Statement on analysis and interpretation of clinical human gastrointestinal microbiome testing using next-generation sequencing in South Africa.

Advances in DNA sequencing technologies and computational tools over the past few years have led to vast improvements in the metagenomic analysis of the human microbiota. While this has also significantly improved our understanding of the role of the host-microbiome interaction in health and disease, the current clinical expectation is that testing, particularly of the gastrointestinal biome, can be used to diagnose, manage and treat patients. The authors outline the available technologies and highlight current limitations of these techniques to address this clinical demand. Through understanding the limitations of and need for more research and data collection, one can improve the appropriate utilisation and interpretation, as well as the current rational clinical application of these techniques.

Chronically elevated bilirubin protects from cardiac reperfusion injury in the male Gunn rat.

AIMS: Bilirubin is associated with reduced risk of cardiovascular disease, as evidenced in conditions of mild hyperbilirubinaemia (Gilbert's Syndrome). Little is known regarding myocardial stress resistance in hyperbilirubinaemic conditions or whether life-long exposure modifies cardiac function, which might contribute to protection from cardiovascular disease. METHODS: Hyperbilirubinaemic rats and littermate controls underwent echocardiography at 3, 6 and 12 months of age, with hearts subsequently assessed for resistance to 30 min of ischaemia. Heart tissue was then collected for assessment of bilirubin content. RESULTS: No difference in baseline cardiac function was evident until 6 months onwards, where Gunn rats demonstrated aortic dilatation and reduced peak ejection velocities. Additionally, duration of ventricular ejection increased progressively, indicating a negative inotropic effect of bilirubin in vivo. Ex vivo analysis of baseline function revealed reduced left ventricular pressure development (LVDP) and contractility in hyperbilirubinaemic rats. Furthermore, stress resistance was improved in Gunn hearts: post-ischaemic recoveries of LVDP (76 ± 22% vs. 29 ± 17% Control, P < 0.01) and coronary flow (96 ± 9% vs. 86 ± 16% Control, P < 0.01) were improved in Gunn hearts, accompanied by reduced infarct area (21 ± 5% vs. 47 ± 15% Control, P < 0.01), and ventricular malondialdehyde and protein carbonyl content. Expression of myocardial nitric oxide-regulating genes including Nos1 and Noa1 were not significantly different. CONCLUSIONS: These data reveal life-long hyperbilirubinaemia induces age-dependent hypocontractility in male Gunn rats, and improved stress resistance. In addition, bilirubin exerts sex-independent effects on vascular structure, myocardial function and ischaemic tolerance, the latter likely mediated via bilirubin's antioxidant properties.

Comparing laboratory costs of smear/culture and Xpert® MTB/RIF-based tuberculosis diagnostic algorithms.

SETTING: Cape Town, South Africa, where Xpert® MTB/RIF was introduced as a screening test for all presumptive tuberculosis (TB) cases. OBJECTIVE: To compare laboratory costs of smear/culture- and Xpert-based tuberculosis (TB) diagnostic algorithms in routine operational conditions. METHODS: Economic costing was undertaken from a laboratory perspective, using an ingredients-based costing approach. Cost allocation was based on reviews of standard operating procedures and laboratory records, timing of test procedures, measurement of laboratory areas and manager interviews. We analysed laboratory test data to assess overall costs and cost per pulmonary TB and multidrug-resistant TB (MDR-TB) case diagnosed. Costs were expressed as 2013 Consumer Price Index-adjusted values. RESULTS: Total TB diagnostic costs increased by 43%, from US$440 967 in the smear/culture-based algorithm (April-June 2011) to US$632 262 in the Xpert-based algorithm (April-June 2013). The cost per TB case diagnosed increased by 157%, from US$48.77 (n = 1601) to US$125.32 (n = 1281). The total cost per MDR-TB case diagnosed was similar, at US$190.14 and US$183.86, with 95 and 107 cases diagnosed in the respective algorithms. CONCLUSION: The introduction of the Xpert-based algorithm resulted in substantial cost increases. This was not matched by the expected increase in TB diagnostic efficacy, calling into question the sustainability of this expensive new technology.

The effects of visceral obesity and androgens on bone: trenbolone protects against loss of femoral bone mineral density and structural strength in viscerally obese and testosterone-deficient male rats.

SUMMARY: In males, visceral obesity and androgen deficiency often present together and result in harmful effects on bone. Our findings show that both factors are independently associated with adverse effects on femoral bone structure and strength, and trenbolone protects rats from diet-induced visceral obesity and consequently normalises femoral bone structural strength. INTRODUCTION: In light of the rapidly increasing incidence of obesity and osteoporosis globally, and recent conjecture regarding the effects of visceral adiposity and testosterone deficiency on bone health, we investigated the effects of increased visceral adipose tissue (VAT) mass on femoral bone mineral density (BMD), structure and strength in normal weight rats with testosterone deficiency. METHODS: Male Wistar rats (n = 50) were fed either standard rat chow (CTRL, n = 10) or a high-fat/high-sugar diet (HF/HS, n = 40). Following 8 weeks of feeding, rats underwent sham surgery (CTRL, n = 10; HF/HS, n = 10) or orchiectomy (HF/HS + ORX, n = 30). Following a 4-week recovery period, mini-osmotic pumps containing either vehicle (CTRL, n = 10; HF/HS, n = 10; HF/HS + ORX, n = 10), 2.0 mg kg day(-1), testosterone (HF/HS + ORX + TEST, n = 10) or 2.0 mg kg day(-1) trenbolone (HF/HS + ORX + TREN, n = 10) were implanted for 8 weeks of treatment. Dual-energy X-ray absorptiometry and three-point bending tests were used to assess bone mass, structure and strength of femora. RESULTS: Diet-induced visceral obesity resulted in decreased bone mineral area (BMA) and content (BMC) and impaired femoral stiffness and strength. Orchiectomy further impaired BMA, BMC and BMD and reduced energy to failure in viscerally obese animals. Both TEST and TREN treatment restored BMA, BMC, BMD and energy to failure. Only TREN reduced visceral adiposity and improved femoral stiffness and strength. CONCLUSIONS: Findings support a role for both visceral adiposity and testosterone deficiency as independent risk factors for femoral osteoporosis, adverse bone geometry and impaired bone strength in male rats. Trenbolone may be a more effective candidate for androgen replacement therapy than testosterone in viscerally obese testosterone-deficient males.

Selective pinning control of the average disease transmissibility in an HIV contact network.

Medication is applied to the HIV-infected nodes of high-risk contact networks with the aim of controlling the spread of disease to a predetermined maximum level. This intervention, known as pinning control, is performed both selectively and randomly in the network. These strategies are applied to 300 independent realizations per reference level of incidence on connected undirectional networks without isolated components and varying in size from 100 to 10,000 nodes per network. It is shown that a selective on-off pinning control strategy can control the networks studied with limited steady-state error and, comparing the medians of the doses from both strategies, uses 51.3% less medication than random pinning of all infected nodes. Selective pinning could possibly be used by public health specialists to identify the maximum level of HIV incidence in a population that can be achieved in a constrained funding environment.

Comparing multidrug-resistant tuberculosis patient costs under molecular diagnostic algorithms in South Africa.

SETTING: Ten primary health care facilities in Cape Town, South Africa, 2010-2013. OBJECTIVE: A comparison of costs incurred by patients in GenoType MDRTBplus line-probe assay (LPA) and Xpert MTB/RIF-based diagnostic algorithms from symptom onset until treatment initiation for multidrug-resistant tuberculosis (MDR-TB). METHODS: Eligible patients identified from laboratory and facility records were interviewed 3-6 months after treatment initiation and a cost questionnaire completed. Direct and indirect costs, individual and household income, loss of individual income and change in household income were recorded in local currency, adjusted to 2013 costs and converted to $US. RESULTS: Median number of visits to initiation of MDR-TB treatment was reduced from 20 to 7 (P < 0.001) and median costs fell from US$68.1 to US$38.3 (P = 0.004) in the Xpert group. From symptom onset to being interviewed, the proportion of unemployed increased from 39% to 73% in the LPA group (P < 0.001) and from 53% to 89% in the Xpert group (P < 0.001). Median household income decreased by 16% in the LPA group and by 13% in the Xpert group. CONCLUSION: The introduction of an Xpert-based algorithm brought relief by reducing the costs incurred by patients, but loss of employment and income persist. Patients require support to mitigate this impact.

Two novel HLA alleles, HLA-A*30:02:01:03 and HLA-C*08:113, identified in a South African bone marrow donor.

Genomic sequencing of HLA-A*30:02:01:03, an intronic variant, and HLA-C*08:113, an exonic variant.

Myocardial structure, function and ischaemic tolerance in a rodent model of obesity with insulin resistance.

Obesity and its comorbidities (dyslipidaemia, insulin resistance and hypertension) that together constitute the metabolic syndrome are all risk factors for ischaemic heart disease. Although obesity has been reported to be an independent risk factor for congestive heart failure, whether obesity-induced heart failure develops in the absence of increased afterload (induced by hypertension) is not clear. We have previously shown that obesity with insulin resistance decreases myocardial tolerance to ischaemia-reperfusion, but the mechanism for this decreased tolerance remains unclear. We hypothesize that obesity with insulin resistance induces adverse cardiac remodelling and pump dysfunction, as well as adverse changes in myocardial prosurvival reperfusion injury salvage kinase (RISK) pathway signalling to reduce myocardial tolerance to ischaemia-reperfusion. Wistar rats were fed an obesogenic (obese group) or a standard rat chow diet (control group) for 32 weeks. Echocardiography was performed over the 32 weeks before isolated Langendorff-perfused hearts were subjected to 40 min coronary artery ligation followed by reperfusion, and functional recovery (rate-pressure product), infarct size and RISK pathway function were assessed (Western blot analysis). Obesity with insulin resistance increased myocardial lipid accumulation but had no effect on in vivo or ex vivo left ventricular structure/function. Hearts from obese rats had lower reperfusion rate-pressure products (13115 ± 562 beats min(-1) mmHg for obese rats versus 17781 ± 1109 beats min(-1) mmHg for control rats, P < 0.05) and larger infarcts (36.3 ± 5.6% of area at risk in obese rats versus 14.1 ± 2.8% of area at risk in control rats, P < 0.01) compared with control hearts. These changes were associated with reductions in RISK pathway function, with 30-50 and 40-60% reductions in Akt and glycogen synthase kinase 3 beta (GSK-3ß) expression and phosphorylation, respectively, in obese rat hearts compared with control hearts. Total endothelial nitric oxide synthase expression was reduced by 25% in obese rats. We conclude that obesity with insulin resistance had no effect on basal cardiac structure or function but decreased myocardial tolerance to ischaemia-reperfusion. This reduction in ischaemic tolerance was likely to be due to compromised RISK pathway function in obese, insulin-resistant animals.

Tuberculosis patients in primary care do not start treatment. What role do health system delays play?

SETTING: Primary health care facilities in five provinces of South Africa. OBJECTIVE: To investigate the association between the proportion of sputum results with a prolonged smear turnaround time and the proportion of smear-positive tuberculosis (TB) cases initially lost to follow-up. DESIGN: The unit of investigation was a primary health care facility and the outcome was the initial loss to follow-up rate per facility, which was calculated by comparing the sputum register with the TB treatment register. A prolonged turnaround time was defined as more than 48 h from when the sputum sample was documented in the sputum register to receipt of the result at the facility. RESULTS: The mean initial loss to follow-up rate was 25% (95%CI 22-28). Smear turnaround time overall was inversely associated with initial loss to follow-up (P = 0.008), when comparing Category 2 (33-66% turnaround time within 48 h) with Category 1 (0-32%) (OR 0.73, 95%CI 0.48-1.13, P = 0.163) and when comparing Category 3 (67-100%) with Category 1 (OR 0.62, 95%CI 0.39-0.99, P = 0.045). The population preventable fraction of initial loss to follow-up (when turnaround time was <48 h in ≥67% of smear results) was 21%. CONCLUSION: Initial loss to follow-up should be reported as part of the TB programme to ensure that patients are initiated on treatment to prevent transmission within communities.

Effect of South African beef production systems on post-mortem muscle energy status and meat quality.

Post-slaughter muscle energy metabolism meat colour of South African production systems were compared; steers (n=182) of Nguni, Simmental Brahman crossbreds were reared on pasture until A-, AB-, or B-age, in feedlot until A-AB-age. After exsanguination carcasses were electrically stimulated (400 V for 15 s). M. longissimus dorsi muscle energy samples were taken at 1, 2, 4 and 20 h. Post-mortem samples for meat quality studies were taken at 1, 7 and 14 days post-mortem. Production systems affected muscle glycogen, glucose, glucose-6-P, lactic acid, ATP, creatine-P glycolytic potential (P<0.05), with the muscles of feedlot carcasses having a faster glycolysis rate than pasture carcasses. Energy metabolites correlated (0.40.5) water holding capacity, drip loss, and Warner Bratzler shear force. Muscle energy only affected muscle contraction of the A-age-pasture system (shortest sarcomere length of 1.66 µm vs 1.75 µm highest WBS of 6 kg vs 5 kg 7 days post-mortem).