Reliable and specific detection of Acanthamoeba spp. in dishcloths using quantitative real-time PCR assay

• Published in: Food microbiology 2024-09, Vol.122, p.104562-104562, Article 104562
• Main Authors: Reyes-Batlle, M., Córdoba-Lanús, E., Domínguez-de-Barros, A., Sifaoui, I., Rodríguez-Expósito, R.L., Mantesa-Rodríguez, S., Piñero, J.E., Lorenzo-Morales, J.
• Format: Article
• Language: English
• Subjects: Acanthamoeba; Acanthamoeba - genetics; Acanthamoeba - isolation & purification; Dishcloths; DNA, Protozoan - analysis; DNA, Protozoan - genetics; Foodborne; Humans; Kitchen; q-PCR; Real-Time Polymerase Chain Reaction - methods; Sensitivity and Specificity
• ISSN: 0740-0020; 1095-9998
• DOI: 10.1016/j.fm.2024.104562

Acanthamoeba spp., are ubiquitous protist which belongs to Free-Living Amoeba (FLA) group, is considered as causal agent of side-threatening keratitis or fatal encephalitis among other human infections. Besides, this parasite has been reported as host for other microorganisms important to human health such as Campylobacter spp. or Vibrio spp. among others. This role of Acanthamoeba as pathogen and environmental phagocyte has increased the reports confirming its presence in human related environments, acting as a water quality indicator. Considering the tide relationship between water and kitchen environments, and the high prevalence of Acanthamoeba in water sources, the present study aims to establish a quick and accurate protocol based on DNA extraction and a real time qPCR assay to detect Acanthamoeba spp. in dishcloths. The procedure has been validated by processing 17 used dishcloths. Our findings demonstrated the high sensitivity of the qPCR assay used which was capable of detecting up to one Acanthamoeba from an in vitro contaminated dishcloth. The protocol accurately detected 64.7% of positive samples for Acanthamoeba spp, (in 4 samples DNA concentrations corresponded to 1-102 amoebae). Our findings demonstrate the importance of FLA surveillance by efficient and sensitive methods since one amoeba is capable of colonizing human related food environments such as kitchens sinks and could be a potential source of infection. [Display omitted] •The developed protocol was demonstrated to be a robust DNA extraction method from dishcloths.•The employed qPCR can detect up to one Acanthamoeba in the in vitro contaminated dishcloths.•From the 17 previously used dishcloths, Acanthamoeba spp. DNA was detected in almost the 65% of the samples.•The Ct values from 6 of the evaluated dishcloths corresponded to 1–100 Acanthamoeba spp. in 1 cm2 of dishcloths.