Temporal shifts in the mycobiome structure and network architecture associated with a rat (Rattus norvegicus) deep partial-thickness cutaneous burn

Abstract

With a diverse physiological interface to colonize, mammalian skin is the first line of defense against pathogen invasion and harbors a consortium of microbes integral in maintenance of epithelial barrier function and disease prevention. While the dynamic roles of skin bacterial residents are expansively studied, contributions of fungal constituents, the mycobiome, are largely overlooked. As a result, their influence during skin injury, such as disruption of skin integrity in burn injury and impairment of host immune defense system, is not clearly delineated. Burn patients experience a high risk of developing hard-to-treat fungal infections in comparison to other hospitalized patients. To discern the changes in the mycobiome profile and network assembly during cutaneous burn-injury, a rat scald burn model was used to survey the mycobiome in healthy (n = 30) (sham-burned) and burned (n = 24) skin over an 11-day period. The healthy skin demonstrated inter-animal heterogeneity over time, while the burned skin mycobiome transitioned toward a temporally stabile community with declining inter-animal variation starting at day 3 post-burn injury. Driven primarily by a significant increase in relative abundance of Candida, fungal species richness and abundance of the burned skin decreased, especially in days 7 and 11 post-burn. The network architecture of rat skin mycobiome displayed community reorganization toward increased network fragility and decreased stability compared to the healthy rat skin fungal network. This study provides the first account of the dynamic diversity observed in the rat skin mycobiome composition, structure, and network assembly associated with postcutaneous burn injury.

Keywords: Rattus norvegicus; burned skin mycobiome; deep-partial thickness burn; rat skin mycobiome; skin fungal community structure; skin fungal network assembly.

Figure 1.

Within-community (alpha) diversity longitudinal analysis of healthy and burned rat skin and exposome mycobiome. (A) With the greatest variability observed in the rat exposome, the species richness estimate of healthy (unburned) skin was slightly higher than the PTB skin community. (B) Fungal species abundance was highest in the healthy rat skin when compared to the mycobiome of the burned skin and the exposome. (C) Species evenness of the healthy skin mycobiome was the highest relative to the burned skin and the exposome. (D) The healthy rat skin mycobiome harbored the most number of unique OTUs per community (richness) than the burned skin. Greatest variability was observed in the rat exposome. (E) Phylogenetic diversity of the PTB skin mycobiome was reduced compared to the healthy skin with greatest variability observed in the rat exposome mycobiome. (F) Species richness estimate generated from alpha diversity rarefactions declined over time in both healthy and PTB skin mycobiome; while healthy skin specimen showed a rise by POD 11, species richness of PTB skin continued to decline. (G) Fungal species evenness showed a pattern of decline in the PTB skin mycobiome starting at POD 3, while the healthy skin specimen showed increased evenness by POD 11. (H) PTB skin showed a decreasing pattern of species abundance that continued through POD 11.

Figure 2.

Phylogenetic profile of rat skin mycobiome. (A) Phyla distribution in healthy rat skin mycobiome was dominated by Ascomycota and Basidiomycota. (B) Healthy rat skin mycobiome showed a degree of heterogeneity in genera distribution that was associated with temporal changes over 11-day study period. (C) Phyla profiling of compromised PTB skin mycobiome was mainly composed of Ascomycota and Basidiomycota. (D) Genera distribution exhibited variations associated with temporal changes including an increase in Candida and a decline in members of Nectriaceae, Capnodiales (Cladosporium and Cercospora), and Wallemia in relative abundance.

Figure 3.

Beta-diversity analysis of fungal communities in rat skin and exposome mycobiome. (A) Bray-Curtis analysis demonstrated a grouping of fungal specimens that separate a group of PTB specimens from remaining specimens. The rat exposome specimens were scattered between burned and unburned rat skin fungal communities. (B) When temporal changes are considered, majority of PTB of POD 7 and 11 skin specimens were separated into a cluster, while PTB specimens of POD 1 and 3 were scattered between the two clustering groups. The healthy skin specimens exhibited inter-animal variation across time. (C) Monocyte levels (%) obtained from CBC analysis of rat blood showed elevated monocyte values associated mostly with the PTB skin specimens of POD 7 and 11. Only a subgroup of rat skin specimens (27 rats) were randomly selected for undergoing CBC analysis. (D) Comparison distance plot of mycobiome sequence abundance (Bray-Curtis) over time for healthy and PTB rat skin specimens (Monte Carlo permutations = 2000) demonstrated a fungal shift between POD 3 and POD 7 in PTB skin relative to healthy skin community. (E) Significant enrichment of Candida in specimens exhibiting elevated monocyte levels (%) (FDR P = .03).

Figure 4.

Rat skin mycobiome network structure analysis in healthy and burned skin. (A) The healthy skin mycobiome network displayed a large connected network component that was dominated by members of Ascomycota. (B) The PTB skin fungal network assembly exhibited one large connected component along with smaller disconnected networks (e.g., singleton, dyad, and triad). Each node is colored by phyla and edges between nodes represent predicted interactions.