Clinical rotation in pathology: description of a case based approach.

BACKGROUND: The implementation of a system based, integrated curriculum at the Faculty of Health Sciences of Stellenbosch University, Western Cape, South Africa, resulted in less contact time for the pathology disciplines during theoretical modules, while a weekly rotation in pathology was introduced during clinical training in the fourth and fifth years. OBJECTIVE: To describe a problem based approach for this rotation. METHODS: Students are presented with a clinical "paper" case daily, integrating as many of the pathology disciplines as possible to demonstrate the interdependence of the various disciplines. They receive chemical pathology tutorials, visit the various laboratories, and receive practical training in fine needle aspiration biopsy. On the final day, the case studies are assessed and discussed. RESULTS: Most students appreciated all activities. This rotation enhanced student interactivity and autonomy and guaranteed immediate feedback. On evaluation of the rotation it was found that the students enjoyed the rotation, learnt something new, and realised the value of group work. CONCLUSIONS: This innovation integrates pathology with clinical practice and illustrates the use of laboratory medicine in the management of common diseases seen in this country. Students appreciate learning practical skills and having to request special investigations under a pathologist's supervision changes their approach to pathology requests. Familiarity with the pathology environment empowers the student to use pathology with greater ease. A bank of case studies that can be expanded to include all medical disciplines will facilitate the application of a problem based approach and enhance communication between the basic science disciplines and the clinical and pathology disciplines.

Iron status of South African women working in a fruit-packing factory.

OBJECTIVE: The aim of this study was to determine the iron status, and the risk factors for iron deficiency (ID) and iron-deficiency anaemia (IDA), of non-pregnant adult women working in a fruit-packing factory. DESIGN: A cross-sectional analytical study was done on 338 women, 18 to 55 years of age. Information on demographic data, risk factors for ID, smoking, and the consumption of red meat, chicken and fish was collected by questionnaire. Height and weight were measured and the body mass index (BMI) calculated. A non-fasting venous blood sample was analysed for haemoglobin (Hb), serum ferritin (SF), serum iron, serum transferrin and C-reactive protein; transferrin saturation (TFS) was calculated. SETTING: Fruit-packing factory in the Western Cape, South Africa. RESULTS: The mean value for Hb was 13.06 (standard deviation (SD) 1.16) g dl-1 and for SF 48.0 (SD 47.8) microg l-1 (geometric mean 26.44 microg l-1). Women were categorised on the basis of iron status: 60% had a normal iron status (NIS); 12.6% had low TFS (<16%) but normal Hb (>or=12 g dl-1) and SF (>or=12 microg l-1) concentrations (LTS); and 27.4% had low iron status (LIS), defined as combinations of low SF (<12 microg l-1 or <20 microg l-1), low TFS (<16%) and low Hb (<12 g dl-1). More than 30% of the women were obese The risk ratio for LIS (LIS vs. NIS) was 3.8 (95% confidence interval (CI) 1.9-7.6) if women were still menstruating or 3.2 (95% CI 1.6-6.2) if they were pregnant during the past 12 months. Women with LIS consumed significantly smaller portions of red meat, chicken and fish than did women in the other two groups. CONCLUSIONS: IDA (low Hb, SF and TFS) and ID (low SF and TFS) did not seem to be a major problem. Women who were still menstruating or were pregnant during the past 12 months were at greater risk for ID. The consumption of smaller portions of red meat, chicken and fish was related to LIS. A high prevalence of obesity, which demonstrated the coexistence of both under- and overnutrition, was observed.

Prenatal diagnosis of congenital leukemia in a fetus at 25 weeks' gestation with Down syndrome: case report and review of the literature.

Leukemia in a hydropic fetus with hepatosplenomegaly and Down syndrome was diagnosed at 25 weeks' gestation. Spontaneous demise occurred 10 days after the cordocentesis. A review of the literature of the seven cases described antenatally and 10 cases described in the immediate neonatal period or on examination of the stillborn baby is presented.

Ferritin assay performed on the Technicon Immuno 1 analyser using serum and plasma samples.

Ferritin assays were performed using both serum and plasma samples on the Bayer Technicon Immuno 1 Analyser and their ferritin test kit as only data for serum samples are available. After evaluation of the test kit and analyser the results were correlated using the Statistica for Windows programme. It was established that the plasma samples gave consistently lower values than the serum samples. Using the obtained conversion factor of 1.191, plasma as well as serum samples may be used for ferritin determinations.

Serum concentrations of some metals and steroids in patients with chronic fatigue syndrome with reference to neurological and cognitive abnormalities.

Chronic fatigue syndrome is defined by the Atlanta Centers for Disease Control (Atlanta, GA, USA) as debilitating fatigue lasting for longer than 6 months. Symptoms include disturbances of cognition. Certain factors have in the past been shown to influence cognition, including metals such as aluminum, iron, and zinc; and steroids such as dehydroepiandrosterone. In the present study, concentrations of these factors were determined in the serum and plasma of patients and their age- and gender-matched healthy controls (10 women and 5 men in each group). In addition, copper, dehydroepiandrosterone sulphate, cortisol, cholesterol, hemoglobin, ferritin and transferrin concentrations, as well as transferrin genetic subtypes were determined in both groups. The results indicate that patients had significantly increased serum aluminum and decreased iron compared to controls. In the females, serum iron and dehydroepiandrosterone sulphate were significantly decreased and correlated. Total cholesterol was significantly increased, and significantly negatively correlated with dehydroepiandrosterone sulphate. There were no differences in zinc, copper, cortisol, hemoglobin, transferrin and ferritin concentrations, or in transferrin genetic subtypes.

Analysis of the NRAMP1 gene implicated in iron transport: association with multiple sclerosis and age effects.

Multiple sclerosis (MS) is believed to be an autoimmune process occurring in genetically susceptible individuals after an appropriate environmental exposure. We have exploited the homogeneous Afrikaner population of European ancestry to investigate the likelihood that iron dysregulation, in association with infectious and/or autoimmune disease susceptibility, may underlie the MS phenotype in a subgroup of patients. The functional Z-DNA forming repeat polymorphism of the natural resistance-associated macrophage protein-1 (NRAMP1) gene was analyzed in 104 patients diagnosed with MS and 522 Caucasian controls. A family-based control group consisting of 32 parental alleles not transmitted to MS offspring was additionally studied to exclude the likelihood of population substructures. Statistically significant differences in allelic distribution were observed between the patient and control samples drawn from the same population (P < 0.01). Evidence is furthermore provided that alleles considered to be detrimental in relation to autoimmune disease susceptibility may be maintained in the population as a consequence of improved survival to reproductive age following infectious disease challenge. Although it remains to be determined whether the disease phenotype in MS patients with allele 5 of the NRAMP1 promoter polymorphism is directly related to dysregulation of iron or modified susceptibility to viral infection and/or autoimmunity, a combination of these processes most likely underlies the disease phenotype in these patients. In view of the emerging role of polymorphic variants in complex diseases and minimizing of possible confounding factors in this association study, we conclude that allelic variation in the NRAMP1 promoter may contribute significantly to MS susceptibility in the South African Caucasian population.

Band 3 Cape Town (E90K) causes severe hereditary spherocytosis in combination with band 3 Prague III.

Hereditary spherocytosis (HS) is an inherited haemolytic anaemia, characterized by spheroidal, osmotically fragile red blood cells. This disorder exhibits heterogeneity in terms of both clinical severity and underlying molecular defect. We have studied a South African Cape Coloured individual with severe HS owing to a band 3 deficiency caused by two mutations, occurring in trans, in the band 3 gene: a novel variant that we have designated band 3 Cape Town and a previously described mutation, band 3 Prague III. Analysis of erythrocyte membrane proteins indicated a deficiency of both band 3 and protein 4.2, as well as a decreased functional capacity of band 3 to transport anions. Band 3 Cape Town is defined by a GAG-->AAG point mutation at codon 90, substituting a glutamic acid with a lysine in the cytoplasmic domain of the molecule, while band 3 Prague III is a codon 870 CGG-->TGG point mutation, replacing an arginine with a tryptophan in the transmembrane region of band 3. mRNA is transcribed from both mutant alleles, implying that mutant proteins are synthesized, but are either degraded prior to membrane incorporation or insertion is impaired. We conclude that the combination of these two mutations exacerbated the clinical presentation of the proband.

Prognostic value of day 14 blast percentage and the absolute blast index in bone marrow of children with acute lymphoblastic leukemia.

The product of the percentage blasts on the bone marrow aspirate (BMA) and the biopsy cellularity has been termed the "absolute blast index aspirate" (ABI-aspirate) and is used to measure disease response on day 7 of induction therapy. The authors compared the event-free survival (EFS) in high-risk and standard-risk patients as identified by the ABI-aspirate and the BMA percentage blasts on day 14 of induction therapy. Both indices identified high-risk cases. EFS of patients categorized as high-risk by these 2 methods and the high-risk criteria used by the authors' service (WCC of > 20 x 10(9)/L, age < 2 and > 8 years and a peripheral blood blast count of > 1.0 x 10(9)/L on day 8 of induction) did not differ. There was concordance between patients identified as high risk by all 3 methods. The results confirmed the prognostic value of the ABI-aspirate and the BMA percentage blasts on day 14 of induction therapy, but these methods were not superior to the high-risk criteria currently in use.

Factor V Leiden mutation and the risk of thrombo-embolic disease in pregnancy: a case report.

Factor V Leiden mutation is a risk factor for the development of thrombo-embolic episodes in pregnancy. A case is presented of a pregnant woman with repeated episodes of venous thrombosis with a complicated clinical course.

2'-Chlorodeoxyadenosine Effectively Induces Complete Remission in Hairy Cell Leukaemia.

Hairy cell leukaemia, previously known as leukaemic reticuloendotheliosis, is an indolent lymphoproliferative disorder of unknown etiology. It typically affects males, causes marked splenomegaly and moderate enlargement of the liver, whilst lymphadenopathy is inconspicuous. Pancytopenia is characteristic with unusually profound monocytopenia, variable reduction in platelets, and the presence in the peripheral blood and marrow of abnormal small lymphocytes having irregular cytoplasmic margins. Ultrastructure, combined with cytochemistry and flow cytometry, have refined diagnosis. A variant exists between this classical entity and B prolymphocytic leukaemia, where blastic transformation or massive lymph node enlargement are found, and this is of ominous significance. In all these patients with this entity conventional chemotherapy is ineffective and shortens survival. Our previous experience with splenectomy results in excellent clinical control for long periods of time, but without disease eradication. There followed a vogue for the use of interferon but this is limited by high cost and dose-dependent side-effects. Contemporary management centres on the purine analogues, where durable responses are possible with fludarabine and deoxycoformycin, but best with 2'chlorodeoxyadenosine (2-CDA). To document the efficacy of the latter agent, we analysed the outcome in seventeen consecutive patients treated over the last five years. Four were ineligible for analysis, although two had 2-CDA. The other thirteen, managed on a standard seven-day course of 0.1 mg/kg 2-CDA given as a continuous intravenous infusion, all responded promptly. Apart from transient leucopenia complications have been minimal, and oral co-trimoxazole prophylaxis for pneumocystis carinii was maintained during the first one year. In all thirteen there was a rapid return to normal of peripheral blood count and marrow on aspiration and trephine biopsy. Even in the longest follow-up clinical and haematologic remission has been maintained and no patients have required retreatment. One individual has relapsed in the marrow at two years. Despite the relative expense of the agent the excellent treatment outcome and patient acceptability, coupled with its safety, leads to the recommendation that in South Africa-as elsewhere in the world-this be regarded as the first line of treatment.

Outbreak of Erwinia carotovora on Zantedischia spp. in South Africa.

In South Africa, summer-flowering Arum lilies are grown for the tuber, potted plant, and cut flower markets. In 1998, an outbreak of soft rot was detected on Zantedischia oculata cv. Black Magic and Z. elliottiana plants from several nurseries. Crop losses of up to 25% were incurred. The initial symptom was wilting of leaves. When plants were lifted from the soil, soft rot of the tuber was found. Tuber rot usually developed on one side, and plants developing from affected tubers wilted and died. No discoloration of leaf or tuber tissues was found. Isolations from diseased tissues consistently yielded bacterial colonies that were translucent, white, and glistening and that had entire margins on nutrient agar. Ten representative isolates were chosen for further characterization. Erwinia carotovora subsp. carotovora strain B56 was included as a reference strain. All isolates were gram-negative rods, oxidase and arginine dihydrolase negative, catalase positive, and facultatively anaerobic. They degraded pectate and rotted potato slices but did not hydrolyze starch. All isolates fermented glucose, reduced nitrates to nitrites, and grew at a maximum temperature of 37°C. Isolates produced acids from D(+)-glucose, D(+)-cellobiose, melibiose, amygdalin, L(+)-arabinose, D-mannitol, L(+)-rhamnose, sucrose, ribose, D(-)xylose, and D(-)glucose but not from D-arabinose, D-sorbitol, or maltose. Isolates liquefied gelatin and used citrate, arbutine, esculin, salicin, and cellobiose as the sole carbon source. Pathogenicity to Zantedischia spp. was tested by injection of tubers with an inoculum suspension containing 108 CFU/ml. Control plants were inoculated with sterile distilled water. Inoculated plants were kept in a greenhouse at 24°C. Symptoms developed 2 days after inoculation with the pathogen and appeared to be identical to those observed on diseased material in nurseries. Control plants did not rot. The bacterium was readily reisolated from diseased plants, confirmed to be the inoculated pathogen, and identified as E. carotovora, based on morphological, biochemical, and physiological characteristics and pathogenicity. E. aroideae has been reported to cause soft rot of rhizomes of winter-flowering Arum lilies (Z. aethiopica) in South Africa (1). However, this is the first report of soft rot caused by E. carotovora subsp. carotovora on tubers of Z. oculata and Z. elliottiana plants in South Africa. Reference: (1) V. Wager. 1970. Flower Garden Diseases and Pests. Purnell, Cape Town, South Africa.

First Report of Crown Gall, Caused by Agrobacterium tumefaciens on Geraldton Wax (Chamelaucium uncinatum) in South Africa.

Geraldton wax (Chamelaucium uncinatum, family Myrtaceae) plants are grown as cutflowers for the export market in South Africa. In July 1998, gall-like structures were observed on collars and roots of Geraldton wax plants in commercial fields in Wellington. The galls were observed after plants exhibited poor growth. The galls varied in size and in texture from soft and spongy to hard. Secondary symptoms involved poor root development and browning of stem tissues near galls. Isolations from the galls yielded nearly pure cultures of a Gram negative, rod-shaped bacterium on Roy Sauer medium (2), typical of an Agrobacterium sp. Carbon source utilization testing with the Biolog GN Bacterial Identification System (version 3.50) confirmed the bacterium as a biovar of A. tumefaciens with a similarity of 0.88. Pathogenicity was confirmed by injecting 4- to 6-week old tomato and tobacco plants and 1-year-old Geraldton wax plants with approximately 5 µl of the bacterial suspension (108 CFU/ml) in sterile, distilled water. Inoculated plants were then transferred to a greenhouse at 25°C. Galls developed 1 month after inoculation. The bacterium was readily reisolated from the inoculated plants. A. tumefaciens is endemic to South Africa and has a very wide host range that includes several ornamentals (1). This is the first report of A. tumefaciens on Geraldton wax plants in South Africa. References: (1) J. F. Bradbury. 1986. Guide to Plant Pathogenic Bacteria. CAB Int., Slough, U.K. (2) N. W. Schaad. 1988. Laboratory Guide for Identification of Plant Pathogenic Bacteria. The American Phytopathological Society, St. Paul, MN.

Analysis of the F8 gene in individuals with high plasma factor VIII: C levels and associated venous thrombosis.

High FVIII:C levels have previously been shown to be an independent risk factor for thrombosis with 4.8 times higher potential risk of thrombosis in individuals with FVIII:C levels greater than 1.5 u/ml. Recently, we found that raised FVIII:C levels are largely attributable to elevated FVIII:Ag levels. The determinants of FVIII:Ag levels are unclear and might be partly genetic. The promoter of the F8 gene has recently been characterised we therefore investigated the promoter and the 3' terminus of the F8 gene for possible polymorphisms associated with raised FVIII:Ag levels in 62 selected individuals with a thrombotic tendency. We confirm previous reports that raised FVIII:C levels are largely attributable to an elevation in FVIII:Ag and this is also associated with elevation of vWF; non-O blood group: relatively short APTT and relatively low APC ratio. We screened 1140 bp of the proximal promoter including the protein binding sites identified by DNase I footprint analysis by SSCP, however no polymorphisms were identified. Direct DNA sequence analysis of the region -542 to +165 failed to identify any sequence polymorphisms. The recently described polymorphism in the polyadenylation cleavage site in the prothrombin gene associated with increased prothrombin activity prompted us to screen the region surrounding the 3' terminus of the F8 gene for polymorphisms but we found none.

Response to an iron fortification programme in relation to vitamin A status in 6-12-year-old school children.

Plasma retinol and indices of iron status were measured in 148 school children (6-12 years) receiving a soup fortified with iron and vitamin C for a period of 15 weeks. The most significant change in serum iron (P = 0.0005) and transferrin saturation (P = 0.0002) was seen in subjects with plasma retinol > or = 40 micrograms/dl, while subjects with plasma retinol < 20 micrograms/dl showed no response. Serum ferritin improved most in the retinol categories < 40 micrograms/dl, suggesting that the absorption of iron was not impaired by marginal vitamin A status, but that it was rather the mobilisation of iron from stores that was affected. Changes in vitamin A status correlated positively and significantly with changes in serum iron (r = 0.37; P = 0.0001) transferrin saturation (r = 0.27; P = 0.004) and haemoglobin (r = 0.21; P = 0.03), but negatively with serum ferritin (r = -0.28; P = 0.003). The presence of marginal vitamin A deficiency in a community may limit the effectiveness of an iron intervention programme and vitamin A status should therefore also be considered when such programmes are planned.

The effect of iron fortification on the fatty acid composition of plasma and erythrocyte membranes in primary school children with and without iron deficiency.

An intervention study was designed to evaluate the fatty acid (FA) status of children aged 6-11 years before and after iron fortification. Iron-deficient (ID) and matched controls without ID (n = 30) were selected. All children received soup (160 ml) fortified with 20 mg iron and 100 mg vitamin C for 15 weeks on school days. Measurements before and after intervention included dietary intake, haematological and iron status and FA composition of plasma and erythrocyte membranes (EMBs). The prevalence of low plasma ferritin concentration and transferrin saturation decreased in the ID children by 40% and 56%, respectively, with intervention. Plasma FAs reflected dietary FA intake. In comparison with controls, the ID group presented with increased percentage total saturated FAs (SFAs; p = 0.0002) in their EMB phosphatidylcholine (PC) and reduced percentage total polyunsaturated FAs (PUFAs; p = 0.0037) before intervention. Lower total n-3 FAs (p = 0.0070), including eicosapentenoic acid (EPA; p = 0.0034), docosapentenoic acid (DPA; p = 0.0048) and docosahexenoic acid (DHA; p = 0.0058), were observed in the ID group. The EMB phosphatidylethanol-amine (PEA) of the ID children presented with lower percentages of alpha-linolenic acid (ALA; p = 0.0001), EPA (p = 0.0051) and DHA (p = 0.0084) compared to controls before intervention. Iron intervention was associated with an increase (p < 0.05) in the percentage of n-3 FAs in the EMB-PC and EMB-PEA of the ID group to percentages comparable to that in the control group. It appears that iron status can influence FA metabolism of specific n-3 FAs in the EMBs of young children.

The effect of iron fortification on the fatty acid composition of plasma and erythrocyte membranes in primary school children with and without iron-deficiency.

An intervention study was designed to evaluate the fatty acid (FA) status of children aged 6-11 years before and after iron fortification. Iron deficient (ID) and matched controls without ID (n = 30) were selected. All children received soup (160 mL) fortified with 20 mg iron and 100 mg vitamin C for 15 weeks on school days. Measurements before and after intervention included dietary intake, haematological and iron status and FA composition of plasma and erythrocyte membranes (EMBs). The prevalence of low plasma ferritin concentration and transferrin saturation decreased in the ID children by 40% and 56%, respectively, with intervention. Plasma FAs reflected dietary FA intake. In comparison with controls, the ID group presented with increased percentage total saturated FAs (SFAs; p = 0.0002) in their EMB phosphatidylcholine (PC) and reduced percentage total polyunsaturated FAs (PUFAs; p = 0.0037) before intervention. Lower total n-3 FAs (p = 0.0070) including eicosapentaenoic acid (EPA; p = 0.0034), docosapentaenoic acid (DPA; p = 0.0048) and docosahexaenoic acid (DHA; p = 0.0058) were observed in the ID group. The EMB phosphatidylethanolamine (PEA) of the ID children presented with lower percentages of alpha-linolenic acid (ALA; p = 0.0001), EPA (p = 0.0051) and DHA (p = 0.0084) compared to controls before intervention. Iron intervention was associated with an increase (p < 0,05) in the percentage of n-3 FAs in the EMB-PC and -PEA of the ID group to percentages comparable to that in the control group. It appears that iron status can influence FA metabolism of specific n-3 FAs in the EMBs of young children.

Haematological changes associated with miliary tuberculosis of the bone marrow.

The peripheral blood and bone marrow findings in 25 patients found to have tuberculous granulomata on bone marrow examination were examined to determine whether specific haematological findings are associated with tuberculous infiltration of the bone marrow. All the patients had one abnormality or more on their full blood count. The presence of a peripheral lymphopaenia was the single factor common to all 25 patients studied. The peripheral lymphopaenia was only associated with bone marrow lymphopaenia in 14% of patients. Other findings of note were an association of peripheral monocytopaenia and absence of giant cells in the granulomata, and decreased iron stores in almost a third of the total number of patients. The relevance of the lymphopaenia, monocytopaenia and decreased iron stores are discussed and we propose that the absence of a peripheral lymphopaenia makes it very unlikely that there will be tuberculous localization in the bone marrow.

CD4+, CD33-, CD13-, CD14- acute monoblastic leukaemia.

We report a case of acute monoblastic leukaemia in which the expression of the CD4 antigen occurred in the absence of myeloid and monocytic lineage specific markers. Unexpected marker profiles have biological and diagnostic implications and we also suggest that the inappropriate expression of the CD4 antigen may be implicated in the poor prognosis of this case.

Scanning Electron Microscopy of Invasion of Apple Leaves and Blossoms by Pseudomonas syringae pv. syringae.

Scanning electron microscopy indicated that Pseudomonas syringae pv. syringae L795 entered leaves through stomata and multiplied in the substomatal chambers. Strain L195 applied to blossoms colonized stigmas and also occurred in intercellular spaces of styles. Nonpathogenic strain L796 failed to colonize blossoms. This study suggests that inoculum of pathogenic P. syringae pv. syringae builds up on apple leaves and blossoms.