MLS 400 Quiz Questions (Exam 1)

The influenza virus, which is a Class V negative (-) sense RNA virus, has a genome that serves directly as mRNA, and can be used to synthesize viral proteins immediately after entry into the host cell cytoplasm.

FALSE mRNA is a (+) sense strand of RNA. The influenza virus, a negative (-) sense RNA virus, has a genome that cannot serve directly as mRNA.

Regarding coronavirus, which of the following statements is NOT correct?

- It is an enveloped virus with a helical nucleocapsid containing a positive-sense DNA genome. - It is a prevalent cause of the common cold. - It is associated with severe acute respiratory syndrome (SARS). - Coronavirus appears to attach to host cells using a viral surface glycoprotein spike that interacts with the angiotensin-converting enzyme 2 (ACE2) found on host cell membranes. answer: It is an enveloped virus with a helical nucleocapsid containing a positive-sense DNA genome. explanation: Coronavirus is an enveloped virus with a helical nucleocapsid containing a positive-sense RNA genome.

Which of the following is NOT a nitrogenous base that may be associated with a deoxyribonucleotide?

Adenine Cytosine Guanine Tyrosine Answer: Tyrosine The fourth nitrogenous base is thymine. Tyrosine is an amino acid.

The Genetic Code Chart

(picture)

Viral Nucleic Acid (Chemical Composition of Viruses)

***Either DNA or RNA (but not both) 1.) Single or double stranded 2.) Circular or linear 3.) Segmented or non-segmented 4.) Double-stranded DNA (dsDNA): "sense" and "antisense" strand, which is the template for mRNA 5.) RNA genome polarity: - Positive (+) sense: Single-stranded RNA that is functionally equivalent to mRNA - Negative (-) sense or antisense: Carries RNA polymerase to transcribe genomic negative-sense RNA into mRNA 6.) Most RNA viruses have a single copy of the genome (haploid), but retroviruses have two copies of the RNA genome (diploid) - Most DNA viruses are double-stranded

Transmission (Clinical Presentation)

***Entry of virus into body 1.)The exposure of a susceptible host to virus under conditions that promote infection - Forms of transmission include: a. Vertical transmission from mother to child can occur through: i. Transplacental (virus transferred from mother to fetus through placenta during gestation) ii. At time of birth iii. Through breast milk b. Horizontal transmission from person to person can involve the following routes: i. Respiratory (droplets or aerosols) - Inhalation is the most common route of viral infections - Depends on: Temperature Air currents Viral particle size which impacts distribution ii. Enteral (fecal-oral route) - Must be acid stable - Usually seen with nonenveloped (naked) viruses iii. Parenteral (acquisition other than through the gastrointestinal tract) - Occurs through the exchange of body fluids, such as: Through breaks in the skin or mucous membranes Arthropod (insect) or animal bites Sexual contact 2.) After entry, virus replicates in cells that express viral receptors and have appropriate biosynthetic machinery

Enzymes

***NOTE: the suffix "-ase" is used to refer to an enzyme 1.) Polymerases: a. Participates in the synthesis of nucleic acids. b. Polymerases are categorized by the nucleic acid that will be synthesized and the nucleic acid used as the template (for example: a DNA-dependent polymerase uses DNA as the template) i. DNA-dependent DNA polymerase: synthesizes DNA from a DNA template ii. DNA-dependent RNA polymerase: synthesize RNA from a DNA template (virus needs to provide). iii. RNA-dependent RNA polymerase: synthesizes RNA from an RNA template iv. RNA-dependent DNA polymerase, also called reverse transcriptase, synthesizes DNA from an RNA template, reversing the normal direction of transcription (virus needs to provide). 2.) Nucleases: a. Digest nucleic acids or nucleotides b. Can be RNA specific (RNase) or DNA specific (DNase) c. Endonuclease: site of digestion is within the strand of nucleic acid - Includes restriction enzymes; degrades in middle of strand. d. Exonuclease: site of digestion is at the end of the strand of nucleic acid Nuclease "chews off" one nucleotide at a time 3.) Ligases: a. Forms phosphodiester bonds between existing DNA strands 4.) Methyltransferases: a. Adds methyl groups to nitrogenous bases. 5.) Deaminases a. Removes amino groups from nitrogenous bases

Specimen Collection

***Selection of appropriate specimen for analysis 1.) Specimen selection depends on disease syndrome and viral etiologies suspected. a. Specimens may need to be collected from multiple body sites if a disseminated infection is involved. b. Similar clinical syndromes can be caused by many different viruses. 2.) Optimal specimen selection should include a proper specimen source, which will include infected cells and viable organisms. 3.) Specimens should be collected as early as possible after the onset of symptomatic disease when viral shedding is highest. 4.) Collection devices should be sterile, leak-proof and non-breakable. - Dacron, rayon or polyester fiber swabs with a plastic or aluminum shaft are acceptable. 5.) Viral transport medium (VTM) a. VTM contains buffered salt solution, protein and saccharide nutrients, as well as a pH indicator and antibiotics to inhibit bacterial and fungal contaminants. b. VTM is designed to stabilize viruses and the patient's infected cells. c. Swabs, nasopharyngeal aspirates and tissue biopsies should be placed in VTM for culture and antigen detection.

Regarding adenoviruses, which of the following statements is NOT correct?

- Adenoviruses have a double-stranded DNA genome, an icosahedral capsid and are nonenveloped. - Virtually all adenovirus infections are symptomatic. - Respiratory diseases and gastroenteritis are commonly associated with adenovirus infections. - Outbreaks of severe, acute respiratory disease related to adenovirus infections have been seen in military recruits. answers: Virtually all adenovirus infections are symptomatic.

With EBV infections, which of the following statements is NOT correct?

- After initially infecting epithelial cells in the nasopharynx, EBV then infects and maintains a latent state in B lymphocytes. - EBV has been associated with infectious mononucleosis, Burkitt's lymphoma and nasopharyngeal carcinoma. - The IgM antibody to the VCA antigen of EBV persists for life, and indicates prior exposure to EBV. - EBV is a common infection, with more than 90% of adults in the US demonstrating antibodies to the virus. answer: The IgM antibody to the VCA antigen of EBV persists for life, and indicates prior exposure to EBV.

Cytopathic Effects (Detection of Virus-Infected Cells)

- Cytopathic effect (CPE): the damage, morphological and functional, inflicted on the cell by the virus and includes: a. Cell lysis or necrosis b. Inclusion body formation - Concentrated regions of viral nucleic acid and protein - May be situated in: Nucleus Cytoplasm Both c. Syncytia formation - The fusion of cells to form multinucleated giant cells d. Cytoplasmic vacuolization - Presence of virus may be detected as a distinctive CPE (shrinking, swelling, rounding of cells, or syncytia) using a light microscope. - Cell culture may support growth of several viruses, each producing a distinct CPE. - The type of cell culture in which the virus grows may be used for presumptive identification. - The rate at which at which CPE progresses may help identify the virus. CPE may develop in 1 to 3 days (HSV) or may not be recognized for 5 to 20 days (RSV, VZV, CMV).

Regarding human papillomaviruses, which of the following statements is NOT correct?

- HPVs cause infections of the skin and mucous membranes. - HPV genital infections are typically asymptomatic and most resolve completely in one to two years. - There are 14 specific high-risk HPV types that have been associated with the development of cervical cancer. - Proteins encoded by the L1 and L2 genes are involved in carcinogenesis by interfering with the p53 and retinoblastoma (RB) tumor suppressor proteins. answer: Proteins encoded by the L1 and L2 genes are involved in carcinogenesis by interfering with the p53 and retinoblastoma (RB) tumor suppressor proteins. explanation:nE6 and E7 genes encode proteins that inactivate proteins from tumor suppressor genes p53 and Retinoblastoma (RB), which are an important step in the process by which a normal cell becomes a cancer cell.

When testing the serum collected from a patient with an acute hepatitis B infection, which of the following markers would NOT be likely to be detected?

- Hepatitis B surface antigen (HBsAg) - Hepatitis B core antigen (HBcAg) - IgM antibody to Hepatitis B core antigen (IgM anti-HBc) - Hepatitis B endogenous antigen (HBeAg) answer: Hepatitis B core antigen (HBcAg) explanation: The Hepatitis B core antigen (HBcAg) is never found in the serum of a patient infected with Hepatitis B Virus. The other markers can all be detected during the acute stage of the infection.

In the diagnosis of infectious mononucleosis caused by EBV, which of the following antibodies would NOT typically be expected to be present in the patient's serum during the acute stage of the disease?

- Heterophile antibodies - IgM VCA antibodies - IgG VCA antibodies - EBNA antibodies answer: EBNA antibodies

Penetration (Virus Entry)

- Internalization of virus into the cell involves interaction between multiple VAPs and host cell receptors - The mechanism of internalization depends on the virion structure and the host cell type, and includes: 1.) Endocytosis - With this receptor-mediated phagocytosis by the host cell, the virus binds to specific receptors on the host cell, which induces the formation of a pit that pinches off from the cell membrane to form a vesicle - Low pH in the vesicle promotes the release of the nucleocapsid and viral genome 2.) Fusion - This process involves fusion of the viral envelope with the host cell membrane. - It not only provides a mechanism for internalizing the virus, but can lead to the fusion between the infected host cell and additional nearby host cells, forming multinucleated cells called syncytia. - Detection of syncytia can be used to determine the presence of virus in cell cultures. - Once internalized, the nucleocapsid is delivered to the site of replication.

An indirect immunoassay can be used to detect viral antigen in patient specimens.

FALSE Indirect immunoassays are immunologic techniques that can identify antibodies for a specific virus in clinical samples. Viral antigens are detected using direct immunologic techniques.

Regarding CMV infection in neonates, which of the following statements is NOT correct?

- It may occur as a congenital infection (in utero), during delivery (perinatal) or after delivery (postnatal) through breast milk or saliva. - Congenital infection is most likely to occur due to reactivation of CMV from a latent maternal infection. - Congenital infection may result in cytomegalic inclusion disease. - Perinatal and postnatal infections may be asymptomatic. answer: Congenital infection is most likely to occur due to reactivation of CMV from a latent maternal infection.

Macromolecule Synthesis

- Macromolecule synthesis involves the production of protein polymers and nucleic acids: - Viral transcription results in the synthesis of viral messenger RNA (mRNA) - mRNA codes for the early proteins (nonstructural elements such as enzymes) and late proteins (structural components) - Replication of viral nucleic acid is necessary to synthesize viral genomes that are incorporated into the progeny virus particles. - The mechanics of macromolecular synthesis varies depending on the characteristics of the viral genome (DNA or RNA, positive or negative sense strand, single- or double-stranded).

Regarding HSV infections, which of the following statements is NOT correct?

- Most primary HSV-1 infections occur during childhood. - An oral HSV infection becomes latent when the virus migrates up the neuron to the trigeminal ganglia. - With reactivation, HSV migrates back down the neuron to the initial site of infection, where it replicates and may produce skin lesions. - HSV-2 infection has been linked to encephalitis, with a lesion appearing in one temporal lobe and a high mortality rate. answer: HSV-2 infection has been linked to encephalitis, with a lesion appearing in one temporal lobe and a high mortality rate.

Nucleic Acid Hybridization Probe Assays

- Nucleic acid probes are oligonucleotides (single-stranded DNA or RNA) with sequences complementary to specific regions of the viral genome - These probes are sensitive and specific tools for identifying viral infectious agents - Involves utilization of a nucleic acid probe (oligonucleotide) specific for a specific viral nucleic acid sequence in the test specimen - Nucleic acid probes, labeled with an enzyme, chemiluminescent molecule, fluorescent molecule or radioisotope, will hybridize (bind) to the complementary sequence in the patient specimen. - The hybridized products can then be detected. *In Situ Hybridization (ISH): - In situ hybridization is a direct probe technique that can be used in anatomic pathology to detect a variety of viral pathogens inside individual cells in tissue biopsy specimens. - When fluorescent detection is used by attaching a fluorochrome to the probe, it is called fluorescent in situ hybridization (FISH)

Viral Protein Synthesis

- Once the viral mRNA is transcribed (from either DNA or RNA), it is translated by host cell ribosomes into early and late proteins: a. "Early" is defined as occurring before the replication of the viral genome - The early proteins include nonstructural elements such as regulatory proteins and enzymes used to replicate viral genome - For RNA viruses, an important early protein is the RNA polymerase that will synthesize many copies of viral genetic material for progeny virus particles b. "Late" is defined as occurring after genome replication - The late proteins include structural elements such as the capsid proteins c. Some viral mRNAs are translated into precursor polypeptides (long protein strands) that must be cleaved (cut) by proteases to produce the functional structural proteins, while other viral mRNAs are translated directly into the functional proteins - At the end of macromolecule synthesis, viral proteins have been synthesized and the genome has been replicated so identical copies of the virion can be generated.

Cytopathogenesis

- Replication of the virus can initiate changes in cells that lead to cell death or to alterations in the cell's appearance, functional properties or antigenicity. - The effects on the cell may result from viral takeover of macromolecular synthesis, accumulation of viral proteins, modification of cellular structures, etc. - The outcomes of a viral infection of a cell include: A. Failed infection (no viral replication) - Do not multiply and therefore disappear - May be associated with a host cell that lacks the appropriate receptor, important enzyme pathway, etc. B. Cell death (lytic infection): 1.) A lytic infection results when virus replication ultimately kills the host cell 2.) May see characteristic changes in the appearance of the target cells - These changes in the host cells due to the viral infection are referred to as cytopathic effect (CPE) and can include: - Inclusion bodies in nucleus or cytoplasm (may be due to the accumulation of viral proteins, etc.) - Presence of inclusion bodies are used in laboratory diagnosis as the nature and location of inclusion bodies may be characteristic of particular viral infections - Vacuolization (a small cavity or vesicle in the cytoplasm containing air or fluid) - Rounding of cells - Fusion of neighboring cells into multinucleated giant cells called syncytia - Allows viral infection to spread cell-to-cell and escape antibody detection C. Persistent infection involving replication of virus without cell death: a. Viruses continue to replicate despite the host's defense mechanisms - Cell might not be destroyed if the virus is released through budding b. Individuals chronically infected can be referred to as carriers 1.) Can serve as reservoirs of infectious virions 2.) Low levels of virus may be detectable with little or no damage (shedding) 3.) In a population with a high proportion of carriers, the disease is endemic D. Presence of virus without viral replication, but with the potential for reactivation of the virus (latent - recurrent infections) - May include: a. Latent infection: limited viral macromolecular synthesis 1.) No clinical symptoms 2.) Virus present in resting or inactive state, with absence of viral replication - Nonproductive response - Viral genetic material persists indefinitely 3.) Cell often functions normally 4.) Virus may be dormant for months or years - Stress and other stimuli can activate cells to viral replication, resulting in: b. Recurrent infection: periods of latency followed by viral production 1.) Reactivation can occur with immune suppression 2.) Results in full viral replication 3.) Recurrence of clinically apparent disease E. Transforming (immortalizing) infections caused by oncogenic viruses: - Viruses are an established cause of cancer in humans - Persistent infections can stimulate uncontrolled cell growth, causing transformation of the cell - Causes alteration in cell morphology and metabolism - Oncogenic viruses have mechanisms for immortalizing cells - Viral transformation is first step, but not sufficient to cause tumor formation - Over time, immortalized cells accumulate other mutations that promote development of tumor

Shell Vials Cultures / Centrifuge-Enhanced Culture (Detection of Virus-Infected Cells)

- Shell vial cell culture is a modification of the conventional cell culture adapted for a few viruses - The viruses can be detected more quickly because infected cells are stained for viral antigens produced soon after infection - This modification is best used for viruses that require long incubation before the characteristic CPE is observed (e.g. CMV and VZV) - A round coverslip is added to bottom of shell vial tube, and monolayers of the cell line are grown on the coverslip. - The specimen is inoculated into the vial and the vials are incubated at 37°C. - The coverslips are removed, stained using fluorescent-labeled antibodies and viewed on a microscope. - Only a single type of virus can be detected per shell vial. - If more than one virus is present in a specimen, it would require one shell vial for each virus - A modification of the technique allows simultaneous recovery of multiple respiratory viruses using R-Mix shell vials - Inoculate specimen to two duplicate vials and incubate at 35° to 37°C for 18 to 24 hours. - Stain one vial with monoclonal antibody pool screening reagent: if result is positive, scrape second shell vial and spot an eight-well slide for specific monoclonal reagent to detect the specific virus present.

Viral Genome Detection: Molecular Methods

- The DNA or RNA genome of the virus can be used to identify the infectious agent. - The advantages of molecular techniques are sensitivity, specificity and safety. - Nucleic acid hybridization and amplification tests detect virus by targeting specific regions of the viral RNA or DNA genome. - Accurate and timely viral diagnosis optimizes patient management. - Nucleic acid-based tests can help identify viruses that propagate slowly or not at all in cell culture. - Quantitative assays can determine the number of viral genome copies in a sample, and are referred to as viral load. - With viruses demonstrating the ability to produce antiviral drug resistance, mutation detection is the method of choice for appropriately selecting antiviral therapy.

DNA Viruses (Macromolecule Synthesis)

- The genomes of most DNA viruses are transcribed by the host DNA-dependent RNA-polymerase in the cell nucleus to produce viral mRNAs, which are exported to the cytoplasm, where the host cell ribosomes are used for translation. a. Class I: Double-stranded DNA genome i. Transcription/Translation - Most dsDNA viruses replicate and assemble in the host cell nucleus - These viruses use the host cell DNA-dependent RNA polymerase to synthesize their mRNA transcripts ii. Viral Genome Replication - DNA-dependent DNA polymerase is used to replicate the viral genome b. Class II: Positive (+) sense single-stranded DNA genome i. Transcription/Translation - The positive DNA sense strand has same sequence as mRNA, and cannot be used as the source of the protein code. - The antisense or (-) sense DNA strand must be synthesized first, which can then be transcribed directly into mRNA using the host cell DNA-dependent RNA polymerase. ii. Viral Genome Replication - The host cell DNA polymerase is used to synthesize the negative (-) sense strand, which will then serve as the template to replicate the viral genome by synthesizing the positive (+) sense strand c. Class VII: Partially double-stranded DNA genome - Hepadnaviruses (Hepatitis B) have a genome that is mostly dsDNA but includes a small section that is single-stranded. - It first uses host DNA polymerase to synthesize the missing portion of the DNA genome, forming a complete double-stranded circular DNA molecule. i. Transcription/Translation - Host cell DNA-dependent RNA polymerase is used to transcribe positive (+) sense RNA strands (viral mRNA) from the negative (-) sense DNA strand of the viral genome. ii. Viral Genome Replication - The positive (+) sense RNA strands synthesized as described above will also serve as the template for replication of the genome, generating a complementary negative (-) sense DNA strand. - This process will require a viral-encoded RNA-dependent DNA polymerase as the host cell does not contain that enzyme. - The negative (-) sense DNA strand that has been generated can then serve as the template for the synthesis of the complementary positive (+) sense DNA strand. - Synthesis of the positive (+) sense strand stops when the genome and core of the viral progeny are enveloped, which results in the partially double-stranded circular DNA that is the genome of the Hepadnavirus.

Plaque Assays (Detection of Virus-Infected Cells)

- The plaque assay can be used for viruses that grow well in tissue culture, and can be used to determine virus concentration a. Monolayers of host cells are inoculated with dilutions of virus b.After adsorption, the cells are overlaid with agar to prevent virus from spreading c. Results demonstrate localized cell killing where cell has been infected with virus d. Multiple replication cycles produce a small area of cellular destruction or plaque where a single virus infects, spreads and kills surrounding cells. - A single plaque can arise from a single clonal infectious particle, termed a plaque-forming unit - Plaques can be counted macroscopically - By counting the plaques and correcting for the dilution factor, the viral titer can be calculated (plaque forming units per milliliter or pfu/mL)

Assembly (Viral Maturation)

- The progeny viral particles are assembled by enclosing the viral genome in a protein capsid. - Virion assembly is analogous to a three-dimensional interlocking puzzle that puts itself together in the box. - The capsid is self-assembled from small, easily manufactured component parts. - Assembly occurs when the concentration of the structural proteins is sufficient to drive the reaction thermodynamically, much like crystallization. - Individual protein structural subunits (protomers) are assembled in a stepwise and orderly fashion into capsomers, which are used to create the capsid

Regarding rotavirus, which of the following statements is NOT correct?

- The viral genome is double-stranded segmented RNA. - Rotavirus is a common cause of viral gastroenteritis with diarrhea. - It is a member of the Hepadnaviridae family. - Use of available vaccines has reduced the incidence of disease, especially in young children. answer: It is a member of the Hepadnaviridae family. explanation: Rotavirus is a member of the Reoviridae family; Hepatitis B is a member of the Hepadnaviridae family.

Regarding Hepatitis B Virus (HBV), which of the following statements is NOT correct?

- The viral genome is partially double-stranded DNA. - Hepatocellular carcinoma (HCC) can occur in individuals with chronic HBV. - Reverse transcriptase, an RNA-dependent DNA polymerase encoded by the host cell genome reverse transcribes a positive sense RNA strand into a negative sense DNA strand as part of the viral genome replication. - Antibody to HBV surface antigen (anti-HBs) is not detectable in the chronic HBV infection. answer: Reverse transcriptase, an RNA-dependent DNA polymerase encoded by the host cell genome reverse transcribes a positive sense RNA strand into a negative sense DNA strand as part of the viral genome replication. explanation: The human host cannot synthesize reverse transcriptase, an RNA-dependent DNA polymerase and cannot reverse transcribe an RNA strand into DNA. This enzyme must be encoded by the viral genome.

Release

- The viral nucleocapsid is released from the host cell by one of two processes: a. Lysis (rupture) of host cell membrane - Lysis may be associated with the release of nonenveloped (naked) viruses b. Budding (enveloped viruses) 1.) Nucleocapsids are enveloped as they bud through the host cell membrane modified by the insertion of viral-encoded glycoproteins 2.) The viral nucleocapsid interacts with the membrane site mediated by the matrix protein 3.) Virus exits the cell, which may or may not be damaged or destroyed in the process - Most RNA viruses bud from the cell membrane and the virus is released without killing the cell. - The HSV nucleocapsid assembles in the nucleus and buds out of the nuclear membrane into the endoplasmic reticulum. The virion is then transported to the cell surface and released. 4.) Released virions may infect adjacent cells or may be carried by fluids to distant cells 5.) Spread of infection generally occurs from virus released into the extracellular environment, but alternatively, a virus, nucleocapsid or genome can be transmitted through cell-to-cell bridges, cell-to-cell fusion or vertically to daughter cells. - These alternative routes can allow the virus to escape antibody detection.

Regarding the hepatitis C Virus (HCV), which of the following statements is NOT correct?

- There are 6 genotypes of HCV and genotype 1 causes a significant number of infections in the US. - Transmission is primarily parenteral, and includes injection drug use, body piercing and tattoos. - Acute HCV infection is often asymptomatic. - HCV rarely results in chronic infection. answer: HCV rarely results in chronic infection. explanation: Chronic infection occurs in about 80% of patients with HCV.

Viremia is a term used to describe a localized viral infection, in which the virus replicates and remains at the primary site of entry.

FALSE Viremia is a term that indicates the virus is present in the blood stream.

Regarding Hepatitis A Virus (HAV), which of the following statements is NOT correct?

- Transmission is associated with fecally contaminated water or food, such as oysters grown in polluted water and eaten raw. - Immunization with the HAV vaccine has reduced the incidence of Hepatitis A infections. - There is a strong association with HAV infection and the development of hepatocellular carcinoma later in life. - The detection of an IgM antibody to HAV is useful in the diagnosis of infection. answer: There is a strong association with HAV infection and the development of hepatocellular carcinoma later in life. explanation: An HAV infection typically resolves without complications or permanent liver damage. Chronic infection does not occur and no carrier state develops. There is no predisposition to the development of hepatocellular carcinoma.

RNA Viruses (Macromolecule Synthesis)

- Unlike the DNA viruses, most RNA viruses replicate and assemble in the host cell cytoplasm. - RNA viruses must provide RNA-dependent RNA polymerases as the host cell has no means of replicating RNA. - The RNA genome structure and polarity of the virus groups will determine how the genome is replicated and how viral messenger RNA (mRNA) is generated and proteins are processed. a. Class III: Double-stranded, segmented RNA genome - Most of the RNA segments code for a single protein i. Transcription/Translation - This process will require a viral-encoded RNA-dependent RNA polymerase as the host cell does not contain that enzyme. - For each of the double-stranded RNA genome segments, the positive (+) sense RNA (mRNA) will be synthesized from the negative (-) sense strands ii. Viral Genome Replication - Excess positive (+) sense strands synthesized as described above will serve as the template for the synthesis of the negative (-) sense strands for the replication of the genome (dsRNA segments) b. Class IV: Positive (+) sense non-segmented RNA genome i. Transcription/Translation - Positive (+) sense strand RNA genomes can serve as mRNA and can be used immediately to synthesize proteins after binding to the host cell ribosomes - Translation typically results in the synthesis of a large polyprotein that is cleaved (cut) into several smaller, functional proteins by protease enzymes ii. Viral Genome Replication - Genome replication requires synthesis of a negative (-) sense RNA intermediate strand using virus-encoded RNA dependent RNA polymerase - This intermediate serves as the template for the synthesis of the positive (+) sense genomic RNA c. Class V: Negative (-) sense RNA genomes - This group of viruses includes non-segmented and segmented genomes - Most segments code for a single protein i. Transcription/Translation - Viral-encoded RNA-dependent RNA polymerase is required to transcribe the positive (+) sense strand RNA (mRNA) using the negative (-) sense strand genomic RNA as a template ii. Viral Genome Replication - Genomic replication of the negative (-) sense strand RNA requires synthesis of a full length positive (+) sense RNA intermediate, which serves as the template for the synthesis of the negative (-) sense RNA genome d. Class VI: Positive (+) sense single-stranded diploid RNA genome - Diploid indicates these viruses have 2 copies of the single-stranded RNA genome - Although the viral genome is single-stranded positive (+) sense strand RNA, it does not serve as mRNA following viral entry into the host cell - Instead, the RNA genome serves as the template for the synthesis of a complementary DNA (cDNA) copy of the genome, manufactured in the host cell cytoplasm - It uses a virus-encoded RNA-dependent DNA polymerase - The cDNA travels to the host cell nucleus and the viral cDNA is integrated into the host cell genome, becoming a "provirus" i. Transcription/Translation - Transcription of the viral cDNA into mRNA and translation into viral proteins proceeds along with host cell processes ii. Viral Genome Replication - The positive (+) sense RNA transcripts generated as described above can be used as the new genomes - Macromolecule synthesis also includes the production of viral proteins

Which of the following statements is NOT a characteristic of the Varicella-Zoster Virus (VZV)?

- VZV can be transmitted by respiratory droplets and initially infects the upper respiratory tract. - The virus infects sensory neurons where it establishes a latent (dormant) infection. - VZV present in shingles lesions may be transmitted by direct contact to susceptible individuals, causing chickenpox. - All of the above are characteristics of VZV. answer: All of the above are characteristics of VZV.

Incubation Period (Clinical Presentation)

- Virus is replicating but has not induced sufficient damage to cause disease 1.) May be asymptomatic, resolved by host's innate immune response 2.) May see early nonspecific symptoms, referred to as prodrome: a. Fever or chills b. Malaise c. Myalgias d. Headache

Viral Replication

- Viruses multiply only in living cells. - A lytic viral response exists when productive viral replication occurs in the host cell, resulting in the release of new viral particles. - The infected host cell acts as a factory, providing energy, synthetic machinery and low molecular weight precursors for the synthesis of viral proteins and replication of the viral genome. - Processes not provided by the cell must be encoded in the genome of the virus. - The manner in which each virus accomplishes the steps of viral replication, transcription and translation is different for each type of virus. - A single round of the viral replication cycle can be separated into several phases: 1.) A single virion (infective viral particle) recognizes an appropriate target cell, attaches to the cell and penetrates the cell membrane. 2.) The viral nucleic acid (viral genome) is released into the host cell (uncoating). 3.) Uncoating of the genome initiates the eclipse period, in which no intact viral particles are present and the infectivity of the virus is eliminated. 4.) In the eclipse period, intense viral macromolecular synthesis occurs: a. Viral genome replication produces multiple copies of viral nucleic acids b. Transcription and translation are initiated, producing numerous viral proteins 5.) Some viral proteins will begin self-assembly to form the viral capsid 6.) The appearance of viral particles in the cell signals the end of the eclipse period. 7.) The virions are released to infect new susceptible host cells. - Viral replication includes the following 6 steps...

Target Amplification Assays

- With amplification techniques, viral target DNA or RNA is amplified many times a. Polymerase Chain Reaction (PCR) - The Polymerase Chain Reaction is used to amplify small amounts of specific viral DNA present in the clinical specimen - The assay uses thermostabile DNA polymerase to produce a twofold amplification of the target DNA with each temperature cycle: - In the denaturation step, the temperature is raised to 90° to 95°, which separates the two strands of target DNA - In the annealing step, the temperature is lowered to 45° to 60°, which anneals (binds) two short DNA oligonucleotides called primers to specific sequences of the target DNA. - In the extension step, the temperature is raised to 72°, which extends the primers by adding nucleotides complementary to the target DNA - The cycle is repeated 30 to 40 times to yield amplification of the target DNA by more than 1010-fold. b. Quantitative Real-Time PCR (QPCR) - Quantitative Real-Time PCR (QPCR) is a variation of a traditional PCR reaction, which is used to assess viral loads and monitor response to therapy for viruses such as HIV, CMV, HCV, HBV and BKV. - Real-TIme PCR uses a fluorescent-labeled probe which generates a fluorescent signal as viral DNA is amplified - The signal from the probes is proportional to the amount of amplified viral DNA present in the reaction - The production of DNA is measured by the increase in the fluorescent signal die to the binding of a probe labeled with a fluorescent molecule to the amplified DNA. c. Reverse Transcriptase Polymerase Chain Reaction (RT-PCR) - Another variation of traditional PCR can be performed using an RNA target, which is called Reverse Transcriptase Polymerase Chain Reaction (RT-PCR). - The enzyme reverse transcriptase is used to reverse transcribe RNA into cDNA (complementary or copy DNA) for subsequent PCR amplification. - Once cDNA has been synthesized, DNA polymerase will amplify the DNA in a traditional PCR reaction - The concentration of the cDNA will double with each PCR cycle.

Hepatitis E Virus (HEV) is one of the most common causes of acute viral hepatitis in the US.

FALSE; HEV infection is uncommon in US, and is usually the result of travel to developing country where HEV is endemic.

HPV DNA is found in 99.7% of cervical cancer specimens, and is the only HPV-associated cancer seen in humans.

FALSE; In addition to the association of HPV in 99.7% of cervical cancers, data from CDC provided the following estimates of HPV-associated cancers: 90% of anal squamous cell carcinomas 65% of vaginal cancers 50% of vulvar cancers 35% of penile cancers 60% of oropharyngeal cancers (95% associated with HPV-16 or HPV-18)

Most HAV infections are symptomatic, usually featuring fever and jaundice.

FALSE; Most HAV infections are asymptomatic and are detected only by presence of antibody.

Acute Viral Infection (Clinical Presentation)

1.) During the acute stage, signs and symptoms of infection are displayed 2.) Initial local infection: near site of entry into host (related to mode of transmission) 3.) Virus may replicate and remain at the primary site (localized infection) 4.) Viremia: virus is present in the blood stream a. May travel free in plasma b. May be carried in specific blood cells (macrophages, lymphocytes, etc.) c. Allows inoculation of tissues and organs distant from the primary site 5.) Systemic disease: virus must cross the mucosal barrier in the respiratory, gastrointestinal or genitourinary tracts. 6.) Symptoms caused by viral replication in target tissues: a. May cause tissue damage due to the cytopathic ability of virus b. Symptoms may be due to interaction of virus and immune response c. Symptoms may continue through convalescence 7.) Specific symptoms depend on: a. Portal of entry b. Route or pathway of spread c. Tropism (propensity of virus to infect a distinct group of cells and tissues) - Availability of virus receptors on surface of host cell that interact with viral attachment proteins d. Host age e. Nutritional status of host f. Presence of genetic polymorphisms that affect susceptibility to infection g. Virulence of the virus

In a double-stranded DNA polymer, which DNA sequence below would be complementary to the sequence 5'GATTCTCAAAGGACT3'?

3' CTAAGAGTTTCCTGA 5'

One megabase equals ____________ base pairs.

1,000,000

Transcription and Translation

1. Genes provide instructions for the production of proteins. 2. The order of base pairs in a gene determines the order of amino acids in the protein. 3. The order of amino acids in the protein determines the final three-dimensional shape and function of that protein. 4. Protein synthesis can be divided into two stages: transcription and translation

Asymptomatic Infections (Clinical Presentation)

1.) A lack of symptoms may be associated with a viral infection if the extent of damage is below a functional threshold 2.) Despite the lack of symptoms, virus-specific antibodies are produced 3.) Asymptomatic infections can be a major source of contagion

DNA Replication

1.) A parent strand of DNA serves as a template for synthesis of the new strand a. Nucleotides form complementary base pairing with the template (parent strand) in the creation of the strand being synthesized b. The enzyme DNA polymerase joins the phosphate at the 5' end of the nucleotide being added to the hydroxyl group attached to the 3' (which number?) carbon of the nucleotide already in the chain c. The covalent bond joining nucleotides to create the sugar-phosphate backbone is called a phosphodiester bond. 2. DNA is synthesized in a 5' to 3' direction, copying a DNA template strand running in a 3' to 5' direction - In the diagram to the right, the upper strand (leading strand) is in the correct orientation to be copied along its length so that the direction of replication is left to right, producing a strand oriented 5' to 3'

Viral Lipid Envelopes (Chemical Composition of Viruses)

1.) Acquired when nucleocapsid buds through the host's cell membrane. 2.) Budding occurs at sites where virus-specific proteins have been inserted. 3.) Envelope is lipid bilayer with proteins and glycoproteins 4.) Most viruses acquire envelope from outer membrane a. Herpesviruses bud through nuclear membrane 5.) Viral Glycoproteins a. Glycoproteins in viral envelope are virus encoded b. Surface glycoproteins protrude outside of the envelope - Attach virus to a target cell by interacting with cellular receptor. c. Glycoproteins serve as viral antigens d. Also function as viral attachment proteins (VAP) - VAPs that bind to erythrocytes are termed hemagglutinins (HAs) e. Neutralizing antibodies to glycoproteins in envelope may prevent infection - Damage to envelope inactivates virus 6.) Enveloped viruses vs naked (nonenveloped) viruses: a. Enveloped: - Susceptible to environment: Drying out High temperature pH below 6 and above 8 Detergents - Transmitted by direct contact: Blood or body fluids May be transmitted by insect or animal bite b. Naked (nonenveloped): - Very stable: resistant to harsh environmental conditions - Most viruses transmitted by fecal-oral route do not have an envelope - All human viruses with a helical nucleocapsid have an envelope - Icosahedral nucleocapsids can be enveloped or naked

Viral Proteins (Chemical Composition of Viruses)

1.) Capsid: protein coat surrounding nucleic acid core a. Composed of repeating protein capsomers 2.) Arrangement of capsomers determines shape of virus a. Icosahedral: 20 equilateral triangles - DNA and RNA viruses b. Helical: a hollow coil that surrounds viral nucleic acid - RNA viruses c. Complex structures d. Advantages of capsid with identical protein subunits called capsomers: - No need for complex genetic information - Promotes viral self-assembly 3.) Protects viral genome from degradation by nucleases 4.) Participates in attachment through viral attachment proteins (VAP) to specific host cell receptors 5.) Determines antigenic characteristics of virus. a. Proteins or glycoproteins exposed on surface stimulates immune response. b. Neutralizing antibodies may prevent infection. 6.) Surface antigens define subcategories of virus (serotypes) a. Antigenic variation b. Type-specific 7.) Capsid plus nucleic acid core: nucleocapsid 8.) Capsid may include enzymes essential for initiation of viral replication

Hemadsorption / Hemagglutination Assay (Detection of Virus-Infected Cells)

1.) Cells infected with certain viruses (influenza, mumps, etc.) express a viral glycoprotein (hemagglutinin) that binds erythrocytes to the infected cell surface (hemadsorption). 2.) Such viruses can be detected in cell culture medium due to the agglutination of erythrocytes (hemagglutination). 3.) The reaction may positive before cytopathic changes are visible 4.) Hemagglutination can be inhibited by antibodies to the hemagglutinin (hemagglutination inhibition)

Central Dogma

1.) Double-stranded DNA is copied through the process of replication. 2.) DNA also serves as the template which transcribes into mRNA which translates into protein, following the instructions that originated with the gene. - This "central dogma" was the explanation of the flow of genetic information within a biological system described by Francis Crick in 1958. - NOTE: The discovery of viruses identified biological agents that did not always follow the "central dogma" 3.) Only one strand of double-stranded DNA is template for synthesis of messenger RNA. - This strand that will be transcribed is the "antisense" or negative (-) sense strand. 4.) The other strand of double-stranded DNA is not transcribed into messenger RNA. - This non-transcribed strand is the "sense" strand or positive (+) sense strand, and has the same sequence as mRNA. 5.) The mRNA transcript created from the template antisense DNA strand is referred to as the sense RNA strand. - The sense or positive (+) sense mRNA strand can then be translated into protein. 6.) An RNA strand that would be complementary to the sense (non-template) DNA strand is called the antisense or negative (-) RNA strand - It has the same sequence as the antisense (template) DNA strand.

Viral Pathogenesis

1.) Pathogenesis is the process whereby a virus interacts with its host to produce disease 2.) Viruses cause disease when they evade the host's immune responses and either destroy cells of an important organ or trigger a destructive immune and inflammatory response. 3.) Viral and host factors determine the severity of the disease: a. The tissue targeted by the virus defines the nature of the disease and its symptoms b. Virulence is the capacity of a virus to produce disease in a susceptible host, and is dependent on viral and host factors, including: i. The rate of viral transmission, which depends on: Population density Number of susceptible individuals Weather ii. Virus strain iii. Ability of virus to escape host defenses iv. Host age and immune status v. Loss of virulence results in attenuation of the virus - The virus is altered so it becomes harmless - Some live virus vaccines use attenuated virus strains c. The host's immune response may contribute to the pathogenesis of the disease 4.) Outbreaks of a viral infection result from introduction of a virus into a new location - The outbreak originates from a common source (such as food preparation) a. Epidemics: viral outbreaks that cause a high proportion of cases in a population, community or region - Generally results from a new strain of virus introduced into a immunologically naive population b. Pandemics: worldwide epidemics (not localized to a community or region)

Deoxyribonucleic Acid (DNA)

1.) Structure: DNA is a double-stranded structure composed of building blocks called deoxyribonucleotides which contain: a. Deoxyribose: the ringed 5-carbon sugar in the DNA polymer - The 5 carbons are numbered sequentially - The five carbons are designated as 1' to 5' - A nitrogenous base is attached to the 1' carbon, a hydroxyl group is attached to the 3' carbon and a phosphate group is attached to the 5' carbon b. Nitrogenous bases in DNA: - Guanine and adenine are purine bases - Cytosine and thymine are pyrimidine bases c. Phosphate group: composed of three phosphate molecules in a string d. Deoxyribonucleoside: consists of the nitrogenous base attached to the deoxyribose sugar e. Deoxyribonucleotide: the nitrogenous base is attached to the 1' carbon of the sugar and phosphate group is bound to the 5' carbon. - The deoxyribonucleotide is the basic structural unit in DNA polymer

Ribonucleic Acid (RNA)

1.) Structure: Ribonucleic acid is a single-stranded structure composed of ribonucleotides which contain: a. Ribose: the ringed 5-carbon sugar in the RNA polymer - The 5 carbons are numbered sequentially in a similar manner to deoxyribose (see above) - RNA differs from DNA due to the presence of a hydroxyl group at the 2' C of the 5-carbon sugar in the ribonucleotide b. Nitrogenous bases in RNA: - Guanine and adenine are purine bases - Cytosine and uracil are pyrimidine bases. - RNA has uracil instead of the base Thymine found in DNA c. Phosphate group: composed of three phosphate molecules in a string d. Ribonucleoside: consists of the nitrogenous base attached to the ribose sugar e. Ribonucleotide: nitrogenous base is attached to the 1' carbon of the sugar and the phosphate group is bound to the 5' carbon - The ribonucleotide is the basic structural unit in the synthesis of the RNA polymer

DNA Strands

1.) The 5' end of a strand of DNA strand has an open and available phosphate group 2.) The 3' end of a strand of DNA has an open and available hydroxyl (OH) group 3.) A strand of DNA can be described as having a 5' (ending in a phosphate group) to 3' (ending in a hydroxyl group) orientation 4.) Complementary base pairing occurs in the creation of a double-stranded DNA polymer: a. When one strand has an adenine nucleotide, the opposite strand has a thymine nucleotide - Two hydrogen bonds form b. When one strand has guanine nucleotide, the opposite strand has a cytosine nucleotide - Three (how many?) hydrogen bonds form c. The two DNA strands anneal or hybridize to each other through hydrogen bonds, referred to as base pairing. d. In the complementary double-stranded DNA helix, one strand runs 5' to 3', while the other runs antiparallel, or 3' to 5' - Example of double-stranded DNA sequence representation: 5' GTCGGATCGTAGCTAT 3' 3' CAGCCTAGCATCGATA 5' e. The double-stranded DNA structure (dsDNA) can be described as having: i. A sugar-phosphate backbone ii. Complementary base pairing iii. Two strands that are antiparallel (oriented in opposite directions) f. Genome: the entire content of DNA contained within a cell nucleus is referred to as genomic DNA g. The length of a double-stranded DNA (dsDNA) polymer is measured in base pairs (bp) - One kilobase (kb) = 1,000 bp. - One megabase (Mb) = 1,000 kb h. Oligonucleotide: is used to describe a short, single-stranded nucleic acid chain (can be DNA or RNA) of at least 20 nucleotides in length - An oligonucleotide can be made synthetically to a specified sequence, in order to perform a specific function

Uncoating

1.) Uncoating is the process by which the capsid is removed. This can be accomplished by: a. Enzymatic degradation (viral or host cell enzymes) b. Simple dissociation 2.) Uncoating releases the viral genome for delivery of the viral DNA or RNA to its intracellular site of replication in the host cell nucleus or cytoplasm.

Attachment (Adsorption)

1.) viral attachment proteins (VAPs) which are proteins or glycoproteins on the surface of the capsid (for naked viruses) or envelope - Viral attachment proteins are conspicuous features on the viral surface and typically extend from the surface of the virus in the form of spikes. 2.) The viral attachment proteins bind to protein or carbohydrate receptors on the host cell surface. 3.) The ability of the VAPs to recognize receptors on the surface of a suitable host cell determines the host range (host specificity), which is the host species a particular virus is capable of infecting - A few viruses can infect cells from different species (a broad range of hosts) but most have a narrow range, and may be limited to a single species. 4.) The specific receptor-mediated reaction of a VAP binding to a cell surface receptor also determines the specific cells and tissues of the host that will support the growth of a particular virus (may be referred to as tissue tropism or organ specificity) 5.) Differences in host range and tissue tropisms are due to the presence or absence of receptors for the viral attachment proteins. - For example, Epstein-Barr virus (EBV) has a very limited host range and tropism because it binds to the CD3 receptor (CR2) present on human B cells.

Introduction to Virology

A. Virus: submicroscopic, obligate intracellular parasite 1.) Do not have a nucleus or cytoplasm 2.) Not capable of independent replication a. Can only replicate in living cells through self-assembly of individual components 3.) Cannot generate metabolic energy (do not have mitochondria) 4.) Cannot synthesize proteins (do not have ribosomes) B. Features: 1.) Genome: either DNA or RNA 2.) Protective protein shell a. May be surrounded by an envelope C. Virion: Viral particle that functions as infectious unit 1.) Virion is inert in the extracellular environment 2.) Viral nucleic acid directs host cell to synthesize virus-specific macromolecules

All of the following statements about immunofluorescent assays are true except:

A. Fluorescent dyes called fluorochromes absorb short wavelength light and emit energy at a longer wavelength. B. These fluorescent dyes can be attached to antibody molecules. C. The reaction is observed using a brightfield microscope. D. A positive reaction will appear brightly illuminated against a dark background. answer: The reaction is observed using a brightfield microscope.

Basis of Viral Classification

A. Morphology: 1.) Size 2.) Shape (viral capsid) 3.) Presence of lipid envelope B. Genome: 1.) Type of nucleic acid 2.) The strategy used in replication and transcription C. Viral proteins: 1.) Functions 2.) Antigenic properties (ability to stimulate an immune response in the host) D. Physiochemical properties: 1.) Susceptibility to physical and chemical agents 2.) Heat stability 3.) pH stability (some viruses can be destroyed by alkaline conditions) E. Biological properties: 1.) Natural host range 2.) Mode of transmission 3.) Pathogenicity 4.) Tissue tropisms F. Taxonomy: 1.) International Committee on Taxonomy of Viruses (ICTV) Classification - Order ends with the suffix -virales - Family ends with the suffix -viridae - Subfamily ends with the suffix -virinae - Genus ends with the suffix -virus - Species name provides unambiguous identification - The Human Immunodeficiency Virus taxonomy as specified by the International Committee on Taxonomy of Viruses (ICTV) 2.) Baltimore classification, based on: a. Nucleic acid type b. Nucleic acid number of strands c. Nucleic acid strand sense d.Methods used to generate mRNA

Cultivation and Detection of Viruses

A. Preparation of Inoculum (substance used for inoculation): 1.) Sterile specimens may be inoculated onto cell cultures directly. 2.) Tissue is washed in media or sterile water and ground into paste. Diluent is added, the tube is centrifuged and the supernatant inoculated. B. Biological Methods: - Requires specimens collected during active viral replication 1.) Embryonated Eggs: a. Still may be used for growth of certain viruses such as influenza (in association with vaccine development) b. Also used for research in specialized laboratories 2. Animal Inoculation: a. Studies of pathogenesis of viral diseases, epidemiology, immune responses and viral oncology b. To test vaccines safety 3.) Cell Cultures: - Cell cultures are host cells grown in a monolayer on the sides of glass or plastic test tubes or petri dishes - Many viruses can be grown in specific types of tissue culture cells - Cell cultures are classified as: a. Primary cultures: prepared directly from parent organ - Mixture is transferred to tubes vials, where cells attach in a monolayer on the glass surface. - Cells are overlaid with a nutrient buffer solution. - Primary cultures have been subcultured only once or twice since harvesting. b. Diploid cells: cultures of a single cell type - Remain viable through 20 to 50 generations. - Cells retain normal diploid chromosome pattern c. Tumor cell lines and immortalized cell lines can be propagated indefinitely - Derived from malignant tissues - Can also be prepared by treatment of primary cells with oncogenic viruses or chemicals - Have altered and irregular numbers of chromosomes (aneuploid) - Unfortunately, most viruses do not grow well in continuous cell lines. d. The cells are immersed in cell culture medium to provide nutrients. e. Cell cultures are incubated at 35°C for one to four weeks.

Immunologic Status of Patient

A. The ultimate goals of functional host antiviral immune responses are to prevent entry, reduce spread and eliminate the virus and the cells harboring the virus. 1.) The immune response is the best and in many cases the only means of controlling a viral infection. Innate, humoral and cellular immune response are all important for antiviral immunity. 2.) The hypersensitivity and inflammatory reactions initiated by antiviral immunity can sometimes also be a major cause of the pathologic manifestations and symptoms of viral diseases (immunopathogenesis) 3.) Immune status a. Naïve: not previously exposed and therefore susceptible to viral infection b. Immunized to virus: previously exposed and possibly protected - Prior immunity activates the specific immune response much sooner and more effectively - Immune responses can be produced by vaccines c. Immunocompromised: the host's immune system is impaired - These patients are more likely to get a viral infection and more likely to have a severe illness - Ultimate goal of immune response: elimination of virus and infected host cells - Failure to resolve both aspects may lead to chronic infection 4.) Intact natural barriers (skin, mucous membranes, gastric acids, fever etc.) are the first opportunity to prevent entry of the virus - Skin and mucous membranes are excellent barriers to infection - The immune system will be activated if the natural barriers are breached 5.) First line of defense: innate (nonspecific) immune system a. Involves cells and mechanisms such as inflammation b. Nonspecific process which attempts to limit and control viral replication and spread 6.) Next line of defense: adaptive or specific immune system - Attempts to resolve the infection by eliminating all infectious virus and virus-infected cells - The two arms of the specific immune response include: a. Humoral immunity: production of specific antibodies - Antibodies are effective against the extracellular virus and are essential to control virus spread to target organs 1.) Neutralizing antibodies: generated toward viral attachment proteins that interact with host cell surface receptors 2.) IgM antibodies: first produced but usually transient (temporary) - Typically drops below detectable limits within 3 to 6 months - Primary and Secondary Immune Responses - Usually present during the acute stage of infection - IgM is an indicator of recent or current infection 3.) IgG antibodies: appear later in infection but are typically produced indefinitely - Indicates an infection sometime in the past b. Cell-mediated immunity: involves T cells that can recognize and destroy viral-infected cells through the activation of cytotoxic T cells c. Disease resolution may occur when specific antibody and cell-mediated immune mechanisms halt continued replication of virus - Resolution requires elimination of free virus and virus-producing cells d. The immune response may be source of pathogenesis for many viral diseases 1.) Antibody interacting with large amounts of viral antigen can lead to immune complex disease 2.) Tissue damage may result from cell-mediated and inflammatory responses 3.) Aggressive NK-cell and T-cell responses in adults may exacerbate diseases benign in children 4.) Viral infections may provide initial activation trigger that allows immune system to respond to self-antigens e. Viruses have developed mechanisms to evade the protective immune response

Which of the following statements would NOT be a characteristic of a latent infection?

A. There are no clinical symptoms observed in the patient. B. The virus is present in an inactive state. C. The virus may be dormant for months or years. D. Viral replication continues to take place in the host cell. answer: Viral replication continues to take place in the host cell.

Mechanisms for the vertical transmission of infection include all of the following except:

A. Transplacental transfer B. During birth C. Through insect bites D. Through breast milk answer: Through insect bites

Which of the following statements is NOT true of hemagglutination inhibition:

A. Viruses such as influenza produce an envelope protein called hemagglutinin (HA). B. Hemagglutinin has the ability to bind to the surface of red blood cells causing them to agglutinate (hemagglutination). C. Antibodies to the viral hemagglutinin neutralize the virus, preventing the attachment of the virus to the red blood cells. D. Hemagglutination is inhibited when antibodies are NOT present. answers: Hemagglutination is inhibited when antibodies are NOT present.

The start codon ________ in the mRNA strand, which codes for the amino acid methionine, initiates translation in protein synthesis.

AUG Translation begins with the AUG sequence in the mRNA strand that is the start codon for translation. The tRNA having the anticodon UAC carries the amino acid methionine (Met).

The term used to describe the orientation of individual strands in a double-stranded DNA polymer is:

Antiparallel

START OF LECTURE MATERIAL

Below this card is all the outlines for Virology

A 22-year-old AIDS patient has symptoms of colitis, with several episodes of bloody diarrhea. Histological staining of a biopsy specimen collected from an ulcerated lesion during the colonoscopy revealed giant cells with large intranuclear inclusions with an owl's eye appearance. Which of the following is the most likely cause of the disease?

CMV Giant cells demonstrating large intranuclear inclusions with an owl's eye appearance is a characteristic of CMV.

A pandemic is an outbreak that results in a high proportion of infected patients in a community or localized region.

FALSE Epidemics are outbreaks in a community or localized region. Pandemics are outbreaks that are spread worldwide.

If a virus has an envelope, it is not easily inactivated by drying out and detergents.

FALSE If a virus has an envelope, it is MORE easily inactivated by drying out and detergents. Viruses that do not have an envelope are very stable and more resistant to harsh environmental conditions.

Nonenveloped viral particles are released from the host cell when they bud through the cell membrane after viral assembly.

FALSE Nonenveloped viral particles are released with host cell lysis (rupture). Nucleocapsids become enveloped when they bud through the host cell membrane modified by the insertion of viral-encoded glycoproteins.

The adenovirus fiber protein stimulates the production of neutralizing antibodies capable of providing lifelong protection from all adenovirus types capable of causing human disease.

FALSE The immune response results in the production of neutralizing antibody which is type-specific only, and does not protect against infection by other types.

With VZV infection, zoster is the primary disease typically occurring in childhood, and varicella is the recurrent infection that occurs later in life.

FALSE With VZV infection, varicella (chickenpox) is the primary disease typically occurring in childhood, and zoster (shingles) is the recurrent infection that occurs later in life.

Which of the following classes of enzyme are responsible for digesting nucleic acids?

Nucleases

Specimen Transport and Storage

Specimen Transport: 1.) Specimens collected for detection of virus should be processed as soon as possible 2.) Every attempt should be made to process the specimen within 12 to 24 hours after collection 3.) Specimens for viral isolation should not be allowed to sit at room or higher temperatures 4.) Specimens are generally kept cool (4°C) and immediately transported to the laboratory. 5.) Viral Transport Media: can be used for primary specimens or swab specimens - Can stabilize viruses and infected cells for up to 48 hours Specimen Storage: 1.) For storage of up to 5 days, specimens can be held at 4°C. 2.) Storage for up to 6 days or longer should be at -20°C. Specimens to be frozen should be placed in viral transport medium.

Binding of the viral attachment protein (VAP) to the receptor on the host cell surface determines the tissue tropism of the virus.

TRUE Binding of the viral attachment protein (VAP) to the receptor on the host cell surface determines the tissue tropism of the virus.

Herpesviruses obtain their envelope by budding from the nuclear membrane.

TRUE Herpesviruses do obtain their envelope by budding from the nuclear membrane.

IgM antibodies are the first antibodies produced on initial exposure to a virus, and are usually transient (temporary), typically dropping below detectable limits in 3 to 6 months.

TRUE IgM antibodies are the first antibodies produced on initial exposure to a virus, and are usually transient (temporary), typically dropping below detectable limits in 3 to 6 months.

The early proteins are regulatory proteins and enzymes, while the late proteins may include the capsid proteins.

TRUE The early proteins are regulatory proteins and enzymes, while the late proteins may include the capsid proteins.

The host range refers to the host species a particular virus is capable of infecting.

TRUE The host range does refer to the host species a particular virus is capable of infecting.

In Real-Time PCR (QPCR), the production of viral DNA is measured by an increase in fluorescent signal due to the binding of a probe labeled with a fluorescent molecule to the amplified DNA.

TRUE In Real-Time PCR (QPCR), the production of viral DNA is measured by an increase in fluorescent signal due to the binding of a probe labeled with a fluorescent molecule to the amplified DNA.

In virus-infected cells, the cytopathic effect (CPE) is the damage, morphological and functional, inflicted on the cell by the virus.

TRUE In virus-infected cells, the cytopathic effect (CPE) is the damage, morphological and functional, inflicted on the cell by the virus.

A genital infection with one of the high-risk (HR) HPV types which persists for years is a primary risk factor for the development of cervical cancer.

TRUE; A genital infection with one of the high-risk (HR) HPV types which persists for years is a primary risk factor for the development of cervical cancer.

A portion of individuals infected with SARS-CoV-2 are asymptomatic, but can still spread the disease.

TRUE; Approximately one third of individuals infected with the virus remains asymptomatic, but can still spread the disease.

Norovirus (e.g. Norwalk virus) is a common cause of viral gastroenteritis in adults.

TRUE; Norovirus (e.g. Norwalk virus) is a common cause of viral gastroenteritis in adults.

With influenza viruses, the internal ribonucleoprotein is the group-specific antigen that distinguishes influenza A, B and C viruses.

TRUE; With influenza viruses, the internal ribonucleoprotein is the group-specific antigen that distinguishes influenza A, B and C viruses.

Electron Microscopy (EM)

a. Can play a significant role in characterizing viral morphology b. Not a standard clinical laboratory technique - used more as a research tool i. Magnetic coils use a beam of accelerated electrons as a source of illumination ii. Wavelengths of electrons allow electron microscopes to have a greater magnification and higher resolving power than light microscopes c. EM is most helpful in detecting viruses that don't grow readily in cell culture d. The cause of newly recognized viral syndromes can be recognized rapidly by identifying characteristic viral morphology in infected tissue

Antigen Detection

a. Direct immunologic techniques can be used to detect viral antigens in clinical samples - Indirect techniques are variations that can identify antibodies for a specific virus in clinical samples b. Direct antigen detection techniques use specific antisera that can detect viral protein in infected cells (such as the hemagglutinin antigen of influenza virus) c. The reagent antibodies used to detect viral antigens can be: i. Polyclonal: heterogeneous antibodies that can recognize many epitopes on a single antigen ii. Monoclonal: recognize individual epitopes on an antigen d. These antigen detection techniques allow detection of viruses that do not readily grow in cell culture e. Methods to detect viral antigen in patient specimens include: i. Fluorescent Antibody Techniques for the Detection of Viral Antigens - Fluorochromes absorb short wavelength light and emit energy at a longer wavelength - These fluorescent dyes (fluorochromes) are attached to antibodies specific for a viral antigen - Reactions are observed using a fluorescent microscope - Viral antigens will appear brightly illuminated against dark background a. Direct Fluorescent Antibody (DFA): - Used to detect viral antigen in the patient's specimen - Reagent antibody to viral antigens is tagged with a fluorescent dye, which can be visualized using a fluorescent microscope ii. Enzyme-Linked Immunosorbent Assay (ELISA) - A capture antibody specific for the virus is bound to solid support - Virus present in specimen (such as blood or body fluids) will bind to capture antibody - Virus-specific reagent antibody linked to an enzyme attaches to bound viral antigen - Substrate molecules capable of interacting with the bound enzyme are added - The enzyme will cause a biochemical change in the substrate, and the resulting color change indicates the presence of virus iii. Immunochromatographic methods - A liquid clinical specimen moves across the cartridge membrane by capillary action. - Viral antigen (Ag) in sample binds to specific conjugated antibody (Ab) in the membrane. - A visible line or dot forms as a complex of Ag-Ab coated colored particles are captured in test region

Regarding Human Immunodeficiency Virus (HIV), which of the following statements is NOT correct?

a. HIV preferentially infects and kills CD4+ T lymphocytes, causing a deterioration of immune functions. b. The envelope glycoprotein gp120 has a high mutation rate, resulting in antigenic variants. c. The three enzymes of HIV are reverse transcriptase, helicase and protease. d. Patients in the late stage of infection (AIDS) are subject to opportunistic infections and cancers such as Kaposi's sarcoma and lymphoma. answer: The three enzymes of HIV are reverse transcriptase, helicase and protease. explanation: The three enzymes of HIV are reverse transcriptase, integrase and protease. Enzymes called helicases unwind and separate the double-stranded DNA helix during DNA replication.

The period of viral replication in the host cell in which there is intense macromolecular synthetic activity producing numerous copies of viral nucleic acid and viral proteins, although there are no intact viral particles detected, is called the:

a. Infectious period b. Assembly period c. Eclipse period d. Activity period answer: Eclipse period

Which of the following is NOT one of the characteristics of a virus?

a. It is an obligate intracellular parasite. b. It cannot replicate within living cells. c. It cannot generate its own metabolic energy. d. It cannot synthesize its own proteins. answer: It cannot replicate within living cells.

Light Microscope

a. Light microscopy uses cytological or histological stains for a morphological study of infected cells or tissues in clinical samples b. The resolving power of brightfield microscopes does not allow visualization of the virus but can be used to detect cytopathic effect (CPE), which is the damage inflicted on the cell by the virus, including: i. Viral inclusions, which are the intracellular structures formed by aggregates of virus or viral components in the infected cell ii. Syncytial cells, which occur due to the fusion of cells to form multinucleated giant cells

Transcription

a. One strand of DNA is the template for synthesis of a complementary strand of messenger RNA (mRNA) using the enzyme RNA polymerase b. Ribonucleotides use complementary base pairing to synthesize RNA - As with DNA replication, RNA is synthesized in a 5' to 3' direction while copying the DNA template running in a 3' to 5' direction c. The ribonucleotides are covalently bonded together by phosphodiester bonds. d. Most genes contain non-coding regions (introns) interspersed among the coding regions (exons), as illustrated in the diagram to the right. e. Introns are excised (removed) and exons are joined together to produce mature mRNA. This process is called RNA processing. f. The mRNA molecule is divided up into codons, three consecutive mRNA bases coding for one specific amino acid. g. The mRNA travels to the ribosomes in the cytoplasm of the cell, where the proteins are assembled in a process called translation.

Your summer research project is to study viruses that cause upper respiratory tract infections. You have isolated a virus from a patient's throat and find that its genome is single-stranded RNA. You also find that the genome is the complement of the viral mRNA within the infected cell. Of the following, which is the most appropriate conclusion you could draw?

a. The RNA genome is segmented. b. The virion has a lipoprotein envelope. c. The RNA genome is circular. d. The virion contains an RNA polymerase. answer: The virion contains an RNA polymerase.

Regarding Human Immunodeficiency Virus (HIV), which of the following statements is NOT correct?

a. The protease enzyme mediates the integration of proviral DNA into the host cell DNA genome. b. The gp160 precursor protein is cleaved to form the envelope glycoproteins gp120 and gp41. c. During clinical latency, HIV is typically not detectable in the blood, but is produced and sequestered in the lymph nodes. d. The main immune response in an HIV infection occurs when the HIV-infected CD4+ cells are attacked by cytotoxic CD8+ lymphocytes. answer: The protease enzyme mediates the integration of proviral DNA into the host cell DNA genome. explanation: The enzyme integrase mediates integration of proviral DNA into the host cell DNA. Protease cleaves precursor polyproteins into functional viral polypeptides.

Viral proteins serve several important functions. Which of the following is NOT considered to be one of the functions of viral proteins?

a. They protect the viral genome from degradation by nucleases. b. They participate in the attachment of the virus particle to susceptible cells through specific host cell receptors. c. They are functionally equivalent to mRNA. d. They determine the antigenic characteristics of the virus. answer: They are functionally equivalent to mRNA.

Translation

a. Transfer RNA (tRNA) is a cloverleaf-shaped RNA structure with a loop containing three bases (anticodon) complementary to the mRNA codons. b. Translation begins with the start codon for translation, AUG. The tRNA having the anticodon UAC carries the amino acid methionine (Met). c. a. tRNAs with the appropriate anticodons pick up specific amino acids, transfer them to the ribosomes, and insert them in their proper position according to the mRNA codon "message." d. The amino acids are incorporated into the growing protein chain with peptide bonds. e. The sequential joining of amino acids to form a three-dimensional, functional protein is known as the elongation step. f. This process continues in the 5' to 3' direction until the ribosome hits a stop codon that terminates the protein chain. - UAA, UAG, and UGA are the stop codons.

Regarding influenza viruses, which of the following statements is NOT correct?

a. Viruses from other species can be the source of RNA segments that encode antigenic shift variants that can cause influenza A epidemics among humans. b. Because influenza B virus is only a human virus, there is no animal source of new RNA segments, so Influenza B virus does not undergo antigenic shifts. c. Hemagglutinin binds to cell surface receptors to initiate infection of the host cell. d. The ability of neuraminidase to spontaneously agglutinate red blood cells is the basis the neuraminidase inhibition test used in the diagnosis of influenza infections. answer: The ability of neuraminidase to spontaneously agglutinate red blood cells is the basis the neuraminidase inhibition test used in the diagnosis of influenza infections. explanation: Hemagglutinin spontaneously agglutinates red blood cells and is the basis the hemagglutination inhibition test used in the diagnosis of influenza infections.

Most genes contain large amounts of non-coding regions, called introns, that are interspersed among the coding regions, called __________, that provide the blueprint for the final protein.

exons