Piperacillin-tazobactam is frequently used in children undergoing liver transplantation (LT), yet its pharmacokinetics (PK) in this population remains poorly characterized. We aimed to develop a population pharmacokinetic model for piperacillin in plasma and peritoneal fluid in children undergoing LT and to optimize dosing regimen for this population. We conducted a retrospective, single-center, observational study including all children receiving piperacillin-tazobactam as prophylaxis for LT and had at least one piperacillin concentration in both plasma and peritoneal fluid. PK analysis was conducted using nonlinear mixed effect modeling (Monolix) and was part of Optimome study. We included 31 patients with a median (range) age of 4 years (4 months-17 years). A total of 172 plasma and 146 peritoneal fluid samples were analyzed. Plasma concentrations were best fitted by a one-compartment model with first-order elimination. A virtual effect site compartment was added to adequately describe piperacillin concentrations in plasma and peritoneal fluid. Allometrically scaled body weight (pre-transplant and current values) and glomerular filtration rate (eGFR) were identified as significant covariates. Median (range) peritoneal/plasma AUC0-24h ratio was 72 (45-94) %. Simulations showed that optimal dosing regimens ranged from 300 mg/kg/day (q6h over 3 hours) to 400 mg/kg/day as a continuous infusion, depending on the PK target (i.e., fT>1 × MIC or fT>4 × MIC) and patient eGFR. In children undergoing LT, body weight and renal function explained some of the between subject variability for piperacillin PK. Higher dosing regimen combined with therapeutic drug monitoring is necessary to optimize exposure in plasma and at surgical site.

OBJECTIVES:To characterize the intrinsic carbapenemase of Sphingobacterium spp. MATERIALS AND METHODS:Sphingobacterium spp. clinical isolates (n = 63) were collected in several French laboratories from 2012 to 2022. Antibiotic susceptibility was assessed by disc diffusion. MICs of last resort antimicrobials (including carbapenems, ceftazidime-avibactam, cefepime-enmetazobactam and cefiderocol) were determined by broth microdilution. WGS was performed on all isolates using the Illumina method. Cloning of the Sphingobacterium spiritivorum β-lactamase encoding gene, blaSPI-1, allowed expression of a novel metallo-β-lactamase (MBL), SPI-1, in Escherichia coli. Enzymatic kinetic parameters were determined with β-lactams as substrates. RESULTS:Among the 64 Sphingobacterium isolates collected we identified six species, of which S. spiritivorum (n = 24) was the most prevalent followed by S. multivorum (n = 20), S. siyangense (n = 16), S. lactis (n = 2), S. thalpophilum (n = 1) and S. faecium (n = 1). Sphingobacterium isolates displayed a phenotype of multidrug resistance including resistance to last resort β-lactams and β-lactam-β-lactamase inhibitors (with MIC50 at 4, 4 and 2 mg/L for ceftazidime-avibactam, cefepime-enmetazobactam and cefiderocol, respectively). The most effective drugs were fluoroquinolones, tigecycline and eravacycline. All S. spiritivorum displayed a carbapenemase activity that correlated with the identification of SPI-1 encoding genes. SPI-like variants (n = 10) shared 93.7% amino-acid identity, but only 50.6% identity with the closest CGB-1 MBL intrinsic of Chryseobacterium gleum. Characterization of SPI-1 showed a strong hydrolytic activity towards most β-lactams including carbapenems. CONCLUSION:S. spiritivorum was the most prevalent species of Sphingobacterium identified in clinical samples. All S. spiritivorum-like species produced a chromosomally encoded SPI-like MBL that might be challenging for the treatment of infections caused by this bacterial species.

OBJECTIVES:Prolonged intravenous (IV) antibiotic therapy remains standard for Staphylococcus aureus bloodstream infections (SAB). However, early oral switch may offer a safe alternative. We evaluated the oral switch strategy in children with SAB. METHODS:All pediatric SAB cases managed at a tertiary hospital between June 2021 and December 2023 were identified by reviewing S. aureus-positive blood cultures through microbiological informatics systems. Characteristics of patients and infectious episodes were compared between the standard full-IV treatment and the oral switch strategy. RESULTS:Among 140 cases, 82 (58.6%) were managed with an oral switch strategy. Bone and joint infections accounted for most oral switch cases (40.2%), while central line-associated bloodstream infections predominated in the full-IV group (86.2%). All endocarditis (n = 2) and 4/10 suppurative thrombophlebitis cases underwent oral switch. Median hospital stay was significantly shorter in the oral switch group (13 days [IQR, 8-24.7] vs 48 days [IQR, 24-78.2]; P = 0.03). Thirty-day failure rates were comparable between oral switch (2/82, 2.4%) and full-IV (2/58, 3.4%) groups (adjusted risk difference 0.5%, 95% CI, -5.4 to 6.4, P = 0.86). CONCLUSION:Oral switch, used in over half of children with SAB, may offer a safe alternative to full IV therapy. Broader prospective studies are warranted to confirm these findings.

OBJECTIVES:The aim is to describe the clinical features, management and outcomes of postoperative mediastinitis in children following cardiac surgery and to identify risk factors for mortality. METHODS:We retrospectively reviewed all cases of mediastinitis in children following congenital heart surgery over a 10-year period (2013-2023) at Necker Hospital, a tertiary care center. RESULTS:Cumulative incidence of mediastinitis was 0.74% [95% confidence interval (CI): 0.56-0.96] with 57 cases of 7665 interventions. Median age at surgery was 12 days (6-146), with 58% of patients younger than 1 month. Thirty-four patients (60%) had delayed sternal closure. The most frequent germs were Staphylococcus spp. (45%), Gram-negative bacteria (36%) and fungi (9%). All patients had surgical debridement: 46 (81%) in the operating room and 11 (19%) in the intensive care unit (ICU). Median length of stay was 21 days (13-30) in the ICU and 35 days (28-48) until hospital discharge. The mortality rate was 27%, with 12 of 15 deaths occurring in the ICU. In univariate analysis, mortality risk factors were surgical revision in the ICU compared with the operating room [odds ratio (OR): 4.9; 95% CI: 1.3-19.9], delayed sternal closure superior to 3 days (OR: 5.0; 95% CI: 1.3-16.5) and fungal mediastinitis (OR: 14.9; 95% CI: 2.0-185.4). In the multivariate analysis, only fungal infection remained a significant risk factor for mortality (OR: 25.4; 95% CI: 2.7-608). CONCLUSION:Mediastinitis is a rare complication of neonatal cardiac surgery, with a low incidence (0.74%) in this tertiary referral center. However, mortality from this condition remains high, with fungal infections identified as the main mortality risk factor.

Once passed into the bloodstream, bacterial pathogens have a limited time to interact with permissive receptors at the surface of host cells. Neisseria meningitidis has developed an extremely effective strategy allowing it to find its receptors in a few seconds. Here, we report that N. meningitidis type IV pili exploit the physical properties of host cells' plasma membranes to promote the formation of early tubular membrane structures essential for initial bacterial adhesion. These tubular structures, which form before any signaling events in host cells, concentrate and trap multiple plasma membrane-associated proteins in the vicinity of bacteria, thereby facilitating the selection, interaction and activation of specific adhesion and signaling receptors by bacterial ligands present on type IV pili. Our results define an additional paradigm for the recruitment of specific receptors by pathogenic bacteria, which depends on the physical property of bacterial pili to induce the formation of tubular plasma membrane structures enriched in integral plasma membrane receptors.

Filamentous phages belong to Inoviruses, a family of non-lytic phages that are mutually beneficial to their bacterial hosts. These phages have a role in bacterial dissemination, biofilm formation or immune evasion. In Neisseria meningitidis, the strains harbouring the MDA filamentous phage are associated with invasive diseases through the MDAΦ key role in epithelial cell colonisation. Using MDAΦ and N. meningitidis as model organisms, we aimed to understand the relationship between filamentous phage infection, type IV pili antigenic variation and bacterial colonisation. The paradigm of filamentous phage infection has been defined for the phages Ff and CTX, where binding to the tips of bacterial type IV pili is a means of infecting their bacterial host. In contrast, we showed that MDAΦ binds N. meningitidis type IV pili along the length of the filament, rather than at the tip, with preferential binding to positively charged variants of PilE (the major fibre-forming pilin) demonstrating a role for antigenic variation in phage infection. Strikingly, bacteria expressing the more positively charged PilE were the most adhesive, meaning that MDAΦ primarily target the most adhesive bacteria. Finally, we showed that adhesion to human cells is sufficient to amplify the phage-positive meningococcal population. Taken together, this study reveals how the propagation strategy of a filamentous phage, aimed at selecting the best coloniser as a host, can promote the selection of hyperadhesive bacterial variants, linking phage infection to bacterial virulence.

Neisseria meningitidis is a human commensal bacterium that can opportunistically invade the bloodstream and cross the blood–brain barrier, where it can cause septicemia and meningitis. These diseases, if left untreated, can be lethal within hours. Hyperinvasive N. meningitidis strains often express a genomically encoded filamentous bacteriophage called MDAΦ, which promotes colonization of mucosal host surfaces to facilitate bacterial invasion. How this phage is organized and how it promotes biofilm formation and infection at the molecular level is unclear. Here, we present an electron cryomicroscopy structure of the MDA phage, showing that MDAΦ is a class I filamentous inovirus, with the major capsid protein (MCP) arranged within the phage as a highly curved and densely packed α-helix. Comparison with other filamentous bacteriophages offers clues about inoviral genome encapsidation mechanisms, providing a framework for understanding the evolutionary diversity of inoviruses. A disordered, N-terminal segment in the MCP presents hydrophobic patches on the surface of assembled phage particles, which, together with electron cryotomography data of phage bundles, furnishes a structural rationale for phage–phage interactions that were seen previously in an epithelium adhesion infection model of N. meningitidis . Taken together, our results shed light on the structure, organization, and higher-order assembly of a biomedically relevant phage encoded in the genome of a human pathogen. Molecular insights gleaned from this study increase our understanding of phage evolution, phage-mediated bacterial adhesion, and pathogenicity.

Filamentous phages are non-lytic phages mutually beneficial to their bacterial hosts. In Neisseria meningitidis, the filamentous phage MDA is associated with invasive diseases thanks to its key role in the formation of biofilm during epithelium colonisation. The infection model for filamentous phages has been defined for phages Ff and CTX. These phages bind to the tips of bacterial pili before being translocated into the periplasm of their hosts. The aim of this study is to investigate the relationship between filamentous phage infection and type IV pili, using the bacterium Neisseria meningitidis and the bacteriophage MDA as model organisms. We show that MDAΦ rather binds to type IV pili along their entire length with preferential binding to positively charged variants of the major fibre-forming pilin, demonstrating a role for antigenic variation in phage infection. Strikingly, bacteria expressing the more positively charged pilin are also the most adhesive, suggesting that MDAΦ primarily target the most adhesive bacteria. Finally, we show that adhesion to human cells is sufficient to amplify the phage-positive meningococcal population. Overall, this study reveals how a filamentous phage can target hyperadhesive bacterial variants and promote their selection, thereby establishing a link between phage infection and bacterial colonisation.

OBJECTIVE:To analyze the changes over the last 2 decades in clinical presentation, initial management, bacterial etiology, and rates of complications of acute mastoiditis (AM) in children. STUDY DESIGN:We conducted a retrospective study including all children diagnosed with AM between 2021 and 2024 in a French tertiary care center. Age at diagnosis, first-line surgical and medical treatment modalities, complications and bacteriological findings were collected. These data were compared with those of a previously published cohort of AM between 2001 and 2008 from the same facility. RESULTS:During the 3-year study period, 223 cases of AM were included (42 cases in 2021-2022; 104 in 2022 -2023; and 77 in 2023-2024). Among the 223 cases, 211 (95%) had a subperiosteal abscess (95%), 65 (29%) an extradural empyema, and 55 (25%) a lateral sinus thrombosis. The main pathogens identified were Streptococcus pyogenes (37%), Streptococcus pneumoniae (23%), and Fusobacterium necrophorum (19%). In cases of AM with intracranial complications, S. pyogenes and F. necrophorum were causative in over 70% compared with 46% in noncomplicated cases (P < .001). Comparing the 2021-2024 cohort with 2001-2008 cohort, we observed an increase in the number of cases (74 cases/year vs 27 cases/year), a rise in rate of intracranial complications (39% vs 4%; P < .001), an increase in performance of mastoidectomy (54% vs 33%; P < .001), and a shift in etiologic agents, with an increase in the proportion of S. pyogenes (P < .001) and F. necrophorum (P = .02) and concomitant decline in S. pneumoniae (P < .001). CONCLUSIONS:This large, single-center study highlights clear changes in the clinical manifestations and bacterial etiology of AM over the past decades. Ongoing surveillance of the bacterial etiology of AM is essential to optimize its initial management.

Aim: Population pharmacokinetics (PK) models may be effective to improve antibiotic exposure with individualized dosing. The aim of the study is to assess cefazolin exposure using a population PK model in critically ill children. Methods: We conducted a single center observational study including children under 18 years old who had cefazolin plasma monitoring before and after the model implementation. First concentration at steady state of each cefazolin course was analyzed. Optimal exposure was defined by concentrations values ranged from free concentration over 4 times the MIC for 100% of the dosing interval to total trough or plateau concentration under 100 mg/L. Results: Fifty-eight patients were included, of whom 39 and 19 children received conventional dosing or model-informed dosing, respectively. Median [range] age was 2.3 [0.1-17] years old and median weight was 14.2 [2.9-72] kg. There were more continuous infusions (CI) in the model group than in the conventional group (n=19/19 (100%) vs n=23/39 (59%)). Compared to conventional dosing, model-informed dosing provided more optimal exposure (n=17/39 (44%) vs n= 15/19 (79%), p=0.01) and less underexposure (n= 18/39 (46%) vs n= 2/19 (10%), p=0.008), without increasing overexposure (n= 4/39 (10%) vs n= 2/19 (11%), p=1). Moreover, the time to reach a 50% decrease of C Reactive Protein levels was significantly shorter in the model group than the conventional group (3 [0.5-13] vs 4 [1-34]; p=0.045. Conclusions: Use of individualized cefazolin model-informed dosing improves critically ill children’s exposure. Further studies are needed to assess the clinical benefit of cefazolin PK model application.

Neisseria meningitidisis a human commensal bacterium that can opportunistically invade the bloodstream and cross the blood-brain barrier, where it can cause septicaemia and meningitis. These diseases, if left untreated, can be lethal within hours. HyperinvasiveN. meningitidisstrains often express a genomically encoded filamentous bacteriophage called MDAΦ, which promotes colonisation of mucosal host surfaces to facilitate invasion. How this phage is organised and how it promotes biofilm formation and infection at the molecular level is unclear. Here, we present an electron cryomicroscopy structure of the MDA phage, showing that MDAΦ is a class I filamentous inovirus, with the major capsid protein arranged within the phage as a highly curved and densely packed α-helix. Comparison with other filamentous bacteriophages offers clues about inoviral genome encapsidation mechanisms, providing a framework for understanding the evolutionary diversity of inoviruses. A disordered, N-terminal segment in the major capsid protein presents hydrophobic patches on the surface of assembled phage particles, which, together with electron cryotomography data of phage bundles, furnishes a structural rationale for phage-phage interactions that were seen previously in an epithelium adhesion infection model ofN. meningitidis. Taken together, our results shed light on the structure, organisation, and higher order assembly of a biomedically relevant phage encoded in the genome of a human pathogen. Molecular insights gleaned from this study increase our understanding of phage evolution, phage-mediated bacterial adhesion and pathogenicity.

The efficient transformation of Neisseria meningitidis and Neisseria gonorrhoeae facilitates the rapid construction of bacterial mutants with insertion of antibiotic resistance cassettes. However, this strategy limits the construction of strains with multiple mutations. Recent advances in markerless strategies for Neisseria species have enabled the construction of mutants without antibiotic resistance markers. However, these innovative approaches have potential limitations related to the selection strategy or the possible occurrence of spontaneous mutations in the selection marker genes. In addition, complementation tools or labelling strategies for N. meningitidis are also lacking. In this study, we have introduced new tools for markerless mutation, genetic complementation and labelling in N. meningitidis , thus improving the research possibilities for understanding and tackling these pathogens. ### Competing Interest Statement The authors have declared no competing interest.

The efficient natural transformation of Neisseria meningitidis allows the rapid construction of bacterial mutants in which the genes of interest are interrupted or replaced by antibiotic-resistance cassettes. However, this proved to be a double-edged sword, i.e., although facilitating the genetic characterization of this important human pathogen, it has limited the development of strategies for constructing markerless mutants without antibiotic-resistance markers. In addition, efficient tools for complementation or labeling are also lacking in N. meningitidis. In this study, we significantly expand the meningococcal genetic toolbox by developing new and efficient tools for the construction of markerless mutants (using a dual counterselection strategy), genetic complementation (using integrative vectors), and cell labeling (using a self-labeling protein tag). This expanded toolbox paves the way for more in-depth genetic characterization of N. meningitidis and might also be useful in other Neisseria species. IMPORTANCE Neisseria meningitidis and Neisseria gonorrhoeae are two important human pathogens. Research focusing on these bacteria requires genetic engineering, which is facilitated by their natural ability to undergo transformation. However, the ease of mutant engineering has led the Neisseria community to neglect the development of more sophisticated tools for gene editing, particularly for N. meningitidis. In this study, we have significantly expanded the meningococcal genetic toolbox by developing novel and efficient tools for markerless mutant construction, genetic complementation, and cell tagging. This expanded toolbox paves the way for more in-depth genetic characterization of N. meningitidis and might also be useful in other Neisseria species.

Mycobacterium abscessus infection is challenging to treat. Extrapulmonary M. abscessus infections (EPMAB) are less common than pulmonary M. abscessus infections. To evaluate treatment regimens, we retrospectively analyzed consecutive microbiologically confirmed EP-MAB cases diagnosed in France during 2012-2020. We studied 45 patients with EP-MAB, including 14 bone and joint infections, 10 skin and soft tissue infections, and 8 lymph node infections. Most (62%) patients had no reported immunodeficiency. In 27 patients, EP-MAB followed healthcare-associated (44%) or environmental (16%) injuries. Of the 45 isolates, 25 were subspecies abscessus, 10 bolletii, and 9 massiliense; 1 was unidentified. Cure was achieved for 36 (80%) patients who received a median antimicrobial regimen of 6 months; 22 (55%) also underwent surgery. Four patients died, and 5 were unavailable for follow-up. EP-MAB predominantly affects immunocompetent patients after an injury; outcomes are favorable. We propose a >6-month regimen of antimicrobial therapy with consideration for surgery and regular patient reassessment.

OBJECTIVES:Children on extracorporeal membrane oxygenation (ECMO) are at high risk of infection that may worsen prognosis. Even though treatment with beta-lactam antibiotics is frequent, dosing is not adapted to altered pharmacokinetic and pharmacodynamic characteristics of children on ECMO. There is, therefore, a risk of inadequate drug levels when using standard dosing. In this study, we aimed to describe beta-lactam exposures of children on ECMO using current dosing and to identify factors associated with inadequate exposure. The optimal pharmacokinetic/pharmacodynamic target was considered as a plasma concentration four times above the minimum inhibitory concentration throughout the dosing interval target. DESIGN:Two-center retrospective cohort study. SETTING:Two PICUs in Paris, France. PATIENTS:Children (from birth to 18 yr) undergoing venovenous or venoarterial ECMO, from 2018 to 2020. INTERVENTIONS:None. MEASUREMENTS AND MAIN RESULTS:There were 57 patients who received 11 different beta-lactams, with 226 plasma concentrations analyzed. A total of 32 infections were documented. Overall, 133 of 226 concentrations (58.8%) were insufficient, primarily in samples from children younger than 28 days (p = 0.035), with low body weight (p = 0.013), or in instances of hypoalbuminemia (p = 0.011) and increased renal clearance (p = 0.032). Supratherapeutic concentrations were observed in 25 of 226 samples (11.1%), associated with being taken from patients with renal impairment (p < 0.01). CONCLUSIONS:In this retrospective cohort of pediatric ECMO cases, there is an associated risk of underexposure when prescribing conventional dosing of beta-lactams, which are likely associated with renal impairment and fluid overload. Prospective testing of therapeutic drug monitoring combined with pharmacokinetic/pharmacodynamic models should be tested as a risk-reduction strategy in this vulnerable population.

Immune reconstitution inflammatory syndrome (IRIS) has been reported in immunocompromised patients with disseminated Mycobacterium genavense. Management relies on high-dose corticosteroids.We describe two cases of late-onset corticosteroid-refractory IRIS related to disseminated infection in a HIV-positive patient and a renal transplant patient who had a favorable outcome with a monoclonal TNF-α blocker.

Objectives: Ceftolozane-tazobactam (C/T) proved its efficacy for the treatment of infections caused by non-carbapenemase producing Pseudomonas aeruginosa and Enterobacterales. Here, we aimed to provide susceptibility data on a large series of Enterobacterales since the revision of EUCAST categorization break-points in 2020.Methods: First, C/T susceptibility was determined on characterized Enterobacterales resistant to third generation cephalosporins (3GCs) (extended spectrum beta-lactamase [ESBL] production or different levels of AmpC overexpression) (n = 213) and carbapenem-resistant Enterobacterales (CRE) (n = 259), includ-ing 170 carbapenemase producers (CPE). Then, 1632 consecutive clinical Enterobacterales responsible for infection were prospectively collected in 23 French hospitals. C/T susceptibility was determined by E -test (R) (biomerieux) and broth microdilution (BMD) (SensititreTM, Thermo Scientific) to perform method comparison.Results: Within the collection isolates, 88% of 3GC resistant strains were susceptible to C/T, with impor-tant variation depending on the resistance mechanism: 93% vs. 13% susceptibility for CTX-M and SHV-ESBL producers, respectively. Only 20% of the CRE were susceptible to C/T. Among CPE, 80% of OXA-48-like producers were susceptible to C/T, whereas all metallo-,B-lactamase producers were resistant. The prospective study revealed that 95.6% of clinical isolates were susceptible to C/T. Method comparison performed on these 1632 clinical isolates demonstrated 99% of categorization agreement between MIC to C/T determined by E-test (R) in comparison with the BMD (reference) and only 74% of essential agreement. Conclusion: Overall, C/T showed good activity against wild-type Enterobacterales, AmpC producers, and ESBL-producing Escherichia coli but is less active against ESBL-producing Klebsiella pneumoniae, and CRE. E-test (R) led to an underestimation of the MICs in comparison to the BMD reference.(c) 2023 The Author(s). Published by Elsevier Ltd on behalf of International Society for Antimicrobial Chemotherapy.This is an open access article under the CC BY license ( http://creativecommons.org/licenses/by/4.0/ )

A 3-year-old male originating from Djibouti presented with a cervical mass evolving for 2 months. Tuberculous lymphadenopathy was suspected based on biopsy results, and he improved quickly on standard antituberculous quadritherapy. Subsequently some features of the mycobacterium that grew in culture were unusual. The isolate was eventually identified as Mycobacterium canettii , a peculiar species of the Mycobacterium tuberculosis complex.