A Novel Exophiala Species Associated With Disseminated Granulomatous Inflammation in a Captive Eastern Hellbender (Cryptobranchus alleganiensis alleganiensis).

Cynthia Hopf,Noha Abou-Madi,Hongxin Fan,Carmita Sanders,Michael M Garner,Connie F C Gibas,Robert Ossiboff,James Mele,Nathan P Wiederhold,Erin A Graham

doi:10.3389/fvets.2020.00025

Abstract

The genus Exophiala is composed of ubiquitous, pigmented, saprotrophic fungi and includes both terrestrial and waterborne species. Though Exophiala species are generally considered opportunistic pathogens, exophialosis can be an important cause of morbidity and mortality in aquatic and semi-aquatic species. Over a 6-year period, a captive 32-year-old male eastern hellbender (Cryptobranchus alleganiensis alleganiensis), was treated for recurring, slow growing, ventral midline cutaneous masses. Excisional biopsies were characterized histologically by granulomatous dermatitis with low numbers of intralesional, pigmented fungal conidia and hyphae. Bacterial and fungal cultures of the masses and skin were negative on two separate submissions. Polymerase chain reaction amplification of a short fragment of the fungal 28S large subunit (LSU) ribosomal RNA was positive with 100% nucleotide sequence identity to several species of Exophiala. Following recurrence after successive rounds of antifungal therapy, euthanasia was elected. At necropsy, similar dermal granulomatous inflammation and intralesional pigmented fungal elements as observed in excisional biopsies formed a thick band in the dermis and extended through the coelomic body wall. Visceral dissemination was noted in the lung and kidney. Postmortem DNA sequence analysis of a large portion of the fungal LSU as well as the internal transcribed spacer (ITS) from a portion of frozen affected dermis identified the fungus as a novel species, Exophiala sp. 1 (UTHSCSA R-5437).

Highlights

Native to the United States, hellbenders (Cryptobranchus alleganiensis) are large aquatic salamanders represented by two subspecies: the eastern hellbender (C. a. alleganiensis) and the Ozark hellbender (C. a. bishopi)
The phylogenetic tree inferred from maximum likelihood (Figure 3) showed our isolate within a monophyletic clade (BPP 0.99/ BT 93%) comprising mostly of waterborne species, i.e., E. opportunistica, E. lacus, E. cancerae, E. psychrophila, E. salmonis, E. aquamarina, E. pisciphila [7] and other species originally isolated from non-aquatic sources, i.e., E. bonariae [22] and E. equina [7]
Phaeohyphomycosis is caused by pigmented filamentous fungi including Exophiala species [23]

Summary

INTRODUCTION

Native to the United States, hellbenders (Cryptobranchus alleganiensis) are large aquatic salamanders represented by two subspecies: the eastern hellbender (C. a. alleganiensis) and the Ozark hellbender (C. a. bishopi). Gross necropsy revealed a chain of multinodular, coalescing, and ulcerated dermal masses of the ventral coelomic body wall (Figure 2A). Several tissues were sampled and fixed in 10% neutral buffered formalin for microscopic examination and frozen for further testing, including the body wall sections of granulomatous lesion, sections of skin and mass, left distal thoracic limb, eye, brain, heart, urinary bladder, lungs, liver, spleen, kidneys, pancreas, intestines, segments of small and large intestine, and testicular tissue. The phylogenetic tree inferred from maximum likelihood (Figure 3) showed our isolate within a monophyletic clade (BPP 0.99/ BT 93%) comprising mostly of waterborne species, i.e., E. opportunistica, E. lacus, E. cancerae, E. psychrophila, E. salmonis, E. aquamarina, E. pisciphila [7] and other species originally isolated from non-aquatic sources, i.e., E. bonariae [22] and E. equina [7]. ITS pairwise comparison of UTHSCSA R-5437 with E. cancerae CBS 120420T (NR_1377664) showed 15 nucleotide base pair differences

DISCUSSION

Findings

ETHICS STATEMENT