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This chart has three columns labeled components, interactions and mictotiter results. In row A the components are the red blood cells which do not interact with anything and show no reaction in a microtiter result. The lack of reaction is seen as a small red dot in the center of the well. In row B the components are viruses and red blood cells. The viruses and red blood cells clump together and this is seen in a microtiter result as redness throughout the well. This is called hemagglutination. In row C, the components are viruses, red blood cells and antibodies. The viruses and antibodies clump together but the red blood cells do not clump with anything. This is again seen as no reaction; this is called hemagglutination inhibition.
This chart shows the possible outcomes of a hemagglutination test. Row A: Erythrocytes do not bind together and will sink to the bottom of the well plate; this becomes visible as a red dot in the center of the well. Row B: Many viruses have hemagglutinins that causes agglutination of erythrocytes; the resulting hemagglutination forms a lattice structure that results in red color throughout the well. Row C: Virus-specific antibody, the viruses, and the erythrocytes are added to the well plate. The virus-specific antibodies inhibit agglutination, as can be seen as a red dot in the bottom of the well. (credit: modification of work by Centers for Disease Control and Prevention)
  • What is the outcome of a positive HIA test?

Nucleic acid amplification test

Nucleic acid amplification tests (NAAT) are used in molecular biology to detect unique nucleic acid sequences of viruses in patient samples. Polymerase chain reaction (PCR) is an NAAT used to detect the presence of viral DNA in a patient’s tissue or body fluid sample. PCR is a technique that amplifies (i.e., synthesizes many copies) of a viral DNA segment of interest. Using PCR, short nucleotide sequences called primers bind to specific sequences of viral DNA, enabling identification of the virus.

Reverse transcriptase-PCR (RT-PCR) is an NAAT used to detect the presence of RNA viruses. RT-PCR differs from PCR in that the enzyme reverse transcriptase (RT) is used to make a cDNA from the small amount of viral RNA in the specimen. The cDNA can then be amplified by PCR. Both PCR and RT-PCR are used to detect and confirm the presence of the viral nucleic acid in patient specimens.

Hpv scare

Michelle, a 21-year-old nursing student, came to the university clinic worried that she might have been exposed to a sexually transmitted disease (STD). Her sexual partner had recently developed several bumps on the base of his penis. He had put off going to the doctor, but Michelle suspects they are genital warts caused by HPV. She is especially concerned because she knows that HPV not only causes warts but is a prominent cause of cervical cancer. She and her partner always use condoms for contraception, but she is not confident that this precaution will protect her from HPV.

Michelle’s physician finds no physical signs of genital warts or any other STDs, but recommends that Michelle get a Pap smear along with an HPV test. The Pap smear will screen for abnormal cervical cells and the CPEs associated with HPV; the HPV test will test for the presence of the virus. If both tests are negative, Michelle can be more assured that she most likely has not become infected with HPV. However, her doctor suggests it might be wise for Michelle to get vaccinated against HPV to protect herself from possible future exposure.

  • Why does Michelle’s physician order two different tests instead of relying on one or the other?

Enzyme immunoassay

Enzyme immunoassays (EIAs) rely on the ability of antibodies to detect and attach to specific biomolecules called antigens. The detecting antibody attaches to the target antigen with a high degree of specificity in what might be a complex mixture of biomolecules. Also included in this type of assay is a colorless enzyme attached to the detecting antibody. The enzyme acts as a tag on the detecting antibody and can interact with a colorless substrate, leading to the production of a colored end product. EIAs often rely on layers of antibodies to capture and react with antigens, all of which are attached to a membrane filter (see [link] ). EIAs for viral antigens are often used as preliminary screening tests. If the results are positive, further confirmation will require tests with even greater sensitivity, such as a western blot or an NAAT . EIAs are discussed in more detail in EIAs and ELISAs .

The explanation of EIA is separated to show what occurs in a positive sample and what occurs in a negative sample. First patient sample is applied to a membrane filter. If the sample contains viruses they are trapped by the filter. Next, antibody with enzyme conjugate is added. Antibody will attach to antigen if present. Next is a wash step. If the virus is present the enzyme binds to the virus, otherwise the enzyme washes away. Finally substrate is added. If the antibody is present (because it is bound to the virus) the attached enzyme causes a color change. If no enzyme linked antibody is present, no color change occurs.
Similar to rapid, over-the-counter pregnancy tests, EIAs for viral antigens require a few drops of diluted patient serum or plasma applied to a membrane filter. The membrane filter has been previously modified and embedded with antibody to viral antigen and internal controls. Antibody conjugate is added to the filter, with the targeted antibody attached to the antigen (in the case of a positive test). Excess conjugate is washed off the filter. Substrate is added to activate the enzyme-mediated reaction to reveal the color change of a positive test. (credit: modification of work by “Cavitri”/Wikimedia Commons)
  • What typically indicates a positive EIA test?

Part 3

Along with the RT/PCR analysis, David’s saliva was also collected for viral cultivation. In general, no single diagnostic test is sufficient for antemortem diagnosis, since the results will depend on the sensitivity of the assay, the quantity of virions present at the time of testing, and the timing of the assay, since release of virions in the saliva can vary. As it turns out, the result was negative for viral cultivation from the saliva. This is not surprising to David’s doctor, because one negative result is not an absolute indication of the absence of infection. It may be that the number of virions in the saliva is low at the time of sampling. It is not unusual to repeat the test at intervals to enhance the chance of detecting higher virus loads.

  • Should David’s doctor modify his course of treatment based on these test results?

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Key concepts and summary

  • Viral cultivation requires the presence of some form of host cell (whole organism, embryo, or cell culture).
  • Viruses can be isolated from samples by filtration.
  • Viral filtrate is a rich source of released virions.
  • Bacteriophages are detected by presence of clear plaques on bacterial lawn.
  • Animal and plant viruses are detected by cytopathic effects , molecular techniques (PCR, RT-PCR), enzyme immunoassays, and serological assays (hemagglutination assay, hemagglutination inhibition assay).

Fill in the blank

Viruses can be diagnosed and observed using a(n) _____________ microscope.

Electron

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Cell abnormalities resulting from a viral infection are called ____________ _____________.

cytopathic effects

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Short answer

Briefly explain the various methods of culturing viruses.

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Questions & Answers

is it better to study microbiology and then medicine it makes no difference to go directly to medicine?
Jessee Reply
Dray's mathdme cell wall konse color k hote he
Jinal Reply
what is dray's mathdme cell wall
Prabhat
I confused. please help me
Karen
just confused
Raj
l don't understand it please explain it for me.
Karen Reply
epitopes are present on the surface of
Rohit Reply
at the tip of variable region on the antibody...where antigen and antibody binding sites combine...
Fiza
The term that is used refer to moving microbes under a microscope are referred to as?
Lee Reply
Members of the genus Neisseria cause which of the folowing human diseases?
Farah Reply
genital infections
Kamaluddeen
gonorrhoea
sandip
gonorrhoea
Jessee
4. Which of the following specimens should not be refrigerated? a. Urine b. Urogenital swab
Zahraa Reply
urine
Muuse
urine
Agatha
Urine
Tean
urine
Yasser
Urine
Ebtehal
urine
anamika
urine
Puja
urine
Inemesit
urine
Samuel
Urine
Muhammad
Details about McConkey agar
Muhammad
urine
SK
urine
what is bacteria
anamika Reply
a member of large number of unicellular microorganism which have cell wall but lack of cell organelles an oranised nucleus including somewhat can cause disease
Sukhdeep
Bacteria are usually composed of one cell onl to that are neither plants nor animals, microscopic, that may cause diseases or may be beneficial(in gut)... it depends upon their weapons. Nearly all animal life is dependent on Bacteria for their survival
Fiza
thanks
anamika
what factor make bacteria colony large and how could we sterlise it in large scale
fatty
nutrient concentration temp gaseous conc ph ion or salt concentration mositure condition factors contribute to make large colony. by autoclaving we will sterilize bactetia
Sukhdeep
Colony is actually visible growth of Bacteria that is as a result of suitable environment for growth i.e optimal conditions for growth, temperature, moisture etc. there're many methods to get rid of bacteria. If We stop giving them optimal conditions for living Bacteria will die soon .
Fiza
what's the difference between an antigen and a pathogen?
Pathogens are organisms that cause disease in other organisms whereas Antigen is a part of a pathogen that triggers the immune response..
Rajat
so it is the antigen that dendritic cells present to the T cells and not the pathogen itself?
no no antigen are the west product or part of the pathogen. in such case bacteria it self fight with over immune response & in another case bacteria release antigens
vasava
& other antigen like pollan grain, dust particles etc.....
vasava
pathogen are microbes that can infect the body and causw illness....antigens are the part of pathogens that alert the body to an infection
Sukhdeep
antigen is a part of blood and pathogen is foreign particle which causes diseases
Yogyata
antigen could be non microorganism.... where as pathogen is mixroorganism
tadesse
Thanks
Karen
a pathogen is a disease causing organism while an antigen is a protein in the white blood cells which combats pathogens.
Jessee
what type of widal test
sobhit Reply
this test determine for typhoid in this test if H,O antigen are present that indicate the positive test bac. are salmonella typhy
vasava
what h.o denotes
Iqra
o: body of bacteria, h: flagellate
Explain Mould
Chinenye Reply
Explain mycoses and it's classification
Chinenye
why do we have hiccups?
Manisha Reply
shakey diaphragm
Curlisse
The antibody binding site is formed primarily by:
Asalla Reply
How many types of MICROORGANISMS do we have?
Hope Reply
Hello friends
Hope
microorganisms are divided into seven type Bacteria archaea protozoa algae fungi virus and multicellular animal parasites
Raj
What's pathogenesity
Usman Reply
something that causes disease
Anietie
who is the father of microbiology?
Hope
antonie van leeuwenhoek
vasava
it is the severity of the effects caused by a pathogen
Jessee

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Source:  OpenStax, Microbiology. OpenStax CNX. Nov 01, 2016 Download for free at http://cnx.org/content/col12087/1.4
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