Dynamic genomic changes in methotrexate-resistant human cancer cell lines beyond DHFR amplification suggest potential new targets for preventing drug resistance.

BACKGROUND: Although DHFR gene amplification has long been known as a major mechanism for methotrexate (MTX) resistance in cancer, the early changes and detailed development of the resistance are not yet fully understood. METHODS: We performed genomic, transcriptional and proteomic analyses of human colon cancer cells with sequentially increasing levels of MTX-resistance. RESULTS: The genomic amplification evolved in three phases (pre-amplification, homogenously staining region (HSR) and extrachromosomal DNA (ecDNA)). We confirm that genomic amplification and increased expression of DHFR, with formation of HSRs and especially ecDNAs, is the major driver of resistance. However, DHFR did not play a detectable role in the early phase. In the late phase (ecDNA), increase in FAM151B protein level may also have an important role by decreasing sensitivity to MTX. In addition, although MSH3 and ZFYVE16 may be subject to different posttranscriptional regulations and therefore protein expressions are decreased in ecDNA stages compared to HSR stages, they still play important roles in MTX resistance. CONCLUSION: The study provides a detailed evolutionary trajectory of MTX-resistance and identifies new targets, especially ecDNAs, which could help to prevent drug resistance. It also presents a proof-of-principal approach which could be applied to other cancer drug resistance studies.

Pseudorabies virus manipulates mitochondrial tryptophanyl-tRNA synthetase 2 for viral replication.

The pseudorabies virus (PRV) is identified as a double-helical DNA virus responsible for causing Aujeszky's disease, which results in considerable economic impacts globally. The enzyme tryptophanyl-tRNA synthetase 2 (WARS2), a mitochondrial protein involved in protein synthesis, is recognized for its broad expression and vital role in the translation process. The findings of our study showed an increase in both mRNA and protein levels of WARS2 following PRV infection in both cell cultures and animal models. Suppressing WARS2 expression via RNA interference in PK-15 cells led to a reduction in PRV infection rates, whereas enhancing WARS2 expression resulted in increased infection rates. Furthermore, the activation of WARS2 in response to PRV was found to be reliant on the cGAS/STING/TBK1/IRF3 signaling pathway and the interferon-alpha receptor-1, highlighting its regulation via the type I interferon signaling pathway. Further analysis revealed that reducing WARS2 levels hindered PRV's ability to promote protein and lipid synthesis. Our research provides novel evidence that WARS2 facilitates PRV infection through its management of protein and lipid levels, presenting new avenues for developing preventative and therapeutic measures against PRV infections.

Inertial Measuring System to Evaluate Gait Parameters and Dynamic Alignments for Lower-Limb Amputation Subjects.

The study aims to construct an inertial measuring system for the application of amputee subjects wearing a prosthesis. A new computation scheme to process inertial data by installing seven wireless inertial sensors on the lower limbs was implemented and validated by comparing it with an optical motion capture system. We applied this system to amputees to verify its performance for gait analysis. The gait parameters are evaluated to objectively assess the amputees' prosthesis-wearing status. The Madgwick algorithm was used in the study to correct the angular velocity deviation using acceleration data and convert it to quaternion. Further, the zero-velocity update method was applied to reconstruct patients' walking trajectories. The combination of computed walking trajectory with pelvic and lower limb joint motion enables sketching the details of motion via a stickman that helps visualize and animate the walk and gait of a test subject. Five participants with above-knee (n = 2) and below-knee (n = 3) amputations were recruited for gait analysis. Kinematic parameters were evaluated during a walking test to assess joint alignment and overall gait characteristics. Our findings support the feasibility of employing simple algorithms to achieve accurate and precise joint angle estimation and gait parameters based on wireless inertial sensor data.

Response to anti-HER2 neoadjuvant chemotherapy in HER2-positive invasive breast cancers with different HER2 FISH patterns.

AIMS: Human epidermal growth factor receptor 2 (HER2)-positive patients with breast cancer may have different HER2/CEP17 ratios and HER2 copy numbers, with inconsistent responses to anti-HER2 neoadjuvant chemotherapy (NACT). Our study aimed to explore the relationship between different HER2 fluorescence in situ hybridisation (FISH) patterns in HER2-positive patients with breast cancer and responses to anti-HER2 NACT. METHODS: 527 patients with HER2-positive invasive breast cancer who received anti-HER2 NACT from 2015 to 2022 were included and divided into three groups by FISH results, namely group A: HER2/CEP17<2.0 and HER2 copy numbers ≥6.0, HER2 immunohistochemistry 2/3+; group B: HER2/CEP17≥2.0 and HER2 copy numbers ≥4.0 and <6.0; group C: HER2/CEP17≥2.0 and HER2 copy numbers ≥6.0. We compared clinicopathological characteristics and pathological complete response (pCR) rates of different groups. RESULTS: According to HER2 FISH results, 12 patients (2.3%, 12/527) were in group A, 40 (7.6%, 40/527) were in group B and 475 (90.1%, 475/527) were in group C. The pCR rate was the lowest in group B (5.0%), while the pCR rates in group A and group C were 33.3% and 44.4%, respectively (p (group A vs. B) =0.021, p (group C vs. B) < 0.001). Both univariate and multivariate analyses revealed that HER2 FISH pattern was correlated with pCR rate (p (group C vs. B) < 0.001, p (group C vs. B) = 0.025). CONCLUSIONS: Patients with HER2/CEP17≥2.0 and HER2 copy numbers ≥4.0 and <6.0 do not benefit to the same extent from current anti-HER2 therapies as FISH-positive patients with other patterns.

Characterization and Analysis of Corrosion Resistance of Rubber Materials for Downhole Tools in a High-Stress Environment with Coupled H2S-CO2.

In the process of constructing deep natural gas wells in Sichuan and Chongqing, gas wells encounter various technical challenges such as high temperature, high pressure, and a corrosive environment containing H2S and CO2. The corrosion of rubber materials in these acidic environments can easily lead to seal failure in downhole tools. To better investigate the corrosion resistance of rubber materials in acidic environments, we utilized a dynamic cyclic corrosion experimental device capable of simulating the service conditions experienced by downhole tools under high-temperature, high-pressure multiphase flow. Corrosion-resistance tests were conducted on fluororubbers (FKM) 1, 2, 3, and HNBR (hydrogenated nitrile-butadiene rubber) under acidic conditions (80 °C and 160 °C), along with sealing corrosion tests on O-rings. These tests aimed to analyze the mechanical properties, hardness, and corrosion resistance before and after exposure to acid media as well as the sealing performance of O-rings. Ultimately, our goal was to identify suitable rubber materials for acidic pressure environments. Experimental results revealed that all four types of rubber exhibited decreased elongation at break after undergoing corrosion testing; however, fluororubber 3 demonstrated significant susceptibility to temperature effects while the other three types showed minimal impact from temperature variations. Fluororubber 1 and fluororubber 3 displayed substantial deformation levels whereas mechanical properties greatly deteriorated for fluororubber 2. Overall, HNBR showcased superior comprehensive performance.

Role of TFEB-autophagy lysosomal pathway in palmitic acid induced renal tubular epithelial cell injury.

Lysosomal dysfunction and impaired autophagic flux are involved in the pathogenesis of lipotoxicity in the kidney. Here, we investigated the role of transcription factor EB (TFEB), a master regulator of autophagy-lysosomal pathway, in palmitic acid induced renal tubular epithelial cells injury. We examined lipid accumulation, autophagic flux, expression of Ps211-TFEB, and nuclear translocation of TFEB in HK-2 cells overloaded with palmitic acid (PA). By utilizing immunohistochemistry, we detected TFEB expression in renal biopsy tissues from patients with diabetic nephropathy and normal renal tissue adjacent to surgically removed renal carcinoma (controls), as well as kidney tissues from rat fed with high-fat diet (HFD) and low-fat diet (LFD). We found significant lipid accumulation, increased apoptosis, accompanied with elevated Ps211-TFEB, decreased nuclear TFEB, reduced lysosome biogenesis and insufficient autophagy in HK-2 cells treated with PA. Kidney tissues from patients with diabetic nephropathy had lower nuclear and total levels of TFEB than that in control kidney tissues. Level of renal nuclear TFEB in HFD rats was also lower than that in LFD rats. Exogenous overexpression of TFEB increased the nuclear TFEB level in HK-2 cells treated with PA, promoted lysosomal biogenesis, improved autophagic flux, reduced lipid accumulation and apoptosis. Our results collectively indicate that PA is a strong inducer for TFEB phosphorylation modification at ser211 accompanied with lower nuclear translocation of TFEB. Impairment of TFEB-mediated lysosomal biogenesis and function by palmitic acid may lead to insufficient autophagy and promote HK-2 cells injury.

Selectively Transport and Removal of Fluoride Ion by Pillar[5]Arene Polymer-Filled Nanochannel Membrane.

Excess fluoride ions in groundwater accumulate through the roots of crops, affecting photosynthesis and inhibiting their growth. Long-term bioaccumulation also threatens human health because it is poorly degradable and toxic. Currently, one of the biggest challenges is developing a unique material that can efficiently remove fluoride ions from the environment. The excellent properties of functionalized pillar[5]arene polymer-filled nanochannel membranes were explored to address this challenge. Constructing a multistage porous nanochannel membrane, consisting of microscale etched nanochannels and nanoscale pillar[5]arene cross-linked polymer voids. A fluoride removal rate of 0.0088 mmol ⋅ L-1 ⋅ min-1 was achieved. Notably, this rate surpassed the rates observed with other control ions by a factor of 6 to 8.8. Our research provides a new direction for developing water fluoride ion removal materials.

Chronic lymphocytic inflammation with pontine perivascular enhancement responsive to steroids (CLIPPERS): contemporary advances and current controversies.

Chronic lymphocytic inflammation with pontine perivascular enhancement responsive to steroids (CLIPPERS) is an inflammatory syndrome with characteristic clinical, radiological, and pathological features, and can be effectively treated with corticosteroid-based immunotherapies. The exact pathogenesis of CLIPPERS remains unclear, and specific diagnostic biomarkers are not available. According to the 2017 diagnostic criteria, probable CLIPPERS should be considered in middle-aged patients with subacute onset of pontocerebellar symptoms and typical punctuate and curvilinear gadolinium enhancement lesions ("salt-and-pepper" appearance) located in the hindbrain (especially pons) on magnetic resonance imaging. In addition, CLIPPERS-mimics, such as central nervous system (CNS) lymphoma, and several antibody-associated autoimmune CNS diseases (e.g., myelin oligodendrocyte glycoprotein antibody-associated disease, autoimmune glial fibrillary acidic protein astrocytopathy, and anti-N-methyl-D-aspartate receptor encephalitis), should be extensively excluded. The prerequisite for definite CLIPPERS is the perivascular T-cell-predominant inflammatory infiltration observed on pathological analysis. A biopsy is strongly suggested when clinical/radiological red flags are present. Most patients with CLIPPERS respond well to corticosteroids and have a good prognosis. Long-term low-dose corticosteroid maintenance therapy or corticosteroids coupled with immunosuppressants are recommended to prevent the recurrence of the syndrome. The potential progression of CLIPPERS to lymphoma has been suggested in some cases; therefore, at least 2-year clinical and radiological follow-up is essential. Here, we critically review the recent developments and provided an update on the clinical characteristics, diagnostic criteria, differential diagnoses, and therapeutic management of CLIPPERS. We also discuss the current controversies in this context that can be resolved in future research studies.

The Validity and Feasibility of Utilizing the Photo-Assisted Dietary Intake Assessment among College Students and Elderly Individuals in China.

Dietary assessments hold significant importance within the field of public health. However, the current methods employed for dietary assessments face certain limitations and challenges that necessitate improvement. The aim of our study was to develop a reliable and practical dietary assessment tool known as photo-assisted dietary intake assessment (PAD). In order to evaluate its validity, we conducted an analysis on a sample of 71 college students' dinners at a buffet in a canteen. We compared estimates of food weights obtained through the 24-h recall (24 HR) or PAD method with those obtained through the weighing method; we also evaluated the feasibility of PAD for recording dinner intakes among a sample of college students (n = 76) and elderly individuals (n = 121). In addition, we successfully identified the dietary factors that have a significant impact on the bias observed in weight estimation. The findings of the study indicated that the PAD method exhibited a higher level of consistency with the weighing method compared to the 24 HR method. The discrepancy in D% values between cereals (14.28% vs. 40.59%, P < 0.05), vegetables (17.67% vs. 44.44%, P < 0.05), and meats (14.29% vs. 33.33%, P < 0.05) was clearly apparent. Moreover, a significant proportion of the food mass value acquired through the PAD method fell within the limits of agreement (LOAs), in closer proximity to the central horizontal line. Furthermore, vegetables, cereals, eggs, and meats, for which the primary importance lies in accuracy, exhibited a considerably higher bias with the 24 HR method compared to the PAD method (P < 0.05), implying that the PAD method has the potential to mitigate the quality bias associated with these food items in the 24 HR method. Additionally, the PAD method was well received and easily implemented by the college students and elderly individuals. In conclusion, the PAD method demonstrates a considerable level of accuracy and feasibility as a dietary assessment method that can be effectively employed across diverse populations.

The association between ambient temperature and semen quality in a Northern Peninsular Province, China.

BACKGROUND: Extreme ambient temperature has been linked to decline in males' semen quality. Although the temperature-semen quality association has been examined in certain cities of South China, how the effect size of the extreme temperature may lag over critical windows in spermatogenesis and whether the strength of association may vary in North China have yet been adequately explored. OBJECTIVES: To quantify the association between air temperature and semen quality, and identify critical exposure windows in a Northern Peninsular Province, China. MATERIALS AND METHODS: Data on semen quality in 2014-2019 were collected from the Human Sperm Bank of Institute of Women, Children and Reproductive Health, Shandong University, China. Daily meteorological data (0.01°×0.01°) were assigned to each subject's residential address. The linear mixed-effect model combined with the distributed lag nonlinear model was used to estimate the effect of ambient temperature over critical spermatogenesis windows. RESULTS: The temperature-semen quality association was inverted V-shaped, with the maximum lag being 0-45 days before ejaculation and the threshold being 9.2°C. Progressively and total motile sperm number, and total sperm number declined more substantially than other semen quality parameters. Semen quality was more sensitive to cold exposure during the epididymal storage period than the sperm motility development period. By contrast, semen quality was insensitive to heat exposure during both critical spermatogenesis windows. Impairment of certain semen quality parameters was more obvious for males with higher educational attainment and those aged over 35 years. DISCUSSION AND CONCLUSION: Exposure to non-optimal temperature is associated with decreased semen quality in North China, with the epididymal storage and sperm motility development periods more sensitive to cold exposure than heat. Older males and those with higher educations may need particular awareness.

Malonate differentially affects cell survival and confers chemoresistance in cancer cells via the induction of p53-dependent autophagy.

Metabolic network intertwines with cancerous signaling and drug responses. Malonate is a prevailing metabolite in cancer and a competitive inhibitor of succinate dehydrogenase (SDH). Recent studies showed that malonate induced reactive oxygen species (ROS)-dependent apoptosis in neuroblastoma cells, but protected cells from ischemia-reperfusion injury. We here revealed that malonate differentially regulated cell death and survival in cancer cells. While high-dose malonate triggered ROS-dependent apoptosis, the low-dose malonate induced autophagy and conferred resistance to multiple chemotherapeutic agents. Mechanistically, our results showed that malonate increased p53 stability and transcriptionally up-regulated autophagy modulator DRAM (damage-regulated autophagy modulator), thus promoting autophagy. We further proved that autophagy is required for malonate-associated chemoresistance. Collectively, our findings suggest that malonate plays a double-edge function in cancer response to stressors, and highlights a pro-cancer impact of p53-induced autophagy in response to malonate.

Safety and Efficacy of Albumin for Pump Priming in Cardiac Surgery: A Meta-Analysis.

OBJECTIVES: To assess the efficacy and safety of albumin as pump priming fluid in cardiac surgery. DESIGN: Meta-analysis of randomized controlled trials. SETTING: Each study was conducted in a surgical center or intensive care unit. PARTICIPANTS: Adult and pediatric patients undergoing cardiac surgery with cardiopulmonary bypass who received circuit priming fluids. INTERVENTIONS: Extracorporeal circuit priming with either albumin or crystalloid. MEASUREMENTS AND RESULTS: Fourteen eligible randomized controlled trials with 741 patients were included in the present meta-analysis. Albumin prime had lower bleeding (CI -202.20 to -142.88 mL, p < 0.00001) and showed a greater advantage in preserving platelet counts (CI 14.85-21.48 × 103 mm-3, p < 0.00001), maintaining colloid osmotic pressure and sustaining negative fluid balance. No significant differences were found in the remaining study outcomes. CONCLUSIONS: Albumin was shown to be safe and efficacious in extracorporeal circulation perfusion. However, its clinical advantages were not clearly highlighted, as there were no significant differences in the number of deaths, length of hospital stay, or intensive care unit duration. The results should be interpreted cautiously, as most included studies were small in scale, and the total number of participants was limited.

Myelin oligodendrocyte glycoprotein antibody and N-methyl-d-aspartate receptor antibody overlapping syndrome: insights from the recent case reports.

The overlapping of two or more types of neural autoantibodies in one patient has increasingly been documented in recent years. The coexistence of myelin oligodendrocyte glycoprotein (MOG) and N-methyl-d-aspartate receptor (NMDAR) antibodies is most common, which leads to a unique condition known as the MOG antibody and NMDAR antibody overlapping syndrome (MNOS). Here, we have reviewed the pathogenesis, clinical manifestations, paraclinical features, and treatment of MNOS. Forty-nine patients with MNOS were included in this study. They were young males with a median onset age of 23 years. No tumors were observed in the patients, and 24 of them reported prodromal symptoms. The most common clinical presentations were psychiatric symptoms (35/49) and seizures (25/49). Abnormalities on magnetic resonance imaging involved the brainstem (11/49), cerebellum (9/49), and parietal lobe (9/49). Most patients mostly responded to immunotherapy and had a good long-term prognosis. However, the overall recurrence rate of MNOS was higher than that of mono antibody-positive diseases. The existence of concurrent NMDAR antibodies should be suspected in patients with MOG antibody-associated disease having psychiatric symptoms, seizures, movement disorders, or autonomic dysfunction. Similarly, serum MOG antibody testing should be performed when patients with anti-NMDAR encephalitis present with atypical clinical manifestations, such as visual impairment and limb weakness, and neuroradiological findings, such as optic nerve, spinal cord, or infratentorial involvement or meningeal enhancement. Early detection of the syndrome and prompt treatment can be beneficial for these patients, and maintenance immunosuppressive therapy is recommended due to the high overall recurrence rate of the syndrome.

Variations in life history parameters, population dynamics, and transcriptome regulation of Brachionus plicatilis exposed to triclosan.

Triclosan (TCS) poses an ecological health risk due to its lipophilic nature, long half-life, and bioconcentration. To evaluate the toxicity of TCS on aquatic organisms, the life history parameters, population dynamics, and transcriptome regulation of Brachionus plicatilis exposed to TCS were investigated. In this study, the fecundity of rotifers was promoted by 25 µg/L of TCS and inhibited by higher concentrations (100 µg/L, 200 µg/L). The reproductive period of rotifers was shortened by 46.24 % but the post-reproductive period was prolonged by 176.47 % in 200 µg/L TCS. Both population growth and life table parameters indicated that a high concentration of TCS (200 µg/L) had negative impacts on population growth. Transcriptomic analysis showed that the effects of TCS on the life history parameters and population dynamics of rotifers were determined by regulating the expression of functional genes in cilium organization and cilium assembly and involved in pathways of focal adhesion.

The Role of Regulatory Cell Death in Vitiligo.

Vitiligo is one of the common chronic autoimmune skin diseases in clinic, which is characterized by localized or generalized depigmentation and seriously affects the physical and mental health of patients. At present, the pathogenesis of vitiligo is not clear; mainly, heredity, autoimmunity, oxidative stress, melanocyte (MC) self-destruction, and the destruction, death, or dysfunction of MCs caused by various reasons are always the core of vitiligo. Regulatory cell death (RCD) is an active and orderly death mode of cells regulated by genes, which widely exists in various life activities, plays a pivotal role in maintaining the homeostasis of the organism, and is closely related to the occurrence and development of many diseases. With the deepening of the research and understanding of RCD, people gradually found that there are many different forms of RCD in the lesions and perilesional skin of vitiligo patients, such as apoptosis, autophagy, pyroptosis, ferroptosis, and so on. Different cell death modes have different mechanisms in vitiligo, and different RCDs can interact and regulate each other. In this article, the mechanism related to RCD in the pathogenesis of vitiligo is reviewed, which provides new ideas for exploring the pathogenesis and targeted treatment of vitiligo.

Analysis of flow field characteristics of sand removal hydrocyclone applicable to solid fluidization exploitation of natural gas hydrate.

With the development of economy and society, the consumption of fossil energy is gradually increasing. In order to solve the current energy dilemma, Natural gas hydrate (NGH) is considered as an ideal alternative energy. At the same time, solid fluidization exploitation is an ideal method. However, in the process of that, sand and hydrate ore bodies enter the closed pipeline together, which will block the pipeline and increase the difficulty of exploitation. Therefore, the pre-separation of sand by hydrocyclone plays an important role in solid fluidization exploitation. In this study, the numerical simulation method was used to study the internal flow field characteristics of the hydrocyclone, and the effects of different flow rate, different flow ratio, different sand content and different particle diameter on the phase distribution were investigated. The results show that: at the same axial position, the increase of flow rate and sand content makes the sand phase more distributed at the edge of the flow field. Under the same working conditions, the sand gradually migrates to the center of the flow field with the increase of the axial distance. By calculation, it is obtained that under the optimum working condition of the flow rate is 4.83m3/h, the flow ratio is 20%, the sand content is 20%, and sand diameter is 80µm, the maximum Es is 22.1% and the minimum is 86.1%. Finally, a comprehensive analysis of the hydrocyclone in this study shows that this hydrocyclone is only applicable to rough pre-separation of sand in the process of solid fluidization exploitation. Through the study of the internal flow field characteristics and phase distribution law of the hydrocyclone, this study provides a reference for the practical engineering application of sand phase pre-separation in the solid fluidization exploitation of NGH.

Combined gas well hydrate prevention and control technology and its application.

The high pressure in some gas wells, such as those in the Xushen gas field in Daqing, China, makes them susceptible to freezing and hydrate blockages. Downhole throttling technology is widely used to reduce costs during well construction, however, due to the limitations of temperature, pressure and depth structure, this technology is sometime applied independently in some gas wells in which freezing and blockages are a frequent problem that can seriously affect production capacity. Moreover, artificial alcohol injection of 'passive plugging' to prevent hydrate formation not only consumes significant amounts of methanol but its efficiency is also dependent on factors such as weather, personnel and equipment, so it is not a continuous solution. In order to solve the above problems, the mechanism of hydrate formation was analyzed in this study, from which a combined mechanical and chemical hydrate control process was developed. OLGA software was used to design the process parameters of the novel mechanical and chemical inhibition technology for hydrate prevention and control, and also to simulate and analyze the wellhead temperature, pressure and hydrate generation once the process was implemented. Based on the results of the parameters calculation, the downhole throttle and hydrate inhibitor automatic filling device are used to realize the functions of downhole throttle depressurization and hydrate inhibitor continuous filling, reduce the wellhead pressure and hydrate generation temperature, and ensure the continuous production of gas well. This novel combination process was subsequently tested in three wells in the Daqing gas oilfield. Measurements showed that the average daily gas increase from a single well was 0.5×104m3, methanol consumption was reduced from the original maximum daily amount of 1750 kg to just 60 kg, the manual maintenance workload was reduced by 80%, and the rate of the well openings was increased from 45% to 100%. These results proved that this technology is feasible and efficient for applications in gas wells with high downhole pressure and low wellhead temperature, and, thus, provides important technical support for the prevention of gas hydrate and improvement of gas well production.

Emergency of the plasmid co-carrying blaKPC-2 and blaNDM-1 genes in carbapenem-resistant hypervirulent Klebsiella pneumoniae.

BACKGROUND: Carbapenem-resistant Klebsiella pneumoniae (CRKP) is a prevalent issue in China, with its spread primarily attributed to the presence of the plasmid-borne carbapenemase genes, blaKPC and blaNDM. However, instances of plasmids containing both blaKPC-2 and blaNDM-1have never been reported. METHODS: In this study, the genomic and microbiological characteristics of hybrid plasmids containing both blaKPC-2 and blaNDM-1 were identified in Chinese clinical CRKP isolates by Illumina combined with ONT nanopore sequencing technology. RESULTS: The newly identified plasmid was formed via IS26-mediated recombination and has been shown to be transferable to Escherichia coli. It substantially elevates the minimum inhibitory concentration (MIC) of meropenem by 4000-fold in E. coli, surpassing the MIC values observed in E. coli strains that carry either blaKPC-2 and blaNDM-1 alone, as previously demonstrated in our study. Notably, the co-occurrence of the KPC-NDM fusion plasmid and a pLVPK-like virulence plasmid was observed in these organisms. In vivo experiments revealed that the isolates harbouring the pLVPK-like virulence plasmid exhibited a significantly higher lethality rate in Galleria mellonella. CONCLUSIONS: The increased antibiotic resistance brought by this novel fusion plasmid and its accompanying virulence factors pose a serious potential threat to human health and deserve our vigilance.