Phage-derived depolymerases represent a promising antibiotic alternative for treating Klebsiella pneumoniae infections. Depolymerases can increase the sensitivity of bacteria to the host immune system and complement-mediated killing by specifically degrading capsular polysaccharides (CPS). However, the impact of depolymerases on bacterial biofilm formation remains unclear. This study found that depolymerase treatment significantly enhances the biofilm formation capability of hypervirulent K. pneumoniae (hvKp) strains, which inherently exhibit relatively weak biofilm formation due to their thick capsular polysaccharide (CPS) layers. Further investigation revealed that depolymerase-mediated CPS degradation relieved its repression on the type 3 fimbriae gene cluster mrkABCDF, thereby promoting biofilm formation. In vivo experiments in mice also showed that CPS can inhibit virulence functions associated with type 3 fimbriae. Furthermore, we found that CPS-mediated biofilm inhibition appears to be a common phenomenon among hvKp strains. In summary, by elucidating the dual role of depolymerase in modulating both virulence and biofilm in hvKp, our work reveals a potential interaction between CPS and type 3 fimbriae, providing deeper insights into the pathogenicity of this clinically important bacterium.

Mpox virus, the causative agent of mpox infections, has been responsible for numerous disease outbreaks globally since 2022, creating an urgent need for rapid and sensitive detection method. This study aimed to develop and validate a droplet digital polymerase chain reaction (ddPCR) assay for mpox virus detection targeting the viral A27L gene, and to systematically compare its performance with that of real-time quantitative polymerase chain reaction (qPCR) using simulated and clinical samples. Optimal reaction conditions for ddPCR were determined to be an annealing temperature of 56 degrees C with primer and probe concentrations of 200 and 100 nM, respectively. ddPCR showed 100 % specificity for each pathogen strain tested within the scope of this study, with a limit of detection (LOD) of 2.0 copies/mu L DNA sample, making it 32.4-fold sensitive than the LOD of qPCR (64.8 copies/ mu L). ddPCR exhibited robust detection repeatability (intra-assay CV: 2.2 %-6.1 %) and reproducibility (inter-assay CV: 2.4 %-10.5 %). Overall, the established A27L-targeting ddPCR assay efficiently and accurately identified mpox virus in various types of clinical and simulated samples with higher sensitivity than qPCR. This assay presents a reliable new method for identifying the causative infections of mpox and guiding treatment options.

Klebsiella pneumoniae (Kpn) is a common opportunistic pathogen, with hypervirulent strains (hvKpn) posing significant risks even to healthy individuals. Traditional clinical diagnostics often use real-time quantitative polymerase chain reaction (PCR) with TaqMan probes, which are costly and unstable due to their reliance on fluorophore and quencher molecules. This study introduces a novel probe using silicon-doped carbon quantum dots (Si-QDs) and single-layer graphene oxides (GOs), forming the Si-QD-probe-GO. This new probe offers excellent selectivity, sensitivity, stability, low photobleaching, and cost-effectiveness. A sensor array targeting khe, peg-344, and iucA genes is developed to detect Kpn and differentiate between classical Kpn (cKpn) and hvKpn. Using a partial least-squares discriminant analysis (PLS-DA) model, the system achieves 100% accuracy in distinguishing hvKpn from cKpn. The study also employs droplet digital PCR (ddPCR) with microfluidic technology to enhance sensitivity and quantification, achieving a detection limit of 5 CFU/mL. In a mouse pneumonia model, the PLS-DA model accurately identifies hvKpn in lung samples. The ddPCR quantification using Si-QD-probe-GO shows strong correlation with traditional culture methods. This research presents a promising ddPCR system for precise detection and virulence identification of Kpn, offering a new direction for clinical diagnostics.

Paraquat (PQ), a widely used non-selective herbicide, poses significant health risks, with as little as 30 mL potentially causing fatal poisoning due to acute lung injury (ALI). Despite frequent cases, effective treatments for PQ poisoning are limited, highlighting the need for in-depth investigation into the mechanisms underlying PQ-induced ALI. Resveratrol was identified as a therapeutic agent based on the key differentially regulated pathways. This study investigates the potential therapeutic effects of Resveratrol on Paraquat-induced acute lung injury (ALI) through both in vivo and in vitro experiments. In the in vivo experiment, Resveratrol significantly prolonged the survival of mice (P < 0.001) compared to the control group, suggesting its protective role in the pathogenesis of ALI. In the in vitro experiment using A549 cells, Resveratrol effectively alleviated Paraquat-induced oxidative stress and iron ion deposition, significantly increasing cell viability (P < 0.05). Through transcriptomic and proteomic analysis, Resveratrol was found to upregulate genes and proteins associated with inflammation, cell death, and lipid peroxidation, while downregulating those related to lipid peroxidation and iron. Furthermore, cell functional analysis revealed that Resveratrol regulates mitochondrial function, the cell cycle, and inflammatory signaling pathways, improving the pathological state of both mice and A549 cells. In conclusion, Resveratrol modulates multiple mechanisms, including lipid metabolism, cell cycle regulation, inflammatory signaling, and cell toxicity pathways, to significantly alleviate Paraquat-induced lung injury. These findings suggest that Resveratrol has broad potential applications in the treatment and prevention of ALI.

Carbapenem-resistant Klebsiella pneumoniae (CRKP) poses a major public health threat worldwide due to the limited treatment options available. There is an urgent need to explore new treatment strategies. Although phages are considered to be an effective treatment, most characterized Klebsiella phages demonstrate narrow host ranges restricted to specific capsular types (KL types), thereby limiting their clinical applicability. In the present study, we isolated a novel broad host range phage, phiA85, with high lytic activity and biofilm inhibitory efficacy against host strains of 13 different KL types and 12 ST types. Among these host strains, six strains were identified as multidrug-resistant (MDR)-CRKP covering five different KL types. To further optimize phage therapy, this study indicated that phage phiA85 in combination with sublethal concentrations of antibiotics had synergistic effects against CRKP in vitro and in vivo. Killing curves and biofilm quantification assays revealed that phiA85-imipenem and phiA85-ciprofloxacin combinations exhibited phage-antibiotic synergy (PAS), demonstrating better bactericidal efficacy and reduction in biofilm formation compared to monotherapy. In a mouse pneumonia model, phage phiA85-imipenem combination treatment reduced mortality and alleviated pneumonia without other side effects. Our findings identify a broad host range phage capable of lysing different KL types and ST types of K. pneumoniae. Notably, phage phiA85-antibiotic combination offers a promising therapeutic option for the clinical treatment against MDR-CRKP. IMPORTANCE:The widespread prevalence of MDR bacteria has become a critical public health threat with limited therapeutic options. This study identifies a novel broad host range phage phiA85, capable of lysing strains across different KL types of CRKP. This characteristic addresses the limitation of narrow host range of capsular-specific phages, thereby significantly expanding their therapeutic potential against multidrug-resistant bacterial infections. Phage phiA85-antibiotic combinations achieve PAS, enhancing lytic activity and biofilm inhibition in vitro and alleviating mouse pneumonia. These findings highlight that phage phiA85-antibiotic combinations are a promising strategy to combat MDR-CRKP infections. Our work provides critical insights into optimizing phage therapy for clinical use against priority pathogens.

We have previously reported that high-alcohol-producing Klebsiella pneumoniae (HiAlc Kpn) in the gut can cause endo-alcoholic fatty liver disease. Here, we discover that 91.2% of Kpn isolates from pulmonary disease samples also produce excess ethanol, which may be associated with respiratory disease severity. To further explore the potential mechanism, a murine model is established with high-dose bacteria. Kpn stimulates granular neutrophils (G0), subsequently transforming them into phagocytic neutrophils (G1). HiAlc Kpn also causes dysfunction of pyrimidine metabolism, leading to neutrophil apoptosis. These changes inhibit phagocytosis of neutrophils and possibly suppress inflammasome-dependent innate immunity. In a persistent infective murine model, HiAlc Kpn induces lung fibrosis and production of reactive oxygen species (ROS), possibly affecting epithelial cell apoptosis and lung function. The results suggest that the subtype of neutrophil is a potential biomarker for the severity of lung injury caused by HiAlc Kpn.

Hypervirulent Klebsiella pneumoniae (hvKp) strains are increasingly recognized for their aggressive nature, which leads to severe clinical outcomes. The emergence of multidrug-resistant strains constitutes a substantial challenge for clinical management. Phage therapy offers a potential solution to the antibiotic resistance crisis. A multidrug-resistant hvKp strain, K2420 (K20 serotype), was used to isolate bacteriophages from hospital sewage. Phage morphology, biological properties, and genome characteristics were analyzed using transmission electron microscopy, plaque assays, and whole-genome sequencing. Therapeutic safety and efficacy were assessed in an acute pneumonia murine model induced by intratracheal injection of K2420. Assessment parameters included bacterial load, phage titer, body temperature, cytokine levels, histopathological findings, and other relevant indicators. Phage PK2420, a member of the Autographiviridae family and Przondovirus genus, was identified. It rapidly lyses K. pneumoniae (K20 serotype), inhibits biofilm formation, and exhibits a burst size of 37.4 plaque-forming units/cell. The phage is stable at temperatures ranging from 0°C to 40°C and pH values between 6 and 9. Its genome, 41,155 bp in length, contains 46 coding sequences. The phage has no genes associated with antibiotic resistance, virulence, or lysogeny. In vivo, PK2420 substantially reduced K. pneumoniae bacterial loads, improved survival rates, and alleviated pneumonia severity without observable side effects. Phage PK2420 exhibits lytic activity against K. pneumoniae both in vitro and in murine models, providing a promising and safe option for the treatment of hvKp infections.IMPORTANCEOur 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.

As the threat of multidrug-resistant Klebsiella pneumoniae strains rises, the potential of phages as promising alternatives to antibiotics is increasingly being demonstrated. In this study, we isolated and characterized phiK2044, a highly specific and efficient lytic phage targeting the model strain K. pneumoniae NTUH-K2044. Demonstrating wide host compatibility, potent lytic activity, and robust environmental adaptability, phiK2044 exhibited exceptional efficacy against hypervirulent subtypes, including hypervirulent K. pneumoniae (45.2%), sequence type 23 (87.5%), and capsular K1 (92.3%). In the mouse model, phiK2044 effectively cleared bacteria without significant side effects, highlighting its therapeutic potential. Mechanistically, we identified wcaJ, a gene encoding a glycosyltransferase essential for capsular synthesis, as the critical determinant of the binding of phiK2044 to the host. Beyond its clinical utility, phiK2044 represents a model for studying phage ecology, host-microbe interactions, and capsule-dependent tropism. Collectively, phiK2044 represents a valuable tool against ST23/K1 K. pneumoniae infections, such as liver abscesses and bacteremia.IMPORTANCEThe rise of multidrug-resistant Klebsiella pneumoniae demands innovative therapies. This study identifies phiK2044, a lytic phage with high specificity and efficacy against hypervirulent subtypes. It safely clears infections in mice and reveals wcaJ-dependent capsule synthesis as the key host interaction mechanism. Beyond its therapeutic promise, phiK2044 serves as a critical tool for studying phage-host dynamics and capsule-mediated tropism, bridging clinical solutions and fundamental research in combating antimicrobial resistance.

Klebsiella pneumoniae has become a major clinical and public health threat due to the increasing prevalence of healthcare-associated infections caused by multidrug-resistant strains. In this study, we demonstrated that the deletion of arcA of ArcAB two-component system diminished the susceptibility of K. pneumoniae to antibiotics, osmotic agents, disinfectants, and structural compounds, which was independent of arcB. RNA-seq analysis revealed a marked upregulation of SMR efflux pump genes kpnEF in the ΔarcA strain compared to the wild-type strain, while the ΔarcB strain exhibited no significant changes. Notably, the deletion of kpnEF in both ΔarcA and wild-type strains abolished their differential susceptibility to antibiotics, osmotic agents, disinfectants, and structural compounds. The EMSA experiments showed that ArcA-P regulated the kpnEF transcriptional expression by directly binding to its promoter region. These findings indicated that ArcA could directly modulate the expression of the KpnEF efflux pump independently of its sensor kinase ArcB. Through phosphorylation level detection and gene knockout experiments, we found that ArcA phosphorylation in the ΔarcB strain was primarily mediated by the AckA-Pta pathway. This study expanded the function of the ArcAB system and provided a critical theoretical foundation for elucidating the mechanisms underlying bacterial antibiotic resistance in K. pneumoniae.IMPORTANCEKlebsiella pneumoniae is an important opportunistic bacterial pathogen, which can acquire a series of antimicrobial resistance (AMR) genes. The emergence of carbapenem-resistant K. pneumoniae (CRKP) posed significant challenges to public health. Polymyxins are often regarded as the last line of defense against CRKP infections. In this study, the deletion of arcA, the regulator of the two-component system ArcAB, increased resistance of K. pneumoniae to antibiotics and decreased susceptibility to osmotic agents, disinfectants, and structural compounds, which was independent of ArcB. Further experiments have shown that ArcA regulated the expression of the small multidrug resistance (SMR) pump KpnEF through direct binding. This process required ArcA phosphorylation, which was independent of ArcB but dependent on the AckA-Pta pathway. This study deepened the regulatory network of ArcAB and provided a new target for the treatment of K. pneumoniae.

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: Capsaicin is commonly used as a flavoring and a riot control agent. However, long-term exposure or high doses can cause acute lung injury in military and police personnel. The mechanisms underlying capsaicin-induced pulmonary toxicity remain unclear. Therefore, this study investigated the molecular mechanisms involved in capsaicin-induced acute lung injury using C57BL/6N mice. Methods: Through both transcriptomic and proteomic analyses of mouse lung tissue, we identified the involvement of the TNF signaling pathway in capsaicin-mediated acute lung injury. Next, we explored the role of TNF signaling in the progression of acute lung injury to identify potential therapeutic targets. In a capsaicin-induced acute lung injury mouse model and A549 cells, we assessed the therapeutic potential of the TNF-α antibody Nerelimomab. Compared with the control group, TNF-α up-regulation was observed, which correlated with increased pathological changes and elevated IL-6 (p < 0.01) and IL-18 (p < 0.01) levels, both in vivo and in vitro. Results: Flow cytometry revealed that compared to the capsaicin group, Nerelimomab treatment reduced the number of capsaicin-induced apoptotic cells (p < 0.001) and was associated with an increased Bcl-2/Bax ratio (p < 0.01) and reduced cleaved caspase 3 expression (p < 0.001). Analysis of A549 cells treated with capsaicin and Nerelimomab corroborated these results. These findings confirm the involvement of the TNF signaling pathway in capsaicin-induced acute lung injury and the apoptosis of alveolar epithelial cells. Conclusions: In conclusion, capsaicin inhalation can cause acute lung injury, and targeting the TNF signaling pathway offers a promising therapeutic strategy. Nerelimomab demonstrates significant potential in alleviating acute lung injury by inhibiting inflammatory mediator release and diminishing apoptosis. Based on transcriptomic and proteomic analyses, this study highlights the crucial role of the TNF signaling pathway in capsaicin-induced acute lung injury and supports the therapeutic efficacy of Nerelimomab in reducing epithelial apoptosis.

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 log10 reduction of the pathogen after treatment with the resveratrol/POL combination. Moreover, adding adenosine disodium triphosphate (Na2ATP) 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.

Hydrogels with rapid wound-healing capabilities and antimicrobial effects are gaining significant interest in related fields. Nonetheless, developing a multifunctional hydrogel wound dressing with injectable self-assembling, self-healing, antimicrobial properties, and efficient skin wound-healing capabilities remained a formidable challenge. In this experiment, we drew inspiration from silkworm cocoons' natural formation and protective mechanisms, employing a novel physical cross-linking method to create an injectable and self-healing quaternary hydrogel successfully. The hydrogel is based on a matrix of silk fibroin/silk sericin (SF/SS), with 1,2-dimyristoyl-sn-glycero-3-phosphate sodium salt (DMPG) serving as a physical cross-linking agent to form the hydrogel network structure, and the incorporation of silver nanoparticles (AgNPs) further enhances its antimicrobial capabilities. Our biomimetic hydrogel, which replicated the chemical properties of silkworm cocoons, demonstrated excellent hydrophilicity with a water contact angle that ranged from 37 to 52°. Its tensile and compressive resistance was approximately four times greater than that of a pure SF hydrogel, and its swelling performance was about three times higher than that of a pure SF hydrogel. Furthermore, the hydrogel exhibited an impressive bacterial inhibition rate of over 98 % in bacterial growth and inhibition experiments, which provided a solid foundation for accelerating wound healing. Likewise, experiments with mice and histological analyses revealed that on day 7, the expression of TNF-α and IL-1β in the wound tissues treated with the SF/SS/AgNPs hydrogel was significantly reduced by >25 % compared to the blank control group. This reduction indicates that the hydrogel could decrease the production of inflammatory cytokines, potentially aiding in the acceleration of wound healing and mitigation of inflammation-related adverse reactions. By day 14, the wounds were healed mainly, with the wound area reduced by 17 % compared to that of the blank group. This demonstrates the significant potential of this cocoon-mimetic hydrogel in accelerating wound healing and providing wound protection.

Candida auris (C. auris) was first discovered in Japan in 2009 and has since spread worldwide. It exhibits strong transmission ability, high multidrug resistance, blood infectivity, and mortality rates. Traditional diagnostic techniques for C. auris have shortcomings, leading to difficulty in its timely diagnosis and identification. Therefore, timely and accurate diagnostic assays for clinical samples are crucial. We developed a novel, rapid recombinase-aided amplification (RAA) assay targeting the 18S rRNA, ITS1, 5.8S rRNA, ITS2, and 28S rRNA genes for C. auris identification. This assay can rapidly amplify DNA at 39 degrees C in 20 min. The analytical sensitivity and specificity were evaluated. From 241 clinical samples collected from pediatric inpatients, none were detected as C. auris-positive. We then prepared simulated clinical samples by adding 10-fold serial dilutions of C. auris into the samples to test the RAA assay's efficacy and compared it with that of real-time PCR. The assay demonstrated an analytical sensitivity of 10 copies/mu L and an analytical specificity of 100%. The lower detection limit of the RAA assay for simulated clinical samples was 10(1) CFU/mL, which was better than that of real-time PCR (10(2)-10(3) CFU/mL), demonstrating that the RAA assay may have a better detection efficacy for clinical samples. In summary, the RAA assay has high sensitivity, specificity, and detection efficacy. This assay is a potential new method for detecting C. auris, with simple reaction condition requirements, thus helping to manage C. auris epidemics.

Veillonella spp. are Gram-negative opportunistic pathogens present in the respiratory, digestive, and reproductive tracts of mammals. An abnormal increase in Veillonella relative abundance in the body is closely associated with periodontitis, inflammatory bowel disease, urinary tract infections, and many other diseases. We designed a pair of primers and a probe based on the 16S rRNA gene sequences of Veillonella and conducted real-time quantitative PCR (qPCR) and droplet digital PCR (ddPCR) to quantify the abundance of Veillonella in fecal samples. These two methods were tested for specificity and sensitivity using simulated clinical samples. The sensitivity of qPCR was 100 copies/μL, allowing for the accurate detection of a wide range of Veillonella concentrations from 103 to 108 CFU/mL. The sensitivity of ddPCR was 11.3 copies/μL, only allowing for the accurate detection of Veillonella concentrations from 101 to 104 CFU/mL because of the limited number of droplets generated by ddPCR. ddPCR is therefore more suitable for the detection of low-abundance Veillonella samples. To characterize the validity of the assay system, clinical samples from children with inflammatory bowel disease were collected and analyzed, and the results were verified using isolation methods. We conclude that molecular assays targeting the 16S rRNA gene provides an important tool for the rapid diagnosis of chronic and infectious diseases caused by Veillonella and also supports the isolation and identification of Veillonella for research purposes. • With suitable primer sets, the qPCR has a wider detection range than ddPCR. • ddPCR is suitable for the detection of low-abundance samples. • Methods successfully guided the isolation of Veillonella in clinical sample.

Klebsiella pneumoniae is a common, conditionally pathogenic bacterium that often has a multidrug-resistant phenotype, leading to failure of antibiotic therapies. It can therefore induce serious diseases, including community-acquired pneumonia and bloodstream infections. As an emerging alternative to antibiotics, phages are considered key to solving the problem of drug-resistant bacterial infections. Here, we report a novel phage, pK3–24, that mainly targets ST447 K. pneumoniae. Phage pK3–24 is a T7-like short-tailed phage with a fast adsorption capacity that forms translucent plaques with halos on bacterial lawns. The optimal multiplicity of infection (MOI) is 0.01, and the average burst size is 50 PFU/mL. Phage pK3–24 shows environmental stability, surviving at below 50 °C and at pH values of 6–10. It has a double-stranded DNA genome of 40,327 bp and carries no antibiotic-resistance, virulence, or lysogeny genes. Phylogenetic analysis assigned phage pK3–24 to the genus Przondovirus as a new species. Phage pK3–24 inhibited the production of biofilm. Moreover, treatment with pK3–24 at doses with an MOI > 1 effectively reduced the mortality of Galleria mellonella larvae infected with ST447 K. pneumoniae.

In this pioneering study, electrostatic spraying (ES) technology with high voltages is proposed to reduce the size of hydrogel microbeads further, aiming to enhance the adsorption rate of cationic methylene blue (MB) dye. The increased voltages, ranging from 0.0 to 13.0 kV, further decreased the size of electrostatically sprayed hydrogel microbeads crosslinked by hydrogen bonds between sodium alginate (SA) and carboxymethyl chitosan (CMCS) in hydrochloric acid. The size of SA/CMCS hydrogel microbeads was successfully reduced from 2000 +/- 121 mu m (SC-2000) to 400 +/- 15 mu m (SC-400). Notably, SC-400 exhibits the highest maximum adsorption capacity (qm) and rate constant (k2) at 840.3 mg/g and 0.0598 g/mg/min, respectively, at pH 9.0 and a temperature of 25 degrees C in the absence of ionic compounds, which is three times higher than that of SC-2000, due to their high specific surface area and pore volume. Through a series of adsorption studies and characterization analyses, SA/CMCS hydrogel microbeads displayed heterogeneous adsorption behaviors towards MB dye through electrostatic interactions between the deprotonated carboxylic groups and cationic MB molecules, where MB adsorption efficiency could be significantly influenced by pH and ionic strength. These findings suggest that ES technology is effective in synthesizing smaller SA/CMCS hydrogel microbeads with enhanced MB removal rates and stable adsorption capacities and their applications could be further explored for removing other organic dyes and toxic metals in subsequent research studies.

Cancer microbiota have recently been demonstrated in several cancer types. The microbiome enhances inflammation in the cancer microenvironment and affects the disease pathology by regulating tumourigenesis, cancer progression, and chemotherapy resistance. Hepatoblastoma (HB), the most common childhood malignant tumour, is a malignant embryonic tumour. However, the pathogenesis and molecular basis of HB remain poorly understood. In this study, to explore the existence and distribution of the microbiome in tumour tissues and adjacent non-tumour tissues of children with HB, we mainly performed 16S rDNA sequencing, and the results showed that the diversity and abundance of the microbiome in children with HB were significantly different between HB tumours and adjacent non-tumour tissues (p < 0.01). At the phylum level, the dominant microbiome in the tumour tissues were Proteobacteria, Bacteroidetes, and Firmicutes. At the genus level, Ruminococcus was more abundant in HB tumours than in the adjacent non-tumour tissues. Simultaneously, the abundances of Bacteroides, Parabacteroides, Lachnospiracea-NK4A136, and Alistipes in HB tumours were lower than those in the adjacent non-tumour tissues. In addition, Romboutsia strongly correlated with alpha-fetoprotein, an important indicator of HB. Sphingomonas was abundant in primary HB tumours, whereas Oscillibacter and Pandoraea were abundant in metastatic HB tumours. However, whether these bacteria are associated with HB needs further evaluation. Therefore, we identified the microbiome that correlated with the occurrence and development of HB. Ruminococcus and Romboutsia were identified as potential bacterial markers of HB tumours. To conclude, we found that HB also contains cancer microbiome, and it is necessary to shed light on the microbiome characteristics of HB in the future.

Klebsiella pneumoniae can enter a viable but nonculturable (VBNC) state to survive in unfavorable environments. Our research found that high-, medium-, and low-alcohol-producing K. pneumoniae strains are associated with nonalcoholic fatty liver disease. However, the presence of the three Kpn strains has not been reported in the VBNC state or during resuscitation. In this study, the effects of different strains, salt concentrations, oxygen concentrations, temperatures, and nutrients in K. pneumoniae VBNC state were evaluated. The results showed that high-alcohol-producing K. pneumoniae induced a slower VBNC state than medium-alcohol-producing K. pneumoniae, and low-alcohol-producing K. pneumoniae. A high-salt concentration and micro-oxygen environment accelerated the loss of culturability. Simultaneously, both real-time quantitative PCR and droplet digital PCR were developed to compare the quantitative comparison of three Kpn strain VBNC states by counting single-copy gene numbers. At 22 degrees C or 37 degrees C, the number of culturable cells decreased significantly from about 10(8) to 10(5)-10(6) CFU/mL. In addition, imipenem, ciprofloxacin, polymyxin, and phiW14 inhibited cell resuscitation but could not kill VBNC-state cells. These results revealed that the different environments evaluated play different roles in the VBNC induction process, and new effective strategies for eliminating VBNC-state cells need to be further studied. These findings provide a better understanding of VBNC-state occurrence, maintenance, detection, and absolute quantification, as well as metabolic studies of resuscitation resistance and ethanol production. IMPORTANCE Bacteria may enter VBNC state under different harsh environments. Pathogenic VBNC bacteria cells in clinical and environmental samples pose a potential threat to public health because cells cannot be found by routine culture. The alcohol-producing Kpn VBNC state was not reported, and the influencing factors were unknown. The formation and recovery of VBNC state is a complete bacterial escape process. We evaluated the influence of multiple induction conditions on the formation of VBNC state and recovery from antibiotic and bacteriophage inhibition, and established a sensitive molecular method to enumerate the VBNC cells single-copy gene. The method can improve the sensitivity of pathogen detection in clinical, food, and environmental contamination monitoring, and outbreak warning. The study of the formation and recovery of VBNC-state cells under different stress environments will also promote the microbiological research on the development, adaptation, and resuscitation in VBNC-state ecology.