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Objectives: Patients undergoing extracorporeal membrane oxygenation (ECMO) are susceptible to fungal infections, also from rare or emerging pathogens. We present a case of a 3-year-old male patient hospitalized for respiratory failure and subjected to open-chest ECMO support, with a fungal infection from a pathogen not identifiable by standard methods. Methods: Although T2Candida panel (T2 Biosystems) resulted negative, blood cultures resulted positive for fungi after 4 days, confirmed by Gram staining. The fungus underwent typing using Bruker matrix-assisted laser desorption ionization-time of flight mass spectrometry system and Autobio Autof ms1000, which could not precisely identify the microorganism. Ultimately, whole genome sequencing (WGS) was performed directly on blood culture. Results: WGS analysis revealed in 5 days the presence of a fungus belonging to the Ustilago genus, a group of fungi commonly found in the environment but rarely causing human diseases. Conclusions: To the best of our knowledge, we presented the first case of an Ustilago spp infection in a pediatric patient undergoing ECMO, not identified by standard techniques but only by WGS performed directly on a blood sample in 5 days. Despite the paucity of literature on Ustilago spp infections treatment, therapy adjustments led to the eradication of the pathogen, underscoring the importance of advanced molecular techniques for the correct and timely identification of these microorganisms.

Background: Infant meningitis, particularly caused by Escherichia coli, remains a life-threatening condition, especially in premature and low-weight infants. Infections of the central nervous system can be fatal, necessitating prompt diagnosis and appropriate treatment. Acute infections caused by various pathogens, including E. coli, often present with similar clinical symptoms. The rapid identification of pathogens and their antimicrobial resistance mechanisms is critical for timely and effective treatment. We report the case of an 8-month-old patient who presented with fever, diarrhea, and convulsive seizures and was subsequently diagnosed with meningitis. Despite initial empirical treatment with ceftriaxone, the patient’s condition worsened. Methods: At Bambino Gesù Children’s Hospital, molecular diagnostic tools, including the FilmArray Meningitis/Encephalitis and Blood Culture Identification panels, were employed. Results: Although the Meningitis panel did not detect any pathogens due to the lack of the specific bacterial target, the off-label use of the Blood Culture Identification panel identified a non-K1 Escherichia coli strain carrying the CTX-M resistance gene, an extended-spectrum beta-lactamase (ESBL). Despite the rapid diagnostic approach and adjustment of antibiotic therapy, the patient succumbed to the infection due to the strain’s high virulence and multidrug resistance. Whole-genome sequencing further characterized the strain, revealing that it belonged to the ST131 group, a highly resistant and virulent strain associated with sepsis. Conclusions: This case highlights the importance of integrating advanced molecular diagnostics, such as whole-genome sequencing, with traditional methods to improve pathogen detection, especially in cases of emerging resistant strains that are not covered by standard diagnostic panels. It also emphasizes the need for the continuous adaptation of diagnostic tools to include non-K1 E. coli strains for more comprehensive and timely meningitis diagnosis.

Cystic fibrosis (CF) is a life-threatening genetic disease characterised by chronic lung infections sustained by opportunistic pathogens such as Pseudomonas aeruginosa. During the chronic long-lasting lung infections, P. aeruginosa adapts to the host environment. Hypermutability, mainly due to defects in the DNA repair system, resulting in an increased spontaneous mutation rate, represents a way to boost the rapid adaptation frequently encountered in CF P. aeruginosa isolates. We selected 609 isolates from 51 patients with CF chronically colonised by P. aeruginosa to study, by full-length genome sequencing, the longitudinal evolution of the bacterium. We recovered at least one hypermutable (mutator) isolate in 57% of patients. By combining genomic information and phenotypic analyses, we followed the evolutionary pathways of the P. aeruginosa mutator strains, identifying their contribution to multi-drug resistance and the emergence of new sub-lineages. By implementing patient clinical data, we observed that mutators preferentially follow a specific evolutionary trajectory in patients with a negative clinical outcome and that maintenance antibiotic polytherapy, based on alternating molecules, apparently reduces the occurrence of hypermutability. Finally, we draw attention to the possibility that modulator-induced changes in the pulmonary environment may be associated with the onset of hypermutability.

(1) Background: The literature reports a low risk of serious bacterial infections (SBIs) in febrile infants presenting with bronchiolitis or respiratory syncytial virus infection, but current microbiological techniques have a higher accuracy. (2) Methods: We assessed the risk of SBIs in neonates and infants with bronchiolitis from 2021 to 2023. We also evaluated C-reactive protein, procalcitonin, and leukocyte values. (3) Results: We included 242 infants. Blood cultures (BCs) were performed in 66/242 patients, with a positivity rate of 9.1% (including one BC with Staphylococcus hominis, considered as a contaminant). The cerebrospinal fluid culture was performed in 6/242 patients, and the results were all negative. Infection markers did not discriminate infants with positive BCs from those with negative ones. (4) Conclusions: Blood cultures should be performed in neonates and young infants with bronchiolitis fever, as the sepsis risk is not negligible. Conversely, our proposed algorithm is to wait for the respiratory panel results before decision-making for a lumbar puncture. Further studies are needed to understand lumbar puncture requirements.

Bacterial infections caused by multidrug-resistant (MDR) Gram-negatives are of great concern worldwide, as they are frequently associated with high mortality and morbidity rates. To date, two cases of VIM-2 metallo-β-lactamase (MBL)-producing Pseudomonas putida bacteremia have been ever reported in France and Spain between 2004 and 2010. Here, we present the first case of VIM-1-like-producing P. putida isolated in blood culture collected from an oncohematological pediatric patient admitted to Bambino Gesù Children’s Hospital (IRCCS) in Rome, Italy.

ABSTRACT Bacterial pneumonia is a challenging coronavirus disease 2019 (COVID-19) complication for intensive care unit (ICU) clinicians. Upon its implementation, the FilmArray pneumonia plus (FA-PP) panel’s practicability for both the diagnosis and antimicrobial therapy management of bacterial pneumonia was assessed in ICU patients with COVID-19. Respiratory samples were collected from patients who were mechanically ventilated at the time bacterial etiology and antimicrobial resistance were determined using both standard-of-care (culture and antimicrobial susceptibility testing [AST]) and FA-PP panel testing methods. Changes to targeted and/or appropriate antimicrobial therapy were reviewed. We tested 212 samples from 150 patients suspected of bacterial pneumonia. Etiologically, 120 samples were positive by both methods, two samples were culture positive but FA-PP negative (i.e., negative for on-panel organisms), and 90 were negative by both methods. FA-PP detected no culture-growing organisms (mostly Staphylococcus aureus or Pseudomonas aeruginosa) in 19 of 120 samples or antimicrobial resistance genes in two culture-negative samples for S. aureus organisms. Fifty-nine (27.8%) of 212 samples were from empirically treated patients. Antibiotics were discontinued in 5 (33.3%) of 15 patients with FA-PP-negative samples and were escalated/deescalated in 39 (88.6%) of 44 patients with FA-PP-positive samples. Overall, antibiotics were initiated in 87 (72.5%) of 120 pneumonia episodes and were not administered in 80 (87.0%) of 92 nonpneumonia episodes. Antimicrobial-resistant organisms caused 78 (60.0%) of 120 episodes. Excluding 19 colistin-resistant Acinetobacter baumannii episodes, AST confirmed appropriate antibiotic receipt in 101 (84.2%) of 120 episodes for one or more FA-PP-detected organisms. Compared to standard-of-care testing, the FA-PP panel may be of great value in the management of COVID-19 patients at risk of developing bacterial pneumonia in the ICU. IMPORTANCE Since bacterial pneumonia is relatively frequent, suspicion of it in COVID-19 patients may prompt ICU clinicians to overuse (broad-spectrum) antibiotics, particularly when empirical antibiotics do not cover the suspected pathogen. We showed that a PCR-based, culture-independent laboratory assay allows not only accurate diagnosis but also streamlining of antimicrobial therapy for bacterial pneumonia episodes. We report on the actual implementation of rapid diagnostics and its real-life impact on patient treatment, which is a gain over previously published studies on the topic. A better understanding of the role of that or similar PCR assays in routine ICU practice may lead us to appreciate the effectiveness of their implementation during the COVID-19 pandemic.

In the present study, the in vitro activity of the sulbactam–durlobactam (SUL–DUR) combination was evaluated against 141 carbapenem-resistant A. baumannii (CRAb) clinical strains collected from six Italian laboratories. Over half (54.6%) of these isolates were resistant to colistin. The SUL–DUR combination was active against these CRAb isolates with MIC50 and MIC90 values of 0.5 mg/L and 4 mg/L, respectively. Only eleven isolates were resistant to SUL–DUR with MIC values ranging from 8 to 128 mg/L. The SUL–DUR resistant A. baumannii exhibited several antimicrobial resistance genes (ARGs) such as blaOXA-20, blaOXA-58, blaOXA-66, blaADC-25, aac(6′)-Ib3 and aac(6′)-Ib-cr and mutations in gyrA (S81L) and parC (V104I, D105E). However, in these isolates, mutations Q488K and Y528H were found in PBP3. Different determinants were also identified in these CRAb isolates, including adeABC, adeFGH, adeIJK, abeS, abaQ and abaR, which encode multidrug efflux pumps associated with resistance to multiple antibacterial agents. This is the first report on the antimicrobial activity of SUL–DUR against carbapenem-resistant A. baumannii isolates selected from multiple regions in Italy.

This study aimed to assess the comparability of in vitro susceptibility testing methods to ceftazidime-avibactam (CZA) and ceftolozane-tazobactam (C/T). Meropenem-resistant and/or carbapenemase-producing clinical isolates of Enterobacterales (Enterobacteriaceae) and Pseudomonas aeruginosa were tested by both bioMérieux ETEST and VITEK-2 AST-N397 card and compared with a Micronaut AST-system broth microdilution (BMD) method. CZA and C/T MICs were interpreted using EUCAST breakpoints. Of the 153 Enterobacteriaceae isolates, 55.6% and 0.0% (VITEK 2) and 56.9% and 0.0% (ETEST and BMD) were susceptible to CZA and C/T, respectively. Of 52 P. aeruginosa isolates, 50.0% and 40.4% (VITEK 2, ETEST, and BMD) were susceptible to CZA and C/T, respectively. The essential agreement (EA) was 96.1% (197/205; VITEK 2 versus BMD) and 95.6% (196/205; ETEST versus BMD) for CZA testing, whereas EA was 98.0% (201/205; VITEK 2 versus BMD) and 96.6% (198/205; ETEST versus BMD) for C/T testing. The categorical agreement (CA) was 98.0% (201/205; VITEK 2 versus BMD) and 100% (ETEST versus BMD) for CZA testing, whereas CA was 100% (VITEK 2 versus BMD) and 100% (ETEST versus BMD) for C/T testing. Categorical errors regarded four Enterobacteriaceae isolates. VITEK 2 and ETEST yielded equivalent CZA and C/T susceptibility testing results, compared to the BMD method, in such a clinical context.