1161

viability could

result in isolated cases that are not thought to be attributable to safe products because they are so few. They are almost never reported in medical publications because all manufacturers and most clinicians would prefer to attribute them to non-therapeutic sources. Any remote epidemiological circumstance concerning the case is hierarchically placed ahead of exposure to a plasma derivative whenever a usually effective method of virus inactivation has been applied. Cash’s suggestion about HCV molecular epidemiology is interesting. The rate of mutation in chronic HCV infection seems to be very slow,4 and self reinfection with the same strain of HCV after orthotopic liver transplantation has been recorded.s Even with many plasma contributors to pooled derivatives, US HCV strains are sufficiently different to be demonstrable in Japanese haemophiliacs treated with American concentrates.6 Within more geographically limited areas, however, pooled plasma derivatives contaminated by several donors may have strains that are fairly similar but diverse enough to make judgment difficult ? Department of Medicine, University of Southern California, Los Angeles, California 90032, USA

JAMES W.

MOSLEY

1. Kreutz W, Auerswald G, Brückmann C, et al. Prevention of hepatitis C virus infection in children with hemophilia A and B and von Willebrand’s disease. Thromb Haemost 1992; 67: 184.

2. Gellis SS, Neefe JR, Stokes J Jr, et al. Chemical, clinical and immunological studies on the products of human plasma fractionation XXXVI, inactivation of the virus of homologous serum hepatitis in solutions of normal human serum albumin by means of heat. J Clin Invest 1948; 27: 239-44. 3. Brackmann H-H, Egli H. Acute hepatitis B infection after treatment with heat-inactivated factor VIII concentrate. Lancet 1988; ii: 967. 4. Ogata N, Alter HJ, Miller RH, Purcell RH. Nucleotide sequence and mutation rate of the H strain of hepatitis C virus. Proc Natl Acad Sci USA 1991; 88: 3392-96. 5. Feray C, Samuel D, Thiers V, et al. Reinfection of liver graft by hepatitis C virus after liver transplantation. J Clin Invest 1992; 89: 1361-65. 6. Hijikata M, Kato N, Mori S, et al. Frequent detection of hepatitis C virus US strain in Japanese hemophiliacs. Jpn J Cancer Res 1990; 81: 1195-97. 7. Li JS, Tong SP, Vitvitski L, Lepot D, Trepo C. Evidence of two major genotypes of hepatitis C virus in France and close relatedness of the predominant one with the prototype virus. J Hepatol 1991; 13 (suppl): S33-37.

Hepatitis C viraemia with normal liver histology in symptomless HIV-1 infection SIR,-Dr Alberti and colleagues (Sept 19,

p

697) suggest that

hepatitis C virus (HCV)-RNA is a marker of liver disease in subjects with antibodies to HCV, independent of alanine aminotransferase (ALT) values, and challenge the idea of the existence of true healthy carriers of HCV. We disagree. We investigated over 6 months 19 subjects (15 were male), aged 19-37 years (mean 27), with anti-HIV-1 antibodies in serum (detected by enzyme immunoassay [EIA], Abbott, and confirmed by western blot, Du Pont de Nemours), no symptoms or signs of liver disease, and persistently normal serum ALT values. 11 subjects had been intravenous drug users, 4 were homosexual men, and 4 were female sexual partners of HIV-1-seropositive individuals. None of these 19 subjects had HIV-1-p24 antigenaemia (EIA). 12 subjects were CDC stage II (symptom-free) of HIV infection; the remaining 7 were CDC III (lymphadenopathy syndrome). Mean CD4-positive lymphocyte count was 392/1!1 (range 128-723); 11 subjects with fewer than 500 CD4-positive cells/ul had taken zidovudine (500 mg daily) for 6-17 months. With the use of a recombinant immunoblotting assay (RIBA-2, Ortho Diagnostics Systems), anti-C22 and anti-C33c reactivity was shown in all 19 subjects, with co-existing anti-ClOO positivity in 8. Serum HCV-RNA (detected by polymerase chain reaction with primers derived from the well-conserved 5’-non-coding region of HCV and nested amplification)’ was positive in all 19 subjects. Percutaneous liver biopsy (after informed consent) was done on the same day as serum samples were obtained, and specimens were tested for HCV-RNA. Liver histology was normal in 8 cases (with no evidence of inflammation, fibrosis, or hepatocyte necrosis), and showed slight changes in 2, chronic persistent hepatitis in 5, and chronic active hepatitis in 4 (with associated cirrhosis in 2 cases). Sex, age, risk group, CDC stage, number of CD4-positive serum

lymphocytes, or zidovudine histological fmdings.

treatment were not

related

to

the

These results show that active HCV infection is not invariably associated with liver disease and suggest that some HIV-1-infected individuals can be healthy carriers of HCV. It remains to be determined whether HIV co-infection affects the rate of HCV replication (as suggested by results of one small study) and why zidovudine treatment, which is associated with remission of HCV-induced chronic active hepatitis in patients with high ALT valueshas no such effect in subjects with normal ALT.

Infectious Diseases Unit, "A Pugliese" Hospital, 88060 Catanzaro, Italy, Department of Infectious Diseases, University of Verona, and 1st Department of Infectious Diseases, University of Genoa

SANDRO VENTO MARIO CRUCIANI GIOVANNI DI PERRI TIZIANA GAROFANO ERCOLE CONCIA TERESA FERRARO DANTE BASSETTI

1. Okamoto

H, Okada S, Sugiyama Y, et al The 5’-terminal sequence of the hepatitis C virus genome. Jpn J Exp Med 1990; 60: 167-77. 2. Watson HG, Zhang LQ, Simmonds P, Ludlam CA. Hepatitis C virus load increases with time after HIV infection. VIII International Conference on AIDS, Amsterdam, July 19-24, 1992, abstr PoB 3629, p B195. 3. Vento S, Garofano T, Di Perri G, Cruciani M, Concia E, Bassetti D. Zidovudine therapy associated with remission of chronic active hepatitis C in HIV-1 carriers. AIDS 1991; 5: 776.

Antibody-capture particle-adherence test for antibody to HIV-1 in urine SIR,-We have used the IgG antibody-capture particletest (GACPAT)l for antibody to HIV-1 on urine samples from intravenous drug users (IVDUs) attending methadone clinics in Hong Kong. Of 1259 samples, 66 (5 -2%) were initially and 37 (2-9%) were repeatedly reactive. Of the 37 repeatedly reactive samples, only 7 remained reactive after being heated in a water bath at 60°C for 20 min. As controls, concurrently heat-treated urine samples from confirmed HIV-positive individuals did not show any alteration in endpoint titres. The 7 repeatedly reactive samples were negative by antibody-capture adherence

enzyme immunosorbent assay and western blot.

Our observation suggests that although GACPAT is economical and simple for use in large scale anti-HIV screening,2 it can give a high number of false-positive reactions in urine samples from IVDUs. Heat treatment of repeatedly reactive samples removed non-specific reactions in most samples. The nature of the heat-labile substance in the urine of IVDUs that causes the non-specific reaction remains to be elucidated. Virus Unit,

Department of Health, Queen Mary Hospital, Pokfulam Road, Hong Kong

W. L. LIM K. C. CHENG

1. Connell

JA, Parry JV, Mortimer PP, et al. Preliminary report: accurate assays for anti-HIV in urine. Lancet 1990; 335: 1366-69. 2. Parry JV, Mortimer PP. An immunoglobulin G antibody capture particle adherence test (GACPAT) for antibody to HIV-1 and HTLV-1 that allows economical large scale screening. AIDS 1989; 3: 173-76.

Serological assessment of Helicobacter pylori eradication SIR,-Various tests are available to assess the success of therapy at eradicating Helicobacter pylori. Upper gastrointestinal endoscopy with antral biopsy is too invasive for routine use; 13C and 14C urea breath-testing are expensive and not readily available to most general practitioners; and a lack of studies on the reliability of serological examination in assessing past from current infection combined with the extravagant number of testing kits have clouded the interpretation of serological assays. However, the diagnostic value of decreasing H pylori antibody titres after eradication of the bacterium has been demonstrated by Kosunen et aP using an acid-glycine extract enzyme-linked immunosorbent assay (ELISA).1 Others2 had different opinions about the value of such serological testing. We can add data from H pylori serological testing in Busselton, a Western Australian rural community (population 9000), to support the findings of Kosunen et al.

aimed

1162

rather than six samples as was used for measuring the more stable second phase. MODY patients have a pattern of a global reduction of insulin responses which is similar to that in mildly hyperglycaemic patients with classic non-insulin-dependent diabetes (NIDDM).4 Previous reports of a specific defect of first-phase insulin secretion in NIDDM is partly spurious because of the use of an oral glucose tolerance test or standard glucose infusion test, in which an impaired first-phase response leads to

Mean H pylori antibody titre after antibiotic and bismuth subcitrate therapy.

With an acid-glycine extract ELISA to detect Hpylori antibody, we have found3 that H pylori antibody in a randomly selected adult community cohort over 21 years was lost only in a small minority of H pylori seropositive subjects. Almost all subjects who lost their seropositivity had taken antibiotics and/or bismuth subcitrate, which might account for the eradication of H pylori.4 The same H pylori serological test has also been used in Busselton to assess the efficacy of therapy aimed at H pylori eradication. Between 1986 and 1991, 425 patients (aged 10-91) who attended general practitioners in Busselton because of dyspepsia and/or peptic ulcer were tested. Of the 214 men (mean age 50-6 years), 95 (44.4%) were seropositive, and of the 211 women (45-6 years), 74 (35-1%) were seropositive. Tests were done on serum from 50 patients before and after attempts at eradication with bismuth subcitrate and antibiotics (mostly amoxycillin plus metronidazole). Of these, 45 (90%) had at least a 50% fall in ELISA titre. Of the 33 subjects followed up for 12 months or longer, 28 (85%) became serologically negative or equivocal. In this group the average time taken for antibody titres to fall by 50% after treatment was 3-4 months (figure). In Kosunen’s study! a 50% or more fall in titre 6 months after treatment for H pylori reliably predicted successful eradication of the organism. Our data provide additional support for the use of H pylori serology to evaluate the effectiveness of therapy for bacterial eradication.

Royal Perth Hospital, Busselton Population Study Group, Perth 6001, Western Australia

D. J. E. CULLEN K. J. CULLEN B. J. COLLINS K. J. CHRISTIANSEN J. EPIS

1. Kosunen TU,

Seppala K, Sarna S, Sipponen P. Diagnostic value of decreasing IgG, IgA, and IgM antibody titres after eradication of Helicobacter pylori. Lancet 1992;

339: 893-95. Zwet AA, Meyer BC, Berrelkamp RJP, et al. Serological tests to monitor treatment of Helicobacter pylori. Lancet 1992; 340: 51. 3. Goodwin CS, Blincow E, Peterson G, et al. Enzyme-linked immunosorbent assay for Campylobacter pyloridis: correlation with presence of Campylobacter pyloridis in the gastric mucosa. J Infect Dis 1987; 155: 488-94. 4. Cullen DJE, Collins BJ, Cullen KJ, et al. Royal Perth Hospital and the Busselton Population Studies Group, Western Australia. A 21 year follow up of Helicobacter pylori infection: the ’cohort theory’ proved,. Proceedings of the Annual Scientific Meeting of the Gastroenterological Society of Australia, April, 1992: 69 (abstr). 2.

van

Beta-cell secretory defect caused by mutations in glucokinase gene SIR,-Dr Velho and colleagues (Aug 22, p 444) report that patients with maturity-onset diabetes of the young (MODY) with mutations in the glucokinase gene have deficient beta-cell secretion that affects mainly the second phase of insulin secretion. We showed in patients with MODYl due to missense mutation arg299gly2 that beta-cell function in response to a glucose infusion test3 was impaired, with 57% normal function compared with 123% normal function in their unaffected, weight-matched relatives, and that both phases of insulin secretion were affected. Inspection of the insulin values that Velho et al show in fig 1 suggests an impaired first phase; the lack of statistical significance might have been partly due to the use of arithmetic rather than log-transformed data, and in part to assessment of the transitory first phase with two plasma samples

secondary hyperglycaemia and a less-impaired second-phase response.5,6 In both NIDDM and in MODY, so-called normal fasting plasma insulin and C-peptide values associated with raised fasting plasma glucose concentrations are compatible with an impaired beta-cell secretory response, and it is doubtful whether different responses to glucose will allow differentiation between NIDDM and MODY. Diabetes Research Laboratories, Radcliffe Infirmary, Oxford OX2 6HE, UK 1. 2.

RENÉE PAGE ANDREW HATTERSLEY ROBERT TURNER

Hattersley AT, Turner RC, Permutt MA, et al. Linkage of type 2 diabetes to the glucokinase gene. Lancet 1992; 339: 1307-10. Stoffel M, Froguel PH, Takeda J, et al. Human glucokinase gene. isolation, characterisation, and identification of two missense mutations linked to early-onset non-insulin dependent (type 2) diabetes mellitus. Proc Natl Acad Sci 1992; 89:

7698-702. 3. Hosker JP, Matthews DR, Rudenski AS, et al. Continuous infusion of glucose with model assessment (CIGMA): measurement of insulin resistance and beta cell function m man. Diabetologia 1985; 28: 401-11. 4. Hosker JP, Rudenski AS, Burnett MA, Matthews DR, Turner RC. Similar reduction of first- and second-phase B-cell responses at three different glucose levels in type II diabetes and the effect of gliclazide therapy. Metabolism 1989; 8: 767-72 5. Perley MJ, Kipnis DM. Plasma insulin responses to oral and intravenous glucose: studies in normal and diabetic subjects. J Clin Invest 1967; 46: 1954-62. 6. Mitrakou A, Kelley D, Mokan M, et al. Role of reduced suppression of glucose production and diminished early insulin release in impaired glucose tolerance. N Engl J Med 1992; 326: 22-29.

*** This letter has been shown to Dr Velho and colleagues, whose reply follows.-ED. L. SIR,- We reported that the first-phase insulin secretion, assessed by an intravenous glucose tolerance test (IGTT), in 9 MODY patients carrying mutations in the glucokinase (GCK) gene was usually well preserved. Indeed, only 2 of 9 patients had plasma insulin values lower than 1 SD below the mean of the control group. Plasma insulin and C-peptide in patients were lower than in controls but did not differ significantly. Page and colleagues suggest that the lack of statistical significance was in part due to assessment of the transitory first-phase with only two samples and in part to the use of arithmetic rather than log-transformed data. To address this issue we have re-computed the statistics of the IGTT data by repeated measures analysis of variance, taking into account either 2 (1 and 3 min) or 3 samples (1, 3, and 5 min) of insulin and C-peptide. In both situations values of insulin and C-peptide did not significantly differ between patients and controls: Insulin

2-value analysis 3-value analysis

p=0069 p=0055

C-peptide p=0076 p=0059

Similarly, non-significant statistical results were also obtained with log transformation of insulin (p = 0-061) and C-peptide (p = 0-065) values. The more severe defect reported by the Oxford groupl might have resulted from the nature of the mutation present in the kindred they studied, from other genetic or environmental factors (eg, duration of diabetes, glucotoxicity) affecting beta-cell secretory profile, or from differences in the methodology (unprimed glucose infusion instead of IGTT). The second issue Page and colleagues raise is the so-called normal fasting plasma insulin and C-peptide values associated with raised fasting plasma glucose that we observed in MODY patients. They remark that this pattern, frequently reported in NIDDM patients, is compatible with an impaired beta-cell secretory response to glucose. However, there is evidence that in our MODY population this pattern is best explained by an upward shift in the threshold of circulating glucose, which induces insulin secretion. First, the