Microimmunology Notes PDF

MICROIMM 2500 NOTES UNIT C: BACTERIOLOGY 2 LECTURE 18 — BACTERIOLOGY INTRODUCTION 2 LECTURE 19 — THE BLACK DEATH 6 LECTURE 20 — ANTIBIOTICS AND ANTIBIOTIC RESISTANCE 9 LECTURE 21 — TUBERCULOSIS AND LEPROSY...

MICROIMM 2500 NOTES UNIT C: BACTERIOLOGY 2 LECTURE 18 — BACTERIOLOGY INTRODUCTION 2 LECTURE 19 — THE BLACK DEATH 6 LECTURE 20 — ANTIBIOTICS AND ANTIBIOTIC RESISTANCE 9 LECTURE 21 — TUBERCULOSIS AND LEPROSY 13 LECTURE 22 — BACTERIAL MENINGITIS 17 LECTURE 23 — BACTERIAL STI 21 LECTURE 24 — LYME DISEASE 25 LECTURE 25 — STREPTOCOCCUS PYOGENES 28 UNIT D: OTHER INFECTIOUS AGENTS 32 LECTURE 26 — PARASITOLOGY INTRODUCTION 32 LECTURE 27 — PROTOZOANS I 36 LECTURE 28 — HELMINTHS 39 LECTURE 29 — MYCOLOGY INTRODUCTION 42 LECTURE 30 — MEDICAL MYCOLOGY 47  UNIT C: BACTERIOLOGY LECTURE 18 — BACTERIOLOGY INTRODUCTION Objectives - Gain an appreciation for the diversity of the microbial world - Recognize basic features and structures of the bacteria cell - Understand the differences between Gram-negative and Gram-positive bacteria - Understand what virulence factors are 3 Domains of Life 1. Bacteria → very diverse (look alike but genetic components are not all the same — humans may look different but are almost genetically the same) 2. Archaea → live in harsh conditions 3. Eukaryote Ex. plants, fungi, humans Prokaryotes - Both bacteria and archaea are prokaryotes → smallest, simplest, most abundant cells - Lack a nucleus and complex organelles Bacteria Can Grow Fast - Reproduce by binary fission → asexual reproduction, don’t exchange DNA - 4 phases of growth: 1. Lag → bacteria adapting to environment 2. Logarithmic → exponential growth 3. Stationary → no growth 4. Death → cells die due to competition (need resources) - Generation time → aka doubling time, time for one generation Bacterial Classification by Shape 1. Coccus 2. Rod 3. Spirillum Bacterial Classification by O2 Utilization - Obligate aerobe → requires O2 for growth - Obligate anaerobe → O2 is toxic for growth - Facultative anaerobe → can use O2 if present, but can also grow without O2 - Aerotolerant anaerobe → doesn’t use O2 but is not toxic - Microaerophile → grows best with low levels of oxygen Taxonomic Ranks Ex. Escherichia coli (E. coli) Basic Bacterial Cellular Structure Cell Component Diagram Description Cell Wall (aka - Rigid structure Peptidoglycan) - Prevents osmotic lysis - Glycan backbone - N-acetylglucosamine (G) - N-acetylmuramic acid (M) - Peptide cross-linkage of N-acetylmuramic acid Nucleoid - NOT the nucleus (prokaryotes, like bacteria, do not have a nucleus) - Single, cellular chromosome (most but all all bacteria) - Haploid genomes (one set of chromosomes) Plasmids - Extra-chromosomal genetic elements - Usually not required for bacterial growth - Often encode for ‘fitness’ factors Ex. antibiotic resistance - Does not contain proteins/genes for growth - Can be transferred from bacteria to bacteria Gram +ve vs Gram -ve Bacteria - Gram stain → staining technique used to classify bacteria as Gram +ve or Gram -ve 1. Flood the heat-fixed smear with crystal violet - Purple dye penetrates all bacteria staining them purple 2. Add iodine solution - Forms a complex w/ crystal violet (enhances binding to bacterial cell wall) 3. Decolourize w/ alcohol  - Gram +ve bacteria = purple due to thick peptidoglycan layer - Gram -ve = lost stain due to thin peptidoglycan layer and outer membrane 4. Counterstain w/ safranin (red dye) - Gram -ve bacteria take up safranin and turn pink/red Gram-Positive Bacteria Cell Envelope Gram-Negative Bacteria Cell Envelope Thick Thin peptidoglycan peptidoglycan layer. layer and outer membrane. Stains purple. Loses purple dye. Counterstained pink/red from safranin. The Human Microbiota - Microbiota → free-living cells in the body - Internal organs are usually sterile - “Surface” tissues have extensive populations of microbes - The collective genome of human microbiota easily contains >100 times as many genes as our own genome Host-Microbe Interactions - Commensalism → one benefits w/o helping the other - Mutualism → both benefit - Parasitism → one benefits (usually microbe) at the expense of the other (usually host) Virulence Factors - Virulence factors → molecules produced by the pathogen that helps it produce disease Surface virulence factor → structural components that help bacteria interact w/ host environment or evade immune defence Virulence Description Factor 1) LPS (toxic) - LPS = lipopolysaccharide - Found in Gram -ve bacteria - Triggers strong immune responses - Often causes inflammation or septic shock 2) Flagella - Structures that allow bacteria to be motile (chemotaxis)  3) Pili - Help bacteria attach to surfaces, host tissue and other bacteria 4) Adhesins - Proteins/molecules that bind to host receptors (aid colonization) 5) Capsules - Usually made of (exo)polysaccharides - Attachment to host tissues - Protection from host immune system - Can be used in vaccines - Formation of biofilms → community of microorganisms that adhere to a surface and to each other (form a structured/protective matrix) Stages: 1. Attachment 2. Microcolony development 3. Biofilm development 4. Maturation 5. Dissolution 6) Secretion - Deliver bacterial proteins to host cells Systems 7) Endospores - Resistant “dormant” (allow bacteria to live until conditions improve for growth) structures formed by certain bacteria to survive - Highly differentiated cells formed within the parent cell (NOT reproductive structures) - Resistant to heat, harsh chemicals and radiation - Most common in soil and Bacillus and Clostridium genera Secreted virulence factor → molecules released by bacteria to harm the host/manipulate the environment Virulence Description Factor 1) Exotoxins - Toxins released by bacteria - Act at specific sites in host body (localized) Ex. hemolysins, toxins that function inside host cells, extracellular enzymes, and superantigens - Some inactivated exotoxins can be used as vaccines Some Bacteria are Intracellular Pathogens - Are taken up and survive in phagocytic cells Ex. macrophages - Some ‘force their own uptake into epithelial cells - Allow bacteria to hide from different components of the immune system LECTURE 19 — THE BLACK DEATH Objectives - Recognize that microbes can cause massive changes to entire societies - Understand the known history of the plague - Comprehend the pathogenic mechanisms of Yersinia pestis and the different forms of the plague Yersinia - Yersinia → Gram -ve, rod bacterium - 3 species are pathogenic for humans: 1. Y. enterocolitis → causes “yersiniosis”, a rare cause of diarrhea and abdominal pain 2. Y. pseudotuberculosis → primarily an animal pathogen that can cause TB-like symptoms in animals, enteritis in humans 3. Y. pestis → cause of the plague Yersinia Pestis - Pestis → pestilence (contagious or infectious epidemic disease) - Discovered in the late 1800s - Incubation of 3-7 days - May cause death in 2-4 days by sepsis and/or overwhelming pneumonia w/ respiratory failure - Virulent pathogen but NOT an efficient colonizer of humans 3 Major Forms of the Plague 1. Bubonic plague - Most common form, transmitted by flea bites - Painfully swollen lymph nodes (“buboes”) - Can develop into septicemic and/or pneumonic plague - 40-60% mortality rate if untreated 2. Septicemic plague - Presence of Y. pestis in the blood - Overwhelming and progressive bacteremia - Fleas can pick up Y. pestis and transmit - Patients experience gangrene and disseminated intravascular coagulation - 50-90% mortality rate if untreated 3. Pneumonic plague - Most dangerous - Transmission via aerosols, coughing up blood, or from septicemic plague (spread to lungs in all cases) - Short incubation - 95-100% mortality rate if untreated (must treat within 24 hours of symptoms) Plague Pandemics Throughout Time 1. The “Plague of Justinian” → the first pandemic - Named after the Eastern Roman Emporer Justinian - 6th century (541-542 AD) - Spread to the Mediterranean, Italy, and throughout Europe - ~50% of the population died - Continued in cycles for another 200 years until 750 AD, then disappeared for ~800 years - Killed ~100 million people 2. The “Black Death” → the second pandemic - Originated in Asia and reached Europe in the late 1340s  - Reduced the global population from ~450 million to ~350 million - Killed ~25 million Europeans (⅓ of the total population) - 1348 — people had no idea what was happening - No (real) treatment, only bloodletting which turned fear into panic - People believed it was God’s anger or Satan's influence - Persecution of strangers, minorities and witches - European social order and feudal system were destroyed - Vacant towns and farms; positions of authority to be filled - Demand for physicians, clergy and gravediggers - New opportunities for peasants 3. Mid-19th Century → the third pandemic - Started in China in the 1850s and spread to all continents - Active until 1959 with >12 million deaths in China and India alone - Reached San Francisco in 1990 — infected rats exchanged fleas w/ local wildlife (Y. pestis is now established in the Southwestern US) Did Y. Pestis Really Cause the Black Death? - Dug-up grave sites - Used next-generation sequencing w/ preserved DNA of bacteria Pathogenesis of Yersinia Pestis - Pathogenesis → how disease originates and develops, and the mechanisms that cause it - Live in rodents, transmitted by fleas - Zoonotic pathogen → animal to human - Biofilm formation in the proventriculus (gut) - “Starving fleas” — regurgitation of organisms (bacteria into humans) - Very low infective dose (~10 cells) - Initially survives and grows in innate immune cells - Replicates in lymphoid organs (this is why lymph nodes swell into buboes) - Kills phagocytes and continues to grow extracellularly - At the terminal stage, blood has increased bacterial cell concentration (essential for transmission as fleas take blood meal) Virulence Factors of Yersinia Pestis 1. Type III secretion → inject effector proteins across the host cell membrane to ‘poison’ the host cell by targeting cell signalling pathways - Found in Gram -ve bacteria 2. Phospholipase → break down phospholipids, allows for survival in fleas 3. Plasminogen activator → convert plasminogen, a host protein, into plasmin which is an enzyme that breaks down blood clots - Allows for dissemination and spread throughout the body 4. Yersiniabactin → iron-binding siderophore, scavenges iron from the host (need for growth) 5. Mutated LPS structure in Y. pestis → helps avoid detection by the host’s innate immune system, allowing bacterium to evade immune response - Mutation in lipid A modifying enzyme The Evolution of Y. Pestis - Evolved form Y. pseudotuberculosis - Acquired new virulence plasmids - All pathogenic Yersinia have pYV → encodes for type III secretion system - Y. pestis can infect fleas and is hypervirulent in humans but doesn’t survive well in animals Transmission of Yersinia Pestis - 4 routes for human disease: 1. Flea bite 2. Inhalation (pneumonic) 3. Handling infected animals 4. Ingesting infected meat - Historically, rat-borne urban pandemics, now mostly wildlife-associated plague w/ sporadic outbreaks Diagnosis, Treatment, Prevention - Rapid diagnosis and treatment are essential - Culture and identification may take 4 days - In endemic regions, there are strains and rapid antigen tests - Isolate pneumonic plague patients - Insecticides kill fleas - Give antibiotics (ex. prophylaxis) to exposed individuals Plague as a Bioterrorism Agent - CDC classifies plague as a “Category A” organism - Easily transmitted and high mortality rate - Potential for major public health impact The Plague Recently - Large plague outbreak in Madagascar in Aug 2017 - ~2400 probable/confirmed cases - >200 deaths LECTURE 20 — ANTIBIOTICS AND ANTIBIOTIC RESISTANCE Objectives - Understand how antibiotics work - Recognize why antibiotic resistance develops and how it occurs - Recognize some of the major bacterial pathogens for which antibiotic resistance is a problem - Describe how antibiotic use can lead to pseudomembranous colitis Antimicrobial Agents - Disinfectants → antimicrobial agents that are applied to inanimate objects Ex. floors, tables, walls, etc. - Antiseptics → antimicrobial agents that are sufficiently nontoxic to be applied to living tissues Ex. hand sanitizers - Antibiotics → antimicrobial agents produced by bacteria and fungi that are exploited by humans (delivered topically and internally) Antibiotics - Most effective therapeutic against bacterial infections - Availability enables cancer chemotherapy, organ transplantation, all invasive surgeries, and treatment of premature infants - 2 major problems: 1. Diminished interest from pharmaceutical companies to develop new antibiotics 2. Bacterial resistance to antibiotics always happens Misuse of Antibiotics - Empiric use → antibiotics prescribed before the exact causative organism is identified - Increased use of broad-spectrum agents → kill beneficial bacteria alongside pathogens, creating selective pressure for resistant strains to thrive - Pediatric use for viral infections → antibiotics are incorrectly prescribed for viral infections (unaffected), encourages resistance and disrupts the microbiome - Patients who do not complete course → sub-lethal doses of antibiotics allow partially resistant bacteria to survive and proliferate - Antibiotics in animal feeds → create an ideal environment for resistance to develop in animal gut bacteria, resistance can transfer to humans Measuring Antibiotic Activity Minimum inhibitory concentration (MIC) → lowest concentration of agent that inhibits growth ← ~0.5 - Series of culture tubes with varying - Antibiotic strips concentrations of agent - Faster and can test multiple antibiotics - Check for visible growth at the same time How do Antibiotics Work? - Antibiotics target essential bacterial components: - Cell wall synthesis - Protein synthesis - DNA/RNA synthesis - Folate synthesis - Cell membrane alteration - Targets are not present (or different) in eukaryotic cells βLactam Antibiotics Penicillin - Contains aβlactam ring - Functions to inhibit cell wall synthesis in bacteria - Bind bacterial penicillin-binding proteins (PBPs) - PBPs → transpeptidases, enzymes that catalyze the formation of peptide bonds between aa residues in a polypeptide chain - No peptide cross-links = weak cell wall = bacterial cell death - Unfortunately, some bacteria can produce βlactamase → an enzyme that destroys the ring and therefore the antibiotic Methicillin - Contains aβlactam ring - Chemically modified penicillin - Can’t be cleaved by βlactamases, BUT unfortunately, some bacteria can produce a different penicillin-binding protein Ex. PBP2a encoded by the ‘mec’ gene which does not bind methicillin (or other βlactams) Vancomycin - Vancomycin → drug of “last resort” - A glycopeptide antibiotic, binds peptide linkage at terminal D-Ala-D-Ala residues and inhibits transpeptidation → the transfer of an aa or peptide residue from one compound to another - Inhibits cell wall synthesis in Gram +ve bacteria - Resistance genes change terminal residues to D-Ala-D-Lac which vancomycin can no longer bind - Resistance is encoded by ‘van’ genes Selection for Antibiotic Resistance - Paradoxically, the use of antibiotics selects for antibiotic-resistant bacteria Bacterial Strategies for Antibiotic Resistance - Prevention of antibiotic entry → Gram -ve outer membrane and mycobacteria cell envelope - Antibiotic modification Ex. βlactamase cleaves βlactam ring in antibiotic - Efflux of antibiotic → actively pump out the antibiotic - Alteration of antibiotic target Ex. PBPs, ribosome modifications  - Bypassing the antibiotic action Ex. use environmental folic acid - Folic acid → essential for bacterial growth (makes aa and nucleotides) - Drugs can target folate synthesis pathway if folic acid is produced in the cell - Use of external folic acid means the drug can’t target the bacteria Antibiotic Resistance Genes - Many mechanisms of antibiotic resistance are genetically encoded Ex. mec, βlactamase, efflux pumps - Can produce very high levels of antibiotic resistance - Often encoded on mobile genetic elements allowing for horizontal gene transfer (creates a “superbug”) Horizontal Gene Transfer - HGT → movement of genetic material between organisms in a manner other than reproduction Type of HGT Description Diagram Bacterial - Recipient takes up foreign DNA transformation from the environment and incorporates it into their genome - Not very efficient Bacterial - Bacteriophage (a virus that infects transduction bacteria) takes up host DNA - DNA is then released into the recipient cell Bacterial - One bacteria transfers genetic conjugation material to another through cell-to-cell contact Klebsiella Pneumoniae - Klebsiella pneumoniae → Gram -ve bacteria - Important cause of nosocomial pneumonia → pneumonia acquired during stay in healthcare facility - Produces a capsule and is commonly resistant to multiple antibiotics - First documented source of NDM-1 (New Delhi Metallo-beta-lactamase-1) - NDM-1 is a carbapenemase - Carbapenem antibiotics → βlactamase resistant βlactams with broad-spectrum activity - NDM-1 is now widespread in other Gram -ve bacteria (CRE = carbapenem-resistant Enterobacteriaceae) Clostridia - Clostridia → Gram +ve bacteria, rod-shaped endospore-formers - Strict anaerobes, vegetative, cells killed by O2 - Generally found in soil and intestinal tracts of animals - Important human pathogens: - Clostridiodes difficile → pseudomembranous colitis - Clostridium tetani → tetanus - Clostridium botulinum → botulism - Clostridium perfringens → food-borne illness and gas gangrene - Can cause life-threatening diseases mediated by exotoxins Cloistridioides Difficile (C. Diff) - Can exist as: - Asymptomatic carrier state in the large interesting - Cause of mild to moderate diarrhea - Cause of life-threatening pseudomembranous colitis - Often found in nursing homes and hospital environments (nosocomial pathogen) - Endospores can be very difficult to eradicate from the environment - Mode of transmission is through the spore: fecal-oral route - C. difficile is not considered an invasive bacterium, but the exotoxins cause damage and inflammation to the intestinal lining of the large intestine - Produces A-B toxins called the large clostridial cytotoxins - A-B serves to designate two domains - A domain → the active portion of the toxin that carries the enzymatic activity - Inactivates key regulatory proteins of host cells Ex. cytoskeletal rearrangements, cell death and inflammation - B domain → the portion of the toxin molecule responsible for binding and uptake by the host cell Pseudomembranous Colitis - Symptom of C. diff - Inflammatory conditions of the large interestine - The most important risk factor is having recently received an antimicrobial agent - Supression of normal microbiota + persistence of C. difficile endospores - After antibiotic is stopped, spores germinate, overgrowth of C. difficile occurs with production of toxins - Lesions can enlarge to cover substantial portions of inflamed mucosa and can be stripped off (the pseudomembrane) Diagnosis and Treatment - History (antibiotic use), symptoms and laboratory tests to confirm C. difficile - Endoscopy and toxin detection assays - Discontinue inciting antibiotic if still being used, use antibiotics more specific for C. diff like oral vancomycin or I.V. metronidazole - Avoid antidiarrheal agents — would cause decreased toxin clearance Fecal Microbiota Transplantation - FMT → stool from healthy donor is introduced into gastrointestinal tract of recipient - Goal: restore balanced gut microbiome LECTURE 21 — TUBERCULOSIS AND LEPROSY Objectives - Understand unique properties of the mycobacteria cell envelope - Understand transmission and progression of tuberculosis and leprosy - Recognize methods and limitations for TB testing - Distinguish key differences between leprosy and tuberculosis Mycobacteria (genus) - M. tuberculosis → causative agent of tuberculosis in humans (“TB” for tubercle bacilli) - M. leprae → causative agent of leprosy in humans - M. bovis → causes tuberculosis in cows, rarely in humans - Humans can be infected by the consumption of unpasteurized milk leading to extrapulmonary tuberculosis - M. avium → can cause tuberculosis-like illness in humans, particularly in patients w/ AIDS Tuberculosis - Infection by M. tuberculosis can be latent (can’t be transmitted to others — just in lungs) or active - ~2 billion people (¼ of the world’s population) likely have latent TB - ~10% of people infected with latent TB will develop active TB in their lifetime - ~1.6 million people die from TB each year Mycobacterium Tuberculosis - Intracellular pathogen (lives in macrophages) - Slow generation time of >15 hours - M. tuberculosis can be grown in the lab on specialized media but takes 4-6 weeks to get small colonies - Have an unusual cell envelope with high concentrations of mycolic acid → wax-like structure - Associated with resistance to some antibiotics, osmotic lysis via complement deposition, and lethal oxidative stress promoting survival inside of macrophages - Impermeability to stains and dyes due to waxy, lipid-rich cell envelope (ex. Gram stain) - Use acid fast stain (a hallmark of mycobacteria) - “Acid-fastness” is due to the presence of mycolic acid - Acid fast stain - Stained with carbol-fuchsin dye to slow heating (melt wax) - Washed with ethanol and HCl - Counterstained with methylene blue - Acid-fast organisms appear red, non-acid fast organisms appear blue - Mycobacterium tuberculosis has a Gram +ve acid fast stain and appears red Spread and Progression of Tuberculosis Stage 1: - Transmission is from inhalation of droplets from an infected host Transmission - Coughing/sneezing generates ~3000 droplet nuclei which contain 90 serotypes of the polysaccharide capsule Vaccine for S. - Previously a 7 serotype pneumococcal vaccine (PCV7) Pneumoniae - Now Prevnar 13 (PCV13) → conjugate capsule vaccine from the 13 most prevalent serotypes of pneumococcus - 23-valent polysaccharide vaccine (PPV23) → vaccine that protects high-risk adults against pneumococcal disease, but poorly immunogenic in children (T-cell independent) Haemophilus Influenzae Type B “Hib” - Haemophilus influenzae → Gram -ve, coccobacullus - Primarly causes meningitis in children under 5 - Tends to follow upper respiratory infection, ear infection or sinusitis - Hib conjugate vaccine available as part of the routine childhood immunization schedules (reduced 99% of invasive Hib disease) - Before 1990s H. influenzae tybe b was the leading cause of bacterial meningitis Listeria Monocytogenes “Listeriosis” - Listeria monocytogenes → Gram +ve, rod - Food-borne pathogen - Can cause gastroenteritis, bacteriemia, meningitis - High rates of mortality for immunocompromised individuals - Able to grow at 4℃ → psychrotroph - Invades intestinal epithelial cells and replicates in cytosol - Actin-based motility and cell-to-cell spread - Causes 10 µm) - Major cause of lyme disease in North America - Other species can cause lyme disease — collectively called B. burgdorferi sensu lato - Have perplasmic flagella called axial filaments - Wrap around cells to produce cork-screw shape - Rotation of axial filament allows bacteria to move through extracellular matrix of host tissues and invade vasculature - Unusual outer membrane - No LPS - Many surface expressed lipoproteins act as adhesins (slows down B. burgdorferi to escape vasculature) - Unusual genetic structure - ‘Linear’ chromosome and multiple plasmids (some circular and some linear) - Plasmids are required for infection but are variable - Limited metabolic capability Reservoir for Borrelia Burgdorferi - B. burgdorferi is transmitted to different mammalian hosts through ticks - Found in their vertebrate or arthropod hosts - White footed mice are a major reservoir Ixodes Tick - In North America, B. burgdorferi is transmitted primarily by Ixodes scapularis and Ixodes pacifica - Ticks suck blood slowly for days (appear grey when engorged) - Removal: - Use tweezers and grasp the tick as close to the skin as possible - Wash the area - Keep the tick for testing - Life cycle: - 3 stages: larvae → nymph → adult - No adult to egg transmission (ticks must acquire B. burgdorferi) Lyme Disease Transmission - Ticks do not fly or jump; they sit on grass/shrubs and wait for a host to pass by - Tick inserts a feeding tube with barbs — secretes local anaesthetic - Compounds in tick saliva are thought to inhibit DC function on multiple levels: decreased phagocytosis, maturation, inflammatory mediators, and antigen presentation - Transmission is thought not to occur within ~24 hours (increases >24 hours) - Nymphs typically transmit the disease - Following initial infection, B. burgdorferi must cause disseminated and persistent infection to propagate thorugh its life cycle - Hematogenous dissemination is a central event in the development of Lyme Borreliosis Stages of Lyme Disease Early - Most common symptom is erythema migrans → painless “bulls-eye” rash (Localized) - ~25% do not have rash Stage - Rash will expand if untreated (>12” diameter) - Occurs 1-2 weeks after tick bite - Flu-like symptoms Early - Days to weeks Disseminated - Occurs in untreated patients Stage - Multiple rashes, heart palpitations (interferance w/ heart signals), meningitis (head stiffness and severe headaches, Bell’s palsy (loss of muscle tone in face) - Transverses the BBB through paracellular traversal Late - Months to years Disseminated - Can cause serious long-term disability: muscle pain, arthritis, Stage neurological problems (~5% of patients) - Response to antibiotics takes longer Post-Treatment - 10-20% experience symptoms following treatment (unknown cause): Lyme Disease fatigue, muscle/joint pain, cognitive defect and sleep disturbance Syndrome - May involve an autoimmune response or possibly persistent infection - Most patients recover after a number of months - Long-term antibiotics are not thought to help Lyme Disease in Canada - Ticks continue to expand: migratory birds, warmer climate - More diagnoses because people are more aware of lyme disease Prevention - Avoid wooded areas endemic w/ lyme disease - Stay on paths and avoid low lying brush/long grass - Wear long pants/sleeves and light colours - Use repellents (containing DEET) - Check for ticks and remove them Diagnosis - Typical symptoms (erythena migrans) or tick bite (or reason to suspect tick bite) - Anti-B. Burgdorferi antibody tests - Detect antibodies to a laboratory strain of B. Burgdorferi  - False negatives occur often due to early testing and genetic diversity of B. Burgdorferi - Submit tick for testing to the National Microbiology Laboratory - Note: lyme disease previously under-diagnosed - Previously incidence of ~30,000 cases, CDC studies indicate ~300,000 cases - Case definition too narrow and many symptoms are not properly recognized - Commercial tests not sensitive enough Treatment - If bitten, watch for rash and be aware of symptoms - Patients diagnosed early will recover from antibiotic treatment (2-4 week course) “Chronic Lyme Disease” - Controversial - Some think it is due to chronic infection by B. Burgdorferi - Can be diagnosed w/o evidence of prior lyme disease - Peristant symptoms: fatigue, headaches, sleep disturbance, cognitive dysfunction and other neurological problems - Long-term antimicrobial therapy is not helpful (indicated by 4 clinical trials) Lyme Disease Vaccine? - LYMErix → based on outer membrane protein - Human trial showed it to be 75% effective - Was considered for those in high risk areas but not recommened - Claims that the vaccine caused arthritis (no evidence) - Pulled from the market due to ‘lack of demand’ — now only approved for dogs LECTURE 25 — STREPTOCOCCUS PYOGENES Objectives - Learn about the basic characteristics of the Streptococcus genus - Identify important infections caused by Streptococcus pyogenes - Understand the key role of the virulence factors of S. pyogenes in the development of infections - Review the mouse model of rheumatic heart disease Streptococcus Classification - Streptococcus → Gram +ve, spherical - Non-motile - Non-spore-forming - Typically arranged in chains or pairs - Disseminated into the environment and/or colonizing humans and animals - Recently, an increase of zoonotic and nosocomial infections by different species - There are at least 100 different species divided into three major categories: 1. ɑ-hemolytic 2. β-hemolytic → Ex. Streptococcus pyogenes 3. 𝛾-hemolytic Streptococcus Pyogenes - Aka Group A Strep (GAS) - Human-specific pathogen - Encapsulated and β-hemolytic - Non-invasive or invasive infections Transmission and Epidemiology - Transmission can occur through airborne droplets, surfaces contaminated w/ bacteria and skin contact - 616 million non-invasive and 1.8 million invasive cases diagnosed annually - More than 500,000 deaths each year Non-Invasive Infections 1. Streptococcal pharyngitis “strep throat” - Most common in children aged 5 to 15 years - Symptoms: - Sore throat - Pain swallowing - Fever - Enlarged tonsils - Red spots on roof of mouth - Diagnosis: throat swab, analyzed by rapid strep test that reports results ~15 mins or bacteria culture 2. Impetigo - Common infection and highly contagious - Most prevalent in children 2-5 years old but can occur at any age - Symptoms: erythematous plaques w/ yellow crust, itchy or painful 3. Scarlet fever “scarlatina” - Rash associated w/bacterial pharyngitis  - S. pyogenes exotoxin (superantigens) are mainly responsible - Symptoms: sore throat, fever, red rash w/ sandpaper-like feel and a whitish coating on the surface of the tongue (strawberry-like) Invasive Infections 1. Meningitis - Infection of the meninges - Symtoms: headache, neck stiffness, fever and altered mental status - Diagnosis: positive culture or bacterial DNA of CSF - Mortality rate of 20% - Recent 21-fold increase in cases generated by GAS in adults 2. Necrotizing fasciitis “flesh-eating bacteria” - Subset of aggressive skin and soft tissue infection that causes necrosis of the muscle fascia and subcutaneous tissues Occurs post-surgery, invasive procedure, or minor procedure - Patients are extremely ill and should be transferred to the intensive care unit 3. Streptococcal toxic shock syndrom (STSS) - Manifests with rapid progression and has the potential to culminate in shock, multi-organ failure and with an acute-elevated mortality Strep Post-Infection Sequelae - Individuals may experience multiple infections by S. pyogenes in the throat (pharyngitis, aka strep throat) or the skin (such as impetigo) - Potential downstream consequences or sequelae (secondary health conditions) that can occur after repeated or untreated infections Virulence Factors GAS Pathogenesis 1. Adhesion → GAS attaches to host tissues, initiates infection - M protein promotes attachment to epithelial surfaces - Fbp facilitates adhesion to extracellular matrix components 2. Invasion → GAS invades deeper into host tissues after attachment - DNases degrade host DNA to aid tissue penetration and escape from neutrophil extracellular traps (NETs) 3. Immune evasion → GAS avoids detection and destruction by the host's immune system - Capsule m imics host tissue, avoiding immune detection - Streptolysins (SLO, SLS) lyse host immune cells 4. Dissemination → GAS spreads from the initial infection site to other parts of the body - Spe toxins (s uperantigens) cause widespread immune activation, leading to systemic diseases such as toxic shock syndrome M Protein - M protein → virulence factor related to adhesion and antiphagocytosis - This protein has been used to seroclassify this S. pyogenes - There are >250 different serotypes of M protein Superantigens - Superantigen → molecule linked to the activation of T-cells leading to cytokine storm - There are at least 16 antigens in S. pyogenes - Superantigens are responsible for the establishment of infections and scarlet fever Streptococcus Pyogenes M1UK - In 2019, the incidence of scarlet fever and invasive infections increase 21-fold - These cases were generated by a new sublineage identified as M1UK - It has 27 chromosomal mutations - Characterized by a 10-fold increase in the SpeA superantigen → bypasses specificity of MHC class II T-cell activation by directly linking molecules on APCs to TCRs, regardless of the antigen specificity Streptococcal DNases - DNases → help bacteria evade the host immune system and spread within tissues - Enzymes that degrade DNA, particularly in neutrophil extracellular traps (NETs), which are part of the host's innate immune defense Treatment - 6-10 day course of amoxicillin (β-lactams) — no confirmed reports of β-lactam resistance - Patients w/ amoxicillin hypersensitivity (rash) requiring antibiotics should receive 10 days clindamycin, erythomycin or clarithromycin Vaccines - There is no available vaccine against GAS - Currently 8 candidates in medical trials - S. pyogenes vaccines include the potential for vaccine-induced rheumatic heart disease due to auto-antibodies Rheumatic Heart Disease (RHD) - RHD → heart disease that results from damage to the heart valves (mitral or aortic) - The host's immune system: - Recognizes GAS antigens - Produces antibodies and T-cells against these antigens - Mistakenly targets host tissues (e.g., heart, joints, and brain) that share structural similarities with GAS antigens - Most commonly acquired heart disease - Has autoimmune inflammatory nature - 34 million people were estimated to be living with RHD Humanized Mouse Model - S. pyogenes is a human-specific pathogen - Regular mice cannot be colonized intranasally - Humanized mice possess MHC class II molecule similar to that of humans, allowing infection to establish in mice  UNIT D: OTHER INFECTIOUS AGENTS LECTURE 26 — PARASITOLOGY INTRODUCTION Objectives - Demonstrate an understanding of parasitology as a field of study - Describe general classifications and characteristics of eukaryotic parasites - Describe mechanisms of entry, survival, and pathogenesis for parasitic protozoans - Describe the parasitic life cycle, structures, mode of transmission, symptoms, and treatment of giardiasis What is a Parasite? - Parasite → any organism that derives metabolic benefits by living on or inside a host of a different species - Include animals, plants, fungi, bacteria and viruses - Two groups of parasites: protozoa and metazoa - Protozoa → “first animals”, single-celled microscopic eukaryotes - Metazoa → “after animals, multicellular macroscopic eukaryotes - Parasitology → involves studying the host, parasite and host-parasite relationship, multidisciplinary field Parasitism as a “Lifestyle” Advantages Disadvantages - Once a host is located, no need for - Extreme host specificity can increase further searching vulnerability to extinction - Food permanently available - Must locate at optimal site on/in host to - Limited requirement for complicated food ensure food/survival capturing mechanisms - Reduced need for food processing - Protection from environmental extremes - Must adapt to host’s internal physiology (internal parasites only) - Protection from predators and diseases - Must overcome host’s immune defenses - Reduced need for dispersal because host - Spread limited by host’s geographic (+ vector) carries the parasite range - Can devote larger proportion of energy - Transmission can be extremely risky, and intake to reproductive output than a most offspring die before establishing in free-living organism new host Different Forms of Parasitism - Facultative Parasite → free-living parasites that do not require host to survive Ex. Naegleria fowleri, free-living but can be parasitic to humans (brain-eating amoeba) if in contact - Obligate Parasite → dependent on host for survival Ex. Plasmodium falciparum, causes malaria and is an endoparasite - Endoparasite → survive inside the host - Ectoparasite → survive outside the host (on the surface) Ex. Head lice Parasitic Protozoa - >200,000 species of protozoa have been described; 35,000 currently living; 10,000 species have adapted for life as parasites; ~70 have been isolated from humans  - Free-living species occult every ecological niche - Parasitic protozoans infect a wide spectrum of vertebrate and invertebrate life Classification of Protozoa - Conventional classifications were according to mechanism of motility: - Flagellates (Mastigophora) → have flagella - Amoeboids (Sarcodina) → use protoplasm to move - Apicomplexans (Sporozoa) → complex parasite, glide - Ciliates (Ciliophora) → have cilia - Modern classification schemes consider motility with metabolism and DNA genotyping Mechanisms of Entry Oral Sexual Inhalation Direct Contact Arthropod Vectors Giaria Duodenalis Trichomonas Toxoplasma gondii Trypanosoma Plasmodium vaginalis cruzi falciparum Parasite Hosts and Reproduction - Three types of hosts: 1. Definitive (final) → host in which the parasite reaches sexual maturity and undergoes reproduction (usually sexually) - For parasites that do not reproduce sexually, this is often the organism crucial for completing the parasite’s lifecycle 2. Intermediate → host in which the parasite undergoes asexual reproduction or develops into its next stage but does not reach sexual maturity 3. Reservoir → host that harbours the parasite and serves as a source of infection for other hosts (may be asymptomatic) - There can be asexual reproduction (mitosis/binary fission), sexual reproduction (meiosis) or both - Monoxenous → one host - Diheteroxenous → two different host species, intermediate host uses asexual reproduction - Triheteroxenous → three different host species Survival and Pathogenesis - Antigenic variation → change surface antigens to avoid recognition by the immune system - Molecular mimicry → produce molecules resembling hosts’ to evade detection - Immune modulation → manipulate host immune responses to their advantage, either by suppressing or altering the immune response - Intracellular habitation → reside within host cells to evade immune surveillance - Encapsulation → produce protective structures, such as cysts, to resist host immune attacks and environmental stress - Protease secretion → secrete proteases to degrade host tissues and facilitate invasion - Thermal tolerance → adapt to and survive febrile responses from the host - Toxin production → produce toxins that damage host cells or tissues - Nutrient deprivation → compete with the host for essential nutrients, leading to malnutrition and weakening of the host Giardia Duodenalis Giardia - G. duodenalis → most prevalent protozoan human intestinal pathogen Duodenalis - Synonyms: G. intestinalis or G. lamblia - Binucleated and inhabits the upper small intestine of vertebrate hosts - Obligate parasite with a monoxenous life cycle consisting of two stages, the trophozoite and cyst - Aerotolerant anaerobe (no mitochondria) - Transmitted to hosts through fecal-oral route Trophozoites and Cysts - Adapted for small intestine - Environmentally stable - 8 µm by 12-15 µm - 5 µm by 7-10 µm - Binucleate, both transcriptionally - Tetranucleate, each diploid active - Facilitates transmission - Four pairs of flagella (8 total, - Metabolic rate is only 10-20% of supported by basal bodies) that of the trophozoite - Adhesive disk allows it to adhere - “Dormant” form, to small intestine developed so it can go out - Mitosomes → reduced form of into environment and be mitochondria, assemble of taken up by new host iron-sulfur clusters to support anaerobic functions Life Cycle 1. Low pH and bile in stomach triggers excystation → cyst opponents up and emerges as a trophozoite form in the small intestine 2. Trophozoites adhere to vili 3. Division/replication (asexual) to make more trophozoites 4. Some are repackaged into cyst (encystation) to go into environment and infect more hosts - Why? Trophozoites are unstable and would die in the environment Epidemiology - G. duadenalis is distributed worldwide and - Cysts survive longer in cool, moist environments Pathogenesis - Trophoizoites adhere tightly to small intestine which can result in atrophy and flattening of the villi - Disripts absorption of nutrients which can kead to weight loss , diarhea and long-term digestion problems  - Proteases and secreted substances → excratory secretory products (ESPs) disrupt tight junctions and attack immune cells - Membrane and surface proteins → variant-specific proteins (VSPs) express different proteins to hude from immune system Diagnosis - “Gold standard” for diagnosis consists of microscopy of fecal sample and - Antigen-capture ELISA or direct fluorescent Treatment antibody test (DFA) - PCR → nucleic acid amplification tests (NAATs) - Nitroimidazoles (ex. metronidazole) are primary drugs used to treat - Produces ROS in anaerobes and damages DNA - Safe for our cells since we don;t have redox potential to reduce the drug to its active form LECTURE 27 — PROTOZOANS I Objectives - Indentify the causative agents of malaria - Describe the parasitic life cycle, modes of transmission, examples of survival and pathogenesis, symptoms, and treatments of malaria - Describe an example of coevolutionary antagonism in the context of malaria - Evaluate measures to prevent the incidence of malaria Malaria - Malaria → “bad air”, thought to have come from inhalation - Actually passed from one human to another by the bite of an infected mosquito - According to WHO, there were ~249 million malaria cases in 85 endemic countries/areas and 608,000 malaria deaths worldwide in 2022 (many were children 100 species of Plasmodium which can infect many animal species Four species of Plasmodium have long been recognized to infect humans Zoonotic malaria in nature: P. falciparum P. vivax P. ovale P. malariae P. knowlesi - Most - Similar in function but differ - Quartan - Naturally severe geographically cycle (fever infects infection - Interact w/ different hosts spikes every macaques and blood types 72 hours) which then - P.vivax = Asia infects - P. ovale = West Africa humans - Have tertian cycle (fever - 24 hour cycle spikes every 48 hours) Anopheles Mosquitoes - Human malaria is transmitted by female mosquitoes of the genus Anopheles - Only 30-40 of the 430 Anopheles species transmit malaria (i.e. are vectors) - Females take blood meals for egg production, this links human and mosquito hosts in the parasites life cycle  - Anterior midgut junction → separates blood meal from snack (sugar) — blood goes into separate pouch - Proboscis → used to take blood meals - Salivary glands → chemicals numb site/thins blood - Crop → sugar storage - Dorsal Diverticulum → digestion of sugar - Stomach - Ovaries → for reproduction - Malpighian tubule → “kidneys”, water balance - Anus → excretion of waste - Hemolymph → the circulatory fluid in invertebrates that function like blood in vertebrates and circulate through their open circulatory system P. Falciparum Mosquito Lifecycle 1. Gametocytes in peripheral blood of host 2. Female mosqito ingests gametocytes (sexual form) - Macrogametocyte = egg (female) - Microgametocyte = sperm (male) 3. Zygote formation - Microgametocyte undergoes exflagellation → sperm comes out - Fertilization occurs in stomach and zygote (diploid) is formed 4. Oocyst formation - Ookinete formed when the diploid replicates DNA (becomes tetraploid) and is motile - Oocyst is formed in wall of stomach 5. Sporozoite formation and release - Sporogony → asexual reproductive process of spore formation (haploid) - Cyst ruptures when there are too many sporozoites - Sporozoites released to hemolymph 6. Sporozoites migrate to salivary glands 7. Sporozoites can now infect another human host when infected mosquito takes another blood meal P. Falciparum Human Lifecycle 1. Sporozoites injected during second blood meal - Enters bloodstream and hides in hepatocytes in liver 2. Schizogony → asexual reproduction by multiple fission - Multiple fission → like binary fission but does not undergo cytokenesis - Called cryptozoites → when the malaria parasite is developing in the liver 3. Exoerythrocytic cycle - Infected liver cell (hepatocyte) ruptures and releases merozoites - Cryptozoites undergo cytokenesis and egress into merozoites - Merozoites → circulate in the blood and can infect RBCs 4. Erythrocytic cycle - Ring form → exists when RBCs are infected by merozoites, early trophozoite - Trophozoites → active feeding and growing stage - Makes RBCs sticky (stick to endothelium and causes blockage in vasculature) - Undergoes more asexual reproduction to form schizonts which cause RBC to rupture and merozoites are released into circulation to infect more RBCs 5. Some gametocytes form and are transmitted to mosquitoes by ingestion during blood meal Other Modes of Transmission 1. Host-vector transmission 2. Vertical transmission → mother to child in utero 3. Blood transfusion, dirty needles and organ transplants Symptoms and Diagnosis - Fever, headache and fatigue among others - Diagnosis: blood smear and microscope, PCR, and rapid testing Survival and Pathogenesis - Host invasion - Injected into bloodstream by Anopheles mosquito - Sporozoite entry into hepatocytes - Merozoite entry into RBCs - Immune evasion - Antigenic variation → change antigens/surface proteins - Intracellular hiding - Toxin production - Hemozoin → crystal structure made of stacked heme (toxic to host cells and parasite) produced by parasites that digest hemoglobin — only toxic to host cells Treatment and Prevention - Treatment should be guided by: - The infecting Plasmodium species - Clnical status of the patient - Expected drug susceptibility of the infecting parasite (determined by geographic area where infection was acquired) - Previous use of antimalarials, including those taken for malaria chemoprophylaxis (high-risk individuals take it as a preventative treatment) - 2 drugs: artemisinin and chloroquine, both interact with heme group and prevent hemozoin formation - Vector control + personal protection - Vaccines for children RTS,S/AS01 (Mosquirix) and R21 (Matrix-M) Incidence of Malaria - Density of suitable habitats for mosquito vector breeding - Prevalence of infected humans that mosquitos can feed on - Susceptibility of humans bitten by an infected mosquito Sickle Cell Trait and Malaria Coevolution - Sickle cell gene → a single aa mutation (glutamic acid to valine) in the beta chain of the hemoglobin gene - Inheritance of mutated gene from both parents leads to sickle cell disease - Individuals who are heterozygous carriers have some protective advantage against malaria - When RBC doesn’t have oxygen, some cells will become sickle due to fibre formation - Affects the ability of malaria to invade RBC (can’t use and access hemoglobin) - Frequency of sickle cell carriers are high in malaria-endemic areas LECTURE 28 — HELMINTHS Objectives - Compare characteristics of parasitic protzoans and helminths - Explore how parasitic helminths are classified - Analyze and compare examples of nematodes and cestodes, including modes of transmission, symptoms and treatments - Describe global health, social, and economic impacts of parasitic diseases and comprea potential strategies for prevention, control, and treatment of parasitic infections Parasitic Metazoa - Metazoan parasites → macroscropic multicellular eukaryotes that can express ecto- and endoparasitic lifestyles and can be obligate or facultative - Can vary in size from mm to m in length - Inlcudes helminths (worms) and arthropods (insects and arachnids) Classification of Helminths - Nematodes → non-segmented roundworms - Cestodes → segmented flat worms - Trematodes → non-segmented flat worms Mechanisms of Entry - Fecal-oral → ingestion of contaminated food, water, or contact with fecally contaminated surfaces - Transdermal → enter the host by penetrating intact skin or through open wounds - Vector-borne → transmitted to hosts through a vector (an organism that carries the pathogen but is not affected by it) - Predatory-prey → consumption of infected intermediate hosts or tissues Mechanisms of Survival - Incorporation of host serum proteins on surface — hide from immune response - Inhibition of the complement system - Secretion of anti-inflammatory molecules - Avoiding direct contact with host tissue (ex. living in the lumen of small intestine) - Pausing life cycle when host develops resistance Ascariasis Ascaris - Ascaris lumbricoides → large nematodes that parasitize the small intestine Lumbricoides - 807 million-1.2 billion are infected (most prevalent helminth infection) - Infection occurs through ingestion of eggs via fecal contamination of soil, foodstuffs, and/or water supplies (fecal-oral route) Life Cycle 1. Mature eggs are ingested 2. Larvae hatch in small intestine, enter the bloodstream and go to liver (food source) 3. Larvae migrate to hear and through pulmonary circulation, reach the lung capillaries 4. Larvase enter alveolar spaces - Pressure triggers them to escape 5. Larvae migrate up trachea and are swallowed back into GI tract 6. Adults mature in small intestine - Can burden and obstruct bowels - Eggs pass out in feces and embryonate in soil Symptoms - People rarely show any symptoms and if they do it is usually mild  abdominal discomfort - Problem occurs when disease is untreated and/or there are other concurrent factors Ex. ascariasis and malaria: - Malaria causes a rising fever, worms dont like heat - They try to escape by travelling up through stomach (choking) or down into colon (obstructs bowel) Treatment - Anthelmintic → substance or drug that is used to treat infections caused by heminths - Mechanisms of action include killing parasites and/or expelling them from the host’s body - Albendazole → binds toβ-tubulin and disrupts microtubules which affects energy and nutrient acquisition for parasite - Same family as drug used to treat Giardia - Ivermectin → binds with high affinity to glutamate-gated chloride channels found in nerve and muscle cells of invertebrates - Causes nerve/muscle damage and paralysis - Isolated from bacteria and is also a cancer drug Taeniasis Taenia - Taenia saginata → aka beef tapeworm, large cestodes that parasitize the saginata small intestine - Hermaphroditic → each section has both male and female reproductive systems - Obligate and diheteroxenous (human = definitive, cow = intermediate) - Cows become infected after feeding in areas that are contaminated with Taenia eggs from human feces - Humans become infected with tapeworms when they eat raw or undercooked beef Life Cycle 1. Cysticerci are ingested with raw or undercooked beef - Cysticerci → larvae stage, encapsulated in cyst form 2. Cysticerci are released from muscle in stomach 3. Worms mature into scolex and live in small intestine - Scolex contains four suckers used to adhere to small intestine, get nutrients and migrate 4. Repeating segments called proglottids , begins to grow and mature - Oldest and most distal proglottids, which are gravid (filled with eggs), detach from the tapeworm’s body and are expelled into the environment via the host’s feces 5. Cows ingests embryonated eggs, hatch, oncospheres migrate to tissues, develops to cysticerci - Oncospheres → first-stage larvae of cestodes Symptoms - Most people have mild symptoms or no symptoms Treatment - Treated with anthelmintic medication praziquantel - Hypothesized to disrupt calcium homeostasis resulting in uncontrolled calcium ion influx Why Are There So Many Parasitic Diseases? - We must drink and eat to survive - We must interact with out environment to survive Parasites and Health Equity - Neglected tropical diseases (NTDs) → group of communicable diseases that disproportionately affect impoverished populations in tropical and subtropical regions - Termed "neglected" because they receive less global attention and funding than other major diseases like HIV/AIDS, tuberculosis, or malaria, despite causing significant health, economic, and social burdens Anti-Parasitic Measures - Improve the efficacy, cost-effectiveness, ecological soundness, and sustainability of disease-vector control - Reduction of areas where vectors breed - Access to bed nets and/or other physical barriers against mosquitoes - Introduction of genetically modified mosquitos Wicked Problems - Complex, difficult, or impossible to solve problems that affect people, communities and businesses - Incomplete, contradictory, and changing requirements - Lack of clarity - Real-world constraints - Unknown number of potential solutions LECTURE 29 — MYCOLOGY INTRODUCTION Objectives - Demonstrate an understanding of mycology as a field of study - Describe general classifications and characteristics of filamentous fungi - Describe general classifications and characteristics of yeast - Differentiate general life cycles of filamentous fungi and yeast - Explore the impact and potential benefits of fungi on humans and the environment Evolution of Fungi - Fungi are closer to humans than plants even though humans came to Earth much later - Prototaxites → type of fungus, tallest structures on Earth in the past and allowed evolution of other species - Today, Armillaria exists, a filamentous fungus and one of the largest organisms on Earth Plant vs Fungal Cells Plant Cell Fungal Cell - Cell wall made of cellulose →β(1→4) - Cell wall made of chitin → β(1→4) linked D-glucose linked N-acetylglucosamine - Usually one - May be uninucleated nucleus per cell or multinucleated - Autotrophs → able - Heterotrophs → to produce its own consumer of energy, energy through photosynthesis cannot make its own - Membranes contain phytosterols - Membranes contain - Store food as starch ergosterol in granules - Store food as glycogen in granules - Starch has two - Glycogen is forms: similar to branched starch but (amylopectin) has more and branching unbranched (amylose) Classification of Fungi - Spore formation - Fungal genomics - Modes of nutrition - Saprophytic → feed on dead organisms - Parasitic → living on other living organisms and absorbing nutrients from host - Symbiotic → have have interdependent relationships with other species in which both are mutually benefited - Lichens → fungi + algae relationship - Mycorrhiza → fungi + plant relationship Fungal Morphology Filamentous fungi → characterized by their growth Yeast → single-celled organisms, form, have long, thread-like structures calledhyphae typically oval or round in shape - Macrofungi → edible form, has a fruit body - Unicellular microfungi - Not all produce an edible fruit body - Also called mold - Microfungi → smaller, also called mold Fungal Reproduction Asexual Sexual Parasexual - Most common mode of - Described as - Involves genetic reproduction in fungi teleomorphs recombination without - Described as anamorphs - Sexual propagules are the requirement of - Asexual propagules are produced by fusion of specific sexual considered spores two nuclei that generally structures produced following undergo meiosis mitosis - Produce genetic diversity - Filamentous Fungi: - Vegetative growth → fungi typically grown as filaments termed hyphae (sing. hypha) which extend only at their extreme tips (apical growth) - Hyphae can be septate or coenocytic: - Septate → division between each of the cells - Coenocytic → no division, cytoplasm and structures can move freely - Fungal hyphae branch repeatedly behind their tips, giving rise to a network termed a mycelium - Yeast: - Fission yeast → divide by binary fission - Budding yeast → parent cell and daughter cells will bud off - Pseudohyphae → some species are dimorphic and can switch between a yeast phase and a hyphal phase in response to environmental conditions Aspergillus Nidulans - Filamentous fungus - Produces asexual and sexual spores during its life cycle Asexual Reproduction: 1. Hyphal growth - A spore (conidium) germinates under favourable conditions, forming a vegetative mycelium composed of a network of hyphae 2. Conidiation - When environmental conditions become suitable, they produces specialized  structures for asexual reproduction - Formation of conidia occurs in conidiophores, which are upright, stalk-like hyphal structures - Conidiophores have chains of conidia at their tips which are genetically identical to the parent and are dispersed into the environment 3. Conidia Germination - When conidia land in a suitable environment, they germinate to form new hyphal growth, starting the cycle Sexual Reproduction 1. Formation of Specialized Structures - Sexual reproduction is triggered by specific environmental conditions (often involves nutrient limitations) - Two mating types of A. nidulans must come together 2. Fusion - Two mating types fuse resulting in the formation of a cell containing two genetically distinct nuclei - The two nuclei undergo fusion resulting in a diploid nucleus 3. Meiosis and Ascopore Formation - Diploid nucleus undergoes meiosis, then mitosis, to formeight haploid ascospores in the sac-like ascus - The ascus is produced in the cleistothecia - Ascospores are the sexual spores and are released into the environment where they can germinate and form new mycelium to complete the sexual life cycle Parasexual Reproduction 1. Formation of Heterokaryons - Heterokaryonsare cells that contain two or more genetically distinct nuclei within a common cytoplasm - Nuclei may come from different individuals or mating types 2. Nuclear Fission and Chromosomal Doubling - Two haploid nuclei exist and divide within the same cell, without undergoing meiosis - Haploid nuclei recombine and undergo genetic exchange (genetic diversity) - The nuclei within the heterokaryon may eventually fuse to form a diploid nucleus, generates a diploid organism 3. Chromsome Loss and Haploidization - Diploid nucleus can undergo mitotic divisions - Due to instability, chromosome loss occurs, leading to the return of a haploid state 4. Production of Recombinant Offspring - Resulting haploid cells carry recombinant genetic material that was created during the parasexual cycle Saccharomyces Cerevisiae “Baker’s Yeast” - A unicellular fungus that has a life cycle involving both asexual and sexual reproduction Asexual Reproducion (Budding) 1. Budding - A small outgrowth (bud) forms on the parent cell - The nucleus divides mitotically, and one nucleus migrates into the bud - Bud grows and eventually separates, forming a genetically identical daughter cell 2. Clonal Propagation - The new cell can immediately begin its own cycle of budding, allowing rapid population growth Sexual Reproduction 1. Mating (Conjugation) - Haploid cells secrete pheromones to attract cells of the opposite mating type - Cells undergo morphological changes to form shmoos , which enable them to fuse 2. Fusion - The cytoplasms of the two mating cells fuse, followed by the fusion of their nuclei, 3. Meiosisresulting in a and diploid zygote Sporulation - Under nutrient starvation or other stressful conditions, form haploiddiploid sporescells undergo meiosis to - Each nucleus is packaged into a protective wall, forming ascospores - The four haploid spores are contained in an ascus - When conditions improve, the ascospores germinate, releasing mate or continue haploid sexual cells, which can reproduction Schizosaccharomyces Pombe “Fission Yeast” - A model organism with a life cycle that involves both asexual and sexual reproduction Asexual Reproduction via Binary Fission 1. DNA replication - The haploid cell replicates its genome 2. Nuclear Division - The nucleus divides mitotically, ensuring that each daughter cell receives an identical copy of the genome 3. Cytokenesis - The cell elongates and separates into two equal daughter cells Sexual Reproduction 1. Mating (Conjugation) - Cells of opposite mating types secrete pheromones that attract each other - Morphological changes occur, allowing cells to fuse 2. Fusion - Cytoplasms of two mating cells fuse and their nuclei fuse to form a diploid nucleus - The diploid cell can undergo a brief period of asexual reproduction by binary fission 3. Meiosis and Sporulation - The diploid nucleus undergoes two meiotic divisions, resulting in four haploid nuclei - Each haploid nucleus is enclosed in a spore wall forming ascospores, these spores are packed in an ascus 4. Germination - When environmental conditions improve, the ascospores germinate to release haploid cells - These cells can resume asexual reproduction or undergo mating to restart the cycle How to Survive on Planet Earth - Sporulation! - Allows for dissemination, reproduction, moving to new food source, survival of periods of adversity, introduction of new genetic combinations and source of inocula for infection Benefits of Fungi - Nutrient cycling - Carbon recycling and climate regulation - Nutrition and food security - Human health - Environmental protection - Sustainable materials Fungi as Food Commercial - Mushrooms have been used for food, medicinal purposes or as a means to Cultivation start a fire - There are hundred of species for which some kind of cultivation system is known — all of these species are saprophytes - There are at least 1,100 species of mushrooms eaten in >80 countries Baking - Most common yeast used for breadmaking is Saccharomyces cerevisiae - Feeds on sugars present in bread dough, producing carbon dioxide gas - Other ingredients affect speed of fermentation - Sugar and eggs speed it up - Fats and salt slow it down - Ethanol is evaporated when the bread bakes Brewing - Ferment sugars present in malted barley to produce alcohol - Several different yeasts are used in brewing beet - Saccharomyces cerevisiae is used to make ale-type beers - Bottom-fermenting yeasts (ex. Saccharomyces pastorianus) are used to make lagers Wine-Making - Alcohol in wine is formed by the fermentation of sugars in grape juice, often by Saccharomyces cerevisiae - Sparkling wine is made by adding further yeast to the wine when it is bottled — carbon dioxide formed is trapped as bubbles Cheese - Filamentous fungi are important in the manufacture and ripening of two types of cheese: - Blue-veined cheeses → mix fungi w/ milk culture and let it grow - Soft-ripened cheeses → prepare cheese and let fungi grow on the outside to make a rind Mycoprotein - Scientists and international aid agencies attempted to develop a protein-rich food source from microbial biomass - Developed a novel food product called Quorn, a mycoprotein - It is produced commercially in chemostat cultures of mycelium from the filementous fungi Fusarium venenatum LECTURE 30 — MEDICAL MYCOLOGY Objectives - Describe general classifications of mycoses - Demonstrate an understanding of dermatophytic fungi - Explore examples of yeast and filamentous fungi as opportunistic mycoses - Demonstrate an awareness of endemic, primary systematic mycoses - Examine classes of antifungal treatments for use in human therapy - Identify existing challenges for developing antifungal treatments Health Risks Posed by Fungi - Fungal infections (mycoses) → diseases caused by a fungus — yeast or mold - Mold → filamentous fungi in microfungi state - Only about 200-300 fungi are reported to causes diseases of humans and other warm-blooded animals - Mycotoxins → toxic chemical compounds produced by certain fungi that grow on food crops and other organic materials - Airborne spores can trigger asthma, allergies, or cause occupational diseases Fungal Virulence - >1 billion people are affected by fungal disease; ~1.5 million deaths attributed - Transient exposure to fungi or fungal colonization occurs without the knowledge of the affected individual (no symptoms and self-limiting) - Most fungi have low virulence - Difficult to diagnostically distinguish between presence and infection Classifying Mycoses - Site of infection — superficial (surface of skin), cutaneous (epidermal/dermal layer), subcutaneous (below the dermal layer), or systemic (penetrates blood and travels to different tissues) - Route of acquisition — exogenous (from outside) or endogenous (from within) - Type of virulence — primary mycoses (can infect immunocompetent individuals) or opportunistic mycoses (infect immunocompromised individuals) Dematophytic Fungi - Most superficial and cutaneous mycoses are caused by dermatophytes - Usually caused by molds of the genera Trichophyton, Microsporum, and Epidermophyton - Infections are usually self-limiting; generally no cellular immune response - Can be treated easily using topical antifungal drugs; in severe cases, oral drugs Ex. athlete’s foot or ringworm Opportunistic Systemic Mycoses - Opportunisitic mycoses → exploit the imbalance between the host and pathogen that occurs in immunocompromised individuals - Two most common are yeast species belonging to the genus Candida and molds belonging to the genus Aspergillus Ex. risk factors: HIV infection/AIDS, solid-organ transplantion, old age, use of broad-spectrum antibiotics, premature birth, anticancer chemotherapy, etc. Candidiasis (yeast) Candidiasis - Large proportion of humans carry Candida species on epithelial surfaces - Most frequently associated human infection is Candida albicans - Superficial candidiasis infections include: - Oropharyngeal candidiasis - Denture stomatitis - Vulvovaginal candidiasis - Chronic mucocutaneous candidiasis Virulence - Adhesins → mediate the attachment of Candida to host cells, tissues, and Factors abiotic surfaces - Dimorphism → can transition between yeast (unicellular) and hyphal (filamentous) forms, depending on environmental conditions - Phenotypic switching → undergo reversible changes in colony morphology and surface antigens - Extracellular hydrolases → secretes enzymes that degrade host tissues and facilitate invasion Invasive - Gastrointestinal surgery using contaminated tools Candidiasis - Species land on gut cells, colonize and change morphology - Very narrow and can penetrate/migrate through tissue - Bud of and travel through bloodstream Symptoms and - Symptoms: fever and chills, low BP, muscle aches, skin rash, weakness, Diagnosis fatigue, etc. - Diagnosis: - Selective media cultures - Detection of anti-Candida antibodies and Candida antigens in blood samples - Epidemiology: DNA fingerprinting, microarrays, and PCR Aspergillosis (fillamentous fungi) Aspergillosis - ~20 Aspergillus species are associated w/ human infections - Saprophytic fungi ubiquitous in the environment - Species most frequently associated w/ human infection is Aspergillus fumigatus Virulence - A. fumigatus conidia (spores from asexual reproduction) are very easily in Factors air and routinely inhaled by humans - Immunocompetent individuals: conidia detected and destroyed by macrophages - Immunocompromised individuals: spores can settle, germinate and invade, ultimately leading to invasive aspergillosis - Thermal tolerance → ability to grow at a wide range of temperatures - Proteinase production → secretion of proteolytic enzymes that degrade host tissues - Gliotoxin production → mycotoxin that has immunosuppressive properties - Environmental stress resistance → withstand hostile conditions Invasive - Similar to Candida except is NOT a yeast, its a filamentous fungi Aspergillosis - Aspergillus conidia (spores) are inhaled into the alveoli of the lungs and adhere to alveolar epithelial cells - Conidia transition into hyphal (filamentous) form which is invasive and helps the fungus penetrate tissues - Hyphae begin to invade the epithelial layer, disrupting alveolar structure - The fungus invades blood vessel walls and eventually is transported via the bloodstream to organs and tissues - The fungus establishes infections in other organs, resulting in systemic invasive aspergillosis Symptoms and - Invasive aspergillosis has a high mortality rate (>50%) Diagnosis - Symptoms are non-specific - Gold standards of diagnosis include histopathological analysis and culture of biopsy and bronchoalveolar lavage fluid taken from the infected area of the lung Endemic Systemic Mycoses - Endemic systemic mycoses → mycoses geographically restricted to certain regions Ex. include: histoplasmosis, coccidioidmycosis, blastomycosis, and paracoccidiomycosis Antifungal Agents Plasma Polyenes Inhibit ergosterol directly Membrane Azoles Inhibits 14-ɑ demethylase that converts lanosterol to ergosterol Cell Wall Echinocandins Inhibits β-1,3-glucan synthase ( β-1,3-glucan is an essential component of the fungal cell wall) Nikkomycin Inhibits chitin synthase directly Nucleic 5-fluorocytosine Interacts w/ nucleic acid Acid & synthesis (thymidine) and Protein protein synthesis Synthesis Sordarin Interferes with protein synthesis by binding to fungal elongation factor 2 (eEF2) Existing Challenges - Early diagnosis - Limited antifungal drugs - Cytotoxicity - Antifungal resistance - Emerging pathogens