Viral Persistence

Viral persistence in vivo: Some Examples Viral Titer (Log10 scale) 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

Determinants of viral persistence can be mapped Reovirus: Respiratory Enteric Orphan Virus (dsRNA genome) 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(s3: major outer capsid protein) and S1(s1 viral attachment protein) from Type 3ts reduce the efficiency of viral entry, thus reducing likelihood of overwhelming lytic infection

Evidence for immune clearance of viral infection: example West Nile Virus: IC injection of 10 6.3 suckling mouse LD50 into adult rats If immune system can clear virus, what accounts for virus persistence Log10 titer per gm No difference in titer in first week, implies difference is due to subsequent immune response (which is abrogated by Cytoxan treatment)

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 (Fig. 7.4) Mouse infected at high dose---> viral persistence Viremia CTL Log10 titer per gm Log10 titer per gm Exhaustion of LCMV-specific 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 a-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

Mechanisms of persistence and escape from immune surveillance 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”

Herpes Simplex Virus Retrograde transport of virions from exposure site to dorsal root ganglion Remains latent Activation results in anterograde transport to epithelial surfaces via peripheral sensory nerves, replication in epithelium results in vesicles

Mechanisms of persistence and escape from immune surveillance 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

Mechanisms of persistence and 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

Mechanisms of persistence and escape from immune surveillance: Smoldering Infections 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

Mechanisms of persistence and escape from immune surveillance: Smoldering Infections 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 Viremia

Mechanisms of persistence and escape from immune surveillance: Smoldering Infections Immunological privileged sites Intracelluar Bridges Suppression of MHC Class I 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 Addition of anti-mouse IgG remove infectivity Virus a-LCMV Y Y Y Y a-mouse IgG

Mechanisms of persistence and escape from immune surveillance: Smoldering Infections Immunological privileged sites Intracelluar Bridges Suppression of MHC Class I 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? E (Effector Cell) CTL MHC Class I HIV antigen T (Target Cell) Y  Staining using a-perforin Abs HIV-specific CTL SAV

Mechanisms of persistence and escape from immune surveillance: Smoldering Infections Immunological privileged sites Intracelluar Bridges Suppression of MHC Class I 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

Mechanisms of persistence and escape from immune surveillance: Smoldering Infections Immunological privileged sites Intracelluar Bridges Suppression of MHC Class I Expression Infectious Immune Complexes Impaired CTL function Antigenic variation Selection for neutralization resistance; allows for viral persistence in the presence of Ab response In vivo selection for neutralization escape variants Ab escape EIAV --Immune sera neutralizes virus from earlier time points but not concurrent virus or virus thereafter --evidence of antigenic drift, and explains viral persistence even in the face of Ab response

Mechanisms of persistence and escape from immune surveillance: Smoldering Infections Indicates humoral immunity plays a role in viral clearance Immunological privileged sites Intracelluar Bridges Suppression of MHC Class I Expression Infectious Immune Complexes Impaired CTL function Antigenic variation Selection for neutralization resistance; allows for viral persistence in the presence of Ab response In vivo selection for neutralization escape variants Ab escape EIAV LCMV Log10 titer/ml Viremia Neutralizing Ab titer

Mechanisms of persistence and escape from immune surveillance: Smoldering Infections Immunological privileged sites Intracelluar Bridges Suppression of MHC Class I Expression Infectious Immune Complexes Impaired CTL function Antigenic variation Selection for neutralization resistance; allows for viral persistence in the presence of Ab response In vivo selection for neutralization escape variants Ab escape EIAV LCMV HIV Viral Env from Day 16 138 212 381 772 X.Wei. et. al. (2003) Antibody neutralization and escape by HIV. Nature 422:307

Mechanisms of persistence and escape from immune surveillance: Smoldering Infections Immunological privileged sites Intracelluar Bridges Suppression of MHC Class I Expression Infectious Immune Complexes Impaired CTL function Antigenic variation Selection for neutralization resistance; allows for viral persistence in the presence of Ab response In vivo selection for neutralization escape variants Ab escape EIAV LCMV HIV CTL escape HIV, LCMV

DNA vaccinated rhesus macaques challenged with pathogenic SHIV 89.6P ? 20 weeks

single amino acid change CTL escape via single amino acid change in immunodominant CTL epitope Viral Load CD4 T-cell count p11C Tetramer (Immunodominant) p41A tetramer p68A tetramer weeks

Diseases associated with persistant viral infections: selected examples