Molecular characterization of macrolide resistance in Haemophilus influenzae and Haemophilus parainfluenzae strains (2018-21).

OBJECTIVES: This study aimed to explore the prevalence of macrolide resistance and the underlying resistance mechanisms in Haemophilus influenzae (n = 2556) and Haemophilus parainfluenzae (n = 510) collected between 2018 and 2021 from Bellvitge University Hospital, Spain. METHODS: Antimicrobial susceptibility was tested by microdilution. Whole-genome sequencing was performed using Illumina MiSeq and Oxford Nanopore technologies, and sequences were examined for macrolide resistance determinants and mobile genetic structures. RESULTS: Macrolide resistance was detected in 67 H. influenzae (2.6%) and 52 (10.2%) H. parainfluenzae strains and associated with resistance to other antimicrobials (co-trimoxazole, chloramphenicol, tetracycline). Differences in macrolide resistance existed between the two species. Acquired resistance genes were more prevalent in H. parainfluenzae (35/52; 67.3%) than in H. influenzae (12/67; 17.9%). Gene mutations and amino acid substitutions were more common in H. influenzae (57/67; 85%) than in H. parainfluenzae (16/52; 30.8%). Substitutions in L22 and in 23S rRNA were only detected in H. influenzae (34.3% and 29.0%, respectively), while substitutions in L4 and AcrAB/AcrR were observed in both species. The MEGA element was identified in 35 (67.3%) H. parainfluenzae strains, five located in an integrative and conjugative element (ICE); by contrast, 11 (16.4%) H. influenzae strains contained the MEGA element (all in an ICE). A new ICEHpaHUB8 was described in H. parainfluenzae. CONCLUSIONS: Macrolide resistance was higher in H. parainfluenzae than in H. influenzae, with differences in the underlying mechanisms. H. parainfluenzae exhibits co-resistance to other antimicrobials, often leading to an extensively drug-resistant phenotype. This highlights the importance of conducting antimicrobial resistance surveillance.

A 30-year perspective of low-dose dexamethasone, a single dose of mannitol and antiseizures prophylaxis on the prognosis of pneumococcal meningitis.

OBJECTIVES: This study details the accumulated experience of more than 31 years using a low-dose systematic dexamethasone protocol with mannitol and antiseizure prophylaxis for the treatment of suspected pneumococcal meningitis. METHODS: Data have been prospectively collected for the period1977-2018. From 1987, patients with suspected pneumococcal meningitis received 12 mg dexamethasone followed by 4 mg/6 h for 48 h, started before or with the first antibiotic dose. They also received (1) a single intravenous dose of 0.5-1 g/Kg mannitol, and (2) antiseizure prophylaxis with phenytoin. RESULTS: In total, 363 episodes of pneumococcal meningitis were recorded. Of these, 242 were treated with the dexamethasone protocol after 1987 and 121 were treated without the protocol. Overall mortality was 11.6% (28/242) among patients treated with dexamethasone and 35% (43/121) among those treated without dexamethasone (p = 0.000). Early mortality was significantly lower at 5.8% (14/242) with dexamethasone and 24% (29/121) without dexamethasone (p = 0.000). Finally, neurological mortality was significantly lower at 7.4% (18/242) with dexamethasone and 23% (28/121) without dexamethasone (p = 0.000). CONCLUSIONS: A low dose of dexamethasone along with a single dose of mannitol and antiseizures prophylaxis might be useful for reducing both overall and early mortality in pneumococcal meningitis in adult patients.

Surveillance of invasive pneumococcal disease in Spain exploring the impact of the COVID-19 pandemic (2019-2023).

OBJECTIVES: Dynamic trends of invasive pneumococcal disease (IPD) including the evolution of prevalent serotypes are very useful to evaluate the impact of current and future pneumococcal conjugate vaccines (PCVs) and the rise of non-vaccine serotypes. In this study, we include epidemiological patterns of S. pneumoniae before and after COVID-19 pandemic. METHODS: We characterized all national IPD isolates from children and adults received at the Spanish Pneumococcal Reference Laboratory during 2019-2023. RESULTS: In the first pandemic year 2020, we found a general reduction in IPD cases across all age groups, followed by a partial resurgence in children in 2021 but not in adults. By 2022, IPD cases in children had returned to pre-pandemic levels, and partially in adults. In 2023, IPD rates surpassed those of the last pre-pandemic year. Notably, the emergence of serotype 3 is of significant concern, becoming the leading cause of IPD in both pediatric and adult populations over the last two years (2022-2023). Increase of serotype 4 in young adults occurred in the last epidemiological years. CONCLUSIONS: The COVID-19 pandemic led to a temporary decline in all IPD cases during 2020 attributable to non-pharmaceutical interventions followed by a subsequent rise. Employing PCVs with broader coverage and/or enhanced immunogenicity may be critical to mitigate the marked increase of IPD.

Evolution of invasive pneumococcal disease by serotype 3 in adults: a Spanish three-decade retrospective study.

Background: Invasive pneumococcal disease due to serotype 3 (S3-IPD) is associated with high mortality rates and long-term adverse effects. The introduction of the 13-valent pneumococcal conjugate vaccine (PCV13) into the Spanish paediatric immunisation programme has not led to a decrease in the adult S3-IPD. We aimed to analyse the incidence, clinical characteristics and genomics of S3-IPD in adults in Spain. Methods: Adult IPD episodes hospitalized in a Southern Barcelona hospital were prospectively collected (1994-2020). For genomic comparison, S3-IPD isolates from six Spanish hospitals (2008-2020) and historical isolates (1989-1993) were analysed by WGS (Illumina and/or MinION). Findings: From 1994 to 2020, 270 S3-IPD episodes were detected. When comparing pre-PCV (1994-2001) and late-PCV13 (2016-2020) periods, only modest changes in S3-IPD were observed (from 1.58 to 1.28 episodes per 100,000 inhabitants year). In this period, the incidence of the two main lineages shifted from 0.38 to 0.67 (CC180-GPSC12) and from 1.18 to 0.55 (CC260-GPSC83). The overall 30-day mortality remained high (24.1%), though a decrease was observed between the pre-PCV (32.4%; 95.0% CI, 22.0-45.0) and the late-PCV13 period (16.7%; 95.0% CI, 7.5-32.0) (p = 0.06). At the same time, comorbidities increased from 77.3% (95.0% CI, 65.0-86.0) to 85.7% (95.0% CI, 71.0-94.0) (p = 0.69). There were no differences in clinical characteristics or 30-day mortality between the two S3 lineages. Although both lineages were genetically homogeneous, the CC180-GPSC12 lineage presented a higher SNP density, a more open pan-genome, and a major presence of prophages and mobile genetic elements carrying resistance genes. Interpretation: Adult S3-IPD remained stable in our area over the study period despite PCV13 introduction in children. However, a clonal shift was observed. The decrease in mortality rates and the increase in comorbidities suggest a change in clinical management and overall population characteristics. The low genetic variability and absence of clinical differences between lineages highlight the role of the S3 capsule in the disease severity. Funding: This study has been funded by Instituto de Salud Carlos III (ISCIII) "PI18/00339", "PI21/01000", "INT22/00096", "FI22/00279", CIBER "CIBERES-CB06/06/0037", "CIBERINFEC-CB21/13/00009" and MSD grant "IISP 60168".

Randomized clinical trial of non-antibiotic prophylaxis with d-Mannose plus Proanthocyanidins vs. Proanthocyanidins alone for urinary tract infections and asymptomatic bacteriuria in de novo kidney transplant recipients: The Manotras study.

BACKGROUND: Studies analyzing non-antibiotic alternatives in kidney transplant UTI's are lacking. d-Mannose, a simple sugar, inhibits bacterial attachment to the urothelium, as does Proanthocyanidins; both could act as a synergic strategy preventing UTI; nonetheless their efficacy and safety have not been evaluated in kidney transplant population yet. METHODS: This is a pilot prospective, double-blind randomized trial. Sixty de novo kidney transplant recipients were randomized (1:1) to receive a prophylactic strategy based on a 24-h prolonged release formulation of d-Mannose plus Proanthocyanidins vs. Proanthocyanidins (PAC) alone. The supplements were taken for the first 3 months after kidney transplant and then followed up for 3 months as well. The main objective of the study was to search if the addition of Mannose to PAC alone reduced the incidence of UTI and/or asymptomatic bacteriuria in the first 6 months post-transplantation. RESULTS: 27% of patients experienced one UTI episode (cystitis or pyelonephritis) while asymptomatic bacteriuria was very common (57%). Incidences according UTI type or AB were: 7% vs. 4% for cystitis episode (p 0.3), 4% vs. 5% for pyelonephritis (p 0.5) and 17% vs. 14% for asymptomatic bacteriuria (p 0.4) for patients in the Mannose+PAC group vs. PAC group respectively. The most frequent bacteria isolated in both groups was Escherichia coli (28% of all episodes), UTI or AB due to E. coli was not different according to study group (30% vs. 23% for Mannose+PAC vs. PAC alone p 0.37). CONCLUSIONS: Non-antibiotic therapy is an unmet need to prevent UTI after kidney transplantation; however, the use of d-Mannose plus PAC does not seem capable to prevent it.

Multicenter clinical evaluation of a novel transcription-mediated amplification assay for SARS-CoV-2 molecular testing / Evaluación clínica multicéntrica de un nuevo ensayo de amplificación mediada por transcripción para la detección molecular de SARS-CoV-2

Introduction The onset and spread of COVID-19 pandemic has forced clinical laboratories to rapidly expand testing capacity for SARS-CoV-2. This study evaluates the clinical performance of the TMA Procleix SARS-CoV-2 assay in comparison to the RT-PCR assay Allplex™ SARS-CoV-2 for the qualitative detection of SARS-CoV-2 RNA. Methods Between November 2020 and February 2021, 610 upper-respiratory specimens received for routine SARS-CoV-2 molecular testing were prospectively collected and selected at the Hospital Universitari Vall d’Hebron and the Hospital Universitari Bellvitge in Barcelona, Spain. All samples were processed in parallel with the TMA and the RT-PCR assays, and results were compared. Discrepancies were retested by an additional RT-PCR method and the clinical history of these patients was reviewed. Results Overall, the level of concordance between both assays was 92.0% (κ, 0.772). Most discordant results (36/38, 94.7%) corresponded to samples testing positive with the TMA assay and negative with the RT-PCR method. Of these discrepant cases, most (28/36, 77.8%) were finally classified as confirmed or probable SARS-CoV-2 cases according to the discrepant analysis. Conclusion In conclusion, the TMA Procleix SARS-CoV-2 assay performed well for the qualitative detection of SARS-CoV-2 RNA in a multisite clinical setting. This novel TMA assay demonstrated a greater sensitivity in comparison to RT-PCR methods for the molecular detection of SARS-CoV-2. This higher sensitivity but also the qualitative feature of this detection of SARS-CoV-2 should be considered when making testing algorithm decisions (AU)

Introducción El inicio y la expansión de la pandemia por COVID-19 han forzado a los laboratorios clínicos a ampliar rápidamente la capacidad de detección de SARS-CoV-2. Evaluamos el rendimiento clínico del ensayo de TMA Procleix SARS-CoV-2 en comparación con el ensayo de RT-PCR Allplex™ SARS-CoV-2 para la detección cualitativa de ARN de SARS-CoV-2. Métodos Entre noviembre de 2020 y febrero de 2021 se seleccionaron prospectivamente 610 muestras del tracto respiratorio superior recibidas de rutina en el Hospital Universitario Vall d’Hebron y el Hospital Universitario de Bellvitge en Barcelona, España, para el diagnóstico molecular de SARS-CoV-2. Todas las muestras fueron procesadas en paralelo con los ensayos de TMA y RT-PCR, y se compararon los resultados. Las discrepancias se estudiaron por un método adicional de RT-PCR y se revisaron las historias clínicas de los pacientes. Resultados En general, la concordancia entre ambos ensayos fue del 92,0% (κ, 0,772). La mayoría de los casos discrepantes (36/38, 94,7%) correspondían a muestras positivas con el ensayo de TMA y negativas con el método de RT-PCR. De estos, la mayoría (28/36, 77,8%) fueron finalmente clasificados como casos confirmados o probables de SARS-CoV-2 de acuerdo al análisis de discrepantes. Conclusión El ensayo de TMA Procleix SARS-CoV-2 funcionó bien para la detección cualitativa de ARN de SARS-CoV-2 en un entorno clínico multicéntrico. Este ensayo TMA demostró una mayor sensibilidad en comparación con métodos de RT-PCR para la detección molecular de SARS-CoV-2. Esta mayor sensibilidad, pero también el carácter cualitativo de esta detección de SARS-CoV-2, se deben considerar en el diagnóstico de la infección (AU)

Development of Meropenem Resistance in a Multidrug-Resistant Campylobacter coli Strain Causing Recurrent Bacteremia in a Hematological Malignancy Patient.

Campylobacter bacteremia is an uncommon disease that mainly occurs in immunocompromised patients and is associated with antibiotic resistance, particularly in Campylobacter coli. We report a patient with persistent blood infection because of a multidrug-resistant (MDR) C. coli strain over a 3-month period. Through this period monotherapy with meropenem was associated with the development of resistance to it. Improving immunity status and a combined therapy for intestinal decolonization were useful to control persistent C. coli infection in this patient.

Multicenter clinical evaluation of a novel transcription-mediated amplification assay for SARS-CoV-2 molecular testing.

INTRODUCTION: The onset and spread of COVID-19 pandemic has forced clinical laboratories to rapidly expand testing capacity for SARS-CoV-2. This study evaluates the clinical performance of the TMA Procleix SARS-CoV-2 assay in comparison to the RT-PCR assay Allplex™ SARS-CoV-2 for the qualitative detection of SARS-CoV-2 RNA. METHODS: Between November 2020 and February 2021, 610 upper-respiratory specimens received for routine SARS-CoV-2 molecular testing were prospectively collected and selected at the Hospital Universitari Vall d'Hebron and the Hospital Universitari Bellvitge in Barcelona, Spain. All samples were processed in parallel with the TMA and the RT-PCR assays, and results were compared. Discrepancies were retested by an additional RT-PCR method and the clinical history of these patients was reviewed. RESULTS: Overall, the level of concordance between both assays was 92.0% (κ, 0.772). Most discordant results (36/38, 94.7%) corresponded to samples testing positive with the TMA assay and negative with the RT-PCR method. Of these discrepant cases, most (28/36, 77.8%) were finally classified as confirmed or probable SARS-CoV-2 cases according to the discrepant analysis. CONCLUSION: In conclusion, the TMA Procleix SARS-CoV-2 assay performed well for the qualitative detection of SARS-CoV-2 RNA in a multisite clinical setting. This novel TMA assay demonstrated a greater sensitivity in comparison to RT-PCR methods for the molecular detection of SARS-CoV-2. This higher sensitivity but also the qualitative feature of this detection of SARS-CoV-2 should be considered when making testing algorithm decisions.

A New Integrative and Mobilizable Element Is a Major Contributor to Tetracycline Resistance in Streptococcus dysgalactiae subsp. equisimilis.

Tetracycline resistance in streptococci is mainly due to ribosomal protection mediated by the tet(M) gene that is usually located in the integrative and conjugative elements (ICEs) of the Tn916-family. In this study, we analyzed the genes involved in tetracycline resistance and the associated mobile genetic elements (MGEs) in Streptococcus dysgalactiae subsp. equisimilis (SDSE) causing invasive disease. SDSE resistant to tetracycline collected from 2012 to 2019 in a single hospital and from 2018 in three other hospitals were analyzed by whole genome sequencing. Out of a total of 84 SDSE isolates, 24 (28.5%) were resistant to tetracycline due to the presence of tet(M) (n = 22), tet(W) (n = 1), or tet(L) plus tet(W) (n = 1). The tet(M) genes were found in the ICEs of the Tn916-family (n = 10) and in a new integrative and mobilizable element (IME; n = 12). Phylogenetic analysis showed a higher genetic diversity among the strains carrying Tn916 than those having the new IME, which were closely related, and all belonged to CC15. In conclusion, tetracycline resistance in SDSE is mostly due to the tet(M) gene associated with ICEs belonging to the Tn916-family and a new IME. This new IME is a major cause of tetracycline resistance in invasive Streptococcus dysgalactiae subsp. equisimilis in our settings.

Genetic Adaptation and Acquisition of Macrolide Resistance in Haemophilus spp. during Persistent Respiratory Tract Colonization in Chronic Obstructive Pulmonary Disease (COPD) Patients Receiving Long-Term Azithromycin Treatment.

Patients with chronic obstructive pulmonary disease (COPD) benefit from the immunomodulatory effect of azithromycin, but long-term administration may alter colonizing bacteria. Our goal was to identify changes in Haemophilus influenzae and Haemophilus parainfluenzae during azithromycin treatment. Fifteen patients were followed while receiving prolonged azithromycin treatment (Hospital Universitari de Bellvitge, Spain). Four patients (P02, P08, P11, and P13) were persistently colonized by H. influenzae for at least 3 months and two (P04 and P11) by H. parainfluenzae. Isolates from these patients (53 H. influenzae and 18 H. parainfluenzae) were included to identify, by whole-genome sequencing, antimicrobial resistance changes and genetic variation accumulated during persistent colonization. All persistent lineages isolated before treatment were azithromycin-susceptible but developed resistance within the first months, apart from those belonging to P02, who discontinued the treatment. H. influenzae isolates from P08-ST107 acquired mutations in 23S rRNA, and those from P11-ST2480 and P13-ST165 had changes in L4 and L22. In H. parainfluenzae, P04 persistent isolates acquired changes in rlmC, and P11 carried genes encoding MefE/MsrD efflux pumps in an integrative conjugative element, which was also identified in H. influenzae P11-ST147. Other genetic variation occurred in genes associated with cell wall and inorganic ion metabolism. Persistent H. influenzae strains all showed changes in licA and hgpB genes. Other genes (lex1, lic3A, hgpC, and fadL) had variation in multiple lineages. Furthermore, persistent strains showed loss, acquisition, or genetic changes in prophage-associated regions. Long-term azithromycin therapy results in macrolide resistance, as well as genetic changes that likely favor bacterial adaptation during persistent respiratory colonization. IMPORTANCE The immunomodulatory properties of azithromycin reduce the frequency of exacerbations and improve the quality of life of COPD patients. However, long-term administration may alter the respiratory microbiota, such as Haemophilus influenzae, an opportunistic respiratory colonizing bacteria that play an important role in exacerbations. This study contributes to a better understanding of COPD progression by characterizing the clinical evolution of H. influenzae in a cohort of patients with prolonged azithromycin treatment. The emergence of macrolide resistance during the first months, combined with the role of Haemophilus parainfluenzae as a reservoir and source of resistance dissemination, is a cause for concern that may lead to therapeutic failure. Furthermore, genetic variations in cell wall and inorganic ion metabolism coding genes likely favor bacterial adaptation to host selective pressures. Therefore, the bacterial pathoadaptive evolution in these severe COPD patients raise our awareness of the possible spread of macrolide resistance and selection of host-adapted clones.

Molecular Epidemiology, Antimicrobial Susceptibility, and Clinical Features of Methicillin-Resistant Staphylococcus aureus Bloodstream Infections over 30 Years in Barcelona, Spain (1990-2019).

Methicillin-resistant Staphylococcus aureus bloodstream infections (MRSA-BSI) are a significant cause of mortality. We analysed the evolution of the molecular and clinical epidemiology of MRSA-BSI (n = 784) in adult patients (Barcelona, 1990−2019). Isolates were tested for antimicrobial susceptibility and genotyped (PFGE), and a selection was sequenced (WGS) to characterise the pangenome and mechanisms underlying antimicrobial resistance. Increases in patient age (60 to 71 years), comorbidities (Charlson's index > 2, 10% to 94%), community-onset healthcare-associated acquisition (9% to 60%), and 30-day mortality (28% to 36%) were observed during the 1990−1995 and 2014−2019 periods. The proportion of catheter-related BSIs fell from 57% to 20%. Current MRSA-BSIs are caused by CC5-IV and an upward trend of CC8-IV and CC22-IV clones. CC5 and CC8 had the lowest core genome proportions. Antimicrobial resistance rates fell, and only ciprofloxacin, tobramycin, and erythromycin remained high (>50%) due to GyrA/GrlA changes, the presence of aminoglycoside-modifying enzymes (AAC(6')-Ie-APH(2″)-Ia and ANT(4')-Ia), and mph(C)/msr(A) or erm (C) genes. Two CC22-IV strains showed daptomycin resistance (MprF substitutions). MRSA-BSI has become healthcare-associated, affecting elderly patients with comorbidities and causing high mortality rates. Clonal replacement with CC5-IV and CC8-IV clones resulted in lower antimicrobial resistance rates. The increased frequency of the successful CC22-IV, associated with daptomycin resistance, should be monitored.

An Overview of Macrolide Resistance in Streptococci: Prevalence, Mobile Elements and Dynamics.

Streptococcal infections are usually treated with beta-lactam antibiotics, but, in case of allergic patients or reduced antibiotic susceptibility, macrolides and fluoroquinolones are the main alternatives. This work focuses on studying macrolide resistance rates, genetic associated determinants and antibiotic consumption data in Spain, Europe and also on a global scale. Macrolide resistance (MR) determinants, such as ribosomal methylases (erm(B), erm(TR), erm(T)) or active antibiotic efflux pumps and ribosomal protectors (mef(A/E)-mrs(D)), are differently distributed worldwide and associated with different clonal lineages and mobile genetic elements. MR rates vary together depending on clonal dynamics and on antibiotic consumption applying selective pressure. Among Streptococcus, higher MR rates are found in the viridans group, Streptococcus pneumoniae and Streptococcus agalactiae, and lower MR rates are described in Streptococcus pyogenes. When considering different geographic areas, higher resistance rates are usually found in East-Asian countries and milder or lower in the US and Europe. Unfortunately, the availability of data varies also between countries; it is scarce in low- and middle- income countries from Africa and South America. Thus, surveillance studies of macrolide resistance rates and the resistance determinants involved should be promoted to complete global knowledge among macrolide resistance dynamics.

Penicillin- and Cephalosporin-Resistant Pneumococcal Meningitis: Treatment in the Real World and in Guidelines.

To report on the therapy used for penicillin- and cephalosporin-resistant pneumococcal meningitis, we conducted an observational cohort study of patients admitted to our hospital with pneumococcal meningitis between 1977 and 2018. According to the European Committee on Antimicrobial Susceptibility Testing (EUCAST) recommendations, we defined pneumococci as susceptible and resistant to penicillin with MIC values of ≤0.06 mg/L and > 0.06 mg/L, respectively; the corresponding values for cefotaxime (CTX) were ≤0.5 mg/L and >0.5 mg/L. We treated 363 episodes of pneumococcal meningitis during the study period. Of these, 24 had no viable strain, leaving 339 episodes with a known MIC for inclusion. Penicillin-susceptible strains accounted for 246 episodes (73%), penicillin-resistant strains for 93 (27%), CTX susceptible for 58, and CTX resistant for 35. Nine patients failed or relapsed and 69 died (20%), of whom 22% were among susceptible cases and 17% were among resistant cases. During the dexamethasone period, mortality was equal (12%) in both susceptible and resistant cases. High-dose CTX (300 mg/Kg/day) helped to treat failed or relapsed cases and protected against failure when used as empirical therapy (P = 0.02), even in CTX-resistant cases. High-dose CTX is a good empirical therapy option for pneumococcal meningitis in the presence of a high prevalence of penicillin and cephalosporin resistance, effectively treating pneumococcal strains with MICs up to 2 mg/L for either penicillin or CTX.

Antimicrobial use and aetiology of bloodstream infections in critically ill patients during early stages of SARS-CoV-2 pandemic.

Background: During early stages of COVID-19 pandemic, antimicrobials were commonly prescribed. Aim: To describe clinical, microbiological and antimicrobial use changes in bloodstream infections (BSI) of ICU patients during the first wave of COVID-19 pandemic compared to pre-COVID-19 era. Methods: Observational cohort study of patients admitted to ICU of Bellvitge University Hospital was conducted during the COVID-19 pandemic (March-June 2020) and before COVID-19 pandemic (March-June 2019). Differences in clinical characteristics, antimicrobial consumption and incidence and aetiology of BSI were measured. Findings: COVID-19 patients had significantly less comorbidities with obesity the only risk factor that increased in frequency. COVID-19 patients more frequently required invasive supportive care measures, had longer median ICU stay and higher mortality rates. The incidence of BSIs was higher in COVID-19 period (RR 3.2 [95%CI 2.2-4.7]), occurred in patients who showed prolonged median ICU stay (21days) and was associated with high mortality rate (47%). The highest increases in the aetiological agents were observed for AmpC-producing bacteria (RR 11.1 [95%CI 2.6-47.9]) and non-fermenting rods (RR 7.0 [95%CI 1.5-31.4]). The emergence of bacteraemia caused by Gram-negative rods resistant to amoxicillin-clavulanate, which was used as empirical therapy during early stages of the pandemic, led to an escalation towards broader-spectrum antimicrobials such as meropenem and colistin which was also associated with the emergence of resistant isolates. Conclusions: The epidemiological shift towards resistant phenotypes in critically ill COVID-19 patients was associated with the selective use of antimicrobials. Our study provides evidence of the impact of empirical therapy on the selection of bacteria and their consequences on BSI over the subsequent months.

Genomic features of predominant non-PCV13 serotypes responsible for adult invasive pneumococcal disease in Spain.

BACKGROUND: Although pneumococcal conjugate vaccines (PCVs) effectively prevent invasive pneumococcal disease (IPD), serotype replacement has occurred. OBJECTIVES: We studied the pangenome, antibiotic resistance mechanisms and presence of mobile elements in predominant non-PCV13 serotypes causing adult IPD after PCV13 vaccine introduction in Spain. METHODS: We conducted a multicentre study comparing three periods in six Spanish hospitals and analysed through whole genome sequencing representative strains collected in the pre-PCV13, early-PCV13 and late-PCV13 periods. RESULTS: Among 2197 cases of adult IPD identified, 110 pneumococci expressing non-PCV13 capsules were sequenced. Seven predominant serotypes accounted for 42.6% of IPD episodes in the late-PCV13 period: serotypes 8 (14.4%), 12F (7.5%), 9N (5.2%), 11A (4.1%), 22F (3.9%), 24F (3.9%) and 16F (3.6%). All predominant non-PCV13 serotypes were highly clonal, comprising one or two clonal complexes (CC). In general, CC538, CC4048, CC3016F, CC43322F and CC669N, related to predominant non-PCV13 serotypes, were antibiotic susceptible. CC15611A was associated with resistance to co-trimoxazole, penicillin and amoxicillin. CC23024F was non-susceptible to penicillin and resistant to erythromycin, clindamycin, and tetracycline. Six composite transposon structures of the Tn5252-family were found in CC23024F, CC98912F and CC3016F carrying different combinations of erm(B), tet(M), and cat. Pangenome analysis revealed differences in accessory genomes among the different CC, with most variety in CC3016F (23.9%) and more conservation in CC15611A (8.5%). CONCLUSIONS: We identified highly clonal predominant serotypes responsible for IPD in adults. The detection of not only conjugative elements carrying resistance determinants but also clones previously associated with vaccine serotypes (CC15611A and CC23024F) highlights the importance of the accessory genome.

Effect of positive microbiological testing on antibiotic de-escalation and outcomes in community-acquired pneumonia: a propensity score analysis.

OBJECTIVES: The usefulness of routine microbiological testing for rationalising antibiotic use in hospitalised patients with community-acquired pneumonia (CAP) continues to be a subject of debate. We aim to determine the effect of positive microbiological testing on antimicrobial de-escalation and clinical outcomes in CAP. METHODS: A retrospective analysis of a prospectively collected cohort of non-immunosuppressed adults hospitalised with CAP was performed. The primary study outcome was antimicrobial de-escalation. Secondary outcomes included 30-day case-fatality rate, adverse events, and CAP recurrence. Adjustment for confounders was performed by inverse probability weighting propensity score, logistic regression, and cause-specific Cox model. RESULTS: Of 3677 patients with CAP, 1924 (52.3%) had any positive microbiological test. Antimicrobial de-escalation was performed in 648/1924 (33.7%) of patients with positive microbiological testing and in 179/1753 (10.2%) of those with non-positive results. When propensity score was entered into the multivariate analysis, positive microbiological testing (adjusted OR (AOR)], 2.59; 1.96-3.41) and clinical stability at day 3 (AOR 1.87; 1.45-2.10) were two of the main factors independently associated with antimicrobial de-escalation. After applying an adjusted cause-specific Cox model, antimicrobial de-escalation was not associated with a higher 30-day case-fatality rate (adjusted hazard ratio (AHR), 0.44 (95% CI, 0.14-1.43)), higher frequency of adverse events (AHR, 0.77 (95% CI, 0.53-1.12)), or CAP recurrence (AHR, 0.65 (95% CI, 0.35-1.14)). DISCUSSION: Antimicrobial de-escalation was more often performed in hospitalised patients with CAP who had positive microbiological tests than in those with non-positive results, and it did not adversely affect relevant clinical outcomes.

Long-lasting adaptive immune memory specific to SARS-CoV-2 in convalescent coronavirus disease 2019 stable people with HIV.

OBJECTIVE: While the course of natural immunization specific to SARS-CoV-2 has been described among convalescent coronavirus disease 2019 (COVID-19) people without HIV (PWOH), a thorough evaluation of long-term serological and functional T- and B-cell immune memory among people with HIV (PWH) has not been reported. METHODS: Eleven stable PWH developing mild ( n  = 5) and severe ( n  = 6) COVID-19 and 39 matched PWOH individuals with mild (MILD) ( n  = 20) and severe (SEV) ( n  = 19) COVID-19 infection were assessed and compared at 3 and 6 months after infection for SARS-CoV-2-specific serology, polyfunctional cytokine (interferon-γ [IFN-γ], interleukin 2 [IL-2], IFN-γ/IL-2, IL-21) producing T-cell frequencies against four main immunogenic antigens and for circulating SARS-CoV-2-specific immunoglobulin G (IgG)-producing memory B-cell (mBc). RESULTS: In all time points, all SARS-COV-2-specific adaptive immune responses were highly driven by the clinical severity of COVID-19 infection, irrespective of HIV disease. Notably, while a higher proportion of mild PWH showed a higher decay on serological detection between the two time points as compared to PWOH, persistently detectable IgG-producing mBc were still detectable in most patients (4/4 (100%) for SEV PWH, 4/5 (80%) for MILD PWH, 10/13 (76.92%) for SEV PWOH and 15/18 (83.33%) for MILD PWOH). Likewise, SARS-CoV-2-specific IFN-γ-producing T-cell frequencies were detected in both PWH and PWOH, although significantly more pronounced among severe COVID-19 (6/6 (100%) for SEV PWH, 3/5 (60%) for MILD PWH, 18/19 (94.74%) for SEV PWOH and 14/19 (73.68%) for MILD PWOH). CONCLUSIONS: PWH develop a comparable short and long-term natural functional cellular and humoral immune response than PWOH convalescent patients, which are highly influenced by the clinical severity of the COVID-19 infection.

SARS-CoV-2 outbreak in a nursing home after vaccination with BNT162b2: A role for the quantification of circulating antibodies.

We describe an outbreak of SARS-CoV-2 (B.1.351) in a nursing home. At the outbreak onset 96% of residents and 76% of HCW had received two doses of BNT162b2. Twenty-eight residents (28/53) and six HCW (6/33) were infected. Infected residents had lower levels of anti-S antibodies compared to those who were not infected (157 vs 552 U/mL). Among 50 residents with available serological status, nineteen (19/25) with serum concentration < 300 U/mL and seven (7/25) with concentration > 300 U/mL acquired SARS-CoV-2 (RR 2.7 [95 %CI 1.4-5.3]). The quantification of circulating antibodies could be useful in detecting people with an impaired immune response who are at high risk of acquiring and spreading SARS-CoV-2.

Multicenter clinical evaluation of a novel transcription-mediated amplification assay for SARS-CoV-2 molecular testing.

INTRODUCTION: The onset and spread of COVID-19 pandemic has forced clinical laboratories to rapidly expand testing capacity for SARS-CoV-2. This study evaluates the clinical performance of the TMA Procleix SARS-CoV-2 assay in comparison to the RT-PCR assay AllplexTM SARS-CoV-2 for the qualitative detection of SARS-CoV-2 RNA. METHODS: Between November 2020 and February 2021, 610 upper-respiratory specimens received for routine SARS-CoV-2 molecular testing were prospectively collected and selected at the Hospital Universitari Vall d'Hebron and the Hospital Universitari Bellvitge in Barcelona, Spain. All samples were processed in parallel with the TMA and the RT-PCR assays, and results were compared. Discrepancies were retested by an additional RT-PCR method and the clinical history of these patients was reviewed. RESULTS: Overall, the level of concordance between both assays was 92.0% (κ, 0.772). Most discordant results (36/38, 94.7%) corresponded to samples testing positive with the TMA assay and negative with the RT-PCR method. Of these discrepant cases, most (28/36, 77.8%) were finally classified as confirmed or probable SARS-CoV-2 cases according to the discrepant analysis. CONCLUSION: In conclusion, the TMA Procleix SARS-CoV-2 assay performed well for the qualitative detection of SARS-CoV-2 RNA in a multisite clinical setting. This novel TMA assay demonstrated a greater sensitivity in comparison to RT-PCR methods for the molecular detection of SARS-CoV-2. This higher sensitivity but also the qualitative feature of this detection of SARS-CoV-2 should be considered when making testing algorithm decisions.