|teoria||4||I semestre||Giovanni Vallini|
|laboratorio [1° turno]||2||I semestre||Silvia Lampis|
|laboratorio [2° turno]||2||I semestre||Silvia Lampis|
The course is designed to introduce students to the basic knowledge of the microbial world as well as to illustrate the main methodological tools for research in microbiology, with a detailed comparison of the properties among the different type of microorganisms, both prokaryotes and eukaryotes, including bacteria, archaea, yeasts, filamentous fungi, and - in a distinct section, since non-cellular organisms – the viruses. During the first series of lectures, general themes will be addressed such as morphological and functional diversity, genetic, biochemical and metabolic features, evolutionary aspects and ecology of microorganisms, including how they interact with specific environmental factors. In the second series of lectures, microorganisms will be discussed as reference systems for fundamental studies dealing with molecular biology as well as biochemistry and metabolic regulatory mechanisms. Also elements will be provided about the methods for microbe cultivation and on strategies for the control of microbial growth and the conditioning of the metabolism. The third part of lectures will cover the detailed study of particularly important microbial groups to be defined with the Students.
This laboratory module is designed to guide students in the acquisition of techniques and in the development of manipulative skills all necessary for the identification and the study of both structural and functional characteristics of the microorganisms of interest, as well as for a proper handling of microbial cultures within research activities. The main objective of the module is to provide students with the basic tools, represented by traditional analytical procedures in microbiology, but also with the knowledge of advanced techniques, based on molecular methods of investigation, useful for basic research but even the definition of the correct approach to issues related to the many themes of applied microbiology.
CONTENTS of THEORETICAL LECTURES
1. MICROBIAL EVOLUTION AND DIVERSITY: Origin and Evolution of the Microbial Life; Bacteria; Archaea; Eukaryotic Cell and Eukaryotic Microorganisms; Microbial Taxonomy and Systematics. 2. PRINCIPLES OF MICROBIOLOGY: Cell Structure and Function in Bacteria, Archaea and Fungi (Yeast and Moulds); 3. MICROBIAL GROWTH: Microbial Nutrition, Culture and Growth Aspects; Antimicrobial Agents and Microbial Growth Control. 4. METABOLIC DIVERSITY: Phototrophy and autotrophy (purple sulfur and nonsulfur bacteria, green sulfur and nonsulfur bacteria, cyanobacteria), Aerobic (EMP pathway, Enther-Doudoroff pathway, Hexose Monophosphate Shunt + electron transport chain and proton motive force generation) and Anaerobic Respiration (denitrification, dissimilative sulfate reduction, acetogenesis, methanogenesis), Fermentations (homolactic and heterolactic, mixed-acid, butane-diol, propionic, butanol/acetone fermentations), Chemolithotrophy (hydrogen oxidizing, sulfur oxidizing, iron oxidizing, ammonia/nitrite oxidizing bacteria, anammox process); Other biosyntheses (ammonia assimilation, nitrogen fixation). 5. VIRAL DIVERSITY: Overview of Different Viral Groups; Bacteriophages. 6. BACTERIAL GENETICS: Mutations; Genetic Recombination: Transformation, Conjugation, Transduction. 7. MOLECULAR BIOLOGY AND GENE EXPRESSION Molecular Biology of Bacteria: Cloning Vectors and Expression Vectors; Regulation of Gene Expression. 8. MICROBIAL ECOLOGY: Methods in Microbial Ecology; Major Microbial Habitats and Diversity; Nutrient Cycles; Microbial Symbioses. 9. FOCUS ON SPECIFIC MICROBIAL GROUPS (Methylotrophs and Methanotrophs, Pseudomonadales, Enteric Bacteria, Myxobacteria, both nonsporulating and endospore-forming Firmicutes, Methanogens, moulds and yeasts).
[A] Basic laboratory techniques for isolation, cultivation, observation and enumeration of microorganisms. 1. Preparation of liquid and agarized culture media (rich and defined media); 2. Culture trasfer techniques and procedures for isolation in pure culture; 3. Enumeration of microbial populations from a complex matrix (e.g. soil, wastewater, compost): i) serial dilution-agar plate procedure and total count of CFUs (Colony Forming Units), ii) MPN (Most Probable Number) method; 4. Microbial staining and use of the compound microscope: i) simple staining, ii) Gram differential staining, iii) acid-fast staining, iv) spore staining.
[B] Characterization of a bacterial isolate from streak-plate procedure with direct complex matrix inoculation. 1. Classic culture-dependent protocols: (a) Microbial growth evaluation in liquid, agarized and slant cultures – (b) Motility tests: i) direct optical microscope observation, ii) recognition in soft agar tube – (c) Biochemical tests: i) sugar fermentation/oxidation, ii) catalase test, iii) oxidase test, iv) nitrate reduction test, v) urease test – (d) Growth in defined liquid medium added with different C substrates: OD measurement at 600 nm; 2. Molecular methods: (a) Total DNA extraction from liquid culture: i) DNA detection by means of agarose gel electrophoresis, ii) DNA quantification on BioPhotometer through analysis of 260/280 and 260/320 ratios – (b) PCR amplification of the 16S rRNA gene sequence – (c) ARDRA (Amplified Ribosomal DNA Restriction Analysis) protocol and observation of digestion profiles by agarose gel electrophoresis – (d) BOX-PCR analysis and observation of BOX profiles by agarose gel electrophoresis – (e) Selection by each Student Group of a specific bacterial isolate (on the basis of ARDRA and BOX profiles as well as of results from classic analytical methods) to be considered for sequencing after elution and quantification of the corresponding 16S rRNA gene sequence – (f) Analysis of the obtained 16S rRNA gene sequence by comparison with genetic sequence data banks: i) NCBI; ii) Ez-Taxon.
Final assignment of course credit by written examination. The test consists in the submission to the student of a multi-page form containing 20 to 25 quizzes, including single answer questions, multiple answer questions, calculation exercises, request of short comments and descriptions. Each question is given a different weight in points. The rating is in thirtieths, based on the percentage of points totalized with the answers correctly provided, also taking into account the score attributed to the laboratory report.
|teoria||Gianni Dehò e Enrica Galli (a cura di)||Biologia dei Microrganismi (Edizione 3)||Casa Editrice Ambrosiana||2018||978-8-808-18623-2|
|teoria||Michael Madigan, John Martinko, Kelly Bender, Daniel Buckley, David Stahl||BROCK Biologia dei microrganismi - Microbiologia generale, ambientale e industriale (Edizione 1)||Pearson Italia||2016||889190094X|
|teoria||Michael T. Madigan, John M. Martinko, Kelly S. Bender, Daniel H. Buckley, David A. Stahl||Brock Biology of Microorganisms - 14th Edition (Edizione 14)||Pearson||2015||978-0-321-89739-8|