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Objective: To investigate the levels of cytomegalovirus (CMV) infection and associated risk factors in corneal transplant recipients who experienced transplant failure. Methods: This was a case-control study. Clinical data from 576 cases (576 eyes) of patients who underwent repeat corneal transplant surgery at the Department of Ophthalmology, Peking University Third Hospital, due to corneal transplant failure from January 2016 to May 2022 were collected. Of these, 305 were male and 271 were female, with a median age of 44.0 (0.7, 91.0) years. The CMV infection rate was analyzed based on the detection of CMV DNA in aqueous humor or corneal tissue during corneal transplant surgery. Patients were divided into the CMV group (CMV DNA positive) and the control group (herpes virus DNA negative). The main research indicators included the CMV infection rate, clinical characteristics, and risk factors in corneal transplant recipients. Chi-square tests and binary logistic analysis were used to compare differences between the two groups in general information, systemic diseases, ocular lesions, ocular surgical history, and local and systemic medications. Odds ratios ( OR ) and 95% confidence intervals ( CI ) were calculated for each CMV infection risk factor. Results: The overall CMV infection rate was 21.9%(126/576), with annual rates ranging from 10.9% to 37.7% from 2016 to 2021. After applying inclusion and exclusion criteria, 378 patients were included in the control trial, with 126 in the CMV group and 252 in the control group. Statistically significant differences between the two groups were observed in systemic immune-related corneal lesions [CMV group: 38 (30.2%), control group: 26 (10.3%)], local immune and inflammatory corneal lesions [CMV group: 46 (36.5%), control group: 40 (15.9%)], congenital corneal opacity [CMV group: 46 (36.5%), control group: 48 (19.0%)] total number of corneal transplants (CMV group: 178 times, control group: 276 times), corneal deep neovascularization crossing the graft [CMV group: 104 (82.5%), control group: 68 (27.0%)] and severe opacity [CMV group: 44 (34.9%), control group: 30 (11.0%)]. Binary logistic regression analysis showed that systemic immune-related corneal lesions ( OR= 4.044, 95% CI 1.810-9.033, P <0.001), local immune and inflammatory corneal lesions ( OR= 3.554, 95% CI 1.569-8.052, P =0.002), congenital corneal opacity ( OR= 2.606, 95% CI 1.216-5.589, P =0.014), total number of corneal transplants ( OR= 3.206, 95% CI 1.753-5.864, P <0.001), corneal deep neovascularization crossing the graft ( OR= 8.347, 95% CI 3.967-17.559, P <0.001), and severe opacity ( OR= 3.063, 95% CI 1.221-7.682, P =0.017) were independent risk factors for CMV infection after corneal transplant. Conclusions: CMV infection was present in more than 1/5 of corneal transplant recipients who experienced transplant failure. CMV infection after corneal transplant may be related to immune rejection reactions and ocular inflammatory responses. Inflammatory corneal lesions associated with systemic or local immune abnormalities, congenital corneal opacity, and multiple corneal transplants may exacerbate the levels of inflammatory factors during the perioperative period of corneal transplant, increasing the risk of post-transplant CMV infection, leading to the infiltration of deep neovascularization and severe opacity in the cornea.

International interest is growing in biodiversity conservation and sustainable use in drylands. Desert ecosystems across arid Central Asia are severely affected by global change. Understanding the changes in a plant community is an essential prerequisite to revealing the community assembly mechanism, vegetation conservation, and management. The knowledge of large-scale spatial variation in plant community structure in different Central Asian deserts is still limited. In this study, we selected the Taukum (TD, Kazakhstan) and the Gurbantunggut (GD, China) deserts as the research area, with similar latitudes despite being nearly 1000 km apart. Thirteen and 15 sampling plots were set up and thoroughly investigated. The differences in community structure depending on multiple plant attributes (individual level: plant height, canopy diameter, and plant volume, and community level: plant density, total cover, and total volume) were systematically studied. TD had a better overall environmental status than GD. A total of 113 species were found, with 68 and 74 in TD and GD, respectively. The number of species and plant attributes was unequally distributed across different families and functional groups between deserts. The values of several plant attributes, such as ephemerals, annuals, dicotyledons, and shrubs with assimilative branches in GD, were significantly lower than those in TD. The Motyka indices of six plant attributes (26.18-38.61%) were higher between the two deserts than the species similarity index (20.4%), indicating a more robust convergence for plant functional attributes. The community structures in the two deserts represented by different plant attribute matrices demonstrated irregular differentiation patterns in ordination diagrams. The most variance in community structure was attributed to soil and climatic factors, while geographic factors had the smallest proportion. Consequently, the community structures of the two distant deserts were both different and similar to an extent. This resulted from the long-term impacts of heterogeneous environments within the same region. Our knowledge is further deepened by understanding the variation in community structure in different deserts on a large spatial scale. This therefore provides valuable insights into conserving regional biodiversity in Central Asia.

Purpose: To report the clinical manifestations, postoperative complications and long-term outcomes of endothelial keratoplasty in VZV-related endothelial decompensation., Methods: In this retrospective study, thirteen eyes undergoing endothelial keratoplasty (EK) for VZV-related endothelial decompensation were compared with controls for Fuchs endothelial dystrophy or pseudophakic bullous keratopathy., Results: Twelve patients did not have typical dermal pain or blisters. Ten patients had obvious iris abnormalities. Glaucoma was noted in eight patients before surgery. The best spectacle-corrected visual acuity improved from 1.12 ± 0.47 to 0.39 ± 0.43 (p = .002), whereas endothelial cell (EC) loss was 65% ±15% at 12 months that higher than that in the controls (p < .05). Postoperative complications included graft detachment (2/13), recurrence of endotheliitis (3/13), neurotrophic ulcer (1/13) and scleritis (1/13)., Conclusions: The onset of VZV-related endothelial decompensation is generally insidious. Iris segmental atrophy, glaucoma and pigment KPs are highly suspected to be associated with VZV. EK is a reasonable option to rehabilitate vision.

Aim: To investigate corneal graft survival rate and endothelial cell density (ECD) loss after keratoplasty in cytomegalovirus (CMV) positive patients., Methods: This was a retrospective cohort study. We analyzed the clinical data of patients who underwent viral DNA detection in aqueous humor/corneal tissue collected during keratoplasty from March 2015 to December 2018 at the Peking University Third Hospital, Beijing, China. To further evaluate the effect of CMV on graft survival rate and ECD loss, patients were divided into three groups: 1) CMV DNA positive (CMV+) group; 2) viral DNA negative (virus-) group, comprising virus- group eyes pairwise matched to eyes in the CMV+ group according to ocular comorbidities; 3) control group, comprising virus- group eyes without ocular comorbidities. The follow-up indicators including graft survival rate, ECD, ECD loss, and central corneal thickness (CCT), were analyzed by Tukey honestly significant difference (HSD) test., Results: Each group included 29 cases. The graft survival rate in CMV+ group were lowest among the three groups ( P =0.000). No significant difference in donor graft ECD was found among three groups ( P =0.54). ECD in the CMV+ group was lower than the virus- group at 12 ( P =0.009), and 24mo ( P =0.002) after keratoplasties. Furthermore, ECD loss was higher in the CMV+ group than in the virus- group in the middle stage (6-12mo) post-keratoplasty ( P =0.017), and significantly higher in the early stage (0-6mo) in the virus- group than in the control group ( P =0.000)., Conclusion: CMV reduces the graft survival rate and exerts persistent detrimental effects on the ECD after keratoplasty. The graft ECD loss associate with CMV infection mainly occurrs in the middle stage (6-12mo postoperatively), while ocular comorbidities mainly affects ECD in the early stage (0-6mo postoperatively)., (International Journal of Ophthalmology Press.)

To examine the effects of extreme drought event and extreme precipitation event on productivity of ephemeral plant, we experimentally reduced and increased growing season precipitation amounts by 65% across four slope positions and aspects along sand dunes in the southern edge of the Gurbantünggüt Desert. The results showed that extreme drought significantly reduced aboveground net primary productivity (ANPP) and belowground net primary productivity (BNPP) by 48.8% and 13.7%, respectively, and that extreme precipitation significantly increased ANPP and BNPP by 37.9% and 23.2%, respectively. The sensitivity of ANPP (0.26 and 0.21 g·m -2 ·mm -1 ) to extreme drought and extreme precipitation was significantly higher than that of BNPP (0.02 and 0.03 g·m -2 ·mm -1 ). In addition, ANPP (24.22 g·m -2 ) and BNPP (5.77 g·m -2 ) on the east side of sand dune were significantly increased by 29.7% and 71.7% compared with those on the west side. There was no significant difference in the sensitivity of ANPP and BNPP to precipitation change among different slope positions and aspects.

A lytic bacteriophage vB&#95;EcoM&#95;DE7 (hereafter designated DE7) that could infect donkey-derived Escherichia coli was isolated. The bacteriophage was examined by transmission electron microscopy, and the result showed that DE7 belonged to the family Myoviridae. The microbiological characterization revealed that DE7 was stable over a broad range of pHs (3 ∼10) at 40-50 °C. The latent period was 10 min, and the burst size was 43 PFUs/infected cell. The whole-genome sequencing showed that DE7 was a dsDNA virus and had a genome of 86,130 bp. The genome contained 124 predicted open reading frames (ORFs), 35 of which had known functions, including DNA replication and modification, transcriptional regulation, structural and packaging proteins, and host cell lysis. Twenty tRNA genes were identified, but no genes associated with bacterial pathogenicity, lysogeny and drug resistance were identified. BLASTN analysis revealed that phage DE7 had a high sequence identity (96%) with Salmonella phage vB&#95;SPuM&#95;SP116, but it could not lyse any Salmonella strain tested in this study. DE7 was classified as a Felix O1-like virus based on its general characterization and genomic information. Since phage DE7 exhibited high efficacy in lysing E. coli and lacked genes associated with bacterial virulence, antimicrobial resistance and lysogeny, it could be potentially used to control foal diarrhoea caused by E. coli., Competing Interests: Conflict of Interest The authors declare that they have no conflict of interest., (Copyright © 2022. Published by Elsevier B.V.)

Chinese kale ( Brassica oleracea var. alboglabra Bailey, CC) is a succulent stem vegetable in the Brassica family. Its allotetraploid (AACC) vegetable germplasm, which was synthesized via distant hybridization with the colloquially named 'yellow turnip' ( B. rapa L. ssp. rapifera Matzg., AA), has a swelling stem similar to CC. To address the molecular mechanism of stem development for CC and AACC, RNA sequencing (RNA-seq) was used to investigate transcriptional regulation of their stem development at three key stages including 28 days, 42 days and the bolting stage (BS) after sowing. As a result, 32,642, 32,665, 33,816, 32,147, 32,293 and 32,275 genes were identified in six corresponding cDNA libraries. Among them, 25,459 genes were co-expressed, while 7,183, 7,206, 8,357, 6,688, 6,834 and 6,814 genes were specifically expressed. Additionally, a total of 29,222 differentially expressed genes (DEGs) were found for functional enrichment as well as many genes involved in plant hormones including gibberellin (GA), abscisic acid (ABA), cytokinin (CTK) and auxin (AUX). Based on gene expression consistency between CC and AACC, the gene families including DELLA, GID, PYR/PYL, PP2C, A-ARR and AUX/IAA might be related to stem development. Among these, eight genes including Bo00834s040 , Bo5g093140 , Bo6g086770 , Bo9g070200 , Bo7g116570 , Bo3g054410 , Bo7g093470 and Bo5g136600 may play important roles in stem development based on their remarkable expression levels as confirmed by qRT-PCR. These findings provide a new theoretical basis for understanding the molecular mechanism of stem development in Brassica vegetable stem breeding., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Zheng, Shi, Zhu, Jin, Chen, Zhang, Zhang, Lin, Zhu, Zang and Wu.)

Methods: Overall, 18 rabbits were randomly divided into control, pacing (600 beats/min), and pacing+sac/val groups. The rabbits in the pacing+sac/val cohort received oral sac/val (10 mg/kg twice daily) across the 21-day investigation period. After three weeks, the atrial effective refractory period (AERP) and AF induction rate were compared. HL-1 cultures were exposed to fast pacing (24 h) with and without LBQ657 (active sacubitril form)/valsartan. Western blots were used for detecting Ca v 1.2 and CaMKII expression within atrial muscles of the rabbits and HL-1 cultures of AF model., Results: In comparison to the sham cohort, the AF induction rate was markedly increased together with AERP reduction within pacing cohort. Such changes were markedly rescued through sac/val treatment in pacing+sac/val cohort. The proteomic expression profiles of CaMKII and Ca v 1.2 showed that the CaMKII expression was markedly upregulated, while Ca v 1.2 expression was downregulated in the pacing cohort. Importantly, these effects were absent in pacing+sac/val cohort., Conclusion: Results of this study show that sac/val treatment regulates the expression of CaMKII/Ca v 1.2 and could alter this pathway in atrial rapid electrical stimulation models. Therefore, this investigation could contribute to a novel strategy in AF therapeutics in clinical settings., Competing Interests: The authors declare that they have no conflicts of interest., (Copyright © 2022 Qi Lou et al.)

Precipitation variability and nitrogen (N) deposition caused by anthropogenic activities could profoundly impact ecosystem productivity and carbon cycling. In desert ecosystems, vegetation is sensitive to changes in precipitation and N deposition. However, the impacts of large changes in precipitation, especially with a concurrent increase in N content, on plant community remain unclear. In this study, we carried out experiments to monitor the impacts of five precipitation levels and two N levels on the plant community function and composition from the Junggar desert in Central Asia during the period 2018-2019. Our results showed that: (1) Aboveground net primary production (ANPP) significantly increased with increasing precipitation, it followed a positive linear model under normal precipitation range, and nonlinear mode under extreme precipitation events; (2) N application led to an increase in ANPP, but did not significantly improve the sensitivity of ANPP to precipitation change; (3) Changes in N content and precipitation, and their impacts on ANPP were mainly driven by plant density. These results provide a theoretical basis for predict the future dynamics of terrestrial vegetation more accurately under climate change and increasing nitrogen deposition., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Zang, Xu, Wu and Yang.)

Background and Purpose: Atrial metabolic remodeling plays a critical role in the pathogenesis of atrial fibrillation (AF). Sirtuin3 (Sirt3) plays an important role in energy homeostasis. However, the effect of Sirt3 agonist Honokiol (HL) on AF is unclear. Therefore, the aim of this study is to determine the effect of HL on atrial metabolic remodeling in AF and to explore possible mechanisms. Experimental Approach: irt3 and glycogen deposition in left atria of AF patients were examined. Twenty-one rabbits were divided into sham, P (pacing for 3 weeks), P + H treatment (honokiol injected with pacing for 3 weeks). The HL-1 cells were subjected to rapid pacing at 5 Hz for 24 h, in the presence or absence of HL and overexpression or siRNA of Sirt3 by transfection. Metabolic factors, circulating metabolites, atrial electrophysiology, ATP level, and glycogens deposition were detected. Acetylated protein and activity of its enzymes were detected. Key Results: Sirt3 was significantly down-regulated in AF patients and rabbit/HL-1cell model, resulting in the abnormal expression of its downstream metabolic key factors, which were significantly restored by HL. Meanwhile, AF induced an increase of the acetylation level in long-chain acyl-CoA dehydrogenase (LCAD), AceCS2 and GDH, following decreasing of activity of it enzymes, resulting in abnormal alterations of metabolites and reducing of ATP, which was inhibited by HL. The Sirt3 could regulate acetylated modification of key metabolic enzymes, and the increase of Sirt3 rescued AF induced atrial metabolic remodeling. Conclusion and Implications: HL inhibited atrial metabolic remodeling in AF via the Sirt3 pathway. The present study may provide a novel therapeutical strategy for AF., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Liu, Xu, Zang, Lou, Hang, Gao, Shi, Liu, Wang, Sun, Liu, Zhang, Liu and Dong.)

Atrial structural and electrical remodelling play important roles in atrial fibrillation (AF). Sacubitril/valsartan attenuates cardiac remodelling in heart failure. However, the effect of sacubitril/valsartan on AF is unclear. The aim of this study was to evaluate the effect of sacubitril/valsartan on atrial electrical and structural remodelling in AF and investigate the underlying mechanism of action. Thirty-three rabbits were randomized into sham, RAP, and sac/val groups. HL-1 cells were subjected to control treatment or rapid pacing with or without LBQ657 and valsartan. Echocardiography, atrial electrophysiology, and histological examination were performed. The concentration of Ca 2+ and expression levels of calcineurin, NFAT, p-NFAT, Cav1.2, collagen Ⅰ and Ⅲ, ANP, BNP, CNP, NT-proBNP, and ST2 in HL-1 cells, and I caL in left atrial cells, were determined. We observed that compared to that in the sham group, the atrium and right ventricle were enlarged, myocardial fibrosis was markedly higher, AF inducibility was significantly elevated, and atrial effective refractory periods were shortened in the RAP group. These effects were significantly reversed by sacubitril/valsartan. Compared to that in the sham group, collagen Ⅰ and Ⅲ, NT-proBNP, ST2, calcineurin, and NFAT were significantly up-regulated, while p-NFAT and Ca v 1.2 were down-regulated in the RAP group, and sacubitril/valsartan inhibited these changes. Ca 2+ concentration increased and I CaL density decreased in in vivo and in vitro AF models, reversed by sacubitril/valsartan. Sacubitril/valsartan attenuates atrial electrical remodelling and ameliorates structure remodelling in AF. This study paves the way for the possibility of clinical use of sacubitril/valsartan in AF patients., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Extreme climatic conditions are major drivers of ecosystem function and dynamics and their frequency is increasing under climate change. Climatic conditions interact with local microtopography, which might either buffer or exacerbate the degree of climatic stress. Here we sought to understand how extremely dry growing seasons affected the composition and productivity of desert ephemeral communities growing in sand dunes from the Gurbantunggut desert in Central Asia, and to which extent did microtopography modulate the response. We set up a rainfall manipulation study on four sand dune microtopographies and, during two consecutive years, we measured soil moisture, nutrients and texture, ephemeral layer composition, plant phenology, biomass accumulation and biomass allocation patterns for the dominant species. We observed significant biomass reductions during the extreme drought but plant community richness and composition were not affected, indicating that the composition of the ephemeral layer in this desert ecosystem may resist under extreme conditions. Additionally, extreme drought increased biomass allocation to reproductive organs of the dominant species. There were also significant microtopographic effects as the sensitivity of biomass to drought in western aspects was larger than in eastern aspects. Our results indicate that previously overlooked microtopographical differences may mediate the impact of climate change on plant communities., Competing Interests: Declaration of competing interest The authors declare that they have no conflict of interest., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Soil degradation resulting from various types of salinity is a major environmental problem, especially in arid and semiarid regions. Exploring the water-related physiological traits of halophytes is useful for understanding the mechanisms of salt tolerance. This knowledge could be used to rehabilitate degraded arid lands. To investigate whether different types of salinity influence the water sources and water-use efficiency of desert plants (Karelinia caspia, Tamarix hohenackeri, Nitraria sibirica, Phragmites australis, Alhagi sparsifolia, Suaeda microphylla, Kalidium foliatum) in natural environments, we measured leaf gas exchange, leaf carbon and xylem oxygen isotope composition and soil oxygen isotope composition at neutral saline-sodic site (NSS) and alkaline saline-sodic site (ASS) in northwest China. The studied plants had different xylem water oxygen isotope compositions (δ 18 O) and foliar carbon isotope compositions (δ 13 C), indicating that desert plants coexist through differentiation in water use patterns. Compared to that at the NSS site, the stem water in K. caspia, A. sparsifolia and S. microphylla was depleted in 18 O at the ASS site, which indicates that plants can switch to obtain water from deeper soil layers when suffering environmental stress from both salinity and alkalinisation. Alhagi sparsifolia had higher δ 13 C at the ASS site than at the NSS site, while K. caspia and S. microphylla had lower δ 13 C, which may have resulted from interspecific differences in plant alkali and salt tolerance ability. Our results suggest that under severe salinity and alkalinity, plants may exploit deeper soil water to avoid ion toxicity resulting from high concentrations of soluble salts in the superficial soil layer. In managed lands, it is vital to select and cultivate different salt-tolerant or alkali-tolerant plant species in light of local conditions., (© 2019 German Society for Plant Sciences and The Royal Botanical Society of the Netherlands.)

The growth and reproduction of shelterbelt plants in arid regions is mainly limited by water availability, which can be alleviated by drip irrigation. We measured water use efficiency (WUE), leaf gas exchange and carbon isotope composition of Tamarix ramosissima in shelterbelt and natural communities in south fringe of Takelamakan Desert to investigate water use efficiency and photosynthetic properties of T. ramosissima differed in water sources. Compared to natural community, T. ramosissima in the drip irrigation community had low water potential due to irrigation and soil salinity. For natural community, T. ramosissima had lower predawn and midday water potential than other plants, indicating that T. ramosissima suffered serious water stress. The net photosynthe-tic rate (P n ), stomatal conductance (g s ), intercellular carbon dioxide concentration (C i ) and maximum photosynthesis rate (P n max ) of T. ramosissima in the drip irrigation community were significantly lower than that in the natural community, indicating the photosynthesis of T. ramosissima in the drip irrigation community was weaker. Compared to the natural community, T. ramosissima in the drip irrigation community had higher long-term water use efficiency, which was related to salt stress caused by irrigation. We concluded that the current irrigation model could meet the water needs of shelterbelt plants. However, the technical defects and soil salt accumulation might affect long-term stability of the shelterbelt community.

In water-limited ecosystems, spatial and temporal partitioning of water sources is an important mechanism that facilitates plant survival and lessens the competition intensity of co-existing plants. Insights into species-specific root functional plasticity and differences in the water sources of co-existing plants under changing water conditions can aid in accurate prediction of the response of desert ecosystems to future climate change. We used stable isotopes of soil water, groundwater and xylem water to determine the seasonal and inter- and intraspecific differences variations in the water sources of six C 3 and C 4 shrubs in the Gurbantonggut desert. We also measured the stem water potentials to determine the water stress levels of each species under varying water conditions. The studied shrubs exhibited similar seasonal water uptake patterns, i.e., all shrubs extracted shallow soil water recharged by snowmelt water during early spring and reverted to deeper water sources during dry summer periods, indicating that all of the studied shrubs have dimorphic root systems that enable them to obtain water sources that differ in space and time. Species in the C 4 shrub community exhibited differences in seasonal water absorption and water status due to differences in topography and rooting depth, demonstrating divergent adaptations to water availability and water stress. Haloxylon ammodendron and T. ramosissima in the C 3 /C 4 mixed community were similar in terms of seasonal water extraction but differed with respect to water potential, which indicated that plant water status is controlled by both root functioning and shoot eco-physiological traits. The two Tamarix species in the C 3 shrub community were similar in terms of water uptake and water status, which suggests functional convergence of the root system and physiological performance under same soil water conditions. In different communities, Haloxylon ammodendron differed in terms of summer water extraction, which suggests that this species exhibits plasticity with respect to rooting depth under different soil water conditions. Shrubs in the Gurbantonggut desert displayed varying adaptations across species and communities through divergent root functioning and shoot eco-physiological traits., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Listeria monocytogenes is an important food-borne pathogenic bacterium that causes human disease, resulting in economic losses worldwide. The current detection methods for L. monocytogenes are not well suited for direct field testing because they involve complicated, time-consuming operations. A simple, efficient method is vital for L. monocytogenes detection. In this study, we combined isothermal recombinase polymerase amplification (RPA) with a lateral flow (LF) strip to rapidly and reliably detect L. monocytogenes. In the presence of biotin- and digoxin-modified primers, RPA produced numerous digoxin- and biotin-attached duplex DNA products. These products were detected on an LF strip via dual immunoreactions (digoxin on the duplex DNA reacted with the anti-digoxin antibody on the gold nanoparticle (Au-NP) and the biotin on the duplex DNA captured by the streptavidin on the LF test zone). The accumulation of Au-NPs produced characteristic bands, enabling the visual detection of L. monocytogenes without instrumentation. This assay could be used to detect L. monocytogenes within 15 min, including DNA amplification with RPA for 10 min at 39 °C and visualization of the amplicons by LF strips for 5 min. Experiments confirmed a detection limit as low as 300 fg of DNA and 1.5 × 10 1 CFU in pure cultures. Furthermore, RPA-LF exhibited no cross-reactions with pathogens. Evaluation of the method with food samples indicated that the detection limit was substantially improved to 1.5 × 10° CFU for the original bacterial content in 25 g/mL samples after enrichment for 6 hr. RPA-LF can be used as a sensitive and rapid detection technique for L. monocytogenes., Practical Application: Recombinase polymerase amplification (RPA) can amplify target DNA at 37 to 42 °C without a thermal cycler. Lateral flow (LF) strips are portable, cheap and easy to operate. RPA combined with LF strips to detect Listeria monocytogenes can be widely used in remote areas., (© 2018 Institute of Food Technologists®.)

Rapid, sensitive, point-of-care detection of bacteria is extremely important in food safety. To address this requirement, we developed a new surface-enhanced Raman scattering (SERS)-based lateral flow (LF) strip biosensor combined with recombinase polymerase amplification (RPA) for simultaneous detection of Listeria monocytogenes and Salmonella enterica serotype Enteritidis. Au MBA @Ag core-shell nanoparticles were used in this SERS-LF. Highly sensitive quantitative detection is achieved by measuring the characteristic peak intensities of SERS tags. Under optimal conditions, the SERS intensities of MBA at 1077 cm -1 on test lines are used to measure S. Enteritidis (y = 1980.6x - 539.3, R 2 = 0.9834) and L. monocytogenes (y = 1696.0x - 844, R 2 = 0.9889), respectively. The limit of detection is 27 CFU/mL for S. Enteritidis and 19 CFU/mL for L. monocytogenes. Significantly, this SERS-LF has high specificity and applicability in the detection of L. monocytogenes and S. Enteritidis in food samples. Therefore, the SERS-LF is a feasible method for the rapid and quantitative detection of a broad range of bacterial pathogens in real food samples.

The efficient and timely detection of pathogens is a major concern worldwide. The aim of this study was to establish a rapid detection method for Salmonella bacteria in food samples to facilitate timely treatment. Widely used detection methods currently include culture-based methods and PCR-based methods. The former are time consuming, requiring 2 to 3 d, whereas the latter have higher accuracy but are typically complicated, requiring expertise and expensive instruments. In this study, a sensitive and rapid approach for the visual and point-of-use detection of Salmonella bacteria based on recombinase polymerase amplification (RPA) and a lateral-flow (LF) nucleic acid strip was established. We designed a pair of primers according to the invA gene of Salmonella bacteria: one was modified with digoxin, and the other was modified with biotin. In the presence of the biotin- and digoxin-modified primers and target DNA, the RPA produced a substantial amount of duplex DNA attached to biotin and digoxin. The products were detected using LF strips through immunoreaction: anti-digoxin antibodies on the gold nanoparticles, digoxin on the duplex, streptavidin on the LF test line, and biotin on the duplex. The developed RPA-LF assay allowed detection of Salmonella genomic DNA in less than 20 min with simple water bath equipment or portable thermal equipment. In addition, the RPA-LF assay was highly sensitive, with a detection limit as low as 20 fg of target DNA or 1.05 × 10 1 cfu of bacteria in pure culture, and highly specific, exhibiting no cross-reaction with Staphylococcus aureus, Escherichia coli, Listeria monocytogenes, Shigella, Enterobacter aerogenes, or Campylobacter jejuni. Importantly, Salmonella could be detected in milk and chicken breast at concentrations as low as 1.05 × 10 0 cfu/mL or 1.05 × 10 0 cfu/g after enrichment for 2 h and in eggs at 1.05 × 10 0 cfu/g after enrichment for 4 h. Furthermore, RPA was more sensitive than PCR, which requires a thermal cycling device. In summary, this study describes a sensitive, simple, and point-of-use detection method for Salmonella bacteria., (Copyright © 2017 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.)

The aim of the present study was to investigate whether the physiological features of Ano1 were affected by enhanced green fluorescent protein (EGFP) fusing at Ano1 C-terminal. The eukaryotic expression vectors of Ano1 and EGFP-Ano1 were constructed, and these plasmids were transfected into Fischer rat thyroid follicular epithelial (FRT) cells using liposome. The expression and location of Ano1 were examined by using inverted fluorescence microscope. The ability of Ano1 to transport iodide was detected by kinetics experiment of fluorescence quenching. The results showed that both Ano1 and EGFP-Ano1 were expressed on FRT cell membrane and could be activated by Ca(2+). There was no significant difference of the ability to transport iodide between Ano1 and EGFP-Ano1. These results suggest Ano1 and EGFP-Ano1 have similar physiological feature.

Glucosinolates (GSs) are an important group of defensive phytochemicals mainly found in Brassicaceae. Plant hormones jasmonic acid (JA) and salicylic acid (SA) are major regulators of plant response to pathogen attack. However, there is little information about the interactive effect of both elicitors on inducing GS biosynthesis in Chinese cabbage (Brassica rapa ssp. pekinensis). In this study, we applied different concentrations of methyl jasmonate (MeJA) and/or SA onto the leaf and root of Chinese cabbage to investigate the time-course interactive profiles of GSs. Regardless of the site of the elicitation and the concentrations of the elicitors, the roots accumulated much more GSs and were more sensitive and more rapidly responsive to the elicitors than leaves. Irrespective of the elicitation site, MeJA had a greater inducing and longer lasting effect on GS accumulation than SA. All three components of indole GS (IGS) were detected along with aliphatic and aromatic GSs. However, IGS was a major component of total GSs that accumulated rapidly in both root and leaf tissues in response to MeJA and SA elicitation. Neoglucobrassicin (neoGBC) did not respond to SA but to MeJA in leaf tissue, while it responded to both SA and MeJA in root tissue. Conversion of glucobrassicin (GBC) to neoGBC occurred at a steady rate over 3 d of elicitation. Increased accumulation of 4-methoxy glucobrassicin (4-MGBC) occurred only in the root irrespective of the type of elicitors and the site of elicitation. Thus, accumulation of IGS is a major metabolic hallmark of SA- and MeJA-mediated systemic response systems. SA exerted an antagonistic effect on the MeJA-induced root GSs irrespective of the site of elicitation. However, SA showed synergistic and antagonistic effects on the MeJA-induced leaf GSs when roots and leaves are elicitated for 3 d, respectively.

This study aimed to explore the impact of depression caused by chronic unpredictable mild stress (CUMS) on in vivo activity of six kinds of CYP450 isoforms in rats. According to 'Katz' method, the model of CUMS was established. Tolbutamide, chlorzoxazone, theophylline, midazolam, omeprazole and dextromethorphan were chosen as probe substrates of CYP2C6, CYP2E1, CYP1A2, CYP3A2, CYP2D1 and CYP2D2 of rats. Plasma concentration of six kinds of CYP450 in control group and model group were determined by LC-MS/MS and computed pharmacokinetic parameters. Consequently, metabolism of theophylline and chlorzoxazone accelerated significantly (P < 0.01), but tolbutamide, dextromethorphan, omeprazole and midazolam had no significant difference. The present study proved that depression caused by CUMS had strong induction to CYP1A2 and medium induction to CYP2E1.

To establish a LC-MS/MS method to determine caffeic acid, chlorogenic acid in rat plasma and study their pharmacokinetics in rats. Six Sprague-Dawley rats were intravenously injected with 4 mL x kg(-1) of Dengzhanxixin injection, respectively. Their drug plasma concentration was determined by LC-MS/MS, with tinidazole as an internal standard. The pharmacokinetic parameters were calculated by DAS 1.0. The linear concentration ranges of caffeic acid, and chlorogenic acid were 2-128 microg x L(-1) (r = 0.998 1) and 3-384 microg x L(-1) (r = 0.998 7), respectively. The methodological test showed conformance to the requirements. The intraday and inter-day variable coefficients were both less than 10.0%, indicating that both of legitimate precise and accuracy were in conformity with the requirements of biological sample analysis. For caffeic acid, the pharmacokinetic parameter t1/2beta AUC0-t, and CL were (130.91 +/- 38.77) min, (4.89 +/- 0.96) mg x min x L(-1) and (0.12 +/- 0.02) L x min(-1) x kg(-1), respectively. For chlorogenic acid, the pharmacokinetic parameter t1/2beta , AUC0-t, and CL were (49.38 +/- 8.85) min, (9.54 +/- 0.95) mg x min x L(-1) and (0.09 +/- 0.003) L x min(-1) x kg(-1), respectively. The LC-MS/MS analysis method established in this study was proved to be so accurate and sensitive that it can be applied to the pharmacokinetic study of caffeic acid and chlorogenic acid.

Glucosinolates play important roles in plant defense against herbivores and microbes, as well as in human nutrition. Some glucosinolate-derived isothiocyanate and nitrile compounds have been clinically proven for their anticarcinogenic activity. To better understand glucosinolate biosynthesis in Brassica rapa, we conducted a comparative genomics study with Arabidopsis thaliana and identified total 56 putative biosynthetic and regulator genes. This established a high colinearity in the glucosinolate biosynthesis pathway between Arabidopsis and B. rapa. Glucosinolate genes in B. rapa share 72-94% nucleotide sequence identity with the Arabidopsis orthologs and exist in different copy numbers. The exon/intron split pattern of B. rapa is almost identical to that of Arabidopsis, although inversion, insertion, deletion and intron size variations commonly occur. Four genes appear to be nonfunctional as a result of the presence of a frame shift mutation and retrotransposon insertion. At least 12 paralogs of desulfoglucosinolate sulfotransferase were found in B. rapa, whereas only three were found in Arabidopsis. The expression of those paralogs was not tissue-specific but varied greatly depending on B. rapa tissue types. Expression was also developmentally regulated in some paralogs but not in other paralogs. Most of the regulator genes are present as triple copies. Accordingly, glucosinolate synthesis and regulation in B. rapa appears to be more complex than that of Arabidopsis. With the isolation and further characterization of the endogenous genes, health-beneficial vegetables or desirable animal feed crops could be developed by metabolically engineering the glucosinolate pathway.

Three Arabidopsis cDNAs, MAM1, CYP79F1, and CYP83A1, required for aliphatic glucosinolate biosynthesis were introduced into Chinese cabbage by Agrobacterium tumefaciens-mediated transformation. The transgenic lines overexpressing MAM1 or CYP83A1 showed wild-type phenotypes. However, all the lines overexpressing CYP79F1 displayed phenotypes different from wild type with respect to the stem thickness as well as leaf width and shape. Glucosinolate contents of the transgenic plants were compared with those of wild type. In the MAM1 line M1-1, accumulation of aliphatic glucosinolates gluconapin and glucobrassicanapin significantly increased. In the CYP83A1 line A1-1, all the aliphatic glucosinolate levels were increased, and the levels of gluconapin and glucobrassicanapin were elevated by 4.5 and 2 fold, respectively. The three CYP79F1 transgenic lines exhibited dissimilar glucosinolate profiles. The F1-1 line accumulated higher levels of gluconapoleiferin, glucobrassicin, and 4-methoxy glucobrassicin. However, F1-2 and F1-3 lines demonstrated a decrease in the levels of gluconapin and glucobrassicanapin and an increased level of 4-hydroxy glucobrassicin.

Indole glucosinolates (IG) play important roles in plant defense, plant-insect interactions, and stress responses in plants. In an attempt to metabolically engineer the IG pathway flux in Chinese cabbage, three important Arabidopsis cDNAs, CYP79B2, CYP79B3, and CYP83B1, were introduced into Chinese cabbage by Agrobacterium-mediated transformation. Overexpression of CYP79B3 or CYP83B1 did not affect IG accumulation levels, and overexpression of CYP79B2 or CYP79B3 prevented the transformed callus from being regenerated, displaying the phenotype of indole-3-acetic acid (IAA) overproduction. However, when CYP83B1 was overexpressed together with CYP79B2 and/or CYP79B3, the transformed calli were regenerated into whole plants that accumulated higher levels of glucobrassicin, 4-hydroxy glucobrassicin, and 4-methoxy glu-cobrassicin than wild-type controls. This result suggests that the flux in Chinese cabbage is predominantly channeled into IAA biosynthesis so that coordinate expression of the two consecutive enzymes is needed to divert the flux into IG biosynthesis. With regard to IG accumulation, overexpression of all three cDNAs was no better than overexpression of the two cDNAs. The content of neoglucobrassicin remained unchanged in all transgenic plants. Although glucobrassicin was most directly affected by overexpression of the transgenes, elevated levels of the parent IG, glucobrassicin, were not always accompanied by increases in 4-hydroxy and 4-methoxy glucobrassicin. However, one transgenic line producing about 8-fold increased glucobrassicin also accumulated at least 2.5 fold more 4-hydroxy and 4-methoxy glucobrassicin. This implies that a large glucobrassicin pool exceeding some threshold level drives the flux into the side chain modification pathway. Aliphatic glucosinolate content was not affected in any of the transgenic plants.