Microbiology

Introduction to microbiology; the classification of living organisms; the optical microscope. Gram staining. The bacterial cell wall. The cell wall of Saccharomyces cerevisiae. The capsule; flagella, cilia and pili; the bacterial endospore. Examples of spore-forming bacteria: Bacillus thuringiensis, Bacillus licheniformis and Clostridium botulinum. The cell membrane and membrane transport. The nutritional and energy needs of microorganisms. The chemical / physical environmental requirements of microorganisms: microorganisms and oxygen. The chemical / physical environmental needs of microorganisms: microorganisms and osmotic pressure (water activity). Bacterial reproduction: the bacterial growth curve; asexual reproduction of Saccharomyces cerevisiae. Culture media: classification based on physical state, chemical composition and function. The concept of sterility and the tools used for work in sterility; the types of inoculum. The determination of the microbial count with direct and indirect measurements: the total microbial count with counting chambers; description and examples of calculation for the viable count by serial decimal dilutions (the concept of CFU, the weighted average for the calculation of the CFU). Eumycetes: filamentous fungi, the classification of eumycetes based on asexual and sexual reproduction. Soil bacteria. Interaction soil-plant-microorganisms: the rhizosphere, the root nodules of legumes, nitrogen fixation, Agrobacterium, mycorrhizae. Introduction to cellular metabolism: ATP and its formation. Free electron transporters (NAD and NADP). The pyruvate production pathways: Embden-Meyerhof, pentose phosphate and Entner-Doudoroff pathways. The definition of the catabolic processes of respiration and fermentation. The different types of fermentation in bacteria (the alcoholic, lactic, mixed acid, butilenglicole, propionic, butyric and omoacetic fermentations). The fructose-6-phosphate phosphoketolase pathway. Lactic acid bacteria and their metabolism; acetic bacteria and their metabolism. Outline of anabolism. Principles of microbial genetics; the central dogma of biology; transcription regulation (the lac operon, the operon, tryptophan operon, maltose regulon); protein synthesis in prokaryotes. The concept of life in biology and viruses; bacteriophages; the plasmids. How microbial biodiversity is generated: horizontal gene transfer. The mechanisms of natural genetic recombination in bacteria: transformation, conjugation and transduction. Ongoing evaluation of student preparation. Polymerase chain reaction. Taxonomy and bacterial identification. The 16S rRNA gene; the 16S rRNA gene in bacterial identification and in the study of bacterial phylogeny.

Program of the laboratory exercitations
During the practical part of the course in the didactic laboratories, the importance and the modalities through which working in sterility in microbiology (explanation of the bunsen burner, oxidizing flame, cone of sterility) are illustrated; the instruments used in classical microbiology are shown and used (loops, spatulas, pipettes, test tubes, slants, shaking flasks, Petri dishes, liquid and agar culture media); it is taught how to prepare serial decimal dilutions starting from a sample of unknown concentration and then continuing with spread plate and encapsulation techniques on selective culture media, following which it is explained how to perform the viable count calculation from the colony forming units grown on the plate; from suspensions of yeasts or bacteria of unknown concentration, it is shown how to perform a calculation of total counts through the counting chamber with an optical microscope; the technique of plate striping is shown to obtain isolated colonies and pure cultures with the use of selective media; examples of biochemical assays to be performed on agar plates are shown to highlight some microbial enzyme activities (catalase, amylase, protease); tests of fermentative metabolism of lactobacilli are carried out with tests of use of different sources of sugars, and how the growth of anaerobic microorganisms is promoted; the Gram staining is explained and performed; the principles of phase contrast optical microscopy are illustrated, with examples of observation of fresh preparations of different bacteria and yeasts, and of bacteria stained from complex food matrices.