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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.

Background Mycoplasma pneumoniae is one of the most common causative pathogens of community-acquired pneumonia (CAP), accounting for as many as 30–50% of CAP during peak years. An early and rapid diagnostic method is key for guiding clinicians in their choice of antibiotics . Methods The recombinase-aided amplification (RAA) assay is a recently developed, rapid detection method that has been used for the detection of several pathogens. The assays were performed in a one-step single tube reaction at 39° Celsius within 15–30 min. In this study, we established an RAA assay for M. pneumoniae using clinical specimens for validation and commercial real-time PCR as the reference method. Results The analytical sensitivity of the RAA assay was 2.23 copies per reaction, and no cross-reactions with any of the other 15 related respiratory bacterial pathogens were observed. Compared with the commercial real-time PCR assay used when testing 311 respiratory specimens, the RAA assay obtained 100% sensitivity and 100% specificity with a kappa value of 1. Conclusions These results demonstrate that the proposed RAA assay will be of benefit as a faster, sensitive, and specific alternative tool for the detection of M. pneumoniae .

Background Infantile cholestasis (IC) is the most common hepatobiliary disease in infants, resulting in elevated direct bilirubin levels. Indeed, hepatointestinal circulation impacts bile acid and bilirubin metabolism. This study evaluates changes in the gut microbiota composition in children with IC and identifies abnormal metabolite profiles associated with microbial alterations. Results The gut microbiota in the IC group exhibits the higher abundance of Veillonella , Streptococcus and Clostridium spp . ( P  < 0.05), compared to healthy infants (CON) group. Moreover, the abundance of Ruminococcus , Vibrio butyricum , Eubacterium coprostanogenes group , Intestinibacter , and Faecalibacterium were lower ( P  < 0.05). In terms of microbiota-derived metabolites, the levels of fatty acids (palmitoleic, α-linolenic, arachidonic, and linoleic) ( P  < 0.05) increased and the levels of amino acids decreased in IC group. Furthermore, the abundances of Ruminococcus , Eubacterium coprostanoligenes group , Intestinibacter and Butyrivibrio are positively correlated with proline, asparagine and aspartic acid, but negatively correlated with the α-linolenic acid, linoleic acid, palmitoleic acid and arachidonic acid. For analysis of the relationship between the microbiota and clinical index, it was found that the abundance of Veillonella and Streptococcus was positively correlated with serum bile acid content ( P  < 0.05), while APTT, PT and INR were negatively correlated with Faecalibalum and Ruminococcus ( P  < 0.05). Conclusion Microbiota dysbiosis happened in IC children, which also can lead to the abnormal metabolism, thus obstructing the absorption of enteral nutrition and aggravating liver cell damage. Veillonella, Ruminococcus and Butyrivibrio may be important microbiome related with IC and need further research.

Background A malignant tracheoesophageal fistula (mTEF) is a complication of primary tumor growth or the recurrence of esophageal tumors or lung carcinoma. Patients with mTEF have lower survival and quality of life than those who do not develop this complication. Esophageal cancer (EC), a common gastrointestinal malignancy, ranks among the world's leading causes of cancer‐related death. The low survival rate in patients with EC is attributed to malnutrition, repeated aspiration, and severe infection; the mean survival duration is 2–4 months after diagnosis. This study investigated the clinical characteristics of patients with EC complicated with mTEF and the efficacy of various treatment regimens. Methods This study was a retrospective analysis of the clinical data of 51 patients with EC complicated with mTEF hospitalized at Guangdong Provincial People's Hospital from February 2007 to May 2021. Patients were divided into three groups according to their treatment regimen: a traditional medical (TM) treatment group, an esophageal intervention (EI) treatment group, and an airway intervention (AI) treatment group. Results Of the 51 patients, 22 received TM treatment, 13 received AI, and 16 received EI. The overall median survival duration was 87 days (TM group, 42 days; AI group, 108 days; EI group, 104 days) and the overall mean survival duration was 130.1 days (TM group, 88.1 days; AI group, 153.5 days; EI group, 166.1 days). Cox regression analysis revealed that the treatment regimen was an independent predictive risk factor for increased survival 1 month after treatment in patients with EC complicated with mTEF, and most symptoms were relieved in the EI and AI groups. Conclusions Interventional treatment of the esophagus and airway in patients with EC complicated with mTEF is an effective approach to improve symptoms and increase short‐term survival.

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.

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.

Mycoplasma pneumoniae (M. pneumoniae) is an important pathogen in community-acquired pneumonia. The community-acquired respiratory distress syndrome (CARDS) toxin is the only known virulence factor of M. pneumoniae. It is worth exploring whether this toxin can be used as a candidate antigen for the serodiagnosis of M. pneumoniae. In this study, the full-length, N-terminal, and C-terminal regions of the CARDS toxin were expressed and purified, and serological reactions were evaluated using ELISA. A total of 184 serum samples were collected and tested using a commercialized test kit. Eighty-seven samples were positive, and 97 samples were negative for infection. The purified recombinant proteins were used as antigens to test the serum via indirect ELISA. The sensitivity of the CARDS toxin, the N-terminal region, and the C-terminal region were 90.8%, 90.8%, and 92.0%, respectively. The specificity of the CARDS toxin, the N-terminal region, and the C-terminal region were 85.6%, 73.2%, and 93.8%, respectively. All three CARDS toxin proteins exhibited good reactivity, of which the C-terminal region had a good discrimination ability in human sera. This may have a potential diagnostic value for M. pneumoniae infections.

Background Pennisetum giganteum , an abundant, fast-growing perennial C 4 grass that belongs to the genus Pennisetum , family Poaceae, has been developed as a source of biomass for mushroom cultivation and production, as a source of forage for cattle and sheep, and as a tool to remedy soil erosion. However, having a chilling-sensitive nature, P. giganteum seedlings need to be protected while overwintering in most temperate climate regions. Results To elucidate the cold stress responses of P. giganteum , we carried out comprehensive full-length transcriptomes from leaf and root tissues under room temperature (RT) and chilling temperature (CT) using PacBio Iso-Seq long reads. We identified 196,124 and 140,766 full-length consensus transcripts in the RT and CT samples, respectively. We then systematically performed functional annotation, transcription factor identification, long non-coding RNAs (lncRNAs) prediction, and simple sequence repeat (SSR) analysis of those full-length transcriptomes. Isoform analysis revealed that alternative splicing events may be induced by cold stress in P. giganteum , and transcript variants may be involved in C18 unsaturated fatty acid biosynthesis and metabolism pathways at chilling temperature in P. giganteum . Furthermore, the fatty acid composition determination and gene expression level analysis supported that C18 unsaturated fatty acid biosynthesis and metabolism pathways may play roles during cold stress in P. giganteum . Conclusions We provide the first comprehensive full-length transcriptomic resource for the abundant and fast-growing perennial grass Pennisetum giganteum . Our results provide a useful transcriptomic resource for exploring the biological pathways involved in the cold stress responses of P. giganteum .

Background The potential role of Klebsiella pneumoniae ( K.pn ) in hypertension development has been emphasized, although the specific mechanisms have not been well understood. Bacterial extracellular vesicles (BEVs) released by Gram-negative bacteria modulate host cell functions by delivering bacterial components to host cells. Endothelial dysfunction is an important early event in the pathogenesis of hypertension, yet the impact of K.pn -secreted EVs ( K.pn EVs) on endothelial function remains unclear. This study aimed to investigate the effects of K.pn EVs on endothelial function and to elucidate the underlying mechanisms. Methods K.pn EVs were purified from the bacterial suspension using ultracentrifugation and characterized by transmission electron microscopy nanoparticle tracking analysis, and EV marker expression. Endothelium-dependent relaxation was measured using a wire myograph after in vivo or ex vivo treatment with K.pn EVs. Superoxide anion production was measured by confocal microscopy and HUVEC senescence was assessed by SA-β-gal activity. SIRT1 overexpression or activator was utilized to investigate the underlying mechanisms. Results Our data showed that K.pn significantly impaired acetylcholine-induced endothelium-dependent relaxation and increased superoxide anion production in endothelial cells in vivo. Similarly, in vivo and ex vivo studies showed that K.pn EVs caused significant endothelial dysfunction, endothelial provocation, and increased blood pressure. Further examination revealed that K.pn EVs reduced the levels of SIRT1 and p-eNOS and increased the levels of NOX2, COX-2, ET-1, and p53 in endothelial cells. Notably, overexpression or activation of SIRT1 attenuated the adverse effects and protein changes induced by K.pn EVs on endothelial cells. Conclusion This study reveals a novel role of K.pn EVs in endothelial dysfunction and dissects the relevant mechanism involved in this process, which will help to establish a comprehensive understanding of K.pn EVs in endothelial dysfunction and hypertension from a new scope.