The presence of drug resistance and extracellular matrix protection in biofilms makes it increasingly difficult to control bacterial infections using antibiotics. Therefore, there is an urgent need to develop new non-antibiotic approaches to eradicate drug-resistant bacterial infections. Here, a bimetal-phenolic framework (Que-Fe-CeMPF) is constructed by direct self-assembly of coordinated Fe3+ and Ce4+ ions with the polyphenol quercetin (Que). Que-Fe-CeMPF enhanced hydroxyl radical (·OH) generation, particularly in an acidic environment and presence of H2O2, compared with single metal-phenolic frameworks (Que-FeMPF and Que-CeMPF). · OH damaged bacterial cell walls, resulting in intracellular protein loss and bacterial cell death. Additionally, Que-Fe-CeMPF effectively dispersed biofilms by degrading matrix eDNA, allowing easier ·OH penetration, resulting in higher killing efficiency compared to Que-FeMPF and Que-CeMPF. Que-Fe-CeMPF stimulated macrophages to adopt an M2-like phenotype, suppressing excessive immune activation and promoting tissue repair at the infection site. As a combined effect of bacterial killing, biofilm degradation, and immune-modulation, the infectious pneumonia caused by Pseudomonas aeruginosa in mice is more effectively eradicated by Que-Fe-CeMPF than by free quercetin or the antibiotic ciprofloxacin. Moreover, Que-Fe-CeMPF is less prone to resistance development in pathogens compared to ciprofloxacin. Thus, Que-Fe-CeMPF is a promising non-antibiotic antimicrobial agent with multimodal activity for controlling drug-resistant bacterial infections.

BACKGROUND:Adolescents undergoing orthodontic fixed appliance treatment face an increased risk of dental caries and developing white spot lesions (WSLs) due to challenges in maintaining effective oral hygiene. Preventive measures, including adherence to clinical guidelines, are important to reduce these risks. OBJECTIVE:To analyse adherence to clinical practice guidelines (CPGs) amongst patients undergoing orthodontic treatment with buccal fixed appliances in the northeast of the Netherlands. METHODS:A survey was presented to 539 adolescents aged 12-17 undergoing buccal fixed appliance treatment. Participants were recruited from ten orthodontic practices. The survey assessed adherence to the six recommendations of the Dutch CPGs. An adherence sum score (range 0 to 6) was calculated. Descriptive statistics and linear regression analyses (1000 bootstrap samples) were performed to analyse the relationships between participants' characteristics and adherence sum scores. RESULTS:In total, 485 adolescents started the survey, of whom 393 (72.9%) could be included (57% female; 48.9% aged 13 or 14). The median adherence sum score was 5 (IQR 4, 5), and 22.6% (n = 89) had an adherence sum score of 6. Males had a lower adherence sum score than females (-0.442, 95% CI: -0.979, -0.234). Older participants had a lower adherence sum score than younger participants (-0.066, 95%CI: -0.136, 0.002) per year of age. Higher educated participants had a lower adherence sum score than participants with lower education (-0.534, 95%CI: -0.953, -0.096). LIMITATIONS:Self-reported data may introduce information bias as participants may give socially desirable answers. CONCLUSIONS:Adherence to the CPGs amongst adolescents with buccal fixed orthodontic appliances is suboptimal, particularly in boys and older adolescents. Tailored educational interventions may address these gaps.

Bacterial biofilm eradication and prevention of re-colonization are critical for effective treatment of biofilm-associated infections. Although significant progress has been made in nanovehicle-assisted antimicrobial platforms for biofilm eradication, strategies to address re-colonization remain underdeveloped. In this study, we constructed a versatile antimicrobial delivery platform based on multimodal interaction polyurea nanogels (MIPN). MIPN demonstrated excellent biocompatibility and could effectively load various antimicrobials with high capacity due to the multiple intermolecular interactions between the antimicrobials and nanocarriers, including hydrogen bonding, electrostatic, and hydrophobic interactions. By incorporating self-synthesized quorum sensing inhibitors (QSI) within MIPN, bacteria re-colonization was successfully prevented by blocking the quorum sensing pathway and disrupting surface-associated bacterial motilities. Furthermore, MIPN coloaded with QSI- and antibiotics showed a synergistic effect on biofilm eradication and re-colonization prevention, significantly enhancing the healing of biofilm-associated infections in chronic wounds.

Background/objectives: This retrospective longitudinal outcome study comparing orthodontic extraction modalities, including extraction of maxillary first or second molars, aimed to compare the three-dimensional tooth movement of maxillary canines (C), premolars (P1, P2), and molars (M1, M2) in Class II division 1 malocclusion treatment with fixed appliances. Methods: A sample of 98 patients (mean age 13.20 ± 1.46 years) was selected for the M1 group, and 64 patients (mean age 13.20 ± 1.36 years) were chosen for the M2 group. Tooth movement was analyzed three-dimensionally on pre-treatment (T0) and post-treatment (T1) digital dental casts. Regression analyses compared the tooth movements (in mm) between the M1 and M2 groups. Results: The mean treatment duration for the M1 group was 2.51 ± 0.55 year, while, for the M2 group, it was 1.53 ± 0.37 year. The data showed limited distal movements of the C, P1, and P2 of approximately 2 mm in the M1 group and 1 mm in the M2 group during orthodontic treatment, but the M1 group exhibited significantly more distal movements than the M2 group (mean difference 1.11 to 1.24 mm). Vertical movements of the C, P1, and P2 in both groups were also minor (0.16 to 1.26 mm). The differences between groups did not exceed 0.2 mm and were not significant. Both treatment modalities resulted in a significant degree of anchorage loss with a distinct mesialization (8.40 ± 1.66 mm) of M2 in the M1 group and limited distalization (0.83 ± 0.98 mm) of M1 in the M2 group. Conclusions: The findings highlight the importance of thorough case evaluation when choosing between extraction modalities in Class II treatment. If a large distal movement of canines and premolars is required, additional anchorage mechanics should be considered.

Bacterial infections pose a major challenge today due to the rise of drug resistance in pathogenic bacteria, creating an urgent need for the development of nonantibiotic therapies. Metal nanomaterials are increasingly recognized as promising antibacterial agents due to their unique physical and chemical properties, which offer strong antibacterial capabilities and broad-spectrum activity against drug-resistant bacteria. Unlike traditional antibacterial therapies, metal nanomaterials rarely induce drug resistance because of their diverse antibacterial mechanisms, which destroy bacteria by directly damaging bacterial cells or generating oxidative stress. Metal nanomaterials have proven effective in treating various drug-resistant bacterial infections. This perspective highlights recent advances in metal nanomaterials for antimicrobial applications. Firstly, bacterial infections and the current dilemma are introduced. Next, recent progress in the antibacterial activity of metal nanomaterials against drug-resistant bacteria is summarized, along with the challenges in their antimicrobial applications. Finally, future prospects and remaining challenges associated with the use of metal nanomaterials to treat drug-resistant bacteria are discussed.

BACKGROUND:Accurate repositioning of the maxilla is a crucial component of orthognathic surgery and can be facilitated by bone-borne (BB) or tooth-bone-borne (TBB) guides. AIMS:The objective of this systematic review and meta-analysis is to compare the accuracy of BB and TBB surgical guides in orthognathic surgery. Accuracy was defined as the difference between the planned and actual position of the maxilla, measured in linear and angular deviations across three axes. Relevant studies were identified up to February 2025. RESULTS:Twenty studies involving 332 patients (223 BB and 109 TBB) were reviewed. BB guides demonstrated slightly better accuracy at the U1 landmark, with deviations of 0.07 mm, 0.10 mm, and 0.24 mm in the X, Y, and Z axes, respectively. Conversely, TBB guides showed superior accuracy at the U6 landmark, with deviations of 0.19 mm and 0.31 mm in the X and Y axes. However, TBB guides were less accurate only in the Z-axis at U6 (deviation of 0.21 mm) when compared to BB guides. TBB guides also demonstrated higher angular accuracy, with differences of 0.35°, 0.39° and 0.02° for pitch, yaw, and roll respectively. However, these differences were generally small and clinically insignificant. CONCLUSION:Both BB and TBB guides are satisfactory choices, and the decision to use one over the other should be based on regulatory and logistical factors. The available evidence suggests that clinicians can be confident that both options yield comparable results, allowing flexibility in the decision-making process. CLINICAL SIGNIFICANCE:This review is the first to directly compare the effectiveness of BB and TBB guides. It provides valuable insights into how guide design influences surgical precision and supports clinical decision-making between the two types of guides.

Three-dimensional (3D) printing has slowly become an integral part of orthognathic surgery. However, there is a lack of studies evaluating accuracy of orthognathic surgical splints fabricated from subtractive milling versus additive 3D printing. The primary aim of this in vitro study was to compare the differences in trueness between milled and 3D-printed splints, while the secondary aim was to compare the differences in clinical fit of these splints. A sample of eight patients was selected, and STL files of the final orthognathic surgical splint were used to fabricate three splints for each of the eight cases. The first splint was fabricated by subtractive milling (SM), whereas the second and third splints were 3D printed with Digital Light Processing (DLP) and Laser Stereolithography (SLA), respectively. Paired superimposition of scans was performed using a reference model. The clinical fit of the splints to the printed models was also assessed. The mean root mean square (RMS) deviations for the SM, SLA, and DLP were 0.11 ± 0.02, 0.16 ± 0.02 and 0.14 ± 0.02 respectively. The post-hoc analysis showed that the SM splints had the highest accuracy (p<0.01). However, DLP splints showed the best clinical fit, followed by SM and SLA. In conclusion, splints fabricated by SM were more accurate than those fabricated by 3D printing, although this difference may not be clinically significant. The site, rather than the magnitude of the errors, may have a greater effect on the clinical usability of splints. In general, SM and DLP splints demonstrated a good clinical fit and were suitable for the fabrication of surgical splints.

OBJECTIVE:To evaluate the efficacy of a plaque-identifying toothpaste in assisting with plaque removal in adolescents undergoing orthodontic treatment with fixed appliances. METHODS:A randomised, double-blind, active-comparator trial was conducted at the Orthodontic Department, University Medical Center Department, University Medical Center Groningen, the Netherlands, from October 2021 to January 2022. Seventy seven healthy adolescents aged 12-16 years, undergoing buccal fixed appliance treatment for at least 3 months, were enrolled. Participants were stratified based on manual or electric brushing habits and randomised to receive either plaque-identifying toothpaste or a non-colouring control toothpaste. The primary outcome measured reduction in dental plaque using Quantitative Light-Induced Fluorescence ∆R30 (QLF) images at baseline (T0) and after 4-7 weeks (T1). Secondary outcomes included self-reported participant experiences. RESULTS:No statistically significant difference in plaque reduction was found between the two groups (F = 0.211, p = 0.647). Subgroup analyses showed that participant-related factors did not significantly influence plaque reduction. Participants using plaque-identifying toothpaste reported a slightly stronger perception of plaque removal (p = 0.018) but encountered slightly more difficulty with the toothpaste colour (p = 0.028). Compliance was high, and no adverse effects were reported. CONCLUSION:Plaque-identifying toothpaste did not lead to significant plaque reduction among adolescents with fixed orthodontic appliances.

A very promising strategy to avoid bacterial drug resistance is to replace antibiotics with artificial nanozymes, but this has not yet been translated to the clinic. Here, we construct a single-atom nanozyme using graphitic carbon nitride nanosheets modified by copper (Cu-g-C3N4). This Cu-g-C3N4 nanosheet possesses both glucose oxidase-like and peroxidase-like activities responsible for reactive-oxygen-species generation by a cascade reaction to eradicate Gram-positive and Gram-negative multidrug-resistant bacteria. Cu-g-C3N4 is introduced into polycaprolactone (PCL) by electrospinning to obtain (Cu-g-C3N4/PCL) nanofibers, which can be used as a dressing for bacterially infected wounds. It is demonstrated that Cu-g-C3N4/PCL nanofiber dressings can eradicate bacterial infections and accelerate wound healing in a mouse model with a skin wound.

To assess the impact of malocclusion on oral health-related quality of life (OHRQoL) and to compare the impact of malocclusion in children with and without hypodontia. Children aged 10–16 years with ≥ 5 missing teeth and without hypodontia completed the Malocclusion Impact Questionnaire (MIQ) to assess the impact of malocclusion on OHRQoL. The Child Perception Questionnaire 11–14 years short form (CPQ11-14-ISF16) was used to verify the validity of the MIQ. Demographic and orthodontic data were collected. Internal consistency and validity of the MIQ were analyzed. MIQ scores were compared using an independent t‑test. Regression analysis was performed to identify predictors of the MIQ score. A total of 92 participants completed the MIQ, and 52 participants the CPQ11-14-ISF16. The MIQ showed excellent internal consistency (Cronbach’s α 0.89) and good criterion validity with CPQ11-14-ISF16 (r = 0.58). No significant difference in the impact of malocclusion on OHRQoL between the groups (p = 0.15) was found. Age (p < 0.05), sex (p < 0.001), and general appearance (p < 0.001) significantly predicted OHRQoL scores in the regression analysis. Multilevel analysis showed that the group and age effects were nonsignificant and that sex and general appearance were predictive for the MIQ score. This study suggests that the MIQ is a useful tool to assess the impact of malocclusion on OHRQoL in the orthodontic field. Differences in the impact of malocclusion on OHRQoL between children with and without hypodontia of ≥ 5 teeth were limited. It may be beneficial delaying treatment until the patient expresses a subjective treatment need which may reduce overtreatment of children with hypodontia.

Owing to their extremely small size, carbon-quantum-dots (CQDs) can cross biological barriers, which makes them attractive for many biomedical and other applications. CQDs can retain key-chemical features and associated functionalities of the molecular sources they are derived from, provided a suitable synthesis method is used at relative mild carbonization temperatures. Here we demonstrate that CQDs hydrothermally-derived from chitosan or 2-hydroxypropyltrimethyl ammonium-chloride (HAC)-chitosan under pressurized conditions at 180 degrees C have a comparable elemental and molecular composition, as determined using X-ray photoelectron spectroscopy and Fourier-transform-infrared spectroscopy. In addition, both types of CQDs generated reactiveoxygen-species as an added functionality alien to their molecular carbon sources. As a result, CQDs exhibited stronger antibacterial properties against a Gram-positive Staphylococcus aureus and a Gram-negative Escherichia coli strain, while both molecular HAC-chitosan as well as CQDs derived from it had stronger antibacterial properties than molecular chitosan and chitosan CQDs due to the possession of quaternary ammonium groups in HAC-chitosan. Therewith, carbonization of chitosan and HAC-chitosan yields enhanced properties that can be beneficial in a high variety of different applications, including promotion of healing and bacterial infection control, preservation of food and beverages, pesticide control in agriculture and horticulture, water treatment and in many cosmetics and personal care products.

The era of relying on antibiotics for curing bacterial infections is rapidly approaching an end, necessitating development of non-antibiotic-based infection-control strategies. Dispersal of infectious biofilms is a potential strategy but yields dispersed bacteria in blood that may cause sepsis. We report a bromide-loaded, core-shell ZnO2-nanoparticle/Ce-based metal-organic framework (ZnO2@CeMOF/Br) of which the ZnO2 core degrades at pH ≤ 6.5, leaving the MOF's Ce node intact. ZnO2-core degradation initially generates a nonradical, relatively stable, low-oxidative hydrogen peroxide that can cleave matrix DNA causing dispersal of Staphylococcus aureus biofilms and reacts with bromide ions to form transient hypobromous acid. Hypobromous acid modulates macrophage polarization toward an M1-like phenotype to clear dispersed bacteria from blood. Subsequently the Ce3+/Ce4+ redox couple forming the Ce node acts as an electron shuttle upon oxidation/reduction to faciltate two catalytic reactions, maintaining hydrolysis of phosphodiester bonds and associated cleavage of matrix DNA as well as modulation of macrophage polarization. Neither growth of tissue cells or macrophages nor hemolysis are negatively affected by exposure to ZnO2@CeMOF/Br nanocatalysts at a ZnO2 nanoparticle over CeMOFs weight ratio ≤ 1.2, up until CeMOF concentrations less than at least 180 μg/mL. Under biosafe, low-turnover catalytic conditions, irrigation of infected wounds in diabetic mice with ZnO2@CeMOF/Br nanocatalysts (90 μg/mL) results in 100% survival, fast recovery of healthy body temperature and weight, lower numbers of CFUs in blood and wound and organ tissues, and macrophage polarization toward an M1-like phenotype, demonstrating potential of ZnO2@CeMOF/Br nanocatalysts for non-antibiotic-based infection control.

Abstract Background External apical root resorption (EARR) is a frequently observed adverse event in patients undergoing fixed appliance therapy. Assessing the patients’ risk during treatment is important, as certain factors are assumed to be associated with an increased likelihood of occurrence. However, their predictive value remains limited, making evidence-based clinical decision-making challenging for orthodontists. To address this issue, the Dutch Association of Orthodontists (NvVO) developed a clinical practice guideline (CPG) for EARR in accordance with the AGREE II instrument (Appraisal of Guidelines for Research and Evaluation II) in 2018. The aim of this study is to get insight into the actual utilization and the practical implementation of the guideline among orthodontists. The hypothesis to be tested was that after its introduction, clinical practice for EARR has changed towards the recommendations in the CPG. Objective To investigate the use of the 2018 clinical practice guidelines for EARR among orthodontists 3 years after its introduction. Methods A questionnaire using a 7-point Likert scale was developed concerning four domains of EARR described in the guideline. The questionnaire was piloted, finalised, and then distributed digitally among Dutch orthodontists. REDCap was used for data collection, starting with an invitation email in June 2021, followed by two reminders. Effect was tested by the Mann–Whitney U test, and the influence of demographic variables was analysed. Results Questionnaires were sent out to all 275 and completed by 133 (response rate 48%); N = 59 females and N = 73 males were included; 81% had their training in the Netherlands, 89% had ≥ 6 years of work experience, and 89% worked in private orthodontic practice. One hundred thirty orthodontists (98.5%) reported changes in clinical practice. The biggest positive change in clinical behaviour regarding EARR occurred if EARR was diagnosed during treatment. Sex, clinical experience, country of specialist training, and working environment of the respondents did not affect clinical practices regarding EARR. Conclusions This questionnaire demonstrated that, 3 years after introduction of the guideline, orthodontists improved their self-reported clinical practices to a more standardised management of root resorption. None of the demographic predictors had a significant effect on the results.

Background: Bone-anchored maxillary protraction (BAMP) aims to correct midfacial deficiencies, with proven positive skeletal changes without potential unwanted side effects. However, the influence of BAMP treatment on facial soft tissues, particularly in subjects with complete unilateral cleft lip, alveolus, and palate (CUCLAP), remains unclear. Methods: This single-center longitudinal cohort study examined the effects of 1.5 years of BAMP treatment on facial soft tissues in growing subjects with complete unilateral cleft lip, alveolus, and palate. The sample consisted of 25 patients, age range 9.7 to 12.6 years. Three-dimensional surface models derived from CBCT scans were superimposed on stable structures of the anterior cranial base and on the occipital area posterior of the foramen magnum to assess three-dimensional changes due to growth and BAMP therapy. Results: The results revealed a moderate positive correlation (Pearson’s correlation coefficient from 0.203 to 0.560) between changes in hard tissue and soft tissue; some correlations were found to be weak (<0.300). Linear changes in soft tissue following BAMP were in the same direction as skeletal changes, showing downward, forward, and outward displacement. The only exception was in the vertical dimension. The lower facial third showed a slight but significant reduction, mainly in lip length (−1.2 mm), whereas the middle facial third showed a small increase (1.1 mm). Conclusions: It was concluded that during BAMP, soft tissue changes occur in the same direction as skeletal changes, although with a larger variability and less pronounced effects.

As a potential alternative to antibiotics, hyperbranched poly(ionic liquid)s (HPILs) have demonstrated significant potential in combating bacterial biofilms. However, their high cation density poses a high risk of toxicity, greatly limiting their in vivo applications. In this study, we constructed a biocompatible HPIL (HPIL-Glu) from a hyperbranched polyurea core with modified terminals featuring charge-convertible ionic liquids. These ionic liquid moieties consist of an ammonium-based cation and a gluconate (Glu) organic counter. HPIL-Glu could form a homogeneous nanoassembly in water and exhibited a pH-responsive charge conversion property. Under neutral conditions, Glu shielded the positively charged surface, minimizing the toxicity. In a mildly acidic environment, Glu protonation exposes cationic moieties to biofilm eradication. Comprehensive antimicrobial assessments demonstrate that HPIL-Glu effectively kills bacteria and promotes the healing of bacteria-infected chronic wounds. Furthermore, prolonged exposure to HPIL-Glu does not induce antimicrobial resistance.

Hybrid-nanozymes are promising in various applications, but comprehensive comparison of hybrid-nanozymes composed of single-atoms or nanoparticles on the same support has never been made. Here, manganese-oxide nanosheets were loaded with Pt-single-atoms or differently-sized nanoparticles and their oxidase- and-peroxidase activities compared. High-resolution Transmission-Electron-Microscopy and corresponding Fast Fourier Transform imaging showed that Pt-nanoparticles (1.5 nm diameter) had no clear (1 1 1) crystal-planes, while larger nanoparticles had clear (1 1 1) crystal-planes. X-ray Photo-electron Spectroscopy demonstrated that unloaded nanosheets were composed of MnO2 with a high number of oxygen vacancies (Vo/Mn 0.4). Loading with 7.0 nm Pt-nanoparticles induced a change to Mn2O3, while loading with 1.5 nm nanoparticles increased the number of vacancies (Vo/Mn 1.2). Nanosheets loaded with 3.0 nm Pt-nanoparticles possessed similarly high catalytic activities as Pt-single-atoms. However, loading with 1.5 nm or 7.0 nm Pt-nanoparticles yielded lower catalytic activities. A model is proposed explaining the low catalytic activity of under- and over-sized Pt-nanoparticles as compared with intermediately-sized (3.0 nm) Pt-nanoparticles and single-atoms. Herewith, catalytic activities of hybrid-nanozymes composed of single-atoms and intermediately-sized nanoparticles are put a par, as confirmed here with respect to bacterial biofilm eradication. This conclusion facilitates a balanced choice between using Pt-single-atoms or nanoparticles in further development and application of hybrid-nanozymes.

The overuse of antibiotics has led to the rapid development of multi-drug resistant bacteria, making antibiotics increasingly ineffective against bacterial infections. Consequently, there is an urgent need to develop alternative strategies to combat multi-drug-resistant bacterial infections. In this study, gold nanoparticles modified with ellagic acid (EA-AuNPs) were prepared using a simple and mild one-pot hydrothermal process. EA-AuNPs demonstrated high bactericidal efficacy and broad-spectrum antimicrobial activities against clinical isolates of the antibiotic-resistant ESKAPE pathogens. Furthermore, EA-AuNPs effectively disperse biofilms of multi-drug-resistant bacteria. Additionally, EA-AuNPs mitigated inflammatory responses at the bacterial infection sites. The combined bactericidal and anti-inflammatory treatment with EA-AuNPs resulted in faster curing of peritonitis caused by Staphylococcus aureus in mice compared to treatment with free EA or gentamicin. Moreover, transcriptome analysis revealed that EA-AuNPs exhibited a multi-targeting mechanism, making resistance development in pathogens more challenging than traditional antibiotics that recognize specific cellular targets. Overall, EA-AuNPs emerged as a promising antimicrobial agent against multi-drug-resistant bacterial infections.

Bacteria can be dead, alive, or exhibit slowed or suspended life forms, making bacterial death difficult to establish. Here, agar-plating, microscopic-counting, SYTO9/propidium-iodide staining, MTT-conversion, and bioluminescence-imaging were used to determine bacterial death upon exposure to different conditions. Rank correlations between pairs of assay outcomes were low, indicating different assays measure different aspects of bacterial death. Principal-component analysis yielded two principal components, named “reproductive-ability” (PC1) and “metabolic-activity” (PC2). Plotting of these principal components in two-dimensional space revealed a dead region, with borders defined by the PC1 and PC2 values. Sensu stricto implies an unpractical reality that all assays determining PC1 and PC2 must be carried out in order to establish bacterial death. Considering this unpracticality, it is suggested that at least one assay determining reproductive activity (PC1) and one assay determining metabolic activity (PC2) should be used to establish bacterial death. Minimally, researchers should specifically describe which dimension of bacterial death is assessed, when addressing bacterial death.

Cascade-reaction containers generating reactive oxygen species (ROS) as an alternative for antibiotic-based strategies for bacterial infection control, require endogenous oxygen-sources and ROS-generation close to or preferably inside target bacteria. Here, this is achieved by cetyltrimethylammonium-chloride (CTAC) assisted in situ metabolic labeling and incorporation of mesoporous SiO2-nanoparticles, dual-loaded with glucose-oxidase and Fe3O4-nanoparticles as cascade-reaction containers, inside bacterial cell walls. First, azide-functionalized D-alanine (D-Ala-N3) was inserted in cell wall peptidoglycan layers of growing Gram-positive pathogens. In Gram-negatives, this could only be achieved after outer lipid-membrane permeabilization, using a low concentration of CTAC. Low concentrations of CTAC had no adverse effect on in vitro blood clotting or hemolysis nor on the health of mice when blood-injected. Next, dibenzocyclooctyne-polyethylene-glycol modified, SiO2-nanoparticles were in situ click-reacted with D-Ala-N3 in bacterial cell wall peptidoglycan layers. Herewith, a two-step cascade-reaction is facilitated inside bacteria, in which glucose-oxidase generates H2O2 at endogenously-available glucose concentrations, while subsequently Fe3O4-nanoparticles catalyze generation of •OH from the H2O2 generated. Generation of •OH inside bacterial cell walls by dual-loaded mesoporous SiO2-nanoparticles yielded more effective in vitro killing of both planktonic Gram-positive and Gram-negative bacteria suspended in 10% plasma than SiO2-nanoparticles solely loaded with glucose-oxidase. Gram-positive or Gram-negative bacterially induced sepsis in mice could be effectively treated by in situ pre-treatment with tail-vein injected CTAC and D-Ala-N3, followed by injection of dual-loaded cascade-reaction containers without using antibiotics. This makes in situ metabolic incorporation of cascade-reaction containers as described attractive for further investigation with respect to the control of other types of infections comprising planktonic bacteria.Statement of SignificanceIn situ metabolic-incorporation of cascade-reaction-containers loaded with glucose-oxidase and Fe3O4 nanoparticles into bacterial cell-wall peptidoglycan is described, yielding ROS-generation from endogenous glucose, non-antibiotically killing bacteria before ROS inactivates. Hitherto, only Gram-positives could be metabolically-labeled, because Gram-negatives possess two lipid-membranes. The outer membrane impedes direct access to the peptidoglycan. This problem was solved by outer-membrane permeabilization using a quaternary-ammonium compound. Several studies on metabolic-labeling perform crucial labeling steps during bacterial-culturing that in real-life should be part of a treatment. In situ metabolic-incorporation as described, can be applied in well-plates during in vitro experiments or in the body as during in vivo animal experiments. Surprisingly, metabolic-incorporation proceeded unhampered in blood and a murine, bacterially-induced sepsis could be well treated.