viral persistence viral persistence in vivo: some examples viral titer (log 10 scale) sites of...

Viral Persistence

Viral persistence in vivo:Some Examples

Virus family Example Host(s)

Site ofpersistence

Cytocidalin

permissivecells

Immuneresponse

HIGH TITER REPLICATION

Arenaviridae LCMV Mouse Macrophage NO Restricted

Hepadnaviridae HBV Human Hepatocyte NO Restricted

LATENT INFECTION

Herpesviridae HSV Human Sensory neuron YES Brisk

Polyomaviridae Papilloma Human Epidermal cells YES Brisk

SMOLDERING INFECTION

Picornaviridae TMEV Mouse

CNSGlial cells YES Normal

Paramyxoviridae Measles Human Neurons YES

Supernormal

Lentiviridae HIV Human CD4 lymphocyte YES Variable

Vir

al T

iter

(L

og10

sca

le)

Sites of persistence are usually terminally differentiated cells

Viral persistence in vivo:Some Examples and Rules RNA and DNA viruses can persist Strategies for persistence can range from “high-titer” replication to

“latency” to “smoldering” infections Corresponding strategies for evasion of host immune responses High titer replication:

Noncytocidal vs rapid replenishment of target cells Ineffective immune clearance due to tolerance, immune complex

formation, viral variation etc. Latency:

Viral genome is maintained in non-replicative mode “Hidden” from immune surveillance

Smoldering infections: Continuous replication at low levels Effective immune clerance is prevented by antigenic variation,

infectious immune complex, transmission via intracellular bridges etc.

Cell culture models of viral persistence

Characteristics of the carrier culture Nonlytic virus Lytic virus

What fraction of the cells are infected? ~100% <100%

Are single cell clones always infected? YES NO

Must antiviral factors be present in the culturemedium to protect the cells?

NO YES (1)

Can the culture be “cured” by adding antiviralantibody or interferon?

NO YES (1)

Does the carrier culture resist superinfectionwith the same virus ?

YES (1) NO (2)

Determinants of viral persistence can be mapped

Predominant gene segments(Type 2wt or Type 3ts)

Gene segments Day 16 Day 230

L1L2L3

22/32

222

M1M2M3

222

222

S1S2S3S4

3/222

3/2

3223

Reovirus is a lytic virus but can be induced to cause persistent infections in cell culture by co-infecting cultures with a lytic wild type virus (Type 2 wt) and a temperature sensitive variant (Type 3 ts).

The virus isolated late after persistent infection is a reassortant carrying the genes of the T2 wt virus except for the S4 and S1 genes of the T3 ts variant virus, and it appears that these two gene segments are responsible for the persistent phenotype

S4(3: major outer capsid protein) and S1(1 viral attachment protein) from Type 3ts reduce the efficiency of viral entry, thus reducing likelihood of overwhelming lytic infection

Reovirus: Respiratory Enteric Orphan Virus (dsRNA genome)

Evidence for immune clearance of viral infection: example

Log

10 ti

ter

per

gm

No difference in titer in first week,implies difference is due to subsequent immune response(which is abrogated by Cytoxan treatment)

If immune system can clear virus, what accounts for virus persistence

West Nile Virus: IC injection of 10 6.3 suckling mouse LD50 into adult rats

Mechanisms of persistence and escape from immune surveillance

High titer persistence Not acutely cytocidal or Target cells are replenished at high rate, and Tolerance (absence of virus-specific immunity)

Deletion of naive T-cell clones Exhaustion of peripheral virus-specific T-cell clones Absence of specific Ab response

E.g. HBV (fig. 6.4), LCMV (fig. 7.4) and HIV

Mechanisms of High Titer Persistence:LCMV & Immune tolerance

Log

10 ti

ter

per

gm

Log

10 ti

ter

per

gm

(Fig. 7.4)

Viremia

Exhaustionof LCMV-specific CTL

Mouse infected at high dose---> viral persistence

CTL

Mechanisms of persistence and escape from immune surveillance High titer persistence

Not acutely cytocidal or Target cells are replenished at high rate, and Tolerance (absence of virus-specific immunity)

Deletion of naive T-cell clones Exhaustion of peripheral virus-specific T-cell clones Absence of specific Ab response

E.g. HBV (fig. 6.4), LCMV (fig. 7.4) and HIV

Nonlytic viruses -LCMV Abs circulate as immune complexes LCMV persistence can be terminated by adoptive transfer of virus specific CTL Similar for HBV ( see fig. 5.11)

“Lytic” viruses SIV/HIV Constant replenishment of target cell pool


Latency (e.g HSV, VZV, EBV, CMV) Virus enters and replicates in permissive cells at portal of

entry, after immune induction, virus appears cleared but

actually becomes latent in another cell type Genome may be maintained chromosomally (integrated) or

episomally If genome is in terminally differentiated cells (e.g. neurons for

HSV), no need to replicate genome, but signals required for re-activation (e.g. fever, sunburn, trigeminal nerve insult)

Axoplasmic spread towards periphery, conducts virus to skin-->”cold sores”

Retrograde transport of virions from exposure siteto dorsal root ganglion

Remains latent

Activation results in anterograde transport to epithelial surfaces via peripheral sensory nerves, replication in epithelium results in vesicles

Herpes Simplex Virus


Latency (e.g HSV, VZV, EBV, CMV) Virus enters and replicates in permissive cells at portal of

entry, after immune induction, virus appears cleared but

actually becomeslatent in another cell type Genome may be maintained chromosomally (integrated) or

episomally If genome is in terminally differentiated cells (e.g. neurons for

HSV), no need to replicate genome, but signals required for re-activation (e.g. fever, sunburn, trigeminal nerve insult)

Latently infected cells express little if any viral proteins, permitting escape from immune surveillance


Smoldering Infections Infectious virus is produced, but at minimal levels Virus continues to spread, may produce

progressive chronic disease Detectable immune response, sometimes immune

response may even be hypernormal (due to chronic viral antigenic challenge)

Paradox: why does virus continue to spread in the presence of a potentially effective immune response

Mechanisms of persistence and escape from immune surveillance:Smoldering Infections

Immunological privileged sites Brain

Blood brain barrier limits trafficking of lymphocytes thru the brain

Neurons express little or no MHC Class I (resulting in little presentation of viral antigens and ineffective CTL response)

Experimental evidence: allogeneic/xenogeneic grafts survive better in the brain than in the skin or other sites

Kidney LCMV is cleared more slowly from

kidney than any other tissues ??inability of lymphocytes to cross

subendothelial basement membrane


Immunological privileged sites Intracelluar Bridges

Cell to cell spread of virus without exposure to immune effector mechanisms (e.g. Abs)

Measles in SSPE: neuron to neuron spread in the presence of high titers of neutralizing antibodies

About 1:100,000 primary measles infection results in SSPE

Virus implicated in SSPE is maturation defective--either mutations in matrix protein or envelope glycoprotein, thus, selects for efficient cell to cell spread


Immunological privileged sites Intracelluar Bridges Suppression of MHC Class I

Expression Virus infected cells rendered

less sensitive to CTL attack Adenovirus E1A SIV/HIV nef V

irem

ia



Expression Infectious Immune Complexes

Ab-coated virus remains infectious Ab-virus complex may be

internalized by Fc receptors on macrophages-- Ab dissociates from virus in vacuoles, permitting infection of macrophages

LCMV, Aleutian Disease Virus can form infectious immune complexes and macrophages are major host cell

Treatment of seraFrom mice persistentlyinfected with LCMV

LCMV titer(log10 LD50 per 0.02 ml)

Anti-mouse immunoglobulin

ControlsNormal rabbit serumAnti-mouse albumin

<1.0

3.73.5

Addition of anti-mouse IgGremove infectivity

YY YY

Virus

-LCMV

-mouse IgG



Expression Infectious Immune Complexes Impaired CTL function

Ag specific CTL may be deficient in effector molecules

HIV specific CTL identified by tetramer staining are deficient in perforin content

What would be your controls?

CTL

MHC Class I HIV antigen

E (Effector Cell)

T (Target Cell)

HIV-specific CTL

SAV

YYY

Y Y

Staining using-perforin Abs



Expression Infectious Immune Complexes Impaired CTL function Antigenic variation

Selection for neutralization resistance; allows for viral persistence in the presence of Ab response

11 X 105

pfu/ml





In vivo selection for neutralization escape variants

Ab escape EIAV

Neutralization index (log10) of serum collected on the indicatedday after infection

Virus isolate (day of infection)

Feverspike

(day ofinfection)

0 days 20 days 44 days 62 days 83 days 155 days

0 days 0 0 0.7 2.5 3.2 3.2 20 days 21 0 0 1.0 1.5 1.5 2.5 44 days 44 0 0 0 3.5 5.4 >5.4 62 days 62 0 0 0 0 2.0 2.0 83 days 83 0 0 0 0 0 3.5 155 days 155 0 0 0 0 0 0

--Immune sera neutralizes virus from earlier time points but not concurrent virus or virus thereafter

--evidence of antigenic drift, andexplains viral persistence even in the face of Ab response






Ab escape EIAV LCMV

Vir

em

iaL

og

10 t

ite

r/m

lN

eu

tra

lizi

ng

Ab

tit

er

Indicates humoral immunityplays a role in viral clearance






Ab escape EIAV LCMV HIV

X.Wei. et. al. (2003) Antibody neutralization and escape by HIV. Nature 422:307

16

138212

381

772

Viral Env from Day


Immunological privileged sites Intracelluar Bridges Suppression of MHC Class I Expression Infectious Immune Complexes Impaired CTL function Antigenic variation



Ab escape EIAV LCMV HIV

CTL escape HIV, LCMV

20 weeks

?

DNA vaccinated rhesus macaques challenged withpathogenic SHIV 89.6P

Viral Load

CD4T-cellcount

p11CTetramer

(Immunodominant)

p41Atetramer

p68Atetramer

weeks

CTL escape viasingle amino acid change

in immunodominantCTL epitope

Diseases associated with persistant viral infections: selected examples

Virus family Virus Host Disease

ONCOGENIC VIRUSES

Retro MuLV Mice Hematopoietic, lymphoreticular neoplasms

Hepadna HBV Humans Hepatocellular Carcinoma

Papilloma HPV Humans Cervival Carcinoma

Herpes EBV Humans Burkitt’s lymphoma

HIGH TITER PERSISTENCE

Arena LCMV Mice Glomerulonephritis, vasculitis

Parvo Aleutian disease Mink Glomerulonephritis, vasculitis

LATENT INFECTIONS

Herpes HSVCMVEBVVZV

HumansHumansHumansHumans

Cold sores, encephalitisPneumonitis, retinitis, encephalitisMononucleosisHerpes zoster

SMOLDERING INFECTIONS

Morbilli MeaslesCDV

HumansDogs

Subacute sclerosing panencephalitisEncephalitis, demyelination

Retro HTLV I Humans Tropical spastic paraparesis (HAM)

Polyoma JC Humans Progressive multifocal leucoencephalopathy

Lenti VMVEIAVHIV

SheepHorses

Humans

Interstitial pneumonitis, demyelinationEpisodic hemolytic anemiaAIDS

viral persistence viral persistence in vivo: some examples viral titer (log 10 scale) sites of...

Documents

viral persistence ctl

mechanisms of persistence

clear virus

tolerance absence of

immune system

viral genome

viral variation

hightiter replication