Diagnostic utility of zinc protoporphyrin to detect iron deficiency in Kenyan preschool children: a community-based survey

Abstract

Background: Zinc protoporphyrin (ZPP) has been used to screen and manage iron deficiency in individual children, but it has also been recommended to assess population iron status. The diagnostic utility of ZPP used in combination with haemoglobin concentration has not been evaluated in pre-school children. We aimed to a) identify factors associated with ZPP in children aged 12-36 months; b) assess the diagnostic performance and utility of ZPP, either alone or in combination with haemoglobin, to detect iron deficiency.

Methods: We used baseline data from 338 Kenyan children enrolled in a community-based randomised trial. To identify factors related to ZZP measured in whole blood or erythrocytes, we used bivariate and multiple linear regression analysis. To assess diagnostic performance, we excluded children with elevated plasma concentrations of C-reactive protein or α1-acid glycoprotein, and with Plasmodium infection, and we analysed receiver operating characteristics (ROC) curves, with iron deficiency defined as plasma ferritin concentration < 12 μg/L. We also developed models to assess the diagnostic utility of ZPP and haemoglobin concentration when used to screen for iron deficiency.

Results: Whole blood ZPP and erythrocyte ZPP were independently associated with haemoglobin concentration, Plasmodium infection and plasma concentrations of soluble transferrin receptor, ferritin, and C-reactive protein. In children without inflammation or Plasmodium infection, the prevalence of true iron deficiency was 32.1%, compared to prevalence of 97.5% and 95.1% when assessed by whole blood ZPP and erythrocyte ZPP with conventional cut-off points (70 μmol/mol and 40 μmol/mol haem, respectively). Addition of whole blood ZPP or erythrocyte ZPP to haemoglobin concentration increased the area-under-the-ROC-curve (84.0%, p = 0.003, and 84.2%, p = 0.001, respectively, versus 62.7%). A diagnostic rule (0.038689 [haemoglobin concentration, g/L] + 0.00694 [whole blood ZPP, μmol/mol haem] >5.93120) correctly ruled out iron deficiency in 37.4%-53.7% of children screened, depending on the true prevalence, with both specificity and negative predictive value ≥90%.

Conclusions: In young children, whole blood ZPP and erythrocyte ZPP have added diagnostic value in detecting iron deficiency compared to haemoglobin concentration alone. A single diagnostic score based on haemoglobin concentration and whole blood ZPP can rule out iron deficiency in a substantial proportion of children screened.

Trial registration: ClinicalTrials.gov NCT02073149 (25 February 2014).

Keywords: Child; Erythrocyte protoporphyrin; Inflammation; Iron deficiency; Kenya; Malaria; Plasmodium; Preschool; Zinc protoporphyrin.

Conflict of interest statement

Competing interests

The authors declare that they have no competing interests.

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Figures

Fig. 1

Ability of zinc protoporphyrin-haem ratio, either alone or in combination with haemoglobin concentration, to discriminate between children with and without iron deficiency. Panel a: Receiver operating characteristics (ROC) curves for various blood markers, used alone, to discriminate between iron-deficient and iron-replete children. Panel b: As in Panel a, with haemoglobin concentration and whole blood ZPP, alone and in combination. Panel c: As in Panel a, with haemoglobin concentration and erythrocyte ZPP, alone and in combination. Panel d: As in Panel a, with combined haemoglobin concentration and whole blood ZPP, versus combined haemoglobin concentration and erythrocyte ZPP. Hb: haemoglobin concentration. Grey diagonal lines in ROC curves indicate a ‘worst’ possible test, which has no discriminatory value and an area-under-the-curve (AUC) of 0.5. An ideal marker would have a curve that runs from the lower-left via the upper-left to the upper-right corner, yielding an AUC of 1.0

Fig. 2

Application of a diagnostic strategy to rule out iron deficiency. [Hb]: haemoglobin concentration, expressed in g/L; [whole blood ZPP]: whole blood ZPP content, expressed in μmol/mol haem. The diagnostic strategy in a screen-and-treat survey is based on two criteria: a) the probability of correctly diagnosing iron deficiency should exceed 90%; and b) iron deficiency can be ruled out if the probability of a negative test result being correct (negative predictive value) exceeds 90%. To meet the first requirement, a cut-off point for each diagnostic test result (top) is selected to yield a sensitivity of 90%; the corresponding specificity value is obtained from the ROC curves (Fig. 1). For haemoglobin concentration and whole blood ZPP, the cut-off points were 122 g/L and 99 μmol/mol haem; the corresponding specificity values were 14.8% and 36.0%, respectively. When these markers were combined in a single diagnostic rule, 0.038689 [Hb] + 0.00694 [whole blood ZPP] > 5.93120 had a specificity of 53.7%. The negative predictive value (top panels, blue lines) depends on sensitivity and specificity values thus fixed, and the prevalence of iron deficiency. The second diagnostic criterion, i.e. the negative predictive value should exceed 90%, applies only within a limited prevalence range (top panels, red rectangle); at prevalence values exceeding this range, the negative predictive value will be below 90% and iron deficiency cannot be ruled out with diagnostic test applied (top). The percentage of children with a negative test result declines linearly with the prevalence of iron deficiency (middle panels, blue lines). The percentage of children for whom iron deficiency can be ruled out (middle panels, Y-intercepts of red rectangles) depends on the prevalence range in which the negative predictive value exceeds 90% (top panels, red rectangles). With fixed sensitivity and specificity values, the positive predictive value (bottom panels, blue lines) increase monotonically with the prevalence of iron deficiency. Within the prevalence range in which the negative predictive value exceeds 90% (top panels, red rectangles), the highest positive predictive value is 54% (for combined use of haemoglobin concentration and whole blood ZPP), indicating that additional tests (i.e. other than haemoglobin concentration and whole blood ZPP) are required to accurately determine iron status. For example, haemoglobin concentration > 122 g/L (left panels) has 90% sensitivity of detecting iron deficiency; at a true prevalence of iron deficiency <14.1%, a negative test result obtained with this decision rules out iron deficiency with a probability of 90% (upper left panel, red rectangle). Depending on the true prevalence of iron deficiency, such a cut-off for haemoglobin concentration would result in iron deficiency being ruled out in 14.1%–14.8% of children who are tested (middle left panel, red rectangle). Similarly, within a prevalence range < 28.6%, whole blood ZPP > 99 μmol/mol haem rules out iron deficiency in 28.6%–36% of children tested. Within a prevalence range of <37.4%, 0.038689 [Hb] + 0.00694 [whole blood ZPP] > 5.93120 rules out iron deficiency in 37.4%–53.7% of children