{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,12]],"date-time":"2026-09-12T14:02:48Z","timestamp":1789221768933,"version":"build-2803163510"},"reference-count":40,"publisher":"Wiley","issue":"12","license":[{"start":{"date-parts":[[2022,11,13]],"date-time":"2022-11-13T00:00:00Z","timestamp":1668297600000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by-nc-nd\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100013041","name":"Vice-Chancellor for Research, Shiraz University of Medical Sciences","doi-asserted-by":"publisher","award":["93\u20107274"],"award-info":[{"award-number":["93\u20107274"]}],"id":[{"id":"10.13039\/501100013041","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100004320","name":"Shiraz University of Medical Sciences","doi-asserted-by":"publisher","id":[{"id":"10.13039\/501100004320","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":["onlinelibrary.wiley.com"],"crossmark-restriction":true},"short-container-title":["Clinical Laboratory Analysis"],"published-print":{"date-parts":[[2022,12]]},"abstract":"<jats:title>Abstract<\/jats:title><jats:sec><jats:title>Background<\/jats:title><jats:p>Fungal species are responsible for 40%\u201350% of all microbial keratitis cases. Due to the low amount of extracted DNA in ocular Formalin\u2010fixed Paraffin\u2010embedded (FFPE) samples, selecting a reliable molecular method is a substantial issue in this field.<\/jats:p><\/jats:sec><jats:sec><jats:title>Methods<\/jats:title><jats:p>Sixty\u2010six samples were collected via the penetrating keratoplasty (PK) technique. Histopathology assays were performed using hematoxylin\u2013eosin (H&amp;E) and periodic acid Schiff (PAS) staining methods. The ITS1\/ITS4 and ITS1\/ITS2 primer pairs were used in a semi\u2010nested polymerase chain reaction (PCR) to target the universal internal transcribed spacer (ITS) region. Some PCR results were validated through sequencing.<\/jats:p><\/jats:sec><jats:sec><jats:title>Results<\/jats:title><jats:p>Fungal DNA was detected in 44 of 66 samples (66.7%), and histopathology was positive for 41 of 66 samples (62.1%). Of 41 histopathologically proven fungal\u2010positive cases, 39 were PCR\u2010positive (95%). Moreover, of 44 PCR\u2010positive samples, 39 (88.6%) were histopathology\u2010positive, and 5 (11.3%) were histopathology\u2010negative. Totally in 39 cases (59%), both histopathology and PCR yielded positive results. The Kappa agreement rate between the two diagnostic methods, including histopathology and PCR, was 0.77. Sensitivity, specificity, positive predictive value, and false predictive value were reported as 88.64%, 90.9%, 95.12%, and 80%, respectively.<\/jats:p><\/jats:sec><jats:sec><jats:title>Conclusion<\/jats:title><jats:p>As we reached the acceptable Kappa agreement rate, we concluded that applying the semi\u2010nested PCR assay is a promising method for supporting the evidence by histopathology. Finally, we suggest targeting more specific gene regions using primer pairs that amplify smaller amplicon sizes and surveying novel molecular methods such as NGS to achieve higher sensitivity and Kappa agreement rates.<\/jats:p><\/jats:sec>","DOI":"10.1002\/jcla.24764","type":"journal-article","created":{"date-parts":[[2022,11,13]],"date-time":"2022-11-13T17:56:20Z","timestamp":1668362180000},"update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":2,"title":["Accompanying a semi\u2010nested <scp>PCR<\/scp> assay to support histopathology findings of fungal keratitis in formalin\u2010fixed paraffin\u2010embedded corneal samples"],"prefix":"10.1002","volume":"36","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-4987-8477","authenticated-orcid":false,"given":"Mohammad Javad","family":"Ashraf","sequence":"first","affiliation":[{"name":"Department of Pathology Shiraz University of Medical Sciences  Shiraz Iran"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-8208-189X","authenticated-orcid":false,"given":"Foroogh","family":"Shamsizadeh","sequence":"additional","affiliation":[{"name":"Department of Parasitology and Mycology, School of Medicine Shiraz University of Medical Sciences  Shiraz Iran"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-3569-0035","authenticated-orcid":false,"given":"Hamid","family":"Morovati","sequence":"additional","affiliation":[{"name":"Department of Parasitology and Mycology, School of Medicine Shiraz University of Medical Sciences  Shiraz Iran"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Safoora","family":"Hejazinia","sequence":"additional","affiliation":[{"name":"Department of Pathology Shiraz University of Medical Sciences  Shiraz Iran"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-8059-1060","authenticated-orcid":false,"given":"Mohammad","family":"Kord","sequence":"additional","affiliation":[{"name":"Department of Parasitology and Mycology, School of Medicine Shiraz University of Medical Sciences  Shiraz Iran"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-1623-5803","authenticated-orcid":false,"given":"Saham","family":"Ansari","sequence":"additional","affiliation":[{"name":"Department of Medical Parasitology and Mycology, School of Medicine Shahid Beheshti University of Medical Sciences  Tehran Iran"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-7809-4690","authenticated-orcid":false,"given":"Keyvan","family":"Pakshir","sequence":"additional","affiliation":[{"name":"Department of Parasitology and Mycology, School of Medicine Shiraz University of Medical Sciences  Shiraz Iran"},{"name":"Basic Sciences in Infectious Diseases Research Center Shiraz University of Medical Sciences  Shiraz Iran"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-4033-8992","authenticated-orcid":false,"given":"Golsa","family":"Shekarkhar","sequence":"additional","affiliation":[{"name":"Department of Pathology Shiraz University of Medical Sciences  Shiraz Iran"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-3338-7112","authenticated-orcid":false,"given":"Kamiar","family":"Zomorodian","sequence":"additional","affiliation":[{"name":"Department of Parasitology and Mycology, School of Medicine Shiraz University of Medical Sciences  Shiraz Iran"},{"name":"Basic Sciences in Infectious Diseases Research Center Shiraz University of Medical Sciences  Shiraz Iran"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"311","published-online":{"date-parts":[[2022,11,13]]},"reference":[{"key":"e_1_2_11_2_1","volume-title":"StatPearls [Internet]","author":"Castano G","year":"2021"},{"key":"e_1_2_11_3_1","doi-asserted-by":"publisher","DOI":"10.1111\/myc.12306"},{"key":"e_1_2_11_4_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.exer.2020.108372"},{"key":"e_1_2_11_5_1","doi-asserted-by":"publisher","DOI":"10.1016\/S1473-3099(20)30448-5"},{"key":"e_1_2_11_6_1","doi-asserted-by":"publisher","DOI":"10.1155\/2019\/6395840"},{"key":"e_1_2_11_7_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.ophtha.2006.05.035"},{"key":"e_1_2_11_8_1","doi-asserted-by":"publisher","DOI":"10.4103\/ijo.IJO_734_18"},{"key":"e_1_2_11_9_1","doi-asserted-by":"publisher","DOI":"10.7883\/yoken.JJID.2018.269"},{"key":"e_1_2_11_10_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.ajo.2008.04.019"},{"key":"e_1_2_11_11_1","doi-asserted-by":"publisher","DOI":"10.3390\/jof7040273"},{"key":"e_1_2_11_12_1","doi-asserted-by":"publisher","DOI":"10.4103\/0301-4738.53051"},{"key":"e_1_2_11_13_1","doi-asserted-by":"publisher","DOI":"10.1111\/j.1439-0507.2007.01480.x"},{"key":"e_1_2_11_14_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.jtos.2022.03.002"},{"key":"e_1_2_11_15_1","doi-asserted-by":"publisher","DOI":"10.3109\/02713683.2012.698357"},{"key":"e_1_2_11_16_1","doi-asserted-by":"publisher","DOI":"10.1007\/s12348-011-0019-9"},{"key":"e_1_2_11_17_1","doi-asserted-by":"publisher","DOI":"10.1111\/1469-0691.12126"},{"key":"e_1_2_11_18_1","doi-asserted-by":"publisher","DOI":"10.1007\/s12281-020-00374-7"},{"key":"e_1_2_11_19_1","doi-asserted-by":"publisher","DOI":"10.1128\/JCM.40.5.1644-1647.2002"},{"key":"e_1_2_11_20_1","doi-asserted-by":"publisher","DOI":"10.1186\/s12866-019-1426-6"},{"key":"e_1_2_11_21_1","doi-asserted-by":"publisher","DOI":"10.4103\/0377-4929.142673"},{"key":"e_1_2_11_22_1","doi-asserted-by":"publisher","DOI":"10.1128\/JCM.38.8.2940-2942.2000"},{"key":"e_1_2_11_23_1","doi-asserted-by":"publisher","DOI":"10.1111\/myc.12837"},{"key":"e_1_2_11_24_1","doi-asserted-by":"publisher","DOI":"10.1099\/jmm.0.001282"},{"key":"e_1_2_11_25_1","doi-asserted-by":"publisher","DOI":"10.1177\/0300985817698207"},{"key":"e_1_2_11_26_1","doi-asserted-by":"publisher","DOI":"10.1136\/jcp.2004.024703"},{"key":"e_1_2_11_27_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.prp.2016.02.010"},{"key":"e_1_2_11_28_1","doi-asserted-by":"publisher","DOI":"10.1007\/s10096-005-0078-7"},{"key":"e_1_2_11_29_1","doi-asserted-by":"publisher","DOI":"10.1128\/JCM.41.2.581-585.2003"},{"issue":"5","key":"e_1_2_11_30_1","first-page":"360","article-title":"Understanding interobserver agreement: the kappa statistic","volume":"37","author":"Viera AJ","year":"2005","journal-title":"Fam Med"},{"key":"e_1_2_11_31_1","volume-title":"StatPearls. Treasure Island (FL): StatPearls Publishing Copyright \u00a9 2020","author":"Castano G","year":"2020"},{"key":"e_1_2_11_32_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.ophtha.2010.03.048"},{"key":"e_1_2_11_33_1","doi-asserted-by":"publisher","DOI":"10.1159\/000313988"},{"issue":"6","key":"e_1_2_11_34_1","doi-asserted-by":"crossref","first-page":"2043","DOI":"10.1128\/JCM.02319-09","article-title":"Molecular detection and identification of zygomycetes species from paraffin\u2010embedded tissues in a murine model of disseminated zygomycosis: a collaborative European Society of Clinical Microbiology and Infectious Diseases (ESCMID) fungal infection study group (EFISG) evaluation","volume":"48","author":"Dannaoui E","year":"2010","journal-title":"J Clin Microbiol"},{"key":"e_1_2_11_35_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.funbio.2015.07.002"},{"key":"e_1_2_11_36_1","doi-asserted-by":"publisher","DOI":"10.1093\/mmy\/myz096"},{"key":"e_1_2_11_37_1","doi-asserted-by":"publisher","DOI":"10.1111\/1469-0691.12571"},{"key":"e_1_2_11_38_1","doi-asserted-by":"publisher","DOI":"10.1002\/jcla.24209"},{"key":"e_1_2_11_39_1","first-page":"773","volume-title":"Involvement of Fusarium Spp. in Fungal Keratiti","author":"D\u00f3czi I","year":"2004"},{"key":"e_1_2_11_40_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.jfo.2017.08.001"},{"key":"e_1_2_11_41_1","doi-asserted-by":"publisher","DOI":"10.1007\/s40588-020-00142-8"}],"container-title":["Journal of Clinical Laboratory Analysis"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/onlinelibrary.wiley.com\/doi\/pdf\/10.1002\/jcla.24764","content-type":"application\/pdf","content-version":"vor","intended-application":"text-mining"},{"URL":"https:\/\/onlinelibrary.wiley.com\/doi\/full-xml\/10.1002\/jcla.24764","content-type":"application\/xml","content-version":"vor","intended-application":"text-mining"},{"URL":"https:\/\/onlinelibrary.wiley.com\/doi\/pdf\/10.1002\/jcla.24764","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2023,8,20]],"date-time":"2023-08-20T11:22:48Z","timestamp":1692530568000},"score":1,"resource":{"primary":{"URL":"https:\/\/onlinelibrary.wiley.com\/doi\/10.1002\/jcla.24764"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2022,11,13]]},"references-count":40,"journal-issue":{"issue":"12","published-print":{"date-parts":[[2022,12]]}},"alternative-id":["10.1002\/jcla.24764"],"URL":"https:\/\/doi.org\/10.1002\/jcla.24764","archive":["Portico"],"relation":{},"ISSN":["0887-8013","1098-2825"],"issn-type":[{"value":"0887-8013","type":"print"},{"value":"1098-2825","type":"electronic"}],"subject":[],"published":{"date-parts":[[2022,11,13]]},"assertion":[{"value":"2022-08-09","order":0,"name":"received","label":"Received","group":{"name":"publication_history","label":"Publication History"}},{"value":"2022-10-31","order":1,"name":"accepted","label":"Accepted","group":{"name":"publication_history","label":"Publication History"}},{"value":"2022-11-13","order":2,"name":"published","label":"Published","group":{"name":"publication_history","label":"Publication History"}}],"article-number":"e24764"}}