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PurposeEstimating the incremental costs of scaling‐up novel technologies in low‐income and middle‐income countries is a methodologically challenging and substantial empirical undertaking, in the absence of routine cost data collection. We demonstrate a best practice pragmatic approach to estimate the incremental costs of new technologies in low‐income and middle‐income countries, using the example of costing the scale‐up of Xpert Mycobacterium tuberculosis (MTB)/resistance to riframpicin (RIF) in South Africa.Materials and methodsWe estimate costs, by applying two distinct approaches of bottom‐up and top‐down costing, together with an assessment of processes and capacity.ResultsThe unit costs measured using the different methods of bottom‐up and top‐down costing, respectively, are $US16.9 and $US33.5 for Xpert MTB/RIF, and $US6.3 and $US8.5 for microscopy. The incremental cost of Xpert MTB/RIF is estimated to be between $US14.7 and $US17.7. While the average cost of Xpert MTB/RIF was higher than previous studies using standard methods, the incremental cost of Xpert MTB/RIF was found to be lower.ConclusionCosts estimates are highly dependent on the method used, so an approach, which clearly identifies resource‐use data collected from a bottom‐up or top‐down perspective, together with capacity measurement, is recommended as a pragmatic approach to capture true incremental cost where routine cost data are scarce.

The human immunoglobulin repertoire is vast, producing billions of unique antibodies from a limited number of germline immunoglobulin genes. The immunoglobulin heavy chain variable region (IGHV) is central to antigen binding and is comprised of 48 functional genes. Here we analyzed whether HIV-1 infected individuals who develop broadly neutralizing antibodies show a distinctive germline IGHV profile. Using both 454 and Illumina technologies we sequenced the IGHV repertoire of 28 HIV-infected South African women from the Center for the AIDS Programme of Research in South African (CAPRISA) 002 and 004 cohorts, 13 of whom developed broadly neutralizing antibodies. Of the 259 IGHV alleles identified in this study, approximately half were not found in the International Immunogenetics Database (IMGT). This included 85 entirely novel alleles and 38 alleles that matched rearranged sequences in non-IMGT databases. Analysis of the rearranged H chain V region genes of monoclonal antibodies isolated from 7 of the CAPRISA women and previously isolated broadly neutralizing antibodies from other donors provided evidence that at least 8 novel or non-IMGT alleles contributed to functional antibodies. Importantly, we found that despite a wide range in the number of IGHV alleles in each individual, including alleles used by known broadly neutralizing antibodies, there were no significant differences in germline IGHV repertoires between individuals who do and do not develop broadly neutralizing antibodies. This study reports novel IGHV repertoires and highlights the importance of a fully comprehensive immunoglobulin database for germline gene usage prediction. Furthermore, these data suggest a lack of genetic bias in broadly neutralizing antibody development in HIV-1 infection, with implications for HIV vaccine design.

e South Africa implemented Xpert MTB/RIF as the initial diagnostic test for pulmonary tuberculosis (TB). Xpert MTB/RIF's accuracy for diagnosing extrapulmonary tuberculosis (EPTB) was investigated. EPTB specimens (n ‫؍‬ 7,916) from hospitalized patients received over a 6-month period at a high-throughput TB referral laboratory in Johannesburg were investigated. Largevolume specimens were centrifuged, tissue biopsy specimens homogenized, and all specimens checked for growth of contaminating bacteria on blood agar. Contaminated samples received NALC-NaOH (N-acetyl-L-cysteine-sodium hydroxide) decontamination prior to liquid culture. Residual specimens (volumes > 1 ml) after inoculation of culture (n ‫؍‬ 1,175) were tested using the Xpert MTB/RIF sputum protocol. Using culture as the reference, Xpert MTB/RIF's overall sensitivity was 59% (95% confidence interval [95% CI], 53% to 65%) and specificity was 92% (CI, 90% to 94%), with the highest sensitivities of 91% (95% CI, 78% to 97%) for pus, 80% (95% CI, 56% to 94%) for lymph node aspirates, and 51% (95% CI, 44% to 58%) for fluids ( ascitic, 59%; pleural, 47%). A difference in sensitivities was noticed between specimens classified as having a thick (87% [95% CI, 76% to 94%]) versus clear (watery) (48% [95% CI, 36% to 61%]) appearance. This was unchanged with traces of blood (52% [95% CI, 44% to 60%]) or precentrifugation (57% [95% CI, 28% to 82%]) among clear specimens. Xpert MTB/RIF generated an additional 124 specimen results that were contaminated by Mycobacterial Growth Indicator Tubes (MGIT; 10.5%) and diagnosed rifampin (RIF) resistance earlier (9.6% [25/260]). Xpert MTB/RIF's performance on EPTB specimens provides very promising results and should be considered for incorporation into national TB guidelines. Xpert MTB/RIF is less affected by contaminating bacteria and reduces laboratory labor and diagnostic delay compared to traditional methods. Evidence from 138 studies published before 2008 suggested that nucleic acid amplification technologies (NAAT) could not replace conventional mycobacterial tests (microscopy, culture) for diagnosing pulmonary and, especially, extrapulmonary tuberculosis (EPTB) (1). Only a few years later, GeneXpert technology (2) has changed this paradigm, with a recent systematic review showing pooled sensitivity of 88% and pooled specificity of 98% (3) for diagnosis of pulmonary TB, but evidence (as of March 2012) for using Xpert MTB/RIF for diagnosing EPTB is still comparatively weak (4). Globally, there is still a dearth of studies involving the use of Xpert MTB/RIF in EPTB specimens, and few provide definitive answers. This is due mostly to the studies having small sample sizes across a range of various specimen types and differences in preprocessing methodologies and in input volumes and to studies having been conducted in different populations (adults, children, HIV infected). In one large published study, the overall sensitivity of Xpert MTB/RIF on tissue biopsy specimens/fineneedle aspirates (FNA), pleural fluid, gastric aspir...

e Early initiation of antiretroviral therapy reduces HIV-related infant mortality. The early peak of pediatric HIV-related deaths in South Africa occurs at 3 months of age, coinciding with the earliest age at which treatment is initiated following PCR testing at 6 weeks of age. Earlier diagnosis is necessary to reduce infant mortality. The performances of the Amplicor DNA PCR, COBAS AmpliPrep/COBAS TaqMan (CAP/CTM), and Aptima assays for detecting early HIV infection (acquired in utero and intrapartum) up to 6 weeks of age were compared. Dried blood spots (DBS) were collected at birth and at 2, 4, and 6 weeks from HIVexposed infants enrolled in an observational cohort study in Johannesburg, South Africa. HIV status was determined at 6 weeks by DNA PCR on whole blood. Serial DBS samples from all HIV-infected infants and two HIV-uninfected, age-matched controls were tested with the 3 assays. Of 710 infants of known HIV status, 38 (5.4%) had in utero (n ‫؍‬ 29) or intrapartum (n ‫؍‬ 9) infections. By 14 weeks, when treatment should have been initiated, 13 (45%) in utero-infected and 2 (22%) intrapartum-infected infants had died or were lost to follow-up. The CAP/CTM and Aptima assays identified 76.3% of all infants with early HIV infections at birth and by 4 weeks were 96% sensitive. DNA PCR demonstrated lower sensitivities at birth and 4 weeks of 68.4% and 87.5%, respectively. All assays had the lowest sensitivity at 2 weeks of age. CAP/CTM was the only assay with 100% specificity at all ages. Testing at birth versus 6 weeks of age identifies a higher total number of HIV-infected infants, irrespective of the assay.

BackgroundExpansion of HIV viral load (VL) testing services are required to meet increased targets for monitoring patients on antiretroviral treatment. South Africa currently tests >4million VLs per annum in 16 highly centralised, automated high-throughput laboratories. The Xpert HIV-1 VL assay (Cepheid) was evaluated against in-country predicates, the Roche Cobas Taqmanv2 and Abbott HIV-1RT, to investigate options for expanding VL testing using GeneXpert’s random access, polyvalent capabilities and already established footprint in South Africa with the Xpert MTB/RIF assay (207 sites). Additionally, the performance of Xpert HIV-1VL on alternative, off-label specimen types, Dried Blood Spots (DBS) and whole blood, was investigated.MethodPrecision, accuracy (agreement) and clinical misclassification (1000cp/ml) of Xpert HIV-1VL plasma was compared to Taqmanv2 (n = 155) and Abbott HIV-1 RT (n = 145). Misclassification of Xpert HIV-1VL was further tested on DBS (n = 145) and whole blood (n = 147).ResultsXpert HIV-1VL demonstrated 100% concordance with predicate platforms on a standardised frozen, plasma panel (n = 42) and low overall percentage similarity CV of 1.5% and 0.9% compared to Taqmanv2 and Abbott HIV-1 RT, respectively. On paired plasma clinical specimens, Xpert HIV-1VL had low bias (SD 0.32–0.37logcp/ml) and 3% misclassification at the 1000cp/ml threshold compared to Taqmanv2 (fresh) and Abbott HIV-1 RT (frozen), respectively. Xpert HIV-1VL on whole blood and DBS increased misclassification (upward) by up to 14% with increased invalid rate. All specimen testing was easy to perform and compatible with concurrent Xpert MTB/RIF Tuberculosis testing on the same instrument.ConclusionThe Xpert HIV-1VL on plasma can be used interchangeably with existing predicate platforms in South Africa. Whole blood and DBS testing requires further investigation, but polyvalency of the GeneXpert offers a solution to extending VL testing services.

IntroductionThe World Health Organization recommends viral load (VL) monitoring at six and twelve months and then annually after initiating antiretroviral treatment for HIV. In many African countries, expansion of VL testing has been slow due to a lack of efficient blood sample transportation networks (STN). To assist Zambia in scaling up testing capacity, we modelled an optimal STN to minimize the cost of a national VL STN.MethodsThe model optimizes a STN in Zambia for the anticipated 1.5 million VL tests that will be needed in 2020, taking into account geography, district political boundaries, and road, laboratory and facility infrastructure. We evaluated all‐inclusive STN costs of two alternative scenarios: (1) optimized status quo: each district provides its own weekly or daily sample transport; and (2) optimized borderless STN: ignores district boundaries, provides weekly or daily sample transport, and reaches all Scenario 1 facilities.ResultsUnder both scenarios, VL testing coverage would increase to from 10% in 2016 to 91% in 2020. The mean transport cost per VL in Scenario 2 was $2.11 per test (SD $0.28), 52% less than the mean cost/test in Scenario 1, $4.37 (SD $0.69), comprising 10% and 19% of the cost of a VL respectively.ConclusionsAn efficient STN that optimizes sample transport on the basis of geography and test volume, rather than political boundaries, can cut the cost of sample transport by more than half, providing a cost savings opportunity for countries that face significant resource constraints.

Introduction Routine viral load testing is the WHO‐recommended method for monitoring HIV‐infected patients on ART, and many countries are rapidly scaling up testing capacity at centralized laboratories. Providing testing access to the most remote populations and facilities (the “last mile”) is especially challenging. Using a geospatial optimization model, we estimated the incremental costs of accessing the most remote 20% of patients in Zambia by expanding the transportation network required to bring blood samples from ART clinics to centralized laboratories and return results to clinics. Methods The model first optimized a sample transportation network (STN) that can transport 80% of anticipated sample volumes to centralized viral load testing laboratories on a daily or weekly basis, in line with Zambia's 2020 targets. Data incorporated into the model included the location and infrastructure of all health facilities providing ART, location of laboratories, measured distances and drive times between the two, expected future viral load demand by health facility, and local cost estimates. We then continued to expand the modelled STN in 5% increments until 100% of all samples could be collected. Results and Discussion The cost per viral load test when reaching 80% patient volumes using centralized viral load testing was a median of $18.99. With an expanded STN, the incremental cost per test rose to $20.29 for 80% to 85% and $20.52 for 85% to 90%. Above 90% coverage, the incremental cost per test increased substantially to $31.57 for 90% to 95% and $51.95 for 95% to 100%. The high numbers of kilometres driven per sample transported and large number of vehicles needed increase costs dramatically for reaching the clinics that serve the last 5% of patients. Conclusions Providing sample transport services to the most remote clinics in low‐ and middle‐income countries is likely to be cost‐prohibitive. Other strategies are needed to reduce the cost and increase the feasibility of making viral load monitoring available to the last 10% of patients. The cost of alternative methods, such as optimal point‐of‐care viral load equipment placement and usage, dried blood/plasma spot specimen utilization, or use of drones in geographically remote facilities, should be evaluated.