{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,6,26]],"date-time":"2026-06-26T05:22:15Z","timestamp":1782451335510,"version":"3.54.5"},"reference-count":31,"publisher":"American Society for Microbiology","issue":"3","license":[{"start":{"date-parts":[[2025,3,4]],"date-time":"2025-03-04T00:00:00Z","timestamp":1741046400000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"},{"start":{"date-parts":[[2025,3,4]],"date-time":"2025-03-04T00:00:00Z","timestamp":1741046400000},"content-version":"tdm","delay-in-days":0,"URL":"https:\/\/journals.asm.org\/non-commercial-tdm-license"}],"content-domain":{"domain":["journals.asm.org"],"crossmark-restriction":true},"short-container-title":["Microbiol Spectr"],"published-print":{"date-parts":[[2025,3,4]]},"abstract":"<jats:title>ABSTRACT<\/jats:title>\n          <jats:sec>\n            <jats:title\/>\n            <jats:p>\n              <jats:italic>Acanthamoeba<\/jats:italic>\n              keratitis (AK) is a sight-threatening corneal infection that is challenging to diagnose and treat due to the resistance of\n              <jats:italic>Acanthamoeba<\/jats:italic>\n              to standard antimicrobial agents. Current detection methods have limitations. This study aimed to develop and validate a sensitive viability PCR (v-PCR) assay using a photoreactive dye to distinguish viable from non-viable\n              <jats:italic>Acanthamoeba<\/jats:italic>\n              for rapid identification of viable\n              <jats:italic>Acanthamoeba<\/jats:italic>\n              trophozoites and cysts. Propidium monoazide (PMAxx) was used as a photoreactive dye. Mixtures containing decreasing percentages of viable\n              <jats:italic>Acanthamoeba<\/jats:italic>\n              , including reference strains\n              <jats:italic>Acanthamoeba polyphaga<\/jats:italic>\n              trophozoites and cysts,\n              <jats:italic>Acanthamoeba castellanii<\/jats:italic>\n              trophozoites, and\n              <jats:italic>Acanthamoeba castellanii<\/jats:italic>\n              trophozoites from a clinical sample, were prepared. Disinfectant efficacy against\n              <jats:italic>Acanthamoeba<\/jats:italic>\n              was also assessed. Samples were divided into PMAxx-treated and non-PMAxx-treated parts, and v-PCR assay was applied to both. The difference in viable\n              <jats:italic>Acanthamoeba<\/jats:italic>\n              was determined by subtracting the cycle threshold (Ct) value of the PMAxx-treated sample from the non-PMAxx-treated sample. Mixtures with decreasing concentrations of viable\n              <jats:italic>Acanthamoeba<\/jats:italic>\n              trophozoites and cysts showed increasingly lower delta Ct values as the percentage of viable\n              <jats:italic>Acanthamoeba<\/jats:italic>\n              decreased, as expected. This relationship was observed across all tested samples. Menicon Progent effectively eliminated\n              <jats:italic>A. polyphaga<\/jats:italic>\n              trophozoites and cysts, while propamidine, chlorhexidine, or their combination resulted in approximately 2-log reductions in\n              <jats:italic>A. polyphaga<\/jats:italic>\n              trophozoites and cysts. In the current study, a rapid v-PCR assay was developed that can distinguish between viable and non-viable\n              <jats:italic>Acanthamoeba<\/jats:italic>\n              , for both trophozoites and cysts, across multiple species. The presence of viable\n              <jats:italic>Acanthamoeba<\/jats:italic>\n              , as determined by v-PCR, allows monitoring of treatment response and efficacy in AK.\n            <\/jats:p>\n            <jats:sec>\n              <jats:title>IMPORTANCE<\/jats:title>\n              <jats:p>\n                The development of a sensitive viability PCR (v-PCR) assay using propidium monoazide (PMAxx) as a photoreactive dye marks a significant advancement in the diagnosis and treatment of\n                <jats:italic>Acanthamoeba<\/jats:italic>\n                keratitis (AK), a severe corneal infection notorious for its resistance to conventional antimicrobials. This innovative assay offers a rapid and accurate method to distinguish viable from non-viable\n                <jats:italic>Acanthamoeba<\/jats:italic>\n                trophozoites and cysts, addressing a critical need in the field. By effectively distinguishing between viable and non-viable\n                <jats:italic>Acanthamoeba<\/jats:italic>\n                , this test enables monitoring of treatment response and efficacy, essential for guiding clinical interventions in AK cases. The successful validation of this v-PCR assay across various\n                <jats:italic>Acanthamoeba<\/jats:italic>\n                species and its ability to assess disinfectant efficacy further underline its potential as a valuable tool for improving diagnostic precision and therapeutic outcomes in the treatment of AK.\n              <\/jats:p>\n            <\/jats:sec>\n          <\/jats:sec>","DOI":"10.1128\/spectrum.01811-24","type":"journal-article","created":{"date-parts":[[2025,2,7]],"date-time":"2025-02-07T14:01:11Z","timestamp":1738936871000},"update-policy":"https:\/\/doi.org\/10.1128\/asmj-crossmark-policy-page","source":"Crossref","is-referenced-by-count":4,"title":["Enhancing\n            <i>Acanthamoeba<\/i>\n            diagnostics: rapid detection of viable\n            <i>Acanthamoeba<\/i>\n            trophozoites and cysts using viability PCR assay"],"prefix":"10.1128","volume":"13","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-0469-5781","authenticated-orcid":true,"given":"J. M. J.","family":"Veugen","sequence":"first","affiliation":[{"name":"University Eye Clinic Maastricht, Maastricht University Medical Center+","place":["Maastricht, the Netherlands"]},{"name":"School for Mental Health and Neuroscience (MHeNs), Maastricht University","place":["Maastricht, the Netherlands"]},{"name":"Department of Medical Microbiology, Infectious Diseases & Infection Prevention, Maastricht University Medical Center+","place":["Maastricht, the Netherlands"]},{"name":"Care and Public Health Research Institute (CAPHRI), Maastricht University","place":["Maastricht, the Netherlands"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"P. H. M.","family":"Savelkoul","sequence":"additional","affiliation":[{"name":"Department of Medical Microbiology, Infectious Diseases & Infection Prevention, Maastricht University Medical Center+","place":["Maastricht, the Netherlands"]},{"name":"Care and Public Health Research Institute (CAPHRI), Maastricht University","place":["Maastricht, the Netherlands"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"R. M. M. A.","family":"Nuijts","sequence":"additional","affiliation":[{"name":"University Eye Clinic Maastricht, Maastricht University Medical Center+","place":["Maastricht, the Netherlands"]},{"name":"School for Mental Health and Neuroscience (MHeNs), Maastricht University","place":["Maastricht, the Netherlands"]},{"name":"Department of Ophthalmology, Zuyderland Medical Center","place":["Heerlen, the Netherlands"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"M. M.","family":"Dickman","sequence":"additional","affiliation":[{"name":"University Eye Clinic Maastricht, Maastricht University Medical Center+","place":["Maastricht, the Netherlands"]},{"name":"School for Mental Health and Neuroscience (MHeNs), Maastricht University","place":["Maastricht, the Netherlands"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-5326-3985","authenticated-orcid":true,"given":"P. F. G.","family":"Wolffs","sequence":"additional","affiliation":[{"name":"Department of Medical Microbiology, Infectious Diseases & Infection Prevention, Maastricht University Medical Center+","place":["Maastricht, the Netherlands"]},{"name":"Care and Public Health Research Institute (CAPHRI), Maastricht University","place":["Maastricht, the Netherlands"]}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"235","reference":[{"key":"e_1_3_3_2_2","doi-asserted-by":"publisher","DOI":"10.1128\/CMR.16.2.273-307.2003"},{"key":"e_1_3_3_3_2","doi-asserted-by":"publisher","DOI":"10.1097\/ICO.0000000000000804"},{"key":"e_1_3_3_4_2","doi-asserted-by":"publisher","DOI":"10.1111\/j.1574-6976.2006.00023.x"},{"key":"e_1_3_3_5_2","doi-asserted-by":"publisher","DOI":"10.1186\/1756-3305-5-6"},{"key":"e_1_3_3_6_2","doi-asserted-by":"publisher","DOI":"10.1007\/s00284-001-0050-4"},{"key":"e_1_3_3_7_2","doi-asserted-by":"publisher","DOI":"10.1016\/s0140-6736(74)90285-2"},{"key":"e_1_3_3_8_2","first-page":"221","article-title":"Acanthamoeba polyphaga keratitis and Acenthamoeba uveitis associated with fatal meningoencephalitis","volume":"95","author":"Jones DB","year":"1975","unstructured":"Jones DB, Visvesvara GS, Robinson NM. 1975. Acanthamoeba polyphaga keratitis and Acenthamoeba uveitis associated with fatal meningoencephalitis. Trans Ophthalmol Soc U K (1962) 95:221\u2013232.","journal-title":"Trans Ophthalmol Soc U K (1962)"},{"key":"e_1_3_3_9_2","doi-asserted-by":"publisher","DOI":"10.1111\/j.1750-3639.1997.tb01076.x"},{"key":"e_1_3_3_10_2","doi-asserted-by":"publisher","DOI":"10.3390\/pathogens10030323"},{"key":"e_1_3_3_11_2","doi-asserted-by":"publisher","DOI":"10.1001\/jama.2021.0424"},{"key":"e_1_3_3_12_2","doi-asserted-by":"publisher","DOI":"10.1111\/ceo.14113"},{"key":"e_1_3_3_13_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.clae.2019.04.009"},{"key":"e_1_3_3_14_2","doi-asserted-by":"publisher","DOI":"10.4103\/0301-4738.40360"},{"key":"e_1_3_3_15_2","doi-asserted-by":"publisher","DOI":"10.1016\/S2214-109X(20)30488-5"},{"key":"e_1_3_3_16_2","doi-asserted-by":"publisher","DOI":"10.1097\/ICO.0000000000001497"},{"key":"e_1_3_3_17_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.mimet.2005.04.008"},{"key":"e_1_3_3_18_2","doi-asserted-by":"publisher","DOI":"10.1186\/s13104-017-2666-x"},{"key":"e_1_3_3_19_2","doi-asserted-by":"publisher","DOI":"10.1128\/JCM.00875-06"},{"key":"e_1_3_3_20_2","doi-asserted-by":"publisher","DOI":"10.1128\/JCM.00908-08"},{"key":"e_1_3_3_21_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.mimet.2006.04.015"},{"key":"e_1_3_3_22_2","doi-asserted-by":"publisher","DOI":"10.1186\/1471-2180-13-216"},{"key":"e_1_3_3_23_2","doi-asserted-by":"publisher","DOI":"10.1371\/journal.pone.0165920"},{"key":"e_1_3_3_24_2","doi-asserted-by":"publisher","DOI":"10.3389\/fmicb.2020.01911"},{"key":"e_1_3_3_25_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.scitotenv.2021.149085"},{"key":"e_1_3_3_26_2","doi-asserted-by":"publisher","DOI":"10.1111\/j.1550-7408.2011.00557.x"},{"key":"e_1_3_3_27_2","doi-asserted-by":"publisher","DOI":"10.1128\/spectrum.00160-24"},{"key":"e_1_3_3_28_2","doi-asserted-by":"publisher","DOI":"10.1097\/ICL.0000000000000925"},{"key":"e_1_3_3_29_2","unstructured":"Nederlandse Vereniging voor Parasitologie NVP. 2020. Therapierichtlijn parasitaire infecties. Available from: https:\/\/adult.nl.antibiotica.app\/sites\/default\/files\/2020-08\/NVP%20therapierichtlijn%202020_15.pdf"},{"key":"e_1_3_3_30_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.ophtha.2014.04.013"},{"key":"e_1_3_3_31_2","doi-asserted-by":"publisher","DOI":"10.1016\/S0161-6420(03)00481-0"},{"key":"e_1_3_3_32_2","doi-asserted-by":"publisher","DOI":"10.1128\/AAC.00686-16"}],"container-title":["Microbiology Spectrum"],"original-title":[],"contributor":[{"given":"Pieter W.","family":"Smit","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"reviewer"}]},{"given":"Mehmet","family":"Aykur","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"reviewer"}]}],"language":"en","link":[{"URL":"https:\/\/journals.asm.org\/doi\/pdf\/10.1128\/spectrum.01811-24","content-type":"application\/pdf","content-version":"vor","intended-application":"text-mining"},{"URL":"https:\/\/journals.asm.org\/doi\/pdf\/10.1128\/spectrum.01811-24","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,3,4]],"date-time":"2025-03-04T14:11:20Z","timestamp":1741097480000},"score":1,"resource":{"primary":{"URL":"https:\/\/journals.asm.org\/doi\/10.1128\/spectrum.01811-24"}},"subtitle":[],"editor":[{"given":"Denis","family":"Sereno","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"editor"}]}],"short-title":[],"issued":{"date-parts":[[2025,3,4]]},"references-count":31,"journal-issue":{"issue":"3","published-print":{"date-parts":[[2025,3,4]]}},"alternative-id":["10.1128\/spectrum.01811-24"],"URL":"https:\/\/doi.org\/10.1128\/spectrum.01811-24","relation":{},"ISSN":["2165-0497"],"issn-type":[{"value":"2165-0497","type":"electronic"}],"subject":[],"published":{"date-parts":[[2025,3,4]]},"assertion":[{"value":"2024-09-09","order":0,"name":"received","label":"Received","group":{"name":"publication_history","label":"Publication History"}},{"value":"2025-01-13","order":2,"name":"accepted","label":"Accepted","group":{"name":"publication_history","label":"Publication History"}},{"value":"2025-02-07","order":3,"name":"published","label":"Published","group":{"name":"publication_history","label":"Publication History"}}],"article-number":"e01811-24"}}