Entomophthoramycosis is a sporadic, slow-growing subcutaneous infection that includes two distinct clinical forms: Basidiobolus ranarum (subcutaneous) and Conidiobolus coronatus (Rhinofacial), common in tropical countries like India. A 20-year-old male presented with painless progressive swelling of the anterolateral aspect of the right thigh, with a history of Incision and Drainage (I&D) not responding to antibiotics. A clinical diagnosis of a soft-tissue tumour was made. Magnetic Resonance Imaging (MRI) showed a subcutaneous lesion with inflammatory changes and overlying skin changes. Core biopsy showed spindle cells. Surgeons proceeded with a wide local excision of the lesion. Histopathology {Haematoxylin and eosin (H&E), Grocott Methenamine Silver stain (GMS), Periodic Acid Schiff stain (PAS)} proved to be subcutaneous zygomycosis showing Splendore-Hoeppli phenomenon. The patient was then started on antifungal treatment followed by skin grafting. In the present case, the granulomatous inflammation was not picked up in multiple random core biopsies and hence, a wide excision was done, leading to an avoidable extensive surgery.
Histopathology, Subcutaneous, Zygomycoses
Case Report
A 20-year-old male zoology student presented with a progressively painless swelling of the anterolateral aspect of the right thigh for the past four months, which had an insidious onset and gradually increased in size. A history of I&D performed in the same region four months prior, followed by treatment with antibiotics, was reported. There was no history of fever or difficulty in walking at the time of presentation. A well-circumscribed swelling measuring 20×18 cm was noted over the anterior aspect of the right thigh. The skin covering the swelling showed hyperpigmentation, with a 0.3 cm linear scar present at the centre of the swelling [Table/Fig-1]. There was no warmth or tenderness, and the skin over the swelling was not pinchable. The surface of the swelling was smooth, and it felt firm to hard in consistency. The swelling showed mobility on both horizontal and vertical axes, but restricted mobility was noted upon muscle contraction.
Swelling over right thigh.
Complete blood count, renal function test, serology {Human Immunodeficiency Virus (HIV), Hepatitis B Virus (HEP-B), and Hepatitis C Virus (HCV)-negative}, urine routine, and total electrolyte panel results were all within normal limits. The patient was immunocompetent. A clinical diagnosis of a soft-tissue tumour was made, and an MRI of the thigh was recommended. The MRI revealed areas of heterogeneously hypointense and hyperintense regions indicating central fat necrosis, along with overlying skin thickening and surrounding inflammatory changes. Multiple linear core biopsies were taken from different areas of the swelling, showing proliferating spindle to plump cells, foamy histiocytes entrapped in adipose tissue, dense fibrocollagen, and chronic inflammatory infiltrate in the background. It was reported as a spindle cell lesion, and excision biopsy was suggested for an accurate diagnosis [Table/Fig-2].
Core biopsy showing adipose tissue and proliferating spindle cells (H&E, 4x). Inlet showing proliferating spindle cells (H&E, 40X).
The surgeons proceeded with a wide local excision of the swelling. Intraoperatively, a hard tumour measuring 20×18 cm was noted in the anterior aspect of the right thigh, extending up to the deep fascia and involving the sartorius muscle at the centre as well as a part of the tensor fascia lata, which was excised. The specimen was sent to Histopathology [Table/Fig-3a-d]. The patient was started on antibiotics and discharged on postoperative day 3, with instructions to return for a review after one week.
a) Excised specimen; b) Surgical surface of excised specimen; c) Thickness of the excised specimen; d) Cut surface showing white areas extending upto the underlying muscle layer.
A circular, disk-like specimen covered with skin and featuring a central bulge, weighing 882 grams and measuring 20×18 cm with a thickness of 3-4.5 cm, was received. The surgical surface exhibited glistening fascia with adherent skeletal muscle tissue in the centre. The cut surface showed many firm and depressed whitish areas mixed with yellow adipose tissue. These whitish areas did not reach the peripheral margin, with a clearance of 2 cm; however, they extended up to the underlying muscle layer in the deep surgical margin [Table/Fig-4a-d].
a) Fungal hyphae with the frill of Splendore-Hoeppli phenomenon surrounded by foreign body giant cells and proliferating spindle cells (H&E, 4X); b) Most hyphae were 8-10 microns in diameter (H&E, 40X); c) GMS staining revealed broad, ribbon-like, pleomorphic, aseptate or pauci-septate fungal elements (GMS, 100X); d) PAS staining showing fungal elements, 8-10 microns in width (PAS, 10X).
Microscopy of multiple sections revealed numerous micro-abscesses, suppurative palisaded epithelioid and foreign body type granulomas predominantly in the subcutaneous tissue, extending from the dermis to the adherent skeletal muscle. Extensive panniculitis with dense acute and chronic inflammation rich in eosinophils and foamy histiocytes was observed. Inflammation was not present in the fibrous deep surgical margin (clearance of 1-2 mm), but it extended into the adherent skeletal muscle. The surgical margin of the muscle was free from inflammation.
Definite angioinvasion by fungal elements was not detected in the multiple sections studied. Based on the clinical, gross, and microscopic findings, the features were suggestive of Entomophthoramycosis (subcutaneous zygomycosis). The patient was started on antifungal therapy (Fluconazole) for four weeks, and the lesion resolved in six months, following which split skin grafting was performed at the surgical site. The patient showed symptomatic improvement during the follow-up visits [Table/Fig-5].
Skin grafting done on follow-up after treatment with Fluconazole.
Discussion
Entomophthoramycosis is a rare fungal infection predominantly seen in tropical and subtropical regions. In Greek, “Entomon” means insect, reflecting that these fungi were originally identified as parasites infecting insects. It is a diagnosis that is often misdiagnosed and not thought of. Entomophthoramycosis is the most common clinical form of basidiobolomycosis, an endemic fungal infection in southern parts of India [1]. Basidiobolus species is a filamentous fungus isolated from amphibians, reptiles, horses, dogs, and bats, along with woodlice, plant debris, and soil. This fungus affects immunocompetent hosts [1] and shows a male predominance (M:F- 3:1), mostly affecting children (80% under the age of 20 years) [2]. Basidiobolus is classified into B. ranarum, B. meristosporus, and B. haptosporus. However, all human-pathogenic isolates belong to B. ranarum [2].
The fungus may be rare in tissue sections and when present, is often fragmented. Additionally, focal hyphae may only appear in part of the specimen. Moreover, fungal elements stain poorly with H&E and are not well demonstrated with fungus-specific tissue stains like periodic acid-Schiff. Examination of the fluorescent dye (Blankophor) wet-mount preparation under fluorescent microscopy increases the sensitivity of diagnosis [2]. Entomophthoramycosis has a predilection for areas with adipose tissue, possibly because these organisms thrive on fatty substances [3]. The disease manifests as disc-shaped, rubbery, mobile masses that can be quite large and are typically located in the shoulders, hips, or thighs [4]. The diagnostic challenge in such diseases is difficult and may initially be missed. The disease is prevalent in subtropical and tropical regions globally, although sporadic cases have been reported in Africa, Asia, the USA, and Latin America [4]. Subcutaneous zygomycosis usually results from the inoculation of fungal spores through the skin, possibly due to minor trauma such as an insect bite, intravenous catheter, or even intramuscular injection [5]. In the present case, there was no history of trauma, but there was a history of I&D. The Zygomycetes class of fungus is subdivided into two orders: Mucorales and Entomophthorales. Basidiobolus ranarum is a type of Entomophthorales and is typically associated with infections of the cutaneous and subcutaneous planes, without dissemination to internal organs [6].
The entomophthoralean fungi trigger a distinctive eosinophilic inflammatory response, with the formation of eosinophilic material around the invasive coenocytic hyphae of Basidiobolus and Conidiobolus species (Splendore-Höeppli phenomenon), which is absent in Mucorales [7]. Diagnosis in the present case was made based on the location of the lesion (thigh) and the eosinophilic reaction (Splendor-Hoepelli) on histopathology evaluation. The demonstration of hyaline, thin-walled, wide, non septate or scarcely septate hyphae, or hyphal fragments in Potassium Hydroxide (KOH) digests or in H&E stained sections of a biopsy from the actively growing part of a lesion is a very characteristic feature [8]. A pathognomonic feature is the presence of thin-walled, often septate hyphae or hyphal fragments (4-10 micrometres in diameter) with a thick eosinophilic sheath (Splendore-Hoeppli phenomenon) staining intense pink in H&E sections and bright red in PAS stained sections [9]. Similarly, based on histopathological findings only (hyphae with the Splendore-Höeppli phenomenon), a case of a five-year-old boy in Sudan with painless progressive swelling in the upper right forearm was reported [10]. The treatment involves prolonged anti-fungal therapy along with surgical debridement. More awareness of this fungus is warranted for definitive diagnosis and implementation of early proper therapeutic strategies [2]. A summary of 16 recent cases documented in English literature is presented in the following [Table/Fig-6] [5,10-21].
Summary of recently reported cases of subcutaneous entomophthoramycosis [5,10-21].
| Authors name | Age/Gender | Site | Duration | Geographical location | History of trauma | Treatment | Results |
|---|
| Verma RK et al., (2012) [10] | 42 years/F | Neck and temporal region-right | 15 days | Chandigarh | - | Intravenous (i.v.) amphotericin B and oral potassium | Complete resolution |
| Kumari PHP et al., (2013) [11] | 6 months/F | Left thigh | 4 months | Andhra Pradesh | Present | Saturated solution of potassium iodide | Not known |
| Jayanth ST et al., (2013) [12] | 58 years/F | Right gluteal region | 2 years | Chattisgarh | Im injection | Oral potassium iodide | Complete regression after 6 months of therapy |
| Anaparthy UR et al., (2014) [13] | 6 months/F | Left knee | 4 months | Andhra Pradesh | Insect bite | Saturated solution of oral potassium iodide | Complete regression after 8 weeks of therapy |
| Mondal AK et al., (2015) [5] | 25 years/F | Left arm | 8 months | West Bengal | - | Saturated solution of oral potassium iodide | Responded remarkably within 2 months |
| Arora P et al., (2015) [14] | 2 and half years/M | Left buttock and upper thigh | 6 months | New Delhi | Im injection | Saturated solution of potassium iodide | Complete resolution |
| Chintangunta SR et al., (2016) [15] | 12 years/M | Right thigh and buttock | 6 months | Telangana | - | Itraconazole | Complete resolution |
| Rajan RJ et al., (2017) [16] | 20 months/M | Left buttock | 6 months | Jharkhand | - | Potassium iodide and cotrimoxazole | Complete regression after 2 months of therapy |
| Luc VC Brun et al., (2018) [17] | 3 years/F | Right flank | 2 months | Buruli ulcer endemic area of Zagnanado | - | - | Lost to follow-up and no fungal treatment given |
| 43 years/M | Right thigh | 6 months | | | | Died unknown cause |
| 3 years/M | Right thigh | 1 month | | -- | -- | Died unknown cause (non ulcerated lesion to ulcerative lesion) |
| Nalini P et al., (2019) [18] | 75 years/M | Left forearm and arm | 6 months | Tamil Nadu | - | Saturated solution | Complete resolution at end of 3 months of therapy |
| Patro P et al., (2019) [19] | 4 years/M | Right upper arm | 4 months | Chhattisgarh | - | Itraconazole | Lesion started to resolve within first month |
| Katari Gopalakrishnan V et al., (2022) [20] | 80 years/M | | 3 months | Chennai | - | Itraconazole | Complete resolution |
| Choubey S et al., (2023) [21] | 56 years/F | Upper Back | 2 years | Bhopal | Wild parrot bite | Itraconazole | Complete resolution |
| Present case | 20 years/M | Right thigh | 6 months | Puducherry | Incision and drainage (I&D) | Fluconazole | Lesion resolved within 6 months and split skin graft was done |
Conclusion(s)
When a patient presents with a subcutaneous swelling in a tropical country like India that is not resolved with antibiotics, a high suspicion is needed to rule out entomophthoramycosis through microbiology culture and histopathological biopsy examination at the time of presentation to prevent misdiagnosis, unnecessary extensive surgery, and disfigurement.
Author Declaration:
Financial or Other Competing Interests: None
Was informed consent obtained from the subjects involved in the study? No
For any images presented appropriate consent has been obtained from the subjects. No
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