Systemic study of the host-pathogen interaction in talaromyces marneffei infection

Abstract

Talaromycosis, caused by Talaromyces marneffei, has emerged as a life-threatening pathogen in different groups of immunocompromised patients. A better understanding of host-pathogen interaction is particularly important for developing strategies for disease control and prevention amongst such patients. However, the pathogenesis – including factors determining disease outcome – remains largely unknown. In the first part of the study, we aimed to describe the roles of both macrophages and respiratory cells upon encountering the airborne T. marneffei. We established an in vitro human peripheral blood monocyte-derived macrophages (PBDMs) model to study the host innate immune responses against T. marneffei during early infection. Moreover, to obtain a global transcriptome profile of macrophages upon T. marneffei infection, we performed a series RNA–Seq using macrophages isolated from three independent donors. As a result, 452 differential expressed genes (DEGs; p < 0.001 and q < 0.05) were found to be significantly up- or down-regulated during the infection course. Gene ontology (GO) enrichment and KEGG functional analysis revealed that most of these genes play a significant part in immune responses and are involved in M1 polarization. Next, by using a green fluorescent protein-tagged T. marneffei strain, we showed the direct interaction of human bronchial epithelial cells (hBECs) with T. marneffei live conidia. Fascinatingly, we found that these live conidia were taken up by hBECs, but the induction of relevant cytokines – previously reported as important in activating airway epithelial cells during Aspergillus fumigatus infection – could not be detected. In addition, these infected cells have been widely observed to undergo a cell division process, and it is most likely that live pathogens are carried by progeny cells. The survival rate of T. marneffei was found to be markedly higher from co-cultures with hBECs than those with PBDMs (p < 0.01), implicating the possible role of hBECs as reservoir cells. In the second part of the study, we aimed to explore the adaptation mechanisms of T. marneffei during early infection and to identify a set of genes that represent phase-specific markers by using a time series RNA–Seq. We identified 520 gene loci (522 DEGs) as being notably up- or down-regulated. Principal component analysis revealed that the expression values over these DEGs explain 57.4% and 23.6% of the total variance, with the effect greatly differing from zero hours post-infection, indicating that T. marneffei early transcriptional levels were affected by events that occurred during a much broader incubation time. These genes were curtailed for GO definitions and also blasted with Saccharomyces cerevisiae for functional annotation. The molecular interactions map showed that these DEGs had significantly higher network connectivity. KEGG pathway mapping analysis and GO enrichment analysis showed that these DEGs were found to be enriched in ribosome biogenesis, endosomal transportation, and metabolic processes. Furthermore, clustering functional analysis revealed a switch of metabolite pathways and an activation of alternative pathways. To our best knowledge, this is the first intracellular transcriptome analysis of T. marneffei in response to macrophages. Mechanisms applied by the fungal pathogen to enhance its survival are also described here.