Noninvasive Combined Diagnosis and Monitoring of Aspergillus and Pseudomonas Infections: Proof of Concept

Abstract

In acutely ill patients, particularly in intensive care units or in mixed infections, time to a microbe-specific diagnosis is critical to a successful outcome of therapy. We report the application of evolving technologies involving mass spectrometry to diagnose and monitor a patient’s course. As proof of this concept, we studied five patients and used two rat models of mono-infection and coinfection. We report the noninvasive combined monitoring of Aspergillus fumigatus and Pseudomonas aeruginosa infection. The invasive coinfection was detected by monitoring the fungal triacetylfusarinine C and ferricrocin siderophore levels and the bacterial metabolites pyoverdin E, pyochelin, and 2-heptyl-4-quinolone, studied in the urine, endotracheal aspirate, or breath condensate. The coinfection was monitored by mass spectrometry followed by isotopic data filtering. In the rat infection model, detection indicated 100-fold more siderophores in urine compared to sera, indicating the diagnostic potential of urine sampling. The tools utilized in our studies can now be examined in large clinical series, where we could expect the accuracy and speed of diagnosis to be competitive with conventional methods and provide advantages in unraveling the complexities of mixed infections.

Diagnosis of Aspergillosis in Horses

“…This would make it necessary to use an integrated diagnostic strategy combining BALF GM measurements with other techniques. We, therefore, investigated the diagnostic utility of multiple non-culture methods that rely on fungal infection biomarkers used in human medicine [ 29 , 35 , 36 , 37 ]. The detection of these biomarkers in equine serum and urine samples warrants further study, and the available serum data are not conclusive other than in the case of serum BDG ( Figure 5 ).…”

“…Gtx is an immunosuppressive fungal toxin that is secreted to the extracellular environment in response to fungal stress [ 40 ]. Studies have shown the utility of siderophores and/or gliotoxin in human blood and urine as useful biomarkers of Aspergillus infection, and we have detailed this recently [ 36 , 41 , 42 , 43 ].…”

Quantitation of small molecules from liquid chromatography—mass spectrometric accurate mass datasets using CycloBranch

“…The dataset of mass spectra was taken from the published literature, 16 where the sample preparation and data collection processes were described in detail. Briefly, the siderophore standards (ferrioxamine E (FoxE) and a mixture of pyoverdines D and E) were purchased from EMC Microcollections GmbH (Tübingen, Germany) and spiked into the human urine and serum samples.…”

“…An HPLC gradient can be found in the reference. 16 The liquid chromatograph was coupled to a 12 T SolariX Fourier transform ion cyclotron resonance (FTICR) mass spectrometer (Bruker Daltonics, Billerica, MA, USA) operating in positive ion mode. All LC-MS datasets were collected in triplicate, and the results were averaged.…”

“…CycloBranch 2.1.21 was used in the experiments. The accuracy of the quantitation methodology was compared with the clinical diagnostic application 16 based on manual data retrieval from Bruker DataAnalysis 5.0 software (Bruker Daltonics, Billerica, USA), and Microsoft Excel (Microsoft Corporation, USA).…”