Download Etiologic significance of Aspergillus terreus in primary cutaneous

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Etiologic significance of Aspergillus terreus in primary cutaneous
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mycosis of an agricultural worker
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Pratibha Dave*, Raj Mahendra**, Mahendra Pal***
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*Department of Dermatology, Welfare Hospital and Research Center, Bharauch-392001,
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Bharauch, Gujarat, India
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** Shashwat Skin Clinic, Bharauch-392001,Gujarat, India
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** Department of Microbiology, Immunology and Public Health, College of Veterinary
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Medicine and Agriculture, Addis Ababa University, P.B.No.34,Debre Zeit, Ethiopia
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Corresponding author: Prof. Mahendra Pal, Email Id: [email protected]
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Abstract
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Aim: This pilot study was aimed to elucidate the growing role of non-dermatophytic filamentous
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moulds in the cutaneous lesions of laborers who worked in agriculture field in the villages of
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Bharauch, Gujarat ,India.
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Materials and Methods: Eighteen patients (13 males and 5 females, aged 21 to 46 years) with
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various skin problems presented to the outpatient department (OPD) of Welfare Hospital and
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Research Center, and Shashwat
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dermatophytic filamentous fungi by using standard mycological techniques. All the specimens
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were treated in 10 % potassium hydroxide solution of direct microscopy; and cultural isolation
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was done onto Sabouraud medium with chloramphenicol. The detailed identification of fungal
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isolates was carried out in “ Narayan” stain. The treatment of patient was attempted with oral
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administration of itraconazole.
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Results: Aspergillus terreus was indentified in the cutaneous lesion of 1 of the 18 patients both
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by direct microscopy as well as by cultural isolation. The cinnamon-brown coloured colonies of
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A.terreus grew in pure culture from the biopsied tissue on Sabouraud dextrose agar with
Skin Clinic, Bharauch, India were examined for the non-
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chloramphenicol. In “Narayan” stain, conidial heads of A.terreus were found compact, biseriate,
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and densely columnar. The patient had received trauma on the skin of right lower leg by wooden
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splinter while working in the field. The oral administration of itraconazole for 12 weeks showed
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good clinical response.
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Conclusion: The demonstration of A.terreus in the skin lesion by direct microscopy and cultural
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isolation, and good response to antifungal drugs clearly indicated that our patient was suffering
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with cutaneous mycosis. The patient with chronic cutaneous lesion, history of the skin trauma,
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and occupational exposure to the soil should be investigated for cutaneous mycosis. The
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immediate attention to traumatic injury to the skin is highly imperative to prevent the further
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complications. This seems to be first report of primary cutaneous mycosis due to A.terreus in an
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immunocompetent patient from this part of India.
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Key words: Aspergillus terreus, Cutaneous lesion, Itraconazole, Narayan stain, Soil, Trauma
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INTRODUCTION
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Cutaneous diseases of multiple etiologies are commonly encountered in human and animal
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clinical practice. In recent years, mycotic infections due to opportunistic fungi are gaining
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significance both in developed and developing countries [1]. Among such group of fungi,
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Aspergillus infections are being reported with increasing frequency in humans and animals from
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many regions of the world including India [2 , 3, 4, 5, 1]. There are about 600 species of
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Aspergillus prevalent in our environment, of which 27 species of Aspergillus are implicated in
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various clinical disorders of humans and animals [1, 6]. Aspergillosis is primarily caused by
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A.fumigatus, however, other species such as A. amstelodami, A. candidus, A.chevallieri,
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A.clvatus ,A.deflectus , A.flavus, A.glaucus, A.nidulans, A.niger, A.ochraceous, A.restrictus,
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A.syowii, A.tamari, A.terreus , A.udagawae, A ustus, and A. versicolor are also incriminated in
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the etiology of disease [7,8,9,1,6,10] . Disseminated aspergillosis is associated with a high
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mortality rate of about 90 %. Maximum cases of aspergillosis are encountered in
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immunocompromised patients [1].The prolonged neutropenia predisposes the humans and
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animals to Aspergillus infection. Therefore, invasive aspergillosis has become a leading cause of
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death in neutropenic patients. The cutaneous form of aspergillosis is rarely encountered in
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immnunocompetent hosts. It may be either primary in origin following traumatic implantation of
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fungi with contaminated objects, or occurs due to haematogenous dissemination of the infection
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from the lungs to other sites [9]. Occasional outbreaks of cutaneous aspergillosis are traced to the
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fungal contaminated biomedical devices [9]. Certain occupational groups such as gardeners,
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agricultural workers, brick manufacturers, etc., who remain in direct contact with the soil are at a
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greater risk of acquiring fungal infections [9, 1]. A plethora of drugs such as amphotericin B,
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caspofungin, itraconazole, posaconazole, terbinafine, and voriconazole have been tried with
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variable success in the management of aspergillosis [ 5,9,1].The paucity of information on
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cutaneous aspergillosis from this region of India prompted us investigate the causative role of
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non-dermatophytic filamentous fungi in primary cutaneous mycosis in persons who were
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engaged in agricultural occupation.
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MATERIALS AND METHODS
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In all, 18 patients of both sexes (13 males and 5 females) and different age groups (21 to 46
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years) with various dermatological disorders were presented at the Skin OPD of Welfare
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Hospital and Research Center, and Shashwat Skin Clinic, Bharauch, Gujarat, India for diagnosis
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and treatment. All the patients belonged to nearby villages of Bharauch, India. The suitable
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clinical materials such as swab, scrapings from the border of the lesion, pus, aspirate, and biopsy
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(punch method) etc., were collected aseptically from all the patients for mycological diagnosis.
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Each specimen was subjected for detailed mycological investigation using standard techniques.
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Direct microscopy was done in 10 % solution of potassium hydroxide (KOH) [1], India ink,
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Gram stain; and the cultural isolation was attempted on nutrient agar, Sabouraud dextrose agar,
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Sabouraud dextrose agar with chloramphenicol, and Pal sunflower seed medium [11].The
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examination of the fungal isolates under light microscope was carried out by preparing mount in
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Narayan stain which contained 6.0 ml of dimethyl sulfoxide (DMSO), 0.5 ml of 3 % aqueous
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solution of methylene blue, and 4.0 ml of glycerin [12].
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RESULTS
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There were 13 males and 5 females, and their age varied from 21 to 46 years. All the patients did
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not use any protective wears while working in the agriculture field. The lesions were observed on
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different parts of the body especially on the legs and hands. Clinical findings in these patients
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included erythema, vesicles, cellulitis,papules, plaques, nodules, and ulcers. Aspergillus terreus
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was demonstrated in the cutaneous lesion of 1 of the 18 patients. The patient who yielded
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A.terreus was 27- years- old male; and he had one ulcerated lesion on the lower part of the right
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leg. As narrated by the patient, he had received injury with wooden splinter when planting in the
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field. The direct microscopy of the punch biopsy sample (taken from the ulcer edge) in 10%
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KOH mounts showed the presence of thin ,hyaline, septate, and dichotomously branched hyphae
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morphologically resembling Aspergillus. However, India ink preparation failed to detect any
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capsule of Cryptococcus neoformans. Similarly, Nocardia was absent in the impression smear
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when stained by Gram’s technique. There was no growth of bacteria, Nocardia, and
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Cryptococcus neoformans on nutrient agar, Sabouraud dextrose agar, and Pal’s sunflower seed
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medium, respectively. However, velvety cinnamon-brown coloured colonies were isolated in
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pure and luxuriant from the ulcer biopsy material on Sabouraud dextrose agar with
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chloramphenicol medium after 3 days of incubation at 37 C. As A.terrerus is sensitive to
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cycloheximide, it should not be incorporated in the medium. The growth of fungal isolate in
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Narayan stain revealed small, smooth walled, globose- shaped conidia, thin walled smooth
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condiophores, hemispherical vesicles, and biseriate sterigmata [1].Based on the gross
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cultural,and microscopic morphology, the fungal isolate was identified as A.terreus. The
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laboratory examination of our patient for HIV, diabetes mellitus, and tuberculosis was non-
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committal indicating that he was not immunocompromised, and his immune status was normal.
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Moreover, the negative culture of blood, and urine on mycological media ruled out the
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possibility of dissemination of A.terreus infection. The patient was put on itraconazole (200 mg
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,12 hourly, orally daily for 4 weeks, followed by 100 mg, 12 hourly orally daily for 8 weeks)
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therapy for the management of cutaneous mycosis. In addition, supportive drugs such as
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multivitamin, multimineral, liver tonic, and unienzyme were also prescribed. The oral therapy
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with itraconazole showed good clinical response in our patient.
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DISCUSSION
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Cutaneous mycosis (dermatomycosis, fungal dermatitis) is an infectious, sporadic, global fungal
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disease caused by a large number of non-dermatophytic fungi which are widely prevalent in our
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environment [1]. Most of the fungi responsible for cutaneous mycosis are opportunistic
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pathogens; and are recovered from a wide variety of natural substrates including the soil [1]. In
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majority of cases, transmission of the infection occurs by the introduction of fungi into the skin
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through traumatic injury from saprobic environment [9, 1].Clinical history, and laboratory
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investigations indicated that our patient was immunocompetent who developed primary
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cutaneous aspergillosis due to A.terreus following traumatic injury in the agriculture field. Cases
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of primary cutaneous aspergillosis in immunocompetent patients have been reported by several
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investigators [13, 14, 15, 16]. Our finding is in accordance with Ozer and co-workers [17] who
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isolated A.terreus from the cutaneous lesions of an immunocompetent patient. As A.terreus
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resembles to other filamentous fungi in direct microscopy, hence cultural isolation of the fungus
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is very essential to confirm the specific diagnosis. Our experience had indicated that the skin
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biopsy by punch method is a very good specimen to establish an unequivocal diagnosis of
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cutaneous mycosis due to non-dermatophytic filamentous fungi. Our patient ignored the skin
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injury, and did not seek medical advice due to financial constraints .However, when the lesion
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became very apparent on the lower part of the right leg after about five months, the patient
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visited the hospital for treatment. The duration of lesion in primary cutaneous aspergillosis in an
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immunocompetent patient was recorded 10 years by some workers [15]. As A.terreus is widely
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prevalent in Indian environment, we believe that our patient probably acquired the infection from
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the fungal contaminated soil after receiving traumatic injury on the skin. The role of trauma in
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the development of cutaneous aspergillosis is described by earlier investigators [13 ,17]. Since
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the lesion was chronic in nature, long duration of therapy was recommended. As A.terreus is
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refractory to treatment with amphotericin B, we tried itraconazole in our patient, and the clinical
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response was encouraging. The drug was well tolerated as our patient did not exhibit any side
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effects. The drug itraconazole has been found effective to treat cutaneous aspergillosis in an
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immunocompetent patient [16]. Moreover, it was observed in our clinical practice of over two
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decades that the patients who were given vitamins, minerals, liver and unienzyme preparations
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along with antibacterial antibiotics or antifungal antibiotics showed better results. The clinical
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efficacy of newer drugs such as caspofungin, posaconazole, and voriconazole should be further
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studied in immunocompromised as well as immunocompetent patients. It is, therefore, advised
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that person with a history of traumatic injury to the skin from the environment must immediately
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visit the physician for medical treatment to avert the further complications of disease. Moreover,
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the high risk groups should be provided protective wears; and they should be educated about the
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skin hygiene. It is emphasized that antifungal therapy is warranted in all patients with localized
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lesions in order to prevent the risk of dissemination. As A.terreus is an emerging human and
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animal pathogen, its etiologic role in various clinical disorders should be further studied.
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ACKNOWLEDGEMENTS
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The authors are thankful to the technical assistance rendered by the staff of the Welfare Hospital
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and Research Center, and also Shashwat Skin Clinic, Bharauch, Gujarat, India. Thanks are also
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due to the patients for their cooperation.
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