{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,8,18]],"date-time":"2026-08-18T01:16:52Z","timestamp":1787015812179,"version":"build-2736575974"},"reference-count":46,"publisher":"Wiley","issue":"11","license":[{"start":{"date-parts":[[2024,11,1]],"date-time":"2024-11-01T00:00:00Z","timestamp":1730419200000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/onlinelibrary.wiley.com\/termsAndConditions#vor"}],"funder":[{"DOI":"10.13039\/501100011033","name":"Agencia Estatal de Investigaci\u00f3n","doi-asserted-by":"publisher","award":["PID2020\u2010114546RB"],"award-info":[{"award-number":["PID2020\u2010114546RB"]}],"id":[{"id":"10.13039\/501100011033","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":["onlinelibrary.wiley.com"],"crossmark-restriction":true},"short-container-title":["Mycoses"],"published-print":{"date-parts":[[2024,11]]},"abstract":"<jats:title>ABSTRACT<\/jats:title>\n                  <jats:sec>\n                    <jats:title>Background<\/jats:title>\n                    <jats:p>\n                      <jats:italic>Candida parapsilosis<\/jats:italic>\n                      is a pathogenic yeast that has reduced susceptibility to echinocandins and ranks as the second or third leading cause of candidaemia, depending on the geographical region. This yeast often causes nosocomial infections, which are frequently detected as outbreaks. In recent years, resistance to azoles in\n                      <jats:italic>C.\u2009parapsilosis<\/jats:italic>\n                      has increased globally, primarily due to the accumulation of mutations in the\n                      <jats:italic>ERG11<\/jats:italic>\n                      gene.\n                    <\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Objectives<\/jats:title>\n                    <jats:p>\n                      In this study, we have developed an assay based on real\u2010time PCR and high\u2010resolution melting (HRM) curve analysis to detect two of the most prevalent mutations at\n                      <jats:italic>ERG11<\/jats:italic>\n                      that confer resistance to fluconazole (Y132F and G458S).\n                    <\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Methods<\/jats:title>\n                    <jats:p>We designed allele\u2010specific oligonucleotides that selectively bind to either the wild type or mutated sequences and optimised the conditions to ensure amplification of the specific allele, followed by detection via high\u2010resolution melting (HRM) analysis.<\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Results<\/jats:title>\n                    <jats:p>\n                      The designed oligonucleotides to detect the Erg11\n                      <jats:sup>Y132F<\/jats:sup>\n                      and Erg11\n                      <jats:sup>G458S<\/jats:sup>\n                      mutations produced specific amplification of either WT or mutated alleles. We conducted a duplex real\u2010time PCR combining oligonucleotides for the wild\u2010type sequences in one mix, and oligonucleotides for the mutated alleles in another. Following this, we performed an analysis of the HRM curve to identify the amplified allele in each case. This technique was blindly evaluated on a set of 114\n                      <jats:italic>C.\u2009parapsilosis<\/jats:italic>\n                      isolates, all of which were unequivocally identified using our approach.\n                    <\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Conclusion<\/jats:title>\n                    <jats:p>\n                      This technique offers a new method for the early detection of azole resistance mechanism in\n                      <jats:italic>C.\u2009parapsilosis.<\/jats:italic>\n                    <\/jats:p>\n                  <\/jats:sec>","DOI":"10.1111\/myc.13811","type":"journal-article","created":{"date-parts":[[2024,11,1]],"date-time":"2024-11-01T23:35:38Z","timestamp":1730504138000},"update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":2,"title":["High\u2010Resolution Melting Assay to Detect the Mutations That Cause the\n                    <scp>Y132F<\/scp>\n                    and\n                    <scp>G458S<\/scp>\n                    Substitutions at the\n                    <i>ERG11<\/i>\n                    Gene Involved in Azole Resistance in\n                    <i>Candida parapsilosis<\/i>"],"prefix":"10.1111","volume":"67","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-3010-6610","authenticated-orcid":false,"given":"Nuria","family":"Trevijano\u2010Contador","sequence":"first","affiliation":[{"name":"Mycology Reference Laboratory National Centre for Microbiology, Instituto de Salud Carlos III  Madrid Spain"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-8024-3595","authenticated-orcid":false,"given":"Elena","family":"L\u00f3pez\u2010Peralta","sequence":"additional","affiliation":[{"name":"Mycology Reference Laboratory National Centre for Microbiology, Instituto de Salud Carlos III  Madrid Spain"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jorge","family":"L\u00f3pez\u2010L\u00f3pez","sequence":"additional","affiliation":[{"name":"Mycology Reference Laboratory National Centre for Microbiology, Instituto de Salud Carlos III  Madrid Spain"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0000-3196-2550","authenticated-orcid":false,"given":"Alejandra","family":"Rold\u00e1n","sequence":"additional","affiliation":[{"name":"Mycology Reference Laboratory National Centre for Microbiology, Instituto de Salud Carlos III  Madrid Spain"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Cristina","family":"de Armentia","sequence":"additional","affiliation":[{"name":"Mycology Reference Laboratory National Centre for Microbiology, Instituto de Salud Carlos III  Madrid 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