Microbiology Flashcards
What is a pathogen?
Organism that causes or is capable of causing disease
What is a commensal?
Organism which colonises the host but causes no disease in normal circumstances
What is an opportunist pathogen?
Microbe that only causes disease if host defences are compromised
What is meant by virulence/ pathogenicity?
The degree to which a given organism is pathogenic
What is asymptomatic carriage?
When a pathogen is carried harmlessly at a tissue site where it causes no disease
What is the resolving power of the eye?
100 micrometers
What is the resolving power of a light microscope?
0.2 micrometers
What is special about chlamydia?
Needs to grow in cells like a virus but is actually a bacteria
What are round bacteria called?
cocci
What are rod shaped bacteria called?
bacillus
What colour are the bacteria if theyre gram positive?
Purple
What colour are the bacteria if theyre gram negative?
Pink
What are the different types of cocci?
Diplococcus
Chain of cocci
Clusters of cocci
What are the different types of rods?
Chain of rods
Curved rod - vibrio
Spiral rod - spirochaete
What are the features of bacteria?
Capsule
Cell wall
Outer membrane
Inner membrane
Chromosome of circular double stranded DNA
pili
Sometime have extra sugar layer
What do motile bacteria have?
Flagella - these spin to allow movement
What stain needs to be done for bacteria that doesnt gram positive or negative stain?
Ziehl-Neelsen stain
What does the Ziehl-Neelsen stain do?
Detects acid- fast bacili
What are the features of a gram positive bacterial cell envelope?
Capsule
peptidoglycan
Lipoteichoic acid
Cytoplasmic membrane
What are the features of a gram negative bacterial cell envelope?
Capsule
Lipopolysaccharide (Endotoxin)
Outer membrane
Lipoprotein
Periplasmic space
Peptidoglycan
Inner membrane
What are the differences between gram positive and gram negative bacterial cell envelopes?
Positive – have phospholipid inner membrane – outside have large peptidoglycan – large peptidoglycan layer linked to phospholipid membrane
Gram negative – have 2 phospholipid membrane –
Lipopolysaccharide (Endotoxin) – can get endotoxic shock
Secreted effector
Protein secretion system
protein toxin or ‘effector’
What are the best areas for bacterial environemnt?
Temperature
<-800C to + 80C (1200C for spores)
pH
<4-9
Water/dessication
2 hours – 3 months (>50 years for spores)
Light
UV
What is the growth rate of bacteria?
Most viruses : Less than 1 hour
E.coli, S.Aureus: 20-30 min
Mycobacterium Tuberculosis: 24 HOURS
Fungi (Candida albicans): 30 min
Mycobacterium leprae: 2 weeks
What is an endotoxin?
Component of the outer membrane of bacteria, eg lipopolysaccharide in Gram negative bacteria
What is an exotoxin?
Secreted proteins of gram positive and gram negative bacteria
Exotoxin features
Composition: Protien
Action: Specific
Effect of heat: Labile
Antigenicity: Strong
Produced by :Gram positive/ Gram negative
Covertibility to toxoid: Yes
Endotoxin features
Composition: lipopolysaccharide
Action: non-specific
Effect of heat: stable
Antigenicity: weak
Produced by : LPS - gram negative
Covertibility to toxoid: No
What is a toxoid?
Toxoid is a toxin treated (usually with formaldehyde) so that it loses its toxicity but retains its antigenicity
What is a bacterial chromosome like?
Bacteria usually have one singular chromosome in a circle made up of DNA – have single RNA polymerase which makes messenger rna
Typically 2-4 10 to the power of 3 kilobases
What is a plasmid?
These can be transcribed and translated to make proteins
Have point mutation in chromosomes
Can be transferred around by different bacteria
What are some genetic variations in bacteria?
Mutation
-Base substitution
-Deletion
-Insertion
Gene transfer
-Transformation eg via plasmid
-Transduction eg via phage – bacterium infected by virus
-Conjugation eg via sex pilus
Classification of bacteria
Please look at slide 49 of ‘bacteria as a cause of disease’ REMEMBER THIS PLEASE
What are some examples of obligate intracellular bacteria?
Rickettsia
-R. rickettsii
-R. prowazekii
-R. conorii
Chlamydia
-C. trachomatis
-C. psittaci
-C. pneumoniae
Coxiella
-C. burnetii
These cannot be cultivated on media
What are some bacteria that may be cultured on artifical media with no cell wall?
Mollicutes
Mycoplasma pneumoniae
M. hominis
Ureaplasma urealyticum
What are the bacteria that can be cultured on artificial media with a cell wall split into?
Growing as filaments and growing as single cells
What are some examples of growing as filaments bacteria?
Actinomyces - A. israelii
Nocardia - N. asteroides
Streptomyces
What are the divisions of growing as single cells?
Rods
Cocci
Spirochaetes
What are spirochaetes eg?
Leptosira
L. . icterohaemorrhagiae
Treponema
T. pallidum
Borrelia
B. burghdorferi
B. recurrentis
What are some examples of gram negative cocci?
Anaerobic: VEILLONELLA
Aerobic: NEISSERIA
N. meningitidis
N. gonorrhoeae
What are some examples of gram positive cocci?
Aerobic: STAPHYLOCOCCUS - S. aureus, S. epidermidis
Streptococcus
Anaerobic: PEPTOSTREP-
TOCOCCUS
What are some examples of streptococcus?
BETA-HAEMOLYTIC
S. pyogenes (A)
S. agalactiae (B)
ALPHA-HAEMOLYTIC
S. pneumoniae
S. oralis
S. milleri
S. sanguis
NON-HAEMOLYTIC
S. bovis
ENTEROCOCCUS
E. faecalis (D)
What are some ZIEHL-NEELSEN
STAIN POSITIVE bacteria?
MYCOBACTERIA
M. tuberculosis
M. leprae
M. avium-intracellulare
M. ulcerans
M. kansasii
Examples of gram positive rods?
ANAEROBIC
CLOSTRIDIUM
C. perfringens
C. tetani
C. botulinum
C. difficile
PROPIONIBACTERIUM
P. acnes
Aerobic: AEROBIC
CORYNEBACTERIUM
C. diphtheriae
etc
LISTERIA
L. monocytogenes
etc
BACILLUS
B. anthracis
B. cereus
etc
If we do a gram film and get a gram + coccus with chains what do we have?
Streptococcus
If we do a gram film and get a gram + coccus with clusters what do we have?
Staphylococcus
What would happen if we did haemolysis on blood agar of streptococcus?
Alpha result: Alpha haemolytic strep ( viridans strep)
Beta result: Beta haemolytic strep Antigenic group (A,B,C,G)
What would happen if we did an optochin test of the alpha haemolytic strep?
If its resistant then its viridians strep
If its sensitive: S. penuomniae
What happens if you do a coagulase or DNAse test on staphylococcus?
Positive = S. aureus
Negative = Coagulase negative staphylococcus
What is coagulase?
Coagulase: enzyme produced
by bacteria that clots blood plasma.
What is the most important sstaphylococci?
S. Aureus
Staphylococci features
at least 40 species:
-‘Coagulase’ + ve or – ve
Fibrin clot formation around bacteria may protect
from phagocytosis.
-Coagulase -ve species, e.g. S. epidermidis only important as opportunistic infections
Normal habitat
-nose and skin
Staphylococcus aureus features?
Spread by aerosol and touch
-carriers & shedders
Virulence factors
-Pore-forming toxins (some strains)
-alpha - haemolysin
-Panton-Valentine Leucocidin ‘PVL’
Proteases
-Exfoliatin
Toxic Shock Syndrome toxin
-(stimulates cytokine release)
Protein A
-(surface protein which binds antibodies in wrong orientation)
MRSA features
resistant to major antibtiocis
Beta-lactams
gentamicin, erythromycin tetracycline
Why is SA a successful pathogen?
Due to a large range of virulence factors.
This includes pore forming toxins.
The PVL toxin is produced by many of the community MRSA strains and causes haemorrhagic pneumonia.
It is encoded by a pro-phage gene that is incorporated into the bacterial genome.
What is alpha haemolysin ?
Low concentration: induces apoptosis in cells by allowing exchange of monovalent ions across the cell membrane.
High concentration: it binds to the lipid membrane and causes massive necrosis
What are some coagulase negative staphylococci?
S.epidermidis:
-Infections are ‘opportunistic’
.immunocompromised,
.prostheses
-Main virulence factor - ability to form persistent biofilms
S.saprophyticus:
-Acute cystitis
.haemagglutinin for adhesion
.urease
What happens in alpha haemolysis on blood agar?
Alpha- haemolysis is due to the production of hydrogen peroxide, which reacts with haemoglobin to form the green compound met-haemoglobin. Get partial greening
S.penumoniae
What happens in beta haemolysis on blood agar?
Beta-haemolysis is due to the production of two pore-forming toxins – streptolysin O and S. Streptolysin O is oxygen sensitive and is very antigenic
e.g. S.pyogenes
What is antigenic sero grouping?
For Beta haemolytic strep only
Carbohydrate cell surface antigens
Lancefield Groups A-H and K-V
Group A - S.pyogenes
throat, skin, post partum
Group B - S.agalactiae neonatal infections
What is the lancefield microbead agglutination test?
Antiserum (antibodies) made that recognise each group
Tagged to tiny plastic beads
added to a suspension of bacteria
Antibodies bind bacteria and beads clump together
Visible to naked eye
What are the virulence factors (enzymes)
Hyaluronidase
- spreading
Streptokinase
- breaks down clots
C5a peptidase
- reduces chemotaxis
What are the virulence factors (Toxins)?
Toxins
Streptolysins O&S
-binds cholesterol
Erythrogenic toxin
-Streptococcal pyrogenic toxin e.g. SPeA – exaggerated response
What are the virulence factors (Surface)?
Capsule - hyaluronic acid
M protein – surface protein - M protein is an antiphagocytic protein and some strains carry a hyaluronic capsule layer, which is non-antigenic.
(encourages complement degradation)
What are some infections caused by S.pyogenes?
Respiratory
-Tonsillitis & pharyngitis - most common
Skin and Soft tissue
-Impetigo
-cellulitis
Scarlet fever
rheumatic fever
Features of S.pneumoniae?
Normal commensal in oro-pharynx ~ 30% of population
Causes - pneumonia, otitis media, sinusitis, meningitis
Predisposing factors
-impaired mucus trapping (e.g. viral infection)
-hypogammaglobulinaemia
-Asplenia
-HIV
What are the virulence factors of S.Pneumoniae?
Capsule
-polysaccharide (84 types), antiphagocytic
.polysaccharide vaccine ‘PPV’ 23 types
.conjugate vaccine ‘PCV’ 13 types
Inflammatory wall constituents
-teichoic acid (choline)
-peptidoglycan
Cytotoxin
-pneumolysin
What are some features of Viridans group streptococci?
alpha- haemolytic (or non-haemolytic)
Optochin resistant
Some cause dental caries & abscesses
Important in infective endocarditits
-S. sanguinis, S. oralis
Cause deep organ abscesses (e.g. brain, liver)
Most virulent are the “milleri group”
-S.intermedius, S.anginosus, S.constellatus
Viridans streptococci is a term used to describe collectively the oral streptococci.
What are some aerobic gram positive bacili?
Listeria monocytogenes
Bacillus anthracis (Anthrax) - spore forming
Corynebacterium diphtheriae
What are some anaerobic gram positive bacili?
Clostridia…: spore forming, survive in environment, produce toxins
C. tetani
Tetanus
C. botulinum
Botulism
C. difficile
antibiotic associated diarrhea
pseudomembranous colitis
Tetanus
From infected wounds
Toxin inhibit GABAs neurotransmission and cause muscle contractions and spasms progressing from head to body
‘Risus sardonicus ‘orrictusgrin
Botulism
From contaminated food (canned) or infected wounds - Botulinum toxin causes paralysis spreading from head to body
What is the model of the bacterial cell envelope in gram negative bacteria?
LPS (‘endotoxin’) forms the outer leaflet of the outer membrane of G-ves, and comprises:
1.Lipid A, the toxic portion of LPS - anchored in the outer leaflet of outer membrane.
2.Core (R) antigen, short chain of sugars, some are unique to LPS.
3.Somatic (O) antigen, a highly antigenic repeating chain of oligosaccharides.
What are some pathogenecity determinants/ virulence factors?
any product or strategy that contributes to pathogenicity/virulence
1.Colonisation factors: adhesins, invasins, nutrient acquisition, defence against the host
2.Toxins (‘effectors’): usually secreted proteins that cause damage, subversion
What are enterobacteria?
Family Enterobacteriaceae = more commonly termed “Enterobacteria”*
Rods, most are motile (peritrichous flagella) – flagella over entire surface of bacteria
Facultatively anaerobic
Some species colonise the intestinal tract (in a good or bad way!)
Family within proteobacteria
Rod shaped and gram negative: most gram negative are rod shaped
What are some cell surface antigens of Gram- negative bacteria?
H antigen (flagellum)
O (somatic) antigen (LPS)
K antigen (exopolysaccharide ‘capsule)
What is a serovar?
Antigenically distinct variants of a single species are referred to as ‘serovars’, i.e. E. coli O157:H7 (EHEC) and E. coli O45:K1:H7 (NMEC) are different serovars of E. coli.
What does amino acid/ carbohydrate sequence variation in cell surface structures cause?
Gives rise to antigenic variation among species AND between different strains of the same species.
What is Escherichia coli?
Commensals - most abundant facultative anaerobe (107-108/g faeces) Peritrichous flagella
What are the principal infections caused by pathogenic Escherichia coli strains?
Wound infections (surgical)
UTIs (cystitis; 75-80% ♀ UTIs - faecal source or sexual activity; catheterisation - most common type of nosocomial infection)
Gastroenteritis
Travellers’ diarrhoea
Bacteraemia (sometimes leading to sepsis syndrome)
Meningitis (infants) - rare in UK
Why are some E.coli strains pathogenic?
Several ‘pathovars’ (distinct pathogenic strategies)
Common ‘core genome’ (most genes are common to all strains/pathovars)
Acquisition of pathogenicity genes
‘lateral gene transfer’
What are Shigella?
Very closely related to Escherichia (= “E. coli + a virulence plasmid”) Four species:
S. dysenteriae, S. flexneri, S. boydii, S. sonnei
What is shigellosis and what does it cause?
Pathology like Enteroinvasive Escherichia coli (EIEC) but with the addition of Shiga toxin
Severe bloody diarrhoea (esp. S. dysenteriae) Frequent passage of stools (>30/d)
pus and blood, prostrating cramps, pain in straining, fever Self-limiting (in adults)
What is the pathogenesis of shigella infection?
Acid tolerant
Person-to-person or contaminated water and food
Entry through colonic M cells* (antigen sampling)
Induced uptake
M cells: overlie lymphoid follicles and deliver antigens to underlying immune cells.
How does invasion of the colonic mucosa occur by Shigella?
Look at slide 21 of gram negative bacteria powerpoint
Shigella bacteria enter M cell from apical surface of the lumen causing apoptotic macrophage to enter the cell and destroy M cell.
More shigella bacteria enter enterocyte (intestinal epithelial cell) from basolateral surface after which causes translocating polymorphonuclear leukocytes to be recruited to site of infection causing further damage to epithelium.
What is the major Shigella virulence determinant?
Shiga toxin (Stx)
What is the structure of Shiga toxin
Think of one big circle in the middle which is A and 5 smaller cirlces around it which is B
A is the catalytic subunit (glycosidase) and B is the receptor binding subunits
It cleaves N-glycosidic bond of adenosine residue in 28S rRNA of the 60S subunit of the ribosome (depurination)
> blocks EF-1 and EF-2 binding > protein synthesis inhibited > cell death
What are the complications of systemic absorption of Stx?
Targets kidney > microvascular thrombosis in kidneys > kidney failure
What are the 2 species of salmonella?
Two species:
S. enterica - responsible for salmonellosis
> 2,500 serovars*
S. bongori - rare (contact with reptiles)
What are some infections caused by S. Enterica?
1.Gastroenteritis/enterocolitis (serovars Enteritidis and Typhimurium)
Frequent cause of food poisoning (milk, poultry meat & eggs) Second highest no. of food-related hospitalisations/deaths (UK) 6-36 hr incubation, resolves (~7 days)
Localised infection, only occasionally systemic
2.Enteric fever - typhoid/paratyphoid fever (serovars Typhi and Paratyphi)
3.Poor quality drinking water/poor sanitation
Systemic disease
~20 million cases, ~200,000 deaths/yr (globally)
Bacteraemia (serovars Cholerasuis and Dublin) Uncommon
What is the pathogenesis of salmonellosis?
Ingestion of contaminated food/water - high I.D. (~106) unless in chocolate!
(‘faecal-oral route’)
1.Invasion of gut epithelium (small intestine)
2.Transcytosed to basolateral membrane
3.Enters submucosal macrophages
4.Intracellular survival/replication
What is the pathogenesis of gastroenteritis?
1.Bacterial-mediated endocytosis into M cell
2.Induction of interleukin-8 release from enterocyte>
3.Neutrophil recruitment and migration>
4.Neutrophil-induced tissue injury>
5.Fluid and electrolyte loss > diarrhoea
6.Inflammation/necrosis of gut mucosa
Look at slide 26 gram negative bacteria
What is the pathogenesis of enteric fever?
S. enterica and sv. Typhil
1.Bacterial-mediated endocytosis into enteric cell
2.Transcytosis to basolateral membrane of M cell
3.Survival in Macrophage >systemic spread
4. Migration to reticuloendothelial organs via lymphatics & blood
Look at slide 26 gram negative bacteria
What is the pathogenesis of typhoid (enteric fever)? (Small intestine)
- Ingestion of S.Typhi via bacterial endocytosis >
- Enters small intestine >
- Inflammation and ulceration of Peyer’s patches >
- Diarrhoea; haemorrhage or perforation (1-3% of
cases)#
What is the pathogenesis of typhoid (enteric fever)? (In general)
- Ingestion of S.Typhi via bacterial endocytosis >
- Enters small intestine >
- Travels from SI in lymphatic system within macrophages to Mesenteric lymph nodes
- From mesenteric lymph nodes enters bloodstream via thoracic duct >
- Goes to transient (primary) bacteraemia
- Multiplication in macrophage in liver, spleen and bone marrow
7a. CAN travel from liver to gall bladder where it stays in a carrier state for 1 year to rest of life
7b. Or bacteria is released into bloodstream (secondary bacteraemia) > Fever, kidney and other organs infected
Asymptomatic until bacteria released into bloodstream as secondary bactereamia
Proteus mirabalilis (pathogenic Enterobacteria)
can differentiate into an elongated hyperflagellated form > surface motility (‘swarming’)
Have swimmer and swarmer cells
catheter-associated UTIs (~30% cases) > pyelonephritis
produces urease (causes urine pH to increase) > calcium phosphate precipitation >
formation of bladder/kidney stones, catheter blockage
Klebsiella pneumoniae
environmental
opportunistic, nosocomial infections (neonates, elderly, compromised)
colonisation of GIT (normal) and oropharynx (less frequent) is benign but can lead to:
UTI, pneumonia (aspiration from oropharynx), surgical wound infections, bacteraemia > sepsis (high mortality).
multi-drug resistant (resistant to carbapenems)
Enterobacter (E. cloacae, E. aerogenes)
opportunistic, nosocomial (originating in hospital) outbreaks (including ICUs)
infections in lungs, urinary tract, abdominal cavity, intravascular devices, sepsis
spread from endogenous gut flora, can survive on skin, patient-to-patient transmission
cephalosporin-resistant forms
Yersinia spp - primary pathogen
Y. enterocolitica - localised to ileum: gastroenteritis
(diarrhoea with abdominal pain and fever)
Y. pestis - systemic: bubonic plague (zoonosis) = ‘black death’
Vibrio cholerae
Facultative anaerobe
Saline environments: commensal to planktonic crustaceans such as copepods
>ingestion by shellfish
> contamination of drinking water due to flooding of coastal areas or poor sanitation
Curved rods with single polar flagellum
What is the most severe diarrhoeal disease?
Cholera
Characterised by pandemics (7 since 1817)
1P-6P, Indian subcontinent;
7P began in Sulawesi (1961) > SE Asia (1963) >
Africa (1970) > Latin America (1991) > Caribbean (2010)
O1 serotype > epidemics (and occasionally O139 variant)
1.4-4.0 million cases/yr, 20,000-140,000 associated deaths
What is the pathogenesis of infection by V. cholerae?
Faecal-oral route (not person-to-person) - high infective dose required
(faecal contaminated water (poor sanitation) or undercooked shellfish from risk areas)
>
Incubation, few hours to 5 days (multiplies in small intestine)
>
Voluminous watery stools (‘secretory’ diarrhoea)
What are the effects of cholera?
Can lose 20 litres fluid/day plus electrolytes
> dehydration/death (hypovolaemic shock)
> 50-60% mortality if untreated
No blood, pus or fever (i.e. not dysenteric)
i.e. no invasion or damage to mucosa#
Most cases can be treated with ORT
What is the major virulence determinant of V. cholerae?
Cholera toxin (CT)
1.CT binds to a glycolipid receptor on epithelial cell (B subunits)
2. A subunit ADP-ribosylates G-protein (Gs) > locked in ‘ON’ state
3. Uncontrolled cAMP (cyclic AMP) production
4. Protein kinase activated
5. CFTR ion transporter activity modified (loss of Cl- & Na+ into gut lumen)
6. Leads to massive loss of H2O
CFTR: cystic fibrosis transmembrane conductance regulator (a Cl- ion transporter)
Pseudomonas aeruginosa
motile - single polar flagellum
ubiquitous, free-living, aerobe
opportunistic (serious cause of nosocomial infections)
resistant to multiple antibiotics (& disinfectants) - very difficult to treat
Infections caused by P. aeruginosa
Acute infections:
Localised:
burn/surgical wounds UTI (catheters) keratitis
Systemic:
(bacteraemic > sepsis)
neutropenic patients (leukaemia, chemotherapy, AIDS)
ICU patients:
(ventilators)
leading cause of nosocomial pneumonia
Chronic infections:
(i) Cystic fibrosis (CF) patients
Common denominator to all infections - compromised host defences
What are the virulence determinants of P. aeruginosa?
Multiple toxins - these are the main virulence determinants in acute infections:
Interferes with cell signalling:
ExoS (exoenzyme S)
ExoU (exoenzyme U)
Cell death/ damage:
ToxA (exotoxin A)
LasB (elastase)
PlcH (phospholipase)
HCN (cyanide)
pyocycanin (generates reactive oxygen species)
Haemophilus influenzae
Exclusively human parasite:
Nasopharyngeal carriage in 25-80% population
Opportunistic infections seen mainly in young children and adult smokers:
Meningitis* (age <5 yrs), 5-10% of adult cases
Bronchopneumonia
Epiglottitis, sinusitis, otitis media
Bacteraemia (often associated with pharyngitis)
Pneumonia in CF, COPD, HIV patients
What are the diagnostic characteristics of H.influenzae?
Fastidious
- requires ‘factor X’ (haem) and ‘factor Y’ (NAD)
therefore, cultured on chocolate agar
non - motile
What are the virulence determinants of H.influenzae?
Capsule -
invasive strains are capsulate (‘encapsulated’)
> can penetrate nasopharyngeal epithelium
>resistance to phagocytosis and complement system
- 6 different capsule serotypes (a-f)
(type b strains are the main cause of meningitis)
‘Hib’ vaccine has reduced the incidence
Commensals and upper respiratory tract pathogens are non-capsulate (‘unencapsulated’)
referred to as ‘non-typeable’ H. influenzae (NTHi)
(ii) LPS (‘endotoxin’) > inflammation
>complement resistance
Legionella pneumophila
Legionnaires’ disease - severe inflammatory pneumonia
Immunocompromised
Severe
Infection from man-made aquatic environments
Replicate within freshwater protozoa - intracellular parasite of amoeba
Can survive and replicate within alveolar macrophages
What is Bordetella pertussis?
pERTUSSIS - Whooping cough
B. parapertussis causes mild pharyngitis)
Short rods (‘coccobacilli’)
Fastidious
Humans - only known reservoir
Highly contagious (low I.D.) - aerosol transmission
Non-specific flu-like symptoms (~7 d), followed by paroxysmal coughing
Non-invasive
What are the toxins of B. pertussis?
Pertussis toxin (PT)
S1 subunit ADP-ribosylates G-protein (Gi) > locked in ‘OFF’ state
(in ON state, Gi inhibits adenylate cyclase)
Therefore, PT > cAMP levels rise
Adenylate cyclase-haemolysin toxin (CyaA) > cAMP levels rise
What is pertussis toxin composed of?
PT is composed of 6 protein subunits.
Subunit S1 has ADP-ribosylase activity.
What does the hypersynthesis of cAMP lead to?
suppression of innate immune functions, particularly phagocytosis by macrophages
What are the 2 species of Neisseria which are of medical importance?
N. meningitidis
N. gonorrhoeae
Non-flagellated diplococci
Fastidious
Two species of medical importance
Humans are the only known reservoir
Diplococci present in Polymorphonuclear leukocytes of CSF (meningitis) or urethral
discharge (gonorrhoea) during
infection
What are the features of N. meningitidis? (meningococcus)
Nasopharyngeal carriage in 5-10% population (asymptomatic)
Rises to 20-90% during outbreaks
Person-to-person (aerosol) transmission (universities, barracks, Haj)
What is the pathogenesis of N. meningitidis?
crosses nasopharyngeal epithelium and enters bloodstream
>low level bacteraemia (asymptomatic) or septicaemia (sepsis)
>meningitis: invasion of the meninges - bacteria enter CSF of subarachnoid space after crossing blood-brain barrier (second most frequent cause of meningitis in young children)
very high mortality of septicaemic form if not treated
> requires rapid diagnosis!
What are the virulent determinants of N. meningitidis?
Capsule serogroup B - 90% cases)
> anti-phagocytic
(noncapsulated N.m. only found in nasopharynx - not pathogenic)
(ii) LPS (membrane ‘blebs’)
>cytokine cascade
> sepsis
What are the features of N. gonorrhoeae?
Gonorrhoea - second most common STD worldwide
Person-to-person only
Infection can be asymptomatic (~10% men, ~50% women)
Usually characterised by urethritis with additional infection of female genitalia.
Serious complications in women - can lead to salpingitis and/or PID if infection ascends.
Features of Campylobacter?
C. jejuni, C. coli
Spiral rods
Unipolar (monotrichous) or bipolar (lophotrichous)
flagella
Most common cause of food poisoning in UK & USA
Mild to severe diarrhoea, often with blood - usually self-limiting (< 1 week)
Campylobacter shed in faeces for ~3 weeks
Features of Helicobacter pylori
Spiral shaped, tuft of polar flagella
Present in ~50% global population, but only a fraction develop disease
Major role in gastritis and peptic ulcer disease (80-90% of ulcers)
Implicated in ~10% cases of gastric adenocarcinoma & mucosa-associated lymphoid tissue lymphoma (linked to production of VacA and CagA toxins)
What are (Phylum) Bacteroidetes?
Non-motile rods
Strict anaerobes
Commensal flora (large intestine) - most abundant
Opportunistic - tissue injury (surgery, perforated appendix or ulcer)
> predominantly peritoneal cavity infections (peritonitis, intraabdominal abscesses are most common) can lead to bacteraemia
Most frequent cause of anaerobic infections, usually B. fragilis (although it is only 0.5-1.0% of total commensal Bacteroides)
Features of chlamydia and chlamydophila?
Very small, non-motile.
Obligate intracellular parasites.
Cannot culture in bacteriological media - detect by serum Abs* or PCR.
What is the life cycle of Chlamydia?
- Elementary bodies (EBs) - rigid, extracellular form, ~0.3 micrometres, dormant
infectious
enter cell through endocytosis
prevent phagosome-lysosome fusion
Differentiates into >
2. Reticulate bodies (RBs) - fragile, intracellular form, ~1.0 micrometres, metabolically active
replicative
non-infectious
acquire nutrients from host cell
Life cycle of chlamydia (part 2)
- Entry by Elementary bodies
2.Conversion of elementary
body to reticulate body - Prevention of phagosome-lysosome fusion
- Multiplication
- Conversion of reticulate body to elementary body
- Cell lysis and release of elementary bodies
(including DNA condensation)
How many biovars does C. trachomatis have?
3
What are the types of C. trachomatis biovars?
- trachoma biovar (serotypes A-C) >trachoma > blindness
(eye-to-eye transmission via hands, fomites or flies) - genital tract biovar (serotypes D-K)
most common STD - infects epithelial cells of mucous membranes of urethra (both sexes) and vagina
- can ascend to uterus and ovaries (PID, infertility)
- usually asymptomatic (i.e. 70-80% cases in women)
-conjunctivitis (STD), hand-to-eye transmission - lympho granuloma venereum (LGV) biovar (serotypes L1-L3)
-causes LGV (an STD) - invasive urogenital or anorectal infection
- endemic to the tropics, cases rising in Europe/N. America
What are features of C. pneuominiae?
respiratory tract (mild or “walking” pneumonia)
- ~10% community acquired pneumonias
What are the features of C. psitacci?
Mainly birds
psittacosis (zoonotic infection), severe pneumonia
Features of spirochaetes
Long, slender, helical, highly flexible
Most are free-living and non-pathogenic
Pathogenic varieties difficult to culture
Modified outer membrane (“outer sheath”)
(Treponema and Borrelia lack LPS, replaced by a different glycolipid)
What is the spirochaete endoflagellum?
Endoflagella located between
peptidoglycan and outer membrane.
Fixed at each end of the bacterium and confers shape.
Overlap in the centre of the bacterium.
Propels bacterium in a corkscrew motion
- swim faster in high viscosity medium
- hides” antigenic flagellum
What is Borrelia burgdorferi?
Lyme disease (zoonosis) (~300 cases in UK)
- bull’s eye rash, flu-like symptoms (fever, fatigue, headache)
- dissemination via lymphatics/blood to other organ (neurological problems in 10-15% patients, joints >arthritis)
What are the features of Leptospira interrogans?
contact of infected animal urine with mucous membrane or abraded skin
- flu-like symptoms
- severe form (Weil’s disease) in 10-15% infected individuals (jaundice, acute renal and hepatic failure, pulmonary distress,
haemorrhage)
What can treponema pallidum lead to?
Syphilis
1.primary stage - localised genital infection (ulcer (“chancre”))
days-weeks post-infection
(highly transmissible phase)
- secondary stage (~50% cases) - systemic
(skin (rash), swollen lymph nodes, joint pains,
muscle aches, headache, fever)
1-3 months post-infection
(still highly transmissible) - tertiary stage (~30% cases) - ‘gummas’ (granulomas) in bone and soft tissue,
cardiovascular syphilis (aorta),
neurosyphilis (brain and spinal cord).
Occurs several years post-infection
(non-infectious form)
What are the four major groups (phyla) of Gram- negative pathogens?
- Proteobacteria - all are rod-shaped (bacilli)* except Neisseria (diplococci)
and the Campylobacter/Helicobacter genera (spiral) - Bacteroidetes (Bacteroides) - rod-shaped (bacilli)
- Chlamydia - round (elementary bodies), pleiomorphic (reticulate bodies)
- Spirochaetae (Spirochaetes) - spiral/helical
*= some are curved, some are short (coccobacilli), but are bacilli nonetheless
What happens in bacterial respiration?
Gut pathogens must be able to survive in low oxygen environments and are
therefore usually facultative or obligate anaerobes.
For example, Bacteroides are obligate anaerobes, while members of the
Enterobacteriaceae and V. cholerae are facultative anaerobes.
Campylobacter/Helicobacter genera are microaerophiles.
Most of the other pathogenic species discussed are aerobic.
Define strain/ isolate
Refers to the name given to a bacterium that has been isolated from a patient to distinguish it from bacteria of the same species that were obtained from other patients.
Although the same species, the genome sequences are invariably different among strains due to random mutation/gene loss or acquisition of new genes. However, in an outbreak situation, for example in a hospital ward, you may expect bacteria isolated from different patients to have identical genome sequences
What is a serovar/ serotype?
are strains that have antigenic properties that differ from other strains of the same species.
What is a pathovar/ pathotype?
are strains that are distinguished by possession of particular pathogenic mechanisms rather than being based on antigenic differences between molecules on the bacterial cell surface (the serotype). However, different serovars may exist among members of each pathovar
What are biovars/ biotypes?
are variant bacterial strains that differ physiologically or biochemically from other strains in a particular species.
What does the M in M. tuberculosis stand for?
Mycobacterium
What is M. leprae?
Leprosy
Associated with deformity or loss of fingers
Tropical illness
Climate is warm
Illness of poverty and lack of medical infrastructure
Skin lesions causes issue – hypopigmented – can be destructive at the nose
Nervous lesions causes problems – damages transition of neural signals – causes loss of sensation – affects distal parts of body like hand and foot – don’t have pain response if you injure there
How many deaths have been caused by tuberculosis in the last 200 years?
1,000,000,000
How many deaths caused TB per year?
1.5m
How many new cases of TB do we get a year?
10.4m new cases per year
Mycobacteria doesnt stain well with what?
Gram staining
What does mycobacteria stain with?
Ziehl-Neelsen/ Acid fast positive
What are the features of Mycobacteria?
Slighly curved, beaded bacili
High lipid content with mycolic acids in cell wall makes Mycobacteria resistant to Gram stain
Ziehl-Neelsen stain
Need 10,000 bacili per ml sputum to diagnose
What is the microbiology of TB?
Aerobic, non spore forming, non motile bacillus
Cell wall: high molecular,weight lipids +++.
Slow growing – M tuberculosis generation time 15-20hr vs 1hr for common bacterial pathogens
Significant part of the 4.4 Mb genome of MTB encode genes involved in lipogensis and lipolysis.
Key components include:
- Mycolic acids
- Lipoarabinomannan
What are the challenges of mycobacterium tuberculosis?
Thick lipid rich cell wall making immune cell killing and penetration of drugs challenging
Slow growth
- Gradual onset of disease
– Takes much longer to diagnose
– Takes longer to treat
How can you catch TB?
Catch TB from someones lungs when theyre coughing?
What happens in primary tuberculosis?
-Initial ‘contact’ made by alveolar macrophages in lungs
- Bacilli taken in lymphatics to hilar lymph nodes
What happens in latent tuberculosis?
-Cell mediated immune response (CMI) from T cells
- Primary infection contained but CMI persists
- Latent TB;
. no clinical disease
* detectable CMI to TB on
tuberculin skin test
What happens in Pulmonary tuberculosis?
- Granulomas forms around bacilli that have settled in apex
- In apex of lungs there is more air and less blood suppy (fewer defending white cells to fight)
- Could occur immediately following
primary infection (post-primary)
or after later reactivation - necrosis results in abscess forming and
caseous material coughed up
What happens in Pulmonary tuberculosis?
- Granulomas forms around bacilli that have settled in apex
- In apex of lungs there is more air and less blood suppy (fewer defending white cells to fight)
- Could occur immediately following
primary infection (post-primary)
or after later reactivation - necrosis results in abscess forming and
caseous material coughed up - TB may spread in lung causing other lesions
- Granuloma + Lymphatics + Lymph nodes = Primary Complex
Causes cough, chest pain, fever
How can TB spread beyond the lungs?
TB meningitis
Miliary TB
Pleural TB
Bone and joint TB
Genito urinary TB
Bacili in lymph nodes
Bacili in lung
How does TB infect us?
- Aerosol transmission
- Primary TB in lung
- Latent TB can remain for decades
- Can spread beyond lungs
What is the immunology to mycobacteria?
- Mycobacteria are phagocytosed by macrophages and traffic to a phagolysosomes/ phagosome.
- The bacterium has adapted to the intracellular environment and aims to withstand phagolysosomal killing and escape to the cytosol
- M. tb eradication can happen by apoptosis mediated cell death, auto phagocytosome, by phagolysosome
- Effective immunity requires CD4 T-cells which generate interferon gamma and
this helps activate intracellular killing by macrophages.
What happens if a granuloma is formed?
Mycobacteria shut down metabolically in order to survive – dormancy
What happens if hallmark granuloma formation fails?
if it fails, e.g. in the lung, this can result in the formation of a cavity full of live
mycobacteria and eventual disseminated disease (consumption)
What does our body do to protect us?
- Primarily controlled by macrophages
- Requires a CD4 T cell response to be controlled
- Involves many cells of immunity - formulation of granulomas
- ## Granuloma stability controls reactivation of TB
How do we get a more rapid diagnosis of tuberculosis?
- GeneXpert MTB/RIF cartridge based test
- Nucleic acid amplification test using PCR
- Purifies and concentrates MTB, sonicates to
release genomic material and then performs
PCR - Rapid result for MTB and detects Rifampicin
resistance using fluorescence - Can detect 131 bacilli/ml
- Sensitivity 88%, Specificity 98%
- Recommended for rapid diagnosis in TB
endemic countries
What does the highly immunogenic nature of mycobacterial lipids do?
Stimulates T-cell responses in 3-9 weeks after exposure to M. tuberculosis
Positive effects: ²+ve effects; macrophage killing of mycobacteria
,containment of infection, formation of tissue
granulomata.
Negative effects: ; hypersensitivity reactions with skin lesions, eye lesions and swelling of joints
This reactivity is measured in the tuberculin skin test where intradermal injection of purified protein derivatives induces skin swelling and redness.
What are some standard therapy anti - TB drugs?
isoniazid (INH), rifampicin (RIF),
pyrazinamide (PZA) and ethambutol (ETH) x 2 months
followed by isoniazid and rifampicin for further 4 months
What are some second line treatments?
injectable agents (streptomycin,
cycloserine, capreomycin)
* Side effects are wide-ranging and severe, include liver
damage
What are some forms of resistance mechanisms?
Drug inactivation:
Mtb produces beta-lactamase
Drug titration:
Target overproduction
Alteration of a drug target:
- missense mutations
Altered cell envelope:
- Increased permeability and drug efflux
What are some multi drug reisistant TBs?
XDR-TB: resistant to four commonly used TB drugs. 6% of all TB cases
Resultant from inadequate TB therapy and failure to clear patients of bacteria
Treatment is lengthy and expensive
How can we treat XDR-TB?
BPaL regimen :
– Bedaquiline
– Pretomanid
– Linezolid
All oral treatments for 6 months
These can fail too with totally drug
resistant (TTR) TB. No known solution as
yet.
How do viruses cause disease?
Direct destruction of host cells
Modification of host cell
“Over-reactivity” of immune system
Damage through cell proliferation
Evasion of host defences
Why are viruses of global importance?
Lower respiratory infections, diarrheal diseases and HIV/AIDS are leading killers in lower income countries in 2016
40 MILLION with HIV - 8 MIL DONT know their status, 750 undiagnosed
What is MERS?
Middle Eastern Respiratory syndrome
MERS 2012-now
2578 cases
888 deaths (35%)
What viruses can cause miscarriage and birth defects?
Cytomegalovirus (CMV),Varicella Zoster Virus (VZV), Herpes Simplex Virus (HSV),
Rubella
What are some recent viral outbreaks?
. Influenza, Measles, Mumps, Norovirus, Covid
What are some cancer causing viruses?
Epstein Barr Virus (EBV) + lymphoma,
Hep B/C + hepatocellular carcinoma,
Human Papilloma Virus (HPV) + cervical/anal cancer,
HIV
What is a virus?
An infectious, obligate intracellular parasite comprising genetic material (DNA or RNA) surrounded by a protein coat/ and or membrane
All have a receptor binding protein to “dock” to cells
All contain genetic material
Virus has poisonous fluid
Comprises of genrtic materia (DNA or RNA) surround by protein coat and/ or membrane
What do we mean by obligate intracellular?
totally dependant on living cells for their replication and existance
What are the different shapes of viruses?
Helical
Icosahedral
Complex
Non-enveloped
-adenovirus, parvovirus
Enveloped
- influenza, HIV
When not in an infected cell what do viruses exist as?
Virions
Consist of:
Genetic material (DNA or RNA)
Protein coat (capsid)
What do human viruses range from?
Human viruses range in size from 20 to 260nm in diameter
What do bacteria have that viruses dont?
Cell walls
Organelles
DNA and RNA
How do viruses replicate?
- Attachment to specific receptors
- Cell entry - Uncoating of virion within cell
- HOST CELL INTERACTION + REPLICATION
- Migration of genome tocell nucleus
- Transcription to mRNA using host materials
- Use cell materials (enzymes, amino acids, nucleotides) for their replication; subvert host cell defence mechanisms.
- Translation of viral mRNA to produce:
- structural proteins, viral genome, non-structural proteins eg enzymes - Assembly of virion
- Release of new virus particles
a.Bursts out > cell death eg rhinovirus
b. budding/exocytosis
e.g. HIV, influenza
What is a virus completely dependent on?
A virus is completely dependent on its host cell’s machinery to exist and replicate, and enters a cell with only its genome +/- viral enzymes
How do viruses cause disease?
- Direct destruction of host cells
- Modification of host cells
- “Over-reactivity” of immune system
- Damage through cell proliferation
- Evasion of host defences
What happens in direct destruction of host cells?
host cell lysis and death after a viral replication period of 4 hours
e.g poliovirus
Polio affects neurons – gets into neurones – destroys neurones – causes weakness and paralysis
What happens in modification of host cell?
E.g. Rotavirus
>atrophies villi and flattens epithelial cells
>decreases small intestine surface area
>nutrients incl sugar not absorbed
>hyperosmotic state
>profuse diarrhoea
What happens in the over reactivity of the immune system?
e.g. Hep B
Infects liver cells and hepatocytes
T lymphocytes comes along and destroys stuff
What happens in damage through cell proliferation?
e.g Human papillovirus (HPV) > Cervical cancer
1. Acquisition through contact
2. Partial viral replication and expression of some HPV proteins
3. Viral DNA integrated into host chromosome
4. Continuous expression of oncoproteins causing cellular DNA mutations
5. Dysplasia and neoplasia >
6. Leads to Cell proliferation and local and metastatic spread
What happens in HPV (detailed)
1.HPV (types 16 or 18) infection of suprabasal layer in genital tract.
2.Partial viral replication including transcription and expression of several early
viral gene products (E1, E2, E4, E5).
3.Gradual movement of cells to mucosal surface through natural wear and tear.
4.At some point during this process the HPV genome may become integrated into
the host cell chromosome. Mutagenic agents (eg nicotine) may increase the
chance of integration.
5.Following integration, control of viral gene expression by the HPV E2 protein
is lost and the HPV E6 and E7 proteins may be expressed.
6.Two cell growth and proliferation suppressor proteins, Rb (retinoblastoma)
and p53 are prevented from operating.
7.Excessive cell growth and proliferation occurs and cervical cell carcinoma
results.
What happens in evasion of host defences?
After primary disease, virus not detectable but viral DNA lies latent and can reactivate, particularly at times of lower immune control (illness/immunosuppression/advancing age)
Can hide in nerve root ganglion, Lymphoid cells, Myeloid cells
Lymphoid cells - T cells, B cells, NK cells
Myeloid cells - monocytes, macrophages, neutrophils, basophils, eosinophils etc
What is Varicella Zoster Virus?
Primary - chickenpox
Reactivation - when immune system suppressed - Shingles
What is cell to cell spread in evasion of host defences?
E.g. Measles and HIV
Direct cell to cell spread has multiple advantages
Avoids random release into the environment
Increased speed of spread within tissues
Avoiding immune system
What happens in the evasion of host defences at the molecular level?
Antigenic variability eg influenza, HIV, rhinovirus
Ability to change the surface antigens in order to evade the host’s immune system
Explains:
- how a host can be re-infected with the same virus e.g. common cold
- why with influenza vaccination is required annually, as each season a different viral strain is circulating
What is prevention of host cell apoptosis?
In response to a viral infection, many cells undergo apoptosis, which reduces the amount of virus released from the cell
Prevention of host cell apoptosis allows the virus to continue replicating within it, so more virus is produced and then released. It also has an essential role into how some viruses are oncogenic (cancer causing).
Flu can recombine itself in different configurations
What is Downregulation of interferon and other intracellular host defence proteins ?
In normal cells:
Interferon synthesis stimulated > Antiviral state activated in neighbouring cells
In affected cells:
Interferon synthesis blocked > Neighbouring cell susceptible to infection
Why do viruses Viruses vary wildly in the range of clinical syndromes they can cause?
> Different host cells and tissues that they can infect
> Different methods of interaction with the host cell
What are the 5 types of immunoglobulins?
IgG
IgM
IgE
IgD
IgA
What happens in active immunity?
Cell-mediated immunity
Antibody-mediated immunity
Vaccination stimulates immune response and memory to a specific antigen/infection
Where IgA produced?
Breastmilk
What happens in Passive immunity?
Protection provided from the transfer of antibodies from immune individuals.
Most commonly cross-placental transfer of antibodies from mother to child (e.g. measles, pertussis)
Or, via transfusion of blood or blood products including immunoglobulin (e.g. Hep B)
Protection is temporary – usually only a few weeks or months.
#Immunity from someone else or a different source
First couple of months baby immunised using mothers breast milk antibodies
Antibodies degrade overtime
Active immunity is different
Active immunity gives you longer lasting immunity
What can vaccines be made from?
inactivated (killed) (e.g. pertussis, inactivated polio)
attenuated live organisms (e.g. yellow fever, MMR, polio, BCG)
secreted products (e.g. tetanus, diphtheria toxoids)
the constituents of cell walls/subunits (e.g. Hep B) or
recombinant components (experimental)
What is herd (population) immunity?
Get population immunity through vaccination or past infection
What is vaccine failure?
No vaccine offers 100% protection
Small proportion of individuals get infected despite vaccination.
Primary vaccine failure – person doesn’t develop immunity from vaccine.
Secondary vaccine failure – initially responds but protection wanes over time.
No such thing as perfect vaccine
Some people don’t have enough immune response
Vaccine expired, don’t get administered the proper way
What are examples of vaccine preventable diseases?
Diptheria
Tetany/ Tetanus
Pertussis/ Whooping cough
Polio
Haemophilus influenzae type B
Meningococcal disease
What is contact tracing?
Contact for meningitis is taken to be any person having close contact with a case in the past 7 days
Close contact includes kissing, sleeping with, spending the night together or spending in excess of eight hours in the same room
Notifiable diseases - scary
Anthrax
Cholera
Plague
Rabies
SARS
Smallpox
Viral haemorrhagic fever
Yellow fever
Notifiable diseases - nasty
Acute encephalitis
Botulism
Brucellosis
Enteric fever (Typhoid & Paratyphoid fever)
Haemolytic Uraemic Syndrome
Notifiable diseases - infectious but vaccine preventable
Acute poliomyelitis
Diphtheria
Measles
Mumps
Rubella
Tetanus
Whooping cough
Acute Meningitis / Meningococcal septicaemia
Notifiable diseases - diseases that need specific control measures
Acute infectious hepatitis
Foodborne
Food poisoning
Botulism
Enteric fevers
Infectious bloody diarrhoea
Scarlet fever
Tuberculosis
What is the role of surveillance in notifying disease?
Detection of any changes in a disease
>Outbreak detection
>Early warning
>Forecasting
Track changes in disease
>Extent and severity of disease
>Risk factors
Allows development of interventions targeted at vulnerable groups
How can we protect our community from diseases?
Investigate: contact tracing, partner notification, lookback exercises, etc…
Identify and protect vulnerable persons: e.g. chemoprophylaxis, immunisation, isolation
Exclude high risk persons or from high risk settings
Educate, inform, raise awareness, health promotion
Coordinate multi-agency responses
What is the route of transmission for disease?
Source > Pathway > Receptor
What are protazoa?
“One celled animals”
Single cell with nucleus
(Eukarytoic)
> 30,000 species
Show characteristics usually associated with animals Eg mobility
What are the 5 major groups of protazoa?
Flaggelates
Amoebae
Sporozoan
Cilliates
Microsporidia
What are some examples of flagellates?
Trypanosoma spp
Leishmania spp
Trichomonas vaginalis
Giardia lamblia
What is African Trypanosomiasis (sleeping sickness)?
Tsetse fly bite
Chancre
Flu like symptoms
CNS involvement -(sleepy, confusion, personality change)
Coma and death
Diagnosed on blood film or CSF
What is American Trypanosomiasis?
Spread by Triatomine Bug
Acute:Flu like symptoms
Chronic:
Cardiomyopathy
Megaoesophagus
Megacolon
Spread through contact with faeces of triatomine “kissing” bug
Also through blood, vertically and eating contaminated food
Millions have undiagnosed Chagas in the Americas
Two phases, both can be asymptomatic or life threatening
Leishmaniasis Leishmania spp
Spread by the bite of the sandfly
> 20 species affect humans
Three clinical pictures:
Cutaneous
Mucocutaneous
Visceral
What is trichomonas vaginalis>
Sexually transmitted
Asymptomatic
Dysuria
Yellow frothy discharge
Treated with Metronidazole
Seen on wet prep of high vaginal swab
STI – endemic to UK
Take swab of vagina and put it on a plate with water
What is giardasis?
Faeco-oral spread
Diarrhoea
Cramps, bloating, flatulence
Recent travel, childcare
Trophozoites/cysts seen in stool
Treated with metronidazole
What is amoebiasis?
Faeco-oral spread
Dysentry, Colitis,Liver and lung abscesses
Trophozoites/cysts seen in stool
Treated with metronidazole
Poor sanitary conditions/tropical countries
MSM (men who have sex with men) at risk
What are examples of Amoebae?
Entamoeba histolytica
Examples of sporozoa?
Entamoeba histolytica
What is Cryptosporidiosis?
Waterborne
Diarrhoea (Watery, no blood)
Vomiting, fever, weight loss
Oocytes seen in stool (acid fast!)
Usually self limiting
Severe disease in immunocompromised
Poor sanitary conditions/tropical countries
MSM
Children
Swimmers
Contaminated recreational water sources
What is Toxoplasmosis/Toxoplasma gondii?
Ingestion of contaminated foodand water/feline faeces
Can cause:
- Disseminated disease
- Toxoplasma Encephalitis
- Chorioretinitis
Acute maternal infection can be devastating in pregnancy
Up to half of the UK population will have the infection at some point.
Occurs worldwide (Except Antarctica).
Acquired through the ingestion of oocytes by contact with cat faeces (having kittens at home, contaminated soil or water), or ingestion of bradyzoites (tissue cysts) in contaminated food (eg rare beef/lamb, shellfish, unpasteurised goats milk)
Cats are the only definitive hosts (full cycle occurs, ingestion to excretion)
Monday: 28 Female attends GP
Fit and well
Complaining of….
Fevers
Abdo discomfort
Myalgia
At GP examination: Tachycardic
Pyrexial (38.7C)
Generalised abdotenderness
Urine dip: blood and leucocytes
Said it was UTI
Then Friday: Went back to GP
No improvement
Ongoing fevers
Antibiotics switched
Monday: Went back to GP
Ongoing fevers
Dehydrated
Dark brown urine
Sent to A&E…
History and examination
Pretty thorough!
Asked about ketamine
BUT
Didn’t ask about travel….
Bloods:
Mild anaemia
Thrombocytopenia
Acute kidney injury
Derranged LFTs
9 months earlier:
Travel to DRC and Ethiopia!
Had pre-travel vaccines &took anti-malarials
Relevant?
Travel with Malaria
What mosquito is malaria transmitted by?
female anopheles mosquito
What are the 5 species of malaria that can cause disease in humans?
Plasmodium falciparum
Plasmodium ovale
Plasmodium vivax
Plasmodium malariae
Plasmodium knowlesi
Which malaria tends to cause most disease?
Falciparium tends to cause the most severe disease and mortality untreated is very high
Early identification and treatment is key to prevent death
Presentation tends to occur within a month for falcip
Vivax and ovale can be up to a year!
Falciparum most prevalent and can kill
Rest don’t cause death but can cause illness
How can we diagnose malaria?
Light microscopy
A blood sample is taken for “thick and thin” films
The thick film is sensitive but low resolution> does this person have malaria? Yes or no
The thin film helps identify the species as each looks different, and also is used to work out the parasitaemia percentage > what species of malaria does this person have
What are the symptoms of malaria?
FEVER
Chills
Headache
Myalgia
Fatigue
Diarrhoea
Vomiting
Abdo pain
What are the signs of malaria?
Anaemia
Jaundice
Hepatosplenomegaly - big spleen and liver
‘Black Water Fever’ - occurs from haemolysis
What is the lifecycle of malaria?
- Within the mosquito the gametocytes are within the midgut, and undergo development, end up as sporozoites in the salivary glands of the mosquito
- Mosquito takes its next blood meal and injects the sporozoites into the human
- Sporozoites injected into blood infect the hepatocytes in the liver (Abdominal pain)
- Develops into a schizont, which then bursts and infects red blood cells (Abdominal pain)
- Within the red blood cell, plasmodium becomes a trophozoite -This is one of the forms we can see on a blood film to diagnose malaria
- The trophozoite develops into a schizont, which then ruptures and re-infects another RBC
This blood stage lasts 48 hours in most of the plasmodium species:
>Cyclical fever when RBCs rupture
>Haemolysis
>Anaemia
>Jaundice from bilirubinaemia - Some of the trophozoites develop into gametocytes
- Whhich are then taken up by another mosquito
What can P. falciparum cause?
OBSTRUCTED MICROCIRCULATION
which can lead to “COMPLICATED MALARIA”
The pathophysiology of complicated malaria is mostly down to the infected RBCs ability to adhere to endothelial cells
What happens to RBC’s infected with p.falcip?
They have proteinaceous knobs on the surface that bind to endothelial cells in the vessels and other RBCs
This can cause small vessels to become obstructed by clumps of red blood cells causing hypoxia of the tissues, microinfarcts in brain and lung
What can happen in the brain for those with P. falcip malaria?
Vascular occlusion> Drowsy, Increased Intercranial pressure, seizures, coma
Hypoglycaemia > drowsiness
How can the lungs be affected by vascular occlusion (caused by P.falcip)?
Anaemia and lactic acidosis can cause a fast respiratory rate (tachypnoea) through compensatory mechanisms.
Increased vascular permeability causes fluid to leak directly into the lungs causing pulmonary oedema.
How can ARDS (acute respiratory distress syndrome) be caused by P falcip?
Shortness of breath due to >
due to anaemia
due to lactic acidosis (compensatory)
due to increased vascular permeability pulmonary oedema
How can renal failure occur due to P.falcip?
A direct effect through vascular occlusion
Hypoperfusion secondary to dehydration (fever) or hypotension
Haemolysis creating products that can be nephrotoxic
Vascular occlusion
Dehydration
Hypotension
Haemoglobulinuria
Haemolysis
These cause:
Proteinuria
Fatigue
Haematuria
How does thrombocytopenia occur due to P. falcip?
Thrombocytopenia occurs through platelet aggregation and therefore reduced circulating platelets
Generalised inflammation activates the coagulation cascade, causing clotting factors to be used up.
This then causes DIC (disseminated intravascular coagulation) – lots of micro clots have formed in the blood, but a lack of circulating clotting factors causes bleeding
This causes:
Epistaxis
Abnormal bleeding
Worsening anaemia
Why can a patient develop shock with malaria?
The heart and circulatory system are affected in lots of ways.
The patient can develop shock due to:
Hypovolaemia: bleeding or vasodilation (inflammatory cascade and sepsis)
Anaemia > inadequate oxygenation to heart
Why do people with malaria often go into shock?
-Pro-inflammatory cascade causing vasodilation
-Anaemia (if bad enough) can cause cardiogenic shock
-Gram negative sepsis can occur from increased vascular permeability in the bowel > so gram negative bacteria colonising the bowel can enter the bloodstream
-Bleeding causes a hypovolaemic shock
-Increased vascular permeability means that intravascular fluid can leak out into the “third space”
How is thrombocytopenia caused by malaria?
Adherence to endothelial cells by the parasitised RBC causes a local expression of cell-adhesion molecules causing red blood cells and platelets to bind causing thrombocytopenia through platelets use, and occlusion of small vessels.
How is hypoglycaemia caused by malaria?
There is a proinflammatory cytokine cascade which:
alters the endothelial cell wall function > Leaky vessels > ARDS, gram –ve sepsis, shock
Switches cells to anaerobic metabolism > hypoxia and lactate increases
Reduces gluconeogenesis > hypoglycaemia
Complicated malaria causes…
Cerebral
ARDS/Pulmonary oedema
Renal failure
Sepsis
Bleeding/Anaemia
What is the treatment for malaria?
Complicated
- IV artesunate
- (IV quinine + doxycycline)
Uncomplicated
- Lots of options!
What are the supportive measures for complicated malaria?
Cerebral: antiepileptics
ARDS: oxygen, diuretics, ventilation
Renal failure: fluids, dialysis
Sepsis: broad spectrum antibiotics
Bleeding/Anaemia: blood products
Exchange transfusion if huge parasite burden
How can you relapse from malaria?
Vivax and ovale species can develop hypnozoites in the liver which can “reactivate”. Ie once a patient recovers from the first episode of malaria, they can suffer relapses of malaria months or years later.
These can lie dormant for years!!
They aren’t treated by the treatment for acute malaria
Require primiquine to kill the hypnozoites and therefore stop a relapse occurring
Primiquine can cause haemolysis if deficient in G6PD
What are the features of the Varicella Zoster virus (VZV)?
One virus – two diseases
Varicella “chickenpox” – PRIMARY INFECTION
Herpes Zoster (HZ) “Shingles” – SECONDARY REACTIVATION
Glycoprotein spikes
Lipid envelope
Double-stranded DNA genome
Nucleocapsid
Tegument
What is the incubation period?
When you know you dont have the virus
When is the risk of transmission the highest with chicken pox?
Risk of transmission is highest to other is 48 hours before you get the spot and first couple of day after the spots emerge
Where does chickenpox replicate in?
Chickenpox replicates in nasopharynx and is in nasal secretions – breathe cough and sneeze so you pass on to others
If someone touches vesicles (spots) get it on your own skin
What are the features of chickenpox?
Common in childhood
Highly contagious,
Usually benign but can be serious in certain groups e.g.
-Immunocompromised and patients who have had transplants
-Adults
-Pregnant women
-Smokers
-Infants
-90% of adults raised in the UK have had chickenpox
What happens with chicken pox rashes?
Macule to papule to vesicle to pustule to crust
Initially just a blob
Rash evolves to papule – run your finger over lesion you feel it
Papule becomes blister filled with blister fluid which is infectious
WBC response – rush to vesicle carrying immune response with them and become pustule – pus filled
Then become crust
Chicken pox likes to divide in warm areas – spares cooler areas
What is centrifugal distribution of a rash?
Smallpox lesions all evolve at the same
time whereas with chickenpox and
measles, lesions at different stages of
progression can appear on the body
concurrently
What important chickenpox information is there to collect?
Consider:
Age of the patient
Onset of rash
Any contacts?
Immuno-suppressed?
Pregnant?
How do we confirm the diagnosis of chicken pox?
-Pop lesion with a sterile needle
(Don’t wipe it with alcohol swab first)
-Absorb vesicle contents onto swab
-Replace swab in cassette and send for VZV/HSV PCR
What are the complications of chicken pox?
-Dehydration
-Haemorrhagic change
-Cerebellar ataxia (common) – unsteady gait – common in children
-Encephalitis
-Varicella pneumonia
.Bacterial empyema
-Skin and soft tissue infection typically with group A strep
.Bone and joint infections: deep sepsis-osteomyelitis/pyomyositis
-Congenital (foetal) varicella syndrome
Seizures
Confusion
Lose motor function
More in adults then children
What is chickenpox pneumonitis?
Chickenpox pneumonitis affects 15% of healthy adults
Risk doubles if underlying lung disease or if a smoker
30% mortality untreated
Mortality 6% even with adequate treatment
What antivirals should we use to treat chickenpox pneumonitis?
Acyclovir
What is a bad sign on a chest x-ray?
Chest x-ray should be black - patchy white shadowing is bad
What is foetal varicella syndrome? (FVS)
Foetal infection occurs in 10-15% of cases of chickenpox in pregnancy
-Usually transient and asymptomatic
-If any manifestations – shingles in the first year of life
-If maternal chickenpox occurs in the
first half of pregnancy, about 2% of infants will develop FVS
What are the potentially severe defects of FVS?
Cicatricial skin scarring
Limb hypoplasia –poorly developed limb
Visceral and ocular lesions
Microcephaly and growth retardation
Right leg lost musculature of calf, no toes, not well defined ankles
Why are archived serum samples useful?
Blood sample
Tube contains gel to separate cells from serum once centrifuged
Small sarsted tubes kept for about 3 years
Take clotted sample from all pregnant women when they book first appointment
Centrifuge sample – serum separation gel risen up and formed barrier between clot at bottom and serum at top – stored for 3 or 4 years
Describe an IgM molecule
IGM molecule – 5 pronged molecule – 10 binding site – first mode of defence from our b cells
IGM response can be non specific – don’t use igm very often to diagnose virus
Describe an IgG molecule
memory antibody – tight bond between antigen and antibody – nearly unbreakable – prodcues memory immune response and defence against that pathogen when its seen again – opposite to igm
What happens in viral dormancy?
Primary infection - widespread chickenpox > Viral dormancy in dorsal root or cerebral ganglion > Localised reactivation – shingles
DNA viruses might have dormant phase
Hepatitis B – good eg – sits in liver cells –evades a lot of immune responses – hard to get rid of once established
Herpes similar –
Single sensory nerve supplies single patch of skin – because its on either side – shingles rash wont migrate across the midline – stays unilateral
Stats of shingles
Most common in the elderly
250,000 people estimated to be affected
annually in England and Wales
Around 1/1000 people over seventy who
have shingles will die of the infection
What is the location of reactivation of shingles/
Thoracic region most commonly involved (50-70% of all cases)
Cervical, lumbar and sacral dermatomes are less frequently involved
10-20% of cases are ophthalmic
-Affecting the ophthalmic division of the trigeminal nerve
Ocular shingles affects the eye
Shingles is painful
Have to use drugs that dampen down the abnormal nerve responses in nerve endings
What is hutchinsons sign?
Lesion at the end of the nose
– nasociliary branch of the ophthalmic division of the trigeminal nerve supplies skin to tip of nose – sign that you have corneal involvement – could affect eye
What happens if a child with eczema gets chicken pox or shingles?
lose fluid into those lesions – lose a loot of blood volume into lesions of the skin – could be life threatening – eczema hepatica
What can cause vasculitis rash with arthritis?
Infections:
Parvovirus B19
Rubella
Hepatitis B
Neisseria gonorrhoeae
Neisseria meningitidis
Streptococci
Spirochaetes
Borrelia burgdorferi
What are enteroviruses?
Hand foot and mouth disease?
Secondary syphillus can do this
Diagnose this just the same way
Enteroviruses get shed in their stool
Can cause viral meningitis or myocarditis
What does Parovirus B19 do?
Not the same thing that affects dog
Targets immature red cells – red cells in marrow has nucleus present display on surface p antigen – parvovirus attaches to those and it prevents normal red cell maturation – red cells die but cant be replaced from marrow
Worry about kids and pregnant women
What are complications of Parovirus?
Marrow with nuclear material around red cell – shows how the red cell formation occurs and how immature red cells an be targeted by that virus
What is Hydrops fetalis?
Can happen through an intrauterine fusion (from IV bag) which can cause a fetus to receive a blood transfusion through the umbilical vein in the placenta
What does the Parovirus rash look like?
Looks reticular in body
Tree like
Vesicular rashes
HSV1 can be both mouth and down below
HSV 2 is just down below
Whatis mucositis?
Shedding of epithelium from inside of mouth – that becomes moist area where bugs can target and get into the body
What is cytomegalovirus? (CMV)
Infection
causes glandular fever type initially
Teens and twenty year olds
Most people have had it once theyre 60
Lymphocytes become a bit destructive and red cells stick to them
What happens in CMV reactivation?
In immunosuppressed patients its common to reactivate
HIV patients retinitis is a problem
Bottom left retina has lots of CMV - pizza pan retina because it looks like cheese
Top right histological hallmark of CMV – lots of nucleus
What happens in measles?
Transmissible
R nought for measles is 16
Real profound rash – becomes dry and desquamating
Measles makes kid miserable
What is an antibiotic?
Agents produced by micro-organisms that kill or inhibit the growth of other micro-organisms in high -dilution
Not necessarily an anti bacterial
Most agents currently used are semi-synthetic derivatives of antibiotics so more correctly termed ‘antimicrobials’.
Antimicrobials include
antifungal, antibacterial, antihelminthic,
antiprotozoal and antiviral agents
BUT, in practice
antibiotic = antibacterial
Penicillin discovered by Alexander Fleming in 1929 was the the first antibiotic.
Better term that kill infectious pathogens is antimicrobials
Some agents are antibacterial and antiprotozoal and antifungal – theyre all 3
How do antibiotics work?
Antibiotics are molecules that work by binding a target site on a bacteria
-defined as points of biochemical reaction crucial to the survival of the bacterium
-the crucial binding site will vary with the antibiotic class
How are antiobiotics defined ?
Where antibiotic binds – defines class of antibiotic it belongs too
Main binding site is bacterial cell wall
What antibiotics work on cell wall synthesis?
Beta Lactams
-Penicillins
-Cephalosporins
-Carbapenems
-Monobactams
Beta lactam ring which is a key part of the molecule which is common to all beta lactam antibiotics – helps antibiotic bind to penicillin binding protein
Vancomyocin
Bacitracin
Cell membrane
- Polymyins
What antiobiotics work on nucleic acid synthesis?
Folate synthesis - Important in production in building blocks to make nucleic acids
- Sulfonamides
- Trimethoprim
Sulphamethoxazole
Trimethoprim
Co-trimoxazole
DNA gyrase - Quinolones
-Metronidazole
-Fluoroquinolones
RNA polymerase
- Rifampin
Ciprofloxacin
Levofloxacin
Moxifloxacin
What antibiotics work on protein synthesis?
50s subunit (ribosome)
- macrolides
- Clindamyacin
- Linezolid
- Chloramphenicol
- Streptogramins
30s subunit
- Tetracyclines - Doxycycline
- Aminoglycosides - Gentamiacin
Lincosamides - Clindamyacin
Macrolides
Erythromycin
Clarithromycin
Azithromycin
Chloramphenicol
Where are cell walls bigger in - gram positive or gram negative bacteria?
Gram positive
Beta lactam and glycopeptides are particulary useful for gram positive bacteria – do have a role in gram negative bacteria
How do beta lactam antiobiotics work?
To bind to the PBPs, the β-lactam antibiotic must first diffuse through the bacterial cell wall.
Key thing in cell wall matrix is peptidoglycan – peptides and sugars
Key binding sites that allow peptidoglycan to work as cement within bricks – that cement where antibiotics work and disrupt
When penicillin binding proteins are disrupted by antibiotics such as penicillin lose integrity of cell wall and get lysis
Bacteria explodes and dies
disrupt peptidoglycan production
by binding covalently and irreversibly to the Penicillin Binding Proteins
cell wall is disrupted and lysis occurs
results in a hypo-osmotic or iso-osmotic environment
Active only against rapidly multiplying organisms
Why is it harder for gram negative bacteria to be penetrated by beta lactams?
Gram-negative organisms have an additional lipopolysaccharide layer that decreases antibiotic penetration. Makes it harder for beta lactams.
Differences in the spectrum and activity of β-lactam antibiotics are due to their relative affinity for different PBPs.
Need chemically engineered beta lactam for gram negative
Why are penicillins ineffective in the treatment of intracellular pathogens?
Because the penicillins poorly penetrate mammalian cells, they are ineffective in the treatment of intracellular pathogens.
Features of a fungal cell
DNA/RNA synthesis, protein synthesis
-Similar to mammalian
-Flucytosine
Cell Wall
-mannoproteins
-Β1,3 glucan
-Β1,6 glucan
-chitin
-Doesn’t exist in humans
-Echinocandins
Plasma membrane
-ergosterol
-Human cell membrane contains cholesterol not ergosterol
-Amphotericin
-Azoles
-Terbinafine
What are we trying to achieve with antiobiotics?
Antibiotics have been around for <100 years, yet human race survived!
Antibiotics give time and support for the immune system to deal with an infection.
How are bacteria pathogenic?
Attach and enter > Local Spread > Multiply > Evade host defences > Shed from body
Consequences:
destroy phagocytes or cells in which bacteria replicate
Exotoxin - Protein production
Endotoxin - Gram negative
Inflammation – e.g. necrotic cells
Immune-pathology – e.g. antibody
Diarrhoea
What are bactericidal antiobiotics?
The agent kills the bacteria
Kill >99.9% in 18-24 hrs
Antibiotics that inhibit cell wall synthesis
Useful if poor penetration (Endocarditis), difficult to treat infections or need to eradicate infection quickly (meningitis
What are the situations where bactericidal antibiotics are important? (Sepsis)
Sepsis – the infection has become severe despite a functioning immune system and is so aggressive that the patient will die before a static antibiotic is able to help
What are the situations where bactericidal antibiotics are important? (Meningitis and encephalitis)
The infection will cause death or irreversible brain damage before a static antibiotic will help
What are the situations where bactericidal antibiotics are important? (Endocarditis)
The infection is in a site where the patient’s own immune system is unable to deal with the bacteria and therefore a static antibiotic won’t eliminate the bacteria as it is working alone (Bacteria within cardiac vegetations are at high concentration, and have lower rates of metabolism and cell division or are dormant, being surrounded by fibrin, platelets, and possibly calcified material. High levels of bactericidal agents are required for a prolonged period)
What are the situations where bactericidal antibiotics are important? (Primary and secondary immunodeficiency)
the patient doesn’t have a properly functioning immune system to work with a static antibiotic e.g. febrile neutropaenia
They either kill the bacteria pretty quickly – usually the cell wall ones – really useful in certain conditions or if it’s a quite hard to treat infection where you need everything you can
What are bacteriostatic antiobiotics?
prevent growth of bacteria
‘inhibitory to growth’
In fact kill >90% in 18-24 hrs
defined as a ratio of Minimum Bactericidal Concentration (MBC) to Minimum inhibitory Concentration (MIC) of > 4
Antibiotics that Inhibit protein synthesis, DNA replication or metabolism
Reduce toxin production and Endotoxin surge less likely
Do actually kill most bacteria
Single bacterium doesn’t last too long anyway
Exotoxin or endotoxins less likely to cause damage
No one antibiotic is only one of those 2 groups – different antibioitcs work in different ways at different times
What can bactericidal antiobiotics do?
Can lead to release of endotoxin (essentially bits of the cell wall) and the resulting increase in antigenic load causes an aggressive and dangerous inflammatory response
What is the MIC?
Minimum inhibitory concentration: testing to see if pathogen is resistant to antibiotics
How do we calculate how much antiobiotic you need?
Step 1: Tube dilation
Step 2: Incubate for 24 hours
Step 3: Determine MIC based on turbidity
What we do is have tubes with broth that promotes growth of bacteria (culture medium but its liquid) – mix that broth with different concentrations of antibiotic your testing – from 0 mls to pure antibiotic – if broth is pure antibiotic and no media imagine the bacteria unlikely to go – vice versa bacteria should grow – can work out at what point conc of antibiotic is enough to inihibit the bacterial growth
Why does lowest MIC not mean best antibiotic necessarily?
Drug must not only attach to its binding target but also must occupy an adequate number of binding sites, which is related to its concentration within the microorganism.
To work effectively, the antibiotic should remain at the binding site for a sufficient period of time in order for the metabolic processes of the bacteria to be sufficiently inhibited.
The two major determinants of anti bacterial effects are the concentration and the time that the antibiotic remains on these binding sites
What is time dependent killing?
Key parameter is the time that serum concentrations remain above the MIC during the dosing interval:
t>MIC
beta-lactams (penicillins, cephalosporins, carbapenems, monobactams),
clindamycin,
macrolides
oxazolidinones
What is concetration - dependent killing?
Key parameter is how high the concentration is above MIC
peak concentration/MIC ratio
aminoglycosides
quinolones
What is pharmacokinetics and how does it work in relation to antibiotics?
The movement of a drug from its administration site to the place of its pharmacologic activity and then its elimination from the body
A function of:
1.Its release from the dosage form;
2.Its absorption from the site of administration into the bloodstream;
3.Its distribution to various parts of the body, including the site of action and
4.Its rate of elimination from the body via metabolism (LIVER) or excretion (KIDNEY) of unchanged drug.
What is the appropriate or available route of administration?
Oral
IV
Suppository - for mucosal absorption
How do antibiotics get distributed to sites of infection?
Which antibiotics will penetrate that site?
What is the pH of the site?
Is the antibiotic lipid soluble?
Lipid soluble
pH affects your distribution
How much fat someones got in their body
Not just your physiological status
If you have collection of pus don’t have system of blood vessels going into middle of that – take antibioitcs to ege and hope it finds its way to the middle
Heart valves don’t have blood supply – get antibioitcs to heart valve have to flow
Size of molecule could be really important
F – small molecule – first thing for SA - Staph aureus
Alternative to F is Vancomyocin – when it has resistance
How do bacteria resist antiobiotics?
Change antibiotic target
Destroy antibiotic
Prevent antibiotic access
Remove antibiotic from bacteria
Beta lactamases
What happens in change antibiotic target?
Target protein is what antibiotic binds to in pathogen – target protein aa sequence determined by the genetic sequence of nucleoside bases in DNA – if this gets changed get diff AA sequence so diff peptide means the protein shape folded is slightly different and antibiotic molecule cant bind to it anymore
Another gene might get switched on that produces a molecule that binds that site and blocks it from the anti microbial but it doesn’t itself affect the function of the molecule it binds to – changing the antibiotic target
Examples of change in antibiotic target?
Flucloxacillin (or methicillin) is no longer able to bind PBP of Staphylococci – Methicillin resistant S. aureus
Wall components change in enterococci and reduce vancomycin binding – Vancomycin resistant Enterococci
Rifampicin activity reduced by changes to RNA polymerase in MTB – MDR-TB$
What happens in destroy antibiotic?
The antibiotic is destroyed or inactivated e.g.
1.Beta lactam ring of Penicillins and cephalosproins hydrolysed by bacterial enzyme ‘Beta lactamase’ now unable to bind PBP
2. Staphylococci produce ‘penicillinase’ so penicillin but not flucloxacillin inactivated
3. Gram negative bacteria phosphorylate and acetylate aminoglycosides (gentamicin)
How does preventing antibiotic access work?
-modify the bacterial membrane porin channel size, numbers and selectivity e.g.
1.Pseudomonas aeruginosa against imipenem,
2.Gram negative bacteria against aminoglycosides
How does removing antibiotic from bacteria work?
-Proteins in bacterial membranes act as an export or efflux pumps - so level of antibiotic is reduced
-S. aureus or S. pneumoniae resistance to fluoroquinolones
-Enterobacteriacae resistance to tetracylines
-Bacteria produces new channel/ pump that pumps out the molecule/ antibiotic
Why do bacteria develop resistance?
Intrinsic > Naturally resistant
Acquired> Spontaneous gene mutation or Horizontal gene transfer
Horizontal gene transfer > Conjugation, transduction, transformation
What happens in intrinsic resistance?
-All subpopulations of a species will be equally resistant
examples:
-Aerobic bacteria are unable to reduce metronidazole to its active form
-Anaerobic bacteria lack oxidative metabolism required to uptake aminoglycosides
-Vancomycin cannot penetrate outer membrane of gram negative bacteria – just too big a molecule to get in
-The PBP in enterococci are not effectively bound by the cephalosporins
What happens in acquired resistance?
A bacterium which was previously susceptible obtains the ability to resist the activity of a particular antibiotic
Only certain strains or subpopulations of a species will be resistant
What happens in spontaneous gene mutation?
New nucleotide base pair
change in amino acid sequence
change to enzyme or cell structure
reduced affinity or activity of antibiotic
These all lead to >
MTB - Point mutations in the rifampin-binding region of rpoB
Why is MRSA resistant?
MRSA - acquisition of mecA genes which is on a mobile genetic element called “staphylococcal cassette chromosome”
What happens in conjugation?
Pre designed method for bacteria to share DNA through plasmid transfer
Mechanism of gene transfer responsible for most concerning aspects of antimicrobrial resistance.
Sex pilus (small tube) forms between 2 bacterial cells through which plasmid is transferred from one to another.
What happens in transduction?
viruses that attack bacteria are moving genetic material from one bacterium to the next by accident – tryna move viral genetic material – main mechanism for mrsa to develop
Bacteriophages (viruses that infect bacteria) mediate transfer of DNA between bacteria via transduction - where DNA from donor bacterium is packaged into a virus particle and transferred into a recipient bacterium during infection
What happens in transformation?
Transformation where bacterial dna can get swallowed up by bacteria and get integrated into its genome
Some bacteria are able to take up free DNA from the environment and incorporate it into their chrosome
What can conjugation lead to?
New Delhi metallo-β-lactamase, ESβLs
What can transduction lead to?
mecA genes for MRSA
What can transformation lead to?
foreign DNA from S. mitis to S. pneumoniae, conferring penicillin resistance
What is MRSA?
Gram-positive bacteria
Methicillin resistant Staphylococcus aureus
Bacteriophage mediated acquisition of Staphylococcal cassette chromosome mec (SCCmec)
contains resistance gene mecA
encodes penicillin-binding protein 2a (PBP2a)
confers resistance to all β-lactam antibiotics in addition to methicillin (= flucloxacillin)
What is VRE?
Gram positive bacteria
vancomycin-resistant enterococci
Plasmid mediated acquisition of gene encoding altered amino acid on peptide chain preventing vancomycin binding
Promoted by cephalosporin use
What is ESBL?
Gram-negative bacteria
-Extended spectrum beta lactamase (ESBL) inhibition
-These hydrolise oxyimino side chains of cephalosporins: cefotaxime,ceftriaxone, andceftazidime and monobactams: aztreonam
-TEM-1 in E. coli, H. influenzae and N. gonorrhoea
-SHV-1 in K. pneumoniae
-An even more extensive ESBL, this time plasmid mediated, is the CTX-M cephalosporinase in Enterobacteriacae
What are ESBL strains remain sensitive to?
beta-lactamase inhibitors
Amoxicillin + Clavulanate =
Co-Amoxiclav
Pipericillin + Tazobactam=
Tazocin
What is AmpC b-lactamase resistance?
-Broad spectrum penicillin, cephalosporin and monobactam resistance
-encoded on the chromosome in bacteria such as Citrobacter spp., Serratia marcescens, Enterobacter spp.
-b-lactamase inhibitor resistant!
-inducible expression (gene only turned on by antibiotic)
What was invented to battle What is AmpC b-lactamase resistance?
Carbapenems
such as ertapenem, imipenem, meropenem
-in contrast to other b-lactams, are highly resistant to degradation by b-lactamases or cephalosporinases.
-often the antimicrobials of last resort to treat infections due to ESBL or AmpC -producing organisms of the Enterobacteriacae family*.
What do Carbapenem Resistant Enterobacteriaceae produce?
carbapenemases
Treatment options are very few
and very toxic
What can group A streptococcus cause?
can cause severe skin infections – can be necrotizing
What factors to consider when deciding if an antibiotic is safe to prescribe?
Intolerance, allergy and anaphylaxis
Side effects
Age
Renal and Liver function
Pregnancy and breast feeding
Drug interactions
Risk of Clostridium difficile
What do beta lactams do?
Theyre cell wall (peptidoglycan) weapons
act on cell wall of bacteria – inhibit cell wall synthesis – bacterial cell undergoes lysis – bacteria dies
Whats the fancy name for cell wall?
Peptidoglycan
Whats the difference between gram negative and gram positive peptidoglycan?
gram negative have thinner layer of peptidoglycan than gram positive
Examples of beta lactams?
Penicillin V
Benzylpenicillin
Flucloxacillin
Amoxicillin
Why are side chains of beta lactams important?
Side chains of beta-lactams will impart different spectrum of activity
What are some cephalosporins?
Cephalexin
Cefuroxime
Ceftriaxone
Cefotaxime
Why are cephalosporins good?
Good for people with penicillin allergy
Work against some resistant bacteria
Get into different parts of the body e.g. meningitis
How are cephalosporins different?
Own special group in beta lactams – for people with non severe penicillin allergy – can be broader – come in different types and generations
How do we identify gram positive bacteria? (1)
Gram stain > Gram > Catalase test > If positive We have staphylococcus if negative we hve streptococcus
Coagulase on staphylococcus > if positive S. aureus
How do we identify gram positive bacteria? (1)
Gram stain > Gram + coccus > streptococcus > haemolysis on blood agar:
If beta we have Lancefield group strep GROUP A C G strep
If alpha > optochin > sensitive = Streptococcus
pneumoniae
What do we need to fight gram positive bacteria?
THICK CELL WALL THEREFORE NEED A SIMPLE CELL WALL WEAPOn
think beta-lactams
What antibiotic is used for S. aureus and Group A, C, G strep?
FLUCLOXACILLIN
What other antibiotic can be used for Group A, C, G strep?
PO PENICILLIN V
IV BENZYLPENCILLIN
NO PENICILLIN RESISTANCE*
What antibiotic can we use for S. pneumoniae?
PO AMOXICILLIN
IV BENZYLPENCILLIN
What does MRSA stand for?
Methicillin*-resistant Staphylococcus aureus
Vancomycin and Teicoplanin
Activity: Gram positive ONLY
Use: Gram positive bacteria resistant to beta-lactams e.g. MRSA, enterococci, some coag neg staph
Use: penicillin allergy
Only given IV
Need to know its level in patients blood
Nephrotoxic – bad for kidneys
What are the 5 functional groups for antibiotic weaponry?
Inhibitors of cell wall synthesis
Inhibitors of protein synthesis
Inhibitors of nucleic acid synthesis
Anti-metabolites
Inhibitors of membrane function
What are examples of macrolides - protein synthesis?
Clarithromycin and erythromycin – oral (& IV)
Activity:
Gram positives (S. aureus, β haemolytic strep) and atypical pneumonia pathogens
Use: penicillin allergy
Use: severe pneumonia
What is atypical pneumonia?
legionella – multiply inside cells, no good having a cell wall active antibiotic as can’t get to it if it’s hiding inside a cell
What are examples of lincosamides - protein synthesis?
Activity:
Gram positives eg S. aureus, β haemolytic strep , anaerobes
Use: cellulitis (if pen allergy)
Use: necrotising fasciitis (remember Elizabeth…)
TURNS OFF NASTY TOXINS MADE BY Gram positive bugs*
Examples of tetracyclines - protein synthesis?
Doxycycline = oral
Activity : Broad spectrum but mainly Gram positive (S. aureus and streps)
Use: cellulitis (if penicillin allergy)
Use: pneumonia
How do we identify gram negative bacteria?
Gram stain > Gram (rods) > Lactose fermentation > if positive Enterobacteriacae (coliforms) like Escherichia coli, Klebsiella > if negative Shigella, salmonella, proteus
Pseudomonas (oxidase positive)
Gentamicin
IV only
Activity : Gram negatives and staphs (use synergistically to treat streps)
Use: urinary tract infections (UTIs)
Use: infective endocarditis (synergistically)
Nephrotic
Use synergistically to treat some gram-positive bacteria
Example of quinolones (DNA synthesis)
Ciprofloxacin = oral (& IV)
Activity : Gram negative» Gram positive
Use: penicillin allergy
Use: UTIs
Use: intra-abdominal infections
Inhibit DNA synthesis
Trimethoprim
anti-metabolite (folate antagonist)
Activity: Broad spectrum but mainly used for Gram negatives
Use: UTIs
Nitrofurantoin
Activity: Gram negatives and gram positives
Use: Lower UTIs
Gram negatives make …
Beta lactamases
Co-amoxiclav (Augmentin)
oral and IV
Activity: S. aureus, streps, enterococci, gram negatives, anaerobes
Use: aspiration pneumonia, severe CAP, more resistant urinary organisms
Piperacillin/tazobactam (Tazocin) - IV
Activity: Few gaps. No cover for ESBLs/AmpCs
Use: HA-pneumonia, systemic Pseudomonas infections, >65s abdominal infection, immunocompromised
Broad spectrum
Doesn’t cover more resistant ESBLs
Cefuroxime - IV
Activity: gram positives and gram negatives. No cover for Pseudomonas, anaerobes and enterococci
Use: some surgical prophylaxis, <65s intra-abdominal infections, non-severe penicillin allergy
Good cover for gram positive and gram negative
Meropenem - IV
Activity: BROAD. Active against ESBLs/AmpCs.
Use: HAI (sickest, most at risk), resistant gram negatives, immunocompromised
Need special code to use – restricted use
Broadest spectrum antibiotic – use against very resistant bacteria
Why is resistance an infectious arms race?
Since AB developed saved countless lives – at point now every new antibiotic has resistance – not enough new weapons to counteract this – new resistance every couple of years
23-year old woman attends with dysuria, frequency and cloudy urine for 36 hours.
Likely diagnosis?
Send samples?
Likely diagnosis
Urinary tract infection (lower)
Bacteria: gram negatives (E. coli, proteus, klebsiella)
And gram positives (staph saprophyticus)
Samples
Consider urine culture if possibility Resistance
✓ Children, pregnancy, men, > 65s, sepsis/pyelonephritis, catheter
66 year old woman
3 day history of fevers and nausea
Leg = hot, red, swollen
Likely bacteria -> diagnosis?
Treatment options?
Cellulitis usually lower limbs - unilateral
Red, hot, painful, tender skin
Spreading
Systemic symptoms
Bacteria: S. aureus and β haemolytic strep (Group A, C & G)
What are fungi?
Eukaryotic
Chitinous cell wall
Heterotrophic
“Move” by means of growth or through the generation of spores (conidia), which are carried through air or water
Move passively
Produce spores that can move through water
What is yeast?
Yeasts are small single celled organisms that divide by budding
-Account for <1% of fungal species but include several highly medically relevant ones
What are moulds?
Moulds form multicellular hyphae and spores
Some fungi exist as both yeasts and moulds switching between the two when conditions suit – dimorphic fungi
What type of fungi is very common?
Superficial
What type of fungi is rare?
Invasive - can be easily missed
Eg of fungal infection
Nappy rash and Vulvovaginal candidiasis
Tinea pedis (athlete’s foot)
Onychomycosis (fungal nail infections)
Otitis externa
What are some invasive / life threatening fungal disease in immunocompromised hosts?
Candida line infections
Invasive aspergillosis
Pneumocystis
Cryptococcosis
Mucormycosis
What are some invasive / life threatening fungal disease in post surgical patients?
Intra-abdominal infection
What are some invasive / life threatening fungal disease in healthy hosts?
Fungal asthma
Travel associated fungal infections
-Dimorphic fungi
Post-influenza aspergillosis
How much does NHS spend on antifungals a year
Less than 150m
Much of this cost is prophylaxis or empirical treatment of possible invasive fungal disease due to poor diagnostics.
What are some diagnostics for fungi?
Radiology
Microscopy
Culture
Molecular
Examples of molecular tests
PCR – mixed results
Antigen tests
-Cryptococcal Ag - excellent
-Galactomannan - insensitive
-1,3 Beta-D-glucan – poorly specific
What are the disadvantages of radiology, microscopy and culture?
R - Insensitive in early stages
M - Usually insensitive
C - OK for yeasts, poor for moulds
What is the aim of an antimicrobial drug?
achieve inhibitory levels of agent at the site of infection without host cell toxicity
What are the identifying molecules with selective toxicity for organism targets?
Target does not exist in humans
Target is significantly different to human analogue
Drug is concentrated in organism cell with respect to humans
Increased permeability to compound
Modification of compound in organism or human cellular environment
Human cells are “rescued” from toxicity by alternative metabolic pathways
Why is it more challenging to treat fungi than bacteria?
Generally much more difficult for fungi than bacteria because they are eukaryotic
More challenging to treat fungi than bacteria – more similar to out own cells – eukaryotic
Trying to kill invading cell without killing normal host cell
Fungal cell features
DNA/RNA synthesis, protein synthesis - similar to mammalian - Use Flucytosine
Cell wall - mannoproteins
Β1,3 glucan
Β1,6 glucan
chitin
Doesn’t exist in humans
Echinocandins used to attack cell wall
Plasma membrane:
ergosterol
Human cell membrane contains cholesterol not ergosterol
Amphotericin
Azoles
Terbinafine are used
Amphotericin B
is very broad spectrum with a high barrier to resistance but:
IV only
Adverse effects+++ (reduced but not eliminated by expensive liposomal formulations)
Renal, biochemical disturbance, infusion reactions
Intravenous – has severe adverse effects – effectively forms a pore in the membrane – leach out salt and membrane protein – nephrotoxic drug – can lead to infusion reaction when first added
Not first line therapy anymore
Echinocandins
Echinocandins are safe and effective drugs against Candida with some activity against Aspergillus but are IV only
Target cell wall – enzyme of cell wall – used in ITU and haematology – active against vast majority of candidates – really safe – intravenous only – safe and effective drugs – primarily used
Azoles
Azoles are the mainstay of antifungal therapy currently
Different drugs have different spectrum of activity
Important toxicities and drug-drug interaction to be aware of
Main stay is the azoles – broad class of drugs – been around for decades – earlier azoles where
Important class of drugs to know about
What are some adverse affects of azoles
Relatively safe
All associated with transaminitis and GI SEs
-Rare severe hepatitis
Alopecia with long term fluconazole
GI symptoms more pronounced with Itra
-Nausea, abdominal pain, diarrhoea
-Discontinuation of drug in 10%
-Rare life threatening liver failure
Voriconazole associated with reversible visual disturbance in 30%
Photosensitivity in 1-2% of patients receiving voriconazole and recent reports of skin malignancy
Why do azole drug- drug interactions occur?
Largely a result of cytochrome P450 isoforms
Itraconazole
potent CYP3A4 inhibitor
As above + steroids, statins, rifamycins,PIs
Fluconazole
hydrophilic and the wat its principally excreted unchanged – less significant interactions
Warfarin, phenytoin, calcineurin inhibitors, anxiolytics
What is posaconazole?
Posaconazole is only a mild CYP3A4 inhibitor.
What is onychomycosis?
Common
Caused by dermatophyte moulds
Trichophyton rubrum is most common
Broad differential diagnosis
Microscopy is most specific test but 30% culture negative
Topical success best in children with thin nails and less than 50% of plate involved. Amorolfine has an approximately 20% complete cure rate
Non-dermatophytes on culture often just colonising
What is the treatment for onychomycosis?
Slightly depressing
Results of sampling can be confusing
Limited treatment options
Topical amorolfine
Systemic itraconazole or terbinafine
Treatment takes ages
High failure rate with all therapies
What is 1 or 3 )-β-D glucan?
Cell wall component of many fungi including common ascomycetous pathogens and Pneumocystis
i.e. not zygomycetes or basidiomycetes
Released into serum during invasive infection
Assay very prone to contamination (80pg/ml cut-off)
Non-specific for individual fungi
What is pneumocystis pathogenesis?
Infection/colonisation of healthy people frequent and occurs early in life
Disease develops only with moderate-severe immunocompromise esp. HIV, transplant, steroids
Frequent clinical presentation of unknown HIV
Think about Pneumocystis when a patient has hypoxia more severe than CXR would suggest especially if gradual onset illness or risk factors.
What is the treatment for pneumocystis pathogenesis?
Co-trimoxazole (plus steroids if hypoxic) is first line treatment
What is the amount of people living with HIV?
38.4 million
How many new HIV infections were there in 2021?
1.5 million
How many deaths were due to AIDS in 2021
650 000
Whos is affected by HIV/ AIDS?
About 58% are in sub-Saharan Africa
About 430 are among children under 15 years of age
About 3600 are among adults aged 15 years and older, of whom:
─ almost 49% are among women
─ about 31% are among young people (15–24)
─ about 19% are among young women (15–24)
What is a normal CD4 count ?
Anything more than 500
What is a viral load?
amount of HIV in your blood
What is UNAIDS 90/90/90 goal to eliminate the epidemic?
global target of
-90% of people living with HIV being diagnosed
-90% diagnosed on ART (antiretroviral therapy)
-90% viral suppression for those on ART by 2020
What is the fast track cities initiative?
Global partnership between 90 high HIV burden cities where everyone works together to tackle HIV
Is the number of people being diagnosed with HIV since 2019 decreasing or increasing?
Decreasing
Who are at most risk of contracting HIV?
Gay / bisexual men
What are the HIV transmission routes?
Sexual
Vertical – mother to child transmission
Blood – don’t see much of this because of blood screening in the country
What is U=U?
Undetectable = Untransmissable
signifies that those who receive effective antiretroviral therapy and have achieved and maintained an undetectable viral load cannot transmit the virus to a sexual partner.
What is Prep?
Pre-exposure prophylaxis of HIV (PreP)
Taking HIV tablets b4 sex
Take a pill a day or as in when (on demand) – 2-4 hours before sex
What is the effectiveness of Prep?
Several RCTs have reported on PreP; providing evidence for the effectiveness of daily dosing and event-based dosing
Effectiveness has been demonstrated in MSM (men who have sex with men), heterosexual serodifferent couples, and injecting drug users
What is Pep?
Post-exposure prophylaxis
PEP = 28 days Combination Antiretroviral Therapy –must be started within 72 hours
Not as effective as PreP
What are the benefits of knowing your HIV status?
Access to appropriate treatment and care
Reduction in morbidity and mortality
Reduction of vertical transmission
Reduction of sexual transmission
Public health /partner notification
Cost-effective
What scenarios would you test HIV for?
Clinician initiated diagnostic testing triggered by clinical indicators of immuno-suppression disease /seroconversion
Routine screening in high prevalence locations
Antenatal screening
Screening in high risk groups
Patient initiated requests for testing
Why do some doctors not test for HIV?
They don’t think of HIV
Underestimate the risk of HIV in their patients
Failure to recognise HIV as a modifiable prognostic indicator
Misconception they need pre-test counselling
Misunderstanding of the implications for insurance, etc
Fear of offending the patient
When should we start to suspect HIV in a patient?
Generalised lymphadenopathy
Acute generalised rash
Glandular fever/ flu-like illnesses
Think about seroconversion
Oral candida
Unexplained weight loss or night sweats
Persistent diarrhoea
Gradually increasing shortness of breath and dry cough
Recurrent bacterial infections including pneumococcal pneumonia
What are some common examples of recurrent medical conditions that someoen with HIV might have?
Multi-dermatomal shingles
Unexplained lymphadenopathy
Unexplained wt loss or diarrhoea, night sweats, PUO
Oral/oesophageal candidiasis or hairy leukoplakia
Flu-like illness, rash, meningitis
Unexplained blood dyscrasias
What type of tests can we do for HIV?
Venous blood sample is preferred
Point of care test
Origins of HIV
Africa
HIV-1 and HIV 2 similar to retro viruses found in monkeys in Central and West Africa
First documented sample from infected human was from DR Congo in 1959
Bushmeat markets
in West and central Africa are a potential route of transmission of simian retroviruses
Features of HIV-2 infection
Natural model of HIV control or “functional cure”: 35-40% are “elite controllers”, who maintain undetectable viral load for a decade or more and have a normal lifespan
What is the HIV genome structure?
Small RNA virus (~ 10KB): expresses just 10 genes
Member of retrovirus family (uses reverse transcriptase to make DNA copy of itself)
Lentivirus: characterized by long incubation period
What is the mechanism of viral replication for HIV?
-Glycoproteins on the HIV molecule (gP160 made of gP120 and gP 41) allow it to dock and fuse onto the CD4 and CCR5 receptors
- HIV fuses to CD4 receptor and passes its contents into the CD4+ cell
-The viral capsid the enters the cell and enzymes and nucleic acid are released
-Using reverse transcriptase single stranded RNA is converted into double stranded DNA
-Viral DNA then is integrated into the cells own DNA by integrase enzyme
-When the infected cell divides the viral DNA is read and long chains of viral proteins are made
-The viral protein chains are cleaved and reassembled
-Budding here immature virus pushes out of the cell taking with it some cell membrane
-Immature virus breaks free to undergo more maturation
-Maturation protein chains in the new viral particle are cut by the protease enzyme into individual proteins that combine to form a working virus
What are the different proteins produced by the HIV virus?
GAG, POL, ENV - Structural
TAT, REV - REGULATORY
VPR, VIF, VPU, NEF - Accessory
What does POL do?
encodes the enzymes: reverse transcriptase, integrase and protease
What does ENV do?
encodes the envelope proteins
What does NEF do?
increases infectivity
What does TAT do?
contributes to viral replication. Enhances production of host transcription factors e.g NF-kB.
What does GAG do?
encodes structural proteins.
Made as a polyprotein and cleaved by HIV protease
What does rev do?
binds to viral RNA and allows export from nucleus and also regulates RNA splicing
What is the primary receptor for HIV?
CD4 cells
What are co receptors for HIV?
are CCR5 and CXCR4 chemokine receptors. CCR5 is used by HIV-1 in early infection, may switch to use CXCR4 later in infection. Once viral integration has occurred, infection persists for life in a reservoir of latently infected cells.
Who are some people with genetic resistance to HIV infection?
1% of Caucasians are homozygous for a 32bp deletion in the CCR5 gene (CCR5D32) necessary for primary HIV-1 infection
People with only one copy of the mutant gene can be infected with HIV but show delayed disease progression
It has been hypothesised that the origin in Caucasians could be related to protection from the Plague
Why is HIV-1 so hard to target?
Mutates rapidly
HIV-1 shows huge viral sequence diversity that is increasing over time
Why does HIV-1 evolve so rapidly?
Error-prone replication (the enzyme reverse transcriptase makes at least 1 error in every replication cycle)
Rapid viral replication (generation time ~2.5 days)
Large population sizes (~1010 new virus particles produced each day)
What happens in acute HIV infection?
Acute HIV infection is early stage of HIV where there is a peak of viremia
It is characterized by high viral load
Immune responses during acute HIV infection are important for subsequent viral control
Detection of very high levels of virus in the blood
Symptoms of Acute Retroviral Syndrome:
“Glandular fever”-like illness
Fever, lymphadenopathy
Sore throat, oral ulcers
Skin rash (upper trunk)
May include neurological features
Why is early use of ART useful?
Reduced risk of transmission
Smaller reservoir, lower set-point, delayed progression
What are the key features of HIV pathogenesis?
HIV is integrated into the DNA of the infected CD4-expressing cells
HIV infects a range of CD4 + immune cells in addition to helper T-cells, (including regulatory T-cells, T follicular helper cells, dendritic cells, macrophages and thymocytes)
However, the number of HIV-infected CD4+ T-cells in the blood does not explain the extent of immune suppression
HIV can pass directly from cell to cell, and so it is relatively inaccessible to antibodies in the blood
The small HIV genome encodes a range of genes that enable the virus to evade human immune system responses
Immune system is constantly activated by HIV
How does immune activation drive HIV pathogenesis?
Activated T-cells in the blood, correlated better with disease progression than viral load or CD4 count
Early in HIV infection, there is a dramatic loss of CD4+ T-cells in the lymphoid tissue in the gut: this makes the gut mucosa leaky, allowing passage of bacteria and products (like LPS), stimulating immune cells and setting up a cycle of chronic immune activation that ultimately exhausts the immune system
Immune activation is also driven directly by HIV, e.g. through a form of inflammatory cell death (pyroptosis), and co-infections, particularly with cytomegalovirus (CMV)
Why does immune response to HIV-1 fail to clear the virus?
-Immune system generates a massive immune response to HIV infection, involving up to 20% of all circulating T and B lymphocytes
-Antibodies develop against most viral proteins, but neutralising antibodies take months to develop and rarely neutralise the primary HIV strains that are transmitted from person to person
- key immune response to HIV-1, from CD4+ T-helper cells, is lost from very early in infection, because these are the cells HIV infects first
-vigorous response from cytotoxic CD8+ T-cells, which provides the major force controlling viral replication but ultimately fail when “immune exhaustion” sets in
What is the surface of the virion derived from?
surface of the virion is derived from the host cell membrane containing only a few (6 – 10) envelope spikes
Whats the issue with HIV-1 envelope spike>
is heavily glycosylated (with sugars resembling human types), which makes it difficult for antibodies to bind to the surface
Why do most HIV-1-infected people fail to make an effective antibody response?
-really critical parts of the viral envelope that are needed to enter CD4+ T-cells are either in deep pockets overhung by sugar molecules or only revealed when the virus docks onto the CD4 molecule
-The envelope (gp120/41) proteins can change substantially without affecting virus function
-Thus the virus can evolve very quickly to avoid antibody recognition (including by the addition of more sugar molecules)
-HIV has small number of spikes on cell surface – enough to get into cells but not so many so that we can make antibodies against it and its antigen
What happens in infected people with HIV-1?
circulating neutralising antibodies rarely recognise their own prevailing viral envelope variants`
What do 20 percent of people with HIV have?
generate broadly neutralising antibodies (BNAbs) that can neutralise multiple HIV strains
Involves considerable somatic mutation of the antibodies (50+ mutations from the original antibody), so hard to generate with a vaccine
What are the targets of BNAbs?
-CD4 binding site
-Membrane-proximal region
-V2V3 conformational epitope
-Envelope glycans
Potential use in therapy and to prevent infection
How do CD8+ cytotoxic T-cells identify cells?
CD8+ cytotoxic T-cells identify cells expressing foreign material (from pathogens or tumours), processed as small fragments of protein (8-11 amino acids in length), presented on the surface of the infected cell by HLA class I molecules
What do different HLA class I molecules do?
Different HLA class I molecules are able to present peptides with different characteristics: a set of three distinct HLA class I molecules (A, B & C) are inherited from each parent
These peptides can come from any part of a pathogen, so include more conserved structural and functional internal proteins (whereas antibody recognition is largely limited to surface proteins)
What do recognition triggers do?
Recognition triggers the release of soluble anti-viral factors and the death of the infected cell
Cytokines – soluble anti-viral factors
CC- chemokines
(compete with HIV for
the receptor CCR5)
Cytotoxic factors
– kill the cell
Cytotoxic T-lymphocytes (CTL) appear when?
CTL appear early in HIV infection, coincident with rapid drop in viral load
CTL exert sufficient selection pressure on the virus for variants which escape CTL recognition to emerge
CTL selection leads to HLA-class I associated “foot-prints” on viral evolution
What are HLA class one molecules associated with?
HLA class I molecules associated with good outcome
How does HIV-1 evade the CTL response ?
HIV can evolve to escape T-cell recognition at several points on the antigen processing and presentation pathway
Mutant HIV variants that evade the T-cell response appear within weeks of primary HIV infection
Initially responses develop to the new variants but these are progressively undermined by the failure of CD4+ cell help and dendritic cell function
What does the HIV-1 nef protein do ?
-Protein that reduces cell-surface expression of HLA class I molecules needed for CTL recognition, whilst at the same time upregulating the “death” molecule Fas that can kill virus-specific CTL before they can kill the virus-infected cell
-HLA- A and B molecules are down-regulated to undermine CTL killing of infected cells but HLA-C expression is maintained to prevent NK cell killing
-Ultimately CTLs develop functional “exhaustion”, associated with expression of inhibitory molecules such as PD-1: levels of expression correlate with viral load
What is the median time to AIDS (without ART) ?
10-11 years
What are Long-term non-progressors (LTNPs)?
defined as survivors with HIV-1 infection for >7-10 years,
no therapy
no symptoms
stable CD4+ T-cell count > 500mm3
RARE - around < 1-5 % of cohorts
What are Elite Controllers?
-Defined as HIV-1 infected individuals with plasma VL <50 copies/ml for over one year without ART:
VERY RARE - 0.35-0.8% of cohorts
-General but not complete overlap: controllers may progress to AIDS even with low viral load, LTNPs may have high Viral load: most eventually develop disease
-Strongly associated with HLA class I alleles, but generally not distinguished by stronger or better immune responses (except preserved HIV-specific CD4+ IL-2+ responses)
Why is life expectancy still reduced in HIV-infected people on cART?
Issues of adherence, side-effects, drug resistance
Increase in non-AIDS-defining illnesses (NADIs): lung, cardiovascular and renal disease
Incidence of NADIs is related to:
-Size of latent HIV reservoir
-Persistent immune activation
-CMV co-infection
Even with ART (Anti retroviral therapy), why are there still problems with those with HIV?
Even with the most effective available ART, a poorly-defined reservoir of latently-infected cells persists for years, and HIV starts replicating again (to pre-treatment levels) within weeks of cessation of therapy
HIV is thought to persist as DNA integrated into “resting” transcriptionally-silent CD4+ T-cells and other cells such as tissue macrophages and T follicular helper cells
HIV may continue replicating in lymphoid tissue and other immune-privileged sites
What does the HIV reservoir size correlate with?
With persistent immune activation, largely driven by translocation of microbial products across the gut (damaged gut-associated lymphoid tissue and leaky gut since primary infection)
Is there a cure for HIV?
Berlin patient - He is the only person on the planet who has been cured of HIV infection using the incredibly toxic treatments he was receiving for leukaemia. This case has given new hope but it has also raised some major ethical dilemmas.
What are the key populations who are affected by HIV the most in modern society?
sex workers and their clients, gay men and other MSM, people who inject drugs,
transgender people - account for >70% new infections globally
Which regions seeing most rapid rise in new HIV infections?
Eastern Europe and Central Asia
What is the link between HIV and COVID pandemics together?
People with HIV (PWH) are 30-50% more likely to die from Covid-19
Who does HIV affect?
1.5m new peole in 2021
Over 96% HIV infections occur in low and middle income countries (LMICs)
25.3m HIV-infected people live in Sub-Saharan Africa
~20% do not know their HIV status
Access to anti-retroviral therapy (ART) for Prevention of Mother-To-Child Transmission of HIV for 81% pregnant women globally
Treatment access has improved, with 69% HIV+ adults in Africa now on ART
1.7m children (<15 yrs) living with HIV
160,000 children born with HIV in 2021
In 2021 only 59% HIV+ children were on ART
Who are at most risk of acquiring HIV?
50% of all new infections occurring world-wide are in 15-24 year olds (women mostly)
What contributes to HIV tranmission
Sexual violence
What are the 3 routes of Paediatric HIV-1 infection?
In utero: transplacental, mostly during the third trimester
Intra partum: exposure to maternal blood and genital secretions during delivery
Breast milk: ingestion of large amounts of contaminated milk
In breast-feeding populations, risk of mother-to-child transmission (MTCT) is up to 45%
Transmission can largely be prevented by ART given to pregnant women and to the infant (MTCT)
What is HIV?
HIV is a lentivirus that uses reverse transcriptase to replicate (retrovirus)
HIV replicates within CD4 cells, and over time decimates their population causing immunodeficiency
HIV viral load increases over time
Uses protease and integrase as well
What are the 2 markers used to monitor
- CD4 cell count
- HIV viral load
Both are important in the prognosis
What is a maculopapular rash?
Red, blotchy areas, some flat and raised bits
How many weeks does it take to develop HIV symptoms?
From point of infection, usually takes 2-4 weeks to develop symptoms (acute HIV syndrome)
During the acute HIV syndrome, HIV replicates rapidly, and CD4 cell count drops (as they are used by HIV to replicate within).
The huge viraemia these patients experience is what causes their symptoms…
What are HIV symptoms similar to?
Glandular fever
Flu
What are some non specific symptoms of HIV ?
Fever
Sore throat
Myalgia
Rash
Vomiting + diarrhoea
Headache
Lymphadenopathy
Weight loss
Encephalitis
Aseptic meningitis
What do you always ask a patient with fever, rash and non-specific symptoms:
Ask about sexual history
Think of HIV seroconversion
What happens in clinical latency?
5-10 years
the immune system will start to take back control and the CD4 cell population increases, and the viral load temporarily decreases
However, over time the CD4 population will slowly decline until the person is immunosuppressed enough to present with constitutional symptoms and opportunistic infections
No symptoms!
May notice some enlarged lymph nodes > Persistent Generalised Lymphadenopathy
What is Persistent Generalised Lymphadenopathy?
Persistent Generalised Lymphadenopathy is defined as enlarged lymph nodes involving at least 2 areas of the body for at least 3 months
What can reactive when CD4 count gets really low (500-200)
-Shingles (reactivation of chickenpox virus)
-Thrush
-Candida
-Oral hairy leucoplakia - similar to thrush - white areas on tongue - cant scrape it off - assosciated with Epstein barr virus
- Molluscum contagiosum - more common in kids
Think about doing a HIV test when faced with a common problem:
In an unexpected patient
That is recurring
That has no clear underlying cause
When do we get AIDS?
CD4 <200
or
“AIDS defining illness” present - TB for eg
What is Pneumocystis Pneumonia (PCP)?
Bihilar infiltrates
Very fine white areas in chest x rays
Sometimes you can also get normal x ray
Fevers, SOB, dry cough, pleuritic chest pain,exertional drop in oxygen saturations
The most common AIDS defining illness
42.6% prevalence in AIDS
Key feature is an exertional drop in oxygen saturations
Fungal
Check severity using arterial blood gas
What antibiotics can we use for PCP?
Fungal, but susceptible to some antibiotics!
Co-trimoxazole
+/- prednisolone (steroids) if hypoxic
Co-trimoxazole is first line = 2 antibiotics: trimethoprim + sulfamethoxazole
Also called septrin
What can a late diagnosis mean for AIDS?
Increased transmission
Increased morbidity
Increased mortality
TB and HIV
All patients with TB require a HIV test
TB in HIV at any CD4 count: AIDS defining
Atypical presentations with lower CD4 count
Sample for Acid Fast Bacilli staining
AIDS defining illness
What are some CNS presentations for TB?
Tuberculoma:
Ring enhancing lesion which can have surrounding oedema
Most common
Can be a single mass with surrounding oedema or multiple small masses as part of military TB
CSF analysis often helps in establishing the diagnosis
Treated with 12 months TB treatment
Ocular TB:
TB can occur anywhere in the eye
Can involve any part of the eye (Eg. Uveitis, retinal, orbit, external eye) and can occur with or without evidence of systemic TB.
It generally develops following haematogenous spread from a primary focus but, in rare cases, it can also occur as a primary infection following an epithelial injury.
What happens in TB meningitis?
Can occur in non-HIV but much higher incidence and mortality in the HIV population
Variable presentation from an acute meningitis to a progressive dementia
Cranial nerve palsies (most commonly 6th nerve)
Insidious onset of headache – variable 1 day – 6 months!
Night sweats and fevers
Vomiting
Active pulmonary TB present in 30-60% of cases
Without treatment proceeds to coma and death
What happens in a CNS lymphoma?
Generally more likely to get any lymphoma due to the link with EBV
Risk of having CNS lymphoma is over 1000x higher(!) if you have a HIV diagnosis compared to the general population
Generally present with headaches and focal neurological symptoms depending on where in the brain the tumour is
Usually a single lesion and ring enhancing
Ultimately diagnosed on biopsy
What happens in CNS toxoplasmosis?
CNS toxoplasmosis is due to reactivation of latent infection (Seroprevalence 15-50%, acquired via cat contact or through contaminated meat or water)
CD4 usually <100
Multiple ring enhancing lesions on MRI
Treated with sulphadiazine + pyrimethamine (+folinic acid)
What happens in CNS toxoplasmosis?
CNS toxoplasmosis is due to reactivation of latent infection (Seroprevalence 15-50%, acquired via cat contact or through contaminated meat or water)
CD4 usually <100
Multiple ring enhancing lesions on MRI
Treated with sulphadiazine + pyrimethamine (+folinic acid)
What is CMV retinotis ?
Most common place for CMV to reactivate in HIV is eyes
Looks like a pizza
Cytomegalovirus is a herpes virus, that causes a glandular fever type illness in the immunocompetent individual.
If someone is immunosuppressed, they can get CMV pneumonitis, retinitis, encephalitis and colitis, but in HIV CMV retinitis seems to be the most common site of reactivation.
Causes visual blurring and blindness. Doesn’t cause pain.
Treated with antiviral agents (valganciclovir or ganciclovir)
What happens in ocular toxoplasmosis?
Toxoplasmosis can also affect the eyes… in 1-2% of those with HIV
Causes inflammation at the back of the eye (retina) and can cause pain, blurred vision and blindness
Acutely there is a necrosising retinochoroiditis
If the optic nerve is affected, direct spread can occur into the brain
Intravitreal antibiotics can be given in addition to systemic.
What about cryptococcal meningitis?
Cryptococcus is a yeast like fungus that can cause a meningitis in those who are immunocompromised.
Usually presents with a gradual onset headache and fever (weeks)
Little to find on examination and CT
LP – very high opening pressure (>40cm H2O)
India ink stain to visualise on microscopy, a “halo” appears around the cryptococci due to the polysaccharide capsule
Treat with Amphoteracin B + flucytosine and serial LPs
Gradual onset headache / fever
High opening pressure on lumbar puncture
India ink used on microscopy
If you have someone with HIV and they have CNS headaches what would you do?
Lumbar puncture
HIV increases the risk of any cancer assosciated with a …
Virus eg
Human Herpesvirus 8 > Kaposi’s sarcoma
Epstein Barr Virus > Lymphomas
Human Papillomavirus > Cervical, anal, penile carcinoma
Hepatitis B/C > Hepatocellulcar carcinoma
What are AIDS defining cancers?
Kaposi’s sarcoma
Lymphomas
Cervical cancer
What happens in Kaposis sarcoma?
Human Herpesvirus 8
Usually associated with HIV
Single or multiple lesionsUsually on the skin
Treated with HAART and chemo/radiotherapy
Kaposi’s usually indicates underlying HIV infection, however can rarely can be genetic
Spectrum of disease – single to multiple lesions
Other sites – mouth, GI tract GI bleed, respiratory tract
Can cause bleeding
What is HAART?
HAART (Highly Active Anti-Retroviral Therapy)
3+ antiretroviral drugs
NIRT + NIRT + OTHER
Act on different pointsin replication cycle tosuppress viral replication
Aim to reduce viral load to undetectable levels and increase CD4 count
What are the type of drugs we use for HIV?
Entry inhibitors e.g. Maraviroc
Fusion inhibitors e.g. Enfuvirtide
After HIV enters the cell, it uses reverse transcriptase to transform from RNA to DNA.
Our next two classes of ARVs inhibit reverse transcriptase to block this step.
3. NRTIs: Nucleoside reverse transcriptase inhibitors (-dines and -bines) - most HAART regimes are made up of a back bone of 2 NRTIs)
and NNRTIs: Non-nucleoside reverse transcriptase inhibitors.
What are integrase inhibitors?
The viral DNA then uses integrase to integrate its genome into the host cell’s DNA. This gives us another opportunity to block HIV replication with…
4. Integrase inhibitors “INSTIs” (-gravir) e.g. raltegravir, these are now used frequently as the 3rd agent within HAART, as they result in a rapid reduction of the viral load. Excitingly there are some new injectable INSTIs that can be used.
How can we use protease inhibitors for HIV?
After hijacking the nuclear DNA, the RNA product then uses protease to exit the CD4 cell.
- Protease inhibitors block HIV proteases to prevent HIV becoming a mature virion (–navir) e.g. darunavir. They are often given with a “boosting” agent. These used to be widely used, however they are associated with increased risk of CV events, so many patients have switched to other agents due to the aging population of patients with HIV.
With current HAART regimes what can happen?
With current HAART regimes, HIV infection is an entirely manageable condition with a good prognosis!
How does HIV develop drug resistance?
1 mutation in every 2 new viruses produced
1 -10 billion new virus particles each day
1-5 billion mutations per day
- Non adherence - missing doses or taking meds late
- Drug - drug interactions eg
- Decreased drug levels
a) Clopidogrel + Boosted PI = ↓ clopidrogel active metabolite
b) Lansoprazole + Rilpivirine = reduced rilpivirine levels +/- resistance
- Increased drug levels
a.Steroids + Ritonavir = risk of Addisonian crisis
b.Recreational drugs + antiretroviral boosting agents = toxicity
Good adherence and avoidance drug interactions are key to:
suppress HIV replication
avoid drug resistance
ALWAYS CHECK DRUG INTERACTIONS
What are the virulence factors (enzymes)
What are the virulence factors (Toxins)?
What are the virulence factors (Toxins)?
Toxins
Streptolysins O&S
-binds cholesterol
Erythrogenic toxin
-Streptococcal pyrogenic toxin e.g. SPeA – exaggerated response
What are the virulence factors (Toxins)
Toxins
Streptolysins O&S
-binds cholesterol
Erythrogenic toxin
-Streptococcal pyrogenic toxin e.g. SPeA – exaggerated response
