Behavioral, virologic, and immunologic factors associated with acquisition and severity of primary Epstein-Barr virus infection in university students

Abstract

Background: University students were studied prospectively to determine the incidence of and risk factors for acquisition of primary Epstein-Barr virus (EBV) infection and the virologic and immune correlates of disease severity.

Methods: EBV antibody-negative freshmen participated in monthly surveillance until graduation. If antibodies developed, proximate samples were assayed for viral load by polymerase chain reaction. Lymphocyte and natural killer (NK) cell numbers and activation were measured by flow cytometry, and plasma cytokine levels were measured by a multiplex assay.

Results: Of 546 students screened, 202 (37%) were antibody negative; 143 antibody-negative students were enrolled. During a median of 3 years of observation, 66 subjects experienced primary infection. Of these, 77% had infectious mononucleosis, 12% had atypical symptoms, and 11% were asymptomatic. Subjects reporting deep kissing with or without coitus had the same higher risk of infection than those reporting no kissing (P < .01). Viremia was transient, but median oral shedding was 175 days. Increases were observed in numbers of NK cells and CD8(+) T-cells but not in numbers of CD4(+) T-cells during acute infection. Severity of illness correlated positively with both blood EBV load (P = .015) and CD8(+) lymphocytosis (P = .0003).

Conclusions: Kissing was a significant risk for primary EBV infection. A total of 89% of infections were symptomatic, and blood viral load and CD8(+) lymphocytosis correlated with disease severity.

Figure 1.

Acquisition of primary Epstein–Barr virus (EBV) infection among 143 EBV-naive students followed prospectively throughout their undergraduate years. The white background represents active school semesters, and the gray areas are semester breaks. Vertical dashes mark censored times, which are the end of follow-up for students with no EBV infection. The incidence of infection during the freshman years (26 cases per 100 person-years) was more than twice the mean incidence during the following 3 years (10 cases per 100 person-years; P = .002).

Figure 2.

Kaplan–Meier estimate of time to incident primary Epstein–Barr virus (EBV) infection, according to self-reported sexual activity status: 68 students reported sexual activity (black line), 39 reported kissing only (gray line), and 11 reported no kissing or sexual activity (dashed line); 25 students did not provide this information. Vertical dashes mark censored times, which are the end of follow-up for students with no EBV infection. Subjects reporting deep kissing with or without coitus were significantly different from the group of subjects who reported no kissing and no coitus and experienced no primary EBV infections (P < .01).

Figure 3.

Severity and duration of illness in primary Epstein–Barr virus (EBV) infection. A, Severity of illness (SOI) score for 66 students with primary EBV infection. The SOI score (0–6) was calculated as described in the Methods. B, SOI score and days of symptoms for each subject. The average duration of symptoms for each SOI group is shown in bars. The averages for patients with SOI scores of 2–6 were significantly longer than averages for patients with a SOI score of 1 (P < .05).

Figure 4.

Kinetic analysis of immunologic and virologic changes during primary Epstein–Barr virus (EBV) infection. Whole blood from all symptomatic subjects at time points from 50 days before to 100 days after symptom onset was evaluated. The numbers of CD3⁺CD8⁺ lymphocytes (A), CD3⁺CD4⁺ lymphocytes (B), and CD3⁻CD56⁺ natural killer cells (C), are shown. D, Viral loads among symptomatic subjects. Black boxes mark the first 2 weeks after onset of symptoms.

Figure 5.

CD8⁺ but not CD4⁺ T-cell activation and blood (but not oral) viral load were associated with more severe symptoms. The severity of illness (SOI) score correlated with numbers of CD8⁺ (A) or CD4⁺ (B) lymphocytes or with viral load measured in the blood (C) or oral wash (D) among 59 subjects with symptomatic primary Epstein–Barr virus infection. The Spearman rank correlation coefficient and P values are shown. Abbreviation: NS, not statistically significant.

Figure 6.

Interleukin 6 (IL-6) but not interferon γ (IFN-γ) correlated with severity of illness (SOI) score. Serum cytokines were measured in select subjects at time points before, during, and after primary Epstein–Barr virus infection. Data for IL-6 (A) and IFN-γ (B) are shown. Boxes mark the first 2 weeks after the onset of symptoms. C and D, Average values from subjects with mild- to-moderate symptoms (SOI score, 0–3) or severe symptoms (SOI score, 4–6). Error bars indicate SDs. Dotted lines indicate baseline values.