Fragenkatalog

• non-enveloped DNA virus

• entry via endocytosis

• replication in nucleus

• immediate early (E1A cell cylce protein →S phase), early (viral polymerase) and late (structural proteins) gene expression

• assembly in nucleus

• release by lysis

• enveloped DNA virus

• ccc DNA

• replication in nucleus

• progenmoic RNA

• reverse transcription in virion (RNA→ DNA)

• release by nonlytic exocytosis

Scheller: fusion of cells, mediated by viral envelope proteins, often seen as CPE in cell cutures infected with enveloped viruses

_______

• enveloped viruses enter their host cell by membrane fusion

• fusion of viral envelope and cell membrane is triggered by the envelope proteins

• after cells are infected with enveloped viruses, they express viral proteins on their surface

• close contact to uninfected cells can lead to cell/cell membrane fusion and the formation of syncytia

Syncytium formation is a classical CPE of enveloped viruses.

= Cytopathic Effect

sigmoid curve, read the concentration in the turning point

______________________________

• (The Endpoint-Dilution Assay)

• Cell culture infectious dose concentration of virus at which point 50% of the cells are infected

Scheller:

physical titer: measurement of viral particles (protein or genome; by ELISA or PCR);

biological titer: determined from infection experiments;

physical titer is usually higher than biological titer:

biological Titer measures only intact/infectious particles and is also dependent on the type of cell culture being used

_____________

• General: The virus titer is the concentration of a virus in a suspension.

• infectious titer/biologial titer

  • concenration of infectious particles in a suspension (depends on test system, e.g. celltype

  • test via Plaque assay, transformation assay or flow cyometry

• physical titer

  • concentration of infectious and noninfectious particles

  • meassure by ELISA or TaqMan-PCR

Scheller:

PCR with 2 primers and a probe. Probe carries fluorochrome and quencher. Taq Pol has exonuclease activity and digests probe if it binds to PCR product. Fluorochrome gets released from quencher and fluorescence is measured in a light cycler

_________________

• specific technique for amplification and simultaneous quantification of DNA content

• specific flurorescence signal by dual-labeled probes ("reporter" and "quencher").

• the reporter is released by 5´-exonuclease activity of the Taq polymerase (hydrolysis of the probe); destroying the conncection to the quencher

  • Release of the reporter leads to fluorescence of the reporter (light is no longer absorbed by the quencher)

  • High specificity (PCR only provides signal if both primers and probe match)

• Standard procedure to quantify viral DNA

• (working with RNA-Viruses requires reverse transcription of the RNA genome into DNA prior to PCR)

Scheller:

receptor: viral entry;

coreceptor: assists in viral entry,

attachment factor: lose aaachment of virus to cell

______________

• Attachment factor or adhesion receptor are common extracellular molecules that make the first contact to a virus which increase the probability of the virus to stay at the cell surface

  • They increase the probability that the virus stays at the cell surface e.g. heparane sulfate or silaic acid

• Receptor: mediates viral entry into the cell, determines tropism of virus (the virus can only enter cells that express this molecule), influence host range e.g. CD4

• Coreceptor: second binding receptor, needed for viral entry e.g. CCR5

Dissociation constant, that describes the binding strenght of a ligand/receptor-binding

HIV:

Binding of gp120 to CD4/CCR5 triggers a conformation change in gp120 that enables the attack of gp41 to the host membrane

Influenza:

A drop in pH in the endosome triggers a conformation change in measles H protein that enables the attack of measles fusion protein to the host membrane

Measles:

after receptor binding, the viral H protein undergoes a conformational change allowing the F protein to attack the host membrane

• Enveloped (taken up by membrane fusion: directly at plasma membrane or in endosome)

  • HIV/measles: fusion with cell membrane

  • influenza: uptake via endocytosis; fusion with endosomal membrane

• Non-enveloped (always taken up by endocytosis)

  • Adenovirusues: uptake by endocytosis and disruption of the endosomal membrane (via protein VI)

Scheller:

RBD. Receptor binding domain, TM: trans membrane domain; extracellular region is usually glycosylated, intracellular not

________________________

Non-enveloped virus: Surface structures (grooves and loops) and fibers like spikes

Enveloped virus: Transmembrane glycoproteins: mushroom with receptor binding domain (klappt weg) and fusion domain (nun chuck domain)

• derived from late endosome after further pH decrease

• contain proteases, nucleases and lipase

• have a pH of 4.5 to 5

How to get to mRNA from the genome

All viruses must produce mRNA that can be translated by cellular ribosomes. In his classification system, the unique pathways from various viral genomes to mRNA define specific virus classes on the basis of the nature and polarity o their genome.

Scheller

HAV, HBV, HCV, HDV, HEV

______________________

What viruses can cause hepatitis?

-Hepatitis A-D (HAV, HBV, HCV, HDV)

-ABV

-CMV

-HSV1

-VZV

-Coxsackie-Viruses

-Yellow Fever Virus

Scheller:

HBV, (HDV), HCV

______________________________

• Hepatitis B, C, D (HBV, HCV, HDV)

  • HBV: 5-10% adults, 90% newborns

  • HCV: 50-80%

  • HDV: Co- 5-10, super 70-95%

    
    

• Hepatitis A: never!

• Hepatitis E: rare (in immunocomprised)

• Sex: HBV, HDV, HCV

• smear infection/oral: HAV,HEV

• drug abuse: HBV, HCV

• HAV, HBV, (HDV)

____________________________________

• Hepatitis A (Active/passive)

• Hepatitis B (Avtive(passive)

• Hepatitis C (no)

• Hepatitis D (via HBV)

• Hepatitis E (no)

• virus contains partly dsDNA,

• DNA is completed in the cell,

  cccDNA persists as episome in nucleus

• genomic RNA transcription from cccDNA,

• reverse transcription of DNA genome from genomic RNA

• DNA incorporation into new particles,

• ds-DNA synthesis stops in virus particle if all nucleotides are being used up

• HIV: RT immediately after infection: gemonic RNA -> cDNA (early)

• HBV: RT late in replication prior to exit: genomic RNA -> genomic DNA (late)

  ____________________________________

• in retroviruses the reverse transcription from RNA to DNA takes place early in the replication cycle, before RNA enters the nucleus

• in hepadnaviruses like HBV reverse transcription takes place late when the virus particle is assembled

• the RT of HIV is important for immune response escape

• HBV RT also has a similar bad copying mechanism, however mutations are usually lethal as the genome is rather small and overlapping

• Nucleoside analogs (RT-inhibitors)

__________________________________________

Valentin

• with reverse transcription inhibitors like Tenofovir

  • it is a nucleotide analogon which lacks the 3’ OH causing chain termination during HBV rplication

  • lifelong therapy because cccDNA is stable in the liver and the treatment does not lead to anti HBS production

• with vaccination

  • with recombinant HBs antigen

  • recommended for infants since 1995

_____________________________Kim

--> no elimination

—> HBV therapy is life-long. It can protect the liver but it cannot eliminate the virus from the body.

1. cccDNA is very stable and last lifelong in the liver

2. The therapy does not trigger anti-HBS antibodies: no immunity is being formed to control HBV replication when therapy is being stopped.

Hepatitis-B-Therapy

Reduction of viral load, preservation of the liver, but no cure.

Substances: Reverse transcriptase inhibitors

Vaccine:

• Vaccination with recombinant HBs antigen (inactivated vaccination)

• For infants: Recommended since 1995 as part of basic immunization

• Recommended for at-risk groups, incl. health care providers

• For adults: Triple vaccination (0 - 1 - 6 months)

• For health care workers and other risk groups: HBs antibody control 1 - 2 months after last vaccination.

• HBs-Ak < 100 IU/l: immediate booster vaccination.

• HBs-Ak ≥ 100 IU/l: titer control after 10 years, booster vaccination if < 100 IU

• +ssRNA genome, gets directly translated at ribosome after infection,

• dsRNA

• new +ssRNA genomes transcribed from the dsRNA,

• incorporation into new particles

• nucleus not involved

  
  

Untranslated region

• similarities: +ssRNA;

• differences: no cap but IRES, no poly A but UTR

  
  

__________________________________

• mRNA has a cap structure, HCV has an IRES site

• mRNA is monocistronic, HCV-RNA is also monocistronic but resulting in a large precurser protein that is cleaved into smaller proteins by proteases

• mRNA has a poly A tail, HCV RNA has no poly A tail but a 3’ UTR

• normal mRNAs

  • 5’ cap

  • 5’ UTR

  • start codon (AUG)

  • ORF

  • stop codon (UAA, UGA, UAG)

  • 3’ UTR

  • PolyAtail

  • usually monocystronic (only contains info for one protein)

  • translation initiation is cap dependent

• ssRNA genome of HCV

  • 5’ UTR

  • no cap but IRES after 5`UTR

  • ORF

  • 3’ UTR

  • polycistronic RNA → individual proteins produced by proteolytic cleavage of large precurser protein

Initiation of translation

• mimics the function of the 5’cap of mRNAs In molecular biology?

• Allows expression of 2 different proteins (not a fusionprotein) under control of one promotor: promotor- geneX-IRES-geneY.

____________________________________________

• An IRES (internal ribosomal entry site)

  • structure within an mRNA molecule that allows the binding of the 40S subunit in the absence of a cap structure

IRES in molecular biology

• used to create polycistronic mRNAs

• frequently used to coexpress a reporter-gene or resistance-gene together with a gene of interest with only one promotor needed

• Nucleoside analogs, protease inhibitors, NS5B- inhibitors

_______________________________________________________________________________

Combination of 2-3 of the following substance classes is used.

• Protease inhibitor (…previr)

• Polymerase inhibitora (…buvir)

• NS5A-Inhibitors (…asvir) (NS5A regulates replication and virus assembly

• Ribavirin (change of the equilibrium of the intracellular nucleotide concentration)

Therapy takes 8-12 weeks, then the patient is cured. Due to lack of any persistence mechanism, HCV is completely eradicated from the body after therapy

• HCV has no mechanism of persistence. If replication is blocked, the virus is gone. HBV has a mechanism of persistence: ccc-DNA persists lifelong in nuclei of hepatocytes

• RT

• X

• surface

• core

Virus with RNA genome, RT, integrates into host genome

___________________________

• viruses that have a RNA genome

• reverse transcribe their genome into DNA

• the DNA genome is integrated into the cellular genome → necessary step of replication

• HIV belongs to the genus Lentiviruses

• Lentiviruses belong to the subfamily of orthoretroviruses containing alpha, beta, gamma, delta and epsilon retroviruses besides Lentiviruses

1. virus particle with RNA

2. entry into cell

3. release of RNA

4. RT of RNA into cDNA

5. integration of cDNA into host genome (provirus)

6. transcription of provirus: translation of viral proteins and transcription of genomic RNA

7. assembly and packaging

8. budding at cell membrane

• RNA-dependent DNA polymerase activity (synthesis of 1st DNA strand)

• DNA-dependent DNA polymerase activity (synthesis of 2nd DNA strand)

• Rnase H (heteroduplex exonuclease) activity (degradation of RNA strand)

• RNA genome

• tRNA primer bind at PBS

• synthesis of first cDNA strand by RT

• degradation of RNA in DNA/RNA heteroduplex by RT

• synthesis of second cDNA strand using parts of the original RNA template as primer and removal of RNA primer by RT

Cleavage of Gag proteins to allow maturation of viral particles

Integration of HIV cDNA into host genome

HIV: RT immediately after infection: gemonic RNA -> cDNA (early).

HBV: RT late in replication prior to exit: genomic RNA -> genomic DNA (late)

___________________

• in HIV: early reverse transcription to DNA, before integration into the host genome // RT early (+) ssRNA genome into cDNA

• in HBV late reverse transcription inside of the assembled virus particle// RT late ssRNA genome into dsDNA

• productive infection: provirus is transcriptionally active so that new viral particles are being produced

• latent infection: provirus is transcriptionally inactive and no viral particles are being produced

• reactivation: switch from latent to productive infection

Zoonitic transmission from chimpanzee (SIVcpz)

HIV infects CD4+ T cells, depletion of CD4_t cells over time, after 10 years (on average) AIDS

_________________

• Pathogenesis:

  • Depletion of CD4 cells in the intestine (acute phase)

  • Continuous loss of CD4 cells due to immune activation/depletion anti-HIV CTL apoptosis

  • High viral load correlates with rapid progression

  • High immune activation correlates with rapid progression

  • Rare: long-term nonprogression (control of infection for decades) 

combination therapy of different antiviral substances

• RT inhibitors

• Integrase inhibitors

• protease inhibitors)

_______________

Therapy:

Therapy is usually being started with one of these combinations

A combination of 3 NRTI is inferior compared to the above-described combinations.

• HIV therapy is always a combination therapy

  • 2 NRTI +1 NNRTI

  • 2 NRTI +1 PI

  • 2 NRTI +1 INI

• Start of therapy immediately after positive diagnosis

• Before therapy start: genotypic resistance test

• PCR to monitor viral load during therapy

• Therapy goal: reduction of viral load below detection limit

• Duration of therapy: unlimited, no cure possible

• Therapy improves immune system (CD4-count in blood increases within months to years to normal values: approx. 1000/ml, CD4-count in intestinal flora increases much slower), prevents AIDS, prevents HAND

• Life expectancy under therapy: normal

• HIV quasispecies with high antigenic diversity

• there is no immunity with HIV after infection

• superinfections possible even after long time

• receptor CD4

• coreceptor CCR5 or CXCR4

• tropism:

  • CD4 pos T cells

  • macrophages

  • dentritic cells

  • microglia

⇒ mainly production in activated T helper cells

• retrovirus genes:

  • gag: encodes capsid and matrix

  • pol: encodes PR, RT and IN

  • env: encodes envelope proteins

• accessory genes

  • tat, nef → regulatory proteins

• transmission: sexual, parenterally, during birth, breast milk

• incubation period: 1-6 weeks

• acute phase with general symptoms

• symptom free phase for 2-12 years with increased T helper cell loss

• without treatment AIDS (100%) and HAND (30%)

The virus (its particles or genomes) remain in the body over a long time or lifelong

• the virus persists but does not replicate

• latency controlled on genome level e.g. circular viral DNA of Herpesvirues or integrated DNA of HIV

• active viral replication

• viral particles are formed

switch from latency back to productive infection

refers to a state „without symptoms“. Not to be confused with virological latency.

Examples: HIV-infection has a period of 1-10 years of „clinical latency“, but with ongoing, productive infection.

1. acute infection (influenza, SARS- CoV-2)

2. chronic infection (HIV, HCV (80%), HBV (5%),

3. latent infection with episodes of reactivation (herpesviruses)

   
   

   => Chronic and latent infections are persistent infections.

4. slow infection (HIV)

alpha-herpes in sensible neuroganglia, beta-herpes in lymphocytes/monocytes and kidney and salivary glands, gamma- herpesviruses in B lymphocytes

____________________________

1. HHV1-8

   • alpha subfamiliy - persistence in neuroganglia (non neural cells in CNS)

     • HHV1 (HSV1)

     • HHV2 (HSV2)

     • HHV3 (VZV)

   • beta subfamily - persistance in lymphocytes/ monocytes in the kideney and salivary glands

     • HHV5 (CMV)

     • HHV6

     • HHV7

   • gamma subfamily- persistence in B lymphocytes

     • HHV4 (EBV)

     • HHV8

herpes labialis, herpes genitalis

______________________

• HSV-1: cold sores

  • primary infection: stomatitis aphtosa (inflammation mundschleimhaut)

  • reactivation: mainly herpes labialis but also herpes genitalis

  ______________________________Zusatz:

• HSV-2: genital ulcer

  • primary infection: vulvovaginitis herpetica (vaginalentzündung)

  • reactivation: mainly herpes genitalis but also labialis

• VZV: chicken pox or varizell zooster (windpocken, Gürtelrose

• attachment to heparane sulfate

• binding to HVEM (herpes virus entry mediator) receptor (TNFa superfamily)

• membrane fusion mediated by envelope

• capsid transport to nucleus

• release of HSV-DNA into nucleus via nuclear pore

Concept of IE and E and late:

HSV-1 packaging:

• The concatemeric genome is pushed by the terminase protein into an empty capsid

• Terminase also cleaves the concatemeric pregenome once a full copy of HSV genome is packed into the capsid

• (packaging can be inhibited for the herpesvirus CMV using the drug Letermovir that is being used as CMV prophylaxis after stem cell transplantation

• concatemeric genome is pushed by the terminase into an empty capsid

• terminase cleaves genome once one full progenome is packed into one capsid

• nucleoside analog lacking 3’-OH

• prodrug

• phospoorylated by alpha-herpes TK (Monophosphate),

• phosphorylated by cellular kinases (triphosphate),

• incorporation into herpes DNA during DNA replication,

• chain termination due to lack of 3’ OH

retrograde transport in neurons, HSV DNA as episome in nulceus, LATs are processed into miRNAs; miRNAs suppress transactivators ICP4 and ICP0, switch from immediate early to early genes not possible, virus stays in latency

_____

Latency and reactivation

• epithelial cells support productive infection

• nerve terminals are infected

• axonal (retrograde) transport of the capsid into the cell body in the ganglia

• here viral genome delivered to nucleus

• viral episome is stably maintained in the neural nucleus

• latent infection established

• upon reactivation new particles produced and transported by (anterograde) transport back to the epithelium

1. During latency, 4 latency-associated transcripts (LAT) are transcribed

2. They are processed into miRNAs that suppress the transactivators ICP4 and ICP0 that normally would activate the transition from IE to E phase.

• EBV → name mononucleosis derived from high number of lymphocytes in the blood during EBV infection

• similar symptoms as with HIV

• swollen lymph nodes

• pharyngitis (Entzündung des Rachens)

• fever

• feeling tired

• Rash (Hautauschlag) when treated with antibiotics

• EBV infection linked to multiple sclerosis

infectious mononucleosis/Pfeiffersches Drüsenfieber/kissing disease : fiever, swallen lympg nodes, sore throat, fatigue, skin rash

rolling circle

• G1

• S

• G2

• M

inhibit cell proliferation (S-phase); loss of function can lead to tumors

p53 and Rb inhibit cell proliferation by controlling S phase

(Protooncogenes: activate cell proliferation; overactivation can lead to tumors; M-phase)

• warts (HPV1 and other)

• condyloma (low risk types HPV 6 and 11 and other)

• cervix carcinoma (high risk types HPV 16 and 18 and other)

viral persistence (10%) and cancer (0.8%)

• when infected with high risk subtype: 10% persistence (integration into cellular genome) and 0.8% cancer formation

  
  

• infection of upper epidermal cells

• E6 and E7 stimulate cell cycle entry from G0 to G1 in the upper epithelial cells

  ________________________

• HPV triggers cell proliferation also in upper layers -The E6 and E7 proteins stimulate cell cycle entry from G0 to G1 in upper epithelial cells. A benign tumor (e.g. wart) forms.

• The E6 and E7 proteins stiulate cell cycle entry from G0 to G1 also in basal cells. A malignent tumor (e.g. cervix carcinoma) forms.

• HPV E6 and E7 inhibit the tumor-suppressors p53 and Rb. This causes cell proliferation and benign tumors.

infection of epithelial cells and basal cells, E6 E7 expression/proliferation also in basal cells

_______________

• E6 and E7 proteins stimulate cell cycle entry from G0 to G1 also in basal cells → malignant tumor

• E6 and E7 inhibit immune response

• infection of basal cells consisting of stem cells

• aging skin cells enter the upper layers of the epidermis → virions are released

  
  

HPV infects the basal layer that consists of stem cells. Normally, the basal layer is the only zone in the epidermis with cell proliferation. As the aging skin cells enter the upper layers of the epidermis, viral gene expression occurs within them, and virions are released from the upper layers of the epidermis.

• episomal genome with unstable mRNA for E6/7, liale E6/7 is formed

• moderate inhibition of p53/Rb, no oncogenesis;

• integrated genome with stable mRNA for E6/7 due to lack of AUUUA, much E6/7, strong inhibition of p53/Rb, oncogenesis

• E6 → p53

  ((E6 supports proteosomal degradation of p53 via E6 AP (E6 associated protein → ubiquitin ligasse) this inhibits apoptosis))

• E7 → pRb

  
  

  ((Rb usually binds to E2F and inhibits it; if E2F is active it stimulates the expression of S phase specific genes; this inhibits cell cycle progression ))

high risk HPV infection can be prevented by vaccination; the vaccine does not protect against CC if infection is already persistent because the vaccine does not contain E6/7, no immunity against E6/7 is formed

______________

• by vaccination

• HVP16 and 18 are responsible for 70% of the cervical carcinomas

• vaccine contains protection from these 2 risk groups and additional HPV that cause 90 % of all condylomas and 90% of all cervical carcinomas→ risk significantly reduced´

yes

..know that isolated +ssRNA genomes are infectious after transfection

• Isolate RNA from the virus

• add isolated RNA on a cell culture

• new virus particles are created

• 3’ PolyA tail

• 5’ Cap

• angiotensin-converting enzyme 2

• entry receptor that binds to the spike protein

• expression in the respiratory epithel of upper airways by Pneumocytes and Alveolarmacrophages, blood vessels and the kidney

• cofactor TMPRSS2

• Example 1

  • as an exception, in picornaviruses RNA replication is dependent on a primer

  • Vpg serves as a primer

  • 3CD assembles on CRE element and Vpg binds; Vpg is then uridylylated by 3Dpol which turns it into a primer→ binds to polyA tail

• Example 2

  • in noroviruses or sapoviruses

  • Vpg serves as a cap replacement

⇒ hier könnte man wenn man Zeit hat noch den genauen Mechanismus verstehen

• virus with non infectious RNA associated with functional polymerase complex

• all enveloped

• max 19 kb genome

• transcription via sequential stop-start mechanism

ssRNA viruses with nonsegmented genomes

• RSV (lower respiratory tract disease)

• parainfluenzavirus (respiratory disease)

• measles virus (measles)

• mumps virus (mumps)

require RNA-dependent RNA Polymerase

_____________

• yes

• minimum transcription of Large protein L, Phosphoprotein P and nukleoprotein N or NP

• ...know that isolated -ssRNA genomes are non-infectious after transfection

  
  

• ...important ARI (acute respiratory infections) viruses:

• influenza,

• Corona,

• RSV,

• Adeno,

• HMPV,

• PIV,

• rhino

• coxsackie

...symptoms: respiratory tract symptoms include rhinitis, pharyngitis, bronchitis, pneumonia

____________________

• rhinitis (Nasenschleimhautentzündung)

• pharyngitis (Entzündung des Rachens)

• bronchitis (Entzündung der Schleimhaut in den Bronchien)

• pneumonia (Lungenentzündung)

...RSV cause of severe respiratory infections and hospitalization in <2 year old children

• winter: influenza, Corona, RSV

• summer: coxsackie virus

• all year: all others (Adeno, HMPV, PIV, rhino)

  
  

  
  

  __________________________

• apical release:

  • RSV

  • influenza

  ⇒ local infection

• basal release

  • VSV

  ⇒ systemic infection

A: aquatic birds,

B: humans arte the only host

point mutations in the genome are called antigen drift;

reassorting is called antigen shift

• influenza genome is segmented;

• infection of one cell with two different strains;

• Segmented influenza genome gets reassorted (e.g the new virus contains 3 segments of strain X and 5 segments of strain Y)

cause of severe respiratory infections and hospitalization in old people

• poultry, pig as “mixing vessel”, spread into human popula%on

humans are the only host

• Serotypes are determined by H and N antigens.

• Different antigen-families exist that can be combined.

• HxNy indicates which antigen families are present in a particular strain

• prevalence: proportion of infected people in a population

  • e.g. prevalence of HIV infections in Germany is 0.1%

• incedence: number of new infections in a population in a certain time span

  • e.g. incidence of HIV infections in Germany is 3500 cases per year

• morbidity: proportion of people who get sick in a population caused by the the infectious agent

  • e.g. 24.000 disease cases per 100.000 people

• mortality: proportion of people who die in a population caused by the infectious agent

  • e.g. 120 deaths per 100000 people

• CFR (case fatality rate) or lethality: proportion of people who die among people with diagnosed infection

  • e.g. 9000 deaths per 300000 diagnosed infections =3%

• endemic: normal/usual occurence of a certain infectious disease in a certain population

  • e.g. influenza/ SARSCoV-2 in 2022

• epidemic: unusally stong and temporally limited occurrence(accumulation of a certain infection disease in a certain area

  • e.g. ebola in west africa in 2014

• Pandemic: an endemic of a new pathogen that spans countiers/continents

  • e.g. SARS-CoV2 in 2019-2020

• Weekly incidence: the number of new infections per week is given. If the number of

  infections increase over time, weekly incidence increases. If the number of infections

  over time declines, weekly incidence decreases.

• Cumulative incidence: the number of new infections in each week is added to the

  number of infections in the past.. If the number of infec%ons increase over %me, cumulative incidence increases. If the number of infections over time declines, cumulative incidence still increases. If the number of infections over time is 0, cumulative incidence stays on a plateau

_____________________________________________________________________

• weekly incidence shows number of cases per week

• cumulative incedence shows total cases over time → adds up number of new cases

  • at first look more impressive, good to see which proportion of individuums already hat contact with the pathogen

  • incedence of 0 looks like a horizontal line

basic reproductive number: average number of infections that are caused by 1 infected individual

KNOW THAT there is a seasonal variation of R0 and herd immunity for respiratory infections

_________________________________________________

• R0: baseline reproduction rate

  • average number of subsequent infections caused by one case of infection in a population that is immunologically naive to the pathogen

  • usually calculated from increase in incidence numbers in the early weeks of a pandemic or epidemic

  • e.g. Masern R0=12-18, Polio 5-7, SARS/HIV 2-5

• Herd immunity is reached if a high-enough amount of people in a population is immune so that the virus can no longer spread in the popultion with a positive R

  
  

  ________________________________________

• herd immunity is archived when a sufficiently high proportion of the population has become immune to a pathogen through infection or vaccination so that the incidence of the new infections declines

  • if reached pathogen can no longer spread

  • but if herd immunity is reached and the pathogen has enough genetic flexibility, immune escape variants wil emerge

• yes

• most respiratory viruses have a higher R0 in winter and a lower R0 in summer

• therefore also herd immunity oscillates → R0 reduced in summer compard to winter by 40%

• archievin herd immunity is dependent on:

  • contagiousness of the pathogen (R0)

  • duration of immunity

  • pathogen variability and its ability to generate immune escape muations

R0-1 / R0

___________

0,8 → 80%

scheller:

Mathematical model to predocit numbers of infections in a population

S = Numbr of susceptible

I = Number of Infected

R = number of restistent/ immune people

_______________________________

• Susceptible, Infected, resistant model

• susceptibe nuber decreases, infectios peak and resistant number increases

• oversimplistic but useful model forthe first weeks of an endemic

  • doesnt respect decreasing immunity

  • doesnt inculde seasonal variation of R0

  • does not include changes in behavior of population

  • does not include effects of microenvironments like kindergardens or eldery homes

  • no long term predictions possible

better mathematical model to predict numbers of infections in a population:

4 key parameters.

S = number of susceptible,

E= exposed but not yet infectious,

I = numbers of infected,

R: numbers of resistant/Immune people

scheller:

Mathematical model to predocit numbers of infections in a population

S = Numbr of susceptible

I = Number of Infected

R = number of restistent/ immune people

_______________________________

• Susceptible, Infected, resistant model

• susceptibe nuber decreases, infectios peak and resistant number increases

• oversimplistic but useful model forthe first weeks of an endemic

  • doesnt respect decreasing immunity

  • doesnt inculde seasonal variation of R0

  • does not include changes in behavior of population

  • does not include effects of microenvironments like kindergardens or eldery homes

  • no long term predictions possible

• more deaths than usually expected

  
  

  example: (deaths per week of this year)-(average

  number of deaths of the same week of the last 4 years)

_______________________________________________________________

• how many more people died in a certain week of a year compared to the average number in the same week of the last few years

  • mostly reference period of last 4 years used

  • calculation: (number of deaths within a week)- average number of deaths in this week of the past 4 years)

What is

• RRR: relative response rate, good parameter for treatment

• .ARR: absolute response rate, good parameter for prophylaxis

• NNT: number needed to treat in order to prevent 1 disease case

• .NNV: number needed to vaccinate in order to prevent 1 infec%on

  4-field analysis: control treatment without disease, control treatment with disease,

  ac%ve treatment without disease, ac%ve treatment with disease (example 90%, 10%

  versus 98% versus 2%)

Calculations from 4-field analysis:

• ...RRR: (10%-2%)/10% = treatment reduces risk by 80%

• ...ARR: 10%-2% = treatment reduces risk by 8%-points

• ...NNT: 1/ARR = 1/0.08 = 12.5

• ...NNV: 1/ARR = 1/0.08 = 12.5

______________________________________

• event = get sick

• RRR= relative response rate: good index for effectiveness when you are already infected

RRR= (0.03-0.05)/0.05= -0.4 → treatment reduces the risk by 40%

ARR=absolute response rate: goog for profilaxe → e.g. malarone → chance to get malaria in germany super low in the first place

ARR=(0.03-0.05)= -0.02 treatment reduces the absolute risk by 2% points

NTT= number needed to treat

NTT= 1/ARR= 1/0.02=50 → 1 of 50 patients profits from treatment

Know that there is a Seasonal variation of R0 and herd immunity for respiratory infections

• RRR is a good parameter for treatment

• ARR is a good parameter for prophylaxis

describes the average time between the infection of a patient and the transmission to the next patient

Attempt to slow down infection spread in a population e.g. by wearing masks etc

total number of deaths/illness until herd immunity is reached is not changed, but peak numbers are lower

concept to protect hospitals from overcrowding

• intrinsic immunity: immune response within a single cell e.g. TLR

• Innate immunity: complex immune response directed at specific structures, e.g. NK killing of missing self

• adaptiv immune response: complex immune response directed at variable structures e.g. T- and B-cell response

• unique ligands or unique loci

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Know how receptors from intrinsic immunity can recognize foreign structures (unique ligands or unique loci)

• unique ligands or unique loci

• inhibition of transcription

• inhibition of 5’ end dependent translation

• Inhibition of ER-to Golgi traffic

• Syncytium formation

• Inhibition of transport of host RNA

• Depolymerization of the cytoskeleton

• TLR3 ds DNA,

  TLR7,8: ssRNA

  
  

  _______________

• Tlr3. dsRNA

• Tlr7/8: ssRNA

• Tlr9 CpG DNA, unmethylated CpG oligonucleotides

⇒ recognize only foreign ligands by only recognizing unique loci or being active at unique locations→ main effector response is interferon and cytokine production

Know that TLR-signaling ultimately results in Interferon I production

• interferon 1 production

• now that Interferon-alpha can trigger an antiviral state

• intrinsic:

  • TLRs, interferon, tetherin, apobec3G, epigenetic silencing, induction apoptosis

• Innate:

  • cellular components (granulocytes, mactrophages, NK cells)

  • humoral components (complement, interleukins)

• Adaptive

  • B and T cells

• MHCI: presents antigens to CD8+ T cells; present on all nucleated cells of the body

• MHCII: presents antigens to CD4+ T cells; present only on antigen-presenting cells

• Antigen-presenting cells: dendritic cells, macrophages, B cells

  
  

  ____________________________________

• MHC1:

  • expressed on all nucleated cells (all cels except erys)

  • presents antigens to CD8+ T cells (CTLs)

  • predominantly loaded with self antigens that have been expressed in the same cell; alternatively MHC1 can also be loaded with self antigens that have been taken up → cross presentation

  • loading results in priming of naive CD8+ T cells by unifected APCs or killing of infected MHC-i presenting cell

  • signals the immune cell. kill me

• MHC2:

  • expressed on APCs (DCs, Macrophages, microglia, B cells, T cells and epithelial cells)

  • presents antigen to CD4+ T cells(helper T cells)

  • predominantly loaded with self antigens that have been taken up, alternatively MHC1 can also be loaded with endogenous self antigens → signals helpme

    
    

• NK cells kill other cells that dont express enough MHC1 (missing self)

• MHC1 inhibits the NK cell from killing

• killing by perforin and granzymes

• many viruses cause downregulation of MHC1

• antiviral state shuts down protein synthesis

• tetherin attaches enveloped viruses upon exit

• APOBEC3G causes G-> A hypermutation

• epigenetic silencing shuts down viral gene expression

• apoptosis kills infected cells

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• antiviral state:

  • triggered by interferon I

  • increased cellular concentration of dsRNA-activated protein kinase Pkr in uninfected bystander cells

  • if these cells become infected with dsRNA viruses, Pkr becomes active and shuts down translation by phosphorylating and thus inhibiting eIF2a

• tetherin

  • interferon induced

  • inhibits release of enveloped viruses by sticking into the membrane bilayers

  • counteracted by HIV-1 vpu protein

• APOBEC3G (apolipoprotein B mRNA editing enzyme, catalytic polypeptide-like 3G)

  • deaminates C to U in ssDNA → this hypermutation introduces multiple stop codons

  • HIV-1 vif counteracts APOBEC3G by degradation of A3G

• epigenetic silencing

  • silencing of new DNA in the nucleus by methylation of DNA or deacetylation of histones

• apoptosis

  • infection of cells with virus increases apoptosis rate for many viruses

  • many viruses counteract this by encoding antiapoptotic proteins

    • cell surface: production of apoptosis inducing cytokines; alteration of membrane integrity

    • cytoplasm: translation inhibitiors, modification of cytoskeleton, disruption of signal transduction

    • Nucleus: DNA damage, altered gene expression, diruption of the cell cycle

      
      

• IFNa leads to increased cellular concentrations of dsRNA-activated protein kinase Pkr in uninfected bystander cells

• if these cells are infected with dsRNA, Pkr becomes active and shuts down translation by phosphorylating eIF2

Know that Interferon I can trigger an antiviral state

• interferon I (alpha and beta); no direct antiviral effect but activation of antiviral pathways; cause flu like symptomes

Clara:

MHCI ist normalerweise mit Antigenen aus der Zelle beladen MHC I mit Antigen außerhalbder Zelle

Cross penetration Ist die Fähigkeitvon Dendritischen Zellen DC MHC I Moleküle mit Antikörper zu beladen welche aufgenommen wurde Cexogeneaus antigens

Valentin

• ability of DCs, macrophages and B cells to load MHC1-molecules with antigens that have been taken up; usually MHC1 is only loaded with antigens that have been expressed in the same cell

• this is an important mechanism for the activation of naive CTLs to effector CTLs → only a few viruses infect DCs but all have to be recognized by CTLs

1. viral antigen taken up by naive B cell via BCR

2. B cell migrates to lymph node and presents antigen via MHC2 effector TH cells

3. Th cell recognizes antigens presented by MHC2 on B cells and releases activating cytokines

4. B cell develops into long living plasma cell that initially produces IgM and later IgG

5. some of these activated B cells develop into long lived memory B cells

6. Upon a secondary contact with the same virus or a booster vaccina%on, these

   memory B cells can quickly develop into new plasma cells (no need for an additional

   costimulatory signal from T cells) and produce high-affinity IgG.

   
   

1. A viral antigen is taken-up by dendritic cells

2. The DC presents parts of this antigen on MHCII and (by cross presentation) on MHCI

3. The DC migrates to the Lymph node (LN) and presents the antigen to naïve T cells

4. 4.a An antigen-specific naïve CD4+ T cell recognizes the an%gen on MHCII and develops into an effector TH cell

   4.b An antigen-specific naïve CD8+ T cell recognizes the an%gen on MHCI and develops into an effector CTL cell

5. The effector TH and CTL leave the LN and migrate to a site of inflamma%on in the body

6. The CTL recognizes infected target cells by the antigen presented on MHCI and kills the cell

7. The TH cell recognizes antigens presented by MHCII on antigen-presenting cells in the tissue (e.g. macrophages that have taken up killed infected cells) and releases ac%va%ng cytokines

8. Most of the effector CTL and TH cells die acer 2-3 days by ac%va%on-induced apoptosis

9. Some of these cells survive and become long-lived memory T cells.

10. Upon a secondary contact with the same virus, these memory cells can quickly develop into new effector cells.

• Every body cell has an intrinsic immunity to viruses or bacteria (part of the innate immune defense).

• A viral infection is recognized by dendritic cells (DCs) or macrophages via TLRs (Toll- like receptors).

• Activation of the TLRs leads to the release of interferon-alpha (IFNa).

• IFNa prepares the surrounding cells for a viral infection ("antiviral state"). Among other things, preparations are made to switch off the cell's protein biosynthesis.

• As soon as these cells are also infected, they switch off their protein biosynthesis.

• This brings viral replication to a standstill.

• In addi%on, apoptosis is triggered in the infected cells.

• In addition to interferon-alpha, the virus-actiated DCs also release TNF-a and IL-6. These cytokines trigger the produc%on of prostaglandin-2 (PGE-2) in epithelial cells, for example. PGE-2 triggers fever in the hypothalamus.

• Usually, a flu-like infection is over acer 2-3 days.

• The virus has been eliminated from the body, the release of PGE-2 decreases and we are healthy again.

• Although we have now recovered, the immune system s%ll processes the previous infection over the following weeks: adaptive immunity develops...

• IgM

  • pentameric

  • B cell receptor

  • serologic signal for fresh infection or reactivation

  • production after first contact, usually disappears again

  • important for complement activation

• IgG

  • monomeric, late production

  • important for neutralizing in plasma

  • titer stay up for long time

  • enters plazenta

  • production if antigen is in peripheral lymph nodes

    Know that IgGs can label virus-infected cells for killing via ADCC, CDC and ADCP

• IgE

• IgA

  • like IgG active in plasma and monomeric

• sIgA

  • dimer

  • important for neutralization of mucosa

  • late production when antigen is presented in mucosal lymph nodes (e.g. peyers patches)

  • Know about the crucial need for mucosal antigen delivery if an sIgA response is wanted

1. live: good CTL response, good antibody response, cannot be used in pregnancy, can cause vaccine disease;

2. antigen: weak CTL response, good antibody response, can be used in pregnancy, cannot cause vaccine disease;

3. gene-based: good CTL response, good antibody response, can be used in pregnancy, cannot cause vaccine disease

substance that activates natural immunity/DCs to trigger an immunogenic immune response.

_____________________

• substances added to vaccine to increase the immunogenicity

• activation of inflammasome → proinflammatory, acitvation of TLRs

• e.g. aluminium hydroide or Aluminium Phosphate

• can increase side effects → fever bone pain, pain at injection site etc.

• lipid nanao particles, modified RNa from mRNA vaccines and particles of vector vaccines (e.g. adenoviral particles) have a proinflammatory adjuvant effect

• proinflammatory LNPs activate DCs,

• mRNA is taken-up by cells,

• cells translate antigen,

• antigen is presented on MHCI and MHCII,

• stimulation of CD8-T cells and CD4 T cells and antibody response

• can bind to surface of pathogen and prevent receptor-binding

• Live vaccines: replication-competent, attenuated virus;

• vector vaccines: replication-incompetent, transgene viral vector encoding for antigen;

• RNA-vaccines: mRNA encoding for antigen with proinflammatory LNPs

• dead vaccine: inactivated whole virus with adjuvants;

• proteint vaccines: recombinant antigen with adjuvants

• conjugate vaccine: polysaccharide-antigen coupled to protein, adjuvans (Al(OH)3 oder AlPO4)

• polysaccharide vaccine: polysaccharide-antigen, adjuvans (Al(OH)3 oder AlPO4)

• dominantly blocks these processes, confers immunological tolerance

• Prime means first vaccination,

  • activating the adaptive immune response,

  • relatively low levels of IgG,

  • formation of memory cells;

• Boost means second (ore more) vaccination,

  • activation of memory cells,

  • high levels of IgG

• passive:

  • transfer of antibodies

  • no long- lasting immunity

  • no T-cell response

• Active:

  • delivery of antigen to the body,

  • activation of adaptive immune response and memory,

  • long-lasting

• serial passaging of wild-type virus on different cell cultures and culture conditons (e.g. temperature).

• Evolution of virus variants that are optimally adapted to cell culture.

• Selection of strains that have lost abilities to optimally replicate in the human body.

• Caution: It is possible that the attenuated vaccine regains pathogenicity in humans (e.g. live polio vaccine).

• create an immunological memory that prevents sympotoms upon the first infection

• SARS-CoV-2: Covid19

• Rhinoviruses: common cold

• HAV: hepatitis

• HCV: hepatitis

• RNA genome is infectious because it can directly be translated at the ribosome

• replication involves dsRNA because +RNA replication requires a -RNA template strand

• homologous recombination: template switching after coinfection

• can directly be translated at the ribosome into proteins

• mRNA: cap, UTR, ORF, UTR, poly A

• viral +ssRNA: often no cap, but: VPg/FAD and IRES, sometimes no poly A

Substitute for mRNA-cap

• examples for viruses that require VPg (picornaviridae, caliciviridae) or FAD (flaviviridae)

viral protein genome-linked (VPg)

Cellular furin proteases cleave S2 protein

• RNA-dependent RNA polymerase for replication, but it’s not part of the virion particle (in contrast to -ssRNA viruses) because it can be directly translated from +ssRNA genome

ACE2

variants of a virus that are genetically similar; they cluster in a phylogenetic tree

variants of a virus that can be controlled by the same immune response

(scenario lifelong immunity: if there is only 1 serotype, you get the disease only 1x in your life. If there are 2 serotypes, you can get the disease 2x in your life)

1. mumps (mumps virus)

2. measles/Masern (measles virus),

3. rubella/Röteln (rubella virus)

4. chickenpox/Windpocken (VZV)

5. Exanthema subitem/Dreitagefieber (HHV6 and HHV7)

6. erythema infectiosum/Ringelröteln (PB19)

7. Hand-Foot-Mouth-disease/HandFussMundkrankheit (enterovirus group A)

• endemic

• highly contagious

• life-long immunity

• infection via tonsils and pharyngeal lymph nodes

• systemic infection

• infection of the lungs

• release of virus via aerosols

VZV, PB19, rubella can harm the unborn child

primary infect causes chickenpox (Windpocken), reactivation causes herpes Zoster(Gürtelrose)

Mathematical model to predict numbers of infections in a populationon:

3 key parameters.

S = number of susceptible ,

I = numbers of infected,

R: numbers of resistant/Immune people