Detection of Aspergillus fumigatus mycotoxins: immunogen synthesis and immunoassay development

Fox, M.; Gray, Gary R.; Kavanagh, Kevin; Lewis, Caroline A.; Doyle, Seán

doi:10.1016/j.mimet.2003.10.009

Cited by 24 publications

(22 citation statements)

References 23 publications

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“…More specifically, gliotoxin detection following RP-HPLC separation is usually carried out at 254 nm [10]. The immunological detection of gliotoxin by ELISA has been demonstrated, and here, the limit of detection was 10 μg mL −1 [14]. In addition, Lewis et al showed that gliotoxin was present in sera and lungs of mice suffering from experimentally induced invasive aspergillosis by LC-MS analysis [15].…”

Section: Introductionmentioning

confidence: 81%

Single-pot derivatisation strategy for enhanced gliotoxin detection by HPLC and MALDI-ToF mass spectrometry

et al. 2011

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Gliotoxin is produced by non-ribosomal peptide synthesis and secreted from certain fungi, including Aspergillus fumigatus. It is an epipolythiodioxopiperazine that contains an intact disulphide bridge and is the focus of intense research as a consequence of its negative immunomodulatory properties. Gliotoxin detection is generally enabled by reversed-phase-high-performance liquid chromatography (RP-HPLC), with absorbance detection (220-280 nm), or liquid chromatography-mass spectrometry, yet detection is not readily achievable by matrix-assisted laser desorption ionisation-time-of-flight mass spectrometry (MALDI-ToF MS). We have developed a single-pot derivatisation strategy which uses sodium borohydride-mediated reduction of gliotoxin followed by immediate alkylation of exposed thiols by 5′-iodoacetamidofluorescein to yield a stable product, diacetamidofluorescein-gliotoxin (GT-(AF) 2 ), of molecular mass 1103.931 Da ((M+H)+). This product is readily detectable by RP-HPLC and exhibits a 6.8-fold increase in molar absorptivity compared with gliotoxin, which results in a higher sensitivity of detection (40 ng; 125 pmoL). GT-(AF) 2 also fluoresces (excitation/emission, 492:518 nm). Unlike free gliotoxin, the product (>800 fmol) is detectable by MALDI-ToF MS. Sporidesmin A can also be detected by RP-HPLC and MALDI-ToF MS (>530 fmol) using this strategy. We also demonstrate that the strategy facilitates detection of gliotoxin (mean± SD = 3.55 ± 0.07 μg 100 μL −1 ; n = 2) produced by A. fumigatus, without the requirement for organic extraction of culture supernatants and associated solvent removal. GT-(AF) 2 is also detectable (150 ng; 460 pmol) by thinlayer chromatography.

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Section: Introductionmentioning

confidence: 81%

Single-pot derivatisation strategy for enhanced gliotoxin detection by HPLC and MALDI-ToF mass spectrometry

et al. 2011

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“…We have previously demonstrated the specific detection of gliotoxin by ELISA (Fox et al, 2004), and gliotoxin can be detected by LC-MS in sera and lungs from mice suffering from experimentally induced IA (Lewis et al, 2005). Gliotoxin can also be detected in cancer patients with either proven or probable IA (Lewis et al, 2005).…”

Section: Biopharmaceutical and Diagnostic Potential Of Nrpsmentioning

confidence: 99%

Nonribosomal peptide synthesis in Aspergillus fumigatus and other fungi

2007

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In fungi, nonribosomal peptide synthetases (NRP synthetases) are large multi-functional enzymes containing adenylation, thiolation (or peptidyl carrier protein, PCP) and condensation domains. These enzymes are often encoded within gene clusters. Multiple NRP synthetase ORFs have also been identified in fungi (14 in Aspergillus fumigatus). LeaA, a methyltransferase, is involved in secondary metabolite gene cluster regulation in Aspergillus spp. The NRP synthetases GliP and FtmA respectively direct the biosynthesis of the toxic metabolites gliotoxin and brevianamide F, a precursor of bioactive prenylated alkaloids. The NRP synthetase Pes1 has been shown to mediate resistance to oxidative stress, and in plant-pathogenic ascomycetes (e.g. Cochliobolus heterostrophus) an NRP synthetase, encoded by the NPS6 gene, significantly contributes to virulence and resistance to oxidative stress. Adenylation (A) domains within NRP synthetases govern the specificity of amino acid incorporation into nonribosomally synthesized peptides. To date there have only been limited demonstrations of A domain specificity (e.g. A. fumigatus GliP and in Beauveria bassiana) in fungi. Indeed, only in silico prediction data are available on A domain specificity of NRP synthetases from most fungi. NRP synthetases are activated by 49-phosphopantetheinylation of serine residues within PCP domains by 49-phosphopantetheinyl transferases (49-PPTases). Coenzyme A acts as the 49-phosphopantetheine donor, and labelled coenzyme A can be used to affinity-label apo-NRP synthetases. Emerging fungal gene disruption and gene cluster expression strategies, allied to proteomic strategies, are poised to facilitate a greater understanding of the coding potential of NRP synthetases in fungi.

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“…Some species of Aspergillus were reported allergens [28] while some mycotoxicant [29]. Many species of Acremonium proverd as opportunistic pathogens [30].…”

Section: Some Aquatic Fungi That Found In Surface Water Are Mentionedmentioning

confidence: 99%

Freshwater Fungal Richness, Their Assessment and Impact on Human Welfare: A Review

2016

IJSR

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Fungi are eukaryotic, heterotrophic organism, including both single-celled yeasts and multi-cellular filamentous fungi which can survive in oligotrophic environments, also called recyclers of organic material. The diversity spectrum and identification method of fungi are reviewed in such order that is adapted to aquatic ecosystem as well as their contribution to the environment and significant to human welfare.

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Detection of Aspergillus fumigatus mycotoxins: immunogen synthesis and immunoassay development

Cited by 24 publications

References 23 publications

Single-pot derivatisation strategy for enhanced gliotoxin detection by HPLC and MALDI-ToF mass spectrometry

Single-pot derivatisation strategy for enhanced gliotoxin detection by HPLC and MALDI-ToF mass spectrometry

Nonribosomal peptide synthesis in Aspergillus fumigatus and other fungi

Freshwater Fungal Richness, Their Assessment and Impact on Human Welfare: A Review

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