Dual roles of BK Polyomavirus in promoting urothelial carcinoma progression via regulating CLDN1.

Uncontrolled productive infection of BK polyomaviruses (BKV) in immunocompromised patients was reported to result in serious diseases, especially renourinary malignancies. However, the mechanism of BKV as a role of human carcinogen is still unknown. In this study, we showed that there is a significant association between BKV infection and metastasis of urothelial carcinoma (UCA). BKV-infected tumor tissues exhibit invasive histologic phenomena with vascular invasion and myometrial invasion. Then we identified that BKV promotes UCA invasion in a mode of dual regulation of tumor cells (TCs) invasion and endothelial cells (ECs) adhesion by encoding miRNAs. In cancer cells, BKV-B1-miR-5p promotes cell motility and invasiveness by directly targeting CLDN1. Moreover, exosomal-BKV-B1-miR-3p derived from BK-infected BC cells would be transferred to ECs and increase its adhesion to tumor cells by switching on the CLDN1 enhancer, which subsequently destroyed endothelial monolayers and increased permeability. In a human urothelial cancer metastasis mouse model, BK-inoculated cells exhibited higher incidence of vascular leakage and liver colonization. However, the vascular leakage and liver metastasis could be reduced when knocking down miRNAs in BK-inoculated cells. Our research delineates the bifunctional impact of BKV-encoded microRNAs on the expression of CLDN1 within both TCs and ECs, which orchestrates the establishment of a pre-metastatic niche in UCA.

Association of Blood Manganese and Preeclampsia: A Systematic Review and Meta-analysis.

A systematic review and meta-analysis was performed to comprehensively evaluate the association between manganese (Mn) level and preeclampsia (PE) during pregnancy. Relevant observational studies were retrieved by searching Medline, Web of Science, Embase, and Cochrane Library from database inception to May 25, 2023. Pooling results was performed using a random-effects model incorporating heterogeneity. This meta-analysis incorporated 18 observational studies, which included 1113 women with PE and 5480 normotensive pregnant women. Pooled results showed that compared to normotensive control, women with PE had significantly lower blood Mn concentration (standardized mean difference: -0.36, 95% confidence interval: -0.50 to -0.22, p < 0.001; I2 = 67%). Subgroup analysis showed that the results were not significantly affected by study country (African, Asian, or Western), timing of blood sampling (before, at, or after the diagnosis of PE), mean blood Mn level of controls, or numbers of confounding factors adjusted (p for subgroup analysis all > 0.05), while methods for measuring blood Mn levels might affect the results (p for subgroup difference < 0.001). Finally, pooled results of three studies showed that a high level of blood Mn was related to a low risk of PE with blood Mn analyzed in continuous (risk ratio [RR]: 0.71, 95% CI: 0.59 to 0.85, p < 0.001; I2 = 0%) and categorized variables (RR: 0.50, 95% CI: 0.30 to 0.82, p = 0.006; I2 = 32%). In conclusion, a low blood level of Mn may be associated with PE in pregnant women.

The microbiome, resistome, and their co-evolution in sewage at a hospital for infectious diseases in Shanghai, China.

The emergence of antibiotic-resistant bacteria (ARB) caused by the overuse of antibiotics severely threatens human health. Hospital sewage may be a key transmission hub for ARB. However, the complex link between the microbiome and resistomeresistance in hospital sewage remains unclear. In this study, metagenomic assembly and binning methods were used to investigate the microbial community, resistome, and association of antibiotic resistance genes (ARGs) with ARB in sewage from 10 representative sites (outpatient building, surgery building, internal medicine buildings [IMB1-4], staff dormitory, laboratory animal building, tuberculosis building [TBB], and hospital wastewater treatment plant) of a hospital in Shanghai from June 2021 to February 2022. A total of 252 ARG subtypes, belonging to 17 antibiotic classes, were identified. The relative abundance of KPC-2 was higher at IMBs and TBB than at other sites. Of the ARG-carrying contigs, 47.3%-62.6% were associated with mobile genetic elements, and the proportion of plasmid-associated ARGs was significantly higher than that of chromosome-associated ARGs. Although a similar microbiome composition was shared, certain bacteria were enriched at different sites. Potential pathogens Enterococcus B faecium and Klebsiella pneumoniae were primarily enriched in IMB2 and IMB4, respectively. The same ARGs were identified in diverse bacterial hosts (especially pathogenic bacteria), and accordingly, the latter possessed multiple ARGs. Furthermore, gene flow was frequently observed in the sewage of different buildings. The results provide crucial information on the characterization profiles of resistomes in hospital sewage in Shanghai.IMPORTANCEEnvironmental antibiotic resistance genes (ARGs) play a critical role in the emergence and spread of antimicrobial resistance, which poses a global health threat. Wastewater from healthcare facilities serves as a significant reservoir for ARGs. Here, we characterized the microbial community along with the resistome (comprising all antibiotic resistance genes) in wastewater from a specialized hospital for infectious diseases in Shanghai. Potential pathogenic bacteria (e.g., Escherichia coli, Pseudomonas aeruginosa, Klebsiella pneumoniae, Enterococcus B faecium) were frequently detected in hospital wastewater and carried multiple ARGs. A complex link between microbiome and resistome was observed in the wastewater of this hospital. The monitoring of ARGs and antibiotic-resistant bacteria (ARB) in hospital wastewater might be of great significance for preventing the spread of ARB.

Recent advances of responsive scaffolds in bone tissue engineering.

The investigation of bone defect repair has been a significant focus in clinical research. The gradual progress and utilization of different scaffolds for bone repair have been facilitated by advancements in material science and tissue engineering. In recent times, the attainment of precise regulation and targeted drug release has emerged as a crucial concern in bone tissue engineering. As a result, we present a comprehensive review of recent developments in responsive scaffolds pertaining to the field of bone defect repair. The objective of this review is to provide a comprehensive summary and forecast of prospects, thereby contributing novel insights to the field of bone defect repair.

Glycolytic enzyme PFKFB3 regulates sphingosine 1-phosphate receptor 1 in proangiogenic glomerular endothelial cells under diabetic condition.

Glomerular angiogenesis is a characteristic feature of diabetic nephropathy (DN). Enhanced glycolysis plays a crucial role in angiogenesis. The present study was designed to investigate the role of glycolysis in glomerular endothelial cells (GECs) in a mouse model of DN. Mouse renal cortex and isolated glomerular cells were collected for single-cell and RNA sequencing. Cultured GECs were exposed to high glucose in the presence (proangiogenic) and absence of a vascular sprouting regimen. MicroRNA-590-3p was delivered by lipofectamine in vivo and in vitro. In the present study, a subgroup of GECs with proangiogenic features was identified in diabetic kidneys by using sequencing analyses. In cultured proangiogenic GECs, high glucose increased glycolysis and phosphofructokinase/fructose bisphosphatase 3 (PFKFB3) protein expression, which were inhibited by overexpressing miRNA-590-3p. Mimics of miRNA-590-3p also increased receptor for sphingosine 1-phosphate (S1pR1) expression, an angiogenesis regulator, in proangiogenic GECs challenged with high glucose. Inhibition of PFKFB3 by pharmacological and genetic approaches upregulated S1pR1 protein in vitro. Mimics of miRNA-590-3p significantly reduced migration and angiogenic potential in proangiogenic GECs challenged with high glucose. Ten-week-old type 2 diabetic mice had elevated urinary albumin levels, reduced renal cortex miRNA-590-3p expression, and disarrangement of glomerular endothelial cell fenestration. Overexpressing miRNA-590-3p via perirenal adipose tissue injection restored endothelial cell fenestration and reduced urinary albumin levels in diabetic mice. Therefore, the present study identifies a subgroup of GECs with proangiogenic features in mice with DN. Local administration of miRNA-590-3p mimics reduces glycolytic rate and upregulates S1pR1 protein expression in proangiogenic GECs. The protective effects of miRNA-590-3p provide therapeutic potential in DN treatment.NEW & NOTEWORTHY Proangiogenetic glomerular endothelial cells (GECs) are activated in diabetic nephropathy. High glucose upregulates glycolytic enzyme phosphofructokinase/fructose bisphosphatase 3 (PFKFB3) in proangiogenetic cells. PFKFB3 protects the glomerular filtration barrier by targeting endothelial S1pR1. MiRNA-590-3p restores endothelial cell function and mitigates diabetic nephropathy.

Regulations of phage therapy across the world.

Phage therapy, a century-long treatment targeting bacterial infection, was widely abandoned after the clinical availability of antibiotics in the mid-20th century. However, the crisis of antimicrobial resistance today led to its revival in many countries. While many articles dive into its clinical application now, little research is presenting phage therapy from a regulatory perspective. Here, we focus on the regulations of phage therapy by dividing sections into Eastern Europe where it was never abandoned and Western Europe, Australia, the United States, India, and China where it only re-attracted researchers' attention in recent decades. New insights about its regulations in China are provided as little English literature has specifically discussed this previously. Ultimately, by introducing the regulations in phage therapy for human health across representative countries, we hope to provide ideas of how countries may borrow each other's adapting legislation in phage therapy to best overcome the current regulatory hurdles.

Development of phage resistance in multidrug-resistant Klebsiella pneumoniae is associated with reduced virulence: a case report of a personalised phage therapy.

OBJECTIVES: Phage-resistant bacteria often emerge rapidly when performing phage therapy. However, the relationship between the emergence of phage-resistant bacteria and improvements in clinical symptoms is still poorly understood. METHODS: An inpatient developed a pulmonary infection caused by multidrug-resistant Klebsiella pneumoniae. He received a first course of treatment with a single nebulized phage (ΦKp_GWPB35) targeted at his bacterial isolate of Kp7450. After 14 days, he received a second course of treatment with a phage cocktail (ΦKp_GWPB35+ΦKp_GWPA139). Antibiotic treatment was continued throughout the course of phage therapy. Whole-genome analysis was used to identify mutations in phage-resistant strains. Mutated genes associated with resistance were further analysed by generating knockouts of Kp7450 and by measuring phage adsorption rates of bacteria treated with proteinase K and periodate. Bacterial virulence was evaluated in mouse and zebrafish infection models. RESULTS: Phage-resistant Klebsiella pneumoniae strains emerged after the second phage treatment. Comparative genomic analyses revealed that fabF was deleted in phage-resistant strains. The fabF knockout strain (Kp7450ΔfabF) resulted in an altered structure of lipopolysaccharide (LPS), which was identified as the host receptor for the therapeutic phages. Virulence evaluations in mice and zebrafish models showed that LPS was the main determinant of virulence in Kp7450 and alteration of LPS structure in Kp7450ΔfabF, and the bacteriophage-resistant strains reduced their virulence at cost. DISCUSSION: This study may shed light on the mechanism by which some patients experience clinical improvement in their symptoms post phage therapy, despite the incomplete elimination of pathogenic bacteria.

Synthesis and Evaluation of Interfacial Properties and Carbon Capture Capacities of the Imidazolium-Based Ionic Liquid Surfactant.

Ionic liquid as a chemical flooding agent has broad application prospect in enhancing oil recovery. In this study, a bifunctional imidazolium-based ionic liquid surfactant was synthesized, and its surface-active, emulsification capacity, and CO2 capture performance were investigated. The results show that the synthesized ionic liquid surfactant combines the characteristics of reducing interfacial tension, emulsification, and CO2 capture. The IFT values for [C12mim][Br], [C14mim][Br], and [C16mim][Br] could decrease from 32.74 mN/m to 3.17, 0.54, and 0.051 mN/m, respectively, with increasing concentration. In addition, the emulsification index values are 0.597 for [C16mim][Br], 0.48 for [C14mim][Br], and 0.259 for [C12mim][Br]. The surface-active and emulsification capacity of ionic liquid surfactants improved with the increase in alkyl chain length. Furthermore, the absorption capacities reach 0.48 mol CO2 per mol of ionic liquid surfactant at 0.1 MPa and 25 °C. This work provides theoretical support for further CCUS-EOR research and the application of ionic liquid surfactants.

Monomethyl fumarate prevents alloimmune rejection in mouse heart transplantation by inducing tolerogenic dendritic cells.

Dendritic cells (DCs) are important targets for eliciting allograft rejection after transplantation. Previous studies have demonstrated that metabolic reprogramming of DCs can transform their immune functions and induce their differentiation into tolerogenic DCs. In this study, we aim to investigate the protective effects and mechanisms of monomethyl fumarate (MMF), a bioactive metabolite of fumaric acid esters, in a mouse model of allogeneic heart transplantation. Bone marrow-derived DCs are harvested and treated with MMF to determine the impact of MMF on the phenotype and immunosuppressive function of DCs by flow cytometry and T-cell proliferation assays. RNA sequencing and Seahorse analyses are performed for mature DCs and MMF-treated DCs (MMF-DCs) to investigate the underlying mechanism. Our results show that MMF prolongs the survival time of heart grafts and inhibits the activation of DCs in vivo. MMF-DCs exhibit a tolerogenic phenotype and function in vitro. RNA sequencing and Seahorse analyses reveal that MMF activates the Nrf2 pathway and mediates metabolic reprogramming. Additionally, MMF-DC infusion prolongs cardiac allograft survival, induces regulatory T cells, and inhibits T-cell activation. MMF prevents allograft rejection in mouse heart transplantation by inducing tolerogenic DCs.

Characterization of a Filamentous Phage, Vaf1, from Vibrio alginolyticus AP-1.

Filamentous phages are ubiquitously distributed in the global oceans. However, little is known about their biological contribution to their host's genetic and phenotypic diversity. In this study, a filamentous phage, Vaf1, was isolated and characterized from the emerging marine pathogen strain Vibrio alginolyticus AP-1. We explored the effects of the resident phage Vaf1 on the host physiology under diverse conditions by precisely deleting the entire phage Vaf1. Our results demonstrate that the presence of phage Vaf1 significantly increased biofilm formation, swarming motility, and contact-dependent competition. Furthermore, the gene expression profile suggests that several phage genes were upregulated in response to low-nutrient conditions. Unexpectedly, an in vivo study of zebrafish shows that fish infected with strain ΔVaf1 survived longer than those infected with wild-type strain AP-1, indicating that Vaf1 contributes to the virulence of V. alginolyticus. Together, our results provide direct evidence for the effect of Vaf1 phage-mediated phenotypic changes in marine bacteria V. alginolyticus. This further emphasizes the impressive complexity and diversity that filamentous phage-host interactions pose and the challenges associated with bacterial disease control in marine aquaculture. IMPORTANCE Non-lytic filamentous phages can replicate without killing their host, establishing long-term persistence within the bacterial host. In contrast to the well-studied CTXφ phage of the human-pathogenic Vibrio cholerae, little is known about the filamentous phage Vaf1 and its biological role in host fitness. In this study, we constructed a filamentous phage-deleted strain, ΔVaf1, and provided direct evidence on how an intact phage, φVaf1, belonging to the family Inoviridae, helps the bacterial host AP-1 to overcome adverse environmental conditions. Our results likely open new avenues for fundamental studies on how filamentous phage-host interactions regulate different aspects of Vibrio cell behaviors.

Inhibition of ALKBH5 attenuates I/R-induced renal injury in male mice by promoting Ccl28 m6A modification and increasing Treg recruitment.

Ischemia reperfusion injury (IRI) is a common cause of acute kidney injury (AKI). The role of N6-methyladenosine (m6A) modification in AKI remains unclear. Here, we characterize the role of AlkB homolog 5 (ALKBH5) and m6A modification in an I/R-induced renal injury model in male mice. Alkbh5-knockout mice exhibit milder pathological damage and better renal function than wild-type mice post-IRI, whereas Alkbh5-knockin mice show contrary results. Also conditional knockout of Alkbh5 in the tubular epithelial cells alleviates I/R-induced AKI and fibrosis. CCL28 is identified as a target of ALKBH5. Furthermore, Ccl28 mRNA stability increases with Alkbh5 deficiency, mediating by the binding of insulin-like growth factor 2 binding protein 2. Treg recruitment is upregulated and inflammatory cells are inhibited by the increased CCL28 level in IRI-Alkbh5fl/flKspCre mice. The ALKBH5 inhibitor IOX1 exhibits protective effects against I/R-induced AKI. In summary, inhibition of ALKBH5 promotes the m6A modifications of Ccl28 mRNA, enhancing its stability, and regulating the Treg/inflammatory cell axis. ALKBH5 and this axis is a potential AKI treatment target.

Rapid point-of-care detection of BK virus in urine by an HFman probe-based loop-mediated isothermal amplification assay and a finger-driven microfluidic chip.

Background: BK virus (BKV)-associated nephropathy (BKVN) is one of the leading causes of renal dysfunction and graft loss in renal transplant recipients. Early monitoring of BKV in urine is crucial to minimize the deleterious effects caused by this virus on preservation of graft function. Methods: We report a simple, rapid, sensitive loop-mediated isothermal amplification (LAMP) assay using an HFman probe for detecting BKV in urine. To evaluate the performance of the assay, a comparison of the HFman probe-based LAMP (HF-LAMP) assay with two qPCR assays was performed using urine samples from 132 HIV-1 infected individuals. We further evaluated the performance of HF-LAMP directly using the urine samples from these HIV-1 infected individuals and 30 kidney transplant recipients without DNA extraction. Furthermore, we combined the HF-LAMP assay with a portable finger-driven microfluidic chip for point-of-care testing (POCT). Results: The assay has high specificity and sensitivity with a limit of detection (LOD) of 12 copies/reaction and can be completed within 30 min. When the DNA was extracted, the HF-LAMP assay showed an equivalent and potentially even higher sensitivity (93.5%) than the qPCR assays (74.2-87.1%) for 132 urine samples from HIV-1 infected individuals. The HF-LAMP assay can be applied in an extraction-free format and can be completed within 45 min using a simple heat block. Although some decreased performance was seen on urine samples from HIV-1 infected individuals, the sensitivity, specificity, and accuracy of the extraction-free BKV HF-LAMP assay were 95%, 100%, and 96.7% for 30 clinical urine samples from kidney transplant recipients, respectively. Conclusion: The assay has high specificity and sensitivity. Combined with a portable finger-driven microfluidic chip for easy detection, this method shows great potential for POCT detection of BKV.

Formyl peptide receptor 2 activation by mitochondrial formyl peptides stimulates the neutrophil proinflammatory response via the ERK pathway and exacerbates ischemia-reperfusion injury.

BACKGROUND: Ischemia-reperfusion injury (IRI) is an inevitable process in renal transplantation that significantly increases the risk of delayed graft function, acute rejection, and even graft loss. Formyl peptide receptor 2 (FPR2) is an important receptor in multiple septic and aseptic injuries, but its functions in kidney IRI are still unclear. This study was designed to reveal the pathological role of FPR2 in kidney IRI and its functional mechanisms. METHODS: To explore the mechanism of FPR2 in kidney IRI, the model rats were sacrificed after IRI surgery. Immunofluorescence, enzyme-linked immunosorbent assays, and western blotting were used to detect differences in the expression of FPR2 and its ligands between the IRI and control groups. WRW4 (WRWWWW-NH2), a specific antagonist of FPR2, was administered to kidney IRI rats. Kidney function and pathological damage were detected to assess kidney injury and recovery. Flow cytometry was used to quantitatively compare neutrophil infiltration among the experimental groups. Mitochondrial formyl peptides (mtFPs) were synthesized and administered to primary rat neutrophils together with the specific FPR family antagonist WRW4 to verify our hypothesis in vitro. Western blotting and cell function assays were used to examine the functions and signaling pathways that FPR2 mediates in neutrophils. RESULTS: FPR2 was activated mainly by mtFPs during the acute phase of IRI, mediating neutrophil migration and reactive oxygen species production in the rat kidney through the ERK1/2 pathway. FPR2 blockade in the early phase protected rat kidneys from IRI. CONCLUSIONS: mtFPs activated FPR2 during the acute phase of IRI and mediated rat kidney injury by activating the migration and reactive oxygen species generation of neutrophils through the ERK1/2 pathway.

First-in-human application of double-stranded RNA bacteriophage in the treatment of pulmonary Pseudomonas aeruginosa infection.

A double-stranded RNA (dsRNA) phage phiYY is able to kill a pyomelanin-producing Pseudomonas aeruginosa strain, which was isolated from a 40-year-old man with interstitial lung disease (ILD) and chronic lung infection. Phage therapy was used as a last resort for this patient. The three-course nebulized phiYY treatment was used to reduce the bacterial burden and clinical symptoms of the patient. Recurrences of P. aeruginosa infections were observed 1-3 days post phage therapy. The recurrent isolates exhibited distinct antibiotic-susceptibility profiles compared with the original strain yet were still susceptible to phiYY. This assay represents the application of dsRNA phage in the treatment of chronic lung infection, albeit the safety and efficacy of the dsRNA phage require further assessment.

Basiliximab for the therapy of acute T cell-mediated rejection in kidney transplant recipient with BK virus infection: A case report.

A 66-year-old Chinese man underwent a deceased donor kidney transplantation. Induction-immunosuppressive protocol consisted of basiliximab (BAS) and methyl prednisolone (MP), followed by maintenance immunosuppression with cyclosporin (CsA), mycophenolate mofetil (MMF), and prednisone (PED). The patient's post-transplantation course was almost uneventful, and the graft was functioning well [serum creatinine (Scr) 2.15 mg/dL]. The MMF and CsA doses were decreased 1-month post-operative as the BK virus activation was serologically positive. His Scr was elevated to 2.45 mg/dL 45 days after the transplant. A graft biopsy showed BKV nephropathy (BKVN) and acute T cell-mediated rejection (TCMR) Banff grade IIA (I2, t2, ptc2, v1, c4d1, g0, and SV40 positive). The conventional anti-rejection therapy could deteriorate his BKVN, therefore, we administered BAS to eliminate activated graft-infiltrating T cells and combined with low-dose steroid. He responded well to the therapy after two doses of BAS were given, and the kidney graft status has been stable (recent Scr 2.1 mg/dL).

Combating the SARS-CoV-2 Omicron (BA.1) and BA.2 with potent bispecific antibodies engineered from non-Omicron neutralizing antibodies.

The highly mutated and transmissible Omicron (BA.1) and its more contagious lineage BA.2 have provoked serious concerns over their decreased sensitivity to the current COVID-19 vaccines and evasion from most anti-SARS-CoV-2 neutralizing antibodies (NAbs). In this study, we explored the possibility of combating the Omicron and BA.2 by constructing bispecific antibodies based on non-Omicron NAbs. We engineered 10 IgG-like bispecific antibodies with non-Omicron NAbs named GW01, 16L9, 4L12, and REGN10987 by fusing the single-chain variable fragments (scFvs) of two antibodies through a linker and then connecting them to the Fc region of IgG1. Surprisingly, 8 out of 10 bispecific antibodies showed high binding affinities to the Omicron receptor-binding domain (RBD) and exhibited extreme breadth and potency against pseudotyped SARS-CoV-2 variants of concern (VOCs) including Omicron and BA.2, with geometric mean of 50% inhibitory concentration (GM IC50) values ranging from 4.5 ng/mL to 103.94 ng/mL, as well as the authentic BA.1.1. Six bispecific antibodies containing the cross-NAb GW01 not only neutralized Omicron and BA.2, but also neutralized the sarbecoviruses including SARS-CoV and SARS-related coronaviruses (SARSr-CoVs) RS3367 and WIV1, with GM IC50 ranging from 11.6 ng/mL to 103.9 ng/mL. Mapping analyses of 42 spike (S) variant single mutants of Omicron and BA.2 elucidated that these bispecific antibodies accommodated the S371L/F mutations, which were resistant to most of the non-Omicron NAbs. A cryo-electron microscopy (cryo-EM) structure study of the representative bispecific antibody GW01-16L9 (FD01) in its native full-length IgG form in complex with the Omicron S trimer revealed 5 distinct trimers and one novel trimer dimer conformation. 16L9 scFv binds the receptor-binding motif (RBM), while GW01 scFv binds a epitope outside the RBM. Two scFvs of the bispecific antibody synergistically induced the RBD-down conformation into 3 RBD-up conformation, improved the affinity between IgG and the Omicron RBD, induced the formation of trimer dimer, and inhibited RBD binding to ACE2. The trimer dimer conformation might induce the aggregation of virions and contribute to the neutralization ability of FD01. These novel bispecific antibodies are strong candidates for the treatment and prevention of infection with the Omicron, BA.2, VOCs, and other sarbecoviruses. Engineering bispecific antibodies based on non-Omicron NAbs could turn the majority of NAbs into a powerful arsenal to aid the battle against the pandemic.

Estimation of Mycophenolic Acid Exposure in Chinese Renal Transplant Patients by a Joint Deep Learning Model.

BACKGROUND: To predict mycophenolic acid (MPA) exposure in renal transplant recipients using a deep learning model based on a convolutional neural network with bilateral long short-term memory and attention methods. METHODS: A total of 172 Chinese renal transplant patients were enrolled in this study. The patients were divided into a training group (n = 138, Ruijin Hospital) and a validation group (n = 34, Zhongshan Hospital). Fourteen days after renal transplantation, rich blood samples were collected 0-12 hours after MPA administration. The plasma concentration of total MPA was measured using an enzyme-multiplied immunoassay technique. A limited sampling strategy based on a convolutional neural network-long short-term memory with attention (CALS) model for the prediction of the area under the concentration curve (AUC) of MPA was established. The established model was verified using the data from the validation group. The model performance was compared with that obtained from multiple linear regression (MLR) and maximum a posteriori (MAP) methods. RESULTS: The MPA AUC 0-12 of the training and validation groups was 54.28 ± 18.42 and 41.25 ± 14.53 µg·ml -1 ·h, respectively. MPA plasma concentration after 2 (C 2 ), 6 (C 6 ), and 8 (C 8 ) hours of administration was the most significant factor for MPA AUC 0-12 . The predictive performance of AUC 0-12 estimated using the CALS model of the validation group was better than the MLR and MAP methods in previous studies (r 2 = 0.71, mean prediction error = 4.79, and mean absolute prediction error = 14.60). CONCLUSIONS: The CALS model established in this study was reliable for predicting MPA AUC 0-12 in Chinese renal transplant patients administered mycophenolate mofetil and enteric-coated mycophenolic acid sodium and may have good generalization ability for application in other data sets.

Characterization of Phage Resistance and Their Impacts on Bacterial Fitness in Pseudomonas aeruginosa.

The emergence and spread of antibiotic resistance pose serious environmental and health challenges. Attention has been drawn to phage therapy as an alternative approach to combat antibiotic resistance with immense potential. However, one of the obstacles to phage therapy is phage resistance, and it can be acquired through genetic mutations, followed by consequences of phenotypic variations. Therefore, understanding the mechanisms underlying phage-host interactions will provide us with greater detail on how to optimize phage therapy. In this study, three lytic phages (phipa2, phipa4, and phipa10) were isolated to investigate phage resistance and the potential fitness trade-offs in Pseudomonas aeruginosa. Specifically, in phage-resistant mutants phipa2-R and phipa4-R, mutations in conferring resistance occurred in genes pilT and pilB, both essential for type IV pili (T4P) biosynthesis. In the phage-resistant mutant phipa10-R, a large chromosomal deletion of ~294 kb, including the hmgA (homogentisate 1,2-dioxygenase) and galU (UTP-glucose-1-phosphate uridylyltransferase) genes, was observed and conferred phage phipa10 resistance. Further, we show examples of associated trade-offs in these phage-resistant mutations, e.g., impaired motility, reduced biofilm formation, and increased antibiotic susceptibility. Collectively, our study sheds light on resistance-mediated genetic mutations and their pleiotropic phenotypes, further emphasizing the impressive complexity and diversity of phage-host interactions and the challenges they pose when controlling bacterial diseases in this important pathogen. IMPORTANCE Battling phage resistance is one of the main challenges faced by phage therapy. To overcome this challenge, detailed information about the mechanisms of phage-host interactions is required to understand the bacterial evolutionary processes. In this study, we identified mutations in key steps of type IV pili (T4P) and O-antigen biosynthesis leading to phage resistance and provided new evidence on how phage predation contributed toward host phenotypes and fitness variations. Together, our results add further fundamental knowledge on phage-host interactions and how they regulate different aspects of Pseudomonas cell behaviors.

Characterization of Pseudomonas aeruginosa Bacteriophage L5 Which Requires Type IV Pili for Infection.

Pseudomonas aeruginosa is a common opportunistic human pathogen. With the emergence of multidrug-resistant (MDR) clinical infection of P. aeruginosa, phage therapy has received renewed attention in treating P. aeruginosa infections. Moreover, a detailed understanding of the host receptor of lytic phage is crucial for selecting proper phages for therapy. Here, we describe the characterization of the P. aeruginosa bacteriophage L5 with a double-stranded DNA genome of 42,925 bp. The genomic characteristics indicate that L5 is a lytic bacteriophage belonging to the subfamily Autographivirinae. In addition, the phage receptors for L5 were also identified as type IV pili, because the mutation of pilZ, which is involved in pili synthesis, resists phage infection, while the complementation of pilZ restored its phage sensitivity. This research reveals that L5 is a potential phage therapy candidate for the treatment of P. aeruginosa infection.