Soil C:N:P stoichiometry in rhizosphere and non-rhizosphere of Pinus sylvestris var. mongolica forests.

The aim of this study was to reveal the stoichiometric characteristics of carbon, nitrogen and phosphorus in rhizosphere and non-rhizosphere soils of Pinus sylvestris var. mongolica in the Hulunbuir desert. We investigated the contents and stoichiometry of organic carbon, total nitrogen, and total phosphorus contents of rhizosphere and non-rhizosphere soils across different stand ages (28, 37 and 46 a) of P. sylvestris var. mongolica plantations, with P. sylvestris var. mongolica natural forest as the control. We analyzed the correlation between soils properties and soil stoichiometry. The results showed that rhizosphere effect significantly affected soil N:P, and stand age significantly affected soil organic carbon content in P. sylvestris var. mongolica plantation. Soil organic carbon content in plantation was significantly lower than that in natural forest. Soil organic carbon and total nitrogen contents of plantations in both rhizosphere and non-rhizosphere soils firstly decreased and then increased with increasing stand age, while total phosphorus firstly increased and then decreased in rhizosphere soils, and firstly decreased and then increased in non-rhizosphere soils. There was significant positive correlations between C:N and C:P in rhizosphere soils but not in non-rhizosphere soils, suggesting that higher synergistic rhizosphere soil N and P limitation. The mean N:P values of rhizosphere and non-rhizosphere soils were 4.98 and 8.40, respectively, indicating that the growth of P. sylvestris var. mongolica was restricted by soil N and the rhizosphere soils were more N-restricted. The C:N:P stoichiometry of rhizosphere and non-rhizosphere soils were significantly influenced by soil properties, with available phosphorus being the most important driver. The growth of P. sylvestris var. mongolica was limited by N in the Hulunbuir desert, and root system played an obvious role in enriching and maintaining soil nutrients. It was recommended that soil nitrogen should be supplemented appropriately during the growth stage of P. sylvestris var. mongolica plantation, and phosphorus should be supplemented appropriately according to the synergistic nature of nitrogen and phosphorus limitation.

Challenges in the treatment of pediatric Mycoplasma pneumoniae pneumonia.

Mycoplasma pneumoniae (MP) is an important cause of community-acquired pneumonia in children and young adolescents. Despite macrolide antibiotics effectiveness as a first-line therapy, persistence of fever and/or clinical deterioration sometimes may complicate treatment and may even lead to severe systemic disease. To date, there is no consensus on alternative treatment options, optimal dosage, and duration for treating severe, progressive, and systemic MP pneumonia after macrolide treatment failure. Macrolide-resistant MP pneumonia and refractory MP pneumonia are the two major complex conditions that are clinically encountered. Currently, the vast majority of MP isolates are resistant to macrolides in East Asia, especially China, whereas in Europe and North America, whereas in Europe and North America prevalence is substantially lower than in Asia, varying across countries. The severity of pneumonia and extrapulmonary presentations may reflect the intensity of the host's immune reaction or the dissemination of bacterial infection. Children infected with macrolide-resistant MP strains who receive macrolide treatment experience persistent fever with extended antibiotic therapy and minimal decrease in MP-DNA load. Alternative second-line agents such as tetracyclines (doxycycline or minocycline) and fluoroquinolones (ciprofloxacin or levofloxacin) may lead to clinical improvement after macrolide treatment failure in children. Refractory MP pneumonia reflects a deterioration of clinical and radiological findings due to excessive immune response against the infection. Immunomodulators such as corticosteroids and intravenous immunoglobulin (IVIG) have shown promising results in treatment of refractory MP pneumonia, particularly when combined with appropriate antimicrobials. Corticosteroid-resistant hyperinflammatory MP pneumonia represents a persistent or recrudescent fever despite corticosteroid therapy with intravenous methylprednisolone at standard dosage. CONCLUSION:  This report summarizes the clinical significance of macrolide-resistant and refractory MP pneumonia and discusses the efficacy and safety of alternative drugs, with a stepwise approach to the management of MP pneumonia recommended from the viewpoint of clinical practice. WHAT IS KNOWN: • Although MP pneumonia is usually a benign self-limited infection with response macrolides as first line therapy, severe life-threatening cases may develop if additional treatment strategies are not effectively implemented. • Macrolide-resistant and refractory MP pneumonia are two conditions that may complicate the clinical course of MP pneumonia, increasing the risk for exacerbation and even death. WHAT IS NEW: • This report summarizes the clinical relevance of macrolide-resistant and refractory MP pneumonia and discusses the efficacy and safety of alternative drug therapies. • A practical stepwise approach to the management of MP pneumonia is developed based on a comprehensive analysis of existing evidence and expert opinion.

Phthalate exposure and blood pressure in U.S. children aged 8-17 years (NHANES 2013-2018).

BACKGROUND: Current evidence from epidemiologic studies suggested that phthalate metabolites might be associated with blood pressure (BP) changes. However, the special relationship between phthalate metabolites and BP changes in children has not been clearly elucidated in existing researches. OBJECTIVES: We investigated the links between phthalate metabolites and various BP parameters, including systolic/diastolic BP, mean arterial pressure (MAP), and the presence of hypertension. METHODS: The population sample consisted of 1036 children aged 8 to 17 years from the 2013-2018 NHANES in the United States. High performance liquid chromatography-electrospray ionization-tandem mass spectrometry was used to measure urinary concentrations of 19 phthalate metabolites. Systolic/diastolic BP were derived from the average of three valid measurements, and MAP was calculated as (systolic BP + 2 × diastolic BP)/3. Hypertension was defined as mean systolic BP and/or diastolic BP that was ≥ 95th percentile for gender, age, and height reference. Linear regression, logistic regression, and weighted quantile sum (WQS) regression models were employed to assess the associations between phthalate exposure and systolic/diastolic BP, MAP, and hypertension. RESULTS: Ten of 19 phthalate metabolites including MCNP, MCOP, MECPP, MBP, MCPP, MEP, MEHHP, MiBP, MEOHP, and MBzP had detection frequencies > 85% with samples more than 1000. MCNP, MCOP, MECPP, MBP, MCPP, MEHHP, MiBP, MEOHP, and MBzP were generally negatively associated with systolic/diastolic BP and MAP, but not protective factors for hypertension. These associations were not modified by age (8-12 and 13-17 years) or sex (boys and girls). The above-mentioned associations were further confirmed by the application of the WQS analysis, and MCOP was identified as the chemical with the highest weight. CONCLUSION: Phthalate metabolites were associated with modest reductions in systolic/diastolic BP, and MAP in children, while appeared not protective factors for hypertension. Given the inconsistent results among existing studies, our findings should be confirmed by other cohort studies.

Patterns and environmental drivers of C, N, and P stoichiometry in the leaf-litter-soil system associated with Mongolian pine forests.

Ecological stoichiometry is an important approach to understand plant nutrient cycling and balance in the forest ecosystem. However, understanding of stoichiometric patterns through the leaf-litter-soil system of Mongolian pine among different stand origins is still scarce. Therefore, to reveal the variations in Mongolian pine carbon (C), nitrogen (N), and phosphorus (P) stoichiometry and stoichiometric homeostasis among different stand origins, we measured C, N, and P concentrations of leaves, litter, and soil, and analyzed the nutrient resorption efficiencies of leaves in differently aged plantations and natural forests from semi-arid and dry sub-humid regions. The results showed that (1) the stand origin had a significant effect on the C-N-P stoichiometry, and also significantly affected leaf N and P reabsorption efficiencies. Leaf N/P ratios indicated that Mongolian pine was co-limited by N and P in the NF, HB and HQ, and was mainly limited by P in MU. (2) With increasing stand age, C concentrations in the leaf-litter-soil system initially increased and then decreased, the N and P concentrations and reabsorption efficiencies in the leaf-litter-soil system were gradually increased. Overall, stand age had a significant effect on N concentrations, C/N and C/P ratios in the leaf-litter-soil system. (3) The C and N elements between the leaf-litter-soil system had a strong coupling relationship, and the P element between litter-soil had a strong coupling relationship. In addition, plantations exhibited greater N/P homeostasis than natural forests, and N/P exhibited greater homeostasis than N and P alone, which may be a nutrient utilization strategy for forests to alleviate N or P limitation. (4) Environmental factors have a significant influence on C-N-P stoichiometry in the leaf-litter-soil system, the most important soil properties and meteorological factors being soil water content and precipitation, respectively. These results will be essential to provide guidance for plantation restoration and management in desert regions.

Joint association of sedentary behavior and physical activity with pulmonary function.

BACKGROUND: Sedentary behavior may influence the respiratory health, but the joint effects of sedentary behavior and physical activity on pulmonary function remains poorly elucidated. We aimed to estimate the association between sedentary behavior and physical activity with pulmonary function. METHODS: A total of 12,343 participants aged 12-79 years were analyzed from the U.S. NHANES 2007-2012. Participants were categorized into 16 groups according to the cross-tabulation of sedentary behavior time (0-4.0, 4.1-8.0, 8.1-12.0, and > 12.0 h/day) and moderate or vigorous physical activity (MVPA) (0, 1-149, 150-299, and ≥ 300 min/week). Generalized linear models were used to test the association of sedentary behavior and MVPA with pulmonary function. RESULTS: Participants with sedentary behavior > 4.0 h/day were negatively related to FEV1 (forced expiratory volume in 1 s) (ß ranging from -0.015 to -0.009, p < 0.05). Compared with the reference group (0 min of MVPA and > 12.0 h/day of sedentary behavior), the negative association of sedentary behavior ≤ 8.0 h/day with FEV1 may be reduced through appropriate MVPA (ß ranging from 0.019 to 0.030, p < 0.05). For sedentary behavior > 8.0 h/day, even MVPA ≥ 300 min/week may not decrease the negative relationships. Similar results were also observed in FVC (forced vital capacity) (ß ranging from 0.018 to 0.030, p < 0.05). In participants aged ≥ 45 years, the associations were more notable. CONCLUSION: This study indicated the sedentary behavior ≤ 4.0 h/day was a relatively healthy lifestyle for pulmonary function. Only below 8.0 h/day of sedentary behavior, the negative association with pulmonary function may be reduced through appropriate MVPA.

Associations of legacy perfluoroalkyl and polyfluoroalkyl substances, alternatives, and isomers with gestational diabetes mellitus and glucose homeostasis among women conceiving through assisted reproduction in Shanghai, China.

Prior research has reported that perfluoroalkyl and polyfluoroalkyl substances (PFAS) may be linked to impaired glucose homeostasis in pregnant women. However, few studies have investigated PFAS alternatives and isomers, and even less is known about the association among women conceiving through assisted reproductive technology (ART). The prospective cohort study aimed to explore associations of legacy PFAS, alternatives and isomers with gestational diabetes mellitus (GDM) and glucose homeostasis during pregnancy among 336 women conceiving through ART. Nineteen PFAS, including nine linear legacy PFAS, four short-chain alternatives, four branched isomers, and two emerging PFAS alternatives, were determined in first-trimester maternal serum. Fasting plasma glucose (FPG), 1-h and 2-h glucose concentrations following the oral glucose tolerance test (OGTT), and glycated hemoglobin (HbA1c) were measured during the second trimester. After adjusting for confounding variables, nearly half of individual PFAS (10/19) and PFAS mixtures were correlated with increased GDM risk or elevated 2-h glucose levels. Among PFAS congeners, emerging PFAS alternatives, chlorinated perfluoroalkyl ether sulfonic acids (Cl-PFESAs), showed a notable association with impaired glucose homeostasis. For example, 6:2 Cl-PFESA exhibited a correlation with GDM (OR = 1.31, 95% CI = 1.02, 1.68) and 2-h glucose concentrations (ß = 0.22, 95% CI = 0.08, 0.36), and contributed most to the overall association with 2-h glucose concentrations. Compared to those diagnosed with male factor infertility, the associations were more pronounced in infertile women with reproductive endocrine diseases. We provide evidence that exposure to PFAS, especially emerging PFAS alternatives, may impair glucose homeostasis and increase the risk of GDM among women conceiving through ART.

Trends in Prevalence of Breastfeeding Initiation and Duration Among US Children, 1999 to 2018.

This cross-sectional study analyzes changes in the prevalence of breastfeeding initiation and duration among US children over the past 20 years.

Temporal Action Segmentation: An Analysis of Modern Techniques.

Temporal action segmentation (TAS) in videos aims at densely identifying video frames in minutes-long videos with multiple action classes. As a long-range video understanding task, researchers have developed an extended collection of methods and examined their performance using various benchmarks. Despite the rapid growth of TAS techniques in recent years, no systematic survey has been conducted in these sectors. This survey analyzes and summarizes the most significant contributions and trends. In particular, we first examine the task definition, common benchmarks, types of supervision, and prevalent evaluation measures. In addition, we systematically investigate two essential techniques of this topic, i.e., frame representation and temporal modeling, which have been studied extensively in the literature. We then conduct a thorough review of existing TAS works categorized by their levels of supervision and conclude our survey by identifying and emphasizing several research gaps.

Fungal complexity and stability across afforestation areas in changing desert environments.

The great achievements in combating desertification are attributed to large-scale afforestation, yet we lack verification of how the stability of the fungal community changes in afforestation areas in desert environments. Here, we present the fungal network structure from different niches (root and bulk soil) of plantations of Mongolian pine, a crucial species for afforestation introduced widely in desertification regions. We assessed changes in community complexity and stability of root-associated fungi (RAF) and soil fungi (SF) among different introduction sites: the Hulunbuir Desert (HB), the Horqin Desert (HQ) and the Mu Us Desert (MU). To illuminate the complexity and stability of the fungal network, the differences in topological properties, fungal function, and vegetation and environmental factors between introduction sites were fully considered. We showed that (1) the SF networks had more nodes and edges than the RAF networks. There was a lower ratio of negative:positive cohesion of RAF networks in HB and MU. For SF but not for RAF, across the three introduction sites, a higher modularity and ratio of negative:positive cohesion indicated higher stability. (2) Ectomycorrhizal (EcM) fungi were the dominant functional group in the RAF network (especially in HQ), and were only significantly correlated with vegetation factor. There was a higher relative abundance and number of OTUs of saprophytic fungi in the SF network and they showed positive correlations with soil nutrients. (3) RAF and SF network complexity and stability showed different responses to environmental and vegetation variables. The key determinant of the complexity and stability of the SF networks in Mongolian pine plantations was soil nutrients, followed by climate conditions. The composition and structure of the RAF community was closely related to host plants. Therefore, clarifying the complexity and stability of fungal communities in afforestation areas in changing desert environments is helpful for understanding the interactions between the environment, plants and fungi.

Precipitation pattern changed the content of non-structural carbohydrates components in different organs of Artemisia ordosica.

BACKGROUND: Non-structural carbohydrates (NSC) play a significant role in plant growth and defense and are an important component of carbon cycling in desert ecosystems. However, regarding global change scenarios, it remains unclear how NSCs in desert plants respond to changing precipitation patterns. [Methods] Three precipitation levels (natural precipitation, a 30% reduction in precipitation, and a 30% increase in precipitation) and two precipitation intervals levels (5 and 15 d) were simulated to study NSC (soluble sugar and starch) responses in the dominant shrub Artemisia ordosica. RESULTS: Precipitation level and interval interact to affect the NSC (both soluble sugar and starch components) content of A. ordosica. The effect of precipitation on NSC content and its components depended on extended precipitation interval. With lower precipitation and extended interval, soluble sugar content in roots increased and starch content decreased, indicating that A. ordosica adapts to external environmental changes by hydrolyzing root starch into soluble sugars. At 5 d interval, lower precipitation increased the NSC content of stems and especially roots. CONCLUSIONS: A. ordosica follows the "preferential allocation principle" to preferentially transport NSC to growing organs, which is an adaptive strategy to maintain a healthy physiological metabolism under drought conditions. The findings help understand the adaptation and survival mechanisms of desert vegetation under the changing precipitation patterns and are important in exploring the impact of carbon cycling in desert systems under global environmental change.

Temporal approach to identifying ectomycorrhizal community associated with Mongolian pine in a desert environment, northern China.

To date, the ectomycorrhizal (EM) fungi community has been widely investigated with aging plantations affected by the pedologic factors. However, knowledge on the effects of phenology induced by climatic factors across the age range remains poorly understood on an intra-annual scale. Here, we sampled the fine roots of Mongolian pine (Pinus sylvestris var. mongolica) plantations at different stages of the growing season (from April to September) across three stand ages (27, 34, and 44 yr) in the Mu Us Desert, Northwest China. We aim to disentangle the community composition and structure of EM fungi, as well as the impact of climate on EM fungi. We observed that the 173 distinct EM fungal operational taxonomic units (OTUs) were identified. Geopora, Inocybe, Tomentella, and Tuber were the most frequent, and their dominance was maintained as stand aging. The richness and community composition were not significantly different with stand aging (P > 0.05). Host phenology and stand age are two important factors that have shaped the EM fungal community. The growing stage affected the beta diversity of the EM fungal community more than stand age, and this variation of the EM fungal community was closely related to seasonal climate, particularly precipitation. This improved information will provide a theoretical basis for the reforestation and rehabilitation of the Mongolian pine plantations using mycorrhizal techniques. IMPORTANCE Ectomycorrhizal (EM) fungi are particularly important for host plants in a desert ecosystem. With a high degree of plasticity, EM fungi are largely influenced by host plant and environmental variables and fundamentally contribute to the ability of individuals to adapt to environmental changes. Therefore, the EM fungi are important for Mongolian pine (Pinus sylvestris var. mongolica) plantation in a desert ecosystem. Although previous studies have concluded that multiple endogenous and exogenous processes ultimately lead to species-specific temporal patterns in EM fungal populations. We still neglect the effect of host phenology on EM fungal activity. The significance of our study is the interplay between climate-driven EM fungi and plant phenology.

Quantifying cell viability through organelle ratiometric probing.

Detecting cell viability is crucial in research involving the precancerous discovery of abnormal cells, the evaluation of treatments, and drug toxicity testing. Although conventional methods afford cumulative results regarding cell viability based on a great number of cells, they do not permit investigating cell viability at the single-cell level. In response, we rationally designed and synthesized a fluorescent probe, PCV-1, to visualize cell viability under the super-resolution technology of structured illumination microscopy. Given its sensitivity to mitochondrial membrane potential and affinity to DNA, PCV-1's ability to stain mitochondria and nucleoli was observed in live and dead cells, respectively. During cell injury induced by drug treatment, PCV-1's migration from mitochondria to the nucleolus was dynamically visualized at the single-cell level. By extension, harnessing PCV-1's excellent photostability and signal-to-noise ratio and by comparing the fluorescence intensity of the two organelles, mitochondria and nucleoli, we developed a powerful analytical assay named organelle ratiometric probing (ORP) that we applied to quantitatively analyze and efficiently assess the viability of individual cells, thereby enabling deeper insights into the potential mechanisms of cell death. In ORP analysis with PCV-1, we identified 0.3 as the cutoff point for assessing whether adding a given drug will cause apparent cytotoxicity, which greatly expands the probe's applicability. To the best of our knowledge, PCV-1 is the first probe to allow visualizing cell death and cell injury under super-resolution imaging, and our proposed analytical assay using it paves the way for quantifying cell viability at the single-cell level.

Association of the Severity of Hypertensive Disorders in Pregnancy with Birthweight, Childhood Obesity, and Blood Pressure at Age 7.

We aimed to examine the effects of various severities of hypertensive disorders in pregnancy on birthweight, blood pressure (BP), and body mass index in offspring at age 7. In the China Labor and Delivery Survey and the United States Collaborative Perinatal Project (CPP), the relationship of the severity of hypertensive disorders and nutritional and cardiovascular outcomes in offspring was assessed using a multivariable logistic and general linear regression model. In both datasets, those with gestational hypertension were more likely to deliver large for gestational age (LGA) and macrosomia (adjusted odds ratios (aOR) ranged from 1.29 to 1.91), as well as low birth weight (LBW) neonates (aOR ranged from 1.23 to 3.56), compared with normotensive mothers. In the CPP, when gestational hypertension was further stratified into mild and severe, only those with mild gestational hypertension (the mild group) were more likely to deliver macrosomia and LGA (aOR ranged from 1.25 to 1.32). Others (severe gestational hypertension and preeclampsia/eclampsia) were closely related to LBW and small for gestational age (aOR ranged from 1.27 to 2.77). Moreover, children of mothers in the mild group tended to be overweight/obese and had elevated diastolic BP. We concluded that the severity of hypertensive disorders had different effects on birthweight, childhood overweight, and BP.

Association of parabens and bisphenols with lung function in children aged 5-12 years from Shanghai, China.

Epidemiological studies have reported potential effects of individual paraben or bisphenol exposure on lung function, but few studies have estimated their joint effects. We conducted a cross sectional survey to investigate the associations of parabens and bisphenols exposure with lung function in 205 children aged 5-12 years from Shanghai, China. Urinary concentrations of six parabens [methyl-, ethyl-, propyl-, butyl-, benzyl-, and heptyl-paraben (MeP, EtP, PrP, BuP, BzP, and HeP)] and seven bisphenols [bisphenol A (BPA), bisphenol AF (BPAF), bisphenol AP (BPAP), bisphenol B (BPB), bisphenol P (BPP), bisphenol S (BPS), and bisphenol Z (BPZ)] were assessed by the high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS). Lung function, including forced expiratory volume in 1 s (FEV1), forced vital capacity (FVC), FEV1/FVC, peak expiratory flow (PEF), and forced expiratory flow between 25% and 75% of forced vital capacity (FEF25-75%), was further measured. Linear regression, bayesian kernel machine regression (BKMR), and weighted quantile sum regression (WQS) evaluated the individual and joint relationships of the parabens and bisphenols with the lung function parameters. Further, the analysis was stratified by child sex. Parabens (MeP, EtP, PrP, and BuP) and bisphenols (BPA, BPAP, BPB, and BPS) with detection rates >75% were included for analyses. In linear regressions, parabens (MeP, PrP, and BuP) were generally negatively associated with FEV1, FVC, PEF, and FEF25-75%, but no associations for bisphenols were found. The association of parabens with lung function was more pronounced in girls. The aforementioned negative associations between parabens and lung function were confirmed by both the BKMR and WQS, with MeP being considered most heavily weighing chemical. Our findings suggested that exposure to parabens, either individuals or as a mixture, were associated with decreased lung function in children aged 5-12 years, and these associations were stronger among girls. Considering the cross-sectional study design, large longitudinal studies are warranted to confirm our findings.

Organophosphate esters, airway inflammation, and lung function among U.S. participants: A nationally representative cross-sectional study from NHANES, 2011-2012.

BACKGROUND: Human beings are widespread exposed to organophosphate esters (OPEs), but little is known about their effects on respiratory health. OBJECTIVES: To investigate the associations of exposure to OPEs with lung function and airway inflammation among U.S. participants from NHANES, 2011-2012. METHODS: A total of 1636 participants aged 6-79 years were included. Concentrations of OPE metabolites were measured in urine and lung function was assessed with spirometry. Fractional exhaled nitric oxide (FeNO) and blood eosinophils (B-Eos), two important inflammatory biomarkers, were also measured. Linear regression was performed to examine the relationships of OPEs with FeNO, B-Eos and lung function. Bayesian kernel machine regression (BKMR) was used to evaluate the joint associations between OPEs mixtures and lung function. RESULTS: Three of seven OPE metabolites had detection frequencies > 80 %, including diphenyl phosphate (DPHP), bis (1,3-dichloro-2-propyl) phosphate (BDCPP), bis-2-chloroethyl phosphate (BCEP). A 10-fold increase in DPHP concentrations were associated with 1.02 mL decreases in FEV1 (ß = -0.01, 95 % CIs = -0.02, -0.003) and FVC (ß = -0.01, 95 % CIs = -0.02, -0.003), respectively, and the similar, modest decreases were seen for BDCPP. For each 10-fold increase in BCEP concentration, FVC was also reduced by 1.02 mL (ß = -0.01, 95 % CIs = -0.02, -0.002). Moreover, the negative associations were only found in non-smokers aged >35 years. The aforementioned associations were confirmed by BKMR, but we cannot definitively identify a constituent driving this association. B-Eos was negatively associated with FEV1 and FEV1/FVC, but not with OPEs. No associations were found of FeNO with OPEs and lung function. CONCLUSIONS: Exposure to OPEs was associated with modest decrements in lung function, although the observed decrease in FVC and FEV1 is unlikely to be of real clinical relevance for the majority of subjects in this series. Moreover, those associations presented age and smoking status-dependent pattern. Unexpectedly, the adverse effect was not mediated by FeNO/B-Eos.

Constant Conversion Rate of Endolysosomes Revealed by a pH-Sensitive Fluorescent Probe.

Endolysosome dynamics plays an important role in autophagosome biogenesis. Hence, imaging the subcellular dynamics of endolysosomes using high-resolution fluorescent imaging techniques would deepen our understanding of autophagy and benefit the development of pharmaceuticals against endosome-related diseases. Taking advantage of the intramolecular charge-transfer mechanism, herein we report a cationic quinolinium-based fluorescent probe (PyQPMe) that exhibits excellent pH-sensitive fluorescence in endolysosomes at different stages of interest. A systematic photophysical and computational study on PyQPMe was carried out to rationalize its highly pH-dependent absorption and emission spectra. The large Stokes shift and strong fluorescence intensity of PyQPMe can effectively reduce the background noise caused by excitation light and microenvironments and provide a high signal-to-noise ratio for high-resolution imaging of endolysosomes. By applying PyQPMe as a small molecular probe in live cells, we were able to reveal a constant conversion rate from early endosomes to late endosomes/lysosomes during autophagy at the submicron level.

Quantifying cell viability through organelle ratiometric probing.

Detecting cell viability is crucial in research involving the precancerous discovery of abnormal cells, the evaluation of treatments, and drug toxicity testing. Although conventional methods afford cumulative results regarding cell viability based on a great number of cells, they do not permit investigating cell viability at the single-cell level. In response, we rationally designed and synthesized a fluorescent probe, PCV-1, to visualize cell viability under the super-resolution technology of structured illumination microscopy. Given its sensitivity to mitochondrial membrane potential and affinity to DNA, PCV-1's ability to stain mitochondria and nucleoli was observed in live and dead cells, respectively. During cell injury induced by drug treatment, PCV-1's migration from mitochondria to the nucleolus was dynamically visualized at the single-cell level. By extension, harnessing PCV-1's excellent photostability and signal-to-noise ratio and by comparing the fluorescence intensity of the two organelles, mitochondria and nucleoli, we developed a powerful analytical assay named organelle ratiometric probing (ORP) that we applied to quantitatively analyze and efficiently assess the viability of individual cells, thereby enabling deeper insights into the potential mechanisms of cell death. In ORP analysis with PCV-1, we identified 0.3 as the cutoff point for assessing whether adding a given drug will cause apparent cytotoxicity, which greatly expands the probe's applicability. To the best of our knowledge, PCV-1 is the first probe to allow visualizing cell death and cell injury under super-resolution imaging, and our proposed analytical assay using it paves the way for quantifying cell viability at the single-cell level.