CRISPR/Cas9-based engineered Escherichia coli biosensor for sensitive and specific detection of Cd(II) in drinking water.

Cadmium (Cd) is a ubiquitous pollutant that poses a potential threat to human health. Monitoring Cd(II) in drinking water has significant implications for preventing potential threats of Cd(II) to human. However, the weak signal output and response to nontarget interference limit the detection of Cd(II) using bacterial biosensors. In this study, to enable sensitive and specific detection of Cd(II) in water, a stable whole-cell biosensor, K12-PMP-luxCDABE-△cysI, was constructed in a dual-promoter mode by fusing the mercury promoter Pmer, regulatory gene merR(m), and luciferase gene luxCDABE into the E.coli chromosome based on CRISPR/Cas9 gene editing technology. By knocking out the cadmium-resistance-gene cysI, the sensitivity of the biosensor to Cd(II) was further enhanced. The constructed E. coli biosensor K12-PMP-luxCDABE-△cysI exhibited good nonlinear responses to 0.005-2 mg/L Cd(II). Notably, among the three constructed E. coli biosensor, it exhibited the strongest fluorescence intensity, with the limit of detection meeting the allowable limit for Cd(II) in drinking water. Simultaneously, it could specifically detect Cd(II). Nontarget metal ions, such as Zn(II), Hg(II), and Pb(II), did not affect its performance. Furthermore, it exhibited superior performance in detecting Cd(II) in real drinking water samples by avoiding background interference, and showed excellent stability with the relative standard deviation under 5%. Thus, K12-PMP-luxCDABE-△cysI holds promise as a potential tool for the detection of Cd(II) in drinking water.

One-year mortality risk in older individuals with femoral intertrochanteric fracture: a tertiary center in China.

BACKGROUND: The accelerated growth of older individuals worldwide has increased the number of patients presenting with fragility hip fractures. Having a hip fracture can cause excess mortality, and patients with hip fracture have a higher risk of death than those without hip fracture. Most studies have treated hip fracture as a single, homogeneous condition, but hip fracture includes two major anatomic types: intertrochanteric fracture and femoral neck fracture. Few studies have specifically evaluated 1-year mortality risk in older individuals with femoral intertrochanteric fracture. The aim of this study was to evaluate 1-year mortality and factors associated with mortality in older individuals with femoral intertrochanteric fracture. METHODS: A retrospective review was conducted of 563 patients ≥ 65 years old who underwent surgery for femoral intertrochanteric fractures at our institution between January 2010 and August 2018. Patient demographics, comorbidities, and treatment were collected by retrospective chart review. Age, sex, Body Mass Index (BMI), American Society of Anesthesiologists (ASA) classification, Charlson comorbidity index (CCI), Arbeitsgemeinschaft Für Osteosynthesefragen (AO) fracture classification, haemoglobin value at admission, time to surgery, operation time, and intraoperative blood loss were risk factors to be tested. Multivariable logistic regression was used to evaluate associations between variables and death. RESULTS: Among the 563 patients, 49 died within 1 year after surgery, and the 1-year mortality rate was 8.7%. Multivariate analysis identified age > 80 years (OR = 4.038, P = 0.011), haemoglobin < 100 g/l (OR = 2.732, P = 0.002), ASA score ≥ 3 (OR = 2.551, P = 0.005), CCI ≥ 3 (OR = 18.412, P = 0.018) and time to surgery > 14 d (OR = 3.907, P = 0.030) as independent risk factors for 1-year mortality. Comorbidities such as myocardial infarction and chronic pulmonary disease were associated with 1-year mortality after adjusting for age > 80 years and time to surgery > 14 days. CONCLUSIONS: Patients over 80 years old with haemoglobin < 100 g/l, ASA score ≥ 3, CCI ≥ 3, and multiple comorbidities, especially myocardial infarction and chronic pulmonary disease before surgery, are at a higher risk of 1-year mortality. Doctors should pay more attention to these vulnerable patients, and a surgical delay greater than 14 days should be avoided.

Emergence of colistin-resistant Stenotrophomonas maltophilia with high virulence in natural aquatic environments.

The presence of Stenotrophomonas maltophilia in aquatic environments poses great health risks to immunocompromised individuals because of its multidrug resistance and resultant high mortality. However, a significant gap exists in the isolation and understanding of colistin-resistant S. maltophilia in aquatic environments. In this study, nine colistin-resistant S. maltophilia strains isolated from natural lakes were explored, and their phylogenetic relationship, biofilm formation, virulence, and antibiotic resistance profiles and underlying genetic determinants were assessed. After genome analysis, besides known multi-locus sequence typing (MLST) of ST532, new assigned ST965 and ST966 which phylogenetically clustered into soil isolates were found firstly. All the isolates exhibited resistance to multiple antibiotics, including aminoglycosides, beta-lactams, tetracyclines, and even colistin, with the highest minimum inhibitory concentration (MIC) against colistin reaching 640 mg/L. Comparative genomic analysis revealed aph(3')-Iic, blaL1, tetT, phoP, mcr-3, arnA, pmrE, and efflux pump genes as the genetic determinants underlying this multidrug resistance. Notably, the biofilm-forming capacities of the newly discovered ST965 and ST966 isolates were significant stronger than those of the known ST532 isolates (p < 0.01), resulting in the death of over 50 % of the Galleria mellonella population within 1 day of injection. The ST965 isolates demonstrated the highest virulence against G. mellonella, followed by the ST966 isolates and ST532 isolates which was phylogenetically clustered with clinical isolates, indicating that the novel S. maltophilia strains of ST965 and ST966 may pose considerable health risks to humans. Our findings provide insights into colistin-resistant S. maltophilia in aquatic environments and raise concerns about the health risks posed by the newly assigned sequence types of colistin-resistant S. maltophilia with potential high virulence in natural aquatic environments.

A mica filter enables bacterial enrichment from large volumes of natural water for sensitive monitoring of pathogens by nanopore sequencing.

Nanopore sequencing is extremely promising for the high-throughput detection of pathogenic bacteria in natural water; these bacteria may be transmitted to humans and cause waterborne infectious diseases. However, the concentration of pathogenic bacteria in natural water is too low to be detected directly by nanopore sequencing. Herein, we developed a mica filter to enrich over 85% of bacteria from > 10 L of natural water in 100 min, which led to a 102-fold improvement in the assay limits of the MinION sequencer for assessing pathogenic bacteria. Correspondingly, the sequencing time of S. Typhi detection at a concentration as low as 105 CFU/L was reduced from traditional 48 h to 3 h. The bacterial adsorption followed pseudo-first-order kinetics and the successful adsorption of bacteria to the mica filter was confirmed by scanning electron microscopy and Fourier infrared spectroscopy et al. The mica filter remained applicable to a range of water samples whose quality parameters were within the EPA standard limits for freshwater water. The mica filter is thus an effective tool for the sensitive and rapid monitoring of pathogenic bacteria by nanopore sequencing, which can provide timely alerts for waterborne transmission events.

Carbonaceous particulate matter promotes the horizontal transfer of antibiotic resistance genes.

There is growing concern about the transfer of antibiotic-resistant bacteria (ARB) and antibiotic resistance genes (ARGs) in airborne particulate matter. In this study, we investigated the effects of various types of carbonaceous particulate matter (CPM) on the transfer of ARGs in vitro. The results showed that CPM promoted the transfer of ARGs, which was related to the concentration and particle size. Compared with the control group, the transfer frequency was 95.5, 74.7, 65.4, 14.7, and 3.8 times higher in G (graphene), CB (carbon black), NGP (nanographite powder), GP1.6 (graphite powder 1.6 micron), and GP45 (graphite powder 45 micron) groups, respectively. Moreover, the transfer frequency gradually increased with the increase in CPM concentration, while there was a negative relationship between the CPM particle size and conjugative transfer frequency. In addition, the results showed that CPM could promote the transfer of ARGs by increasing ROS, as well as activating the SOS response and expression of conjugative transfer-related genes (trbBp, trfAp, korA, kroB, and trbA). These findings are indicative of the potential risk of CPM for the transfer of ARGs in the environment, enriching our understanding of environmental pollution and further raising awareness of environmental protection.

Comparative analysis of chlorine-resistant bacteria after chlorination and chloramination in drinking water treatment plants.

Chlorine-resistant bacteria (CRB) in drinking water treatment plants (DWTPs) jeopardize water quality and pose a potential risk to human health. However, the specific response of CRB to chlorination and chloramination remains uncharacterized. Therefore, we analyzed 16 S rRNA sequencing data from water samples before and after chlorination and chloramination taken between January and December 2020. Proteobacteria and Firmicutes dominated all finished water samples. After chloramination, Acinetobacter, Pseudomonas, Methylobacterium, Ralstonia, and Sphingomonas were the dominant CRB, whereas Ralstonia, Bacillus, Acinetobacter, Pseudomonas, and Enterococcus were prevalent after chlorination. Over 75% of the CRB e.g. Acinetobacter, Pseudomonas, Bacillus, and Enterococcus were shared between the chlorination and chloramination, involving potentially pathogens, such as Acinetobacter baumannii and Pseudomonas aeruginosa. Notably, certain genera such as Faecalibacterium, Geobacter, and Megasphaera were enriched as strong CRB after chloramination, whereas Vogesella, Flavobacterium, Thalassolituus, Pseudoalteromonas, and others were enriched after chlorination according to LEfSe analysis. The shared CRB correlated with temperature, pH, and turbidity, displaying a seasonal pattern with varying sensitivity to chlorination and chloramination in cold and warm seasons. These findings enhance our knowledge of the drinking water microbiome and microbial health risks, thus enabling better infectious disease control through enhanced disinfection strategies in DWTPs.

Cross-border dual-channel supply chain decision-making under random demand and tariff conditions.

Given the cross-border e-commerce import tariff and random demands, this study establishes a pricing decision model for cross-border e-commerce dual-channel supply chain, which is composed of domestic manufacturers and overseas retailers, so as to analyze the effects of import tariff and random demand on the pricing, demand and profit of cross-border e-commerce. According to the research, import tariffs have a positive correlation with retailers' retail prices and a negative correlation with manufacturers' direct prices, wholesale prices, demand and profit from direct channels, and profit from retail channels. The export tax rebate policy will lessen the negative effects of import tariffs and maximize the best choices made by manufacturers and retailers.

Nanozyme-Enhanced Electrochemical Biosensors: Mechanisms and Applications.

Nanozymes, as innovative materials, have demonstrated remarkable potential in the field of electrochemical biosensors. This article provides an overview of the mechanisms and extensive practical applications of nanozymes in electrochemical biosensors. First, the definition and characteristics of nanozymes are introduced, emphasizing their significant role in constructing efficient sensors. Subsequently, several common categories of nanozyme materials are delved into, including metal-based, carbon-based, metal-organic framework, and layered double hydroxide nanostructures, discussing their applications in electrochemical biosensors. Regarding their mechanisms, two key roles of nanozymes are particularly focused in electrochemical biosensors: selective enhancement and signal amplification, which crucially support the enhancement of sensor performance. In terms of practical applications, the widespread use of nanozyme-based electrochemical biosensors are showcased in various domains. From detecting biomolecules, pollutants, nucleic acids, proteins, to cells, providing robust means for high-sensitivity detection. Furthermore, insights into the future development of nanozyme-based electrochemical biosensors is provided, encompassing improvements and optimizations of nanozyme materials, innovative sensor design and integration, and the expansion of application fields through interdisciplinary collaboration. In conclusion, this article systematically presents the mechanisms and applications of nanozymes in electrochemical biosensors, offering valuable references and prospects for research and development in this field.

Non-invasive brain stimulation for fibromyalgia: current trends and future perspectives.

Fibromyalgia, a common and enduring pain disorder, ranks as the second most prevalent rheumatic disease after osteoarthritis. Recent years have witnessed successful treatment using non-invasive brain stimulation. Transcranial magnetic stimulation, transcranial direct current stimulation, and electroconvulsion therapy have shown promise in treating chronic pain. This article reviews the literature concerning non-invasive stimulation for fibromyalgia treatment, its mechanisms, and establishes a scientific basis for rehabilitation, and discusses the future directions for research and development prospects of these techniques are discussed.

Landscape of antibiotic resistance genes and bacterial communities in groundwater on the Tibetan Plateau, and distinguishing their difference with low-altitude counterparts.

Groundwater is a vital source of drinking water for Tibetans. Antibiotic resistance genes (ARGs) and bacterial communities in groundwater on the Tibetan Plateau remain unclear. Furthermore, the characterization of their differences between high-altitude and low-altitude groundwater is still unrevealed. Herein, 32 groundwater samples were collected on the plateau, and intra- and extracellular ARGs (iARGs and eARGs), and bacterial communities were characterised through qPCR assays to 19 ARGs and 16S rRNA sequencing. It showed top four abundant intra- and extracellular last-resort ARGs (LARGs) were blaOXA-48, mcr-1, vanA, and vanB, whereas dominant common ARGs (CARGs) were tetA and ermB, respectively. CARGs had higher abundances than LARGs, and iARGs were more frequently detected than eARGs. Proteobacteria, an invasive resident phylum, and Firmicutes dominated eDNA release. Network analysis revealed all observed LARGs co-occurred with pathogenic and non-pathogenic bacteria. Community diversity was significantly associated with longitude and elevation, while nitrate correlated with ARGs. Comparative analysis demonstrated eARG frequencies and abundances were higher at high altitudes than at low altitudes. Additionally, Acinetobacter and Pseudomonas specifically dominated at high altitudes. This study reveals the widespread prevalence of ARGs, particularly LARGs, in groundwater on the less-disturbed Tibetan Plateau and underlines the potential risks associated with the LARG-carrying bacteria. ENVIRONMENTAL IMPLICATION: Antibiotic resistance genes (ARGs), which are defined as emerging environmental contaminants, are becoming a global concern due to their ability to confer antibiotic resistance to pathogens. Our findings highlight the prevalence of ARGs, particularly LARGs, in groundwater on the Tibetan Plateau, and the possibility that naturally-occurring pathogenic and non-pathogenic bacteria carry multiple LARGs. In addition, we further reveal differences in the distribution of ARGs and bacterial community between high-altitude and low-altitude groundwater. Collectively, our findings offer an important insight into the potential public risks related to groundwater on the Tibetan Plateau.

Incidence, clinical features, and risk factors for acute pancreatitis following posterior instrumented fusion surgery for lumbar degenerative disease: a single-center, retrospective analysis of 20,929 patients.

PURPOSE: The aim of this study is to identify the incidence, clinical features, and risk factors for postoperative acute pancreatitis (PAP) after lumbar surgery. METHODS: We retrospectively analyzed patients who developed PAP after posterior lumbar fusion surgery. For each PAP patient, data were collected for four controls who underwent procedures in the same period and did not develop PAP. Statistical methods included univariate and multivariate analyses. RESULTS: Totally, 21 out of 20,929 patients were diagnosed with PAP (0.10%) after posterior lumbar fusion surgery. Patients with degenerative lumbar scoliosis were at higher risk of developing PAP (P < 0.05). With atypical clinical features, PAP occurred within 3 days (0-5) after surgery. PAP patients had significantly higher incidence of osteoporosis (47.6 vs. 22.6%, P = 0.030) and fusion of L1/2(42.9 vs. 4.3%, P = 0.010), lower albumin (42.2 ± 4.1 vs. 44.3 ± 3.2 g/L, P = 0.010), more fusion segments (median 4 vs. 3, P = 0.022), larger surgical invasiveness index (median 9 vs. 8, P = 0.007), longer operation duration (232 ± 109 vs. 185 ± 90 min, P = 0.041), greater estimated blood loss (median 600 vs. 400 mL, P = 0.025), lower intraoperative mean arterial pressure (87.2 ± 9.9 vs. 92.1 ± 8.8 mmHg, P = 0.024). Multivariate logistic regression analysis found three independent risk factors: fusion of L1/2, surgical invasiveness index > 8, and intraoperative mean arterial pressure < 90 mmHg. All patients were treated with conservative therapy and fully recovered after 8.1 (4-22) days. CONCLUSION: The incidence of PAP following posterior surgery for degenerative lumbar disease was 0.10%, and its clinical features were not typical. The fusion of L1/2, high surgical invasiveness index, and low intraoperative mean arterial pressure were independent risk factors for PAP after surgery for lumbar degenerative disease.

Spatial and temporal analysis of the seasonal dynamics of antibiotic resistance gene occurrence in recreational marine water.

People who engage in water sports in recreational marine water may be at high risk of exposure to hazardous antibiotic-resistant bacteria (ARB). However, information on the contribution of specific sources to ARB contamination in recreational marine water is still lacking. Here, we carried out monthly analyses of antibiotic resistance genes (ARGs), pathogenic bacteria and 16S rRNA sequencing data at the First Bathing Beach in Qingdao. The sampling sites were divided into four areas: swimming area, intermediate area, polluted area, and sewage outlet. Correlations between ARGs and bacterial communities among sampling sites were explored by spatial and temporal analysis. We found that all of 21 important ARG types were detected in the swimming area, with aadA (1.3 × 106 ± 2.7 × 106 genomic copies/L) and sul2 (4.3 × 105 ± 5.9 × 105 genomic copies/L) at the highest concentration. Most ARGs were detected at highest frequency and concentration in the sewage outlet and decreased from there to the swimming area. ARG correlation between these two areas was positive only in the cold season, suggesting that sewage was the main source of ARG pollution in the swimming area during that period. The ARGs ermA(1) and vanA were detected at highest frequency and concentration in the swimming area and were significantly correlated with the intestinal pathogen Enterococcus, which was more abundant here than in the surrounding areas during the warm season. Co-occurrence analysis of bacterial genera and ARGs showed that six genera were commonly correlated with ARGs in all sampling areas in the cold season, while none were found in the warm season. Our findings indicate that ARG pollution in the swimming area was also driven by sources other than sewage, especially in the warm season, which is the peak tourist season in Qingdao. These results provide a valuable basis for the implementation of effective strategies to control ARG risks in recreational waters.

A Novel Classification of Cervical Spine Trauma in Ankylosing Spondylitis and Corresponding Surgical Outcomes.

OBJECTIVE: Currently, there are no reports on the specific classification of cervical spine trauma (CST) in ankylosing spondylitis (AS) based on the trauma mechanism. In this study, we aimed to describe a novel classification of CST in AS with more details, and put forward the corresponding surgical outcomes related to different types, hoping to provide a practical reference for clinical decision-making and academic communication. METHODS: From January 2008 to December 2021, AS patients who experienced CST were retrospectively reviewed and included. Clinical data including gender, age, reason of trauma, time interval between AS diagnosis and trauma were collected. The American Spinal Injury Association (ASIA) grade system was used to describe patients' neurological status. Based on the combination of surgical experience and follow-up observation, the lower cervical spine trauma in AS patients was divided into three main types, namely single level fracture-dislocation (type 1), spinal cord injury without fracture-dislocation (type 2), and Andersson lesion (type 3). Furthermore, we performed detailed subtypes according to whether cervical spine was completely fused and the location of injury. Meanwhile, according to different approaches, surgical methods mainly included Anterior Cervical Discectomy and Fusion (ACDF), Anterior Cervical Corpectomy and Fusion (ACCF), Posterior Expansive Open-door Cervical Laminoplasty (PEOLP), Posterior Cervical Laminectomy Decompression and Fusion (PCLDF), and their combination. Postoperative general and surgery-related complications were also recorded. RESULTS: A total of 102 patients were enrolled, including 91 males and 11 females, with an average age of 51.9 years. Their average interval time between AS diagnosis and injury was 27.8 years. Patients with high-energy and low-energy trauma were 54 and 48 respectively. There were 79 patients suffering spinal cord nerve impairment after trauma. With regard to the distribution of different types, the number of patients in type 1, type 2, and type 3 were 86, 14, and two, respectively. For different types, PCLDF was the most commonly used surgical method, accounting for 55.9%, while ACCF was only applied for one time. In type 1, the frequencies of ACDF, ACCF, PCLDF, and ACDF+PCLDF were 10.5%, 1.2%, 55.8%, and 32.5%. In type 2, the frequencies of ACDF, PCLDF, ACDF+PCLDF, and PEOLP were 7.1%, 50.0%, 7.1%, 35.8%. Postoperatively, 21 patients achieved neurological function improvement. The incidences of general and surgery-related complications were 19.6% and 5.9%, respectively. All patients achieved bone fusion and durable decompression at the last follow-up. CONCLUSIONS: Our novel classification could enrich the scope of CST in AS patients and provide valuable references to the corresponding clinical management. Besides, there are strict indications of different surgical methods, factors like patient's physical condition, trauma type, surgical purpose, and expected efficacy were all required to consider before making a clinical decision.

BMSC-Derived Exosomes Alleviate Sepsis-Associated Acute Respiratory Distress Syndrome by Activating the Nrf2 Pathway to Reverse Mitochondrial Dysfunction.

Type II alveolar epithelial cell (AECII) apoptosis is one of the most vital causes of sepsis-induced acute respiratory distress syndrome (ARDS). Recent evidence has proved that bone mesenchymal stem cell-derived exosomes (BMSC-exos) can effectively reduce sepsis-induced ARDS. However, the function and molecular mechanism of BMSC-exos in sepsis-induced AECII apoptosis remain to be elucidated. In the present study, a more significant number of AECII apoptosis, high mitochondrial fission p-Drp1 protein levels, and low levels of mitochondrial biogenesis-related PGC1α, Tfam, and Nrf1 proteins accompanied with ATP content depression were confirmed in AECIIs in response to sepsis. Surprisingly, BMSC-exos successfully recovered mitochondrial biogenesis, including the upregulated expression of PGC1α, Tfam, Nrf1 proteins, and ATP contents, and prohibited p-Drp1-mediated mitochondrial fission by promoting Nrf2 expression. However, the aforementioned BMSC-exo reversal of mitochondrial dysfunction in AECIIs can be blocked by Nrf2 inhibitor ML385. Finally, BMSC-exos ameliorated the mortality rate, AECII apoptosis, inflammatory cytokine storm including HMGB1 and IL-6, and pathological lung damage in sepsis mice, which also could be prevented by ML385. These findings reveal a new mechanism of BMSC-exos in reversing mitochondrial dysfunction to alleviate AECII apoptosis, which may provide novel strategies for preventing and treating sepsis-induced ARDS.

Alpha-linolenic acid pretreatment alleviates NETs-induced alveolar macrophage pyroptosis by inhibiting pyrin inflammasome activation in a mouse model of sepsis-induced ALI/ARDS.

Background: Neutrophil extracellular traps (NETs) can cause acute lung injury (ALI)/acute respiratory distress syndrome (ARDS) by inducing macrophage pyroptosis. The purpose of this study was to find out whether pretreatment of alpha-linolenic acid (ALA) could inhibit NETs-induced macrophage pyroptosis in sepsis-induced ALI/ARDS, as well as to identify which inflammasome is involved in this process. Methods: LPS was instilled into the trachea to establish sepsis-induced ALI/ARDS in a mouse model. ​Lung injury was assessed by microscopic examination of lung tissue after hematoxylin and eosin staining, pathology score, and bronchoalveolar lavage fluid (BALF) total protein concentration. The level of NETs in lung tissue was detected by MPO-DNA ELISA. Purified NETs, extracted from peritoneal neutrophils, induced macrophage pyroptosis in vitro. Expression of pyroptosis-related proteins (Cl-caspase-1, Cl-GSDMD, ASC) and IL-1ß in the lung tissue and bone marrow-derived macrophages (BMDMs) were determined by western blotting or ELISA. Specks of Pyrin/ASC were examined by confocal immunofluorescence microscopy. Mefv (Pyrin)-/- mice were used to study the role of Pyrin in the process of sepsis-induced ALI/ARDS. Results: ALA alleviated LPS-induced lung injury. ALA reduced the level of NETs, pyroptosis-related proteins (Cl-caspase-1, Cl-GSDMD, ASC), and IL-1ß in the lung tissue of sepsis mice. In vitro, NETs increased the expression of pyroptosis-related proteins (Cl-caspase-1, Cl-GSDMD, ASC) and IL-1ß significantly in BMDMs. Pyrin protein was found to be higher and form the inflammasome with ASC in NETs challenged-BMDMs. Knockout of Mefv (Pyrin) gene fully restored the increased expression of pyroptosis-related proteins (Cl-caspase-1, Cl-GSDMD, ASC) and IL-1ß in vitro and in vivo. Lung injury was alleviated significantly in Mefv (Pyrin)-/- mice as well.​ ALA suppresses all the NETs-induced changes as mentioned above. Conclusion: Our study is the first to demonstrate Pyrin inflammasome driving NETs-induced macrophage pyroptosis, and ALA may reduce ALI/ARDS by inhibiting the activation of the Pyrin inflammasome-driven macrophage pyroptosis.

Antidepressant exposure as a source of disinfectant resistance in waterborne bacteria.

The emergence of disinfectant-resistant pathogens in water is a major threat to public health. However, whether human-consumed pharmaceuticals can induce bacterial resistance to disinfectants remains unclear. Herein, Escherichia coli was exposed to 12 antidepressants, and susceptibility of antidepressant-induced chloramphenicol (CHL)-resistant mutants to disinfectants was tested. Whole genome sequencing, global transcriptomic sequencing, and real-time quantitative polymerase chain reaction were used to elucidate the underlying mechanisms. We observed that duloxetine, fluoxetine, amitriptyline, and sertraline significantly increased the mutation frequency of E. coli against CHL by 15- to 2948-fold. The resultant mutants increased the average MIC50 of sodium hypochlorite, benzalkonium bromide, and triclosan roughly 2- to 8-fold. Consistently, marRAB and acrAB-tolC genes, together with ABC transporter genes (e.g., yddA, yadG, yojI, and mdlA), were triggered to increase the efflux of disinfectants out of the cell, while ompF was inhibited, reducing disinfectant penetration into the cell. Additionally, the occurrence of DNA mutations in marR and acrR in the mutants was observed, potentially resulting in increased synthesis of the AcrAB-TolC pump. This study indicates that pharmaceutical exposure may create disinfectant-resistant bacteria, which may then be released into water systems, providing novel insights into the potential source of water-borne disinfectant-resistant pathogens.

Simulated Gastric Acid Promotes the Horizontal Transfer of Multidrug Resistance Genes across Bacteria in the Gastrointestinal Tract at Elevated pH Levels.

The assessment of factors that can promote the transmission of antibiotic resistance genes (ARGs) across bacteria in the gastrointestinal tract is in great demand to understand the occurrence of infections related to antibiotic-resistant bacteria (ARB) in humans. However, whether acid-resistant enteric bacteria can promote ARG transmission in gastric fluid under high-pH conditions remains unknown. This study assessed the effects of simulated gastric fluid (SGF) at different pH levels on the RP4 plasmid-mediated conjugative transfer of ARGs. Moreover, transcriptomic analysis, measurement of reactive oxygen species (ROS) levels, assessment of cell membrane permeability, and real-time quantitative assessment of the expression of key genes were performed to identify the underlying mechanisms. The frequency of conjugative transfer was the highest in SGF at pH 4.5. Antidepressant consumption and certain dietary factors further negatively impacted this situation, with 5.66-fold and 4.26-fold increases in the conjugative transfer frequency being noted upon the addition of sertraline and 10% glucose, respectively, compared with that in the control group without any additives. The induction of ROS generation, the activation of cellular antioxidant systems, increases in cell membrane permeability, and the promotion of adhesive pilus formation were factors potentially contributing to the increased transfer frequency. These findings indicate that conjugative transfer could be enhanced under certain circumstances in SGF at elevated pH levels, thereby facilitating ARG transmission in the gastrointestinal tract. IMPORTANCE The low pH of gastric acid kills unwanted microorganisms, in turn affecting their inhabitation in the intestine. Hence, studies on the factors that influence antibiotic resistance gene (ARG) propagation in the gastrointestinal tract and on the underlying mechanisms are limited. In this study, we constructed a conjugative transfer model in the presence of simulated gastric fluid (SGF) and found that SGF could promote the dissemination of ARGs under high-pH conditions. Furthermore, antidepressant consumption and certain dietary factors could negatively impact this situation. Transcriptomic analysis and a reactive oxygen species assay revealed the overproduction of reactive oxygen species as a potential mechanism by which SGF could promote conjugative transfer. This finding can help provide a comprehensive understanding of the bloom of antibiotic-resistant bacteria in the body and create awareness regarding the risk of ARG transmission due to certain diseases or an improper diet and the subsequent decrease in gastric acid levels.

Trochanteric and subtrochanteric fractures irreducible by closed reduction: a retrospective study.

AIM: To explore the preoperative radiographic features and reduction methods of irreducible trochanteric and subtrochanteric fractures of the femur and to compare the perioperative characteristics and prognoses of irreducible and reducible fractures. METHODS: The data of 1235 patients with femoral trochanteric fractures surgically treated in our hospital between January 2010 and January 2021 were retrospectively analyzed. According to the inclusion criteria and exclusion criteria, 1163 cases of femoral trochanteric and subtrochanteric fractures were included in this study. Fractures in which good or acceptable reduction could not be reached by closed manipulation were defined as irreducible fractures. The preoperative radiographic features, fracture displacement patterns after closed manipulation and intraoperative reduction methods used to treat irreducible fractures were analyzed, and the perioperative characteristics and prognoses of irreducible fractures and reducible fractures were compared. RESULTS: There were 224 patients in the irreducible group and 939 patients in the reducible group. According to the radiographic features of fractures, irreducible fractures could be divided into four types: those with interlocking of the fracture, sagging of the femoral shaft, splitting of the lateral wall or medial wall, and comminution of the subtrochanteric area. Various kinds of reduction techniques were needed for different types. CONCLUSIONS: The incidence of irreducible trochanteric fractures was 15.4%, while the incidence of irreducible subtrochanteric fractures was 84.6%. According to the radiographic features of fractures, they can be divided into four types. It is important to identify irreducible fractures preoperatively and make comprehensive plans to the greatest extent possible to shorten the operation time, reduce intraoperative blood loss, and reduce the incidence of complications.

Extended chloramination significantly enriched intracellular antibiotic resistance genes in drinking water treatment plants.

Chloramination and chlorination are both strong barriers that prevent the transmission of potential pathogens to humans through drinking water. However, the comparative effects of chloramination and chlorination on the occurrence of antibiotic resistance genes (ARGs) in drinking water treatment plants (DWTPs) remain unknown. Herein, the antibiotic resistome in water before and after chloramination or chlorination was analyzed through metagenomic sequencing and then verified through quantitative real-time polymerase chain reaction (qPCR). After the treatment of 90 min, chloramination led to higher enrichment of the total relative abundance of intracellular ARGs (iARGs) in water than chlorination, whereas chlorination facilitated the release of more extracellular ARGs (eARGs) than chloramination. According to redundancy and Pearson's analyses, the total concentration of the observed iARGs in the finished water exhibited a strong positive correlation with ammonium nitrogen (NH4+-N) concentration, presenting a linear upward trend with an increase in the NH4+-N concentration. This indicated that NH4+-N is a crucial driving factor for iARG accumulation during chloramination. iARG enrichment ceases if the duration of chloramination is shortened to 40 min, suggesting that shortening the duration would be a better strategy for controlling iARG enrichment in drinking water. These findings emphasized the potential risk of antibiotic resistance after extended chloramination, shedding light on the control of transmission of antibiotic-resistant bacteria through water by optimizing disinfection procedures in DWTPs.

Comparison of femoral neck system to multiple cancellous screws and dynamic hip screws in the treatment of femoral neck fractures.

OBJECTIVE: To compare the short-term outcomes of the femoral neck system (FNS) with multiple cancellous screws (MCS) and dynamic hip screws (DHS) in the treatment of femoral neck fractures. METHODS: A retrospective analysis was performed on 157 patients with fresh femoral neck fractures treated with FNS (from September 2019 to October 2020), MCS, and DHS (from January 2018 to October 2020). According to internal fixation methods, all patients were divided into the FNS group (54 cases), MCS group (51 cases), and DHS group (52 cases). Demographic data were also collected, recorded, and compared, including the follow-up time, days of hospitalization, operation time, blood loss, intraoperative fluoroscopy times (IFT), weight-bearing time (WBT), length of femoral neck shortening (LFNS), Harris hip score, and complications (such as internal fixation failure, bone nonunion, and avascular necrosis of the femoral head) between the three groups. RESULTS: Overall, 157 patients with a mean age of 61.8 (range, 18-89) years were analyzed. In the FNS group, IFT (median 10.5) was significantly lower than that in the MCS group (median 21) and DHS group (median 20.5) (P<0.05), and WBT was significantly earlier than that of the MCS group (P<0.05). In the FNS group, the median hospitalization time, operation time, and blood loss were 2 (2, 4) days, 45 (40-59) min, and 30 (20, 50) ml, respectively. They were all significantly less than 3.5 (3, 6) days, 72 (55-88.75) min, and 50 (30, 50) ml in the DHS group, respectively (P < 0.05). There was no statistical difference in LFNS, quality of reduction, internal fixation failure rate, and Harris hip score at the latest follow-up (P>0.05). No surgical complications such as incision infection, deep infection, pulmonary embolism, or femoral head necrosis were found in any of the three groups. CONCLUSION: FNS, MCS, and DHS are all effective for femoral neck fractures. However, a lower IFT of FNS shortens the operation time. Compared to MCS, FNS has the advantages of being a simple operation, with early WBT; compared to DHS, a minimally invasive operation is the main advantage of FNS.