TROPICAL GROVE

Subcutaneous due to in a Filipino child responding to : a case report

Maria Christina Corazon C. Gonzalez, MD, DPDSa, Eleanor L. Letran, MD, FPDSb

ABSTRACT Subcutaneous zygomycosis is caused by Basidiobolus ranarum (B. ranarum) which is endemic in parts of Africa, India and other parts of Asia. We report an evolving case of a Filipino male child who presented with chronic and persistent subcutaneous nodules and plaques on the left extremity. Dense cell infiltrates consisting of lymphocytes, histiocytes and eosinophils were seen on histology with non-septated hyphal structures on Gomori’s methenamine silver stain. Fungal culture of the lesion yielded B. ranarum. Complete resolution of the lesions were observed after 6 months of itraconazole therapy alone.

Key Words: Zygomycosis, , , Basidiobolus ranarum, Itraconazole, Pediatric dermatology

INTRODUCTION CASE REPORT ubcutaneous zygomycosis or basidiobolomycosis A 12-year-old Filipino male, from Bataan, Philippines, is caused by a saprophytic fungi Basidiobolus initially presented with five 1 x 1 cm painless S ranarum (B. ranarum). It presents clinically as subcutaneous nodules on the left arm of 1 month a slowly growing painless subcutaneous nodule duration sought consult with a dermatologist and more commonly on the extremity, trunk and rarely was assessed with a provisional diagnosis of lupus on other parts of the body. Mode of infection has not panniculitis. Patient had already taken a week of been established but most likely follows traumatic prednisone at 0.5 mg/kg/day and amoxicillin-clavulanic implantation. We report a gradually evolving case of acid at 40 mg/kg/day as prescribed, with no noticeable basidiobolomycosis in a 12-year-old immunocompetent change. He was subsequently referred to our institution Filipino male with no history of trauma who responded for skin punch biopsy and management. Patient denies dramatically to itraconazole. trauma nor any other systemic symptoms.

Patient appeared well, not in distress, with his height (z score <1) and body mass index (z score <0) normal for his age. The skin appeared normal over the 10 x 6 cm deeply palpable non-tender, movable, firm, ten 1 x 1 cm aResident graduate, Department of Dermatology, University subcutaneous nodules some coalescing into plaques on of Santo Tomas Hospital bConsultant dermatologist, Department of Dermatology, the lower third posterolateral aspect of left arm (Fig. 1a). University of Santo Tomas Hospital There were no palpable lymphadenopathy, peripheral Department of Dermatology, University of Santo Tomas nerve enlargements, muscle atrophy, contractures, Hospital, España, Manila anhidrosis, motor and sensory deficits. The rest of the systemic examination was unremarkable. Differential Corresponding Author: Maria Christina Corazon C. diagnoses such as Hansen’s disease, tuberculosis or Gonzalez, MD deep fungal infection were considered. A deep double +63 (02) 712-1422 skin punch biopsy on the nodule revealed a densely [email protected] infiltrated subcutaneous layer with lymphocytes, Conflict of interest: None histiocytes, occasional epithelioid giant cells and Source of funding: None several eosinophils (Fig. 2a). An initial diagnosis of granulomatous panniculitis was given. Both Fite-Faraco and Periodic acid-Schiff stains (PAS) were negative.

78 J Phil Dermatol Soc · November 2020 · ISSN 2094-201X a b c On Blood Agar Plate, creamy, radially folded colonies with aerial hyphae were seen (Fig. 3b). Lactophenol cotton blue wet mount preparation showed numerous zygospores (Fig. 3c). The was identified as B. ranarum.

Patient was started with Itraconazole 10 mg/kg body weight daily (Fig. 4a). The lesions began regressing 2 weeks after initiation of therapy. Monthly follow-up visits revealed aminotransferase levels within normal ranges. Significant decrease in limb girth were observed Figure 1. Clinical photographs showing evolution of every month with complete healing of the entire lesion multiple subcutaneous nodules coalescing by 12 weeks of therapy (Fig. 4b). Itraconazole was into plaques on the left arm (a) at time of continued for 12 more weeks after compete resolution presentation, (b) at 4 months, and (c) at11 without any adverse events. Post-inflammatory months.

Urinalysis, fecalysis, serum creatinine, fasting blood a b sugar, alanine and aspartate aminotransferase liver enzymes, chest and left arm radiography were within normal range. Antinuclear and anti-double stranded DNA tests were negative. Mild peripheral eosinophilia and 12.5 times elevated total IgE were noted.

He came back a month later, with previous lesions increasing in size to 10.5 x 6 cm extending distally to the upper third of forearm. This time, an incisional Figure 2. (a) Densely infiltrated subcutaneous layer with biopsy of the nodule revealed acute and chronic lymphocytes, histiocytes, occasional epithelioid inflammation, fibrosis, granulation tissue, foreign body and several eosinophils. (H&E, x400) (b) Gomori’s giant cell reaction and increased tissue eosinophilia. methenamine silver (GMS) stain revealed non- Polarized light to detect foreign particles was suggested septated hyphal structures (black solid arrow). but was not done due to financial constraints. Patient (GMS, x400) came back three months later with appearance of five hypopigmentation was noted on previously affected new erythematous 1 x 1 cm slightly tender nodules areas. Hypertrophic scarring was noted on previous and progression in size of previous lesions with largest biopsy sites. No recurrence was noted 30 months post- plaque now measuring 11 x 10 cm which became treatment. harder on palpation. There was also a noticeable 2 cm increase in limb girth (Fig. 1b). Purified protein DISCUSSION derivative skin test was negative. He came back seven months later with some lesions becoming more Zygomycosis describes a group of infections caused by erythematous and swollen spreading further distally fungi of orders Entomophthorales and of to half of left forearm measuring 11 x 11.5 cm with the class . The former has three important draining seropurulent discharge (Fig. 1c) accompanied pathogenic species namely B. ranarum, Conidiobolus by occasional moderate pruritus. As a deep bacterial or coronatus and Conidiobolus incongruous.¹ Recent fungal infection (mycetoma) was strongly considered, taxonomic studies based on antigenic analysis, a 3 x 5 cm excisional biopsy was done revealing dense isoenzyme banding, and restriction enzyme analysis of cell infiltrates consisting of lymphocytes, histiocytes recombinant DNA indicate that all human pathogens and many eosinophils. PAS stain was negative for fungal belong to B. ranarum.² This is the causative agent of elements, but Gomori’s methenamine silver (GMS) subcutaneous zygomycosis (basidiobolomycosis). This stain revealed non-septated hyphal structures (Fig. 2b). uncommon disease is mainly encountered in tropical Routine aerobic culture showed no growth after 7 days areas of Asia, Africa and South America.³ B. ranarum are of incubation. Fungal culture on Saboraud’s Dextrose saprophytes found in soil, decaying vegetable matter, Agar (SDA) grew flat, waxy, whitish, centrally heaped and and of amphibians, reptiles, fish, radially folded aerial hyphae and earthy odor (Fig. 3a). and bats.¹

J Phil Dermatol Soc · November 2020 · ISSN 2094-201X 79 Clinically, the disease begins as a hardened firm that may reveal presence foreign material are crucial in nodule on extremities or trunk which may progress evaluating granulomatous inflammation. Hematoxylin to expand locally to form a hard, painless, non-tender and eosin (H&E) staining of the biopsy often but not swelling.4 The mass does not adhere to deeper tissues, always reveals a hyphae surrounded by eosinophilic but involvement of muscle were reported previously.5 materials a phenomenon known as Splendore- Although the clinical appearance of the lesions of Hoeppli. This has been hypothesized to be due to a basidiobolomycosis is very suggestive, presence of predominant T-helper 2 type of immune response with chronic subcutaneous nodules and histopathologic the release of the cytokines IL-4 and IL-10 which in turn findings of granulomatous inflammation does not recruits eosinophils to the affected site.² In addition, entirely rule out other granulomatous diseases such when tissue components are sparse, fungi are easily as tuberculosis and Hansen’s disease which remain overlooked in H&E sections. Special stains are therefore endemic in the Philippines.6,7 The definitive diagnosis important for histopathologic evaluation of unexplained requires isolation of the causative agent. Peripheral inflammatory processes, as well as further delineating blood eosinophilia was evident in this case, but it is the type of culture to be used. PAS and GMS stains a nonspecific indicator of several diseases requiring are the two commonly used stains to identify fungi in

a b c

Figure 3. (a) Growth of Basidiobolus ranarum on Saboraud’s dextrose agar showing flat, waxy, whitish, centrally heaped and radially folded with aerial hyphae. (b) Growth of Basidiobolus ranarum on Blood agar plate showing creamy, radially folded colonies with aerial hyphae. (c) Photomicrograph of lactophenol cotton blue wet mount preparation showing numerous zygospores (original magnification: x400)

investigation of an underlying cause.⁸ While direct a b examination may suggest the diagnosis, culture remains the gold standard for diagnosis⁹. However, recovery of fungi could also be difficult.10 In a report by Davis et al, despite the typical clinical manifestation of the case, fungal hyphae were not identified in the tissue sections from the initial debridement and were only identified after repeated screening of deeper sections11, which caused a delay in diagnosis. This may be due to the aggressive processing of the specimen that occurs before plating, where fungal hyphae are often damaged and become non-viable.12 Due to these difficulties in culture techniques and because successful management relies on early diagnosis, microscopic identification of characteristic fungi should be considered significant Figure 4. (a) Clinical photograph documenting increase despite failure to recover the offending fungus in in size and number of subcutaneous nodules culture.10,12 with some excoriations and hypetrophic scarring on previous biopsy sites pre-treatment, and The need for cultures and special stains to identify (b) return of limb to normal after 3 months of fungal and bacterial causes, as well as polarized light itraconazole therapy.

80 J Phil Dermatol Soc · November 2020 · ISSN 2094-201X tissues and cytology specimens. In screening for fungi, vomiting, or abdominal pain; other reactions include PAS stain demonstrates fungal morphology better than rash, dizziness, sleepiness, headache, and abnormalities the silver stain. Moreover, PAS can stain degenerated in liver function tests. No fatal reactions have been fungi that may not be visible on H&E stain. GMS, on the noted and toxicity has rarely led to a discontinuation of other hand, provides better contrast, and stains even therapy.25,26 degenerated and non-viable fungi that are sometimes refractory to PAS stains.13 The PAS stains in the present Patients treated with appropriate antifungals show case were done twice. This underscores the importance complete resolution of the disease. Patro et al of requesting both stains in histologic evaluation of reported a case of a 4-year old male child with a large granulomatous panniculitis to include or exclude the subcutaneous lesion on the right distal arm, which presence of fungi. Similar to a previous report14, SDA was initially misdiagnosed as a soft tissue tumor. culture of the present case also showed white waxy and Excellent respons were noted with oral itraconazole glaborous colony with many radial folds. On lactophenol therapy alone wherein lesions started to resolve cotton blue wet mount, zygospores of basidiobolus within a month, but patient was lost to follow up 20 typically possess two lateral appendages which are after 2 months. Sackey et al also reported success in conjugation beaks.15 treatment of basidiobolomycosis in a 3-year old girl as manifested by marked reduction in right leg size and No underlying disease seems to predispose patients to inguinal lymphadenopathy after several weeks of oral infection.1,16 The mode of infection is not known but is itraconazole, and complete healing of ulcers after 10 assumed to be through the skin after insect bites, minor weeks of treatment.21 Mahamaytakit et al reported two skin abrasions17, or sometimes iatrogenic.14 However, a cases; a 2-year old female with with slow expanding history of previous trauma is seldom obtained.4,15 While mass at the buttock, and a 10-month old male with the patient confirms occasional exposure to grassy areas prolonged high fever and rapidly expanding perioral in his school, he denies sustaining any skin trauma prior ulcerated plaque.22 Both responded significantly after to development of lesions. Unfortunately, due to poor 2 weeks of oral itraconazole therapy with no adverse follow-up and limited resources, testing and diagnosis events noted. Treatment was given for 3 and 5 months were delayed leading to a full evolutionary and marked respectively. progression of lesions over several months. Surgical debridement has been suggested for extensive Treatment of basidiobolomycosis is not always deforming lesions12, but this may cause worsening of successful, and no single drug has proved effective. the lesions and spread of infection. Patient responded To date, no single therapy is recommended for the to itraconazole within 2 weeks with his arm returning treatment of this uncommon infection. Several to normal size with no untoward sequelae in 3 months pharmaceutical agents have been used to successfully time. Based from previous literatures, we extended his treat basidiobolomycosis including potassium iodide, treatment for a total of 6 months. trimethoprim-sulfamethoxazole, , oral azoles.16 However, because of the rarity of these CONCLUSION infections, the dosage, duration, and even the best antifungal drug selection remains unclear.18 Most patients This case demonstrated the gradual progression of the respond very well to oral potassium iodide therapy4,14,19 disease, and that the lack of a history of trauma does not as well as to azoles, particularly itraconazole.15,20-22 exclude its diagnosis. Subcutaneous zygomycosis should The likelihood, however of adverse events from be highly suspected in chronic painless subcutaneous potassium iodide in children e.g suppression of thyroid nodules with swelling, and its early recognition is crucial functions15, makes itraconazole the best option in terms to prevent progression of the disease. In patients whom of response to treatment, low side effects and safety.23 an infectious cause is suspected, appropriate cultures, Oral itraconazole and oral at a dose of special staining, and sometimes repeat biopsies of 400 mg once daily or 7 - 10 mg/kg once daily for two new cutaneous lesions may be helpful in determining to six months have been reported for management of or ruling out the associated disease. Furthermore, basidiobolomycosis24, while others report use of lower performing PAS stain alone to identify the presence dose of 5 mg/kg/day.20-22 In children, itraconazole has or absence of a fungi is insufficient, rather, additional demonstrated an excellent safety profile with frequency staining such as GMS should be done to heighten of adverse effects being reported between 1.9% and detection of fungal elements. 3.5%. The most common adverse events are usually mild and transient gastrointestinal reactions such as nausea,

J Phil Dermatol Soc · November 2020 · ISSN 2094-201X 81 REFERENCES

1. Gugnani HC. A review of zygomycosis due to Basidiobolus ranarum. Eur J 15. Mathew R, Kumaravel S, Kuruvilla S, Varghese RG, Shashikala, Srinivasan pidemiol. 1999; 15: 923-929. S, Mani MZ. Successful treatment of extensive basidiobolomycosis with 2. Sujatha S, Sheeladevi C, Khyriem AB, Parija SC, Thappa DM. Subcutaneous oral itraconazole in a child. Int J Dermatol. 2005; 44 (7): 572–575. zygomycosis caused by Basidiobolus ranarum—a case report. Indian J 16. Verma RK, Shivaprakash MR, Shanker A, Panda NK. Subcutaneous Med Microbiol. 2003; 21(3): 205–206. zygomycosis of the cervicotemporal region: due to Basidiobolus ranaram. 3. Anand M, Deshmukh SD, Pande DP, Naik S, Ghadage DP. Subcutaneous Med Mycol Case Rep. 2012; 1(1): 59–62. zygomycosis due to Basidiobolus ranarum: a case report from 17. Goyal A, Gupta N, Das S, Jain S. Basidiobolomycosis of the nose and face: Maharastra, India. J Trop Med. 2010; 2010: Article ID 950390, 3 pages. a case report and a mini-review of unusual cases of basidiobolomycosis. doi: 10.1155/2010/950390. Mycopathologia. 2010; 170:165–168. 18. Ostrosky-Zeichner L, Smith M, Michael R. McGinnis. Zygomycosis. In: 4. Mondal AK, Saha A, Seth J, Mukherjee S. Subcutaneous zygomycosis: a Anaissie EJ, McGinnis MR, Pfaller MA, eds. Clinical Mycology 2nd edition. report of one case responding excellently to potassium iodide. Indian J London, United Kingdom: Churchill Livingstone; 2009: 305. Dermatol. 2015; 60(5): 500–502. 19. Thotan SP, Kumar V, Gupta A, Mallya A, Rao S. Subcutaneous - 5. Kamalam A, Thambiah AS. Muscle invasion by Basidiobolus haptosporus fungal infection mimicking a soft tissue tumor: a case report and review of following IM injection. Sabouraudia. 1984; 22: 273-277. literature. J Trop Pediatr. 2009; 56(1): 65–66. 6. Department of Health, Republic of the Philippines [Internet]. National 20. Patro P, Das P, Sachdev D, Borkar N, Ganguly S, Hussain N. An instance Tuberculosis Control Program; c2020 [cited on 2020 April 27]. Available of excellent response of subcutaneous zygomycosis to itraconazole from: http://www.ntp.doh.gov.ph/magnitudeTB.php monotherapy. Med Mycol Case Rep. 2019; 24: 13–17. 7. Department of Health, Republic of the Philippines [Internet]. National 21. Sackey A, Ghartey N, Gyasi R. Subcutaneous basidiobolomycosis: a case Leprosy Control Program; [cited on 2020 April 27]. Available from: http:// report. Ghana Med J. 2017; 51(1): 43-46. www.doh.gov.ph/leprosy-control-program 22. Mahamaytakit N, Singalavanija S, Limpongsanurak W. Subcutaneous 8. Leru PM. Eosinophilic disorders: evaluation of current classification and zygomycosis in children: 2 case reports. J Med Assoc Thai. 2014; 97 (6): diagnostic criteria, proposal of a practical diagnostic algorithm. Clin Transl 248-253. Allergy. 2019; 9: 36. 23. Kumaravel S, Bharath K, Rajesh NG, Singh R, Kar R. Delay and misdiagnosis 9. Anaparthy UR, Deepika G. A case of subcutaneous zygomycosis. Indian of basidiobolomycosis in tropical South India: case series and review of Dermatol Online J. 2014; 5 (1): 51-41. the literature. J Paediatr Child Health. 2016; 36 (1): 52-57. 10. El-Shabrawi MHF, Arnaout H, Madkour L, Kamal NM. 24. Brun LVC, Roux JJ, Sopoh GE, Aguiar J, Eddyani M, et al. Subcutaneous Entomophthoromycosis: a challenging emerging disease. Mycoses. 2014; granulomatous inflammation due to Basidiobolomycosis: case reports of 57 (3): 132–137. 3 patients in buruli ulcer endemic areas in Benin. Case Rep Pathol. 2018; 11. Davis SR, Ellis DH, Goldwater P, Dimitriou S, Byard R. First human culture- 1351694. proven australian case of entomophthoromycosis caused by Basidiobolus 25. Gupta AK, Cooper EA, Ginter G. Efficacy and safety of itraconazole use in ranarum. J Mycol Med. 1994; 32 (3): 225–230. children. Dermatol Clin. 2003; 21: 521–535 12. Ribes JA, Vanover-Sams CL, Baker DJ. Zygomycetes in human disease. Clin 26. Tucker RM, Haq Y, Denning DW, Stevens DA. Adverse events associated Microbiol Rev. 2000; 13: 236. with itraconazole in 189 patients on chronic therapy. J Antimicrob 13. Haque A. Special stains use in fungal infections. Connection. 2010; 187- Chemother. 1990; 26: 561–566. 194. 14. Jayanth ST, Gaikwad P, Promila M, Muthusami JC. The sinus that breeds fungus: subcutaneous zygomycosis caused by Basidiobolus ranarum at the injection site. Case Reports in Infectious Diseases. 2013; 2013: Article ID 534192, 2 pages. doi: 10.1155/2013/534192

82 J Phil Dermatol Soc · November 2020 · ISSN 2094-201X