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Cited by 9Malaria Research and ControlHemoglobinopathies and Related DisordersDigital Imaging for Blood Diseases

Diagnosis of placental malaria in poorly fixed and processed placental tissue

Open access
Liu, Yunhao, Griffin, Jennifer B., Muehlenbachs, Atis, Rogerson, Stephen J., Bailis, Anya J., Sharma, Rajni, Sullivan, David J., Tshefu, Antoinette K. +4 more·2016·Malaria Journal
DOI 10.1186/s12936-016-1314-6PMC PMC4863337
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Abstract

BACKGROUND: Placental histopathology has been considered the gold standard for diagnosis of malaria during pregnancy. However, in under-resourced areas placental tissue is often improperly fixed and processed; the resulting formalin pigment is difficult to distinguish from malaria pigment. This study examines two alternative diagnostic methods: polymerase chain reaction (PCR) and a novel immunohistochemistry (IHC)-based method using an antibody against histidine-rich protein 2 (HRP2). METHODS: Placental histopathology from 151 pregnant women in Kinshasa was assessed by two blinded microscopists and compared with peripheral blood PCR and IHC for HRP2. The Cohen's kappa coefficients were calculated to assess the test agreement. The sensitivity and specificity of individual tests were calculated using PCR or IHC as the reference standard as well as latent class analysis (LCA). RESULTS: PCR and IHC correlated fairly well. The correlation between the two blinded microscopists was poor, as there was widespread formalin pigment. Using LCA, all of the tests had high specificities. The most sensitive test was IHC (67.7 %), with PCR as second-best (56.1 %). CONCLUSIONS: PCR and/or IHC are suitable diagnostics when the presence of formalin pigment substantially compromises placental histopathology.

AI snapshot

Written from the paper’s full text.

TL;DR

In 151 placental biopsies from Kinshasa with abundant formalin pigment, histopathology agreement between two blinded microscopists was poor, while PCR and HRP2 immunohistochemistry (IHC) correlated fairly well. Latent class analysis found all tests highly specific, with IHC the most sensitive (67.7%) and PCR second (56.1%), supporting PCR and/or IHC when placental histopathology is compromised.

Key findings

  • All placental biopsy specimens had moderate to high formalin pigment; the two blinded histopathology readers had poor agreement (Cohen’s κ = −0.0462, p = 0.5448).
  • IHC and PCR correlated fairly well (κ = 0.3176, p < 0.0001); histopathology reader P2 correlated with IHC (κ = 0.2775) and PCR (κ = 0.2916).
  • Using LCA, specificities were high for all tests: histopathology P1 96.5%, P2 96.8%, IHC 94.9%, PCR 98.4%. Sensitivities were IHC 67.7%, PCR 56.1%, histopathology P2 54.4%, and histopathology P1 0%.
  • Prevalence by method among 151 samples: histopathology P1 5 positives (3.31%), P2 10 (6.62%), IHC 14 (9.27%), PCR 8 (5.30%), LCA 6.71%; 44 women (29.1%) were ever qPCR-positive during pregnancy.

Methods

  • Prospective cohort in Kinshasa, Democratic Republic of Congo, 2005–2006: 182 pregnant women enrolled, with 151 placental biopsy specimens available for this analysis; peripheral dried blood spots and formalin-fixed placental tissue were collected.
  • PCR was real-time qPCR targeting Plasmodium 18S rRNA from dried blood spots; histopathology used Giemsa-stained placental sections read by two blinded microscopists; IHC used monoclonal antibody 3A4 against P. falciparum HRP2.
  • Agreement was assessed with Cohen’s kappa; sensitivity and specificity were calculated using PCR or IHC as reference standards and by latent class analysis (LCA) using PROC LCA, confirmed with poLCA in R.

Limitations

  • The paper states there was no gold standard; formalin pigment was abundant and both pathologists considered the paraffin sections low quality and unreliable for reading.
  • The paper notes PCR was done at the last antenatal visit and infections could have cleared by delivery, and that low malaria prevalence was likely due to intermittent preventive therapy and treatment.
  • Evident from the text: positive counts were small (5–14 by individual methods), LCA sensitivity confidence intervals were very wide (e.g., IHC 10.5–100%), and LCA assumes conditional independence though the paper states it is robust to violations.

Model-written summary. Check it against the paper before relying on it.

Integrity signals

Registry

Crossref · 10.1186/s12936-016-1314-6

Cites

Statistics

Nothing to recompute.

Claims the paper makes

0 of 8 cited sources checked · 0 supported · 0 flagged

  • Findingown data

    The correlation between the two blinded microscopists was poor.

    “The correlation between the two blinded microscopists was poor, as there was widespread formalin pigment.”
    • Also stated in: § Correlations among tests · plain
      “There was poor correlation between the two histopathology readers.”
    • Also stated in: § Discussion · plain
      “the findings of the two microscopists did not correlate at all.”
    Quote in the paperSubstantial claim
  • Findingown data

    Histopathology by reader P2 correlated fairly well with IHC and PCR.

    “Readings by P2, on the other hand, correlated fairly well with IHC and PCR.”
    Quote in the paperSubstantial claim
  • Findingown data

    With PCR as the reference standard, all other tests had high specificities but low sensitivities.

    “With PCR as the reference standard, all of the other tests had high specificities, but low sensitivities.”
    Quote in the paperSubstantial claim
  • Findingown data

    PCR and IHC correlated fairly well.

    “PCR and IHC correlated fairly well.”
    • Also stated in: § Correlations among tests · plain
      “Agreement between IHC and PCR was fairly good as well.”
    Quote in the paperSubstantial claim
  • Findingown data

    All placental biopsy specimens in this study had a moderate to high level of formalin pigment deposition.

    “All placental biopsy specimens had a moderate to high level of formalin pigment deposition (Fig. 1).”
    • Also stated in: § Discussion · plain
      “both histopathologists noted an abundance of formalin pigment”
    Quote in the paperSubstantial claim
  • May combine two claims. 4
  • Classified as probably not a substantial claim. 6

  • May combine two claims.

    Findingown data

    Using latent class analysis, IHC had the highest sensitivity (67.7%), followed by PCR (56.1%).

    “The most sensitive test was IHC (67.7 %), with PCR as second-best (56.1 %).”
    • Also stated in: § Sensitivity and specificity of diagnostic methods · plain
      “on sensitivity, IHC slightly outperformed PCR and histopathology by reader P2.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    With IHC as the reference standard, both histopathology by P2 and PCR showed poor sensitivity, missing more than 70% of infections.

    “Surprisingly, with IHC as the referent, both histopathology (P2) and PCR showed poor sensitivity, missing >70 % of infections.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    With PCR as the reference standard, IHC was the most sensitive other test but missed 50% of malaria infections diagnosed by PCR.

    “The most sensitive other test was IHC, but it missed 50 % of the malaria infections diagnosed by PCR.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Premisecitation

    Formalin pigment has similar optical properties to malaria pigment and can lead to misclassification in placental histopathology.

    “since formalin pigment has similar optical properties to malaria pigment and can lead to misclassification [6].”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [6] Romagosa C, Menendez C, Ismail MR, Quinto L, Ferrer B, Alonso PL, et al. Polarisation microscopy increases the sensitivity of hemozoin and Plasmodium detection in the histological assessment of placental malaria. Acta Trop. 2004;90:277–84. source

      Not checked in the free run.

  • Classified as probably not a substantial claim.

    Study design and conductnone given

    The stained placental sections were examined by two blinded microscopists.

    “The stained sections were examined by two blinded microscopists under light microscopy to assess parasitaemia and malaria pigment.”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Study design and conductown data

    Blood samples were collected from study participants 0 to 33 days before delivery, with one exception at 77 days before delivery.

    “Blood samples were collected from study participants 0 to 33 days before delivery with one exception (one woman gave blood 77 days before delivery).”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Study design and conductnone given

    The results from Proc LCA were confirmed using the poLCA package for R.

    “The results from Proc LCA were confirmed using the poLCA package for R (R Foundation for Statistical Computing, Vienna, Austria).”
    Quote in the paperNot a substantial claim

  • Classified as probably not a substantial claim.

    Premisecitation

    Latent class analysis does not assume a gold standard but constructs one based on the results of the individual tests.

    “LCA does not assume a gold standard but constructs one based on the results of the individual tests [10].”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [10] Hui SL, Walter SD. Estimating the error rates of diagnostic tests. Biometrics. 1980;36:167–71. source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Study design and conductnone given

    Of 182 women in the study, 151 placental biopsy specimens were available for the current analysis.

    “Of 182 women in the study, 151 placental biopsy specimens were available for the current analysis.”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Study design and conductreasoning

    Due to the large number of missing peripheral blood samples from delivery, the final antenatal dried blood spot was used.

    “Due to the large number of missing peripheral blood samples from delivery, the final antenatal dried blood spot obtained (usually in the late third trimester) was used.”
    Quote in the paperNot a substantial claim
10 of 10

  • The authors state this as an assumption.

    Latent class analysis assumes that diagnostic tests are conditionally independent, given the true disease status of the subject.

    “LCA assumes that diagnostic tests are conditionally independent, given the true disease status of the subject.”

  • The paper relies on this without support.

    Residual formalin pigment is common in under-resourced areas due to the use of inexpensive, low-quality fixative.

    “Unfortunately, residual formalin pigment is common in under-resourced areas due to the use of inexpensive, low-quality fixative.”
2 of 2

Claims other papers make about this one

Each is checked against this paper’s own text: does it say what the citing paper says it does?

Self-check

Percentages, sums and reported differences re-computed from the paper’s own numbers.

Each contradiction between two passages listed here was confirmed by an independent check.

  • IHC specificity does not exceed 95%

    Minor

    The discussion says all tests had specificity above 95%, but Table 6 reports IHC specificity of 94.9%.

1 of 1

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References 25 · Cited by 9

Authors
Liu, Y., Griffin, J. B., Muehlenbachs, A., Rogerson, S. J., Bailis, A. J., Sharma, R., Sullivan, D. J., Tshefu, A. K., Landis, S. H., Kabongo, J.-M. M., Taylor, S. M., & Meshnick, S. R.
Affiliations
  • University of North Carolina at Chapel Hill (US)
  • RTI International (US)
  • University of Washington (US)
  • The University of Melbourne (AU)
  • Johns Hopkins University (US)
  • Ecole de santé publique de Kinshasa (CD)
  • GlaxoSmithKline (United Kingdom) (GB)
  • University of Kinshasa (CD)
  • Duke University (US)
Corresponding author
Liu, Yunhao
ORCID
Venue
Malaria Journal
Publisher
Springer Science and Business Media LLC
Published
2016-05-10
Volume
15
Issue
1
Pages
272-272
ISSN
1475-2875 (ISSN-L)
DOI
10.1186/s12936-016-1314-6
PMC
PMC4863337
OpenAlex
W2389841531
Cited by
9
References
25
Citations by year
2025: 1 · 2023: 2 · 2022: 2 · 2020: 1 · 2019: 1 · 2018: 1 · 2016: 1
FWCI
0.99
Type
journal article
Language
en
Open access
Licence
Locations
  • Malaria Journal (publishedVersion, cc-by)
  • PubMed Central (publishedVersion, cc-by)
  • PubMed (publishedVersion)
  • Minerva Access (University of Melbourne) (submittedVersion, cc-by)
  • Carolina Digital Repository (University of North Carolina at Chapel Hill) (submittedVersion)
  • UNC Libraries
Funders
National Institutes of Health (5-T32-AI070114, 5-R01-AI107949)
Topics
  • Health Sciences › Medicine › Public Health, Environmental and Occupational Health › Malaria Research and Control
  • Health Sciences › Medicine › Genetics › Hemoglobinopathies and Related Disorders
  • Physical Sciences › Computer Science › Computer Vision and Pattern Recognition › Digital Imaging for Blood Diseases
Keywords
placental malaria, malaria in pregnancy, placental pathology, HRP-2, PCR diagnosis, sensitivity and specificity, latent class analysis, resource-limited settings
MeSH terms
Adolescent, Adult, Diagnostic Tests, Routine, Female, Humans, Immunohistochemistry, Malaria, Placenta, Placenta Diseases, Pregnancy, Prospective Studies, Sensitivity and Specificity, Democratic Republic of the Congo, Polymerase Chain Reaction, Young Adult

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Claims read by deepseek-v4.1-flash under definition version 3 (extract-v3) on 2026-10-10.

Classified by jev-1.13-20260917 (gold-guideline-v0.2) on 2026-10-10.

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