A Rare Human Pathogenic Fungi: Cladosporium

A Rare Human Pathogenic Fungi: Cladosporium

Cedric NDINGA MUNIANIA

Western Illinois University

My name is Cedric NDINGA MUNIANIA a graduate student in the Biological Science Department at Western Illinois.  This is focus on in fungal infections in general and in particular infections caused by Cladosporium sp. I hope this would raise awareness about fungal diseases and will increase your interest in the subject.

                                                                Cladosporium spp.

Figure 1: Cladosporium conidiophores with chain of blastoconidiahttp://www.microbiologybook.org/mycology/mycology-5.htm

 Taxonomy

Domain: Eukarya

Kingdom: Fungi

Phylum: Ascomycota

Class: Dothideomycetes

Order: Capnodiales

Family: Cladosporiaceae

Genus: Cladosporium

Species: ~60

General Description

Cladosporium is a genus that belongs to the phylum Ascomycota and includes about 60 species (12). Generally found in the environment as a saprobe (4), this genus is considered as common mold and an airborne fungus along with Alternaria and Aspergilus (11). Cladosporium is usually isolated as an environmental contaminant in the laboratory and food industry (11) and rigorous testing are required before it can be determined as a causative agent of fungal infections. The Cladosporium genus is a very diverse group that include species specific to certain geographic areas (12) and can be isolated as plants pathogen or causing opportunistic human infections (14). Despite the fact that incidence of fungal infections are low, some species have been associtated with asthma, rhinitis, chromablastomycosis, phaehyphomycosis, intrabronchial lesions and cladosporiosis (6,12,14).  

The most common species of Cladosporium are:

_Cladosporium bantiana  associated with central nervous system cladosporiosis (6)

_ Cladosporium cladosporioides : can cause pulmonary and cutaneous opportunistic infections in  human and animals (13).

_Cladosporium herbarum causes mold allergies and can also cause skin infection in human and animals (1, 13).

_Cladosporium oxysporum is associated with keratitis and other skin infection in human (13)

_ Cladosporium sphaerospermum is associated with cases of corneal ulcer, bronchial infections and onychomycosis (13)

 _Cladosporium trichoid both are associated with case of CNS phaehyphomycosis (4).

Macroscopic features

Several species of this genus are moderate fast growers, 4-6 days (10) and growth better at 25°C with a preference for SDA and PDA (11, 14). However some species are thermotolerant with an optimum temperature at 42°C (9). Most Cladosporium spp. are usually classified within the group dematicae due to the melanization of their cell wall (11). Many species of this genus form pigmented molds ranging from dull olive green to black in coloration (1) (Figure 2).

Figure 2: Culture of Cladosporium herbarum in Petri dish on PDA medium  http://www.livne.co.il/thesis/fungi_pictures/misc/index.html

Microscopic features

Cladosporium spp. are characterized by having branched pigmented hyphae from which chains of blastococonidia will rise (11). Most conidia in the chains measuring 3µm X 6.4 µm (4) will have scars and the tips of the terminal conidiophore usually has ramoconidia or branched conidia  (12). Figure 3 denote the bud-like chain of blastoconidia formed by Cladosporium.

                                       Figure 3: Holoblastic conidiogenesis associated with Cladosporium                                         http://www.environix.com/mold-iaq-library/mold/cladosporium/.

           

Seasonal variability

            Though Cladosporium spp.  release spores throughout the entire year, the amount of spores released follow a seasonal pattern with the peak being during the summer months (5). Furthermore Cladosporium releases spores during both wet and dry conditions, but a higher number of spores is released during the wet and rainy season because spores are easily dispersed by rain splash (7).

Geographic Distribution:

This fungus is ubiquitous in the environment and distributed worldwide (11). However some species are limited or dominant to specific environments. That is the case for C. herbarum that is mainly found in temperate regions (12) and C. carrionii mainly found in East Asia (15).

Habitat

Being a saprobes in nature, Cladosporium spp  are usually isolated from soil, decaying organic matter and  dead plants (11). In addition to soil, these massive producers of spore can be isolated from air and water sample (12). Some species have been isolated from water pipe line, glass fiber and as a food and textile contaminant (11).

Main infections and related fungi:

Lower respiratory tract diseases:

               Indeed most Cladosporium spp. produce a lot of spores as well as allergens (12) leading to allergic reactions with extended exposure. For some people inhaling the allergens will lead to more complicated conditions such as asthma and rhinitis (3). C. cladosporioides has been reported as one of the main cause of air-borne allergies due to fungus (3). However allergens are effectively produced as the spore are being produced are but they are very low when the fungi is in the to form (3).

Mycotoxins poisoning

Along with Aspergillus, Cladosporium is one of the most carcinogenic fungi (11), thus producing high amounts of toxins. C. cladosporioides produces two strong mycotoxins cladosporin and emodin as well as some other less toxic compounds (11). Because Cladosporium can coexist with plants as endophytes (12), it is more likely that consumption of fruits or leaves of an infected plant will result in mycotoxin poisoning. Furthermore mycotoxin are resistant to heat which means that consumption of thoroughly cooked infected food product could still cause health issue in human (14)

Phaehyphomycosis:

It is a general fungal infection caused by pigmented mold also called dematiaceous fungi, which includes Cladosporium spp, Cladophora spp, Exophiala spp, Alternaria spp and Ramichloridium spp (9). This disease starts as a pulmonary infection and can become disseminated and infect the skin, and could extend to the central nervous system (9). This particular infection is characterized by the growth of pigmented hyphae on the infected tissues (6). The Cladosporium spp associated with this disease includes C. cladosporioides, Cladosporium oxysporum and Cladosporium sphaerospermum. (8,10,11,13) (Figure 4).

Figure 4: Cutaneous Phaehyphomycosis. http://www.hindawi.com/journals/cridm/2011/385803/fig3/

Chromoblastomycosis

This is a fungal infection that mainly infects the skin and subcutaneous tissues and is characterized by chronic ulcerative granuloma especially on the feet and the legs (9). It is mainly caused by Cladophora spp, and Exophiala spp, (9) but recent cases have reported Cladosporium as the causative agent of this disease including species such as C. cladosporioides , Cladosporium bantiana  and Cladosporium trichoides (6) (Figure 5).

Figure 5: chromoblastomycosis  caused by cladosporium spp http://www.biomedsearch.com/nih/Chromoblastomycosis-due-to-Cladosporium-carrionii/21814409.html

Central nervous system cladosporiosis

            This is a rare fungal infection that mainly infects the central nervous system (CNS) and is caused by Cladosporium, but more precisely Cladosporium bantiana (6). The defining symptoms for this infection includes fever and intracranial abscesses, and if not treated , it can lead to diffuse meningoencephalitis with extensive necrosis (6).    

Site of entry and Population at risk

            Cladosporium is an important air-borne fungus producing a lot of spores (11). Therefore depending of the type of infection, the fungus enters the body mainly due to inhalation of spores or allergens (14)

            Just like for other fungal infection, the population at risk for Cladosporium infections includes immuno-compromised patients  and steroid users (6), patients in intensive care unit such as patients on peritoneal dialysis (11) and on immuno-deficient patients (2). For many reported cases, the infected patient were farmers or were involves in a lot of outdoors activities (6). However Cladosporium is not always pathogenic and can infect healthy patient with no history of previous fungal  infection (8).

Diagnosis and identification

            Diagnosis of Cladosporium infection can require a combination of technique and collection of information about patient recent activities and occupations. In most cases doctor use a combination of direct microscopy and growth of the fungi on a plate (6). To begin, tissues from the lesions are harvested and stained using periodic acid-Schiff (10). Observation of the specimen under the microscope will reveal branched hyphae darkly pigmented with chains of blastococonidia that are clustered (10) (Figure 3). Also, samples from the lesion site are plated on SDA or PDA at both 25°C and 37°C for at least two weeks (8). Positive identification Cladosporium should result in a formation of dark gray to black mold on the plate (8). In some extreme cases DNA from the sample could be isolated and PCR and sent out for sequencing of the ITS region for exact identification (10)  

Treatment

            The treatment for Cladosporium infection is very hard and depends on the severity of the infections. Most antifungal drugs available on the market have limited effect against many species of Cladosporium (6). This resistance of the fungus though unclear, seems to be related to the presence of melanin in their cell wall (11). Indeed melanin has been considered to increase the virulence of pathogens by reducing their susceptibility to host immune system, inhibiting some of the host response mechanisms as well as the action of some antifungal drugs (16). Furthermore, some Cladosporium species can have innate resistance to Amphotericin B (Amp B) (6) and thus requires combination of different drugs to heal the patient.

            Nevertheless less severe respiratory infection or allergies would just light symptoms or flu like symptoms and will disappear (11). Mild infections can be treated with azoles which sometimes need to be combined with flucytosine (4). For example cutaneous infection due to  C. cladosporioides can be treated with itraconazole and ketoconazole (10).  However the best approach for systemic infections or deeper organ infections is surgical removal of infected tissue combined with cure of Amp B associated with azoles (13).

Clinical cases

Case study I:  Intrabronchial lesions

            A healthy 58-year-old woman with no previous record of asthma or respiratory issues arrived at the hospital complaining about persistent dry cough  that had lasted for a month prior coming to the hospital. Blood tests were performed and came back normal. Serum IgE and the multiple antigen simultaneous test (MAST) for Aspergillus were also performed and they all came back negative. In addition to these tests,  routine bacteriological, fungal, and mycobacterial cultures of sputum were performed and the results were negative. That led the doctors to do chest X-ray  (Figure 6a) and CT scanning that revealed a nodular opacity in the right right basal bronchus (Figure 6b). Based on this results doctor decided to perform a transbronchial biopsy that revealed presence of hyphae-like structure on the necrotic tissues. Samples from the lesion were then collected and cultured on Sabouraud glucose agar for seven day. After incubation, three grayish green cotton-like colonies were growing on the plates. Examination of these colonies under the microscope using lactophenol cotton blue stain revealed brown budding conidia and globular conidiophores, the defining characteristics of C. sphaerosporum.  Thus the patient was diafnosed with Intrabronchial lesion due to Cladosporium sphaerospermum and was treated with intrabronchial infusion of amphotericin B (60 mg)  followed by oral administration of prednisolone (10 mg/day) for two weeks before being switched to oral itraconazole (100 mg/day). The patient condition really improved after the treatment.

Figure 6: (a) Chest radiograph showing a nodular opacity of the right basal bronchus. (b) CT scan of the thorax showing an intrabronchially branching, band-like opacity in the right basal bronchus.  (Yano et al. 2003)

Case study II

            A 66-year-old woman that was a gardener and had history of trauma was admitted to the hospital due to multiple nodular erythematous lesions on her right leg (Figure 7a). These lesions had a reddish brown coloration and were soft and elastic. The patient reported that the lesions started as papules a year ago and had joined forming nodules. Her medical record revealed history of high blood pressure, hyperthyroidism and sensory motor polyneuropathy. She also showed symptoms of Cushing syndrome but previous tested came back negative. Biopsy of the lesions along with histopathological examinations were realized and revealed normal skin morphology but staining with periodic acid-Schiff showed septate hyphae and yeast like vesicles. At the same time, sample from the lesions were inoculated on Sabouraud glucose agar with cycloheximide (an antibiotic) and incubated at 28 °C. After 6 days of incubation, colonies that velvet olive green on top and greenish black at the bottom were obersved. Observation of these colonies under the microscope revealed clustered of blastoconidia attached to a central branched hyphae (Figure 7b), defining characteristic of Cladosporium oxysporum. The patient was thus diagnosed with cutaneous phaeohyphomycosis caused by Cladosporium oxysporum. The doctors proposed surgical lesion of the excision but the patient did not agree. Consequently, she was treated with itraconazole (200mg/day) for 3 months. After the treatment the nodular lesions highly leaving some minor scars. However examination fibrotic tissue as check-up test revealed presence of spetate hyphae similar to the one seen before. Immunohistochemicals examination showed increased in level of  CD8 and CD1a and HLA-Dr, which means that inflammatory cells were activated. While the hormone profile indicated increase in urine free cortisol and ACTH level. Based on the first diagnosis and he latest results, the second diagnosis was pituitary adenoma. This time a surgical removal of the adenoma had to be performed and itraconazole was replaced with ketoconazole (3dose/day of 100mg each) for three month. The patient recover after the treatment, and no recurrence of the infection was seen after 6 months follow-up.

Figure 7: (a) Nodular lesions on the right leg of the patient. (b) Clusters of conidia typical of Cladosporium oxysporum.

Taxonomy challenges

This anarmorphic genus present some challenge for taxonomy due to the fact that most species have no known teleomorphs  (12) and are suspected to have lost the ability to reproduce sexually (1), while other species are associated with teleomorphic species, Mycosphaerella (11).  Also some of the species previously known as Cladosporium have been moved to the genera Cladophora and Exophiala (12). That is the case for Cladosporium devriesii, Cladosporium carrionii and Cladosporium mansonii which have been renamed respectively to Cladophialophora devriesii, Cladophialophora carrionii and Exophiala castellanii (12).

Reference

1.     De Hoog GS, Gueho E, Masclaux F, Gerrits van den Ende AH, Kwon-Chung KJ, McGinnis MR. 1995. Nutritional physiology and taxonomy of human- pathogenic Cladosporum-Xylohyphy species. J Med Vet Mycol. 33: 339-47.

2.      Aglawe V., Tamrakar M., Singh S.M. and Sontakke H. 2013. Systemic phaeohyphomycosis caused by Cladosporium cladosporioides: in vitro sensitivity and its serological diagnosis. Advances in Life Science and Technology. 8: 16-20.

3.     Bouziane H, Latge JP, Fitting C, Mecheri S, Lelong M, David B. 2005. Comparison of the allergenic potency of spores and mycelium of Cladosporium. Allergol Immunopathol (Madr) .33: 125-30.

4.     Dixon DM, Walsh TJ, Merz WG, McGinnis MR. 1989. Infections due to Xylohypha bantiana (Cladosporium trichoides). Rev Infect Dis. 11: 515-25.

5.     Fairs A, Wardlaw AJ, Thompson J, Pashley CH. 2010. Guidelines on ambient intramural airborne fungal spores. J Investig Allergol Clin Immunol. 20: 490-8.

6.     Garg N, Devi IB, Vajramani GV, Nagarathna S, Sampath S, Chandramouli B A, Chandramuki A, Shankar S K. 2007. Central nervous system cladosporiosis: An account of ten culture-proven cases. Neurol India. 55:282-8

7.     Knutsen AP, Bush RK, & Demain JG . 2012. Fungi and allergic lower respiratory tract diseases. J Allergy Clin Immunol. 129: 280-91.

8.     Patterson JW Warren NG, Kelly LW. 1999. Cutaneous phaeohyphomycosis due to Cladophialophora bantiana. J Am Acad Dermatol. 40: 364-6.

9.     Reiss E., Shadomy H. J. & Lyon M G. 2012. Fundamental of Medical Mycology. Hoboken, NJ: Wiley-Blackwell.

10. Romano C, Bilenchi R, Alessandrini C, Miracco C. 1999. Cutaneous phaeohyphomycosis caused by Cladosporium oxysporum. Mycoses..42(1-2):111-5.

11. Tasic S & Miladinovic N. 2007. Cladosporium spp. – Cause of opportunistic mycoses. Institute of Microbiology Faculty of Medecin. 24 : 15-19

12. Webster J. & Weber R. W. S. 2007. Introduction to fungi 3^rdedition. New York, NY: Cambridge University press.

13. Yano S, Koyabashi K, Kato K. 2003. Intrabronchial lesion due to Cladosporium sphaerospermum in a healthy, non-asthmatic woman. Mycoses. 46: 348-50.

14. Ogórek R, Lejman A, Pusz W, Miłuch A, & Miodyńska P. 2012. Characteristics and taxonomy of Cladosporium fungi. Mikologia Lekarska. 19: 80-85.

15. De Hoog GS, Queiroz-Telles F, Haase G, Fernandez-Zeppenfeldt G, Angelis DA, van den Ende A, Matos T, Peltrohe-Llacsahuanga H, Pizzirani-Kleiner AA rainer J, Richard-Yegres N, Vicente V, Yerges F. 2000. Black fungi: clinical and pathogenic approaches. Med Mycol. 38: 243-50.