1. Viremia Presence of viruses in the blood stream – biphasic  Primary (prodromal phase of infection)  Secondary replication in target organs

2. Cell free viremia  Free virus particles in plasma  Accessible to antibodies and immune cells  Parvoviruses  Enteroviruses  Togaviruses  Flaviviruses

3. Cell associated viremia  Virus is hidden in blood cells  Protected against antibodies  Slow virus clearing  Monocytes  Herpesviruses  Retroviruses  Distemper

4.  Lymphocytes  Marek´s disease virus  EB virus  HIV  Erythrocytes  Bluetongue virus (erythroblasts)  Rift Valley fever virus  African swine fever virus  Neutrophils  Short half-life  Anti-microbial mechanisms  May contain phagocyted viruses

5. Monocytes - macrophages  Prevent access of viruses in the blood and tissues by ingestion of viruses  Antigen presenting cells  Virus replications in macrophages = Trojan horse mechanism  Virulence factor

6. Viruses replicating in macrophages  Retroviruses  Circoviruses  Flaviviruses  Coronaviruses  Arenaviruses  Togaviruses  Reoviruses

7. Samples  Serum samples  Whole blood (EDTA, heparin…)  Intermittent virus shedding

8. Respiratory tract  Primary replication  Tonsil (Aujeszky)  Epithelial cells (Influenza virus)  Alveolar macrophages (PRRS)  Secondary replication  Epithelial cells  Alveolar macrophages

9. Respiratory tract - samples  Nasal swabs (samples from upper resp. tract are often sufficient)  Conjunctival swabs  Serum (virus + antibodies)  Transport medium  Rapid transport

10. Enteric tract  Primary replication  Tonsil (enteroviruses)  Enterocytes (parvoviruses, coronaviruses)  Secondary replication  Mature enterocytes  Usually short term shedding  Some viruses replicate in the ET without causing disease (enteroviruses, FeCOv)

11. Enteric tract - samples  Rectal swabs  Feces

12. Genital tract  Transplacental infection  Cell associated viremia  Endothelial tropism  Infertility (porcine enteroviruses, BVDv)  Abortion (EHV-1, EVA, PRRS, PPV, CHV)

13. Genital tract - samples  Aborted fetuses (EHV-1, EVA, PPV, PRRS, BVDv)  Placenta (EHV-1)  Serum (virus or antibodies)

14. Infection of skin  Protection of skin surface  Keratinisation  Low pH  Permanent renewing  Infection through skin  abrasions, wounds  microtraumatisation  blood sucking insect  Langerhans cells (epidermis)  Lymphatic system, nerve endings

15. Primary skin infections  Papillomaviruses  Ovine Poxviruses  Vesicular swine disease

16. Secondary skin infections  generalised infections, hematogenous spread (poxviruses, FMDV, distemper…)  nerves (herpes simplex, herpes zooster)  Marek´s disease virus –virus dissemination by infected keratinised cells

17. Passive role of skin in virus infections  Entry for viruses transmitted by blood sucking insect  Equine infectious anemia  Myxoma virus  African swine fever virus  Equine encephalitis  Ski lesions due to immunopathologic reactions  PDNS (porcine circovirus)

18. Skin infection - samples  Tissue for histology (papillomaviruses)  Vesicles, vesicular fluid (FMDv)  Serum samples

19. CNS infections  Crossing hematoencephal barrier  By neuronal axons  Infection of endothelial cells  Through capillaries  Infected leukocytes (rare)

20. Some viruses causing encephalitis  Rabies  Distemper (old dog encephalitis)  Tick borne encepalitis  Herpesviral encephalitis  EHV-1  Aujeszky disease virus  Maedi-Visna  Teschoviruses  Borna virus disease

21. CNS infections - samples  Serum (antibodies)  Cerebrospinal fluid (antibodies or virus) Occasional samples  Saliva (rabies)  Section samples are usually necessary

22. Eye infection  Conjunctiva  Distemper, herpesviruses, EVA  Virus replication in the eye  EHV-1, EHV-2  Immunocomplex  CAV-1, La piedad, EIA Samples: swabs, serum

23. When to take samples? Viremia NK cell killing

24. When to take samples? Viremia IgM IgG

25. Diagnostic virology How do we diagnose viral diseases? This can be achieved : Directly – detecting the virus or viral products (proteins, nucleic acids) Indirectly – detecting an immunological response to the virus (antibodies)

26. Direct methods  Virus isolation  Virus visualisation (EM)  Direct antigen detection  DNA/RNA detection

27. Indirect methods  Antibody detection (serology)  Lymphocyte activation  Cytokine release

28. Virus isolation  Virus has to remain alive  Transport medium  Rapid transport  Keep the sample at 4 o C or freeze it at low temperature (at least -50 o C)

30. Virus visualisation - EM  Suitable for viruses with characteristic morphological features  Highly concentrated virus (rota, corona, astroviruses…)

31. Direct antigen detection

32. DNA/RNA detection

33. Antibody detection