Mgy 277 Study guide.pdf

Unit 1:Perspectives on Microbiology Know what the major contributors to the Germ theory of disease contributed and how they arrived at their conclusions. 1. Akshamsaddin Akshamsaddin, claimed that diseases, like plants and animals, have "invisible seeds." However, this idea doesn't align with the co...

Unit 1:Perspectives on Microbiology Know what the major contributors to the Germ theory of disease contributed and how they arrived at their conclusions. 1. Akshamsaddin Akshamsaddin, claimed that diseases, like plants and animals, have "invisible seeds." However, this idea doesn't align with the contributions of major figures to the Germ Theory of Disease. Akshamsaddin's views were more in line with pre-scientific ideas rather than the scientific developments that led to the Germ Theory. 2. van Leeuwenhoek(1632-1723): Van Leeuwenhoek is often credited as the first person to observe microorganisms using a microscope that he designed and built himself. He discovered a variety of microorganisms, including bacteria, protozoa, and even sperm cells, by carefully observing samples of water, plaque from his teeth, and other substances. His observations laid the groundwork for the idea that there is a hidden world of tiny living organisms not visible to the naked eye. While he didn't directly contribute to the Germ Theory of Disease, his work paved the way for later scientists to explore the microbial world. 3. Semmelweis (1818-1865) -Pioneer of antiseptic procedures -He was a Hungarian obstetrician working in Vienna. “Children fever” typically killed 10% of women who gave birth in hospitals at that time. During bad outbreaks, it killed 50% of new mothers – some died days later from infections after labour caused by strep bacteria invading the uterus. He saw that the clinic with doctors killed more patients than the clinic run by midwives. The procedure was simple, but the midwives did not perform the autopsy. Doctors were performing autopsies on dead patients and then attending deliveries of babies without washing – sometimes with blood still on their hands.Doctors themselves also died from childbed fever when performing autopsies of women with the disease. Semmelweis suggested that something was being transferred from the woman’s dead body to the doctors.Doctors ignored his advice and dismissed him from the hospital where he worked. When a mandatory hand washing policy was in place, the deaths decreased. The hand washing consisted of washing the hands with bleach and water after performing the autopsy and before delivering a baby. 4. Bassi (1773-1856) He was the first to demonstrate that a microbe can cause disease by noticing that if you isolate worms, they remained healthy. Hence, the source of disease must come from outside the worm. He was looking at the muscardine disease that killed silkworms that forced many silk factories to close down. He found a fungus that caused it, and he cultured it to introduce it to healthy worms to prove his hypothesis. 5. Koch (1843-1910) He was a German physician and founder of ‘modern bacteriology’ along with Louis Pasteur. He discovered the causes of ANTHRAX, CHOLERA and TUBERCULOSIS He developed modern methods for working with microbes in the laboratory, by using media or petri dishes to cultivate bacteria, separate different species among different colonies by looking at their different colours. Koch’s Postulates -The microbe must be found in all organisms suffering from the disease, but should not be found in healthy organisms. -The microbe must be isolated from a diseased organism and grow in a pure culture. -The cultured microorganism should cause disease when introduced back into a healthy organism. -The microbe must be isolated again from the newly diseased animal and identified as being identical to the original microbe. 6. Pasteur (1822-1895) ● He was French chemist and microbiologist ● He disproved ‘spontaneous generation’ of microbes. Thus he eliminated a ● commonly-held hypothesis of the time. ● He is considered the biggest contributor toward the germ theory of disease ● and he developed techniques for vaccine production that are still used today. ● He figured the mechanism behind fermentation and the role of oxygen in food ● production. ● He also found out how food got spoiled: microorganisms coming from outside ● air. Microbes do not just appear out of nowhere on sterile broth. They come ● from other microbes. ● Pasteurization: The brief heating of food or milk to kill microbes that ● cause spoilage. He developed… ● Vaccines for rabies – pointed toward the discovery of viruses ● Vaccines for anthrax ● Pasteurization for food and milk ● He discovered… ● Staphylococcus aureus ● Streptococcus pyogenes ● Pneumococcus ● Anaerobic fermentation by microbes ● New pathogens of silkworms ● He had three major contributions: microbes discovery, development of ● vaccines and lab techniques – the only scientist that had equivalent ● contribution was Isaac Newton. 7. Lister (1827-1912) ● He was a physician who helped pioneer antiseptic surgery. ● Surgical equipment and room were kept routinely sterilized before use ● He developed carbolic acid (phenol) as antiseptic for sterilization of wound and equipment – it was applied to damaged tissue, which prevented infection. Phenolic compounds are a class of germicidal compounds that kill a wide range of bacteria and non-toxic enough to be added to personal care products (mouthwash). o There was dramatic decrease infection rate and death rate. • How did the discoveries of the major contributors to the Germ theory of disease build upon one another? The discoveries of major contributors to the Germ theory of disease built upon one another in a chronological progression: ● Semmelweis and Lister: Identified the importance of hygiene in preventing infections during medical procedures, even though they did not identify specific microbes. ● Pasteur: Disproved spontaneous generation and demonstrated that microorganisms cause fermentation. He proposed the germ theory by showing that microorganisms are responsible for the spoilage of liquids. ● Koch: Developed Koch's postulates, a set of criteria to establish a causal relationship between a microbe and a disease. He used these postulates to identify specific bacteria as the causative agents of diseases. These contributions collectively established the germ theory, which states that microorganisms cause many diseases, laying the foundation for understanding and combating infectious diseases. • What impact has infectious disease (and the control of infectious disease) has had on the human population? Infectious diseases have had a profound impact on the human population throughout history. They have caused massive mortality, altered the course of civilizations, and influenced demographic patterns. The control of infectious diseases, through measures like vaccines, antibiotics, and public health practices, has led to increased life expectancy, reduced morbidity, and improved overall well-being • What are the major categories of microbes? The major categories of microbes include: ● Bacteria: Unicellular organisms with a rigid cell wall, often motile with flagella, and reproduce by binary fission. ● Archaea: Prokaryotic organisms similar to bacteria but with distinct genetic, biochemical, and structural differences. ● Fungi: Eukaryotic organisms, including yeasts and molds, with a rigid cell wall, involved in decomposing organic material. ● Protozoa: Unicellular eukaryotic organisms, typically motile, and often parasitic. ● Helminths: Multicellular parasitic worms, including nematodes, cestodes, and trematodes. ● Viruses: Non-cellular infectious agents consisting of genetic material (DNA or RNA) enclosed in a protein coat. • How can microbes be useful? Microbes have various useful roles, including: ● Biotechnology: Microbes are used in biotechnological processes, such as the production of insulin using recombinant DNA technology. ● Food Production: Microbes are involved in fermentation processes for food production, like yogurt and beer. ● Environmental Processes: Microbes play a crucial role in environmental processes, such as biodegradation and nitrogen fixation. ● Medicine: Microbes are essential in the development of vaccines, antibiotics, and other pharmaceuticals. • What are the key differences between Bacteria and Archaea? What about the Similarities? Differences: ● Cell Wall Composition: Bacteria have peptidoglycan in their cell walls, while Archaea lack peptidoglycan. ● Lipids in Membrane: Archaea have different lipids in their cell membranes compared to bacteria. ● Genetic Differences: Archaea have a distinct rRNA sequence, transcription process, and ribosome structure compared to bacteria. Similarities: ● ● ● ● Prokaryotic: Both Bacteria and Archaea are prokaryotic organisms. Size and Shape: They share similarities in size, shape, and appearance. Motility: Both can be motile, using flagella for movement. Binary Fission: Both reproduce through binary fission. • What are the defining features of fungi, protozoa and helminths? ● Fungi: Eukaryotic organisms with a rigid cell wall, often multicellular (molds) or unicellular (yeasts), involved in decomposing organic material through absorption. ● Protozoa: Unicellular eukaryotic organisms, often motile, involved in various ecological roles, including some that are parasitic. ● Helminths: Multicellular parasitic worms, including nematodes (roundworms), cestodes (tapeworms), and trematodes (flukes). • Why are some non-microbes part of the study of microbiology? Non-microbes, such as helminths (parasitic worms) and viruses, are part of the study of microbiology because they can cause infectious diseases. Although larger in size compared to bacteria and archaea, these organisms have significant impacts on human health, and understanding their biology and mechanisms of infection is crucial for effective disease control and prevention. • What is the evidence for the two-domain system? The evidence for the two-domain system (Bacteria and Archaea) comes from genetic information, particularly ribosomal RNA (rRNA) sequences. Carl Woese, based on DNA evidence, proposed that some prokaryotes were distinct from bacteria in many ways, leading to the classification of Archaea as a separate domain. Genetic studies, including rRNA sequence divergence, provided strong support for the separation of Archaea from Bacteria and the establishment of the two-domain system. Unit 2: Bacteria Know the definitions of magnification, refraction, resolution and contrast and be able to apply them when looking at an image Magnification – The increase in the apparent size of the object compared to the size of the actual object (x = how many times larger). Refraction – When light rays change direction due to a change in the medium though which they travel – bend at the boundary of two different materials. ● Refractory index – The measure of relative speed of light as it passes through a medium (air or water) – how fast the light can travel through that particular medium. ● Lenses use refraction to focus on light – they are shaped to take advantage of refraction to focus light. ● Light bends when it hits a change in materials when the refractive index of each differs significantly. The bend is sharper when the light ray hits the interface between the two materials at an angle. Resolution – The ability to see objects (or points) as distinct, instead of as a blur that combines them – The minimum distance at which two points can be distinguished as individuals. Poor resolution indicates that when it is magnified, they look like one big object. Contrast – The ability to see objects against the background Techniques to Improve Contrast: ● Changing the microscope optics – dark-field microscopy, phase contrast microscopy, and differential interference. Manipulate/focus the light differently to make the object jump out from the background. ● Staining the sample – positive staining (cells are darker), negative staining (background darker), and fluorescent stains. Make the object or the background darker. • Know the lenses of a bright-field microscope (ocular, objective, condenser) Bright-Field Microscope – The basic type of microscopy ● Ocular – It magnifies an additional 10X ● Objective – It can magnify from 4X to 100X – the main magnifying lens on the microscope ● Condenser – It focus light coming from the bottom to provide an even white background underneath the sample, but it does not magnify the sample. • Know how to calculate the total magnification in a light microscope given the power of the different lenses The total magnification is the product of the magnifying power of the ocular and objective lenses. Example: 10X (ocular lens) x 40X (objective lens) = 400X total magnification • Know why and when oil is used in a light microscope ● The 100X lens on a bright field microscope requires that a drop of immersion oil is placed between the slide and the lens since it displaces air between lens and specimen. ● 100X cannot do that good of a job because it is too sensitive to the sudden changes of direction of light when the light enters the glass slide, then enters the air, then to glass lens. The refraction is making the light bent all over the place. ● The 40X can tolerate the small changes of light direction without too much distortion – therefore, the oil does not need to be used. ● Oil has the same refractive index as glass – the oil prevents light from missing objective lens so the image can be clear and undistorted. • Know the difference between bright and dark field light microscopy Bright-Field Microscope ● This is the most common microscope in a lab. ● Light passes through specimen, then a series of magnifying lenses, which evenly illuminates the entire field of view. ● It has two types of magnifying lenses: ocular and objective and one condenser to focus the light from the lamp, however, it does not magnify the object. ● 0.2 nanometers resolution maximum, which is a physical limitation of visible light and cannot be improved with powerful lenses. ● It can see the shape of bacteria but it cannot see the details. Also, it cannot see viruses. ● It is the worst when it comes to contrast, because many bacteria are clear against a bright, white background. This makes it difficult to distinguish the objects. Dark-Field Microscope ● It directs light towards specimen at an angle, instead of projecting light straight up to the sample like in bright-field microscope. ● Only the light scattered by specimen enters objective (background light does not enter the object lenses directly. ● Cells stand out as bright against a dark background. ● These microscopes require a wet-mount preparation – this can keep the cells alive so that motility is visible ● Wet mount = slide + drop of liquid (water or nutrient broth) + cover ● (usually thin) • Know the fundamental differences between electron and light microscopy and the advantages and disadvantages of each Electron Microscope Electromagnetic lenses, electrons and fluorescent screen replace glass lenses, visible light, and the eye Image can be captured on film to create an electron micrograph Wavelength of electrons of about 1000 shorter than light Light microscopy is limited to 0.2 micron because of the light wavelength. Electrons have much shorter wavelength Resolving power of about 1000 fold greater than bright field microscope which is about 0.3 nm. Resolution is much better and it can be used to resolve details at a much higher magnification, and it can be used to visualize individual proteins or DNA. It can magnify images 100,000 X Disadvantages of Electron Microscopy It is very expensive and large It is complicated to operate High maintenance costs Sample must be dead (if alive it will damage the sample as it needs to be kept in a vacuum) Stains use toxic/ radioactive metals (to improve contrast – uranium) Difficult sample operation No color, no movement *A light microscope can do just as much or more, but it cannot need high resolution. • Know the different images obtained from scanning and transmission EM One drawback is that lenses and specimen must be in a vacuum because air molecules would interfere with the electrons. This results in large, expensive unit and complex specimen preparation. There are two main types: transmission EM (TEM) and scanning EM (SEM). Transmission EM These are used to observe the fine detail of the cell structure and it works by directing electrons that either pass through or scatter Dark areas on image correspond to dense portions of specimen Thin-sectioning – allows you to see internal details but process can distort cells Scanning EM It is used to observe surface details, as the surface is coated with thin film of metal A beam of electrons is scanned over the surface of specimen Electrons released from the specimen are reflected and observed in viewing chamber Relatively large specimens can be viewed and it yields a 3D effect (dust mite) • Know the steps of a Gram-stain and the basic reason why some cells stain purple while others do not This is the most commonly used stain for bacteria – it distinguishes gram positive from gram negative bacteria (which allows to tell the difference between cell wall composition). A differential stain is a staining procedure that can distinguish one type of cell from another. Steps 1. Crystal violet (primary stain) which makes the cells purple 2. Iodine (mordant) is added to combine the crystal violet to make the crystal violet much less soluble – the cells remain purple – without this step, the cells would easily wash away. 3. Alcohol (decolorizer) is used to wash out excess crystal violet and iodine – gram-positive cells remain purple and gram-negative cells become colorless 4. Safranin (counterstain) is used to stain the gram-negative cells that the crystal violet did not stain after washed off by ethanol – gram-positive cells remain purple and gram-negative cells appear pink. • Know what the acid-fast stain is used for ● It detects a small group of organisms that do not readily take up the stain – mycobacterium ● Cell wall contains high concentrations of mycolic acid ● Waxy fatty acid that prevents uptake of dyes ● Harsh methods needed ● A dye called carbol fuchsin is used – cells are treated with heat to take up the dye, then washed with mixture of ethanol and acid, which quickly decolorizes most cells, but mycobacteria ● typically retains the stain. ● Gram stain or acid-fast stain depends on whether the cell can retain the dye during the washing step ● Gram-positive bacteria – species that can retain gram iodine and crystal violet mix ● Acid-fast bacteria – species that can retain carbol fuchsin after the ● ethanol and acid wash • Know the difference between differential and simple staining ● Simple staining involves one dye ● Differential staining distinguishes different types of bacteria • Know what fluorescence microscopy is (vs. simple light microscopy) and what makes immunofluorescence techniques so powerful Fluorescence Microscopes It requires the use of microscopes that have lamps that can excite fluorophores at one wavelength of light and detect light at a different wavelength They are more complicated and expensive than the conventional bright-field microscopes Yet, the information that you can get from this microscope is more effective and powerful Immunofluorescence It is the use of fluorescent molecules (fluorophores) that have been chemically attached to antibodies that can bind specific cellular components. Antibodies bound with fluorophores can emit different colours and show different parts of the cell all at the same time. Technique used to tag specific proteins with a fluorescent compound by using an antibody to deliver the fluorescent tag. Tagging a protein unique to a microbe can detect that organism. • Know the terms for the shapes and groupings of bacterial cells (cocci, bacilli, etc) The shapes of bacteria are the first way to classify them – but this a terrible way of classification because it does not tell the relation among bacteria. The two most common types are the coccus (spherical) and the rod/ bacillus (cylindrical) Rod shaped bacteria examples are E. coli, listeria and salmonella. There can also be other shapes such as vibrio (rod-shaped with a bend), spirillum (spiral shaped) and spirochete (cork screw shaped - can swim by twisting themselves around) Groupings Most of the prokaryotes divide by binary fission – the cells often stick together following division They form characteristic groupings: ● Neisseria gonorrhoeae – diplococcus ● Streptococcus – long chains ● Sarcina – cubical packets ● Staphylococcus – grapelike clusters However, the cell clustering is also an unreliable way to identify bacteria – growth conditions changes can cause bacteria to clump or cluster in different ways. • Know how molecules get across lipid bilayers (diffusion and passive, active, and facilitated transport) Permeability of the Cell Membrane Cytoplasmic membrane is selectively permeable – blocks ions such as Na+, K+ and Clo O2, CO2, N2, small hydrophobic molecules and water pass freely. Some cells facilitate water passage with aquaporins – water passes really slowly. Other molecules must be moved across membrane via transport systems – the bigger the compound, the slower it gets. Most molecules must pass through proteins functioning as selective gates. Termed transport systems (permeases or carrier) – move nutrients, small molecules, waste and other compounds o Membrane-spanning o Highly specific – carriers transport certain molecule type Facilitated Diffusion Form of passive transport Movement down the gradient; no energy required Not useful in low-nutrient environments Rarely used by prokaryotes Equilibrium forces move molecules from high concentration to low concentration. Active Transport The movement against a concentration gradient Requires energy It uses proton motive force (H+ gradient) Uses ATP (ABC transporter) Commonly used by bacteria *In = nutrients, things the bacteria need to live + Out = waste product, signals to other bacteria, large proteins and DNA Protein Secretion Active movement of proteins out of the cell Examples: Extracellular enzymes and external structures Proteins tagged for secretion via a signal sequence of amino acids Prokaryotes use a variety of secretion systems Summary of Transports Diffusion Transport – A transport that does not require any energy and is a movement across a concentration gradient Passive Transport – A transport that does not require any energy. Active Transport – Movement against a concentration gradient and requires energy. Commonly used by bacteria. Facilitated transport - (A form of Passive transport) Movement down gradient —No energy required. Rarely used by prokaryotes and not useful in low-nutrient environments. • Know the fundamental structure of a Gram-positive and Gram-negative bacterial cell Molecules get across the outer membrane of gram-negative bacteria by using The non-specific porin portion is located on the outer membrane.Gram-negative have a thin layer of peptidoglycan then surrounded by a second membrane called the outer membrane (this outer membrane is a unique lipid bilayer embedded with protein – it is far away from the cytoplasm so there is no ATP, H+ motor force, or any source of cellular energy in periplasm and outer membrane. Porins on the outer membrane allow passage of AAs and simple sugars, not as specific as transport in the inner membrane. They are actually open channels that allow different molecules access the periplasm (the space between the two layers). Gram-positive have one single layer of cytoplasmic membrane surrounded by a thick peptidoglycan cell wall. • Know the basic structure of peptidoglycan (NAG-NAM chains and wall peptides). ● ● ● ● A rigid cell wall is made from peptidoglycan – found only in bacteria Peptido = contains peptide + glucan = contains sugar Alternating series of subunits form glycan chains N-acetylmuramic acid (NAM) ● ● ● ● ● ● ● ● N-acetylglucosamine (NAG) Tetrapeptide chain (string of four amino acids) “L” shaped Alternating series of subunits form glycan chains Wall glycan: NAG (Two sugars) and NAM (It is a derivative of NAG that can link to the N-terminus of the wall peptide – Only in bacteria) Wall Peptide: A tetra peptide chain (string of four amino acids) **The amino acids end with 2 D-alanines • Know the very basic structure of LPS (Lipid A, core, O-antigen) and its importance ● ● ● ● ● ● ● ● ● This can cause symptoms characteristic of infection by live bacteria Components of LPS include… Lipid A – Part of LPS recognized by immune system, it anchors LPS to lipid bilayer Fatty acid linked to pair of sugar – closest to the interior. This is the most conserved component of LPS O antigen – It can be used to identify species or strains, and it is composed of sugar molecules (number and type vary) Furthest to cell interior This is the least conserved component of LPS It also helps bacteria stay one step ahead of our adaptive immune systems Core polysaccharide – short stretch of sugars arranged in very unique patterns in between Lipid A and antigen-O Unit 3: Viruses What are the contributions of Beijerink, Ivanovsky, Loeffler and Frosch? Beijerink and Ivanovsky: Beijerink and Ivanovsky independently contributed to the discovery of viruses. Beijerink coined the term "virus" to describe a filterable infectious agent, and Ivanovsky identified the tobacco mosaic virus, the first virus discovered. Loeffler and Frosch: Loeffler and Frosch discovered that foot-and-mouth disease in cattle is caused by a virus. Their work laid the foundation for understanding viral diseases in animals. • What are the physical and genomic features of viruses? ● They are notable for their small size ● RBC is far greater than the size of any virus known (mimivirus is the 3rd ● largest virus). ● Viruses are about 10 to 1000 times smaller than the cell that they ● infect ● The smallest virus is a spherical virus called parvovirus – it only requires two genes to replicate and produce virsuses ● Virion is nucleic acid + protein coat (capsid) ● Nucleocapsid compromised of a capsid containing a genome – capsid has several functions such as protecting the nucleic acid from degradation ● Viruses have protein components for attachment (‘spikes’) ● They attach to specific receptor sites and enter the host cell • Why were we able to eradicate smallpox? What viruses is it related to? Smallpox was eradicated through a global vaccination campaign led by the World Health Organization (WHO). The virus responsible for smallpox is related to the Variola virus. The success of eradication was possible due to the availability of an effective vaccine and a coordinated global effort. • How are viruses classified and grouped? Viruses are classified based on various criteria, including the type of nucleic acid (DNA or RNA), presence of an envelope, symmetry of capsid, and dimensions of virion. They are grouped into orders, families, sub-families, and genera. • What makes viruses challenging to study in the laboratory setting? Viruses are challenging to study because they require a host cell for replication. They are intracellular parasites, making it necessary to cultivate them in cells for laboratory study. • What are the basic structures of enveloped and non-enveloped animal viruses, and what functions do they serve? (helical, icosahedral, complex, spike proteins) Enveloped Viruses: Have an outer lipid envelope derived from the host cell membrane. Examples include Influenza and Herpes viruses. Non-enveloped Viruses: Lack a lipid envelope. Examples include Adenovirus and Tobacco Mosaic Virus. Structures: Viruses can have helical, icosahedral, or complex capsid symmetry. Spike proteins on the envelope can facilitate attachment to host cells. • What is the relationship between shingles and chickenpox? Where does the varicella zoster virus reside in the body? Both are caused by the Varicella-Zoster virus. Chickenpox occurs initially, and the virus may become latent in nerve cells. Shingles (Herpes Zoster) occurs when the virus reactivates, causing a painful rash along the affected nerve pathway. • Know the difference between acute and persistent infections. Acute Infections: Short-lived infections with a rapid onset, often resulting in the host's recovery or death. Persistent Infections: Last for an extended period, divided into chronic (continual virus production) and latent (periods of inactivity followed by reactivation). • How can viruses contribute to cancer? Some viruses can contribute to cancer by integrating their genetic material into the host cell's DNA, disrupting normal cell cycle control. Examples include Human Papillomavirus (HPV) and Hepatitis B and C viruses. • What is a virion? A virion is a complete, infectious viral particle outside a host cell. It consists of the viral genome (DNA or RNA) and a protective protein coat (capsid). • Know the five steps of viral infection life cycle. Attachment: Viral particles bind to host cell receptors. Penetration and Uncoating: Entry into the host cell and release of viral genetic material. Synthesis of Viral Proteins and Replication: Replication of viral genome and production of viral proteins. Assembly and Maturation: Viral components come together to form new virions. Release: Newly formed virions exit the host cell to infect new cells. • Know the basic strategies of replication and how they relate to the type of nucleic acid the virus has (DNA or RNA - single stranded or double stranded - positive or negative sense) Strategies vary based on the type of nucleic acid (DNA or RNA) and its sense (positive or negative). DNA viruses often replicate in the host cell nucleus, while RNA viruses replicate in the cytoplasm. Retroviruses (e.g., HIV) use reverse transcription. • Define latent, acute, chronic infections Latent Infection: Periods of inactivity followed by reactivation. Acute Infection: Short-lived with a rapid onset. Chronic Infection: Continual virus production. • What is a cytopathic effect? How does it manifest? CPE refers to the visible damage to host cells caused by viral infection. It may manifest as cell rounding, cell fusion, or cell death. Unit 4: Eukaryotic Microbes Know the difference between an opportunistic and primary pathogen. Opportunistic pathogens typically cause disease in hosts with compromised immune systems, while primary pathogens can cause disease in healthy individuals. • Know what differentiates a eukaryotic cell from a prokaryotic cell. Eukaryotic cells have a true nucleus and membrane-bound organelles, while prokaryotic cells lack a true nucleus and membrane-bound organelles. • What are the defining characteristics of fungi? Eukaryotic organisms with chitin-containing cell walls, ergosterol in cell membranes, and existing in forms like yeast, hyphae, or spores. • What unique fungal cell pathways make attractive targets for antifungal drugs? Ergosterol biosynthesis is a major target for antifungal drugs. • What are the defining characteristics of protozoa and how they contribute to human disease? Microscopic, single-celled eukaryotes with various forms. They can cause disease through direct infection or by serving as hosts for other pathogens. • Know the three major types of helminths, their differentiating features, and routes of human infection. a.Nematodes (Roundworms): Cylindrical, digestive system, ingestion. b. Cestodes (Tapeworms): Segmented, absorb nutrients, ingestion of undercooked meat. c. Trematodes (Flukes): Flat, absorb nutrients, penetration or ingestion. • Know the definition of intermediate and definitive hosts Intermediate hosts support asexual reproduction, while definitive hosts support sexual reproduction of parasites. • What is a prion and how does it propagate? Prions are misfolded proteins (PrPSc) that induce normal proteins (PrPC) to adopt the abnormal shape, leading to the accumulation of PrPSc. • What is the prion “species barrier”? The difficulty in transmitting prions between different species due to differences in amino acid sequences of PrP. • How can prions be detected? Techniques like RT-QuIC (Real-Time Quaking-Induced Conversion) can be used to detect minute quantities of prions. • Know the basics of the following eukaryotic pathogens (including risk factors for acquiring, if relevant) ● Histoplasma capsulatum: Causes histoplasmosis; found in soil with bird or bat droppings. ● Pneumocystis jiroveci: Causes pneumonia; common in immunocompromised individuals. ● Cryptococcus gattii and neoformans: Cause cryptococcosis; associated with bird droppings. ● Candida auris and albicans: Yeast causing candidiasis; C. auris is multidrug-resistant. ● Acanthamoeba keratitis: Protozoan causing eye infection; associated with contact lens use. ● Cryptosporidium: Protozoan causing gastrointestinal illness; waterborne transmission. ● Toxoplasma gondii: Causes toxoplasmosis; transmitted by contaminated food or cat feces. ● Tapeworms (Cestodes): Various species; transmitted by ingesting undercooked meat. ● Ascaris: Roundworm causing ascariasis; ingestion of contaminated food/water. ● Naegleria fowleri: Amoeba causing fatal brain infection; found in warm freshwater. ● Schistosoma mansoni: Blood fluke causing schistosomiasis; freshwater snail intermediate host. Unit 5: Controlling Microbial Growth Know the definition and characteristics of psychrophile, psychrotroph, mesophile, thermophile, hyperthermophile, neutrophile, acidophile, alkalophile, halophile, and halotolerant bacteria Psychrophile: A psychrophile is a microorganism that thrives at low temperatures, typically ranging from -20°C to 10°C. Psychrotroph: Psychrotrophs are organisms that can grow at cold temperatures but have an optimal temperature range higher than that of psychrophiles, usually around 20-30°C. Mesophile: Mesophiles are microorganisms that prefer moderate temperatures, typically thriving between 20°C and 45°C. Most common bacteria and pathogens are mesophiles. Thermophile: Thermophiles are microorganisms that thrive at high temperatures, usually between 45°C and 80°C. Hyperthermophile: Hyperthermophiles are extremophiles that grow optimally in extremely high temperatures, often above 80°C. Neutrophile: Neutrophiles are microorganisms that grow optimally at a neutral pH, around 7. Acidophile: Acidophiles are microorganisms that prefer acidic environments, thriving at pH levels below 5. Alkalophile: Alkalophiles are microorganisms that prefer alkaline environments, thriving at pH levels above 9. Halophile: Halophiles are microorganisms that thrive in high-salt environments. Halotolerant: Halotolerant microorganisms can tolerate some level of salinity but do not necessarily require it for growth • Know the differences between facultative anaerobes, obligate aerobes, obligate anaerobes, microaerophiles, aerotolerant anaerobes Facultative Anaerobes: Facultative anaerobes can grow with or without oxygen, but they usually prefer oxygen. Obligate Aerobes: Obligate aerobes require oxygen for growth. Obligate Anaerobes: Obligate anaerobes cannot survive in the presence of oxygen and only grow in anaerobic conditions. Microaerophiles: Microaerophiles require oxygen, but at lower concentrations than found in the atmosphere. Aerotolerant Anaerobes: Aerotolerant anaerobes can survive in the presence of oxygen but do not use it for growth. • What is catalase? What is superoxide dismutase? Catalase: Catalase is an enzyme that catalyzes the breakdown of hydrogen peroxide into water and oxygen, protecting cells from oxidative damage. Superoxide Dismutase: Superoxide dismutase is an enzyme that converts superoxide radicals into hydrogen peroxide and oxygen, playing a role in antioxidant defense. • Describe the principles of sterilization, disinfection, pasteurization, decontamination, sanitization, and preservation. Sterilization: Sterilization is the complete elimination of all forms of microbial life, including spores. Disinfection: Disinfection refers to the elimination of most or all pathogens on inanimate surfaces, reducing the microbial population to a safe level. Pasteurization: Pasteurization is a process of heat treatment to reduce or eliminate pathogens in food and beverages, without affecting their quality. Decontamination: Decontamination involves the reduction of microbial populations to safe levels, reducing the risk of infection. Sanitization: Sanitization is the process of reducing microbial populations to levels considered safe by public health standards. Preservation: Preservation involves inhibiting the growth of microorganisms to extend the shelf life of perishable products. • Understand when the following physical methods would be used to control microbial growth: Pasteurization: Used in the food industry to eliminate pathogens and reduce spoilage organisms without compromising food quality. Autoclaving: Utilized to sterilize equipment and media by applying steam under pressure. Dry Heat: Sterilization method using hot air, suitable for materials that may be damaged by moisture. Filtration: Involves passing liquids or gases through a filter to physically remove microorganisms. Radiation (Ionizing vs Ultraviolet): Ionizing Radiation (e.g., gamma rays) damages DNA and is used for sterilization. Ultraviolet (UV) radiation is effective for disinfection, damaging DNA to prevent microbial replication. High Pressure: Destroys microorganisms by altering their molecular structure, often used in food preservation. Pascalization: Involves applying high pressure to foods to eliminate pathogens and extend shelf life. • Understand when the following classes of chemicals would be used to control microbial growth: Alcohols: Used as disinfectants and antiseptics, effective against a wide range of microorganisms. Aldehydes: Formaldehyde and glutaraldehyde are examples used for disinfection and sterilization. Ethylene Oxide Gas: A sterilizing agent for heat-sensitive materials in the healthcare industry. Halogens: Chlorine and iodine are common halogens used for disinfection and water treatment. Peroxygens: Hydrogen peroxide is an example, used as a disinfectant and bleach. Phenolic Compounds: Phenol and derivatives (e.g., triclosan) are disinfectants with antimicrobial properties. Quats (Quaternary Ammonium Compounds): Used as disinfectants, detergents, and antiseptics. • What important factors are there to consider when selecting an appropriate germicidal chemical? Efficacy: The chemical's effectiveness against the target microorganisms. Safety: The chemical should not harm humans or surfaces. Compatibility: Compatibility with the material being disinfected. Residue: Some chemicals leave residues that may be undesirable. Environmental Impact: Consideration of the chemical's impact on the environment. • Compare and contrast chemical preservatives, low-temperature storage, and reducing the available water as methods to preserve perishable products. Chemical Preservatives: Additives that inhibit microbial growth and extend the shelf life of products. Low-Temperature Storage: Refrigeration and freezing slow down microbial growth and preserve food. Reducing Available Water: Reducing water availability through dehydration or adding solutes inhibits microbial growth. • What is a biofilm? Biofilm: Biofilms are communities of microorganisms that adhere to surfaces and produce a protective extracellular matrix. They are resistant to antibiotics and immune responses, causing persistent infections. Unit 6: Microbial Classification and Identification What is the difference between a strain and a species? Species: In microbiology, a species is a fundamental taxonomic rank. It represents a group of organisms capable of interbreeding and producing fertile offspring. Strain: A strain refers to a group of microorganisms derived from a single original isolate. Strains within a species share many characteristics but may exhibit variations. • Describe how prokaryotes have been/are identified, classified, and assigned names Prokaryotes Identification: ● Microscopic Morphology ● Culture Characteristics ● Metabolic Capabilities ● Serology ● Fatty Acid Analysis Prokaryotes Classification: ● Morphology, Gram Stain ● Immunological Methods ● Molecular Techniques (16S rRNA Sequencing, DNA Hybridization) • Describe how phenotypic characteristics—including microscopic morphology, culture characteristics, metabolic capabilities, serology, and fatty acid analysis—can be used to identify prokaryotes. Microscopic Morphology: ● Description: Examining cellular structures and arrangements under a microscope. ● Application: Distinguishing between bacteria based on cell shape, size, and arrangements (e.g., cocci, bacilli, spirilla). Culture Characteristics: ● Description: Observing growth patterns, color, texture, and other features in laboratory culture. ● Application: Differentiating bacteria based on colony morphology, growth requirements, and environmental adaptations. Metabolic Capabilities: ● Description: Assessing the ability of microorganisms to utilize specific substrates or carry out metabolic reactions. ● Application: Utilizing biochemical tests to identify metabolic pathways, aiding in the classification of bacteria. Serology: ● Description: Using antibodies to detect and identify specific antigens on the surface of microorganisms. ● Application: Typing and classifying bacteria based on their antigenic properties, facilitating serological identification. Fatty Acid Analysis: ● Description: Analyzing the composition and structure of fatty acids in bacterial cell membranes. ● Application: Profiling fatty acid patterns for bacterial identification and classification based on lipid content. • Understand the differences between strategies used to distinguish different strains – biochemical, serological, molecular typing, phage typing, antibiograms Distinguishing Strains: Biochemical Tests: Analyzing metabolic pathways and enzymatic activities. Serological Typing: Using antibodies to identify specific antigens on the surface of bacteria. Molecular Typing: Examining DNA, including methods like PCR, RFLP, MLST. Phage Typing: Using bacteriophages to differentiate bacterial strains. Antibiograms: Assessing the susceptibility of bacterial strains to antibiotics. • What methods can be used to classify species and strains and how do they work? When are they appropriate to use? Advantages/disadvantages? (including those listed above) 16S rRNA/rDNA Sequencing: Sequencing the 16S ribosomal RNA gene to identify and classify bacteria. Appropriateness: Excellent for bacterial identification and phylogenetic analysis. Advantages: Highly conserved, universal across bacteria. Disadvantages: Limited resolution for closely related strains. Serological Typing: Identifying bacteria based on their antigenic properties using antibodies. Appropriateness: Useful for certain species with well-defined antigens. Advantages: Specific, established for certain pathogens. Disadvantages: Limited to known antigens. MLST (Multi-Locus Sequence Typing): Analyzing sequences of multiple housekeeping genes for strain typing. Appropriateness: Useful for studying genetic diversity and population structure. Advantages: High discriminatory power. Disadvantages: Requires sequencing multiple genes. RFLP (Restriction Fragment Length Polymorphism): Analyzing variations in DNA fragment lengths after digestion with restriction enzymes. Appropriateness: Useful for detecting genetic variations. Advantages: Simple, cost-effective. Disadvantages: Requires a considerable amount of DNA. DNA Hybridization: Assessing DNA sequence similarity through hybridization. Appropriateness: Suitable for comparing genetic relatedness. Advantages: Quantitative measure of DNA similarity. Disadvantages: Sensitivity to hybridization conditions. Biochemical Tests/API Tests:Evaluating metabolic capabilities and enzyme activities. Appropriateness: Initial screening for bacterial identification. Advantages: Rapid, cost-effective. Disadvantages: Limited to observable biochemical traits. MALDI/TOF MS (Matrix-Assisted Laser Desorption/Ionization Time of Flight Mass Spectrometry): Profiling microbial proteins for identification. Appropriateness: Rapid identification in clinical microbiology. Advantages: Fast, cost-effective. Disadvantages: Database-dependent. Real-Time PCR:Quantifying DNA in real-time during PCR. Appropriateness: Rapid detection of specific DNA sequences. Advantages: High sensitivity, quantitative. Disadvantages: Requires target sequence knowledge. WGS (Whole Genome Sequencing): Sequencing the entire genome for comprehensive analysis. Appropriateness: Broad applications in microbial genomics. Advantages: High resolution, detailed genomic information. Disadvantages: Cost, computational demands. Unit 7: Antimicrobials and Resistance What species of microbes produce antibiotics? Antibiotics are primarily produced by bacteria and fungi. Examples include Streptomyces species (bacteria) and Penicillium species (fungi). Define: Selective toxicity: The ability of an antimicrobial agent to selectively target and inhibit or kill the pathogen without causing significant harm to the host. Antimicrobial action: The ability of a substance to inhibit or kill microorganisms. Spectrum of activity: The range of microorganisms that an antimicrobial agent is effective against. Tissue distribution/metabolism/excretion: Processes involving the absorption, distribution, metabolism, and elimination of antimicrobial drugs within the body. Effects of combinations: The outcomes and synergistic effects when using multiple antimicrobial agents simultaneously. Adverse effects: Undesirable and harmful effects associated with the use of antimicrobial drugs. Resistance to antimicrobials: The ability of microorganisms to survive and grow in the presence of concentrations of an antimicrobial agent that would normally inhibit or kill them. • What is a “cidal” drug? What is a “static” drug? Cidal drug: A drug that kills the microorganisms. Static drug: A drug that inhibits the growth of microorganisms without necessarily killing them. • Why use combination therapy? What are the benefits? Increase efficacy. Broaden the spectrum of activity. Prevent resistance. Reduce toxicity. Treat polymicrobial infections. • What is a “therapeutic index”? The therapeutic index is a ratio comparing the dose of a drug that causes a toxic effect to the dose that causes a therapeutic effect. It reflects the safety of a drug. • Describe the ß-lactam drugs and other antimicrobials that inhibit cell wall synthesis. ß-lactam drugs: Include penicillins, cephalosporins, carbapenems, and monobactams. They inhibit cell wall synthesis. Other antimicrobials: Vancomycin and bacitracin also inhibit cell wall synthesis. • Describe the antimicrobial drugs that: Inhibit protein synthesis: Tetracyclines, macrolides (e.g., azithromycin, erythromycin). Inhibit nucleic acid synthesis: Fluoroquinolones, rifamycins. Inhibit metabolic pathways: Sulfonamides, trimethoprim. Interfere with cell membrane integrity: Daptomycin, polymyxin B. • Describe how the minimum inhibitory concentration (MIC) and the minimum bactericidal concentration (MBC) are determined. MIC: The lowest concentration of an antimicrobial agent that inhibits visible growth of a microorganism. MBC: The lowest concentration of an antimicrobial agent that kills 99.9% of the bacterial cells. • Compare and contrast the Kirby-Bauer disc diffusion test with commercial modifications of antimicrobial susceptibility testing. Kirby-Bauer: Uses paper discs impregnated with antimicrobial agents to test susceptibility. Commercial modifications: May involve automated systems, broth microdilution, or E-test strips for more precise measurements. • Be able to interpret a disc diffusion assay and an MIC assay Disc diffusion assay: Measures the diameter of the zone of inhibition to determine susceptibility. MIC assay: Determines the lowest concentration of an antimicrobial agent that inhibits growth. • Describe four general mechanisms of antimicrobial resistance. Drug inactivation/enzyme modification. Alteration in target molecule. Decreased uptake of the drug. Increased elimination of the drug. • Describe how antimicrobial resistance can be acquired to specific drugs Through spontaneous mutations during replication. Via gene transfer, often through plasmids carrying resistance genes. • What is the difference between intrinsic and acquired resistance? Intrinsic resistance: Naturally occurring, inherent resistance in some microbes. Acquired resistance: Results from genetic changes or the acquisition of resistance genes • Describe how the emergence and spread of antimicrobial resistance can be slowed. Proper use of antimicrobials. Combination therapy. Global cooperation. Public education. Surveillance and monitoring. • Describe economic factors that are slowing the development of new drugs. High costs and risks associated with drug development. Limited market potential for new antibiotics compared to chronic medications. The challenge of overcoming resistance and finding novel targets. Unit 8: Epidemiology and Disease Transmission Communicable vs. Non-communicable: Communicable diseases are those that can be transmitted from one host to another, such as measles or influenza. Non-communicable diseases do not spread from person to person. Legionella and tetanus are considered non-communicable because they do not transmit directly from person to person but rather have environmental sources. Prevalence: Prevalence is the total number of cases at any time or for a specific period in a given population. It reflects the overall impact of the disease on society. Incidence:Incidence is the number of new cases occurring over a specific period in a given population. It measures the risk of an individual contracting a disease. Case-Fatality Rate:Case-fatality rate is the percentage of the population that dies from a specific disease. It is calculated by dividing the number of deaths by the total number of cases. Morbidity:Morbidity reflects the burden of disease in a population at risk. Mortality Rate: Mortality rate is the overall death rate in a population. Common/Single-Source vs. Propagated Epidemics: Common/Single-Source Epidemic: Rapid rise in cases from exposure to a single source of the pathogen. Propagated Epidemic: Slow rise in cases suggesting contagious disease spreading in the population. Epidemic/Pandemic/Endemic: Epidemic: Unusually large number of cases, usually over a larger region. Pandemic: Global outbreak. Endemic: Diseases constantly present in a population. Outbreak:Outbreak is a higher than expected cluster of disease cases over a specific time in a population. Reservoir:Reservoir is the natural habitat in which a pathogen lives, which could be in or on an animal, human, or the environment. Vector:Vector is a living organism, often an arthropod, that can carry and transmit a pathogen. Contact Tracing: Contact tracing is the process of aggressively tracking down individuals who may have been exposed to a contagious disease to prevent further spread. Vertical Transmission vs. Horizontal Transmission: Vertical Transmission: From pregnant woman to fetus or mother to infant during childbirth. Horizontal Transmission: Person to person via air, physical contact, ingestion of food or water, or vector. Mechanisms of Spread: Fomites: Transmission via inanimate objects. Droplet Transmission: Respiratory droplets from an infected person. Droplet Nuclei: Microbes attached to dried material, remaining suspended in the air. Natural Host/Transmission Host/Terminal Host: Natural Host: The host in which a pathogen typically resides. Transmission Host: The host in which a pathogen undergoes changes but doesn't necessarily cause disease. Terminal Host: The host in which the pathogen reaches maturity and causes disease. Prospective Study/Cross-Sectional Study/Retrospective Study: Prospective Study: Looks ahead and predicts the tendency to develop a disease. Cross-Sectional Study: Surveys a range of people at a single point in time to identify associations between risk factors and disease. Retrospective Study: Compares actions and events following an outbreak to identify causative chain of events. • Explain how characteristics of a pathogen or of a host can influence the epidemiology of a disease. Characteristics of a pathogen (e.g., virulence, dose, incubation period) and host (e.g., immunity, general health, age) can influence the epidemiology of a disease. • Understand the commonalities and differences between descriptive studies, analytical studies, and experimental studies. Descriptive Studies: Collect data to characterize the occurrence of an outbreak. Analytical Studies: Determine relevance of risk factors identified in descriptive studies. Experimental Studies: Involve manipulation and intervention to study cause and effect. • Describe how a common-source epidemic can be distinguished from a propagated epidemic. Common-Source Epidemic: Rapid rise in cases from exposure to a single source. Propagated Epidemic: Slow rise in cases indicating contagious disease spreading in the population. • Describe the zoonotic origin of many emerging human infectious diseases. Many emerging human infectious diseases have a zoonotic origin, meaning they are transmitted from animals to humans. • Know the three main factors that contribute to the emergence of a disease Human, environmental, and microbial factors contribute to the emergence of diseases. • Be able to classify scenarios by the factors that contribute to the emergence of a disease Scenarios can be classified based on factors like human demographics, behavior, climate, economic development, and more, contributing to disease emergence. Unit 9: Immunity and Vaccines Difference between First-line Defenses, Innate Immune Responses, and Adaptive Immune Responses: First-line Defenses: These are physical and chemical barriers, like the skin and mucous membranes, that prevent pathogens from entering the body. Innate Immune Responses: These are immediate, non-specific responses, including phagocytosis by neutrophils and macrophages, inflammation, and the complement system. Adaptive Immune Responses: These are specific responses involving B and T lymphocytes that target specific pathogens. It includes the humoral (antibody-mediated) and cell-mediated immunity. Ways in Which First-line Defenses Protect from Microbial Invasion: Physical barriers like the skin and mucous membranes. Chemical barriers such as stomach acid, enzymes, and antimicrobial substances. How the Immune System Distinguishes Between Self and Non-self: Through the recognition of self-antigens and the elimination or inactivation of immune cells that react against self. Three Pathways of Complement Cascade Activation, Convergence, and Major Outcomes: Classical, lectin, and alternative pathways. Converge at the activation of C3. Major outcomes include opsonization, inflammation, and cell lysis. Major Immune Cell Subsets and Functions: Neutrophils and macrophages: Phagocytosis. Dendritic cells: Antigen presentation. T cells: Cell-mediated immunity. B cells: Humoral immunity (antibody production). PRRs and PAMPs: Pattern Recognition Receptors (PRRs) recognize Pathogen-Associated Molecular Patterns (PAMPs). PAMPs are molecular patterns unique to pathogens that the immune system recognizes. Inflammatory Response: A defensive response to infection or injury. Causes redness, heat, swelling, and pain. Involves vasodilation, increased permeability, and recruitment of immune cells. Consequences include pathogen destruction and tissue repair. Adaptive Immunity vs. Innate Immunity: Adaptive is specific, with memory. Innate is non-specific, immediate. Humoral Immunity vs. Cell-mediated Immunity: Humoral involves antibodies produced by B cells. Cell-mediated involves the actions of T cells. Antigens and Epitopes: Antigens are substances that can induce an immune response. Epitopes are specific regions on antigens recognized by antibodies. Characteristics of Primary and Secondary Antibody Response: Primary response is the first encounter, slower. Functions of Antibodies: Opsonization, neutralization, complement activation, and antibody-dependent cellular cytotoxicity. Antigen Presentation: Displaying antigen fragments on cell surfaces for recognition by T cells. Principles of Active and Passive Immunity: Active: Immune response triggered by exposure. Passive: Transfer of antibodies (temporary). Vaccine Characteristics and Types: Effective vaccines are safe, long-lasting, and induce immunity. Types include attenuated, inactivated, toxoid, recombinant, and subunit vaccines. Sterilizing Immunity and Herd Immunity: Sterilizing immunity prevents infection completely. Herd immunity occurs when a sufficient portion of the population is immune, preventing widespread disease spread. Attenuated vs. Inactivated Vaccines: Attenuated contains weakened live pathogens. Inactivated contains killed pathogens. Advantages and disadvantages depend on factors like stability and safety. Types of Vaccines and Diseases: Purified antigens (toxoids/subunit vaccines) are poorly immunogenic – lack danger signal to activate dendritic cells Necessitates use of adjuvants which mimic danger signal to stimulate an immune response and/or allow slow but constant release of the antigen Example: alum, PAMPs, lipid-water emulsions Seropositive It is an individual that has been exposed o It has produced specific antibodies to pathogen o Seroconversion, process of producing antibodies, it takes about 7-10 days; rise in titer is characteristic of infection (even when microbe not detected directly) Small, steady antibody level indicates previous exposure Seronegative individuals do not have antibodies to a particular pathogen. Unit 10:Principles of Pathogens Define the terms primary pathogen, opportunist, and virulence: Primary pathogen: A microorganism capable of causing disease in healthy individuals, often associated with specific diseases. Opportunist: A microorganism that normally does not cause disease in healthy individuals but can do so under certain conditions, such as a weakened immune system. Virulence: The degree of pathogenicity of a microorganism, indicating its ability to cause disease. Virulence factors contribute to the severity of the disease. Describe genomic islands and pathogenicity islands: Genomic islands: Large DNA segments in a microbial genome that have features distinguishing them from the rest of the genome. They often carry genes related to specific functions or adaptations. Pathogenicity islands: Specific genomic islands in the bacterial chromosome associated with the virulence of a pathogen. They contain genes that contribute to the pathogen's ability to cause disease. How do these islands explain why O157:H7 is deadly and strain HS is not? The presence or absence of specific pathogenicity islands can determine the virulence of a strain. In the case of O157:H7, it possesses pathogenicity islands that encode factors contributing to its virulence, making it more deadly compared to strain HS, which lacks these specific virulence factors. Describe conjugation, competence/transformation, and transduction: Conjugation: The t