CT694 and pgp3 as serological tools for monitoring trachoma programs

Abstract

Background: Defining endpoints for trachoma programs can be a challenge as clinical signs of infection may persist in the absence of detectable bacteria. Antibody-based tests may provide an alternative testing strategy for surveillance during terminal phases of the program. Antibody-based assays, in particular ELISAs, have been shown to be useful to document C. trachomatis genital infections, but have not been explored extensively for ocular C. trachomatis infections.

Methodology/principal findings: An antibody-based multiplex assay was used to test two C. trachomatis antigens, pgp3 and CT694, for detection of trachoma antibodies in bloodspots from Tanzanian children (n = 160) collected after multiple rounds of mass azithromycin treatment. Using samples from C. trachomatis-positive (by PCR) children from Tanzania (n = 11) and control sera from a non-endemic group of U.S. children (n = 122), IgG responses to both pgp3 and CT694 were determined to be 91% sensitive and 98% specific. Antibody responses of Tanzanian children were analyzed with regard to clinical trachoma, PCR positivity, and age. In general, children with more intense ocular pathology (TF/TI = 2 or most severe) had a higher median antibody response to pgp3 (p = 0.0041) and CT694 (p = 0.0282) than those with normal exams (TF/TI = 0). However, 44% of children with no ocular pathology tested positive for antibody, suggesting prior infection. The median titer of antibody responses for children less than three years of age was significantly lower than those of older children. (p<0.0001 for both antigens).

Conclusions/significance: The antibody-based multiplex assay is a sensitive and specific additional tool for evaluating trachoma transmission. The assay can also be expanded to include antigens representing different diseases, allowing for a robust assay for monitoring across NTD programs.

Figure 1. Antibody response to pgp3 and CT694 by country.

Median responses are shown in median fluorescence intensity minus background (MFI-BG) by country for (A) pgp3 and (B) CT694. Sera from Haitian children (n = 86) and from United States children (n = 122) were used.

Figure 2. Antibody response to pgp3 and CT694 by village.

Median responses are shown in median fluorescence intensity minus background (MFI-BG) by village for (A) pgp3 and (B) CT694.

Figure 3. Antibody response by clinical diagnosis and PCR.

Median responses are shown in median fluorescence intensity minus background (MFI-BG) for antibody response to (A) pgp3 and (B) CT694 in relation to clinical diagnosis. Antibody response to (C) pgp3 and (D) CT694 is shown by PCR positivity. Responses shown in red indicate PCR positivity. Antibody responses to pgp3 differed between children with no clinical signs and PCR positive children (p = 0.0008) and between children with no clinical signs and those with a TF/TI score of 2 (p = 0.0041). For CT694, antibody responses for children with no clinical signs were lower than for PCR positive children (p = 0.0024) and for those with a TF/TI score of 2 (p = 0.0282).

Figure 4. Antibody response in association with age.

Median responses are shown in median fluorescence intensity minus background (MFI-BG) for (A) pgp3 and (B) CT694 antibody positive responses by age ranges of less than 3 years old (n = 42), between 3 and less than 6 years old (n = 65), and 6 years or older (n = 52). Responses shown in red indicate PCR positivity. Responses in green are children with ocular pathology (shown only in the <3 age group). Children with both trachoma and PCR positivity are indicated by a red dot with a green border. For pgp3, children younger than three years of age had lower antibody responses than those who were between three and six and those who were older than six years of age (p<0.0001 and p = 0.0062, respectively). For CT694, children younger than three years of age had lower antibody responses than children between three and six (p = 0.0291) and children older than six (p = 0.0001).