Single dose pharmacodynamics of amphotericin B against Aspergillus species in an in vitro pharmacokinetic/pharmacodynamic model

Conventional MIC testing of amphotericin B results in narrow MIC ranges challenging the detection of resistant strains. In order to discern amphotericin B pharmacodynamics, the in vitro activity of amphotericin B was studied against Aspergillus isolates with the same MIC with a new in vitro pharmacokinetic/pharmacodynamic (PK/PD) model that simulates amphotericin B human plasma levels. Clinical isolates of A. fumigatus, A. terreus and A flavus with the same CLSI modal MICs of 1 mg/l were exposed to amphotericin B concentrations following the plasma concentration-time profile after single bolus administration with Cmax 0.6, 1.2, 2.4 and 4.8 mg/L. Fungal growth was monitored up to 72h based on galactomannan production. Complete growth inhibition was observed only against A. fumigatus with amphotericin B Cmax ≥2.4 mg/L. At lower Cmaxs 0.6 and 1.2 mg/L, a significant growth delay of 34h and 52h was observed, respectively (pA flavus>A. terreus in the in vitro PK/PD model possibly reflecting the different concentration- and time-dependent inhibitory/killing activities amphotericin B exerting against these species

Fundamental niche prediction of the pathogenic yeasts cryptococcus neoformans and cryptococcus gattii in europe

Fundamental niche prediction of Cryptococcus neoformans and Cryptococcus gattii in Europe is an important tool to understand where these pathogenic yeasts have a high probability to survive in the environment and therefore to identify the areas with high risk of infection. In this study, occurrence data for C. neoformans and C. gattii were compared by MaxEnt software with several bioclimatic conditions as well as with soil characteristics and land use. The results showed that C. gattii distribution can be predicted with high probability along the Mediterranean coast. The analysis of variables showed that its distribution is limited by low temperatures during the coldest season, and by heavy precipitations in the driest season. C. neoformans var. grubii is able to colonize the same areas of C. gattii but is more tolerant to cold winter temperatures and summer precipitations. In contrast, the C. neoformans var. neoformans map was completely different. The best conditions for its survival were displayed in sub-continental areas and not along the Mediterranean coasts. In conclusion, we produced for the first time detailed prediction maps of the species and varieties of the C. neoformans and C. gattii species complex in Europe andMediterranean area

International Society of Human and Animal Mycology (ISHAM)-ITS reference DNA barcoding database - the quality controlled standard tool for routine identification of human and animal pathogenic fungi

Human and animal fungal pathogens are a growing threat worldwide leading to emerging infections and creating new risks for established ones. There is a growing need for a rapid and accurate identification of pathogens to enable early diagnosis and targeted antifungal therapy. Morphological and biochemical identification methods are time-consuming and require trained experts. Alternatively, molecular methods, such as DNA barcoding, a powerful and easy tool for rapid monophasic identification, offer a practical approach for species identification and less demanding in terms of taxonomical expertise. However, its wide-spread use is still limited by a lack of quality-controlled reference databases and the evolving recognition and definition of new fungal species/complexes. An international consortium of medical mycology laboratories was formed aiming to establish a quality controlled ITS database under the umbrella of the ISHAM working group on "DNA barcoding of human and animal pathogenic fungi." A new database, containing 2800 ITS sequences representing 421 fungal species, providing the medical community with a freely accessible tool at http://www.isham.org and http://its.mycologylab.org/ to rapidly and reliably identify most agents of mycoses, was established. The generated sequences included in the new database were used to evaluate the variation and overall utility of the ITS region for the identification of pathogenic fungi at intra-and interspecies level. The average intraspecies variation ranged from 0 to 2.25%. This highlighted selected pathogenic fungal species, such as the dermatophytes and emerging yeast, for which additional molecular methods/genetic markers are required for their reliable identification from clinical and veterinary specimens.This study was supported by an National Health and Medical Research Council of Australia (NH&MRC) grant [#APP1031952] to W Meyer, S Chen, V Robert, and D Ellis; CNPq [350338/2000-0] and FAPERJ [E-26/103.157/2011] grants to RM Zancope-Oliveira; CNPq [308011/2010-4] and FAPESP [2007/08575-1] Fundacao de Amparo Pesquisa do Estado de So Paulo (FAPESP) grants to AL Colombo; PEst-OE/BIA/UI4050/2014 from Fundacao para a Ciencia e Tecnologia (FCT) to C Pais; the Belgian Science Policy Office (Belspo) to BCCM/IHEM; the MEXBOL program of CONACyT-Mexico, [ref. number: 1228961 to ML Taylor and [122481] to C Toriello; the Institut Pasteur and Institut de Veil le Sanitaire to F Dromer and D Garcia-Hermoso; and the grants from the Conselho Nacional de Desenvolvimento Cientifico e Tecnologico (CNPq) and the Fundacao de Amparo a Pesquisa do Estado de Goias (FAPEG) to CM de Almeida Soares and JA Parente Rocha. I Arthur would like to thank G Cherian, A Higgins and the staff of the Molecular Diagnostics Laboratory, Division of Microbiology and Infectious Diseases, Path West, QEII Medial Centre. Dromer would like to thank for the technical help of the sequencing facility and specifically that of I, Diancourt, A-S Delannoy-Vieillard, J-M Thiberge (Genotyping of Pathogens and Public Health, Institut Pasteur). RM Zancope-Oliveira would like to thank the Genomic/DNA Sequencing Platform at Fundacao Oswaldo Cruz-PDTIS/FIOCRUZ [RPT01A], Brazil for the sequencing. B Robbertse and CL Schoch acknowledge support from the Intramural Research Program of the NIH, National Library of Medicine. T Sorrell's work is funded by the NH&MRC of Australia; she is a Sydney Medical School Foundation Fellow.info:eu-repo/semantics/publishedVersio

Study of Toll-like receptors (TLR) expression on haematopoietic and stromal bone marrow cells of patients with myelodysplastic syndromes and hypoplastic neutropenias

Background: Excessive pro-inflammatory cytokine production in the bone marrow (BM) has been recognized as a prominent pathogenetic mechanism for the disturbed hematopoiesis in patients with Myelodysplastic Syndromes (MDS) and Chronic Idiopathic Neutropenia (CIN). However, the upstream pathways, the exact cellular source and the triggering events related to cytokine excess in MDS BM remain unknown.Furthermore, in patients with MDS, infection vulnerability does not always reflect the severity of neutropenia. This observation suggests that additional factors may predispose MDS patients to infections.Aim: The first objective of this study was to investigate the possible involvement of toll-like receptors (TLRs) and their endogenous ligands in the induction/maintenance of the inflammatory process in patients’ BM.Besides, based on available evidence suggesting that the rs4986790 and rs4986791 Single Nucleotide Polymorphisms (SNP) of human TLR4 may be associated with increased susceptibility to bacterial infections, we sought to examine the possible association between TLR4 SNPs and infection susceptibility in MDS patients from a homogeneous Greek population.Methods: TLR expression was evaluated in the BM of patients and healthy controls using flow-cytometry. Quantitative PCR analysis of 84 genes related to TLR-mediated signal transduction was performed, using a commercially available PCR array, in immunomagnetically sorted CD14+ BM cells of patients and controls. The results were confirmed by testing separately three significantly over-expressed genes. The levels of various cytokines produced by patient monocytes treated with autologous plasma in the presence/absence of a TLR4 inhibitor were measured with chemiluminescence and the levels of the high mobility group box 1 (HMGB1) protein, a TLR4 endogenous ligand, in long-term BM culture supernatants were assayed with ELISA. To further examine if the excess of HMGB1 in the BM of MDS patients is due to macrophage dysfunction, a fluorescent microscopy-based assay was developed to estimate macrophage capacity to phagocytose apoptotic cells, and the time-course of HMGB1 release in a co-culture system of patient macrophages with different numbers of apoptotic cells was monitored.Genotyping for TLR4 rs4986790 and rs4986791 SNPs was performed by PCR – Restriction Fragment Length Polymorphisms (RFLPs) as described previously, using the NcoI and HinfI restriction enzymes, which digest the G allele-containing and T allele-containing PCR products of the rs4986790 and rs4986791 SNPs, respectively. To examine the possible correlation of the rs4986790 and rs4986791 polymorphic genotypes with MDS patient susceptibility to infections, we recorded the number of infections per patient tested, since the day of diagnosis.Results: A statistically significant increase in the proportion of TLR4+ cells within the CD14+ BM cells was observed in both patient groups compared to controls along with an up-regulation of TLR4 expression as was indicated by the TLR4 Μean Ratio of relative fluorescence intensity (MRFI) in patients. The TLR signaling gene array study in purified BM CD14+ cells showed that many of the TLR-related genes displayed at least two-fold increase in patients compared to controls. When examined separately, the relative mRNA expression of MyD88, TRIF/TICAM1 and TRAM/TICAM2, three adaptor molecules necessary for MyD88-dependent and MyD88-independent TLR4 signaling, was also found significantly increased in patients compared to controls confirming the results obtained from the PCR-arrays. Incubation of patient monocytes with autologous BM plasma resulted in the TLR4 dependent production of the cytokines measured, as been indicated by the significant decrease of IL-1, IL-6 and TNFα levels in the presence of the TLR4 inhibitor compared to cultures treated with the BM plasma alone. HMGB1 levels were significantly increased in long-term BM cultures of patients compared to controls suggesting that HMGB1 might constitute an endogenous TLR4-activating ligand in MDS and CIN BM. MDS-derived macrophages displayed impaired capacity to engulf apoptotic cells in comparison to healthy individuals. HMGB1 release by MDS BM macrophages loaded with increasing numbers of apoptotic cells for different time periods was dependent on the apoptotic cell load and incubation time.Regarding the genetic study, there was no statistically significant difference in the frequency of infectious episodes between MDS patients carrying the risk allele and the MDS patients that did not carry the risk allele for either SNP studied. However, it is interesting, that the frequency of the minor allele G of the rs4986790 SNP was found significantly increased in MDS patients compared to healthy controls. Furthermore, the heterozygous polymorphic genotype A/G seemed to be more frequent among MDS patients compared to controls. The frequency of the minor allele T and the polymorphic genotype C/T of rs4986791 SNP was also higher in MDS patients, although not at a statistically significant level.Conclusions: These data demonstrate a significant role of BM monocytes in the pathophysiology of CIN and MDS through the production of pro-inflammatory cytokines in a TLR4-mediated mechanism under the influence of endogenous ligands such as HMGB1. A primary apoptotic cell clearance defect of BM macrophages in MDS might contribute to the inflammatory process through aberrant release of HMGB1 from the late apoptotic/necrotic cells.The presence of the polymorphic genotype of rs4986790 does not seem to correlate with MDS patients’ vulnerability to infections. However, the frequency of the mutant allele G and the A/G genotype is significant higher in MDS patients compared to healthy controls, representing probably a genetically defined risk factor for MDS development

Myeloid-Derived Suppressor Cells: New Insights into the Pathogenesis and Therapy of MDS

Myelodysplastic syndromes (MDS) are hematopoietic malignancies characterized by the clonal expansion of hematopoietic stem cells, bone marrow failure manifested by cytopenias, and increased risk for evolving to acute myeloid leukemia. Despite the fact that the acquisition of somatic mutations is considered key for the initiation of the disease, the bone marrow microenvironment also plays significant roles in MDS by providing the right niche and even shaping the malignant clone. Aberrant immune responses are frequent in MDS and are implicated in many aspects of MDS pathogenesis. Recently, myeloid-derived suppressor cells (MDSCs) have gained attention for their possible implication in the immune dysregulation associated with MDS. Here, we summarize the key findings regarding the expansion of MDSCs in MDS, their role in MDS pathogenesis and immune dysregulation, as well their potential as a new therapeutic target for MDS

Αγωγή και προαγωγή υγείας σε μαθητές της πρωτοβάθμιας εκπαίδευση «Διαχείριση του στρες και εγκατάσταση υγιεινών συμπεριφορών δια της Πυθαγορείου Αυτογνωσίας»

Η παρούσα μελέτη αποτελεί μια παρεμβατική μελέτη ενός σκέλους η οποία διερευνά την αποτελεσματικότητα ενός ολιστικού προγράμματος (PSAI) για τη διαχείριση του στρες και εγκατάσταση υγιεινών συμπεριφορών σε παιδιά και εφήβους. Η έρευνα πραγματοποιήθηκε στην πρωτοβάθμια εκπαίδευση στο 4ο δημοτικό σχολείο Χαλανδρίου με διάρκεια δύο μήνες. Η παρέμβαση πραγματοποιήθηκε διαμέσου της τεχνικής της Πυθαγορείου Αυτογνωσίας η οποία αποτελεί μια τεχνική γνωστικής αναδόμησης με σκοπό τη διαχείριση του στρες, της ποιότητας του ύπνου και τη βελτίωση της μνήμης. Στο πρώτο κεφάλαιο αναφέρονται οι στρεσογόνοι παράγοντες με τους οπούς έρχονται αντιμέτωπα τα παιδιά στο σχολικό περιβάλλον. Πιο συγκεκριμένα παρουσιάζονται οι επιπτώσεις του στρες στην παιδική ηλικία και ο αντίκτυπος που έχουν στην ενήλικη ζωή. Στο δεύτερο κεφάλαιο περιγράφεται η τεχνική της Πυθαγορείου Αυτογνωσίας ,οι εβδομαδιαίες συνεδρίες, τα βήματα καθώς και τα οφέλη της τεχνικής. Στο τρίτο κεφάλαιο παρουσιάζονται η μεθοδολογία της έρευνας ,οι πρωταρχικοί -δευτερεύοντες στόχοι, η συλλογή των δεδομένων και οι μετρήσεις που πραγματοποιήθηκαν κατά τη διάρκεια του προγράμματος συμπεριλαμβανομένων τις σωματομετρήσεις και τα σταθμισμένα ψυχομετρικά εργαλεία. Επίσης γίνεται αναφορά στη διαδικασία του προγράμματος. Στο τέταρτο κεφάλαιο αναφέρεται η στάθμισή του ερωτηματολογίου που πραγματοποιήθηκε για τις ανάγκες της έρευνα. Στο πέμπτο κεφάλαιο παρουσιάζονται τα αποτελέσματα της έρευνας και παρατίθενται οι πίνακες. Στο έκτο κεφάλαιο γίνεται σύνοψη και συζήτηση επί των αποτελεσμάτων. Συστήνονται μελλοντικές προτάσεις για την διερεύνηση της αποτελεσματικότητας των ολιστικού και η αναγκαιότητα της προαγωγής της υγείας.Introduction: Stress is common in childhood and an important factor that affects behavior later in adulthood. The aim of this study was to assess the effects of a school-based stress-management intervention program on children’s stress. Materials and methods: This is a one-arm clinical trial-pilot study assessing the effects of the Pythagorean Self Awareness Intervention (PSAI), a novel cognitive-based stress management intervention on stress and anxiety, sleep quality, adherence in Mediterranean diet, and bullying behavior of children in primary school Results: Participants were 33 second-grade students. Self-report measures were applied for the evaluation of various variables during the beginning and the end of the 8-week monitoring period. There was a statistically significant reduction of stress levels, nightmares, body mass indexhair cortisol concentrations, bully scale, and Mediterranean diet quality. Daily routine and sleep were significantly improved. Hair cortisol concentrations increased. The social scale was significantly increased. Bedtime remained the same after the intervention. Conclusions: This pilot trial showed that the PSAI established healthy behaviors and lifestyle in children while it reduced their stress levels. Further research on the implementation of this holist program is suggeste

Changing Epidemiology of Invasive Candidiasis in Children during a 10-Year Period

Candida species are a common cause of invasive infection in neonates and children. The aim of our study was to evaluate the epidemiology and microbiology of invasive candidiasis (IC) in the largest tertiary Greek pediatric hospital during a 10-year period. A retrospective cohort study was performed from January 2008 to December 2017. Identification of species and antifungal susceptibility testing was performed according to the Clinical and Laboratory Standards Institute (CLSI) methodology. During the study period, 178 cases of IC were recorded. The tissue distribution included blood (87.1%), cerebrospinal (7.9%), peritoneal (3.9%) and pleural fluids (1.1%). Candida albicans and Candida parapsilosis (sensu lato) were the most frequently isolated species (47.8% and 28.7% respectively). From period 2008⁻2012 to period 2013⁻2017, a significant decrease in IC rates was detected (0.21 cases/1000 hospitalization days VS 0.11 cases/1000 hospitalization days, P = 0.040), while median minimum inhibitory concentrations (MICs) of amphotericin B were significantly increased for both C. albicans and C. parapsilosis (sl) (P = 0.037 and P = 0.004 respectively). The decrease in IC rates may reflect the increased awareness as well as the effective infection control initiatives and antifungal interventions. However, the significant increase in the MICs for amphotericin B and echinocandins such as caspofungin, raises concerns about their common use as first-line treatment. Epidemiologic monitoring is, therefore, critically important in order to evaluate and optimize therapeutic protocols for IC in pediatric populations

Reliability and validity of the Peer Relation Questionnaire in a sample of Greek school children and adolescents.

Bullying among children and adolescents can have detrimental effects on their physical and mental health. The first step in the identification of bullying is the detection of relevant signs and behaviours, with the use of appropriate instruments. The widely used Peer Relation Questionnaire (PRQ)has been developed to measure bullying behaviour in children and adolescents. The aim of this study was to translate and validate this instrument in the Greek language in a school sample of children and adolescents. A sample of 193 students (mean age ± SD 11.44 ± 1.26 years, girls 49.7%), was recruited from elementary and junior high schools of the public and private sector, in the capital of Athens, Greece. Along with the Peer Relation Questionnaire, the Stress in Children questionnaire and a structured inquiry regarding satisfaction from various aspects of life were used to assess the criterion validity of the Peer Relation Questionnaire. Results revealed good psychometric properties of the questionnaire in the Greek language. The principal component analysis resulted in two subscales that demonstrated an adequate internal consistency and a significant correlation with stress and satisfaction parameters, suggesting satisfying criterion validity. Meaningful associations between demographic characteristics and the subscales were also found. The Greek version of the Peer Relation Questionnaire-short form can be used as a valid tool to measure bullying behaviour in children and adolescents