Functionality of Dengue Virus Specific Memory T Cell Responses in Individuals Who Were Hospitalized or Who Had Mild or Subclinical Dengue Infection

<div><p>Background</p><p>Although antibody responses to dengue virus (DENV) in naturally infected individuals have been extensively studied, the functionality of DENV specific memory T cell responses in relation to clinical disease severity is incompletely understood.</p><p>Methodology/Principal findings</p><p>Using ex vivo IFNγ ELISpot assays, and by determining cytokines produced in ELISpot supernatants, we investigated the functionality of DENV-specific memory T cell responses in a large cohort of individuals from Sri Lanka (n=338), who were naturally infected and were either hospitalized due to dengue or had mild or sub clinical dengue infection. We found that T cells of individuals with both past mild or sub clinical dengue infection and who were hospitalized produced multiple cytokines when stimulated with DENV-NS3 peptides. However, while DENV-NS3 specific T cells of those with mild/sub clinical dengue infection were more likely to produce only granzyme B (p=0.02), those who were hospitalized were more likely to produce both TNFα and IFNγ (p=0.03) or TNFα alone.</p><p>We have also investigated the usefulness of a novel T cell based assay, which can be used to determine the past infecting DENV serotype. 92.4% of DENV seropositive individuals responded to at least one DENV serotype of this assay and none of the seronegatives responded. Individuals who were seronegative, but had received the Japanese encephalitis vaccine too made no responses, suggesting that the peptides used in this assay did not cross react with the Japanese encephalitis virus.</p><p>Conclusions/significance</p><p>The types of cytokines produced by DENV-specific memory T cells appear to influence the outcome of clinical disease severity. The novel T cell based assay, is likely to be useful in determining the past infecting DENV serotype in immune-epidemiological studies and also in dengue vaccine trials.</p></div

Cross reactive dengue-specific immune responses and disease severity.

<p>A: Circulating NS3-specific IFNγ ex vivo ELISpot responses were measured in individuals who were hospitalized due to dengue and in those with past mild/sub clinical dengue infection. B: Granzyme B production by PBMCs from individuals who were hospitalized due to dengue and who had a past mild/sub clinical dengue infection, following stimulation with DENV-NS3 overlapping peptides. C: TNFα production by PBMCs from individuals who were hospitalized due to dengue and those with past mild/sub clinical dengue infection following stimulation with DENV-NS3 overlapping peptides.</p

Number and percentage of individuals with mild/sub clinical dengue infection and those who were hospitalized due to dengue who produced of different cytokines when PBMCs were stimulated DENV-NS3 overlapping peptides and JE Live vaccine.

<p>Number and percentage of individuals with mild/sub clinical dengue infection and those who were hospitalized due to dengue who produced of different cytokines when PBMCs were stimulated DENV-NS3 overlapping peptides and JE Live vaccine.</p

Serotype specific dengue immune responses and disease severity.

<p>a) The association of the number of DENV serotypes individual responded to with age of the individual assessed by cultured ELISpot assays. b) The numbers of responses to DENV serotype specific peptides of those who were hospitalized due to dengue and those with past mild/sub clinical dengue infection assessed by cultured ELISpot assays. c) The association of DENV-NS3 specific IFNγ-producing ex vivo T cell responses with age (Spearmans r = 0.05, p = 0.42). d) Percentage of individuals who had been hospitalized due to dengue and those with past mild/sub clinical dengue infection who responded to different serotypes of the DENV assessed by cultured ELISpot assays.</p