DOCSLIB.ORG

Candida Glabrata and Candida Parapsilosis and Its Consequences on the Infection Process

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
Candida Glabrata and Candida Parapsilosis and Its Consequences on the Infection Process Variability of biofilm formation in Candida glabrata and Candida parapsilosis and its consequences on the infection process Dissertation In partial fulfillment of the requirements for the degree “Doctor of Philosophy (PhD)” at the Georg-August-Universität Göttingen within the “Biology” Doctoral program at the Georg-August University School of Science (GAUSS) submitted by Emilia Gómez Molero born in Albacete, Spain Göttingen, 2019 To my Family “Todo hombre puede ser, si se lo propone, el escultor de su propio cerebro” Dr. Santiago Ramón y Cajal (1852-1934) I Supervisors: Dr. rer. nat. Oliver Bader (Main Supervisor) (Institute for Medical Microbiology, Department of Medical Microbiology, University Medical Center Göttingen) Prof. Dr. med. Michael Weig (Second Supervisor) (Institute for Medical Microbiology, Department of Medical Microbiology, University Medical Center Göttingen) Members of the Thesis Committee: Prof. Dr. med. Uwe Groß (Institute for Medical Microbiology, Department of Medical Microbiology, University Medical Center Göttingen) Prof. Dr. Gerhard Braus (Institute for Microbiology and Genetics, Department of Molecular Microbiology and Genetics, University Göttingen) Dr. rer. nat. Oliver Bader (Supervisor) (Institute for Medical Microbiology, Department of Medical Microbiology, University Medical Center Göttingen) Members of the Examination Board: Prof. Dr. med. Uwe Groß (1st Reviewer) (Institute for Medical Microbiology, Department of Medical Microbiology, University Medical Center Göttingen) Prof. Dr. Gerhard Braus (2nd Reviewer) (Institute for Microbiology and Genetics, Department of Molecular Microbiology and Genetics, University Göttingen) Further Members of the Examination Board: Prof. Dr. rer. nat. Carsten G. K. Lüder (Institute for Medical Microbiology, Department of Medical Microbiology, University Medical Center Göttingen) PD Dr. Michael Hoppert (Institute for Microbiology and Genetics, Department of General Microbiology, University Göttingen) Prof. Dr. Fabian Commichau (Institute for Microbiology and Genetics, Department of General Microbiology, University Göttingen) Prof. Dr. Kai Heimel (Institute for Microbiology and Genetics, Department of Molecular Microbiology and Genetics, University Göttingen) Date of Disputation: II TABLE OF CONTENTS TABLE OF CONTENTS ........................................................................................................ III ACKNOWLEDGEMENTS ................................................................................................... VII LIST OF TABLES ................................................................................................................ XI LIST OF FIGURES .............................................................................................................. XII LIST OF ABREVIATIONS ................................................................................................... XV ABSTRACT ....................................................................................................................... XX 1. INTRODUCTION ................................................................................................... 1 1.1 Epidemiology of Candida spp. .................................................................................... 1 1.2 Features and phylogeny of the genus Candida ......................................................... 5 1.3 C. albicans infections and host-pathogen interactions ............................................. 7 1.4 Candida biofilms ....................................................................................................... 10 1.5 The fungal cell wall ................................................................................................... 13 1.6 Candida spp. cell wall proteins (CWP) ..................................................................... 14 1.7 GPI-anchored Cell Wall proteins (adhesins) ............................................................ 15 1.8 Candida glabrata ...................................................................................................... 18 1.8.1 Candida glabrata cell wall proteins ......................................................................... 20 1.9 Candida parapsilosis ................................................................................................ 21 1.9.1 Candida parapsilosis cell wall proteins .................................................................... 22 1.10 Candida species interaction in host invasion ........................................................... 23 1.11 Aims of the study ..................................................................................................... 25 2. MATERIALS AND METHODS ............................................................................... 26 2.1 Materials .................................................................................................................. 26 2.1.1 Synthetic oligonucleotides ....................................................................................... 30 2.1.2 Candida spp. clinical isolates .................................................................................... 30 2.1.3 Candida spp. reference strains ................................................................................ 31 2.2 Methods ................................................................................................................... 32 2.2.1 Routine diagnostic procedures ................................................................................ 32 2.2.2 Candida spp. strain maintenance and growth conditions ....................................... 32 2.2.3 MALDI-TOF analyses species-identification ............................................................. 33 2.2.4 C. glabrata biofilm formation capacity to polystyrol ............................................... 33 2.2.5 C. glabrata biofilm formation capacity to silicone elastomers ................................ 35 III 2.2.6 C. parapsilosis biofilm formation capacity to polystyrol .......................................... 36 2.2.7 Antifungal susceptibility test .................................................................................... 38 2.2.8 Agar invasion capacity .............................................................................................. 39 2.2.9 Optical and phase-contrast microscopy .................................................................. 39 2.2.10 Sedimentation assay of clinical morphotypes ......................................................... 40 2.2.11 Electrophoretic karyotyping of colony morphotypes .............................................. 40 2.2.12 Genomic DNA isolation ............................................................................................ 41 2.2.13 Polymerase chain reaction analyses of adhesin-encoding genes in clinical morphotypes ............................................................................................................ 42 2.2.14 DNA Sanger sequencing ........................................................................................... 43 2.2.15 Candida cell wall isolation ........................................................................................ 43 2.2.16 Candida spp. genome sequencing analyses ............................................................. 44 2.2.17 In vivo infection analyses. Galleria mellonella animal model .................................. 44 2.2.18 C. albicans and C. glabrata identification from patient blood cultures .................. 45 2.2.19 C. albicans and C. glabrata interaction under phase-contrast microscopy ............. 45 3. RESULTS ............................................................................................................ 47 3.1 Genomic plasticity enhances phenotypic diversity in Candida glabrata clinical isolates from different clades .................................................................................. 47 3.1.1 Changes in the genome of hyper-biofilm-forming C. glabrata isolates .................. 47 3.1.2 Genomic changes between time-resolved C. glabrata isolates .............................. 53 3.2 Phenotypic variability in C. glabrata clinical isolates correlates with variations in cell wall proteome and infection-relevant parameters ........................................... 55 3.2.1 Biofilm formation capacity to polystyrol of a large C. glabrata clinical strain collection .................................................................................................................. 58 3.2.2 Virulence of C. glabrata isolates in the Galleria mellonella model ......................... 59 3.2.3 Variations in C. glabrata biofilm formation capacity to silicone elastomers .......... 62 3.2.4 Different adhesins are present in hyper biofilm-forming clinical isolates ............... 64 3.2.5 Hyper biofilm-forming C. glabrata clinical isolates’ adherence to C. albicans. ....... 69 3.2.6 C. albicans and C. glabrata mixed culture analysis .................................................. 71 3.3 Pheno- and genotypic analyses of Candida parapsilosis clinical isolates ................ 76 3.3.1 Establishing a classification reference ..................................................................... 76 IV 3.4 Candida parapsilosis polymorphism drives differences at morphological and genotypic level in isolates from a single patient ...................................................... 80 3.4.1 The five clinical isolates are distributed into
Load more

Recommended publications
  • Evolution and Application of Inteins in Candida Species: a Review
    fmicb-07-01585 October 6, 2016 Time: 13:7 # 1 View metadata, citation and similar papers at core.ac.uk brought to you by CORE provided by Frontiers - Publisher Connector REVIEW published: 10 October 2016 doi: 10.3389/fmicb.2016.01585 Evolution and Application of Inteins in Candida species: A Review José A. L. Fernandes1†, Tâmara H. R. Prandini2†, Maria da Conceição A. Castro1, Thales D. Arantes1,3, Juliana Giacobino2, Eduardo Bagagli2 and Raquel C. Theodoro1* 1 Institute of Tropical Medicine of Rio Grande do Norte, Universidade Federal do Rio Grande do Norte, Natal, Brazil, 2 Department of Microbiology and Immunology, Institute of Biosciences, Universidade Estadual Paulista Julio de Mesquita Filho, Botucatu, Brazil, 3 Post-graduation Program in Biochemistry, Universidade Federal do Rio Grande do Norte, Natal, Brazil Inteins are invasive intervening sequences that perform an autocatalytic splicing from their host proteins. Among eukaryotes, these elements are present in many fungal species, including those considered opportunistic or primary pathogens, such as Candida spp. Here we reviewed and updated the list of Candida species containing inteins in the genes VMA, THRRS and GLT1 and pointed out the importance of these elements as molecular markers for molecular epidemiological researches and species- specific diagnosis, since the presence, as well as the size of these inteins, is polymorphic Edited by: among the different species. Although absent in Candida albicans, these elements are Joshua D. Nosanchuk, present in different sizes, in some environmental Candida spp. and also in most of the Albert Einstein College of Medicine, USA non-albicans Candida spp. considered emergent opportunistic pathogens.
    [Show full text]
  • Genome Diversity and Evolution in the Budding Yeasts (Saccharomycotina)
    | YEASTBOOK GENOME ORGANIZATION AND INTEGRITY Genome Diversity and Evolution in the Budding Yeasts (Saccharomycotina) Bernard A. Dujon*,†,1 and Edward J. Louis‡,§ *Department Genomes and Genetics, Institut Pasteur, Centre National de la Recherche Scientifique UMR3525, 75724-CEDEX15 Paris, France, †University Pierre and Marie Curie UFR927, 75005 Paris, France, ‡Centre for Genetic Architecture of Complex Traits, and xDepartment of Genetics, University of Leicester, LE1 7RH, United Kingdom ORCID ID: 0000-0003-1157-3608 (E.J.L.) ABSTRACT Considerable progress in our understanding of yeast genomes and their evolution has been made over the last decade with the sequencing, analysis, and comparisons of numerous species, strains, or isolates of diverse origins. The role played by yeasts in natural environments as well as in artificial manufactures, combined with the importance of some species as model experimental systems sustained this effort. At the same time, their enormous evolutionary diversity (there are yeast species in every subphylum of Dikarya) sparked curiosity but necessitated further efforts to obtain appropriate reference genomes. Today, yeast genomes have been very informative about basic mechanisms of evolution, speciation, hybridization, domestication, as well as about the molecular machineries underlying them. They are also irreplaceable to investigate in detail the complex relationship between genotypes and phenotypes with both theoretical and practical implications. This review examines these questions at two distinct levels offered by the broad evolutionary range of yeasts: inside the best-studied Saccharomyces species complex, and across the entire and diversified subphylum of Saccharomycotina. While obviously revealing evolutionary histories at different scales, data converge to a remarkably coherent picture in which one can estimate the relative importance of intrinsic genome dynamics, including gene birth and loss, vs.
    [Show full text]
  • 10-ELS-OXF Kurtzman1610423 CH002 7..20
    Part II Importance of Yeasts Kurtzman 978-0-444-52149-1 00002 Kurtzman 978-0-444-52149-1 00002 Chapter 2 c0002 Yeasts Pathogenic to Humans Chester R. Cooper, Jr. regularly encounter the organisms described below. In fact, many s0010 1. INTRODUCTION TO THE MEDICALLY medical mycologists spend entire careers without direct clinical expo- IMPORTANT YEASTS sure to many of these fungi. Rather, the purpose of this review is to enlighten the non-medical mycologist as to the diversity of yeast and p0010 Prior to global emergence of the human immunodeficiency virus mold species regularly associated with human and animal disease (HIV), which is the causative agent of acquired immunodeficiency that also, at least in part, present a unicellular mode of growth in vivo. syndrome (AIDS), approximately 200 fungal pathogens were recog- The following descriptions present a concise overview of the key p0025 nized from among the more than 100,000 then-known fungal spe- biological and clinical features of these fungi. Where appropriate, refer- cies (Kwon-Chung and Bennett 1992, Rippon 1988). About 50 of ences to recent reviews of particular disease agents and their patholo- these species were regularly associated with fungal disease (myco- gies are provided. For a global perspective of fungal diseases, including sis). Since then, there has been a concurrent dramatic increase in in-depth clinical discussions of specific pathologies, diagnoses, and both the number of known fungal species and the incidence of treatments, the reader is referred to several outstanding and recently mycoses that they cause. Moreover, the spectrum of pathogenic fungi published texts (Anaissie et al.
    [Show full text]
  • Candida Parapsilosis: a Review of Its Epidemiology, Pathogenesis, Clinical Aspects, Typing and Antimicrobial Susceptibility
    Critical Reviews in Microbiology Critical Reviews in Microbiology, 2009; 35(4): 283–309 2009 REVIEW ARTICLE Candida parapsilosis: a review of its epidemiology, pathogenesis, clinical aspects, typing and antimicrobial susceptibility Eveline C. van Asbeck1,2, Karl V. Clemons1, David A. Stevens1 1Division of Infectious Diseases, Santa Clara Valley Medical Center, and California Institute for Medical Research, San Jose, CA 95128 USA and Division of Infectious Diseases and Geographic Medicine, Stanford University, Stanford, CA 94305, and 2Eijkman-Winkler Institute for Medical and Clinical Microbiology, University Medical Center Utrecht, Utrecht, The Netherlands Abstract The Candida parapsilosis family has emerged as a major opportunistic and nosocomial pathogen. It causes multifaceted pathology in immuno-compromised and normal hosts, notably low birth weight neonates. Its emergence may relate to an ability to colonize the skin, proliferate in glucose-containing solutions, and adhere to plastic. When clusters appear, determination of genetic relatedness among strains and identifica- tion of a common source are important. Its virulence appears associated with a capacity to produce biofilm and production of phospholipase and aspartyl protease. Further investigations of the host-pathogen inter- actions are needed. This review summarizes basic science, clinical and experimental information about C. parapsilosis. Keywords: Candida parapsilosis, epidermiology, strain differentiation, clinical aspects, pathogenesis, For personal use only. antifungal susceptibility Introduction The organism was first described in 1928 (Ashford 1928), and early reports of C. parapsilosis described the organ- Candida bloodstream infections (BSI) remain an ism as a relatively non-pathogenic yeast in the normal exceedingly common life-threatening fungal disease flora of healthy individuals that was of minor clinical and are now recognized as a major cause of hospital- significance (Weems 1992).
    [Show full text]
  • Candida Parapsilosis Complex Francesco Barchiesi1* , Elena Orsetti1, Patrizia Osimani3, Carlo Catassi2, Fabio Santelli4 and Esther Manso5
    Barchiesi et al. BMC Infectious Diseases (2016) 16:387 DOI 10.1186/s12879-016-1704-y RESEARCH ARTICLE Open Access Factors related to outcome of bloodstream infections due to Candida parapsilosis complex Francesco Barchiesi1* , Elena Orsetti1, Patrizia Osimani3, Carlo Catassi2, Fabio Santelli4 and Esther Manso5 Abstract Background: Although Candida albicans is the most common cause of fungal blood stream infections (BSIs), infections due to Candida species other than C. albicans are rising. Candida parapsilosis complex has emerged as an important fungal pathogen and became one of the main causes of fungemia in specific geographical areas. We analyzed the factors related to outcome of candidemia due to C. parapsilosis in a single tertiary referral hospital over a five-year period. Methods: A retrospective observational study of all cases of candidemia was carried out at a 980-bedded University Hospital in Italy. Data regarding demographic characteristics and clinical risk factors were collected from the patient’s medical records. Antifungal susceptibility testing was performed and MIC results were interpreted according to CLSI species-specific clinical breakpoints. Results: Of 270 patients diagnosed with Candida BSIs during the study period, 63 (23 %) were infected with isolates of C. parapsilosis complex which represented the second most frequently isolated yeast after C. albicans. The overall incidence rate was 0.4 episodes/1000 hospital admissions. All the strains were in vitro susceptible to all antifungal agents. The overall crude mortality at 30 days was 27 % (17/63), which was significantly lower than that reported for C. albicans BSIs (42 % [61/146], p = 0.042). Being hospitalized in ICU resulted independently associated with a significant higher risk of mortality (HR 4.625 [CI95% 1.015–21.080], p = 0.048).
    [Show full text]
  • Nails: Tales, Fails and What Prevails in Treating Onychomycosis
    J. Hibler, D.O. OhioHealth - O’Bleness Memorial Hospital, Athens, Ohio AOCD Annual Conference Orlando, Florida 10.18.15 A) Onychodystrophy B) Onychogryphosis C)“Question Onychomycosis dogma” – Michael Conroy, MD D) All the above E) None of the above Nail development begins at 8-10 weeks EGA Complete by 5th month Keratinization ~11 weeks No granular layer Nail plate growth: Fingernails 3 mm/month, toenails 1 mm/month Faster in summer or winter? Summer! Index finger or 5th digit nail grows faster? Index finger! Faster growth to middle or lateral edge of each nail? Lateral! Elkonyxis Mee’s lines Aka leukonychia striata Arsenic poisoning Trauma Medications Illness Psoriasis flare Muerhrcke’s bands Hypoalbuminemia Chemotherapy Half & half nails Aka Lindsay’s nails Chronic renal disease Terry’s nails Liver failure, Cirrhosis Malnutrition Diabetes Cardiovascular disease True or False: Onychomycosis = Tinea Unguium? FALSE. Onychomycosis: A fungal disease of the nails (all causes) Dermatophytes, yeasts, molds Tinea unguium: A fungal disease of nail caused by dermatophyte fungi Onychodystrophy ≠ onychomycosis Accounts for up to 50% of all nail disorders Prevalence; 14-28% of > 60 year-olds Variety of subtypes; know them! Sequelae What is the most common cause of onychomycosis? A) Epidermophyton floccosum B) Microsporum spp C) Trichophyton mentagrophytes D) Trichophyton rubrum -Account for ~90% of infections Dermatophytes Trichophyton rubrum Trichophyton mentagrophytes Trichophyton tonsurans, Microsporum canis, Epidermophyton floccosum Nondermatophyte molds Acremonium spp, Fusarium spp Scopulariopsis spp, Sytalidium spp, Aspergillus spp Yeast Candida parapsilosis Candida albicans Candida spp Distal/lateral subungal Proximal subungual onychomycosis onychomycosis (DLSO) (PSO) Most common; T. rubrum Often in immunosuppressed patients T.
    [Show full text]
  • Candida Species – Morphology, Medical Aspects and Pathogenic Spectrum
    European Journal of Molecular & Clinical Medicine ISSN 2515-8260 Volume 07, Issue 07, 2020 Candida Species – Morphology, Medical Aspects And Pathogenic Spectrum. Shubham Koundal1, Louis Cojandaraj2 1 Assistant Professor, Department of Medical Laboratory Sciences, Chandigarh University, Punjab. 2Assistant Professor, Department of Medical Laboratory Sciences, Lovely Professional University, Punjab. Email Id: [email protected] ABSTRACT Emergence of candidal infections are increasing from decades and found to be a leading cause of human disease and mortality. Candida spp. is one of the communal of human body and is known to cause opportunistic superficial and invasive infections. Many of mycoses-related deaths were due to Candida spp. Major shift of Candida infection towards NAC (non-albicans Candida) is matter of concern worldwide. In this study we had given a systemic review about medically important Candidaspp. Along with their morphological features, treatment and drugs. Spectrum of the pathogen is also discussed. Morphology of Different Medically Important Candida Species with their medical aspects along and pathogenic spectrum. Corn meal agar morphology along with anti-candida drugs has been discussed. The study is done after considering various published review’s and the mycological studies. Key words: Candida, Yeast, C.albicans, C. tropicalis, C. parapsilosis, C. glabrata, C. krusei and C. lusitaniae 1. INTRODUCTION Yeasts are unicellular, sometimes dimorphic fungi. It can give rise to wide range of infections in humans commonly called fungal infections. Yeast infections varies from superficial cutaneous/skin infections, mucosa related infections to multi-organ disseminated infections.(Sardi et al., 2013)Cutaneous and mucosal yeast infections can infect a number of regions in human body including the skin, nails, oral cavity, gastrointestinal tract, female genital tract and esophageal part and lead to chronic nature.
    [Show full text]
  • Candida Species (Volatile) Metabotyping Through Advanced Comprehensive Two-Dimensional Gas Chromatography
    microorganisms Article Candida Species (Volatile) Metabotyping through Advanced Comprehensive Two-Dimensional Gas Chromatography Carina Pedrosa Costa 1 , Ana Rita Bezerra 2, Adelaide Almeida 3,* and Sílvia M. Rocha 1,* 1 Department of Chemistry & LAQV-REQUIMTE, University of Aveiro, Campus Universitário Santiago, 3810-193 Aveiro, Portugal; [email protected] 2 Health Sciences Department, Institute for Biomedicine—iBiMED, University of Aveiro, Campus Universitário Santiago, 3810-193 Aveiro, Portugal; [email protected] 3 Department of Biology & CESAM, University of Aveiro, Campus Universitário Santiago, 3810-193 Aveiro, Portugal * Correspondence: [email protected] (A.A.); [email protected] (S.M.R.); Tel.: +351-234-370784 (A.A.); +351-234-401524 (S.M.R.) Received: 5 November 2020; Accepted: 29 November 2020; Published: 30 November 2020 Abstract: Microbial metabolomics is a challenge strategy that allows a comprehensive analysis of metabolites within a microorganism and may support a new approach in microbial research, including the microbial diagnosis. Thus, the aim of this research was to in-depth explore a metabolomics strategy based on the use of an advanced multidimensional gas chromatography for the comprehensive mapping of cellular metabolites of C. albicans and non-C. albicans (C. glabrata and C. tropicalis) and therefore contributing for the development of a comprehensive platform for fungal detection management and for species distinction in early growth times (6 h). The volatile fraction comprises 126 putatively identified metabolites distributed over several chemical families: acids, alcohols, aldehydes, hydrocarbons, esters, ketones, monoterpenic and sesquiterpenic compounds, norisoprenoids, phenols and sulphur compounds. These metabolites may be related with different metabolic pathways, such as amino acid metabolism and biosynthesis, fatty acids metabolism, aromatic compounds degradation, mono and sesquiterpenoid synthesis and carotenoid cleavage.
    [Show full text]
  • The Quest for a General and Reliable Fungal DNA Barcode
    The Open Applied Informatics Journal, 2011, 5, (Suppl 1-M6) 45-61 45 Open Access The Quest for a General and Reliable Fungal DNA Barcode Vincent Robert*,1, Szaniszlo Szöke1, Ursula Eberhardt1, Gianluigi Cardinali2, Wieland Meyer3, Keith A. Seifert4, C. Andre Lévesque4 and Chris T. Lewis4 1CBS-KNAW Fungal Biodiversity Centre, Utrecht, The Netherlands 2Dipartimento Biologia Applicata- Microbiologia, Università degli Studi di Perugia, Perugia, Italy 3Molecular Mycology Research Laboratory, CIDM, Westmead Millennium Institute, SEIB, Sydney Medical School - Westmead Hospital, The University of Sydney, Sydney, Australia 4Biodiversity (Mycology & Botany), Eastern Cereal and Oilseed Research Centre, Agriculture & Agri-Food Canada, Ottawa, Canada Abstract: DNA sequences are key elements for both identification and classification of living organisms. Mainly for historical reasons, a limited number of genes are currently used for this purpose. From a mathematical point of view, any DNA segment, at any location, even outside of coding regions and even if they do not align, could be used as long as PCR primers could be designed to amplify them. This paper describes two methods to search genomic data for the most efficient DNA segments that can be used for identification and classification. Keywords: Genome, molecular, sequences, barcoding, identification, classification, fungi. 1. INTRODUCTION When first large molecular phylogenetic studies were completed, it was obvious that many clades were poorly Since the early days of classification, taxonomists have supported statistically when only one or two genes were struggled with the available information and characteristics used. Recently, several authors explored possibilities for of their organisms of interest to develop systems that reflect analyzing several genes to obtain the true phylogeny [1-7].
    [Show full text]
  • Fungal Infections in Gastroenterology
    Current Trends in Gastroenterology L UPINE PUBLISHERS and Hepatology Open Access DOI: 10.32474/CTGH.2018.01.000114 ISSN: 2641-1652 Review article Fungal Infections in Gastroenterology Michael AB Naafs MD* MD, Endocrinologist, Health Consultant at Naafs, International Health Consultancy, Rhodoslaan, Oldenzaal, The Netherlands Received: August 13, 2018; Published: August 20, 2018 *Corresponding author: Michael AB Naafs, Dutch Internist Endocrinologist, Health Consultant at Naafs, International Health Consultancy, Rhodoslaan 20,7577KN, Oldenzaal, The Netherlands Abstract Fungal infections are increasing in gastroenterology. Cirrhotic patients, liver transplantation recipients, and patients with The advent of sequencing technology can identify now previously unculturable fungi. Dysbiosis of the mycobiome and microbiome inflammatory bowel disease (IBD) are vulnerable to these infections. The origin of these fungal infections is frequently the GI tract. can uncheck normal commensal fungi and turn pathogenic by largely unknown mechanisms Resistance to antifungals is becoming a global problem. New antifungals are in development and they are badly needed. In this mini-review the enteric mycobiota and fungal infections in GI disease are discussed. Introduction to be increased in Crohn’s disease (CD) patients [14-17]. Most Fungi are normally present in the gastrointestinal tract as part Candida ssp. can also be detected in the feces or intestinal mucosa of the gut microbiome. With the advent of sequencing technology of patients with ulcerative colitis (UC)- [18,19]. To characterize the microbes can be studied now that were previous non-culturable. mycebiota by sequencing variable portions of the fungi genome are Fungal species, also called the “silent population” are known as targeted, which characterize the fungi to the genes level, such as the mycobiome [1].
    [Show full text]
  • Molecular Markers of Antifungal Resistance: Potential Uses in Routine Practice and Future Perspectives
    Journal of Fungi Review Molecular Markers of Antifungal Resistance: Potential Uses in Routine Practice and Future Perspectives Guillermo Garcia-Effron 1,2 1 Laboratorio de Micología y Diagnóstico Molecular, Cátedra de Parasitología y Micología, Facultad de Bioquímica y Ciencias Biológicas, Universidad Nacional del Litoral, Santa Fe CP3000, Argentina; [email protected]; Tel.: +54-9342-4575209 (ext. 135) 2 Consejo Nacional de Investigaciones Científicas y Tecnológicas, Santa Fe CP3000, Argentina Abstract: Antifungal susceptibility testing (AST) has come to establish itself as a mandatory routine in clinical practice. At the same time, the mycological diagnosis seems to have headed in the direction of non-culture-based methodologies. The downside of these developments is that the strains that cause these infections are not able to be studied for their sensitivity to antifungals. Therefore, at present, the mycological diagnosis is correctly based on laboratory evidence, but the antifungal treatment is undergoing a growing tendency to revert back to being empirical, as it was in the last century. One of the explored options to circumvent these problems is to couple non-cultured based diagnostics with molecular-based detection of intrinsically resistant organisms and the identification of molecular mechanisms of resistance (secondary resistance). The aim of this work is to review the available molecular tools for antifungal resistance detection, their limitations, and their advantages. A comprehensive description of commercially available and in-house methods is included. In addition, gaps in the development of these molecular technologies are discussed. Citation: Garcia-Effron, G. Molecular Keywords: antifungal resistance; molecular tools; intrinsic resistance; secondary resistance; Cyp51A; Markers of Antifungal Resistance: FKS; Candida; Aspergillus Potential Uses in Routine Practice and Future Perspectives.
    [Show full text]
  • Validation of a Simplex PCR Assay Enabling Reliable Identification Of
    Fidler et al. BMC Infectious Diseases (2018) 18:393 https://doi.org/10.1186/s12879-018-3283-6 RESEARCH ARTICLE Open Access Validation of a simplex PCR assay enabling reliable identification of clinically relevant Candida species Gabor Fidler1, Eva Leiter2, Sandor Kocsube3, Sandor Biro1 and Melinda Paholcsek1* Abstract Background: Fungal bloodstream infections (BSI) may be serious and are associated with drastic rise in mortality and health care costs. Candida spp. are the predominant etiological agent of fungal sepsis. The prompt and species-level identification of Candida may influence patient outcome and survival. The aim of this study was to develop and evaluate the CanTub-simplex PCR assay coupled with Tm calling and subsequent high resolution melting (HRM) analysis to barcode seven clinically relevant Candida species. Methods: Efficiency, coefficient of correlation and the limit of reliable detection were estimated on purified Candida EDTA-whole blood (WB) reference panels seeded with Candida albicans, Candida glabrata, Candida parapsilosis, Candida tropicalis, Candida krusei, Candida guilliermondii, Candida dubliniensis cells in a 6-log range. Discriminatory power was measured on EDTA-WB clinical panels on three different PCR platforms; LightCycler®96, LightCycler® Nano, LightCycler® 2.0. Inter- and intra assay consistencies were also calculated. Results: The limit of reliable detection proved to be 0.2–2 genomic equivalent and the method was reliable on broad concentration ranges (106–10 CFU) providing distinctive melting peaks and characteristic HRM curves. The diagnostic accuracy of the discrimination proved to be the best on Roche LightCycler®2.0 platform. Repeatability was tested and proved to be % C.V.: 0.14 ± 0.06 on reference- and % C.V.: 0.14 ± 0.02 on clinical-plates accounting for a very high accuracy.
    [Show full text]