496A A A S L D A B S T R A C T S HEPATOLOGY O c t o b e r 1995

1557 MOLECULAR ANALYSIS OF THE HCV 5'UR/CORE REGION IN COMBINATION WITH PHYLOGENETIC ANALYSIS OF THE NSSB REGION PROVIDES EVIDENCE FOR 10 TYPES AND AT LEAST 52 SUBTYPES. L Stuvver 1, A Wyseud, W van Arnheml. F Lunel 2 P Laurent-Puig 3. B Kleter 4. L Bassit s. J Nkengasong 6 ̀L-J van Doorn 7, and G. Maertens ~. qnnogenetics, Ghent, Belgium, 2Groupe Hospitalier Piti&Salp6tri&e, Paris, 3CHU Bic~tre, Kremlin- Bic~tre, France, 4EUR, Rotterdam, the Netherlands, SFunda(;fio Pro Sangue, Sao Paulo, Brazil, 6Institute of Tropical Medicine, Antwerp, Belgium, and 7SSDZ, Delft, The Netherlands.

To test the theoretical possibility of 5'UR mistyping between hepatitis C virus subtypes la and lb, we combined a 5' UR/Core Line Probe Assay (LiPA) with a nested PCR system and retested 183 sera, including non-typeable cases from previous studies, previously genotyped as type la or lb, and which originated mainly from Western Europe. Based on this method, 8% of these were wrongly subtyped and 3 additional subtypes of type 1 were discovered (ld-lf). Randomly selected European type 2 sera (n=18) were tested with an analogous type 2 5'UR/Core LiPA. Surprisingly, most of these coufd be classified as subtype 2c. Among serum samples originating from South-East Asia, several additional genotypes (7a, 7c, 7d, and 9a) were detected. Based on 13,203 pairwise comparisons in the 340-bp NS5B region, classification into types, subtypes, and isolates was obtained in 99.8% of all cases by using the phylogenetic border value of 0.328 for subtypes/types and 0.127 for isolates/subtypes; and evidence for a 10th major type of HCV was provided. Combination of all available HCV sequence data from the 447-bp Core/E1 region and the NS5B 340-bp and 222-bp regions provided evidence for the existence of t 0 types including at least 52 subtypes.

1558 A SECOND GENERATION LINE PROBE ASSAY (LiPA) FOR THE DETECTION OF THE 6 MOST COMMON HEPATITIS C VIRUS GENOTYPES. L Stuyver. A Wyseur. G Verpooten. and G Maertens. Innogenetics, Ghent, Belgium.

We previously reported a reverse hybridization method allowing the identification of genotypes 1, 2, 3 and 4/5. We now developed INNO-LiPA HCV II which allows discrimination of types 4, 5, and 6, in addition to improved classification of type 1 and 2 subtypes.

From the 5' untranslated region (5'UR), 2 type-specific probes were selected for types 11 2 and 5; three type-specific probes for types 3 and 4; and one type-specific probe for genotype 6. Four subtyping probes for type 1 and two new subtype-specific probes for type 2 allowed more specific recognition of subtypes. No subtype- specific probes for type 4, 5, or 6 were applied. Hybridization and stringent wash was carried out in 3x SSC/0.1% SDS for 1 hour at 50°C, followed by the previously described LiPA procedure. The specificity of the typing probes was established with PCR fragments from 5' UR clones of all different genotypes, after which 184 type 1, 28 type 2, 21 type 3, 19 type 4, 8 type 5, and 6 type 6 serum samples were analyzed. Since each of these probes iricluded 2 mismatches compared with all other probes (0ne,type 5 probe contained only 1 mismatch), a very high specificity was detected and confirmed by reactivity to Core probes and Core/E1/NS5B sequencing. A two-log difference in viral load was still detectable in artificially mixed infections.

Sensitivity and specificity of HCV genotyping has considerably improved, especially for type 1 subtyping and for discrimination of types 4, 5, and 6. This assay performs well in Europe, Japan, and the Americas, but type 7, 8, or 9 (present in e.g. Vietnam and Thailand) can not be distinguished from type 1.

1559 Outcome of Prolonged Arabinoside A Therapy of Chronic Active Hepatitis: a 10 year follow-up study. N. Kanaga Sundaram and Willem H. tenHove; Division of Gastroenterologyy, Department of Medicine, UMD-New Jersey Medical School, Newark, NJ 07103

Arabinoside A (AraA) when given IV for 4 weeks causes transient improve- ment in patients with chronic hepatitis B (CI-IB). A study was undertaken to evaluate the efficacy of AraA given for a period of 120 days covering the life span of normal hepatocytes. Patients with biopsy proven C-HB were given 10 mg/kg/d for 10 days followed by 5 mg/kg three times a week for 14 weeks. No new patients were included in the study after the first 18 months because of the publication of high incidence of peripheral neuropathy. Patients symptoms, weight, liver size, pre and post treatment Indocyanin clearance (ICG) and serial aminotransferases were monitored for a period of 10 years.

Of the six CHB patients (1 homosexual, 1 IVDA, 2 post transfusion (one diabetic) and 2 with unknown cause) none had a change in the HBsAg status at the end of the therapy. Three developed pain and heaviness of legs but showed no change in the nerve conduction studies. Two recovered in 2 months and one patient with diabetes had a protracted disability even though ascites, joint effusion, edema, albumin and insulin requirement improved. All patients showed a persistent improvement in the aminotransferases and ICG clearance at the end of the study. In two, deterioration occurred during follow-up (one due to resumption of IVDA and stopped follow up visits, and the other posttransfusion) and showed delta hepatitis and hepatitis C respectively. One patient gave birth to a HBsAg negative full term healthy baby after a normal pregnancy. One died of AIDS. Of the four followed for 10 years all developed core Ab, three developed cAb and serum HBVDNA negative by PCR, and one lost sAg and developed sAb. The diabetic posttransfusion CHB patient never lost the eAg and is positive for sAg and HBVDNA. All four have no evidence of hepatoma.

Prolonged IV AraA therapy induces a sustained clinical improvement in CHB. Three out of four patients show no evidence of HBVDNA. The neuropathy is reversible and no long term side effects are seen. We propose that AraA therapy should be considered in all patients who fail interferon A therapy using HBVDNA by PCR to m0niter its efficacy.

1560 STABILITY OF CEREBRAL PERFUSION PRESSURE AND ICP DURING ORTHOTOPIC LIVER TRANSPLANTATION WITH PRESERVATION OF TIIE IVC AND TEMPORARY PORTACAVAL SHUNT. TS Suttnn~ CI Weiss~ S McClure~ S Shenny~ J Lowell~ TK Howard. Depts. of Anesthesiology and Surgery, Washington University, St. Louis, MO 63110-1093.

Patients with fulminant hepatic failure (FHP) and raised intracranial pressure (ICP) need to have attention paid to surgical techniques that alter ICP and cerebral perfusion pressure (CPP). The surgical technique of orthotopic liver transplantation (OLT) with preservation of portal and caval flows may facilitate stability of CPP during the anhepatic phase (1). This is the first report of the stability of CPP and ICP during OLT using this surgical technique.

f [ ~ : A 30 year old man presented with fulminant hepatic tylenol overdose. Head CT showed cerebral swelling. An

ICP monitor was placed preoperatively. Anesthesia for OLT included sodium thiopental, isoflorane, fentanyl, midazolam, and vecuronium. Exhaled carbon dioxide tension was 25-30 millimeters of mercury.

Table 1. Hemodynamic Parameters and ICP During OLT with Preservation of Porta-Caval Flow.

CVP MAP ICP CPP Baseline 10 83 11 72 PV XC 9 90 11 79

PV XCR 7 91 11 80 NeoheDatic 9 77 10 67

Portal Vein Cross Clamp / Release (PV XC/R); Central Venous Pressure (CVP); Mean Arterial Pressure (MAP). All units are in millimeters of mercury.

a n s i : Cardiovascular hemodynamics, cerebral perfusion pressure, re stable during the anhepatic phase.

Conclusions:l)Porta-caval shunts can be constructed without s i g m ~ r e m e n t in CVP or MAP. 2) CVP and ICP are stable with restoration of portal flow. 3)MAP and CPP are stable during the anhepatic phase. 4)Preservation of the IVC with temporary PC shunt should be considered during OLT'in patients with fulminant hepatic failure and elevated ICP. References:l) Cherqui D, et al. Orthotopic liver transplantation with p r ~ of the caval and portal flow. Transplantation 1994;58:793-6.