{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"institution":[{"name":"bioRxiv"}],"indexed":{"date-parts":[[2026,7,30]],"date-time":"2026-07-30T17:08:55Z","timestamp":1785431335329,"version":"3.56.0"},"posted":{"date-parts":[[2026,6,15]]},"group-title":"Microbiology","reference-count":63,"publisher":"openRxiv","license":[{"start":{"date-parts":[[2026,6,15]],"date-time":"2026-06-15T00:00:00Z","timestamp":1781481600000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by-nc-nd\/4.0\/"}],"funder":[{"award":["GM132037"],"award-info":[{"award-number":["GM132037"]}],"id":[{"id":"https:\/\/ror.org\/01cwqze88","id-type":"ROR","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"accepted":{"date-parts":[[2026,6,15]]},"abstract":"<jats:title>Abstract<\/jats:title>\n                <jats:p>\n                  <jats:italic>Trichomonas vaginalis<\/jats:italic>\n                  is the causative agent of trichomoniasis, the most common non-viral sexually transmitted infection (STI). Despite its prevalence, low levels of public knowledge and research funding and the absence of\n                  <jats:italic>T. vaginalis<\/jats:italic>\n                  screening or control programs have led to its categorization as a \u201cneglected\u201d STI. Unlike other STIs in the USA, prevalence increases with age, peaking among individuals in their 40s. Both a motile trophozoite stage and a non-motile pseudocyst state have been described for the parasite, although it is debated whether the latter is a quiescent stage or a degenerate form on its way to cell death. Here we characterize the\n                  <jats:italic>T. vaginalis<\/jats:italic>\n                  pseudocyst by flow cytometry, membrane integrity assays, transcriptomics, and reversion studies. Pseudocysts were induced by culturing trophozoites in acidic media or in iron depleted media, with a variety of resulting survival rates. Flow cytometry studies showed that pseudocysts have intact cell membranes and express phosphatidylserine on their cell surface. Fluorescence-activated cell sorting studies also identified distinct sub-populations of parasites, revealing the importance of using pure live pseudocyst cultures in reversion studies. Pseudocysts were transcriptionally active for several days and had consistent subsets of genes with increased expression compared to trophozoites, although decreased transcription of genes involved in metabolism. Comparative transcriptomics of pseudocysts and trophozoites of two\n                  <jats:italic>T. vaginalis<\/jats:italic>\n                  strains revealed distinct cell states. Combined, our results provide evidence that the pseudocyst cell state is a stress-induced quiescent stage of\n                  <jats:italic>T. vaginalis<\/jats:italic>\n                  that can remain viable for days, with implications for a role in persistent infections.\n                <\/jats:p>\n                <jats:sec>\n                  <jats:title>Author Summary<\/jats:title>\n                  <jats:p>\n                    The sexually transmitted parasite\n                    <jats:italic>Trichomonas vaginalis<\/jats:italic>\n                    has two well-known cell forms: a free-swimming, flagellated trophozoite and an amoeboid form adhered to host epithelia. A third morphology, the pseudocyst, has been described, but it is unclear whether this is a quiescent stage capable of facilitating persistent infections, or a degenerate form indicating cell death. Here we describe experiments revealing that pseudocysts have overall decreased gene expression compared to trophozoites but exhibit stage-specific gene transcription and plasma membrane integrity for days. Moreover, pseudocysts maintain externalized phosphatidylserine for multiple days. Taken together, our results suggest that pseudocysts are a viable cell stage in\n                    <jats:italic>T. vaginalis<\/jats:italic>\n                    distinct from cell death. Further research into pseudocysts and particularly their interactions with host immune cells is needed to reveal what role they may play in\n                    <jats:italic>T. vaginalis<\/jats:italic>\n                    pathogenesis and persistent or asymptomatic infections.\n                  <\/jats:p>\n                <\/jats:sec>","DOI":"10.64898\/2026.06.14.730637","type":"posted-content","created":{"date-parts":[[2026,6,16]],"date-time":"2026-06-16T05:55:15Z","timestamp":1781589315000},"source":"Crossref","is-referenced-by-count":0,"title":["<i>Trichomonas vaginalis<\/i>\n                  pseudocysts are a quiescent cell stage characterized by a differentially regulated transcriptome and intact membranes"],"prefix":"10.64898","author":[{"given":"Mari","family":"Shiratori","sequence":"first","affiliation":[{"name":"Center for Genomics and Systems Biology, Department of Biology, New York University, New York City, NY, United States of America"},{"name":"Department of Molecular Microbiology and Immunology, Bloomberg School of Public Health, Johns Hopkins University, Baltimore, MD 21205, United States of America"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Francisco","family":"Callejas-Hern\u00e1ndez","sequence":"additional","affiliation":[{"name":"Department of Molecular Microbiology and Immunology, Bloomberg School of Public Health, Johns Hopkins University, Baltimore, MD 21205, United States of America"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jordan C.","family":"Orosco","sequence":"additional","affiliation":[{"name":"Department of Molecular Microbiology and Immunology, Bloomberg School of Public Health, Johns Hopkins University, Baltimore, MD 21205, United States of America"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Steven A.","family":"Sullivan","sequence":"additional","affiliation":[{"name":"Department of Molecular Microbiology and Immunology, Bloomberg School of Public Health, Johns Hopkins University, Baltimore, MD 21205, United States of America"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-5238-6852","authenticated-orcid":false,"given":"Carlos","family":"Carmona Fontaine","sequence":"additional","affiliation":[{"name":"Center for Genomics and Systems Biology, Department of Biology, New York University, New York City, NY, United States of America"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-9340-5020","authenticated-orcid":false,"given":"Jane M.","family":"Carlton","sequence":"additional","affiliation":[{"name":"Department of Molecular Microbiology and Immunology, Bloomberg School of Public Health, Johns Hopkins University, Baltimore, MD 21205, United States of America"},{"name":"Department of Biomedical Engineering, Whiting School of Engineering, Johns Hopkins University, Baltimore, MD 21218 USA"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"54368","reference":[{"key":"2026061704500783000_2026.06.14.730637v1.1","unstructured":"Recommendations for the treatment of Trichomonas vaginalis, Mycoplasma genitalium, Candida albicans, bacterial vaginosis and human papillomavirus (anogenital warts). 2024, Geneva: World Health Organization."},{"key":"2026061704500783000_2026.06.14.730637v1.2","doi-asserted-by":"publisher","DOI":"10.1128\/JCM.00748-12"},{"key":"2026061704500783000_2026.06.14.730637v1.3","doi-asserted-by":"publisher","DOI":"10.1136\/sextrans-2013-051075"},{"key":"2026061704500783000_2026.06.14.730637v1.4","doi-asserted-by":"crossref","unstructured":"Gashi, A.M. , Trichomonas Vaginalis-A Clinical Image. 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