1. Update on West Nile virus and Blood Safety November 3, 2005 Hira Nakhasi, Ph.D. Director, DETTD/OBRR CBER, FDA

2. Background Information WNV is an enveloped single stranded RNA virus WNV is a mosquito-borne Flavivirus – Primarily infects birds –Occasionally infects humans and other animals About 80% of human infection is asymptomatic, and 20% develop mild febrile illness (flu-like illness) Approximately 1 in 150 infections results in meningitis or encephalitis –Advanced age is by far the most significant risk factor for severe neurologic disease Viremic period can occur up to 2 weeks prior to symptoms and last up to more than a month from the initiation of the infection

3. Background Information……. The 2002 US outbreak of WNV resulted in the identification of other modes of transmission including: – blood transmission (RBCs, plasma and platelets), transplantation, breast-feeding, transplacental and occupational by percutaneous injury The magnitude of the risk of WNV from transfusion is unknown. Virus titer in blood is low compared to other transmissible viruses. –Viremia in encephalitis patients can be as high IgM can persist for a long time in some cases up to 2 years No chronic stage of WNV infection has been reported

4. West Nile virus in the Americas United States 1999 Canada 2000 Mexico 2002 Dominican republic 2002 El Salvador 2003 Jamaica 2003 Cuba 2005

5. WNV Human Cases 1999-2005 Total Human Cases 19287 Neuroinvasive Diseases 8151 Deaths 748

6. 2005 West Nile Virus Activity in the United States Reported to CDC as of November 01, 2005 Total Human Cases 2581 Neuroinvasive Diseases 1053 Deaths 83

7. November 01 2005 WNV Viremic Blood Donor Activity in the U. S. Reported to CDC as of November 01, 2005 West Nile Virus Infections in Organ Transplant Recipients: Three cases were identified in New York and Pennsylvania, August - September, 2005 2002-2005: 34 cases of transmission by blood transfusion and transplantation

8. West Nile Virus and Blood Safety: Nationwide testing of WNV under FDA approved INDs since July 1, 2003 using GenProbe and Roche NAT Tests resulted in : –Donations interdicted from asymptomatic donors with confirmed or suspected WNV infections –In 2003, 818 WNV presumptive viremic blood donors officially reported to CDC’s ArboNet 6 confirmed T-T cases (4/6 had low viremia ~0.11 pfu/ml) –In 2004 a total of 224 WNV presumptive viremic donors officially reported to CDC’s ArboNet using MP-NAT as well as ID-NAT in select areas starting May ’04 one reported case of T-T (detectable only by ID-NAT) –As of November 1, 2005; 374 presumptive viremic donors officially reported to CDC’s ArboNet using MP-NAT as well as ID-NAT in select areas. No reported T-T cases West Nile Virus Infections in Organ Transplant Recipients: Three cases were identified

9. WNV Blood Safety Measures FDA’s current recommendations for donor deferral –FDA most recent recommendations targeting prevention of transmission by donors with current or recent symptoms was issued on June 23, 2005, which provides revision to May 1, 2003 guidance. –During the epidemic season of May 1 to November 30: Defer donors suspected of having WNV infection or diagnosis with WNV infection for 120 days after diagnosis or onset of illness, which ever is later. Donors be deferred on the basis of a reactive investigational screening test for WNV. Blood establishments at their own discretion, may enter such donors after 120 days from the reactive donation. Although no additional testing of the donor during 120 day deferral period is recommended, IDNAT for WNV on F/U sample obtained during the 120 day deferral period will provide useful additional scientific info on the duration of WNV viremia. If such a F/U sample is reactive for WNV, FDA recommends that the donor be deferred for an additional 120 days from the date the sample was collected.

10. West Nile Virus and Blood Safety: FDA’s Actions to Date FDA is continuing to work closely with test kit manufacturers to expedite test licensure FDA is continuing to participate in biweekly meetings with the task force established by blood banking community, which includes CDC and NIH to coordinate the epidemiological data on WNV infection and to monitor the out come of testing.

11. West Nile Virus and Blood Safety: Gaps in knowledge Genetic variations in WNV strains – limited data in human disease cases –Detection by currently available WNV assays Determination of residual risk of WNV infection in the presence of antibody –MP-NAT low titer, MP-NAT (-), ID-NAT (+) units Usefulness of WNV surveillance data to predict epidemic –Detection in birds, mosquitoes, equines, symptoms in human, etc. –Severity of epidemic from year to year

12. Studies on genetic evolution of WNV in the US epidemics Rios et. al. Aliquots of human plasmas that had been identified by WNV NAT screening were used for viral isolation: 33 isolates were obtained Structural genes have been sequenced in 25 isolates: 7 from 2002 9 from 2003 7 from 2004 2 from 2005 Five isolates have been completely sequenced: 2 from 2002 1 from 2003 2 from 2005

13. Evolution of WNV in the United States GENETIC SHIFT 2001-2002 New genotype emerged in the USA in 2001: 55% of all isolates in 2002, 85% of all isolates in 2003 Displacement of NY99 genotype by new dominant genotype may be due to differences in mosquito transmission efficiency (Ebel et al. 2004. AJTMH) –Significantly higher proportion of mosquitoes became infected (infectious virus in bodies) and developed disseminated infections (infectious virus in legs) following feeding with dominant genotype compared to NY99 –And higher proportion of mosquitoes were able to transmit (infectious virus in salivary secretions) at days 5 and 7 post-feed with dominant genotype GENETIC DRIFT 2002-2005 Majority of nucleotide changes are silent transitions (U <> C, A<>G). The small numbers of mutations in structural genes may suggest the absence of a strong immune selective pressure and shows limited evolution of West Nile virus during 2002-2005 epidemics.

14. In vitro infectivity of WNV NAT positive plasmas with or without antibodies Cultures (positive / tested) Antibody NegativeVeroMacrophageCombined MP+ ID+27/309/927/30 MP+/- ID+0/22/2 29 of 32 RNA+ Ab- samples infected cells in culture Cultures (positive / tested) Antibody PositiveVeroMacrophageCombined MP+ ID+4/94/46/9 MP- ID+2/72/44/7 10 of 16 RNA+ Ab+ samples infected cells in culture

15. Conclusion 10/16 WNV positive plasmas containing IgG and/or IgM were infectious for Vero cells and/or human MDM in vitro. Although, in vitro infectivity does not imply infectivity in vivo, it demonstrates the presence of live virus and therefore raises concern about a potential risk for transfusion transmission. The potential transfusion risk from low titer/antibody positive donations needs further study either through recipient lookback or an inoculation study in non-human primates.

16. Acknowledgments CDC AABB Task Force ABC ARC BSL Roche GenProbe DOD FDA