MLS 3312 - Module 4 ALL

Baltimore Classification *Class IV*

(+) ssRNA (-) ssRNA

Baltimore Classification *Class V*

(-) ssRNA

HIV Early Therapy:

(1987) Early therapies only targeted one portion of the HIV lifecycle - AZT (zidovudine) - first antiviral used for treatment, though resistant strains were seen weeks to months after introduction. (Now see HAART)

HAART:

(1995) Saquinavir was introduced (protease inhibitor) and combined with available drugs to create the first combination therapy (HAART) The introduction of HAART has changed the natural history of HIV-1 infection and has *increased the average life span of individuals infected with HIV by decades* *Consists of 3 or more antiviral drugs*

Eclipse Phase (HIV Acute Infection):

(7-21 days) The *period between when the first cell is infected to when HIV is detectable* in the blood - a subclinical phase where HIV sets up low levels of replication in the mucosa, submucosa, and draining lymphatics. During this period HIV makes its way to the *gut associated lymphatic tissues (GALT)*, this is a key anatomical site in acute & chronic HIV infection

Hepatotrophic viruses (tropism):

(Viral hepatitis) Viruses that specifically seek out cells of the liver (tissue tropism) - Hepatitis A, B, C, D, & E

HIV Natural history of infection:

(graph) First couple of weeks - Primary infection, acute HIV syndrome, Wide dissemination of virus, seeding of lymphoid organs --> 9ish weeks to years - Clinical latency, constitutional symptoms, opportunistic diseases, death

Viral set point:

*A stable plasma viral load that is predictive of HIV disease progression to AIDS* - the *CD8 T cell (CTL) response controls HIV replication* to this stable plasma viral load following peak HIV viremia - may persist for years

Capsid:

*Contains viral genetic material* In the case of an enveloped virus, the virion is the entire virus and the nucleocapsid is the capsid and nucleic acid, minus the envelope.

Hepatitis Diagnostic laboratory results:

*Elevated levels of ALT, AST, Bilirubin, PT* (clotting time increased)

HEV Transmission:

*Fecal-oral* - rare in the US, usually associated with *travel*

Mixed essential cyroglobulinemia:

*HCV infection is the most common cause*, Immune complexes form with virus, antibody, complement, and *rheumatoid factor*, Cause symptoms of type III hypersensitivity (rashes and joint pain)

Raynaud's phenomena:

*If exposed to the cold*, HCV patients can develop - pain, numbness, and tingling of the fingers and toes

Hepatitis Common causes:

*Inflammation of the liver*, the specific cause can vary (viruses, alcohol, drugs, toxins, autoimmunity)

**Reverse transcription:

*Initiation of (-) strand DNA synthesis* second template exchange is facilitated by annealing of PBS sequences

Variolation:

*Inoculation* of healthy individuals with material from smallpox pustules into a scratch was widespread in China and India ~1000 A.D.

HDV

*Requires co-infection with HBV* - uses HBV surface antigens to make viral envelope, subviral satellite

HIV At risk populations:

*Risk factors vary by country and the overall economic status of those countries* [MSM, IV drug use, unprotected sex, multiple sex partners, sex workers]; Globally, young woman are twice as likely to be infected than men and HIV-1 is the leading cause of death in women of reproductive age

HLA homozygosity:

*Speeds progression* to AIDS

Nucleocapsid:

*The protein coat (capsid) and the nucleic acid together*

Icosahedral symmetry (capsid symmetry):

*most efficient way of forming a spherical shell to maximize internal volume* (the simplest case is an icosahedral symmetry with 20 triangular faces)

Viron:

*refers to the entire virus* - a complete virus particle that consists of an RNA or DNA core with a protein coat sometimes with external envelopes and that is the extracellular infective form of a virus

Most viruses appear:

*rod-shaped or spherical* under electron microscopy

Chronic carrier (HBV):

- After acute infection is cleared virus persists - Patients can be asymptomatic for 20 - 30 years - Chronic infection leads to liver scarring (cirrhosis) - Hepatoma (liver cancer) results in 15 - 25% of chronically infected - Both cirrhosis and hepatoma can lead to death

Viral history:

- Greece/Mesopotamia (The Iliad, Homer describes Hector as RABID; ~ 1000 B.C. Mesopotamian law dictates how to be responsible for RABID dogs) - Egyptian hieroglyphs depict withered legs, typical of poliomyelitis - Evidence of Smallpox being ENDEMIC in the Ganges River basin ~ 500 B.C. (later to decimate the native peoples of the Americas ~1400 A.D.) - Agriculture and the domestication of animals changed the ways humans and viruses interact

HCV Clinical testing (diagnostic algorithm using EIA, RIBA, NAT:

- Molecular testing for HCV RNA are both *qualitative and quantitative* - After positive molecular test, genotyping is performed because different types respond differently to treatment - Liver biopsy is performed to assess cirrhosis and hepatoma

HCV Treatment:

- No postexposure prophylaxis is available - No vaccine is available - *Pegylated interferon-α + ribavrin* is effective in ~55% of patients - Patients with cirrhosis do not respond to treatment - Transplantation can be performed in liver failure

HIV Antiretroviral Drug classes:

1) *Protease Inhibitor* 2) *Nucleoside Analog Reverse-Transcriptase Inhibitor (NRTI)* 3) *Non-Nucleoside Reverse-Transcriptase Inhibitor (NNRTI)*

Polyproteins (Main):

1) *gag* (group-specific antigen) - core and matrix proteins 2) *pol* (polymerase) - reverse transcriptase, protease, integrase 3) *env* (envelope) - transmembrane glycoproteins, gp120, gp41 Accessory Proteins: tat, rev, vif, vpr, vpu, nef

HBV Viron:

1) Enveloped virus 2) Nucleocapsid core protein: HBc/HBcAg 3) Envelope surface antigen: HBsAg 4) Structural E antigen: HBeAg

HIV Viral diversity and ancestral lineage:

1) Fast replication (10^10 virions/day) with high rate or error (3x10-5 / base / cycle) result in a *high amount of viral diversity* 2) HIV is a *zoonotic infection that crossed into humans* from different primate species (simian immunodeficiency virus - SIV) - HIV-1 : Chimapnzee (SIVcpz) - HIV-2 : Sooty mangabey (SIVsmm) 3) HIV-1 is found throughout the world and the more virulent of the two main strains that infects humans (HIV-2, West Africa and Asia) - HIV-1 has been isolated from African patients in the 1950s 4) HIV-1 is further divided into: - Groups (MNO) - Group M most prevalent and subdivided into 8 clades

Tobacco mosaic disease (TMV):

1) First virus discovered 1892, Ivanovsky 2) Beijerinck discovered the agent causing this disease was ultrafilterable (smaller than bacteria, virus = poison)

HBV At risk populations:

1) Having sex with an infected person 2) MSM 3) People with multiple sex partners 4) Healthcare workers 5) Hemodialysis patients 6) Travelers 7) IV drug users 8) Newborns from infected mother

Shared properties of viruses:

1) Infectious obligate intracellular parasites 2) Genome is either DNA or RNA 2) Viral genome is replicated and directs synthesis, by cellular systems, of other viral components 4) Progeny virus particles, VIRIONS, are formed by de novo assembly from synthesized components in host 5) A progeny virion assembled during the infectious cycle is the vehicle of transmission of the viral genome to the next host 6) TWO principle life cycles - lytic & lysogenic 7) NOT the simplest biologically active agents - viroids & prions

HBV Treatment:

1) No treatment is given for acute HBV infection due to spontaneous recovery 2) Antivirals are given to severely ill patients but little evidence they work 3) Chronic infection with active replication is treated with: - Interferon-α - Reverse transcriptase inhibitors - Severely cancerous or cirrhosis can be treated with liver transplant along with anti-HBV immunoglobulin 4) Five available vaccines in the US

HEV Clinical testing:

1) Rarely performed in the US 2) Immunoassays to detect IgM & IgG 3) RT-PCR (reverse transcriptase polymerase chain reaction) for detection of viral genome No vaccine or treatment is currently available

HIV Structure:

1) Surface (SU) 2) Strands of mRNA (+) 3) Lipid bilayer 4) Integrase (IN) 5) Reverse transcriptase (RT) 6) Capsid (CA) 7) Nucleocapsid (NC) 8) Matrix (MA) 9) Protease (PR) 10) Transmembrane (TM)

HBV Global prevelance:

1.5 cases per 100,000 US population - greatly reduced by vaccination program started in 1991; 2 Billion people infected worldwide // ~350 million develop chronic infection; HBV tenth major cause of worldwide mortality

HAV Vaccine introduced in:

1995 - greatly reduced prevalence, 1 case per 100,000 in US

AIDS:

<200 CD4+ Tcells / uL

HAV Clinical testing:

A variety of immunoassays exist for testing: - Indirect enzyme immunoassays - Competitive direct enzyme immunoassays - Capture immunoassays

28

Acute HAV occurs after an incubation of _______ days.

HCV Confirmatory Testing

After a positive screen, would perform: 1) Recombinant Immunoblot Assay (*RIBA*) or 2) *NAT*

B, C, and D

Blood borne Hepatitis

HDV Transmission:

Blood borne/Body Fluids

No

Can HAV become chronic?

HDV Clinical Testing

Can be detected by antibody product to HDV antigens and RT-PCR *Can be prevented with HBV vaccine*

HDV Genome

Circular ssRNA (only known *animal virus*) - only codes for two proteins (long and small delta antigen)

Capsid symmetry forms:

Constructed from a small number of protein subunits

ssDNA, dsDNA

DNA viruses contain both _______ and _______

ds (double stranded)

DNA viruses contain mostly ___ DNA

anti-HAV IgM

Diagnosis of HAV usually involves _______

Bacteriophage (phage=eating):

Discovered by Twort 1915 and named for their ability to *lyse bacteria on agar plates*

No

Does HEV become chronic?

milkmaids

Edward Jenner (1790's) observed that _______ were protected against smallpox

Founder Strain (HIV Acute Infection):

Establishes a local site of infection

cowpox lesions

Extracts from _____ _____ protected individuals against smallpox

HAV Transmission:

Fecal-oral

Hepatotrophic viruses are spread through 2 distinct routes:

Fecal-oral: A and E Blood-borne: B,C, and D

HIV Clinical testing:

Generally performed in a *two-step classical fashion* - Screening test followed by confirmatory test HIV-1 Ab ELISA was the first assay developed for screening blood products in 1985 New guidelines issued by the CDC in 2014 to screen for HIV-1/2 antibody and antigen, along with NAT testing

Class IV

HAV falls under which Baltimore Classification?

rash, joint pain

HBV symptoms are similar to other hepatitis infections with the addition of _______ and _______

Chronic cyroglobulinemia:

HCV Chronically infected patients: 75 - 85% develop chronic infection, 60 - 70% will progress to chronic liver disease (5 - 20% will progress to liver cirrhosis or hepatoma [~20 years])

A and E

Hepatitis spread through fecal-oral route

6-7 weeks

How long is the incubation period for HCV?

4 million

How many people are chronically infected with HCV in the US?

HIV

Human Immunodeficiency Virus

HCV At risk populations

IV *drug use* (60%), Blood transfusions prior to implementation of standard testing in 1992

asymptomatic

In HBV most people are _______

Australian Antigens

In HBV, excess HBsAg is produced and forms particles called _______ _______

Viral load vs CD4 cell counts:

Individuals with *Low set points and High CD4 counts* will *survive LONGER and progress SLOWER* to AIDS than people with a high set point and low CD4 count

Yes

Is HAV reportable to the state health department?

Vaccination (vacca = cow):

Louis Pasteur coined the term _______ in the 1800's in honor of Jenner

HAV At risk populations:

Men who have sex with men (MsM), International travelers, Illegal drug users

HIV History of US epidemic/world pandemic:

Most new (~ 70%) HIV-1 infections are transmitted heterosexually and of the ~ 30 Million people world wide infected with HIV-1 half are women

Hepatitis General symptoms (Physical):

Nausea, Abdominal pain, Fever, Malaise, Anorexia, Dark urine, Clay-colored stool, Jaundice - yellowing of skin and whites of eyes (Hepatitis with jaundice - icteric, Hepatitis without jaundice - anicteric)

HIV Clinical definition of Acquired Immune Deficiency Syndrome (AIDS):

Positive HIV-1 infection (ELISA, Western blot, RT-PCR) with EITHER: 1) *CD4 T-cell count < 200 cells/uL* or < 15% of total lymphocyte population 2) One of the defining illnesses (Cryptococcosis, Pneumocystis, Kaposi's Sarcoma, other *opportunistic infections* and rare or aggressive cancers)

HIV Transmission:

Predominately sexually transmitted infection (STI) can be blood-borne / body fluids

Differences in HIV progression:

Progression to AIDS is variable based on an individuals genetics or pre-existing immune states 1) *Rapid Progressors (RP)* 2) *Long-term Non-progressors (LTNP, viraemic controllers)* 3) *Elite Controllers (EC, elite suppressors)*

ssRNA

RNA viruses are mostly _______ molecules and can be (-), (+), or ambisense

HIV Life cycle:

Receptor recognition (CD4, CCR5 [dual binding], CXCR4) --> Membrane fusion --> Infuse viral RNA --> RT --> Viral DNA (variation) --> Integration (host DNA, provirus DNA) --> Transcription --> Assembly (viral proteins, RNA) --> Budding --> Maturation

RIBA (HCV):

Similar to a western blot, uses synthetic antigens placed on a nitrocellulose strip (C33c, NS5, 5-1-1, c100, and c22 peptides) - *Indirect immunoassay* is performed with *enzyme labeled secondary antibodies*

Diagnostic algorithm:

Since 1989, the diagnostic algorithm for HIV testing in the United States recommended by CDC and the Association of Public Health Laboratories (APHL) initiated testing with a sensitive HIV-1 antibody immunoassay. Specimens with repeatedly reactive initial immunoassays were then tested with a more specific HIV-1 antibody test, either the HIV-1 Western blot or HIV-1 indirect immunofluorescence assay (IFA), to validate those results

HBV Clinical testing:

The levels of antibody are different in acute resolving patients and acute becoming chronic patients. Both antigens and antibodies are detected and have distinct clinical outcomes - HBsAg - Anti-HBs+ (anti-HBsAg) - HBcAg - Anti-HBc+ (anti-HBcAg) - HBeAg - Anti-HBe+ (anti-HBeAg)

HIV Tropism:

There are two HIV-1 tropisms: 1) *Macrophage Tropic (M-tropic, non-syncitia-inducing strain, R5)* - these strains infect cells that express *CD4 and CCR5*, typically *macrophages and effector T helper* cells 2) *T-Topic (syncitia-inducing, X4)* - these strains infect cells expressing *CD4 and CXCR4*, broad range of cells notable long lived *central memory cells and lymphoid progenitors*

HIV Acute Infection:

Usually across mucosal layers (e.g. genital); once exposes, there is a 10 day "Eclipse phase" - cannot detect, can *detect RNA first*

mRNA, Baltimore Classification

Viruses are classified based on their strategies to create _______. This is called the _______.

Acquired Immune Deficiency Syndrome (AIDS) -

Was first described in homosexual men in the US in 1981; HIV-1 was identified as the causative agent in 1983

40%

What % of HBV patients need hospitalization?

In 1% of cases the disease is fatal - Pregnant women infected and in their third trimester have a nearly 30% mortality - If previous liver damage present mortality can be up 70% in infected people

What % of HEV cases are fatal?

40%

What % of HEV infected individuals become ill?

70%

What % of acutely infected HCV patients asymptomatic?

Class VII

What Baltimore Classification does HBV belong to?

Class IV

What Baltimore Classification does HEV belong to?

Class VI

What Baltimore Classification does HIV fall under?

Class IV

What Baltimore Classification is HCV under?

Hepeviridae

What Family does HEV belong to?

Total anti-HAV

What can be used to assess HAV immune status?

HIV1/2 Ag/Ab combinations

What does the HIV screening method test for?

Picornaviridae

What family does HAV belong to?

Hepadnaviridae

What family does HBV belong to?

Flavivirdae

What family is HCV part of?

90 days

What is the incubation period for HBV?

2 weeks to 2 months

What is the incubation period for HEV?

0.8%

What is the mortality rate for HAV?

0.5-1%

What is the mortality rate for people with HBV?

NAT

What method is used for HIV confirmatory testing?

5%

What percentage of HBV infections become chronic?

35%

What percentage of people infected with HAV are hospitalized?

Stool

What specimen are high levels of the HAV virus found in?

2 weeks prior to symptoms, 1 week post symptoms

When are high levels of HAV found in the stool?

HLA-B*27 and HLA-B*57

Which HLA variations *slow HIV disease progression*?

HAV

Which form of hepatitis can people NOT be re-infected with?

Symptomatic

_______ HCV patients are less likely to progress to chronic infection (like HBV)

CCR5-Δ32

_______ is a 32 bp deletion in the CCR5 gene which infers *HIV resistance* (predominate in northern European populations)

cowpox, vaccinia

_______ virus and _______ virus are not the same. At some point during the history of passing cowpox from animal to human vaccinia appeared... no one definitively knows how

*Non-Nucleoside Reverse-Transcriptase Inhibitor (NNRTI)*

binds allosterically at a site away from the active RT pocket

HBV Transmission:

blood-borne / body fluids

HCV Transmission:

blood-borne / body fluids

Tropism:

cells that express specific surface receptors, which make them permissive to infection by a particular virus or bacteria

Naked viral particle:

does not have an envelope

Baltimore Classification *Class I*

dsDNA

Baltimore Classification *Class VII*

dsDNA-RT

Baltimore Classification *Class III*

dsRNA

Complex particles (capsid symmetry):

form complex capsids with structures that are poorly understood (e.g. Retroviridae,)

Enveloped viral particle:

has a *lipid membrane* acquired from the host that surrounds the capsid

HDV Also called:

hepatitis delta virus

Positive strand / SENSE (+):

immediately translatable information

Lytic

infection resulting in lysis

Lysogenic

infection results in the integration of viral DNA into host genome (PROPHAGE)

Viroids

infectious agents of plants, a single molecule of RNA

Prions

infectious protein molecules

*Nucleoside Analog Reverse-Transcriptase Inhibitor (NRTI)*

nucleoside analog that blocks RT binding pocket

HBV Genome:

partial dsDNA

Protease Inhibitor

prevents the cleavage of viral polyproteins which prevents the formation of a mature HIV virion

Helical symmetry

rod-shaped

HAV Genome

ss(+)RNA

HEV Genome:

ss(+)RNA

HCV Genome:

ss(+)RNA - six genotypes with 50+ subgroups

HIV Genome

ss(+)RNA (two copies)

Baltimore Classification *Class II*

ssDNA dsDNA

Baltimore Classification *Class VI*

ssRNA-RT DNA/RNA dsDNA

Platonic Polyhedra Symmetry

tetrahedron, icosahedron, etc. (spherical)

Negative Strand / SENSE (-)

the *complement* to a positive sense DNA or RNA molecule

HCV Screening Testing

third-generation *indirect enzyme immunoassay* using a mixture of *viral antigens on microbeads* - measures structural and nonstructural antigens *qualitatively*

Helical symmetry (capsid symmetry):

two-dimensional lattice and rolled into a cylindrical structure

Nonhepatotropic viruses:

viruses that primarily infect other cells but can infect cells of the liver - *Herpesviridae:* Epstein-Barr Virus (EBV-Mono), varicella zoster (chicken pox), cytomegalovirus (CMV)

*Rapid Progressors (RP)*

will progress to AIDS in ~ 2 years or less

*Elite Controllers (EC, elite suppressors)*

~ 1 in 300 of infected will not progress to AIDS and maintain undetectable viral loads

*Long-term Non-progressors (LTNP, viraemic controllers)*

~ 7% of infected will not progression to AIDS even after decades of low-level viraemia in the absence of treatment (< 2,000 copies/mL)