Intro to Virology Pt 2 Flashcards
Encounter and Entry-Transmission
- Transmission of virus from infected host to susceptible individual
- Human to human transmission from acutely ill persons, chronic carriers,
mother to fetus - Direct contact - sexually transmitted diseases
- Environmental - fecal-oral route in diarrhea
- Aerosols – respiratory - Chickenpox
- Direct inoculation - infected needles or blood products
- Hepatitis C
chronic carriers like those who are infected with hepatitis B sheeding virus and at low levels.
Trace contacts, and involved public health to trace hep B
Vertical transmission to fetus
Respiratory route
Aerosol droplets, nasal secretions or saliva
* Via coughing or sneezing
* A sneeze may generate up to 2,000,000 aerosol particles, a cough
90,000
* Epstein-Barr virus spread by saliva
* Kissing disease
You don’t necessarily need to sneeze it out. But you know the touching or sharing of saliva or spreading by saliva
Common cold
Caused by rhinoviruses
* Not spread by aerosols
* Spread from hands to eyes, nose or mouth
* Cycle can be interrupted by hand washing
Gastrointestinal Route
- Viruses shed in feces contaminate food or water
- Ingested by a susceptible individual
- “Fecal-oral spread”
- Poor personal hygiene: stool-tainted hands
*Day care centers and institutions for mentally impaired have difficulty
maintaining hygiene - Examples: Norovirus, Hepatitis A/E, Rotavirus
Transcutaneous route
west nile virus
- Skin provides a physical and biological barrier and virus entry.
- Direct inoculation via insect or animal bites or via mechanical devices
such as needles - Animal/insect vectors
- Rabies or West Nile virus, complicated disease cycle
West nile virus
- the mosquito can then bite, you know.
- and other animals that can invite us in when it infects other animals say, for example, horses, the the virus replicates, but it doesn’t produce what we call varymia, or acute illness that we can physically see with our own eyes.
We can’t tell if this this animal is sick, for example. But essentially the virus is replicating, and that animal hosts Another susceptible mosquitoes say it doesn’t carry the muscle virus to bite that host, and then draw some of that
infected blood and carry the virus as well. And then humans essentially become infected when those mosquitoes come and bite us
Most often cases are asymptomatic patients, typically, you know, present with like a very short febrile illness, immunocomp or preg pt more at risk
animal host, like the horse or the humans as Dead End hosts meaning, we can’t pass on the virus from human to human, necessarily or animal to animal. You requires really that mosquito vector for it to transmit from one human to the next, unless
you have direct inoculation, sharing of needles,
Transcutaneous route
- Iatrogenic inoculation - induced by medical or diagnostic procedures
- Contaminated blood products: Hepatitis B, Cytomegalovirus, HIV
- Infected donor material in transplantation
- Pituitary tissues used to prepare growth hormone
- May be purposeful as in parenteral vaccination using live attenuated
virus: Measles or Sabin polio vaccines
Sexual Transmission
Entry across the genitourinary or rectal
mucosa
* Virus may be spread to other parts of the
body or remain near the site of entry
* Ex. HIV, Hepatitis B, Cytomegalovirus, Herpes
Simplex Type 2 (or HSV-1), Mpoxx
Types of transmission
- Horizontal
- Transmission of virus between members of a susceptible host population
- Vertical
- Infection of a fetus in utero through
- Virus carried in the germ cell line
- Virus infecting the placenta
- Virus in the birth canal
Multiplication and Spread
- For some viruses, entry, primary replication and tissue tropism all
occur at the same anatomic site - Rhinovirus – common cold
- Rotavirus – enteritis
- Papillomavirus – warts
- Other viruses enter at one site and spread to a distant area to
produce disease - Enterovirus – meningitis
- Varicella zoster – chickenpox
Host factors in defense and damage
- Age and genetic make-up
- Newborns are susceptible to severe disseminated herpes simplex virus infection
- Some infections are more severe in older individuals, Epstein-Barr
(mononucleosis), poliovirus - General health - nutritional status, stress
- Physical Barriers
- In-tact skin
- Cell-mediated immunity
Antibody response
- Do not usually play a primary
role in terminating acute viral
infections but are very
important in preventing reinfection - Neutralizing antibodies reduce
infectivity by inhibiting
attachment, penetration or
uncoating of virus
Complement
Viruses can trigger the activation of the alternative
and classic pathways of complement
* Activated complement enhances phagocytosis of
viruses
* Leads to lysis of enveloped viruses or virus-infected
cells
Cell-mediated immunity
- Major factor in the termination of viral infections and the
pathogenesis of these diseases - Infected cells are susceptible to the action of lymphocytes
that recognize viral antigens on their surface - Virus-infected cells can be lysed by several types of
lymphoid cells
Identification of viruses (3)
- Isolation of virus in animals or tissue culture
- Detection of virus-specific antigens or nucleic acids in tissues or
body fluids - Demonstration of specific serologic responses
* Identification requires appropriate specimen obtained during a suitable phase
of illness
* Specimens must be rapidly transported with clinical information to the
laboratory
Lab Diagnosis
Standard Culture
ok
What is CPE?
- CPE applies to virus induced cellular
changes that are visible by light
microscope - Most effective way to visualize the
changes is by staining by chromatic
dyes - Cell monolayers are fixed and stained
by acid or basic dyes that accentuate
the nature and location of the change - haemotoxylin (basic dye) and eosin
(acidic dye) (H&E staining)
Direct Detection Fluorescent Antigen
Used for viruses that create
lesions (HSV, VZV)
* Vigorous scraping of the lesion
base is essential. Remove scab
before sampling.
* Uses HSV antigen-specific
murine MAbs (fluorescein
labeled) for detection
* apple-green fluorescence
* Observe under UV microscope
You add some
antigen specific marine fluorescent antibodies, so the antibodies are tied to fluorescence. They bind specifically to the antigen of interest, and under a UV. Microscope you can see these apple green fluorescence over here, indicative of a certain viral infection that you’re inquiring about.
Viral Culture
benefits (3)
pitfalls (5)
Benefits
1. Can differentiate between
viable and non-viable viral
particles
2. Able to detect unknown viral
etiologies
3. Inexpensive compared to PCR
Pitfalls
1. Relatively slow
2. Does not support
growth for all types of
viruses
3. Toxins can resemble
cytopathic effects
4. Contamination due to
bacteria, fungus and
mycoplasma reduce the
sensitivity of viral
detection
5. Endogenous virus from
cells can be mistaken as
the infectious agent
Electron microscopy
- Identification of viral
particles - Cell culture
- Tissue samples
- Skin scrapings
- Stool samples
Serology
- A fourfold or greater increase in the antibody titre to
a specific viral agent in an acute and convalescent
sera (3 to 4 weeks later) - Neutralizing, complement fixation, hemagglutinationinhibition
- Single sample demonstrating specific IgM antibody
patient presented with an acute illness, maybe a few days or weeks ago. And now you’re trying to
identify, maybe, what they had.
You want to look for their serological markers, seeing if they have ig as opposed to Igm. So ig. Being a marker of past infection that can give you an idea.
when the patients actually infectious, presenting acutely
on. Well, you can collect serum then, and look for Igm
a marker of the acute infection.
after the acute illness they’ve likely translated into now a past infection something that they’ve done within the past, and you’d look for your Ig marker at that point 3 to 4 weeks later (convalescent sera)
Molecular studies
- Detection of even single copies of virus genomes in tissue or cells
from body fluids - Restriction enzyme analysis of DNA genomes
- Oligonucleotide fingerprinting of RNA viruses
- In situ hybridization, polymerase chain reaction techniques
- Commercial available kits allow for rapid and sensitive diagnosis
Polymerase Chain Reaction (PCR)
in order for you to replicate numbers and millions of copies of of nucleic acid, you start with this double stranded DNA that separated, using heat, and then you have proteins that bind to keep the 2 different strands separated. The temperature is then reduced to 50 degrees, and you have what we call these primers, which are small and nucleotide sequences that attach to the separated DNA strands, and the polymerase is then bind to those primers and create a complementary sequence and elongate the strand.
Detection of amplified products
Agarose gel analysis for detecting the presence of amplified
products for conventional PCR
Real Time PCR
with real time. Pcr: You cut out this whole gel electrophoresis step, and you visualize that product, multiplying in real time
essentially attached to the primers are these reporters and clenchers, and when these
the replication process occurs, when the polymerase binds to the the nucleic acid strand and creates a complementary sequence. It causes the release of the reporter on that primer to fluoresce, and that’s captured by a computer. And you can essentially have this real time detection of nucleic acid in your your target sample.
Syndromic multiplex testing panels
often what’s very ideal is having these multiplex testing panels in-house are commercially available where you target multiple viruses, ideological agents at the same time.
respiratory pathogen panel that we use actually and at the provincial lab, targeting a whole number of viruses and bacterial targets influenza a respect to our sensitive virus pair influenza viruses your endemic coronaviruses.
because once you extract your once you receive your sample, extract it within about 3 h we got results.
Viral Amplification
Benefits (4)
pitfalls (5)
Benefits
1. Can detect very low number of
viruses
2. Rapid
3. High throughput
4. Multiplexed assays
Pitfalls
1. Contamination due to
high concentration of
amplicons
2. Detection of dead
organisms
3. False positive and
False negative results
4. Expensive
5. Special training
Dead organism - Detection of nucleic acid does not necessarily mean we’re detecting live virus
How to overcome contamination problems
- Physical separation
- Dedicated set of supplies
- Aliquot PCR reagents for single use
- Plugged tips to prevent carryover between samples
- Negative and Positive controls
- Maintenance of thermocyclers
- Meticulous lab techniques
Rapid Diagnostic Test
- Quick, easy-to-use tests
- Results within a few minutes
- Immunochromatographic tests
containing bound antibody that will
react to presence of antigen on a
nitrocellulose membrane - Ex. COVID Rapid Antigen
Johnny comes to the pharmacy and wants
antibiotics to treat his “cold”
- His lab results are:
- DFA: NEG
- RTPCR: POS for
rhino/entero on
syndromic panel - What does this mean?
- Should he be given
antivirals/antibiotics?
