Nomogram for predicting invasive lung adenocarcinoma in small solitary pulmonary nodules.

Background: This study aimed to construct a clinical prediction model and nomogram to differentiate invasive from non-invasive pulmonary adenocarcinoma in solitary pulmonary nodules (SPNs). Method: We analyzed computed tomography and clinical features as well as preoperative biomarkers in 1,106 patients with SPN who underwent pulmonary resection with definite pathology at Qilu Hospital of Shandong University between January 2020 and December 2021. Clinical parameters and imaging characteristics were analyzed using univariate and multivariate logistic regression analyses. Predictive models and nomograms were developed and their recognition abilities were evaluated using receiver operating characteristic (ROC) curves. The clinical utility of the nomogram was evaluated using decision curve analysis (DCA). Result: The final regression analysis selected age, carcinoembryonic antigen, bronchus sign, lobulation, pleural adhesion, maximum diameter, and the consolidation-to-tumor ratio as associated factors. The areas under the ROC curves were 0.844 (95% confidence interval [CI], 0.817-0.871) and 0.812 (95% CI, 0.766-0.857) for patients in the training and validation cohorts, respectively. The predictive model calibration curve revealed good calibration for both cohorts. The DCA results confirmed that the clinical prediction model was useful in clinical practice. Bias-corrected C-indices for the training and validation cohorts were 0.844 and 0.814, respectively. Conclusion: Our predictive model and nomogram might be useful for guiding clinical decisions regarding personalized surgical intervention and treatment options.

Starch and sucrose metabolism plays an important role in the stem development in Medicago sativa.

The forage quality of alfalfa (Medicago sativa ) stems is greater than the leaves. Sucrose hydrolysis provides energy for stem development, with starch being enzymatically converted into sucrose to maintain energy homeostasis. To understand the physiological and molecular networks controlling stem development, morphological characteristics and transcriptome profiles in the stems of two alfalfa cultivars (Zhungeer and WL168) were investigated. Based on transcriptome data, we analysed starch and sugar contents, and enzyme activity related to starch-sugar interconversion. Zhungeer stems were shorter and sturdier than WL168, resulting in significantly higher mechanical strength. Transcriptome analysis showed that starch and sucrose metabolism were significant enriched in the differentially expressed genes of stems development in both cultivars. Genes encoding INV , bglX , HK , TPS and glgC downregulated with the development of stems, while the gene encoding was AMY upregulated. Weighted gene co-expression network analysis revealed that the gene encoding glgC was pivotal in determining the variations in starch and sucrose contents between the two cultivars. Soluble carbohydrate, sucrose, and starch content of WL168 were higher than Zhungeer. Enzyme activities related to sucrose synthesis and hydrolysis (INV, bglX, HK, TPS) showed a downward trend. The change trend of enzyme activity was consistent with gene expression. WL168 stems had higher carbohydrate content than Zhungeer, which accounted for more rapid growth and taller plants. WL168 formed hollow stems were formed during rapid growth, which may be related to the redistribution of carbohydrates in the pith tissue. These results indicated that starch and sucrose metabolism play important roles in the stem development in alfalfa.

Clinical Study of a Four-Step Program for the Treatment of Plantar Fasciitis with Bone Spurs.

OBJECTIVE: The most common causes of plantar and heel pain are plantar fasciitis and calcaneal spurs, and they often co-exist. Surgery is a recognized treatment for refractory plantar fasciitis. However, few studies have proposed treatment options for patients with metatarsophalangeal fasciitis with bone spurs. Accordingly, this study's purpose was to propose a four-step surgical regimen, and to improve the surgical outcome of plantar fasciitis with osteophytes and to establish a procedure for surgical treatment. METHODS: Retrospective analysis of 45 patients suffering from plantar fasciitis with bone spurs from 2020 to 2023. All patients underwent a four-step procedure, including plantar fascia release, calcaneal spur grinding, inflammatory tissue removal, and calcaneal burr decompression. The imaging parameters and functional scores were recorded before and after the operation. The objective evaluation included the measurement of calcaneal spur length on radiographs. Clinical evaluation included the American Orthopaedic Foot and Ankle Society (AOFAS), the Visual Analog Scale (VAS), and the Foot and Ankle Outcome Scale (FAOS). Measurement data that conformed to normal distribution were expressed as (x2 ± s), and pre-and postoperative AOFAS, FAOS, and VAS scores were compared using repeated-measures ANOVA, and preoperative and postoperative spur lengths were compared using paired t-tests. RESULTS: The 45 patients were followed up for 3 to 30 months, (17.72 ± 8.53) months, at final follow-up, the patient's AOFAS score improved from preoperative (74.93 ± 5.56) to (94.78 ± 3.98), FAOS score increased from preoperative (76.42 ± 3.37) to (96.16 ± 2.74), the VAS score decreased from (3.18 ± 0.54) to (1.07 ± 1.20) (p < 0.05), the length of spur decreased from (0.72 ± 1.81) cm to (0.23 ± 1.19) cm, and there were significant differences before and after operation (p < 0.05). CONCLUSION: The four-step surgical regimen is an appropriate and effective surgical procedure to treat plantar fasciitis with bone spurs.

Fabrication, structure, characterization and emulsion application of citrate agar.

In this study, citric acid successfully reacted with agar through the dry heat method, and citrate agar (CA) gel was used to stabilize O/W emulsions. The mechanisms of the CA structure and emulsion pH that affected emulsion stabilization were analyzed, and the application of CA gel emulsion (CAGE) was explored. Compared with native agar (NA), CA showed lower gel strength, higher transparency, and higher water contact angle. These changes indicate that a cross-linking reaction occurred, and it was demonstrated via FTIR and NMR. The emulsion properties were evaluated using particle size, ζ-potential, and the emulsification activity index. Results showed that CAGEs had a smaller particle size and lower ζ-potential than the native agar gel emulsion (NAGE). Meanwhile, confocal laser scanning microscopy confirmed that the CA gels stabilized the emulsions by forming a protective film around the oil droplets. Stability experiments revealed that CAGE (prepared with CA gel [DS = 0.145]) exhibited better stability than NAGE in the pH range of 3-11, and the rheological results further confirmed that the stability of the emulsions was influenced by the network structure and oil droplet interaction forces. Afterward, the application prospect of CAGE was evaluated by encapsulating vitamin D3 and curcumin.

Construction and validation of a predictive model of invasive adenocarcinoma in pure ground-glass nodules less than 2 cm in diameter.

OBJECTIVES: In this study, we aimed to develop a multiparameter prediction model to improve the diagnostic accuracy of invasive adenocarcinoma in pulmonary pure glass nodules. METHOD: We included patients with pulmonary pure glass nodules who underwent lung resection and had a clear pathology between January 2020 and January 2022 at the Qilu Hospital of Shandong University. We collected data on the clinical characteristics of the patients as well as their preoperative biomarker results and computed tomography features. Thereafter, we performed univariate and multivariate logistic regression analyses to identify independent risk factors, which were then used to develop a prediction model and nomogram. We then evaluated the recognition ability of the model via receiver operating characteristic (ROC) curve analysis and assessed its calibration ability using the Hosmer-Lemeshow test and calibration curves. Further, to assess the clinical utility of the nomogram, we performed decision curve analysis. RESULT: We included 563 patients, comprising 174 and 389 cases of invasive and non-invasive adenocarcinoma, respectively, and identified seven independent risk factors, namely, maximum tumor diameter, age, serum amyloid level, pleural effusion sign, bronchial sign, tumor location, and lobulation. The area under the ROC curve was 0.839 (95% CI: 0.798-0.879) for the training cohort and 0.782 (95% CI: 0.706-0.858) for the validation cohort, indicating a relatively high predictive accuracy for the nomogram. Calibration curves for the prediction model also showed good calibration for both cohorts, and decision curve analysis showed that the clinical prediction model has clinical utility. CONCLUSION: The novel nomogram thus constructed for identifying invasive adenocarcinoma in patients with isolated pulmonary pure glass nodules exhibited excellent discriminatory power, calibration capacity, and clinical utility.

The role of adenocarcinoma subtypes and immunohistochemistry in predicting lymph node metastasis in early invasive lung adenocarcinoma.

BACKGROUND: Identifying lymph node metastasis areas during surgery for early invasive lung adenocarcinoma remains challenging. The aim of this study was to develop a nomogram mathematical model before the end of surgery for predicting lymph node metastasis in patients with early invasive lung adenocarcinoma. METHODS: In this study, we included patients with invasive lung adenocarcinoma measuring ≤ 2 cm who underwent pulmonary resection with definite pathology at Qilu Hospital of Shandong University from January 2020 to January 2022. Preoperative biomarker results, clinical features, and computed tomography characteristics were collected. The enrolled patients were randomized into a training cohort and a validation cohort in a 7:3 ratio. The training cohort was used to construct the predictive model, while the validation cohort was used to test the model independently. Univariate and multivariate logistic regression analyses were performed to identify independent risk factors. The prediction model and nomogram were established based on the independent risk factors. Recipient operating characteristic (ROC) curves were used to assess the discrimination ability of the model. Calibration capability was assessed using the Hosmer-Lemeshow test and calibration curves. The clinical utility of the nomogram was assessed using decision curve analysis (DCA). RESULTS: The overall incidence of lymph node metastasis was 13.23% (61/461). Six indicators were finally determined to be independently associated with lymph node metastasis. These six indicators were: age (P < 0.001), serum amyloid (SA) (P = 0.008); carcinoma antigen 125 (CA125) (P = 0. 042); mucus composition (P = 0.003); novel aspartic proteinase of the pepsin family A (Napsin A) (P = 0.007); and cytokeratin 5/6 (CK5/6) (P = 0.042). The area under the ROC curve (AUC) was 0.843 (95% CI: 0.779-0.908) in the training cohort and 0.838 (95% CI: 0.748-0.927) in the validation cohort. the P-value of the Hosmer-Lemeshow test was 0.0613 in the training cohort and 0.8628 in the validation cohort. the bias of the training cohort corrected C-index was 0.8444 and the bias-corrected C-index for the validation cohort was 0.8375. demonstrating that the prediction model has good discriminative power and good calibration. CONCLUSIONS: The column line graphs created showed excellent discrimination and calibration to predict lymph node status in patients with ≤ 2 cm invasive lung adenocarcinoma. In addition, the predictive model has predictive potential before the end of surgery and can inform clinical decision making.

Prognostic significance and survival benefits of postoperative adjuvant chemotherapy in patients with stage IA lung adenocarcinoma with non-predominant micropapillary components.

BACKGROUND: The prognostic significance of adjuvant chemotherapy (ACT) for patients with stage IA micropapillary non-predominant (MPNP) lung adenocarcinoma (LUAD) remains unknown. This study aimed to investigate the effects of postoperative ACT in patients with stage IA MPNP-LUAD. METHODS: A total of 149 patients with pathological stage IA MPNP-LUAD who underwent surgery at our center were retrospectively analyzed. Propensity score matching (PSM) analysis was conducted to reduce potential selection bias. Kaplan-Meier analyses were used to assess the impact of ACT on recurrence-free survival (RFS), overall survival (OS), and disease-specific survival (DSS). Subgroup analyses were performed for the survival outcomes based on the percentage of micropapillary components. Cox proportional hazards regression analyses were applied to identify risk factors associated with survival. RESULTS: The receipt or non-receipt of postoperative ACT had no significant effect on RFS, OS, and DSS among all enrolled patients with stage IA MPNP-LUAD (P > 0.05). For patients with a micropapillary component > 5%, the 5-year rates of RFS, OS, and DSS were significantly higher in the ACT group compared to the observation group, both before and after PSM (P < 0.05). However, the differences between the two groups were not significant for patients with a micropapillary component ≤ 5% (P > 0.05). The resection range (HR = 0.071; 95% CI: 0.020-0.251; P < 0.001), tumor size (HR = 2.929; 95% CI: 1.171-7.330; P = 0.022), and ACT (HR = 0.122; 95% CI: 0.037-0.403; P = 0.001) were identified as independent prognostic factors for RFS through Cox regression analysis. CONCLUSION: Patients with stage IA MPNP-LUAD who have a micropapillary component greater than 5% might benefit from postoperative ACT, while those with a micropapillary component ≤ 5% did not appear to derive the same benefit from postoperative ACT.

Nomogram combining clinical and radiological characteristics for predicting the malignant probability of solitary pulmonary nodules measuring ≤ 2 cm.

Background: At present, how to identify the benign or malignant nature of small (≤ 2 cm) solitary pulmonary nodules (SPN) are an urgent clinical challenge. This retrospective study aimed to develop a clinical prediction model combining clinical and radiological characteristics for assessing the probability of malignancy in SPNs measuring ≤ 2 cm. Method: In this study, we included patients with SPNs measuring ≤ 2 cm who underwent pulmonary resection with definite pathology at Qilu Hospital of Shandong University from January 2020 to December 2021. Clinical features, preoperative biomarker results, and computed tomography characteristics were collected. The enrolled patients were randomized at a ratio of 7:3 into a training cohort of 775 and a validation cohort of 331. The training cohort was used to construct the predictive model, while the validation cohort was used to test the model independently. Univariate and multivariate logistic regression analyses were performed to identify independent risk factors. The prediction model and nomogram were established based on the independent risk factors. The receiver operating characteristic (ROC) curve was used to evaluate the identification ability of the model. The calibration power was evaluated using the Hosmer-Lemeshow test and calibration curve. The clinical utility of the nomogram was also assessed by decision curve analysis (DCA). Result: A total of 1,106 patients were included in this study. Among them, the malignancy rate of SPNs was 85.08% (941/1,106). We finally identified the following six independent risk factors by logistic regression: age, carcinoembryonic antigen, nodule shape, calcification, maximum diameter, and consolidation-to-tumor ratio. The area under the ROC curve (AUC) for the training cohort was 0.764 (95% confidence interval [CI]: 0.714-0.814), and the AUC for the validation cohort was 0.729 (95% CI: 0.647-0.811), indicating that the prediction accuracy of nomogram was relatively good. The calibration curve of the predictive model also demonstrated a good calibration in both cohorts. DCA proved that the clinical prediction model was useful in clinical practice. Conclusion: We developed and validated a predictive model and nomogram for estimating the probability of malignancy in SPNs measuring ≤ 2 cm. With the application of predictive models, thoracic surgeons can make more rational clinical decisions while avoiding overtreatment and wasting medical resources.

Clinical characteristics and risk factors of lower extremity amputation in the diabetic inpatients with foot ulcers.

Objectives: To analyze clinical characteristics of the diabetic inpatients with foot ulcers and explore the risk factors of lower extremity amputation (LEA) in West China Hospital of Sichuan University. Methods: A retrospective analysis was performed based on the clinical data of the patients with diabetic foot ulcer (DFU) hospitalized in West China Hospital of Sichuan University from January 1, 2012 to December 31, 2020. The DFU patients were divided into three groups: non-amputation, minor amputation, and major amputation groups. The ordinal logistic regression analysis was used to identify the risk factors for LEA. Results: 992 diabetic patients (622 males and 370 females) with DFU were hospitalized in the Diabetic Foot Care Center of Sichuan University. Among them, 72 (7.3%) (55 minor amputations and 17 major amputations) cases experienced amputation, and 21(2.1%) refused amputation. Excluding the patients who refused amputation, the mean age and duration of diabetes of and HbA1c the 971 patients with DFU, were 65.1 ± 12.3 years old, 11.1 ± 7.6 years, and 8.6 ± 2.3% respectively. The patients in the major amputation group were older and had longer course of diabetes for a longer period of time than those in the non-amputation and minor amputation groups. Compared with the non-amputation patients (55.1%), more patients with amputation (minor amputation (63.5%) and major amputation (88.2%)) suffered from peripheral arterial disease (P=0.019). The amputated patients had statistically lower hemoglobin, serum albumin and ankle brachial index (ABI), but higher white blood cell, platelet counts, fibrinogen and C-reactive protein levels. The patients with amputation had a higher incidence of osteomyelitis (P = 0.006), foot gangrene (P < 0.001), and a history of prior amputations (P < 0.001) than those without amputation. Furthermore, a history of prior amputation (odds ratio 10.194; 95% CI, 2.646-39.279; P=0.001), foot gangrene (odds ratio 6.466; 95% CI, 1.576-26.539; P=0.010) and ABI (odds ratio 0.791; 95% CI, 0.639-0.980; P = 0.032) were significantly associated with LEAs. Conclusions: The DFU inpatients with amputation were older with long duration of diabetes, poorly glycemic control, malnutrition, PAD, severe foot ulcers with infection. A history of prior amputation, foot gangrene and a low ABI level were the independent predictors of LEA. Multidisciplinary intervention for DFU is essential to avoid amputation of the diabetic patients with foot ulcer.

mRNA therapeutics for disease therapy: principles, delivery, and clinical translation.

Messenger RNA (mRNA) has become a key focus in the development of therapeutic agents, showing significant potential in preventing and treating a wide range of diseases. The COVID-19 pandemic in 2020 has accelerated the development of mRNA nucleic therapeutics and attracted significant investment from global biopharmaceutical companies. These therapeutics deliver genetic information into cells without altering the host genome, making them a promising treatment option. However, their clinical applications have been limited by issues such as instability, inefficient in vivo delivery, and low translational efficiency. Recent advances in molecular design and nanotechnology have helped overcome these challenges, and several mRNA formulations have demonstrated promising results in both animal and human testing against infectious diseases and cancer. This review provides an overview of the latest research progress in structural optimization strategies and delivery systems, and discusses key considerations for their future clinical use.

[Characteristics of Inflammatory Markers in Diabetic Foot Patients and Their Relationship With Prognosis of Diabetic Foot Ulcers]. / ç³–å°¿ç— è¶³æ‚£è€ ç‚Žç—‡æ ‡å¿—ç‰©ç‰¹å¾åŠå ¶ä¸Žè¶³æºƒç–¡é¢„åŽå ³ç³».

Objective: To explore the characteristics of baseline inflammatory markers in diabetic foot patients and their relationship with the prognosis of diabetic foot ulcers. Methods: The clinical data of diabetic foot patients (n=495) admitted to West China Hospital, Sichuan University since 2016 were retrospectively collected through the hospital electronic medical record system to analyze the characteristics of inflammatory markers and their relationship with the prognosis of diabetic foot ulcers. Results: White blood cell count (WBC), erythrocyte sedimentation rate (ESR), C-reactive protein (CRP), procalcitonin (PCT), and interleukin-6 (IL-6) levels were significantly higher in patients defined as grade 4 on the Wagner Scale than those in patients defined as grade 0-3 on the Wagner Scale. Neutrophil percentage (NE%) was higher in Wagner grade-4 patients than those in Wagner grade-0 and grade-1 patients and higher in Wagner grade-3 patients than those in Wagner grade-0 patients. NE%, CRP, PCT, and IL-6 levels were positively correlated with the severity of diabetic foot, with the respective odds ratio (OR) at 95% confidence interval (CI) being 1.038 (1.019-1.056), 1.019 (1.012-1.026), 8.225 (2.015-33.576), and 1.017 (1.008-1.025). Using Wagner grade-0 patients as the reference, patients with higher WBC were more likely to progress to Wagner grade 2, 3, and 4, with the respective OR (95% CI) values being 1.260 (1.096-1.447), 1.188 (1.041-1.356), and 1.301 (1.137-1.490); patients with higher ESR were more likely to progress to Wagner grade 3 and 4, with the respective OR (95% CI) values being 1.030 (1.006-1.054) and 1.045 (1.019-1.071). Baseline ESR (P=0.008), CRP (P=0.039), and IL-6 (P=0.033) levels were lower in patients who had received antibiotics prior to their admission than those in patients who had not received antibiotics before admission. The levels of WBC, NE%, ESR, PCT, and IL-6 were lower in the full recovery group than those in the group of patients who did not respond to treatment. The higher the levels of NE% and IL-6, the worse the prognosis of diabetic foot ulcers became, with the respective OR (95% CI) values being 1.030 (1.010-1.051) and 1.008 (1.002-1.013). Conclusion: The severity of diabetic foot ulcers increased with the rise in baseline levels of inflammatory markers. Elevated baseline NE% and IL-6 levels suggest a poor prognosis. Our findings suggest that early assessment of diabetic foot infection and standardized antibiotic therapy should be implemented to improve the prognosis.

Changes in soil properties and the phoD-harboring bacteria of the alfalfa field in response to phosphite treatment.

Phosphite, a reduced form of orthophosphate, is characterized by high solubility, and transportation efficiency and can be used as potential phosphorus fertilizer, plant biostimulant and supplemental fertilizer in agriculture. However, the effects of phosphite fertilizer on soil properties and microorganisms are poorly understood. This study evaluated the effects of phosphate and phosphite fertilizers on the different forms of phosphorus, alkaline phosphatase (ALP) activity, and phoD-harboring bacterial community in the alfalfa (Medicago sativa) field. The study used four concentrations (30, 60, 90, and 120 mg P2O5 kg-1 soil) of phosphate (KH2PO4) and phosphite (KH2PO3) fertilizers for the alfalfa field treatment. The results showed that both phosphite and phosphate fertilizers increased the total phosphorus (TP) and available phosphorus (AP) contents in the soil. The phosphorus content of the phosphite-treated soil was lower than that of the phosphate-treated one. TP, inorganic phosphate (Pi), and AP negatively regulated ALP activity, which decreased with increasing phosphate and phosphite fertilizers concentrations. Furthermore, high-throughput sequencing analysis identified 6 phyla and 29 families, which were classified from the altered operational taxonomic units (OTUs) of the soil samples. The redundancy analysis (RDA) revealed that pH, TP, AP and Pi were significantly related to the phoD-harboring bacterial community constructure. The different fertilizer treatments altered the key families, contributing to soil ALP activities. Frankiaceae, Sphingomonadaceae, and Rhizobiaceae positively correlated with ALP activity in phosphite-treated soil. Moreover, the structural equation model (SEM) revealed that ALP activity was affected by the phoD-harboring bacterial community through altered organic phosphorus (Po), AP, total nitrogen (TN), soil organic carbon (SOC), and pH levels under phosphate fertilizer treatment. However, the effect was achieved through positive regulation of pH and AP under phosphite fertilizer. Thus, the changes in soil properties and phoD-harboring bacteria in response to phosphate and phosphite treatments differed in the alfalfa field. This study is the first to report the effects of phosphite on the soil properties of an alfalfa field and provides a strong basis for phosphite utilization in the future. Highlights: - Phosphite and phosphate increase the total phosphorus and available phosphorus.- The pH was the dominant factor influencing the phoD-harboring bacterial community under phosphite fertilizer.- The response of soil properties and phoD-harboring bacterial community to phosphate and phosphite fertilizers differed in the alfalfa field.

First report of alfalfa root rot caused by Trichothecium roseum in China.

Alfalfa (Medicago sativa L.) is perennial leguminous forage, which is cultivated throughout the world due to its high yield, high quality, satisfactory palatability, and wide adaptability. With the increase of planting area in China, root diseases caused by Fusarium spp., Sclerotium rolfsii, Phytophthora spp. (Yang et al. 2022), and new pathogens have been found that reduce the yield and quality of alfalfa and cause economic losses (Li at al. 2019). In 2021, an alfalfa disease occurred under conditions of high temperature and high humidity at the Jiaozhou Experimental Base of Qingdao Agricultural University (Jiaozhou Modern Agricultural Science and Technology Demonstration Park, 36.33°N 120.40°E, Qingdao, Shandong, China), and about 2 ha of alfalfa were infected. The disease affected up to 35% of the plants and caused grass spots. Infected plants developed black-brown lesions with irregular shapes on roots with yellowing of the foliage; the leaves of the whole plant turned yellow. In the late stage of the disease, defoliation occurred and the plants stopped growing, wilted and died. Ten infected plants with typical symptom were collected for isolation and identification of pathogen. The infected roots were cut into 3-5 mm2 sections and then soaked in 75% ethanol for 30 s, followed by a 3-minute immersion in 2% sodium hypochlorite for surface sterilization. Next, the tissues were rinsed in sterile water five times and then placed on potato dextrose agar (PDA) medium. After three subcultures and subsequent single spore isolation, one representative strain named as DC1 was isolated from the infected roots. Based on morphological observation, the colony of DC1 was flat, granular, and powdery in appearance. Four days after inoculation on PDA medium, the size of the colony were 2.1-2.6 cm. After 8 to 20 days, the colonies were initially white and gradually change a light pink to peach color. The conidia are two-celled (Hamid et al. 2014), elliptic to pear-shaped, colorless or translucent, smooth to slightly rough with thick walls. The size of conidia ranged from 11.3 to 23.5 µm long × 6.1 to 12.7 µm wide (n =30). For the identification, the rDNA--ITS gene of the fungus was amplified using the primers ITS1/ITS4 (White et al.1990), and the EF1α gene was amplified using primers EF1-983F/EF1-2218R (Rehner and Buckley 2005). Then the PCR amplicons were cloned into the pCE2 TA/Blunt-Zero vector. The results of the rDNA-ITS (OM049197.1, 515 bp) and EF1α (OM069381.1, 926 bp) sequences were deposited in GenBank. DNA analysis showed that the two sequences were 100% similar to the rDNA-ITS sequence (MN882763.1) and EF1α sequence (DQ676610.1) of Trichothecium roseum, respectively. A pathogenicity test was done by placing one piece (0.5 cm in diameter) of fungal culture (PDA plug) 1cm below the crown of 40-day-old healthy alfalfa (cv. Zhongmu No.3) plants, 3 replicates and 20 plants in each replicate. PDA plug without the pathogen were used for control. All plants were cultivated in a growth chamber at 25±1°C with a light cycle of 15 h (90% relative humidity). After 18 days, the roots of inoculated plants had dark brown lesions and the leaves of their plants turn yellow, while those control plants had no symptoms. To fulfilling Koch's postulates, the same pathogen was re-isolated from necrotic root tissue of inoculated plants and confirmed by morphology and the rDNA-ITS and EF1-α sequences. Based on disease symptoms, morphological characteristics DNA sequences and pathogenicity, the pathogen of alfalfa disease in Jiaozhou Experimental Base of Qingdao Agricultural University was identified as T. roseum. To our knowledge, this is first report of T. roseum causing alfalfa root rot. The newly emerging disease may pose threat to alfalfa production of central and southern China in future.

Dissection of the response mechanism of alfalfa under phosphite stress based on metabolomic and transcriptomic data.

Phosphite, a reduced form of phosphate, inhibits the growth and even has toxic effect on plants. To learn more about the mechanism of alfalfa responses to phosphite, the morphological and physiological characteristics, and the metabolites and transcript levels were comprehensively analyzed following the exposure of alfalfa seedlings to phosphite and phosphate under greenhouse conditions. The results showed that phosphite inhibited seedling growth and photosynthesis. However, the absorption efficiency of phosphite was higher than that of phosphate in roots, which was supported by increased total phosphorus concentration of 16.29% and 52.30% on days 8 and 12. Moreover, phosphite stress affected the synthesis of lipids and carbohydrates, which were reflected in enhanced glycolipid and sulfolipid in roots and amylose in shoots. Phosphite stress resulted in a decrease in indole acetic acid (IAA) in the whole plant and zeatin in the shoots, which could enable alfalfa to adapt to the phosphite environment. Some genes involved in phosphate starvation response included SPX, phosphate response regulator2, and inorganic phosphate transporter 1-4 (PHT1;4) in roots were affected by phosphite stress. In addition, some genes that are involved in stress responses and DNA repair were induced by phosphite stress. These observations together suggest that alfalfa responds to phosphite stress by inhibiting growth, regulating the genes induced by phosphate starvation, improving oxidative protection, promoting DNA repair, and adjusting the IAA and zeatin signaling transductions. Our findings provide novel insights into the molecular response to phosphite stress in alfalfa.

Humidity Sensing of Stretchable and Transparent Hydrogel Films for Wireless Respiration Monitoring.

Respiratory monitoring plays a pivotal role in health assessment and provides an important application prospect for flexible humidity sensors. However, traditional humidity sensors suffer from a trade-off between deformability, sensitivity, and transparency, and thus the development of high-performance, stretchable, and low-cost humidity sensors is urgently needed as wearable electronics. Here, ultrasensitive, highly deformable, and transparent humidity sensors are fabricated based on cost-effective polyacrylamide-based double network hydrogels. Concomitantly, a general method for preparing hydrogel films with controllable thickness is proposed to boost the sensitivity of hydrogel-based sensors due to the extensively increased specific surface area, which can be applied to different polymer networks and facilitate the development of flexible integrated electronics. In addition, sustainable tapioca rich in hydrophilic polar groups is introduced for the first time as a second cross-linked network, exhibiting excellent water adsorption capacity. Through the synergistic optimization of structure and composition, the obtained hydrogel film exhibits an ultrahigh sensitivity of 13,462.1%/%RH, which is unprecedented. Moreover, the hydrogel film-based sensor exhibits excellent repeatability and the ability to work normally under stretching with even enhanced sensitivity. As a proof of concept, we integrate the stretchable sensor with a specially designed wireless circuit and mask to fabricate a wireless respiratory interruption detection system with Bluetooth transmission, enabling real-time monitoring of human health status. This work provides a general strategy to construct high-performance, stretchable, and miniaturized hydrogel-based sensors as next-generation wearable devices for real-time monitoring of various physiological signals.

Integrated mRNA and microRNA expression analysis of root response to phosphate deficiency in Medicago sativa.

The deficiency of available phosphate significantly limits plant growth and development. This study sought to investigate how alfalfa (Medicago sativa), a high-yielding and high-quality forage widely cultivated worldwide, responds to phosphate deficiency stress by integrating transcriptional and post-transcriptional data. In this study, 6,041 differentially expressed genes (DEGs) were identified in alfalfa roots under phosphate deficiency conditions. Furthermore, psRNATarget, RNAhybrid, and TargetFinder were used to predict the target genes of 137 differentially expressed miRNAs (DEMs) in the root. In total, 3,912 DEGs were predicted as target genes. Pearson correlation analysis revealed 423 pairs of miRNA-mRNA regulatory relationships. MiRNA negatively regulates mRNA involved in regulatory pathways of phosphate deficiency responses in alfalfa. miR156e targeted squamosa promoter-binding-like protein 13A (SPL13), miR160c targeted auxin response factor 18 (ARF18), and miR2587a controlled glycolysis and citrate cycle via Phosphoenolpyruvate carboxykinase (ATP) (PCKA). Novel-miR27 regulated SPX domain-containing protein that controls phosphate transport in alfalfa root, novel-miR3-targeted sulfoquinovosyl transferase SQD2 controlled sulfolipid synthesis and glutathione S-transferase (GST; mediated by miR169j/k and novel-miR159) regulated glutathione metabolism. miR399l regulated auxin-responsive protein SAUR72 involved in IAA signal transduction, while abscisic acid receptor PYL4 (regulated by novel-miR205 and novel-miR83) participated in ABA signal transduction. Combined miRNA-mRNA enrichment analysis showed that most miRNAs regulate the phosphate starvation response of alfalfa by modulating target genes involved in carbohydrate metabolism, sulfolipid metabolism, glutathione metabolism, and hormone signal transduction. Therefore, this study provides new insights into the post-transcriptional regulation mechanism of phosphate deficiency responses and new perspectives on phosphate assimilation pathways in alfalfa and other legumes.

Effect of Light Intensity on Morphology, Photosynthesis and Carbon Metabolism of Alfalfa (Medicago sativa) Seedlings.

To understand how light intensity influences plant morphology and photosynthesis in the forage crop alfalfa (Medicago sativa L. cv. Zhongmu 1), we investigated changes in leaf angle orientation, chlorophyll fluorescence, parameters of photosynthesis and expression of genes related to enzymes involved in photosynthesis, the Calvin cycle and carbon metabolism in alfalfa seedlings exposed to five light intensities (100, 200, 300, 400 and 500 µmol m-2 s-1) under hydroponic conditions. Seedlings grown under low light intensities had significantly increased plant height, leaf hyponasty, specific leaf area, photosynthetic pigments, leaf nitrogen content and maximal PSII quantum yield, but the increased light-capturing capacity generated a carbon resource cost (e.g., decreased carbohydrates and biomass accumulation). Increased light intensity significantly improved leaf orientation toward the sun and upregulated the genes for Calvin cycle enzymes, thereby increasing photosynthetic capacity. Furthermore, high light (400 and 500 µmol m-2 s-1) significantly enhanced carbohydrate accumulation, accompanied by gene upregulation and increased activity of sucrose and starch-synthesis-related enzymes and those involved in carbon metabolism. Together, these results advance our understanding of morphological and physiological regulation in shade avoidance in alfalfa, which would guide the identification of suitable spatial planting patterns in the agricultural system.

Ultrasensitive, stretchable, and transparent humidity sensor based on ion-conductive double-network hydrogel thin films.

Ion-conductive hydrogels with intrinsic biocompatibility, stretchability, and stimuli-responsive capability have attracted considerable attention because of their extensive application potential in wearable sensing devices. The miniaturization and integration of hydrogel-based devices are currently expected to achieve breakthroughs in device performance and promote their practical application. However, currently, hydrogel film is rarely reported because it can be easily wrinkled, torn, and dehydrated, which severely hinders its development in microelectronics. Herein, thin, stretchable, and transparent ion-conductive double-network hydrogel films with controllable thickness are integrated with stretchable elastomer substrates, which show good environmental stability and ultrahigh sensitivity to humidity (78,785.5%/% relative humidity (RH)). Benefiting from the ultrahigh surface-area-to-volume ratio, abundant active sites, and short diffusion distance, the hydrogel film humidity sensor exhibits 2 × 105 times increased response to 98% RH, as well as 5.9 and 7.6 times accelerated response and recovery speeds compared with the bulk counterpart, indicating its remarkable thickness-dependent humidity-sensing properties. The humidity-sensing mechanism reveals that the adsorption of water improves the ion migration and dielectric constant, as well as establishes the electrical double layer. Furthermore, the noncontact human-machine interaction and real-time respiratory frequency detection are enabled by the sensors. This work provides an innovative strategy to achieve further breakthroughs in device performance and promote the development of hydrogel-based miniaturized and integrated electronics. Electronic Supplementary Material: Supplementary material is available in the online version of this article at 10.1007/s40843-021-2022-1.

Transcriptome Analysis of Fusarium Root-Rot-Resistant and -Susceptible Alfalfa (Medicago sativa L.) Plants during Plant-Pathogen Interactions.

Alfalfa (Medicago sativa L.) is a perennial leguminous forage cultivated globally. Fusarium spp.-induced root rot is a chronic and devastating disease affecting alfalfa that occurs in most production fields. Studying the disease resistance regulatory network and investigating the key genes involved in plant-pathogen resistance can provide vital information for breeding alfalfa that are resistant to Fusarium spp. In this study, a resistant and susceptible clonal line of alfalfa was inoculated with Fusarium proliferatum L1 and sampled at 24 h, 48 h, 72 h, and 7 d post-inoculation for RNA-seq analysis. Among the differentially expressed genes (DEGs) detected between the two clonal lines at the four time points after inoculation, approximately 81.8% were detected at 24 h and 7 d after inoculation. Many DEGs in the two inoculated clonal lines participated in PAMP-triggered immunity (PTI) and effector-triggered immunity (ETI) mechanisms. In addition, transcription factor families such as bHLH, SBP, AP2, WRKY, and MYB were detected in response to infection. These results are an important supplement to the few existing studies on the resistance regulatory network of alfalfa against Fusarium root rot and will help to understand the evolution of host-pathogen interactions.