429

characteristics, and medication in the previous 2 weeks. Stools were examined for ova and parasites by direct wet preparation and specimens with faecal leucocytes were cultured on selective media for shigella and salmonella. 248 specimens (25%) contained B hominis, 211 contained Entamoeba histolytica, and 62 were positive for Giardia intestinalis;

Shigella and Salmonella spp accounted for 9% of specimens. No previous medication was reported for 747 (75%) samples. Tinidazole/metronidazole was reported for 107, tetracycline for 56, and co-trimoxazole for 47. Use of ampicillin, dicloxacillin, erythromycin, diiodohydroxyquine, diloxanide furoate, quinacrine, or mebendazole was reported for small numbers. Previous treatment with tinidazole/metronidazole did not affect the positivity rate for B hominis; as expected, the percentages of E histolytica and G intestinalis recovered were lower. Tetracycline use decreased the recovery rate of B hominis (25% vs 14%; p 0-08) but the striking result was for co-trimoxazole and B hominis when only 1 sample of 47 was positive (p 00004): =

=

Co-t

Organism

(n = 47)

Previous treatment Tet Tin/Met

(n = 56)

B hominis 2% 14% E histolytica 26% 21% Gmmunafu 4% 5% Co-t=co-tnmoxazo!e, Tet = tetracycline,

None

(n=107) (n=747) 23% 10%

27% 22% 3% 7% Tin/Met = tlnldazole/

metronidazole

Several antiparasitic therapies (eg, metronidazole) have been recommended for B hominis infection but their efficacy is unclear and no controlled trials have been done.1 In-vitro studies have shown an inhibitory effect against B hominis for the following drugs: emetine > metronidazole > furazolidone > co-trimoxazole > iodochlorhydroxyquine > pentamidine isethionate.2 In our study, previous use of tinidazole or metronidazole had no effect on recovery of B hominis but co-trimozazole almost completely eliminated it, while not effecting the other pathogens (ie, this result cannot be attributed to elimination of a coexisting organism). Kain et aP demonstrated a significant reduction of B hominis in patients who had recently taken salazopyrine, another sulphate-containing drug. Our findings suggest that prospective studies on the efficacy of co-trimoxazole may be warranted should B hominis be more clearly shown to be a pathogen. CIWEC Clinic,

Kathmandu, Nepal American Peace Kathmandu

ELI SCHWARTZ*

Corps Clinic,

ROBIN HOUSTON†

*,†Present addresses. *Medical Centre for the Traveler, Misgav Ladach Hospital, Jerusalem 93190, Israel; t Emory School of Public Health, Atlanta, Georgia, USA. 1. Markell EK, Udkow MP. Blastocystis hominis: pathogen or fellow traveler? Am J Trop Med Hyg 1986; 35: 1023-26 2. Zierdt CH, Swan JC, Hosseini J. In vitro response of Blastocystis hominis to antiprotozoal drugs. J Protozool 1983; 30: 332-34. 3 Kain KC, Noble MA, Freeman HKJ, Barteluk RL. Epidemiology and clinical features associated with Blastocystis hominis infection. Diagn Microbiol Infect Dis

1987; 8: 235-44.

False-positive legionella serology in campylobacter infection SIR,-Dr Boswell and Dr Kudesia (Jan 18, p 191) report the frequent detection of antilegionella antibodies in patients with campylobacter infection. We report findings that corroborate and extend their observations. In February, 1991, a 57-year-old man with microbiologically confirmed Campylobacterjejuni diarrhoea was incidentally found to have an antibody titre of 512 to Legionella pneumophila serogroup 1 by the indirect immunofluorescent antibody test (IFAT) with formalised yolk-sac antigen. The reactivity in IFAT was abolished by absorption with heat-killed Cjejuni. This absorption was specific since it had no effect on the titre of other antibodies present in the patient’s serum. IFAT-positive serum samples from patients with legionnaires’ disease were also unaffected. Since then we have obtained serum samples from 16 further patients with campylobacter infection in the acute and convalescent stages-12 with Cjejuni, 3 with C coli, and 1 untyped. None was

positive by IFAT in acute-phase samples but 4 patients were positive in samples collected 4-10 days later. Positive samples had titres of 16, 32, 128, and 1024. 2 of these were from patients with C coli, 1 from C jejuni, and 1 untyped infection. Preliminary immunoblotting with these serum samples shows that the main increase in antilegionella antibody appears as a broad lowmolecular-weight band, suggesting that lipopolysaccharide is the principal cross-reactive antigen. These epitopes elicit IgM and IgG antibody responses. Although the frequency of false-positive legionella IFAT tests in campylobacter infection (25%) was significantly lower than that recorded by Boswell and Kudesia (71 %), this nevertheless remains a potentially important source of diagnostic error, which may result in both inappropriate clinical management and epidemological investigation. We recommend that any patient with a positive legionella IFAT after diarrhoeal illness that is not associated with pneumonia has, in addition to a stool culture, a repeat IFAT test after absorption of their serum samples with heat-killed C jejuni. Formalin yolk-sac antigen was supplied by the Public Health Laboratory Service, London.

Department of Medical Microbiology, Royal Liverpool University Hospital, Liverpool L7 8XP, UK Infectious Diseases Unit, Public Health Laboratory,

J. S. CHEESBROUGH T. MAKIN B. C. TAXMAN

Regional

Fazakerley Hospital, Liverpool

N. K.

J. BEECHING J. MUTTON

Immunofluorescence technique for the identification of polioviruses SiR,—To investigate the efficacy of oral poliomyelitis vaccine or the inactivated vaccine in the induction of gut immunity to polioviruses in Gambian infants, an immunofluorescence technique with monoclonal antibodies has been developed to detect

polioviruses in stool specimens. 435 stool specimens were obtained from immunised infants one week after challenge with a standard dose of trivalent or monovalent type-1oral poliomyelitis vaccine. Poliovirus was isolated on HEP-2 Cincinnati cells and identified by the virus neutralisation technique by standard methods1 and by a new indirect immunofluorescence technique. Briefly, the Hep-2 cells were infected with a 1 in 10 dilution of a second passage of the virus in question. When 25-50% of the cells showed a cytopathic effect, they were removed from the culture vessel with phosphate-buffered saline and edetic acid (EDTA), washed and pelleted by centrifugation, and resuspended at 2 x 106 cells/ml in phosphate-buffered saline. 25 at samples of this suspension were placed on each spot of a multispot slide, air-dried, fixed in cold acetone for 5 min, and then stored desiccated in silica gel at -20°C until used. Mouse monoclonal antibodies 1588 (anti-polio type 1), 267 (anti-polio type 2), and 204 (anti-polio type 3) were used at a dilution of 1 in 300 and bound antibodies were detected by ultraviolet-light microscopy with fluorosceinisothiocyanate-labelled F(ab’)2 fragment of goat anti-mouse IgG antibody (full details available from the authors). Preliminary experiments with reference virus strains showed that the monoclonal antibodies were poliovirus type-specific and did not crossreact with the 2 coxsackie viruses (types A9 and B2) or the 2 echoviruses (types 7 and 30) tested. To see if the antibodies reacted type-specifically against both vaccine and wild-type polioviruses, we tested them by immunofluorescence against several intratypic strains of virus that had been characterised by molecular sequencing and by neutralisation with monoclonal antibodies. Monoclonal 1588 detected all 16 wild and 15 vaccine type-1strains. Antibody 267 detected 9 of 10 wild and 8 of 10 vaccine type-2 strains, and antibody 204 detected 6 of 8 wild and 10 of 10 vaccine type-3 strains. Each antibody proved type-specific and did not crossreact with heterologous wild or vaccine strains. Cytopathic effect was noted in 105 of the 435 cultures of stool, and the results of examination of these positive cultures by virus neutralisation or immunofluorescence were compared. Overall, immunofluorescence identified 64 polioviruses (39 type 1,16 type 2,

430

and 9 type 3), whereas virus neutralisation identified only 50 polioviruses (34 type 1, 13 type 2, and 3 type 3) in the same stool specimens. In each case, immunofluorescence identified the same type of poliovirus as did the viral neutralisation test but, in addition, it identified multiple poliovirus types in three specimens in which virus neutralisation only identified 1 type. Immunofluorescence also identified 6 type-1 polioviruses and 3 type-3 polioviruses in specimens in which virus neutralisation showed only the presence of an enterovirus. These reactions were not false positives since, when each type of poliovirus in the mixture was individually neutralised with its own type-specific antiserum, repeat immunofluorescence testing for that type was negative. Thus, immunofluorescence proved more sensitive, faster, and

easier to do than the classic virus neutralisation. It should be useful for epidemiological studies, especially in developing countries where the high prevalence of endemic non-polio/enteroviruses makes the virus neutralisation test arduous, time-consuming, and sometimes difficult to interpret. We thank Dr Merag Ferguson, National Institute for Biological Standards and Control, UK, for the mouse monoclonal antibodies. Medical Research Council Laboratories, PO Box 273, Banjul, The Gambia

HILTON WHITTLE DAN HAZLETT

National Institute for Biological Standards and Control, Potters Bar, Herts, UK

DAVID WOOD

Expanded Programme on Immunisation, World Health Organisation, Geneva, Switzerland

ELEANOR BELL

Organisation, Expanded Programme of Immunisation and Division of Communicable Diseases manual for the virological investigation of poliomyelitis. Geneva: WHO, 1990.

1. World Health

Co-transplantation of peripheral

nerve and adrenal medulla in Parkinson’s disease

SiR,—Transplant of adrenal mesencephalon are being tested

medulla

and

fetal

ventral

therapeutic alternatives in Parkinson’s disease. The lack of both common protocols and a controlled, multicentre trial has impeded the comparison of results and created confusion. At the Clinica Puerta de Hierro, Madrid, a controlled study is underway to establish which donor tissue, if any, is best for implantation in parkinsonian patients. An open surgery technique is used. Since 1987, by implanting perfused adrenal medulla or fetal ventral mesencephalon into right caudate nucleus of grade IV-V parkinsonian patients, we have achieved varying degrees of clinical improvement.1,2 Animal models of Parkinson’s disease3-s show that after co-implants of adrenal medulla and peripheral nerve and implants of adrenal medulla plus long-term nerve growth factor (NGF) infusion, the chromaffm cells of adrenal medulla survive better, tend to be of neuronal phenotype, and enhance sprouting of host fibres. Taking these reports into account, we present the first results on clinical progress of three grade IV Parkinson’s patients 5 months after co-implantation of adrenal as

medulla and intercostal nerve. We used the clinical protocol reported earlier.1,2 The mean age of the patients was 60 years, and the mean duration of their disease was 10-5 years; they had been on levodopa for 9-5 years. They were given the optimum medication until 2 days before surgery, at which time dopaminergic agonists were tapered off. Right adrenal gland was removed retroperitoneally via a posterior subcostal approach. The gland (unperfused) was cut into slices of about 5 mm and incubated.6 Viability was over 71 %. The 12th intercostal nerve was dissected, minced, and then incubated with adrenal medulla in enriched minimum essential medium until use about 1 h later 6 After right frontal craniectomy, 15-20 pieces of adrenal medulla and nerve ranging in size from 1 mm to 3 mm were implanted into right caudate nucleus.’z2 The patients had fewer surgical complications than did those whose gland was removed transperitoneally. No systemic or neurological complications were seen, but two patients had psychiatric complications (confusion in one, hallucinations in two, and delusions in one) that were slightly milder than they had been in

Clinical response to implant in

on

and off periods.

the first series.1 All had dyskinesias, levodopa being gradually reduced from 950 (SD 204) mg daily before surgery to 500 (200) mg by the second month. The complications seem to be secondary to the medication (ie, there were dose and time relations), and may also have been influenced by the handling of the gland. For the first 2 weeks, major improvements were noted in rigidity, facies, and ease of movement, with no appreciable changes in gait, postural stability, or other symptoms. Subsequently, as in the first series,’ the patients experienced gradual deterioration, principally with respect to kinesia. In this period, we found no significant difference in the number and duration of "on/offperiods. Once home, the patients were assessed monthly.1 The figure shows Unified Parkinson’s Disease Rating Scale data, up to 5 months postoperatively, in on and off stages. The preliminary results of this study seem to show that the clinical course, slope of improvement, and reduction of medication parallel to those observed for the first 5 months in perfused adrenal medulla transplants,’ and suggest that progress is better than it is with implants of untreated adrenal medulla. There are so far no statistically significant clinical differences to indicate whether handling the tissue is an improvement over our earlier procedure (implants of perfused adrenal medulla). If co-transplantation of nerve and medulla increases adrenal tissue viability and/or enhances and maintains the sprouting of host nigrostriatal fibres, this procedure may prolong or increase the improvement produced in parkinsonian patients by other techniques. A recent report from Oslon et al7 offers hope in this respect. JUAN J. LÓPEZ LOZANO GONZALO BRAVO JAVIER ABASCAL B. BRERA Department of Neurology, Neurosurgery, H. SANTOS Surgery, and Experimental Surgery, C. GOMEZ-ANGULO, Clinica Puerta de Hierro, Universidad Autonoma, 28035-Madrid, Spain

and CHP Neural

Transplantation Group

López-Lozano JJ, Bravo G, Abascal J, and CPH neural transplantation group. Grafting of perfused adrenal medulla tissue into the caudate nucleus of patients with Parkinson disease. J Neurosurg 1991; 75: 234-43. 2. Lopez-Lozano JJ, Bravo G, Brera B, et al. Can an analogy be drawn between the clinical evolution of Parkinson’s patients who undergo antoimplantation of adrenal medulla and those of fetal ventral mesencephalon transplant recipients? In: Lindvall O, Bjorklund A, Widner H, eds. Intracerebral transplantation in movement disorders. Amsterdam: Elsevier, 1991: 83-94. 3. Date I, Felten SY, Felten DL. Cografts of adrenal medulla with peripheral nerve enhance the survivability of transplanted adrenal chromaffin cells and recovery of the host nigrostriatal dopaminergic system m MPTP-treated young adult mice. Brain Res 1990; 537: 33-39. 4. Kordower JH, Fiandaca MS, Notter MFD, et al. Peripheral nerve provides NGF-like trophic support for grafted Rhesus adrenal chromaffin cells. J Neurosurg 1990; 73: 1.

418-28. Herrera-Marschitz M, Ungerstendt U, et al. Chronic implants of chromaffin tissue into dopamine-denervated striatum. Effects of NGF on graft survival, fiber growth and rotational behavior. Exp Brain Res 1985; 60: 335-19. 6. López-Lozano JJ, Brera B, Abascal J, Bravo G. Preparation of adrenal medullary tissue for transplantation in Parkinson’s patients: a new procedure. J Neurosurg 1989; 71: 552-55. 7. Olson L, Backlund EL, Ebendal T, et al Intraputaminal infusion of nerve growth factor to support adrenal medullary autografts in Parkinson’s disease: one year follow up of first clinical trial Arch Neurol 1991; 48: 373-81. 5.

Stromberg I,

Immunofluorescence technique for the identification of polioviruses.

429 characteristics, and medication in the previous 2 weeks. Stools were examined for ova and parasites by direct wet preparation and specimens with...
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