Transmissible Disease Testing Canadian Blood Services

1. Transmissible Disease TestingCanadian Blood Services Transfusion Medicine Residents March 16, 2010 Dr. Margaret Fearon & Mr. Vito Scalia

3. Donor Selection • Donor health assessment questionnaire • Questions 1-13 completed by donor alone • Questions 14-29 administered orally by nurse • Donor asked about ~ 85 different items related to health, medication, travel, lifestyle. Identical at each donation

4. Blood Donor Screening • Donor testing • HIV1/2 Antibody (Ab) and nucleic acid testing (NAT) • HBV HBsAg, anti-HBc • HCV Ab and NAT • HTLV1/2 Ab • WNV NAT • Syphilis Ab • CMV Ab (selected units) • Chagas Ab (selective donor testing) May 2010

5. Confirmed TD Positive Allogeneic Donors 2002 - 2008

6. Estimated Risk of Transfusion Transmitted Diseases - Residual Risk (per million donations) 95% CI • HIV 1: 7.8 million • Hepatitis C 1: 2.3 million • Hepatitis B 1: 153,000 • HTLV 1: 4.3 million

8. Hepatitis B Virus

9. Hepatitis B • DNA virus, hepadnavirus family • Transmission • Sexual – most common • Household contact • Perinatal (mother to baby) • Injection drug use • Nosocomial (needlestick injury in health care workers)

10. Hepatitis B • Clinical • Incubation 45-180 days (avg. 60-90 days) • Asymptomatic in 50 – 70% • Symptomatic – anorexia, nausea, vomiting, jaundice • Chronic carriage in 0.1 – 20% (90% in infected infants) • 15 – 25% of chronic carriers develop cirrhosis or hepatocellular carcinoma • Prevention and Treatment • Hepatitis B vaccine • Hepatitis B Immune Globulin (HBIG) • Treatment with antiretroviral agents, interferon (some success in chronic carriers)

11. WHO estimates more than 2 billion infected worldwide

12. Hepatitis B Markers SEROLOGICAL MARKERS • Hepatitis B surface antigen (HBsAG)1 • Hepatitis B surface antibody (anti-HBs)2 • Hepatitis B core antibody (anti-HBc)1 • Hepatitis B core IgM (aHBcIgM) • Hepatitis B e antigen (HBeAg) • Hepatitis B e antibody (aHBe) VIRAL DNA • Hepatitis B DNA (HBV DNA)2 1CBS Screening test 2Supplemental test

13. Hepatitis B Serological Profile Resolved Infection

14. Hepatitis C Virus capsid envelope protein protease/helicase RNA-dependent RNA polymerase c22 c-100 33c 5’ 3’ NS3 NS5 core E1 E2 NS2 NS4 hypervariable region

16. Sources of Infection forPersons With Hepatitis C Injecting drug use 60% Sexual 15% Injection drug use 60% Transfusion 10% (before screening) Occupational 4% Other 1%* Unknown10% Source: Centers for Disease Control and Prevention * Nosocomial; iatrogenic; perinatal

17. Hepatitis C - Clinical Features Incubation period: Average 6-7 wks Range 2-26 wks Clinical illness (jaundice): 30-40% (20-30%) Chronic hepatitis: 70% Persistent infection: 85-100% Immunity: No protective antibody response identified

18. Hepatitis C Virus Infection Typical Serologic Course anti-HCV HCV antibody Symptoms Titre ALT ALT Normal Months Years 6 1 2 3 4 0 1 2 3 4 5 Time after Exposure

19. HIV

20. HIV • Acquired Immunodeficiency syndrome first described in 1981 • HIV-1 isolated in 1984, and HIV-2 in 1986 • Enveloped RNA retrovirus • Clinical • Seroconversion – flu-like illnesss – approx. 10% of patients, 2 – 3 wks post exposure • Asymptomatic ‘latent period’ – several months to years • AIDS related complex • AIDS

21. HIV Serological Profile IgM p24 antigen ENV ENV GAG POL Viral load CD4 pre-antibody Exposure Infection 3 Months

22. HTLV I and II • HTLV I • Caribbean, southern Japan, parts of Africa • Prevalence increases with age, 2X commoner in females • Associated with: • adult T cell leukemia (1:500 who are seropositive, develops after 15 – 20 years) • Tropical spastic paraperesis • HTLV II • IVDU

23. Syphilis • Primary Syphilis • Primary lesion – chancre (painless) at site of innoculation • Secondary or Disseminated Syphilis • Rash - Macular, maculopapular, papular or pustular • Latent Syphilis • Early latent • Late latent • Late or Tertiary Syphilis • Neurosyphilis – asymptomatic or symptomatic (delusions, hallucinations, personality change, seizures, ataxia) • Cardiovascular syphilis – aortic aneurysm • Gummatous syphilis – skin, bone, mucous membranes Syphilis (bacteria) spirochaetes by Immunoflourescence

24. Immune Response in Syphilis Primary Anti-lipoidal antibody Anti-treponemal IgG Anti-treponemal IgM Secondary Early Latent Late Latent Late

26. Screen Testing vs Confirmatory Testing Screen Tests are designed to be highly sensitive goal is to not miss any positives however false reactive results can occur even when the donor was never exposed to the particular infection Confirmatory Testing is highly specific This is used for : Donor counselling Reporting to public health Initiating Lookback Donors are deferred based on screening test results. 26

27. Confirmatory TestingHIV-1 Western Blot Individual proteins of HIV-1 lysate separated according to size by polyacrylamide gel electrophoresis. The viral proteins are then transferred onto nitrocellulose paper and reacted with the donor’s sample. The results are: NEGATIVE (no bands present), INDETERMINATE (any bands present but pattern does not meet criteria of positive) POSITIVE (must have two or more of bands at p24, gp41 and gp120/160) based on the pattern which is present. Western Blot also used for HIV-2, HTLVI/II 27

28. HIV-1 Western Blot 28

29. Confirmatory and Supplemental TestingHepatitis B HBsAg Neutralization confirms the presence of HBsAg by means of specific antibody neutralization. Anti-HBs EIA for the qualitative and quantitative detection of antibodies to the Hepatitis B surface antigen. HBV DNA qualitative test for the direct detection of HBV using PCR methodology 29

30. Confirmatory Testing HCV Qualitative immunoblot assay - RIBA. Utilizes recombinant HCV encoded antigens and synthetic HCV encoded peptides that are immobilized as individual bands onto test strips. The possible serological profiles defined by this assay include the following: Negative, Positive, Indeterminate 30

31. RIBA 31

32. Confirmatory TestingSyphilis Repeat reactive samples are referred to the Public Health Laboratory (Alberta or Ontario) for confirmatory testing. EIA screening test, followed by: Flourescent Treponemal Antibody-Absorbed (FTA- ABS) Western Blot MicroHAemagglutination-Treponema Pallidum (MHA- TP) 32

34. Anti-HIV-1/2 Confirmatory Algorithm 34

35. Anti-HCV Confirmatory Algorithm 35

36. Anti-HTLV-I/II Confirmatory Algorithm 36

37. HBsAg Confirmatory Algorithm 37

38. Anti-HBc Algorithm 38

40. Future Alternate Algorithms Use of HCV and HIV-1 NAT in confirmatory algorithm: Already being performed as a screening test for all donations NAT results integrated into donor counselling Sensitivity and specificity is high relative to confirmatory assays used even though NAT is performed in pools For NAT positive donor samples, HCV RIBA or HIV-1 WB is not needed. Revised screening strategy for anti-HBc: Algorithm has been changed to a one strike algorithm the same as for other TD markers (2005 - Jan. 2010 aHBc positive donors were allowed to return to donate as long as not aHBs or HBV DNA positive) 40

41. West Nile Virus Transmission Cycle Mosquito vector Incidental infections West Nile virus West Nile virus Incidental infections Bird reservoir hosts

42. West Nile Infection - Clinical • Incubation 3 - 15 days • Asymptomatic or mild febrile illness + rash • Elderly often more severely ill with encephalitis: • Headache, stiff neck, nausea, vomiting • Altered level of consciousness, profound muscle weakness • CSF shows pleocytosis, elevated protein, normal glucose

43. West Nile VirusTransmission by organ transplantation and blood transfusion 1st reported case of WNV transmission in U.S. by organ donation August 2002 – 2 kidney, 1heart, 1 liver recipients infected 23 patients confirmed to have acquired WNV infection via RBCs, platelets, FFP in 2002 5 cases of reported WNV in Canada (2002) had received blood transfusion within 28 days: Total 4 probable cases of TT-WNV 43

44. Strategy for Single Unit Testing2005 - 2009 CBS began screening for WNV in blood donors using a Roche WNV NAT assay in 2003 by mini-pool (6) Single unit testing was initiated in 2004 to enhance sensitivity in areas of WNV activity: SUT initiated in a health region when one WNV positive donor is identified, or The number of new confirmed community cases reported in a health region reaches the level of 1/1,000 (rural areas) or 1/2,500 (urban) for the past 2 consecutive weeks. SUT discontinued if no more positive donors or # of community cases fall below population trigger 2005 - SUT discontinued after 14 days 2006 – 2009 SUT discontinued after 7 days 44

45. WNV NAT Screen Reactive Donors by Province(to Jun. 1 2007- Oct. 15, 2009) 1 1 false positive donor (alt. NAT and antibody negative) 22 donors with travel history 31 positive donor in 2008 - Winnipeg 45

47. Chagas Disease – What is it? Infection caused by a protozoan parasite, spread by triatomine bugs endemic to Central & South America, Mexico Estimated that 16-18 million people are infected ~50,000 people die annually from Chagas Also spread by blood transfusion, organ transplants, rarely mother-child (transplacental), 47

48. From CDC 48

49. Reduvid Bug – aka ‘Kissing Bug’ Where they like to hang out. If your Holiday Inn looks like this – move to another hotel! WHO/TDR 49

50. Where is Chagas Disease Found? Primarily found in Latin America Increased infections are being detected in the United States 50