秒后,Y样品的成核密度增加了149.53/mm2。此外,方差分析表明,Y的加入提高了低冷却速率条件下7MoSASS凝固组织的均匀性。Johnson-Mehl-Avrami-Kolmogorov(JMAK)理论结果表明,当固相比为0.5时,Y样品的成核模式从饱和位点成核过渡到饱和位点成核+Avrami成核。YAlO3与奥氏体的晶格失准值低,使其成为促进奥氏体早期形核的合适异质形核。在凝固的后期,Y积聚在残留的液相中,提供更大程度的成分过冷。SEM-EDS分析表明Y有助于7MoSASS凝固组织的细化,析出相的比例降低约7.5%。Cr和Mo是7MoSASS中表现出正偏析的主要元素,Y样品中Cr偏析率增加,而Mo偏析率下降。
    The effects of Y on the solidification process of 7Mo super austenitic stainless steel (7MoSASS) under low cooling rate conditions (10 °C/min) were investigated using high-temperature confocal laser scanning microscopy (HT-CLSM). The in situ observation results indicate that Y samples promote an increase in austenite nucleation density. After 10 s of nucleation, the nucleation density increased by 149.53/mm2 for the Y sample. Furthermore, variance analysis indicated that Y addition improved the uniformity of the 7MoSASS solidification microstructure under low cooling rate conditions. The Johnson-Mehl-Avrami-Kolmogorov (JMAK) theory results showed that when the solid phase ratio was 0.5, the nucleation mode of the Y sample transitioned from saturation site nucleation to saturation site nucleation + Avrami nucleation. YAlO3 has a low lattice disregistry value with austenite, making it a suitable heterogeneous nucleation core for promoting the early nucleation of austenite. During the late stages of solidification, Y accumulates in the residual liquid phase, providing a greater degree of compositional undercooling. SEM-EDS analysis showed that Y contributed to the refinement of the 7MoSASS solidification microstructure, with the proportion of precipitated phases decreasing by approximately 7.5%. Cr and Mo were the main elements exhibiting positive segregation in 7MoSASS, and the Cr segregation ratio increased in the Y sample, while the Mo segregation ratio decreased.
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  • 文章类型: Journal Article
    背景:胞外多糖(EPS)是口腔生物膜内细胞外基质的必需成分,并且受局部微环境的显着影响。本研究旨在调查两种不同的抗菌药物的影响,DJK-5和氯己定(CHX),对口腔生物膜中EPS体积和pH水平的影响。
    方法:将从两个供体获得的口腔生物膜在羟基磷灰石圆盘上培养3天,1周,2周,3周,和4周。随后,这些生物膜用10μg/mLDJK-5或2%CHX处理3分钟。这些抗菌治疗对死亡细菌比例等因素的影响,原位pH值,使用相应的荧光探针评估生物膜内的EPS体积。使用共聚焦激光扫描显微镜进行检查,并将所得图像集中在生物膜的上层和下层进行分析,分别。
    结果:DJK-5在生物膜的3天至4周持续时间内表现出比CHX更有效的杀菌作用(P<0.05)。生物膜是酸性的,上层的酸性低于下层(P<0.05)。两种抗菌剂都增加了pH值,但DJK-5的作用大于CHX(P<0.05)。与CHX相比,DJK-5处理的生物膜中EPS的体积显着降低,无论年龄或层(P<0.05)。
    结论:DJK-5在减少活菌和EPS体积方面表现出优异的效果,以及提高细胞外pH值,与氯己定相比。
    Exopolysaccharides (EPS) are essential constituents of the extracellular matrix within oral biofilms and are significantly influenced by the local microenvironment. This study aimed to investigate the impact of two distinct antimicrobial agents, DJK-5 and chlorhexidine (CHX), on the EPS volume and pH levels in oral biofilms.
    Oral biofilms obtained from two donors were cultured on hydroxyapatite discs for durations of 3 days, 1 week, 2 weeks, 3 weeks, and 4 weeks. Subsequently, these biofilms were subjected to treatment with 10 µg/mL DJK-5 or 2% CHX for 3 min. The impact of these antimicrobial treatments on factors such as the proportion of dead bacterial, in situ pH, and EPS volume within the biofilms was assessed using corresponding fluorescent probes. The examination was carried out utilizing confocal laser scanning microscopy, and the resulting images were analyzed with a focus on the upper and lower layers of the biofilm, respectively.
    DJK-5 exhibited a more potent bactericidal effect compared to CHX across the 3-day to 4-week duration of the biofilm (P < 0.05). The biofilms were acidic, with the upper layer being less acidic than the lower layer (P < 0.05). Both antimicrobial agents increased the pH, but DJK-5 had a greater effect than CHX (P < 0.05). The volume of EPS was significantly lower in DJK-5 treated biofilms compared to that of CHX, regardless of age or layer (P < 0.05).
    DJK-5 exhibited superior effectiveness in reducing viable bacteria and EPS volume, as well as in raising extracellular pH, as compared to chlorhexidine.
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  • 文章类型: Journal Article
    单聚合物纤维的去污力测定对于筛选用于洗衣的有效洗涤剂具有重要意义。但仍然具有挑战性。在这里,我们展示了一种使用共聚焦激光扫描显微镜(CLSM)量化单聚合物纤维去污力的新颖有效方法。它用于可视化单个聚合物纤维的除油过程,从而评估各种洗涤剂的去污力。选择了四种典型的表面活性剂进行比较,和含有多种组分的复合洗涤剂(例如,阴离子和非离子表面活性剂,酶)被证明是最有效,最强大的去污洗涤剂,因为棉纤维上超过50%的油可以很容易地去除。此外,三种纤维的除油过程(即棉花,粘胶,和聚酯)通过共聚焦显微镜成像和监测。发现单根聚酯纤维的去污力百分比超过70%,远高于棉和粘胶纤维(〜50%),这可能是由于其相对光滑的表面。与传统方法相比,CLSM成像方法对单根高分子纤维洗涤剂的去污力测定更为可行和有效。
    Detergency determination for single polymeric fibers is of significant importance to screening effective detergents for laundry, but remains challenging. Herein, we demonstrate a novel and effective method to quantify the detergency for single polymeric fibers using a confocal laser scanning microscope (CLSM). It was applied to visualize the oil-removing process of single polymeric fibers and thus assess the detergency of various detergents. Four typical surfactants were selected for comparison, and a compounded detergent containing multiple components (e.g., anionic and nonionic surfactants, enzymes) was demonstrated to be the most effective and powerful soil-removing detergent because more than 50% of oil on the cotton fiber could be easily removed. Moreover, the oil removal process of three kinds of fibers (i.e., cotton, viscose, and polyester) was imaged and monitored by confocal microscopy. It was found that the percentage of the detergency of a single polyester fiber exceeded 70%, which is much higher than that of cotton and viscose fibers (~50%), which may be due to its relatively smooth surface. Compared to traditional methods, the CLSM imaging method is more feasible and effective to determine the detergency of detergents for single polymeric fibers.
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  • 文章类型: Journal Article
    This study aimed at evaluating the in vitro antibacterial efficacy of photodynamic therapy (PDT) on planktonic E. faecalis and its biofilm in the root canal of infected deciduous teeth.
    Forty root canals of maxillary deciduous anterior teeth were enlarged up to #35 K-file and inoculated with E. faecalis for 21 days. The root canals were randomly assigned into four groups (n = 10): The normal saline group (control), 1% NaClO group, PDT group, and the 1% NaClO + PDT group. Paper point samples were obtained at baseline (S1) and after treatment (S2). The colony-forming units (CFU) were counted, and the bacterial growth rate calculated. From each subgroup, 5 samples were randomly selected after treatment and a scanning laser confocal microscope (CLSM) used to determine the distribution of dead / living bacteria on the biofilm surface of each subgroup. A scanning electron microscope (SEM) was used to observe bacterial morphologies in the root canal walls of the remaining 5 samples in each subgroup. The Kruskal-Wallis test and Dunn test with boferroni adjustment were used to analyze the effect of the different treatment techniques on the E. faecalis in root canals.
    Compared to the saline group, PDT significantly reduced bacterial counts in the root canal (p < 0.05). The CFU counts were lowest (p < 0.05) in the 1% NaClO and in 1% NaClO + PDT groups. The rate of bacterial death on the surface of the biofilm in the PDT group was significantly increased after treatment (p < 0.05), and the rate of bacterial death was highest in 1%NaClO group and 1%NaClO + PDT group (p < 0.05).
    PDT has an antibacterial activity against E. faecalis in the root canal of deciduous teeth. Its activity against planktonic E. faecalis is better than the activity on the intact biofilm. The antibacterial activity of PDT on E. faecalis in root canals of deciduous teeth is lower compared to that of 1% NaClO.
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  • 文章类型: Journal Article
    目的:动态评估四种根管封闭剂对牙本质小管内生物膜的杀伤作用。
    方法:为牙本质小管感染准备牙本质块。培养粪肠球菌VP3-181和来自两个供体的多物种细菌。经过3天的孵化,感染的牙本质标本用无菌水冲洗1分钟并接受治疗。此外,将来自供体1的多物种细菌孵育3周以允许生物膜成熟,然后对样本进行处理。Gutta-percha处理的牙本质标本包括对照组。根管密封剂(生物陶瓷密封剂:EndoSequenceBC密封剂,ProRootEndoSealer,或GuttaFlowBioseal;以及传统的基于有机硅的密封剂:Guttaflow2)散布到牙本质的管壁上。在7、30或60天用共聚焦激光扫描显微镜检查样本。
    结果:在3日龄生物膜组中,从治疗的前7天到治疗的60天,所有封闭剂的死亡细菌比例显着下降(p<0.05)。在3周大的生物膜组中,与生物膜的7天暴露相比,暴露于生物陶瓷密封剂60天导致更显著的死亡细菌(p<0.05)。生物陶瓷密封剂在杀灭细菌方面比GuttaFlow2密封剂更有效(p<0.05)。
    结论:硅酸钙基密封剂对牙本质小管内的生物膜显示出良好的抗菌作用,尤其是在年轻生物膜的第一周。对于所检查的根管封闭剂,没有观察到针对年轻牙本质小管生物膜的实质性抗菌活性。
    结论:所检查的生物陶瓷根管封闭剂在长期暴露于年轻生物膜后显示出最小的额外抗菌作用。
    OBJECTIVE: To dynamically evaluate the effect of four root canal sealers on the killing of biofilms within dentinal tubules.
    METHODS: Dentin blocks were prepared for infection of the dentinal tubules. Enterococcus faecalis VP3-181 and multi-species bacteria from two donors were cultured. After 3 days of incubation, the infected dentin specimens were rinsed with sterile water for 1 min and subjected to treatment. Additionally, multi-species bacteria from donor 1 were incubated for 3 weeks to allow biofilm maturation and then the specimens were subjected to treatment. Gutta-percha-treated dentin specimens comprised the control group. A root canal sealer (bioceramic sealers: EndoSequence BC Sealer, ProRoot Endo Sealer, or GuttaFlow Bioseal; and a traditional silicone-based sealer: Guttaflow 2) was spread onto the canal walls of the dentin. The specimens were examined with confocal laser scanning microscopy at 7, 30, or 60 days.
    RESULTS: In the 3-day-old biofilm group, the proportion of killed bacteria decreased significantly from the first 7 days of treatment to 60 days of treatment for all sealers (p < 0.05). In the 3-week-old biofilm group, 60 days of exposure to bioceramic sealers resulted in more significant dead bacteria than 7-day exposures of the biofilms (p < 0.05). Bioceramic sealers were more effective in killing bacteria than the GuttaFlow 2 sealer (p < 0.05).
    CONCLUSIONS: Calcium silicate-based sealers showed good antimicrobial effects against biofilms within dentinal tubules, especially in the first week in young biofilms. There is no substantive antibacterial activity observed for the examined root canal sealers against young dentinal tubule biofilms.
    CONCLUSIONS: The bioceramic root canal sealers examined demonstrate minimal additional antibacterial effects after long-term exposure to young biofilms.
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  • 文章类型: Journal Article
    To evaluate early degradation at resin-dentin interface using non-invasive swept-source optical coherence tomography (SS-OCT) and confocal laser scanning microscope (CLSM).
    Self-etch adhesives and resin-composites containing bisphenol-glycidyl-dimethacrylate (Bis-GMA), which is one of the most widely used monomers in restorative materials, were investigated in this study. Forty cervical cavities were prepared in bovine incisors and applied by the adhesive with/without Bis-GMA (AdhesiveBG/Adhesive), filled by the resin with/without Bis-GMA (ResinBG/Resin) and then challenged by cariogenic biofilm (37 °C, 24 h). Gap Formation and dentin demineralization around resin-composites were observed by SS-OCT and CLSM.
    Three types of resin-dentin interfacial degradation could be detected from SS-OCT. Type I-dentin demineralization around resin without gap, showing feather-shaped dark zones without bright scattered lines at resin-dentin interfaces. Type II-dentin demineralization around resin with adhesive-dentin bonded gaps, showing feather-shaped dark zones with bright scattered lines at resin-dentin interfaces. Type III-dentin demineralization around resin with adhesive-dentin debonded gaps, showing edge-shaped dark zones with bright scattered lines at resin-dentin interface. From CLSM, the groups were compared in gap scale (GS), gap depth (GD), gap width (GW) and dentin wall lesion depth (WLD). Bis-GMA-containing adhesive groups showed significantly lower GS than Bis-GMA-free adhesive groups. Bis-GMA-containing resin groups showed significantly lower WLD than Bis-GMA-free resin groups. However, they did not show significant differences in GD and GW.
    Three types of early degradation at resin-dentin interface can be noninvasively detected by SS-OCT. Bis-GMA-containing and Bis-GMA-free restorative materials show differences in gap scale and dentin wall lesion depth.
    SS-OCT can nondestructively detect early resin-dentin interfacial degradation. Gap scale can be used as a parameter to evaluate the risk factor of gaps.
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  • 文章类型: Journal Article
    BACKGROUND: To evaluate the in vitro antibacterial effect of flavonoids-rich Ziziphus jujuba Mill. extract (FZM) against the formation of bacterial biofilms (BBFs) in Staphylococcus aureus.
    RESULTS: FZM can effectively inhibit the formation of S. aureus biofilms in vitro. Morphological observation showed a decrease in both biofilm adhesion and thickness. Results of confocal laser scanning microscopy used to detect the thickness of the BBFs showed that FZM treatment reduced the thickness of the BBFs. Furthermore, after the Image-Pro Plus v.6.0 analysis of the fluorescence intensity, FZM treatment reduced the thickness of the BBFs as well as the proportion of green fluorescence. Scanning electron microscopy showed that FZM can disrupt the channels available for substance exchange in the biofilm, thus exposing the bacterial cells and damaging its three-dimensional structures.
    CONCLUSIONS: FZM can inhibit biofilm formation, improve the bacterial pH environment, and eliminate the hydrophobic effect of reactive oxygen species and flavonoids.
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  • 文章类型: Comparative Study
    Acidithiobacillus thiooxidans (A. thiooxidans) is often used for sulfur-bearing ores bioleaching, but its adaptive mechanism to harsh environments remains unclear. Here, we explored the adaptive mechanism of A. thiooxidans in the process of low-grade chalcopyrite bioleaching based on the physiology and comparative transcriptome analysis. It was indicated that A. thiooxidans maintains intracellular pH homeostasis by regulating unsaturated fatty acids, especially cyclopropane fatty acids, intracellular ATP, amino acid metabolism, and antioxidant factors. Comparative transcriptome analysis indicated that the key genes involved in sulfur oxidation, sor and soxABXYZ, were significantly up-regulated, generating more energy to resist extreme environmental stress by more active sulfur metabolism. Confocal laser scanning microscope analysis found that down-regulation of flagellar-related genes was likely to promote the biofilm formation. System-level understanding of leaching microorganisms under extreme stress can contribute to the evolution of these extremophiles via genetic engineering modification work, which further improves bioleaching in future.
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  • 文章类型: Journal Article
    The present study deals with the proteomics analysis of crude squid ink isolated from Sepia esculenta for their antibacterial, antifungal, antibiofilm and cytotoxic properties. To achieve this, SDS-PAGE was used to separate proteins as bands, In-gel trypsin digested and analyzed by MALDI-TOF mass spectrometry. A total of 4 bands were identified by MASCOT search analysis namely astacin-like squid metalloprotease type I (ASMT-I), 70 kDa neurofilament protein (NP), uncharacterized protein LOC106181966 isoform X1 (UP-Iso-X1) and Ommochrome-binding protein (Oc-BP). Further, the obtained crude squid proteins were subjected to antimicrobial and antibiofilm activities against pathogenic bacterial and fungal strains respectively. Further, MTT assay was also carried out to deliberately explain the cytotoxic ability of crude squid ink protein against MCF-7 breast cancer cell lines. The results from the study revealed that, the proteins are shown to be toxic against pathogenic strains and breast cancer cell lines in a dose-dependent manner. More importantly, the proteins are well enough to eradicate biofilms substantiated by light and confocal laser scanning microscopic observations. Altogether, the crude squid ink proteins hampered the growth of breast cancer cells with an IC50 value of 65.3 ± 0.46 μg mL-1. In conclusion, it is believed that the proteins from crude squid ink will provide new insights in hampering bacterial biofilms and cancer in near future.
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