Persistence of SARS-CoV-2 infection and viral intra- and inter-host evolution in COVID-19 hospitalized patients.

Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) persistence in COVID-19 patients could play a key role in the emergence of variants of concern. The rapid intra-host evolution of SARS-CoV-2 may result in an increased transmissibility, immune and therapeutic escape which could be a direct consequence of COVID-19 epidemic currents. In this context, a longitudinal retrospective study on eight consecutive COVID-19 patients with persistent SARS-CoV-2 infection, from January 2022 to March 2023, was conducted. To characterize the intra- and inter-host viral evolution, whole genome sequencing and phylogenetic analysis were performed on nasopharyngeal samples collected at different time points. Phylogenetic reconstruction revealed an accelerated SARS-CoV-2 intra-host evolution and emergence of antigenically divergent variants. The Bayesian inference and principal coordinate analysis analysis showed a host-based genomic structuring among antigenically divergent variants, that might reflect the positive effect of containment practices, within the critical hospital area. All longitudinal antigenically divergent isolates shared a wide range of amino acidic (aa) changes, particularly in the Spike (S) glycoprotein, that increased viral transmissibility (K417N, S477N, N501Y and Q498R), enhanced infectivity (R346T, S373P, R408S, T478K, Q498R, Y505H, D614G, H655Y, N679K and P681H), caused host immune escape (S371L, S375F, T376A, K417N, and K444T/R) and displayed partial or complete resistance to treatments (G339D, R346K/T, S371F/L, S375F, T376A, D405N, N440K, G446S, N460K, E484A, F486V, Q493R, G496S and Q498R). These results suggest that multiple novel variants which emerge in the patient during persistent infection, might spread to another individual and continue to evolve. A pro-active genomic surveillance of persistent SARS-CoV-2 infected patients is recommended to identify genetically divergent lineages before their diffusion.

Early initiation of combined therapy in severely immunocompromised patients with COVID-19: a retrospective cohort study.

This single-centre retrospective cohort study reports on the results of a descriptive (non-comparative) retrospective cohort study of early initiation of antivirals and combined monoclonal antibody therapy (mAbs) in 48 severely immunocompromised patients with COVID-19. The study assessed the outcomes and the duration of viral shedding. The patients started early combined therapy (ECT) a median of 2 days (interquartile range [IQR]: 1-3 days) after the diagnosis of SARS-CoV-2 infection. Except for 1 patient who died due COVID-19-related respiratory failure, patients had their first negative nasopharyngeal swab result after a median of 11 days (IQR: 6-17 days) after starting combined therapy. There were no reports of severe side effects. During a follow-up period of 512 days (interquartile range [IQR]: 413-575 days), 6 patients (12.5%) died and 16 (33.3%) were admitted to hospital. Moreover, 12 patients (25%) were diagnosed with SARS-CoV-2 reinfection a median of 245 days (IQR: 138-401 days) after starting combined treatment. No relapses were reported. Although there was no comparison group, these results compare favourably with the outcomes of severely immunocompromised patients with COVID-19 reported in the literature.

SARS CoV-2 spike protein-guided exosome isolation facilitates detection of potential miRNA biomarkers in COVID-19 infections.

OBJECTIVES: Nearly three years into the pandemic, SARS-CoV-2 infections are occurring in vaccinated and naturally infected populations. While humoral and cellular responses in COVID-19 are being characterized, novel immune biomarkers also being identified. Recently, an increase in angiotensin-converting enzyme 2 expressing (aka, ACE2 positive) circulating exosomes (ExoACE2) were identified in the plasma of COVID-19 patients (El-Shennawy et al.). In this pilot study, we describe a method to characterize the exosome-associated microRNA (exo-miRNA) signature in ACE2-positive and ACE2-negative exosomal populations (non-ExoACE2). METHODS: We performed a sorting protocol using the recombinant biotin-conjugated SARS CoV-2 spike protein containing the receptor binding domain (RBD) on plasma samples from six patients. Following purification, exo-miRNA were characterized for ACE2-positive and ACE2-negative exosome subpopulations by RT-PCR. RESULTS: We identified differential expression of several miRNA. Specifically let-7g-5p and hsa-miR-4454+miR-7975 were upregulated, while hsa-miR-208a-3p and has-miR-323-3p were downregulated in ExoACE2 vs. non-ExoACE2. CONCLUSIONS: The SARS CoV-2 spike-protein guided exosome isolation permits isolation of ExoACE2 exosomes. Such purification facilitates detailed characterization of potential biomarkers (e.g. exo-miRNA) for COVID-19 patients. This method could be used for future studies to further the understanding mechanisms of host response against SARS CoV-2.

Role of the T2Dx magnetic resonance assay in patients with suspected bloodstream infection: a single-centre real-world experience.

BACKGROUND: T2Dx was approved by the US Food and Drug Administration for the rapid detection of a modified panel of ESKAPE bacterial species or Candida spp. causing bloodstream infection (BSI). PATIENTS AND METHODS: We performed a retrospective, observational study from January 1, 2018 to December 31, 2019 of all hospitalised patients with suspected BSI who underwent assessment using T2Dx in addition to standard blood culture (BC). T2-positive patients (cases) were compared to a matched group of patients with BSI documented only by BC (1:2 ratio) to investigate the possible impact of T2Dx on the appropriateness of empirical antimicrobial therapy and 21-day mortality. RESULTS: In total, 78 T2Dx-analysed samples (49 patients) were analysed. The T2Dx assay result was positive for18 patients and negative for 31 patients. The concordance rates of the T2Bacteria Panel and T2Candida Panel results with those of standard BC were 74.4% and 91.4%, respectively. In the matched analysis, inappropriate empiric antimicrobial therapy administration was significantly less frequent in cases than in comparators (5.5% vs. 38.8%). The 21-day mortality rate was twofold lower in cases than in comparators (22.2% vs. 44.4%), although the difference was not significant. No other analysed variables were significantly different between the two groups. CONCLUSIONS: This study illustrated that T2Dx might be associated with an increase in the appropriateness of empiric antimicrobial therapy in patients with BSI. Further studies are needed to evaluate whether the T2Dx assay can improve patient outcomes.

Lessons learned and implications of early therapies for coronavirus disease in a territorial service centre in the Calabria region: a retrospective study.

BACKGROUND: Monoclonal antibodies (mAbs) and antivirals have been approved for early therapy of coronavirus disease (COVID-19), however, in the real-life setting, there are difficulties to prescribe these therapies within few days from symptom onset as recommended, and effectiveness of combined use of these drugs have been hypothesised in most-at-risk patients (such as those immunocompromised) but data supporting this strategy are limited. METHODS: We describe the real-life experience of SARS-CoV-2 antivirals and/or monoclonal antibodies (mAbs) and focus on the hospitalisation rate due to the progression of COVID-19. Clinical results obtained through our risk-stratification algorithm and benefits achieved through a strategic proximity territorial centre are provided. We also report a case series with an in-depth evaluation of SARS-CoV-2 genome in relationship with treatment strategy and clinical evolution of patients. RESULTS: Two hundred eighty-eight patients were analysed; 94/288 (32.6%) patients were treated with mAb monotherapy, 171/288 (59.4%) patients were treated with antivirals, and 23/288 (8%) patients received both mAbs and one antiviral drug. Haematological malignancies were more frequent in patients treated with combination therapy than in the other groups (p = 0.0003). There was a substantial increase in the number of treated patients since the opening of the centre dedicated to early therapies for COVID-19. The provided disease-management and treatment appeared to be effective since 98.6% patients recovered without hospital admission. Moreover, combination therapy with mAbs and antivirals seemed successful because all patients admitted to the hospital for COVID-19 did not receive such therapies, while none of the most-at-risk patients treated with combination therapy were hospitalized or reported adverse events. CONCLUSIONS: A low rate of COVID-19 progression requiring hospital admission was observed in patients included in this study. The dedicated COVID-19 proximity territorial service appeared to strengthen the regional sanitary system, avoiding the overwhelming of other services. Importantly, our results also support early combination therapy: it is possible that this strategy reduces the emergence of escape mutants of SARS-CoV-2, thereby increasing efficacy of early treatment, especially in immunocompromised individuals.

Characteristics, Management, and Outcomes of Elderly Patients with Diabetes in a Covid-19 Unit: Lessons Learned from a Pilot Study.

Background and objectives: Diabetes may affect in-hospital mortality of patients with Coronavirus disease 2019 (COVID-19). We have retrospectively evaluated clinical characteristics, diabetes management, and outcomes in a sample of COVID-19 patients with diabetes admitted to our hospital. Materials and Methods: All patients admitted to the Infectious Diseases Unit from 28 March 2020, to 16 June 2020, were enrolled. Clinical information and biochemical parameters were collected at the time of admission. Patients were ranked according to diabetes and death. Results: Sixty-one patients with COVID-19 were analyzed. Most of them were from a long-term health care facility. Mean age was 77 ± 16 years, and 19 had type 2 diabetes (T2D). Eighteen patients died, including 8 with T2D and 10 without T2D (p = 0.15). Patients with diabetes were significantly older, had a higher prevalence of cardiovascular diseases, and a significantly lower lymphocyte count. No significant relationship was found between diabetes and in-hospital mortality (Odds Ratio OR 2.3; Confidence Interval CI 0.73-7.38, p = 0.15). Patients with diabetes were treated with insulin titration algorithm. Severe hypoglycemic events, ketoacidosis and hyperosmolar hyperglycemias did not occur during hospitalization. Mean pre-meal capillary blood glucose was 157 ± 45 mg/dL, and the coefficient of variation of glycaemia was 29%. Conclusions: Our study, albeit limited by the small number of subjects, did not describe any significant association between T2D diabetes and mortality. Clinical characteristics of patients, and acceptable glucose control prior and during hospitalization may have influenced the result. The use of an insulin titration algorithm should be pursued during hospitalization.

Virological and Clinical Outcome of DAA Containing Regimens in a Cohort of Patients in Calabria Region (Southern Italy).

Background and objectives: In Italy, Hepatitis C Virus (HCV) infections are most prevalent in people older than 50 years of age, who often experience multi-morbidities, take co-medications, and have a long history of liver disease. These characteristics could potentially affect tolerability of HCV treatments and adherence in this subgroup. After achievement of sustained virological response (SVR), retention into care is very important both to detect the onset of possible complications and prevent further infections. In this study, SVR rates and retention into care of patients treated with directly acting antivirals (DAAs) of a single-center cohort in Southern Italy were evaluated. Materials and Methods: Patients treated with directly acting antivirals from 2014 to 2018 were included. Patients were stratified by age (i.e., <65 vs. ≥65 years) and by cirrhosis presence (i.e., liver stiffness >14.6 KPa or clinical/ultrasound cirrhosis vs. absence of these criteria). Primary outcome was availability of SVR at Weeks 12-24 after the end of treatment. Inter- and intra-group comparisons were performed along the follow-up for significant laboratory parameters. Results: In total, 212 patients were treated; 184 (87%) obtained SVR after the first treatment course and 4 patients after retreatment. Twenty-two (10.4%) patients were lost to follow-up before assessment of SVR, and two patients died before the end of treatment for liver decompensation. Considering only the first treatment episode, per protocol analysis (i.e., excluding patients lost to follow-up) showed the following rates of SVR: 97% (overall), 97% (older age group), 96% (age group <65 years), 94% (cirrhotics), and 100% (non-cirrhotics). By contrast, at the intention to treat analysis (i.e., patients lost were computed as failures), SVR percentages were significantly lower for patients <65 years of age (80%) and for non-cirrhotics (85%). Conclusions: High rates of SVR were obtained. However, younger patients and those without cirrhosis displayed an apparent high risk of being lost to follow-up. This may have important implications: since those who are lost may transmit HCV in case SVR is not achieved, these subpopulations should receive appropriate counselling during treatment.

Antibiotic Treatment of Carbapenem-Resistant Acinetobacter baumannii Infections in View of the Newly Developed ß-Lactams: A Narrative Review of the Existing Evidence.

It is estimated that antimicrobial resistance (AMR) is responsible for nearly 5 million human deaths worldwide each year and will reach 10 million by 2050. Carbapenem-resistant Acinetobacter baumannii (CRAB) infections represent the fourth-leading cause of death attributable to antimicrobial resistance globally, but a standardized therapy is still lacking. Among the antibiotics under consideration, Sulbactam/durlobactam seems to be the best candidate to replace current back-bone agents. Cefiderocol could play a pivotal role within combination therapy regimens. Due to toxicity and the pharmacokinetics/pharmacodynamics (PK/PD) limitations, colistin (or polymyxin B) should be used as an alternative agent (when no other options are available). Tigecycline (or minocycline) and fosfomycin could represent suitable partners for both NBLs. Randomized clinical trials (RCTs) are needed to better evaluate the role of NBLs in CRAB infection treatment and to compare the efficacy of tigecycline and fosfomycin as partner antibiotics. Synergism should be tested between NBLs and "old" drugs (rifampicin and trimethoprim/sulfamethoxazole). Huge efforts should be made to accelerate pre-clinical and clinical studies on safer polymyxin candidates with improved lung activity, as well as on the iv rifabutin formulation. In this narrative review, we focused the antibiotic treatment of CRAB infections in view of newly developed ß-lactam agents (NBLs).

Interleukin-62/lymphocyte as a proposed predictive index for COVID-19 patients treated with monoclonal antibodies.

In a convenience sample of 93 patients treated with monoclonal antibodies (moAbs) against SARS-CoV-2, the interleukin-62/lymphocyte count ratio (IL-62/LC) was able to predict clinical worsening both in early stages of COVID-19 and in oxygen-requiring patients. Moreover, we analysed 18 most at-risk patients with asymptomatic or mild disease treated with both moAbs and antiviral treatment and found that only 2 had clinical progression, while patients with a similar risk were reported to have an unfavourable outcome in most cases from recent data. In only one of our 18 patients, clinical progression was attributable to COVID-19, and in the other cases, clinical progression was observed despite IL-62/LC being above the risk cut-off. In conclusion, IL-62/LC may be a valuable method to identify patients requiring more aggressive treatments both in earlier and later stages of the disease; however, most at-risk patients can be protected from clinical worsening by combining moAbs and antivirals, even if levels of the IL-62/LC biomarker are lower than the risk cut-off.

Evolution of in-hospital patient characteristics and predictors of death in the COVID-19 pandemic across four waves: are they moving targets with implications for patient care?

Objectives: The aim of this work was to study characteristics, outcomes and predictors of all-cause death in inpatients with SARS-CoV-2 infection across the pandemic waves in one large teaching hospital in Italy to optimize disease management. Methods: All patients with SARS-CoV-2 infection admitted to our center from March 2020 to June 2022 were included in this retrospective observational cohort study. Both descriptive and regression tree analyses were applied to identify factors influencing all-cause mortality. Results: 527 patients were included in the study (65.3% with moderate and 34.7% with severe COVID-19). Significant evolutions of patient characteristics were found, and mortality increased in the last wave with respect to the third wave notwithstanding vaccination. Regression tree analysis showed that in-patients with severe COVID-19 had the greatest mortality across all waves, especially the older adults, while prognosis depended on the pandemic waves in patients with moderate COVID-19: during the first wave, dyspnea was the main predictor, while chronic kidney disease emerged as determinant factor afterwards. Conclusion: Patients with severe COVID-19, especially the older adults during all waves, as well as those with moderate COVID-19 and concomitant chronic kidney disease during the most recent waves require more attention for monitoring and care. Therefore, our study drives attention towards the importance of co-morbidities and their clinical impact in patients with COVID-19 admitted to hospital, indicating that the healthcare system should adapt to the evolving features of the epidemic.

Clinical Characteristics and Outcome of Hospitalized COVID-19 Patients Treated with Standard Dose of Dexamethasone or High Dose of Methylprednisolone.

The hyperinflammatory phase represents the main cause for the clinical worsening of acute respiratory distress syndrome (ARDS) in Coronavirus disease 2019 (COVID-19), leading to the hypothesis that steroid therapy could be a mainstream treatment in COVID-19 patients. This is an observational study including all consecutive patients admitted to two Italian University Hospitals for COVID-19 from March 2020 to December 2021. The aim of this study was to describe clinical characteristics and outcome parameters of hospitalized COVID-19 patients treated with dexamethasone 6 mg once daily (standard-dose group) or methylprednisolone 40 mg twice daily (high-dose group). The primary outcome was the impact of these different steroid treatments on 30-day mortality. During the study period, 990 patients were evaluated: 695 (70.2%) receiving standard dosage of dexamethasone and 295 (29.8%) receiving a high dose of methylprednisolone. Cox regression analysis showed that chronic obstructive pulmonary disease (HR 1.98, CI95% 1.34−9.81, p = 0.002), chronic kidney disease (HR 5.21, CI95% 1.48−22.23, p = 0.001), oncologic disease (HR 2.81, CI95% 1.45−19.8, p = 0.005) and high-flow nasal cannula, continuous positive airway pressure or non-invasive ventilation oxygen therapy (HR 61.1, CI95% 5.12−511.1, p < 0.001) were independently associated with 30-day mortality; conversely, high-dose steroid therapy was associated with survival (HR 0.42, CI95% 0.38−0.86, p = 0.002) at 30 days. Kaplan−Meier curves for 30-day survival displayed a statistically significant better survival rate in patients treated with high-dose steroid therapy (p = 0.018). The results of this study highlighted that the use of high-dose methylprednisolone, compared to dexamethasone 6 mg once daily, in hospitalized patients with COVID-19 may be associated with a significant reduction in mortality.

Prevalence of Antibiotic Resistance Over Time in a Third-Level University Hospital.

This study evaluated the spread and possible changes in resistance patterns of ESKAPE bacteria to first-choice antibiotics from 2015 to 2019 at a third-level university hospital after persuasive stewardship measures were implemented. Isolates were divided into three groups (group 1, low drug-resistant; group 2, multidrug/extremely drug-resistant; and group 3, pan-resistant bacteria) and a chi-squared test (χ2) was applied to determine differences in their distributions. Among the 2,521 isolates, Klebsiella pneumoniae was the most frequently detected (31.1%). From 2015 to 2019, the frequency of isolates in groups 2 and 3 decreased from 70.1% to 48.6% (χ2 = 63.439; p < 0.0001). Stratifying isolates by bacterial species, for K. pneumoniae, the frequency of PDR isolates decreased from 20% to 1.3% (χ2 = 15.885; p = 0.003). For Acinetobacter baumannii, a statistically significant decrease was found in groups 2 and 3: from 100% to 83.3% (χ2 = 27.721; p < 0.001). Also, for Pseudomonas aeruginosa and Enterobacter spp., the frequency of groups 2 and 3 decreased from 100% to 28.3% (χ2 = 225.287; p < 0.001) and from 75% to 48.7% (χ2 = 15.408; p = 0.003), respectively. These results indicate that a program consisting of persuasive stewardship measures, which were rolled out during the time frame of our study, may be useful to control drug-resistant bacteria in a hospital setting.

Clinical characteristics and predictors of mortality associated with COVID-19 in elderly patients from a long-term care facility.

Since December 2019, coronavirus disease 2019 (COVID-19) pandemic has spread from China all over the world and many COVID-19 outbreaks have been reported in long-term care facilities (LCTF). However, data on clinical characteristics and prognostic factors in such settings are scarce. We conducted a retrospective, observational cohort study to assess clinical characteristics and baseline predictors of mortality of COVID-19 patients hospitalized after an outbreak of SARS-CoV-2 infection in a LTCF. A total of 50 patients were included. Mean age was 80 years (SD, 12 years), and 24/50 (57.1%) patients were males. The overall in-hospital mortality rate was 32%. At Cox regression analysis, significant predictors of in-hospital mortality were: hypernatremia (HR 9.12), lymphocyte count < 1000 cells/µL (HR 7.45), cardiovascular diseases other than hypertension (HR 6.41), and higher levels of serum interleukin-6 (IL-6, pg/mL) (HR 1.005). Our study shows a high in-hospital mortality rate in a cohort of elderly patients with COVID-19 and hypernatremia, lymphopenia, CVD other than hypertension, and higher IL-6 serum levels were identified as independent predictors of in-hospital mortality. Given the small population size as major limitation of our study, further investigations are necessary to better understand and confirm our findings in elderly patients.