Candidemia in intensive care unit: a nationwide prospective observational survey (GISIA-3 study) and review of the European literature from 2000 through 2013.

BACKGROUND: Candida bloodstream infections (BSI) represent an important problem in Intensive Care Units (ICUs). The epidemiology of candidemia is changing with an increase in the proportion of Candida (C.) non-albicans. OBJECTIVES: An Italian 2-year observational survey on ICU was conducted to evaluate the species distribution and possible differences between BSI caused by C. albicans and C. non-albicans. For comparative purposes, we performed a European literature-based review to evaluate distribution and frequency of Candida spp. causing ICU candidemia, during the period 2000-2013. MATERIALS AND METHODS: This laboratory-based survey involved 15 microbiology centers (GISIA-3 study). All candidemia episodes in adult patients were considered. Data were prospectively collected from 2007 to 2008. PubMed was searched for peer-reviewed articles. RESULTS: In total, 462 candidemia episodes were collected. C. albicans accounted for 49.4% of the isolates, followed by C. parapsilosis (26.2%) and C. glabrata (10.4%). Mortality was higher in patients with C. non-albicans than C. albicans (47.3% vs. 32.4 %, p > 0.05). Among risk factors, parenteral nutrition was more common (p = 0.02) in non-albicans candidemia, while surgery was more frequent (p = 0.02) in C. albicans candidemia. Twenty-four relevant articles were identified. C. albicans was the predominant species in almost all studies (range 37.9% -76.3%). C. glabrata was commonly isolated in the German-speaking countries, France, UK and North Europe; C. parapsilosis in Turkey, Greece and Spain. CONCLUSIONS: Although C. non-albicans BSI is increasing, our study shows that C. albicans is still the predominant species in ICU candidemia. There are differences in the epidemiology of Candida BSI among European countries, with a prevalence of C. glabrata and C. parapsilosis in Northern and Southern countries, respectively.

Fungal infections in ICU patients: epidemiology and the role of diagnostics.

Invasive fungal infections (IFIs) are on the increase not only among oncology and transplant patients but also among patients admitted to intensive care units (ICU). The rise in ICU IFIs can be attributed to the growing use of complex surgical procedures, invasive medical devices, and long-term, broad-spectrum antibiotic therapy. The majority of these life-threatening infections are caused by the well-known opportunistic pathogens Candida albicans and Aspergillus fumigatus, but new opportunistic pathogens, including yeast-like and other filamentous fungi, have emerged as additional causes. Invasive Candida infections, particularly candidemia, represent the most common IFI in critically ill patients. The species that cause candidemia markedly differ in their responses to antifungal drugs; for this reason, therapy must be tailored to the susceptibility characteristics of the infectious agent. Candidemia caused by non-albicans Candida species is increasing worldwide, and these infections are generally associated with high mortality rates, particularly bloodstream infections caused by C. krusei, which is innately resistant to fluconazole, or C. glabrata, which easily develops azole resistance. Although invasive yeast infections can be considered the most important causes of morbidity and mortality in ICU patients, pulmonary aspergillosis has recently emerged as an additional complication. Diagnosis of IFIs can be achieved using conventional approaches (microscopy, culture, and serology) and newer methods, including antigen detection and polymerase chain reaction (PCR) assays. Because most of the conventional approaches lack sensitivity, antigen detection and PCR assays could represent a valid alternative; however, these procedures need to be standardized and evaluated in a large number of patients.

Zygomycosis in Italy: a survey of FIMUA-ECMM (Federazione Italiana di Micopatologia Umana ed Animale and European Confederation of Medical Mycology).

The aims of the study were to analyze the clinical and epidemiological characteristics and treatments for patients who developed zygomycosis enrolled in Italy during the European Confederation of Medical Mycology of medical mycology survey. This prospective multicenter study was performed between 2004 and 2007 at 49 italian Departments. 60 cases of zygomycosis were enrolled: the median age was 59.5 years (range 1-87), with a prevalence of males (70%). The majority of cases were immunocompromised patients (42 cases, 70%), mainly hematological malignancies (37). Among non-immunocompromised (18 cases, 30%), the main category was represented by patients with penetrating trauma (7/18, 39%). The most common sites of infection were sinus (35%) with/without CNS involvement, lung alone (25%), skin (20%), but in 11 cases (18%) dissemination was observed. According to EORTC criteria, the diagnosis of zygomycosis was proven in 46 patients (77%) and in most of them it was made in vivo (40/46 patients, 87%); in the remaining 14 cases (23%) the diagnosis was probable. 51 patients received antifungal therapy and in 30 of them surgical debridement was also performed. The most commonly used antifungal drug was liposomal amphotericin B (L-AmB), administered in 44 patients: 36 of these patients (82%) responded to therapy. Altogether an attributable mortality rate of 32% (19/60) was registered, which was reduced to 18% in patients treated with L-AmB (8/44). Zygomycosis is a rare and aggressive filamentous fungal infection, still associated with a high mortality rate. This study indicates an inversion of this trend, with a better prognosis and significantly lower mortality than that reported in the literature. It is possible that new extensive, aggressive diagnostic and therapeutic procedures, such as the use of L-AmB and surgery, have improved the prognosis of these patients.

In vitro testing of Aspergillus fumigatus clinical isolates for susceptibility to voriconazole, amphotericin B and itraconazole: comparison of sensititre versus NCCLS M38-A using two different inocula.

Voriconazole, amphotericin B and itraconazole were tested in vitro against 18 strains of Aspergillus fumigatus isolated from cystic fibrosis patients. Susceptibility was tested with the broth microdilution method (M38-A protocol-NCCLS). Results of this reference method were compared with those of an experimental commercial microdilution broth method (Sensititre). Two different inocula, prepared from 2- and 7-day cultures, were used. Minimum inhibitory concentrations (MICs) of the reference method ranged from 0.25 to 2 microg/ml for voriconazole, 0.06 to 1 microg/ml for amphotericin B, 0.016 to >16 microg/ml for itraconazole. There were no significant differences in the MIC ranges or MIC90 values obtained with the two testing methods or with the two types of inocula. These findings confirm the good in vitro activity of voriconazole, itraconazole and amphotericin B against A. fumigatus. They also indicate that reliable susceptibility data can be generated more rapidly by commercial systems and use of 2-day cultures for inoculum preparation.

Mapping of protein domains of hepatitis A virus 3AB essential for interaction with 3CD and viral RNA.

The small hydrophobic protein 3AB of the picornaviruses, encompassing the replication primer 3B, has been suggested to anchor the viral replication complex to membranes. For hepatitis A virus (HAV) 3AB, we have previously demonstrated its ability to form stable homodimers, to bind to membranes, and to interact specifically with RNA, implicating its multiple involvement in viral replication. In the present report, we show that HAV 3AB additionally interacts with HAV protein 3CD, a feature also described for the corresponding polypeptide of poliovirus. By assessing the interactions of three deletion mutants, distinct domains of HAV 3AB were mapped. The hydrophobic domain and the 3B moiety were found to be essential for the 3AB interaction with 3CD. Both electrostatic and hydrophobic forces are involved in this interaction. The cluster of charged amino acid residues at the C terminus of 3A seems to determine the specificity of 3AB interaction with RNA structures formed at either terminus of the HAV genome. Furthermore, our data implicate that 3A can interact with HAV RNA. Compared with poliovirus 3AB, which by itself is a nonspecific RNA-binding protein, HAV 3AB specifically recognizes HAV RNA structures that might be of relevance for initiation of viral RNA replication.

Retrospective study of candidemia in patients with hematological malignancies. Clinical features, risk factors and outcome of 76 episodes.

A retrospective study of 76 episodes of candidemia in 73 patients with underlying hematological malignancy, from 1988 until 1997, has been conducted to evaluate the clinical characteristics and to ascertain the variables related to the onset and the outcome of candidemia. The most frequent malignancy was acute myeloid leukemia (29 episodes). Candidemia developed mainly during aplasia in patients refractory to chemotherapy (42%). In 65 episodes (86%) the patients were neutropenic (ANC <1 x 10(9)/l) before the candidemia diagnosis for a median time of 13 d, and in 53 episodes (70%) at microbiological diagnosis of candidemia ANC was <1 x 10(9)/l. Candida albicans was the most frequently isolated etiologic agent (31 episodes), but C. non-albicans species sustained the majority of candidemia. Seventeen candidemias developed during azoles prophylaxis. One month after the diagnosis of candidemia, 26 patients died. In 19 cases, death was attributable to candidemia. The case-control study demonstrated, at univariate analysis, that the colonization with Candida. spp. (p=0.004), antimycotic prophylaxis (p=0.01), presence of central venous catheter (p=0.01), neutropenia (p=0.002), and the use of glycopeptide (p=0.0001) increased the risk of candidemia. Using multivariate regression analysis only colonization with Candida spp. and the previous therapy with glycopeptide were associated with a significantly increased risk. Acute mortality, expressed by a cumulative probability of survival at 30 d from diagnosis of candidemia, was 0.67 (95% C.I. 0.55-0.77) and was significantly reduced in patients with neutrophils <1 x 10(9)/l when compared to those with neutrophils >1 x 10(9)/l (p at Mantel-Cox=0.029). Overall cumulative probability of survival at 1 yr was 0.38 (95% C.I. 0.27-0.49) and only the treatment with Amfotericin B significantly reduced the risk of death.

Role of protease inhibitors in preventing recurrent oral candidosis in patients with HIV infection: a prospective case-control study.

This study was conducted to evaluate the efficacy of highly active anti-retroviral therapy (HAART) in preventing recurrence of oral candidosis (OC) associated with HIV. A prospective case-controlled observational study was performed in an inner-city university-hospital HIV/AIDS clinic. Ninety-three HIV-positive study subjects with a history of recurrent OC were divided into two groups: protease inhibitors (PI)-treated patients (group 1, n = 30) and non-PI-treated patients (group 2, n = 63). Study subjects were matched for sex, age, stage of HIV infection, and peripheral CD4+ T-cell counts. The non-PI-treated group was further subdivided into the following three subgroups: HIV-positive study subjects treated with reverse transcriptase inhibitors (RTI; groups 2a and 2c) and HIV-positive study subjects not treated with RTIs (group 2b). Group 2c met the same inclusion criteria as group 2a had but was matched 6 months after the beginning of the study. We also assessed in vitro peripheral blood mononuclear cells (PBMC) and their lymphoproliferative response, as well as cutaneous delayed-type hypersensitivity (DTH) response to Candida-associated antigens in a randomly selected sample of study subjects divided into those treated with PIs and those who were not. During a 1-year follow-up, OC was diagnosed in 2 (7%) PI-treated and 23 (36%) non-PI-treated patients (p<.001). In addition to comparing findings in group 1 with those in group 2c, OC was detected in 14 (50%) non-PI-treated patients compared with no HAART-treated study subjects (p<.001). Only 41% of PI-treated study subjects had positive lymphoproliferative response in PBMCs and none was positive in terms of DTH to Candida antigens (p = not significant versus non-PI-treated study subjects). While objectively demonstrating a beneficial effect of HAART in preventing recurrence of OC infections, our findings suggest this effect cannot be not fully accounted for by reconstitution of anti-Candida cell-mediated immunity, given that other mechanisms, even of a nonimmune nature, could have some effect.

PCR-restriction enzyme analysis for detection of Candida DNA in blood from febrile patients with hematological malignancies.

Blood samples were drawn daily from 72 patients who had hematological malignancies, neutropenia, and fever and who had failed to respond to broad-spectrum antibiotics. Each sample was used for conventional fungal blood cultures and for detection and identification of Candida DNA by a PCR method with subsequent restriction enzyme analysis (REA) recently developed in our laboratory. The PCR method was able to detect five CFU of Candida spp. per ml of blood, and subsequent REA of the amplicons allowed the identification of the Candida species most commonly implicated in cases of candidiasis. Thirty-one patients were PCR-REA positive, and four of these patients were also culture positive. The ultimate diagnosis for 13 of these patients and 1 patient who was PCR-REA negative was disseminated candidiasis (confirmed by clinical data, multiple cultures, histology, autopsy, and/or ultrasonographic evidence of hepatosplenic candidiasis). The molecular method is significantly more sensitive than conventional fungal blood cultures and has a high negative predictive value (97.5%) for the development of disseminated candidiasis in neutropenic patients.

Polypeptide 3AB of hepatitis A virus is a transmembrane protein.

Hepatitis A virus (HAV) protein 3AB is a membrane-interacting protein containing a stretch of 21 hydrophobic amino acid residues. The nature of its membrane association was studied in detail by analysing various deletion mutants. In vivo and in vitro expression of the wild-type protein and its mutants allowed to demonstrate that the hydrophobic domain interacts with membranes and to define the portions essential for this feature. Furthermore, the results suggest that 3AB behaves as an integral membrane protein. Expression in Escherichia coli showed that 3AB can be isolated, in association with membranes, both in monomeric and in dimeric form. This finding was confirmed in vitro after post-translational incubation of the protein with microsomal membranes. Analysis of deletion mutants demonstrated that the dimerization region colocalises with the hydrophobic transmembrane domain, implicating that HAV 3AB could form oligomers mediated by the interaction of transmembrane alpha-helices.

Comparative analysis of prognostic indicators of aspergillosis in haematological malignancies and HIV infection.

The objective of this study was to identify the prognostic factors influencing the outcome of aspergillosis in two models of immunodeficiency, namely haematological malignancies and HIV infection. The study is based on a 5 year prospective logistic regression analysis of risk factors, clinical features, radiological findings and therapy affecting the prognosis of aspergillosis in 43 patients, i.e. 27 haematological neoplastic patients (group A) and 16 HIV infected patients (group B). Univariate analysis indicated that neutropenia (P = 0.02), haemoptysis (P = 0.03) and concomitant AIDS (P = 0.02), negatively influenced the prognosis of aspergillosis. Comparing the two groups of patients, significant differences emerged in the prognostic indicators. In particular respiratory failure (P = 0.02) and radiological bilateral involvement of the lungs were associated with a poor prognosis in group A (P = 0.04) and low (2100/mm3) T CD4+ cell count in group B (P = 0.02). At variance, a better prognosis was documented in patients treated with sequential therapy (amphotericin B and itraconazole) only within the group of haematological patients (P = 0.003). On multivariate analysis sequential therapy (P = 0.01) and AIDS (P = 0.03) were independent prognostic indicators of aspergillosis. In conclusion, our prospective study indicates that aspergillosis, although an uncommon event in patients with HIV infection, has a more severe prognosis in comparison to haematological patients. Future prospective clinical trials are necessary to confirm the real importance of the sequential therapy, with amphotericin B and itraconazole, in patients with aspergillosis.

The role of bronchoalveolar lavage in the microbiological diagnosis of pneumonia in patients with haematological malignancies.

In the aetiological diagnosis of pulmonary infections in patients affected by haematological malignancies we evaluated the utility of bronchoalveolar lavage (BAL). One hundred and twenty-seven BAL were performed in 119 patients. In our series, we identified the agent of pneumonia in 53.5% of episodes with the best results in aspergillosis, very common in these patients. The previous empirical anti-infective treatment was modified in 14 episodes (11%). The procedure was generally well tolerated and only one patient bled. We maintain that BAL is a useful diagnostic tool for detecting the agents of pulmonary infections in patients with haematological malignancies, especially when the routine microbiological procedures fail, and it also represents a good alternative to more invasive procedures.

Interaction of hepatitis A virus (HAV) precursor proteins 3AB and 3ABC with the 5' and 3' termini of the HAV RNA.

RNA secondary structures within the terminal nontranslated regions of entero- and rhinoviral genomes interact specifically with viral nonstructural proteins and are required in cis for viral RNA replication. Here we show that recombinant hepatitis A virus (HAV) polypeptide 3ABC specifically interacts in vitro with secondary RNA structures formed at both the 5' and 3' terminus of the viral genome. Similar to protein 3AB, HAV 3ABC bound to the 3' terminal RNA structure which did not interact with the mature proteinase 3C. In contrast to 3AB, 3ABC interacted with RNA stem-loop IIa and combinations of individual secondary structure elements of the 5' noncoding region. RNA binding of the precursor polypeptide 3ABC was 50 times stronger than that of 3AB and 3C, implicating a specific role of this stable processing intermediate in viral genome replication.

Adjuvant therapy with rhGM-CSF for the treatment of Blastoschizomyces capitatus systemic infection in a patient with acute myeloid leukemia.

We report a patient with acute myeloid leukemia and Blastoschizomyces capitatus sepsis who developed multiple abscesses when neutrophils recovered. The patient did not respond to antifungal therapy and her clinical condition showed an improvement only after rhGM-CSF was added to the treatment.

Recombinant expression of hepatitis A virus protein 3A: interaction with membranes.

The function of hepatitis A virus (HAV) protein 3A and its structural requirements were studied in vitro and in a bacterial expression system by comparing the polypeptide precursor 3AB derived from a cytopathogenic strain with that of an attenuated strain. Although the precursor polypeptides 3AB of both HAV strains bind to microsomal membranes after translation in vitro they differ in inducing membrane permeability when expression is induced in bacteria. Intake and release of macromolecules was dramatically increased when 3AB of the cytopathogenic strain was expressed. Amino acid sequence alignments suggest that membrane binding might be due to a hydrophobic stretch near the C-terminus of 3A found in all picornaviruses whereas the ability to induce permeability of E. coli membranes is determined by an amphipathic helix formed at the N-terminus of 3A of HAVFG.

In vitro susceptibility of Candida species isolated from patients with haematological malignancies.

Candida spp. (83 isolates including C. (Torulopsis) glabrata) were tested in vitro for their susceptibility to 5-fluorocytosine, amphotericin B, ketoconazole, itraconazole, fluconazole, and miconazole. The yeasts were isolated from clinical specimens, mostly from the lower respiratory tract, of 30 oncologic patients, 27/30 with haematological malignancies, during a 6-month period (December 1991-May 1992). Minimal inhibitory concentration (MIC) and minimal fungicidal concentration (MFC) values of the 6 drugs were obtained for each yeast using a microdilution broth method developed in our laboratory. Amphotericin B, and 5-fluorocytosine were active against the majority of the yeasts with MIC90/MFC90 values within achievable serum concentrations (3.12/6.25 micrograms ml(-1) and 0.625/0.625 micrograms ml(-1) respectively). Azole derivatives showed a species-specific activity. MFC values were two to four times higher than those of the MICs, confirming the fungistatic rather than fungicidal activity of azole derivatives. An interesting correlation was found when the in vitro susceptibility values of the isolates were compared with data of patients with or without antifungal prophylaxis or therapy during that period. In general, with respect to fluconazole, C. albicans strains isolated from patients who received no treatment showed MIC and MFC values lower than those obtained from patients who were under prophylaxis or treatment with this drug. Fluconazole administration appears to influence in vitro susceptibility testing.

Fluconazole for primary prophylaxis of AIDS-associated cryptococcosis: a case-control study.

In order to verify whether fluconazole has a prophylactive effect against the occurrence of cryptococcosis in HIV-infected patients and to identify other factors capable of increasing or reducing the risk of this infection, we arranged a case-control study of 17 patients with cryptococcal infection. 34 controls were selected, matched by presence of an AIDS-defining event, CD4 cell count, and date of T-cell phenotyping. No significant difference in exposure to fluconazole, in total days of treatment, or in total dose administered was observed between cases and controls. However, control patients took a significantly higher average daily dosage of fluconazole and a linear tendency in risk reduction (p = 0.04) in relation to increasing dosage was observed. Antiretroviral therapy and an average daily fluconazole dose exceeding 150 mg both each reduced the risk of a cryptococcal infection.

[The euthyroid sick syndrome. Its incidence and clinical significance in an internal medicine department]. / Sindrome del malato eutiroideo. Incidenza e significato clinico in un reparto di medicina interna.

In this paper the authors have evaluated the incidence and the clinical implications of sick euthyroid syndrome (SES) in a group of 144 patients in a department of internal medicine. SES is an alteration of thyroid hormone values in the absence of a thyroid disease, which is seen in patients suffering from serious diseases. Having classified SES into 3 subgroups according to the different alterations seen in the values of T3, T4, FT3, FT4, TSH, rT3 and TBG, they show the hypotheses that explain the biochemical mechanisms which are at the basis of these hormonal alterations. Fourteen of the 144 patients under observation were excluded as they were suffering from ascertained or subclinical thyroid disease. Thirty (23% of cases) of the remaining 130 patients had alterations of the thyroid hormones in accordance with SES diagnosis. Of these 30 patients, 19 had hormone values found in SES type I (63%), 2 in SES type II (6.5%) and 9 in SES type III (30.5%). In SES type I the diseases seen, in order of frequency, were: obstructive chronic bronchopneumopathy with acute respiratory failure, diabetic ketoacidosis, neoplasms, ischemic heart disease, cardiac failure, chronic renal failure, liver diseases, acute cerebral vasculopathies, sepsis and collagenopathies. The disease seen in the 2 cases of SES type II was obstructive chronic bronchopneumopathy with acute respiratory failure. In SES type III the diseases seen were, in order of frequency: diabetic ketoacidosis, lung diseases, ischemic heart disease, cardiac failure, peripheral arteriopathies, acute cerebral vasculopathies, neoplasms, liver diseases, acute renal failure. The incidence of SES in 23% of the admitted to hospital patients was found to be slightly higher than in other studies; this could be explained by a stricter selection of inpatients: in fact self-sufficient patients or those not needing urgent admission, were sent to an efficient out patient clinic where necessary examinations were quickly carried out, hospitalization being reserved for patients with more serious illnesses. We would like to underline how the incidence of SES is much greater than that of what is known as thyroid disease (23% compared to 5%), thereby confirming that it is the most frequent cause of alterations of thyroid hormones. With regard to the pathogenetical hypotheses, it is confirmed that in SES, the reduction of T3 values is accompanied by an increase in the values of rT3 as for reduced activity of 5-desiodinasis enzyme. In SES type III the increase of T4 values is due to the increase of TBG resulting in an increase in the link for T4 and therefore a reduced peripheral hormone activity.(ABSTRACT TRUNCATED AT 400 WORDS)