Bacterial infections in critically ill patients with SARS-2-COVID-19 infection: results of a prospective observational multicenter study.

PURPOSE: To investigate the prevalence, incidence and characteristics of bacterial infections and their impact on outcome in critically ill patients infected with COVID-19. METHODS: We conducted a prospective observational study in eight Italian ICUs from February to May 2020; data were collected through an interactive electronic database. Kaplan-Meier analysis (limit product method) was used to identify the occurrence of infections and risk of acquisition. RESULTS: During the study period 248 patients were recruited in the eight participating ICUs. Ninety (36.3%) patients developed at least one episode of secondary infection. An ICU length of stay between 7 and 14 days was characterized by a higher occurrence of infectious complications, with ventilator-associated pneumonia being the most frequent. At least one course of antibiotic therapy was given to 161 (64.9%) patients. Overall ICU and hospital mortality were 33.9% and 42.9%, respectively. Patients developing bacteremia had a higher risk of ICU mortality [45.9% vs. 31.6%, odds ratio 1.8 (95% CI 0.9-3.7), p = 0.069] and hospital mortality [56.8% vs. 40.3%, odds ratio 1.9 (95% CI 1.1-3.9), p = 0.04]. CONCLUSION: In critically ill patients infected with COVID-19 the incidence of bacterial infections is high and associated with worse outcomes. Regular microbiological surveillance and strict infection control measures are mandated.

Tracheostomy in critically ill patients with SARS 2 COVID-19 infection: a prospective observational multi-center study of short- and long-term outcomes.

Background: We report the characteristics, timing, and factors related to the decision to perform a tracheostomy in patients with confirmed COVID-19 infection admitted to eight Italian intensive care units (ICUs). Materials and methods: Prospective observational cohort study of patients with COVID-19 disease on mechanical ventilation. Long-term functional impairment (up to 180 days' post-hospital discharge) was assessed using the Karnofsky scale. Kaplan-Meier analysis assessed differences in survival and freedom from tracheostomy in relation to ICU stay. Cox regression model was used to assess which variables impacted on tracheostomy as a categorical outcome. Results: A total of 248 patients were recruited in the eight participating ICUs. Patients undergoing tracheostomy (n = 128) had longer ICU (25 (18-36) vs. 10 (7-16), P = 0.001) and hospital (37 (26.5-50) vs. 19 (8.5-34.5) P = 0.02) stays. ICU and hospital mortality of patients tracheostomized was 34% and 37%, respectively. Cumulative survival Kaplan-Meier analysis documented improved survival rates in patients undergoing tracheostomy (Log-Rank, Mantel-Cox = 4.8, P = 0.028). Median Karnofsky scale values improved over time but were similar between survivors receiving or not receiving tracheostomy. No healthcare worker involved in the tracheostomy procedure developed COVID-19 infection during the study period. Conclusions: Patients with COVID-19 infection who underwent tracheostomy had a better cumulative survival but similar long-term functional outcomes at 30, 60, and 180 days after hospital discharge.

Extending rnaSPAdes functionality for hybrid transcriptome assembly.

BACKGROUND: De novo RNA-Seq assembly is a powerful method for analysing transcriptomes when the reference genome is not available or poorly annotated. However, due to the short length of Illumina reads it is usually impossible to reconstruct complete sequences of complex genes and alternative isoforms. Recently emerged possibility to generate long RNA reads, such as PacBio and Oxford Nanopores, may dramatically improve the assembly quality, and thus the consecutive analysis. While reference-based tools for analysing long RNA reads were recently developed, there is no established pipeline for de novo assembly of such data. RESULTS: In this work we present a novel method that allows to perform high-quality de novo transcriptome assemblies by combining accuracy and reliability of short reads with exon structure information carried out from long error-prone reads. The algorithm is designed by incorporating existing hybridSPAdes approach into rnaSPAdes pipeline and adapting it for transcriptomic data. CONCLUSION: To evaluate the benefit of using long RNA reads we selected several datasets containing both Illumina and Iso-seq or Oxford Nanopore Technologies (ONT) reads. Using an existing quality assessment software, we show that hybrid assemblies performed with rnaSPAdes contain more full-length genes and alternative isoforms comparing to the case when only short-read data is used.

Hybrid transcriptome sequencing approach improved assembly and gene annotation in Cynara cardunculus (L.).

BACKGROUND: The investigation of transcriptome profiles using short reads in non-model organisms, which lack of well-annotated genomes, is limited by partial gene reconstruction and isoform detection. In contrast, long-reads sequencing techniques revealed their potential to generate complete transcript assemblies even when a reference genome is lacking. Cynara cardunculus var. altilis (DC) (cultivated cardoon) is a perennial hardy crop adapted to dry environments with many industrial and nutraceutical applications due to the richness of secondary metabolites mostly produced in flower heads. The investigation of this species benefited from the recent release of a draft genome, but the transcriptome profile during the capitula formation still remains unexplored. In the present study we show a transcriptome analysis of vegetative and inflorescence organs of cultivated cardoon through a novel hybrid RNA-seq assembly approach utilizing both long and short RNA-seq reads. RESULTS: The inclusion of a single Nanopore flow-cell output in a hybrid sequencing approach determined an increase of 15% complete assembled genes and 18% transcript isoforms respect to short reads alone. Among 25,463 assembled unigenes, we identified 578 new genes and updated 13,039 gene models, 11,169 of which were alternatively spliced isoforms. During capitulum development, 3424 genes were differentially expressed and approximately two-thirds were identified as transcription factors including bHLH, MYB, NAC, C2H2 and MADS-box which were highly expressed especially after capitulum opening. We also show the expression dynamics of key genes involved in the production of valuable secondary metabolites of which capitulum is rich such as phenylpropanoids, flavonoids and sesquiterpene lactones. Most of their biosynthetic genes were strongly transcribed in the flower heads with alternative isoforms exhibiting differentially expression levels across the tissues. CONCLUSIONS: This novel hybrid sequencing approach allowed to improve the transcriptome assembly, to update more than half of annotated genes and to identify many novel genes and different alternatively spliced isoforms. This study provides new insights on the flowering cycle in an Asteraceae plant, a valuable resource for plant biology and breeding in Cynara and an effective method for improving gene annotation.

A dynamic causal modeling of the second outbreak of COVID-19 in Italy.

While the vaccination campaign against COVID-19 is having its positive impact, we retrospectively analyze the causal impact of some decisions made by the Italian government on the second outbreak of the SARS-CoV-2 pandemic in Italy, when no vaccine was available. First, we analyze the causal impact of reopenings after the first lockdown in 2020. In addition, we also analyze the impact of reopening schools in September 2020. Our results provide an unprecedented opportunity to evaluate the causal relationship between the relaxation of restrictions and the transmission in the community of a highly contagious respiratory virus that causes severe illness in the absence of prophylactic vaccination programs. We present a purely data-analytic approach based on a Bayesian methodology and discuss possible interpretations of the results obtained and implications for policy makers.

Treatment with ß-blockers and incidence of post-traumatic stress disorder after cardiac surgery: a prospective observational study.

OBJECTIVE: The aim was to investigate perioperative factors associated with the development of post-traumatic stress disorder (PTSD) in patients who underwent cardiac surgery. DESIGN: Prospective observational study. SETTING: Single academic center. PARTICIPANTS: One hundred twenty-eight consecutive patients scheduled for elective cardiac surgery with cardiopulmonary bypass. INTERVENTIONS: Patients were interviewed within the surgical unit 1 to 3 days before cardiac surgery. MEASUREMENTS AND MAIN RESULTS: Six months after surgery, participants were mailed the modified version of the Posttraumatic Stress Symptom Inventory 10. Of the 71 patients who completed the questionnaire and mailed it back at follow-up, 14 (19.7%) received a diagnosis of PTSD. Seven of 13 female patients who were not treated with ß-blockers received a diagnosis of PTSD compared with 0 of 12 who were treated with ß-blockers (p = 0.005, Fisher exact test). In a general linear model, including sex and ß-blocker treatment as predictors, the Posttraumatic Stress Symptom Inventory 10 score was significantly predicted by ß-blockade (F = 4.74, p = 0.033), with a significant interaction between sex and ß-blockade (F = 9.72, p = 0.003). CONCLUSIONS: These findings suggest that the use of ß-blockers might be protective against the development of PTSD in women after cardiac surgery.

A performance evaluation of despiking algorithms for eddy covariance data.

Spike detection for raw high-frequency eddy covariance time series is a challenging task because of the confounding effect caused by complex dynamics and the high level of noise affecting such data. To cope with these features, a new despiking procedure rooted on robust functionals is proposed. By processing simulated data, it is demonstrated that the proposed procedure performs better than the existing algorithms and can be therefore considered as a candidate for the implementation in data center environmental monitoring systems, where the availability of automatic procedures ensuring a high quality standard of released products constitutes an essential prerequisite.

Levosimendan to facilitate weaning from cardiorespiratory support in critically ill patients: current evidence and future directions.

Appropriate weaning is of crucial importance for critically ill patients requiring respiratory support. However, a remarkable proportion of them are difficult to wean. Levosimendan is a positive inotropic agent characterized by vasodilatory properties, which is used for the treatment of acute decompensated heart failure or in patients needing inotropic treatment, including cardiogenic shock, septic shock, pulmonary hypertension and right ventricular dysfunction, needed for hemodynamic support in patients with diuretic resistance, and weaning either from ventilator or from extracorporeal membrane oxygenation. This position paper will discuss the use of levosimendan in facilitating weaning from cardiorespiratory support in critically ill patients, according to available evidence and the personal experience of a group of Italian Experts.

A plasmid-borne blaOXA-58 gene confers imipenem resistance to Acinetobacter baumannii isolates from a Lebanese hospital.

We investigated the basis of the carbapenem resistance of 17 multidrug-resistant Acinetobacter baumannii clinical isolates collected from 2004 to 2005 at the Saint George University Hospital in Beirut, Lebanon. A. baumannii isolates were clonally related and were susceptible to colistin and trimethoprim-sulfamethoxazole, susceptible or intermediate to ampicillin-sulbactam and meropenem, and resistant to all other antimicrobials. Conjugation experiments demonstrated that resistance to imipenem could be transferred along with a plasmid containing the carbapenem-hydrolyzing oxacillinase bla(OXA-58) gene. The plasmid that we called pABIR was 29,823 bp in size and showed a novel mosaic structure composed of two origins of replication, four insertion sequence (IS) elements, and 28 open reading frames. The bla(OXA-58) gene was flanked by IS18 and ISAba3 elements at the 5' and 3' ends, respectively. The production of the carbapenem-hydrolyzing oxacillinase OXA-58 was apparently the only mechanism for carbapenem resistance in A. baumannii isolates causing the outbreak at the Lebanese Hospital.

Multiorgan failure is an adaptive, endocrine-mediated, metabolic response to overwhelming systemic inflammation.

Sepsis and other critical illnesses produce a biphasic inflammatory, immune, hormonal, and metabolic response. The acute phase is marked by an abrupt rise in the secretion of so-called stress hormones with an associated increase in mitochondrial and metabolic activity. The combination of severe inflammation and secondary changes in endocrine profile diminish energy production, metabolic rate, and normal cellular processes, leading to multiple organ dysfunction. This perceived failure of organs might instead be a potentially protective mechanism, because reduced cellular metabolism could increase the chances of survival of cells, and thus organs, in the face of an overwhelming insult. We propose that, first, multiple organ failure induced by critical illness is primarily a functional, rather than structural, abnormality. Indeed, it may not be failure as such, but a potentially protective, reactive mechanism. Second, the decline in organ function is triggered by a decrease in mitochondrial activity and oxidative phosphorylation, leading to reduced cellular metabolism. Third, this effect on mitochondria might be the consequence of acute-phase changes in hormones and inflammatory mediators.

Ion Torrent next-generation sequencing for routine identification of clinically relevant mutations in colorectal cancer patients.

AIMS: To evaluate the accuracy, consumable cost and time around testing (TAT) of a next-generation sequencing (NGS) assay, the Ion Torrent AmpliSeq Colon and Lung Cancer Panel, as an alternative to Sanger sequencing to genotype KRAS, NRAS and BRAF in colorectal cancer patients. METHODS: The Ion Torrent panel was first verified on cell lines and on control samples and then prospectively applied to routine specimens (n=114), with Sanger sequencing as reference. RESULTS: The Ion Torrent panel detected mutant alleles at the 5% level on cell lines and correctly classified all control tissues. The Ion Torrent assay was successfully carried out on most (95.6%) routine diagnostic samples. Of these, 12 (11%) harboured mutations in the BRAF gene and 47 (43%) in either of the two RAS genes, in two cases with a low abundance of RAS mutant allele which was missed by Sanger sequencing. The mean TAT, from sample receipt to reporting, was 10.4 (Sanger) and 13.0 (Ion Torrent) working days. The consumable cost for genotyping KRAS, NRAS and BRAF was €196 (Sanger) and €187 (Ion Torrent). CONCLUSIONS: Ion Torrent AmpliSeq Colon and Lung Cancer Panel sequencing is as robust as Sanger sequencing in routine diagnostics to select patients for anti-epidermal growth factor receptor (EGFR) therapy for metastatic colorectal cancer.

Ivabradine: potential clinical applications in critically ill patients.

It has been extensively demonstrated that an elevated heart rate is a modifiable, independent risk factor for cardiovascular events. A high heart rate increases myocardial oxygen consumption and reduces diastolic perfusion time. It can also increase ventricular diastolic pressures and induce ventricular arrhythmias. Critical care patients are prone to develop a stress induced cardiac impairment and consequently an increase in sympathetic tone. This in turn increases heart rate. In this setting, however, heart rate lowering might be difficult because the effects of inotropic drugs could be hindered by heart rate reducing drugs like beta-blockers. Ivabradine is a new selective antagonist of funny channels. It lowers heart rate, reducing the diastolic depolarization slope. Moreover, ivabradine is not active on sympathetic pathways, thus avoiding any interference with inotropic amines. We reviewed the literature available regarding heart rate control in critical care patients, focusing our interest on the use of ivabradine to assess the potential benefits of the drug in this particular setting.

Mitochondrial diabetes in children: seek and you will find it.

Maternally Inherited Diabetes and Deafness (MIDD) is a rare form of diabetes due to defects in mitochondrial DNA (mtDNA). 3243 A>G is the mutation most frequently associated with this condition, but other mtDNA variants have been linked with a diabetic phenotype suggestive of MIDD. From 1989 to 2009, we clinically diagnosed mitochondrial diabetes in 11 diabetic children. Diagnosis was based on the presence of one or more of the following criteria: 1) maculopathy; 2) hearing impairment; 3) maternal heritability of diabetes/impaired fasting glucose and/or hearing impairment and/or maculopathy in three consecutive generations (or in two generations if 2 or 3 members of a family were affected). We sequenced the mtDNA in the 11 probands, in their mothers and in 80 controls. We identified 33 diabetes-suspected mutations, 1/33 was 3243A>G. Most patients (91%) and their mothers had mutations in complex I and/or IV of the respiratory chain. We measured the activity of these two enzymes and found that they were less active in mutated patients and their mothers than in the healthy control pool. The prevalence of hearing loss (36% vs 75-98%) and macular dystrophy (54% vs 86%) was lower in our mitochondrial diabetic adolescents than reported in adults. Moreover, we found a hitherto unknown association between mitochondrial diabetes and celiac disease. In conclusion, mitochondrial diabetes should be considered a complex syndrome with several phenotypic variants. Moreover, deafness is not an essential component of the disease in children. The whole mtDNA should be screened because the 3243A>G variant is not as frequent in children as in adults. In fact, 91% of our patients were mutated in the complex I and/or IV genes. The enzymatic assay may be a useful tool with which to confirm the pathogenic significance of detected variants.

Recombinant activated factor VII for refractory bleeding after acute aortic dissection surgery: a propensity score analysis.

OBJECTIVE: The aim of this study was to assess safety and effectiveness of recombinant activated factor VII (rFVIIa) in patients with refractory bleeding undergoing acute aortic dissection surgery with deep hypothermic circulatory arrest. DESIGN: Propensity score-matched analysis. SETTING: University hospital. PATIENTS: Twenty-three cardiac surgery patients receiving rFVIIa compared with 23 matched controls. INTERVENTIONS: An intravenous bolus of rFVIIa (70 microg/kg) was administered at the end of a complete transfusion protocol. Five patients received rFVIIa in the operating room, and 18 patients received rFVIIa in the intensive care unit. Four of the intensive care unit patients required a second dose. MEASUREMENTS AND MAIN RESULTS: Blood loss and transfusion requirements were significantly reduced in the period after rFVIIa administration. A highly significant reduction in hourly blood loss was found at -1 hr vs. 0 hrs and 0 hrs vs. 1 hr (-194 and -77.5 mL, respectively; both adjusted p < .001). In addition, significant improvements of international normalized ratio (p < .001), partial thromboplastin time (p < .001), platelet count (p < .001), fibrinogen (p < .001), and antithrombin (p < .001) were detected after rFVIIa administration. The two groups did not differ regarding adverse events. CONCLUSIONS: rFVIIa was successfully used as an additional therapy both during and after acute aortic dissection surgery with deep hypothermic circulatory arrest, when bleeding was refractory to conventional methods. Randomized studies are necessary to confirm the safety and efficacy of rFVIIa in this setting.

Comparision of the inotropic effects of levosimendan, rolipram, and dobutamine on human atrial trabeculae.

The aim of this study was to compare the positive inotropic effects of 3 different agents with 3 different mechanisms of actions-levosimendan, rolipram, and dobutamine-on human atrial trabecular muscles. Samples of right atrial appendage (1 cm, 500-1000 mg) were removed and immersed in preoxygenated and modified Tyrode solution. In oxygenated Tyrode solution, preparations were used to investigate the concentration-effect relationship of levosimendan, dobutamine, and rolipram on percentage developed tension (DT), from 10 to 10 M, each concentration for 15 minutes. All 3 agents produced concentration-dependent increments in DT. We found that levosimendan was the most efficacious positive inotropic agent on isolated human atrial trabeculae. Both the sensitivity (pD2) and maximum response (Emax) of human atrial trabeculae to levosimendan (6.711 +/- 0.26 and 23.2 +/- 2.2 mN, respectively) were significantly greater than those of dobutamine (6.663 +/- 0.19 and 17.6 +/- 2.8 mN) and rolipram (6.497 +/- 0.18 and 15.0 +/- 1.0 mN). pD2 and Emax values for dobutamine were significantly higher than those for rolipram. It was suggested that because of its potential to enhance cardiac performance without predisposition to calcium-induced arrhythmias, levosimendan might be more useful as a positive inotropic agent in clinical practice.