THE CAUSATIVE AGENTS, DIAGNOSIS AND
MONITORING OF CHRONICALLY INFECTED PATIENTS
Poljak Mario
Institute of Microbiology and Immunology Medical Faculty of Ljubljana, Slovenia
VIRAL HEPATITIS
THE CAUSATIVE AGENTS, DIAGNOSIS AND
MONITORING OF CHRONICALLY INFECTED PATIENTS
PRIMARY hepatitis A virus hepatitis B virus hepatitis C virus hepatitis D virus hepatitis E virus
SECONDARY Epstein-Barr virus cytomegalovirus herpes simplex viruses varicella-zoster virus coxcackieviruses adenoviruses rubella virus measles virus mumps virus hepatitis viruses
A, B, C, D, E A, E
B, C, D
• only acute hepatitis, NO chronic disease
• usually mild clinical course
• life-long immunity? Hepatitis A virus
- nonenveloped - single-stranded RNA virus - icosahedral capsid, 27 nm - family Picornaviridae - genus Hepatovirus - linear positive-strand RNA, 7.5 kb
Hepatitis A diagnosis
Anti-HAV total
Anti-HAV IgM
Hepatitis A Vaccination
• Several inactivated and attenuated vaccines available
• Two injections (0 and 6 - 12 month)
• Antibody titer 10 - 100 fold higher after natural infection compared to vaccination
• Lifelong immunity after vaccination is anticipated but has to be proven Hepatitis E virus
- nonenveloped - single-stranded RNA virus - linear positive-strand RNA, 7.5 kb - family Caliciviridae - “hepatitis E-virus like” genus
Hepatitis E diagnosis
Anti-HEV IgG
Anti-HEV IgM
parenterally transmissible hepatitis viruses
B, C, D
HIV HBV TRANSMISSION TRANSMISSION
HCV TRANSMISSION
1/10,000,000 1/1,000,000 1/100,000 1/10,000 1/1,000 1/100
TUNDERBOLT FATAL MURDER CAR DEATH DUE TO DEATH AIRPLANE DEATH ACCIDENT UNEXPECTED ACCIDENT DEATH DRUG ADVERSE REACTION IN HOSPITAL
Safety of injections
Estimated total burden of infection 8-16 million hepatitis B cases per annum attributable to unsafe 2-4.5 million hepatitis C cases injection practices 75.000-150.000 HIV infections
Estimated annual public health 26 millions of years of life cost d cost generated by unsafe irect med. cost of USD 535 M injection practices
Estimated annual deaths due to 1.3 million deaths unsafe injection practices WHO Bulletin, 2002
Hep C Hep B CIRRHOSIS Hep D 30
20 % 10 FIBROSIS 0 0 5 10 15 20 Years Liver transplantation in Europe (2004)
l 59% hepatitis C
l 28% hepatitis B
l 7% hepatitis D
Prevalence of HBV carriers
Percentage long-term HBV carriers (350 million long-term carriers):
< 2% - Low 2-7% - Intermediate > 8% - High Hepatitis B virus
- partially double-stranded DNA virus that replicates through an RNA intermediate - family Hepadnaviridae, genus Orthohepadnavirus - circular partially double-strand DNA, 3.2 kb
Hepatitis B – “serology” “markers” ü HBsAg ü anti-HBs (quantitative)
ü anti-HBc total antibodies ü anti-HBc IgM
ü HBeAg ü anti-HBe
quantitative measurement of HBV DNA =
HBV DNA viral load r = 0.60, p=0.001 8,8
8,4
Poljak M, Marin IJ, Seme K, Brinovec V, 8,0 Matičič M, Meglič-Volkar J. Second- generation Hybrid capture test and Cobas Amplicor 7,6 Monitor test generate highly correlated hepatitis B virus DNA levels. 7,2 J Virol Methods 2001; 97: 165-9.
6,8 Hybrid-Capture log HBV DNA (copies/ml) Hybrid-Capture
6,4 6,0 6,4 6,8 7,2 7,6 8,0 8,4 8,8 9,2 Marin IJ, Poljak M, Seme K, Meglič-Volkar J, Amplicor log HBV DNA (copies/ml) Matičič M, Lešničar G, Brinovec V. Comparative evaluation of semi-automated Cobas Amplicor Hepatitis B Virus Monitor Test and manual microwell 10 plate-based Amplicor HBV Monitor Test. r = 0.976; p < 0.0001 9 J Clin Microbiol 2001; 39: 758-61.
8
7 Marin IJ, Poljak M, Seme K, Brinovec V, Matičič M, 6 Meglič-Volkar J, Lešničar G, Gradišek P. Comparative evaluation of three commercial assays for 5 quantitative measurement of log HBV DNA (copies/ml) DNA HBV log hepatitis B virus DNA in serum samples.
Hybrid Capture II HBV DNA Test DNA HBV II Capture Hybrid 4 Hepato-Gastroenterol 2002; 49: 1390-1394 3 3 4 5 6 7 8 9 10 11 Cobas Amplicor HBV Monitor log HBV DNA (copies/ml) quantitative measurement of HBV DNA =
HBV DNA viral load
pg/ml copies/ml IU/ml Diagnosis of HBV infection
Acute hepatitis B
HBV DNA detection or quantification is not necessary for the diagnosis of acute hepatitis B, which is based on serological testing Diagnosis of HBV infection
Problematic samples
detection of HBV DNA useful in resolving diagnostic uncertainties following serological testing for markers of HBV infection that are caused by HBV genetic variations
isolated anti-HBc positivity isolated HBsAg positivity simultaneous HBsAg and anti-HBs positivity HBsAg NEGATIVE anti-HBs NEGATIVE anti-HBc total POSITIVE anti-HBc IgM NEGATIVE HBe-Ag NEGATIVE anti-HBe NEGATIVE anti-HBc - alone status anti-HBc - alone status
HBV DNA !!!!! HBsAg POSITIVE anti-HBs NEGATIVE anti-HBc total POSITIVE anti-HBc IgM NEGATIVE
HBe-Ag NEGATIVE anti-HBe POSITIVE HBeAg-negative patient
“inactive HBsAg carrier”
HBeAg-negative chronic hepatitis HBeAg-negative patient
“inactive HBsAg carrier” presence of HBsAg in serum without HBeAg more than 6 months without aminotransferase elevations and with HBV DNA levels less than 105 copies/ml
HBeAg-negative chronic hepatitis presence of HBsAg in serum without HBeAg more than 6 months with elevated aminotransferases and/or with HBV DNA levels more than 105 copies/ml Disease severity L
☺
Viral load
Hepatitis B treatment
FDA approved • interferon α-2b (1992)
• lamivudine (1998)
• adefovir (2002)
• entecavir (2005)
• pegylated interferon α-2a (2005) Treatment of HBV infection
Decision to treat
Selection of optimal therapy
Treatment monitoring
Prevention of Hepatitis B
• HBV infection can be prevented in non-infected individuals by vaccination with HBV vaccine
• series of 3 injections at 0, 1 and 6 months
• vaccination is effective in 90-95% of recipients Recommendations for post-exposure vaccination
• infants born to HBsAg+ mothers or to mothers who had acute hepatitis B during pregnancy
• healthcare workers after needlestick injury involving blood from a HBV infected patient
• sexual partners of individuals with HBV infection
Hepatitis D virus
- defective RNA virus - infections only in HBV infected patients - enveloped single-stranded RNA virus - genus Deltavirus - resembles plant viroids HBV HDV
Hepatitis D diagnostic methods
HbsAg + anti-HDV total antibodies anti-HDV IgM antibodies HDV Ag
HDV RNA
Hepatitis C virus
- enveloped single-stranded RNA virus
- family Flaviviridae, genus Hepacivirus
- linear positive-strand RNA, 9.4 kb
Detection of HCV markers during window phase following infection
EIA 3.0 EIA 2.0 EIA 1.0
0 70 80 150 (DAYS)
Hepatitis C diagnostic pathway
anti-HCV screening test Hepatitis C diagnostic pathway
anti-HCV screening test
anti-HCV confirmatory test
NIH Consensus Statement Management of Hepatitis C June 2002
Hepatology 2002; 36: S3-S20 anti-HCV screening test anti-HCV screening test negative anti-HCV screening test
negative no HCV infection anti-HCV screening test
negative positive no HCV infection anti-HCV screening test
negative positive no HCV infection qualitative NAT
anti-HCV screening test
negative positive no HCV infection qualitative NAT
negative anti-HCV screening test
negative positive no HCV infection qualitative NAT
negative
anti-HCV confirmation or qualit. NAT after 3-6 months ?? anti-HCV screening test
negative positive no HCV infection qualitative NAT
negative positive Envelope Proteins HCV Core Protein (HCV Ag)
HCV RNA
Hepatitis C diagnostic pathway
qualitative NAT positive
70
60
50
40
30
20
10 Sustained response Sustained virologic
0
IFN CS-IFN IFN/RIB PEG-INF PEG-INF/RIB
Therapy Hepatitis C diagnostic pathway
qualitative NAT positive
if therapy considered
end of treatment response (ETR) sustained viral response (SVR) early viral response (EVR) Early viral response (EVR)
• a minimum 2 log decrease in viral load during the first 12 weeks of treatment is predictive of sustained viral response (SVR) in patients with genotype 1
• patients who fail to achieve an EVR at week 12 of treatment have only a small chance of achieving an SVR even if therapy is continued for a full year
• treatment need not be extended in these patients
NIH Consensus Statement, Management of Hepatitis C, June 2002 Virological monitoring algorithm for patients with chronic hepatitis C on treatment
GBV-C Virus / Hepatitis G virus
Prevalence of GBV-C RNA
Blood donors 0 - 2.3 %
Haemophiliacs 8 - 38% Haemodialysis patients 15 - 57% IVDUs 16 - 38% p value, p > 0.01 p value, p > 0.01 p = please be pathogenic GBV-C Virus/Hepatitis G virus
• most infections asymptomatic, transient, self-limiting, with slight or no elevation of ALT levels
• majority of cases resolve after loss of serum RNA with a concomitant appearance of anti-E2 antibodies
• infection with GBV-C doesn’t affect a clinical course in patients with hepatitis A, B, or C GBV-C or Hepatitis G virus
• virus in search of a disease (Pessoa & Wright) • human orphan virus (Theodore & Lemon) ) • accidental tourist (Miyakawa & Mayumi Xiang J, et al. Effect of confection with GB virus C on survival among patients with HIV infection. N Engl J Med 2001; 345: 707-14.
Tillmann HJ et al. Infection with GBV-C and reduced mortality among HIV-infected patients. N Engl J Med 2001; 345: 715-24.
GBV-C as a therapeutic approach for HIV? hepatitis viruses
A, B, C, D, E