Poor rates of linkage to care for those with low CD4+ T-cell counts, eligible for antiretroviral therapy (ART), have been reported in several African cohort studies.^1,2,3,4,5

Several attrition steps exist in the continuum of care pathway: patients lost to care between testing HIV-positive and going for a CD4 test⁶; CD4 test result not available and/or lost^7,8; patient not returning for their CD4 test result; and lack of uptake of care from eligibility to initiation of ART even in those who return for test results.⁹ These challenges may be overcome by point-of-care (POC) testing¹⁰, resulting in less attrition over time.^2,4 It has been suggested that POC CD4 testing in those who do not return for their results would potentially increase enrolment pre-ART.⁹ POC CD4 testing was shown to modestly increase linkage to care and reduce pre-treatment loss to follow-up in fixed and mobile clinics.^2,11,12,13 Factors contributing to pre-treatment loss to follow-up have been previously documented.^{8,14,15,16,17}

The Alere PIMA™ POC has been evaluated against the ‘gold standard’ flow cytometry platforms, for example, Beckman Coulter using panleucogating (PLG)¹⁸; BD FACS count^19,20,21; PARTEC Cytoflow™^19,20; Guava and BD FACS Calibur^{20,21,22,23,24} for the enumeration of CD4+ T-cells in HIV-1-infected adults and in HIV-1-infected pregnant women.²²

This study assessed the accuracy, sensitivity and specificity of the Alere PIMA™ POC analyser in CD4+ T-cell count enumeration compared to the predicate South African National Health Laboratory Services (NHLS) flow cytometry test (Beckman Coulter) and its potential operational role as a predictor of ART eligibility in a primary healthcare clinic (PHC) in Durban, South Africa.

The study occurred at Lancers Road PHC, a facility under the eThekwini Health Unit, situated in the centre of the convergence of the taxi rank in the city centre of Durban. This PHC offers HIV Counselling and Testing (HCT) – approximately 900 per month to walk-in patients who receive pre- and post-test counselling and CD4 testing for the staging of HIV-1-infected disease to determine eligibility for ART. Patients are advised to return after 7 days for their CD4 results. As per the SA HIV and AIDS guidelines²⁵ operating at the time of this study, patients with a CD4+ T-cell count ≤ 350 cells/mm³, upon their return, were medically assessed, and education and counselling undertaken prior to ART initiation. Those ineligible for ART, viz CD4+ count > 350 cells/mm³, were counselled to return 6 monthly for CD4+ T-cell count testing and for further medical assessment. Eligible patients, who did not return for results, were contacted telephonically to ascertain whether they had been initiated on ART elsewhere, and if not, they were encouraged to return for further care.

Conventional flow cytometry to determine CD4 counts usually requires that samples be sent to a central laboratory, which may be off-site. Although the turn-around time for a CD4 test result by the NHLS is 24–72 hours, HIV-1-infected patients are counselled to return to the PHC within 1 week for receipt of these results. POC technologies can reduce these delays resulting in rapid linkage to care. This study demonstrated a high PPV and sensitivity and acceptable specificity in predicting ART eligibility (≤ 500 cells/mm³) using the PIMA™ POC analyser as compared to the NHLS CD4 test.

The majority of HIV-1-positive individuals undergoing CD4 testing were women, of whom 25% were 25–29 years old, whereas the majority of men were older than 30 years of age. There were no significant differences in the median CD4+ T-cell count in men versus women performed by the NHLS versus the PIMA™ POC analyser, although the median count was higher in the latter. Overall, according to NHLS versus PIMA™ POC, 81% versus 80% of individuals were eligible for ART initiation (≤ 500 cells/mm³), of whom 82% versus 84% were men and 81% versus 79% were women, respectively.

There was high reproducibility in all three PIMA™ POC analysers using normal and low beads with coefficient of variation < 5% over time (10 and 165 days). The PIMA™ POC analyser slightly overestimates NHLS flow cytometry in CD4+ T-cell enumeration in this study, which corroborates most studies using capillary or venous blood.^26,30,31,32 This overestimation is minimal (mean bias 17 cells/mm³) and is not clinically significant. Differences have been reported on conventional CD4 testing platforms between the BD FACS count versus the BD FACS Calibur²³ where the mean bias between the two platforms was -76 cells/mm³ (95% CI LOA -316.0–163.0).

The adequate correlation between the PIMA™ POC analyser and FACS Calibur (0.97) corroborates similar findings in another study.²¹ Although a correlation of > 0.90 was observed between the three platforms, these differences are due to variability of instrument settings, antibodies and fluorochromes used, gating strategies and sample volume input.

The overall sensitivity of the PIMA™ POC CD4 test in HIV-1-infected adults and pregnant mothers to determine their eligibility for ART has been reported at 96.3% in individuals with a CD4+ T-cell count of ≤ 250 cells/mm³,²⁴ and 92% and 91% in those with ≤ 350 cells/mm³,^20,22. The total misclassifications have been documented in several studies using the PIMA™ POC analyser: 31%,¹⁸ 17%,¹² 5.2%,³³ 6.7% – 14%,³¹ 10%,²² 11.4%³⁴ and 9%.¹⁹ This study found 13% misclassifications, of which 27/35 were FNs at ≤ 350 CD4+ T-cells/mm³. At a CD4+ T-cell threshold of ≤ 500 cells/mm³, 91% of patients were correctly classified as either eligible or ineligible for ART. In the 13 FNs, the mean bias observed was 149 cells/mm³. The PPV of 95% indicates that only 5% of those who are diagnosed as eligible for ART according to the PIMA™ POC analyser would not be needing treatment according to the NHLS CD4 test result. A high sensitivity of 94% was observed at the sacrifice of lower specificity of 78%. This high sensitivity corroborates recent findings^21,22,34 and fits in well with current SA HIV and AIDS guidelines³⁵ where all those eligible for treatment will be initiated but it will come at a cost of low specificity, whereby individuals not needing treatment will be commenced on ART. However, in light of ART-lowering viral loads and reducing horizontal transmission,^36,37 this downside is minimised. A recent study has demonstrated that as household ART coverage is increased, there is a decrease in HIV acquisition.³⁸ The agreement in these data between the PIMA™ POC analyser and NHLS laboratory-based flow cytometry appears to decline with increasing CD4+ T-cell count ≥ 500 cells/mm³. This is not of concern as these HIV-1-infected individuals are ineligible for ART under current guidelines.

From the operational perspective in the use of the PIMA™ POC analyser, similar to other studies using venous or capillary blood,^{18,20,21,22,33} we also experienced reading errors (8%) mostly because of movement and vibration.³⁴ The ‘operator’ used in our study was a trained laboratory technician compared to health professionals, for example, nurse or counsellor. Several studies have reported that the PIMA™ POC is interchangeable with conventional platforms,^{18,20,22,30,31,33,39} although a study in Kenya²³ found it to be unreliable due to the high coefficient of repeatability and misclassification in favour of undertreatment compared to the FACS Calibur.

Under the standard SA HIV and AIDS guidelines operating at the time of the study, we observed that the median time for patients to return for their CD4 results was 8 days and 7 days in those with ≤ 200 cells/mm³, with a median of 49 days regardless of CD4+ T-cell count from CD4 testing to ART initiation. The use of the PIMA™ POC analysers could facilitate the fast tracking of patients with CD4+ T-cell count ≤ 200 cells/mm³ onto ART within 7 days. In this study, the provision of immediate CD4 test results to patients would have prevented the 35/268 not having access to their results (through them not returning), in whom over half (57%) were eligible for ART.

The high rates (61%) of ‘walk–in’ patients found in this study who were eligible (≤ 350 cells/mm³) for immediate ART at the time of the HIV test, half of whom had CD4+ T-cell counts ≤ 248 cells/mm³, and the time lapse to ART initiation undergirds the urgent need for the use of the rapid PIMA™ POC technology. At a threshold of ≤ 500 cell/mm³, 75% of patients had a median CD4+ T-cell count of 444 cells/mm³ at the time of the HIV test. A recent study reported that providing same-day POC CD4 testing that is not rapid has no benefit in health outcomes.⁹ As suggested by others,^11,18,40,41 we agree that using existing infrastructure and based on demand, the integration of a PHC POC mini-laboratory run by dedicated personnel (laboratory technician) is possible, offering tests for staging and pathology that assess ART eligibility.^42,43 However, as suggested in a recent systematic review,⁴⁴ this needs to be supported by streamlining services through minimising patient clinic visits,²² addressing psychosocial issues and barriers to healthcare,⁴⁵ optimising the opportunity for patient empowerment through counselling and peer support,⁴⁶ emphasising the importance of starting and adhering to ART if eligible,⁶ positive health-seeking behaviours and encouragement for patient ownership of their health. A family-centred model of integrated healthcare incorporating most of the above-mentioned health system changes has previously been shown, in a similar population, to yield high adherence (94%) and retention in the care and management of HIV-1-positive individuals.^47,48 In this study, similar reasons for not linking into care were given as found previously⁴⁵; of those eligible for ART who did not access treatment (33%), the reasons given upon telephonic communication were economic (no money to cover transport costs), social (too busy to come to the clinic), structural (cannot take time off work) and emotional (were not ready to take ART and they were still feeling well). We would anticipate that there would be an increase in loss of uptake of care at the higher CD4+ T-cell count threshold of ≤ 500 cells/mm³ because of the reasons re-iterated. Previous studies have shown that provision of immediate CD4+ T-cell count results increased the number of patients linking into care.^2,4,6,11,13

In summary, the overall agreement between PIMA™ POC analyser and NHLS CD4+ T-cell count enumeration in adult HIV-1-positive individuals was acceptable with clinically insignificant mean bias. Together with high PPV and sensitivity and acceptable specificity, the PIMA™ POC CD4 test has the potential role for CD4+ T-cell enumeration in PHC settings and lends itself to be an excellent facilitator in rapid linkage to care in ART programmes, particularly that it has been demonstrated in simulated cohort models of HIV-1-infected adults and pregnant women, to result in not only better clinical outcomes but also to cost savings in the long term.^49,50 Even in the era of ‘test and treat’,⁵¹ PIMA™ POC CD4 testing would facilitate the fast tracking of patients with low CD4+ T-cell counts (< 200 cells/mm³) for the administration of cotrimoxazole prophylaxis as well as in screening for cryptococcal infection in patients with < 100 cells/mm³. The operational role of the PIMA™ POC CD4 test in provision of immediate CD4+ T-cell count results combined with integrated health system changes and interventions such as mobile phone technology and provision of incentives need to be evaluated in a variety of settings across the HIV cascade, to determine its implementation effectiveness in linkage to care, time to ART initiation and retention in HIV care.