Generation And Characterization Of Candida Parapsilosis Over Expression Mutant Strains
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Generation And Characterization Of Candida Parapsilosis Over Expression Mutant Strains
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Author : S.E. Pu00e1l
language : en
Publisher:
Release Date : 2017
Generation And Characterization Of Candida Parapsilosis Over Expression Mutant Strains written by S.E. Pu00e1l and has been published by this book supported file pdf, txt, epub, kindle and other format this book has been release on 2017 with categories.
Candida parapsilosis is an opportunistic human fungal pathogen, threatening a wide range of patient group ranging from low birth-weight neonates to the critically ill and severely immune suppressed patients treated at intensive care units. Furthermore, C. parapsilosis infection rates can even exceed the candidaemia episodes caused by C. albicans at certain geographical regions. Therefore, excessive research is now focused on the investigation of C. parapsilosisu2019 pathogenesis. In order to search for genes potentially influencing the virulence of C. parapsilosis, we aimed to generate and characterize over-expression mutant strains of ORFs that, according to our preliminary experiments, are thought to influence fungal pathogenesis.To identify such set of genes, transcriptional analysis of fungi co-incubated with THP-1 human monocytes was performed. Differentially expressed kinases and transcriptional factors and additional orthologous genes of C. albicans virulence regulatory ORFs (adherence, biofilm formation, antifungal resistance) were sentenced to over-expression. We used the Thermo Fisher Scientific Gateway cloning technology to create over-expression mutant strains. Transformants were validated by colony-PCR, Southern blot and real-time PCR. The fitness of the verified mutants was monitored in vitro at various temperatures and under conditions that are thought to influence the virulence of C. parapsilosis. These included different pH conditions, minimal and complex media and also solid media supplemented with osmotic, oxidative or cell wall stressors. Fitness under metal iron limiting conditions was also examined.During our work, we successfully generated 36 over-expression mutant strains with 2 parallels and added unique 20-mer nucleotide barcode sequences to each in order to be able to identify them during later co-infection studies. Integration of the over-expression constructs was confirmed by both colony PCR and real-time PCR. To exclude the possibility of ectopic integrations, Southern-blot analysis was also applied. We confirmed that all mutant strains contained only one copy of the integrating constitutive promoter containing over-expression cassette. Thereafter, all available mutant strainsu2019 viability was examined under the aforementioned conditions. During the characterization, altered phenotypes were detected mainly in the presence of SDS, Calcofluor white and Congo Red. Under these conditions, mutants showed growth disabilities when compared to the parental strain. In this study, we successfully generated 36 C. parapsilosis over-expression mutant strains. According to our results, several of these showed a phenotype different from the wild type under conditions that are thought to impact the pathogenicity of this species. In order to supplement this hypothesis we further aim to perform virulence studies including both in vitro and in vivo infection models.This project was funded by NKFIH K123952 and GINOP-2.3.2-15-2016-00015.
Generation And Characterization Of Candida Parapsilosis Over Expression Mutant Strains
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Author : Sára E. Pál
language : en
Publisher:
Release Date : 2017
Generation And Characterization Of Candida Parapsilosis Over Expression Mutant Strains written by Sára E. Pál and has been published by this book supported file pdf, txt, epub, kindle and other format this book has been release on 2017 with categories.
Candida parapsilosis is an opportunistic human fungal pathogen, threatening a wide range of patient group ranging from low birth-weight neonates to the critically ill and severely immune suppressed patients treated at intensive care units. Furthermore, C. parapsilosis infection rates can even exceed the candidaemia episodes caused by C. albicans at certain geographical regions. Therefore, excessive research is now focused on the investigation of C. parapsilosisu2019 pathogenesis. In order to search for genes potentially influencing the virulence of C. parapsilosis, we aimed to generate and characterize over-expression mutant strains of ORFs that, according to our preliminary experiments, are thought to influence fungal pathogenesis.To identify such set of genes, transcriptional analysis of fungi co-incubated with THP-1 human monocytes was performed. Differentially expressed kinases and transcriptional factors and additional orthologous genes of C. albicans virulence regulatory ORFs (adherence, biofilm formation, antifungal resistance) were sentenced to over-expression. We used the Thermo Fisher Scientific Gateway cloning technology to create over-expression mutant strains. Transformants were validated by colony-PCR, Southern blot and real-time PCR. The fitness of the verified mutants was monitored in vitro at various temperatures and under conditions that are thought to influence the virulence of C. parapsilosis. These included different pH conditions, minimal and complex media and also solid media supplemented with osmotic, oxidative or cell wall stressors. Fitness under metal iron limiting conditions was also examined.During our work, we successfully generated 36 over-expression mutant strains with 2 parallels and added unique 20-mer nucleotide barcode sequences to each in order to be able to identify them during later co-infection studies. Integration of the over-expression constructs was confirmed by both colony PCR and real-time PCR. To exclude the possibility of ectopic integrations, Southern-blot analysis was also applied. We confirmed that all mutant strains contained only one copy of the integrating constitutive promoter containing over-expression cassette. Thereafter, all available mutant strainsu2019 viability was examined under the aforementioned conditions. During the characterization, altered phenotypes were detected mainly in the presence of SDS, Calcofluor white and Congo Red. Under these conditions, mutants showed growth disabilities when compared to the parental strain. In this study, we successfully generated 36 C. parapsilosis over-expression mutant strains. According to our results, several of these showed a phenotype different from the wild type under conditions that are thought to impact the pathogenicity of this species. In order to supplement this hypothesis we further aim to perform virulence studies including both in vitro and in vivo infection models.This project was funded by NKFIH K123952 and GINOP-2.3.2-15-2016-00015.
Drosophila Protocols
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Author : William Sullivan
language : en
Publisher: CSHL Press
Release Date : 2000
Drosophila Protocols written by William Sullivan and has been published by CSHL Press this book supported file pdf, txt, epub, kindle and other format this book has been release on 2000 with Medical categories.
This exceptional laboratory manual describes thirty-seven procedures most likely to be used in the next decade for molecular, biochemical, and cellular studies on Drosophila. They were selected after extensive consultation with the research community and rigorously edited for clarity, uniformity, and conciseness.The methods included permit investigation of chromosomes, cell biology, molecular biology, genomes, biochemistry, and development. Each protocol includes the basic information needed by novices, with sufficient detail to be valuable to experienced investigators. Each method is carefully introduced and illustrated with figures, tables, illustrations, and examples of the data obtainable. The book's appendices include key aspects of Drosophila biology, essential solutions, buffers, and recipes.An evolution of Michael Ashburner's 1989 classic Drosophila: A Laboratory Manual, this book is an essential addition to the personal library of Drosophila investigators and an incomparable resource for other research groups with goals likely to require fly-based technical approaches.
The Fungal Kingdom
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Author : Joseph Heitman
language : en
Publisher: John Wiley & Sons
Release Date : 2020-07-10
The Fungal Kingdom written by Joseph Heitman and has been published by John Wiley & Sons this book supported file pdf, txt, epub, kindle and other format this book has been release on 2020-07-10 with Medical categories.
Fungi research and knowledge grew rapidly following recent advances in genetics and genomics. This book synthesizes new knowledge with existing information to stimulate new scientific questions and propel fungal scientists on to the next stages of research. This book is a comprehensive guide on fungi, environmental sensing, genetics, genomics, interactions with microbes, plants, insects, and humans, technological applications, and natural product development.
Candida And Candidiasis
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Author : Richard A. Calderone
language : en
Publisher: American Society for Microbiology Press
Release Date : 2011-12-07
Candida And Candidiasis written by Richard A. Calderone and has been published by American Society for Microbiology Press this book supported file pdf, txt, epub, kindle and other format this book has been release on 2011-12-07 with Medical categories.
The underlying mechanisms of Candida and candidiasis and promising new directions in drug discovery and treatment. • Reviews all aspects of this common fungal pathogen and its impact on human health, from the basic biology of Candida albicans to the clinical management of candidiasis. • Reviews the latest basic and clinical research, focusing on findings in genome variability, host-pathogen interactions, antifungal resistance and drug discovery, and diagnostics to foster better understanding and treatment of candidiasis. • Examines recent discoveries that have shed light on morphogenesis and the cell cycle, including how new findings on host responses may have applications for the diagnosis of blood-borne candidiasis.
Non Conventional Yeasts In Genetics Biochemistry And Biotechnology
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Author : Klaus Wolf
language : en
Publisher: Springer Science & Business Media
Release Date : 2003-03-04
Non Conventional Yeasts In Genetics Biochemistry And Biotechnology written by Klaus Wolf and has been published by Springer Science & Business Media this book supported file pdf, txt, epub, kindle and other format this book has been release on 2003-03-04 with Medical categories.
Most information on yeasts derives from experiments with the conventional yeasts Saccaromyces cerevisiae and Schizossaccharomyces pombe, the complete nuclear and mitochondrial genome of which has also been sequenced. For all other non-conventional yeasts, investigations are in progress and the rapid development of molecular techniques has allowed an insight also into a variety of non-conventional yeasts. In this bench manual, over 70 practical protocols using 15 different non-conventional yeast species and in addition several protocols of general use are described in detail. All of these experiments on the genetics, biochemistry and biotechnology of yeasts have been contributed by renowned laboratories and have been reproduced many times. The reliable protocols are thus ideally suited also for undergraduate and graduate practical courses.
Pathogenic Yeasts
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Author : Ruth Ashbee
language : en
Publisher: Springer Science & Business Media
Release Date : 2009-10-27
Pathogenic Yeasts written by Ruth Ashbee and has been published by Springer Science & Business Media this book supported file pdf, txt, epub, kindle and other format this book has been release on 2009-10-27 with Science categories.
Mycological studies of yeasts are entering a new phase, with the sequencing of multiple fungal genomes informing our understanding of their ability to cause disease and interact with the host. At the same time, the ongoing use of traditional methods in many clinical mycology laboratories continues to provide information for the diagnosis and treatment of patients. This volume reviews various aspects of pathogenic yeasts and what is known about their molecular and cellular biology and virulence, in addition to looking at clinical and laboratory findings. As each chapter is written by a leading expert in the field, this book summarizes in one volume much of the latest research on several pathogenic yeasts, including Candida, Cryptococcus, Malassezia and yeasts of emerging importance. The importance of laboratory diagnosis, antifungal susceptibility testing, antifungal resistance and yeast diseases in animals are reviewed.
Yeasts In Biotechnology And Human Health
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Author : Isabel Sá-Correia
language : en
Publisher: Springer
Release Date : 2019-03-25
Yeasts In Biotechnology And Human Health written by Isabel Sá-Correia and has been published by Springer this book supported file pdf, txt, epub, kindle and other format this book has been release on 2019-03-25 with Science categories.
This book discusses genome-based strategies to provide a holistic understanding of yeasts in Human Health and as model organisms in basic research or industrial production. Using numerous Saccharomyces cerevisiae strains and various non-conventional yeast species isolated from diverse origins, it describes essential biological processes, the biotechnological exploitation of yeast and pathogenesis control. It also demonstrates how functional and comparative genomics and the development of genome engineering tools are used in modern yeast research. The use of yeasts as experimental eukaryotic models increasingly gained prominence when several Nobel Prizes in Physiology/Medicine and Chemistry were awarded for innovative research, using yeast strains to elucidate molecular mechanisms in a wide range of human physiological processes and diseases, such as autophagy, cell cycle regulation and telomerase activity. This book offers useful insights for scientists in yeast research, clinical scientists working with yeast infectious models and for industrial researchers using applied microbiology.
Fungi
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Author : Kevin Kavanagh
language : en
Publisher: John Wiley & Sons
Release Date : 2011-08-04
Fungi written by Kevin Kavanagh and has been published by John Wiley & Sons this book supported file pdf, txt, epub, kindle and other format this book has been release on 2011-08-04 with Science categories.
Fungi: Biology and Applications, Second Edition provides a comprehensive treatment of fungi, covering biochemistry, genetics and the medical and economic significance of these organisms at introductory level. With no prior knowledge of the subject assumed, the opening chapters offer a broad overview of the basics of fungal biology, in particular the physiology and genetics of fungi and also a new chapter on the application of genomics to fungi. Later chapters move on to include more detailed coverage of topics such as antibiotic and chemical commodities from fungi, new chapters on biotechnological use of fungal enzymes and fungal proteomics, and fungal diseases of humans, antifungal agents for use in human therapy and fungal pathogens of plants.
The Yeasts
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Author : Cletus Kurtzman
language : en
Publisher: Elsevier
Release Date : 2011-05-09
The Yeasts written by Cletus Kurtzman and has been published by Elsevier this book supported file pdf, txt, epub, kindle and other format this book has been release on 2011-05-09 with Science categories.
The Yeasts: A Taxonomic Study is a three-volume book that covers the taxonomic aspect of yeasts. The main goal of this book is to provide important information about the identification of yeasts. It also discusses the growth tests that can be used to identify different species of yeasts, and it examines how the more important species of yeasts provide information for the selection of species needed for biotechnology. • Volume 1 discusses the identification, classification and importance of yeasts in the field of biotechnology. • Volume 2 focuses on the identification and classification of ascomycetous yeasts. • Volume 3 deals with the identification and classification of basidiomycetous yeasts, along with the genus Prototheca. High-quality photomicrographs and line drawings Detailed phylogenetic trees Up-to-date, clearly presented yeast taxonomy and systematic, easy-to-use reference sequence accession numbers to allow for correct identification