Melatonin reduces the need for sedation in ICU patients: a randomized controlled trial.

BACKGROUND: Critically ill patients suffer from physiological sleep deprivation and have reduced blood melatonin levels. This study was designed to determine whether nocturnal melatonin supplementation would reduce the need for sedation in patients with critical illness. METHODS: A single-center, double-blind randomized placebo-controlled trial was carried out from July 2007 to December 2009, in a mixed medical-surgical Intensive Care Unit of a University hospital, without any form of external funding. Of 1158 patients admitted to ICU and treated with conscious enteral sedation, 82 critically-ill with mechanical ventilation >48 hours and Simplified Acute Physiology Score II>32 points were randomized 1:1 to receive, at eight p.m. and midnight, melatonin (3+3mg) or placebo, from the third ICU day until ICU discharge. Primary outcome was total amount of enteral hydroxyzine administered. RESULTS: Melatonin treated patients received lower amount of enteral hydroxyzine. Other neurological indicators (amount of some neuroactive drugs, pain, agitation, anxiety, sleep observed by nurses, need for restraints, need for extra sedation, nurse evaluation of sedation adequacy) seemed improved, with reduced cost for neuroactive drugs. Post-traumatic stress disorder prevalence did not differ between groups, nor did ICU or hospital mortality. Study limitations include the differences between groups before intervention, the small sample size, and the single-center observation. CONCLUSION: Long-term enteral melatonin supplementation may result in a decreased need for sedation, with improved neurological indicators and cost reduction. Further multicenter evaluations are required to confirm these results with different sedation protocols.

Aerophagia increases the risk of ventilator-associated pneumonia in critically-ill patients.

BACKGROUND: Gastric residual volume in ventilated critically ill may complicate gut function. Over the years studies suggested to tolerate progressively higher residuals. The relationship between such volumes and the development of ventilator-associated pneumonia (VAP) is still under debate. No reports deal with the relevant anecdotal finding of air in the stomach. Aim of the present study is to test the role of air in the development of VAPs. METHODS: Prospective observational trial in consecutive patients with a predicted length of ICU stay >3 days. The first 8 days of stay were studied. Sedation was targeted to have awake/cooperative patients. Early enteral nutrition was attempted. Gastric content was measured every 4 hours by 60 mL-syringe suction. Upper digestive intolerance (UDI) was defined as >2 consecutive findings of liquid >200 mL, aerophagia was defined as >2 consecutive findings of air >150 mL. RESULTS: Three hundred sixty-four patients enrolled, 43 developed VAP (11.8%). Patients were sedated with enteral (76% total time), intravenous (6%) or both (28%) drugs. Conscious sedation was achieved in 54% of the observations. 326 patients began enteral nutrition during the first 24 hours (1000 kcal median calorie intake). 10% developed UDI, 15% had aerophagia. No association was found between VAP and UDI (P=0.78), while significant association was found between VAP and aerophagia (OR=2.88, P<0.01). A sensitivity analysis, excluding patients admitted with respiratory infection, confirmed the results. CONCLUSION: High volumes of air in the stomach significantly increased the risk of developing VAP, while gastric residual volumes were not associated with the incidence of pneumonia.

Performance assessment of a glucose control protocol in septic patients with an automated intermittent plasma glucose monitoring device.

BACKGROUND & AIMS: The optimal level and modality of glucose control in critically ill patients is still debated. A protocolized approach and the use of nearly-continuous technologies are recommended to manage hyperglycemia, hypoglycemia and glycemic variability. We recently proposed a pato-physiology-based glucose control protocol which takes into account patient glucose/carbohydrate intake and insulin resistance. Aim of the present investigation was to assess the performance of our protocol with an automated intermittent plasma glucose monitoring device (OptiScanner™ 5000). METHODS: OptiScanner™ was used in 6 septic patients, providing glucose measurement every 15' from a side-port of an indwelling central venous catheter. Target level of glucose was 80-150 mg/dL. Insulin infusion and kcal with nutritional support were also recorded. RESULTS: 6 septic patients were studied for 319 h (1277 measurements); 58 [45-65] hours for each patient (measurements/patient: 231 [172-265]). Blood glucose was at target for 93 [90-98]% of study time. Mean plasma glucose was 126 ± 11 mg/dL. Only 3 hypoglycemic episodes (78, 78, 69 mg/dL) were recorded. Glucose variability was limited: plasma glucose coefficient of variation was 11.7 ± 4.0% and plasma glucose standard deviation was 14.3 ± 5.5 mg/dL. CONCLUSIONS: The local glucose control protocol achieved satisfactory glucose control in septic patients along with a high degree of safeness. Automated intermittent plasma glucose monitoring seemed useful to assess the performance of the protocol.

Intensive care of patients requiring open abdomen treatment: a case-series analysis.

BACKGROUND: This retrospective overview examines the management of patients with temporary open abdomen (OA). METHODS: The clinical characteristics and intensive care treatment of 34 consecutive patients with OA (1996-2012) were reviewed. RESULTS: Average age was 61 years, SAPS II score 43, SOFA 8. Two patients had non-contaminated abdomen; 12 had intact gut (only 8 later during stay); 7 repaired gut (only later 4); 13 cutaneous stoma (later 14), and 2 entero-atmospheric fistula (later 8+1 entero-enteral). The median ICU stay was 48 [36-94] days. One quarter of the 2376 ICU-days were classified as severe sepsis/septic shock (antibiotics were given for two thirds of the stay); three quarters were with ventilation; in 95% of days sedatives were given (mainly enterally). Continuous cavity lavage was done in three quarters of days; in 3% of days patients were fasted whereas >20 kcal/kg was given for 74% of days; we fed the gut in 95% of fed-days, in half of them combined with parenteral nutrition. Complications are discussed; mortality was 32.4%, limited to the ICU stay. CONCLUSION: The intensive care of patients with OA is challenging but can achieve better outcomes than expected. Continuous abdominal lavage improves the evacuation of contaminated fluid or debris and, coupled with antiseptics and low antibiotic pressure, reinforces the control of infection. The gut can be used for nutrition (even without gastrointestinal continuity), and long-term light sedation (mainly enteral) with minimal impact on perfusion, ventilation and gut motility.

Time course of endogenous nitric oxide inhibitors in severe sepsis in humans.

AIM: Asymmetric and symmetric dimethylarginines (ADMA and SDMA, respectively) are protein breakdown markers; both compete with arginine for cellular transport and both are excreted in urine. Moreover, ADMA is a non-selective inhibitor of nitric oxide (NO) synthase that is metabolized by a specific hydrolase in which the activity during stress remains controversial. While an increase in ADMA is known to be associated with adverse events, little is known about SDMA. We investigated plasma ADMA and SDMA levels during ICU stay to reveal the time course of endogenous NO inhibition in patients with sepsis. METHODS: A post hoc analysis from a prospective random controlled trial conducted in three ICUs was performed to study the pathophysiological pathways of sepsis. ADMA, SDMA, the ratio of ADMA/SDMA (a marker of ADMA catabolism), arginine, interleukin-6 (IL-6), tumor necrosis factor-alpha (TNF-alpha), and C reactive protein (CRP) were measured on days 1, 3, 6, 9, 12 and at discharge in 72 consecutive severely septic patients. RESULTS: Fasting basal glycemia, creatinine, IL-6, TNF-alpha, CRP, ADMA, and SDMA were higher than normal. The ADMA/SDMA ratio was decreased by 50%, and arginine levels were low. ADMA levels were related to the total Sequential Organ Failure Assessment (SOFA) scores and arginine levels, and inversely related to IL-6 and CRP levels. SDMA levels were related to Simplified Acute Physiologic Scores II (SAPS II), SOFA scores, blood urea, creatinine, and arginine levels. The ADMA/SDMA ratio was inversely related to IL-6 levels. In 58 ICU survivors, creatinine, IL-6, and CRP levels decreased over time; ADMA levels increased, SDMA levels remained stable, and the ADMA/SDMA ratio increased. In 14 non-survivors, creatinine, IL-6, TNF-alpha, CRP, and ADMA levels were stable, whereas the SDMA levels increased and the ADMA/SDMA ratio remained low. In both ICU survivors and non-survivors, the levels on the last ICU day confirmed the data trends. SDMA, but not ADMA, was associated with ICU mortality. CONCLUSION: ADMA catabolism appears to be activated by inflammation; its increase during the advanced septic phase in surviving patients may suggest an endogenous inhibition of NO synthesis during the full-blown septic phase. In severe sepsis, SDMA, but not ADMA, appears to be a marker of alterations in vital functions and mortality.

Reclamation and agricultural reuse of wastewater: the experience of the Cagliari sewage treatment plant (Sardinia, Italy).

In Sardinia, as in many other Mediterranean regions, recurrent droughts and climate change have dramatically reduced available water resources. As a result of this critical situation, in 1995 the Italian Government declared a state of emergency and drew up a program for financial support by the State and local authorities with the aim of reducing this serious deficit. One of the actions focused on reclaiming and reusing the effluent from the sewage treatment plant of Cagliari. This article reports on the multidisciplinary preliminary study performed by the Ente Acque della Sardegna (ENAS) to evaluate the suitability of reusing Is Arenas effluent for irrigation and on the operation of the tertiary treatment plant.

Effects of recombinant human activated protein C on the fibrinolytic system of patients undergoing conventional or tight glycemic control.

AIM: Recombinant human activated protein C (rh-APC) and tight glycemic control (TGC) have been shown to reduce mortality in septic patients. Both interventions can reduce the plasma concentration and/or activity of the most powerful suppressor of fibrinolysis, plasminogen activator inhibitor-1 (PAI-1). Our aim was to evaluate the effects on the fibrinolytic system after the administration of rh-APC in septic patients undergoing conventional or TGC. METHODS: Posthoc analysis of data was collected from 90 patients with severe sepsis/septic shock, randomized to either conventional or TGC groups. Independent of these treatments, patients with at least two organ dysfunctions simultaneously received rh-APC. Plasma levels of multiple biochemical markers for fibrinolysis, coagulation, and inflammation were determined every day for the 1st week and then on study days 9, 11, 13, 18, 23, and 28. Clinical data and sepsis-related organ failure assessment (SOFA) scores were also recorded. RESULTS: Patients who had received rh-APC exhibited significantly more impairments in fibrinolysis at baseline (PAI-1 activity 49.76 [24.61-71.82] vs 21.92 [6.47-55-83] IU/mL, P=0.03). The reductions in plasma PAI-1 activity over time associated with rh-APC treatment were different according to whether the treatment was administered to patients undergoing conventional or TGC (P=0.01). However, the most prominent reductions were in patients undergoing conventional glycemic control. Significant interactions between the two study interventions were also found for PAI-1 concentration (P<0.001), C-reactive protein (P=0.02), and interleukin-6 levels (P<0.001). CONCLUSIONS: Both rh-APC and TGC appear to improve fibrinolysis in septic patients. The reduction in the impairment of fibrinolysis associated with rh-APC treatment seems greater in patients undergoing conventional glycemic control than in those undergoing TGC.

Metabolic treatment of critically ill patients: energy balance and substrate disposal.

Oxidation of substrates is the main biochemical process used by the human body to produce energy. Different substrates (carbohydrates, lipids, and proteins) have different effects on oxygen consumption and carbon dioxide production: during the critical phase of pathologies it could be relevant pay attention to the use of various nutrients, that have some altered effect respect to the normal subjects metabolism, and during the length of metabolic treatment, too. Generally, nutrition lead to replenish body stores, while endogenous substrates are used to be oxidized. Critically ill patients show a preference for prompt energy availability (i.e. glucose) to avoid endogenous protein catabolism; lipids are shown to have a more pronounced storage effect. Adequate amount of energy intake in carbohydrates determine an increase of RQ, that means a shift from a more lipid-based to a more glucose-based oxidation. Composition of dietary intake can be usefully different for each pathology, and also for different periods of the same pathology, because critically ill patients have a variety of metabolic needs during their stay in ICU.

Transcriptional regulation of components of the type III secretion system and effectors in Pseudomonas syringae pv. phaseolicola.

Quantitative real-time polymerase chain reaction was used with specific TaqMan probes to examine transcription of selected hrp and effector genes in Pseudomonas syringae pv. phaseolicola strains 1448A (race 6) and 1449B (race 7). Transcripts examined were from genes encoding the regulators hrpR and hrpL, core structural components of the type III secretion system (TTSS) hrcC, hrcJ, hrcN, hrcU, and hrpA; the first open-reading frame of each hrp operon, including hrpF, hrpJ, hrpP, and hrpY, and also secreted effectors hrpZ, avrPphE, avrPphF, and virPphA. All genes were induced by incubation in a minimal medium and showed patterns of expression indicating regulation by HrpRS and HrpL. Basal mRNA levels and the timing of accumulation of transcripts after induction differed significantly, suggesting the operation of additional regulatory elements. However, no clear transcriptional hierarchy emerged to explain the ordered construction of the TTSS. Quantitative analysis confirmed that the rates and levels of transcript accumulation within the first 2 h after inoculation were considerably higher in planta than in vitro, and indicated that plant cell wall contact may enhance transcription of TTSS and effector genes in P. syringae pv. phaseolicola. The low-abundance hrcU mRNA had a half-life of 16.5 min, whereas other transcripts had half-lives between 3 and 8 min.

Different types of hysterectomy in the radio-surgical treatment of early cervical cancer (FIGO Ib-IIa).

PURPOSE OF INVESTIGATION: Surgery and radiotherapy achieved equivalent results for FIGO stages Ib-IIa invasive cervical carcinoma. The integration of radiotherapy and surgery provided the same results for a selected series of patients without increasing the rate of complications. The aim of the study was to verify if, applying a radio-surgical protocol, the reduction of the surgery extension on the parametrium in one of two consecutive series might achieve the same results in terms of survival and recurrence rates with fewer complications. MATERIAL AND METHODS: We analysed actuarial survival (with >10-year follow-up), local control rates and morbidity of 390 patients who had different the kinds of surgery applied in the radio-surgical treatment protocol: Protocol A: brachytherapy plus type III radical hysterectomy vs Protocol B: brachytherapy plus type II radical hysterectomy. Patients were all included in an analysis of complications according to the French-Italian glossary. RESULTS: Analyses showed no differences in terms of survival, patterns of recurrences and onset time within the two protocols. Urinary complications were more frequent and severe in protocol A vs protocol B (G2: 26.5% vs. 6.1%; G3: 5.3% vs. 3.6%). CONCLUSION: Our study pointed out that the reduction of the surgery extension allowed the same overall survival and relapses with fewer complications particularly in terms of grade of severity.

The hydrophobin HCf-1 of Cladosporium fulvum is required for efficient water-mediated dispersal of conidia.

Six hydrophobin genes (HCf-1 to -6) have thus far been identified in the tomato pathogen Cladosporium fulvum. HCf-1 to -4 are Class I hydrophobins and HCf-5 and -6 are Class II hydrophobins. In this paper we describe the isolation of deletion mutants that lack HCf-1, HCf-2, or both these genes. Global down-regulation of the expression of Class I hydrophobins is achieved by homology-dependent gene silencing. Analysis of the mutant strains shows that HCf-1 confers hydrophilic character to the conidia and this facilitates the dissemination of conidia on the surface of water droplets. Other Class I hydrophobins, such as HCf-3 or HCf-4, may be involved in the development and germination of conidia.

HCf-6, a novel class II hydrophobin from Cladosporium fulvum.

C. fulvum, a fungal tomato pathogen, has previously been shown to express a complex family of hydrophobin genes including four class I hydrophobins and one class II hydrophobin. Here we describe a gene for HCf-6, a sixth member of the hydrophobin family and the second class II gene. The protein is predicted to consist of a signal sequence, an N-terminus rich in glycine and asparagine and a C-terminal hydrophobic domain which bears the hall-marks of hydrophobins. In contrast to the previously described class II hydrophobin HCf-5, HCf-6 is expressed in mycelium growing in pure culture and mRNA levels do not increase during sporulation. It is down-regulated by carbon starvation but not by depletion of nitrogen in the growth medium.

Stable transformation of erysiphe graminis an obligate biotrophic pathogen of barley.

Barley powdery mildew, Erysiphe graminis f.sp. hordei, is an obligate biotrophic pathogen and as such cannot complete its life cycle without a living host. The inability to transform this fungus and manipulate its genome has constrained research towards understanding its life cycle and pathogenicity. Here we describe an in planta transformation system based on delivery of DNA using a gold-particle gun and selection using benomyl or bialaphos. Using this method, we consistently obtained stable transformants with efficiencies comparable to other filamentous fungi. Stable expression of the beta-glucuronidase in E. graminis was demonstrated by co-transforming the uidA gene with the selectable markers.

Isolation and characterisation of five different hydrophobin-encoding cDNAs from the fungal tomato pathogen Cladosporium fulvum.

Five different hydrophobin-encoding cDNA clones from Cladosporium fulvum were isolated from cDNA libraries, made from nutrient-depleted mycelium. One cDNA clone was identical to the previously isolated hydrophobin HCf-1. The other clones were named HCf-2, -3, -4 and -5. HCf-1, -2, -3 and -4 show a high degree of identity, and are predicted to encode class I hydrophobins. HCf-5 encodes a class II hydrophobin. The expression patterns of these hydrophobins at various stages of development, and in liquid media lacking either carbon or nitrogen, or both, showed clear differences. All hydrophobins were more strongly expressed during sporulation than before, with HCf-4 and -5 showing the highest increase. Expression of HCf genes in infected plants was also higher at 16 days than at 10 days after infection. The expression of HCf-5 in sporulating mycelium was much lower in planta than in vitro. All HCf genes were upregulated under conditions of nutrient deprivation. HCf-1, -2, -3 and -4 showed highest levels of transcription in medium lacking both carbon and nitrogen. Expression of HCf-5 was highest in medium lacking nitrogen but containing carbon. HCf-1 was generally the most abundant hydrophobin. The introduction of multiple copies of HCf-1, which caused co-suppression of the endogenous HCf-1 gene, was shown to affect the expression of HCf-2, -3 and -4 also. Expression of HCf-4 was suppressed, but expression of HCf-2 and -3 was upregulated. Expression of HCf-5 was not changed.

Co-suppression of the hydrophobin gene HCf-1 is correlated with antisense RNA biosynthesis in Cladosporium fulvum.

Transformation of Cladosporium fulvum with DNA containing a truncated copy of the hydrophobin gene HCf-1 causes co-suppression of hydrophobin synthesis in 30% of the transformants. The co-suppressed isolates have a hydrophilic phenotype, lower levels of HCf-1 mRNA than wild type and contain multiple copies of the plasmid integrated as tandem repeats at ectopic sites in the genome. Gene silencing is not associated with DNA cytosine methylation. Nuclear run-off experiments reveal that transcription rate of HCf-1 in the co-suppressed isolates is higher than in the untransformed strains, suggesting that silencing acts at the post-transcriptional level. We show, for the first time in fungi, that co-suppression is correlated with the presence of antisense RNA, and that this is synthesised on a DNA template. Derivatives showing reversion to the wild-type phenotype and restoration of HCf-1 gene expression were also observed. Reversion is associated with loss of some copies of the transgene. We propose that co-suppression is due to ectopic integration of the transgene next to promoters which initiate transcription to form antisense RNA and that this in turn determines down-regulation of HCf-1.

Use of monoclonal antibodies to determine biomass of Cladosporium fulvum in infected tomato leaves.

A monoclonal antibody, OX-CH1, was raised against surface washings of Cladosporium herbarum. This antibody recognizes an epitope that is found in various fungi belonging to the genus Cladosporium, including C. fulvum, the causal agent of tomato leaf mold. The epitope is present at comparable levels in two different races of C. fulvum and in transgenic isolates derived from them. The epitope is heat-and protease-resistant but sensitive to oxidation with periodate and it is constitutively expressed in C. fulvum grown in pure culture and on the plant. C. fulvum can be detected in infected tissues at levels starting from around 1 mg fresh weight of fungus per g fresh weight of leaf tissue. Noninfected tomato leaves do not cross-react with OX-CH1. We have developed an enzyme-linked immunosorbent assay (ELISA) for fungal biomass in tomato leaves and compared it with the assay based on measurements of beta-glucuronidase (GUS) activity in tissues infected with a transgenic isolate of C. fulvum race 4 carrying a uidA gene; the two assays give similar results.

[Ciaglia's percutaneous dilatative tracheostomy in intensive care. Perioperative complications and long-term results]. / Tracheostomia percutanea dilatativa secondo ciaglia in terapia intensiva. Complicanze perioperatorie ed esiti a distanza.

BACKGROUND AND AIM: The introduction of percutaneous tracheostomic techniques using dilatation (PDT) has led to the gradual disuse of conventional methods of surgery. The aim of this study was to evaluate ventilatory function in critical patients during the execution of PDT using Ciaglia's method as well as, in the postoperative phase, the long-term results, namely endotracheal lesions and cosmetic deformities of the stoma. EXPERIMENTAL DESIGN: A prospective study was carried out on a consecutive series of adult patients. SETTING: The intensive care ward of a 600-bed teaching hospital. PATIENTS: A group of 50 critical patients undergoing PDT from 1993 to 1996. Valuations of ventilatory function: expiratory volume (EV), PaO2 and PaCO2, were calculated in 40 patients undergoing PDT. Endoscopic controls of the trachea were performed in 21 surviving patients 60 days after the removal of the tracheostomic tube and a cosmetic evaluation of the tube insertion site was also made in the same patients. RESULTS: During PDT a mean reduction of EV was observed of 1.41/min and PaO2 values also diminished 15% accompanied by a 14.2% increase in PaCO2. The endoscopic control performed after 60 days in 21 out of 50 patients revealed a subglottal stenosis in 1 patient, the presence of nonstenosing cicatricial granuloma in 2 patients, edema in 2 patients and dysepithelisation of the tracheal mucosa in a further 2 patients. The remaining 66% showed flattening of the mucosa and complete restitutium ad integrum. No usurpations and cicatricial tractions of the tube insertion site were observed. CONCLUSIONS: This prospective study of intraoperative ventilatory function and the final outcome of PDT according to Ciaglia confirms that this is a reliable technique for the ventilatory management of critical patients with a low percentage of long-term complications.