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Předmět Praktická cvičení z mikrobiologie je navazujícím předmětem pro předmět Mikrobiologie. Důraz v tomto předmětu je primárně kladen na metodické přístupy volené v rámci mikrobiologických laboratoří. Student během studia tohoto předmětu získá přehled a praktické dovednosti ohledně metodik, které se váží jak k problematice farmaceutické mikrobiologie, tak i lékařské mikrobiologie. Cílem výuky je připravit studenty rozvíjet získané poznatky při dalším studiu a aplikovat je ve zdravotnické praxi.
Poslední úprava: Konečná Klára, RNDr., Ph.D. (14.02.2025)
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Podmínky pro udělení zápočtu:
Poslední úprava: Konečná Klára, RNDr., Ph.D. (14.02.2025)
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Doporučená:
Poslední úprava: Konečná Klára, RNDr., Ph.D. (14.02.2025)
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- PPT prezentace, semináře - diskuse na dané téma - praktická cvičení - úvod do základních technik využívaných v mikrobiologické laboratoři Poslední úprava: Konečná Klára, RNDr., Ph.D. (14.02.2025)
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Znalosti k problematice metodických přístupů volených v mikrobiologických laboratořích - problematika dezinfekce, sterilizace, mikrobiologické kontroly čistoty prostředí - mikroskopické techniky, nativní a barvený preparát, barvící techniky - kultivace, podmínky kultivace, typy kultivačních půd, využití kultivačních půd, očkování kultivačních půd - testování citlivosti mikroorganismů vůči antimikrobním látkám - laboratorní diagnostika gram-pozitivních, gram-negativních bakterií - laboratorní diagnostika mykotických agens - laboratorní diagnostika parazitárních agens - metody identifikace mikrobiálních původců Znalosti definované otázkami v dokumentu Výstupy učení Poslední úprava: Konečná Klára, RNDr., Ph.D. (14.02.2025)
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Metodické přístupy v mikrobiologické laboratoři - dezinfekce, sterilizace, stěrové metody - mikroskopické techniky, nativní a barvený preparát, barvící techniky - kultivace, podmínky kultivace, typy kultivačních půd, využití kultivačních půd - testování citlivosti mikroorganismů vůči antimikrobním látkám - laboratorní diagnostika gram-pozitivních, gram-negativních bakterií - laboratorní diagnostika mykotických agens - laboratorní diagnostika parazitárních agens - metody identifikace mikrobiálních původců Poslední úprava: Konečná Klára, RNDr., Ph.D. (14.02.2025)
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Study Materials | Intranet FAF (cuni.cz) Department of Biological and Medical Sciences | Intranet FAF (cuni.cz) Department of Biological and Medical Sciences | Intranet FAF (cuni.cz) Poslední úprava: Konečná Klára, RNDr., Ph.D. (14.02.2025)
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Department of Biological and Medical Sciences, Charles University, Faculty of Pharmacy in Hradec Králové
Course Title: GF397 Microbiology-Practical Classes (Pharmacy, 2nd Year) Topic: Safety at work, disinfection, sterilization, microbiological laboratories, collection and types of biological material Syllabus – occupational health and safety in microbiological laboratory, protective equipment, laboratories and their classification according to BSL (Biosafety Levels), laminar flow box, the concept of disinfection and sterilization, physical sterilization – dry-heat sterilizer, autoclave, chemical sterilization – ethylene oxide, formaldehyde, pre-sterilization preparation of materials, sterilization control, types of samples/clinical specimens for microbiological investigation, sterile and non-sterile biological material, collection tools, material transport, results delivery Learning Objective – Students should become familiar with terms such as sterilization and disinfection. They should distinguish between naturally sterile and non-sterile clinical specimens, understand the concept of BSL, and know about the laminar box. In the practical part, they will learn how to interpret the results of the swab method (studying microbial contamination of spaces/surfaces, and assess the correctness of hand disinfection procedures. Task for Knowledge: · Determine whether it is appropriate to eat or drink within a microbiological laboratory. · Name at least three personal protective equipment items for work in a microbiological laboratory. · Determine what the acronym BSL stands for. Identify which BSL level of laboratory has the highest technical requirements and why. Determine whether it is necessary to work with common non-pathogenic agents in BSL3 or BSL4 laboratories. · Explain the purpose of disinfection and describe, what the goal of the disinfection process is. · Explain what the term sterilization means and what the goal of sterilization is. · Specify the criteria for choosing disinfection methods. · List the methods of physical sterilization. Define the conditions under which the sterilization cycle occurs in a dry-heat sterilizer and in an autoclave. Explain the fundamental difference between these two sterilization approaches. · Provide at least two examples of chemical sterilization. · Name at least five different clinical specimens for microbiological investigation. · Identify which clinical specimens are considered naturally sterile and which are naturally non-sterile.
Topic: Introduction to Microscopic Techniques and Staining Syllabus – microscopy, microscopic identification, types of microscopy – bright field, dark field, phase contrast, fluorescence microscopy, electron microscopy - TEM, SEM, dry system, oil immersion system, working distance of the objective lens, resolution, numerical aperture, basic shapes and formations of bacterial cells, microscopic preparations/mounts – wet, stained, indicative staining, diagnostic staining, staining of acid-fast bacteria and structures, visualization of capsules, Giemsa staining, Wirtz-Conklin staining Learning Objective –Students should become familiar with microscopic and staining techniques that can be used for microbiological investigations. Additionally, they should learn how to prepare wet mounts and identify microbial strains stained according to the Gram method.
Task for Knowledge: · Specify the difference between bright field microscopy, dark field microscopy, and phase contrast microscopy – describe how the studied object appears in each type. · Determine whether phase contrast microscopy is suitable (ideal) for observing bacteria. · Specify what should be used when preparing samples for fluorescence microscopy in order to visualize the objects. · Determine, for what observations is used transmission electron microscopy (TEM) and scanning electron microscopy (SEM). Name which type of electron microscopy provides greater magnification. · Explain the terms dry system and oil immersion system. · Determine the relationship between numerical aperture and working distance for observing an object. · Specify how total magnification is expressed. · Name the basic shapes and basic formations of bacterial cells. · Determine the purpose of a wet mounts observations. · Name whether wet or stained preparations are considered permanent preparations. · Name possible methods for fixing microbial agents to a slide for subsequent preparation of stained mounts. · Determine whether the movement of microbial agents can be observed in wet or fixed preparations. · Specify the difference between indicative and diagnostic staining. · Decide whether wet mounts are most suitable for microscopic observation of bacteria. Specify for which microbial agents this type of preparation is appropriate. · Explain the principle of Gram staining technique. Explain why some bacteria are stained blue (blue-violet) and others are stained pink (pink-red). · Specify the color of Gram-positive bacteria stained by Gram technique, and the color Gram-negative bacteria. · Name which agents are stained using the Ziehl-Neelsen staining technique. Specify the color appearance of acid-fast structures. · Explain why it is not easy to stain mycobacteria using the Gram method and name the staining technique used to visualize these agents microscopically. · Specify the purpose of use of the Burri staining technique. Describe how the structures visualized by this technique are distinguished (how they appear). · Name the microbiological discipline, where the Giemsa staining technique is the most commonly used. · Name what can be visualized using the Wirtz-Conklin staining technique. · Determine whether wet mounts are commonly observed under a school microscope at a total magnification of 1000X.
Topic: Cultivation of Infectious Agents – Cultivation Conditions, Types of Inoculation, Classification of Cultivation Media, Types of Cultivation Syllabus – cultivation, L. Pasteur, R. Koch, cultivation conditions, nutrition, appropriate temperatures for cultivation, terms: psychrophile, mesophile, thermophile, role of water for nutrient absorption, optimal pH for cultivation, term: acidophile, role of osmotic pressure, term: halophile, halotolerant bacteria, positive and negative redox potential, oxygen dependence, terms: aerobic, anaerobic, facultative anaerobic, microaerophilic, inoculation of solid and liquid media, isolative inoculation, classification of media by composition, classification of media by consistency, classification of media by purpose and use, McConkey agar, Loewenstein-Jensen agar, chromogenic media, types of cultivation – surface, submerse, static, dynamic, anaerobic cultivation, anaerostat, Fortner ‘s plate, macroscopic description of colonies, biochemical tests – KOH test, catalase production test, oxidase test. Learning Objectives – students should be able to explain the principles of cultivation and the selection of appropriate conditions for cultivation. They should also be able to describe the macroscopic characteristics of microbial colonies, perform basic biochemical tests – evaluate the ability to produce catalase or oxidase, and distinguish whether the studied strain is Gram-positive or Gram-negative based on the KOH test. Task for Knowledge:: · Explain the concept of cultivation. · Name the person who first introduced cultivation in liquid media, and the person who first introduced cultivation on solid media. · Identify the primary source of carbon and energy used by most pathogenic microorganisms. · State the temperature suitable for cultivating pathogenic bacteria. · State the temperature suitable for cultivating most pathogenic fungi and yeasts. · Explain the terms psychrophilic, mesophilic, and thermophilic microorganisms. · Explain why humidity is essential for the growth and multiplication of microorganisms. · Determine the optimal pH for most pathogenic microorganisms. · Explain the term acidophilic bacteria. · Explain the term halophilic and halotolerant microorganisms. · Determine the relationship of microorganisms to oxygen, explain terms - aerobic, anaerobic, and facultative anaerobic bacteria. · Name the methods for inoculating solid media for the purpose of isolation of different microbial strains. · Explain the term isolative inoculation. · Name how media can be classified by composition. · Name how media can be classified by consistency. · Name the categories into which media can be classified according to their purpose and use. · Classify blood agar and Sabouraud agar into the appropriate category based on their purpose and use. State for which microbial agents are these media used. · Explain the use of diagnostic media. · Explain the use of selective media, and list at least two components they may contain. · Determine the use of selective-diagnostic media. Name at least two media from this category. · Explain how McConkey agar can distinguish between lactose-positive and lactose-negative bacteria. · Name microbial agents cultivated on Loewenstein-Jensen agar. · Explain the concept of chromogenic media. · Identify the difference between static and dynamic cultivation, submerse and surface cultivation. · Name the methods for performing anaerobic cultivation. · Explain the terms anaerostat and Fortner ‘s plate. · Describe how the KOH test is performed. Determine, how the test is evaluated as positive or negative. Indicate, whether Gram-positive bacteria give a positive or negative result within the KOH test. · Describe, how the catalase test is performed. Identify the group of bacteria for which this test is primarily performed. · Describe how the oxidase test is performed. Identify the group of bacteria for which this test is primarily performed.
Topic: Laboratory Screening of susceptibility to antimicrobial agents Syllabus – antibiotics, anti-infectives, antibiotic resistance, MRSA, VRSA, VRE, mycobacteria, classification of antibiotics based on their mechanism of action, classification based on intensity of action, factors influencing the effectiveness of antibiotics in vitro, methods for evaluation of antibiotic susceptibility, classification of methods based on results and the environment/medium, quantitative and qualitative tests, dilution and diffusion tests, disk diffusion test, diffusion test – E-test, MIC, MBC, EUCAST.
Learning Objectives – Students should be able to define the term antibiotic drug. Students should be able to classify methods based on the results and the environment/medium in which they are performed. Students should be able to define the abbreviations MIC, and MBC. Students will gain practical experience in evaluating the disk diffusion test, microdilution broth test, and evaluation according to EUCAST.
Task for Knowledge: · Define the term antibiotic drug. · Explain the significance of antibiotic resistance for mankind. · Identify what the abbreviations MRSA, VRSA, and VRE mean. · Explain what contributes to the development and spread of resistance. · State the optimal pH for media used for in vitro antibiotic susceptibility testing. · Classify methods for determining antibiotic susceptibility based on the results and the environment/medium. Explain the terms dilution method/dilution test and diffusion method/diffusion test. Name examples of methods that belong to individual categories. · Explain the principle of the disk diffusion test. Identify ,which cultivation medium is used for this method. · Explain how the results of the disk diffusion test are evaluated. · Explain the principle of the E-test. Identify, into which group of methods this test belongs. · Explain the abbreviation MIC. Desribe, how this abbreviation is defined. · Explain the abbreviations MBC. Define, how this abbreviation is defined. · Describe, how to read the MIC value. State, how is evaluated. · State, how the information available on the EUCAST website can be used.
Topic: Laboratory Diagnostics of G+/G- cocci and G- rods Syllabus – clinically significant groups of G+ cocci, catalase test, genus Streptococcus, classification of streptococci based on hemolysis, classification of streptococci based on antigenic structure (Lancefield classification), genus Staphylococcus, classification of staphylococci based on coagulase production, classification of staphylococci based on hemolysis, clinically significant groups of G- cocci, genus Neisseria, test for cytochrome oxidase production, laboratory diagnostics of Neisseria, antigenic typing of Neisseria. classification of G- bacteria based on lactose production, biochemical identification of bacteria, HISU test/HISU series, serotyping of Salmonella, auxanogram.
Learning Objectives – Students will become familiar with the basic knowledge regarding clinically significant G+/G- cocci and G- rods. Students will learn biochemical identification techniques for bacteria, determining hemolysis types, and the biochemical identification of yeasts using auxanograms. This topic provides essential information, how to identify clinically significant bacteria, focusing on the differentiation of G+ cocci, G- cocci, and G- rods, as well as understanding laboratory techniques for characterizing microbial species and their biochemical properties.
Task for Knowledge: · Name three clinically significant genera of facultative anaerobic G+ cocci. · Determine the purpose of the catalase test. Explain the principle of this biochemical reaction. Name, which genus of bacteria (staphylococci or streptococci) produce catalase. · Describe the types of hemolysis observed in streptococci. Explain the arrangement of streptococcal cells that can be observed under the microscope. · Describe the arrangement of cells observed under the microscope for staphylococci. · State the purpose of the coagulase test. Determine, which clinically significant bacterium gives a positive reaction within the coagulase test. · Name two clinically significant species of the genus Neisseria. Describe the shape and arrangement of cells typical for these bacteria. · Determine, if non-pathogenic Neisseria species can be found in the oral cavity. · State, if serotyping is performed for Neisseria and identify the clinical specimen used for examination. · State the basis on which clinically significant G- rods can be classified. Identify the group to which Escherichia coli, Klebsiella spp., Salmonella spp., and Shigella spp. belong. · Name which G- intestinal bacteria are considered obligate pathogens and which are opportunistic pathogens. · State the method used for identifying G- rods. · Explain the principle of biochemical identification. · Name at least three selective diagnostic media used for biochemical identification of G- rods. · State the reason for performing Salmonella serotyping. Explain the principle of serotyping. Name the surface antigens of Salmonella that are distinguished. · State the purpose of an auxanogram. Explain the principle of this test, and explain, how the results are evaluated. · Describe, how biochemical tests (such as Enterotest, Staphytest) are evaluated. · Describe, how an auxanogram is evaluated. · Describe the difference between viridation and complete hemolysis in streptococci.
Topic: Mycology, Parasitology Syllabus – diagnostics of fungal and parasitic infections, microscopic techniques in mycology and parasitology, KOH (potassium hydroxide) preparation/mounts, fluorescence microscopy, detection of Cryptococcus capsules, Grocott silver staining technique, cultivation of fungal agents, cultivation media and cultivation conditions, biochemical identification of yeasts, identification of molds, gyrification and guttation, sample collection for parasitological examination, types of clinical specimens, microscopic techniques in parasitology. Learning Objectives – students should become familiar with the basic approaches for diagnosis of fungal and parasitic infections. Students will learn the techniques of cultivation of fungal agents, microscopy for fungal agents, and the microscopic detection of helminth infections. This topic provides essential knowledge for diagnosis of mycotic and parasitic infections, using microscopy and cultivation methods. Practical techniques and methods for the identification of fungal and parasitic agents, as well as the microscopic recognition of infections caused by helminths will be introduced.
Task for Knowledge: · Determine which laboratory investigations can support the diagnostics of mycotic infections. · Name the microscopic techniques commonly used for diagnostics of fungal infections. · Determine whether is possible to stain yeasts by Gram method. · State the purpose of KOH (potassium hydroxide) mounts in mycology. Name the fluorescent dye used in mycology for the microscopic detection of fungal agents. · Identify the role of ink in mycology. · Explain the purpose of the Grocott silver staining technique. · Name a universal medium used in mycology. State how this medium is enriched to prevent bacterial growth. · Determine how a universal medium for fungal agents is enriched when culturing dermatophytes. · State the use of corn/rice agar and Czapek-Dox medium. · Name the temperature used for cultivating Candida, Aspergillus, or Mucor, and the temperature for cultivating other fungal agents. · State the duration of fungal cultivation, distinguishing between yeasts and dermatophytes. · Determine whether biochemical tests like auxanograms or zymograms are commonly used for identification of molds. · Explain the term "auxanogram." · Explain the terms "gyrification" and "guttation." · State what type of clinical specimen is collected in case of suspected parasitic infections. · Explain the use of Graham and Schüffner’s technique for mount preparation. Describe how the preparation is made. · State the purpose of the thick smear technique according to Kato. Describe how the preparation is made. · State the material collected when malaria is suspected. Name the technique used to prepare samples for malaria diagnosis.
Poslední úprava: Konečná Klára, RNDr., Ph.D. (31.03.2025)
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