Prenatal determination of fetal RhD type

and colleagues (April 23, p 993) conclude that they had recorded Alzheimer’s disease (AD)-like changes in tau protein processing in 7 of 15 renal dialysis patients. All shown to react positively with this antiserum were assumed to be hyperposphorylated tau. However, none of the abnormal phosphorylation sites of tau are known to reside in the amino terminal 16-aminoacid residues. Furthermore, the presence of a few individual paired helical filaments (PHF) shown in figure 3 might represent ageassociated changes. They rightfully dismiss the role of aluminium in P-amyloid because of the lack of an association between degrees of exposure to aluminium and frequency of plaques. The presence of many Ap deposits in the form of neuritic plaques are known to be essential for diagnosis of AD. Taken together, Harrington’s data support the current notion that aluminium does not play a part in the formation of neuritic plaques or neurofibrillary tangles in AD.

polypeptides

pathology, a feature regarded by many as being prerequisite for the clinical symptoms of AD.

a

necessary

Charles R Harrington, Claude M Wischik, Fiona K McArthur, Geoff A Taylor, James A Edwardson, John M Candy Cambridge Brain Bank Laboratory, Department of Psychiatry, University of Cambridge Clinical School, MRC Centre, Cambridge 1

2

3

4

CB2

2QH,

UK

Goedert M, Spillantini MG, Cairns NJ, Crowther RA. Tau proteins of Alzheimer paired helical filaments: abnormal phosphorylation of all six brain isoforms. Neuron 1992; 8: 159-68. Novak M, Kabat J, Wischik CM. Molecular characterization of the minimal protease resistant tau of the Alzheimer’s disease paired helical filament. EMBO J 1993; 12: 365-70. Bondareff W, Harrington CR, McDaniel SW, Wischik CM, Roth M. Presence of axonal paired helical filament-tau in Alzheimer’s disease: submicroscopic localization. J Neurosci Res (in press). Mukaetova-Ladinska EB, Harrington CR, Roth M, Wischik CM. Biochemical and anatomical redistribution of tau protein in Alzheimer’s disease. Am J Pathol 1993; 143: 565-78.

Henry M Wisniewski Institute for Basic Research in USA

Authors’

Developmental Disabilities, Staten Island, NY 10314,

reply

SiR-Wisniewski is correct to point out that the antiserum BR133 does not recognise phosphorylation-dependent epitopes in tau protein. This antibody was used to visualise tau proteins that are abnormal in two respects: first, in their sarkosyl insolubility, and second, in their decreased electrophoretic mobility. Normal tau proteins were not noted in this sarkosyl-insoluble fraction. Although we have not yet identified the phosphorylation sites in tau from renal dialysis patients, the physicochemical properties of this tau remain similar to the hyperphosphorylated tau characteristic of AD.’1 The presence of hyperphosphorylated tau, however, was just one of several abnormal changes in tau processing that were associated with brain aluminium accumulation in dialysis patients. Truncation of tau at Glu-391, a characteristic feature of tau protein within the PHF-core2 was recorded in both supernatant and PHF-core fractions from white matter. Normal tau protein values were depleted in grey matter, and PHF-tau was correlated with a depletion of normal tau in white matter. Finally, PHFs were detected in those cases with the highest values of PHF-core tau and phosphorylated tau. We speculated that the white matter changes represent early stages in the formation of PHFs, and we have lately identified PHF-tau in axons in AD.3 The changes in tau processing were seen in dialysis cases (mean age 57-1 years) with no evidence of a family history of AD; they did not occur in age-matched controls. Furthermore, PHFs were detected in frontal cortex, a region usually spared in non-demented controls, and at a frequency greater than expected for AD in this age group. We think it likely, therefore, that these are aluminium-associated rather than age-associated changes. Garruto and Brown in their commentary (April 23, p 989) suggest that our findings of the age-related loss of normal tau protein4 might account for dementia in AD. This suggestion seems unlikely, since several features distinguish AD from normal ageing: normal tau is depleted still further in AD than in elderly controls; AD cases, but not controls, are associated with the accumulation of PHF-core tau; and there is an extensive redistribution of tau protein from the axon in controls to insoluble tau in the somatodendritic compartment in AD.4 Although aluminium does not cause AD in dialysis patients, our data suggest that it could trigger AD-like changes in tau processing. Thus, for individuals with a genetic predisposition to AD, environmental toxic factors might influence the rate of development of PHF-type

Prenatal determination of fetal RhD type SiR-Since the molecular cloning of the genes encoding rhesus (Rh) antigens, efforts have been directed towards the development of methods for DNA-based Rh typing, particularly for the prenatal assessment of fetal RhD status. The polymerase chain reaction (PCR) may allow early’ and even non-invasive2 determination of fetal RhD type, since the RhD-positive/negative polymorphism is reported to correlate with the presence or absence of the entire RHD gene in all samples studied.3 The two published methods have assayed for the presence of exon 10 of the RHD gene, since it is here that differences between it and the related RHCE gene are greatest. However, doubts have been expressed about the reliability of these assays, as discrepancies between serological and PCR results have been found. Our results have revealed alternative molecular mechanisms responsible for lack of RhD expression, and that serologically RhD-negative donors may retain significant portions of the RHD gene. Moreover, we have found that genes lacking certain RHD exons may be capable of RhD antigen expression. The Black RhD-negative complex dCces contains an internally deleted RHD gene in which exon 10 gives a normal PCR product.5 A hybrid transcript corresponding approximately to this internal deletion has also been found in a white donor with the phenotype dCe (our unpublished results). We note that the phenotypes of the false-positive samples found by Simsek et al4 (dC[weak]cee and dCee) could result from the genotypes dccesldce and dCe/dCe, respectively. Our findings explain some of the discrepancies found with use of RHD exon 10 primers, and also suggest why intron 4 RHD PCR primers were more reliable in

detecting RHD-negative genomes.4 Furthermore, RHD transcripts with deletions corresponding to exons 7-9 and a UGA stop codon in exon 5 have been isolated from an individual of phenotype cde (our unpublished observations). Within some oriental populations, our unpublished work has distinguished two classes of D-negatives; those with a full RHD deletion and totally absent D antigen, and those with a grossly intact RHD gene and very low levels of antigen detectable only by specialised elution assays. More alarming to clinicians is the demonstration of false-negative results by Simsek et al. In certain RhD-positive donors we have found rearranged genes in which RHD exons are either absent or replaced by RHCE exons. One of these rearrangements involves exon 10. Our knowledge of the structure of the RH locus is far from complete, and variation in gene organisation in different populations is emerging. It is therefore essential

205

that there

are more

genotyping

method

studies to by PCR.

assess

the

validity

of any RHD

B Carritt, F J Steers, N D Avent MRC Human Biochemical Genetics Unit & MRC Blood Group Unit, Galton Laboratory, University College London, London NW1 2HF, UK; and International Blood Group Reference Laboratory, Bristol

1

2

3

4

5

Bennett PR, Le Van Kim C, Colin Y, et al. Prenatal determination of fetal RhD type by DNA amplification. N Engl J Med 1993; 329: 607-10. Lo Y-MD, Bowell PJ, Selllinger M, et al. Prenatal determination of fetal RhD status by analysis of peripheral blood of rhesus negative mothers. Lancet 1993; 341: 1147-48. Colin Y, Cherif-Zhar B, La Van Kim C, Raynal V, Van Huffel V, Cartron J-P. Genetic basis of the RhD-positive and RhD-negative blood group polymorphism as determined by Southern analysis. Blood

1991; 78: 2747-52. Simsek S, Bleeker PMM, von den Borne AEGK. Prenatal determination of fetal RhD type. N Engl J Med 1994; 330: 795. Blunt T, Daniels G, Carritt B. Serotype switching in a partially deleted RHD gene. Vox Sang (in press).

Haemolytic-uraemic syndrome in adults with resistant Shigella dysenteriae type I SiR-The haemolytic-uraemic syndrome (HUS) is a welldescribed complication of Shigella dysenteriae type I infection in childhood.’ It is defined by the triad of acute renal insufficiency, microangiopathic haemolytic anaemia, and thrombocytopenia. HUS associated with S dysenteriae type I in adults is rarely We report 7 cases of HUS in adults associated with an outbreak of resistant S dysenteriae type I at Shongwe Hospital in the Eastern Transvaal province of South Africa. The hospital is situated in a rural area close to the borders of Swaziland and Mozambique. Between February, 1994, and May, 1994, we isolated S dysenteriae type I resistant to ampicillin, co-trimoxazole, and chloramphenicol from the stools of 34 patients and from the blood of 2 patients admitted to Shongwe Hospital with bloody diarrhoea. All the patients were previously healthy. The organism was sensitive to two oral agents-nalidixic acid and ciprofloxacin-as well as to the parenteral agent ceftriaxone. The mean (SD) patient age was 32 (13) years. All patients presented with abdominal cramps and dysentery. The median duration of symptoms before presentation to hospital was 3 days. 5 patients had upper gastrointestinal symptoms as part of their presenting complaint. 9 were hyponatraemic during their admission to hospital (sodium 112-129 mmol/L). 2 patients required hypertonic saline to correct severe hyponatraemia. 5 had hypokalaemia of less than 3-0 mmol/L on presentation. Nosocomial transmission from patient to a nurse was documented in 1 case. The most striking complication of the epidemic was the development of HUS in 7 patients. In 4 of these patients S dysenteriae type I was isolated. The 3 other patients, admitted the during epidemic with dysentery,

thrombocytopenia, renal failure, and microangiopathic haemolytic anaemia, were identified from a review of hospital records. Supportive diagnostic criteria included a substantial drop in haemoglobin concentration, evidence of red-cell fragmentation, thrombocytopenia, reticulocytosis, normal coagulation studies, negative Coombs’ test, unconjugated hyperbilirubinaemia, raised serum lactate dehydrogenase, a negative malaria smear, and renal failure in the absence of prerenal azotaemia. Clinical features of the 7 patients are summarised in the table. One 22-year-old patient who was treated with antibiotics to which the shigella was resistant died. None of the patients required dialysis. 2 patients required blood transfusions. S dysenteriae has until recently been the rarest of the shigella species encountered in South Africa. In 1990-91 S dysenteriae was identified only twice (27%) among 73 shigella isolates received by the South African Institute for Medical Research. During 1992-93, S dysenteriae isolations had increased to 9 of 101 (89%). This is the first epidemic of S dysenteriae type I reported in South Africa. S dysenteriae type I is responsible for outbreaks of bloody dysentery in neighbouring countries including Zimbabwe and Mozambique. Susceptibility studies indicate resistance to all commonly available oral agents except nalidixic acid and ciprofloxacin. Medical practitioners and health-care workers treating patients from Southern Africa with bloody diarrhoea should be aware of the resistance pattern of this organism. It is of note that this outbreak of S dysenteriae type I in the Eastern Transvaal has been primarily associated with morbidity and mortality in an adult population due to HUS. No cases of HUS were detected in the infant or paediatric wards of our hospital during the same period. Bloom, A P MacPhail, K Klugman, M Louw, C Raubenheimer, C Fischer

P D

Shongwe Hospital, Eastern Transvaal; and Departments of Medicine and Medical Microbiology, University of the Witwatersrand, Parktown 2193, South Africa; and South African Institute for Medical Research, Johannesburg, South Afric

2

F, Levin JL, Walker L, et al. Hemolytic-uremic syndrome after shigellosis. N Engl J Med 1977; 298: 927-33. Neild GH. Haemolytic-uraemic syndrome in practice. Lancet 1994;

3

343: 398-401. O’Riordan T, Kavanagh P, Mellotte G,

Koster

1

et al. Haemolytic uraemic syndrome with shigella. Ir Med J 1990; 83: 72-73.

CORRECTIONS

Diagnosis of growth-hormone deficiency m adults-In this article by D M Hoffman and colleagues (April 30, p 1064), the y axis of panel C, figure 2, was incorrectly labelled. The correct figure is printed here. In addition, line 8 of the first full paragraph on p 1065 should have read "testosterone enanthate 250 mg intramuscularly every three or four weeks for men". HCV and

*Platelet counts may be overestimated of red-cell fragments.

by automated

Table: Patients with features of HUS

206

counters because of presence

Gammagard n France-In the news item by J-M Bader 1628), there was a typographical error. In the last sentence of the penultimate paragraph the reference to polymerase chain reaction applied to HCV, not HIV. (June 25,

p