After a 75-year-old male, who pursued gardening as a hobby, noticed decreased vision in his left eye, he visited an ophthalmologist in September of 2015. He had a history of hypertension, colon cancer and metastatic hepatic tumor. As an ophthalmologic examination revealed macular edema in his left retina resulting from branch retinal vein occlusion (BRVO), he underwent a posterior STTA injection in his left eye to treat the macular edema. After routine disinfection of the lid margins and conjunctival sac with 10% and 0.625% (16-fold dilution) povidone iodine, respectively, the procedure was performed uneventfully. The patient once again visited his ophthalmologist in November due to complaints of ocular pain and hyperemia in his left eye. The symptoms did not improve even though he received oral and topical antibiotic treatment for 3 days. Computed tomography of the orbit was performed and revealed a peribulbar high density lesion of the left eyeball. Due to a suspected peribulbar infection, he was subsequently referred to our hospital in the middle of November.\nAt his first visit, best corrected visual acuity (BCVA) (decimal) was 1.2 in the right eye and 0.15 in the left eye. Intraocular pressures were normal. Although the upper and lower eyelids of his left eye were swollen, we observed no restrictions of his eye movement. Slit lamp examination of the left eye showed ciliary injection, cells in the anterior chamber and a subtenon abscess in the superotemporal quadrant, which corresponded to the site of the posterior STTA (Fig. ). While our fundus examination did not reveal any intravitreal involvement, it did find macular edema and BRVO, which had been reported by his previous doctor. The right eye exhibited no abnormal findings, including both the anterior and posterior segments. The patient had no fever. The orbital magnetic resonance imaging (MRI) revealed the presence of a peribulbar high intensity lesion that corresponded to the subtenon abscess (Fig. ). The MRI did not detect any paranasal sinus or intracranial invasion. The serologic test, which included white blood cells, C-reactive protein and beta-D-glucan, only revealed a slight elevation of C-reactive protein (1.11 mg/dL). To determine the causative pathogen, incision and drainage of the subtenon abscess were performed, with the obtained purulent fluid submitted for bacterial and fungal cultures. While the smears revealed polymorphonuclear leukocytes and a large number of round or oval conidia-like structures, a fungal filamentous appearance was not observed (Fig. ). Based on these results, we suspected fungal infectious scleritis induced by STTA. Therefore, the patient was initially started on 1% voriconazole (VRCZ) and 1.5% levofloxacin (LVFX) eye drops that were each administered six times per day in an outpatient department. Inflammation of the anterior chamber of the left eye worsened after 4 days, with choroidal detachment observed in the temporal region of the fundus. The fungal culture found an early colony of a filamentous fungus. Based on the definitive diagnosis of infectious scleritis by filamentous fungi, we then performed debridement of the infected sclera and Tenon’s capsule. The resected scleral tissue was submitted for further pathological examination. The conjunctival sac of the left eye was irrigated with 0.625% povidone iodine, followed by hospitalization for treatments with 1% natamycin ointment, 1% atropine eye drops and intravenous VRCZ. Ten days after the initial visit, we noted improvement in both his ocular pain and inflammation of the anterior chamber. As a result, we switched the voriconazole from an intravenous to an oral administration. Histopathological examination using Grocott staining revealed infiltration of branching hyphae into the sclera (Fig. ). Due to a novel subtenon abscess of the adjacent upper region of the left eyeball that developed at 18 days after the initial visit, we once again performed scleral debridement. The fungal species, Scedosporium apiospermum, was identified by DNA sequencing of the internal transcribed spacer (ITS) region of ribosomal DNA amplified by polymerase chain reaction (PCR) with ITS1F and ITS1R primers []. The strain was sent to Teikyo University for complete species identification. The susceptibility testing of the antifungal agents was performed using the broth microdilution method based on CLSI M38-A2 []. Results indicated low minimum inhibitory concentrations against micafungin, VRCZ and miconazole (Table ). Based on the antifungal susceptibility test findings, the natamycin ointment was discontinued. Topical and systemic voriconazole administrations were continued for 2 and 1 months, respectively, and then stopped once the ciliary injection subsided. However, 1 month after the discontinuation of topical VRCZ, the subtenon abscess recurred at the same place (superotemporal quadrant). Incision and drainage of the abscess was performed three times. Light microscopic examination of the smears of the subtenon abscess revealed that septate hyphae were still present. As a result, we resumed the 1% VRCZ and 1.5% LVFX eye drops for 2 weeks and then stopped the administration. No recurrence was observed after the final treatment. Due to the prolonged macular edema, however, the final BCVA (decimal) of the left eye was 0.2 as of April 2016 (Fig. ).\nCharacteristics of the colonies found on the potato dextrose agar incubated at 28 °C for 21 days included a grayish white and cottony appearance with an umbonate center on the surface, and a reverse side showing a brownish dark wrinkle appearance in the center and a yellow-white appearance on the margin (Fig. ). Microscopic features observed when using the slide culture technique on potato dextrose agar at 10 days included septate hyphae that were 2 μm in diameter and branching irregularly, along with the production of lateral and terminal conidia, which were round or oval (3 to 5 by 5 to 10 μm) (Fig. ). We also observed a Graphium synanamorph that produced a brush of cylindrical conidia (3 by 12 μm) (Fig. ).\nIn order to confirm the species identification using a molecular approach, the BUT locus of the β tubulin gene was also amplified by PCR using Bt1a/Bt1b primers [, ] and then applied to DNA sequencing [, ]. The obtained DNA sequencing was compared by using the Basic Local Alignment Search Tool. Both the ITS and the BUT exhibited 100% homology with that reported by Zouhair et al. for Scedosporium apiospermum (GenBank accession no. JQ690950 and JQ691056) []. Based on the current and previous results, we were finally able to identify the causative fungal strain as Scedosporium apiospermum sensu stricto.
A 69-year-old Japanese woman had a macular oedema in her right eye, which occurred secondary to BRVO and was unresponsive to intravitreal injection of anti-vascular endothelial growth factor aflibercept. She was subsequently treated by STTA injection at the referral hospital twice. During the treatments, her eye was anaesthetized with 4% topical lidocaine. A sterilized eyelid speculum was used, and the superior-temporal conjunctiva was incised using sterilized scissors to inject 20 mg of triamcinolone acetonide into the posterior subtenon space. After the injection, the conjunctiva was not sutured because this step is usually unnecessary. These procedures were performed in an outpatient treatment room with wearing sterilized gloves. A 1.5% topical levofloxacin ophthalmic solution was administered four times daily for 3 days post-injection. Four months after the second injection, the patient noted a conjunctival mass that became progressively larger with a little pain and redness. She was then referred to Kindai University with a clinical diagnosis of orbital fat herniation.\nDuring her first visit, best-corrected visual acuity (BCVA) was 6/30 in the right eye and 6/6 in the left eye. Intraocular pressure was normal in both eyes. Slit-lamp microscopic examination revealed no inflammation in both the anterior chamber and vitreous. A soft, well-demarcated mass mimicking herniated orbital fat was observed in the super-temporal quadrant, which corresponded to the posterior STTA site (Fig. a). Magnetic resonance imaging (MRI) of the orbits revealed a hypo-intensity lesion on T2-weighted imaging at the super-temporal peribulbar area (Fig. b). Her white cell count and C reactive protein levels were normal.\nTo excise the conjunctival mass, first, we made an incision in the conjunctiva, and found that yellowish pus was discharged. The local administration of antibiotics was performed empirically based on the presumed diagnosis of bacterial infection. Although a strain of dematiaceous fungi was isolated from the pus 1 month later, the antifungal drug susceptibility of the strain was not determined at that time. We did not immediately administer antifungal drugs because there were no significant changes in clinical findings initially. The abscess was debrided again 2 months after the surgery through a superior forniceal incision because the orbital abscess at the same lesion had developed gradually. During the second surgery, the wound was irrigated with 0.1% amphotericin B after three large abscesses were removed. Postoperatively, she was treated with 400 mg/day of oral voriconazole because of Fungiflora Y staining of the pus (Fig. a) and the presence of a large amount of fungi by pathological examination of the abscess by Grocott’s staining (Fig. b).\nWe obtained the pathogenic strain from the abscess during the second surgery and observed it macroscopically and microscopically. The colony on potato dextrose agar medium (PDA; Difco, Becton, Dickinson and Company, Sparks, USA) had a diameter of 10 mm 14 days after inoculation and with incubation at 25 °C. The colony appeared as a moist-form pigmented yeast-like colony (Fig. a). The slide culture using lactophenol cotton blue staining revealed cylindrical conidogenous cells producing ellipsoidal conidia (Fig. b). From the Basic Local Alignment Search Tool search using the sequence of the ribosomal RNA gene internal transcribed spacer domain base, the sequence (the accession number LC566592 in the DDBJ) of this strain showed 100% similarity to those of some strains of E. dermatitidis. The isolate was thus identified as E. dermatitidis on the basis of morphology and phylogeny. The pathogenic strain in the current case was deposited at the Medical Mycology Center, Chiba University in Japan, as IFM 65961.\nMinimum inhibitory concentrations (MICs) of several antifungal drugs and the minimal effective concentration of micafungin against the strain were determined using of the broth dilution method according to M38-A2 of the Clinical and Laboratory Standard Institute (Table ). The inhibitory concentrations of amphotericin B, itraconazole, voriconazole, and miconazole against the pathogenic strain were found to be low. Based on these results, topical and systemic voriconazole were administered for 3 months. The patient’s serum beta-D-glucan levels were normal during the follow-up periods since the first surgery, and no recurrence was observed after the second drainage.