barbic Flashcards
1
Q
reservoir/carrier definition
A
a reservoir is any living or non-living area where a pathogen naturally lives, grows, and multiplies. They’re long-term hosts, so can provide persist environment for them to live and spread to susceptible hosts via direct/vectors
o It harbours a specific infectious agent and shedding it in at least minimum infective dose without observable clinical disease
2
Q
types of reservoir
A
o Animal Reservoirs: Animals that harbour pathogens capable of infecting humans or other animals. Many zoonotic diseases originate from animal reservoirs (eg Rodents - reservoirs for causing plague)
o Human Reservoirs: Humans can also serve as reservoirs, either through symptomatic or asymptomatic infections. (Eg Humans are for M. tuberculosis = tuberculosis)
o Environmental Reservoirs: Certain pathogens can live in soil, water, etc (eg Soil reservoir C.tetani = tetanus_
3
Q
latent infection - example +
A
- A latent infection in animals occurs when a pathogen, persists in the host’s body in a dormant or inactive state after the initial infection. (doesn’t cause active disease symptoms)
- Some situ’s: such as stress, immunosuppression, etc , the pathogen can reactivate, leading to an active infection and symptom recurrence.
feline, bovine and equine herpes virus
4
Q
passive immunity
A
- Natural (colostral)
- Artificial (passive immunisation)
- Passive immunisation and passive immunotherapy = transfer of antibodies from one individual to another, providing immediate, short-term immunity. Doesn’t involve recipients immune response and antibodies are eventually degraded (unlike active immunity which is developed by themselves by exposure or vaccination)
5
Q
classification of passive immunity
A
- Classification: homologous, heterologous, immune system, hyperimmune serum, gamma globulin products, polyclonal and monoclonal
o Immune serum (Homologous antibody,)
o Homologous hyperimmune serum (homologous hyperimmune globulin )
o Heterologous hyperimmune serum.
6
Q
colostrum
A
- crucial for neonates especially: dogs, cats, and horses, where MDA don’t pass through placenta so rely on colostrum to get antibodies
- colostrum’s very rich in immunoglobulins (IgG)nutrients, and growth factors. Provides passive immunity by coating the gut and entering bloodstream of newborn. Anything mother has been exposed to, baby will be immune too
o IgG (systemic immunity), IgA (mucosal immunity) and IgM - Newborn animals have specialised cells in their intestines that can absorb large antibody molecules directly into the bloodstream.
o Only for 12-24 hrs after birth, highest after the first few hours, after they lose ability to absorb and anymore consumed are broken down in gut and don’t go to blood - The success of passive immunity depends on several factors:
o Timing, quality (varies on health, age and vaccination status), quantity, health of neonate, whether the area is clean and stress free
7
Q
tuberuclinisation
A
- Allergolocial (immunological) method
- Gold standard for tuberculosis
- Based on: during infection, animal becomes hypersensitive to protein antigen (late phase) = cellular type hypersensitivity type 4
8
Q
bovine tuberculinisation - individual skin test
A
- Shave area of 5cm2 on left side of neck
- Measure thickness of skin folds with cutimetre
- Apply tuberculin (purified protein derivative), 0.1ml intradermally
- Check by palpation if correctly applied, if wrong 10-15cm cranially
- Shouldn’t be done on in cows with immunosuppressive drugs
- Check 72hrs after: clinical exam of infection site + regional LN + measure skin thickness
- -ve reaction = only limited swelling, without clinical signs, increased skin fold <2mm
- Doubtful reaction = no change of LN + no clinical sign, increased skin fold 2-4mm
- +ve reaction = inflamed LN + site of application, clinical signs, increased skin folds >4mm
- if doubtful/+ve reaction = comparative intradermal test (earliest 42 days after)
9
Q
Bovine – comparative intradermal test
A
- right side of neck, cut hair with scissors in 2 areas 4-5cm2 + measure skin fold
- area of avian tuberculin hair is cut about 10cm from ridge of the neck and avian tuberculin is 15cm lower, in a line parallel with line of shoulder
- apply tuberculin avian 0.1ml intradermal, bovine 0.1ml, 15cm lower than avian
- same signs as the individual test
- doubtful/+ve reaction = culling
10
Q
why use avian and bovine tuberculin
A
- Differentiation of Infections: The use of avian tuberculin helps distinguish between bovine tuberculosis and infections caused by Mycobacterium avium. This differentiation is crucial for accurate diagnosis and management of the diseases.
- Cross-Reactivity: Cattle can show cross-reactivity to both bovine and avian tuberculin. By using avian tuberculin, veterinarians can identify animals that are specifically infected with Mycobacterium avium, thus avoiding misinterpretation of the results.
- Regulatory Compliance: In many regions, regulations require the use of avian tuberculin as part of a testing strategy to ensure that cattle herds are free from tuberculosis and to control the spread of disease.
- Public Health: Ensuring accurate diagnosis in cattle populations helps protect public health, as bovine tuberculosis can be transmitted to humans through contaminated milk or meat.
11
Q
antibody titres
A
- Antibody titres are a measure of the concentration of antibodies in the blood, indicating the immune response to a particular antigen or infection. They are often expressed as a dilution factor, representing the highest dilution of serum at which antibodies can still be detected.
- antibody titres in various immunological methods:
o ELISA: This method quantifies antibodies in serum. Results are often expressed in units or as titres (e.g., 1:100, 1:200), indicating the dilution at which antibodies are still detectable.
o HA Test: used for viral infections (e.g., influenza). The titre is the highest dilution of serum that prevents agglutination of red blood cells, indicating the presence of specific antibodies
o Neutralisation Tests: Used for viruses and toxins, the titre is the highest serum dilution that still neutralizes the pathogen’s effects, showing protective antibody levels.
o Western Blot: While not typically reported as a titre, positive results indicate the presence of specific antibodies against particular proteins, often following an initial screening test like ELISA.
o Immunofluorescence Assays (IFA): Antibody titers can also be determined through IFA, where dilutions of serum are tested against cells or tissues to assess the highest dilution showing fluorescence.
12
Q
types of vaccine
A
- Infectious (live)
o Attenuated, Modified, Recombinant vector - Non-infectious (killed)
o Whole pathogen, Subunit, Conjugated, Toxoid, DNA/RNA
Vaccination - Core vaccines
o ALL dogs and cats, regardless of circumstances or geographical location should receive
o CATS:
Feline parvovirus (FPV)
Feline calicivirus (FCV)
Feline herpesvirus -1 (FHV-1)
Rabies?
o DOGS:
Canine distemper virus (CDV)
Canine adenovirus (CAV)
Canine parvovirus type 2 (CPV-2)
Rabies? - Non-core vaccines
o DOGS:
Leptospirosis, Lyme disease, Parainfluenza, Bordetella bronchiseptica
o CATS:
FeLV, Chlamydia felis, Bordetella bronchiseptica - Not recommended vaccines
o Canine coronavirus (CCV), giardia, leishmania, piroplasmosis
o FIV, FIP
13
Q
primary SOI
A
- Where microorganisms exist before outbreak of infectious diseases
- Individual animal, SOI is place where at least one minimal infectious dose of pathogen is present
- Sick animal as primary source of infection
- Carrier animal as a source of infection
- Sick or carrier man as a source of infection
- Animal products (sick or carrier animal)
- Animal by-products (sick or carrier animal)
- Animal carcasses
- Wild animals (vertebrates)
- Arthropods
14
Q
arthropods as SOI
A
Vector-borne diseases (equine infectious anaemia)
o Increase in incidence, due to increase in vector populations due to the environmental change
o In TBE, west Nile – arthropods have a central role due to multiplication of pathogen in vector
ow Transovarial transmission occurs in certain arthropod vectors
o Transstadial transmission – pathogen remains with the vector from one life stage to the next
15
Q
wild animals as SOI
A
o Important SOI for many zoonoses
o High potential for spreading infectious diseases due to long-range migration or high population density
16
Q
carcasses as SOI
A
o Death/sacrifice of animal that had infectious diseases leads to inactivation of pathogen
o Inactivation depends on pathogen nature, duration of exposure, environmental conditions etc
o After a certain time, carcass is auto-sterilised with respect to pathogen
Carcasses are important SOI for pathogen that create spores (anthrax) and some viruses (African swine fever, etc)
17
Q
animal by-products as SOI
A
o Humans and animals don’t consume but it goes to rendering plants
o Anthrax, BSE
18
Q
animal products as SOI
A
o Human/animal eating the infected meat/milk/etc can contaminate them
o TB, brucellosis, q-fever and listeriosis
19
Q
sick or carrier man as SOI
A
o Zoonoses are any disease or infection that’s naturally transmissible from vertebrate animals to humans
o Reverse zoonoses is humans to animals
o Transmission is from prolonged exposure to pathogens or closer social contact and poor hygiene conditions
20
Q
sick animal as SOI
A
o Sick animal = displays clinical symptoms (as a result of pathogen multiplication)
o Shedding multiplied pathogens, sick animal = SOI for other susceptible animals
o Infectious period – time interval during which host is capable of directly or indirectly transmitting pathogens to another susceptible host
21
Q
secondary SOI
A
- Environment and objects (inanimate) contaminated with the pathogen
- Contamination occurs through intermediate sources of infection
- Soil
- Water
- Air
- Food (forages)
- Contaminated objects
- Contaminated premises (spaces)
- Contaminated means of transport (vehicles)
22
Q
soil as SOI
A
o Permanent SOI for anthrax, malignant oedema, black leg and tetanus
o Temporary SOI for foot-rot in sheep and listeriosis
o Soil is important SOI for pathogens resistant to environmental conditions
23
Q
water as SOI
A
Standing waters can be a SOI as it’s easier to achieve larger amount of pathogen in it
o SOI in water = leptospirosis
24
Q
air as SOI
A
o Any infectious disease that’s transmitted by air
o Aerosol (suspension of tiny particles/ droplets in the air) generated by primary sources (coughing/sneezing)
o Air is SOI for limited time due to precipitation of the droplets into dust. Dust can be blown up again and make the air contaminated from secondary SOI (ground)
o Q-fever
25
food as SOI
Caused by contamination of food and occurs at any stage of food production chain
o Animal products originating from infected/carrier animals used for feeding could be SOI (aujeszky, African swine fever, classical swine fever)
o Occurs in 2 ways:
Contamination on the field where it’s cultivated (anthrax)
Contamination during preparing and storage (listeriosis, botulism)
26
epizootiology
- = science that studies origin, distribution, frequency, development and extinction of epizooties (why, where, when and who)
27
epizootic (epidemic)
= outbreak of animal disease, temporarily prevalent and widespread among animal population
28
enzootic (endemic)
= prevailing among or afflicting animals in a particular locality at any time, without tendency of spreading widely
often affecting animals in a particular place
29
panzootic (pandemic)
= occurs in a widespread outbreak among a large number of animals, usually affecting more than one species
30
sporadic
= occurring occasionally, singly or in irregular or random
31
microbiological method
- Based on principle of objective proof and identification of causative agent in diagnostic material taken from a sick animals, or from tissue samples from a dead animals
- comprise isolation and growth of the causative agent in optimal conditions
- Microbiological methods during life
- Microbiological methods after death
- Direct proof of the causative agent
32
what is parameters for microbiological analysis
- parameters per gram, this usually refers to the colony-forming units (CFUs) per gram of the sample. It quantifies the number of viable bacteria present in a sample and helps in determining the bacterial load.
33
what are the 3 conditions that needs to be met for koch's postulates?
three basic conditions must be met:
* the pathogen must be found in all changed tissue, and it must not be found in other infectious diseases
* the pathogen must be isolated and cultured in a pure culture
* Inoculation of an isolated causative agent into a healthy animal must cause the same disease and it must be possible to re-isolate it from that animal.
34
microbiological methods during life
o Demonstrating or isolation of causative agent from clinical diagnostic material
o Blood, swabs of MM/skin or samples of various tissues and secretions and excretions
35
microbiological methods after life
Send sample materials in as short a time period as possible after death occurs
36
direct proof of causative agent
Presumes preparing preparations from diagnostic material in which the causative agent is visible and recognisable under a light or electronic microscope using stains
o Eg can identify causative agent of feline infectious anaemia using Giemsa stain – look at position on erythrocyte and its morphology
o Microbiological evidence of the causative agent by growing it in the appropriate culture medium is the most frequent use of microbiological methods and gold standard for most infectious diseases (founded on Koch’s postulates)
which states that for a causative agent to be proven as the causative agent of a specific infectious disease, three basic conditions must be met
37
definition of vaccine
a microorganism in a weakened, live or killed state, or proteins or toxins from the organism // a biological suspension that provides active acquired immunity to a particular infectious disease
38
definition of vaccination
administration of a vaccine to help the immune system develop protection from a specific disease
39
active immunity mechanism
Antigen Recognition:
- When a dog or cat’s immune system encounters a pathogen, immune cells called antigen-presenting cells (APCs) recognize and capture the pathogen’s antigens.
Activation of Lymphocytes:
- B cells produce specific antibodies that bind to and neutralise the pathogen.
- T cells help by either directly attacking infected cells or by supporting other immune cells in fighting the infection.
Formation of Memory Cells:
- After the initial infection or vaccination, some B and T cells become memory cells, which "remember" the pathogen.
-This immunological memory enables a faster, stronger response if the animal is exposed to the pathogen again
40
advantages and disadvantages of active immunity
- Advantages: Long-lasting Protection, defence against future infections, Herd Immunity
- Disadvantages: Time to Develop, possible Side Effects, not Immediate, booster Shots
41
dog/cat vaccination protocol
- Maternally Derived Antibodies (MDA) – 8- 12 weeks
- Puppies with poor MDA are capable of responding to vaccination at an earlier age
- Some puppies may possess MDA at such high titres that they are incapable of responding to vaccination until >12 weeks of age
- Initial core vaccination = 6-8 weeks of age
- Then every 2 weeks
- Last vaccination 16 weeks of age or older
- Number of puppy primary core vaccinations will be determined by the age at which vaccination is stared and the selected interval between vaccinations
Revaccination of adult dogs
42
booster vaccines
- Aim of this vaccine is to ensure that a protective immune response develops in any dog that may have failed to respond to any of the vaccines in the primary core series
- “boosting” the immune system
- 26-52 weeks of age (6-12months)
43
revaccination of adult dogs
- After booster, revaccination = every 3 years (vaccination with MLV core vaccines)
- Every 3 years doesn’t apply to killed core vaccines (except for rabies), to the non-core vaccines and not to vaccines containing bacterial antigens
- Leptospira, Bordetella, borrelia etc products require more frequent boosters for reliable protection
- Non-core vaccinations = annually
44
rabies vaccine
Rabies vaccination
- Mandated by law
- First vaccination at age of 3 months and revaccination every 3 years
- the rabies vaccine is the ENTIRE VIRUS INACTIVATED, but we can use attenuated with oral canid vaccination
45
revaccination of adult cats
- FPV vaccine every 3 years
- FCV/FHV-1 vaccine
Low risk = every 3 years
Higher risk = every year
46
vaccine diseases
- Injury, toxicity or hypersensitivity reaction associated with vaccination
- Adverse events should be reported
- Allergic reactions – Hypersensitivity reaction type 1
- Post vaccinal encephalitis
- Polyarthritis
- Feline injection site sarcoma
o A localised chronic inflammatory reaction initiates malignant transformation of mesenchymal cells and that this process has some genetic basis
o “left leg leukaemia, right leg rabies”
o Non-adjuvanted vaccines should be administered to cats wherever possible
o Vaccines (particularly adjuvanted products) or other injectables should not be administered into the interscapular region
o Always administer subcutaneous
o Vaccines should be administered into a different site on each occasion
47
general infectious syndrome
A general infectious syndrome refers to a set of symptoms that can occur due to various infectious diseases. These symptoms often include:
- Fever: Elevated body temperature, often a sign of infection.
- Fatigue: A general feeling of tiredness or lack of energy
- Muscle and Joint Pain: Discomfort in muscles or joints, often due to systemic inflammation
- Nausea and Vomiting:
- Cough: A reflex action to clear the airways, often present in respiratory infections.
48
complement fixation test
- Serological
- Detects antibodies/causative agent
- Quantitative/qualitative
- High specificity and good sensitivity
- Based on ability of antigen-antibody complex to activate a complement
- 2 stages: main reaction + secondary (indicator reaction)
- Performed with v bottom wells
- 1. Complement inactivated by heat for 30 minutes at 56oC
- 2. Antigen added to inactivated sample and defined quantity of titre complement
- 3. Incubation of wells for 1hr at 37oC, immune complexes are created which activate the complement
- 2nd stage
o Erythrocytes are added
o If complement in main reaction wasn’t activated it’s seen in indicator reaction by lysis of erythrocytes uniform reddish turbidity
o If complement was activated then RBCs remain intact and sink to the bottom
o +ve when no lysis of erythrocytes
49
epizootiology - research methods
- Descriptive method
o Examining distribution of disease in a population and observing features of its distribution
- Historical comparative method
- Analytical (statistical method)
o Testing a hypothesis – how exposure relates to the disease
- Experimental method
- Sociological method
- Theoretical method
50
morbidity rate
ratio between number of sick individuals within a period over total
51
mortality rate
number of deaths over total population
52
incidence
number of new cases within a period
53
lethality rate
number of deaths in sick population
54
prevalence
number of cases in population at a given time
55
factors of general disposition
Factors of general disposition
- Species disposition
o Result of phylogenetic development
o Some infectious diseases occur in multiple species, some only one species
o Parvo – some species more susceptible to specific pathogens due to biological factors
- Breed
o Genetic engineering – making breeds that are resistant to some diseases
- Age disposition
o Colibacillosis and aujeszky disease
o FIP – in young cats as kittens are vulnerable due to developing immune system
- Gender disposition
o real difference in susceptibility related to gender doesn’t exist
o difference in morbidity of females is related to bigger exposure to the infection
- geographic
o west Nile virus – mosquitos vary by region
- vaccination rates - rabies
56
factors of individual disposition
Factors of individual disposition
- Constitution
- Keeping of animals (free range, pasture ecosystem, stable, farm system)
- Nutrition
- Exploitation
o Excessive: breeding and reproduction, forced production and work
- Comorbidity (organic, parasitic, infectious disease)
o Higher susceptibility, worse clinical form of disease and primary diseases
- Tissue damage (mechanical, chemical, physical, biological)
- Transport
- Stress (internal/external)
- Number of susceptible animals
- Immunity
- Environment – climate
57
GDP test
- Serological/immunological method, using agar gel
- Antibody/antigen detection
- Quantitative and qualitative
- Based on antibodies and antigens that are soluble in water and can be diffused, creating ICs
- Formation of precipitation line depends on concentration of antigen and antibody
- If there’s more antibodies, they’ll bind to individual molecules on antigen and ICs will not be visible (prozone effect)
- Concentration of antibody and antigen = optimal – precipitation line
- High specificity
- Single radial immunodiffusion
- double radial immunodiffusion
58
double radial immunodiffusion
so equine infectious anaemia (EIA) (gold standard) and bovine infectious leukosis
o specific and highly sensitive
o both antigen and antibodies diffuse from the well
o known participant is in centre, tested samples in the 6 around
o +ve result = creating line of precipitation between central and peripheral wall
o Controls should form precipitation line
59
single radial immunodiffusion
o Only 1 of serological participants is diffused from a well in the gel
o Other is already present in the gel, in known concentration diffusion is radial
o Will diffuse radially in all directions with concentration decreased in proportion to distance
o Around well --> circular line of precipitation where the concentration = optimal
o used to determine total quantity of Ig in serum/individual fractions
60
first epizootiological factor
- They help determine not only how diseases spread through populations but also the severity and outcomes of infections. Effective management strategies can be developed by considering these factors, ultimately aiding in disease prevention and control efforts.
= Pathogen – dosis and virulence
- Dosis
- pathogen dose is the quantity of pathogen that enters the host.
- This amount is the infective dose (ID) required to establish an infection in a host.
- there is a minimum infective dose below which the pathogen cannot establish an infection
- Salmonella spp. may require a relatively high dose to cause disease in healthy animals.
- Mycobacterium tuberculosis has a low infective dose, meaning even a small exposure can- initiate an infection in a susceptible animal.
- Virulence = measure of the severity or harmfulness of the pathogen once it establishes an infection.
o Virulence factors, such as toxins, adhesion factors, or the ability to evade the host immune response, determine how severe the disease can be and the pathogen's overall impact on host health
o The susceptibility of the host can modulate the effects of virulence
o Transmission Dynamics: Highly virulent pathogens may spread more easily within populations, leading to outbreaks.
61
notification and registration of disease
- Two global organisations: WHO (world health organisation) and OIE (organisation for animal health)
- Needs to be reported urgently if there's a confirmation or suspicion of an infected disease for the interest for the economy and human health
- Responsibility of animal health is obligation of the keeper
- Notifying diseases makes it possible to monitor outbreaks and movements of diseases in animals in the country, but also on a global level, which is the foundation for drawing up the measures necessary to prevent the spread, to control, monitor or eradicate those diseases
- Duties and obligations of keepers of animals
o They’re the first to notice changes to the animals, if they notice changes:
Immediately and without delay notify vet in a vet organisation
Immediately restrict movement
Prevent contact between people and animals
After examination take all the prescribed measures
- Duties and obligations of vets
o Suspicion of a disease
2/+ sick/dead animals with same/similar signs
Sudden death of animals without visible cause
Results of lab tests, clinical picture/ epidemiological circumstances
o Location of an outbreak of disease (confirmed case of the disease)
- If regulations for specific disease prescribe the mandatory reporting of a suspicion, the vet must notify the vet and food safety directorate and the competent vet inspector of this without delay and at the latest within 24hours. Each confirmed case no later than 24 hours, need to do weekly reports until the registration of the end of the outbreak at that location
62
subunit vaccine
- Killed vaccine
- Contains purified pieces of the pathogen (proteins/sugars), elicits an immune response without causing the disease
- natural antigen derived from that virus or organism
- advantages
o Unable to infect, replicate or induce pathology/clinical signs
o Induce a strong immune response
o Rare allergic reaction
- Disadvantages
o Reduced immunogenicity compared to attenuated vaccines
o require an adjuvant (substance than enhances immune response)to increase their potency
63
vector vaccine
- Infectious (live) vaccine
- Uses harmless virus/bacterium (vector) to give specific pieces of genetic material from a pathogen, inducing immune response
- Advantages
o Isn’t relevant to, or pathogenic in dog/cat
o Can carry genetic material from more than 1 pathogen
o Produces strong and lasting immune response
- Disadvantages
o Possibility of immunopathological response
o Risk of transmission
o Cost of production
64
vogralik's chain
- Used to describe and understand the process of infectious diseases transmission
- 1. Pathogen – dosis and virulence
- 2) Source of infection
- 3) Transmission
- 4) Way of entry into the body
- 5) Susceptibility
Of host is ability or affinity of acquiring of some pathogen microorganism and to allow/enable to this pathogen to maintain itself within the host
65
fully virulent/ attenuated vaccine
- use live pathogens that are still capable of replication but have been weakened or modified to reduce their virulence
- aim to elicit a strong and long-lasting immune response without causing the disease in healthy individuals.
- been altered to lose their pathogenicity but retain their ability to replicate.
- Strong Immune Response: robust immune response, including both humoral (antibody-mediated) and cellular immunity.
- Live attenuated vaccines typically provide long-lasting immunity, often requiring fewer doses compared to inactivated or subunit vaccines.
- Not Suitable for Everyone: Live attenuated vaccines may not be appropriate for immunocompromised individuals or those with certain health conditions due to the risk of the vaccine strain causing disease.
66
infection
= invasion of an organisms body tissues by pathogenic microorganism, their multiplication and reaction of host tissues to the infectious agents and toxins they produce
67
recurrent infection
= infections that are too great in number/too severe/ too long lasting
68
reinfection
= additional infection with the same microorganism, occurring after the first infection has resolved
69
relapse
= 2nd episode of disease due to re-emergence of the original infection
70
superinfection
= additional infection with same microorganism, occurring during the course of an existing infection
71
latent infection
= infection that is inactive or dormant
72
incubation period
= period between exposure and onset of clinical symptoms. Pathogen begins active replication without producing recognisable symptoms in the host
73
anamnesis
- Anamnesis refers to the beginning, course and outcome of the disease / the management, feeding, care and use of a sick animal, its treatment and vaccinations given by the owner or person dealing with the sick animal
- Purpose : to direct the clinical examination in order to make an accurate objcetive diagnosis as soon as possible
- Questions about the beginning, symptoms, course and outcome of the disease
o Why, when
- Questions about how the animals are kept and fed
o In case of a conditioned disease – how many animals there is so can see if there’s cohabitation or a chance of spreading between animals
- Questions about previous treatment and vaccination of the animals
o Whether animal’s been treated, what with, if it was suitable
o When it’s vaccinated, by what, dose, how applied, expiration date
- Questions about Epizootioloigcal data, in narrow and broad sense
o Narrow sense = whether owner has other sick animals, whether raised by owner/purchased by legal/illegal manner, which illnesses
o Wider sense = occurrence of illness in neighbourhood/village, whether there's other sick animals around, whether people have been ill
74
MAT
- Reference test for serodiagnosis for leptospirosis
- Based on agglutination reaction between antigens and antibodies (IgG/ IgM) where creation occurs of insoluble complexes – agglutinates observed under dark field microscope
- Detects antibodies to specific serovars using live leptospiral antigens
- Use panel of antigens because of diversity of bacteria
o Composed of strains of refence serovars
o Aligned with species of animal, very important as lack of cross reactivity between groups
- Qualitative and quantitative method
- Quantitative part established whether antibodies to specific Leptospira serovar exist in basic test serum dilution
- Qualitative part = may be positive or negative
- If result is positive = quantitative MAT is performed to establish quantity of antibodies in titres
o Double series of serum dilutions are made for serovars that’s produced +ve result
o Final antibody titre is determined from highest serum dilution in which +ve reaction was present
o Any serum >50% of Leptospira agglutinated is deemed +ve
- Establish antibodies to Leptospira which appear in serum 5-10 days after the start
- If test is -ve in 1st week, test paired sera samples 1-2 weeks after to see seroconversion/4-fold increase (signs of acute leptospirosis)
- In initial phase, antibodies in sera react with several serovars = cross reaction
- Serovars with highest titre = presumptive infective serovar
- Other serovars which have lowest tire = coagglutination
75
advantages and disadvantages of attenuated vaccine
- Advantages
o 1/few doses required
o Long lasting protection
o Both humoral and cellular response
o Can be administered directly to mucosal sites
o Rare allergic reaction
- Disadvantages
o Controlled attenuation normally required
o Poorly define composition
o Risk of reversion to pathogenicity
o Certain risk of transmission
76
ELISA
- Immunological method
- Used to show specific antibodies/antigens
- Qualitative or quantitative
- If only antigens/antibodies present is sought for = qualitative
- If it’s only to determine concentration of antibodies/antigens = quantitative
- Certain quantity of antigen is fixed on a surface then specific enzyme-labelled antibodies are added to it
- Before adding enzyme labelled antibodies, blocking solution is added to the well (contains soluble proteins to bind to the free space so specific antibodies don’t)
- After antigen is fixed in medium, specific antibodies bind to it
- Enzyme labelled antibodies bind specifically to antibodies bound to the antigen
- Between each step, need to rinse to remove all unbound antigens/antibodies
- Add substrate to give reaction colour spectrophotometer to measure absorbance
- Direct ELISA, indirect ELISA, sandwich ELISA and competitive
o Indirect = determining antibodies
o Sandwich = determining antigens
o Competitive = how much antigen is present
77
specificity
= focuses on confirming the absence of the disease
o Ability of a test to correctly identify individuals without the disease or condition (true negatives) (RULES OUT condition)
78
sensitivity
= focuses on detection of the disease
o Ability of a test to correctly identify individuals with a disease or condition (true positive) good at detecting the disease
79
advantages of ELISA
- Advantages
o Sensitivity = can detect low concentrations of substance = highly sensitive
o Specificity = allows for accurate identification of target molecules
o Can make large scale testing
o Easy to perform
80
disadvantages of ELISA
- Disadvantage
o False positive/negative = cross reactivity and can lead to inaccurate results
o Requires specific antibodies
o Stability of reagents
81
PCR
- Molecular diagnostics involve analysis of nucleic acids for diagnostics
PCR (1)
- Advantages = quick availability of results, high specificity and sensitivity
- Based on discovery and then amplification of one or more nucleotide sequences of DNA or RNA
- Before PCR it’s necessary to extract the entire DNA or RNA
- Need to create PCR reaction solution in sterile lab conditions, which consists of:
o Buffer for reaction to take place
o Mixture of DNA in equal measures
o Primers (short strands of DNA ~20 nucleotides) –needs to be specific for part of genome of the target causative agent. Primers are the boundaries of the section of DNA that’s amplified in the reaction
o Heat resistant enzyme Taq polymerase -DNA polymerase extracted from bacteria that lives in thermal springs at temps of 70oC
- Positive control is PCR sample in which DNA is added that was extracted from clinical material previously proved to be positive for the specific microorganism
- Negative control is a sample of purified water which is added instead of DNA or DNA extracted from clinical sample which was previously proven to be negative
- PCR reaction is based on cyclical changes in temperatures to which each individual PCR sample is subjected. It’s consisting of three steps (repeated 30-35times)
o Denaturation or separation of the double stranded DNA into 2 single stranded chains (93-95oC)
o Hybridisation or annealing of the primer (50-60oC) – it’s cooled. Primer binds to complementary base sequence on strands = annealing. Primers determine the target DNA fragment that is amplified in the PCR reaction
o Extension of the DNA chain (72oC)– Taq polymerase binds to free dNTPs and creates new chains, which is complementary to chain of which it’s a copy (extension).
- 2 hours, there's 30-35 cycles and target DNA fragment is copied 2n times (n is number of cycles)
- Amplification of target DNA fragment takes place in a PCR thermocycler
Electrophoresis in agarose gel (2)
- After chain reaction, region of DNA amplified by polymerase = PCR product, which is then subjected to electrophoresis to establish the success of the amplification process of the target region of the DNA
- Does further analysis of DNA (purification, determining its concentration or separation of more DNA molecules)
- Electrophoresis in agarose gel is used for visualisation of DNA amplified in PCR reaction
- 1. PCR product is put in wells, mixed with bromophenol blue dye (which stains PCR product and lowers it to the bottom of the well)
- 2. PCR product travels through the agarose gel by electric current
- The speed of which is moves, depends on length/mass of PCR product
- DNA sample is visible due to ethidium bromide or chemically synthesis dye of DNA added to the gel as it was being prepared
o Dye binds to 2-strranded DNA
- Amplified DNA sequence is shown on the agarose gel in form of a band
- Length of PCR product is expressed in pb units
- One of the wells of gel a DNA marker is placed
o Which contains parts of DNA molecules of a precisely known length
- Visualisation = use chamber with UV source, connected to camera and computer
- +ve PCR reaction = appearance of a signal in the gel. Comparing signals obtained in wells with the DNA marker, can define approx. size of amplified DNA fragments
Reverse transcription PCR (RT-PCR)
- Consists of reverse transcription of RNA into complementary cDNA and its amplification by PCR reaction
- Serves to amplify segments of RNA molecule from a test sample
- Need to add reverse transcriptase enzyme to PCR reaction mixture
- Enzyme converts single chain RNA to a double chain molecule (=cDNA) and amplification reaction of target part of RNA takes place according to the principles of a PCR reaction with a DNA sample
Real-time PCR (or qPCR)
- Follow the creation of PCR product on computer screen in an individual sample in real time
- Make it possible to quantify the amplified part of DNA/RNA in test sample
- Newly created PCR products are marked with fluorescent dye
- Quantity of fluorescence is proportional to the quantity of the PCR product produced
Multiplex PCR
- Instead of single pair of primers, two or more pairs of primers are used, and as a result instead of 1 DNA fragment, several of them are amplified
- Method is more difficult to optimise than simple PCR method, but save significant amounts of chemicals and time needed for the reaction
Nest PCR reaction
- Used when we don’t obtain sufficient PCR product from the PCR reaction to be visible by electrophoresis in gel
- 2 reactions are performed with 2 pairs of primers
- Product of the 1st PCR reaction serves as a mould for 2nd reaction
- Primers used in 2nd reaction are located Inside the fragment amplified in 1st PCR reaction
Restriction fragment length polymorphism (RFLP method)
- Molecular method used to detect variations in homologous DNA molecule sequences
- Compare the polymorphism of homologous sequences of a DNA molecule obtained by cleavage with restriction endonucleases
- Restriction endonucleases are enzymes which cleave DNA fragments in various places, known as restriction sites
- Point mutation/polymorphism changes sequence of nucleotides of an individual DNA fragment, so that the restriction site is lost for a specific enzyme or a new one is created resulting in different cleavage patterns of the DNA fragments analysed
- Used in DNA profiling
- Conducted in several stages:
o Extracting DNA from the sample
o Restriction digestion of a test sample of DNA (using restriction endonucleases – fragments of DNA sample are obtained)
o Agarose gel electrophoresis – DNA fragments obtained are separated lengthwise by electrophoresis in gel
o Hybridisation and visualisation – the fragments are transferred to filter membrane and labelled by specific DNA probes. Length of fragments is according to their complementariness with the probe
RFLP PCR
- Restriction endonuclease cleaves product of PCR reaction
- If there’s no cleavage site, PCR product remains uncloven
- Analysis after cleavage is by electrophoresis
- DNA fragment size is compared in restriction reaction with restriction enzyme and without it
- Polymorphisms and point mutations are detected and analysed in the tested PCR product
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HA
- Serological method
- It’s a neutralisation test
- Based on inhibition of ability of individual viruses to agglutinate the erythrocytes
- Parvo, morbillivirus (distemper) and influenza (gold standard)
- Quantitative specific serum antibodies
- Need virus in concentration of 4-haemagglutination units + erythrocyte suspension
- Concentration of virus in a quantity of 4HA units added in 2-fold serial dilution to 2 rows of microtitration plate + incubated 30 minutes at room temperature
- Plate tilted to 45o and monitor agglutination/settling or erythrocytes
- Presence of infection = agglutination
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mRNA vaccine
- Mechanism:
o mRNA vaccines contain mRNA that encodes a portion of the target pathogen's protein
o Once injected, the mRNA is taken up by cells, which then use their own ribosomes to translate the mRNA into the target protein.
- Immune Response:
o The expressed protein is recognised as foreign by the immune system, prompting an immune response. This includes the activation of T cells (which destroy infected cells) and B cells (which produce antibodies).
o If the vaccinated individual is later exposed to the actual virus, their immune system is primed to recognize and combat it effectively.
- Advantages:
o Rapid development and production.
o No live virus needed, reducing the risk of infection.
o Strong immune response.
Synthetic Vaccines
- Mechanism:
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synthetic vaccine
o Synthetic vaccines can include a variety of components, such as inactivated viruses, recombinant proteins, or peptides designed to mimic parts of the pathogen.
o These vaccines are developed using synthetic biology techniques, allowing precise design of antigens.
- Immune Response:
o Like mRNA vaccines, synthetic vaccines aim to induce an immune response without causing disease. The body learns to recognize the synthetic antigens and prepares to respond to actual infections.
- Advantages:
o Flexibility in design and formulation.
o Ability to target specific parts of the pathogen, potentially improving efficacy and safety.
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division of vaccine
- divided into two main categories: live (attenuated) vaccines and inactivated (or dead) vaccines.
- Live (Attenuated) Vaccines = Live vaccines contain a weakened form of the pathogen that can still replicate but does not cause disease in healthy individuals.
o Eg: canine distemper, feline viral rhinotracheitis, feline parvo + parvo, equine influenza, BVD vaccine
o Advantages
generate a strong and long-lasting immune response, often mimicking natural infection.
both humoral (antibody) and cell-mediated immunity (T cell response).
Single Dose: Often require fewer doses for full immunity compared to inactivated vaccines.
o Disadvantages:
Risk of Infection, Storage Requirements, Not Suitable for Everyone: May not be suitable for individuals with weakened immune systems or certain medical conditions.
- Inactivated (Dead) Vaccines
o contain pathogens that have been killed or inactivated so they can’t cause disease
o eg: leptospirosis, calicivirus, west nile, rabies
o Advantages:
no risk of causing disease
more stable
Suitable for a Wide Range of Patients
o Disadvantages:
Weaker Immune Response,
They usually generate a stronger antibody response but less robust cell-mediated immunity.
Adjuvants Needed
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modes of transmission
- Ways of spreading of pathogenic microorganism from SOI to a suitable portal of entry of the susceptible host
- Successful = transmission of the pathogenic microorganism in at least one minimal infectious dose
- Direct contact, indirect contact and human activities
Factors influencing possible ways of spreading a particular infectious disease
- Pathogen environmental tenacity
o The resistance of microorganisms to environmental conditions
- Environmental conditions
- Minimum infectious dose = Lowest amount of pathogenic microorganism sufficient to cause infection
- Time duration of spread of pathogenic microorganism from SOI to susceptible host
Success of pathogenic transmission
- High pathogenic tenacity, favourable environmental conditions, higher minimum infectious dose and shorter duration of transmission
- Direct contact
- Indirect contact
-Vertebrates
- arthropods
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arthropods spread of disease
- B) Arthropods
o vector borne diseases.
o Arthropods that spread infectious disease are called vectors
o There are two basic ways of spreading infectious diseases by arthropods:
Biological
* Transmission of pathogen microorganism that multiples within it (WNV)
Mechanical
* Transmitting pathogen to contaminated surface of insect
* Hematophagous insects – proboscis is contaminated with blood during feeding on sick animal and then feeds on susceptible animal and spreads
Use repellents to kill – for good prophylaxis, or use nets/protections
Mosquitoes west Nile virus, heart worm
Ticks Lyme disease, anaplasmosis, babesiosis
Flies bluetongue
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vertebrates spread of disease
o Various animal species may be involved in the spread of the pathogen.
o The spread of infection by indirect contact through an animal of the same species is possible in the case when the intermediary’s animal is resistant to infection due to previous illness or Immunoprophylaxis.
o wild animals can also play this intermediaries role. In natural conditions, it is often the case that wild animals, dismember the carcass of a sick animal participate in the spread of infection.
o Hard to prevent entry into certain areas, esp birds and they all travel far
o Prophylaxis by vertebrates spread is preventing entry of them and contact
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indirect contact
- Requires some intermediary that allows a pathogen to remain infective, in at least 1 infectious dose
- Spread of infection by indirect contact allows transmission over a long distance
- Transmission by indirect contact may occur through inanimate and live intermediaries
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direct contact
- Between sick and carrier animals and susceptible host
- Direct body contact = very effective as environmental conditions don’t affect infectivity of pathogen as the spread is instantaneous (ie rabies). Prophylactic = reduce contact
- Faecal-oral and droplet transmission = susceptible animals comes to contact with contaminated parts. Prophylactic = separate sick and susceptible animals
- Transplacental = microorganism crosses placenta from mother to foetus (vertical spread. Prophylactic = general prophylaxis and Immunoprophylaxis of female
- Transovarial = in birds and arthropod eggs
- Cohabitation = living together in close coexistence (intensive farming). Prophylactic = separating is hard, so Immunoprophylaxis most commonly used
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immunologic ablation
- Immunologic Ablation
o The intentional reduction or elimination of specific immune responses, often through therapies like monoclonal antibodies or other immune-modulating treatments.
o Commonly employed in organ transplantation to prevent rejection and in certain autoimmune diseases to dampen harmful immune activity.
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type of infection
- Types of Infections
o Bacterial Infections: examples include pneumonia and tuberculosis.
o Viral Infections
o Fungal Infections: examples include candidiasis and histoplasmosis.
o Parasitic Infections: Caused by parasites; examples include malaria and giardiasis.
o Opportunistic Infections: Occur more frequently in individuals with weakened immune systems
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immunosuppression
o A reduced immune response due to various factors, including medical treatments, diseases or organ transplantation.
o Consequences: Increased risk of infections, especially opportunistic pathogens, and potentially reduced vaccine efficacy.
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immunotolerance
- Immunotolerance
o A state in which the immune system does not mount an attack against specific antigens, often to prevent autoimmunity. This can occur naturally or be induced therapeutically.
o Mechanisms: Can involve regulatory T cells and various cytokines that inhibit immune responses against specific tissues or foreign antigens.
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immunosuppression vs immunotolerance
- Immunosuppression vs. Immunotolerance: Immunosuppression involves a general decrease in immune response, while immunotolerance is a specific lack of response to particular antigens to prevent autoimmunity
