Feasibility of detecting fungal DNA in exhaled breath condensate by the Luminex Multiplex xTAG fungal PCR assay in lung transplant recipients

Alya Bhimji

Abstract: 

immunocompromised patients, partly due to the difficulty in diagnosing this infection. Invasive
sampling procedures (i.e. bronchoscopy) are often precluded in patients and conventional diagnostic
techniques lack sensitivity. Exhaled breath condensate (EBC) collection provides a noninvasive
means of sampling the airway lining fluid. We assessed the suitability of EBC to detect fungal DNA.
Material/methods: A single-center prospective study was conducted to investigate the feasibility of
detecting clinically relevant fungi (23 species and one internal control) in EBC specimens from lung
transplant recipients using the Luminex multiplex xTAG fungal ASR assay. Specifically, the assay
targets seven Candida species, including albicans, glabrata, lusitaniae, tropicalis, parapsilosis, krusei
and guilliermondii; and four Aspergillus species, terreus, fumigatus, flavus, and niger. Other targets
include Zygomycetes (specifically Rhizopus arrhizus, Rhizopus microsporus, Cunninghamella
bertholletiae, and Mucor indicuss), Pneumocystis jirovecii, Histoplasma capsulatum, Coccidioides
immitis, Cryptococcus neoformans, Blastomyces dermatitidis, Scedosporium prolificans and
apiospermum, and Fusarium. Tremella fuciformis was used as an internal control. PCR results in EBC
specimens were compared with conventional diagnostic tests, such as culture and galactomannan
(GM), in concomitant bronchoalveolar lavage (BAL) samples.
Results: We tested 202 EBCs negative (n = 121) or positive (n = 81) for fungi (see figure). The xTAG
ASR assay in EBC specimens and BAL fungal cultures yielded concurrent negative results in 162
(80%) samples and positive results in 19 (9%) samples. The xTAG ASR assay yielded sensitivity,
specificity, positive and negative predictive values of 82.6%, 90.5%, 52.8%, and 97.6%. Identification
of species was provided by the xTAG ASR assay for 17 unspeciated BAL fungal cultures [A. flavus (n
= 3), A. fumigatus (n = 1), A. terreus (n = 1), C. albicans (n = 1), C. guilliermondii (n = 2), C.
parapsilosis (n = 2), H. capsulatum (n = 3), M. indicus (n = 1), C. parapsilosis + A. flavus (n = 1), C.
parapsilosis + C. bertholletiae (n = 1), C. parapsilosis + S. apiospermum (n = 1)]. Discordant results
were obtained in 4 samples which were culture positive for A. fumigatus, but negative for the xTAG
ASR assay. ITS2 primers were used to sequence clinical isolates from samples with dissonant results.
EBC PCR results were also compared to BAL GM EIA values (n = 190). Concurrent negative results
were seen in 158 (83%) samples and positive results in 6 (3%) samples. The xTAG ASR assay
displayed sensitivity, specificity, positive and negative predictive values of 100%, 85.9%, 16.1%, and
100% when compared to GM EIA. PCR was also performed in matching BAL samples.
Conclusions: These results provide proof-of-concept that detection of fungal DNA is feasible in EBC.
Further evaluation to determine its role in prophylaxis and treatment is warranted.

Tables: 

2016

abstract No: 

#1984

Full conference title: 

26th European Congress of Clinical Microbiology and Infectious Diseases
    • ECCMID 26th (2016)