Acidic Urine pH and Clinical Outcome of Lower Urinary Tract Infection in Kidney Transplant Recipients Treated with Ciprofloxacin and Fosfomycin.

Different factors, including antimicrobial resistance, may diminish the effectiveness of antibiotic therapy, challenging the management of post-transplant urinary tract infection (UTI). The association of acidic urine pH with microbiological and clinical outcomes was evaluated after fosfomycin or ciprofloxacin therapy in 184 kidney transplant recipients (KTRs) with UTI episodes by Escherichia coli (N = 115) and Klebsiella pneumoniae (N = 69). Initial urine pH, antimicrobial therapy, and clinical and microbiological outcomes, and one- and six-month follow-up were assessed. Fosfomycin was prescribed in 88 (76.5%) E. coli and 46 (66.7%) K. pneumoniae UTI episodes in the total cohort. When the urine pH ≤ 6, fosfomycin was prescribed in 60 (52.2%) E. coli and 29 (42.0%) K. pneumoniae. Initial urine pH ≤ 6 in E. coli UTI was associated with symptomatic episodes (8/60 vs. 0/55, p = 0.04) at one-month follow-up, with a similar trend in those patients receiving fosfomycin (7/47 vs. 0/41, p = 0.09). Acidic urine pH was not associated with microbiological or clinical cure in K. pneumoniae UTI. At pH 5, the ciprofloxacin MIC90 increased from 8 to >8 mg/L in E. coli and from 4 to >8 mg/L in K. pneumoniae. At pH 5, the fosfomycin MIC90 decreased from 8 to 4 mg/L in E. coli and from 512 to 128 mg/L in K. pneumoniae. Acidic urine is not associated with the microbiological efficacy of fosfomycin and ciprofloxacin in KTRs with UTI, but it is associated with symptomatic UTI episodes at one-month follow-up in E. coli episodes.

Analysis of bacterial pangenomes reduces CRISPR dark matter and reveals strong association between membranome and CRISPR-Cas systems.

CRISPR-Cas systems are prokaryotic acquired immunity mechanisms, which are found in 40% of bacterial genomes. They prevent viral infections through small DNA fragments called spacers. However, the vast majority of these spacers have not yet been associated with the virus they recognize, and it has been named CRISPR dark matter. By analyzing the spacers of tens of thousands of genomes from six bacterial species, we have been able to reduce the CRISPR dark matter from 80% to as low as 15% in some of the species. In addition, we have observed that, when a genome presents CRISPR-Cas systems, this is accompanied by particular sets of membrane proteins. Our results suggest that when bacteria present membrane proteins that make it compete better in its environment and these proteins are, in turn, receptors for specific phages, they would be forced to acquire CRISPR-Cas.

Clinical and Ecological Impact of an Educational Program to Optimize Antibiotic Treatments in Nursing Homes (PROA-SENIOR): A Cluster, Randomized, Controlled Trial and Interrupted Time-Series Analysis.

BACKGROUND: Antimicrobial stewardship programs (ASPs) are recommended in nursing homes (NHs), although data are limited. We aimed to determine the clinical and ecological impact of an ASP for NHs. METHODS: We performed a cluster, randomized, controlled trial and a before-after study with interrupted time-series analyses in 14 NHs for 30 consecutive months from July 2018 to December 2020 in Andalusia, Spain. Seven facilities implemented an ASP with a bundle of 5 educational measures (general ASP) and 7 added 1-to-1 educational interviews (experimental ASP). The primary outcome was the overall use of antimicrobials, calculated monthly as defined daily doses (DDD) per 1000 resident days (DRD). RESULTS: The total mean antimicrobial consumption decreased by 31.2% (-16.72 DRD; P = .045) with respect to the preintervention period; the overall use of quinolones and amoxicillin-clavulanic acid dropped by 52.2% (P = .001) and 42.5% (P = .006), respectively; and the overall prevalence of multidrug-resistant organisms (MDROs) decreased from 24.7% to 17.4% (P = .012). During the intervention period, 12.5 educational interviews per doctor were performed in the experimental ASP group; no differences were found in the total mean antimicrobial use between groups (-14.62 DRD; P = .25). Two unexpected coronavirus disease 2019 waves affected the centers increasing the overall mean use of antimicrobials by 40% (51.56 DRD; P < .0001). CONCLUSIONS: This study suggests that an ASP for NHs appears to be associated with a decrease in total consumption of antimicrobials and prevalence of MDROs. This trial did not find benefits associated with educational interviews, probably due to the coronavirus disease 2019 pandemic. Clinical Trials Registration. NCT03543605.

Study protocol for a randomized clinical trial to assess 7 versus 14-days of treatment for Pseudomonas aeruginosa bloodstream infections (SHORTEN-2 trial).

BACKGROUND: Research priorities in Antimicrobial Stewardship (AMS) have rapidly evolved in the last decade. The need for a more efficient use of antimicrobials have fueled plenty of studies to define the optimal duration for antibiotic treatments, and yet, there still are large areas of uncertainty in common clinical scenarios. Pseudomonas aeruginosa has been pointed as a priority for clinical research, but it has been unattended by most randomized trials tackling the effectiveness of short treatments. The study protocol of the SHORTEN-2 trial is presented as a practical example of new ways to approach common obstacles for clinical research in AMS. OBJECTIVE: To determine whether a 7-day course of antibiotics is superior to 14-day schemes for treating bloodstream infections by P. aeruginosa (BSI-PA). METHODS: A superiority, open-label, randomized controlled trial will be performed across 30 Spanish hospitals. Adult patients with uncomplicated BSI-PA will be randomized to receive a 7 versus 14-day course of any active antibiotic. The primary endpoint will be the probability for the 7-day group of achieving better outcomes than the control group, assessing altogether clinical effectiveness, severe adverse events, and antibiotic exposure through a DOOR/RADAR analysis. Main secondary endpoints include treatment failure, BSI-PA relapses, and mortality. A superiority design was set for the primary endpoint and non-inferiority for treatment failure, resulting in a sample size of 304 patients. CONCLUSIONS: SHORTEN-2 trial aligns with some of the priorities for clinical research in AMS. The implementation of several methodological innovations allowed overcoming common obstacles, like feasible sample sizes or measuring the clinical impact and unintended effects. TRIAL REGISTRATION: EudraCt: 2021-003847-10; ClinicalTrials.gov: NCT05210439.

Carbapenem Combinations for Infections Caused by Carbapenemase-Producing Pseudomonas aeruginosa: Experimental In Vitro and In Vivo Analysis.

In the context of difficult-to-treat carbapenem-resistant Pseudomonas aeruginosa infections, we evaluated imipenem, meropenem, and doripenem combinations against eleven carbapenemase-producing P. aeruginosa isolates. According to the widespread global distribution of high-risk clones and carbapenemases, four representative isolates were selected: ST175 (OXA-2/VIM-20), ST175 (VIM-2), ST235 (GES-5), and ST111 (IMP-33), for efficacy studies using a sepsis murine model. Minimum inhibitory concentration (mg/L) ranges were 64-256 for imipenem and 16-128 for meropenem and doripenem. In vitro, imipenem plus meropenem was synergistic against 72% of isolates and doripenem plus meropenem or imipenem against 55% and 45%, respectively. All combinations were synergistic against the ST175, ST235, and ST155 clones. In vivo, meropenem diminished the spleen and blood bacterial concentrations of four and three isolates, respectively, with better efficacy than imipenem or doripenem. The combinations did not show efficacy compared with the more active monotherapies, except for imipenem plus meropenem, which reduced the ST235 bacterial spleen concentration. Mortality decreased with imipenem plus meropenem or doripenem for the ST175 isolate. Results suggest that carbapenem combinations are not an alternative for severe infections by carbapenemase-producing P. aeruginosa. Meropenem monotherapy showed in vivo efficacy despite its high MIC, probably because its dosage allowed a sufficient antimicrobial exposure at the infection sites.

IgM-enriched immunoglobulin improves colistin efficacy in a pneumonia model by Pseudomonas aeruginosa.

We evaluated the efficacy of ceftazidime or colistin in combination with polyclonal IgM-enriched immunoglobulin (IgM-IG), in an experimental pneumonia model (C57BL/6J male mice) using two multidrug-resistant Pseudomonas aeruginosa strains, both ceftazidime-susceptible and one colistin-resistant. Pharmacodynamically optimised antimicrobials were administered for 72 h, and intravenous IgM-IG was given as a single dose. Bacterial tissues count and the mortality were analysed. Ceftazidime was more effective than colistin for both strains. In mice infected with the colistin-susceptible strain, ceftazidime reduced the bacterial concentration in the lungs and blood (-2.42 and -3.87 log10 CFU/ml) compared with colistin (-0.55 and -1.23 log10 CFU/ml, respectively) and with the controls. Colistin plus IgM-IG reduced the bacterial lung concentrations of both colistin-susceptible and resistant strains (-2.91 and -1.73 log10 CFU/g, respectively) and the bacteraemia rate of the colistin-resistant strain (-44%). These results suggest that IgM-IG might be useful as an adjuvant to colistin in the treatment of pneumonia caused by multidrug-resistant P. aeruginosa.

Prevalence and Risk Factors for Multidrug-Resistant Organisms Colonization in Long-Term Care Facilities Around the World: A Review.

Elderly people confined to chronic care facilities face an increased risk of acquiring infections by multidrug-resistant organisms (MDROs). This review presents the current knowledge of the prevalence and risk factors for colonization by MDROs in long-term care facilities (LTCF), thereby providing a useful reference to establish objectives for implementing successful antimicrobial stewardship programs (ASPs). We searched in PubMed and Scopus for studies examining the prevalence of MDROs and/or risk factors for the acquisition of MDROs in LTCF. One hundred and thirty-four studies published from 1987 to 2020 were included. The prevalence of MDROs in LTCF varies between the different continents, where Asia reported the highest prevalence of extended-spectrum ß-lactamase (ESBL) Enterobacterales (71.6%), carbapenem resistant (CR) Enterobacterales (6.9%) and methicillin-resistant Staphylococcus aureus (MRSA) (25.6%) and North America the highest prevalence to MDR Pseudomonas aeruginosa (5.4%), MDR Acinetobacter baumannii (15.0%), vancomycin-resistant Enterococcus spp. (VRE) (4.0%), and Clostridioides difficile (26.1%). Furthermore, MDRO prevalence has experienced changes over time, with increases in MDR P. aeruginosa and extended spectrum ß-lactamase producing Enterobacterales observed starting in 2015 and decreases of CR Enterobacterales, MDR A. baumannii, VRE, MRSA and C. difficile. Several risk factors have been found, such as male sex, chronic wounds, the use of medical devices, and previous antibiotic use. The last of these aspects represents one of the most important modifiable factors for reducing colonization with MDROs through implementing ASPs in LTCF.

Are antimicrobial stewardship interventions effective and safe in long-term care facilities? A systematic review and meta-analysis.

BACKGROUND: Long-term care facilities (LTCFs) are health-care settings with high antimicrobial consumption and hence need to develop effective antimicrobial stewardship programmes (ASPs). OBJECTIVE: To assess the effects of ASPs on care-related, clinical and ecological outcomes in LTCFs. METHODS: Data sources were PubMed, EMBASE, CINAHL and SCOPUS. Study eligibility criteria were original research articles (controlled clinical trials or controlled before and after studies) published up to 1 October 2020. Participants were adult residents of LTCFs, residential aged-care facilities, nursing homes, veterans' homes, skilled nursing facilities and assisted living facilities for older people. Interventions included ASPs versus standard care. Outcomes assessed were antimicrobial consumption and appropriateness, infections, hospital admissions and mortality. Available data were pooled in a meta-analysis, and inconsistency between studies was evaluated using the I2 statistic. Certainty of evidence was assessed using the GRADE approach. RESULTS: Of the 3111 papers identified, 12 studies met the inclusion criteria. All of them analysed the impact of interventions on antimicrobial use based on consumption-related variables (n = 8) and/or percentage of inappropriate prescriptions (n = 6). Pooled data showed a mean difference of -0.47 prescriptions per 1000 resident-days in favour of ASPs (95% CI -0.87 to -0.07, I2 = 71%). Five studies analysed the clinical effect of ASPs on the number of hospital admissions and/or resident mortality. The meta-analysis showed a mean difference of 0.17 hospital admissions per 1000 resident-days (95% CI -0.07 to 0.41, I2 = 17%) and a mean difference of -0.02 deaths per 1000 resident-days (95% CI -0.14 to 0.09, I2 = 0%). Only two studies included infections as a study outcome. CONCLUSIONS: ASPs appear to improve antimicrobial use in this setting without increasing hospital admissions or deaths, indicating that these programmes do not lead to under-treatment of infections. Nonetheless, further higher-quality clinical trials are required to understand the effects of ASPs in LTCFs. PROSPERO REGISTRATION NUMBER: CRD42021225127.

Impact of Treating Asymptomatic Bacteriuria in Kidney Transplant Recipients: A Prospective Cohort Study.

This study aims to define the epidemiologic, clinical, and microbiological features of asymptomatic bacteriuria (AB) and cystitis in kidney transplantation recipients (KTRs), and to determine the impact of antimicrobial therapy of AB and the risk factors of cystitis. We conducted a prospective observational study of AB and cystitis in KTRs from January to June 2017. One-hundred ninety seven KTRs were included: 175 (88.8%) with AB and 22 (11.2%) with cystitis. The most frequent etiologies were Escherichia coli, Klebsiellapneumoniae, Enterococcusfaecalis, and Pseudomonas aeruginosa. No differences were observed regarding the etiologies, antimicrobial susceptibility patterns, and microbiologic outcomes in AB vs. cystitis. The treatment of AB diminished the microbiological cure and increased the rates of microbiologic relapses and reinfections; in addition, treated AB patients showed a trend of developing symptomatic urinary tract infection in the following six months. The analysis of the data identified the following independent risk factors for cystitis during the six months of follow-up: AB treatment, thymoglobulin induction, previous acute pyelonephritis, and time since transplantation < 1 year. In summary, considering the lack of clinical benefits of treating AB and its impact on cystitis development in the follow-up, we support the recommendation of not screening for or treating AB.

In Vitro Activity of Pentamidine Alone and in Combination with Antibiotics against Multidrug-Resistant Clinical Pseudomonas aeruginosa Strains.

Multidrug-resistant (MDR) Pseudomonas aeruginosa is a public health problem causing both community and hospital-acquired infections, and thus the development of new therapies for these infections is critical. The objective of this study was to analyze in vitro the activity of pentamidine as adjuvant in combinations to antibiotics against seven clinical P. aeruginosa strains. The Minimum Inhibitory Concentration (MIC) was determined following standard protocols, and the results were interpreted according to the European Committee on Antimicrobial Susceptibility Testing (EUCAST) breakpoints; however, the gentamicin activity was interpreted according to the Clinical and Laboratory Standards Institute (CLSI) recommendations. The bactericidal in vitro activity was studied at 1×MIC concentrations by time-kill curves, and also performed in three selected strains at 1/2×MIC of pentamidine. All studies were performed in triplicate. The pentamidine MIC range was 400-1600 µg/mL. Four of the strains were MDR, and the other three were resistant to two antibiotic families. The combinations of pentamidine at 1×MIC showed synergistic activity against all the tested strains, except for pentamidine plus colistin. Pentamidine plus imipenem and meropenem were the combinations that showed synergistic activity against the most strains. At 1/2×MIC, pentamidine plus antibiotics were synergistic with all three analyzed strains. In summary, pentamidine in combination with antibiotics showed in vitro synergy against multidrug-resistant P. aeruginosa clinical strains, which suggests its possible use as adjuvant to antibiotics for the therapy of infections from MDR P. aeruginosa.

Virulence role of the outer membrane protein CarO in carbapenem-resistant Acinetobacter baumannii.

Novel approaches to treat carbapenem-resistant Acinetobacter baumannii (CRAB) infections are urgently needed and anti-virulence drugs represent promising alternatives, but our knowledge on potential targets is scarce. We searched for potential A. baumannii virulence factors by whole-genome sequencing-based comparisons of CRAB clinical isolates causing bloodstream infections secondary to ventilator-associated pneumonia from demographics and clinically homogeneous patients, who received optimal treatment but with different clinical outcomes. Thus, the carO gene was interrupted in CRAB isolates from surviving patients, while it was intact in isolates from non-surviving patients, and proteomic/immunoblot techniques corroborated it. When the virulence role of A. baumannii CarO was analyzed in model systems, isogenic ΔcarO mutants and a CRAB clinical isolate with truncated CarO, showed lower ability to adhere and invade A549 cells and in vivo virulence. This unnoticed virulence role for CarO postulate this A. baumannii outer membrane protein as a potential target for new therapies against CRAB infections.

Discovery of Novel Substituted N-(4-Amino-2-chlorophenyl)-5-chloro-2-hydroxybenzamide Analogues as Potent Human Adenovirus Inhibitors.

An effective therapy for human adenovirus (HAdV) infections in immunocompromised patients and healthy individuals with community-acquired pneumonia remains an unmet medical need. We herein reported a series of novel substituted N-(4-amino-2-chlorophenyl)-5-chloro-2-hydroxybenzamide analogues as potent HAdV inhibitors. Compounds 6, 15, 29, 40, 43, 46, 47, and 54 exhibited increased selectivity indexes (SI > 100) compared to the lead compound niclosamide, while maintaining sub-micromolar to low micromolar potency against HAdV. The preliminary mechanistic studies indicated that compounds 6 and 43 possibly target the HAdV DNA replication process, while compounds 46 and 47 suppress later steps of HAdV life cycle. Notably, among these derivatives, compound 15 showed improved anti-HAdV activity (IC50 = 0.27 µM), significantly decreased cytotoxicity (CC50 = 156.8 µM), and low in vivo toxicity (maximum tolerated dose = 150 mg/kg in hamster) as compared with niclosamide, supporting its further in vivo efficacy studies for the treatment of HAdV infections.

Assessing the impact on intestinal microbiome and clinical outcomes of antibiotherapy optimisation strategies in haematopoietic stem cell transplant recipients: study protocol for the prospective multicentre OptimBioma study.

INTRODUCTION: Haematopoietic stem cell transplantation (HSCT) is a life-saving treatment for a number of haematological diseases. Graft versus host disease (GVHD) is its main complication and hampers survival. There is strong evidence that intestinal microbiota diversity of the recipient may increase the risk of GVHD worsening survival. Antibiotic regimens used during the early phase of the transplant may influence clinical outcomes by reducing intestinal microbiota diversity. Present guidelines of European Conference on Infections in Leukaemia exhort to optimising antibiotic use in haematological patients including HSCT recipients. The present study aims to investigate if, in HSCT recipients, the optimisation of antibacterial use may preserve intestinal microbiota composition reducing the incidence and severity of acute GVHD and improving relevant clinical outcomes. METHODS AND ANALYSIS: This is a prospective longitudinal observational study of two cohorts of HSCT recipients: (1) the intervention cohort includes patients treated in centres in which a predefined strategy of antibiotherapy optimisation is implemented, with the objective of optimising and reducing antibiotic administration according to clinical criteria and (2) the control cohort includes patients treated in centres in which a classic permissive strategy of antibiotic prophylaxis and treatment is used. Adult patient receiving a first HSCT as a treatment for any haematological condition are included. Clinical variables are prospectively recorded and up to five faecal samples are collected for microbiota characterisation at prestablished peritransplant time points. Patients are followed since the preconditioning phase throughout 1-year post-transplant and four follow-up visits are scheduled. Faecal microbiota composition and diversity will be compared between both cohorts along with acute GVHD incidence and severity, severe infections rate, mortality and overall and disease-free survival. ETHICS AND DISSEMINATION: The study was approved between 2017 and 2018 by the Ethical Committees of participant centres. Study results will be disseminated through peer-reviewed journals and national and international scientific conferences. TRIAL REGISTRATION NUMBER: NCT03727113.

Exploration of piperazine-derived thioureas as antibacterial and anti-inflammatory agents. In vitro evaluation against clinical isolates of colistin-resistant Acinetobacter baumannii.

A. baumannii is one of the most important multidrug-resistant microorganisms in hospital units. It is resistant to many classes of antibiotics and the development of new therapeutic strategies is necessary. The aim of this study was to evaluate the antibacterial activity of a set of piperazine-derived thioureas against 13 clinical strains of colistin-resistant A. baumannii. Six derivatives were identified to inhibit bacterial growth of 46% of the A. baumannii strains at low micromolar concentrations (Minimum Inhibitory Concentration from 1.56 to 6.25 µM). A common structural feature in most active compounds was the presence of a 3,5-bis-trifluoromethyl phenyl ring at the thiourea function. In addition, the ability of the compounds to inhibit production of nitric oxide (NO) was examined in RAW 264.7 murine macrophages, highlighting the potential of piperazine-derived thioureas as promising scaffolds for the design of new combined anti-bacterial/anti-inflammatory agents.

Role of PstS in the Pathogenesis of Acinetobacter baumannii Under Microaerobiosis and Normoxia.

Acinetobacter baumannii is a successful pathogen responsible for infections with high mortality rate. During the course of infection it can be found in microaerobic environments, which influences virulence factor expression. From a previous transcriptomic analysis of A. baumannii ATCC 17978 under microaerobiosis, we know the gene pstS is overexpressed under microaerobiosis. Here, we studied its role in A. baumannii virulence. pstS loss significantly decreased bacterial adherence and invasion into A549 cells and increased A549 cell viability. pstS loss also reduced motility and biofilm-forming ability of A. baumannii. In a peritoneal sepsis murine model, the minimum lethal dose required by A. baumannii ATCC 17978 ΔpstS was lower compared to the wild type (4.3 vs 3.2 log colony forming units/mL, respectively), and the bacterial burden in tissues and fluids was lower. Thus, the loss of the phosphate sensor PstS produced a decrease in A. baumannii pathogenesis, supporting its role as a virulence factor.

Extended-spectrum resistance to ß-lactams/ß-lactamase inhibitors (ESRI) evolved from low-level resistant Escherichia coli.

OBJECTIVES: Escherichia coli is characterized by three resistance patterns to ß-lactams/ß-lactamase inhibitors (BLs/BLIs): (i) resistance to ampicillin/sulbactam and susceptibility to amoxicillin/clavulanic acid and piperacillin/tazobactam (RSS); (ii) resistance to ampicillin/sulbactam and amoxicillin/clavulanic acid, and susceptibility to piperacillin/tazobactam (RRS); and (iii) resistance to ampicillin/sulbactam, amoxicillin/clavulanic acid and piperacillin/tazobactam (RRR). These resistance patterns are acquired consecutively, indicating a potential risk of developing resistance to piperacillin/tazobactam, but the precise mechanism of this process is not completely understood. METHODS: Clinical isolates incrementally pressured by piperacillin/tazobactam selection in vitro and in vivo were used. We determined the MIC of piperacillin/tazobactam in the presence and absence of piperacillin/tazobactam pressure. We deciphered the role of the blaTEM genes in the new concept of extended-spectrum resistance to BLs/BLIs (ESRI) using genomic analysis. The activity of ß-lactamase was quantified in these isolates. RESULTS: We show that piperacillin/tazobactam resistance is induced in E. coli carrying blaTEM genes. This resistance is due to the increase in copy numbers and transcription levels of the blaTEM gene, thus increasing ß-lactamase activity and consequently increasing piperacillin/tazobactam MICs. Genome sequencing of two blaTEM-carrying representative isolates showed that piperacillin/tazobactam treatment produced two types of duplications of blaTEM (8 and 60 copies, respectively). In the clinical setting, piperacillin/tazobactam treatment of patients infected by E. coli carrying blaTEM is associated with a risk of therapeutic failure. CONCLUSIONS: This study describes for the first time the ESRI in E. coli. This new concept is very important in the understanding of the mechanism involved in the acquisition of resistance to BLs/BLIs.

Pangenome of Acinetobacter baumannii uncovers two groups of genomes, one of them with genes involved in CRISPR/Cas defence systems associated with the absence of plasmids and exclusive genes for biofilm formation.

Acinetobacter baumannii is an opportunistic bacterium that causes hospital-acquired infections with a high mortality and morbidity, since there are strains resistant to virtually any kind of antibiotic. The chase to find novel strategies to fight against this microbe can be favoured by knowledge of the complete catalogue of genes of the species, and their relationship with the specific characteristics of different isolates. In this work, we performed a genomics analysis of almost 2500 strains. Two different groups of genomes were found based on the number of shared genes. One of these groups rarely has plasmids, and bears clustered regularly interspaced short palindromic repeat (CRISPR) sequences, in addition to CRISPR-associated genes (cas genes) or restriction-modification system genes. This fact strongly supports the lack of plasmids. Furthermore, the scarce plasmids in this group also bear CRISPR sequences, and specifically contain genes involved in prokaryotic toxin-antitoxin systems that could either act as the still little known CRISPR type IV system or be the precursors of other novel CRISPR/Cas systems. In addition, a limited set of strains present a new cas9-like gene, which may complement the other cas genes in inhibiting the entrance of new plasmids into the bacteria. Finally, this group has exclusive genes involved in biofilm formation, which would connect CRISPR systems to the biogenesis of these bacterial resistance structures.

Prevalence and clinical impact of Streptococcus pneumoniae nasopharyngeal carriage in solid organ transplant recipients.

BACKGROUND: S. pneumoniae is the leading cause of community-acquired pneumonia in the solid organ transplant recipient (SOTR); nevertheless, the prevalence of colonization and of the colonizing/infecting serotypes has not been studied in this population. In this context, the aim of the present study was to describe the rate, characteristics, and clinical impact of S. pneumoniae nasopharyngeal carriage. METHODS: A prospective observational cohort of Solid Organ Transplant recipients (SOTR) was held at the University Hospital Virgen del Rocío, Seville, Spain with the aim to evaluate the S. pneumoniae colonization and the serotype prevalence in SOTR. Two different pharyngeal swabs samples from 500 patients were included in two different seasonal periods winter and spring/summer. Optochin and bile solubility tests were performed for the isolation of thew strains. Antimicrobial susceptibility studies (MICs, mg/l) of levofloxacin, trimethoprim-sulfamethoxazole, penicillin, amoxicillin, cefotaxime, ceftriaxone, erythromycin, azithromycin and vancomycin for each isolate were determined by E-test strips. Capsular typing was done by sequential multiplex PCR reactions. A multivariate logistic regression analysis of factors potentially associated with pneumococcal nasopharyngeal carriage and disease was performed. RESULTS: Twenty-six (5.6%) and fifteen (3.2%) patients were colonized in winter and spring/summer periods, respectively. Colonized SOT recipients compared to non-colonized patients were more frequently men (79.5% vs. 63.1%, P < 0.05) and cohabitated regularly with children (59% vs. 32.2%, P < 0.001). The most prevalent serotype in both studied periods was 35B. Forty-five percent of total isolates were included in the pneumococcal vaccine PPV23. Trimethoprim-sulfamethoxazole and macrolides were the less active antibiotics. Three patients had non-bacteremic pneumococcal pneumonia, and two of them died. CONCLUSIONS: Pneumococcal colonization in SOTR is low with the most colonizing serotypes not included in the pneumococcal vaccines.

The anthelmintic oxyclozanide restores the activity of colistin against colistin-resistant Gram-negative bacilli.

Due to the significant increase in antimicrobial resistance in Gram-negative bacilli (GNB), development of non-antimicrobial therapeutic alternatives, which can be used together with the few and non-optimal available antimicrobial agents such as colistin, has become an urgent need. In this context, dysregulation of the bacterial cell wall could be a therapeutic adjuvant to the activity of colistin. The aim of this study was to analyse the activity of oxyclozanide, an anthelmintic drug, in combination with colistin against colistin-susceptible (Col-S) and colistin-resistant (Col-R) GNB. Three Col-S reference strains and 13 clinical isolates (1 Col-S, 12 Col-R) of Acinetobacter baumannii, Pseudomonas aeruginosa and Klebsiella pneumoniae were studied. Microdilution assays and time-kill curves were performed to examine the activity of oxyclozanide in combination with colistin. The outer membrane protein (OMP) profile, membrane permeability and cell wall structure of Col-S and Col-R A. baumannii, P. aeruginosa and K. pneumoniae in the presence of oxyclozanide were assessed by SDS-PAGE, fluorescence microscopy and transmission electron microscopy, respectively. Oxyclozanide in combination with colistin increased the activity of colistin against Col-S and Col-R A. baumannii, P. aeruginosa and K. pneumoniae. Time-kill curves showed synergistic activity between oxyclozanide and colistin against these bacterial isolates. Moreover, Col-R A. baumannii, P. aeruginosa and K. pneumoniae in the presence of oxyclozanide presented greater permeability and disruption of their cell wall than Col-S strains, without modification of their OMP profile. These data suggest that combination of oxyclozanide and colistin may be a new alternative for the treatment of Col-R GNB infections.