Kein Folientitel

1
Correlation of HBsAg and HBV DNA Michael Chudy,
Paul-Ehrlich-Institut SoGAT XVIII, Bethesda
MD 24-25 May 2005
2
HBsAg and HBV DNA

• Most sensitive HBsAg assay detects0.01 ng/ml
  (PEI HBsAg standard ad)
• HBsAg cut-off correspond to HBV DNA
• 114 IU/ml (WHO DNA standard)
• 461 copies/ml (five SC panels ZeptoMetrix)

Good correlation to published data from Biswas et
al. Transfusion 43788-798 (2003)
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Virus and HBsAg Particles
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HBsAg reactive polypeptides

• Most sensitive ELISA detects 10 pg HBsAg/ml
• Virion-HBsAg
• One HBV particle 20 ag HBsAg
• 10 pg HBsAg correspond to about 500,000 HBV
  particles
• 500 particles/ml (HBsAg cutoff of SC panels) vs.
  500,000 HBV particles/ml
• Viral surface antigen reactive polypeptides
• Virion envelope
• Incomplete viral forms
• Excessive incomplete viral forms are present
  (about 11,000)
• Average numbers of molecules of LHBs,
  MHBs, and SHBs 30, 30, and 350, respectively
  Calculation correlate well with data presented by
  Prof. Gerlich (EPFA meeting 2002)

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Correlation of HBsAg and HBV DNA

• Is that true for all geno-/subtypes, stages of
  HBV infection?
• Can HBV NAT replace the HBsAg testing in blood
  donors?

6
HBsAg and HBV DNA

• HBV DNA reverse transcription from 3.35 kb
  pre-genomic mRNA
• HBsAg forms translated from
• LHBs 2.4 kb mRNA
• MHBs/SHBs 2.1 kb mRNA
• Level of regulation
• Transcription
• Post-transcription
• e.g. Transport of unspliced mRNAs from nucleus to
  cytoplasm
• Cellular factors

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Ratio of total HBsAg to Virion-HBsAgat different
HBV stages

• Early period of HBV infection
• Genotype A
• Genotype G
• Chronic HBV infection

8
Total HBsAg/Virion-HBsAg ratio in samples of SC
panel PHM911 (BBI)
21 d
data performed in 1995
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HBV genotype G

• Only few infections are known (all co-infections
  with genotype A)
• First report of HBV infection and transmission
  based exclusively on genotype G (2003 in Germany)
• No escape variant (subtype adw2)
• HBeg minus variant

Studies were performed in cooperation with the
Institute of Transfusion Medicine and
Immunohematology, GRC, JW Goethe University,
Frankfurt (WK Roth and co-workers)
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HBV transmission exclusively by genotype G-virus
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Virion-HBsAg/total HBsAg ratio in genotype G

• Sample from R3 (2003-09-23) with about 9 Mio
  cps/ml (positive for HBsAg, negative for
  anti-HBc, HBeAg/anti-HBe)
• Titration in Prism HBsAg test
• Virus concentration at HBsAg cut-off24,000
  copies/ml
• Ratio of virion-HBsAg to total HBsAg of 120
• vs. 11,000 and more in genotype A samples of
  early HBV infection
• Significant decrease of excessive HBsAg in
  infection with genotype G

12
HBsAg and HBV DNA in chronic HBV infection

• Lack of correlation between HBsAg and HBV DNA in
  HBsAg/anti-HBc positive tested samples (Kuhns et
  al. 2004, Transfusion 44,1332-1339)

13
HBsAg and HBV DNA in chronic HBV carriers

• Investigation of 106 chronic carriers tested
  positive for HBsAg, anti-HBc, and anti-HBe
• Only 84 (89/106) HBV NAT reactive with the
  Procleix Ultrio Assay (Gen-Probe)
• s/co gt20 5x
• s/co 15-20 65x
• s/co 10-15 9x
• s/co 5-10 7x
• s/co lt5 3x
• No correlation to s/co values of HBsAg test
  (Prism)

14
17 HBsAg positive/HBV DNA negative samples of
chronic HBV carriers
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Summary Correlation ofHBsAg and HBV DNA

• Virion-HBsAg of one particle (20 ag) correspond
  to one copy of HBV DNA
• Significant excess of HBsAg particles in the
  early period of HBV infection (genotype A)
• Variable ratio of Virion-HBsAg to total HBsAg in
  the course of infection (genotype A)
• Genotype G infection shows a significant decrease
  in synthesis of incomplete viral forms
• Lack of correlation between HBsAg and HBV DNA in
  chronic infection
• HBsAg carrier (integrative phase) can be
  non-reactive by HBV NAT
• Results indicate caution in any consideration of
  dropping HBsAg screening