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Xavier Pomares1, Concepción Montón1, Mateu Espasa2, Jordi Casabon1, Eduard Monsó1,3, Miguel Gallego1,31Pneumology Service, 2Laboratory Service, Corporació Parc Taulí, Universitat Autònoma de Barcelona, Sabadell, Spain; 3Ciber de Enfermedades Respiratorias – CibeRes, Bunyola, SpainBackground: The aim of this study was to determine whether long-term intermittent azithromycin therapy reduces the frequency of exacerbation in severe chronic obstructive pulmonary disease (COPD).Methods: We retrospectively investigated the clinical benefits of long-term azithromycin (500 mg orally three times per week) over 12 months in patients with severe COPD and a minimum of four acute exacerbations (AECOPD) per year or chronic bronchial colonization by Pseudomonas aeruginosa, comparing the number of AECOPD, hospitalizations due to respiratory disease, days of hospital stay, and bacterial infections during azithromycin treatment and in the year prior to this therapy.Results: Twenty patients who completed the 12-month treatment period were analyzed. No clinically significant adverse events were observed during azithromycin treatment. Compared with baseline data, azithromycin therapy significantly reduced the number of AECOPD (2.8 ± 2.5 versus 6.8 ± 2.8, P < 0.001), hospitalizations (1.4 ± 1.5 versus 3.6 ± 1.4, P < 0.001), and cumulative annual days of hospital stay (25 ± 32.2 versus 43.7 ± 21.4, P = 0.01). The improvement was particularly significant in patients with exacerbations caused by common potentially pathogenic microorganisms, who had 70% fewer AECOPD and hospitalizations. Patients colonized by P. aeruginosa had reductions of 43% in AECOPD and 47% in hospitalizations.Conclusion: Long-term azithromycin is well tolerated and associated with significant reductions in AECOPD, hospitalizations, and length of hospital stay in patients with severe COPD.Keywords: azithromycin, chronic obstructive pulmonary disease, exacerbation, macrolides

Genetic redundancy in bacteria plays a crucial role in enhancing adaptability and accelerating evolution in response to selective pressures, particularly those associated with rapid environmental changes. Aminopenicillins like ampicillin are important therapeutic options for Enterococcus infections in both humans and animals, with resistance mostly associated with pbp5 gene mutations or overexpression. While the occurrence of redundant pbp5 genes has been occasionally reported, the advantages for the host bacteria have not been explored in detail. During a whole-genome sequencing project of Enterococcus faecium from bacteremic patients, we identified an ST592 strain (Efm57) with redundant pbp5 genes. This presented an opportunity to investigate the prevalence and implications of multiple pbp5 acquisitions in diverse Enterococcus species across various sources, geographical regions, and timeframes. The analysis of 618 complete Enterococcus genomes from public databases revealed that 3.2 % harbored redundant pbp5 genes, located on chromosomes or plasmids across different species from diverse epidemiological backgrounds. The proteins encoded by these genes showed homologies ranging from 51.1 % to 97.5 % compared to native copies. Phylogenetic analysis grouped redundant PBP5 amino acid sequences into three distinct clades, with insertion sequences (mostly IS6-like) facilitating their recent spread to diverse plasmids with varying genetic backbones. The presence of multiple antibiotic resistance genes on pbp5-plasmids, including those conferring resistance to linezolid, underscores their involvement in co-selection and recombination events with other clinically-relevant antibiotics. Conjugation experiments confirmed the transferability of a specific 24 kb pbp5-plasmid from the Efm57 strain. This plasmid was associated with higher minimum inhibitory concentrations of ampicillin and conferred bacteria growth advantages at 22 °C. In conclusion, the widespread distribution of redundant pbp5 genes among Enterococcus spp. highlights the complex interplay between genetic mobility, environmental factors, and multidrug resistance in overlapping ecosystems emphasizing the importance of understanding these dynamics to mitigate antibiotic resistance spread within the One Health framework., (Copyright © 2024. Published by Elsevier B.V.)

Objectives: This study aims to identify infection etiology in febrile neutropenia (FN) is vital. This study explores different microbiological approaches and their impact on diagnosing infections in patients with hematologic malignancies and FN., Methods: This is a retrospective analysis conducted at the Hospital Clinic of Barcelona details microbiological testing strategies used to diagnose infections at FN onset between January 2020 and July 2022., Results: A total of 4520 microbiological tests were ordered in 462 FN episodes, achieving a 10% test positivity rate, with 200 (43.3%) episodes showing microbiological documentation of infection. Blood cultures (40.4%), non-culture blood tests (21.2%), and respiratory tract samples (16.2%) were the most requested. Blood cultures exhibited the highest (16.9%) test positivity rates, whereas non-culture blood tests showed the lowest (3.3%). Bacterial infections were present in 149 of 462 (32.3%) FN episodes. Viral infections (66 of 462, 14.3%)-notably, respiratory viruses-were also frequent. Mortality rate at 60 days was 9.1%; documented infections were associated with a higher risk (15%)., Conclusions: In the current landscape of antimicrobial diagnostics, our findings revealed the highest reported rate of microbiologically documented infections at FN onset. Bacterial infections are common; however, our data reiterate the significance of viral infections in causing fever. Optimizing FN management during respiratory viral infections remains a challenge for antimicrobial de-escalation. The low positivity rates observed in certain diagnostic tests emphasize the need for cost-effective diagnostic stewardship., Competing Interests: Declarations of competing interest CG-V has received honoraria for talks on behalf of Gilead Science, MSD, Pfizer, Jannsen, Novartis, Basilea, GSK, Shionogi, AbbVie, and Advanz Pharma and grant support from Gilead Science, Pfizer, GSK, MSD, and PharmaMar. PP-A has received honoraria for talks on behalf of Merck Sharp and Dohme, Lilly, ViiV Healthcare, and Gilead Science., (Copyright © 2024. Published by Elsevier Ltd.)

Objectives: We aimed to report the emergence of azole-resistant invasive aspergillosis in hematologic patients admitted to a tertiary hospital in Spain during the last 4 months., Methods: Prospective, descriptive study was performed to describe and follow all consecutive proven and probable invasive aspergillosis resistant to azoles from hematological cohort during the last 4 months. All patients had fungal cultures and antifungal susceptibility or real-time PCR detection for Aspergillus species and real-time PCR detection for azole-resistant mutation., Results: Four cases of invasive aspergillosis were diagnosed in 4 months. Three of them had azole-resistant aspergillosis. Microbiological diagnosis was achieved in three cases by means of fungal culture isolation and subsequent antifungal susceptibility whereas one case was diagnosed by PCR-based aspergillus and azole resistance detection. All the azole-resistant aspergillosis presented TR34/L98H mutation. Patients with azole-resistant aspergillosis had different hematologic diseases: multiple myeloma, lymphoblastic acute leukemia, and angioimmunoblastic T lymphoma. Regarding risk factors, one had prolonged neutropenia, two had corticosteroids, and two had viral co-infection. Two of the patients developed aspergillosis under treatment with azoles., Conclusion: We have observed a heightened risk of azole-resistant aspergillosis caused by A. fumigatus harboring the TR 34 /L98H mutation in patients with hematologic malignancies. The emergence of azole-resistant aspergillosis raises concerns for the community, highlighting the urgent need for increased surveillance and the importance of susceptibility testing and new drugs development., (© 2024. The Author(s).)

Early detection of disseminating vancomycin-resistant Enterococcus faecium (VREfm) in ICU wards is crucial for outbreak identification and the implementation of prompt infection control measures. Genotypic methods like pulsed-field gel electrophoresis (PFGE) and whole-genome sequencing (WGS) are costly and time-consuming, hindering rapid response due to batch dependency. Fourier-transform infrared spectroscopy (FT-IR) offers the potential for real-time outbreak detection and reliable strain typing. We utilized FT-IR to identify clonal VREfm dissemination and compared its performance to PFGE and WGS. Between February through October 2023, an unusually high number of VREfm were recovered at a tertiary hospital in Barcelona. Isolates were examined for antimicrobial susceptibility, carriage of vanA/vanB genes and clonality was also studied using FT-IR, PFGE, and WGS. Routine FT-IR inspections revealed recurring VREfm clustering during the outbreak's initial weeks. In total, 104 isolates were recovered from 75 patients and from multiple wards. However, only one isolate was recovered from an environmental sample, suggesting the absence of environmental reservoirs. An ST80 vancomycin-resistant ( vanA ) E. faecium strain was the main strain responsible for the outbreak, although a few additional VREfm strains were also identified, all belonging to CC17. PFGE and cgMLST (WGS) yielded identical clustering results to FT-IR, and WGS confirmed vanA/vanB gene carriage in all VREfm isolates. Infection control measures led to a rapid decline in VREfm isolates, with no isolates detected in November. FT-IR spectroscopy offers rapid turnaround times, sensitivity, and reproducibility, comparable to standard typing methods. It proved as an effective tool for monitoring VREfm dissemination and early outbreak detection.

Staphylococcus pseudintermedius , a commensal opportunistic bacterium predominantly residing in the skin of companion animals, particularly dogs, has the potential to induce skin and soft tissue infections in pets, and zoonotic infections, including catheter-related complications. This study documents four cases of S. pseudintermedius infection or colonization in patients who had close contact with dogs or cats. Identification of the bacterial species was performed using MALDI-TOF mass spectrometry, and antibiotic susceptibility was determined using microdilution assay. DNA was sequenced using Nanopore technology followed by in silico analysis. Three isolates were multidrug resistant, including resistance to methicillin, with one belonging to the prevalent European lineage ST551, and the other two were attributed to a novel multilocus sequence type, ST2672. The remaining isolate was attributed to the novel multilocus sequence type ST2673 and was methicillin susceptible. All four isolates exhibited an array of virulence factors that contributed to colonization, damage to host immune cells, and biofilm formation. All the ST551 isolates included in the comparative analysis displayed clonality within the European continent. The importance of describing zoonotic infections associated with S. pseudintermedius resides in the scarcity of available scientific literature, further accentuated by its heightened resistance profile and potential complications, particularly in the context of catheter-related infections.

Genomic analyses of bacterial isolates are effective to compare the prevalence of antibiotic resistance genes and virulence determinants in different contexts. This study provides a comprehensive genomic description of 339 Staphylococcus aureus strains isolated from patients with bacteremia (2014-2022). Nosocomial acquisition accounted for 56.6% of cases, with vascular catheters being the main infection source (31.8%). Fatality (27.4%), persistent bacteremia (19.5%), and septic emboli (24.2%) were documented. During the COVID-19 pandemic, S. aureus bacteremia episodes increased by 140%. Genetic features in pandemic isolates revealed higher prevalence of methicillin ( mecA ) and macrolide ( msrA and mphC ) resistance genes. Additionally, genes encoding clumping factors A and B, involved in fibrinogen binding, were more prevalent. This was linked to extensive macrolide use in COVID-19 accessory therapy and elevated fibrinogen levels in SARS-CoV-2 infection. These findings highlight S. aureus adaptation to COVID-19 selective pressures and the value of whole-genome sequencing in molecular epidemiology studies., Competing Interests: The authors declare no competing interests., (© 2024 The Author(s).)

Background: Pandemic preparedness is critical to respond effectively to existing and emerging/new viral pathogens. Important lessons have been learned during the last pandemic at various levels. This revision discusses some of the major challenges and potential ways to address them in the likely event of future pandemics., Objectives: To identify critical points of readiness that may help us accelerate the response to future pandemics from a clinical microbiology laboratory perspective with a focus on viral diagnostics and genomic sequencing. The potential areas of improvement identified are discussed from the sample collection to information reporting., Sources: Microbiologists and researchers from five countries reflect on challenges encountered during the COVID-19 pandemic, review published literature on prior and current pandemics, and suggest potential solutions in preparation for future outbreaks., Content: Major challenges identified in the pre-analytic and post-analytic phases from sample collection to result reporting are discussed. From the perspective of clinical microbiology laboratories, the preparedness for a new pandemic should focus on zoonotic viruses. Laboratory readiness for scalability is critical and should include elements related to material procurement, training personnel, specific funding programmes, and regulatory issues to rapidly implement "in-house" tests. Laboratories across various countries should establish (or re-use) operational networks to communicate to respond effectively, ensuring the presence of agile circuits with full traceability of samples., Implications: Laboratory preparedness is paramount to respond effectively to emerging and re-emerging viral infections and to limit the clinical and societal impact of new potential pandemics. Agile and fully traceable methods for sample collection to report are the cornerstone of a successful response. Expert group communication and early involvement of information technology personnel are critical for preparedness. A specific budget for pandemic preparedness should be ring-fenced and added to the national health budgets., Competing Interests: Transparency declaration The authors declare that they have no conflicts of interest., (Copyright © 2023 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Introduction: Malaria is one of the most prevalent infectious diseases in sub-Saharan Africa, with 247 million cases reported worldwide in 2021 according to the World Health Organization. Optical microscopy remains the gold standard technique for malaria diagnosis, however, it requires expertise, is time-consuming and difficult to reproduce. Therefore, new diagnostic techniques based on digital image analysis using artificial intelligence tools can improve diagnosis and help automate it., Methods: In this study, a dataset of 2571 labeled thick blood smear images were created. YOLOv5x, Faster R-CNN, SSD, and RetinaNet object detection neural networks were trained on the same dataset to evaluate their performance in Plasmodium parasite detection. Attention modules were applied and compared with YOLOv5x results. To automate the entire diagnostic process, a prototype of 3D-printed pieces was designed for the robotization of conventional optical microscopy, capable of auto-focusing the sample and tracking the entire slide., Results: Comparative analysis yielded a performance for YOLOv5x on a test set of 92.10% precision, 93.50% recall, 92.79% F-score, and 94.40% mAP0.5 for leukocyte, early and mature Plasmodium trophozoites overall detection. F-score values of each category were 99.0% for leukocytes, 88.6% for early trophozoites and 87.3% for mature trophozoites detection. Attention modules performance show non-significant statistical differences when compared to YOLOv5x original trained model. The predictive models were integrated into a smartphone-computer application for the purpose of image-based diagnostics in the laboratory. The system can perform a fully automated diagnosis by the auto-focus and X-Y movements of the robotized microscope, the CNN models trained for digital image analysis, and the smartphone device. The new prototype would determine whether a Giemsa-stained thick blood smear sample is positive/negative for Plasmodium infection and its parasite levels. The whole system was integrated into the iMAGING smartphone application., Conclusion: The coalescence of the fully-automated system via auto-focus and slide movements and the autonomous detection of Plasmodium parasites in digital images with a smartphone software and AI algorithms confers the prototype the optimal features to join the global effort against malaria, neglected tropical diseases and other infectious diseases., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Maturana, de Oliveira, Nadal, Serrat, Sulleiro, Ruiz, Bilalli, Veiga, Espasa, Abelló, Suñé, Segú, López-Codina, Clols and Joseph-Munné.)

The aim of this study was to validate the detection of anti-nucleocapsid protein (N protein) antibodies for the diagnosis of SARS-CoV-2 infection in light of the fact that most COVID-19 vaccines use the spike (S) protein as the antigen. Here, 3550 healthcare workers (HCWs) were enrolled from May 2020 (when no S protein vaccines were available). We defined SARS-CoV-2 infection if HCWs were found to be positive by RT-PCR or found to be positive in at least two different serological immunoassays. Serum samples from Biobanc I3PT-CERCA were analyzed by Roche Elecsys ® (N protein) and Vircell IgG (N and S proteins) immunoassays. Discordant samples were reanalyzed with other commercial immunoassays. Roche Elecsys ® showed the positivity of 539 (15.2%) HCWs, 664 (18.7%) were found to be positive by Vircell IgG immunoassays, and 164 samples (4.6%) showed discrepant results. According to our SARS-CoV-2 infection criteria, 563 HCWs had SARS-CoV-2 infection. The Roche Elecsys ® immunoassay has a sensitivity, specificity, accuracy, and concordance with the presence of infection of 94.7%, 99.8%, 99.3%, and 0.96, respectively. Similar results were observed in a validation cohort of vaccinated HCWs. We conclude that the Roche Elecsys ® SARS-CoV-2 N protein immunoassay demonstrated good performance in diagnosing previous SARS-CoV-2 infection in a large cohort of HCWs.

Immune imprinting or original antigenic sin (OAS) is the process by which the humoral memory response to an antigen can inhibit the response to new epitopes of that antigen originating from a second encounter with the pathogen. Given the situation of the COVID-19 pandemic, multiple vaccines have been developed against SARS-CoV-2 infection. These vaccines are directed to the spike protein (S protein) of the original variant of Wuhan D614G. Vaccine memory immune response against S protein in noninfected subjects could inhibit, through the OAS mechanism, the response to new epitopes of SARS-CoV-2 after infection. The present study analyzes whether the memory antibody B cell response generated by mRNA vaccines against S protein can inhibit the primary antibody immune response to other SARS-CoV-2 antigens, such as nucleocapsid protein (N protein). SARS-CoV-2 primary infection in vaccinated healthcare workers (HCWs) produced significantly lower titers of anti-N antibodies than that in nonvaccinated HCWs: 5.7 (IQR 2.3-15.2) versus 12.2 (IQR 4.2-32.0), respectively ( p = 0.005). Therefore, spike protein vaccine-induced immune imprinting (original antigenic sin) reduces N protein antibody response., Competing Interests: The authors declare that they have no conflicts of interest., (Copyright © 2022 Juan F. Delgado et al.)