Microbiology
summary of vaccine types and examples of each
Live attenuated - MMR Whole virus with reduced virulence So similar so give lifelong immunity but can cause some problems Inactivated - Hep A, Flu Dead form of the pathogen Long lasting but not lifelong immunity Subunit - Hep B Strong immunity to the antigens in the vaccine Toxoid - Tetanus Injection of the toxin that causes the disease - no protection against the pathogen itself Conjugate - Pneumococcus, Hib The binding of a "weak" antigen to a "strong" antigen, so that the response to the "weak" antigen is bigger
Which vaccine type induces both T cell and antibody responses?
Live attenuated vaccines
Types of vaccine antigens
Live attenutaed, killed whole organism, purified component, toxoids, polysaccharide conjugates Increasingly, purified component vaccines and polysaccharide conjugates are used
Give pros and cons of live vs inactivated vaccine
Live vaccine pros: oRapid broad, long lived immunity oDose sparing oCellular immunity (B cell and T cell)- best in terms of immunity Live vaccine cons: oRequires attenuation (need to make sure that virus does not replicate and become virulent again), but also not over-attenuate so it's ineffective oMay revert Inactivated vaccine pros: oSafe oCan be made from wild type virus (The naturally occurring, non-mutated strain of a virus) Inactivated vaccine cons: oFrequent boosting required oHigh doses needed
What mycotoxin will you find produced by liberty caps during the British autumn?
Psilocybin produced by Psilocybe
How do antifungals that target DNA synthesis work? Give an example. (brainscape)
Pyrimidine analogues are used to interfere with DNA synthesis. e.g. flucytosine (used for cryptococcus)
What is another way of classifying bacteria apart from gram-positive and negative (brainscape)
intracellular and extracellular pathogens
MERS
Middle East Respiratory Syndrome · Coronavirus · Health tourism and business travel spreading the virus beyond Midde East · High seropositivity rate in camels · Originates from bats and is endemic in camels · Closely related to HKU4 and HKU5 - coronaviruses in bats · Causes ARDS (acute respiratory distress syndrome) in older people but can be asympotomatic, which can make difficult to map spreading · Zoonosis from camels · Uses DPP4 receptor in the lungs. largely in middle east, but can get it in other areas e.g. in korea in 2015.
3 places of replication for bacteria
intracellular, extracellular, obligate intracellular
What are the 3 source and routes of bacterial infection
intrinsic, extrinsic, mythical
Where are most fungal infections acquired from?
Most fungal infections are opportunistic and are acquired from the patients environment.
Bacteridical
kills bacteria
Genetic basis of polio attenuation
know why forward genetic produced vaccine works because of gene sequencing of vaccine, virulent parent and revertant (isolated from child who received vaccine but got ill from vaccine). By comparing these three across the 7500 nucleotides, and sequencing, can see where changes are and see which ones are key ones for generating attenuation. So now can engineer these back into polio virus and use as a vaccine.
Sepsis
life threatening organ dysfunction secondary to infection
Examples of human virus vaccines
live attenuated - smallpox, MMR, flu Inactivated - polio, rabies, hep a Purified subunit - flu Cloned subunit - hep B, HPV
What type of vaccine does rotavirus have? (brainscape)
live attenuated reassortant
An example of a disease that has been unchecked by vaccination
malaria
Describe the type of tropsim exhibited by measles (brainscape)
measles bind to SLAM on immune cells when they enter host, and bind to nectin 4 on airway epithelia when they leave the host
two species of neisseria cause?
meningitis and gonorrhea
The two species of neisseria cause which two diseases?
meningitis, gonorrhea
Phage transduction process
most common) bacteriophages. E.g. many gram neg and gram pos use this · When phages invade bacteria, it replicates its DNA in the bacterium and cuts the bacterial DNA into small pieces · Some bacterial DNA may be packaged into the phage heads · New phage particles are released · The phage particles injects the bacterial DNA from the previous bacterium it infected into the next bacterium. · Injected DNA may be incorporated into the bacterial chromosome.
Zika virus symptoms
new strain evolved causing outbreak in brazil in 2015 fever, rash, joint pain, headache, red eyes, conjunctivitis, diarrhoea
Live natural viral vaccines
(e.g. smallpox - attenuated) A natural virus with its genome inside the capsid, which has had its virulence reduced (attenuated) so it replicates and only produces a mild infection and kick starts the immune response that would've occurred if person got the actual virus and recovered. No associated symptoms.
Cloning vaccines
(live virus vector vaccine, DNA vaccine, Virus-like particle vaccine) o Virulent parts of the original viral genome are cloned inside bacteria o You can then put the DNA into another virus-like particle (e.g. HPV vaccine) = live vector vaccine. Then immune system becomes immune to both viruses o You may just inject viral DNA into people's muscles (DNA vaccine) - so own cells make protein from DNA and then B cells recognise that o Or might express protein in bacteria/ yeast/ food --> eat it/take it and then protein originating from original virus is in system and recognised by immune system ○ Protein might be given as individual pieces or might build protein into particles, because immune system responds better to multivalent particles - lots of copies of same thing all over a structure (e.g. lots of capsid proteins). Viruses are multivalent. Hence, best recombinant vaccines are those built into virus-like particles, which can't replicate/cause disease but LOOK like virus.
Virus quasispecies that occur in host
A collection of isolates (usually viruses) from a common source of infection that have evolved into many different types within one host. · Exist as swarm of quasi-species - viruses replicate with own polymerase that make a lot of errors. Each one copy of genome may contain errors and be different than all the others. So if sequence whole genome will see consensus (most have same at any one position). If condition changes i.e. drug, then quasispecies will change. Is shaped by repeated bottlenecks. Over time of virus in body, diversity increases. · Within host evolution of HIV. The quasispecies that exists within a single infected person contains every single mutation at every single position in the genome. · Within a single infected person, you will get HIV genomes which are slightly different because they evolve while they are in the host. · This is called a QUASISPECIES · Some mutations will improve viral replication and some will emerge to be more predominant than others · Bottlenecks will occur every now and then through which only one or two of the quasispecies genomes will make it through. · The impact of evolution can be seen by transferring the virus into a different environment (e.g. in the presence of a drug) A drug resistant virus may infect a patient who is not taking drugs and so will revert to the wildtype
Virus dissemination (spread) from site of entry
1. Local infection - local infection, apical release (apical is outside, basolateral is inside side of epithelium and closer to blood)- in via apical and out via basal site and out e.g. rhinovirus. 2. Primary viraemia (spread of virus in blood)- dissemination, basal release 3. Amplification - replicate. Systemic infection, haematogenous spread (in blood) or neural spread 4. Secondary viraemia (spread in blood again to another place) 5. Target organ
Influenza replication cycle
1. Spike proteins are used to attach to the surface of the cell (BINDING) 2. It is engulfed and taken into an endosome (ENDOCYTOSIS) 3. Viral protein coat (HA) and intracellular vesicle then fuse and releases its RNA genome (ACIDIFICATION AND FUSION) 4. Genome enters the nucleus where it replicates and directs synthesis of the proteins 5. These (virus genome and proteins) come together to form new virus particles. Which bud out of cell 6. From a single cell - potential 100s of viruses can come out
ebola virus
1. Starts off as a worm like virus outside the cell 2. It attaches to a cell receptor and is engulfed by macropinocytosis (big piece of cell membrane wraps itself around the virus and takes it in) 3. In the early endosome it undergoes various cleavages which allows it to interact with different receptors 4. The genome is released and then copied into new genomes and into mRNA which produces proteins 5. Proteins and genomes assemble and bud out to form new virus particles
Procedure of plaque assay for viruses?
1. Take a sample from the patient that contains the virus (urine, blood, nasal wash) 2. Make serial 10 fold dilutions 3. Take a known volume of the dilutions and put it onto the susceptible cells 4. There will be plaques on the susceptible cells showing how many viruses there are in the dilution (should see 10x less in each one) NOTE: this is sort of like a titration
Genomes of bacterial pathogens can encode between ---- and ---- proteins depending on species
1000 and 5000
What is the average size of a virus? (brainscape)
100nm
Gram negative bacterial structure
2 membranes - peptidogylcan periplasm in-between two membranes. Core lipid-polysaccharide in outer membrane = LIPOPOLYSACCHARIDE (LPS) LPS is extremely potent agonist for our innate immune response. Too much LPS in blood is bad as our immune system, through our toll-like receptors esp. LR4 , are very sensitive receptors for LPS and stimulate pro-inflammatory immune response Both membranes have embedded channel proteins LPS IS A PAMP (pathogen-associated molecular pattern molecules) - microbial molecule that share general patterns that alert immune cells to destroy intruding pathogens • IMPORTANT: LPS IS ONLY PRESENT IN GRAM NEGATIVE BACTERIAL CELL WALLS
what is the size of a virus?
20nm -1micrometre. Smallest being yellow fever to the largest being the megavirus. average 100nm
Preparation of conjugate vaccine
3 preparation pathways leads to different structures of vaccines: o Pathway A: ○ Polysaccharide chain activated at various residues along length and then conjugated to protein --> lattice and matrix o Pathway B and C: Activated at one (pathway C) or both (B) ends --> smaller structure and often single unit of protein coated in polysaccharide chains
What are integrons?
A mobile genetic element that can 'recognise and capture antibiotic resistance genes by site-specific recombination'. Gene cassettes that occur in clusters
Live Attenutated Vaccine
A vaccine prepared from live microorganisms or functional viruses whose disease-producing ability has been weakened but whose immunogenic properties have not.
Epidemic
A widespread outbreak of an infectious disease.
Epstein-Barr virus link with cancer
o A gamma herpes virus, most common virus infection of mankind o 95% of us are infected with Epstein-Barr Virus o Most people --> lytic infection in childhood or infectious mononucleosis (glandular fever) in young adulthood. And then remains latent in B cells. o Passed on in saliva o Some people infected with EBV will go on to develop cancer (combined with other circumstances): · Burkitt's Lymphoma · Hodgkin's Lymphoma · Nasopharyngeal Carcinoma o Incidence of these varies geographically suggesting other predisposing factors to these outcomes e.g. there's link between malaria and EBV.
Non-culture methods in fungal diagnosis
o Antibody and Antigen-based assays can be used to detect fungal polysaccharides: · Glucan · Mannan · Endolase . Proteinase (PEMG) Chitin cannot be detected by the Ig-detected assay. The presence of any of the other proteins can help detect the pathogen that is infecting a host.
Consequences of DNA virus genome
o Can be big (100s kb) because DNA is more stable o There is space for accessory genes - things that viruses don't inherently need to survive but may give some advantage e.g. suppressing the immune system o Accessory genes are lost in culture because there is no immune system in culture - this is a way of producing attenuated vaccines o Some viruses have their genome in one long strand, others have it in several little pieces - SEGMENTED GENOME o EXAMPLE: Influenza has 8 RNA segments; Rotavirus has 11 o BAD - pieces of RNA must be gathered together when the virus tries to leave the cell and move to another cell; impose more difficult packaging strategies GOOD - allows opportunity to pick up new genes and evolve; allows additional easy form of recombination called REASSORTMENT
Examples of opportunistic fungi
o Candida o Aspergillus o Cryptococcus neoformans o Fusarium
Consequences of infection via the GI tract
o DIARRHOEA and vomiting · Dysentery = bloody diarrhoea o Bacteremia/Systemic Infections · Typhoid (S. typhi) · Listeriosis (Lysteria monocytogenes) · Salmonellosis and septic arthritis, aortitis o Toxin-mediated disease · D&V e.g. aureus enterotoxin · Neurological e.g. botulism o Listeria isn't a major cause of diarrhoea and vomiting but if it gets into the blood stream it can cause serious disease in neonates, the elderly and people who are immunocompromised.
What are the different viral groups under baltimore classification? (brainscape)
o DNA viruses (single stranded, double stranded) o RNA viruses (positive sense, negative sense, double stranded) o DNA and RNA viruses (retroviruses, double stranded DNA (RT))
Give some examples of intrinsic bacteria that can infect via the urogenital tract (brainscape)
o E. Coli o Klebsiella o Candida
Give 2 examples of gram-positive opportunistic bacteria (brainscape)
o E. faecalis o Staphylococcus epidermidis
Give some examples of gram-negative pathogenic bacteria and the diseases they cause (brainscape)
o E.Coli - diarrhoea, dysentery, kidney failure o Salmonella - food typhoid, typhoid o Shigella - dysentery o Neisseria - meningitis, gonorrhoea o Vibrio cholera - cholera
Example of mycotoxicoses
o E.g. Psilocybin (psilocybe semilanceata) · Symptoms: · visual distortions of colour, depth and form, progressing to visual hallucinations. · The effects are similar to the experience following consumption of LSD, although milder. · Therapy: · Time! Can potentially be used to treat depression by rebooting brain
Examples of gram negative bacteria
o Escherichia coli (transmitted by food/water/faecal-oral; EPEC - diarrhea, EHEC - produces toxin that goes into blood, dysentry and kidney failure) o Salmonella (typhimurium - food poisoning, typhi - typhoid) o Shigella (dysentery) o Vibrio cholerae (cholera) o Neisseria (meningitidis- meningitis, gonorrhoeae- gonorrhea) - Respiratory, endotoxin that damages RBCs/causes shock. Has polysaccharide capsule (anti-phagocytic)
What is sepsis?
o Essentially, it is SIRS + infection o Sepsis and septic shock are systemic inflammatory responses to infection o Caused by immune and vascular system overreaction and dysregulation o Very complicated S&S: Skin colour/warmth Pain Mental ability Breathing difficulty
What are the two different of portals of entry?
o Expected E.g. normal (harmless) microbiota entering via an expected route (newborn infant and maternal microbiota) o Unexpected e.g. normal (harmless) microbiota entering unusual site OR pathogenic microbiota entering any site
Describe the replication cycle of HIV-1. (brainscape)
o GP120 receptors on the HIV bind to CD4 receptors and bind to co-receptors (CCR5 and CXCR4) allowing the membranes to fuse and the viral contents to enter the cell. o some copies of the virus genome gets replicated o some gets reverse transcribed to viral DNA, which is integrated into the host genome. It is then transcribed and translated into proteins. o the proteins and copies of the genome then assemble to form new virus particles, which exit the cell
Gastro-intestinal to urogenital tract.. to neonates
o Genital tract colonisation with group B strep (from GI tract)- Harmless 30-40% of women ....leading to invasive group B strep infection in neonates: meningitis, septicaemia, death
Give four examples of oncogenic viruses and the types of cancer they cause. (brainscape)
o HHV8 - Kaposi sarcoma o HTLV-1 - leukaemia o Merkel cell polyoma virus - merkel cell polyoma o HPV - cervical cancer
Give an example of co-infections affecting the outcome of an infection. (brainscape)
o HHV8 can cause kaposi sarcoma in people who are infected with HIV and hence are immunocompromsied o Hepatitis delta virus only infects people with Hep B
Give 2 examples of subunit vaccines (brainscape)
o HPV and Hep B virus
Vibrio cholerae pathogenecity
o HUGE INFECTIVE DOSE o Use flagella to penetrate mucus o Makes 2 component toxins A + B o These bind to GM gangliosides on gut o Triggers production of cAMP o CHLORIDE EFFLUX o Sodium ions and water flood out leading to diarrhoea: "rice water stools"
Consequences of infection via broken skin
o If the infection spreads across the skin layer it's called CELLULITIS - you get red inflammation of the skin o Abscess - pus filled pocket o Necrotic Infection - cell death underneath the superficial layer o Myositis - infection spread deeper into the muscle and causes inflammation o Gangrene/Necrotic Infection - any layer of skin or soft tissue can be subject to necrosis o All of these infections can lead to bacteraemia and then sepsis STAPHYLOCOCCUS AUREUS IS THE MAIN CAUSE OF ALL OF THIS
Characteristics of Neisseria meningitidis
o It binds CD46 (complement regulatory protein) o It is host-specific o Tfp is necessary for adhesion o Septicaemia caused by the bacterium is fatal in 10% of cases o Gram negative
What are the two strategies for inhibiting influenza? (brainscape)
o M2 channel blocking o neuraminidase inhibition
Microscopy in fungal diagnosis
o Need to have a well established fungal infection to be able to see it down the microscope o Gold standard for diagnosis o Rapid o cheap
Give examples of some extracellular pathogens (brainscape)
o Neisseria o Yersinia o Staphylococcus o Streptococcus
Gram positive bacteria structure
o ONE cytoplasmic membrane. o cytoplasmic membrane with a THICK LAYER of peptidoglycan on top which retains the dye and stains DEEP VIOLET
Culture methods in fungal diagnosis
o Once it has been identified under the microscope, it can be cultured o This allows susceptibility testing o Positive ID o Slow o Prone to contamination o Requires skilled sample collection
Purified subunit vaccine
o Original parental genome has been taken and treated with proteases to chop it into little pieces o These are subunits of the virus which contains antigens that can trigger an immune response
Why do viruses evolve fast?
o Replicate Fast o Replicate in Large Numbers o High Mutation Rate This makes viral evolution in responses to selective pressure very fast. e.g. Oseltamivir resistance during H51 hospitalisation associated with fatal outcome
sepsis stages
o SIRS - systemic inflammatory response syndrome o Sepsis o Severe sepsis o Septic shock o MODS - multiple organ dysfunction syndrome
severe sepsis
o Severe sepsis = Sepsis + lactic acidosis/SBP<90/SBP drop by 40 or more o Septic shock = severe sepsis + hypotension despite fluid resus o (shock = poor tissue perfusion)
Where would you expect to find bacteria in body?
o Skin - staphylococcus epidermidis, Staphylococcus aureus o Mouth - streptococcus spp o Nasal cavity, sinuses and upper respiratory tract - staphylococcus spp, s. pneumoniae o Stomach - helicobacter pylori, lactobacilli o Small intestine/large intesine - E.Coli, strep group B o Lower genital tract (vagina) - lactobacillus, strep group B These are all: "normal microbiota"
Factors affecting pathogenicity
o Transmission to host o Ability to colonise host o Ability to find unique niche o Ability to replicate o Immune evasion
List all sources of bacteria in different routes of infection
o Upper respiratory tract (intrinsic + extrinsic) o Lower GI tract (intrinsic + extrinsic) oSexual/Urogenital tract (intrinsic + extrinsic) o Skin (intrinsic + extrinsic) oNosocomial/Hospital Acquired (all of the above) o Food and Water oAnimals
List all portals of entry/routes of infection
o Upper respiratory tract (mouth - S. pyogenes/pharyngitis), (S. pneumonia/pneumonia) o Faeco-oral (enteric) - (Vibrio cholerae/cholera) o Urogenital tract - (Group B Streptococcus/neonatal meningitis) o Broken skin - (S. aureus/wound infections) o Intravenous
Consequences of genomes of RNA viruses and retroviruses
o Use their own polymerase to replicate o These polymerases are pretty error prone and these lack proofreading capacity o Leads to a HIGH MUTATION RATE - hence fast evolution o RNA viral genomes are limited in size due to the instability of RNA vs DNA o Largest RNA virus = 30kb - e.g. Coronavirus They often use complicated coding strategies to encode more proteins than expected from a small RNA genome (e.g. some viruses use all three reading frames)
Inactivated virus vaccine
o You take the parental virus and treat it with chemicals and heat to destroy the genome so it is no longer infectious o However, if injected into a person, the viral proteins and shape will still be recognised and an immune response will be triggered o Getting an immune response from this vaccine is more difficult so you may need to add adjuvants
Describe the role of the type III secretion system (brainscape)
o a protein machine assembles which provides a channel through which virulence proteins can be injected into host cell o the virulence proteins then stimulate actin polymerisation and membrane ruffling which allows bacterial internalisation o gram-positive bacteria don't have the type III secretion system
Define virulence (brainscape)
o ability of pathogen to cause disease
State some consequences of infection via broken skin (brainscape)
o abscess formation o bacteraemia o necrotic infection
What is known as the most carcinogenic natural compounds and what produces it? (brainscape)
o aflatoxin produced by aspergillus flavus
At what point does varicella-zoster infection become symptomatic? (brainscape)
o after secondary viraemia
What are the four main mechanisms of antibiotic resistance? (brainscape)
o altered target site o inactivation of antibiotic o decreased metabolism o decreased drug accumulation
Name 7 types of antibiotic (brainscape)
o aminoglycosides - affect RNA proofreading leading to misfolded proteins, some of these proteins get incorporated into the membrane and cause leakage, e.g. gentamycin and streptomycin o beta-lactams - penicillin and methicillin, they have a beta-lactam ring that is a similar shape to a precursor of peptidoglycan in bacterial cell wall and hence interferes with the synthesis of the cell wall o chloramphenicol - inhibits the peptidyl transfer step, binds to the 50S subunit o quinolones - inhibit the function of DNA gyrase (gram-negative) and topoisomerase (gram-positive) hence hampering the unravelling of DNA during replication o macrolides - only gram-positive infections, targets the 50s ribosomal subunit and prevents the peptidyl transfer step, e.g. erythromycin o sulphonamides - Inhibits the folate pathway o tetracycline - prevents amino-acyl tRNAs from binding to the mRNA/ribosome complex
How do aminoglycosides work? Give some examples (brainscape)
o aminoglycosides affect RNA proofreading leading to misfolded proteins o some of these proteins get incorporated into the membrane and cause leakage o e.g. gentamycin and streptomycin
What are the two main phyla of fungi? (brainscape)
o ascomycota o basidiomycota
Describe another way in which actin in manipulated by bacteria (brainscape)
o bacteria (such as listeria and shigella) can polymerise actin at one pole of the bacterium forming comet tails. This polymerisation propels the bacterium through the cytoplasm
What is the main classification of viruses and what is it based on? (brainscape)
o baltimore classification based on virus' genome
What are the main risk factors for systemic candida infections? (brainscape)
o catheters o gut-related surgery o chemotherapy
What is the classic triad of rubella? (brainscape)
o deafness o eye abnormalities (e.g. cataracts) o congenital heart disease
What are some some consequences of infection via GIT (brainscape)
o diarrhoea o bacteraemia/ systemic infections
What are the three main targets for antifungal therapy? (brainscape)
o dna synthesis o cell membrane o cell wall
What are the three phases of growth of a virus? (brainscape)
o eclipse o logarithmic o cell death
What are the two broad types of virus morphology? (brainscape)
o enveloped: membrane derived from host membrane o non-enveloped: protein capside
Motility and invasion require which two multi-protein machines? (brainscape)
o flagella o type III secretion system
How do antifungals that target the cell membrane work? Give some examples (brainscape)
o fungal cell membranes contain ergosterol instead of cholesterol. o some antifungals inhibit ergosterol synthesis o e.g. azole (itraconazole), polyene antibiotics (amphotericin B and nystatin)
What are the good and bad aspect of having a good segmented genome? (brainscape)
o good - allows an opp for exchanges of genetic material and fast evolution o bad - all the segments need to be assembled when the virus leaves the cell
Describe the type of tropism exhibited by HIV (brainscape)
o gp120 on HIV interacts with CD4 on the T-cell o it attaches to co-receptors CCR5 and CXCR4
What is the difference between gram-negative and gram-positive bacteria? (Brainscape)
o gram-negative: thin peptidoglycan layer with two membranes (cytoplasmic and outer membranes) o gram positive: thick peptidoglycan layer, which retains the dye well. Only has one membrane
Describe the type of tropism exhibited by influenza (brainscape)
o haemaggltutinin on influenza binds to sialic acid to gain entry to the cell o sialic acid is ubiquitous, so in theory, influenza can infect all cells o once it's in the endosome, the low endosomal pH allows a massive conformational change that is required for the virus to fuse with the endosome membrane and uncoat o the cleaving of the protein of the virus is essential for the viral genome to gain access to the host cell genetic material o so influenza can only infect cells that have the necessary proteases to cleave the protein o the fluid that lines our lungs have these proteases present
Describe the latency of the herpes simplex virus (brainscape)
o herpes simplex virus can remain latent on nerve cells and then become activated in an attack of shingles o the virus then moves to the end of neurones and replicates through the skin (forming a cold sore)
How can influenza tropism be changed? (brainscape)
o influenza tropism is extended by mutation of the HA cleavage site. o the mutation could change the cleavage site meaning that more proteases, found in other parts of the body, can cleave the protein
How does chloramphenicol work? (brainscape)
o inhibits the peptidyl transfer step o binds to the 50S subunit
State two treatments for hep c (brainscape)
o interferons o ribavirin
What is the best available broad antiviral therapy and what the limitations of it? (brainscape)
o interferons - it activates inflammation and fever and can make the patient feel even more ill
Describe how vibrio cholerae causes disease (brainscape)
o it uses its flagella to propel itself into the mucosal membrane of the intestines o it then begins producing toxins A and B, which bind to GM gangliosides and triggers production of cAMP o this leads to chloride efflux and hence movement of water into the lumen from the cells
What are some common features among DNA viruses? (brainscape)
o larger because DNA is more stable so there is space for accessory genes
What are the four broad types of virus vaccines (brainscape)
o live attenuated o inactivated o purified subunit o cloning
What types of vaccines exit for polio? (brainscape)
o live attenuated (SABINE) o inactivated (SALK)
What are some differences between live attenuated vaccines and inactivated vaccines? (brainscape)
o live attenuated - give rapid, broad, long-living immunity; dose-sparing o inactivated - often require booster, high doses needed, safe
Variation in vertically transmitted DNA can occur due to what?
Mutation
Tuberculosis caused by?
Mycobacterium tuberculosis
Examples of bacteria that can be either gram positive or negative
Mycobacterium tuberculosis (TB) Mycobacterium leprae (leprosy)
Shape of Herpes virus
NOTE: Herpes virus is a combination of capsid AND envelope surrounding tegument (viral matrix), contains lots of viral proteins to take over host cell
How are pathogenicity islands acquired?
Pathogenicity Isalnds are a class of non-core genomes that are acquired by HORIZONTAL TRANSMISSION; they can be revealed by aligning pathogenic genomic DNA and a closely-relate non-pathogen.
When do SARS patients become contagious?
Patients did not become contagious until quite late into the infection once they had become symptomatic (10 days post infection)- you can see who has SARS and isolate them
ASSERTION: Penicillin is effective in treating Aspergillus infections REASON: Penicillin inhibits the synthesis of peptidoglycan.
Penicillin is effective against infections due to bacteria, but NOT fungi, such as Aspergillus. The reason statement is true; penicillin does act by inhibiting the synthesis of peptidoglycans, which are a component of the bacterial cell wall.
Gram positive bacteria appearance
Peptidoglycan (sugar-peptide molecule gives rigidity and shape) in cell wall retains dye. High peptidoglycan = deep violet
How is viral genetic evolution displayed?
Phylogenetic trees
What disease is best associated with the following statement: Two vaccines, Sabin and Salk, for this disease.
Poliomyelitis Sabin vaccine type is live attenuated virus. Salk vaccine type is inactive virus.
What is the issue with treating fungi?
Problem is that our metabolism is similar to fungi
What is chikingunya associated with?
Prolonged arthralgia
Salmonella example of invading bacteria
ACTIVELY INVADING BACTERIA. Swims to pm, injects proteins, form pore, polymerise actin cytoskeleton into filaments, this ruffles the plasma membrane and bacteria get stuck in this ruffle, as process dies down and returns to normal, the trapped bacteria is internalised Salmonella surface structures enable motility (flagellum) and invasion (needle-like structures - to pump virulence proteins into cell.) Salmonella uses evolutionary related protein machines: 1) flagellum (movement) - are filamentous structures and they rotate and are organised as a series of rods and filaments 2) injectisome (Type III secretion system) (causes actin polymerisation by injecting salmonella proteins, membrane ruffling, bacterial internalisation = transfer of virulence proteins into host cells - goes through 3 membranes (2 bacterial membranes and 1 host cell))
What is the most carcinogenic natural compound known?
AFLATOXIN, produced by Aspergillus flavus
In which classes of anti-fungal drugs can resistance occur?
ALL OF THEM E.g. plant resistance caused by anti-fungals used in treatment could cause human fungal resistance. Resistance in azoles increasing through time.
nosocomial infection
Acquired in hospital
Why was smallpox eradicated?
Smallpox virus eradication in 1977: those w/ cowpox didn't get smallpox = basis. Why smallpox could be eradicated: • No animal reservoir (uniquely human virus; not zoonotic) • No latent or persistent infection (is an acute infection - either kills you/ get better and get rid of it) • Smallpox was an easily recognised disease (characteristic pustules --> when saw symptoms did ring vaccination) • The vaccine was effective against all strains of virus (no antigenically distinct strains) • Vaccine properties: Potency, low cost, abundance, heat stability, easy administration (scrape on skin). Live natural vaccine (attenuated) • WHO determination to eradicate it • $250 million cost to get rid of it (SUPER CHEAP)
Where can bacteria divide?
Some bacteria replicate inside blood or in lymph. Other bacteria divide in cells. • This has no correspondence with the gram stain system. • Bacteria invaginate the membrane of a host cell and gains entry into the cell - they remain in a membrane bound vesicle (endosome) and find ways of avoiding being lysed. • Three Methods of Surviving in the Host Cell: o Prevent endosome fusion with Lysosomes - Salmonella, Mycobacteri and Chlamydia o Escape enodosome- Listeria and Shigella break down the vesicle and escape into the cytoplasm o Survive in Phagolysosome - Coxiella can just survive in the harsh environment inside a phagolysosome
What are the different 'ways' to see a virus?
Some viruses can see in microscope if label with FLUORESCENT MARKERS (red pic - little green dots = viruses). Others can see by indirect effect: Picture on RHS is HAEMAGLUTINATION assay, in each well - chicken RBC, if left on own will form dot at bottom; if virus there it causes clumping of RBC and get haemaglutination effect - cheap and more quantitative
What does diagnostic virology depend on?
Source of the specimen Purpose of test (therapy or surveillance) Who will use it and where What stage is the disease (active replication (for acute response) or antibody response)
Why can SARS bind to loads of different receptors?
Spike proteins are HIGHLY PLASTIC and can adapt
H7N9 influenza
Spread from birds (often poultry) to humans and may occasionally spread person to person; more virulent than 1918 strain · 177 deaths since 2013 · Potential to cause a pandemic · Potential to become resistant 5 waves of H7N9 occurred in china - if undergo reassortment could cross human barrier and potentially come to humans
Examples of gram positive bacteria
Staphylococcus aureus (skin diseases, endocarditis, bacteraemia, joint diseases, pneumonia) - Respiratory Release of enzymes & toxins - cause multiple symptoms Streptococcus pneumoniae (pneumonia, meningitis, otitis media) Streptococcus pyogenes (tonsilitis, necrotizing fasciitis, bacteremia, scarlet fever)
Which pathogen causes pneumonia?
Strep pneumonia
Which pathogen causes tonsilitis?
Strep pyogenes
Pneumonia caused by?
Streptococcus pneumoniae
Also known as 'flesh-eating bacterium'.
Streptococcus pyogenes
Example of rapidly dividing bacteiria
Streptococcus pyogenes - responsible for necrotising fasciitis (flesh-eating bacteria) • massive source of genetic variation • rapid generation time + • selective pressure approx 1000,000,000 years worth of evolution
What is candida an example of?
Superficial mycoses
Systemic inflammatory response syndrome (SIRS)
Sytemic response to infection, burns, trauma, or cancer. temp >38C <36, WBC >10,000 or <4000, resting HR>90, RR>20 (consider infection, need 2/4)
What is conjugate vaccine important in stimulating?
T cell dependent antigens. B cells get T cell help due to conjugated polysaccharide. Hence, get boostable response, affinity maturation and mainly IgG response
What are the characteristic sings of systemic fungal infection?
TRICK QUESTION! There are few signs and symptoms in patients that are specific for systemic fungal infection, begin with high index of suspicion.
Can HIV be cured?
Timothy brown cured because had HIV and leukaemia. Was given entire regeneration of immune system by bone marrow transplant. Doctor chose donor who had CCR5- delta 32 mutation, so HIV couldn't dock onto cell and infect. So is now completely negative.
In the transfer of antibiotic resistance, what can move small DNA elements into bacterial chromosomes or onto plasmids? .
Transposons Transposons are sequences of DNA that can move around to different parts of a cell's genome, carrying with them resistance genes. Plasmids are circular, extrochromosomal, autonomously-replicating DNA and integrons are gene cassettes which occur in clusters. Organons have nothing to do with this process . Bacteriophages are bacterial viruses, which can move DNA from one bacterium to another and integrate into the chromosome
What are transpoons?
Transposons are sequences of DNA that can move around to different parts of a cell's genome, carrying with them resistance genes.
ASSERTION: Immunity to influenza is relatively short-lived REASON: New influenza virus variants arise regularly.
True, true - reason is correct explanation
ASSERTION: Some antibiotics are able to target protein synthesis by ribosomes REASON: Ribosomes in prokaryotic and eukaryotic cells are structurally different.
True, true - reason is correct explanation
What type of secretion system is an injectisome?
Type 3 secretion system
Virulence factors
Virulence - features that enhance disease causation - enhance the bugs ability to make you unwell o Toxins o Enzymes (degrade host molecules) o Interrupt normal host processes o Complete immune evasion
What is found inside the pathogenicity island?
Virulence genes and some insertion sequences
Virus replication cycle
You start with the virus on the outside where it is inert, then finds cell 1) Entry: The virus will have a protein on its coat called the viral attachment protein which binds to a receptor on the host cell surface · This allows the virus to dock down onto the cell surface · Some viruses enter the cell by fusing with the host cell membrane and injecting their DNA or RNA into the cytoplasm · Other viruses enter via a series of vesicles such as endosomes 2) Entry: Once the virus is in the cell, it needs to uncoat/fall apart so genome and anything else exits · It must be made into messenger RNA if it isn't already 3) Translation: The viruses uses the host ribosomes and translational machinery to translate its own mRNA and produce proteins (early (regulatory) proteins and late (structural) proteins, which may form capsid of new virus) 4) Replication: At the same time the virus begins to replicate its own genome either using its own polymerase or host cell polymerase 5) Assembly and exit: The copies of the genome and the newly synthesised viral proteins assemble to form new virus particles which then leave the cell (by bursting/budding)
Zoonosis
a disease which can be transmitted from animals to humans
Fungal mycotoxicoses definition
a toxic reaction due to the ingestion/inhalation of toxins produced by fungi
Define infectivity (brainscape)
ability of pathogen to establish infection
antibiotic
antimicrobial agent produced by a microorganism that kills or inhibits other microorganisms. Most antibiotics in use today are produced by soil-dwelling fungi (Penicillium and Cephalosporium) or bacteria (Streptomyces and Bacillus). However, antibiotics commonly used today encompass a range of natural, semi-synthetic and synthetic chemicals with antimicrobial activity.
Development of antibiotic resistance
antimicrobial therapy provides a selection pressure for the spread of antimicrobial resistance Some isolates of S. aureus were resistant to penicillin from the start! Routine use of penicillin provided selective pressure for the acquisition and maintenance of resistance genes.
3 domains
archaea, bacteria, eukarya
Why are viruses difficult to target?
are OBLIGATE INTRACELLULAR PARASITES - many things that viruses do, you do e.g. protein synthesis. Cannot target these without harming host. Viruses don't have as many distinctive features as bacteria which we can target. There isn't very much that is UNIQUE to a virus They're not as difficult as developing cancer drugs, but harder than for bacteria and poisons in animal pests
What type of fungal infection is a big problem in transplant settings? (brainscape)
aspegillosis
acid fast bacteria
bacteria that are resistant to the Gram staining method and are therefore stained with the acid-fast stain method
Listeria example
causes food poisoning and more serious diseases in the immunocompromised, elderly and pregnant women. Listeria bacteria can swim around and replicate in cell cytoplasm by using actin cytoskeleton - it causes actin polymerisation for this reason. • It breaks out of the vacuole • It then assembles/polymerises actin at one pole of the bacterial cell • This polymerisation of actin generates force which propels the bacterium through the cytoplasm • This leads to the spread of the bacterium from one cell to another • These streams of actin are known as COMET TAILS Listeria and Shigella can both do this Overall: invasion, intracellular movement, cell-cell spread
antiseptic
chemical that kills or inhibits microbes that is usually used topically to prevent infection.
antimicrobial
chemical that selectively kills or inhibits microbes (bacteria, fungi, viruses
Varicella Zoster Virus
chicken pox and shingles 1. Virus enters the body through the respiratory route 2. VZV can infect many cell types including peripheral blood mononuclear cells (PMBCs) and skin cells 3. Leads to mild self limiting illness in most childhood cases 4. From the skin site it can infect terminally differentiated non-dividing sensory neurones in PNS where it remains latent 5. Virus can be reactivated in sensory neurone when cellular immunity is impaired in adulthood causing a painful rash at nerve endings - SHINGLES or herpes zoster · You only get symptoms after secondary viraemia Incubation Period: 14 days
Examples of obligate intracellular replicating bacteria
chlamydia and coxiella
Subcutaneous mycoses definition
chronic, localised infections of the skin and subcutaneous tissue following traumatic implantation of the aetilogic agent
What are subcutaneous mycoses? (brainscape)
chronic, localised infections of the skin and subcutaneous tissue following traumatic implantation of the aetiological agent
rhinoviruses
common cold
Endemic
confined to a particular country or area
What are the three mechanisms of horizontal gene transmission? (brainscape)
conjugation, transduction, transformation
Conjugate vaccines
created by attaching bacterial capsule polysaccharides to a immunogenic protein because polysaccharides on bacteria are poorly immunogenic Polysaccharides are T cell independent antigens - they directly cross-link to Ig receptors on B cell surface, which stimulates them to produce Ig - but these have low avidity due to haven't had the benefit of affinity maturity. Doesn't require T cells and doesn't lead to good immune or memory response. T cell recognition of protein carriers enhances B cell activation Promotes efficient antibody response to polysaccharide capsule
Which fungus out of the basidiomycetes causes the largest burden of disease? (brainscape)
cryptococcus neoformans and cryptococcus gatii. cause cryptococcal meningitis
What is the cytopathic effect? (brainscape)
death of a cell as a result of being infected by a virus
Other names for cutaneous mycoses
dermatophytocis and dermatomycosis
What is tropism of HIV dependent on?
determined by receptor use Primary receptor is CD4 so HIV infects T cells · The viral gp120 on HIV interacts with CD4 on the T cell · It then attaches to a co-receptor (CCR5 or CXCR4) · Some people have mutation in CCR5 (delta 32 = 32 nucleotides deleted in CCR5 gene --> out of frame) which means they don't make a proper CCR5 receptor and hence are resistant to HIV · Some people who are exposed and unaffected produce a large amount of chemokine which blocks the use of the co-receptors · You can get a tropism switch during HIV replication - viruses evolve to bind to other receptors (e.g. from CCR5 to CXCR4) CCR5 (macrophages)and CXCR4 co-receptors (t cells): Chemokine receptors that sit on immune cells. In general viruses transmitted through mucosal routes (e.g. unsafe sex) and macrophages, which are first cells to get infected in HIV event, hence if no CCR5 but have unprotected sex will not get infected. BUT if have blood transfusion, then HIV may enter into T cells in blood directly through CXCR4 co-receptor.
reverse vaccinology
development of new vaccines using only specific proteins of pathogens Look at genome sequence, predict antigens, produce recombinant antigen, try in mouse , develop in vaccine in clinical trials for people. Used when developing new men b vaccine
Septic shock
develops after sepsis (40% mortality) as opposed to 10% of just sepsis. require innate immune response to fight sepsis
What form the edges of pathogenicity islands?
direct repeats (2 or more repeats of specific sequence)
Define mycosis (brainscape)
disease caused by fungi that is classified based on the level of tissue affected
Mycoses definition
disease caused by fungi which is classified by the level of tissue affected (superficial , cutaneous - skin, subcutaneous - under skin, systemic) E.g. frog virus swimming fungus starts at skin, burrows and secretes proteases to become subcutaneous and disrupts skin function.
commensals
don't cause disease e.g. lactobacillus in vagina and GI
Shigella causes?
dysentery
What does shigella cause?
dysentery
What do segmented genomes in viruses allow?
easy recombination e.g. rotavirus, flu
What is cell-mediated immunity particularly important for?
eliminating intracellular bacteria e.g. TB, typhoid, legionella § Interaction of the reactive T lymphocytes and the macrophage is key to clearance of infection o In reality, protection is often result of combo of humoral and cell-mediated immunity
Bacteria can divide as quickly as...
every 20 minutes
How can influenza tropism be extended?
extended by mutation at the HA cleavage site · Some chicken viruses (H5, H7) have mutated the region where the protease cleaves · This allows it to be cleaved by different proteases (FURIN which is ubiquitous) found in other areas of the body - deadly as infects every cell and can be activated everywhere · Infected chickens can allow virus replication in every organ of the body
How do fungi digest their food?
extracellularly · They produce hydrolytic enzymes which are pumped out into the environment · They are powerful polymer degrading substances which rot the material around them - so the fungus is effectively suspended in its food source, they then absorb products This means that they are SAPROPHYTES
Virulence
features that promote infection and damage (e.g. toxins, immune evasion). capacity to cause disease
Hep A transmission route
fecal-oral
Good things about fungi
food, carbon recycling, symbioses e.g. algae and fungi, produce antimicrobials, yeast used in making wine
Intrinsic source meaning
from inside body
Extrinsic source meaning
from outside body e.g. ingestion of contaminated water or food, coughs, sexual transmission, nosocomial
Summary of modes of viral transmission
germline, respiratory, oral, fecal-oral, blood, urine, direct skin contact
BCG vaccine
gives some protection against tuberculosis in children but is ineffective against adult pulmonary disease new vaccine strategies based on genome information very effective live attenuated vaccines are being developed for enteric pathogens (e.g. Salmonella typhi)
What components are found in fungal cell walls but not in mammalian cell walls? (brainscape)
glucan and chitin
What kind of bacteria do you find in the SI? (brainscape)
gram-negative only
Majority of bacteria are..?
harmless or beneficial (commensal)
All human viruses...
have enveloped virions into which are inserted spike proteins
how are most accessory genes gained?
horizontal transfer from (mainly) unknown sources
What is a pathogenicity island? (brainscape)
horizontally acquired genes that contribute to virulence
Define virus (brainscape)
infectious obligate intracellular parasite
What are arboviruses
A class of viruses transmitted to humans by arthropods such as mosquitoes and ticks. · EXAMPLES: o Yellow Fever o Dengue o West Nile o Zika oChikingunya
Preventing emergence of drug resistant bacteria and nosocomial infections
Adressing resistance • Prescribing strategies - tighter controls, temporary withdrawal of certain classes. Restriction of ABs for certain serious infections • Reduce use of broad-spectrum antibiotics • Quicker identification of infections (diagnosis) caused by resistant strains • Combination therapy - use more than 1 antibiotic. Difficult because some are antagonistic (might not work well with others) and poor evidence . Knowledge of local strains/resistance patterns Overcoming resistance: • Modification of existing medications to e.g. Prevent cleavage (beta-lactams) or enhance efficacy. E.g. Methicillin. • Combinations of antibiotic + inhibitor of e.g. Beta-lactams. E.g. Augmentin. • However, this is a reactive approach in response to emergence of resistance! Infection-control measures work!!
Name a highly carcinogenic mycotoxin and what fungus produces it?
Aflatoxin produced by Aspergillus flavus
Vaccination basis
After a week - get immune response from effector T cells --> memory, so next time w/ same pathogen get more potent and rapid response --> mild/no symptoms Successful virus vaccination: In 1940s, incidence increasing due to people moving to cities and sharing water (polio spreads by water) , then decreased with vaccines and now near eradication
Actions of reverse transcriptase in HIV replication.
After the viral capsid has entered the cell, Reverse Transcriptase liberates the single-stranded RNA from attached viral proteins and produces complementary DNA (cDNA), which then becomes double-stranded DNA.
germ line transmission
Agent is transmitted as part of the genome. Part of the host genome (eg intergrated retrovirus)
What do Polio, Measles, Influenza and Human Immunodeficiency Virus (HIV) have in common?
All are single stranded RNA viruses
Horizontal transmission
All other forms of transmission but from parent to offspring
Summary of fungal pathogens
Allergies E.g. Rhinitis, Dermatitis, Asthma Sensitisation to the spores -> allergic reaction Mycotoxicoses Inhalation or ingestion of a mycotoxin (secondary metabolite of moulds) Typically eating poisonous mushrooms Certain crops can lead to production of Aspergillus flavus, which produces Aflatoxin, which causes around 28% of Hepatic cell carcinomas worldwide Mycoses: Candida Recent increase in infections Superficial - Rash, in all age groups, responds readily to treatment Mucosal - Occurs in HIV/immunosuppressed patients particularly Systemic - Not seen in healthy individuals, associated with catheters, gut surgery, and chemotherapy
Alzheimer's link with viruses
Alzheimer's recent article- amyloid beta is an antimicrobial response that coats herpes virus glycoproteins. Herpes simplex virus (HSV) in neurones stimulates amyloid beta production and plaques; but if virus reaches brain then plaque form and lead to alzheimers. So now trying acyclovir (anti-HSV drug) to see if slows alzehimers
Koch's Postulates
Carry out this technique to prove that a virus causes disease - norovirus was proven to be the cause of disease using Koch's Postulates 1. The microorganism must be found in large numbers in all diseased animals, but not in healthy ones. 2. The organism must be isolated from a diseased animal and grown outside the body in a pure culture. 3. When the isolated microorganism is injected onto other healthy animals, it must produce the same disease. 4. The suspected microorganism must be recovered from the experimental hosts, isolated, compared to the first microorganism, and found to be identical. E.g. used in yellow fever when it was spread by mosquitoes during the construction of the Panama canal. But these days tend to use molecular basis.
Chikingunya
Chikingunya is another type of arbovirus. Is an alpha virus. Get muscle aches and- joint pain. Different to dengue fever which is acute. Chikunguya is more chronic.
The following fungus is not opportunistic
Coccidioides immitis
What flanks the pathogenicity island on either side?
Core host genes
What type of virus is SARS?
Coronavirus - positive-sense RNA genome · Enveloped spike protein Receptor in human: ACE-2
Out of the basidiomycetes, what is the largest burden of disease?
Cryptococcal meningitis caused by Cryptococcus neoformans and Cryptococcus gatii
Name the basidiomycete fungal pathogen that causes meningitis
Cryptococcus neoformans
Gram negative bacteria appearance
Dye is lost from thinner peptidoglcan The cells absorb counterstain making them appear pink.
Non-pathogenic forms of this bacterium are part of the normal intestinal flora
E. Coli
Risk benefit analysis of vaccinating the immunocompromised
E.g, when giving attenuated vaccine to someone immunodeficient - need to see whether attenuated vaccine will actually be able to cope with it and not get infection. If give component or conjugate vaccine there is no risk of the infection
How can AB therapy impair commensal flora (microbiota)?
E.g. C.difficile found in body and is usually kept at bay by microbiota. AB affects microbiota and then get it. In health, commensal organisms can out-compete pathogen WRT adhesion, metabolism, growth. Pathogen cannot colonise at levels sufficient for infection. Trying to reduce broad spectrum antibiotics due to affect microbiota but difficult because broad spectrum targets many diseases
ebola virus shape
EXAMPLE of typical shape virus:
Sepsis Treatment
Early detection is key Target the infection Antibiotics - broad spectrum and then organism specific Blood products - more RBCs -> better perfusion Try to increase the BP to prevent shock: Fluids Vasopressors
In which stage of VIRUS REPLICATION does the replication of the nucleic acid occur?
Eclipse Phase. During the Eclipse Phase, virus proteins are expressed and nucleic acids are replicated- this is highly regulated!
Invasive aspergillosis
Emerged as the major clinical problem of common mycology e.g. aspergillus fumigatus Get in lung brain kidney heart Get haziness in lung - a ball of fungus that needs to be surgically removed It is responsible for 60,000 deaths per year worldwide Susceptible patients · Neutropenic / transplant · HIV/AIDS · Diabetic · Congenital susceptibility e.g. CARD-9, Dectin-1, IL-17 · Influenza · Leukaemia - due to tyrosine inhibitors given in chemo, but these can knockdown macrophage response causing susceptibility
Endemic, epidemic, pandemic meaning
Endemic: a disease that exists PERMANENTLY in a particular region or population. Malaria is a constant worry in parts of Africa Epidemic: An OUTBREAK of disease that attacks many peoples at about the same time and may spread through one or several communities. Pandemic: When an epidemic spreads throughout the world. GLOBAL.
What do fungi use instead of cholesterol in cell membrane?
Ergosterol
Bacterium that use a Type III secretion system to inject its own receptor into the plasma membrane and leads to diseases such as 'traveller's diarrhoea'.
Escherichia coli
What group are fungi part of?
Eukarya They have a similar metabolism to us - so anything that works on yeast is likely to have a similar effect on us
Explain individual differences and the genetic role on susceptibility to infection
Factors: o Genetic - Genetics determines susceptibility to particular pathogens E.g. HIV, Hep B/C, Malaria Main genetic factor is HLA/MHC Determines which antigens are recognised o Microbiome - Gut, skin o Nutrition - Vit A o Social networks o Stress - negative effect o Splenectomy o Immunosuppression - transplant, autoimmune
ARQ: A: Bacterial type 3 secretion systems deliver DNA for horizontal gene transfer R: Horizontally acquired DNA contributes to bacterial virulence
False, true Type 3 secretion systems in injectisomes of bacteria are used when injecting host cell to insert virulence proteins in order to be taken up by cell.
ASSERTION: Influenza virus does not undergo recombination REASON: Influenza virus has a segmented genome.
False, true.
What is herd immunity
Form of immunity that occurs when the vaccination of a significant portion of the population provides a measure of protection for individuals who have not yet developed immunity. R0 = basic reproduction number of organism in vaccine effectiveness
Cryptococcus diagnosis
From lumbar puncture to dipstick diagnosis. Cheap and quicker result. But antifungal drugs after are expensive
Vertical transmission
From parent to offspring
Ebola outbreak 2014
Fruit bat in tree --> everyone. To prevent community spread of ebola in nigeria they traced contact and contained it. Hugging is a bit part of religious worship --> can lead to spread When humans, bats and domestic animals come together --> hub for viruses.
General strategy of designing drugs which target infectious agents but not patient's own cells
General strategy (rational drug design): 1) Identify elements of viral life cycle that differ from host's 2) Identify the site of drug action 3) Design the drug to be highly specific for the site 4) Block spread/ replication/ dissemination of the virus
Which type of bacteria don't have injectisome?
Gram positive
How to identify different bacterial cell walls
Gram stain.
Example of zoonoses that have lead to pandemic?
HIV
examples of viruses whose tropism is determined by receptors
HIV and measles
What part of flu virus binds to sialic acid?
Haemagglutinin spike protein
Iatrogenic transmission
Health Care workers eg contaminated needles
Genetically-engineered subunit vaccine that in 1989 replaced the previous vaccine made from human plasma.
Hepatitis B
Examples of subunit vaccines
Hepatitis B virus sAg (surface antigen) cloned and expressed in yeast; easy to make multiple copies and give vaccine. Recommended for healthcare workers as it is blood transmitted Human Papillomavirus (cause warts, some cause cervical cancer) Virus Like Particles from recombinant coat proteins Gardasil and Cervarix
What type of cancer do hepatitis B and C cause? (brainscape)
Hepatocellular carcinoma
What virus is the most common cause of cold sores around the mouth?
Herpes Simplex Virus 1 (HSV-1) causes cold sores around the mouth and other parts of the face. It can be spread by skin-to-skin contact.
Acyclovir and cidofovir are used to treat what?
Herpes virus
Examples of conjugate vaccine
Hib - Haemophilus influenzae type b, Men C, pneumococcal
HAART
Highly Active Anti-Retroviral Therapy. uses three or four different drugs in combination. This prevents the virus from being able to generate resistance mutants but does lead to difficult drug regimens and is associated with significant side effects.
What is a pathogenicity island?
Horizontally Acquired DNA that contributed to virulence - either by encoding for injectisome, producing toxin or something like that to bacteria. Are critical for causing disease. These are the main drivers of evolution of bacterial pathogens and their origin is usually unknown. Flanked on either side by core host genes. Found near trna genes. On edges of island are direct repeats. On the inside are virulence genes and insertion sequences.
Pathogenic candida species
Hot topic - candida auris - new species arising in world. Trying to do genome sequences to see epidemiological connections. Is very resistant to drugs and disinfectants
Injectisome structure and function
IMPORTANT: bacterial virulence proteins (effectors) induce actin polymerisation, membrane ruffling and bacterial internalisation • Hence the type III secretory system is needed to deliver the effectors, which allow the internalisation of the bacteria. • This needle structure is evolutionarily linked to the flagellum • Instead of making a flagellum, a bacterium can make this needle shaped structure, which allows effector proteins to be injected into the host cell through translocase. • Many gram negative bacteria have this feature but NO GRAM POSITIVE BACTERIA HAVE THIS • This is a translocon, which inserts into the plasma membrane of the host cell. • By making a passage into the cell, it allows effector proteins to be transferred from the bacterial cell to the host cell. • The effector proteins lead to actin polymerisation, which causes the uptake of bacteria into the host cell. Actin polymerisation is induced by Salmonella proteins --> mimic activation system to cause membrane ruffling and bacterial internalisation
How is vaccine safety and efficacy assessed?
In clinical trials. Phase 1 - safety Phase 2 - efficacy - assess immune response Phase 3 - larger scale. placebo double-blind trials
Where do gram positive bacteria retain the violet iodine dye?
In peptidoglycan of the cell wall
What are targets for antiviral drugs?
In reality most antiviral drugs target viral enzymes ( as easier to design), often acting as substrate analogues (look like substrate but different so mess up the enzyme pathway), or are cancer-like drugs- nucleoside analogues (block replication). nucleoside or substrate analogues
Label an Human immunodeficiency virus.
Include RNA , reverse transcriptase, p18 (viral envelope), p24 (capsid), gp41, gp120 (glycoproteins). Size = 110 nm
Infectious dose is affected by
Infectious Dose is affected by: · Route of Transmission - e.g. stomach acid means that higher infectious dose is usually required, hence people who take antacids might be more at risk of infection · Ability to Colonise Host - EPEC (enteropathogenic E. Coli type 3 secretion for adherence) · Tropism and Motility - vibrio cholera- high infective dose (around 10 million CFU) but falgella motility may help reach gut epithelium · Replication Speed · Immune Evasion - Mycobacterium TV have low infective dose (around 10 CFU). Low replication rate. Able to survive inside macrophages and resist killing NOTE: if the pathogen can kill or subvert phagocytes then their infectious dose is lower
Why is rubella harmful in early stage pregnancy?
Infects dividing fetal neuronal tissue leading to deafness
Rimantidine and Tamiflu are used to treat what?
Influenza
There are two common routes of antigenic variation: antigenic drift and antigenic shift. Which virus uses these processes to cause new epidemics?
Influenza
Deep/systemic myosis
· Coccidioides immitis- in new world e.g. inhaling desert air · Histoplasma capsulatum - in cavers a lot · Opportunistic pathogens - opportunistically infect us - they are resistant to alveolar macrophages so they sort of accidentally infect us - they don't intend to infect us Patients tend to have some sort of immunosuppression. More people die from top 10 fungal diseases than Malaria or TB Fungal infections are a big problem in TRANSPLANT settings (aspergillosis) There is a very high mortality for systemic mycoses Fungal infections are on the rise with a very high mortality rate
describe each of the types of virus vaccines (brainscape)
o live attenuated - live virus has its virulence reduced and is then adminstered to patient o inactivated - the virus is taken and its genome is destroyed so that it still stimulated a response but can no longer be infectious - given with an adjuvant (They potentiate the immune response (humoral, cellular or both). Two groups - delivery system and immune potentiators). o purified subunits - the viral genome is taken and treated with proteases which chops it into small pieces. These subunits have antigens that can trigger an immune response. o cloning - viral genome is cloned in a bacterium and the copies of the genome are either: injected into people, put in to virus-like particles, a new virus is made with a little segment of virulent material from the original virus
Summary of pathogens entering different parts of body and diseases they cause
o mouth (S. pyogenes/pharyngitis) o skin (S. aureus/wound infections) o faeco-oral (Vibrio cholerae/cholera), o respiratory tract (S.pneumonia/pneumonia) o vagina-neonate (Group B Streptococcus/neonatal meningitis)
Give examples of some mycobacteria and the diseases they cause (brainscape)
o mycobacterium tuberculosis (TB) o mycobacterium leprae (leprosy)
What three types of illness do fungi cause? (brainscape)
o mycoses o mycotoxicoses o allergies
What are superficial mycoses? give some examples. (brainscape)
o mycoses that affect the skin and hair. no invasion of tissue hence no cellular response o e.g. tinea nigra, white piedra, black piedra, dandruff
Give some examples of bacteria and viruses that use the upper respiratory tract as a portal of entry (brainscape)
o neisseria meningitidis o influenza o parainfluenza o streptococcus pneumonia o streptococcus pyogenes o staphylococcus aureus
Describe neuraminidase inhibition in influenza. (brainscape)
o neuraminidase is usually produced by influenza to destroy sialic acid on the surface of the host cell thus meaning that the virus doesn't bind to the same cell again. o by blocking neuraminidase you can limit the spread of the virus to other cells
Define infectious dose (brainscape)
o number of bacteria needed to cause infection
What are the three types of mucocutaneous candidiasis that occurs in people with HIV? (brainscape)
o oesophageal o oropharyngeal o vulvovaginal
How do macroloides work? Give an example (brainscape)
o only gram-positive infections o targets the 50s ribosomal subunit and prevents the peptidyl transfer steo o e.g. erythromycin
Give two examples of neuraminidse inhibitors (brainscape)
o oseltamivir (tamiflu) o zanamivir (relenza)
How do beta-lactams work? Give some examples (brainscape)
o penicillin and methicillin o they have a beta-lactam ring that is a similar shape to a precursor of peptidoglycan in bacterial cell wall and hence interferes with the synthesis of the cell wall
How can virus plaques be used to quantify the amount of virus in a sample? (brainscape)
o plaque assay - the virus undergoes serial 10-fold dilutions and is then spread on a monolayer of susceptible cells o a plaque will appear where an individual virus has killed some cells o the number of plaques can be counted and scaled up to quantify the amount of virus in a sample
State what type of vaccine has been produced for each of these diseases (brainscape)
o polio - inactivated and live attenuated o smallpox - live attenuated o rubella - live attenuated o hep B - cloned subunit o influenza - inactivated and live attenuated o HPV - cloned subunit
What is the difference between positive sense RNA and negative sense RNA? (brainscape)
o positive can be translated straight away o negative needs to be transcribed into a positive sense copy
What are cutaneous mycoses? give some examples (brainscape)
o produce keratinases - causes inflammation. o trychopyton and microsporum are main examples o other examples: tinea capitis, tinea corporis, tinea pedis
Give 2 examples of gram-negative opportunistic bacteria (brainscape)
o pseudomonas aeruginosa o acinetobacter baumanii
Name some major antibiotic resistance gram-negative bacterial pathogens (brainscape)
o pseudomonas aeruginosa o klebsiella o salmonella o acinetobacter baumanii o neisseria gonorrheae
What types of vaccines exist for influenza? (brainscape)
o purified subunit o live attenuated (nasal spray) o inactivated
How do quinolones work? Give some examples (brainscape)
o quinolones inhibit the function of DNA gyrase (gram-negative) and topoisomerase (gram-positive) hence hampering the unravelling of DNA during replication
State whether each of the following are gram-positive or gram-negative (brainscape)
o shigella - negative o streptococcus - positive o listeria - positive o clostridium - positive o vibrio cholerae - negative o salmonella - negative o staphylococcus - positive o neisseria - negative o haemophilus influenzae - negative
What are some common features among RNA viruses and retroviruses? (brainscape)
o small genome limited by instability of RNA o no proofreading capacity - high mutation rate
Name some major antibiotic resistance gram-positive bacterial pathogens (brainscape)
o staphylococcus aureus (MRSA) o streptococcus pneumoniae o clostridium dificile o enterococcus spp. o mycobacterium tuberculosis
Give examples of some gram-positive pathogenic bacteria and the diseases they cause (brainscape)
o staphylococcus aureus - skin infections, endocarditis, bacteraemia, pneumonia o streptococcus pneumoniae - pneumonia, meningitis, otitis media o o streptococcus pyogenes - tonsilitis, necrotisisng fasciitis, scarlet fever
Give some examples of bacteria that can infect via broken skin (brainscape)
o staphylococcys aureus o streptococcus pyogenes
Give 4 examples of gram-positive bacteria (brainscape)
o streptococcus - positive o listeria - positive o clostridium - positive o staphylococcus - positive
What are the four types of mycosis?(brainscape)
o superficial o cutaneous o subcutaneous o systemic
What three factors are tropisms based on? (brainscape)
o susceptibility - receptor interactions o permissivity - ability to use the host cell to complete replication o accessibility - ability of virus to reach the tissue
Give some examples of extrinsic bacteria that can infect via the urogenital tract (brainscape)
o syphilis o chlamydia o neisseria gonorrheae
What are the three types of candida infection that can occur in the immunocompromised? (brainscape)
o systemic o mucosal o superficial
How are live attenuated vaccines made? (brainscape)
o the virus is passed through the wrong type/wrong species of cells. this makes the virus evolve and change its virulence e.g. if a virus is passed through monkey cells then it will become a monkey virus and it will no longer be as virulent to humans
How do antifungals that target the cell work? give an example (brainscape)
o they inhibit the assembly of fungal cell wall components such as glucans and chitin e.g. caspofungin (an echinocandin)
Define mycotoxicosis (brainscape)
o toxic reaction following inhalation of ingestion of toxins produced by fungi
gangliosides
o type of sphingolipids (sphingosine or sphingoid backbone) GLYCOLIPIDS o glycosidic linages - no phosphate group; polar head group is oligosaccharides at the terminus this contains a *negative charge* o play a role in cell interaction, recognition, and signal transduction
Give some examples of bacteria and virsuses that infect via the GIT (brainscape)
o vibrio cholera o salmonella o listeria o shigella o E. Coli o Campylobacter jejuni
What are the five techniques used to diagnose a viral infection? (brainscape)
o viral particles (electron microscopy, haemagglutination assay) o antibodies to the viruses o cytopathic effect o genome (PCR) o antigen (indirect fluorescence antibody, ELISA)
What two main factors affect pathogenicity? (brainscape)
o virulence o infectivity
Describe the inhibition of the M2 channel in influenza (brainscape)
o when endocytosed, influenza enters the endosome o the low pH of the endosome opens the M2 channel allowing protons to move into the endosome and break the bonds holding together the protein capsid of influenza o this allows influenza to uncoat and release its contents into the cytoplasm of the cell o adamantes (rimantadine and amanatadine) can fit in the M2 channel and prevent protons from moving in, thus meaning that the influenza is locked in its protein shell
Give 7 examples of HIV drugs (brainscape)
o zidovudine o stavudine o efavirenz o viramune o atazanavir o reltagravir o maraviroc
Host range definition
organisms that can be infected by a disease. Some hosts are dead-end hosts, meaning that they can't pass the disease on.
Timeline of antifungal drug development
polyenes (cell membrane function) --> azoles (cell membrane ergosterol biosynthesis) --> echinocandins (cell wall synthesis)
Define tropism (brainscape)
predilection of virus to infect some tissues and not others
What is prophylaxis?
preventing disease before the aetiologic agent is acquired, by vaccination or giving drug before infection
Define prophylaxis (brainscape)
preventing the disease before the aetiological agent is acquired, by vaccination or giving the drug before infection
How does tetracycline work? (brainscape)
prevents amino-acyl tRNAs from binding to the mRNA/ribosome complex
HIV drugs mechanisms and side effects
protease inhibitors, integrase inhibitors, nucleoside analogues, reverse transcriptase inhibits, fusion and entry inhibitors • NRTI (Nucleoside Reverse Transcriptase Inhibitor) o Zidovudine o Stavudine • NNRTI (Non-Nucleoside Reverse Transcriptase Inhibitor) Binds enzymes but not at the active site o Efavirenz o Viramune · Integrase Inhibitor o Raltegravir · Entry Inhibitors o Maraviroc - blocks gp120 - CXCR4 interaction o Enfuvirtide - binds gp41 - prevents membrane fusion · Protease Inhibitors Atazanavir (taken with Ritonavir) Overall: Some anti-retroviral drugs do have side effects so not a perfect solution but prevents people dying.
spike proteins
protruding structures of virus that are recognized by antibodies
what is RT-PCR
reverse transcriptase PCR: • Synthesize cDNA from RNA using "reverse transcriptase" • Use cDNA as "template" for PCR (see cDNA synthesis)
shape of vibrio cholerae
rod-shaped GRAM NEGATIVE
Give an example of viral load affecting the outcome of an infection (brainscape)
second child in a household infected with chicken px will have a worse infection because they'll get a large dose from the first child
What are mycotoxins?
secondary metabolites of moulds that exert toxic effects on animals and humans
What is the genome of fungi like?
several chromosomes and have a massive complex genome
SARS
severe acute respiratory syndrome · Chain of human transmission began November 2002, Guandong province China. · Late February 2003 international spread. · July 2003 last chain of human transmission broken. · The virus was containable so within a year of its emergence from bats --> cats --> people it had been contained by isolation · It was transmitted via respiratory droplets; concentrated sources on lift buttons and in bathroom drains · Amplified by: Nebulizers in hospitals brought up virus from deep down in the lungs · Elderly people showed high mortality · Children experienced mild disease, patients over 60 years have 55% mortality. · Patients did not become contagious until quite late into the infection once they had become symptomatic (10 days post infection)- you can see who has SARS and isolate them · Destruction of lung tissue from overexuberant immune response. · Emergency response coordinated, international, rapid. · Flu is the opposite - they become symptomatic after they are contagious so you can't contain it · SARS is a CORONAVIRUS - positive-sense RNA genome · Enveloped spike protein · Receptor in human: ACE-2 · A virus almost identical isolated from masked palm civets and raccoon dogs in wet markets. Chinese horseshoe bats harbour SARS-like coronaviruses that can use bat and human ACE2 as receptors · There are lots of other coronaviruses in bats which can emerge and become pandemic · The spike protein is HIGHLY PLASTIC and can adapt to bind to different receptors · Comes from civets, raccoons and bats
Historically, variolation was used as a method of immunization against this disease.
smallpox
bacteriostatic
stops bacterial growth
What was the last pandemic we had?
swine flu pandemic (H1N1) in 2009: looked like 1918 spanish virus, and hence the older people were actually least affected due to had encountered it before
What are two other ways of detecting the presence of virus in a sample apart from plaque assay and cytopathic effect (brainscape)
syncytia formation and immunostaining
Where are many pathogenicity islands found near?
tRNA genes
Pathogenicity
the ability of an organism to cause disease
Infectivity
the ability of an organism to get into host system and establish themselves to initiate infection (transmission, colonisation, replication)
Infectious dose
the average number of organisms required to initiate infection - this directly influences infectivity
What is the breakpoint? (brainscape)
the concentration of antibiotics than can be achieved in a clinical setting. if the bacteria can divide at a conc at or higher than the breakpoint, it is deemed resistant
MIC (minimum inhibitory concentration)
the lowest concentration of antibiotic required to inhibit growth
What is variolation?
the obsolete process of inoculating a susceptible person with material taken from a vesicle of a person who has smallpox
How are recombinant attenuated virus vaccines made? (brainscape)
the virulence gene is either mutated or deleted
When do flu patients become symptomatic?
they become symptomatic after they are contagious so you can't contain it
What is therapy?
treating the disease after the host has been infected
Salmonella typhi causes?
typhoid
What does salmonella typhi cause
typhoid fever
Portals of entry in upper-respiratory tract
usually extrinsically-acquired from respiratory tract droplets or airborne. Hand transmission can act as an intermediate. e.g. Viruses: influenza, measles, varicella (chicken pox) e.g. Bacteria: streptococcus spp (dental bacteria)/pneumoniae/pyogenes, neisseria menigitidis, staphylococcus aureus (MRSA and MSSA)
How to propagate viruses in the lab?
· We can passage viruses in the lab by providing permissive cells (allow infection), often continuous lines of transformed cell cultures · Viruses may accumulate mutations that adapt them to the new host (e.g. losing some accessory genes) · This can lead to ATTENUATION (become good at infecting cells in dish but now not as good as infecting cells in body) and was the basis of generation of vaccines in the past e.g. louis pasteur with rabies, polio virus vaccine (generated by growing virus in wrong cells - so good at those viruses but not in human cells) · For some viruses there are no permissive cell lines (e.g. norovirus - winter vomiting disease' and hepatitis C) so they are difficult to work with and difficult to study in lab.
Portal of entry - broken skin
· You need a break in the skin to acquire an infection through the skin · Surgery/wound · Skin diseases (Varicella - chicken pox causes areas of broken skin, Eczema, Pressure Sores) · Insect bites - because people scratch and break skin · I.V. Cannulae · IDU - Injecting Drug Use · Unexpected Causes: o Human bites Infections Targeting Broken Skin: o Staphylococcus aureus - good at living on the skin - there is a lot in the nose and mouth o Streptococcus pyogenes - very good at getting into skin lesions · Most people have Staph. aureus in our nose and throat · Group A strep = Strep. pyogenes o If you've recently had antibiotics, you may get MRSA or pseudomonas o Surgery/bowel flora exposure (staph aureus, strep pyogenes, and gram negative bacteria from gut) Unusual pathogens - e.g. from medicinal leeches, soil, dog bites
How do new viruses emerge?
· Zoonosis - from animals and crosses into humans e.g. ebola from fruit bats/monkeys · Genetic Variation - antigenic drift · Increased Exposure - travel or world population · Increased Exposure - spread of vector e.g. increased insects --> increased arboviruses New Discoveries
Why do bacteria occupy huge component of biodiversity in world?
• As they have access to a huge variety of DNA through horizontal gene transmission, • The high reproductive rate and ability to mix DNA means that there is huge selection pressure among bacteria, which is responsible for the sophisticated processes that bacteria possess. So there are loads of bacterial genomes and the phage which infects them, which are all mixable and mutatable and bacteria grow with a very rapid generation time
Process of gram staining
• Bacteria are stained with a violet dye and iodine, rinsed in alcohol, and then stained with a red dye. Violet dye + iodine -> alcohol rinse -> red dye Stain indicates whether or not you have Gram-negative or Gram-positive bacteria.
BCG vaccine against TB baceria
○ Attenuated strain of Mycobacterium bovis ○ Serial passage, 231 times over 13 years to make it attenuated ○ Phenotypic changes - rough/dry to moist/smooth ○ Genotypic changes - loss of RD1 region encoding 9 proteins
Killed whole cell cholera vaccine (bacteria)
○ Killed whole cell parenteral vaccine § Poor efficacy § Short-lived protective response § Strain specific ○ Killed whole cell oral vaccine § Good efficacy in cholera endemic area (Vietnam trials) § Oral delivery § Safe § >85% show >4-fold rise in vibriocidal antibodies (<3% in controls) § Protective efficacy 66% after two doses § WHO mediated tech transfer ○ Dukoral© vaccine § Killed whole cell + CtxB (cholera enterotoxin subunit B precursor) oral vaccine § Drink formulation (3 doses at weekly intervals, booster every 2 years) May also protect against ETEC (enterotoxogenic E.Coli due to cross reactivity between toxins)
Typhoid vaccine (bacteria)
○ Vivotif - live attenuated vaccine based on strain Ty21a § Gal.E mutant of S.typhi Ty2 § Isolated by NTG mutagenesis - this was done before restriction enzymes and PCR existed § Formulated in enteric-coated capsules ○ Vi polysaccharide - VI capsule from typhoid made into vaccine § Offered to those travelling to typhoid endemic areas § Only suitable for >2 year olds § Conjugates in development - primarily in endemic areas e.g. India
How can dengue antibodies either neutralise or enhance infection?
· Dengue virus usually gets into the cell by binding to specific receptors · If you have homologous antibodies (perfect match to virus serotype) then the viral antigens will bind the antibodies TIGHTLY and stop it from entering the cell - neutralises virus · If infected by a different viral serotype - the antibodies bind loosely to the antigens but don't block it. By binding to the wrong anti-dengue antibodies, the viruses are given another way of getting into the cells. · Then the Fc portion of antibodies can bind to Fc receptors on immune cells thus carrying the virus into immune cells (e.g. macrophages) it normally wouldn't enter This leads to a cytokine storm and dengue haemorrhagic fever (blood under skin, loss of fluid, increased Hct, give IV fluid)
Portal of entry: The gastro-intestinal tract (faeco-oral route)
· Viruses o Hepatitis A o Norovirus o Hepatitis E · Bacteria o E. coli - travellers diarrhoea o Campylobacter jejuni food poisoning o Shigella (dysentery) o Vibrio cholerae o Salmonella enterica (food poisoning) o Salmonella typhi (typhoid) o Listeria o Clostridium difficile - hospital bug causing diarrhoea · Toxins o Enterotoxins o Parasites eg worms
Bad things about fungi :(
cause amphibian extinctions, cause decline in bats, forest loss (Namerican chestnut), crop failures and famines, human disease
Two groups of adjuvants
1) Delivery systems, usually a depot of antigen which is slowly released - help in transport: o Mineral salts o Surface active agents o Synthetic microparticles o Oil-water emulsions o Liposomes 2) Immune potentiators - molecules that improve/change immune response o Toxins and lipids o Nucleic acids (CpG) o Peptidoglycan o Carbs o Peptides o Cytokines and hormones
List 3 types of viruses, when classified according to the type of their genetic material (Baltimore's classification)?
Double stranded DNA Single stranded DNA Antisense strand RNA Sense strand RNA
Staphylococcus is a major.... pathogen
-skin- Produces a family of leukocydins - toxins which destroy neutrophils producing characteristic pus. can lead to small abscess or big
Fungi (differences from bacteria)
. Membrane bound nucleus . Cell wall . DNA vs RNA Main effect is that fungal infections are harder to treat
Mechanisms of antibiotic resistance
1) Altered target site • Can arise via acquisition of alternative gene or a gene that encodes a target-modifying enzyme. • Methicillin-resistant Staphylococcus aureus (MRSA) encodes an alternative PBP (PBP2a) with low affinity for beta-lactams. • Streptococcus pneumoniae resistance to erythromycin occurs via the acquisition of the erm gene, which encodes an enzyme that methylates the AB target site in the 50S ribosomal subunit, so antibiotic can no longer recognise proteins produced by bacteria • quinolones 2) Inactivation of antibiotic • Enzymatic degradation or alteration, rendering antibiotic ineffective. • Examples include beta-lactamase (bla) and chloramphenicol acetyl-transferase (cat). • ESBL and NDM-1 are examples of broad-spectrum beta-lactamases (can degrade a wide range of beta-lactams, including newest). Inactivation can be enzyme-independent - was shown that bacteria could release bits of themselves as decoys to distract antibiotics 3) Altered metabolism • Increased production of enzyme substrate can out-compete antibiotic inhibitor (e.g. increased production of PABA confers resistance to sulfonamides). Alternatively, bacteria switch to other metabolic pathways, reducing requirement for PABA. 4) Decreased drug accumulation • Reduced penetration of AB into bacterial cell (permeability) and/or increased efflux of AB out of the cell - drug does not reach concentration required to be effective. Gram negatives are difficult to target due to two cell membranes and LPS on outer layer surface as well
Summary of antimicrobials
1) Amniglycosides (type of antibiotic) • E.g. Gentamicin, streptomycin. • Bactericidal. • Target protein synthesis (30S ribosomal subunit), RNA proofreading and cause damage to cell membrane. In addition to preventing it. It corrupts protein synthesis --> misfolding --> break down bacterial membrane • Toxicity has limited use, but resistance to other antibiotics has led to increasing use. 2) Macrolides • E.g. Erythromycin, azithromycin. • Gram-positive and some Gram-negative infections. • Targets 50S ribosomal subunit preventing amino-acyl transfer and thus truncation of polypeptides. 3) Quinolones • Synthetic, broad spectrum, bactericidal. Was hoped bacteria wouldn't form resistance to completely synthesised AB but it didn't work • Act on DNA replication: Target DNA gyrase in Gm-ve and topoisomerase IV in Gm+ve. So DNA can't unwind to replicate. 4) Beta-lactams: Inhibit cell wall (peptidoglycan) synthesis Penicillin, methicillin 5) Tetracycline: Inhibits protein synthesis Bacteriostatic 6) Chloramphenicol: Inhibit protein synthesis Bacteriostatic Higher toxicity 7) Sulphonamides: Bacteriostatic Combination therapy Sulpha-methoxazole Specific antibiotic types 1) Rifampicin antibiotic • Bactericidal. • Targets RpoB subunit of RNA polymerase. • Spontaneous resistance is frequent. • Makes secretions go orange/red - affects compliance. • E.g. for TB 2) Vancomycin antibiotic • Bactericidal. NOT beta lactam • Targets Lipid II component of cell wall biosynthesis, as well as wall crosslinking via D-ala residues • Toxicity has limited use, but resistance to other antibiotics has led to increasing use e.g. against MRSA and VRE 3) Linezolid antibiotic • Bacteriostatic. • Inhibits the initiation of protein synthesis by binding to the 50S rRNA subunit. • Only has gram-positive spectrum of activity. 4) Daptomycin antibiotic • Bactericidal. • Targets bacterial cell membrane. • Gram-positive spectrum of activity. • Toxicity limits dose - given IV and patient needs support . 5) Chloramphenicol - inhibits peptidyl transferase in dna translation - disrupt protein synthesis 6) tetracycline - disrupt protein synthesis and prevent AA-tRNA complex forming
What are the three main elements in vaccines?
1) Antigen o To stimulate immune response to the target disease ·2) Adjuvant o To enhance and modulate the immune response 3) Excipients o Buffer, salts, saccharides and proteins to maintain the pH, osmolarity and stability of the vaccine Preservative e.g. phenoxyethanol, thiomersal etc. generally only done in multidose vials. Generally single dose vials without preservatives are usually used
Bacterial pathogens have to be able to...
1) Attach and Colonize (surface structures such as pili and fimbriae allow them to cling to surfaces) 2) Persist (avoid, subvert, or circumvent host defenses in or outside cells) 3) Replicate (acquire nutrients such as iron, energy sources etc) 4) Disseminate within cells, tissues between organs and hosts (bacterial and host cell motility, through aerosols, faeces etc) - 5) Cause disease (produce toxins that kill host cells, induce diarrhea, dysregulate immune responses) 6) transmission to new host
List 3 other bacterial vaccines
1) BCG (against TB) ○ Attenuated strain of Mycobacterium bovis ○ Serial passage, 231 times over 13 years to make it attenuated ○ Phenotypic changes - rough/dry to moist/smooth ○ Genotypic changes - loss of RD1 region encoding 9 proteins 2) Typhoid vaccine ○ Vivotif - live attenuated vaccine based on strain Ty21a § Gal.E mutant of S.typhi Ty2 § Isolated by NTG mutagenesis - this was done before restriction enzymes and PCR existed § Formulated in enteric-coated capsules ○ Vi polysaccharide - VI capsule from typhoid made into vaccine § Offered to those travelling to typhoid endemic areas § Only suitable for >2 year olds § Conjugates in development - primarily in endemic areas e.g. India 3) Killed whole cholera vaccine ○ Killed whole cell parenteral vaccine § Poor efficacy § Short-lived protective response § Strain specific ○ Killed whole cell oral vaccine § Good efficacy in cholera endemic area (Vietnam trials) § Oral delivery § Safe § >85% show >4-fold rise in vibriocidal antibodies (<3% in controls) § Protective efficacy 66% after two doses § WHO mediated tech transfer ○ Dukoral© vaccine § Killed whole cell + CtxB (cholera enterotoxin subunit B precursor) oral vaccine § Drink formulation (3 doses at weekly intervals, booster every 2 years) May also protect against ETEC (enterotoxogenic E.Coli due to cross reactivity between toxins)
How were accessory genes acquired?
1) Bacteria divide by binary fission: (vertical transmission of DNA) 2) Also undergo horizontal gene transfer (HGT): primary drivers of bacterial evolution 3 basic mechanisms: Transformation, conjugation, transduction
Summary of the 5 mechanisms of resistance of antibiotic resistance for certain antibiotics.
1) Blocking uptake/ decreased influx e.g. sulfonamide resistance 2) Increased efflux/ membrane pumps e.g. tetracycline resistance 3) Target site modification e.g. in peniciilin resistance: alternative penicillin-binding proteins as found in MRSA 4) Drug inactivation e.g. in penicillin resistance: beta-lactamase 5) Target amplification e.g. in metronidazole resistance
Viral diagnosis
1) Detecting Viral Genome o PCR (very common) o Have primers which are complementary to a specific part of the viral genome which identifies the virus o Use sets of primers and the one which shows up positive will identify the virus 2) Detecting Viral Antigen o Indirect Fluorescence Antibody (IFA) - using antibodies for viral antigen o ELISA - using antibodies for antigen 3) Detecting Viral Particles o Electron Microscopy - difficult, expensive, not very sensitive - rare only if new virus and don't know what looking for o Haemagglutination Assay - some viruses bind to proteins on RBC surface and cross-link RBCs together - visual assay, doesn't tell which virus it is though 4) Detecting virus cytopathic effect in cultured cells o Virus isolation, e.g. for Zika - look at neurones, influenza - cells derived from respiratory tract 5) Detecting antibodies to virus o Serology (in blood), needs to be done quickly though
Two ways in which vaccinates work?
1) Direct protection - immunisation prevents them from getting infection 2) Herd immunity - prevent transmission amongst population (indirect protection)
Antiviral treatment options
1) Interferons (body's own natural antiviral response) to induce the hosts natural antiviral response 2) Drugs with specific antiviral activity 3) Treatment that alleviate symptoms but do not inhibit virus replication. And that's important because many symptoms of virus replication are bodys own immune system turning on itself. May need to prevent/tone it down to prevent harming themselves in effort to get rid of virus
What are the different types of viral vaccines?
1) Live natural virus vaccine 2) Inactivated virus vaccine 3) Purified subunit vaccine 4) Cloning --> live virus vector vaccine or dna vaccine or protein (virus-like/subunit vaccine)
What are the 4 classes of antifungal drugs?
1) Membrane function - polyenes e.g. Nystatin and amphotericin B work by stimulating ergosterol, also they act on the fungal cytochrome P450 enzymes 2) Nucleic acid synthesis - 5-Flucytosine. Pyrimidine analogues Used in combination therapy (not too effective alone) 3) Cell wall synthesis - echinocandins. (𝛽-glucan synthesis) 4)Membrane ergosterol biosynthesis - azoles (e.g. Ketoconazole and itraconazole), terbinafine, fenpropimorph (FAT)
Diagnosis of fungal infections
1) Sample acquisition 2) Microscopy 3) Culture 4) Non-culture methods - e.g. antibody and antigen-based assays
Name 3 ways viruses can enter the body
1) Through the epithelial layers; respiratory tract, GI tract, genital tract 2) Directly into the blood through a bite or needle 3) Through the skin, often following abrasion
Consequences of urogenital tract infection
1) Urinary tract infection (cystitis (limited to bladder), pyelonephritis (spreads up to kidney by ureters)) --> spread to bloodstream e.g. E.Coli bacteraemia Genital tract infection e.g. gonococcal urethritis--> pregnancy-related infection (travel via fallopian tube) e.g. neonatal group B strep infection --> spread to bloodstream
Stages in life cycle of virus which could potentially allow therapeutic intervention
1) Viral binding and entry 2) Viral replication: Transcription *Reverse transcription* *Integration* Translation Replication 3)Viral assembly 4) Viral exit
Sepsis pathophysiology
1) WBC recruitment because of pathogens 2) Release of NO for increased permeability/vasodilation (Permeability - to allow for the cells to migrate into the tissue Vasodilation - to reduce blood flow, also to allow for the cells to migrate) 3) Reduced BF contributes to shock 4) Increased permeability leads to fluid loss to interstitial space and therefore less perfusion and hypotension 5) Chemical release from WBC leads to endothelial damage -> permeability increases 6) Increases in CO to compensate for low TPR
Picture of targets for antifungal therapy
1) cell membrane 2) dna synthesis 3) cell wall
Investigating viruses in the lab
1) cytopathic effect 2) plaque assay 3) syncytia 4) immunostaining
Give examples of bacteria using the three methods by which bacteria survive in the host cell (brainscape)
1) escape - shigella, listeria 2) preventing fusion with lysosome - salmonella, mycobacteria, chlamydia 3) surviving in phagolysosome - coxiella
What are the three methods by which bacteria survive in the host cell, give examples (brainscape)
1) escape - shigella, listeria 2) preventing fusion with lysosome - salmonella, mycobacteria, chlamydia 3) surviving in phagolysosome - coxiella
Explain each of the three mechanisms for horizontal gene transmission (brainscape)
1) transformation - the uptake of naked DNA from the environment 2) transduction - bacteriophages infect a bacterium and take up some of bacterial DNA. The bacteriophage then carries the bacterial DNA to another bacterium 3) conjugation - transfer of genetic material in the form of a plasmid via a conjugation tube
Classes of antiviral drugs
1)Interferons 2)Nucleoside analogues e.g.Acyclovir 3)Fusion/entry inhibitors 4)Non-nucleoside reverse transcriptase inhibitors (NNRTIs) 5)Integrase inhibitors 6)Protease inhibitors (viral exit) 7)Many others
Patterns of viral infection
1. Acute - followed by viral clearance, sometimes can get 'accidental' tissue infected with permanent damage despite viral clearance e.g. rhinovirus 2. Persistent - latent, slow, transforming (transform cells and lead to oncogenesis) e.g. papilloma virus in warts 3. Long incubations (last for years and then come back out again) e.g. herpes simplex virus, varicella zoster virus 4. Oncogenesis - affect the way our cells control themselves e.g. EBV --> burkitt's lymphona, HIV --> kaposhi's sarcoma; hep b/c --> hepatocellular carcinoma 5) Slow infection - Cause an asymptomatic primary infection. Have a long incubation period ranging from months to years. Follow a slow but relentless progressive course leading to death. Tend to have a genetic predisposition Often re-emerge from latency if the host becomes immuno-compromised e.g. measles virus SSPE, HIV, Human T-lymphotropic virus KNOW THE EXAMPLES IN THE DIAGRAM
New vaccines against group B meningococcal disease
Another problem w/ conjugate is that can't always be used With men group B, polysaccharides on bacteria capsule are very similar to polysaccharide antigens on our own cells - so potentially self-antigen in vaccination So use outer membrane vesicle (OMV) vaccine - OMVs are released spontaneously by meningococci in vivo and in culture - they contain all the protein antigens usually associated with the OM. OMVs can be harvested and used in vaccine - vaccines are produced by detergent extraction to reduce endotoxin content Hyper-producing mutants have been isolated Vaccines used in brazil, norway etc. with various efficacies These vaccines only offer good protection against the homologous strain - only against the same strain they're made against - good for strain causing epidemic but not good for in paediatric programme New MenB vaccine targets non-capsular antigens: So new licensed vaccine Bexsero consists of OMVs and heparin binding antigen and factor H binding and NadA to give broader protection - 4-valent. Introduced at time when disease incidence was dropping anyway, but after vaccine introduced, it decreased further showing effectiveness of around 83%. Clinical trial showed that unlikely to offer herd protection though. Incidence ratio decreased.
Macrolides
Antibiotic types • E.g. Erythromycin, azithromycin. • Gram-positive and some Gram-negative infections. Targets 50S ribosomal subunit preventing amino-acyl transfer and thus truncation of polypeptides.
Mechanisms of antibiotics
Antibiotics target many different bacterial processes and are selectively toxic . The large number of differences between mammals and bacteria result in multiple targets for antibiotic therapy - selective toxicity (much more toxic to bacteria than to us). Toxicity is dose dependent so if bacteria is resistant can't keep on increasing dose, have to switch to new antibiotic
antigenic drift vs antigenic shift
Antigenic drift is the process of subtle changes eg amino acid mutations, whereas antigenic shift involves drastic changes e.g. reassortment of genetic information.
Interferons
Antiviral proteins secreted by T cells. Hosts natural antiviral system
Zidovudine (AZT)
Antiviral: nucleoside inhibitor of HIV reverse transcriptase
How do viruses form plaques in monolayers of cells?
As a result of this death - viruses form plaques on cell monolayers (hole in the layer of cells it was growing in where virus budded out and killed cell) · Plaques - the result of an INDIVIDUAL virus infecting one cell and then infecting other cells · Each plaque is the result of one single virus that was originally put onto the cell monolayer · If you count the plaques you can measure how many viruses there were in the sample o Someone who is freshly infected will have fewer plaques than someone who has had it for a while · This is used to find out how much virus is in the patient and is done using Plaque Assay Can also stain with antibody to see viral proteins (pic A)
Name an ascomycete fungal pathogen that causes hospital-acquired fungal infections
Aspergillus fumigatus or Candida sp.
Which cell type does the Epstein Barr Virus most commonly affect?
B lymphocytes. The Epstein Barr Virus causes Infectious Mononucleosis (also known as 'glandular fever' or 'mono') by causing lymphocytosis, which means there is an abnormally larger number of B lymphocytes in the blood.
Examples of different shaped viruses
Bacteriophage = virus that infects bacteria, don't infect humans TMV infects plants Rabies virus (rhabdobirus (VSV)) Rota virus causes diarrhoea
What kind of genetic material does Herpes Simplex virus have?
Double-stranded DNA
What are bacteriophages?
Bacteriophages are bacterial viruses, which can move DNA from one bacterium to another and integrate into the chromosome
How are viruses classified?
Based on genome
What are the two phyla in dikarya?
Basidiomycota Asomycota
Which animals are an important reservoir of coronaviruses?
Bats
Nucleoside analogues
Best antiviral that we have are nucleoside analogues • Viruses have to replicate their genome at some point - hence they have to add nucleotides or nucleosides to their growing RNA chain • We know what normal bases look like • N.b. zidovudine = AZT • Nucleoside analogues look like normal bases but have a little bit missing - e.g. acyclovir looks like guanosine but is completely missing the bottom half of the ring • Work by chain termination: modified nucleoside incorporated into DNA lack of 3' -OH prevents phosphodiester bond formation
Who are the important hosts in the spread of arboviruses such as West Nile Virus?
Birds
What are sterile sites in body?
Brain, trachea, lungs, liver, heart, spleen, kidney, pancreas, bladder, gall bladder. Would NOT expect to find bacteria here
What types of cancer can Epstein-Barr Virus cause? (brainscape)
Burkitt's lymphoma, Hodgkin's lymphoma, nasopharyngeal carcinoma
How can pathogenicity islands be revealed?
By aligning pathogenic genomic DNA and a closely-relate non-pathogen.
Spread of antibiotic resistance genes
By transduction, transformation, conjugation. E.g. MRSA most likely got AB res genes by phage transduction
Co-receptors CCR5 and CXCR4 in HIV tropism
CCR5 (macrophages)and CXCR4 co-receptors (t cells): Chemokine receptors that sit on immune cells. In general viruses transmitted through mucosal routes (e.g. unsafe sex) and macrophages, which are first cells to get infected in HIV event, hence if no CCR5 but have unprotected sex will not get infected. BUT if have blood transfusion, then HIV may enter into T cells in blood directly through CXCR4 co-receptor.
What does the respiratory tract use as non-immune host defence?
CILIARY clearance, or muco-ciliary clearance, involves the action of ciliated mucous-secreting cells. In addition to ciliary clearance, the respiratory tract has MACROPHAGE-LINED ALVEOLI and HAIRS in nostrils which filter large particles.
Virus morphology
Can be non-enveloped or enveloped: 1) non-enveloped o Have a protein capsid o Tend to be more symmetrical EXAMPLES: Adenovirus, Picornavirus, Calicivirus 2) enveloped Have proteins around their genome o Have a lipid envelope around that which is derived from the host membrane (it is not encoded in the genome) o Enveloped viruses are pleomorphic - have lots of different shapes e.g. measles virus EXAMPLE of typical shape virus: Ebola virus (loopy) NOTE: Herpes virus is a combination of capsid AND envelope surrounding tegument (viral matrix), contains lots of viral proteins to take over host cell
What gives bacteria immense potential to adapt and overcome our immune system?
Can take up DNA from environment
Candida infections
Candida Infections NOTE: Candida is an example of superficial mycosis · Candida albicans is an opportunistic commensal · Pretty much all of us have it in our gastrointestinal and genitourinary tracts · When we are immunocompromised, C. albicans can colonise and invade host tissues · Superficial Candida Infections o Mouth, throat, skin, scalp, vagina, fingers, nails, bronchi, lungs or the GI tract o Usually due to impaired epithelial barrier function o Occur in all age groups o Most common in the new born and elderly o Respond readily to treatment · Mucosal Candida Infections o Symptomatic infection e.g. white tongue o Occurs in the new born and the elderly o Mucocutaneous candidiasis occurs in three forms in people with HIV: · Oropharyngeal · Oesophageal · Vulvovaginal Once this disease becomes systemic - you might see high mortality rates · Systemic Candida Infections o Not seen in normal healthy individuals o Main Risk Factors: · Chemotherapy · Gut-related surgery · Catheters o Cause about 10% of hospital-acquired blood streatm infections and 47% mortality
Transformation process
Done by: Neisseria, Streptococcus naked DNA uptake and integration into recipient's chromosome e.g. neisseria and streptococcus • Sucking up DNA from outside • They can recognise naked DNA (e.g. from the lysis of other bacteria) They have transport mechanisms which allow uptake of this DNA and incorporation into the bacterial chromosome
Fungal mycotoxicoses
DEFINITION: a toxic reaction due to the ingestion/inhalation of toxins produced by fungi o Mycotoxins are secondary metabolites of moulds that exert toxic effects on animals and humans o Symptoms: · Breathing problems · Dizziness · Severe vomiting · Diarrhoea · Dehydration · Hepatic and renal failure 6 days later o Treatment: · Gastric Lavage and charcoal - wash out the toxins · Liver transplant o AFLATOXIN, produced by Aspergillus flavus, is the most carcinogenic natural compound known · Contaminates grain e.g. mouldy corn and peanuts · If you get aflatoxin poisoning and you have liver damage from hepatitis B then you are at particular risk of cancer · Majority of cases in Africa and South-East Asia - higher rates of liver cancer possibly due to greater exposure to aflatoxin o E.g. Psilocybin (psilocybe semilanceata) · Symptoms: · visual distortions of colour, depth and form, progressing to visual hallucinations. · The effects are similar to the experience following consumption of LSD, although milder. · Therapy: · Time! · Can potentially be used to treat depression by rebooting brain
Paediatric combination vaccines
DTaP-Hib-IPV: diphtheria, tetanus and acellular pertussis (whooping cough) and inactivated polio vaccine Tetanus and Diptheria Toxoids o Chemical inactivation of bacterial exotoxin · Simple to produce · Relatively pure - what's secreted from organism is purified and reproduced in fermenter · Safe · High protective efficacy o Very immunogenic o Appropriate immune response i.e. toxin neutralising antibodies block activity Bordetella pertussis - whooping cough o Caused by bordetella pertussis infected ciliated epithelium and releasing toxin - more complex to target due to multiple virulence determinants o Condition identified by characteristic convulsive cough o Recovery coincides with antibody production o There was also a whole cell pertussis vaccine which was effective (efficacy rate >90%). But was associated with a number of adverse reactions (anaphylaxis, prolonged crying, febrile seizures, acute encephalopathy (not shown to due to mutations within a sodium channel gene). Development of acellular (component) vaccine driven by acceptance of whole cell vaccine o B.pertussis components associated with virulence: ○ Attachment to ciliated epithelium: § Filamentous haemagglutinin (FHA) § Fimbriae § Pertactin ○ Adherence and complement resistance § BrkA ○ Toxins § Pertussis toxin (PT) § Adneylate cyclase § Tracheal cytotoxin § Heat labile toxin § Endotoxin o Acellular vaccines ○ Multicomponent (bi-,tri- and pentavalent formulations) ○ Pertussis toxin, FHA, pertactin, fimbrial antigens 2 and 3 ○ Safe and efficacious (75-90%) - safer but bit lower efficacy than whole cell vaccine ○ Acellular vaccine introduced few years ago, but disease seemed to increase - there were 10 deaths in newborns in 2012. some suggestions were to give additional booster doses, improved vaccine needed, offer vaccination to pregnant women (28-38 weeks) so there's passive immunity to baby. ○ Rationale for this comes from acellular pertussis (aP) study dhoeing it didn't provide adequate herd immunity. · Haemophilus influenzae type b o H influenza type b causes meningitis and septiciaemia in the young o Bacterial surface covered by a polyribosyl-ribitol phosphate capsule o Polysacchardes are poorly immunogenic - bad antigens and don't provoke good immune responses o The Hib component is a conjugate vaccine - vaccine dramatically reduced when introduced in 1990s, then resurgence in 2000s but then booster introduced which dampened it again · Summary: o Examples include Pediacel (sanofi) and infanrix (GSK) o Antigens: ○ Tetanus toxoid ○ Diptheria toxoid ○ Pertussis components: § Pertussis toxid § Filamentous haemagglutinin § Pertactin § Fimbriae types 2 and 3 ○ Hib PRP-conjugate ○ Inactivated polio virus types 1,2 and 3 o Adjuvant: aluminium phosphate
UK Child immunisation programme
DTaP-IB= diphtheria, tetanus and pertussis (whooping cough) and inactivated polio vaccine
What is the higher fungi kingdom called?
Dikarya. which consists of two phyla - · Basidiomycota Ascomycota
What are distinguishing features of virulence genes found on pathogenicity islands?
Direct repeats and insertion sequences
Pandemic
Disease that occurs over a wide geographic area and affects a very high proportion of the population. Global.
Conjugative transfer
Done by many gram positive and negative self-transmissible plasmid process. You get a physical bridge between bacteria, which allows the transfer of a plasmid between the bacteria.
Influenza tropism
Influenza is enveloped in haemagglutinin (HA) - tropism is not determined by receptor use. Envelope virus with spike proteins that will bind to what it wants to on cell surface · HA helps influenza latch onto the surface of cells · It does this by HA binding sialic acid (sugar NOT Protein) - terminal sugar on all glycoproteins in all cells · SIALIC ACID IS UBIQUITOUS - it is everywhere · In theory, influenza can enter any cell of your body · Influenza only tends to affect the respiratory route - partly due to accessibility · Once bound onto sialic acid, it enters the cell via the endosome · LOW endosomal pH allows a massive conformational change - this is essential for the virus to fuse with the endosome membrane and uncoat Fusion with the membrane and uncoating can only happen if the HA protein of the virus has been snipped in two at a particular point (tip of red arrow). Protease that cleaves HA virus is only found in certain tissues of body · This requirement of cutting the protein into two pieces is what determines the tropism of the virus · It is only in the fluid that lines our lungs that the right proteases are present. · Viruses mill around until they come into contact with the appropriate proteases which can chop the HA in two and activate the virus. IMPORTANT: Influenza tropism is determined by availability of host proteases (which is in lungs) NOTE: HAT and TMPSSR2 proteases are found in the respiratory tract secretions to chop up foreign bodies; but also is used by viruses to get activated. So, flu virus can infect any cell, but in some places e.g. liver where there are no proteases secreted, there the virus cannot get activated, hence although virus comes in the virus that comes out is inactive as no HA cleavage
How do sulphonamides work? (brainscape)
Inhibits the folate pathway
Active immunisation
Intentionally expose someone to a form of the antigen that does not produce disease Vaccines = active immunisation; switching on immune system and asking b cell and t cells to work
What type of pathogen is salmonella?
Intracellular
Why do fungi grow on nails?
Keratin in toe nail is food source for fungi
Adjuvants
Key element of effective vaccines. They potentiate the immune response (humoral, cellular or both) Two groups - delivery system and immune potentiators
What technique is used to prove that a virus is causing a particular disease? (brainscape)
Koch's postulates: 1. The microorganism must be found in large numbers in all diseased animals, but not in healthy ones. 2. The organism must be isolated from a diseased animal and grown outside the body in a pure culture. 3. When the isolated microorganism is injected onto other healthy animals, it must produce the same disease. 4. The suspected microorganism must be recovered from the experimental hosts, isolated, compared to the first microorganism, and found to be identical.
Why do hospitals provide pressure for ab resistance?
Large numbers of infected people receiving high doses of antibiotics - strong selective pressure for emergence/maintenance of AB resistance
What feature is found only on gram-negative cell walls? (brainscape)
Lipopolysaccharide
Acyclovir specificity
Looks like guanosine • Highly specific - due to given as prodrug (un-phosphorylated form), nucleotides need to be triphosphorylated and needs viral thymidine kinase to do this and then viral dna polymerase to integrate into virus replication • Only activated inside virus infected cells • Higher affinity for viral DNA polymerase than for host cell polymerase • Resistance is rare but maps to thymidine kinase - if thymidine stops working then prodrug won't be activated and virus still grows Overall: Acyclovir and related nucleotide and nucleoside analogues are in effect "biochemical confidence tricks" whereby the viral thymidine kinase has a high affinity for acyclovir (unlike cellular thymidine kinase) and converts it to acycloGMP, and hence (via cellular kinases) generates large amounts of acycloGTP in infected cells. This in turn is an excellent substrate for the viral DNA polymerase and so is incorporated into the growing DNA strand of viral DNA. Unfortunately for the virus acycloGTP blocks further polymerisation of DNA because no 3' OH group is available for the production of the next phosphate bond. Thus, viral DNA replication is blocked and hence generation of new virus prevented. On the other hand acycloGTP is a poor substrate for the host cell DNA polymerase and so is less often incorporated into the cell's replicating DNA. Thus, the cell is spared from the most potent effects of the drug.
Two examples of coronaviruses
MERS and SARS (both originating from bats)
What could be the next pandemic?
MERS: Limited transmission Diverse clinical signs No vaccine, no antiviral H7N9 Limited transmission No vaccine but technology to make one is known Antivirals but resistance tolerated Pandemic could also occur due to humans creating pandemic virus. E.g. rabbit virus (myxoma virus) made to control rabbits, then rabbits evolved and rabbits came back again. Also genetic manipulation of virus created a transmissable H151 flu virus.
Changing landscape of antibiotic resistance
MRSA incidence decreased slightly in western europe from 2010-2013 due to infection control measures. But for klibella pneumoniae and e.coli has increased. Today we are much more worried about gram negative organisms that contain beta lactamases that degrade beta-lactam Generally those countries who use more antibiotics get more antibiotic resistant drugs
How did MRSA most likely get Ab res genes?
MRSA most likely got AB res genes by phage transduction
How is ab resistance measured?
Measurements made in vitro may not reflect in vivo. Typically measured in lab (takes about a day). Do agar, PCR for resistance genes and broth micro-dilution
Men C vaccine
Men C season is winter. Introduction was phased - protect people most at risk first (late teens and infants). Protection against disease correlates with serum bactericidal antibody. As more vaccine produced introduced to other age groups. Men C declines rapidly since vaccination. In unimmunised pop (>18), disease also went down - herd immunity. Efficacy = 93.5%. Herd effect = 66.8%. But as mainly young people getting vaccinated, there's need for booster dose; introduced 4-valent vaccine (against the 4 serotypes) to address increase in disease alter in later generations.
Active retroviral therapy for HIV
Need to understand replication cycle of virus. At each stage, a drug can target. The proteins, genes and enzymes used - use crystal structures. Drugs that block virus entry, early stages of infection, integration of viral DNA into host DNA, processing proteins and budding out of cell. Good thing is these stages are different enzymes which HIV uses is diff to humans e.g. reverse transcriptase is key target.
Bacterium for which there is currently no vaccine against some serogroups
Neisseria meningitidis
Example of extracellular pathogens
Neisseria, Staphylococcus, Streptococcus, Yersinia
Neuraminidase enzyme structure
Neuraminidase = One of the spike proteins on the outside of the influenza virus and the crystal structure: 4 subunit enzyme - 4 identical copies- we know how substrate (sialic acid) binds into enzyme and how it works. So drugs developed which looked like sialic acid substrate but were chemically different - they entered into pocket of enzyme and never came out again. Orange box in diagram below depicts chemical pocket in enzyme. Similarity of the two drugs to sialic acid; relenza has positive guanidine which sticks into negatively charged pocket. tamiflu sticks nicely into hydrophobic pocket because hydrophobic amino acids on it.
Neuraminidase
Neuraminidase is produced by influenza virus • Virus enters the cell by binding to sialic acid • During infection and replication, the host cell gradually dies • New viruses leave the cell and want to move onto new healthier cells • So, virus produces neuraminidase which moves to the cell surface and destroys sialic acid so the original cell is less likely to get infected by the daughter viruses • IF WE INHIBIT NEURAMINIDASE - the enzymes will latch back down to the old cell and will not spread so the infection will be contained within the patient • The structure of neuraminidase is well known allowing the design of a substrate analogue which avidly sits in the neuraminidase binding pocket and acts as an inhibitor • Neuraminidase Inhibitors: o Zanamivir (Relenza) o Oseltamivir (Tamiflu)
Is Ebola an arbovirus?
No
What are the best antiviral drugs?
Nucleoside analogues
What kind of bacteria found in small intestine?
ONLY GRAM NEGATIVE BACTERIA
Where did MERS come from?
Originates in bats and is endemic in camels
What makes it easier in Gram negative bacteria to remove the iodine?
Outer cell membrane
What do modern adjuvants relate to?
PAMPs - because they stimulate innate immune response. At time of giving antigen, if stimulate innate immune response. Through stimulating toll-like receptors on cells - get different types t helper responses and steer immune responses by cytokines produced by t helper cells - tailor immune response to needs
Man's intervention as the cause of emerging viruses
Pandemic could also occur due to humans creating pandemic virus. E.g. rabbit virus (myxoma virus) made to control rabbits, then rabbits evolved and rabbits came back again. Also genetic manipulation of virus created a transmissable H151 flu virus.
What diagnostic test would you use to detect for coronavirus?
RT-PCR. is the largest RNA genome in viruses
What can increased understanding of viral components allow?
Rational drug design
New anti-viral drug baloxavir
Rationally designed - 3d structure of polymerase known and drug binds to it to stop replication Baloxavir decreased virus shedding- may interrupt transmission. Works a bit better than oseltamivir in terms of time period to alleviate symptoms and reducing viral load. Did double-blind randomised placebo RCT of drug. Had to see which had flu. Had to show superiority to get drug onto market as well and outdo alternatives.
What is the tropism of kaposis sarcoma herpes virus (KSHV) dependent on?
Receptor use
How can two or more viruses give rise to a new virus with new properties?
Recombination of two or more viruses can give rise to a new virus with new properties · There is pandemic potential of influenza virus due to reassortment · 2009 swine flu pandemic has an antigenic subtype of H1N1 · H1N1 is the same as a virus which had been circulating in humans · This can happen because of ANTIGENIC DRIFT · As a virus circulates in humans it changes a little bit at a time - after 10 years the virus will be completely different to how it was at the start · You do not get antigenic drift in pigs because pigs don't live long enough to be re-infected by the virus so the virus stays the same · Humans live long enough to be re-infected so their antibodies can drive evolution · By 2009, the H1 virus in pigs was totally different to the H1 virus in humans Avian influenza h5n1 - worried will get another outbreak. Worry is that it will mutate and transfer to other species.
How can arbovirus spread be controlled?
Release mosquitoes with a bacteria known as woolbachia
When does diarrhoea occur when infected by vibrio cholerae?
There is SUDDEN (not late) onset of watery diarrhoea
Examples of viral routes of transmission
Respiratory --> influenza Faeco-oral --> rotavirus Contact --> Herpes simplex (HSV) Zoonoses (from the animals) --> Rabies Blood --> HIV, Hep B/C Needles Insect bites Sexual --> HIV, HPV Maternal --> Hep B Germ line (to the offspring) --> retroviruses
How is SARS transmitted?
Respiratory droplets; concentrated sources on lift buttons and in bathroom drains
List 4 stages of the HIV life cycle that can be inhibited by current antiviral drugs? (2 points)
Reverse transcription DNA integration Entry HIV protease inhibitors
Examples of intracellular bacterial pathogens
Salmonella, mycobacterium, shigella, listeria
septic shock
Septic shock = severe sepsis + hypotension despite fluid resus (shock = poor tissue perfusion)
When there is chickenpox outbreak at school...
Siblings are at high risk of contracting infection
What kind of genetic material does HIV have?
Single-stranded RNA
What kind of genetic material does Measles have?
Single-stranded RNA
What kind of genetic material does Polio have?
Single-stranded RNA
What kind of genetic material does influenza have?
Single-stranded RNA
What is viral tropism?
Site of replication
What effect does cholera toxin have on ion channels in host's epithelium?
The cholera toxin causes opening of ions channels
Infective dose
The minimum amount of a pathogen that is required to establish disease Influenced by factors mentioned on last slide Measured in CFU (colony-forming units)
What is the tropism of a virus?
The place where it replicates. Can be dependent on receptors (HIV and CD4 on T cells), intracellular host cell components (poliovirus with mutations in their 5' noncoding regions cannot utilize neuronal host cell factors to translate their mRNAs) or extracellular factors (influenza HA protein needs to be cleaved by host encoded protease expressed in respiratory secretions)
Tropism definition
The predilection of viruses to infect certain tissues and not others. This is based on: · Susceptibility - receptor interactions · Permissivity - ability to use the host cell to complete replication . Accessibility - ability of the virus to reach the tissue
What is vaccine efficacy?
The reduction in the incidence of disease among people who have received a vaccine compared to the incidence in unvaccinated people.
perfusion
The supply of oxygen to and removal of wastes from the cells and tissues of the body as a result of the flow of blood through the capillaries.
What are most human fungal infections?
There are about 150 serious fungal infections of humans · MOST ARE ASCOMYCETES - 90% of all human fungal infections · Basidiomycetes consists of many of the mushrooms that we find · Out of the basidiomycetes, Cryptococcus neoformans and Cryptococcus gatii cause the largest burden of disease - CRYPTOCOCCAL MENINGITIS · These cryptococci can be inhaled into the lungs - alveolar macrophages are usually good at mopping up fungal cells as we inhale them · Cryptococcus can get into the brain (a sugar rich environment) and cause meningitis
Acute viral infections
Typically: • Over matter of days. Virus comes in and overcome innate defences to establish infection, grow and adaptive response kicks in (T cells then B cells), virus disappears and then have memory 3 outcomes: Acute infection and clearance 1. Colds and influenza - many unapparent or asymptomatic infections 2. Adaptive immune response provides immunity 3. Viruses continue to circulate in populations by antigenic variation Acute infection and death 1. Smallpox caused by variola virus- Mortality 20-80%. Infection of skin. Viral growth factors induce proliferation of skin resulting in pox. 2. Dengue (arbovirus)- Dengue haemorrhagic fever. Responsible for 500,00 hospitalizations each year with 5% fatalities. Leakage of blood plasma from capillaries. Detected by increase in red cell count and decrease in platelet and protein level in blood. Tendency to severe bruising, and bleeding. Patient deteriorates even after fever drops; shock. Treat with iv fluid replacement. Acute infection with accidental pathogenesis 1. Polio - fecal-oral route, causing localised infection of small intestine. Poliomyelitis. Neurovirulent virus infects motor neurones and can cause paralysis. Now don't really see cos of polio vaccine 2. Rubella - causes mild rash except in early stage fetus where virus has strong tropism for dividing neuronal tissue Leads to classic triad: o Deafness o Eye abnormalities (cataracts) o Congenital heart disease
Using viruses as vectors in vaccination
Use a virus against another virus
Antivirals targeting influenza virus replication
Use drugs: zanamivir and oseltamivir. Inhibit neuraminidase enzyme aka (sialidase) on virus. Viral neuraminidase cleaves terminal neuraminic acid (also called sialic acid) on host cell surface. Neuraminidase cleaves the sialic acid molecule, thereby freeing the virus to infect other cells in the host organism. Ribarvirin interrupts replication by acting on RNA polymerase Amantidine can affect protein synthesis Amanatidine - prevents virus escaping endocytic vesicle into cell
Ebola vaccine
Vaccines = active immunisation; switching on immune system and asking b cell and t cells to work GSK: Chimpanzee adenovirus vectored vaccine that expresses Ebola G protein Merck : Vesicular Stomatitis Virus vectored vaccine expressing Ebola virus G protein. Passive immunization: (make antibodies somewhere else, purify and then transfer) Zmapp : A cocktail of 3 monoclonal antibodies raised against G protein of a previous strain of Ebola virus given as passive immunotherapy Unusually production in plants: limited capacity. Serum therapy: blood from survivors
Two ways to defend ourselves against viruses
Vaccines: • Prophylactic • Live or inactive • Public health strategy --> Herd immunity or defined target group? • Safety > efficacy (RSV vaccine in 1970s!) • Governments and WHO Antiviral drugs: • Therapeutic • Random screen or rational design • Define target group: very sick or over the counter? • Individuals
Baltimore classification
Viral families by nature of the GENOME (useful for diagnostics and treatment). · Viruses are classified by the type of genome that the viruses have · They can be made from DNA or RNA (and in various forms) · Some DNA viruses have Double Stranded DNA whereas others have Single Stranded DNA · RNA viruses carry their RNA as: o Positive Strand - sense strand which can be translated directly o Negative Strand - antisense strand o Double Stranded RNA · Some viruses have DNA or RNA but follow a different cycle for making it - e.g. carrying an RNA genome but going through a DNA intermediate inside the cell to make more RNA (retroviruses) · The classification based on genome is known as the BALTIMORE CLASSIFICATION
Replication cycle of HIV-1
Virus is on the outside 1. GP120 glycoprotein on virus capsule interacts with CD4 on the host cell membrane 2. Once it interacts with CD4 it can interact with a co-receptor (CCR5 or CXCR4) which brings the virus within close proximity of the cell membrane 3. Virus membrane and host membrane will fuse and viral content (HIV RNA, reverse transcriptase, integrate and other viral proteins) will be inserted into the cell 4. Enzymatic copying of viral genome by reverse transcriptase (found in virus) to produce viral DNA 5. The DNA moves into the nucleus and is integrated into the host genome 6. Host replicates viral DNA and transcribes into mRNA and, subsequently, translated into proteins. 7. All of these then come together at the cell membrane and produce a new immature HIV virus particle. proteases hydrolyses proteins, important in final steps of HIV maturation, mature HIV leaves the cell Can target reverse transcriptase in drugs, as humans don't have this enzyme. So: gp120, cd4, ccr5/cxcr4, fusion, preintegration complex (viral RNA, viral proteins and host proteins --> reverse transcription by reverse transcriptase), integrase and protease
Virus definition
Viruses are infectious OBLIGATE intracellular PARASITES. · Obligate = cannot complete its life-cycle without being inside a host cell (uses the host cell's genetic machinery)
Features of viruses
Viruses are infectious OBLIGATE intracellular PARASITES. · Obligate = cannot complete its life-cycle without being inside a host cell (uses the host cell's genetic machinery) · Virus genome comprises of DNA or RNA. · Within an appropriate cell, the viral genome is replicated and directs the synthesis, by cellular systems, of more viral components and genomes. · The components affect the transport of replicated viral genomes through the environment to new host cells. · Average size of a virus = 100nm, smallest ones = 20nm, largest about 1 micrometre. · can only see with electron microscope Bacteria visible through light microscope
Cytopathic effects of viruses
Viruses have to be grown inside host cells · Cells need to be grown in the laboratory before being infected by a virus · If you put a virus on healthy cells you get a change in cell shape and you get cell death · This is the CYTOPATHIC EFFECT (CPE) which is defined as the death of the cell as a result of being infected by a virus · The CPE could be due to the virus taking over your genetic machinery so that the cell is no longer able to produce the proteins that it needs to survive - the cell is destined to die · Death is probably due to apoptosis · In other words: virus has to compete with host proteins synthesis - they do this in such an aggressive way that it destroys the capability of the host cell to use its own machinery · Lack of stain in dead cells
Central dogma (to classify viruses and how to treat virus)
Viruses may have DNA as starting, or just start with RNA itself. Some who carry RNA as negative sense RNA, it is complement of mRNA (complement of bases and runs in opposite direction) DNA --> RNA --> Protein · Most of the time, living things have a DNA genome which is transcribed into RNA and then translated into protein · Some viruses carry their RNA as a positive sense strand - this means that as soon as it enters the cell, ribosomes can translate that into a protein · Some viruses carry their RNA as a negative sense strand - this cannot be translated straight away o It needs to be transcribed back into a positive sense complimentary copy o Viruses with negative sense RNA must carry proteins/enzymes/machinery that will convert the genomes back into sense RNA o Otherwise, they are inert
Viral oncogenesis
Viruses that cause cancer · May encode an oncogene · Interferes with the host cell cycle to enhance their own replication (e.g. always want cell in S phase so can replicate DNA) By making the host cell into a cancer cell, the virus can replicate more and keep moving onto new cells. · EXAMPLES: o Papillomaviruses (cause warts) encode inhibitors of tumour suppressor p53, E6 and E7 genes - forces cells into S phase o Kaposi Sarcoma-Associated Herpes Virus (KSHV or HHV8) and Merkel Cell Polyoma were discovered by finding non-human genetic material in tumours o HTLV-1 (retorovirus) causes adult T cell leukaemia · Hepatitis B and C viruses cause hepatocellular carcinoma o 350 million carriers of HBV o 600,000 annual deaths o HBV is a hepadnavirus (has a DNA genome) but it uses reverse transcriptase during a stage in its life cycle o Transmitted through blood and semen - much more infectious than HIV o There is a vaccine with sAg expressed from yeast o HCV is a blood borne virus o Hep C infected lot of people in 70s and 80s before blood was screened. o Now seeing a rise in liver cancer resulting from these chronic infections o 4% of people with HCV will go on to get hepatocellular carcinoma o Antiviral therapies for HCV are available (hep C). Hep B still no really good drugs · Epstein-Barr Virus o A gamma herpes virus, most common virus infection of mankind o 95% of us are infected with Epstein-Barr Virus o Most people --> lytic infection in childhood or infectious mononucleosis (glandular fever) in young adulthood. And then remains latent in B cells. o Passed on in saliva o Some people infected with EBV will go on to develop cancer (combined with other circumstances): · Burkitt's Lymphoma · Hodgkin's Lymphoma · Nasopharyngeal Carcinoma o Incidence of these varies geographically suggesting other predisposing factors to these outcomes e.g. there's link between malaria and EBV.
Syncytia
Viruses with surface proteins that can fuse at neutral pH often fuse cells together. In pic: Healthy cells on outside, inside infected cells all fused together. There is a big bundle of cells in the middle which have stuck together (effectively becoming one massive cell with many nuclei) Some viruses don't make plaques - they do other things to the cell such as syncytia formation · HIV has the ability to fuse at the cell membrane · When a cell has been infected by HIV, it begins expressing HIV glycoproteins on their surface · One cell can fuse with another cell because of the interaction between the HIV protein and the receptor on the opposite cell · In the laboratory, this is way you can visualise whether there is a virus or not in a sample · You can take a sample of blood, drop it on some susceptible cells and if you see syncytian formation, it suggests that there is virus present
Fungal allergies
WORLDWIDE PROBLEM Inhalation of fungal spores (numerous, diversified air-borne microorganisms) may produce a wide range of allergic diseases: · Rhinitis · Dermatitis · Asthma · ALLERGIC BRONCHO-PULMONARY ASPERGILLOSIS (ABPA) caused by aspergillus fumigatus - found in compost. Occurs in 2.5% of asthmatics o Allergic responses differ by individual and by fungal species
Example of interferon treatment
Was a few years ago the only option for some viruses, eg. Pegylated (chemical modification helps protein survive longer) IFN given with ribavirin for hepatitis C virus - only treatment available then.
Examples of emerging viruses
West Nile Fever re-emergence in NY; birds in Bronx Zoo; they are not sure how this came to the USA Illegal birds, infected mosquito, viraemic human Dengue Wild aedes aegypti. 50-100 million cases of DF and 300 000 cases of DHF. Zika mosquitos in Brazil Ebole, Hendra, Nipah -> they can't infect efficiently so they pop up every now and then. Chimpansees, Bats, Civets SARS Severe acute respiratory syndrome. In China in 2002, 8422 case. Chinese horseshoe bats. Civets and racoon in markets MERS found in camels in Middle East, H1N1, H3N2 , avian flue due to influenza virus being able to rearrange and change. Some variants are restriced to human, swines and birds. If the mix, then the dangerous variants appear.
What does outcome of virus infection depend on?
balance between virus virulence and host response · VIRAL SEQUENCE- · Two strains of polio virus may vary in virulence · One mutations may turn it into a live attenuated virus (sabin) · Another mutation may make it invade the motor neurone and causes flaccid paralysis (poliomyelitis) · VIRUS LOAD - · The first child in a family to contract chicken pox often has a milder illness · This may be because the second child is in closer contact to an infected individual and hence gets infected by a higher dose Might be affected by route of transmission · HOST IMMUNE RESPONSE/STATUS · HOST CO-MORBIDITY - . Asthmatics and respiratory viruses · Obesity · Immunosuppression · Immunodeficiency · Elderly · Diabetes Mellitus · Pregnancy - immunosuppression in pregnancy · Potentially gender - is man flu real? · CO-INFECTIONS- · Secondary bacterial infection following influenza (morens and taubenberger), big cause of death - compromises respiratory tract and immune system · HHV8 causes Kaposi Sarcoma in HIV infected individuals: o Hepatitis Delta Virus (a small defective RNA virus) only infects people with HBV (hep b virus) infection - it suppresses HBV replication but causes severe liver disease with rapid progression to cirrhosis and hepatic decompensation · OTHER MEDICATIONS · HOST GENETICS - Genetic Resistance and Susceptibility · CCR5 delta 32 mutation protects against HIV-1 infection · Killer-cell immunoglobulin-like receptor (KIRs) e.g. NK cells can determine the outcome of a hepatitis C virus infection · Interferon-induced transmembrane protein 3 (IFITM3) was associated with a severe outcome in 2009 H1N1 pandemic. Most people ended up in hospital were those with the CC genotype even though in population CC was less common . HOST AGE/GENDER
Hep B and C transmission
blood
How is Ebola transmitted?
bodily fluids
true pathogen
can cause disease in normal, healthy people e.g. staphylococcus aureus from skin causing abscess and bacteremia
opportunistic pathogen
can only cause disease when they are given the chance E.g. Staphylococcus epidermidis is a weedy pathogen but if it gets into a prosthetic hip it can colonise the metal in the hip and cause infection.
Summary of how viruses are detected, cultivated and manipulated
visualized with the electron microscope. Lab culture effects (cytopathic, syncytia, plaque assays). Antibody (fluorescent). Viral proteins (ELISA). Viral genetic material (PCR). Manipulation using cDNA representing viral genome, introducing into cells, getting altered virus out. 1) 'see' viruses by their effects on their hosts. This can be a plant or animal (in vivo) or cells cultured in the laboratory (in vitro). Viruses may produce effects on the cells as they replicate such as CYTOPATHIC effect, cpe, or SYNCTIA, inclusion bodies, membrane blebbing. Dilutions of virus can be ASSAYED to find the end point at which they no longer produce such effects. This allows their quantification. e.g. plaque assay in which foci of infection can be counted. 2) Some viruses produce little visible change on the cells but can be detected using ANTIBODIES to their proteins that will be expressed during replication. Immune FLUORESCENE or immunohistochemistry allow the antibody binding to be seen. 3) Techniques for 'seeing' viruses that do not use cell cultures include detection of the particles or the VIRAL PROTEINS, or the virally encoded nucleic acid. Viruses that attach to red blood cells can be visualized using the HAEMAGGLUTINATION assay. Cells infected by viruses can be lysed and antibodies to viral antigens used in Western Blot or enzyme-linked immunosorbent assay (ELISA). 4) Nucleic acid detection is usually by the polymerase chain reaction (PCR). Viruses can be manipulated in the laboratory in order to study them. Classical genetic techniques involve growing the virus under different conditions e.g. temp or w/ drug, that may induce changes in the genome and studying their effects. Reverse genetics techniques involve engineering cDNA that represents the sequence of the virus genome, introducing it into cells and recovering an altered virus.
Give an example of genetic resistance to a viral disease (brainscape)
with HIV. delta 32 mutation causes formation of a different CCR5 receptor and so these are people are naturally resistant HIV
Superficial mycoses
· Affecting skin or hair shaft · No living tissue is invaded - no cellular response from host e.g. dandruff by malassezia globosa
What are the three types of illness caused by fungi?
· Allergies · Mycotoxicoses . Mycoses Fungi can be classified by the level of tissue affected as superficial (live on skin), subcutaneous and systemic (deep-seated into body)
Immunostaining virus infected cells
· Antibodies generated in the lab to unique virus proteins indicate which cells are infected or where in the cell the virus proteins are located. · We can visualise a virus because it produces things that are different inside the cell · If we suspect a person has RSV, we can inoculate their sample onto a cell and wait for the cell to start producing a protein that is unique to the virus · You then use an antibody which can detect that protein In the image above - the antibody shows up green if the protein is present in the cell (can see speckles of viral protein as well)
West Nile Virus
· Belongs to Japanese encephalitis group of flaviviruses - cause disease by going to the brain · 1999 - West Nile Virus was found in New York - bad year for mosquitoes in NY · Some elderly people succumbed to a brain disease - crows and birds at the zoo became ill · Subtractive differential analysis used to diagnose a human brain sample revealed WNV RNA · RT-PCR of WNV RNA showed that the virus originated from Israel - could be by illegally imported bird? Viraemic human? Infected mosquito? · Vector: culex tarsalis; horses and humans are dead end hosts - it doesn't spread any further · West Nile Virus arrived in the USA 2002 and has spread to every state by 2008 Infection of mosquitoes. BIRDS ARE NATURAL HOSTS
Cutaneous mycoses
· Caused by dermatophytes or keratinophilic fungi · Produce exrtacellular keratinase enzymes which are capable of hydrolysing keratin · Inflammation is caused by host response to metabolic by-products · Trychopyton and Microsporum are particular examples of this · Cutaneous mycosis is also called dermatophytocis and dermatomycosis · Initially thought to be caused by wormlike parasites · Therefore given the name Tinea - latin for worm · Tinea Capitis- head/neck (scalp, eyebrows, eyelashes) § The most common paediatric dermatophyte infection § Esp. attack hair shafts and follicles § Mainly in Africa § Disfiguring and affects children's self esteem and school time · Tinea pedis- foot = athelete's foot § Incredibly infectious - hence should spray yoga mats before using § Trichophyton rubrum = world's most prevalent dermatophyte § 70% of the population will be infected with Tinea pedis at some point · Tinea corporis - body § Tricophyton sp. AKA ring worm § Treated with antigungal creams or orally · Tinea cruris - groin · Tinea unquium - finger/toe nails Account for 10-20% visits to dermatologist
Persistent viral infections
· Chronic - low level replication in tissues which regenerate (EXAMPLE: papillomaviruses in warts). Also, Chronic carriers of help B and hep C viruses. And sometimes viruses in nervous system, because it is an immunoprivileged site so don't get much surveillance from T cells and B cells · Latent - viral genomes are maintained but no virus is seen until episodes of reactivation at times at which you are immunocompromised. EXAMPLE: herpes viruses (can get cold sores), varicella zoster Strategies of Viral Persistence There are lots of accessory genes, which allow viruses to evade our immune responses. · Evading immune surveillance o MHC Downregulation. Compensation for lost MHC class I; HCMV cytomegalovirus · CTL escape by mutation - Hepatitis C virus · Infecting tissues with reduced immune surveillance: o CNS; measles SSPE, herpes virus oSkin; papillomavirus Latency · E.g Herpes simplex virus · No virions or viral products detected, residing inactive in the trigeminal ganglion of face (only Latency Associated transcripts or LATs detected) · Neurons do not divide (live till you die), and contain hundreds of copies of HSV genome · So if then nerve gets infected it may contain lots of copies of viral genome and every now and then it will get reactivated. Reactivation frequency varies between hosts EXAMPLE of latency: Herpes Simplex Virus · The virus first enters through the skin epithelial cells = primary site of infection and replicates) · It gets into the sensory neurone (secondary site of infection) by retrograde transport and sits there in a latent state · When you get reactivation of the virus, it begins to make new copies of itself and travels back to the end of the nerve to the skin (anteretrograde transport) and replicates through the skin (forming a cold sore)
Examples of deep mycoses
· Coccidioides immitis- in new world e.g. inhaling desert air · Histoplasma capsulatum - in cavers a lot
Example of latent persistent viral infection
· E.g Herpes simplex virus · No virions or viral products detected, residing inactive in the trigeminal ganglion of face (only Latency Associated transcripts or LATs detected) · Neurons do not divide (live till you die), and contain hundreds of copies of HSV genome · So if then nerve gets infected it may contain lots of copies of viral genome and every now and then it will get reactivated. Reactivation frequency varies between hosts EXAMPLE of latency: Herpes Simplex Virus · The virus first enters through the skin epithelial cells = primary site of infection and replicates) · It gets into the sensory neurone (secondary site of infection) by retrograde transport and sits there in a latent state · When you get reactivation of the virus, it begins to make new copies of itself and travels back to the end of the nerve to the skin (anteretrograde transport) and replicates through the skin (forming a cold sore)
Subcutaenous mycoses examples
· E.g. sporotrichosis - rare but ongoing epidemic of cat-transmitted (by scratch) sporotrichosis in brzeil E.g. sporotrichosis, chromoblastomycosis and Mycetoma - chronic infection of the skin, subcutaneous tissue and sometimes bone, characterised by discharging sinuses filled with organisms - increasingly seen in combat- related blast wounds and thorn scratches
Examples of arboviruses
· EXAMPLES: o Yellow Fever o Dengue o West Nile o Zika o Chikingunya · Many of these are flaviviruses and alphaviruses- single strand positive-sense RNA genomes · Mosquito host (replicate within the mosquito) · CONCERN: global warming leads to an increase in the global distribution of mosquitoes · Culex tarsalis is the vector of the West Nile Virus With some of these arboviruses, humans and horses are dead end hosts
Global influences of emerging viral infections
· Environmental modifications/demographics · World population · Climate change · Travel · Farming practices; monocultures · Immunosuppressed humans Medical progress
Norovirus propogation new information
· For many years scientists not been able to culture norovirus in lab · Even inoculating virus onto primary cultures from gut has not worked · Scientists recently figures out how to culture norovirus in the lab · Norovirus seems to infect B CELLS IN OUR GUT · Norovirus needs to bind to ENTERIC BACTERIA in order to infect those cells - symbiosis between virus and bacteria It may be that the bacteria transport the virus across the intestinal epithelium to gain access to target cells below.
Spore dispersal in fungi
· Fungi produce a large number of spores · They are dispersed over large distances · Humans are constantly exposed to fungal spores · Commensal organisms & skin colonisers, transmitted by contact e.g. athlete's foot
Measles virus receptors, tropism and pathogenesis
· Has H receptor (hemagglutinin) which binds to different receptors that it binds to at different times in replication cycle while in host: o SLAM (CD155) - found on dendritic cell (survey environment and capture invading pathogens) o Nectin 4 · The vaccine strain of measles uses CD55, evolved to use different receptor, can make virus that is attenuated when passaged in lab (mentioned earlier). Doesn't cause same infection as doesn't use same receptor as normal virus. · SLAM and Nectin 4 are used at different stages · When infecting someone for the first time, measles enters via the respiratory tract · Virus binds to SLAM on immune cells (e.g. dendritic cells) which are surveying the area · By doing this, they hijack a ride to the lymph nodes · At the lymph nodes, the virus is exposed to more cells expressing SLAM which it then infects causing immunosuppression (lasts for about 3 years), also amplification here · When measles wants to leave the host and move on to another host, it uses NECTIN 4 · Virus is carried from the lymph nodes to the lungs · It crosses from the blood to the airspace by binding to Nectin 4 which is on the bottom of the airway epithelial cells (basal side) · Viruses replicate in airway epithelial cells and burst out into the airspace to spread via the respiratory route · Overall: entry to new host = SLAM on immune cells (immunosuppression) o Exit from infected host = nectin 4 on airway epithelia
Hep B and C viruses link with cancer
· Hepatitis B and C viruses cause hepatocellular carcinoma o 350 million carriers of HBV o 600,000 annual deaths o HBV is a hepadnavirus (has a DNA genome) but it uses reverse transcriptase during a stage in its life cycle o Transmitted through blood and semen - much more infectious than HIV o There is a vaccine with sAg expressed from yeast o HCV is a blood borne virus o Hep C infected lot of people in 70s and 80s before blood was screened. o Now seeing a rise in liver cancer resulting from these chronic infections o 4% of people with HCV will go on to get hepatocellular carcinoma Antiviral therapies for HCV are available (hep C). Hep B still no really good drugs
New viruses recently detected/discovered
· Hepatitis C (discovered as a 'non A non B hepatitis') · HPV 16 and 18 - cause cervical cancer · HHV8 (KSHV) - cause of Kaposi sarcoma, noticed during AIDS pandemic Merkel cell polyoma virus - cause Merkel cell carcinoma; identified in tumours as non human sequence
How do antibodies exert selection pressure on viruses?
· If a person is infected with a virus for which the patient already has antibodies for HA, then the person is immune and protected · BUT, if the person infected has a subneutralising amount of antibody - the virus will replicate in that person and only the fittest viruses survive - ones which change their spike proteins · This is ANTIGENIC DRIFT - antigens gradually change over a period of time due to pressure exerted by antibodies. Hence for influenza, vaccine updated every year to best represent the circulating strains NOTE: RHINOVIRUSES don't show antigenic drift but there are so many of them circulating at the same time so you are likely to catch many of them in your lifetime.
Staphylococcus aureus virulence factors
· Interference with Host Cell Function - e.g. S. aureus makes superantigens interfere with normal T cell function, causes them to proliferate loads Immune Evasion - e.g. S. aureus makes leukocydins which causes neutrophil death and abscess formation
Portal of entry- genital tract
· Intrinsic pathogens - from Large intestine (strep group B (pregnancy)), candida/yeast · Extrinsic - Sexually transmitted (neisseria gonorrhoeae, chlamydia trachomatis, treponema pallidum (syphilis), HIV, HSV
Bacteria general properties
• Small • Unicellular • No internal organelles • Haploid • Some have flagella Forms: cocci (round), bacilli (rod), sprilli (spiral)
Dengue haemorrhagic fever (DHF)
· Most serious thing to get with dengue fever. Get it in endemic areas. · 3 billion people live in high risk areas. Case fatality of DHF = 5% · There are FOUR SEROTYPES of dengue virus - antibodies against one will bind to the other one but will not protect you; cross reactivity but no cross protection. By binding and not protecting makes it a lot worse. · First time you get dengue you are sick but not that sick · If, the second time you get infected, you get infected by a different serotype - then the antibodies you developed the first time will make you MORE sick · This is called ANTIBODY DEPENDENT ENHANCEMENT OF THE INFECTION · Becomes increasing problem due to much more increased mosquito population · Risk Factors for DHF: o Pre-existing anti-dengue antibody • previous infection • maternal antibodies in infants o Virus strain o Age - increases likelihood of having pre-existing antibody o Higher risk in secondary infections o Higher risk in areas where there are two or more serotypes circulating simultaneously at high levels (hyperendemic transmission) · DHF = 5% mortality · Spread of DHF correlates with upsurge in mosquito population Dengue antibodies can either neutralise infection or enhance infection
How to make recombinant attenuated virus vaccine?
· Pathogenic virus genome typically consists of receptor-binding gene, virulence gene and capsid protein genes · You can either mutate the virulence gene or delete the virulence gene You then get a virus which is IMMUNOGENIC but NOT VIRULENT
Sample acquisition in fungal diagnosis
· Sample Acquisition - useful for CF diagnosis o Skin o Sputum o Bronchoalveolar Lavage o Blood o Vaginal swab/smear - e.g. candida o Spinal fluid - e.g. cryptococcus causing death in HIV patients, determining whether bacterial or fungal meningitis o Tissue biopsy
Properties of a good vaccine
· Stimulates an effective immune response · Safe and does not cause adverse reactions · Inexpensive · Stable · Easy to administer Simple for both manufacturer and regulatory authorities to control In summary: a good vaccine provides substantial benefit to health at low cost and low risk
Pneumococcal conjugate vaccine
· Streptococcus pneumoniae is significant cause of meningitis and septicaemia worldwide · But also pneumonia (mucosal infection) · More than 90 serotypes determined by capsular polysaccharide o Antibiotic resistant associated with some serotypes · Vaccines: o Pneumovax II™, 23-valent plain (unconjugated) polysaccharide vaccine - used in older people with mature immune response and respond better to polysaccharides o Prevenar 7™, Prevenar 13™ and Synflorix™ conjugate vaccines - for younger people who respond poorly to polysaccharides · 97% efficacy, and estimated efficacy of 18% against pneumonia. Reduction in otitis media by around 10%. Now because of herd effects has even bigger effect · One of problems with using only 13 serotypes of vaccine out of 100, is get serotype replacement phenomenon: So although major impact on reducing disease, there's some replacement disease: decline in 13 valent vacciens, but increase in non-vaccine serotypes.
What is tropism dependant on?
· Susceptibility - receptor interactions · Permissivity - ability to use the host cell to complete replication Accessibility - ability of the virus to reach the tissue
Naming a virus
· The disease - Poliovirus, Rabies · The person who discovered it - Epstein Barr Virus · The place it was discovered - Coxsackievirus · The part of the body affected by the virus - Rhinovirus (nose), Adenovirus, Hepatitis The way it was spread - Dengue, Influenza
Preventing drug resistance in HIV
· There are drugs which target different parts of the HIV life cycle: proteases, integrases, fusion, CCR5 · If you give the patient a protease inhibitor, a single nucleotide switch may make the protease inhibitor ineffective · So you must give a combination of antiviral drugs because in that case, for a virus to be resistant, it would need 5 or so simultaneous mutations, which is unlikely. · Error rate is roughly 1 per 10,000 nucleotides and as each genome is roughly 10,000 nucleotides long, each HIV genome is likely to be slightly different at a single position. · Therefore, monotherapy would almost certainly result in proliferation of a resistant population of HIV. NOTE: HAART = Highly Active Anti-Retroviral Therapy. uses three or four different drugs in combination. This prevents the virus from being able to generate resistance mutants but does lead to difficult drug regimens and is associated with significant side effects.
S. pneumoniae virulence factors
· Toxin Production - e.g. pneumococcus makes a toxin which destroys bits of the lungs (pneumolysin). Pneumolysin : cholesterol-dependent pore-forming toxin affecting lung architecture, thought to be reason why so much lung damage from pneumonia Degradation of Host Molecules
Consequences of bacterial infection acquired via upper respiratory tract
· Upper Respiratory Tract Infection oPharyngitis o Tonsilitis (inflammation of tonsils) o Sinusitis (as it goes up through sinuses via air channels) · Lower Respiratory Tract Infection oBronchitis oPneumonia oPneumonitis oEmphysema · Spread to Adjacent Tissues o Brain abscess oMeningitis o Middle ear infection (oditis) · Spread to Blood Stream o Bacteriaemia e.g. pneumococcal, meningococcal bacteraemia
Portal of entry - urinary tract
· Urinary system connected to outside world by urethra. Urethra is close to anus. In females distance is shorter and hence get more bladder infections, hence bacteria from LI spread and cause infections. · Intrinsic - bacteria from the large intestine (usually intrinsic bateria that infect urinary tract) o E. coli o Klebsiella o Candida · Extrinsic oNosocomial (hospital-acquired) Urinary catheters to help pass urine- allows E.Coli, Klebsiella spp to enter
Manipulating viruses
· Virus genomes are so small that they can be synthesised · When introduced into permissive cells they direct synthesis of all their components and new viruses are made de novo · This allows reverse genetics - the production of new viruses with engineered mutations in their genomes ·
What is the single step growth kinetics of virus
· Virus has been put onto cells and at various times, samples have been collected and titrated out · The amount of virus present is plotted on the y axis and time on the x axis · At the start of the infection there is a known amount of virus · If you take a sample quite early on, there is NO virus at all - because virus has gone inside the cell This is the ECLIPSE phase because we can't see it. · After a while the virus begins making new copies and exiting the cell and becomes detectable · You get logarithmic increase in the number of viruses in the sample (logarithmic phase) It then tails off because cells start dying (cell death) Overall: eclipse --> log --> death
How could complex bacterial invasion have formed?
• Bacteria have much larger genomes than viruses: 500 - 4500 proteins • The majority of the proteins are involved in housekeeping functions e.g. DNA replication, making cell walls • Only a subset of these proteins are involved in virulence • Core Genome (40%) - all the bacteria of a species have these genes - these are housekeeping genes • If you code one bacterium of E. Coli you will get 60% non-core genes, which are different for different bacteria. Makes you consider what is a species. • Bacterial genomes are changing constantly • Bacteria replicated by binary fission - Vertical Gene Transmission - the daughter cells have an identical genome to the parent cell • Core genes (40%) and accessory genes (vary considerably between diff strains) = gene strains Accessory genes are the interesting ones for understanding bacterial pathogens
Vancomycin
• Bactericidal antibiotic • Targets Lipid II component of cell wall biosynthesis, as well as wall crosslinking via D-ala residues Toxicity has limited use, but resistance to other antibiotics has led to increasing use e.g. against MRSA and VRE
rifampicin
• Bactericidal antibiotic • Targets RpoB subunit of RNA polymerase. • Spontaneous resistance is frequent. • Makes secretions go orange/red - affects compliance. E.g. for TB
Daptomycin
• Bactericidal antibiotic • Targets bacterial cell membrane. • Gram-positive spectrum of activity. Toxicity limits dose - given IV and patient needs support .
Linezolid antibiotic
• Bacteriostatic antibiotic • Inhibits the initiation of protein synthesis by binding to the 50S rRNA subunit. Only has gram-positive spectrum of activity.
Other non-genetic mechanisms of bacterial survival (apart from antibiotic resistance)
• Biofilm - form big sticky masses • Intracellular location - hide inside other cells e.g. TB hides inside macrophages • Slow growth - much harder to kills e.g. TB • Spores • Persisters - essentially dormant. AB inhibiting these cells don't kill bacteria
Process after gram staining
• Culture and microscopy • Biochemical and serological tests • Sensitivities to antibiotics • DNA techniques: PCR, sequencing Clinicians need to know which antibiotic to give - Antibiotics might work differently between gram negative and gram positive bacteria because drug might find it difficult to get through two cell membranes in gram negative bacteria
Why are interferons not really used as antiviral treatment?
• Discovered by Isaacs and Lindemann. 1957. • Recombinant interferon 1981-1982 • If virus infects cell, cell recognises virus as foreign due to it's different actions (i.e. protein synthesis), cell switches on interferon response, secretes interferon, acts and warns neighbouring cells to warn them and ramp up interferon stimulating genes and this creates inflammatory response and is protective against any viruses coming. Don't really use as therapy today because it switches on everything including inflammation and fever By giving interferons - you would suppress the virus but you would make the patient feel even worse. • Side effects due to induction of cytokines and other ISGs • Was a few years ago the only option for some viruses, eg. Pegylated (chemical modification helps protein survive longer) IFN given with ribavirin for hepatitis C virus - only treatment available then.
Amniglycosides antibiotics
• E.g. Gentamicin, streptomycin. • Bactericidal. • Target protein synthesis (30S ribosomal subunit), RNA proofreading and cause damage to cell membrane. In addition to preventing it. It corrupts protein synthesis --> misfolding --> break down bacterial membrane • Toxicity has limited use, but resistance to other antibiotics has led to increasing use.
Important multi-drug resistant pathogens
• Gram-negative: o Pseudomonas aeruginosa- Cystic fibrosis, burn wound infections. Survives on abiotic surfaces. Good at colonising e.g. in ventilators o E. Coli (ESBL) GI infect., neonatal meningitis, septicaemia, UTI. o E. coli, Klebsiella spp (NDM-1) As above. o Salmonella spp. (MDR) GI infect. , typhoid fever. o Acinetobacter baumannii (MDRAB) Opportunistic, wounds, UTI, pneumonia (VAP). Survives on abiotic surfaces. o Neisseria gonorrhoeae Gonorrhoea. PEESAN • Gram-positive: o Staphylococcus aureus (MRSA, VISA) Wound and skin infect. pneumonia, septicaemia, infective endocarditis. o Streptococcus pneumoniae Pneumonia, septicaemia. oClostridium difficle Pseudomembranous colitis, antibiotic-associated diarrhoea. Often causes diseases in those receiving antibiotics. o Enterococcus spp (VRE) UTI, bacteraemia, infective endocarditis. SECS Also: o Mycobacterium tuberculosis (MDRTB, XDRTB) Tuberculosis
Reasons for hospital acquired infections
• High number of ill people! (immunosuppression) • Crowded wards • Presence of pathogens • Broken skin - surgical wound/IV catheter • Indwelling devices - intubation • AB therapy may suppress normal flora - useful also in excluding pathogens . Transmission by staff (staff vectors)- contact with multiple patients
Quinolones
• Synthetic, broad spectrum, bactericidal antibiotics. Was hoped bacteria wouldn't form resistance to completely synthesised AB but it didn't work • Act on DNA replication: Target DNA gyrase in Gm-ve and topoisomerase IV in Gm+ve. So DNA can't unwind to replicate.
Viral rashes features and causes
• Systemic viral infection • Virus leaves blood and enters skin • Cells destroyed by virus replication • Koplik spots (small white spots) in the mouth on mucous membranes from measles infection . Lesion on mucus membranes of soft respiratory tract tissues occur before skin rashes: Measles (R0= 30, which means how many people 1 person can infect) and chicken pox spread through the air before the rash is visible.
Antiviral therapy for Hep C virus
• Highly mutatable and need many drugs • HCV is a hepatotropic flavivirus that was spread widely in the 1970s in blood products before screening was put in place. • 170 million people are chronically infected and 4% will proceed to hepatocellular carcinoma. • For more than 20 years therapy relied on interferon treatment with ribavirin. • This was only effective (produced a sustained virological response SVR) in 50% patients infected with the most common genotype 1 of the virus, and carried unpleasant side effects. Now use direct acting antivirals against hepatitis C virus: because of understanding of virus replication, enzymes and processes • Can inhibit HCV proteases, nucleoside polymerase, non-nucleoside polymerae, NS5A (key protein for viral replication) • NS3-4A is the virally encoded protease that processes the polyprotein • First generation protease inhibitors were telaprivir and boceprivir • Monotherapy results in rapid resistance • They have been used successfully with interferon and ribavirin • New Targets: o NS5B is the viral polymerase that replicates the RNA genome o NS5A is the phosphoprotein required by the virus for replication o Structures of these have been solved using crystallography allowing rational based drug design for Directly Acting Antivirals (DAA) o Daclastivir for NS5A • New therapy combines: o Protease Inhibitor - asunaprevir o NS5A inhibitor - daclastivir o With or without: IFN and RBVN Sustained Virological Response (SVR) - not only are the patients feeling better, their viral load is going down
What vaccines are there for influenza?
• Inactivated virus or the subunit vaccine consists only of the spike proteins (HA) • Given to people are risk - e.g. over 65, asthmatics, diabetes, CVD • Also given to healthcare workers • IT DOES NOT GIVE YOU FLU • New strategy of influenza vaccination for children: o Live Attenuated Influenza Vaccine o Cold Adapted - can replicate at 34 degrees (nose) but not at 37 degrees o FluMist delivered intranasally o Updated regualrly and Introudced for children in 2013 Influenza vaccines must be updated regularly become influenza evolves fast
Why might treatment fail for bacteria (apart from ab resistance)
• Inappropriate choice for organism • Poor penetration of AB into target site • Inappropriate dose (half life) • Inappropriate administration (oral vs IV) • Presence of AB resistance within commensal flora e.g. secretion of beta-lactamase
Consequences of antibiotic resistance (increased mortality, morbidity and cost)
• Increased time to effective therapy. • Requirement for additional approaches - e.g. surgery. • Use of expensive therapy (newer drugs). • Use of more toxic drugs e.g. vancomycin for MRSA Use of less effective 'second choice' antibiotics - hence takes longer for patients to recover
Process of attenuating a virus to make a live virus vaccine
• Isolate pathogenic virus from patient • Grow in human cell culture • Take cultured virus and infect monkey cells Gradually the genome of the virus will adapt to the monkey cells and it will become a monkey virus • The virus will no longer grow well in human cells , doesn't replicate as fast, and now attenuated. This is forward genetic procedure: driving virus forward using evolution, without really knowing how many mutations virus has or which mutation is key one that causes attenuation THEORY: Take a virus and pass it through the wrong cell/wrong species - you will make the virus evolve so it is no longer virulent to humans Alternative hypothesis driven way: reverse genetics: understand virus, which mutation is the key one making people sick. Genetically engineer virus to make key and known mutations to stop it making people sick but it still replicates in humans.
Examples of hospital-acquired infections
• Methicillin-resistant S. aureus (MRSA) • Vancomycin-insensitive S. aureus (VISA) • Clostridium difficle • Vancomycin-resistant enterococci (VRE) • E. coli (ESBL/NDM-1) • P. aeruginosa • Acinetobacter baumanii • Stenotrophomonas maltophilia Incidence of drug resistant higher in critical care units in hospitals due to higher dependence and use of antibiotic resistance
Sources of antibiotic resistance genes
• Plasmids - extra-chromosomal circular DNA, often multiple copy. Often carry multiple AB res genes - selection for one maintains resistance to all. • Transposons (mobile pieces of DNA that mobilise other pieces of DNA). Integrate into chromosomal DNA. Allow transfer of genes from plasmid to chromosome and vice versa. . Naked DNA. DNA from dead bacteria released into environment.
Misconceptions of antibiotics
• Resistance against more than one class of antibiotics at the same time would not occur. • Horizontal gene transfer would not occur. • Resistant organisms would be significantly less 'fit' (sometimes true, sometimes not) to cause infection. • Resistance usually emerges soon after the arrival of a new Antibiotic. Two clear exceptions: vancomycin (not great selective pressure as not used much cos toxic) and erythromycin
Rotavirus vaccine
• Rotarix is a live attenuated rotavirus reassortant vaccine • Can massively reduce deaths due to rotavirus infection that leads to diarrhoea and vomiting • UK - 1 in 50 babies admitted to hospital because of rotavirus infection in the first 5 years of life • Early use of the vaccine in the US showed that it can cause diarrheoa and intussusception (bowel blockage) in older babies (>3 months). So the vaccine is only given to babies <15 weeks
What vaccines are there for polio?
• Salk inactivated vaccine o Preparation of virus which has been treated so it can no longer replicate o Isn't a particularly good vaccine - need a large dose • Sabin live attenuated vaccine o This is better o 1 in 7 million vaccinations result in poliomyelitis o If this vaccine was given to people who are immunosuppressed they get a PERSISTING INFECTION - they are reservoirs of live polio virus Salk will be required in long-term for the 'end game'.
Shingles vaccine
• Shingles is a painful rash resulting from the reactivation of a latent varicella zoster virus infection (chicken pox). • Shingles occurs in people after stress, and is more common and more serious in the elderly as their immune system wanes. • Even after the rash has gone (7-10days) pain can remain as Post Herpetic Neuralgia (PHN). • The live attenuated vaccine is similar but distinct from the chicken pox vaccine given to children in some countries (not routinely given in UK due to cost-effectiveness). Shingles vaccine introduced in September 2013 and available today but only for those aged 70 or 78.
Virus routes of entry to the body
• Skin • Mucosal surfaces (respiratory, enteric, genital tract) • Conjunctiva • Blood . Bites - arboviruses spread by insects