Week 6 Flashcards
Define immunological memory
What is the role of lymph nodes
filter & trap pathogens
What is the role of the spleen
filter blood
remove damaged RBCs
store platelets
What is the role of tonsils
first-line defence against ingested or inhaled pathogens
What is the role of thymus
maturation & education of T cells
What is the role of bone marrow
site of blood cell production (including lymphocytes)
What is the role of Peyers patches
important for gut associated lymphoid tissue (GALT) & immune response in digestive system
Describe major histocompatibility complex
MHC1 presents intracellular antigens to cytotoxic T cells
MHC2 presents extracellular antigens to helper T cells
Explain the roles of cytokines
Explain MHC class 1
Explain MHC class 2
What are the roles of T lymphocytes
What are the roles of B lymphocytes
What is GALT
Lymphoid tissue associated with lining of intestinal tract
Located immediately below epithelium of mucosal tissue
Primary lymphoid organ in ruminants (B cell proliferation)
Secondary lymphoid organ in other species
What does GALT contain
organized structures such as:
- Peyer’s Patches (PP)
- isolated lymphoid follicles
- Cryptopatches (aggregates of lymphoids)
scattered lymphocytes (intra-epithelial & in lamina propria) in intestinal villi
Where can Peyer’s patches be found
Located in jejunum and ileum
They can be visible inside lumen of intestines during inflammation
Describe peyers patches in group 1 species (ruminants, pigs, horses, dogs, humans)
Ileum
- 80-90% of Peyer’s Patches
- Form single continuous structure that extends forward from ileocecal junction
- Densely packed lymphoid follicles, contain only B-cells
- Reach maximal size & maturity before birth (prior to encounter of gut microbiota)
Jejunum
- Multiple discrete accumulations of follicles
- Persist throughout life
- Mainly B-cells, up to 30% T-cells
Describe peyers patches in group 2 species (rabbits, rodents)
Peyer’s Patches located at random intervals in ileum & jejunum
Development starts 2-4 weeks after birth & persist into old age
Development dependent on gut microbiota
Label the Peyers patches histology
Name the immunocytological content of GALT
Intestinal villi
- intraepithelial lymphocytes & lymphocytes in lamina propria
Peyers patches
Follicle associated epithelium
- M cells in between villous epithelium
sub-epithelial dome
- dendritic cells, macrophages, neutrophils, B & T cells
interfollicle areas
- T cells
follicle areas with germinal centres
- B cells
Lamina propria
- T cells
Describe M cells (part of GALT)
Regular distribution of M cells across follicle associated epithelium
Have apical microfolds with adhesion molecules (so antigens can be transported)
M-cells sample antigens & transport them to basolateral pocket to facilitate contact with immune cells on other side of barrier
They are transporter cells which transport antigens to dendritic cells which can present the antigen to lymphocytes
Compare Peyer’s patches to lymph nodes
Describe antigen sampling in GALT
Follicle associated epithelium sample soluble antigens
M cells & dendritic cells sample particulate antigens (eg bacteria)
Antigen is captured by dendritic cells & presented to T cells (in local LN if naïve animal)
Antigen specific T cells will be activated by second co-stimulatory signals
Activated Th cells secrete cytokines
- Promote differentiation & proliferation of:
*B cells –> plasma cells
*T cells –> CTL
How do lymphocytes of GALT stay at site of infection if infection is intestinal?
(Activated T & B cells normally recirculate from PP to LN into circulation)
During infection, mucosal adhesion molecules are upregulated on local endothelial cells
Activated T cells express mucosal adhesion receptor (done by dendritic cells)
As antigen specific lymphocytes re-circulate through local LN & site of infection, they adhere & remain at local site (& all other mucosal tissue) ie homing
Describe mucosal B-lymphocytes in GALT
- B cells are activated by cytokines from Th lymphocytes
- Differentiate into plasma cells
- IgM secreted first (but very large & vulnerable to intestinal enzymes)
- Cytokines stimulate plasma cells to class switch their antibody isotype from IgM to secretory IgA (protected from enzymatic digestion)
What are the 2 isotypes of IgA
IgA1 - predominant IgA subclass found in serum
IgA2 - predominant IgA subclass found in secretory lymphoid tissues e.g., GALT
- Secretory component is added to IgA antibody in lamina propria as it passes through mucosal epithelial cells
- –> protected from enzymatic digestion
Describe how the secretory component is added to IgA in GALT
Secretory IgA captures antigens in lumen:
- IgA binds to receptor on basolateral aspect of epithelial cell
- IgA-receptor complex is internalised into epithelial cell
- IgA-receptor complex is translocated to apical aspect where secretory component is wrapped around IgA molecule
- On mucosal surface of cells secretory IgA is either released into lumen or captures antigen directly
- Secretory IgA interferes with binding of pathogen to receptors on the epithelial cells to prevent pathogen invasion
- –> Immune exclusion rather than immune elimination
Describe IgG in GALT
Mucosal immunity also stimulates IgG production in circulation which can go into lumen but…
Mostly in serum & lamina propria
Neutralises bacterial toxins
- limit toxic effects
Opsonises invading pathogens, enhancing their phagocytosis or complement mediated lysis
- reduce numbers or eliminate pathogen
- –> Immune elimination
Name some parasites that exploit M cells & use them as portals of entry into body
Salmonella
Listeria
Yersinia
Some E. coli
Rotavirus
Cryptosporidium
Define hypersensitivity
when an animals immune response reacts to normally innocuous environmental antigens
Define type 1, 2, 3 & 4 hypersensitivity
Describe the phases of type 1 hypersensitivity
phase 1:
- sensitisation phase (no clinical signs yet)
1. allergen exposure via respiratory or skin
2. antigen presenting cells capture antigens
3. causes naive T cells to differentiate
4. release cytokines causing B cell proliferation
5. class switch for B cells from IgM to IgE
6. IgE binds to mast cells
phase 2:
1. same allergen encountered
2. primed IgE coated mast cells bind to antigen of allergen
3. release of cytoplasmic granules
4. immediate hypersensitivity - pruritis, broncho-constriction, oedema
Describe anaphylaxis (example of type 1 hypersensitivity)
What kind of hypersensitivity is atopic dermatitis in cats
type 1 hypersensitivity
Describe type 2 hypersensitivity
- mediated by IgG antibodies that recognise antigens on cell surface
- intrinsic or extrinsic antigens - activates classical complement pathway
- destruction of cells (phagocytosis)
How is type 2 hypersensitivity different to type 1
Describe Myasthenia Gravis (example of type 2 hypersensitivity)
Describe type 3 hypersensitivity
Describe equine recurrent airway obstruction (RAO) (example of type 3 hypersensitivity)
Describe canine blue eye (example of type 3 hypersensitivity)
Describe type 4 hypersensitivity
How is TB (type 4 hypersensitivity) diagnosed?
What are the 5 main clinical signs of inflammation
Describe chronic inflammations
Can occur if there is no return to steady state conditions following acute inflammation
Often infectious cause, may be immune mediated
- e.g. chronic colitis/inflammatory bowel disease in dogs
- –> Constant immune output occurs
Granuloma may form –> e.g. enteric granuloma seen commonly in ruminant Johne’s disease (paratuberculosis)
Granuloma may take long time to develop & is e.g. of delayed type hypersensitivity (Type IV) response
give examples of causes of chronic inflammation
Autoimmune disorders
- e.g. diabetes, rheumatoid arthritis, IBD
Exposure to toxins
- e.g. pollution, exposure to chemicals
Chronic exposure to irritant or foreign material
- e.g. foreign body granuloma
Auto-Inflammatory (Hyper-Inflammatory) Syndromes
- Hyper-inflammatory Syndrome in Weimaraner
Infection
- e.g. bacteria:
* e.g. gingivitis, Johne’s disease
Trauma
- e.g. injury
Give some examples of both
Describe granuloma formation
- Macrophages (Histiocytes – macrophages in tissues) fail to clear foreign agent & become chronically infected
- Activated macrophages constantly secrete TNF-α & other proinflammatory cytokines which promote influx of T-cells
- Removing TNF-α causes breakdown of granuloma - T cells around periphery secrete IFN-γ
- Stimulated macrophages can further mature into epithelioid cells
- Epithelioid cells sustain granuloma
- Prolonged stimulation leads epithelioid cells to fuse together & form multinucleated ‘Giant Cells’ or Langhans cells
- Giant cells are capable of phagocytosis & cytokine secretion
- T-cell & macrophages stimulate fibroblasts promoting fibrosis which can lead to organ damage
What are the pros and cons of granuloma formation
pros:
1. Granuloma form focal killing point in which micro-organisms may eventually be killed
2. Granuloma prevent dissemination since it ‘walls off’ micro-organisms
cons:
1. Break-down of granuloma can have serious effects since bacteria can disseminate
2. Granuloma may cause severe tissue necrosis & hence affect organ function
3. Granuloma can cause fibrosis & tissue thickening impairing organ function
What is Chronic enterocolitis in dogs & cats
Chronic Diarrhoea (>14 days)
Causes:
- Idiopathic
- Chronic infection
- Allergic response (e.g. food)
* –> thus, immunosuppressive therapy may be effective
- May be a genetic component
Although granuloma may not be present, cytokines & immune cell infiltrate & maintain chronic inflammatory state
Chronic enteropathy is most common gastrointestinal disorder in older cats, with rising prevalence over past decade
Classified into 4 forms
What are the 4 Forms of chronic enterocolitis in dogs & cats
Describe lymphocytic plasmocytic enteritis (form of chronic enterocolitis in dogs & cats)
Most common form of chronic enterocolitis in dogs & cats (commonly middle aged)
Characterised by increased lymphocyte & plasma cell infiltrate into lamina propria (Ileum & colon common)
In cats it may be difficult to differentiate from small cell lymphoma
Describe eosinophilic (gastro)enteritis (form of chronic enterocolitis in dogs & cats)
Less common that lymphocytic plasmocytic enteritis
No. of different breeds- usually younger animals
Infiltrate is predominantly eosinophils
Chronic parasitic infections/allergy e.g. food.
Describe neutrophilic colitis (form of chronic enterocolitis in dogs & cats) & causes
Neutrophil infiltrate
Possible causes:
- Idiopathic
- Response to normal bacterial microbiota
- Invasion of Campylobacter spp
- Trichomonas foetus infection
What is Hypereosinophilic syndrome in cats?
A variant of eosinophilic enteritis
Overproduction of eosinophils in bone marrow
Widely disseminated eosinophilic infiltration of multiple organs
Organs commonly affected include:
- Bone marrow
- Small intestine
- Liver
- Spleen
- mesenteric & peripheral lymph nodes
- Skin lesions
Poor prognosis
Describe granulomatous colitis (form of chronic enterocolitis in dogs & cats)
Rare - Seen in young Boxer dogs, French bulldogs
Thickened, partially obstructed segment of bowel (ileum & colon most commonly)
Large foamy, Periodic acid-Schiff (PAS) +ve macrophages present in colonic mucosa (pro-inflammatory macrophage type)
True granulomas are not present.
Caused by E. coli
What are the methods of antigen sampling in gut
How are lymphocytes recruited to peripheral lymphatic organs such as GALT?
Lymphocytes leave LN, enter blood, then emigrate back from capillaries to intestine
Addressins (adhesion molecules) on high endothelial venules interact with homing receptors on lymphocytes to ensure they reach mucosal surface
How is bacterial DNA recognised in mammals
Bacterial DNA CpG dinucleotides are un-methylated
Most mammalian CpG di-nucleotides are methylated
TLR-9 in mammals recognises un-methylated CpG in bacterial DNA & initiates inflammation
Describe bacterial chromosomes & plasmids
Describe plasmids
Typically circular
DsDNA
Separate from the chromosomal DNA
Some have a suicide gene to ensure maintenance
Some are transmissible by conjugation
Variable in sizes
Describe bacterial replication
Binary fission
They have a generation time
- Length of time required for single bacterial cell to divide to 2 daughter cells
Binary fission leads to clones of bacteria
If bacterial replication leads to clones how do bacteria evolve?
Mutation of genes
Transfer of genes
Rearrange their DNA
Delete genes
New genes inserted
What are the methods of transferring genes between & within bacteria
Introduction of new DNA into and between bacteria:
- Transformation
- Conjugation
- Transduction
- Recombination
Movement of DNA already in bacteria:
- Transposition
- Recombination
Describe transformation in bacteria
Transformation is the uptake of naked DNA
DNA contacts the bacteria and is taken up
Some bacteria are…
- Naturally competent. They can take up DNA without treatment. Have competence mechanisms
- Require treatment to become competent (laboratory competent)
Once naked DNA is taken up into cells there are two options:
- It is degraded by nucleases
- It recombines into the genome
Describe conjugation in bacteria
Bacterial conjugation is transfer of genetic material between bacteria through direct cell-to-cell contact (horizontal gene transfer)
It’s transfer of genetic information from a donor cell to a recipient
donor must host conjugative or mobilisable genetic element (plasmid or transposon)
Most conjugative plasmids have systems ensuring that recipient cell doesn’t already contain similar element
Describe transduction by bacteriophages
Viruses that infect bacteria are known as bacteriophage
Each virus is specific for a specific species of bacteria
They need to bind (specific receptor)
Then inject their genome and this infects the bacteria
What are the types of bacteriophage & transduction
Lytic and generalised transduction
Temperate phage and specialised transduction
Describe generalised transduction by lytic bacteriophage
Process by which any bacterial DNA may be transferred to another bacterium via a bacteriophage
Describe specialised transduction by lysogenic bacteriophage
Lysogenic (viruses that insert their DNA into bacterial chromosome)
Bacterial genes that get transferred depend on where phage genome is inserted on chromosome
When prophage excises it may do this imprecisely or package adjacent bacterial DNA
These packaged particles can then infect another bacteria with bacterial DNA
What happens to DNA that enters bacteria
Degraded – non specific (no recombination general nucleases degrade DNA)
Degraded – by specific restriction endonucleases (DNA restriction)
It manages to integrates with genome (DNA recombination)
Describe Homologous, directed & non-homologous recombination/insertion
Homologous recombination - Where 2 DNA sequences are same they can intercalate their strands. During replication these can get spliced
Non homologous – less efficient, DNA doesn’t match but can be in random positions
Directed - Insertion elements have repeats which lead to homologous/directed insertions in specific sites or enzymes that direct integration at set sites
- These have hot spots for insertions & are more effective
What is DNA restriction in bacteria
It’s not good for bacteria to constantly take up DNA so they have restriction system
They have Restriction endonuclease(s)
Which degrade DNA not made in their own cell
These restriction nucleases don’t degrade self-DNA as they act on specific sites
Bacteria methylates its own DNA where its own restriction nuclease cut so blocking enzyme (They don’t chew up their own genome)
Works on any DNA including non-self plasmids
What are frameshift mutations
Either addition or loss of single base moving reading frame in an out of position
Frame shift adds or removes base to change codon position relative to reading frame
What is transposition
Transposons are sequences of DNA that can move around to different positions within genome
Transposons encode their own transposases
Transposase cut out & re-insert the DNA flanked by inverted repeats (IR)
The process directed by these enzymes is transposition
They pop into (integrate) and out of DNA
If they insert into a gene they will disrupt it
What are insertion sequences
IS elements are a type of transposon. They are….
- short DNA sequence that acts as simple transposable element
- are small relative to other transposable elements
- only code for proteins implicated in the transposition activity
- are different from transposons, which carry accessory genes
What is the clinical relevance of mutations
If a mutations is beneficial to bacteria this will become more fit
Alteration of antibiotic binding sites
- E.g. mutations ribosomal binding sites can reduce binding of ribosome targeting drugs
Avoidance of vaccines
- If mutations alters amino acid sequence of epitope that antibodies bind to then this can reduce or abrogate binding
Alteration of metabolism
- Can be no. of reasons mutations in promotors can upregulate or down regulate critical functions. Making bacteria less or more competitive
Define genetic drift
Changes of over time by mutations of genomes
What are antimicrobial growth promoters and what is the problem with them
substances added to the feed or water of animals to promote growth and improve feed efficiency
Prolonged and large scale use will select for antibiotic resistance
Leading to a reservoir of transferable antibiotic resistance
Banned in UK and EU
Describe bacterial metabolism of xenobiotics (drugs)
Gut flora bacteria can transform (alter) compounds. This can have different effects
- Drugs activated
- Drugs inactivated
- Toxic metabolites (and carcinogens)
Different compounds in food can also alter composition of gut flora
- Can lead to increased bacteria associated either with ‘improved’ or ‘reduced’ gut health
What are methods used to enumerate bacteria
Qualitative
- General assessment of level low, med, high
Quantitative
- Enumeration normally vs. volume or weight
Visual examination
- microscopy total count or general assessment
Culture:
- Viable counts where you dilute count colonies & calculate original no. of bacteria
Molecular detection
- Detecting species by quantitation of their nucleic acid
What are methods for assessing presence of bacteria
Slides/smears
- May work for some very characteristic spirochetes but rods & cocci can’t be distinguished in faeces
- Useful in discrete normally body sites/tissues where you would expect sterility or only single common morphologies (you can spot abnormal types or mixed infections)
Culture:
- For faeces, the most common method is culture.
What can impact gut flora
Age
Stress
Use of drugs
Genetics
Diet
Illness
What genes are essential for bacterial colonisation
Surface structures, for adhesion and stop wash out
LPS and membrane integrity for tolerance of harmful factors
Motility to get to target niches
General gene regulation for stress responses to turn on genes required to adapt to GI-environment
Specific transporters to take up GI-specific nutrients
What is natural competitive exclusion and competitive exclusion products
“Natural” competitive exclusion: Existing bacterial flora present competitive barrier to incoming bacteria so can exclude & prevent colonisation &/or infection
Competitive exclusion products: Deliberate use of desired microbial cultures to colonise animals that out-compete pathogens from colonising specific niches (so can exclude & prevent colonisation &/or infection)
- Applied commercially
Describe competitive exclusion
Presence of one bacteria inhibits other due to it competing & winning in acquisition of growth factors
‘Nutrition’ is NOT free
There is Competition (for….)
- Space
- co factors elements etc..
- Electron acceptors (Humans Oxygen)
- Carbon sources
Only compete if they require same type of nutrition
Describe how antibiotics can disrupt flora
Antimicrobials can remove sensitive organisms from a particular niche
Space left is then repopulated by intrinsically resistant bacteria
Thus antibiotics can disrupt existing flora or allow over growth of undesirable organisms
Name viruses that cause major clinical signs associated with GI tract
What are key features of parvoviruses
What are parvoviruses of veterinary importance
Describe feline parvovirus infection
also known feline infectious enteritis (FIE) or feline panleukopenia
faecal-oral transmission
infects lymph nodes of naso- & oro-pharynx & then spreads to other tissues
- needs rapidly dividing cells to propagate
then infects intestinal cells & bone marrow
kittens most susceptible
Describe parvoviruses pathogenesis
panleukopenia, enteritis & cerebellar hypoplasia
What are the clinical signs & treatment of feline parvovirus
How is feline parvovirus controlled
How is feline parvovirus diagnosed
Describe canine parvovirus 2 infection
Describe corona viruses
enteric & respiratory pathogens
commonly mutate
Describe transmissible gastroenteritis virus (TGEV) (porcine corona virus)
Describe porcine epidemic diarrhoea virus (PED) (porcine corona virus)
Describe Porcine delta coronavirus (PDCoV) (porcine corona virus)
Describe swine acute diarrhoea syndrome coronavirus (SADS-CoV) (porcine corona virus)
Describe betacoronaviruses (bovine corona virus)
Describe canine coronavirus
Describe feline coronavirus
What are the 2 main forms of feline infectious peritonitis
wet FIP
- effusions
dry FIP
- pyogranulomatous lesions
What might lead you to suspect a case of wet FIP (feline infectious peritonitis)
What are the features of pestiviruses
Describe bovine viral diarrhoea virus (pestivirus)
Causes:
- diarrhoea
- decreased fertility/milk yield
- abortion/congenital defects
- immunosuppression
- mucosal disease
2 genotypes:
- BVDV 1 - classical form of disease
- BVDV 2 - haemorrhagic syndrome (severe acute BVD)
Describe epidemiology of mucosal disease (BVDV)
BVDV can cross placenta
infection during pregnancy may have various effects on foetus
- if infected before development of foetal immunocompetance calf aborted or PI calf
- if infected after calf aborted or born with congenital defects
mucosal disease only develops in PI (infected in utero) animals around 2 years of age
- invariably fatal
How is BVD (bovine viral diarrhoea) controlled
Describe the structre of paramyxoviridae
describe rinderpest virus (Morbillivirus) & its clinical signs
cattle plague
highly infectious
respiratory & alimentary tract disease
high mortality
clinical signs:
- nasal discharge
- pyrexia
- oral & nasal erosions & ulcerations
- diarrhoea with mucus, blood & debris
- dehydration
describe canine distemper (morbilillivirus)
young dogs
transmitted by direct contact
replicates in urinary tract
spreads to tonsils & lymph nodes & then CNS
clinical signs:
- pyrexia
- depression
- ocular & nnasal discharge
- vomiting
- diarrhoea
- hyperkeratosis of nose/pads
- neurological signs if poor immune response
Describe Newcastle disease
What are some strains of Newcastle disease
What are viral membrane glycoproteins
Compare lentogenic & mesogenic strains of Newcastle virus
How is Newcastle disease diagnosed
- egg inoculation
- harvesting allantoic fluid after 7 days
- Haemagglutination test to detect viruses in allantoic fluid
How is Newcastle disease controlled
acid-fast staining
Describe method of staining Mycobacteria (acid-fast staining)
How long do Mycobacteria take to grow & what problem does this pose for diagnosis?
slow (2-16 weeks)
takes very long to culture so disease can spread while waiting
what is Johne’s disease
What are the clinical signs of Johnes disease?
What are the clinical characteristics/pathogenesis of Johne’s disease?
- MAP colonises in intestinal epithelial cells by attaching to surface receptors
- body forms granulomas to attempt to contain infection
- causes disruption of normal digestive functions & leads to malabsorption
- intestinal mucosa becomes damaged which leads to diarrhoea
- contributes to weight loss
Describe oesophageal infections
Relatively uncommon due to rapid passage of material through oesophagus & tough stratified epithelium
Some viral infections can cause ulcers
Most notable infections:
- BVDV (Bovine viral Diarrhoea virus) & mucosal disease (Ovine Herpes virus 2)
- Newcastle disease in poultry
How can infections spread from GIT to other body systems
Liver & spleen, GI Tract is a route for infection
1. Portal vein
2. Hepatic artery (billiary system)
3. Contagious spread from adjacent system
Describe Campylobacter jejuni
Zoonotic
Associated with diarrhoea in dogs
Gram negative
Flagellated and Motile
Microaerophilic
Optimum growth temperature for C. jejuni is 42ºC.
Grown on nutrient media or isolation using Campylobacter selective medium
Can be seen as commensals in intestinal tract of warm-blooded animals
The genera contains a number of species
- C. jejuni, C. coli, C. lari
Important because it is a common contaminant of meat
How can GI tract diseases enter food chain
How can campylobacter be identified (sample collection)
Samples depending on stage of production
On farm:
- Environmental samples
- Faecal swabs
- Boot swabs
At abattoir:
- Gut contents
- Neck Swabs from birds
- Skin samples
At retail:
- Swabs
- Juice from meat packages
- Whole meat samples
What is Multi Locus Sequence Typing (MLST)
MLST looks at single base pair changes
Done by sequencing 7 housekeeping (essential) genes & clustering strains based on differences
These clusters are called clades
Researchers have collected lots of strains from different sources & compared their MLS
So if clinical isolate clusters in MLST clade it indicates potential source of infection
MSLT systems exists for range of bacteria
They get developed where there is clinical need to understand sources of infection
Describe campylobacter in birds
Mainly infects lower intestinal tract
There is an inflammatory response during initial colonisation
A correlation with Campylobacter colonisation and
- reduced weight gain
- Hock burn (which is associated with wet bedding i..e more watery faeces)
Are campylobacter sp. Clear pathogen in dogs?
Diarrhoeas in dogs & other domesticated animals has been associated with Campylobacter
However, Campylobacter is often found in healthy animals so what does this mean?
Presence of large number of Campylobacter may indicate infection
In older dogs evidence is unclear for campylobacter causing diarrhoea but there is some evidence in puppies
It may be that disease is associated more with young or debilitated animals which are often increased at risk
How can brachyspira (spirochete that causes disease in pigs & poultry) be diagnosed
Selective media - Blood agar plus selective antibiotics
Stained faecal smear for spirochaetes
Silver stain faecal smear
PCR tests
What is Lawsonia and how can it be diagnosed
Curved Gram-negative rod
Obligate intracellular pathogen
Microaerophilic
In piglets & foals
Diagnosis:
- Clinical signs
- Demonstration in mucosa or faeces by immunofluorescence or PCR
- Staining of tissue sections
- Serological tests
Describe Lawsonia intracellularis
Invades enterocytes
Causes lack of differentiation
Undeveloped cells do not shed
Leads to pseudo-stratified epithelium
- Thickening of gut & thus poor absorption
What are enterobacteriaceae
Gram negative rods which are oxidase negative, facultative anaerobes & grow on MacConkey
Major enteric pathogens
- Escherichia coli (also commensal)
- Salmonella enterica
- Yersinia species
Opportunistic pathogens
How can you differentiate between different enterobacteriaceae
Differentiation by growth characteristics and biochemistry
- Motility Test
- Citrate utilization Test
- Indole Test
- Methyl Red (MR) Test
- Voges–Proskauer (VP) Test
- Triple Sugar Iron (TSI) Agar Test
- Urease Test
More common now for molecular tests to confirm species by:
- PCR of unique genes
- Whole genome sequence (WGS)
- Ribosomal genome sequence
- Mass Spectrophotomerty
Describe E. coli
Most members of this genus are low virulence
They may cause opportunistic infections
Pathogenic strains possess virulence factors which allow them to cause disease
Basic genomes of E. coli are same but they will have gained some very specific properties.
3 main criteria
- Adhesion factors
*Presence or absence of LEE island
- Type of enterotoxin present
- Species associated with
What factors can lead to pathogenic E. coli in young farm animals
Poor colostrum immunity
ETEC receptors present
Build-up of pathogenic E. coli
Stress and environment
What are the 2 species of Salmonella
S. bongori
S. enterica
What are the sources of Salmonella infection
Organisms found in:
- Contaminated water sources
- Carrier animals
- Faecal contaminated environment
Foodborne
- Eggs (from infected birds)
GI content contaminated food:
- Raw Meat
- Offal
- Animal Feed
- Plant material (Faecal contamination and uptake)
What are clinical signs of Enteric Salmonella
Profuse foul-smelling diarrhoea
Fever
Depression
Anorexia
Severely affected animals may become recumbent
Describe Salmonella invasion
- TTSS-1 injects effector proteins into host enterocyte
- Triggers actin changes that leads to engulfment of bacteria into vacuole
- There are multiple TTSS-1 effectors but overall effect of different effector proteins is to
- Trigger invasion by actin and tubulin remodelling
- Trigger cytokine response
- trigger Enteropathogenic response (fluid secretion) - Once bacteria is internalised in vacuole, host will try & attack bacteria moving antimicrobial vacuoles to form phagolysozome out of Salmonella containing vacuole.
- These fusion is inhibited by TTSS-2
Describe Salmonella intracellular survival (in vacuole)
- TTSS-2 injects SPI-2 effectors
- These jam up in cytoskeletal movements in cell so cell can’t mature vacuoles
- This means Salmonella is in protected space
- Salmonella replicates in controlled vacuole
- In this space it’s safe from adaptive responses as antibodies only survey outside cells
Describe Septicaemic salmonellosis
Common to specific host serotype combinations
Host may vary in susceptibility with age
Sudden high fever, depression, recumbence, rapid death
Mucosa is route to deeper tissues Salmonella can spread in blood & lymph inside macrophages
Organisms can spread to other organs liver & spleen (oviduct in birds)
Describe Salmonella in eggs
Carrier state observed after recovery from enteric infection of
- S. Enteritidis
- S. Gallinarum
Bacteria found in macrophages
These reach oviducts and contaminate eggs
Biosecurity & vaccination have reduced infection & egg contamination
FSA guidance is that Lion Brand Eggs (or equivalent) in UK are now considered Salmonella ‘free’
Describe phylum firmicutes
The ‘majority’ Gram positive
Found in various environments
The group includes commensals and pathogens
BUT exceptions
- Mycoplasmas lack cell walls (e.g. of genetic reduction)
Some Gram positive genera:
- Produce endospores
- Have porous pseudo-outer-membrane that causes them to stain Gram-negative
Give examples of Firmicutes
Describe phylum bacteroidetes + e.g.
Gram negative
Wide distribution in the environment, (soil, water GI tracts)
Rarely pathogenic
examples:
- Bacteroides (abundant in the faeces)
- Porphyromonas, (oral cavity)
Describe phylum proteobacteria
All Gram Negative
A major Phylum (with subgroups)
- Alphaproteobacteria
- Betaproteobacteria
- Gammaproteobacteria
- Deltaproteobacteria
- Epsilonproteobacteria
Includes commensals and pathogens
Members can be facultative/obligate anaerobes or even microaerophilic
Notable Genera clinically relevant to GI tract:
- Escherichia, Salmonella, Vibrio (in humans), Helicobacter, Campylobacter
Name an important family of gammaproteobacteria
Pseudomonas found in lots of environments & can cause ear infections, burn infections, wool/skin in sheep infections
It’s good at forming biofilms in infections as it often produces exopolysaccharides (slime made form sugars)
These films can make it more intransigent to treatment
Name important epsilonproteobacteria
Both are microaerophilic
Helicobacter is important in stomach infections
Campylobacter causes GI infection in humans & is foodborne zoonotic as its carried by number of large animals
Give examples from phyla actinomycetota
Describe Bifidobacterium
Gram Positive, Anaerobes
Can be branched
Common commensal bacterium
Niche gastrointestinal tract of mammals
A major constituents of probiotic food supplements
Describe Genus bacteroides
A major component of mammalian gastrointestinal flora
Gram negative, bacilli (morphology as in rod), anaerobes
Non-endospore-forming
Variable motility between species
Will use simple sugars when available, but main source of energy is polysaccharides from plant sources
Describe Genus Lactobacillus
Gram Positive
Facultative anaerobic / microaerophilic bacteria
Major part of group can convert lactose & other sugars to lactic acid
Common and benign
Present on mucosa of reproductive and GI tract
Production of lactic acid makes its environment acidic which inhibits growth of some harmful bacteria
This ‘MAY’ help protect the gut against less pH tolerant bacteria
Describe Genus streptococci
Gram Positive, Coccoid, facultative-anaerobes
Members of phylum Firmicutes
Cellular division on a single axis so they grow in chains or pairs
Found in many body sites as commensals
Can cause opportunistic disease
Describe Enterococcus sp. (faecalis & faecium)
Gram Positive, Facultative anaerobe
Commensal bacterium
Niche gastrointestinal tracts mammals
A major constituents of some probiotic food supplements
Can cause opportunistic infections
One of the ESKAPE pathogens
What are ESKAPE pathogens
6 nosocomial pathogens that exhibit multidrug resistance & virulence
Enterococcus faecium, Staphylococcus aureus, Enterobacter Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa & spp.
Describe genus Clostridia
Gram Positive, Endospore forming, Rod shape, obligate anaerobes
What is genome
Genetic material of animals, plants & bacteria consists of DNA
Genome includes both genes (coding regions) & noncoding DNA
In viruses genome can be double stranded or single stranded RNA or DNA
Describe mitochondrial DNA (eukaryotes)
In animals mitochondrial genome is typically single circular chromosome that is approx. 16-kb long & has 37 genes
Not all proteins necessary for mitochondrial function are encoded by mitochondrial genome
Mitochondria divide by binary fission similar to bacterial cell division
Mitochondria & thus mitochondrial DNA, usually comes from egg only
Sperm’s mitochondria enter egg but don’t contribute genetic information to embryo – Maternal inheritance
define microbiome
Combined genetic material of microorganisms in particular environment – population level
What is metagenomics
study of structure & function of entire nucleotide sequences in bulk sample
Describe the intestinal microbiome
Gut composed of widely divergent microbial lineages whose genomes contain functionally similar sets of genes that would give rise to coordinated single metabolic outcome
Diversity & abundance level of microbes, genes, proteins & metabolites will influence energy balance, gut motility, inflammatory tone, mucosal integrity, appetite & signalling
Gut key player (i.e., pathogens) may also negatively influence gut barrier
- –> promoting inflammation
Good flora can produce good products & also exclude detrimental bacteria
Detrimental bacteria may produce toxins or interact with cells stimulating unwanted responses in gut mucosa
Describe Influence of Microbiota gut brain axis (MGBA) on behaviour
via cell structural components (ie LPS) & with release of microbial metabolites which act on intestinal epithelium, then released into bloodstream & cross blood-brain barrier
production of pro-inflammatory cytokines by immune cells
stimulate enteric nervous system & its sensory neurons or induce secretion of neuropeptides by entero-endocrine cells
Affects:
- anxiety-like behaviour
- memory capacities
- social behaviour
- feeding behaviour
What are coding regions of a gene
portion of gene’s DNA or RNA that is composed of exons
What are Long interspersed nuclear elements & short interspersed nuclear elements
Long interspersed nuclear elements (LINEs):
- expressed as RNA & encode reverse transcriptase that creates DsDNA copy that can then insert
Short interspersed nuclear element (SINE) sequences:
- Widely distributed in eukaryotic genomes & have crucial roles in genome organization, genome evolution & modulating gene expression
What are the Applications of PCR & qPCR in veterinary medicine
pathogen detection
quanitification of gene expression
detection of gene mutations
Why might diarrhoea occur more commonly in younger animals
not very developed immune system
GI tract not fully developed
Incomplete gut flora
lack of exposure
exploring environment with mouth
How might an animal ingest viruses shed within faeces
Viruses shed in diarrhoea can create aerosols
foodstuffs, drinking water & fomites can be contaminated & spread disease
poor disinfection of environment
Define fomite
objects or materials which are likely to carry infection
Describe common features of enteric virus infections
How can enteric virus infections cause malabsorption
What are the 4 main mechanisms of viral diarrhoea
What are the main viruses that affect GIT
Describe rotaviruses
What is important about viruses with a segmented genome?
have their genetic material divided into separate segments (each encoding for different genes)
allows for genetic diversity (each segment can undergo reassortment to create new viral variants)
increases adaptability to survive in different conditions
some have antigenic variation (encode for proteins that undergo frequent changes to avoid immune system)
Where do rotaviruses infect & effect
What is the importance of rotavirus
How is rotavirus diagnosed
How is rotavirus controlled
What are the steps of PCR
- reaction mix is heated to 95 degrees
- hydrogen bonds of DNA nucleotides are broken down
- double stranded DNA will be denatured
- reaction mix is cooled down to annealing temperature
- primers align with complementary sequence of target DNA
- primers form hydrogen bonds with target DNA
- reaction mix is heated to reach optimum temperature for DNA polymerase
- heat stable DNA polymerase extends primers in 5’->3’ direction
- temperature raised to 95 degrees again to start next cycle
Compare microbial culture to PCR
Compare PCR to immunodiagnostics (ELISAs & snap tests).
1. What do they detect?
2. How quick are they?
3. How accurate are they?
- PCR detects pathogen DNA & immunodiagnostics detect Ab that animal produced when in contact with pathogen
- snap tests immediate. PCR & ELISA 24-72h
- PCR more accurate
Define silent or synonymous mutation
define missense mutation
nucleotide substitution that leads to change of an amino acid
Impact can vary:
- new amino acid might have similar properties to the old one or the substitution is in a part of the protein that does not affect its structure or function and has no impact
- It can cause a major change in the protein
Define nonsense mutation
A point mutation that changes a codon for an amino acid into a stop codon and causes the premature termination of translation. If that happens close the 3’ end of the strand than that might not have much impact on the proteins.
Most non-sense mutations lead to non-functional proteins.
Which organism causes Johne’s disease
How is canine coronavirus transmitted
Define diarrhoea
Increase in frequency, fluidity, volume & water content of faeces
What factors can cause disease
Secretion of exotoxin
Invasion of target cells leading to cell death
Invasion of host cells & lymphatic’s leading to systemic disease
What is hypersecretion (pathogenic mechanism)
A functional disturbance of the intestinal epithelial cells
Increased out flow or decrease re-absorption of water
crypt cells can pump chloride ions (Cl-) into the crypt space
Cl- ions attract sodium ions (Na+) into crypt space, increasing local osmotic pressure
As osmotic pressure increases, water is pulled into intestine
Some enterotoxins can trigger & lock ON chloride secretion leading to hypersecretory diarrhoea
Done by activating adenylate cyclase
Example cause:
- ETEC (enterotoxigenic E. coli)
What are examples of pathogenic mechanisms
Hypersecretion
Villous atrophy
Infiltrative and proliferative distortion of mucosa
Mucosal necrosis
What is villus atrophy (pathogenic mechanism) + example pathogens
Destruction of the epithelial cells on villi or in the crypts
Villi become often stunted or fused
Example pathogens:
- Attaching and effacing E. coli (EHEC, VTEC)
- Rotavirus (mild villous atrophy)
- Canine parvovirus (severe villous atrophy)
What is infiltrative & proliferative distortion of mucosa (pathogenic mechanism)
Recruitment of large numbers of Macrophage & T-lymphocytes in lamina propria & sub-mucosa
Leading to crypt compression and villous distortion
Therefore reduce absorptive capacity
Example Paratuberculosis (Johne’s disease)
What is mucosal necrosis (pathogenic mechanism)
Infection leads to cell death
Severity and extent depend on virulence of pathogen
Majority of Diarrhoea from reduced absorption
Example pathogens:
- Brachyspira hyodysenteriae
What are the advantages & disadvantages of preventative vaccination
