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Stud Mycol 59(1): 75-88 2007
DOI: 10.3114/sim.2007.59.10
Copyright © 2007 CBS Fungal Biodiversity Centre
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Polyphasic taxonomy of Aspergillus section Candidi based on molecular, morphological and physiological data

J. Varga1,3,*, J.C. Frisvad2 and R.A. Samson1

1 CBS Fungal Biodiversity Centre, Uppsalalaan 8, NL-3584 CT Utrecht, the Netherlands
2 BioCentrum-DTU, Building 221, Technical University of Denmark, DK-2800 Kgs. Lyngby, Denmark
3 Department of Microbiology, Faculty of Science and Informatics, University of Szeged, H-6701 Szeged, P.O. Box 533, Hungary

* Correspondence: János Varga, j.varga{at}cbs.knaw.nl


   Abstract
TOP
 Abstract
INTRODUCTION
 MATERIALS AND METHODS
 RESULTS AND DISCUSSION
 References
 
Aspergillus section Candidi historically included a single white-spored species, A. candidus. Later studies clarified that other species may also belong to this section. In this study, we examined isolates of species tentatively assigned to section Candidi using a polyphasic approach. The characters examined include sequence analysis of partial β-tubulin, calmodulin and ITS sequences of the isolates, morphological and physiological tests, and examination of the extrolite profiles. Our data indicate that the revised section Candidi includes 4 species: A. candidus, A. campestris, A. taichungensis and A. tritici. This is strongly supported by all the morphological characteristics that are characteristic of section Candidi: slow growing colonies with globose conidial heads having white to yellowish conidia, conidiophores smooth, small conidiophores common, metulae present and covering the entire vesicle, some large Aspergillus heads with large metulae, presence of diminutive heads in all species, conidia smooth or nearly so with a subglobose to ovoid shape, and the presence of sclerotia in three species (A. candidus, A. taichungensis and A. tritici). Aspergillus tritici has been suggested to be the synonym of A. candidus previously, however, sequence data indicate that this is a valid species and includes isolates came from soil, wheat grain, flour and drums from India, Ghana, Sweden, The Netherlands and Hungary, making it a relatively widespread species. All species produce terphenyllins and candidusins and three species (A. candidus, A. campestris and A. tritici) produce chlorflavonins. Xanthoascins have only been found in A. candidus. Each of the species in section Candidi produce several other species specific extrolites, and none of these have been found in any other Aspergillus species. A. candidus has often been listed as a human pathogenic species, but this is unlikely as this species cannot grow at 37 °C. The pathogenic species may be A. tritici or white mutants of Aspergillus flavus.

Taxonomic novelty: revalidation of Aspergillus tritici Mehrotra & Basu.

Keywords Ascomycetes / Aspergillus section Candidi / β-tubulin / calmodulin / Eurotiales / extrolites / ITS / polyphasic taxonomy


   INTRODUCTION
TOP
 Abstract
 INTRODUCTION
MATERIALS AND METHODS
 RESULTS AND DISCUSSION
 References
 
Aspergillus section Candidi (Gams et al. 1995; A. candidus species group according to Raper & Fennell 1965) was established by Thom & Raper (1945) to accomodate a single white-spored species, A. candidus Link. This species frequently contaminates stored food and feeding stuff (Kozakiewicz 1989; Park et al. 2005). A. candidus is moderately xerophilic, and able to grow on stored grains with 15 % moisture content (Lacey & Magan 1991), raising the moisture level of the infested grain to 18 percent or higher, and the temperature to up to 55 °C. This species is one of the most frequently encountered mould in cereal grains and flour (Rabie et al. 1997; Weidenbörner et al. 2000; Ismail et al. 2004; Hocking 2003). A. candidus causes loss of viability and germ discolouration in cereals (Papavizas & Christensen 1960; Battacharya & Raha 2002; Lugauskas et al. 2006). It also occurs in soil, usually on seeds or in the rhizosphere, and also in milk (Raper & Fennell 1965; Kozakiewicz 1989; Moreau 1976).

A. candidus enzymes has also been used in the fermentation industry for the production of galacto-oligosaccharides (Zheng et al. 2006), and D-mannitol (Smiley et al. 1969), while some A. candidus metabolites including terphenyllins has antioxidant and anti-inflammatory activities (Yen et al. 2001, 2003). A. candidus is also used in the meat industry for spontaneous sausage ripening (Gracia et al. 1986; Sunesen & Stahnke 2003).

A. candidus is claimed to be involved in a wide range of human infections including invasive aspergillosis (Rippon 1988; Ribeiro et al. 2005), otomycosis (Yasin et al. 1978; Falser 1983), brain granuloma (Linares et al. 1971) and onychomycosis (Schonborn & Schmoranzer 1970; Zaror & Moreno 1980; Piraccini et al. 2002). A. candidus has also caused various disorders in pigs (Moreau 1979) and was found to be the second most prevalent Aspergillus species in a hospital surveillance project in the U.S.A. (Curtis et al. 2005). Concentration of A. candidus conidia can reach alarming levels in grain dust and was suggested to contribute to the development of the so-called organic dust toxic syndrome (Weber et al. 1993; Krysinska-Traczyk & Dutkiewicz 2000). A. candidus is able to induce both cellular and humoral response in animals (Krysinska-Traczyk & Dutkiewicz 2000). A. candidus metabolites including terphenyl compounds and terprenins exhibit immunomodulating capabilities and are highly cytotoxic (Shanan et al. 1998; Krysinka & Dutkiewicz 2000). There is some evidence that A. candidus might be toxic to chickens and rats (Marasas & Smalley 1972) and has also been isolated from birds (Saez 1970, Sharma et al. 1971). A. candidus has been reported to produce several secondary metabolites including candidusins (Kobayashi et al. 1982; Rahbaek et al. 2000), terprenins (Kamigauchi et al. 1998), chlorflavonin (Bird & Marshall 1969), dechlorochlorflavonin (Marchelli & Vining 1973), xanthoascin (Takahashi et al. 1976b), kojic acid (Kinosita & Shikata 1969, Saruno et al. 1979, Cole & Cox 1981), 3-nitro-propionic acid (Kinosita et al. 1968), and 6-sulfoaminopenicillanic acid (Yamashita et al. 1983). A. candidus is reported to produce citrinin but the first report of citrinin production by an Aspergillus confused A. niveus with A. candidus (Timonin & Rouatt 1944; Raper & Fennell 1965). However, some later reports indicate that some isolates may produce citrinin (Kinosita & Shikata 1969; Cole & Cox 1981).

The description of A. candidus is admittedly broad, encompassing considerable variability among the isolates (Raper & Fennell 1965, Kozakiewicz 1989). A. candidus is characterised by white conidial heads, globose to subglobose vesicles, biseriate large and uniseriate small conidial heads, and smooth conidiophores and conidia (Raper & Fennell 1965, Kozakiewicz 1989). Several white-spored Aspergillus species described in the past have been synonymised with A. candidus, including A. albus, A. okazakii, or A. dubius (Raper & Fennell 1965). Raper & Fennell (1965) also stated that "it is possible that our current concept of A. candidus is too broad". Recent studies indicated that other species including A. campestris (Christensen 1982; Rahbaek et al. 2000; Peterson 2000; Varga et al. 2000) and A. taichungensis (Yaguchi et al. 1995, Rahbaek et al. 2000) are also members of section Candidi. Besides, two other white-spored species, A. tritici (as A. triticus, Mehrotra & Basu 1976) and A. implicatus (Maggi & Persiani 1994) have also been suggested to belong to this section.

In this study, we examined available isolates of the species, proposed to belong to section Candidi, to clarify the taxonomic status of this section. The methods used include sequence analysis of the ITS region (including internal transcribed spacer regions 1 and 2, and the 5.8 S rRNA gene of the rRNA gene cluster), and parts of the β-tubulin and calmodulin genes, macro- and micromorphological analysis, and analysis of extrolite profiles of the isolates.


   MATERIALS AND METHODS
TOP
 Abstract
 INTRODUCTION
 MATERIALS AND METHODS
RESULTS AND DISCUSSION
 References
 
Morphological examinations
The strains examined are listed in Table 1. The strains were grown for 7 d as 3-point inoculations on Czapek agar, Czapek yeast autolysate agar (CYA), malt extract agar (MEA), and oat meal agar (OA) at 25 °C (medium compositions in Samson et al. 2004).


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  		Table 1. The Aspergillus section Candidi isolates examined in this study.

 

Analysis for secondary metabolites
The cultures were analysed according to the HPLC-diode array detection method of Frisvad & Thrane (1987, 1993) as modified by Smedsgaard (1997). The isolates were analyzed on CYA and YES agar using three agar plugs (Smedsgaard 1997). The secondary metabolite production was confirmed by identical UV spectra with those of standards and by comparison to retention indices and retention times in pure compound standards (Rahbaek et al. 2000).

Isolation and analysis of nucleic acids
The cultures used for the molecular studies were grown on malt peptone (MP) broth using 10 % (v/v) of malt extract (Oxoid) and 0.1 % (w/v) bacto peptone (Difco), 2 mL of medium in 15 mL tubes. The cultures were incubated at 25 °C for 7 d. DNA was extracted from the cells using the MasterpureTM yeast DNA purification kit (Epicentre Biotechnol.) according to the instructions of the manufacturer. Fragments containing the ITS region were amplified using primers ITS1 and ITS4 as described previously (White et al. 1990). Amplification of part of the β-tubulin gene was performed using the primers Bt2a and Bt2b (Glass & Donaldson 1995). Amplifications of the partial calmodulin gene were set up as described previously (Hong et al. 2005). Sequence analysis was performed with the Big Dye Terminator Cycle Sequencing Ready Reaction Kit for both strands, and the sequences were aligned with the MT Navigator software (Applied Biosystems). All the sequencing reactions were purified by gel filtration through Sephadex G-50 (Amersham Pharmacia Biotech, Piscataway, NJ) equilibrated in double-distilled water and analyzed on the ABI PRISM 310 Genetic Analyzer (Applied Biosystems). The unique ITS, β-tubulin, and calmodulin sequences were deposited at the GenBank nucleotide sequence database under accession numbers EU076291 [GenBank] -EU076311.


Figure 1
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  		Fig. 1. Neighbour-joining tree based on β-tubulin sequence data of Aspergillus section Candidi. Numbers above branches are bootstrap values. Only values above 70 % are indicated.

 
Data analysis
The sequence data was optimised using the software package Seqman from DNAStar Inc. Sequence alignments were performed by using CLUSTAL-X (Thompson et al. 1997) and improved manually. The neighbour-joining (NJ) method was used for the phylogenetic analysis. For NJ analysis, the data were first analysed using the Tamura-Nei parameter distance calculation model with gamma-distributed substitution rates (Tamura & Nei 1993), which were then used to construct the NJ tree with MEGA v. 3.1 (Kumar et al. 2004). To determine the support for each clade, a bootstrap analysis was performed with 1000 replications.

For parsimony analysis, the PAUP v. 4.0 software was used (Swofford 2002). Alignment gaps were treated as a fifth character state and all characters were unordered and of equal weight. Maximum parsimony analysis was performed for all data sets using the heuristic search option with 100 random taxa additions and tree bisection and reconstruction (TBR) as the branch-swapping algorithm. Branches of zero length were collapsed and all multiple, equally parsimonious trees were saved. The robustness of the trees obtained was evaluated by 1000 bootstrap replications (Hillis & Bull 1993). An A. flavus isolate was used as outgroup in these experiments.


   RESULTS AND DISCUSSION
TOP
 Abstract
 INTRODUCTION
 MATERIALS AND METHODS
 RESULTS AND DISCUSSION
References
 
Phylogeny
We examined the genetic relatedness of section Candidi isolates using sequence analysis of the ITS region of the ribosomal RNA gene cluster, and parts of the calmodulin and β-tubulin genes. During analysis of part of the β-tubulin gene, 496 characters were analyzed, among which 68 were found to be parsimony informative. The Neighbour-joining tree based on partial β-tubulin genes sequences is shown in Fig. 1. The topology of the tree is the same as the single maximum parsimony tree constructed by the PAUP program (length: 240 steps, consistency index: 0.8833, retention index: 0.9263). The calmodulin data set included 532 characters, with 43 parsimony informative characters (Fig. 2). The topology of the Neighbour-joining tree was the same as that of one of the 78 maximum parsimony trees (tree length: 300, consistency index: 0.9633, retention index: 0.9396). The ITS data set included 492 characters with 5 parsimony informative characters. The Neighbour joining tree shown in Fig. 3 has the same topology as one of the more than 105 maximum parsimony trees (tree length: 35, consistency index: 1 0000, retention index: 1 0000).


Figure 2
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  		Fig. 2. Neighbour-joining tree based on calmodulin sequence data of Aspergillus section Candidi. Numbers above branches are bootstrap values. Only values above 70 % are indicated.

 

Figure 3
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  		Fig. 3. Neighbour-joining tree based on ITS sequence data of Aspergillus section Candidi. Numbers above branches are bootstrap values. Only values above 70 % are indicated.

 

Phylogenetic analysis of both β-tubulin and calmodulin sequence data indicated that Aspergillus section Candidi includes 4 species, namely: A. candidus, A. campestris, A. taichungensis and A. tritici. Interestingly, the reference strain of A. candidus, CBS 283.95 was found to belong to the A. tritici species. Isolates CBS 597.65 and CBS 112449 were found to be related to the A. taichungensis type strain based on β-tubulin sequence data, and formed a distinct clade on the tree based on calmodulin sequences. Further studies are needed to clarify the taxonomic position of these isolates.

Comparison of our ITS sequence data to those available on the web site of the Japan Society for Culture Collections (http://www.nbrc.nite.go.jp/jscc/idb/search) indicated that several strains held as A. candidus represent other species. Three strains (NBRC 4389 = IFO 4389, NBRC 4037 = IFO 4037, and NBRC 4322 = IFO 4322) were found to be actually white-spored A. oryzae isolates, NBRC 5468(= IFO 5468) and NBRC 33019(= IFO 33019 = CBS 283.95 = SRRC 310) belong to A. tritici, while NBRC 32248 (= IFO 32248) has identical ITS sequence to A. campestris. However, further loci should also be analyzed to confirm their assignment. Other isolates including NBRC 8816, NBRC 4309, NBRC 4310 and NBRC 4311 are representatives of the A. candidus species based on their identical ITS sequences.

Aspergillus implicatus, another species previously assigned to this section (Maggi & Persiani 1994), was found to be more closely related to A. anthodesmis based on sequence data, which places this species close to Aspergillus section Sparsi (data not shown). Further studies are needed to clarify the taxonomic position of this white-spored species within the Aspergillus genus.

Chemotaxonomy
All strains of species in section Candidi produced terphenyllins and candidusins. Aspergillus candidus isolates produced candidusins A and B, terphenyllin, 3-hydroxyterphenyllin and some isolates also produced chlorflavonin and a chlorflavonin analogue. A. tritici isolates differed from A. candidus in not producing candidusin A and chlorflavonin. A. taichungensis produced candidusin C, terphenyllin, and 3-hydroxyterphenyllin, while the type strain of A. campestris also produced chlorflavonin. Xanthoascin was only found in some strains of A. candidus and not in any other species in Candidi. Each species produced a large number of as yet not structure elucidated extrolites. These extrolites, including terphenyllins, candidusins, chlorflavonins and xanthoascin, have only been found in section Candidi and not in any other aspergilli, except for A. ellipticus, that produces terphenyllin and candidusin (Samson et al. 2004, 2007).

Morphology
Aspergillus candidus is a wide-spread species throughout the world. According to Raper & Fennell (1965), "a typical strain of A. candidus differs little from members of the A. niger group except for the absence of both pigmentation and roughening in the conidia". Another interesting feature observed in A. candidus is the production of diminutive conidial heads which are frequently uniseriate in contrast with the biseriate large heads. Colonies on CYA and MEA usually slow growing, colonies white to cream coloured, reverse usually uncoloured. Conidial heads usually biseriate, white to cream coloured, at first globose, with spore chains later adherent in loose divergent columns, diminutive heads commonly produced, conidiophores varying with the strain from less than 500 µm to up to 1000 µm long, thick walled, smooth, occasionally septate, vesicles globose to subglobose, ranging from 40 µm or more in diam in very large heads to less than 10 µm in small heads, typically fertile over the whole surface, phialides occasionally uniseriate in small heads but typically in two series, colourless, conidia globose or subglobose in most strains to elliptical in others, thin walled, 2.5-3.5 µm or occasionally 4 µm, smooth, colourless. Sclerotia, when produced, at first white, quickly becoming reddish purple to black, consisting of thick-walled parenchyma-like cells. A. candidus is unable to grow at 37 °C.

Aspergillus taichungensis was described by Yaguchi et al. (1995) from soil, Taiwan. The species is characterised by restricted growth on CZA and MEA at 25 °C, colonies white to pale yellow, velvety, reverse uncoloured. Conidial heads radiate, biseriate, conidiophores smooth, 300-450 µm long, often diminutive (90-250 µm long, biseriate), vesicles hemispherical to elongate, 5-20 µm in diam, fertile over the upper half to two-thirds, conidia hyaline, yellow in mass, globose to subglobose, microverrucose, 3-4 µm. Dark brown sclerotia which appear on MEA after more that 25 d incubation. A. taichungensis is able to grow at 37 °C on CYA.

Aspergillus campestris was described by Christensen (1982) from native prairie soil, North Dakota. The species is characterised by its restricted growth on CZA and MEA at 25 °C, colonies velvety, sulphur yellow, reverse uncoloured. Conidial heads biseriate, radiate, conidiophores usually 400-800 µm but can be up to 1 300 µm long, smooth, often diminutive (up to 100 µm long, biseriate), vesicles globose to slightly elongate, 25-40 µm in diam, fertile over the entire surface, conidia thin-walled, hyaline, pale yellow in mass, slightly ellipsoidal, 3-4 x 2.3-3 µm. Sclerotia not observed. A. campestris is unable to grow at 37 °C on any media tested.

Aspergillus tritici was described as A. triticus by Mehrotra & Basu (1976) from wheat grains, India. Colonies are slow-growing on CZA and MEA, white to light cream coloured, reverse light brown. Conidial heads are biseriate, radiate, conidiophores thick-walled, septate, 130-700 µm long, often diminutive (10-75 µm, sometimes uniseriate), vesicles elongated, small (5-11 µm), conidia globose to subglobose, slightly roughened, 2.7-3.5 µm. At maturity conidia are embedded in a water drop giving the conidial heads a "slimy" appearance. The sclerotia are at first white, later becoming purple to black. A. tritici grows well at 37 °C.

Based on a polyphasic investigation of Aspergillus section Candidi, the section includes four species: A. candidus, A. campestris, A. taichungensis and A. tritici. Phenotypic characteristics of these species are shown in Table 2. A. campestris was placed in section Circumdati because of its yellowish white conidia and it was not considered closely related to A. candidus by Christensen (1982). A. taichungensis was equivocally placed in either section Versicolores, Terrei or Flavipedes (Yaguchi et al. 1995). However, the phylogenetic and chemotaxonomic evidence presented here indicates that both species belong to section Candidi. This is strongly supported by all the morphological characteristics that are characteristic of the section Candidi: slow growing colonies with globose conidial heads having white to yellowish conidia, conidiophores smooth, small conidiophores common, metulae present and covering the entire vesicle, some large Aspergillus heads with large metulae, conidia smooth or nearly so with a subglobose to ovoid shape (albeit slightly ellipsoidal in A. campestris), and sclerotia present in A. taichungensis, A. candidus and A. tritici. Sclerotia have not been observed in A. campestris, but have been observed in A. candidus (light cream coloured turning purple to black in age). Aspergillus tritici has been suggested to be the synonym of A. candidus by Samson (1979). However, sequence data indicate that this is a valid species and includes isolates from soil, wheat grain, flour and drums from India, Ghana, Sweden, The Netherlands and Hungary, making it a relatively widespread species.


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  		Table 2. Phenotypic characteristics of species in Aspergillus section Candidi.

 

Aspergillus campestris Christensen, Mycologia 74: 212. 1982. Fig. 4.


Figure 4
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  		Fig. 4. Aspergillus candidus. A-B Colonies after 7 d at 25 °C A. CYA. B. MEA. C, G. Conidial heads. D-F, H-K. Conidiophores. H. Sclerotia. L. Conidia. Scale bars = 10 µm.

 


Figure 5
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  		Fig. 5 Aspergillus campestris. A-B Colonies after 7 d at 25 °C A. CYA. B. MEA. C. Conidial heads. D-H. Conidiophores. I. Conidia. Scale bars = 10 µm.

 
Type: CBS 348.81, from soil from native prairie, North Dakota, U.S.A.

Other no. of the type: IBT 27921 = IBT 13382

Description
Colony diam: CZA25: 10-12 mm; CYA25: 10-15 mm, MEA25: 7-10 mm, YES25: 18-24 mm, OA25: 9-12 mm, CYA37: 0 mm, CREA25: poor growth, no acid production

Colony colour: sulphur yellow to pinard yellow

Conidiation: abundant

Reverse colour (CZA): uncoloured

Colony texture: velvety

Conidial head: radiate, splitting in age

Stipe: 400-800(-1300) x 7-12 µm

Vesicle diam/shape: (18-)24-36(-46) µm, globose to subglobose Conidium size/shape/surface texture: 3-4 x 2.3-3 µm, ellipsoidal to egg-shaped, smooth

Cultures examined: KACC 42091, KACC 42090 = IBT 27920, KACC 41955 = IBT 3016, UAMH 1324 (from mouse, Canada, as A. sulphureus), IBT 17867

Diagnostic features: restricted growth on all media, sulphur yellow colony colour and diminutive conidial heads

Similar species: -

Ecology and habitats: soil

Distribution: U.S.A., Canada

Extrolites: candidusin C, terphenyllins, chlorflavonin (Rahbaek et al. 2000), confirmed in this study

Pathogenicity: not reported

Note: Diminutive conidial heads commonly produced (100 x 10-12 µm)

Aspergillus candidus Link, Mag. Ges. Naturf. Freunde Berlin 3: 16. 1809. Fig. 5.

Type: CBS 566.65, from Westerdijk, 1909

Other no. of the type: ATCC 1002; IMI 091889; LSHB Ac27; NCTC 595; NRRL 303; QM 1995; WB 303

Description
Colony diam: CZA25: 15-30 mm; CYA25: 13-20 mm, MEA25: 8-14 mm, YES25: 19-33 mm, OA25: 9-18 mm, CYA37: 0 mm, CREA25: poor growth and no acid production

Colony colour: white

Conidiation: limited

Reverse colour (CZA): uncoloured to pale yellow

Colony texture: submerged

Conidial head: diminutive, with few divergent spore chains

Stipe: 500-1000 x 5-10(- 20) µm, walled, smooth, occasionally septate, colourless or slightly yellowed in age

Vesicle diam/shape: 10-40 µm, globose to subglobose

Conidium size/shape/surface texture: 2.5-3.5(- 4) µm, globose to subglobose, smooth

Cultures examined: CBS 119.28, CBS 116945, CBS 175.68, CBS 114385, CBS 120.38, CBS 225.80, CBS 102.13, CBS 118.28, CBS 566.65, 1-F9, 13-C4, 17-C2, 25-I1, IMI 091889, CBS 283.95, NRRL 5214

Diagnostic features: phialides clustered on one side of the vesicle, echinulate conidia, slow growth rate and cream-yellow reverse on CYA; unable to grow at 37 °C

Similar species: A. tritici

Distribution: worldwide (Bangladesh, Pakistan, Kuwait, Sri Lanka, Japan, South Africa, Somalia, Chad, Libya, Egypt, Syria, Israel, Argentina, Bahama Islands, New Guinea, Solomon Islands, China, Central America, Chile, Russia, Nepal, U.S.A., Spain, Italy, Hungary, Austria, Czechoslovakia, Germany, France, Britain, Ireland, Netherlands, Denmark)

Ecology and habitats: stored products, especially cereals, soil, dried fruits, dung, dried fish, indoor air

Extrolites: terphenyllin, 3-hydroxyterphenyllin (Rahbaek et al. 2000), prenylterphenyllin, 4''''-deoxyprenylterphenyllin, 4''''-deoxyisoterprenin, 4''''-deoxyterprenin (Wei et al. 2007), and other terphenyl-type compounds (Marchelli & Vining 1975 Kurobane et al. 1979; Kobayashi et al. 1985; Takahashi et al. 1976b), including candidusins (A & B) (Kobayashi et al. 1982) and other terprenins (Kamigauchi et al. 1998), chlorflavonin (Bird & Marshall 1969; Munden et al. 1970), dechlorochlorflavonin (Marchelli & Vining 1973), and xanthoascin (Takahashi et al. 1976a). The production of terphenyllin, 3-hydroxyterphenyllin, candidusin A, candidusin B, chlorflavonin and xanthoascin was confirmed by HPLC-DAD.

Extrolites not produced by A. candidus: kojic acid (Kinosita & Shikata 1969; Cole & Cox 1981), and 3-nitro-propionic acid (Kinosita et al. 1968) were reported from the same strain of A. candidus of which ATCC 44054 is representative. A re-examination of that strain showed that it was a white-spored mutant of Aspergillus flavus, a known producer of these two metabolites. The asterriquinone analogs, neoasterriquinone and isoasterriquinone (Alvi et al. 1999) have not been found in any strains of A. candidus by us. These asterriquinone analogues are probably produced by A. niveus, but this has to be confirmed by examination of isolates of the latter species. Citrinin production was observed in some studies (Timonin & Rouatt 1944; Kinosita & Shikata 1969), but the producing fungus was later identified as A. niveus (NRRL 1955, Raper and Fennell, 1965). The production of 6-sulfoaminopenicillanic acid by A. candidus (Yamashita et al. 1983) has not been confirmed

Pathogenicity: Pathogenicity of A. candidus is rather improbable, as this species cannot grow at 37 °C., however pathogenicity has often been reported: A. candidus has been claimed to be involved in a wide range of human infections including invasive aspergillosis (Rippon 1988; Ribeiro et al. 2005), pulmonary aspergillosis (Iwasaki et al. 1991), aspergilloma (Avanzini et al. 1991), otomycosis (Yasin et al. 1978; Falser 1983), brain granuloma (Linares et al. 1971) and onychomycosis (Kaben 1962; Fragner & Kubackova 1974; Cornere & Eastman 1975; Piraccini et al. 2002; Schonborn & Schmoranzer 1970; Zaror & Moreno 1980); also caused various disorders in pigs (Moreau 1979). In these cases it is more likely caused by white spored mutants of A. flavus or by A. tritici

Note: young heads varying in the same culture from globose masses 200 to 300 µm in diam to small heads less than 100 µm in diam; some isolates produce purple to black sclerotia

Aspergillus taichungensis Yaguchi, Someya & Udagawa, Mycoscience 36: 421. 1995. Fig. 6.


Figure 6
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  		Fig. 6. Aspergillus taichungensis. A-B Colonies after 7 d at 25 °C A. CYA. B. MEA. C. Conidial heads. D-H Conidiophores. I. Conidia. Scale bars = 10 µm.

 


Figure 7
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  		Fig. 7. Aspergillus tritici. A-B Colonies after 7 d at 25 °C A. CYA. B. MEA. C. Conidial heads. D-F, G-H.Conidiophores. I. Sclerota. J. Conidia. Scale bars = 10 µm.

 
Type: PF1167, from soil, Taiwan

Other no. of the type: IBT 19404

Description
Colony diam: CZA25: 12-15 mm; CYA25: 17-20 mm, MEA25: 9-13 mm in 7 d, YES25: 25-28 mm, OA25: 12-16 mm, CYA37: 7-10 mm, CREA25: poor growth, no acid production

Colony colour: yellowish white to primrose

Conidiation: moderate

Reverse colour (CZA): colourless (CZA), light yellow to pale luteous (MEA)

Colony texture: floccose (MEA)

Conidial head: loose radiate

Stipe: 300-440 x 5-9 µm

Vesicle diam/shape: 5-20 µm, hemispherical to elongate

Conidium size/shape/surface texture: 3-4 µm, globose to subglobose; sometimes ovoid, 3-5 x 3-4.5 µm, microverrucose

Cultures examined: IBT 19404, CBS 567.65, CBS 112449

Diagnostic features: slow growing colonies with globose conidial heads having white to yellowish conidia, presence of diminutive conidiophores and dark brown sclerotia

Similar species: A. candidus, A. tritici

Ecology and habitats: soil, air

Distribution: Taiwan, Brazil, Germany

Extrolites: candidusin C, terphenyllin, 3-hydroxyterphenyllin (Rahbaek et al. 2000, and confirmed in this study). A large number of additional extrolites, until now only found in this species, were also produced. These have not yet been structure elucidated, but had characteristic UV spectra

Pathogenicity: not reported

Notes:the type strain produces dark brown sclerotia 300-500 x 200-400 µm in size in 30 d (Yaguchi et al. 1995; Rahbaek et al. 2000); diminutive conidiophores present, 90-250 x 2-3 µm in size

Aspergillus tritici Mehrotra & Basu, Nova Hedwigia 27: 599, 1976. Fig. 7.

Type: CBS 266.81, from wheat grain, India

Other no. of the type: No. A x 194

Morphological characteristics
Colony diam (7 d): CZA25: 18-23 mm; CYA25: 16-29 mm, MEA25: 11-17 mm, YES25: 18-41 mm, OA25: 13-25 mm, CYA37: 7-21 mm, CREA25: poor growth, no acid production

Colony colour: white to light cream coloured

Conidiation: moderate

Reverse colour (CZA): light yellow to light brown with age

Colony texture: radially furrowed

Conidial head: short radiate

Stipe: 130-700 x 4-8 µm (diminutive stipes 10-75 x 1.5-3.5 µm), septate

Vesicle diam, shape: 4.8-11 µm, small, only slightly enlarged at the end

Conidium size, shape, surface texture: 2.7-3.5 µm, globose to subglobose, slightly roughened

Cultures examined: CBS 119225, CBS 117270, CBS 266.81, CBS 112.34, 11-H7, SZMC 0565, CBS 283.95, SZMC 0897, IBT 23116, IBT 24170

Diagnostic features: colonies more yellowish than those of A. candidus; able to grow at 37 °C

Similar species: A. candidus

Distribution: India, Ghana, Sweden, Hungary, Slovenia, South Africa

Ecology and habitats: wheat, soil

Extrolites: candidusin B, candidusin analogue, terphenyllin, 3-hydroxyterphenyllin, chlorflavonin (Rahbaek et al. 2000, and confirmed in this study)

Pathogenicity: not reported, but since this species is able to grow at 37 °C, it may have caused some of the mycoses listed under A. candidus

Notes: some isolates produce sclerotia purple to black in colour; in some isolates conidia are embedded in a water drop with age ("slimy" appearance) and produces diminutive heads


   References
TOP
 Abstract
 INTRODUCTION
 MATERIALS AND METHODS
 RESULTS AND DISCUSSION
 References
 

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