Maribavir treatment for resistant cytomegalovirus disseminated disease in kidney transplant recipients: A case-based scoping review of real life data in literature.

The treatment of refractory CMV is often associated with high toxicity. Maribavir (MBV) is a novel oral antiviral, known for its favourable safety profile in fragile patients. We describe a case of CMV disease with end organ damage following kidney transplantation at high risk, for recipient-donor serological mismatch. A 54-year-old female with history of obesity, hypertension, and chronic kidney disease, on prednisone and tacrolimus after kidney transplantation in November 2022, soon after developed primary CMV infection, treated with Valganciclovir and CMV Ig. In January 2023 the patient presented with fever and dyspnea. Pulmonary miliary opacities and right-upper lobe consolidation were found at CT-scan along with CMV-DNA positivity on BAL and serum. Lung biopsy confirmed CMV infection. Antiviral was switched to Ganciclovir. Despite initial benefit, fever and respiratory failure happened 8 days later, leading to intubation at day 15. Due to slow decrease serum CMV-DNA and detection of UL97 mutation, conferring resistance to valganciclovir and ganciclovir, the patient was started on foscarnet and letermovir. She was extubated after a gradual respiratory improvement and discharged from ICU to rehabilitation department with HFNC; reduction in serum CMV-DNA, but persistently elevated CMV-DNA on BAL were documented. At week 8, MBV was started and letermovir continued, for a 8 weeks course, without notable adverse effects. Respiratory function improved but soon after septic shock occurred. A bone marrow biopsy resulted in lymphoma, without indications for treatment: the patient developed coma and died 6 months after admission. MBV has recently been approved in Europe for treatment of R/R CMV in HSCT and SOT recipients. MBV showed superior rates of viraemia clearance after 8 weeks compared to SOC, demonstrating also a favourable safety profile with fewer patients discontinuing treatment and being affected by nephrotoxicity and neutropenia. Its main side effects are taste impairment, gastro-intestinal symptoms and asthenia. Based on actual promising perspectives regarding antiviral stewardship, more data are required to corroborate benefit of MBV in terms of toxicity and impact on mortality in highly fragile populations as SOT recipients. MBV received approval for the treatment of refractory or resistant CMV infections to other antiviral agents. Nevertheless, real-life data on efficacy and safety of MBV are still lacking. We conducted a narrative review of the current literature on MBV as treatment for CMV infection in kidney transplant recipients to understand clinical characteristics, safety and outcomes of MBV in this population. A search was run on the main scientific databases. 194 papers were identified, of which 188 were excluded by title and abstract evaluation. Subsequently, 6 papers were included. We performed descriptive statistics on the entire study population. The studies included in our analysis showed a higher prevalence of male subjects. The median age was 57 year. CKD was the most frequently reported comorbidity. Seven patients reported a donor/recipient mismatch (D+/R-). The case report and the cohort of patients collected from the literature show that MBV was used as an option in R/R CMV, notably for the presence or suspicion of CMV resistance to previous treatment. The clinical presentation of CMV in kidney SOT was heterogenous and varied from isolated reactivation of CMV-DNAemia, isolated fever or gastrointestinal involvement. For mild to moderate CMV disease, as with the cases reported in our review, or for proven ganciclovir, foscarnet or cidofovir resistance, MBV could be a valuable option. Outcomes of the patients treated with MBV were not reported in all the studies; however, where reported, 45.4% of the cases developed virological failure during MBV treatment with the development of specific resistance to MBV. MBV was generally well-tolerated, with low rates of toxicity, normally reversible. The introduction of new oral antivirals, such as MBV, could improve treatment, prophylaxis and preemptive treatment strategies, especially in anti-CMV treatment experienced patients.

Epidemiology and Impact of Anti-Pneumococcal Vaccination and COVID-19 on Resistance of Streptococcus pneumoniae Causing Invasive Disease in Piedmont, Italy.

BACKGROUND: The international surveillance of antimicrobial resistance (AMR) reports S. pneumoniae as one of leading causes of death associated with AMR. Against invasive disease, several vaccinations are available and a reduction in AMR in S. pneumoniae has been observed. Here, we evaluated the impact of anti-pneumococcal vaccination policy and the SARS-CoV2 outbreak on AMR in S. pneumoniae causing invasive disease. METHODS: We collected all strains of S. pneumoniae causing invasive disease from 2008 in the Piedmont region (Italy). Each strain was typed in order to identify the serogroup and data about AMR were collected. The population under surveillance was classified as infants, children, adults, and the old population. RESULTS: We collected n = 2076 S. pneumoniae strains, with 21.9% and 40.3% being resistant to penicillin G and erythromycin, respectively. We reported an increased risk of infection with penicillin-resistant S. pneumoniae among all populations and evaluated whether the infection was caused by a serotype included in the vaccine formulation. A similar increase was observed after the SARS-CoV2 outbreak. CONCLUSIONS: In the Piedmont region, subsequently to the introduction of anti-pneumococcal vaccination, a significant increase in the risk of penicillin G-resistant invasive pneumococcal disease among infants and old population was reported. No significant impact was found for the SARS-CoV2 outbreak.

QuantiFERON CMV Test and CMV Serostatus in Lung Transplant: Stratification Risk for Infection, Chronic and Acute Allograft Rejection.

The QuantiFERON CMV (QCMV) test evaluates specific adaptive immune system activity against CMV by measuring IFN-γ released by activated CD8+ T lymphocytes. We aimed to evaluate the QCMV test as a predictive tool for CMV manifestations and acute or chronic lung allograft rejection (AR and CLAD) in lung transplant (LTx) patients. A total of 73 patients were divided into four groups based on donor and recipient (D/R) serology for CMV and QCMV assay: group A low-risk for CMV infection and disease (D-/R-); group B and C at intermediate-risk (R+), group B with non-reactive QCMV and group C with reactive QCMV; group D at high-risk (D+/R-). Group D patients experienced higher viral replication; no differences were observed among R+ patients of groups B and C. D+/R- patients had a higher number of AR events and group C presented a lower incidence of AR. Prevalence of CLAD at 24 months was higher in group B with a higher risk of CLAD development (OR 6.33). The QCMV test allows us to identify R+ non-reactive QCMV population as the most exposed to onset of CLAD. This population had a higher, although non-significant, susceptibility to AR compared to the R+ population with reactive QCMV.

Molecular diagnosis of Cytomegalovirus infection: clinical performance of the Aptima transcription-mediated amplification assay toward conventional qPCR chemistry on whole blood samples.

Human Cytomegalovirus (HCMV) infection is life-threatening for immunocompromised patients. Quantitative molecular assays on whole blood or plasma are the gold standard for the diagnosis of invasive HCMV infection and for monitoring antiviral treatment in individuals at risk of HCMV disease. For these reasons, an accurate standardization toward the WHO 1st International Standard among different centers and diagnostic kits represents an effort for better clinical management of HCMV-positive patients. Herein, we evaluate, for the first time, the performance of a new transcription-mediated amplification (TMA) assay versus quantitative polymerase chain reaction (qPCR) chemistry, used as a routine method, on whole blood samples. A total of 755 clinical whole blood specimens were collected and tested simultaneously with TMA and qPCR assays. The data showed a qualitative agreement of 99.27% for positive quantified samples and 89.39% for those undetected between the two tested methods. Evaluation of viremia in positive samples highlighted a good correlation between TMA and qPCR chemistries in terms of International Units (ΔLog10 IU/mL: -0.29 ± 0.40). The TMA assay showed a significant correlation with qPCR in patients monitored for up to 3 months, thus allowing an accurate assessment of viremia in transplant patients. Therefore, TMA chemistry showed good agreement with qPCR testing, used as a current diagnostic routine. It also offers important advantages, such as FDA approval on plasma and In Vitro Diagnostic (IVD) on both plasma and whole blood, automated workflow with minimal hands-on time, and random access loading, thus enabling a rapid and reliable diagnostic in HCMV-infected patients. IMPORTANCE: In this paper, we describe the clinical performance of a novel transcription-mediated amplification (TMA) assay for the detection and quantification of human Cytomegalovirus (HCMV) DNA from whole blood samples. This is a pivotal analysis in immunocompromised patients [transplanted, HIV-positive, and Hematopoietic Stem Cell (HSC) recipients], and molecular tests with high sensitivity and specificity are necessary to evaluate the HCMV viral load in these patients. To our knowledge, this is the first in-depth evaluation of TMA chemistry for HCMV diagnosis on whole blood samples. Moreover, also technical aspects of this assay make it suitable for clinical diagnostics.

Herpes Virus Infection in Lung Transplantation: Diagnosis, Treatment and Prevention Strategies.

Lung transplantation is an ultimate treatment option for some end-stage lung diseases; due to the intense immunosuppression needed to reduce the risk of developing acute and chronic allograft failure, infectious complications are highly incident. Viral infections represent nearly 30% of all infectious complications, with herpes viruses playing an important role in the development of acute and chronic diseases. Among them, cytomegalovirus (CMV) is a major cause of morbidity and mortality, being associated with an increased risk of chronic lung allograft failure. Epstein-Barr virus (EBV) is associated with transformation of infected B cells with the development of post-transplantation lymphoproliferative disorders (PTLDs). Similarly, herpes simplex virus (HSV), varicella zoster virus and human herpesviruses 6 and 7 can also be responsible for acute manifestations in lung transplant patients. During these last years, new, highly sensitive and specific diagnostic tests have been developed, and preventive and prophylactic strategies have been studied aiming to reduce and prevent the incidence of these viral infections. In this narrative review, we explore epidemiology, diagnosis and treatment options for more frequent herpes virus infections in lung transplant patients.

Antiviral Approach to Cytomegalovirus Infection: An Overview of Conventional and Novel Strategies.

Human cytomegalovirus (HCMV) is a herpesvirus capable of establishing a lifelong persistence in the host through a chronic state of infection and remains an essential global concern due to its distinct life cycle, mutations, and latency. It represents a life-threatening pathogen for immunocompromised patients, such as solid organ transplanted patients, HIV-positive individuals, and hematopoietic stem cell recipients. Multiple antiviral approaches are currently available and administered in order to prevent or manage viral infections in the early stages. However, limitations due to side effects and the onset of antidrug resistance are a hurdle to their efficacy, especially for long-term therapies. Novel antiviral molecules, together with innovative approaches (e.g., genetic editing and RNA interference) are currently in study, with promising results performed in vitro and in vivo. Since HCMV is a virus able to establish latent infection, with a consequential risk of reactivation, infection management could benefit from preventive treatment for critical patients, such as immunocompromised individuals and seronegative pregnant women. This review will provide an overview of conventional antiviral clinical approaches and their mechanisms of action. Additionally, an overview of proposed and developing new molecules is provided, including nucleic-acid-based therapies and immune-mediated approaches.

Performance Evaluation of BD Phoenix and MicroScan WalkAway Plus for Determination of Fosfomycin Susceptibility in Enterobacterales.

BACKGROUND: Fosfomycin is an old bactericidal drug that has gained increasing interest in the last decade for its potential use in multi-drug resistant gram-negative infections. However, evidence on fosfomycin susceptibility testing reports a poor correlation between commercial methods vs. reference agar dilution (AD) for Enterobacterales (EB). The study aimed at assessing the performance of two automated systems for the determination of fosfomycin susceptibility in EB clinical isolates. METHODS: Fosfomycin susceptibility testing results of two collections of 100 non-duplicate clinical EB strains obtained using two different platforms (BD Phoenix and MicroScan WalkAway Plus) were compared with those obtained by AD. Categorical agreement (CA), major error (ME) and very major error (VME) rates were calculated. RESULTS: BD Phoenix exhibited a 6.9% rate of false-resistant results and achieved a CA of 69%, whereas MicroScan WalkAway Plus achieved 3.7% of false-resistant results and 72% of CA. Both automated systems showed poor detection of resistant isolates, with 49.1% and 56.2% of false-susceptible results for BD Phoenix and Microscan WalkAway Plus, respectively. CONCLUSIONS: Overall, agar dilution remains the most suitable method for routine laboratory antimicrobial susceptibility testing of fosfomycin on Enterobacterales strains, given the poor performance of automated systems. The application of both automated systems, in the clinical laboratories reporting of fosfomycin, should be reviewed in light of the accuracy results falling below the acceptable threshold.

Application of FT-IR Spectroscopy for Mycobacterium abscessus complex subspecies differentiation.

Mycobacterium abscessus complex (MABSC) subspecies differentiation improves patients' therapy and outcome. Fourier-Transform-Infrared Spectroscopy (FT-IRS) was applied for subspecies discrimination of 15 strains on different media: Löwenstein-Jensen showed the best resolution power; Linear Discriminant Analysis model differentiated M. abscessus susbsp. abscessus from M. abscessus subsp. massiliense. FT-IRS has a potential role in rapidly MABSC subspecies identification.

Extraction Bottleneck in the Diagnosis of SARS-CoV-2: Evaluation of an Alternative Protocol Derived from Veterinary Use.

The COVID-19 pandemic represented a challenge for health-care systems, and a major bottleneck in SARS-CoV-2 diagnosis was the unavailability of extraction reagents. To overcome this limitation, we performed a comparative analysis to evaluate the performance of an alternative extraction protocol derived from veterinary use adapted to an open robotic platform (Testing method). A total of 73 nasopharyngeal swabs collected for diagnosis of SARS-CoV-2 infection were simultaneously extracted with the Testing protocol and the laboratory Standard of Care in order to assess the performance of the first one. The Cohen's coefficient between both procedures was excellent (K Value = 0.955). Analysis of cycle threshold and linear regression showed a significant correlation between the two methods for each tested genetic target. Although validated for veterinary applications, the Testing method showed excellent performances in RNA extraction, with several advantages: lower sample input volume, the possibility to overcome the lack of deep-well plates and adaptability to robotic liquid handlers.

Seroprevalence of hepatitis E virus in liver transplant patients in Turin, Italy.

BACKGROUND: Acute E hepatitis is usually a self-limited non-progressive disease; however, acute liver failure and death can occur in the presence of conditions such as pregnancy and chronic liver diseases. In immunocompromised individuals, such as transplant patients, acute hepatitis E virus (HEV) infection may evolve to chronic hepatitis with rapid progression to liver decompensation. At our center, serology for HEV is not routinely performed in transplant patients and serological status is investigated only based on clinical judgement. METHODS: In this study, seroprevalence of HEV was evaluated in 217 patients (120 liver transplant recipients and 97 individuals diagnosed with acute or chronic hepatitis). Molecular evaluation of HEV-RNA was also performed. RESULTS: Thirteen patients (6%) showed positivity for HEV-IgG; in particular, 10/120 (8.3%), with concomitant presence of IgM and IgG in six and 3/97 (3.1%). None of the plasma samples tested by HEV-RNA was positive. CONCLUSIONS: As the detectable RNA window is narrow and an undetectable HEV-RNA result does not exclude recent infection and the transplant context per se represents a risk factor for chronic infection in patients infected with HEV, a routine diagnostic workflow including HEV should be taken into consideration, increasing awareness and knowledge of the basic and clinical aspects of the disease.

Comparison of Different Chemical and Mechanical Modalities for Implant Surface Decontamination: Activity against Biofilm and Influence on Cellular Regrowth-An In Vitro Study.

Objectives: The aim of this in vitro study was to compare the efficacy of chemical and mechanical methods for decontamination of titanium dental implant surfaces previously infected with polymicrobial biofilms in a model simulating a peri-implant defect. Furthermore, the effect of each decontamination protocol on MG-63 osteoblast-like cells morphology and adhesion to the treated implants was assessed. Background: Peri-implantitis is a growing issue in dentistry, and evidence about implant surface decontamination procedures is lacking and inconclusive. Methods: A total of 40 previously biofilm-contaminated implants were placed into a custom-made model simulating a peri-implant defect and randomly assigned to five treatment groups: (C) control (no treatment); (AW) air abrasion without any powder; (ESC) air abrasion with powder of erythritol, amorphous silica, and 0.3% chlorhexidine; (HBX) decontamination with a sulfonic/sulfuric acid solution in gel; and (HBX + ESC) a combination of HBX and ESC. Microbiological analysis was performed on five implants per treatment group, and the residual viable bacterial load measured in log 10 CFU/mL was counted for each bacterial strain and for the total number of colonies. The remaining three implants per group and three noncontaminated (NC) implants were used to assess surface biocompatibility using a scanning electron microscope and a backscattered electron microscope after seeding with MG-63 cells. Results: A significant decontaminant effect was achieved using HBX or HBX + ESC, while no differences were observed among other groups. The percentage of implant surface covered by adherent MG-63 cells was influenced by the treatment method. Progressive increases in covered surfaces were observed in groups C, AW, ESC, HBX, HBX + ESC, and NC. Conclusions: A combination of mechanical and chemical decontamination may provide more predictable results than mechanical cleaning alone.

In vitro antimicrobial activity of ozonated oil in liposome eyedrop against multidrug-resistant bacteria.

Bacteria are the most common causative agents of ocular infections. Treatment with topical broad-spectrum antibiotics is recommended in severe cases. However, antibiotic resistance has become a major concern in recent years, although antibiotics are generally effective in treating ocular infections. Antibacterial compound screening is performed to identify alternative therapeutic options to antibiotics. The aim of this study was to assess the in vitro antimicrobial activity of an ophthalmic solution containing ozonated oil. Strains of bacterial species with a multidrug resistance profile, which are responsible for a large proportion of ocular infections, were isolated and selected from different biological samples. The bacterial isolates were cultured, and ozonated oil was used to evaluate the inhibition zones at different time points. The treatment exhibited antibacterial activity against all the tested species. The effect was lower against the strains of Pseudomonas aeruginosa and more evident against Staphylococcus and Streptococcus spp. Our results suggest that the administration of ozonated oil may be a candidate agent to treat some infections of the ocular surface with a potential role in antimicrobial prophylaxis.

Can we trust in Sars-CoV-2 rapid antigen testing? Preliminary results from a paediatric cohort in the emergency department.

BACKGROUND: Rapid identification of Covid-19 in the paediatric emergency department is critical; Antigen tests are fast but poorly investigated in children. AIMS: To investigate Sars-CoV-2 antigen rapid test in children. METHODS: We compare the performance of LumiraDx with molecular tests in a paediatric emergency department. RESULTS: A retrospective cohort of 191 patients with AT and PCR tests performed in the same episode was analysed; 16% resulted positive for Sars-CoV-2. Using the PCR test as the gold standard, we calculated antigen testing overall sensitivity of 94.1%, specificity of 91.9%, and NPV of 99.4%. Only one false-negative test was found. CONCLUSIONS: AT may be helpful in the initial screening of patients at PED.

Clinical features of respiratory syncytial virus bronchiolitis in an infant: rapid and fatal brain involvement.

BACKGROUND: Respiratory Syncytial Virus (RSV) infection is a significant cause of bronchiolitis and pneumonia, mostly responsible for hospitalization and infant death worldwide. However, in recent years the importance of extrapulmonary RSV manifestations, especially at neurological level, have become evident. Seizures, lethargy, ataxia and status epilepticus are suggestive of brain involvement, but also in their absence a direct neurological damage RSV-related need to be evaluated. CASE PRESENTATION: A 40-day old male infant was admitted to the Emergency Department with severe bronchiolitis and dyspnea. The patient was reported to be coughing for a week with a vomiting episode in the previous two days. The nasopharyngeal swab confirmed the diagnosis of RSV infection and blood gas test showed hypoxemia and respiratory acidosis. For these reasons, the patient was provided with oxygen therapy. A few hours later, after an initial improvement in clinical parameters, a worsening of respiratory dynamics occurred and the patient was prepared for endotracheal intubation, but in the meantime death occurred. During all the observation period in the Emergency Room, no signs of neuropathological damage were evident. Post mortem examination showed lungs congestion with alveolar atelectasis and white matter degradation with severe edema at brain level. Microbiological analysis performed on autoptic samples confirmed the presence of RSV genome in tracheobronchial aspirate, meningeal swabs, pericardic and abdominal fluids, lung and brain biopsies. CONCLUSIONS: RSV is usually associated with respiratory diseases, however, as reported by an increasingly number of studies, the systemic dissemination of virus during severe disease can lead to a sudden infant death. The clinical picture herein reported showed a severe bronchiolitis resulting in a fatal and underestimated cerebral involvement due to RSV neurotropic behaviour and underline the need for clinicians to pay more attention to neurological involvement of RSV infection, even in absence of cerebral damage evidence.

Effectiveness of liposomal ozonized oil in reducing ocular microbial flora in patients undergoing cataract surgery.

PURPOSE: To evaluate the antimicrobial effectiveness of a liposomal ozonized oil solution used as a home therapy in patients undergoing cataract surgery. Antimicrobial efficacy was evaluated as the reduction in the bacterial load of the most common bacteria isolated from cases with endophthalmitis. SETTING: 20 Italian experimental centers of the Effectiveness of Liposomal Ozonized oil on Ocular Microbial flora before cataract surgery study group. DESIGN: Interventional, nonrandomized, paired-eye designed, phase 4 clinical study. METHODS: A total of 174 patients undergoing cataract surgery were divided into 2 groups: the study group (174 eyes) underwent surgery and received an isotonic ophthalmic solution of 0.5% ozonized oil in liposomes plus hypromellose treatment (2 drops 4 times/d), and the control group (174 contralateral eyes) was treated with saline solution. The treatment lasted for 3 days. Subconjunctival swabs were taken from both eyes of each patient at T0 (the day before starting the treatment and 4 days preoperatively) and at T4 (after 3 days of treatment and 10 min preoperatively) and sent to the laboratory within 24 hours of collection for microbiological analysis. RESULTS: 30% of 696 swabs taken at T0 were sterile. Contaminated swabs had a high prevalence of coagulase-negative staphylococci, including Staphylococcus epidermidis, and more than 60 different bacterial species were isolated. A significant reduction in microbial load was observed after treatment (>90% of the samples). The microbial load in the control group remained unchanged. CONCLUSIONS: Liposomal ozonized oil reduced the microbial burden after topical administration in a large study population.

Carbapenemase detection testing in the era of ceftazidime/avibactam-resistant KPC-producing Enterobacterales: A 2-year experience.

OBJECTIVES: The aim of this study was to investigate the prevalence of ceftazidime/avibactam (CZA) resistance among carbapenemase-producing Enterobacterales (CPE) blood culture isolates as well as the performance of the main carbapenemase phenotypic detection methods to identify KPC variants associated with CZA resistance. METHODS: Non-duplicate CPE strains isolated from blood cultures during 2018-2020 were tested for antimicrobial susceptibility. Molecular testing was used to identify carbapenemase-producers. Strains harbouring blaKPC and with a CZA minimum inhibitory concentration (MIC) ≥8 mg/L were investigated by sequencing. Subsequentially, five phenotypic carbapenemase detection methods were evaluated on these strains, namely the modified carbapenem inactivation method (mCIM), Rapidec® Carba NP, the disk diffusion synergy test, NG-Test CARBA® 5 and RESIST-5 O.O.K.N.V. RESULTS: Overall, the CZA resistance rate was high (13.7%) and remained relevant (5.9%) excluding metallo-ß-lactamases-producers. All isolates harbouringblaKPC mutants (n = 8) were associated with reduced carbapenem MICs and negative results by all detection methods based on revelation of enzyme activity. Lateral flow immunoassays failed to detect KPC-31 (n = 4) and KPC-33 (n = 2) but correctly identified KPC-14 (n = 2). Conversely, isolates harbouring wild-type KPC genes (n = 3) were associated with high-level CZA resistance and carbapenem resistance and tested positive by all of the evaluated methods. CONCLUSION: In the era of CZA-based therapies, molecular blaKPC identification followed by a carbapenem hydrolysis-based phenotypic assay could be the most reasonable diagnostic algorithm to detect all KPC-producers and to identify mutants associated with impaired carbapenemase activity and CZA resistance.

Detection of Carbapenemase and CTX-M Encoding Genes Directly from Bronchoalveolar Lavage Using the CRE and ESBL ELITe MGB Assays: Toward Early and Optimal Antibiotic Therapy Management of Critically Ill Patients with Pneumonia.

The detection of carbapenemase extended-spectrum ß-lactamase (ESBL)-producing Enterobacterales (EB) has become a major issue among critically ill patients, especially due to their impact on appropriate antimicrobial therapy. This study aimed at evaluating the potential contribution of molecular assays to early optimization of empirical antibiotic therapy among critically ill patients with carbapenemase- and/or CTX-M-producing EB pneumonia. The CRE and ESBL ELITe MGB® assays were evaluated directly on 197 bronchoalveolar lavage (BAL) samples obtained from 120 patients. Molecular results were then compared to routine culture-based diagnostic results, and a retrospective analysis of the therapeutic antimicrobial management was performed. Among the 197 clinical specimens, blaKPC-like and blaCTX-M-like were detected in 20 (10.2%) and 12 (6.1%) specimens belonging to 15 and 11 patients, respectively. Positive predictive value (PPV) and negative predictive value (NPV) of the CRE ELITe MGB Kit were 85% [95% confidence interval [CI]: 64.9-94.6] and 100%, respectively. PPV and NPV of the ESBL ELITe MGB Kit were 75% [95% CI: 49.4-90.2] and 100%, respectively. Retrospective analysis of the therapeutic antimicrobial management at the time of BAL collection showed that in ∼50% of patients with carbapenemase- and CTX-M-producing EB pneumonia empirical antibiotic therapy could have been optimized at least 48-72 hr earlier if positive molecular data had been used. The CRE and ESBL ELITe MGB assays might be an interesting tool for expediting optimization of empirical antibiotic therapy in critically ill patients with pneumonia, depending on local epidemiology of antibiotic resistance, patient risk stratification for EB infection, and availability of an antimicrobial stewardship team.

Direct Ethylenediaminetetraaceticacid-Modified ß-Lactam Inactivation Method: An Improved Method to Identify Serine-Carbapenemase-, Metallo-ß-Lactamase-, and Extended-Spectrum-ß-Lactamase-Producing Enterobacterales Directly from Positive Blood Culture.

Rapid identification of extended-spectrum-ß-lactamase (ESBL)-, serine carbapenemase-, and metallo-ß-lactamase (MBL)-producing Enterobacterales directly from positive blood culture (BC) bottles is of paramount importance to early optimize antimicrobial management and infection control measures. In this study, we describe and evaluate an improved variant of direct ß-lactam inactivation method, named direct ethylenediaminetetraaceticacid-modified-ß-lactam inactivation method (deBLIM). The deBLIM test is designed to detect ESBL or carbapenemase activity and to differentiate serine-carbapenemases from MBLs directly from Enterobacterale-positive BCs. The deBLIM was evaluated on both aerobic and anaerobic BCs spiked with 167 characterized Enterobacterale isolates. The deBLIM showed 100% sensitivity in detecting KPC and OXA-48-like serine carbapenemase, CTX-M, SHV variants, and TEM-10 ESBLs both in aerobic and anaerobic conditions. In contrast, a significant discrepancy between aerobic and anaerobic BCs was observed in detecting MBLs. The sensitivity rate in aerobic BCs was of 100% for all metalloenzyme types, whereas only 56.1% and 80% of VIM and NDM producers were detected in anaerobic bottles, respectively. IMP-producing Escherichia coli NCTC 13476 was also not detected in the anaerobic BC. No false positive result was observed among ESBL producers and broad-spectrum-ß-lactamase nonproducers, demonstrating an overall specificity of 100%. The deBLIM could be a cost-effective screening method for the identification of ESBLs, serine carbapenemases, and MBLs directly from Enterobacterale-positive BCs on the same day of bottle positivity detection. Nevertheless, it must be considered its poor performance in detecting MBLs in the anaerobic condition.

A new ophthalmic formulation containing antiseptics and dexpanthenol: In vitro antimicrobial activity and effects on corneal and conjunctival epithelial cells.

Antibiotic resistance is increasing even in ocular pathogens, therefore the interest towards antiseptics in Ophthalmology is growing. The aim of this study was to analyze the in vitro antimicrobial efficacy and the in vitro effects of an ophthalmic formulation containing hexamidine diisethionate 0.05%, polyhexamethylene biguanide (PHMB) 0.0001% disodium edetate (EDTA) 0.01%, dexpanthenol 5% and polyvinyl alcohol 1.25% (Keratosept, Bruschettini, Genova, Italy) on cultured human corneal and conjunctival cells. The in vitro antimicrobial activity was tested on Staphylococcus aureus, Methicillin-Resistant Staphylococcus aureus (MRSA), Pseudomonas aeruginosa, Streptococcus pneumoniae, Streptococcus pyogenes and Streptococcus mitis. For each microbial strain 10 µL of a 0.5 MacFarland standardized bacterial inoculum were incubated at 25 °C with 100 µL of ophthalmic solution for up to 6 h. After different periods of time, samples were inoculated on blood agar with 5% sheep blood. Moreover, a 0.5 MacFarland bacterial inoculum was seeded in triplicate on Mueller-Hinton Agar or on Mueller-Hinton Fastidious Agar; then a cellulose disc soaked with 50 µL of ophthalmic solution was applied on the surface of agar and plates were incubated for 18 h at 37 °C, in order to evaluate the inhibition of bacterial growth around the disc. Human corneal and conjunctival epithelial cells in vitro were incubated for 5, 10 and 15 min with Keratosept or its components. The cytotoxicity was assessed through the release of cytoplasmic enzyme lactate dehydrogenase (LDH) into the medium immediately after exposure to the drugs; the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay was performed to evaluate the metabolic cell activity. Our results show that Keratosept ophthalmic solution gave an average logarithmic (log) reduction of bacterial load of 2.14 ± 0.35 within 6 h of exposure (p-value < 0.05 versus control saline solution). On agar plates, all microbial strains, excluding P. Aeruginosa, showed an inhibition zone of growth around the Keratosept-soaked discs. Keratosept and its components after 5 and 10 min did not show any cytotoxic effect on cultured corneal and conjunctival cells, and only after 15 min a significant reduction of cell viability and an increase of cytotoxicity compared to control (vehicle) was seen; dexpanthenol 5% and polyvinyl alcohol accelerated the wounding of corneal cells in vitro. In conclusion, Keratosept showed good antimicrobial activity on the tested strains; the ophthalmic solution and its components were safe and non-toxic for the corneal and conjunctival epithelial cells for 5 and 10 min at the concentrations analyzed, and dexpanthenol 5% and polyvinyl alcohol promoted the wounding of corneal cells.