Identification and validation of diagnostic cut-offs of the ELISpot assay for the diagnosis of invasive aspergillosis in high-risk patients

PLoS One. 2024 Jul 9;19(7):e0306728. doi: 10.1371/journal.pone.0306728. eCollection 2024.

Abstract

Objective: We investigated the performance of enzyme linked immunospot (ELISpot) assay for the diagnosis of invasive aspergillosis (IA) in high-risk patients with hematologic malignancies.

Methods: We prospectively enrolled two cohorts of patients undergoing intensive myelosuppressive or immunosuppressive treatments at high risk for IA. ELISpot was performed to detect Aspergillus-specific T cells producing Interleukin-10.

Results: In the discovery cohort, a derived cut-off of 40 spot forming cells (SFCs)/106 PBMCs has shown to correctly classify IA cases with a sensitivity and specificity of 89.5% and 88.6%, respectively. This cut-off is lowered to 25 SFC when considering the subset of possible IA patients, with sensitivity and specificity of 76% and 93%, respectively. The application of the 40 SFCs cut-off to the validation cohort resulted in a positivity rate of 83.3% in proven/probable cases and a negativity rate of 92.5% in possible/non-IA cases. Adopting the 25 SCFs cut-off, the assay resulted positive in 83.3% of proven/probable cases while it resulted negative in 66.7% of possible/non-IA cases.

Conclusions: ELISpot shows promises in the diagnosis of IA and the possibility to use two distinct cut-offs with similar diagnostic performances according to patients' different pre-test probability of infection can widen its use in patients at risk.

Publication types

  • Validation Study

MeSH terms

  • Adult
  • Aged
  • Aspergillosis / diagnosis
  • Aspergillosis / immunology
  • Enzyme-Linked Immunospot Assay* / methods
  • Female
  • Hematologic Neoplasms / diagnosis
  • Hematologic Neoplasms / immunology
  • Humans
  • Interleukin-10 / immunology
  • Male
  • Middle Aged
  • Prospective Studies
  • Sensitivity and Specificity
  • T-Lymphocytes / immunology

Substances

  • Interleukin-10

Grants and funding

This work was supported by grants to M.L. from Gilead Sciences, Fellowship Program 2014 and 2018; PNRR CN3 Terapia Genica-Spoke 2 (Project #CN00000041); the “Charity Dinner Initiative” in memory of Alberto Fontana for Associazione Italiana Lotta alle Leucemie, Linfoma e Mieloma (AIL)—Sezione ‘Luciano Pavarotti’—Modena-ONLUS; Fondazione IRIS CERAMICA GROUP. This work was also supported by grants to P.C. from the Fondazione Regionale per la Ricerca Biomedica Project N_ CP2_10/2018; Regione Lombardia, Progetto POR-FESR 2526393 “Force4Cure”; Fondazione IRCCS Policlinico San Matteo di Pavia (Ricerca Corrente 08069119 and 08045818); Fondazione Just Italia. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.