{"title":"分子检测与真菌学方法诊断犀牛眼眶毛霉病：120例COVID-19患者临床标本分析","authors":"Sajedeh Soltani, Mahzad Erami, Kazem Ahmadikia, Shima Aboutalebian, Faezeh Rouhi, Mojtaba Fakhrehi, Reza Mohammadi Manesh, Hossein Mirhendi","doi":"10.1007/s11046-025-00937-7","DOIUrl":null,"url":null,"abstract":"<p><strong>Background: </strong>Mucormycosis, a fungal emergency, poses a serious threat to both COVID-19 and non-COVID-19 individuals due to its invasive nature, rapid progression, and high rates of morbidity and mortality. This underscores the crucial need for timely detection and management. In this study, we investigated the utility of real-time PCR (RT-qPCR) assays for detecting Mucorales in clinical specimens, and assessed the performance of both SYBR Green and TaqMan probe RT-qPCR in amplifying Mucorales-specific 18S rDNA genes. We conducted accuracy analyses using direct examination with KOH as a standard for the laboratory diagnosis of mucormycosis. Additionally, we compared the results with culture and duplex PCR.</p><p><strong>Patients/methods: </strong>Both SYBR Green and TaqMan RT-qPCR were optimized using Mucorales-specific oligonucleotides to amplify the conserved 18S rDNA targets. DNAs extracted from 120 rhino sinus specimens, which all were collected from COVID-19 patients upon suspicion of invasive fungal infections, were used for molecular diagnosis. The results of both RT-qPCR assays were compared with the result of direct microscopy, culture, and duplex Mucorales-specific PCR assay.</p><p><strong>Results: </strong>SYBR Green real-time PCR produced a distinct melting temperature (Tm) pattern (80.24 ± 0.70 °C) and detected Mucorales in 51 out of 120 clinical samples. When compared to direct examination with KOH, the standard method for diagnosing mucormycosis, SYBR Green PCR demonstrated a sensitivity of 91.67% (95% confidence interval (CI): 86.7-96.5%) and a specificity of 90.28% (95% CI: 84.9-95.5%). In contrast, TaqMan-probe PCR identified Mucorales in 34 out of 120 samples, with a sensitivity of 64.58% (95% CI: 56-73.1%) and a specificity of 95.83% (95% CI: 92.26-99.39%).</p><p><strong>Conclusion: </strong>SYBR Green-based PCR can be used as a reliable confirmatory test for diagnosing mucormycosis, particularly in cases with atypical hyphae, mixed infections (featuring both septate and non-septate hyphae), or when the direct examination is positive but culture results are negative. The lower sensitivity of the TaqMan-probe PCR may be attributed to factors such as using a degenerate probe, which can lead to false-negative results.</p>","PeriodicalId":19017,"journal":{"name":"Mycopathologia","volume":"190 2","pages":"30"},"PeriodicalIF":3.6000,"publicationDate":"2025-03-05","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":"{\"title\":\"Molecular Assays Versus Mycological Methods for Diagnosis of Rhino Orbital Mucormycosis: Analysis of 120 Clinical Specimens from COVID-19 Patients.\",\"authors\":\"Sajedeh Soltani, Mahzad Erami, Kazem Ahmadikia, Shima Aboutalebian, Faezeh Rouhi, Mojtaba Fakhrehi, Reza Mohammadi Manesh, Hossein Mirhendi\",\"doi\":\"10.1007/s11046-025-00937-7\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<p><strong>Background: </strong>Mucormycosis, a fungal emergency, poses a serious threat to both COVID-19 and non-COVID-19 individuals due to its invasive nature, rapid progression, and high rates of morbidity and mortality. This underscores the crucial need for timely detection and management. In this study, we investigated the utility of real-time PCR (RT-qPCR) assays for detecting Mucorales in clinical specimens, and assessed the performance of both SYBR Green and TaqMan probe RT-qPCR in amplifying Mucorales-specific 18S rDNA genes. We conducted accuracy analyses using direct examination with KOH as a standard for the laboratory diagnosis of mucormycosis. Additionally, we compared the results with culture and duplex PCR.</p><p><strong>Patients/methods: </strong>Both SYBR Green and TaqMan RT-qPCR were optimized using Mucorales-specific oligonucleotides to amplify the conserved 18S rDNA targets. DNAs extracted from 120 rhino sinus specimens, which all were collected from COVID-19 patients upon suspicion of invasive fungal infections, were used for molecular diagnosis. The results of both RT-qPCR assays were compared with the result of direct microscopy, culture, and duplex Mucorales-specific PCR assay.</p><p><strong>Results: </strong>SYBR Green real-time PCR produced a distinct melting temperature (Tm) pattern (80.24 ± 0.70 °C) and detected Mucorales in 51 out of 120 clinical samples. When compared to direct examination with KOH, the standard method for diagnosing mucormycosis, SYBR Green PCR demonstrated a sensitivity of 91.67% (95% confidence interval (CI): 86.7-96.5%) and a specificity of 90.28% (95% CI: 84.9-95.5%). In contrast, TaqMan-probe PCR identified Mucorales in 34 out of 120 samples, with a sensitivity of 64.58% (95% CI: 56-73.1%) and a specificity of 95.83% (95% CI: 92.26-99.39%).</p><p><strong>Conclusion: </strong>SYBR Green-based PCR can be used as a reliable confirmatory test for diagnosing mucormycosis, particularly in cases with atypical hyphae, mixed infections (featuring both septate and non-septate hyphae), or when the direct examination is positive but culture results are negative. The lower sensitivity of the TaqMan-probe PCR may be attributed to factors such as using a degenerate probe, which can lead to false-negative results.</p>\",\"PeriodicalId\":19017,\"journal\":{\"name\":\"Mycopathologia\",\"volume\":\"190 2\",\"pages\":\"30\"},\"PeriodicalIF\":3.6000,\"publicationDate\":\"2025-03-05\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Mycopathologia\",\"FirstCategoryId\":\"99\",\"ListUrlMain\":\"https://doi.org/10.1007/s11046-025-00937-7\",\"RegionNum\":3,\"RegionCategory\":\"生物学\",\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"Q2\",\"JCRName\":\"MYCOLOGY\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Mycopathologia","FirstCategoryId":"99","ListUrlMain":"https://doi.org/10.1007/s11046-025-00937-7","RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q2","JCRName":"MYCOLOGY","Score":null,"Total":0}
Molecular Assays Versus Mycological Methods for Diagnosis of Rhino Orbital Mucormycosis: Analysis of 120 Clinical Specimens from COVID-19 Patients.
Background: Mucormycosis, a fungal emergency, poses a serious threat to both COVID-19 and non-COVID-19 individuals due to its invasive nature, rapid progression, and high rates of morbidity and mortality. This underscores the crucial need for timely detection and management. In this study, we investigated the utility of real-time PCR (RT-qPCR) assays for detecting Mucorales in clinical specimens, and assessed the performance of both SYBR Green and TaqMan probe RT-qPCR in amplifying Mucorales-specific 18S rDNA genes. We conducted accuracy analyses using direct examination with KOH as a standard for the laboratory diagnosis of mucormycosis. Additionally, we compared the results with culture and duplex PCR.
Patients/methods: Both SYBR Green and TaqMan RT-qPCR were optimized using Mucorales-specific oligonucleotides to amplify the conserved 18S rDNA targets. DNAs extracted from 120 rhino sinus specimens, which all were collected from COVID-19 patients upon suspicion of invasive fungal infections, were used for molecular diagnosis. The results of both RT-qPCR assays were compared with the result of direct microscopy, culture, and duplex Mucorales-specific PCR assay.
Results: SYBR Green real-time PCR produced a distinct melting temperature (Tm) pattern (80.24 ± 0.70 °C) and detected Mucorales in 51 out of 120 clinical samples. When compared to direct examination with KOH, the standard method for diagnosing mucormycosis, SYBR Green PCR demonstrated a sensitivity of 91.67% (95% confidence interval (CI): 86.7-96.5%) and a specificity of 90.28% (95% CI: 84.9-95.5%). In contrast, TaqMan-probe PCR identified Mucorales in 34 out of 120 samples, with a sensitivity of 64.58% (95% CI: 56-73.1%) and a specificity of 95.83% (95% CI: 92.26-99.39%).
Conclusion: SYBR Green-based PCR can be used as a reliable confirmatory test for diagnosing mucormycosis, particularly in cases with atypical hyphae, mixed infections (featuring both septate and non-septate hyphae), or when the direct examination is positive but culture results are negative. The lower sensitivity of the TaqMan-probe PCR may be attributed to factors such as using a degenerate probe, which can lead to false-negative results.
期刊介绍：
Mycopathologia is an official journal of the International Union of Microbiological Societies (IUMS). Mycopathologia was founded in 1938 with the mission to ‘diffuse the understanding of fungal diseases in man and animals among mycologists’. Many of the milestones discoveries in the field of medical mycology have been communicated through the pages of this journal. Mycopathologia covers a diverse, interdisciplinary range of topics that is unique in breadth and depth. The journal publishes peer-reviewed, original articles highlighting important developments concerning medically important fungi and fungal diseases. The journal highlights important developments in fungal systematics and taxonomy, laboratory diagnosis of fungal infections, antifungal drugs, clinical presentation and treatment, and epidemiology of fungal diseases globally. Timely opinion articles, mini-reviews, and other communications are usually invited at the discretion of the editorial board. Unique case reports highlighting unprecedented progress in the diagnosis and treatment of fungal infections, are published in every issue of the journal. MycopathologiaIMAGE is another regular feature for a brief clinical report of potential interest to a mixed audience of physicians and laboratory scientists. MycopathologiaGENOME is designed for the rapid publication of new genomes of human and animal pathogenic fungi using a checklist-based, standardized format.
求助内容：
应助结果提醒方式：
京公网安备 11010802042870号
