Fortin et al. Acta Vet Scand (2017) 59:63 DOI 10.1186/s13028-017-0330-z Acta Veterinaria Scandinavica CASE REPORT Open Access Sublingual pythiosis in a cat Jessica Sonia Fortin1, Michael John Calcutt2 and Dae Young Kim1* Abstract Background: Pythiosis is a potentially fatal but non-contagious disease afecting humans and animals living in tropi- cal and subtropical climates, but is also reasonably widespread in temperate climates, throughout the world. The most commonly reported afected animal species with pythiosis are equine and canine, with fewer cases in bovine and feline. Extracutaneous infections caused by Pythium insidiosum have been rarely described in the cat. Case presentation: Sublingual pythiosis was diagnosed in a 2-year-old, male, Domestic Shorthair cat. The cat had a multilobulated, sublingual mass present for 3 months. Histopathological examination revealed severe multifocal coalescing eosinophilic granulomatous infammation. Centers of the infammation contained hyphae that were 3–7 μm-wide, non-parallel, uncommonly septate and rarely branching. The fungal-like organism was identifed as P. insid- iosum by polymerase chain reaction and subsequent amplicon sequencing. Conclusions: Only a few feline pythiosis cases have been reported and, when encountered, it usually causes granu- lomatous diseases of the skin or gastrointestinal tract. This case presents an unusual manifestation of feline pythiosis, representing the frst involving the oral cavity in cats or dogs. Keywords: Eosinophilic granulomatous infammation, Feline, Hyphae, Pythium insidiosum, Sublingual mass Background trauma in Afghanistan [5]. Animals afected by the dis- Pythiosis is a potentially fatal but non-contagious disease ease are often younger and exposed to warm, freshwa- afecting humans and animals living in tropical and sub- ter habitats [1]. Te infective form is the asexual motile tropical climates, but that is emerging and now reason- zoospore produced in aquatic environs hypothetically in ably widespread in temperate climates, throughout the association with plant materials that presumably causes world. Notably, it has been reported in other countries infection by encysting on injured skin or a mucosal sur- such as the Republic of Korea, Japan and Haiti [1]. Te face [1, 2, 4, 6]. Defnitive identifcation relies on serology, infection is contracted typically during the warm months, immunohistochemistry and molecular diagnostics, but from early spring to late fall [2]. Pythiosis, caused by not solely on morphological features, since histopatho- Pythium insidiosum, represents a life-threatening infec- logical features of fungal pathogens such as coenocytic tion in humans and mammals. Members of the Pythium fungi are similar [7, 8]. genus, class Oomycota, are water or soil dwelling and Pythiosis was frst identifed in the horse during the phylogenetically distinct from fungi, rather closely 19th century [4]. Te frst reported case of pythiosis in related to diatoms and algae [3]. Many Pythium sp. are a dog, cat and human occurred respectively in 1971 [9], economically important plant pathogens [1, 4]. Pythium 1991 [10] and 1989 [11]. In animals, the infection exhibits insidiosum is the only mammalian pathogen implicated diferent clinical presentation in cutaneous, subcutane- previously in this genus. Over the last decade, a case of ous, intestinal, and disseminated manifestations depend- Pythium aphanidermatum invasive wound infection ing on the route of entry. Skin infection is most common has been reported in an injured man following combat in animals. Interestingly, in dogs, gastrointestinal pythio- sis occurs more frequently than the cutaneous/subcu- taneous clinical presentation [6]. Pythiosis occurs most *Correspondence: [email protected] frequently in horses and dogs, and only a “handful” of 1 Veterinary Medical Diagnostic Laboratory, University of Missouri, 810 E Campus Loop, Columbia, MO, USA feline cases have been reported. Te purpose of this case Full list of author information is available at the end of the article report is to describe an unusual manifestation of feline © The Author(s) 2017. This article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/ publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated. Fortin et al. Acta Vet Scand (2017) 59:63 Page 2 of 5 pythiosis, representing the frst involving the oral cav- hyphae. Te hyphae were 3–7 μm-wide, non-parallel, ity in pet animals. Te general aim is to promote global uncommonly septate and rarely branching. Hyphal struc- monitoring of potentially emerging fungal-like diseases tures were partially evidentiated with a Gomori methe- in uncommonly afected animal species since moving and namine silver (GMS) stain (Fig. 2c). Te hyphae are travelling with pets is common presently. morphologically similar to those of oomycetes. Based on the microscopic characteristics of the infammation and Case presentation intralesional hyphal structures, P. insidiosum infection A 2-year-old, male, Domestic Shorthair cat was pre- was highly suspected. Although not encountered in the sented at a central Missouri veterinary clinic with a mul- cat and infection of the oral cavity has not been reported, tilobular, irregular and frm sublingual mass measuring lagenidiosis was also considered as a diferential diagno- 2.5 × 2 × 1 cm (Fig. 1). Te cat was an indoor/outdoor sis because of close similarities of microscopic lesions cat and had no travel history. Te mass was present for and hyphae to pythiosis [1]. Te infammation containing 3 months of duration, according to the owner, prior hyphal organisms reached the surgical margins so that it to consultation. No pharmacological treatment was was unclear whether the lesion was completely excised or attempted. Incisional biopsy was performed to obtain a not. Te histopathologic evaluation was summarized as diagnosis. Te formalin-fxed specimen from the mass a severe eosinophilic granulomatous infammation with was submitted for histopathologic evaluation to the Uni- intralesional fungal hyphae. versity of Missouri Veterinary Medical Diagnostic Labo- Te identifcation of the fungal-like organism as P. ratory, USA. insidiosum was carried out by polymerase chain reaction Te mass was fxed in 10% neutral bufered formalin, (PCR) and amplicon sequencing using DNA extracted trimmed and embedded in parafn. Parafn sections from formalin-fxed parafn-embedded tissue [12]. PCR (4 µm) were stained with hematoxylin and eosin (HE). was carried out using the Expand Long Template PCR Histopathological examination revealed severe multifocal System (Roche, Indianapolis, IN, USA) with oligonucleo- to coalescing areas of eosinophilic granulomatous infam- tide primers that amplify the complete ribosomal Internal mation, characterized by multifocal central areas of Transcribed Spacer 1 (ITS1) region. Te resulting ampli- necrosis with degenerated eosinophils, which were sur- con was purifed by spin chromatography (QIAquick rounded by large numbers of macrophages, epithelioid PCR Purifcation Kit, Qiagen, Valencia, CA, USA) and macrophages, lymphocytes and plasma cells and occa- nucleotide sequence determined using BigDye termina- sional multinucleated cells (Fig. 2a, b). Te infammation tor chemistry with the amplifcation primers at the Uni- extended to the underlying skeletal muscle. Te central versity of Missouri DNA Core Facility. Sequence analysis areas of necrosis contained faintly stained fungal-like disclosed the identity of the pathogen as P. insidiosum. Fig. 1 Lateral views of the sublingual mass 5 days after the biopsy (a right, b left). A 2-year-old, male, Domestic Shorthair cat had a multilobulated, sublingual mass present for 3 months. Sublingual pythiosis was diagnosed following histopathological examination from a biopsy specimen Fortin et al. Acta Vet Scand (2017) 59:63 Page 3 of 5 Fig. 2 Photomicrographs of the intralesional fungal hyphae (black arrow) within the severe sublingual eosinophilic granulomatous infammation. a Lower magnifcation of the sublingual mass composed of eosinophilic granulomatous infammation. b The hyphae were 3–7 μm-wide, non- parallel, occasionally septate with rare branching. Hematoxylin and eosin (HE) stain. c Hyphal structures are evidentiated with Gomori methenamine silver (GMS) stain Te sequence of the complete ITS1 region amplifed from of pythiosis involving the hard palate and nasal cavity in the cat tissue (GenBank accession MF928597) was > 99% a sheep from southwestern Paraná, southern Brazil, was identical to multiple GenBank entries from P. insidio- confrmed by indirect enzyme-linked immunosorbent sum (identifed by using Blastn). ITS1 sequences from assay and immunohistochemistry [14]. Interestingly, two three strains submitted to GenBank as Pythium destruens cases of canine chronic esophagitis caused by P. insidio- exhibited 93–94% identity to that described herein and sum occurred previously in rural Tennessee, USA [15]. those from Pythium grandisporangium were < 82% iden- Cases of oral cavity pythiosis have not been diagnosed tical to the sequence obtained from the cat lesion. in cats,