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Our previous studies have shown that high alcohol-producing Klebsiella pneumoniae (HiAlc Kpn ) in the intestinal microbiome could be one of the causes of non-alcoholic fatty liver disease (NAFLD). Considering antimicrobial resistance of K. pneumoniae and dysbacteriosis caused by antibiotics, phage therapy might have potential in treatment of HiAlc Kpn -induced NAFLD, because of the specificity targeting the bacteria. Here, we clarified the effectiveness of phage therapy in male mice with HiAlc Kpn -induced steatohepatitis. Comprehensive investigations including transcriptomes and metabolomes revealed that treatment with HiAlc Kpn -specific phage was able to alleviate steatohepatitis caused by HiAlc Kpn , including hepatic dysfunction and expression of cytokines and lipogenic genes. In contrast, such treatment did not cause significantly pathological changes, either in functions of liver and kidney, or in components of gut microbiota. In addition to reducing alcohol attack, phage therapy also regulated inflammation, and lipid and carbohydrate metabolism. Our data suggest that phage therapy targeting gut microbiota is an alternative to antibiotics, with potential efficacy and safety, at least in HiAlc Kpn -caused NAFLD. Previous studies have shown that high alcohol-producing Klebsiella pneumoniae (HiAlc Kpn ) in the intestinal microbiome could be one of the causes of non-alcoholic fatty liver disease (NAFLD). Here, the authors show the effectiveness of phage in mice with HiAlc Kpn -induced NAFLD indicating phage therapy targeting gut microbiota may be an alternative to antibiotics, with potential efficacy and safety.

Capsular polysaccharides are critical virulence factors of Klebsiella pneumoniae, enabling the bacterium to evade host immune recognition and exacerbate infection. Phage-derived depolymerases, which specifically degrade these capsular polysaccharides, are increasingly recognized as a highly promising strategy for the treatment of bacterial infections. In the present study, we isolated and characterized a lytic Klebsiella pneumoniae phage, named phiTH1, and sequenced its genome. The K30-type capsular polysaccharide was identified as the receptor for phiTH1 infection. A tail fiber protein with a pectate lyase domain, Dop5, was then recognized as a potential K30-type depolymerase. Therefore, the recombinant protein Dop5 was expressed in Escherichia coli and purified, and its in vitro capsular depolymerase activity was demonstrated. Further, by using a murine aspiration pneumonia model induced by K30-type Klebsiella pneumoniae TH1, we found that Dop5 protected 80% of mice from lethal challenge with Klebsiella pneumoniae. After Dop5 treatment, the pathological damage in multiple organs of mice was alleviated, the bacterial load was reduced, and serum levels of inflammatory cytokines and complement C3 decreased, along with a significant reduction in the pathological score of the lungs. Hence, this study revealed the potential of the depolymerase Dop5 for the treatment of Klebsiella pneumoniae infections.

The adenoids and tonsils are important immune organs of the nasopharynx that often become hypertrophic in childhood because of recurrent pathogen infection. However, the differences in the immune microenvironment of adenoid hypertrophy (AH) and tonsil hypertrophy (TH) are unclear. Here, we show the epidemiological characteristics and peripheral blood cell indices of 1209 pediatric patients (1–15 years old) diagnosed with AH, and find that AH is often accompanied by TH and characterized by specific changes in immune cell types. Single-cell RNA sequencing analysis show that 12 paired AH and TH samples contain large numbers of B, T cells and some exhausted effector memory CD4 + T cells. Compared with matched TH, AH have more naïve B cells and regulatory CD4 + T cells and less plasma B cells. Weaker antigen presentation and more significant immunosuppression are also observed in AH. In contrast, the number and cytotoxicity of cytotoxic CD8 + T cells decrease with AH grade. These findings will help our understanding of the immune response to nasopharyngeal infection. Adenoids and tonsils in children can become hypertrophic because of multiple infections. Here the authors use single cell transcriptomics to assess the immune cell composition of these hypertrophic organs and find B, T and NK cells and exhausted memory CD4 T cells indicating immune changes in these organs associated with respiratory infection.

Background Klebsiella aerogenes can cause ventilator-associated pneumonia by forming biofilms, and it is frequently associated with multidrug resistance. Phages are good antibiotic alternatives with unique advantages. There has been a lack of phage therapeutic explorations, kinetic studies, and interaction mechanism research targeting K. aerogenes . Methods Plaque assay, transmission electron microscopy and whole-genome sequencing were used to determine the biology, morphology, and genomic characteristics of the phage. A mouse pneumonia model was constructed by intratracheal/endobronchial delivery of K. aerogenes to assess the therapeutic effect of phage in vivo. Bioinformatics analysis and a prokaryotic protein expression system were used to predict and identify a novel capsule depolymerase. Confocal laser scanning microscopy, Galleria mellonella larvae infection models and other experiments were performed to clarify the function of the capsule depolymerase. Results A novel lytic phage (pK4-26) was isolated from hospital sewage. It was typical of the Podoviridae family and exhibited serotype specificity, high lytic activity, and high environmental adaptability. The whole genome is 40,234 bp in length and contains 49 coding domain sequences. Genomic data show that the phage does not carry antibiotic resistance, virulence, or lysogenic genes. The phage effectively lysed K. aerogenes in vivo, reducing mortality and alleviating pneumonia without promoting obvious side effects. A novel phage-derived depolymerase was predicted and proven to be able to digest the capsule, remove biofilms, reduce bacterial virulence, and sensitize the bacteria to serum killing. Conclusions The phage pK4-26 is a good antibiotic alternative and can effectively relieve pneumonia caused by multidrug-resistant K. aerogenes . It carries a depolymerase that removes biofilms, reduces virulence, and improves intrinsic immune sensitivity.

Background/Aims: The aim of our study is to investigate the molecular mechanism by which mammalian STE20-like kinase 1 (Mst1) participates in renal I/R injury through modifying mitophagy and the AMPK-YAP signalling pathway. Methods: WT mice and Mst1-knockout mice were subjected to renal ischaemia-reperfusion (I/R) in vivo. In vitro, the hypoxia-reoxygenation model was used with renal tubular epithelial cells to mimic renal I/R injury. Mitochondrial function was monitored via western blotting and immunofluorescence. Pathway blocker and siRNA knockout technology were used to establish the role of the AMPK-YAP signalling pathway in Mst1-mediated mitochondrial apoptosis in the setting of renal I/R injury. Results: Our data demonstrated that Mst1 expression was upregulated in response to renal I/R injury in vivo, and a higher Mst1 content was positively associated with renal dysfunction and more tubular epithelial cell apoptosis. However, genetic ablation of Mst1 improved renal function, alleviated reperfusion-mediated tubular epithelial cell apoptosis, and attenuated the vulnerability of kidney to I/R injury. In vitro, Mst1 upregulation induced mitochondrial damage including mitochondrial potential reduction, ROS overloading, cyt-c liberation and caspase-9 apoptotic pathway activation. At the molecular levels, I/R-mediated mitochondrial damage via repressing mitophagy and Mst1 suppressed mitophagy via inactivating AMPK signalling pathway and dowregulating OPA1 expression. Re-activation of AMPK-YAP-OPA1 signalling pathway provided a survival advantage for the tubular epithelial cell in the context of renal I/R injury by repressing mitochondrial fission. Conclusion: Overall, our results demonstrate that the pathogenesis of renal I/R injury is closely associated with an increase in Mst1 expression and the inactive AMPK-YAP-OPA1 signalling pathway. Based on this, strategies to repress Mst1 expression and activate mitophagy could serve as therapeutic targets to treat kidney ischaemia-reperfusion injury.

Bacterial infections caused by multidrug-resistant (MDR) gram-negative strains carrying the mobile colistin resistance gene mcr-1 are serious threats to world public health due to the lack of effective treatments. Inhibition of the ATP synthase makes bacteria such as Staphylococcus aureus and Klebsiella pneumoniae more sensitive to polymyxin. This provides new strategies for treating infections caused by polymyxins-resistant bacteria carrying mcr-1 . Six mcr-1 -positive strains were isolated from clinical samples, and all were identified as Escherichia coli . Here we investigated several ATP synthase inhibitors, N , N ’-dicyclohexylcarbodiimide (DCCD), resveratrol, and piceatannol, for their antibacterial effects against the mcr-1 -positive strains combined with polymyxin B (POL). Checkerboard assay, time-kill assay, biofilm inhibition and eradication assay indicated the significant synergistic effect of ATP synthase inhibitors/POL combination in vitro. Meanwhile, mouse infection model experiment was also performed, showing a 5 log 10 reduction of the pathogen after treatment with the resveratrol/POL combination. Moreover, adding adenosine disodium triphosphate (Na 2 ATP) could inhibit the antibacterial effect of the ATP synthase inhibitors/POL combination. In conclusion, our study confirmed that inhibition of ATP production could increase the susceptibility of bacteria carrying mcr-1 to polymyxins. This provides a new strategy against polymyxins-resistant bacteria infection.

Our investigation provides insights into the interaction mechanism among hypervirulent Klebsiella pneumoniae (hvKp) (K20 serotype), phage, and the host in a mouse pneumonia model, offering a valuable reference for future research on phage pharmacokinetics. This study demonstrated that bacteriophage PK2420 exhibits promising biosafety and therapeutic efficacy against hvKp-induced pulmonary infections and dissemination in a murine model. These findings suggest that phage PK2420 may be a potential option for the clinical treatment of hvKp infections.

The objective of this paper is to perform drip irrigation scheduling for spinach (Spinacia oleracea L.) in a plastic greenhouse using the standard 20 cm evaporation pan. A drip irrigation experiment with four levels of irrigation, i.e., 0.6, 0.8, 1.0, and 1.2 times the cumulative evaporation of a 20 cm standard pan (Epan) were set up. The irrigation interval was controlled using a 20 mm Epan, and optimal irrigation water amounts of 0.8 Epan and 0.6 Epan were recommended for the spring and autumn growing seasons, respectively. Overirrigation (1.2 Epan) also led to yield losses, particularly for stem growth. In addition, a proper greenhouse index, defined as the ratio of the cumulative Epan inside and outside the greenhouse, could predict the Epan inside the greenhouse using the external Epan to a high degree of accuracy (daily data with r2 = 0.85, root mean square error (RMSE = 0.68 mm d−1), for a 4-day interval with r2 = 0.95, RMSE = 1.81 mm 4 day−1, and for the entire growth period with r2 = 1.0, RMSE = 2.40 mm). A simple and low-cost greenhouse index method could be used to formulate drip irrigation schedules for spinach in low-technology plastic greenhouses using a 20 cm standard pan outside the greenhouse.

A recent, unprecedented outbreak of human mpox virus infection has led to cases in non-African nations, and the number of confirmed or suspected cases outside of Africa has exceeded 1,000 within 5 weeks. Mpox may pose a double threat to public health in the context of the ongoing COVID-19 pandemic. It is difficult to distinguish mpox virus infection from other diseases in the early stages, and patients are contagious from the onset of nonspecific symptoms; therefore, it is crucial to develop rapid and specific diagnostic methods. The diagnosis of mpox relies on real-time polymerase chain reaction, a time-consuming method that requires a highly sophisticated thermal cycler, which makes it unsuitable for widespread use in underdeveloped areas, where the outbreak is still severe. In this study, we developed a recombinase-aided amplification (RAA) assay that can detect mpox virus within 5–10 minutes. The conserved regions of the A27L gene and F3L gene were selected as targets, as they amplify well from different mpox virus clades with no cross-reaction from other pathogens. The sensitivity of this RAA assay is 10 copies/reaction for the A27L gene and 10 2 copies/reaction for the F3L gene. When applied to simulated clinical samples, both targets showed 100% specificity, and the detection limits were consistent with the sensitivity results. Moreover, through clinical blinded sample detection, RAA exhibits the same detection power as RT-PCR. In summary, the RAA mpox assay described here exhibits rapid detection, high sensitivity and specificity, and low operational difficulty, making it suitable for mpox virus detection in less developed countries and regions.

Q fever is a worldwide zoonosis caused by Coxiella burnetii. Human Q fever is typically acquired through inhalation of contaminated aerosols, resulting in an initial pulmonary infection. In this study, BALB/c mice were infected with C. burnetii via an intratracheal (IT) route using a non-invasive aerosol pulmonary delivery device to directly place the living C. burnetii organisms into the lungs of the mice. The bacterial loads, pathological lesions, and antibody and cellular responses were analyzed and compared with those of mice infected via an intraperitoneal (IP) route. Compared with mice infected via an IP route, mice infected via an IT route exhibited a higher bacterial load and more severe pathological lesions in the heart and lungs at days 3 and 7 post-infection (pi). The levels of interferon-γ and IL-12p70 in the serum of mice infected via the IT route were significantly higher than those of mice infected via the IP route at day 3 pi. In conclusion, this murine model of acute C. burnetii infection via IT inoculation closely resembles the natural route of C. burnetii infection than that of IP injection. Thus, this newly developed model will be useful for investigating the pathogenesis and immunity of C. burnetii aerosol infection, as well as for the evaluation of therapeutic drugs and preventive vaccines of Q fever.