123 Introduction Mycoses in the hyalohyphomycosis group are heterogeneous, defined by the presence of hyaline hyphae in tissues. The number of organisms causing hyalohyphomycosis is increasing and the most clinically important genera are Fusarium spp., Scedosporium spp., Acremonium  spp., Scopulariopsis  spp., Purpureocillium and Paecilomyces spp. (Tortorano et  al. 2014). Fusarium (order Hypocreales) is a large genus with at least 200 species divided into 20 complexes, and many members are commonly found in the environment, where they are isolated from soil, plants and water systems, distributed worldwide and encompassing saprotrophic, biotrophic‑pathogenic or endophytic fungi. Agents of any type of fusariosis are mainly found in three species complexes: F. solani complex, F. oxysporum complex and F. fujikuroi complex. Fusarium  spp. are notorious as pathogen to plants, animals and humans and as a producer of secondary metabolites causing toxicoses (fumonisin mycotoxins, toxic and/or carcinogenic to human and domesticated animals) or invasive disease (Brown and Proctor 2013, Nuñez Otaño et  al. 2014, Wellehan and Divers 2019). Human isolates mostly cause a broad spectrum of opportunistic superficial infections in immunocompetent individuals (i.e. onychomycosis or keratitis), but in recent years they have been increasingly associated with invasive and disseminated infections, predominantly in severely immunocompromised patients, diabetics, cancer patients taking cytotoxic drugs, and burn patients, becoming the leading mycosis affecting immunocompromised patients, and the second most common cause of filamentous fungi infections after aspergillosis, with high morbidity and mortality rates (Montali et  al. 1981, Brown and Proctor 2013, Al‑Hatmi et  al. 2018, Leu et  al. 1995, Alastruey‑Izquierdo et al. 2008). Fusarium  spp. are frequently isolated from marine mammals: Fusarium  solani in captive California sea lions (Zalophus californianus) and grey seals (Halichoerus grypus) (Montali et al. 1981), white‑sided dolphin (Lagenarhynchus acutus), pigmy sperm whale (Kogia breviceps) and harbour deals (Phoca 1Department of Veterinary Medicine, University of Bari ‘Aldo Moro’, Bari, Italy. 2Centro Recupero Tartarughe Marine ‘Luigi Cantoro’, Torre Guaceto, Brindisi, Italy. 3Dipartimento di Medicina Animale, Produzioni e Salute, University of Padova, Padova, Italy. *Corresponding author at: Department of Veterinary Medicine, University of Bari ‘Aldo Moro’, Bari, Italy. E‑mail: antonella.tinelli@uniba.it . Keywords Fusarium solani, Hyalohyphomycosis, Caretta caretta, Multidrug resistance, Zoonosis. Summary Fusarium spp. are pathogens plants, animals and humans, isolated from soil, plants and water systems. They are distributed worldwide and include saprotrophic, biotrophic‑pathogenic or endophytic fungi, or producers of mycotoxins (fumonisins). Human isolates are becoming the leading mycosis affecting immunocompromised patients and frequently involved in mycoses of aquatic mammals and reptiles, included sea turtles or their eggs. Here reported are three severe cases of unusual localizations of Fusarium in loggerhead sea turtle (Caretta caretta) and their diagnostic, therapeutic and clinical output. In the clinical practice, correct genus‑level identification of Fusarium species is critically important to enable correct treatment as in vitro antifungal susceptibility testing is mandatory for each Fusarium‑like isolate. For this reason, susceptibility testing can significantly help the practitioner in choosing the most appropriate therapeutic protocol. Olimpia Lai1, Antonella Tinelli1*, Simona Soloperto2, Giacomo Marzano2, Massimiliano Tosches1, Rosa Leone1, Donatella Gelli3, Chiara Belloli1 and Giuseppe Crescenzo1 Fusarium solani hyalohyphomycosis in loggerhead sea turtles (Caretta caretta): a diagnostic and therapeutical challenge Veterinaria Italiana 2020, 56 (2), 123‑132. doi: 10.12834/VetIt.2035.13422.1 Accepted: 18.05.2020 | Available on line: 08.10.2020 CASE REPORT 124 Veterinaria Italiana 2020, 56 (2), 123‑132. doi: 10.12834/VetIt.2035.13422.1 Fusarium in Caretta caretta Lai et al. with geographic region (Wang et  al. 2011) and different species have different drug susceptibility patterns (Nucci and Anaissie 2007), accurate species assignment is important for epidemiological studies and to guide clinical management. F. solani and F. verticillioides are usually resistant to azoles and exhibit higher amphotericin B minimal inhibitory concentration (MICs) than other Fusarium  spp. (Nucci and Anaissie 2007, Wang et  al. 2011). In contrast, F.  oxysporum may be susceptible to voriconazole and posaconazole. (Nucci and Anaissie 2007, Tanaka et al. 2012). Under this respect, in  vitro antifungal susceptibility testing is mandatory for each Fusarium‑like isolate causing a confirmed systemic infection (Lass‑Flörl et  al. 2010, Summerbell 2002, Johnson 2008, Perkhofer 2010). Itraconazole and other azole drugs may need to be replaced, supplemented with amphotericin B, or discontinued as useless against the pathogen. Empirically, a combination of terbinafine with either posaconazole or voriconazole may be used while identification, speciation, and sensitivity results are pending, but intrinsic resistance interferes with antifungal therapy, clearly challenging treatment owing to the limited therapeutic options, and finally resulting in high mortality rates in patient (Brown and Proctor 2013, Sharma et al. 2017, Al‑Hatmi et al. 2018, Mader 2019). Whenever feasible, infected tissue should be excised surgically (Wellehan and Divers 2019). Case history, clinical signs, pathological findings, and diagnosis Case 1 A juvenile cold stunned loggerhead sea turtle (Caretta caretta) stranded in February on the coast of Adriatic Sea near to San Foca (Lecce, Italy), with average sea temperature of 9  °C. The animal was referred to ‘Torre Guaceto’ Marine Turtle Rescue Centre (Carovigno, Italy), and immediately moved to the Department of Veterinary Medicine of Bari (Italy) for diagnostics and treatment because of severe signs ascribable to cold stunning (hypothermia). At admission the turtle weighted 5 kg, and the straight carapace length resulted 33 cm. The animal was lethargic and severely emaciated. Blood samples were collected in heparinized tubes from cervical sinus for complete haematological and plasma chemistry analyses. Afterward, X‑ray examination was performed in order to check for foreign objects and/or pneumonia because of abnormal buoyancy. Both resulted negative. After determination of glycemia (resulted within the ranges), fluids were intracoelomically administered vitulina) (Frasca et al. 1996) or other marine organisms: American horseshoe crabs Limulus polyphemus (Tuxbury et  al. 2014), wild narrow‑clawed crayfish Astacus leptodactylus (Salighehzadeh et  al. 2019), black gill disease in cultured Kuruma prawn Penaeus japonicas (Bian and Egusa 1981) and giant tiger prawn P. monodon (Khoa et  al. 2004), captive lined seahorses Hippocampus erectus (Salter et  al. 2012). Fusarial infections in freshwater and marine fish include deep mycoses, ocular and skin lesions, and fatal ulceration and necrohemorrhagic dermatitis (Yanong 2003, Noga 2010). Fusarium  spp. has been identified as causative agent of cutaneous hyalohyphomycosis in captive loggerhead sea turtles, where the lesions were described as superficial, white and scaly or ulcerative (Austwick et al. 1981, Cabañes et al. 1997), and in a stranded Kemp’s ridley sea turtle with pulmonary hyalohyphomycosis (Orós et  al. 2004), where fungal pneumonia was associated with cold stunning (Knotek and Divers 2019). Fusarium  solani has also been found in sea turtle eggs, associated with mass mortalities (up to 83.3%) in natural and relocated nests of the sea turtle Caretta caretta (Sarmiento‑Ramìrez et al. 2014a). Colonization of eggs with Fusarium is considered among the main causes of globally declining turtle populations (Chelonia mydas, Caretta caretta, Eretmochelys imbricata, Lepidochelys olivacea, Dermochelys coriacea, and Natator depressus) (Sarmiento‑Ramìrez et al. 2014b). Treatment of fusariomycosis is very challenging. Fusarium  spp. shows a remarkably high degree of intrinsic multidrug resistance to a wide spectrum of commonly used antifungals azoles, echinocandins and polyenes. The triazole group of antifungals works by inhibiting the fungal lanosterol 14α‑demethylase in the ergosterol biosynthesis pathway. The polyenes include amphotericin B, which binds to ergosterol thereby forming pores and damaging the cell membrane. Finally, the echinocandins, namely caspofungin, micafungin and anidulafungin, affect cell wall synthesis by inhibiting 1,3‑β‑D‑glucan synthase. Fusarium species share their high degree of intrinsic resistance to most currently used antifungal agents with Acremonium and Trichoderma, which belong to the same order Hypocreales, and with Microascus, Scopulariopsis, Scedosporium and Lomentospora in the adjacent order Microascales. Intrinsic multiresistance of these fungi is unique among the Ascomycota and suggests that this property was acquired in a common ancestor of the two orders (Al‑Hatmi et al. 2018). For practical purposes in the clinical practice, correct genus‑level identification of Fusarium  species is a critically important action to enable correct treatment and infection control. Since the distribution of Fusarium  species varies 125Veterinaria Italiana 2020, 56 (2), 123‑132. doi: 10.12834/VetIt.2035.13422.1 Lai et al. Fusarium in Caretta caretta incubated at 25  °C in air for 14 days. Whitish gray cottony colonies suggestive of Fusarium  spp. were isolated from all samples. Successive subcultures performed on potato dextrose agar in the dark showed sickle‑shaped multiseptated macroconidia, and one‑ to two‑celled microconidia formed from unbranched phialides, conidiophores, and chlamydospores typical of Fusarium  solani (De  Hoog 1995) (Figure 2). This evidence, its reported in  vitro and in  vivo resistance to most of the available antifungal drugs, together with the lack of clinical outcome of the current therapy, suggested to perform susceptibility test on the isolate. Voriconazole, posaconazole, itraconazole, ketoconazole, fluconazole and amphotericin B were tested. Isolates of F. solani resulted resistant to all the antimycotic drugs tested, so itraconazole was discontinued, and only topical iodopovidone ointment was administered. Tissue samples submitted to the Unit of Pathology for histology were fixed in 10% neutral buffered formalin for routinely process, embedded in paraffin, sectioned at 5 microns, stained with hematoxylin and eosin (HE) and examined by light microscopy. Histological analysis revealed necrosis and the presence of inflammatory cells (heterophils, monocytes, and lymphocytes) at the level of the basement membrane and in the dermis, together (1 part dextrose 5%, 1 part saline, 1 part electrolytic rehydrating solution, 1 part sterile water) at 1% b.w., then the body temperature was increased 3 °C/day until it reached 24  °C. Antimicrobial and antifungal therapy was initiated when the turtle’s temperature reached 19 °C to 20 °C (Norton and Walsh 2012). The turtle presented several head, shell and plastron traumatic injuries, probably resulting from repeated dashes against rocks. The shell showed several disseminated superficial and ulcerous lesions, involving keratinized scales and the underlying bone plates, and one large lesion in the column region that continued on the right and left side of carapace, with necrosis of column bones, carapacial bone plates and ribs too (Figure 1a). Also, the head and plastron showed deep lesions with loss of tissue and exposition of bone superficies. Lesions were sampled using a sterile lancet collecting material from the advancing edges of the lesions, after disinfection of the surface with iodopovidone and local anaesthesia induced by injection of a solution of 2% lidocaine hydrochloride; the bioptic samples were used for microbial and mycological culturing, and for histologic examination. Standard prophylactic antimicrobial and antimycotic treatment was started (Mader 2019, Manire et  al. 2003, Paré and Jacobson 2007, McArthur et  al. 2004), with marbofloxacin (2 mg/kg, IM q 24 h; Lai et  al. 2009) and itraconazole (5 mg/kg, PO, q 24  h administered with assisted feeding; Mader 2019). Topical iodopovidone ointment was applied on shell and plastron lesions. The samples for bacteriological testing were cultured on tryptic soy blood agar and Mac Conkey agar, incubated for 24 h at 37 °C. On the basis of biochemical reaction and morphological characteristics, two different isolates were noticed, then identified with the biochemical system API 20 NE. Vibrio fluvialis and Aeromonas hydrofila were isolated from lesions and tested for susceptibility to several antibiotic drugs (ampicillin, ceftazidime, cephalothin, cefuroxime, doxycycline, norfloxacin, ciprofloxacin, enrofloxacin, marbofloxacin). The isolates resulted susceptible to ceftazidime, norfloxacin, ciprofloxacin, enrofloxacin, marbofloxacin and resistant to ampicillin, cephalothin, cefuroxime, doxycycline. On the basis of that result, marbofloxacin was continued for 60  days, and suspended after bacteriological cultures resulted negative. The samples for mycological examination were stained with calcofluor white and examined with fluorescent microscope. Few single septate nonbranching hyphae bright greenish stained were noticed, so the presence of a fungal infections was supposed. Thus, samples were cultured onto Sabouraud agar chloramphenicol 0.05% and Figure 1. a. Carapacial lesions of the loggerhead sea turtle at admission; b. at release (case 1). 126 Veterinaria Italiana 2020, 56 (2), 123‑132. doi: 10.12834/VetIt.2035.13422.1 Fusarium in Caretta caretta Lai et al. animal was lethargic and severely emaciated. The standard procedure for the treatment of cold stunning was started (see above). Samples of the lesions were obtained for microbial and mycological culturing, and histologic examination (see above), together with blood samples for haematological and biochemical exams (Tables I and II). Standard prophylactic antimicrobial and antimycotic treatment was started (see above). Once recovered from hypothermia, the young turtles started feeding voluntarily, and never stopped. Prevotella rettseri, Serratia marcescens, Pseudomonas aeruginosa, Proteus spp. were isolated from lesions and tested for susceptibility (see above); all of them resulted susceptible to fluoroquinolones. On the basis of that result, marbofloxacin was continued for 50 days, then suspended after bacteriological cultures resulted negative. In about a month the lesion extended as area and depth until it involved the underlying bone planes, with multifocal necrosis and severe haemorrhages, with presence of moderately basophilic hyphae; consequently an elective stain with stain periodic acid‑Schiff (PAS) was decided. PAS staining revealed hyphae morphologically similar in all samples, with thin, generally parallel walls and variably spaced septa, frequent acute and right angle branching septate hyaline hyphae (Figure 3), with optional sporulation, and occasional invasion of blood vessels During the period of hospitalization, the conditions of the turtle worsened. After two months of autonomous feeding with an average daily assumption of 200 g anchovies, the turtle became progressively anorectic due to the progressive avulsion of upper and lower ramphoteca, so a permanent oesophagostomy tube was applied under general anaesthesia (Figure 4) (Di Bello et  al. 2011). Prophylactic marbofloxacin (2 mg/kg PO, q 24 h, Marìn et al. 2009) was administered for as long as the tube was maintained, together with the food (2‑3% of b.w. of homogenized fish supplemented with vitamins). The permanent oesophagostomy tube was well tolerated by the turtle, which began voluntary feeding in one month, after the complete avulsion of ramphoteca, which leaved the maxillary and mandibular bones exposed and partially eroded on the right side (Figure 5a). The tube was left in place for further 7 days, then it was removed after a light sedation with medetomidine (0.05 mg/ kg, IM) reversed with atipamezole (0.25 mg/kg, IM) in order to apply stitches to close oesophagostomy. The recovery was uneventful. Fungal testing, performed every month, resulted negative after 4 months since the admission, then all the lesions progressively healed, including the beak, which started to show signs of new growth of the corneal part. Complete beak repair took further 4 months (Figure 5b), and the carapace lesions repaired with scar tissue to cover the bone plates (Figure 1b). At that time, the turtle was back to ‘Torre Guaceto’ Marine Turtle Rescue Centre for summer release. Case 2 A juvenile cold stunned loggerhead sea turtle (Caretta caretta) stranded in August on the coast of Adriatic Sea near to Jesolo (Venice, Italy), with an unusually low average sea temperature of 15 °C. The animal was referred to ‘Torre Guaceto’ Marine Turtle Rescue Centre, and immediately after moved to the Department of Veterinary Medicine of Bari for diagnostics and treatment because of extensive shell lesion (probably boat impact) limited to corneal scales. At admission the turtle weighted 1.2 kg, and the straight carapace length resulted 25 cm. The Figure 2. Culture on potato dextrose agar showing sickle-shaped multiseptated macroconidia and one/two celled microconidia formed from unbranched phialides, conidiophore and chlamydospores. Figure 3. Micrograph of necrotic tissue with Fusarium solani hyphae invasion (arrows). Periodic Acid Schiff stain, 40X (case 1). 127Veterinaria Italiana 2020, 56 (2), 123‑132. doi: 10.12834/VetIt.2035.13422.1 Lai et al. Fusarium in Caretta caretta which remained uncovered after the corneal scales detached (Figure 6). Mycological culturing isolated Fusarium  solani from all samples. Also, in this case susceptibility test on the isolate was performed, and F. solani resulted resistant to all the antimycotic drugs tested, so itraconazole was discontinued, and only topical iodopovidone ointment was administered. Histological analysis revealed necrosis and a general situation similar to case 1, as in both subjects the colonization interested corneal structures. Fungal testing, performed every month, resulted negative after 5 months since the admission, and the lesion gradually repaired with scar tissue. At that time, the turtle was back to ‘Torre Guaceto’ Marine Turtle Rescue Centre for summer release. Case 3 A juvenile cold stunned loggerhead sea turtle (Caretta caretta) stranded in March on the coast of Adriatic Sea near to Lecce (Italy), with average sea Figure 4. Permanent oesophagostomy tube in place (case 1). Figure 5. a. Eroded maxillary and mandibulary bones after complete avulsion of upper and lower ramphoteca; b. complete repair of beak 4 months after ramphoteca avulsion (case 1). Table I. Haematological results. Turtle 1 Turtle 2 Turtle 3 Reference values* Median Min Max HCT (%):17 18.00 24.00 23.00 28 17 45 RBC RBC (x106/µL):17 2 2.66 2.54 1.87 0.3 6 WBC WBC (x10³/µL):17 3.20 4.30 5.80 5.9 2 18.9 Heterophils (%):18 64 73 79 75.8 51.61 88.6 Lymphocytes (%):18 30 22 16 18.41 4.4 30.92 Monocytes (%):18 0 1 1 1.2 0 5.3 Eosinophils (%):18 6 4 4 4.5 0 29.4 Basophils (%):18 0 0 0 0 0 0 PLT Estimated adequate adequate adequate Table II. Plasma chemistry results. Turtle 1 Turtle 2 Turtle 3 Reference values* Median Min Max AST (IU/L):17 460 653 389 194 < 10 844 ALT (IU/L):17 14 35 18 24 < 10 258 ALP (IU/L):17 45 71 64 67 51 562 CPK(IU/L):20 188 538 874 534 3 1,899 Total bilirubin (mg/dL):17 0.21 0.32 0.01 0.2 0.2 0.5 Glucose (mg/dL):17 79 98 77 129 19.8 291.9 Total cholesterol (mg/dL):17 63 73 52 139 50.2 397.7 Uric acid (mg/dL):20 0.8 0.9 1.9 2.4 0.7 4.2 Creatinin (mg/dL):17 0.32 0.45 0.22 0.4 0.3 0.8 Total protein (g/dL):17 2.60 4.3 1.9 2.4 2 11 Albumin (g/dL):17 1.2 1.3 0.8 1.1 1 1.4 Calcium (mg/dL):17 6.8 7.2 8.1 8 2.8 12.4 Phosforum (mg/dL):17 4.6 4.5 5.1 6.4 4.1 7.9 Ionized Na (mEq/L):20 146 151 134 156 135 175 Ionized K (mEq/L):20 3.7 5.8 3.2 5.1 3.3 13.9 Ionized Cl (mEq/L):20 112 125 109 130 107 158 128 Veterinaria Italiana 2020, 56 (2), 123‑132. doi: 10.12834/VetIt.2035.13422.1 Fusarium in Caretta caretta Lai et al. samples were sent to the Pathology Unit of Department of Veterinary Medicine of Bari for histologic examination. Aeromonas hydrofila was isolated from lesions and tested for susceptibility (see above), resulted susceptible to fluoroquinolones. On the basis of that result, marbofloxacin PO was continued for 40 days, and then suspended after bacteriological cultures resulted negative. Mycological culturing isolated Fusarium solani from all samples. In this case too susceptibility test on the isolate was performed, and F. solani resulted resistant to all the antimycotic drugs tested, so itraconazole was discontinued, and only topical iodopovidone ointment was administered. In case 3 the lesions were very advanced, with extensive destruction of soft and bone tissues, replaced by repair tissue colonized in depth by the fungal hyphae and with extensive microvasculitis caused by the invasion of the microcirculation. The extensive injuries did not recover, so the proposal of live food (crustaceans and fish) was attempted before considering euthanasia. The subject responded positively, by actively preying on living preys, so the tube was removed, and the turtle was back to ‘Torre Guaceto’ Marine Turtle Rescue Centre for summer release. Discussion Dermatomycoses occur regularly in reptiles and are largely underdiagnosed, as lesions are indistinguishable from those caused by bacterial infections at a gross examination, so they are often misdiagnosed as such. Mixed fungal and bacterial infections are also common, and it may be difficult to establish which of the two is the primary offender, so both microbiological and mycological diagnostics have always to be recommended (Paré and Jacobson 2007). temperature of 11  °C. The animal was referred to ‘Torre Guaceto’ Marine Turtle Rescue Centre, and immediately moved to the Department of Veterinary Medicine of Bari for diagnostics and treatment because of severe signs ascribable to cold stunning. At admission the turtle weighted 8.7 kg, and the straight carapace length resulted 45 cm. The animal was lethargic and emaciated. The standard procedure for the treatment of cold stunning and prophylactic antimicrobial and antimycotic treatment were started (see above). The severe emaciation was probably due to the extensive injury to the rostrum and the nasal cavities that the animal presented to admission, with complete absence of soft and cartilaginous tissues of the nose and of the rearward conchae (Figure 7). In addition to the usual hematologic, bacteriological and mycological procedures, X‑ray and computed tomographic examination of the head were performed. X‑ray exam revealed extensive loss of bone tissues in the nasal cavity with absence of the nasal conchae. For the CT exam, images were acquired with the turtle placed in ventral recumbency under general anaesthesia (propofol 2 mg/kg IV; Mader 2019). Contiguous transverse slices were obtained from the basisphenoid to the external nares, showing severe osteolysis of the prefrontal bone, destruction of the cartilage of nasal septum, erosive changes of the dorsal and ventral conchae, and soft tissue opacification of the right dorsal nasal concha sinus and left ventral concha sinus. The turtle rejected various proposed foods (anchovies, squid, cuttlefish, mussels) for several days, probably because of complete loss of the olfactory capacity, so the esophagogastric tube was applied. Prophylactic marbofloxacin (2 mg/kg PO, q 24 h; Marìn et al. 2009) was administrated. Samples of the lesions were obtained for microbial and mycological culturing. Some of the bioptic Figure 7. Extensive injury to the rostrum and the nasal cavities (case 3). Figure 6. Shell damages after corneal scales detachment (case 2). 129Veterinaria Italiana 2020, 56 (2), 123‑132. doi: 10.12834/VetIt.2035.13422.1 Lai et al. Fusarium in Caretta caretta In the reported cases, no particular immunosuppressed status has been noticed in hematologic exams (Tables 1 and 2). In fact, all the parameters were within the normal ranges, if compared with bibliographic data on juvenile loggerhead sea turtles (Casal et al. 2007, Casal et al. 2009, Gelli et  al. 2009) and comparable with data reported for stranded loggerheads (Deem et  al. 2009). No particularly compromised scenery was depicted, so the severity of the Fusarium  solani infection has to be ascribed to other potential factors that may facilitate fungal invasion: concomitant viral or bacterial infections, bacteria‑infected traumatic lesions, large areas of devitalized skin, nutritional nitrogen imbalance and stressors associated with stranding (Frasca et al. 1996, Lass‑Flörl 2010). Unlike most fungi species, in which conidiation is stimulated by emergence of the fungus through the water‑air interface, the conidiation of Fusarium spp. typically occurs both in submersion and upon exposure to air. This explains the rapid seeding of these infections to numerous capillary beds, especially in the skin, a process that gives rise to the widespread ecthyma gangrenosum‑like lesions that characterize disseminated fusarial infections (Summerbell 2002). Angioinvasion, with vascular thrombosis and tissue infarction, explains the evolution of lesions to necrotic and bone‑involving levels. Therefore, Fusarium  spp. infection has not to be undervalued as it can generate extensive and ravaging lesions (Salter et al. 2012). Due to the critical role of immune response in the outcome of fusariosis, the optimal treatment strategy for patients with severe Fusarium  spp. infection remains unclear. Reversal of immunosuppression is recommended whenever possible. Early therapy of localized lesions (including surgical debridement) is important to prevent progression to a more aggressive or disseminated infection (Tortorano et al. 2014). The definitive diagnosis requires isolation of Fusarium spp. from infected sites (culture, histology, molecular probes). The repeatedly positive cultures of biopsies from the lesions for Fusarium  solani, along with evidence of tissue invasion with mycelial elements having the configuration of Fusarium spp., indicate that this fungus was acting as a pathogen in the three turtles. In invasive infection Fusarium  solani can also be isolated from blood cultures in up to 40‑60% of human cases (Tortorano et al. 2014), but in the present cases blood was not used to attempt isolation. Culture identification is important because of the histopathological similarities between Fusarium spp. and other hyalohyphomycetes. Although the genus Fusarium  can be identified by culture by the Mycoses seem particularly prevalent in sea turtles (Mader 2019, Manire et al. 2003, Paré and Jacobson 2007, Cabañes et  al. 1997, Duguy et  al. 1998), most commonly related to immunosuppressed status, traumatic lesions, captivity, and cold‑stunning. In particular, some authors (Cafarchia et  al. 2019) have postulated that from an epidemiological point of view the origin of this opportunistic infection has to be considered related to the presence of F. solani in rescue center tanks and to immunosuppression due to the traumatic lesions suffered, surgical treatment applied, and other stressful conditions associated with transportation or rehabilitation of these marine turtles. The finding that animals admitted to the center for more than 20 days were more frequently colonized also suggested an association between Fusarium‑like organisms and the skin lesions that occurred, given the presence of F.  solani in the tank at the rescue centers, a recognized risk factor of infection in receptive adult turtles. This finding suggested that the environmental conditions and management at the rescue center might favor Fusarium  spp. growth and might be the source of colonization. This latter hypothesis was supported by the fact that sand from the filter of the two cited centers was positive for Fusarium spp. On the contrary, in the present cases the lesions were already present and invaded by the Fusarium  spp. before admission to the center. In particular, analysis of the sands of the center's filtering plant (equipped with a skimmer and UV passage for the sterilization of the filtered water before returning to the tanks) did not reveal any contaminating pathogenic organisms, so the hypothesis posed by the cited authors cannot be accepted and should not be generalized. Otherwise, sudden drops in seawater temperature often result in hypothermia in juveniles or sub‑adult sea turtles. These cold‑stunned turtles typically become weak, float, and strand, with different possible types of wounds and lesions, and their rehabilitation can become long and difficult because of multiple metabolic disorders and opportunistic infections due to immunosuppression. Severe mycotic infections can be common sequelae in these animals, and antifungal drugs are part of the standard pharmacological treatment of this syndrome (Mader 2019, Manire et al. 2003, Paré and Jacobson 2007). Fungi as Fusarium spp. in particular are often implicated, but other species of scarce or little‑known pathogenicity, such as Colletotrichum acutatum, have caused disseminated mycoses in cold‑stunned turtles (Manire et  al. 2003), indicating how severely immunocompromised these animals may become. Nonetheless, fungal infections have also been reported in wild stranded sea turtles in Florida unrelated to cold stunning events (Paré and Jacobson 2007). 130 Veterinaria Italiana 2020, 56 (2), 123‑132. doi: 10.12834/VetIt.2035.13422.1 Fusarium in Caretta caretta Lai et al. Finally, the receptivity of man to fusariosis must not be underestimated, as Fusarium  spp. can act as zoonotic agent for the operators or the visitors of sea turtle rescue centres. Opportunistic Fusarium  species cause a broad spectrum of infections predominantly in immunocompromised individuals via direct inoculation and airborne uptake as the most common routes of infections. The clinical signs of fusariosis depend largely on the immune status of the host and the portal of entry, which include paranasal sinuses, lungs and skin, with neutropenia as one of the most important risk factors for acquiring disseminated disease (Tortorano et al. 2014); therefore the operators of the centres must be informed of the risk and equipped accordingly for the management of the infected subjects, while the visitors must not be admitted to the tanks of subjects with active fusariosis. production of hyaline, crescent or banana‑shaped multicellular macroconidia, species identification is difficult and may require molecular methods, although they should be used only to supplement conventional laboratory tests (Tortorano et al. 2014). Since the distribution of Fusarium  species varies with geographic region 51 and different species have different drug susceptibility patterns (Nucci and Anaissie 2007) accurate species assignment is important for epidemiological studies and to guide clinical management. Even though there is no correlation between F.  solani species complex and antifungal susceptibility, F. solani and F. verticillioides are usually resistant to azoles and exhibit higher amphotericin B minimal inhibitory concentration (MICs) than other Fusarium spp. (Nucci and Anaissie 2007, Wang et al. 2011). In contrast, F. oxysporum may be susceptible to voriconazole and posaconazole (Nucci and Anaissie 2007, Tanaka et al. 2012). 131Veterinaria Italiana 2020, 56 (2), 123‑132. doi: 10.12834/VetIt.2035.13422.1 Lai et al. Fusarium in Caretta caretta Al‑Hatmi A.M.S., Bonifaz A., Ranque S., Sybren de Hoog G., Verweij P.E. & Meis J.F. 2018. Current antifungal treatment of fusariosis. 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