{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,5]],"date-time":"2026-09-05T00:36:14Z","timestamp":1788568574419,"version":"build-2803163510"},"reference-count":58,"publisher":"MDPI AG","issue":"19","license":[{"start":{"date-parts":[[2025,9,26]],"date-time":"2025-09-26T00:00:00Z","timestamp":1758844800000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100003593","name":"Conselho Nacional de Desenvolvimento Cient\u00edfico e Tecnol\u00f3gico","doi-asserted-by":"crossref","award":["402387\/2020-1"],"award-info":[{"award-number":["402387\/2020-1"]}],"id":[{"id":"10.13039\/501100003593","id-type":"DOI","asserted-by":"crossref"}]},{"DOI":"10.13039\/501100002322","name":"Coordena\u00e7\u00e3o de Aperfei\u00e7oamento de Pessoal de Nivel Superior","doi-asserted-by":"crossref","award":["Financial Code 01"],"award-info":[{"award-number":["Financial Code 01"]}],"id":[{"id":"10.13039\/501100002322","id-type":"DOI","asserted-by":"crossref"}]},{"DOI":"10.13039\/501100004612","name":"Funda\u00e7\u00e3o Arauc\u00e1ria-PR agreement","doi-asserted-by":"publisher","award":["2019\/2023 PDI"],"award-info":[{"award-number":["2019\/2023 PDI"]}],"id":[{"id":"10.13039\/501100004612","id-type":"DOI","asserted-by":"publisher"}]},{"name":"Superintend\u00eancia de Ci\u00eancia, Tecnologia e Ensino Superior (SETI)\/UEL","award":["Edital PROPPG 06\/2025"],"award-info":[{"award-number":["Edital PROPPG 06\/2025"]}]}],"content-domain":{"domain":["www.mdpi.com"],"crossmark-restriction":true},"short-container-title":["IJMS"],"abstract":"<jats:p>Candida species are the primary fungal pathogens of invasive infections associated with high morbidity and mortality. The identification of these microorganisms is critical for therapeutic management and control of hospital infection. Herein, assays targeting the Intergenic Spacer 2 (IGS2) and Internal Transcribed Spacer 1 (ITS1) from the rDNA locus were developed to differentiate Candida species. Based on consensus nucleotide sequences, specific primers and positive controls were designed, and standard PCR and real-time PCR (qPCR) assays were performed. All primers resulted in specific amplification of the molecular targets of each species with no amplifications of the negative template control. Furthermore, the primers were highly specific when tested with a range of fungal DNAs and no cross-reactivity was observed among Candida species. The assays presented a limit of detection (LoD) of 10 copies of positive control per reaction for all specific primers designed. Overall, our results showed that qPCR assays employing primers targeting the regions IGS2 and ITS1 completely differentiated between Candida albicans, Candida auris, Candida parapsilosis, Candida tropicalis, and Nakazeomyces glabratus, with great accuracy and no amplification of DNA from other fungal species.<\/jats:p>","DOI":"10.3390\/ijms26199411","type":"journal-article","created":{"date-parts":[[2025,9,26]],"date-time":"2025-09-26T09:35:49Z","timestamp":1758879349000},"page":"9411","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":2,"title":["Development of Melting-Curve-Based Real-Time PCR for Differentiating Medically Important Candida Species"],"prefix":"10.3390","volume":"26","author":[{"given":"Eliandro Reis","family":"Tavares","sequence":"first","affiliation":[{"name":"Departamento de Microbiologia, Centro de Ci\u00eancias Biol\u00f3gicas, Universidade Estadual de Londrina, Londrina CEP 86057-970, PR, Brazil"},{"name":"Escola de Medicina, Pontif\u00edcia Universidade Cat\u00f3lica do Paran\u00e1, Londrina CEP 86067-000, PR, Brazil"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Virginia Prezzi","family":"Santos","sequence":"additional","affiliation":[{"name":"Programa de P\u00f3s-Gradua\u00e7\u00e3o em Microbiologia, Departamento de Microbiologia, Centro de Ci\u00eancias Biol\u00f3gicas, Universidade Estadual de Londrina, Londrina CEP 86057-970, PR, Brazil"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Isabela Madeira de","family":"Castro","sequence":"additional","affiliation":[{"name":"Programa de P\u00f3s-Gradua\u00e7\u00e3o em Microbiologia, Departamento de Microbiologia, Centro de Ci\u00eancias Biol\u00f3gicas, Universidade Estadual de Londrina, Londrina CEP 86057-970, PR, Brazil"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-5412-053X","authenticated-orcid":false,"given":"Paulo Henrique 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Brazil"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-3497-5362","authenticated-orcid":false,"given":"Guilherme","family":"Bartolomeu-Gon\u00e7alves","sequence":"additional","affiliation":[{"name":"Programa de P\u00f3s-Gradua\u00e7\u00e3o em Fisiopatologia Cl\u00ednica e Laboratorial, Centro de Ci\u00eancias da Sa\u00fade, Universidade Estadual de Londrina, Londrina CEP 86038-440, PR, Brazil"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jussevania Pereira","family":"Santos","sequence":"additional","affiliation":[{"name":"Departamento de Microbiologia, Centro de Ci\u00eancias Biol\u00f3gicas, Universidade Estadual de Londrina, Londrina CEP 86057-970, PR, Brazil"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-9975-4627","authenticated-orcid":false,"given":"Alexandre Tadachi","family":"Morey","sequence":"additional","affiliation":[{"name":"Instituto Federal de 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Infect. Dis."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"273","DOI":"10.1016\/j.bjid.2018.07.008","article-title":"Secular trends of candidemia at a Brazilian tertiary care teaching hospital","volume":"22","author":"Braga","year":"2018","journal-title":"Braz. J. Infect. Dis."},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"165","DOI":"10.1016\/j.jped.2016.05.007","article-title":"Risk factors for candidemia mortality in hospitalized children","volume":"93","author":"Motta","year":"2017","journal-title":"J. Pediatr."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"132","DOI":"10.1016\/j.mycmed.2019.02.002","article-title":"Virulence factors of Candida spp. obtained from blood cultures of patients with candidemia attended at tertiary hospitals in Northeast Brazil","volume":"29","author":"Ferreira","year":"2019","journal-title":"J. Mycol. Med."},{"key":"ref_5","doi-asserted-by":"crossref","unstructured":"Khan, S., Cai, L., Bilal, H., Khan, M.N., Fang, W., Zhang, D., Yao, F., Wang, X., Wang, Q., and Hou, B. (2025). An 11-year retrospective analysis of candidiasis epidemiology, risk factors, and antifungal susceptibility in a tertiary care hospital in China. Sci. Rep., 15.","DOI":"10.1038\/s41598-025-92100-x"},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"17","DOI":"10.1007\/978-1-4939-6515-1_2","article-title":"The changing epidemiology of invasive fungal infections","volume":"1508","author":"Enoch","year":"2017","journal-title":"Methods Mol. Biol."},{"key":"ref_7","first-page":"1","article-title":"Nosocomial candidiasis: Antifungal stewardship and the importance of rapid diagnosis","volume":"54","author":"Pfaller","year":"2016","journal-title":"Med. Mycol."},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"7","DOI":"10.1016\/j.mib.2020.05.006","article-title":"The gut, the bad and the harmless: Candida albicans as a commensal and opportunistic pathogen in the intestine","volume":"56","author":"Kumamoto","year":"2020","journal-title":"Curr. Opin. Microbiol."},{"key":"ref_9","doi-asserted-by":"crossref","unstructured":"Romo, J.A., and Kumamoto, C.A. (2020). On commensalism of Candida. J. Fungi, 6.","DOI":"10.3390\/jof6010016"},{"key":"ref_10","doi-asserted-by":"crossref","unstructured":"Doi, A.M., Pignatari, A.C., Edmond, M.B., Marra, A.R., Camargo, L.F., Siqueira, R.A., da Mota, V.P., and Colombo, A.L. (2016). Epidemiology and microbiologic characterization of nosocomial candidemia from a Brazilian National Surveillance Program. PLoS ONE, 11.","DOI":"10.1371\/journal.pone.0146909"},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"62","DOI":"10.1186\/s13613-019-0539-2","article-title":"Candidemia in critically ill immunocompromised patients: Report of a retrospective multicenter cohort study","volume":"9","author":"Ghrenassia","year":"2019","journal-title":"Ann. Intensive Care"},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"e87","DOI":"10.1038\/emi.2017.74","article-title":"Epidemiology and antifungal susceptibility of candidemia isolates of non-albicans Candida species from cancer patients","volume":"6","author":"Wu","year":"2017","journal-title":"Emerg. Microbes Infect."},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"20","DOI":"10.1038\/s41572-024-00503-3","article-title":"Invasive candidiasis","volume":"10","author":"Kanj","year":"2024","journal-title":"Nat. Rev. Dis. Primers"},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"e428","DOI":"10.1016\/S1473-3099(23)00692-8","article-title":"Global incidence and mortality of severe fungal disease","volume":"24","author":"Denning","year":"2024","journal-title":"Lancet Infect. Dis."},{"key":"ref_15","doi-asserted-by":"crossref","unstructured":"Agnelli, C., Guimar\u00e3es, T., Sukiennik, T., Lima, P.R.P., Salles, M.J., Breda, G.L., Queiroz-Telles, F., Chaves Magri, M.M., Mendes, A.V., and Camargo, L.F.A. (2023). Prognostic trends and current challenges in candidemia: A comparative analysis of two multicenter cohorts within the past decade. J. Fungi, 9.","DOI":"10.3390\/jof9040468"},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"myae040","DOI":"10.1093\/mmy\/myae040","article-title":"Candida tropicalis-A systematic review to inform the World Health Organization of a fungal priority pathogens list","volume":"62","author":"Keighley","year":"2024","journal-title":"Med. Mycol."},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"101041","DOI":"10.1016\/j.bjid.2020.11.006","article-title":"Clinical and epidemiological aspects of candidemia in eight medical centers in the state of Parana, Brazil: Parana Candidemia Network","volume":"25","author":"Colombo","year":"2021","journal-title":"Braz. J. Infect. Dis."},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"1437","DOI":"10.1007\/s11274-013-1574-z","article-title":"Candida identification: A journey from conventional to molecular methods in medical mycology","volume":"30","author":"Alam","year":"2014","journal-title":"World J. Microbiol. Biotechnol."},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"e01909-17","DOI":"10.1128\/JCM.01909-17","article-title":"Diagnosing invasive candidiasis","volume":"56","author":"Clancy","year":"2018","journal-title":"J. Clin. Microbiol."},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"329","DOI":"10.1111\/odi.12123","article-title":"Identification of Candida species in the clinical laboratory: A review of conventional, commercial, and molecular techniques","volume":"20","author":"Neppelenbroek","year":"2014","journal-title":"Oral Dis."},{"key":"ref_21","doi-asserted-by":"crossref","unstructured":"Higashi, Y., Niimi, H., Sakamaki, I., Yamamoto, Y., and Kitajima, I. (2020). Rapid identification of Candida species in candidemia directly from blood samples using imperfect match probes. Sci. Rep., 10.","DOI":"10.1038\/s41598-020-62276-5"},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"543","DOI":"10.1159\/000493426","article-title":"Rapid and accurate identification of Candida albicans and Candida dubliniensis by Real-Time PCR and melting curve analysis","volume":"27","author":"Asadzadeh","year":"2018","journal-title":"Med. Princ. Pract."},{"key":"ref_23","doi-asserted-by":"crossref","unstructured":"Alkhars, N., Al Jallad, N., Wu, T.T., and Xiao, J. (2024). Multilocus sequence typing of Candida albicans oral isolates reveals high genetic relatedness of mother-child dyads in early life. PLoS ONE, 19.","DOI":"10.1371\/journal.pone.0290938"},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"391","DOI":"10.1016\/j.ccm.2009.02.014","article-title":"Fungal molecular diagnostics","volume":"30","author":"Wengenack","year":"2009","journal-title":"Clin. Chest Med."},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"59","DOI":"10.1111\/j.1574-6976.2009.00196.x","article-title":"Ribosomal RNA genes in eukaryotic microorganisms: Witnesses of phylogeny?","volume":"34","author":"Cevallos","year":"2010","journal-title":"FEMS Microbiol. Rev."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"2302","DOI":"10.1128\/JCM.38.6.2302-2310.2000","article-title":"Identification of medically important yeasts using PCR-based detection of DNA sequence polymorphisms in the internal transcribed spacer 2 region of the rRNA genes","volume":"38","author":"Chen","year":"2000","journal-title":"J. Clin. Microbiol."},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"1846","DOI":"10.1128\/JCM.37.6.1846-1851.1999","article-title":"Rapid identification of fungi by using the ITS2 genetic region and an automated fluorescent capillary electrophoresis system","volume":"37","author":"Turenne","year":"1999","journal-title":"J. Clin. Microbiol."},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"29","DOI":"10.1046\/j.1439-0507.2003.00949.x","article-title":"Identification of medically important fungi by the Pyrosequencing technology","volume":"47","author":"Gharizadeh","year":"2004","journal-title":"Mycoses"},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"175","DOI":"10.1023\/B:ANTO.0000020154.56649.0f","article-title":"Identification of species of the genus Candida by analysis of the 5.8S rRNA gene and the two ribosomal internal transcribed spacers","volume":"85","author":"Querol","year":"2004","journal-title":"Antonie Van Leeuwenhoek"},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"1353","DOI":"10.1128\/jcm.35.6.1353-1360.1997","article-title":"Detection and identification of fungal pathogens in blood by using molecular probes","volume":"35","author":"Einsele","year":"1997","journal-title":"J. Clin. Microbiol."},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"12","DOI":"10.1016\/j.jmoldx.2010.11.014","article-title":"Molecular identification of closely related Candida species using two ribosomal intergenic spacer fingerprinting methods","volume":"13","author":"Cornet","year":"2011","journal-title":"J. Mol. Diagn."},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"506","DOI":"10.1016\/j.mycmed.2017.07.007","article-title":"The identification of Meyerozyma guilliermondii from blood cultures and surveillance samples in a university hospital in Northeast Turkey: A ten-year survey","volume":"27","author":"Tosun","year":"2017","journal-title":"J. Mycol. Med."},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"517","DOI":"10.1093\/femsre\/fuz015","article-title":"Recent Trends in Molecular Diagnostics of Yeast Infections: From PCR to NGS","volume":"43","year":"2019","journal-title":"FEMS Microbiol. Rev."},{"key":"ref_34","unstructured":"World Health Organization (2018). Chapter 6: Data Analysis. Measles and Rubella Laboratory Network Manual, WHO. Available online: https:\/\/www.who.int\/docs\/default-source\/immunization\/vpd_surveillance\/lab_networks\/measles_rubella\/manual\/chapter-6.pdf."},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"1040","DOI":"10.1099\/jmm.0.000121","article-title":"Rapid Detection of Mycobacterium tuberculosis Complex by Real-Time PCR in Sputum Samples and Its Use in the Routine Diagnosis of a Reference Laboratory","volume":"64","author":"Pinhata","year":"2015","journal-title":"J. Med. Microbiol."},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"e2206525","DOI":"10.1002\/adma.202206525","article-title":"Toward rapid and accurate molecular diagnostics at home","volume":"35","author":"Liu","year":"2023","journal-title":"Adv. Mater."},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"a019299","DOI":"10.1101\/cshperspect.a019299","article-title":"Fungal diagnostics","volume":"4","author":"Kozel","year":"2014","journal-title":"Cold Spring Harb. Perspect. Med."},{"key":"ref_38","doi-asserted-by":"crossref","unstructured":"Sidiq, F., Hoostal, M., and Rogers, S.O. (2016). Rapid identification of fungi in culture-negative clinical blood and respiratory samples by DNA sequence analyses. BMC Res. Notes, 9.","DOI":"10.1186\/s13104-016-2097-0"},{"key":"ref_39","doi-asserted-by":"crossref","first-page":"787","DOI":"10.1111\/myc.12532","article-title":"Molecular identification and phylogenetical analysis of dermatophyte fungi from Latin America","volume":"59","author":"Hrycyk","year":"2016","journal-title":"Mycoses"},{"key":"ref_40","first-page":"100","article-title":"Contribution of the internal transcribed spacer regions to the detection and identification of human fungal pathogens","volume":"67","author":"Trabelsi","year":"2019","journal-title":"Curr. Res. Transl. Med."},{"key":"ref_41","doi-asserted-by":"crossref","first-page":"5135","DOI":"10.1038\/s41467-018-07556-5","article-title":"Molecular diagnostics in medical mycology","volume":"9","author":"Wickes","year":"2018","journal-title":"Nat. Commun."},{"key":"ref_42","first-page":"89","article-title":"Accurate and sensitive real-time PCR assays using intergenic spacer 1 region to differentiate Cryptococcus gattii sensu lato and Cryptococcus neoformans sensu lato","volume":"54","author":"Tavares","year":"2016","journal-title":"Med. Mycol."},{"key":"ref_43","doi-asserted-by":"crossref","first-page":"568","DOI":"10.1016\/j.mycmed.2018.06.007","article-title":"Candida auris: An emerging drug resistant yeast\u2014A mini-review","volume":"28","author":"Bidaud","year":"2018","journal-title":"J. Mycol. Med."},{"key":"ref_44","doi-asserted-by":"crossref","unstructured":"Du, H., Bing, J., Hu, T., Ennis, C.L., Nobile, C.J., and Huang, G. (2020). Candida auris: Epidemiology, biology, antifungal resistance, and virulence. PLoS Pathog., 16.","DOI":"10.1371\/journal.ppat.1008921"},{"key":"ref_45","doi-asserted-by":"crossref","first-page":"306","DOI":"10.1093\/cid\/cix744","article-title":"Approach to the investigation and management of patients with Candida auris, an emerging multidrug-resistant yeast","volume":"66","author":"Tsay","year":"2018","journal-title":"Clin. Infect. Dis."},{"key":"ref_46","doi-asserted-by":"crossref","unstructured":"Granada, M., Cook, E., Sherlock, G., and Rosenzweig, F. (2024). Microbe profile: Candida glabrata\u2014A master of deception. Microbiology, 170.","DOI":"10.1099\/mic.0.001518"},{"key":"ref_47","doi-asserted-by":"crossref","unstructured":"Dai, Z., Lan, X., Cai, M., Liao, Y., Zhang, J., Ye, N., Lu, X., Wang, J., Xiao, Y., and Zhang, Y. (2025). Nineteen years retrospective analysis of epidemiology, antifungal resistance and a nomogram model for 30-day mortality in nosocomial candidemia patients. Front. Cell. Infect. Microbiol., 15.","DOI":"10.3389\/fcimb.2025.1504866"},{"key":"ref_48","doi-asserted-by":"crossref","unstructured":"Peixoto, P.H., Silva, M.L., Portela, F.V., da Silva, B., Milanez, E., de Oliveira, D., Ribeiro, A., de Almeida, H., Lima-Neto, R., and Guedes, G.M. (2023). Clinical, epidemiological and laboratory features of invasive Candida parapsilosis complex infections in a brazilian pediatric reference hospital during the COVID-19 pandemic. J. Fungi, 9.","DOI":"10.3390\/jof9080844"},{"key":"ref_49","doi-asserted-by":"crossref","unstructured":"Lima, R., Ribeiro, F.C., Colombo, A.L., and de Almeida, J.N. (2022). The emerging threat antifungal-resistant Candida tropicalis in humans, animals, and environment. Front. Fungal Biol., 3.","DOI":"10.3389\/ffunb.2022.957021"},{"key":"ref_50","doi-asserted-by":"crossref","unstructured":"Otaguiri, E.S., Morguette, A.E.B., Morey, A.T., Tavares, E.R., Kerbauy, G., de Almeida Torres, R.S.L., Chaves J\u00fanior, M., Tognim, M.C.B., G\u00f3es, V.M., and Krieger, M.A. (2018). Development of a melting-curve based multiplex real-time PCR assay for simultaneous detection of Streptococcus agalactiae and genes encoding resistance to macrolides and lincosamides. BMC Pregn. Childbirth, 18.","DOI":"10.1186\/s12884-018-1774-5"},{"key":"ref_51","doi-asserted-by":"crossref","first-page":"191","DOI":"10.1186\/s12936-023-04617-z","article-title":"Performance of a novel melting curve-based qPCR assay for malaria parasites in routine clinical practice in non-endemic setting","volume":"22","author":"Stuitje","year":"2023","journal-title":"Malar. J."},{"key":"ref_52","doi-asserted-by":"crossref","unstructured":"Wan, Z., Zhang, Y., He, Z., Liu, J., Lan, K., Hu, Y., and Zhang, C. (2016). A melting curve-based multiplex RT-qPCR assay for simultaneous detection of four Human Coronaviruses. Int. J. Mol. Sci., 17.","DOI":"10.3390\/ijms17111880"},{"key":"ref_53","doi-asserted-by":"crossref","unstructured":"Tavares, E.R., de Lima, T.F., Bartolomeu-Gon\u00e7alves, G., de Castro, I.M., de Lima, D.G., Borges, P.H.G., Nakazato, G., Kobayashi, R.K.T., Venancio, E.J., and Tarley, C.R.T. (2023). Development of a melting-curve-based multiplex real-time PCR assay for the simultaneous detection of viruses causing respiratory infection. Microorganisms, 11.","DOI":"10.3390\/microorganisms11112692"},{"key":"ref_54","first-page":"336","article-title":"Exploring rapid molecular methods for diagnosing Candida species infecting humans: A narrative review","volume":"5","author":"Dubey","year":"2023","journal-title":"Med. Infect. Dis."},{"key":"ref_55","doi-asserted-by":"crossref","first-page":"8","DOI":"10.37201\/req\/094.2024","article-title":"Diagnostic and prognostic value of time to positivity in blood cultures. An opinion paper","volume":"38","author":"Maffezzoli","year":"2025","journal-title":"Rev. Esp. Quimioter."},{"key":"ref_56","doi-asserted-by":"crossref","unstructured":"Zacharioudakis, I.M., Zervou, F.N., and Mylonakis, E. (2018). T2 Magnetic Resonance Assay: Overview of Available Data and Clinical Implications. J. Fungi, 4.","DOI":"10.3390\/jof4020045"},{"key":"ref_57","doi-asserted-by":"crossref","first-page":"305","DOI":"10.1007\/978-1-4939-6515-1_18","article-title":"T2 Magnetic Resonance for Fungal Diagnosis","volume":"1508","author":"Zervou","year":"2017","journal-title":"Methods Mol. Biol."},{"key":"ref_58","unstructured":"Thermo Fisher Scientific (2008). Creating Standard Curves with Genomic DNA or Plasmid DNA Templates for Use in Quantitative PCR, Application Note; Thermo Fisher Scientific. Available online: https:\/\/assets.thermofisher.com\/TFS-Assets\/LSG\/Application-Notes\/cms_042486.pdf."}],"container-title":["International Journal of Molecular Sciences"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/1422-0067\/26\/19\/9411\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,9,28]],"date-time":"2025-09-28T04:27:28Z","timestamp":1759033648000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/1422-0067\/26\/19\/9411"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2025,9,26]]},"references-count":58,"journal-issue":{"issue":"19","published-online":{"date-parts":[[2025,10]]}},"alternative-id":["ijms26199411"],"URL":"https:\/\/doi.org\/10.3390\/ijms26199411","relation":{},"ISSN":["1422-0067"],"issn-type":[{"value":"1422-0067","type":"electronic"}],"subject":[],"published":{"date-parts":[[2025,9,26]]}}}