Evaluation of a Novel Mitochondrial Pan-Mucorales Marker for the Detection, Identification, Quantification, and Growth Stage Determination of Mucormycetes

Caramalho, Rita; Madl, Lisa; Rosam, Katharina; Rambach, Günter; Speth, Cornelia; Pallua, Johannes; Larentis, Thomas; Araújo, Ricardo; Alastruey‐Izquierdo, Ana; Lass‐Flörl, Cornelia; Lackner, Michaela

doi:10.3390/jof5040098

Cited by 25 publications

(17 citation statements)

References 60 publications

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“…Recent studies have confirmed the value of qPCR in BAL, with regard to earlier diagnosis and better outcomes in cases of pulmonary mucormycosis [ 102 , 103 , 104 , 105 , 106 , 107 , 108 , 109 , 110 , 111 , 112 , 113 ].…”

Section: Diagnosismentioning

confidence: 94%

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Epidemiology and Diagnosis of Mucormycosis: An Update

Skiada

Pavleas

Drogari‐Apiranthitou

2020

JoF

518

508

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Mucormycosis is an angioinvasive fungal infection, due to fungi of the order Mucorales. Its incidence cannot be measured exactly, since there are few population-based studies, but multiple studies have shown that it is increasing. The prevalence of mucormycosis in India is about 80 times the prevalence in developed countries, being approximately 0.14 cases per 1000 population. Diabetes mellitus is the main underlying disease globally, especially in low and middle-income countries. In developed countries the most common underlying diseases are hematological malignancies and transplantation. Τhe epidemiology of mucormycosis is evolving as new immunomodulating agents are used in the treatment of cancer and autoimmune diseases, and as the modern diagnostic tools lead to the identification of previously uncommon genera/species such as Apophysomyces or Saksenaea complex. In addition, new risk factors are reported from Asia, including post-pulmonary tuberculosis and chronic kidney disease. New emerging species include Rhizopus homothallicus, Thamnostylum lucknowense, Mucor irregularis and Saksenaea erythrospora. Diagnosis of mucormycosis remains challenging. Clinical approach to diagnosis has a low sensitivity and specificity, it helps however in raising suspicion and prompting the initiation of laboratory testing. Histopathology, direct examination and culture remain essential tools, although the molecular methods are improving. The internal transcribed spacer (ITS) region is the most widely sequenced DNA region for fungi and it is recommended as a first-line method for species identification of Mucorales. New molecular platforms are being investigated and new fungal genetic targets are being explored. Molecular-based methods have gained acceptance for confirmation of the infection when applied on tissues. Methods on the detection of Mucorales DNA in blood have shown promising results for earlier and rapid diagnosis and could be used as screening tests in high-risk patients, but have to be validated in clinical studies. More, much needed, rapid methods that do not require invasive procedures, such as serology-based point-of-care, or metabolomics-based breath tests, are being developed and hopefully will be evaluated in the near future.

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Section: Diagnosismentioning

confidence: 94%

“…Most of the molecular tests target the 18S ribosomal RNA genes, but also other targets have been investigated. These include the 28S rDNA [ 104 ], the mitochondrial gene rnl [ 105 ], the cytochrome b gene [ 106 ] or the Mucorales-specific CotH gene [ 107 ].…”

Section: Diagnosismentioning

confidence: 99%

Epidemiology and Diagnosis of Mucormycosis: An Update

Skiada

Pavleas

Drogari‐Apiranthitou

2020

JoF

518

508

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“…We were able to detect and quantify genomic DNA of mucormycetes in small quantities of murine organ homogenates (20 mg). The primers were found to be specific for mucormycetes [13].…”

Section: Fungal Load and Dissemination After Inhalative Infection Witmentioning

confidence: 99%

Comparative immunopathogenesis in a murine model of inhalative infection with the mucormycetes Lichtheimia corymbifera and Rhizopus arrhizus

et al. 2020

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Pathogenic mucormycetes induce diseases with considerable morbidity and mortality in immunocompromised patients. Virulence data comparing different Mucorales species and various underlying risk factors are limited. We therefore compared the pathogenesis of inhalative infection by Rhizopus (R.) arrhizus and Lichtheimia (L.) corymbifera in murine models for predominant risk factors for onset of infection. Mice with diabetes or treated with cyclophosphamide or cortisone acetate were challenged via the intranasal route with an isolate of R. arrhizus or L. corymbifera, respectively. Clinical, immunological and inflammation parameters as well as efficacy of posaconazole prophylaxis were monitored over 14 days. Whereas immunocompetent mice showed no clinical symptoms after mucormycete infection, mice treated with either cyclophosphamide (CP) or cortisone acetate (CA) were highly susceptible. Animals infected with the isolate of R. arrhizus showed prolonged survival and lower mortality, compared to those exposed to the L. corymbifera isolate. This lower virulence of R. arrhizus was risk factor-dependent, since diabetic mice died only after infection with Rhizopus, whereas all Lichtheimia-infected diabetic animals survived. Under posaconazole prophylaxis, both mucormycetes were able to establish breakthrough infections in CA-and CP-treated mice, but the course of infection was significantly delayed. Detailed analysis revealed that susceptibility of CA-and CP-treated mice could not be mimicked by exclusive lack or downmodulation of neutrophils, platelets or complement, but can be supposed to be the consequence of a broad immunosuppressive effect induced by the drugs. Both Lichtheimia corymbifera and Rhizopus arrhizus induce invasive mycoses in immunocompromised hosts after inhalative infection. Key parameters such as virulence and immunopathogenesis vary strongly according to fungal species and underlying risk group. Selected neutropenia is no sufficient risk factor for onset of inhalative mucormycosis.

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“…They evaluated their method using experimentally infected mice for plasma, urine, and bronchoalveolar lavages as well as urine from patients with proven mucormycosis and concluded that the CotH gene was a promising target for diagnostics in urine samples [14]. Further, Caramalho et al, (2019) tested the mitochondrial rnl gene, which encodes for the large subunit of the rRNA, for diagnostic purposes, using a qPCR + HRM assay [35]. The authors found a 100% rate of correct identification of culture isolates and a relatively high sensitivity for the detection of Mucorales in FFPE samples of 71%.…”

Section: Target Genes and Sequence Databasesmentioning

confidence: 99%

“…The authors found a 100% rate of correct identification of culture isolates and a relatively high sensitivity for the detection of Mucorales in FFPE samples of 71%. Mitochondrial genes are more protected from degradation and are present in higher copy numbers than nuclear DNA, which makes them promising candidates for diagnostic assays [ 35 ]. Finally, the cytochrome b gene was targeted by a probe-based qPCR + HRM Mucorales-specific assay, which was evaluated for culture isolates, fresh tissue, and FFPE tissue, resulting in sensitivities of 100%, 100%, and 56%, respectively [ 36 ].…”

Section: Dna-based Diagnosismentioning

confidence: 99%

Microbiological and Molecular Diagnosis of Mucormycosis: From Old to New

2021

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Members of the order Mucorales may cause severe invasive fungal infections (mucormycosis) in immune-compromised and otherwise ill patients. Diagnosis of Mucorales infections and discrimination from other filamentous fungi are crucial for correct management. Here, we present an overview of current state-of-the-art mucormycosis diagnoses, with a focus on recent developments in the molecular field. Classical diagnostic methods comprise histology/microscopy as well as culture and are still the gold standard. Newer molecular methods are evolving quickly and display great potential in early diagnosis, although standardization is still missing. Among them, quantitative PCR assays with or without melt curve analysis are most widely used to detect fungal DNA in clinical samples. Depending on the respective assay, sequencing of the resulting PCR product can be necessary for genus or even species identification. Further, DNA-based methods include microarrays and PCR-ESI-MS. However, general laboratory standards are still in development, meaning that molecular methods are currently limited to add-on analytics to culture and microscopy.

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Evaluation of a Novel Mitochondrial Pan-Mucorales Marker for the Detection, Identification, Quantification, and Growth Stage Determination of Mucormycetes

Cited by 25 publications

References 60 publications

Epidemiology and Diagnosis of Mucormycosis: An Update

Epidemiology and Diagnosis of Mucormycosis: An Update

Comparative immunopathogenesis in a murine model of inhalative infection with the mucormycetes Lichtheimia corymbifera and Rhizopus arrhizus

Microbiological and Molecular Diagnosis of Mucormycosis: From Old to New

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