1. Evidence of international transmission of HCV in pan-European study of HIV- positive men who have sex with men (MSM). M Danta 1,9 *, T van de Laar 4 *, D Brown 1, O Pybus 3, S Bhagani 2, M Vogel 5, S Neifer 6, A Baumgarten 6, H Gőtz 7, J Rockstroh 5, S Bruisten 4, G Dusheiko 1, R Coutinho 4,8 1 UCL Institute of Hepatology, London, UK; 2 Department of HIV Medicine, Royal Free and University College, London, UK; 3 Department of Zoology, University of Oxford, UK; 4 Cluster of Infectious Diseases, Health Service, Amsterdam, Netherlands; 5 University of Bonn, Germany; 6 Practice Dupke / Carganico / Baumgarten, Berlin, Germany; 7 Department of Infectious Diseases, Health Service Rotterdam, Netherlands; 8 Centre for Infectious Disease Control, National Institute of Public Health and the Environment, Bilthoven, Netherlands; 9 St Vincent’s Clinical School, UNSW, Australia. *Joint first author.

2. Acute HCV in HIV-positive men in Europe N= 389

3. UK temporal data of acute HCV in HIV- positive MSM Giraudon STI 2007 (in press) Overall incidence: 9.1/1000 patient years

4. UK temporal data of acute HCV in HIV- positive MSM Giraudon STI 2007 (in press)

5. Transmission risk factors Identified factors: 1.High number of sexual partners. 2.Mucosally traumatic sexual practices. 3.Group sex practices. 4.Ulcerative sexually transmitted infections (STIs). 5.Recreational drug use and sharing (intranasal and intrarectal). Permucosal NOT parenteral transmission. Danta AIDS 2007, Van de Laar JID 2007, Serpaggi AIDS 2006, Gotz AIDS 2005, Brown STI 2004

6. Aim To determine if there is evidence of a HCV transmission network among European MSM using molecular phylogenetic analysis.

7. Study population HIV-positive men who have sex with men (MSM) diagnosed with recent HCV since 2000. Cases from European cohorts: 1.UK: n=107 Danta AIDS 2007 2.Netherlands: n=51 Van de Laar JID 2007 3.France: n=8 Serpaggi AIDS 2006 4.Germany: n=24 Vogel AASLD 2005 Recent hepatitis C defined as: –documented anti-HCV seroconversion within prior 12 months, or –a positive HCV-RNA by polymerase chain reaction assay (PCR) following a negative assay within prior 12 months previously, or –a positive HCV-RNA PCR and a serum alanine aminotransferase (ALT) level >10-times the upper limit of normal (ULN) with documented normal ALT during the preceding year.

8. Baseline characteristics of cohort NumberAge (years) MSMCD4 (µ/ml) HAART (%) UK11136all55067 Danta AIDS 2007 Dutch3440allna Van de Laar JID 2007 France1240all62575 Serpaggi AIDS 2006 Germany3138all447na Vogel AASLD 2005

9. Phylogenetic methods E1/E2 sequence Phylogenetic comparison to other sequences (cases and Genbank) Construction of neighbour- joining trees (PAUP* software) Reverse Transcription PCR HCV genome c E1E2 p7 2345a5b 5’ UTR3’ UTR

10. Phylogenetic trees (>70) A B C GENETIC DISTANCE Bootstrap value

11. Genotyping 1a1b23a4dNT EnglishN = 107536491520 DutchN = 51263-3181 GermanN = 24171--42 FrenchN = 8---161 Total (%) N =19096 (50) 10 (5) 4 (2) 13 (7) 43 (23) 24 (13) 166 sequences in phylogenetic analysis

12. Genotype 1a (1) HCV phylogenetic tree Cluster 1 n=3 (94) Cluster 2 n=11 (84) Cluster 3 n=16 (90) Cluster 4 n=4 (95) UK Netherlands French German E3_64

13. Genotype 1a (2) HCV phylogenetic tree Cluster 5 n=17 (84) Cluster 6 n=37 (72) UK Netherlands French German

14. Genotype 1a - cluster 5 84 86 85 E27 E32 E62 E63 E67 E72 E87 G13 E34 E33 E42 E18 G16 G21 G22 G3 G7 UK German

15. Genotype 3a HCV phylogenetic tree UK Netherlands French German Cluster 2 n=6 (90) Cluster 1 n=3 (53)

16. Genotype 4 HCV phylogenetic tree Cluster 1 n=31 (93) Cluster 2 n=12 (83) UK Netherlands French German

17. Comparison of individual sequences 90% (149/166) of MSM have HCV genotypes 1 or 4 (Difficult treatment genotype SVR < 40%). 88% (147/166) of MSM have HCV strains similar to ≥ 1 MSM. 73% (122/166) of MSM have HCV closely related strains to ≥ 1 MSM of another European country.

18. Comparison of cluster size by country mixing No country mixing 2 countries>2 countries Cluster (genotype) 3 (G1a)4 (G1a)11 (G1a) 3 (G3)6 (G3a)16 (G1a) 12 (G4)17 (1a)31 (G4) 37 (G1a) Median3623.5 Kruskal-Wallis p=0.11

19. Conclusion 1.Molecular analysis reveals large HCV transmission network among HIV-positive MSM in Europe. 2.Travel important factor in HCV transmission in this population. 3.The molecular HCV clusters, which cross subtype and genotype, suggests this epidemic is not due to a hepatitis C viral change, but rather behavioural and/or environmental factors. 4.Suggestion that increased country mixing with larger cluster size consistent with regional spread.

20. Conclusion National public health agencies need targeted preventive strategies including screening of high-risk HIV-positive MSM to mitigate spread of HCV in this population.

21. Acknowledgements UK: HIV and Acute HCV (HAAC) group: M Nelson, Y Gileece, M Aitkins, D Asboe, R Browne Department of HIV Medicine, Chelsea and Westminster Hospitals S Bhagani, M Johnson, G Mulhoney, T Fernandez, Department of HIV Medicine AM Geretti, Department of Virology D Brown, G Dusheiko, M Jacobs, Centre for Hepatology C Sabin, Department of Primary Care and Population Sciences Royal Free and Hampstead Trust/Royal Free and University College Medical School M Fisher, N Perry, A Phillips Department of HIV Medicine, Brighton and Sussex University Hospitals Trust O Pybus Department of Zoology, Oxford University Netherlands: Cluster of Infectious Diseases, Health Service, Amsterdam Centre for Infectious Disease Control, National Institute of Public Health and the Environment, Bilthoven Department of Infectious Diseases, Health Service Rotterdam, Netherlands; Germany: University of Bonn Practice Dupke / Carganico / Baumgarten, Berlin France: Institut de Veille Sanitaire, Paris Hospital Necker, Paris