(Miles Laboratories Ltd, Stoke Poges) ’TDA Flourostat’ method, a substrate-labelled fluorescent immunoassay introduced for the measurement of plasma theophylline concentration, to measure the theophylline concentration on dried blood spots collected by the patient using an ’Autolet’ (Owen Mumford Ltd, Woodstock, Oxon). We have just completed the validation of this procedure and plan to publish methodological details elsewhere. The accuracy, precision, and speed of this fluorometric technique make it clinically useful. We obtained a significant correlation between capillary (dried blood) and plasma theophylline concentrations (r= 0-985; y=0-78x+0-61; n=43; p<0-001). Sample collection and dispatch to the laboratory by the patient living at home allows a 24 h theophylline profile to be determined, avoids time-consuming visits for repeated venesection, and therapy can be adjusted by telephone as necessary. A theophylline assay service on dried blood spots6 therefore provides a convenient method for confidently tailoring theophylline therapy to individual patient needs. G. F. BATSTONE E. J. COOMBES T. R. GAMLEN
Department of Chemical Pathology, Salisbury General Infirmary, Wiltshire SP2 7SX
DRUG REACTIONS AND THE POOR METABOLISER
SIR,-The past few years have witnessed an increasing number of publications on polymorphism in drug metabolism and the use of test compounds such as debrisoquine to identify subpopulations. The word seems to be being used less and less rigorously- There are, indeed, several good examples of metabolic polymorphism (eg, isoniazid, debrisoquine, and sparteine) where the population at large can be divided into discrete subgroups based on pharmacokinetic indices such as area under the plasma curve (AUC), half-life, or urinary drug/metabolite ratio. There are, however, other drugs whose pharmacokinetic indices show only a skewed distribution in the population without evidence of a discontinuity. Propranolol and metoprolol are good examples,9,10 but the list includes anticonvulsants, neuroleptics, and 1
It is possible, therefore, for a few individuals to display much greater plasma drug concentrations or AUC values than the mode. However, such instances are not polymorphic unless a distinct subpopulation can be clearly identified. Debrisoquine may be a useful tool to detect polymorphism when the principal route of elimination is hydroxylation but if this pathway is a minor one then, even though polymorphism has been demonstrated for that pathway, the overall elimination cannot be described as
polymorphic. Professor Smith and his colleagues (June 18, p 1388) believe that the clinical significance of polymorphic oxidation has been demonstrated, but this is not generally accepted. Polymorphic oxidation may be important for clinically useful drugs with a low therapeutic index when hydroxylation is the major route of elimination, but for most drugs no such claim can be made. Of the drugs which Smith and his colleagues mention the beta-blockers have a wide therapeutic index, debrisoquine is one of the less widely used antihypertensives, and phenacetin has been banned in the UK since 1979. Once-a-day treatment with a fixed dose of drug in all patients is likely to remain an unattainable ideal for some time to come, and the 6. Albani M, Toseland determination of
simple rapid gaschromatographic
Neuropadiatrie 1978; 9:
dried whole blood
method for the filter paper cards.
7. Idle JR, Oates
NS, Shah RR, Smith RL Protecting poor metabolisers, a group at high risk of adverse drug reactions. Lancet 1983, i: 1388. 8. Idle JR, Sever PS Treatment of angina pectoris with nifedipine. Br Med J 1983; 286: 1978-79
Jack DB, Quarterman CP, Zaman R, Kendall MJ. Variability of beta-blocker pharmacokinetics in young volunteers. Eur J Clin Pharmacol 1982, 23: 37-42. 10. Jack DB, Wilkins MR, Quarterman CP. Lack of evidence for polymorphism in metoprolol metabolism. Br J Clin Pharmacol (in press). 11. Koch Weser J Serum drug concentrations in clinical perspective. In: Richens A, Marks V. eds Therapeutic drug monitoring. Edinburgh: Churchill Livingstone, 9
careful physician will always try to adjust the dose to suit the individual patient. We believe that it is unrealistic to call for the setting up of regional centres to offer a phenotyping service. The money for this would come, presumably, from the National Health Service, and there are better ways it could be spent. Finally, to suggest that polymorphic or other variations in the metabolic handling of a drug are sufficient grounds to contramdicate its use or further development seems most unwise. Such criteria would, in the past, have denied us propranolol,
isoniazid, phenytoin, and many tricyclic antidepresants. Department of Thrapeutics and Clinical Pharmacology,
D. B. JACK M. J. KENDALL M. R. WILKINS
University of Birmingham, Birmingham B15 2TJ
AIDS AND AFRICAN SWINE FEVER
SiR,—Dr Teas (April 23, p 923) suggests that a variant of African swine fever virus (ASFV) could be the cause of acquired
immunodeficiency syndrome (AIDS). "Perhaps an infected pig was eaten as uncooked or undercooked meat. One of the people eating the meat who was both immunocompromised and homosexual would be the pivotal point, allowing for the disease to spread to vacationing gay tourists in Haiti." In December, 1982, we looked for antibody to ASFV in serum from Haitian patients with AIDS. In the serum of eight patients (and four normal controls) there was no evidence of antibody to ASFV by immunoelectro-osmophoresis or by indirect immunofluorescence. ASFV antibody can be detected in serum of infected swine by these two methods. If ASFV was related to AIDS we would expect to have found specific antibodies to the virus in at least some of our patients. Our findings argue strongly against ASFV being the AIDS agent. We are now looking for ASFV in necropsy or biopsy material from AIDS patients. The hypothesis that AIDS originated in Haiti has been raised at least twice without any scientific basis. Such speculation is damaging to Haiti and to Haitian communities abroad. EMMANUEL ARNOUX JEAN MICHEL GUERIN RODOLPHE MALEBRANCHE ROBERT ELIE A. CLAUDE LAROCHE GERARD PIERRE
de Recherche sur les Maladies Immunitaires en Haiti
MAX MILLIEN FAROUK M. HAMDY
Veterinary Laboratory, Port-au-Prince, Haiti
HAEMORRHAGIC FEVER WITH RENAL SYNDROME RELATED VIRUS IN INDIGENOUS WILD RODENTS IN BELGIUM
SIR,-Using a modification of the indirect immunofluorescent antibody technique (IFA) of McCormick et allwhich will be described elsewhere, we have detected antibody against Hantaan (haemorrhagic fever with renal syndrome, HFRS) virus in the sera of wild rodents (1 Apodemus sylvaticus and 27 Clethrionomys glareolus) trapped in non-port areas of Belgium. Animals were trapped in five different areas of Belgium-2 at Leuven (in Brabant Province, north Belgium), 98 at Buzenol (Luxembourg, south), 110 at Hasselt (Limburg, north-east), 48 at Den Haan (West-Vlaanderen, north-west), and 1077 at Turnhout (Antwerp, north). So far, all wild rodents (71) trapped in the Antwerp harbour area were negative when tested at 1:16 dilution. 139 animals
in 1980, the rest in 1982-83. 10444
1. Hess WR. African swine fever: a reassessment. Adv Vet Sci Comp Med 1981; 25: 39-67 McCormick JB, Palmer EL, Sasso DR, Kiley MP. Morphological identification of the agent of Korean haemorrhagic fever (Hantaan virus) a member of the bunyaviridae
1982; i: 765-68.
111 316 Cglareolus, 16 Rattus rattus, and 2 unknown were tested. The positive sera, with titres varying between 1:16 and 1:2048, were all from the Turnhout area. Further evidence for HFRS in Belgian wild rodents was obtained bv detection, using a modification of the enzyme-linked immunoassay described previously,3of HFRS antigen in lung tissue of 20 of 56 (35%) Cglareolus trapped in Turnhout in 1982 and in Mav. 1983. Lung suspensions (one lung in 0 - 75 mg phosphatebuffered saline with 1:1000 formaldehyde) were incubated overnight at 4°C in wells of microtitre plates precoated with 50 g/ml purified IgG of a convalescent human serum of an HFRS patient from Bashkiria (European focus of HFRS in the USSR) or purified IgG from a control. After repeat washings the antigen was detected by incubation for 60 min at 300C with peroxidase-labelled purified IgG of the same convalescent patient. Lung suspensions were tested in duplicate, and the average optical density (OD) on both immune IgG and normal IgG was determined. The ELISA test was taken as positive when the OD of the lung suspension in the presence of immune IgG was significantly larger than the OD of the lung suspension in the presence of normal IgG. The significance was evaluated by Student’s t test. analysed ELISA results correlated well with naked-eye readings, the background for lung suspensions on normal IgG-coated wells being very low. In the absence of conjugate, no endogenous peroxidase could be detected in the lung suspension. For 20 animals the presence or absence of antibodies against Hantaan virus paralleled completely the presence or absence of HFRS antigen in the lungs. For 3 animals with a weak antibody titre (1:16) no antigen could be detected. 3 very strongly antigenpositive animals showed no antibodies of the IgG type; perhaps this indicates recent infection. The positive rodents were trapped on a nature reserve at Turnhout and there was a pronounced correlation between Hantaan positive animals and wet parts in the area studied. The reactivity of the rodent sera with Hantaan virus and of the antigen in the lungs with convalescent sera of a European USSR HFRS patient, indicated the existence of a new subtype of HFRS virus in wild rodents in Europe. Further efforts are underway to isolate the virus in cell culture.
7:’jncM, 28 A flavicolis,
of Tropical Medicine, Antwerp, Belgium
Institute of Poliomyelitis and Virus Encephalitides of the USSR, Academy of Medical Sciences, Moscow USSR Laboratorium
Rijksuniversitair Centrum, Antwerp
E. A. TKACHENKO A. P. IVANOV R. VERHAGEN
GARDNERELLA VAGINALIS: PATHOGEN OR COMMENSAL?
attempt to clarify the possible pathogenic role of vaginalis in female patients complaining of vaginal discharge, all vaginal swabs submitted to this laboratory since April 1, 1982, have been examined for the presence of G vaginalis and other recognised vaginal pathogens. Specimens from the department of genitourinary medicine were not included. Vaginal discharge was collected on cottonwool swabs and transported to the laboratory in Amies transport medium. A wet
examined for pus cells and Trichomonas vagina lis.. All swabs were inoculated onto the following culture media: Columbia agar (CA) containing horse blood, CA + human blood, CA + lysed horse blood with antibiotics selective for Neisseria gonorrhoeae, and malt extract agar selective for Candida spp. All culture plates were incubated for 48 h at 37°C in an atmosphere containing 5% C02Selective and non-selective media for anaerobic bacteria were included only if the clinical information was relevant (eg, pelvic mflammatory disease, infected intrauterine contraceptive device, postnatal, postoperative). Isolation of Chlamydia trachomatis and’ mount was
3. Tkachenko EA.
Ivanov AP, Dzagurova TK, Donets MA, Rezaphin GV, Leshchinskaya EV Immunosorbent assays for diagnosis of haemorrhagic fever with renal syndrome Lancet 1981; i: 257-58.
4. Arkin H, Colton RR. Tables for statisticians. New York: Barnes Noble 121.
TABLE I-ORGANISMS ASSOCIATED WITH G VAGINALIS
TABLE II-RESPONSE AFTER TREATMENT WITH METRONIDAZOLE
Mvcoplasma spp was not attempted. Herpes simplex culture was done, in another laboratory, when specifically requested by the clinician. After 48 h incubation, colonies showing &bgr;-haemolysis on human blood agar were gram-stained to confirm gram-variable bacilli. Isolates were identified as G vaginalis if subsequent disc sensitivity testing on human blood agar incubated anaerobically revealed resistance to metronidazole 5 g, sensitivity to metrpnidazole 50 fg, and, latterly, sensitivity to trimethoprim 5
Long, unpublished)-included the additional of catalase, peroxide inhibition, and fermentation of glucose, maltose, and starch; with experience, these were not found to be in this laboratory (F. E.
essential in our routine identification scheme. Between April 1, 1982 and April 1, 1983, 9225 vaginal swabs were submitted for culture. G vaginalis was cultured from 623 patients (6’7%). Table i shows the organisms found in association with G vaginalis. Anaerobic bacteria coexisted with G vaginalis in 25 of 42 (60%) specimens from which they were specifically sought. Whenever G vaginalis was cultured from a vaginal swab, treatment with metronidazole 400 mg three times daily by mouth for 5 days was recommended, and a follow-up swab after treatment was requested. One or more follow-up swabs were received from 218 of the 623 patients (35%). All but 5 of these patients had been treated with metronidazole (3 received erythromycin, 1 topical sulphonamides, and 1 received no antimicrobial). The main reason for alternative therapy was the complaint of nausea attributed to metronidazole. Many patients were lost to follow-up, their symptoms having resolved. They consulted their physician only if the discharge recurred. Table n correlates the clinical response with the presence of G vaginalis in post-treatment specimens. Symptomatic, improvement followed the eradication of G vaginalis from the vagina in 50% of the treated patients. There was no improvement, irrespective of the isolation of G vaginalis, in 22% of patients. Of those patients whose symptoms improved shortly after metronidazole therapy, 15 subsequently relapsed from a few days to several months after treatment. G vaginalis was reisolated in 9 of these cases. In those cases where G vaginalis was still present, failure to respond may have been due to reinfection by the sexual partner (treatment of males has been found to be beneficial). 5 strains of G vaginalis cultured from patients whose symptoms persisted showed in-vitro resistance to metronidazole 50 µg. An additional factor responsible for the persistence or recurrence of symptoms was the frequent acquisition of vaginal candidosis after metronidazole therapy. Other vaginal pathogens such as Chlamydia, which we did not investigate, may have been important aetiological agents and would not have been eradicated by metronidazole. It would seem that in a large number of women with symptoms of vaginal discharge, a symptomatic improvement can be expected following the use of metronidazole. A prospective study is now in hand to assess the pathogenicity of G vaginalis in a small group of patients who will be followed up over many months. Department of Microbiology and Public Health Laboratory, Greenbank Hospital, Plymouth, Devon PL4 8NN
SHEENA REILLY R. P. HUMAN