Transcriptome profiling of lung immune responses potentially related to acute respiratory distress syndrome in forest musk deer

Abstract

Background: Forest musk deer is an endangered species globally. The death of captive forest musk deer can be caused by certain respiratory system diseases. Acute respiratory distress syndrome (ARDS) is a huge threat to the life of forest muck deer that breed in our department.

Methods: Lung histopathologic analysis was conducted by hematoxylin and eosin (HE) staining. The lung gene changes triggered by ARDS were examined by RNA sequencing and related bioinformatics analysis in forest musk deer. The potential functions of unigenes were investigated by NR, SwissProt KOG, GO, and KEGG annotation analyses. Vital biological processes or pathways in ARDS were examined by GO and KEGG enrichment analyses.

Results: A total of 3265 unigenes were differentially expressed (|log₂fold-change|> 2 and adjusted P value < 0.01) in lung tissues of 3 forest musk deer with ARDS compared with normal lung tissues of the non-ARDS group. These differentially expressed unigenes (DEGs) played crucial roles in immunity and defense responses to pathogens. Moreover, we identified the DEGs related to one or more of the following biological processes: lung development, immunity, and bacterial/viral/fungal infection. And six DEGs that might be involved in lung injury caused by immune dysregulation or viral/fungal infection were identified.

Conclusion: ARDS-mediated lung gene alterations were identified in forest musk deer. Moreover, multiple genes involved in lung development and lung defense responses to bacteria/viruses/fungi in ARDS were filtered out in forest musk deer.

Keywords: Bacterium; Forest musk deer; Fungus; Immunity; Lung; RNA sequencing; Virus.

Fig. 1

Anatomical and histopathologic analysis of forest musk deer lung tissues. (A-C) The clinical-pathological alterations of lung tissues of 3 forest musk deer that died of ARDS. (D) HE staining image of normal lung tissue. Black arrow: alveoli; Red arrow: alveolar wall. (E-G) HE staining images of lung tissues of forest musk deer with ARDS. The black arrow in Fig. 1E-1G represents the pus cells in the alveolar space. The red arrow in Fig. 1E indicates the exudates in the alveolar space. The red arrow in Fig. 1F denotes the alveolar wall capillary congestion. The red arrow in Fig. 1G indicates pulmonary interstitial congestion. The yellow arrow in Fig. 1G indicates a large number of pus cells in the bronchi

Fig. 2

Statistical analysis of unigenes. (A) Length distribution patterns ofunigenes. (B) GC content frequency distribution patterns of unigenes

Fig. 3

Species and functional annotations of unigenes. (A) Species distribution of unigenes annotated by the NR database. (B) GO annotation analysis for unigenes. (C) KEGG annotation analysis for unigenes

Fig. 4

Identification of DEGs and related GO/KEGG enrichment analysis. (A) Heatmap of DEGsin diseased lung tissues of forest musk deer compared to the normal lung tissue group.The R package v3.6.3 and ComplexHeatmapR package v2.2.0 were used to create the heatmap. (B) Top 20 KEGG pathways enriched by DEGs [20]

Fig. 5

Venn analytical outcomes of unigene sets in Tables S6-S10. The upper section shows the Venn diagram of unigenes in Tables S6-S10. The middle histogram presents the number of unigenes in Tables S6-S10.The chart below displays the number of specific (1) elements of one list or shared elements by 2, 3, … lists. For instance, a total of 332 unigenes were specific to one list. In addition, 79 unigenes, 20unigenes, and 1 unigenewere shared by 2, 3, and 4 of the 5 lists, respectively