Anatomical Pathology

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Analysis of hereditary haemochromatosis and clinical correlations in the elderly
(Stellenbosch : Stellenbosch University, 2000-12) Bouwens, C. S. H.; Kotze, Maritha J.; Maritz, F. J.; De Villiers, J. N. P.; Stellenbosch University. Faculty of Medicine & Health Sciences. Dept. of Pathology.
ENGLISH ABSTRACT: Hereditary haemochromatosis (HH) is an autosomal recessive iron storage disease where the accumulation of iron in parenchymal organs may lead to diabetes, heart failure, liver cirrhosis, arthropathy, weakness and a variety of other ailments if preventive measures are not taken. HH is often not considered as a cause of these conditions, particularly not in the elderly where the background frequencies of type II diabetes, osteoarthritis and heart failure are generally high. Heterozygosity for C282Y, the HFE-mutation causing HH in approximately 80% of affected individuals worldwide, has been linked to a raised incidence of malignancies of the colon and rectum, stomach and the haematological system. One of the highest carrier-frequencies (116) in the world for this mutation has been reported in the South-African Afrikaner population, resulting in C282Y-homozygosity in approximately 1 in every 115 people in this group. A sample of 197 elderly Afrikaner volunteers was recruited for genotype/phenotype association studies. Their clinical presentation was denoted, biochemical iron-status determined and HFE genotyping performed. Either an increase or decrease in survival, or both, were proposed, depending on possible gender effects. HH has been positively associated with various cancer types, but may also protect against iron-deficiency anaemia which is by far the most frequent cause of anaemia in the older person. This study has led to the following findings: 1. The carrier frequency of mutation C282Y was found to be 1/8 in the elderly population (similar in males and females), which is slightly lower than the 1/6 reported in younger adults from the same population. Only one C282Y homozygote and two C282YIH63D compound heterozygotes were detected, all of them female. 2. The prevalence of diabetes, heart disease, arthropathy or a combination of these conditions did not differ significantly in C282Y heterozygotes and the mutationnegative group. 3. Among 24 C282Y heterozygotes only one individual with rectal carcmoma was detected compared with two cases with rectal- and seven with colonic malignancies in 153 mutation-negative individuals. The single female C282Y homozygote identified suffered from both rectal and colon carcinoma and died approximately 6 months ago as a consequence of her colon malignancy. 4. Serum ferritin appears to be a highly unreliable parameter of iron status, particularly in the elderly where a variety of factors that may influence the levels are often present in elderly individuals. This may be due to ageing alone or as a result of multiple comorbidities. 5. Serum ferritin levels were lower than expected in elderly subjects with mutation C282Y and compound heterozygotes with both C282Y and H63D, which may be related to a variable penetrance of the HFE gene mutations. It is possible that variation in other genes exist that confer protection against iron-loading by gene-gene interaction. The probability that environmental factors (e.g. a low iron diet) are more important in this respect cannot be excluded, although this is considered less likely in the light of the fact that the same trend was observed in all mutation-positive elderly individuals. It is therefore highly likely that C282Y -positive subjects with significant iron loading have died before reaching their seventies, particularly since none of the males included in this study were homozygous or compound heterozygous for the mutations analysed. In conclusion, possession of a mutant HFE gene does not appear to confer a survival advantage in old age, neither does it seem that mutation carriers with significant ironloading are overlooked by the medical fraternity. Further investigations are warranted to shed more light on the contributions of gene-gene and gene-environment interaction in the clinical manifestation of Hll, and how these processes can be manipulated to prevent the symptoms of this largely underdiagnosed disease.
Analysis of the clinical utility of gene expression profiling in relation to conventional prognostic markers in South African patients with breast carcinoma
(Stellenbosch : Stellenbosch University, 2014-12) Grant, Kathleen Ann; Kotze, Maritha J.; Wright, Colleen A.; Apffelstaedt, Justus P.; Stellenbosch University. Faculty of Medicine and Health Sciences. Dept. of Pathology. Anatomical Pathology.
ENGLISH ABSTRACT: Breast cancer is a heterogeneous disease characterised by marked inter-individual variability in presentation, prognosis and clinical outcome. The recognition that morphological assessment has limited utility in stratifying patients into prognostic subgroups led to clinico-pathological classification of tumour biology, based on receptor expression using immunohistochemical (IHC) techniques. This standard is currently complemented by the development of gene expression profiling methodology that led to the identification of intrinsic molecular subtypes, reflecting tumour genetics as the true driver of biological activity in breast cancer. The study was based on the hypothesis that molecular classification of breast carcinomas integrated with established clinico-pathological risk factors will improve current diagnostic and risk management algorithms used in clinical decision-making. A pathology-supported genetic testing strategy was used to evaluate microarray-based gene profiling against diagnostic pathology techniques as the current standard. Clinico-pathological factors including age, number of positive axillary nodes, tumour size, grade, proliferation index and hormone receptor status was documented for 141 breast cancer patients (143 tumours) referred for microarray-based gene expression profiling between 2007 and 2014. Subsets of patients were selected from the database based on the inclusion criteria defined for three phases in which the study was performed, in order to determine 1) the percentage of patients stratified as having a low as opposed to high risk of distant recurrence using the 70-gene MammaPrint profile within the inclusion criteria, 2) correlation of HER2 status as determined by IHC and fluorescence in situ hybridisation (FISH) with microarray-based mRNA readout (TargetPrint), and 3) the relationship between hormone receptor determination as reported by standard IHC and molecular subtyping using the 80-gene BluePrint profile. Similar distribution patterns for MammaPrint low- and high-risk profiles were obtained irrespective of whether fresh tumour biopsies or formalin-fixed paraffin embedded (FFPE) tissue was used. During the first phase of the study, 60% of the 106 tumour specimens analysed with MammaPrint were classified as low-risk and 40% as high-risk using a newly-developed MammaPrint pre-screen algorithm (MPA) aimed at cost-saving. In the second phase of the study, performed in 102 breast tumours, discordant or equivocal HER2 results were found in four cases. Reflex testing confirmed the TargetPrint results in discordant cases, achieving 100% concordance regardless of whether fresh tumour or FFPE tissue was used for microarray analysis. For the third phase of the study 74 HER2-negative tumour samples were selected for comparative analysis. Statistically significant positive correlations were found between protein expression (IHC score) and mRNA (TargetPrint) levels for estrogen receptor (ER) (R=0.53, p<0.0001) as well as progesterone receptor (PR) (R=0.62, p<0.0001), while combined ER/PR tumour status was reported concordantly in 82.4% of these tumours. BluePrint was essential for interpretation of these results used in treatment decision-making. The MPA developed in South Africa in 2009 was validated in this study as an appropriate strategy to prevent chemotherapy overtreatment in patients with early-stage breast cancer. The use of microarray-based analysis proved to be a reliable ancillary method of assessing HER2 status in breast cancer patients. Risk reclassification based on the TargetPrint results helped to avoid unnecessary high treatment costs in false-positive cases, in addition to providing potentially life-saving treatment to those for whom it was indicated. While neither IHC nor TargetPrint estimation of intrinsic subtype correlated independently with the molecular subtype as indicated by BluePrint profiling, the ability to distinguish between basal-like and luminal tumours was enhanced when the combined protein and mRNA values was considered. Genomic profiling provided information over and above that obtained from routine clinico-pathological assessments. This finding supports the relevance of a pathology-supported genetic testing approach to breast cancer management, whereby advanced genomic testing is combined with existing clinico-pathological risk stratification methods for improved patient management.
Automated sputum screening using the BD FocalPointTM Slide Profiler : correlation with transbronchial and transthoracic needle aspirates in a high risk population
(Stellenbosch : Stellenbosch University, 2014-04) Neethling, Greta Sophie; Schubert, Pawel T.; Koegelenberg, C. F. N.; Diacon, A. H.; Wright, C. A.; Stellenbosch University. Faculty of Medicine and Health Sciences. Dept. of Pathology, Division of Anatomical Pathology.
ENGLISH ABSTRACT: Background: Sputum is a non-invasive, economic investigation whereby bronchogenic carcinoma can be identified. Manual cytological screening is labour intensive, time-consuming and requires a continuous high level of alertness. Automation has recently been successfully introduced in gynaecological cytology. Since sputum samples are similar to cervical smears, the question arises as to whether they are also suitable for automated screening. Objective: This study presented with various objectives: 1) To test automated sputum screening using the BD FocalPoint™ Slide Profiler (FP) and compare with manual sputum screening. 2) To determine the sensitivity and specificity of sputum in identification of bronchogenic carcinoma. 3) To ascertain if any clinical, radiological or bronchoscopy findings would be predictors for bronchogenic carcinoma. 4) To determine the significance of adequacy. Method: Sputum samples were collected prospectively from patients attending the Division of Pulmonology at Tygerberg hospital for a transbronchial fine needle aspiration biopsy (TBNA) or a transthoracic fine needle aspiration biopsy (TTNA) for the period from 2010 to 2012. A pre-bronchoscopy sputum was collected and submitted for processing. Stained slides were put through the FP for automated screening. After slides were qualified, sputum slides were put back in the routine screening pool. Correlation was done using the TBNA/TTNA result as the standard to evaluate the sputum results. Results: 108 sputum samples were included in this study. Of the 84.3% malignant (n=91) and 15.7% benign (n=17) cases confirmed with a diagnostic procedure, sputum cytology had a sensitivity of 38.5% (35/91 malignant cases), and a specificity of 100% (17/17 benign cases). Automated screening had a better sensitivity of 94.3% (33/35 positive sputum cases), while manual screening showed a sensitivity of 74.3% (26/35 positive sputum cases) when compared to the final sputum result. Individual parameters with a significant association with positive sputum included the presence of an endobronchial tumour, partial airway obstruction / stenosis, round mass, spiculated mass (negative association), loss of weight (negative association) and squamous cell carcinoma as the histological subtype. Adequacy was not as significant as hypothesised since 85.3% of true positive sputum, but also 65.5% of false negative sputum, had large numbers of alveolar macrophages present. Conclusion: Sputum cytology remains an important part of the screening programme for bronchogenic carcinoma in the public health sector of South Africa. Results confirm that sputum cytology is very specific, and automated screening improves sensitivity. Automated screening proved to be more time efficient, resulting in 83.1% reduction (p<0.0001) in the screening time spent per case by a cytotechnologist. Results confirm that the quantity of alveolar macrophages is not directly proprtional to pathology representation. Positive sputum results did however improve with sputum adequacy, but had no significant association. Recommendations from this study include adopting automated sputum screening.
A biobank to support HIV malignancy research for sub-Saharan Africa
(Health & Medical Publishing Group, 2016) Schneider, Johann Wilhelm; Sanderson, Micheline; Geiger, Dieter; Nokta, Mostafa; Silver, Sylvia
Sub-Saharan Africa has one of the highest incidences of infection with HIV globally, but more people in this region are living longer owing to increased access to antiretroviral therapy. However, along with increased care and treatment, this population is expected to have an increase in HIV-associated cancers, as is being seen in the USA and other developed countries. To support translational research in HIV-associated cancers, Stellenbosch University in Cape Town, South Africa, was funded to house the state-of-the-art AIDS and Cancer Specimen Resource Sub-Saharan Africa Regional Biorepository (SSA RBR) to proactively obtain, manage and process biospecimens and associated clinical data representing both AIDS-defining and non-AIDS-defining cancers for research. The SSA RBR furthermore functions as the biorepository for AIDS Malignancy Consortium sub-Saharan clinical trial activities in this region. Although the site had much experience with cryopreservation and storage of specimens, capacity building revolving around operations under International Society for Environmental and Biological Resources/National Cancer Institute best practices took place in such areas as custodianship v. ownership, data sharing and facilities management. The process from selection until launch took 14 months.
Cardiomyopathy in crowded rabbits : a preliminary report
(Health & Medical Publishing Group, 1973) Weber, H. W.; Van Der Walt, J. J.
Rabbits were crowded 4 to a cage for 2 wk, then released for 1 wk, crowded again for 1 wk, and so on. Of 44 rabbits subjected to intermittent crowding only 9 survived longer than 10 mth, 20 died during the 1st month and 15 died between the 2nd and 9th mth of the experiment. Autopsy findings were indicative of heart failure. Light microscopical sections of the myocardium showed myocytolysis, interstitial edema, and an increased amount of acid mucopolysaccharides in rabbits surviving for 2 wk and more. Some rabbits surviving longer than 2 wk had in addition coagulative myocardial necrosis. The accumulations of acid mucopolysaccharides apparently were unrelated to necrotic foci. Longtime survivors frequently showed myocardial fibrosis and endocardial fibro elastosis, as well as basophilic degeneration of myocardial fibers. The lesions observed were similar to those described in idiopathic endomyocardiopathy in Southern Africa. Therefore, the rabbit may be of some value for research in cardiomyopathies.
Chemotherapy naive breast cancer: a correlation study between BD Cytorich™ Red cell blocks and formalin- fixed paraffinembedded tissue blocks
(Stellenbosch : Stellenbosch University, 2019-04) Van Rooyen, Evelyn; Schubert, Pawel T.; Schneider, Johann; Stellenbosch University. Faculty of Medicine and Health Sciences. Dept. of Pathology. Anatomical Pathology.
Background: The ever increasing burden of breast cancer, the most common cancer among women, demands a diagnostic test that is rapid, reliable, informative and cost-effective; particularly in countries with limited financial and medical resources. FNAB cytology and cell block combination has gained worldwide utility and has been described to be accurate and reliable. Aim: Henceforth the aim of this study was to retrospectively correlate the expression of prognostication markers, namely, estrogen receptor (ER), progesterone receptor (PR) and human epidermal growth factor receptor 2 (HER2) performed on cell blocks (using the BD CytorichTM Red method) and formalin-fixed, paraffin-embedded cell blocks (FFPET) in chemotherapy naive breast carcinomas, by immunochemistry (immunocytochemistry and immunohistochemistry respectively) and to perform fluorescence in-situ hybridization (FISH) testing for over- expression of the HER2 gene. Methods: Between 2013 and 2016, 132 cases of primary breast carcinoma were identified that had both cytology (including Cytorich™ Red cell blocks) and histology specimens that were both chemotherapy naive. Immunostaining for ER, PR and HER2 was performed. The staining was scored according to the Allred scoring system for histology specimen and this system was slightly adapted for cytology specimens, which also took the American Society of Clinical Oncology (ASCO)/College of American Pathologists (CAP) guidelines into account. FISH for HER2 over-expression was performed. The grade of the carcinoma was also analysed on both the cytology specimens (using the Robertson’s grading system) and the histology specimens (using the Modified Elston & Ellis system). Results: ER and PR performed on cell blocks had good correlation with FFPET with 91% and 85% sensitivity, respectively. HER2 on cell blocks had an agreement of 88% with FFPET. 87.88% of cell blocks had more than a 100 tumour cells present on H&E sections and cytological grading had an agreement of 41.41% with histological grading. Conclusion: The cell block technique continues to play a vital role in the diagnosis of primary, recurrent and metastatic breast carcinoma, allowing assessment of prognosis and prediction of response to therapy.
Colonic perineurioma (benign fibroblastic polyp) : case report and review of the literature
(BioMed Central, 2018) Van Wyk, Abraham Christoffel; Van Zyl, Hennie; Rigby, Jonathan
Background: Colorectal perineuriomas are uncommon benign mucosal-based proliferations of mesenchymal cells that express perineurial markers, often associated with colonic crypts displaying a serrated/hyperplastic architecture. The vast majority of cases arise distal to the splenic flexure and have been described as sessile polyps. Using molecular analysis, BRAF mutations have been demonstrated in the serrated crypt epithelium. We report a new case of perineurioma presenting as a pedunculated polyp in the transverse colon, with prominent hemosiderin deposits in the uninvolved lamina propria that separated the perineurial proliferation from the surface epithelium, a previously unreported histological finding. By using immunohistochemistry, we demonstrated the presence of BRAF V600E mutated protein in the serrated crypt epithelium. In addition, a review of the literature on colorectal perineurioma is provided. Case presentation: A 5 mm pedunculated polyp was removed from the transverse colon of a 42 year old man who presented with epigastric pain, weight loss and rectal bleeding. A proliferation of uniform plump spindled cells expanded the lamina propria and separated serrated colonic crypts. The epithelial component closely resembled microvesicular hyperplastic polyp. Immunohistochemical stains for epithelial membrane antigen (EMA), glucose transporter 1 (GLUT1) and collagen IV were positive in the stromal proliferation. A mutation-specific monoclonal antibody directed against BRAF V600E showed positive cytoplasmic staining in the serrated crypt epithelium but not in the perineurial proliferation. Conspicuous hemosiderin deposition was seen in the inflamed lamina propria between the perineurial proliferation and the surface epithelium. Conclusion: Although the majority of colorectal perineuriomas occur in the sigmoid colon and rectum and are described as sessile polyps, colorectal perineurioma can present as a pedunculated polyp proximal to the splenic flexure as described in this case. Conspicuous hemosiderin deposition can be seen in the superficial lamina propria. BRAF mutations are limited to the serrated crypt epithelium.
Comparison of Xpert® Breast cancer STRAT4 assay and immunohistochemistry for the evaluation of breast cancer biomarkers in South African patients
(Stellenbosch : Stellenbosch University, 2020-12) Dube, Welile Vumile; De Jager, Louis; Kotze, Maritha J.; Stellenbosch University. Faculty of Medicine and Health Sciences. Dept. of Pathology: Anatomical Pathology.
Background: Breast cancer is one of the most common cancers diagnosed in women and approximately 60% of breast cancer related deaths are reported in low-and middle-income countries. Breast cancer is a highly heterogeneousdisease, and molecular subtyping is paramount foreffective treatment of patients. Therefore, it is important to validate new molecular methods for assessing cancer biomarkers for cost-effective use in resource-poor settings.Aim:Aretrospective study was performed to determine the concordance between aQuantitativeReverse TranscriptionPolymerase Chain Reaction(RT-qPCR) CE-IVD assay (Xpert® Breast Cancer STRAT4*) and the current gold standard methods of immunohistochemistry (IHC) and fluorescence in situ hybridization (FISH) for determining estrogenreceptor (ER),progesterone receptor(PR), human epidermal growth factor receptor 2 (HER2)andproliferation index(KI-67)expression in breast carcinomas.Method: One hundred and one cases of breast carcinoma were retrieved from the archives of the Division of Anatomical Pathology, Tygerberg Academic Hospital. The original stained slides were reviewed and IHC expression of ER, PR, HER2 and KI-67 scored. Three-micron sections were cut from formalin-fixed paraffin embedded (FFPE) tissue blocks and processed according to the instructions of the manufacturer. The assay was run on the resultant lysates.Results:The overall percentageagreement between the Xpert® STRAT4 assay and IHC / FISH results were 85.15% for ESR, 89.90% for PGR,91.09% for ERBB2, 90.72% for MKI67 (when using a cut off of 10%) and 84.54% for MKI67 (when using a cut-off of 20%). The positive percentage agreement for ESR, PGR, ERBB2, MKI67 with 10% cut-off andMKI67 with 20% cut-offwere 82.76%, 94.64%, 68.97%, 91.30% and 96.05%, respectively, and the negative percentage agreement were 100%, 84.09%, 91.67%, 80.00% and 42.86%, respectively. Conclusion:The studyhas shownthattheXpert® BreastCancer STRAT4 assay shows good concordance with IHC and FISH in detecting breast cancer biomarkers, and may become a supplementary or alternativestandard of care aftervalidation studies are performed.
Complex patterns of genomic admixture within Southern Africa
(PLOS, 2013-03) Petersen, Desiree C.; Libiger, Ondrej; Tindall, Elizabeth A.; Hardie, Rae-Anne; Hannick, Linda I.; Glashoff, Richard H.; Mukerji, Mitali; Indian Genome Variation Consortium; Fernandez, Pedro; Haacke, Wilfrid; Schork, Nicholas J.; Hayes, Vanessa M.
Within-population genetic diversity is greatest within Africa, while between-population genetic diversity is directly proportional to geographic distance. The most divergent contemporary human populations include the click-speaking forager peoples of southern Africa, broadly defined as Khoesan. Both intra- (Bantu expansion) and inter-continental migration (European-driven colonization) have resulted in complex patterns of admixture between ancient geographically isolated Khoesan and more recently diverged populations. Using gender-specific analysis and almost 1 million autosomal markers, we determine the significance of estimated ancestral contributions that have shaped five contemporary southern African populations in a cohort of 103 individuals. Limited by lack of available data for homogenous Khoesan representation, we identify the Ju/’hoan (n = 19) as a distinct early diverging human lineage with little to no significant non- Khoesan contribution. In contrast to the Ju/’hoan, we identify ancient signatures of Khoesan and Bantu unions resulting in significant Khoesan- and Bantu-derived contributions to the Southern Bantu amaXhosa (n = 15) and Khoesan !Xun (n = 14), respectively. Our data further suggests that contemporary !Xun represent distinct Khoesan prehistories. Khoesan assimilation with European settlement at the most southern tip of Africa resulted in significant ancestral Khoesan contributions to the Coloured (n = 25) and Baster (n = 30) populations. The latter populations were further impacted by 170 years of East Indian slave trade and intra-continental migrations resulting in a complex pattern of genetic variation (admixture). The populations of southern Africa provide a unique opportunity to investigate the genomic variability from some of the oldest human lineages to the implications of complex admixture patterns including ancient and recently diverged human lineages.
Congenital tuberculosis : a report of a probable case
(Health & Medical Publishing Group, 1978) Le Roux, F. B.; Schwersenski , J.; Greeff, M. J.
Extensive miliary pulmonary tuberculosis in a 17-day-old infant born to a mother with pulmonary tuberculosis is described. It was assumed that the patient probably had congenital tuberculosis. We report our findings to draw attention to a rare disease where a high index of suspicion, timely diagnosis and adequate management are essential if treatment is to be successful.
The contribution of fine needle aspiration biopsy in the diagnosis of Mycobacterial Lymphadenopathy with particular reference to children
(Stellenbosch : University of Stellenbosch, 2009-12) Wright, Colleen Anne; Warren, R. M.; Marais, B. J.; University of Stellenbosch. Faculty of Health Sciences. Dept. of Pathology. Anatomical Pathology.
ENGLISH ABSTRACT: Expediting a diagnosis of tuberculosis in children, particularly those who are immunocompromised due to HIV/AIDS, is essential, as they are vulnerable to develop severe forms of disease due to their immature or compromised immune systems. A significant percentage of children (8 to 10%) with TB have TB lymphadenitis, in isolation, or in combination with other disease manifestations. Fine needle aspiration biopsy (FNAB) is a simple and minimally invasive procedure well tolerated by children. It may be performed as an outpatient procedure by clinicians as well as nurses, and excellent results can be achieved with training in the correct procedure. The aim of this dissertation was to demonstrate that FNAB may contribute significantly to the diagnosis of mycobacterial lymphadenitis, with particular reference to children TB suspects. We first established that TB lymphadenitis is a common clinical problem in children in TB endemic areas and that FNAB is an efficient simple and effective diagnostic modality in children with peripheral lymphadenopathy. We then proceeded to document the diagnostic yield and time to diagnosis of FNAB compared to conventional laboratory specimens collected in children. We investigated the value of additional diagnostic modalities such as autofluorescence in improving the ability of cytology to make a definitive diagnosis of mycobacterial infection based on cytomorphology and identification of the organism. In countries where organisms such as Mycobacterium bovis BCG and nontuberculous mycobacteria are prevalent, culture with subsequent speciation is essential. The amount of material harvested during FNAB is minuscule, and requires immediate bedside inoculation for optimal yields. We developed an inexpensive and effective transport medium to facilitate mycobacterial culture from FNAB, even if this is collected at an outside facility. It is ideally suited for use in clinics and rural hospitals as it is stable at room temperature, maintains viability of the organism for seven days, and the closed lid format reduces contamination. Mycobacterial culture even using liquid-based media, takes up to 6 weeks, and this delay is unacceptable particularly in children. We developed a Nucleic Acid Amplification Technique (NAAT) using High Resolution Melt Analysis and applied this novel technique to FNAB specimens submitted in transport medium. Although sensitivity remained suboptimal, the technique is highly specific, simple and rapid. Its use could be incorporated into routine microbiology laboratories, to assist with rapid diagnosis while cultures are pending. We collected a solid body of evidence, which will promote the use of FNAB in suspected mycobacterial lymphadenopathy, particularly in children in resource-limited countries. The utilisation of the diagnostic methods identified will expedite speciation and allow early and appropriate initiation of therapy. This is in keeping with Millennium Development Goal 6: to combat TB by early detection of new cases and effective treatment.
The contribution of the placenta to the diagnosis of congenital tuberculosis
(Stellenbosch : Stellenbosch University, 2014-04) Rabie, Ursula; Wright, Colleen Anne; Warren, Robin Mark; Hoek, Kim Gilberte Pauline; Bekker, Adrie; Schubert, Pawel T.; Stellenbosch University. Faculty of Medicine and Health Sciences. Dept. of Pathology.
ENGLISH ABSTRACT: The aim of this pilot project was to determine whether mothers with laboratory confirmed or clinically suspected tuberculosis (TB) had evidence of TB in the placenta. A secondary objective was to correlate evidence of placental TB with neonatal outcome. A total of 56 placentas were examined to determine if there were any specific histopathological features predictive of tuberculosis together with Ziehl-Neelsen (ZN) staining. A total of 30 cases were positive for maternal TB and one case was a false positive maternal diagnosis of TB, whilst 25 cases were negative for maternal TB. Biopsies from these 56 placentas were collected for conventional PCR from the paraffin embedded tissue blocks. The performance of these two diagnostic modalities (histopathology and PCR) was assessed coll ectively and individually, and compared to the neonatal outcome (presence or absence of active clinical mycobacterial tuberculosis infection) and evidence of maternal pulmonary and extra pulmonary tuberculosis. The recognition of specific sites of lesions in the placenta (e.g. membranes vs. intervillous space) may lead to an understanding of the pathogenic mechanisms involved in matern alfetal transmission of tuberculosis, and thereby pave the way for further studies in understanding the pathogenesis of congenital TB. Invaluable knowledge was obtained in the diagnoses of M.tuberculosis in the placenta as it was found that micro abscesses and intervillositis were strong indicators of TB infection in the placenta, however, ZN staining still remains the gold standard for diagnosing M.tuberculosis infection in the placenta. PCR is found to have limitations, because only M.tuberculosis DNA is amplified and does not distinguish live from dead bacteria. The conclusion reached is that PCR is of limited value in the diagnosis of active M.tuberculosis infection in the placenta using FFPE tissue, while certain histological changes may be indicative of such infection; however confirmation of the organism by ZN staining is still essential.
Correlating p16INK4a /Ki-67 co-expression and gene methylation with HIV infection and high-risk HPV in abnormal cervical squamous intraepithelial cells
(Stellenbosch : Stellenbosch University, 2023-03) Louw, Meagan; Sanderson-November, Micheline; Neethling, Greta; Stellenbosch University. Faculty of Medicine and Health Sciences. Dept. of Pathology: Anatomical Pathology.
ENGLISH SUMMARY: Globally, cervical cancer is the fourth most common cancer amongst women. Persistent infection with high-risk human papillomavirus (HPV) is shown as the causal factor in cervical cancer development. Women living with HIV is six times more prone to cervical cancer development. The aim of this study was to identify the HR-HPV types, investigate the simultaneous expression of p16INK4a and Ki-67 and evaluate the methylation status of CADM1, MAL and miR124-2 genes in cytology samples from HIV-positive and HIV-negative women with LSIL, HSIL, ASC-US, and ASC-H Pap smear results. Study participants were women between the ages of 21 years to 60 years referred to the Colposcopy clinic at Tygerberg Academic Hospital. Exfoliated cervical intraepithelial cells were collected in Surepath medium and HR-HPV types were determined using the Hybrispot HPV direct flow chip kit, whereas the co- expression of p16INK4a/Ki-67 proteins was evaluated with the CINtec® Plus cytology immunocytochemistry kit. For methylation assays, DNA was isolated from the left-over exfoliated cells followed by the assessment of quantity and purity of isolated DNA using fluorometry and spectrophotometry, respectively. Isolated DNA was bisulfite converted and the methylation assays for the CADM 1, MAL and miR-124-2 genes were done using the respective EpiMelt assays. HPV DNA detection results were associated with cytological diagnosis as well as HIV status. In our study group 74 % (51/69; 95% CI: 62,1%-83%) of woman tested positive for HPV of which 70.6% (36/51) were of WLWH and 29.4% (15/51) of HIV negative women. Our results showed that p16INK4a /Ki-67 dual-staining was detected in 43.8% (25/57) of the samples with the HSIL cytology showing the highest p16INK4a /Ki-67 co-expression rate of 64% (16/25) compared to the other cytology groups. The proportion of the LSIL group with p16INK4a /Ki- 67 dual staining was 33,3% (4/12), whereas that of the ASC-US and ASC-H groups were 25% (2/8) and 23% (3/13) ASC-H, respectively. CADM1 methylation was detected in 12.3% (7/57) of samples, while MAL and the miR-124-2 genes showed methylation in 14% (8/57) and 12.3% (7/57)of the samples, respectively. HPV infection was detected in 28.1% (16/57) of the samples with methylated CADM1, MAL and miR-124-2 genes. A significant relationship was found between HR-HPV and miR-124-2 methylation. The logistic regression model analysis employing predictors such HR-HPV, p16INK4a and Ki- 67 co-expression, as well as the methylation status of the CADM1, MAL, and miR-124-2 genes, for LSIL cytology showed low sensitivity and high specificity, contrasting to that of the HSIL model with high sensitivity and low specificity. Therefore, we conclude that a larger study is warranted, with removing or adding predictors for model improvement.
CYP2D6 genotyping and use of antidepressants in breast cancer patients : test development for clinical application
(Springer Verlag, 2012-05) Van der Merwe, Nicole; Bouwens, Christianne S. H.; Pienaar, Rika; Van der Merwe, Lize; Yako, Yandiswa Y.; Geiger, Dieter H.; Kotze, Maritha J.
Approximately 25 % of clinically important drugs and numerous environmental carcinogens are metabolised by CYP2D6. Variation in the CYP2D6 gene and concomitant use of tamoxifen (TAM) with certain antidepressants may increase recurrence risk in breast cancer patients due to reduced enzyme activity. In this study we determined the appropriateness of adding CYP2D6 genotyping to the breast cancer genetic testing options already available in South Africa, which include BRCA mutation screening and transcriptional profiling to assess estrogen receptor (ER) status. A total of 114 South African breast cancer patients, including 52 Caucasian and 62 Coloured (Mixed ancestry), and 63 Caucasian control individuals were genotyped for the most common inactivating allele (CYP2D6*4, rs3892097) previously identified in the CYP2D6 gene. In the initial validation data set consisting of 25 Caucasian and 62 Coloured patients, the CYP2D6*4 allele frequency was significantly higher in Caucasian compared to Coloured patients (24 % vs. 3 %, p≤0.001), similar to previous findings in the general South African population. Extended CYP2D6 genotyping was subsequently performed in an implementation data set of 27 Caucasian breast cancer patients, to determine the prevalence of depression and use of antidepressants in a clinical setting. A medical history of depression and/or use of antidepressants was reported in 37 % (10/27) of these breast cancer patients genotyped for CYP2D6*4. This translational research study has led to increased awareness among clinicians of the potential benefits of CYP2D6 genotyping to facilitate prevention of cumulative risk in a high-risk genetic subgroup of breast cancer patients considered for concomitant treatment of TAM and antidepressants that may reduce enzyme function. © Springer Science+Business Media, LLC 2012.
Cytokines and tuberculosis : an investigation of tuberculous lung tissue and a comparison with sarcoidosis
(University of Stellenbosch. Faculty of Health Sciences. Dept. of Biomedical Sciences., 2005-12) Bezuidenhout, Juanita; Walzl, Gerhard; University of Stellenbosch. Faculty of Health Sciences. Dept. of Pathology. Anatomical Pathology.
The formation of granulomas at the site of antigen presentation in both tuberculosis and sarcoidosis is an essential component of host immunity for controlling inflammation. Granuloma formation is a complex process that also requires recruitment and activation of lymphocytes and macrophages to the site of infection and arrangement into a granuloma. It is dependant on the activation of especially IFNγ secreting CD4+ T cells, resulting in a Th1 profile. However, it is suggested that a persistently high IFNγ is responsible for the damage caused by granulomatous disease and that moderating cytokines, resulting in a Th0 profile, are necessary to down-regulate the IFNγ response to more appropriate levels later in the disease process, after the antigen has been effectively contained. I propose that: “Cytokine profiles determine clinical and histopathological phenotypes of disease. This thesis tests the hypothesis that it will be reflected by cytokine expression profiles in granulomas in different forms of tuberculosis and in sarcoidosis.” To examine this, biopsy tissue was obtained from patients with pulmonary cavitary tuberculosis, pleural tuberculosis in HIV sero-negative and sero-positive patients, and sarcoidosis. The diagnosis of tuberculosis or sarcoidosis was confirmed, granulomas were characterised as necrotic or non-necrotic, sarcoidosis cases were graded histologically and in situ hybridisation was performed for IL-12-, IFNγ-, TNFα- and IL-4-mRNA. In all patients with pleural tuberculosis, a Th0 profile was noted, while necrotic granulomas were more evident in HIV positive than HIV negative patients. There was a clear association between TNFα and necrosis in tuberculous granulomas that may be ascribed to the increased apoptotic activity of TNFα. An increase in IFNγ correlated with an increase in necrosis, supporting the theory that high IFNγ levels later in disease is detrimental. This effect may be enhanced by a strong presence of TNFα positive cells. An increase in both Th1 and Th2 cytokine mRNA in HIV positive patients supports the theory that an overproduction of cytokines may be a mechanism to compensate for the failure of another immune effector mechanism. Findings in pulmonary tuberculosis were similar to those in pleural tuberculosis. In all sarcoidosis cases the presence of a very strong Th1 and TNFα, but no Th0 response was confirmed. None of the differences in either the histological grading, or the clinical outcome of patients were reflected in the cytokine profile. It is possible that this profile does not reflect the histological grade of disease or that it may reflect various stages of disease. These findings support the theory that a strong Th1 presence later in disease, in conjunction with TNFα may induce fibrosis, as most of these cases showed signs of at least focal fibrosis. Numerous aspects, including a T helper response are involved in granulomatous inflammation. The earlier dogma of good, beneficial (Th1) versus evil, detrimental (Th2), is an oversimplification of a very complex process. It is clear that the effect of a cytokine depends at least partially on the stage of disease. The balance between the various cytokines, and the levels of these cytokines contribute to their role in resolution or disease progression. An early, pure Th1 response may be beneficial if effectively clearing the granuloma-inducing antigen. At this stage, a Th2 presence will be harmful as clearing of the antigen will not be as effective. In chronic disease where failure to remove the antigen results in progression of granulomas with subsequent necrosis and/or fibrosis, a proinflammatory Th1 response may be detrimental and minimising of this effect is needed. An overly strong presence of the various cytokines may also be detrimental, while lower levels will be beneficial.
Determining the suitability of bio-specimens obtained by fine needle aspiration biopsy at a tertiary hospital in Malawi for immunocytochemical assessment
(Stellenbosch : Stellenbosch University, 2022-04) Mulenga, Maurice; Schneider, Johann Werner; Stellenbosch University. Faculty of Medicine and Health Sciences. Dept. of Pathology.
ENGLISH SUMMARY: Fine-needle aspiration biopsy (FNAB) is a quick, economical, least invasive and easy to perform a minor surgical procedure. In resource-limited settings, FNAB is of utmost importance in providing a rapid diagnosis that facilitates timely and correct institution of treatment. The FNAB smear preparation provides an opportunity for either rapid on-site evaluation or routine diagnosis if ancillary tests are necessary to establish a specific diagnosis. Cell blocks (CB) prepared from FNAB specimens improve the diagnostic yield, increase the sensitivity and reduce false-positive interpretations of detecting a malignant neoplasm. In addition, CB allow for additional morphological evaluation with a better architectural pattern, enable the performance of numerous ancillary diagnostic studies, including immunocytochemistry and molecular studies and offer the storage of material that can be used for future research studies. Delays in fixing the cell block have been challenges in various cell block preparatory techniques. However, a special alcohol-based fixative, commercially available solution called CytoRich Red® (CRR) has been described to be comparative to liquid-based cytology due to its effectiveness in lysing red blood cells, reducing background material, and improving staining qualities of the nucleus and cytoplasm in routine preparations of non-gynaecological material in suspension or fluids. Despite this breakthrough, there is a paucity of data on the suitability of CRR cell blocks for immunocytochemical and DNA assessment from FNAB material obtained from solid tumours. This study aimed to establish and confirm the suitability of CytoRich Red® Cell Blocks and FNAB biospecimens obtained and prepared at Kamuzu Central Hospital, Lilongwe, Malawi, for cytomorphological and immunocytochemical assessment. This study analysed 144 cell blocks and 128 FNAB smears. It is one of the first within sub-Saharan Africa to describe diagnostic efficacy from FNAB specimens obtained from various superficial and deep masses fixed in CRR. It describes the advantage of using an alcohol-based fixative immediately to reduce pre-fixation time lag. This study showed that CRR-fixed cell blocks improve sensitivity and architectural preservation, and immunocytochemical staining characteristics of the aspirate compared to routine FNAB smears. It is envisioned that CRR-fixed cell blocks will be a source of extractable, stable and usable DNA that supports research in biorepositories and biobanks.
Development and application of a pathology supported pharmacogenetic test for improved clinical management of South African patients with breast cancer and associated co-morbidities
(Stellenbosch : Stellenbosch University, 2016-03) Van der Merwe, Nicole; Kotze, Maritha J.; Pienaar, Fredrieka; Janse van Rensburg, Susan; Bezuidenhout, Juanita; Stellenbosch University. Faculty of Medicine and Health Sciences. Dept. of Pathology. Anatomical Pathology.
ENGLISH ABSTRACT: Three major challenges in the field of breast cancer have been identified as research priorities for this study. The first is the need to combine genetic testing of high-risk patients with familial breast cancer with pharmacogenetics to reduce recurrence risk in cancer survivors due to drug failure as a consequence of anti-cancer treatment that does not match the patient’s genotype. The second is the delineation of key pathways through which genes implicated in breast cancer and associated co-morbidities can serve as nutritional and drug targets across diagnostic boundaries. The third is the discovery of genetic alterations underlying familial breast cancer not attributed to mutations in the two major tumour suppressor genes, BRCA1 and BRCA2. The study population consisted of 164 breast cancer patients (60 Coloured/Mixed Ancestry and 104 Caucasian), of whom 88 patients were selected from a total of 813 individuals who provided informed consent for inclusion of their data in a genomics database resource generated at the interface between the laboratory and routine clinical practice. In addition, DNA samples of 101 cancer-free individuals above the age of 65 years were available for clinical validation of potentially causative variants in an extended female control group. In the first phase of this study, real-time polymerase chain reaction (PCR) TaqMan© technology was used to confirm the potential value of adding pharmacogenetic testing (CYP2D6 allele 4) to standard immunohistochemistry (IHC)-based breast tumour subtyping complemented by BRCA mutation screening and/or microarray gene profiling in eligible patients. In phase two of the study, common genetic risk factors for cardiovascular disease (CVD) were shown to be significantly associated with earlier age (10 years on average) of breast cancer onset/diagnosis (APOE E4 allele p=0.003; 95% CI: 4-15) and body mass index (BMI) (MTHFR 1298 A>C; p=0.01; 95% CI: 3-14) in patients stratified according to estrogen receptor (ER) status, after adjustment for potential confounders. Age at diagnosis/onset of breast cancer was significantly lower in patients with ER-negative versus ER-positive tumours, after adjustment for ethnicity (p=0.022), while BMI was significantly higher in patiens with ER-positive compared to ERnegative tumours after adjustment for age, ethnicity, and family history of cancer (p=0.035). These findings contributed to the development of an exome pre-screening algorithm (EPA) used in part 3 of this study to select three genetically uncharacterized breast cancer patients for whole exome sequencing (WES) performed in comparison with three ethnically concordant cancer-free controls. WES followed by variant calling using both the standard human genome reference sequence (hg19) and an ethnically concordant major allele reference genome (MARS) revealed a more than 20% discrepancy in the number of gene variants identified in the same samples. After exclusion of a large number of false-positives caused by minor alleles in hg19, two rare missense mutations (<1%) were identified in a family with ER-positive breast cancer: RAD50 R385C and MUC1 Q67E. Three different bioinformatics tools were used to predict functionality and both mutations were confirmed by Sanger sequencing and/or real-time PCR in the Pathology Research Facility (PRF) laboratory. Neither the RAD50 nor the MUC1 missense mutation were identified in the exomes of an unrelated breast cancer patient with triple-negative breast cancer or three population-matched control individuals. This study led to the development of a pathology-supported genetic testing framework for WES beyond the limitations of single-gene BRCA mutation screening in South African breast cancer patients. Our findings support previous WES results indicating that the majority of genetically uncharacterised familial breast cancer may be caused by a combination of low-moderate penetrance mutations exerting their effect in a high-risk environment reflected by high BMI. WES enables identification of genetic risk factors of relevance to both cancer development and tailored therapeutic intervention in a single genetic test.
Development of a novel pre-screen algorithm for cardio-metabolic risk management using a genomics database resource
(Stellenbosch : Stellenbosch University, 2016-03) Luckhoff, Hilmar Klaus; Kotze, Maritha J.; Janse van Rensburg, Susan; Stellenbosch University. Faculty of Medicine and Health Sciences. Dept. of Pathology. Anatomical Pathology.
ENGLISH ABSTRACT: The timely assessment and treatment of dyslipidaemia is an important component of cardiovascular risk screening and intervention. The apolipoprotein E (APOE) ε-2/ε-3/ε-4 polymorphism associated with impaired lipid homeostasis provides a genetic link between cardiovascular disease (CVD) and late-onset Alzheimer’s disease (AD). Realization that the phenotypic expression of the risk associated APOEε-2 and ε-4 alleles may be dependent on non-genetic factors supports the inclusion of APOE genotyping in chronic disease screening programs. The lack of well-defined selection criteria for APOE genotyping, however, limits the use of this biomarker in clinical practice. The aim of the present study was to develop a pre-screen algorithm for identification of a target population most likely to benefit from APOE genotyping, performed in conjunction with a clinical and lifestyle assessment. Towards this goal, comprehensive patient data were evaluated from a total of 580 unrelated Caucasians enrolled in a chronic disease screening program over a five-year period (2010-2015), using an ethically approved study questionnaire. Biochemical tests performed according to standard laboratory protocols were extracted from the research database. All study participants were genotyped for the APOE ε- 2/ε-3/ε-4 polymorphism. APOE genotype distribution differed significantly (p<0.05) between study participants with and without a family history of AD. A positive association between dietary fat intake and lowdensity lipoprotein (LDL) cholesterol (p=0.001), as well as an inverse association with highdensity lipoprotein (HDL) cholesterol (p=0.002), were observed in patients with a family history of AD. Body mass index (BMI) was positively associated with LDL cholesterol and inversely associated with HDL cholesterol levels (p<0.001), irrespectively of an AD family history. Smoking was associated with higher triglycerides (p<0.001) and lower HDL cholesterol levels (p=0.004) in the total study group. Alcohol intake was positively associated with BMI (p=0.008) as well as triglyceride levels (p=0.021) in patients with a positive family history of AD. The clinical expression of a hypercholesterolaemic phenotype in APOE ε-4 allele carriers, as well as apparent mitigation by regular physical activity, were dependent on the interaction between a family history of AD and APOE genotype (p<0.001). APOE ε-2 carriers without an AD family history showed a significant increase in triglyceride levels (p=0.014). The modulating influence of APOE ε-4 on the relationship between alcohol intake and BMI as well as total cholesterol levels was also dependent on the presence or absence of AD family history (p<0.05). This study resulted in the addition of a family history of AD as a novel component to the prescreen algorithm developed for selection of at-risk individuals prior to APOE genotyping performed as part of a chronic disease screening program. The lifestyle questionnaire used in this study furthermore facilitated interpretation of the clinical relevance of variation detected in the APOE gene. This is important to prioritize the use of lipid-lowering medication towards patients with severe subtypes of dyslipidaemia such as familial hypercholesterolaemia (FH), which remains largely undiagnosed and untreated in the highrisk South African population. Incorporating the research findings into clinical practice would suggest that physical activity may be the most effective risk reduction strategy in carriers of the APOEε-4 allele, as supported by international studies.
The development of malignancies in renal allograft recipients with special emphasis on Kaposi's sarcoma
(Stellenbosch : Stellenbosch University, 2002-03) Moosa, M. R.; Du Toit, D. F.; Wranz, P. A. B.; Stellenbosch University. Faculty of Medicine and Health Sciences. Dept. of Pathology. Anatomical Pathology.
ENGLISH ABSTRACT: Renal transplantation is undoubtedly the best treatment for patients with irreversible renal failure. As a prelude to establishing the nature of malignancies in renal transplant patients we sought to determine factors influencing the outcome of renal transplantation. The survival of renal allografts and of recipients is influenced by a number of demographic, clinical and therapeutic factors. Some of these factors have been better studied than others, and we sought to establish the influence of particular factors on our own patients and allografts. The total number and nature of malignancies developing in these patients subsequent to transplantation was also established. All patients transplanted in our unit between April 1, 1976 and March 31, 1999 were included in the study. In the study period, 542 patients received 623 renal allografts. Demographic details were analysed. Patient and graft outcomes were assessed using Kaplan-Meier survival analysis. The survival curves were compared using univariate analysis; results that were significant were subjected to multivariate analysis. The influence of a number of factors on graft and patient survival were assessed and compared. The impact of a variety of variables on the number and behaviour of malignancies was also established. Patient and graft survival were superior in recipients who were aged less than 40 years; cyclosporine improved graft survival but not patient survival. Early graft loss was associated with a high patient mortality rate. Contrary to the experience elsewhere, black and white patients had similar outcomes after renal transplantation. Of the 542 recipients 41(8.1%) developed malignancies with Kaposi's sarcoma occurring, in 21 patients and skin cancers in 13 patients. The relative risk for the Kaposi's sarcoma development was 235. Kaposi's sarcoma was the most common tumour in non-white patients (accounting for 79% of malignancies in this group) and occurred less than 2 years after transplantation. Kaposi's sarcoma was equally common in male and female recipients. Under cyclosporine the latent period to malignancies was reduced but the frequency remained unaffected. Kaposi's sarcoma skin lesions were present in all the affected patients, with the lower limbs the most common site of involvement. Kaposi's sarcoma responded to reduction of immunosuppression without the need for complete discontinuation, and with preservation of renal function. Extracutaneous involvement occurred in over one quarter of the patients and invariably proved fatal in all patients with visceral organ involvement. The histopathology of posttransplant Kaposi's sarcoma was the same as that described in the other epidemiological forms of the disease. White male recipients were at the greatest risk of developing skin cancers after renal transplantation. Squamous cell carcinomas were relatively more common and were found in sun-exposed areas. The lesions were treated only by local excision and none metastasized. Malignant lymphoma, breast cancer and lung cancer occurred in individual patients but the relative risk of all these lesions were close to unity. Patients with preexisting cancers did not develop recurrences following transplantation. SECTION 2 Both immunosuppression and immunostimulation are thought to play a role in the development of Kaposi's sarcoma after renal transplantation. We investigated the quantitative and qualitative aspects of the immune system of patients who had developed Kaposi's sarcoma. The lymphocyte phenotypes were established using flow cytometry while transformation studies were performed using mitogens. Pokeweed was used as the B-cell mitogen, and concanavalin A and phytahaemagglutinin were the T-cell mitogens. Cell mediated immunity was also tested using delayed type hypersensitivity skin tests and the serum immunoglobulin levels were estimated. Firstly, with regard to humoral immunity, 2/3 of the patients had normal serum immunoglobulin levels, although the B-cell count was reduced in all the patients on immunosuppression. B-cell transformation tests with pokeweed mitogen revealed that B-cell function was not impaired in patients with Kaposi's sarcoma. The patients with decreased immunoglobulin levels also appeared to be malnourished as evidenced by low albumin levels. Secondly, CD3 and CD4, but not CD8, cell counts were reduced in patients with Kaposi's sarcoma. The transformation analyses revealed significant differences compared to controls, with reduced responses in patients with Kaposi's sarcoma. Thirdly, natural killer (NK) cell numbers were also reduced in patients with Kaposi's sarcoma. There were no significant differences in delayed type hypersensitivity skin reactions that could not be accounted for by racial differences. Cellular immunity is impaired in patients with Kaposi's sarcoma with a reduction in the number of NK cells. Both of these components of the immune system are important in protection against malignant transformation. SECTION 3 Kaposi's sarcoma is an important complication of renal transplantation. If the human herpesvirus 8 (HHV-8) causes Kaposi's sarcoma, the virus should be present in all Kaposi's sarcoma lesions and be drastically reduced or cleared from involved tissue on remission of the Kaposi's sarcoma. Fourteen renal transplant patients with cutaneous Kaposi's sarcoma, including autopsy material from two cases, were investigated for the presence of HHV-8. A second skin biopsy was taken from 11 survivors, after remission of Kaposi's sarcoma, from normal skin in the same anatomical region as the first biopsy. Remission was induced by reduction or cessation of immunosuppression. A peripheral blood sample was collected simultaneously with the repeat biopsy. A nested polymerase chain reaction assay was used to detect HHV-8 DNA in the biopsy tissue and peripheral blood mononuclear cells followed by direct sequencing of polymerase chain reaction product to detect any nucleotide changes. HHV-8 DNA was detected in all the cutaneous Kaposi's sarcoma and all the visceral Kaposi's sarcoma samples, as well as a number of Kaposi's sarcoma-free organs including the thyroid, salivary gland, and myocardium that have not been described before. Mutations in the viral DNA could be demonstrated in all patients. The mutations found were related more to that seen in AIDS-Kaposi's sarcoma cases than that found in African endemic Kaposi's sarcoma cases. HHV-8 sequences could be detected in follow-up frozen skin biopsies of five patients but were negative in the equivalent formalin-fixed specimens. Viral DNA was also detected in 2 of 11 peripheral blood mononuclear cell samples collected at the time of the follow-up skin biopsies. Reduction or withdrawal of immunosuppression allows the immune system to recover sufficiently to reduce viral replication with subsequent viral persistence and low-grade viral replication that coincides with clinical remission of the Kaposi's sarcoma lesions. This provides further evidence for the important etiological role played by HHV-8 in the pathogenesis of posttransplant Kaposi's sarcoma. SECTION 4 The recently discovered HHV-8 is an important factor in the aetiopathogenesis of Kaposi’s sarcoma. The reason for the exceptionally high prevalence of Kaposi's sarcoma in our area, as well as that of other developing countries, remains unexplained. We investigated the seroprevalence of the virus in the different healthy subjects as well as organ donor-recipient pairs. All recipients were tested at the time of transplantation, as were the paired donors. Control subjects tested were healthy blood donors, Renal Unit staff, and household contacts of patients with Kaposi's sarcoma. An enzyme-linked immunoassay (ELISA) to the whole virus was used for screening and all positives were confirmed using ELISA to the latent ORF 73 antigen. The prevalence of HHV-8 was similar in all groups and averaged less than 6%. After transplantation the seroprevalence increased to almost 20% but neither the transplanted kidney nor blood transfused perioperatively could account for the increase. Kaposi's sarcoma developed in 3 of the 116 patients transplanted. All patients with Kaposi's sarcoma were proven to be HHV-8 seropositive before the development of the disease. Two of the patients who developed Kaposi's sarcoma were seropositive before transplantation. No patient who received a graft from a seropositive donor developed Kaposi's sarcoma. We refute the notion that a high prevalence of HHV-8 in the general population is responsible for the high prevalence of Kaposi's sarcoma in our population or that the donor organ is a major source of infection in renal transplant recipients. Reactivation, rather than primary infection appears to be the source of the virus after renal transplantation.
Diagnostic yield of fine needle aspiration biopsy in HIV-infected adults with suspected mycobacterial lymphadenitis
(Health & Medical Publishing Group, 2014-01) Razack, R.; Louw, M.; Wright, C. A.
Background: Fine needle aspiration biopsy (FNAB) has been shown to be the diagnostic procedure of choice for superficial lymphadenitis in tuberculosis endemic regions. Methods: We conducted a retrospective laboratory-based study to determine the bacteriological yield of clinically suspected mycobacterial tuberculous lymphadenitis following FNAB in adults, and specifically HIV-positive patients, to determine the need for the introduction of automated nucleic acid amplification tests (NAATs) such as the Xpert MTB/RIF assay as the initial diagnostic modality. Results: A diagnostic yield of 80% was achieved, significantly higher in HIV-positive v. HIV-negative patients (84% v. 52%, respectively; p<0.001). Conclusion: The results justify using automated NAATs such as the Xpert MTB/RIF assay as the initial diagnostic modality to expedite management in HIV-infected patients.

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