Viability PCR shows that non-ocular surfaces could contribute to transmission of Chlamydia trachomatis infection in trachoma

Background The presence of Chlamydia trachomatis (Ct) DNA at non-ocular sites suggests that these sites may represent plausible routes of Ct transmission in trachoma. However, qPCR cannot discriminate between DNA from viable and non-viable bacteria. Here we use a propodium monoazide based viability PCR to investigate how long Ct remains viable at non-ocular sites under laboratory-controlled conditions. Methods Cultured Ct stocks (strain A2497) were diluted to final concentrations of 1000, 100, 10 and 1 omcB copies/μL and applied to plastic, woven mat, cotton cloth and pig skin. Swabs were then systemically collected from each surface and tested for the presence Ct DNA using qPCR. If Ct DNA was recovered, Ct viability was assessed over time by spiking multiple areas of the same surface type with the same final concentrations. Swabs were collected from each surface at 0, 2, 4, 6, 8 and 24 hours after spiking. Viability PCR was used to determine Ct viability at each timepoint. Results We were able to detect Ct DNA on all surfaces except the woven mat. Total Ct DNA remained detectable and stable over 24 hours for all concentrations applied to plastic, pig skin and cotton cloth. The amount of viable Ct decreased over time. For plastic and skin surfaces, only those where concentrations of 100 or 1000 omcB copies/μL were applied still had viable loads detectable after 24 hours. Cotton cloth showed a more rapid decrease and only those where concentrations of 1000 omcB copies/μL were applied still had viable DNA detectable after 24 hours. Conclusion Plastic, cotton cloth and skin may contribute to transmission of the Ct strains that cause trachoma, by acting as sites where reservoirs of bacteria are deposited and later collected and transferred mechanically into previously uninfected eyes.
Author summary Trachoma elimination efforts are hampered by limited understanding of Ct transmission routes. We have recently demonstrated the presence of Ct DNA at non-ocular sites in individuals living in households in Ethiopia where at least one resident had an ocular Ct infection detectable by quantitative PCR (qPCR). Ct DNA was most frequently detected on faces, hands and clothing, being found in such locations in 10–16% of samples tested. However, qPCR cannot discriminate between DNA from viable and non-viable organisms, and potentially misinform our understanding of Ct transmission routes. In this study, we used a propidium monoazide based viability PCR to investigate how long Ct remains viable on non-ocular sites by spiking different surfaces including pig skin, plastic and cotton cloth. These surfaces mimic non-ocular sites previously found to be positive for Ct DNA using standard qPCR. The results of our study show that viable Ct DNA could be recovered from plastic, cotton cloth and skin surfaces for up to 24 hours suggesting that these surfaces a role in ocular Ct transmission.