Integrative regulatory mechanisms of stomatal movements under changing climate.

Global climate change-caused drought stress, high temperatures and other extreme weather profoundly impact plant growth and development, restricting sustainable crop production. To cope with various environmental stimuli, plants can optimize the opening and closing of stomata to balance CO2 uptake for photosynthesis and water loss from leaves. Guard cells perceive and integrate various signals to adjust stomatal pores through turgor pressure regulation. Molecular mechanisms and signaling networks underlying the stomatal movements in response to environmental stresses have been extensively studied and elucidated. This review focuses on the molecular mechanisms of stomatal movements mediated by abscisic acid, light, CO2 , reactive oxygen species, pathogens, temperature, and other phytohormones. We discussed the significance of elucidating the integrative mechanisms that regulate stomatal movements in helping design smart crops with enhanced water use efficiency and resilience in a climate-changing world.

[Effect of Shoutai Pills on expression of key glycolytic proteins and apoptosis related factors at maternal fetal interface in mouse model of threatened abortion with syndrome of kidney deficiency].

This study aims to explore the mechanism of Shoutai Pills in treating threatened abortion. According to the random number table method, ICR female mice were randomized into a normal group, a model group, a dydrogesterone group, and a Shoutai Pills group, with 15 mice in each group. Mice were administrated with normal saline(normal and model groups) or the suspension of Shoutai Pills or dydrogesterone by gavage at 9:00 am every day. At 16:00 every day, mice in the normal group were administrated with an equal volume of distilled water, while those in the model, Shoutai Pills, and dydrogesterone groups were administrated with hydrocortisone solution by gavage for 4 consecutive days. ICR female and male mice were caged in a ratio of 2∶1 during the pre-estrous or estrous period. From the first day of pregnancy, drug administration was continued for 5 consecutive days. On day 6, mice were administrated with mifepristone by gavage to establish the model of kidney deficiency-induced abortion. On day 6 of pregnancy, 10 female ICR mice were randomly selected from each group, and the uterus was collected for observation of the pathological changes of trophoblasts at the maternal-fetal interface by hematoxylin-eosin(HE) staining. The protein levels of key enzymes of glycolysis, hexokinase 2(HK2), enolase 1(ENO1), pyruvate kinase M2(PKM2), and lactate dehydrogenase A(LDHA), were determined by Western blot and immunofluorescence. The expression of apoptosis-related proteins including B cell lymphoma-2(Bcl-2), Bcl-2-associated protein X(Bax), and cysteinyl aspartate-specific proteinase-3(caspase-3) was determined by Western blot and real-time PCR. Terminal-deoxynucleoitidyl transferase-mediated nick-end labeling was employed to examine apoptosis. The embryo loss rate of the remaining five female mice was calculated by trypan blue staining method on day 14 of pregnancy. On day 14 of pregnancy, the embryo loss rate of the normal group was 5.00%, which was lower than that(27.78%) in the model group(P<0.05). Dydrogesterone and Shoutai Pills groups showed reduced embryo loss rates(10.26% and 7.50%, respectively) compared with the model group. On day 6 of pregnancy, compared with the normal group, the model group showed down-regulated expression of HK2, ENO1, PKM2, LDHA, and Bcl-2 and up-regulated expression of Bax and caspase-3(P<0.05). Compared with the model group, dydrogesterone and Shoutai Pills up-regulated the expression of HK2, ENO1, PKM2, LDHA, and Bcl-2 and down-regulated the expression of Bax and caspase-3(P<0.05). Compared with that in the normal group, the apoptosis rate in the model group increased(P<0.05). Compared with the model group, dydrogesterone and Shoutai Pills reduced the apoptosis rate(P<0.05). In conclusion, Shoutai Pills can reduce the embryo loss rate and protect embryos by promoting aerobic glycolysis at the maternal-fetal interface and inhibiting the apoptosis of trophoblasts in mice.

OsSLA1 functions in leaf angle regulation by enhancing the interaction between OsBRI1 and OsBAK1 in rice.

Leaf angle is an important trait in plants. Here, we demonstrate that the leucine-rich repeat receptor-like kinase OsSLA1 plays an important role in leaf angle regulation in rice (Oryza sativa). OsSLA1 mutant plants exhibited a small leaf angle phenotype due to changes of adaxial cells in the lamina joint. GUS staining revealed that OsSLA1 was highly expressed in adaxial cells of the lamina joint. The OsSLA1 mutant plants were insensitive to exogenous epibrassinolide (eBL) and showed upregulated expression of DWARF and CPD, but downregulated expression of BU1, BUL1, and ILI1, indicating that brassinosteroid (BR) signal transduction was blocked. Fluorescence microscopy showed that OsSLA1 was localized to the plasma membrane and nearby periplasmic vesicles. Further study showed that OsSLA1 interacts with OsBRI1 and OsBAK1 via its intracellular domain and promotes the interaction between OsBRI1 and OsBAK1. In addition, phosphorylation experiments revealed that OsSLA1 does not possess kinase activity, but that it can be phosphorylated by OsBRI1 in vitro. Knockout of OsSLA1 in the context of d61 caused exacerbation of the mutant phenotype. These results demonstrate that OsSLA1 regulates leaf angle formation via positive regulation of BR signaling by enhancing the interaction of OsBRI1 with OsBAK1.

The Microbiota in Hematologic Malignancies.

OPINION STATEMENT: There are approximately 1.2 million new hematologic malignancy cases resulting in ~ 690,000 deaths each year worldwide, and hematologic malignancies remain the most commonly occurring cancer in children. Even though advances in anticancer treatment regimens in recent decades have considerably improved survival rates, their cytotoxic effects and the resulting long-term complications pose a significant burden on the patients and the health care system. Therefore, non-toxic treatment modalities are needed to decrease side effects. The human body is the host to approximately 40 trillion microbes, known as the human microbiota. The large majority of the microbiota is located in the gastrointestinal tract, and is primarily composed of bacteria. The microbiota plays several important physiological roles, ranging from digestive functions to immunological and neural development. Investigating the microbiota in patients with hematologic malignancies has several important implications. The microbiota affects hematopoiesis, and influences the efficacies of chemotherapy and antimicrobial treatments. Determination of the microbiota composition and diversity could be an important part of risk stratification in the future, and may also take part to personalize antimicrobial treatments. Modulation of the microbiota via probiotics or fecal transplant can potentially be involved in reducing side effects of chemotherapy, and eliminating multiple drug resistant strains in patients with hematologic malignancies.

Visual detection of bacterial DNA using activated paper stripe.

A rapid and accurate detection of pathogens is essential for bedside or on-site diagnosis. Filter paper is an ideal diagnostic tool as it requires no equipment, possesses a high surface-area-to-volume ratio and a high capacity of capillary force. The functionalization of the surface of cellulose filter paper was explored by using glutaric anhydride, N-hydroxysuccinimide, and N,N'-dicyclohexylcarbodiimide. The activated surface systems enable aminated DNA to be immobilized on the surface of filter paper. Both synthetic oligonucleotides and bacterial genomic DNA of Staphylococcus aureus, Escherichia coli, and Campylobacter jejuni were detected successfully. The system produces a clear, consistent and highly visible brown signal within 1-5 min. The digital image can also be analyzed quantitatively due to the brown color resulting from the presence of magnetic beads. Bacterial DNA detection was accomplished by using 16S rDNA probe on the activated paper surface for universal bacterial diagnosis. The method is stable and repeatable. It can detect at least 0.5 pmol of a 120-base synthetic oligonucleotide per assay and 5-10 ng of bacterial DNA per assay. Graphical abstract Schematic representation of the method: a. functionalization of cellulose filter paper, b. printing of aminated probes, c. incubation, d. blocking of unreacted functional groups (as dots shown), e. a visual detection of targets, f. quantitative analysis of image.

Characterization of Spectinomycin Resistance in Streptococcus suis Leads to Two Novel Insights into Drug Resistance Formation and Dissemination Mechanism.

Spectinomycin is an aminocyclitol antibiotic used clinically to treat a variety of infections in animals. Here, we characterized drug resistance prevalence in clinical Streptococcus suis isolates and discovered a novel resistance mechanism in which the s5 mutation (Gly26Asp) results in high spectinomycin resistance. Additionally, a novel integrative and conjugative element encompassing a multidrug resistance spw_like-aadE-lnu(B)-lsa(E) cluster and a cadmium resistance operon were identified, suggesting a possible cause for the wide dissemination of spectinomycin resistance in S. suis.

Visual detection of DNA on paper chips.

On-site DNA analysis for diagnostic or forensic purposes is much anticipated in the future of molecular testing. Yet the challenges to achieve this goal remain large with rapid and inexpensive detection and visualization being key factors for any portable analysis system. We have developed a filter paper-based nucleic acid assay, which is able to identify and distinguish dog and human genomic and mitochondrial samples in a forensic setting. The filter paper material allows for transport by capillary force of the sample DNA through the detection surface, allowing the targets to hybridize specifically to their complementary capture sequences. Coupling micrometer-sized beads to DNA allows the results to be visualized by the naked eye, enabling instant, cost-efficient, and on-site detection, while eliminating the need for advanced expensive instrumentation.

Dynamic Homogenization of Internal Strain in Multi-Principal Element Alloy via High-Concentration Doping of Oxygen with Large Mobility.

Internal strain and its distribution within the crystal lattice play crucial roles in modulating dislocation activities, thereby affecting mechanical properties of materials. Through the synergistic application of integrated differential phase contrast, in situ transmission electron microscopy characterizations, and computational simulations, a method is unveiled for homogenizing dislocation pinning in NiCoCr multi-principal element alloy (MPEA) through the introduction of a high concentration of oxygen atoms with high diffusion mobility. The doping of massive oxygen atoms creates a high density of strong local pinning points for dislocation motion. Notably, oxygen interstitials exhibit remarkable diffusion and mobility across different octahedral and tetrahedral sites within the distorted crystal lattice of NiCoCrO alloy, even at room temperature. The capability allows for the release of severe stress concentrations arising from dislocation entanglement and the establishment of new strong local pinning points at alternative locations in a uniform way, enabling the material with high strength and outstanding deformability. These findings suggest that interstitial atoms can exhibit significant mobility, even at ambient temperature, in complex MPEAs with spreading lattice distortion, opening new possibilities for dislocation engineering.

Effect of 2-deoxyglucose-mediated inhibition of glycolysis on migration and invasion of HTR-8/SVneo trophoblast cells.

The proper invasion of trophoblasts is crucial for embryo implantation and placental development, which is helpful to establish a correct maternal-fetal relationship. Trophoblasts can produce a large amount of lactate through aerobic glycolysis during early pregnancy. Lactate creates a low pH microenvironment around the embryo to help uterine tissue decompose and promote the invasion of trophoblasts. The purpose of this study is to reveal the the potential mechanism of aerobic glycolysis regulating the invasiveness of trophoblasts by investigating the effect of 2-Deoxy-D-glucose (2-DG), a glycolysis inhibitor, on the biological function of HTR-8/SVneo trophoblast cells, the expressions of epithelial mesenchymal transformation (EMT) markers and invasion-related factors. 2-DG could inhibit the aerobic glycolysis of trophoblasts and decrease the activity of trophoblasts in a dose-dependent manner. Moreover, 2-DG inhibited the EMT of HTR-8/SVneo cells, down-regulated the expression of invasion-related factors matrix metalloproteinase 2/9 (MMP2/9) and up-regulated the expression of tissue inhibitor of matrix metalloproteinases 1/2 (TIMP1/2), thus inhibiting cell migration and invasion. This paper provides a foundation in the significance of aerobic glycolysis of trophoblasts in the process of invasion, and also provides ideas and insights for the promotion of embryo implantation.

Genome and Transcriptome Analysis of Ascochyta pisi Provides Insights into the Pathogenesis of Ascochyta Blight of Pea.

Ascochyta blight caused by Ascochyta pisi is a major constraint to pea (Pisum sativum L.) production worldwide. Deciphering the pathogenic mechanism of A. pisi on peas will help in breeding resistant pea varieties and developing effective approaches for disease management. However, little is known about the genomic features and pathogenic factors of A. pisi. In this study, we first report that A. pisi is one of the causal agents of ascochyta blight disease of pea in China. The genome of the representative isolate A. pisi HNA23 was sequenced using PacBio and Illumina sequencing technologies. The HNA23 genome assembly is almost 41.5 Mb in size and harbors 10,796 putative protein-encoding genes. We predicted 555 carbohydrate-active enzymes (CAZymes), 1,008 secreted proteins, 74 small secreted cysteine-rich proteins (SSCPs), and 26 secondary metabolite biosynthetic gene clusters (SMGCs). A comparison of A. pisi genome features with the features of 6 other available genomes of Ascochyta species showed that CAZymes, the secretome, and SMGCs of this genus are considerably conserved. Importantly, the transcriptomes of HNA23 during infection of peas at three stages were further analyzed. We found that 245 CAZymes and 29 SSCPs were upregulated at all three tested infection stages. SMGCs were also trigged, but most of them were induced at only one stage of infection. Together, our results provide important genomic information on Ascochyta spp. and offer insights into the pathogenesis of A. pisi. IMPORTANCE Ascochyta blight is a major disease of legumes worldwide. Ascochyta pisi and other Ascochyta species have been identified as pathogens of ascochyta blight. Here, we first report that A. pisi causes ascochyta blight of pea in China, and we report the high-quality, fully annotated genome of A. pisi. Comparative genome analysis was performed to elucidate the differences and similarities among 7 Ascochyta species. We predict abundant CAZymes (569 per species), secreted proteins (851 per species), and prolific secondary metabolite gene clusters (29 per species) in these species. We identified a set of genes that may be responsible for fungal virulence based on transcriptomes in planta, including CAZymes, SSCPs, and secondary metabolites. The findings from the comparative genome analysis highlight the genetic diversity and help in understanding the evolutionary relationship of Ascochyta species. In planta transcriptome analysis provides reliable information for further investigation of the mechanism of the interaction between Ascochyta spp. and legumes.

An entomopathogenic fungus exploits its host humoral antibacterial immunity to minimize bacterial competition in the hemolymph.

BACKGROUND: The insect hemolymph (blood-equivalent fluid), composed of a large number of hemocytes (blood cells) and a variety of soluble immune effectors, is hostile for pathogens including fungi. In order to survive in the insect hemocoel (body cavity), the entomopathogenic fungus (EPF) has evolved two classical coping strategies, namely evasion and suppression of the host immune reactions. However, it remains unclear whether EPF has other ways of coping with host immunity. RESULTS: In this study, we demonstrated that Metarhizium rileyi (an EPF) infection by injection of blastospores into the hemocoel enhanced the plasma antibacterial activity of cotton bollworm (Helicoverpa armigera), which was partially due to the enhanced expression of antimicrobial peptides (AMPs). The early stage of M. rileyi infection induced the translocation of gut bacteria into the hemocoel, where they were subsequently cleared due to the enhanced plasma antibacterial activity. Further, we showed that the enhanced plasma antibacterial activity and AMP expression were attributable to M. rileyi but not the invasive gut bacteria (opportunistic bacteria). Elevated ecdysone (major steroid hormone in insects) levels in the hemolymph at 48 h post-M. rileyi infection might contribute to the enhanced expression of AMPs. The fungus-elicited AMPs, such as cecropin 3 or lebocin, exhibited potent inhibitory activity against the opportunistic bacteria but not against hyphal bodies. In addition, the opportunistic bacteria competed with hyphal bodies for amino acid nutrients. CONCLUSIONS: M. rileyi infection induced the translocation of gut bacteria, and then the fungi activated and exploited its host humoral antibacterial immunity to eliminate opportunistic bacteria, preventing them from competing for nutrients in the hemolymph. Unlike the classical strategies, EPF utilizes to evade or suppress host immunity, our findings reveal a novel strategy of interaction between EPF and host immunity. Video Abstract.

Shoutai Wan regulates glycolysis imbalance at the maternal-fetal interface in threatened abortion mice.

ETHNOPHARMACOLOGICAL RELEVANCE: Threatened abortion is a common disease among women of childbearing age. Its high incidence rate and unclear etiology, seriously threaten women's physical and mental health. Shoutai Wan (STW) is a traditional Chinese medicine decoction for treating abortion. It has a long history of treating threatened abortion by tonifying the kidney and calming the fetus. However, the mechanism of STW remains unclear. AIM OF STUDY: To study the mechanism and potential benefit of STW in pregnant mice with hydrocortisone and mifepristone-induced threatened abortion. MATERIALS AND METHODS: The STW compounds were identified using gas chromatography-mass spectrometry analysis. STW-H, STW-M, or STW-L was separately given 3 mg/ml, 1.5 mg/ml and 0.75 mg/ml STW in the morning, and 2 mg/ml hydrocortisone in the afternoon from gestation day (D) 1-9 and once with 0.4 mg/kg mifepristone on D10. Didroxyprogesterone (0.1 mg/ml) and equal dose pure water were used to replace STW in didroxyprogesterone (DYD) group and model group respectively. The control group used pure water to replace STW, hydrocortisone, and mifepristone. We performed morphological and histological analyses of the maternal-fetal interface on day 10. RESULTS: The embryo loss rate in the STW-H and DYD groups was lower than that in the model group. Hematoxylin and eosin (HE) staining suggested that the morphology of maternal-fetal interface was improved in the STW-H and DYD groups. Immunohistochemical (IHC), Quantitative Reverse Transcription Polymerase Chain Reactionstaining (qRT-PCR), and Western blot (WB) results indicated that HIF-1α expression in the maternal-fetal interface of the STW-H and DYD groups was higher than that in model group. The activities of HK, PKM, LDH and the concentration of lactic acid in the STW-H and DYD groups were higher than those in model group. Furthermore, the protein and mRNA levels of HK2, PKM2, LDHA, MCT4, and GPR81 were higher in the STW-H and DYD groups than those in the model group. CONCLUSIONS: STW can reduce the pregnancy loss rate by regulating the glycolysis balance at the maternal-fetal interface of kidney deficiency threatened abortion model mice.

An IncN-ST7 epidemic plasmid mediates the dissemination of carbapenem-resistant Klebsiella pneumoniae in a neonatal intensive care unit in China over 10 years.

OBJECTIVES: Carbapenem-resistant Klebsiella pneumoniae (CRKP) has widely disseminated globally, but its epidemiological characterization and clinical significance in paediatric patients are not well understood. In this study, we aimed to trace the dissemination dynamics of CRKP in the neonatal intensive care unit (NICU) of a tertiary hospital over a 10-y period. METHODS: We collected 67 non-duplicate K. pneumoniae species complex isolates from the NICU with patient metadata during 2009-2018. Antimicrobial susceptibility was determined by the agar or broth microdilution method. Risk factors for CRKP-positive patients were identified by univariate and multivariate analysis. Genetic characterization was dissected by whole-genome sequencing. Plasmid transmissibility, stability, and fitness were assessed. RESULTS: Thirty-four of 67 isolates (50.75%) were identified as CRKP. Premature rupture of membranes, gestational age, and invasive procedures are independent risk factors for CRKP-positive patients. The annual isolation rate of CRKP varied between 0% and 88.9%, and multiple clonal replacements were observed during the study period, which could be largely due to the division of the NICU. All but one CRKP produced IMP-4 carbapenemase, which was encoded by an IncN-ST7 epidemic plasmid, suggesting that the IncN-ST7 plasmid mediated the CRKP dissemination in the NICU over 10 y. The same plasmid was found in several CRKP isolates from adult patients, of which two ST17 isolates from the neurosurgery department shared a high homology with the ST17 isolates from the NICU, indicating possible cross-departmental transmission. CONCLUSION: Our study highlights the urgent need for infection control measures targeting high-risk plasmids like IncN-ST7.

Associations of Combined Lifestyle Factors with MAFLD and the Specific Subtypes in Middle-Aged and Elderly Adults: The Dongfeng-Tongji Cohort Study.

Metabolic dysfunction-associated fatty liver disease (MAFLD) is the crucial pathogenesis for intra-hepatic and extra-hepatic diseases, especially in elderly adults. Lifestyle management may be a modifiable cost-effective measure for MAFLD prevention, but the evidence is limited. A total of 23,408 middle-aged and elderly individuals were included in a longitudinal study from 2008 to 2018. Combined lifestyle scores (range 0-6) were evaluated by BMI, smoking, drinking, diet, physical activity, and sleep. Logistic regression models were used to calculate ORs for the risks of MAFLD and specific subtypes. The mean age of participants was 61.7 years, and 44.5% were men. Compared with poor lifestyle (scores 0-2), ORs (95% CIs) of the ideal lifestyle (scores 5-6) were 0.62 (0.57-0.68) for MAFLD, 0.31 (0.28-0.34) for MAFLD with excess weight and obesity, 0.97 (0.75-1.26) for MAFLD with diabetes, and 0.56 (0.51-0.62) for MAFLD with metabolic dysregulation. Additionally, lifestyle improvement was associated with lower risks of MAFLD (OR, 0.76; 95% CI, 0.68-0.86), MAFLD with excess weight and obesity (OR, 0.72; 95% CI, 0.63-0.81), MAFLD with diabetes (OR, 0.74; 95% CI, 0.54-1.02) and MAFLD with metabolic dysregulation (OR, 0.49; 95% CI, 0.43-0.55), respectively. Our findings suggest that adherence to a combined healthy lifestyle was associated with lower risks of MAFLD, particularly in excess weight/obese individuals or those with metabolic dysregulation.

The Impact of Social Isolation and Environmental Deprivation on Blood Pressure and Depression-Like Behavior in Young Male and Female Mice.

Background: Social isolation (SI) and loneliness are major adult and adolescent health concerns, particularly in the coronavirus disease 2019 (COVID-19) era. Recent prospective cohort studies indicate that older women who experienced both SI and loneliness had a significantly higher risk of cardiovascular disease (CVD). Hypertension, a well-established risk factor for CVD, is more prevalent in elderly women than men. Furthermore, a lack of social relationships is strongly associated with an increased risk of hypertension in middle-aged and elderly women compared to men. Although this has not been extensively studied, adolescents and young adults who experience loneliness or SI may also be at risk for CVD and depression. The purpose of this study was to examine the effect of SI on blood pressure and depression-like behavior in young male and female mice. Methods: Weaned C57BL/6 mice were randomly assigned (n = 6/group/sex) to either group housing (GH) or SI. Animals in the SI group were housed in individual cages for 8 weeks with no view of other animals. The cages were kept in ventilated racks to prevent pheromone exposure and socially isolated animals had no cage enrichment. Results: SI increased systolic, diastolic, and mean arterial blood pressure in females and elevated heart rate in both sexes. Body weight gain was dramatically increased in socially isolated females but tended to decrease in socially isolated males. In the forced swim test, which detects depression-like behavior, there was no difference between groups in total immobility time. The latency to immobility, however, was significantly decreased in socially isolated females. Serum concentrations of corticosterone and metanephrine did not differ between socially isolated and group-housed females, but corticosterone levels were significantly reduced in socially isolated males. Conclusions: Our results indicate that 8 weeks of SI leads to significant changes in blood pressure and heart rate and mild changes in depression-like behavior in young mice, with females affected more than males.

Activated paper surfaces for the rapid hybridization of DNA through capillary transport.

The development of low-cost, accurate, and equipment-free diagnostic tests is crucial to many clinical, laboratory, and field applications, including forensics and medical diagnostics. Cellulose fiber-based paper is an inexpensive, biodegradable, and renewable resource, the use of which as a biomolecule detection matrix and support confers several advantages compared to traditional materials such as glass. In this context, a new, facile method for the preparation of surface functionalized papers bearing single-stranded probe DNA (ssDNA) for rapid target hybridization via capillary transport is presented. Optimized reaction conditions were developed that allowed the direct, one-step activation of standard laboratory filters by the inexpensive and readily available bifunctional linking reagent, 1,4-phenylenediisothiocyanate. Such papers were thus amenable to subsequent coupling of amine-labeled ssDNA under standard conditions widely used for glass-based supports. The intrinsic wicking ability of the paper matrix facilitated rapid sample elution through arrays of probe DNA, leading to significant, detectable hybridization in the time required for the sample liquid to transit the vertical length of the strip (less than 2 min). The broad applicability of these paper test strips as rapid and specific diagnostics in "real-life" situations was exemplified by the discrimination of amplicons generated from canine and human mitochondrial and genomic DNA in mock forensic samples.

Rapid DNA visual detection of polymicrobial bloodstream infection using filter paper.

Bloodstream infection (BSI) is a major complication in patients with cancers due to therapy-induced neutropenia and underlying conditions, which increases hospitalization time and mortality rate. Targeted and timely antimicrobial management is crucial to save the patients' lives and reduce the social and economic burdens. Blood culture is a routine clinical diagnostic method of BSI with a long turnaround time, and generally identifies monomicrobial BSI. Thus, polymicrobial BSI often goes undetected although it occurs more frequently in these patients and results in more severe outcomes compared to monomicrobial BSI. In this work, we apply glutaric anhydride, N-hydroxysuccinimide and N,N'-dicyclohexylcarbodiimide to fabricate a functional surface on cellulose filter paper. Targeting three pathogens (Escherichia coli, Saccharomyces cerevisiae, and human cytomegalovirus) commonly occurring in BSI in neutropenic patients, we demonstrate rapid and accurate triplex pathogen DNA detection using the functionalized paper. All three pathogen DNA was identified in 1-5 min with a detection limit of 0.1-0.5 ng/µL. The developed test tool has the potential to provide rapid polymicrobial BSI diagnosis in support of timely, accurate antimicrobial treatment, and could be integrated into an automatic sample-to-result portable equipment.

Microbial and host factors contribute to bloodstream infection in a pediatric acute lymphocytic leukemia mouse model.

Background: Hematological malignancies are the most common cancers in the pediatric population, and T-cell acute lymphocytic leukemia (T-ALL) is the most common hematological malignancy in children. Bloodstream infection (BSI) is a commonly occurring complication in leukemia due to underlying conditions and therapy-induced neutropenia. Several studies identified the gut microbiome as a major source of BSI due to bacterial translocation. This study aimed to investigate changes in the intestinal and fecal microbiome, and their roles in the pathophysiology of BSI in a pediatric T-ALL mouse model using high-throughput shotgun metagenomics sequencing, and metabolomics. Results: Our results show that BSI in ALL is characterized by an increase of a mucin degrading bacterium (Akkermansia muciniphila) and a decrease of butyrate producer Clostridia spp., along with a decrease in short-chain fatty acid (SCFA) concentrations and differential expression of tight junction proteins in the small intestine. Functional analysis of the small intestinal microbiome indicated a reduced capability of SCFA synthesis, while SCFA supplementation ameliorated the development of BSI in ALL. Conclusions: Our data indicates that changes in the microbiome, and the resulting changes in levels of SCFAs contribute significantly to the pathogenesis of bloodstream infection in ALL. Our study provides tailored preventive or therapeutic approaches to reduce BSI-associated mortality in ALL.

Shoutai Wan Improves Embryo Survival by Regulating Aerobic Glycolysis of Trophoblast Cells in a Mouse Model of Recurrent Spontaneous Abortion.

Background: During embryo implantation, the blastocyst exhibits a high capacity for aerobic glycolysis, which results in a unique microenvironment of high lactate/low pH at the maternal-fetal interface. Shoutai Wan (STW) is an effective Chinese herbal formula widely used in the clinical treatment of recurrent spontaneous abortion (RSA). However, the specific molecular mechanism by which STW prevents abortion is yet to be elucidated. Methods: Female CBA/J mice were allocated into six groups randomly and then mated with BALB/c mice as the control group, DBA/2 mice as the RSA model, CBA/J×DBA/2 mice treated with dydrogesterone as the DQYT group, or CBA/J×DBA/2 mice treated with low, medium, and high-dose STW as the STW-L, STW-M, and STW-H groups, respectively. Drug administration started 14 days before mating and ended on the 14th day of pregnancy. The embryo loss rate of each group was calculated on day 14 of gestation, and the pregnancy outcomes of the mice in each group were observed. The mouse serum was collected to determine the levels of progesterone (P) and chorionic gonadotropin (CG). The activities of HK2, PKM2, and LDHA, the key glycolytic enzymes in each group, were detected. The expressions of lactate, ATP, HK2, PKM2, LDHA, MCT4, GLUT1, and GPR81 as well as the morphology of trophoblast cells were examined. Results: The embryo loss rate and adverse pregnancy outcomes were significantly increased (P < 0.05) in the RSA model group. After dydrogesterone or different doses of STW treatment, the embryo loss rate and adverse pregnancy outcomes were rescued to varying degrees (P < 0.05). Interestingly, there was no significant difference among the groups in terms of serum P and CG (P < 0.05). Moreover, the activities of key glycolytic enzymes, lactate, ATP, HK2, PKM2, LDHA, MCT4, GLUT1, GPR81 protein or mRNA expression, and morphological abnormalities of trophoblast cells improved significantly in the RSA mice after dydrogesterone or different doses of STW treatment (P < 0.05). Conclusion: STW can promote aerobic glycolysis in trophoblast cells of RSA mouse embryos, thereby improving the microenvironment of the maternal-fetal interface and enhancing embryo implantation.

Reducing the Periplasmic Glutathione Content Makes Escherichia coli Resistant to Trimethoprim and Other Antimicrobial Drugs.

Although glutathione (GSH) has been shown to influence the antimicrobial effects of many kinds of antibiotics, little is known about its role in relation to trimethoprim (TMP), a widely used antifolate. In this study, several genes related to glutathione metabolism were deleted in different Escherichia coli strains (i.e., O157:H7 and ATCC 25922), and their effects on susceptibility to TMP were tested. The results showed that deleting gshA, gshB, grxA, and cydD caused TMP resistance, and deleting cydD also caused resistance to other drugs. Meanwhile, deleting gshA, grxA, and cydD resulted in a significant decrease of the periplasmic glutathione content. Supplementing exogenous GSH or further deleting glutathione importer genes (gsiB and ggt) restored TMP sensitivity to ΔcydD. Subsequently, the results of quantitative-reverse transcription PCR experiments showed that expression levels of acrA, acrB, and tolC were significantly upregulated in both ΔgrxA and ΔcydD. Correspondingly, deleting cydD led to a decreased accumulation of TMP within bacterial cells, and further deleting acrA, acrB, or tolC restored TMP sensitivity to ΔcydD. Inactivation of CpxR and SoxS, two transcriptional factors that modulate the transcription of acrAB-tolC, restored TMP sensitivity to ΔcydD. Furthermore, mutations of gshA, gshB, grxA, cydC, and cydD are highly prevalent in E. coli clinical strains. Collectively, these data suggest that reducing the periplasmic glutathione content of E. coli leads to increased expression of acrAB-tolC with the involvement of CpxR and SoxS, ultimately causing drug resistance. To the best of our knowledge, this is the first report showing a linkage between periplasmic GSH and drug resistance in bacteria. IMPORTANCE After being used extensively for decades, trimethoprim still remains one of the key accessible antimicrobials recommended by the World Health Organization. A better understanding of the mechanisms of resistance would be beneficial for the future utilization of this drug. It has been shown that the AcrAB-TolC efflux pump is associated with trimethoprim resistance in E. coli clinical strains. In this study, we show that E. coli can sense the periplasmic glutathione content with the involvement of the CpxAR two-component system. As a result, reducing the periplasmic glutathione content leads to increased expression of acrA, acrB, and tolC via CpxR and SoxS, causing resistance to antimicrobials, including trimethoprim. Meanwhile, mutations in the genes responsible for periplasmic glutathione content maintenance are highly prevalent in E. coli clinical isolates, indicating a potential correlation of the periplasmic glutathione content and clinical antimicrobial resistance, which merits further investigation.