Determination of the mycobiome in the lower respiratory tract of horses with equine asthma

Abstract

Fungal particles are important allergenic components involved in the development of equine asthma. The aim of this study was to evaluate the mycobiome composition of the lower respiratory tract in asthmatic horses using fungal culture, quantitative multiplex real-time PCR analysis (FungiMultiPlex) and Next-Generation Sequencing approach. Bronchoalveolar lavage fluid (BALF) samples obtained from 45 client-owned horses diagnosed with equine asthma were analysed by fungal culture (19 samples), FungiMultiPlex (34 samples), and Next-Generation Sequencing (14 samples). The fungal culture was positive in 11/19 (58%) cases, and FungiMultiPlex was positive in 19/34 (56%) cases. No fungal PCR product was detected by Next-Generation Sequencing analysis. Fungal culture and FungiMultiPlex methods were performed simultaneously on eight horses only. Association of results of these methods was calculated using Phi coefficient (φ= 0.333), and concordance between the methods was not confirmed (P = 0.420). The results of this study suggest that the fungal culture and quantitative multiplex real-time PCR might be considered diagnostically useful to assess the presence of fungi in BALF in a semiquantitative and quantitative manner, respectively. The Next-Generation Sequencing method seems to be less diagnostically suitable due to technical obstacles pertinent to both the low concentration of microbial agents in the rather diluted BALF samples, and, also, due to the relatively high environmental background contamination of the collected material. Based on our data, we advocate the use of the combination of quantitative multiplex real-time PCR and fungal culture in a routine clinical diagnostic setting.