Journal of Threatened Taxa | www.threatenedtaxa.org | 26
March 2020 | 12(4): 15530–15534
ISSN 0974-7907 (Online) | ISSN 0974-7893 (Print)
doi: https://doi.org/10.11609/jott.5091.12.4.15530-15534
Molecular characterization of stinkhorn fungus Aseroë
coccinea Imazeki et Yoshimi ex Kasuya 2007 (Basidiomycota: Agaricomycetes: Phallales) from India
Vivek Bobade
1 & Neelesh Dahanukar
2
1 Department of
Microbiology, Modern College of Arts, Science and Commerce, Shivajinagar, Pune,
Maharashtra 411005, India.
2 Indian
Institute of Science Education and Research, Dr. Homi Bhabha Road, Pashan, Pune,
Maharashtra 411008, India.
2 Zoo Outreach
Organization, No. 12 Thiruvannamalai Nagar, Saravanampatti, Coimbatore, Tamil Nadu 641035, India.
1 viveknbobade@gmail.com, 2 n.dahanukar@iiserpune.ac.in
(corresponding author)
Editor: Anonymity requested. Date
of publication: 26 March 2020 (online & print)
Citation: Bobade, V. & N. Dahanukar. (2020). Molecular
characterization of stinkhorn fungus Aseroë
coccinea Imazeki et Yoshimi ex Kasuya 2007 (Basidiomycota: Agaricomycetes: Phallales) from India. Journal of Threatened Taxa 12(4): 15530–15534. https://doi.org/10.11609/jott.5091.12.4.15530-15534
Copyright: © Bobade & Dahanukar 2020. Creative Commons Attribution 4.0 International
License. JoTT
allows unrestricted use, reproduction, and distribution of this article in any
medium by providing adequate credit to the author(s) and the source of
publication.
Funding: None.
Competing interests: The authors declare no competing interests. The views expressed are those of the author.
Acknowledgements: VB is thankful to the principal and head, Department
of Microbiology, Modern College of Arts, Science and Commerce, Shivajinagar,
Pune. We are grateful to Dr. Milind Watve for
encouragement.
The
fungal family Phallaceae, commonly known as stinkhorn
mushrooms, is a member of order Phallales in the
division Basidiomycota. Although the Dictionary
of fungi (Kirk et al. 2008) recognized 77 species under 21
genera, subsequent description of a new genus (Cabral et al. 2012), new species
(Gogoi & Parkash 2015; Trierveiler-Pereira et al. 2017) and new distributional
records (Gogoi & Parkash
2014; Kour et al. 2016) points to knowledge gap
regarding the diversity and distribution of members within Phallaceae. One of the interesting genera of Phallaceae is the pantropical Aseroë,
which is morphologically characterized as having a fruiting body consisting of
pseudo-stipe that is partly covered at the top by a disc from the margin of
which spring numerous, long, acute, fundamentally paired arms, which adopt a
horizontal position at maturity; its gleba is located
on the upper surface of the disc and adaxial faces of the arms (Dring 1980). While
describing the species A. coccinea, Kasuya
(2007) considered four valid species within the genus, namely, A. arachnoidea, A. coccinea, A. floriformis and A. rubra,
however, recent phylogenetic studies (Cabral et al. 2012; Trierveiler-Pereira
et al. 2014) suggested that Aseroë is not
monophyletic and A. arachnoidea was
transferred to Lysurus (Trierveiler-Pereira
et al. 2014), while A. floriformis was
transferred to Abrachium (Cabral et al.
2012). While there is no issue regarding
the generic status of the name bearing type Aseroë
rubra, generic status of A. coccinea has
not been assessed using molecular methods.
In
India, there are reports of Lysurus arachnoideus and A. rubra
(Narasimhan 1932; Iyengar & Krishnamurthy 1954;
Vasudeva 1962; Mohanan 2011a,b; Pradhan et al. 2012),
however, there are no records of A. coccinea. In fact, to our knowledge, there are no
reports of A. coccinea from anywhere outside its type locality in
Japan. In the current communication, we
provide the first report of A. coccinea from northern Western Ghats of
India and provide its phylogenetic placement based on nuclear internal
transcribed spacer region.
Six
specimens of A. coccinea were observed at Khandobacha
Mal (18.252ºN, 73.674ºE, 830m), at the base of Rajgad
fort, Pune, India. Two specimens were
collected in a clean bottle. A small
piece of stipe from each specimen was preserved in absolute ethanol for genetic
study, while the specimens were dried for long term preservation. One of the collected specimens is deposited
in the culture collection of Ajrekar Mycological
Herbarium (AMH), National Fungal Culture Collection of India (NFCCI & FIS),
Biodiversity and Palaeobiology Group, MACS-Agharkar Research Institute, Pune, India, under the
accession number AMH 9967.
DNA
was extracted from two specimens using QIAamp DNA
Mini Kit following manufacturer’s protocol.
The nuclear gene encoding small-subunit ribosomal RNA (18S rRNA) was
amplified using the primer pair A (5’-CCA ACC TGG TTG ATC CTG CCA GT-3’) and B
(5’-GAT CCT TCT GCA GGT TCA CCT AC-3’) (Berger et al. 1998). The internal transcribed spacer (ITS) region
in the nuclear ribosomal repeat unit was amplified using primer pair ITS1f
(5’-CTT GGT CAT TTA GAG CGA AGT A-3’) (Gardes & Bruns 1993) and ITS4 (5’-TCC TCC GCT TAT TGA TAT GC-3’)
(White et al. 1990), which included partial 18S, complete ITS1, complete 5.8S,
complete ITS2 and partial 28S. Protocol for PCR amplification, PCR product
purification and DNA sequencing follow Suranse et al.
(2017) with the annealing temperature 55ºC for 18S and 50ºC for ITS. Sequences generated as a part of this study
are deposited in GenBank under the accession numbers MK543504–MK543505 for 18S
and MK541641–MK541642 for ITS.
Since
limited genetic data were available for 18S gene, genetic analysis was
performed only for ITS region. Additional ITS sequences for Phallales
were downloaded from NCBI database (https://www.ncbi.nlm.nih.gov/).
Gomphus ludovicianus
(Gomphales: Gomphaceae) was
used as an outgroup following Trierveiler-Pereira et
al. (2014). Sequences were aligned using MUSCLE (Edgar 2004) implemented in
MEGA 7 (Kumar et al. 2016). Model for
nucleotide substitution pattern was determined using ModelFinder
(Kalyaanamoorthy et al. 2014) based on Bayesian
Information Criterion (Schwarz 1978) and was used for constructing maximum
likelihood tree in IQTree (Nguyen et al. 2015) with
ultrafast bootstrap (Hoang et al. 2018) support for 1000 iterations. Phylogenetic tree was viewed and edited in FigTree (Rambaut 2009). Raw
genetic p distance was calculated using MEGA 7 (Kumar et al. 2016).
Six
fruiting bodies of A. coccinea were found growing on donkey dung in dry
deciduous forest floor at Khandobacha Mal. Basidiomes of four specimens are shown in Image 1a–d. Observed specimens showed following
morphology. Basidiome terrestrial, gastrocarpic appearance, approximately 30–60 mm in height; gleba horizontally expanded, origin at the base of the
arms, and at the top of the stipe covering the disc at the upper surface of
receptacle, olivaceous green to dark green in color, mucoid, granular;
receptacle white in color, cylindrical, spongy, hollow with single chamber, at
the apex flattened to form a disc, about 10–12 mm in diameter; arms arise from
the tip of receptacle, 25–50 mm in length, single arms arise from the margin of
horizontal discoid portion, diameter at the base 5 mm in the middle 3mm and at
the tip less than 1mm, single chambered (Image 1e), nonbifurcating,
9 (n = 1), 11 (n = 4) or 13 (n = 1) arms, it diameter at the base about 5mm, in
the middle about 3mm, at the tip less than 1mm, red color on the dorsal
surface, pale red to white ventrally, at maturity the arms are fully expanded;
basidiospores hyaline (Image 1f), cylindrical or elongated in shape, mean spore
dimensions were 4.4 (sd 0.3) × 1.9 (sd 0.2) µm , average spore quotient (length/width) of 2.4 (sd 0.2); spread gregarious as well as solitary;
saprophytic, growing on donkey dung among the grasses close to the ground
(epigeal).
Morphologically,
the species closely resembles the original description of A. coccinea
(see Kasuya 2007) except for the number of arms that
varied from 9–13 in our specimens as appose to 7–9 arms reported in the
original description. Further, the
specimens we observed were slightly larger than the type of A. coccinea. The specimens in our collection differ from A.
rubra in having nonbifurcating
and single chambered arms versus bifurcating and several chambered arms in A.
rubra (Kasuya 2007; Hemmes & Desjardin 2009). Further, the specimens in our collection
differs from morphologically closely related species Lysurus
arachnoideus in having dorsally reddish arms
versus white arms in L. arachnoideus (Kasuya 2007; Hemmes &
Desjardin 2009) and basidiospores larger and ellipsoid to
cylindrical (Image 1f) 3.6–5.0 × 1.5–2.4 µm versus 2.5–3.5× 1.5 µm in L.
arachnoideus (Kasuya
2007).
Our
communication provides the first report of A. coccinea from northern
Western Ghats of India and first report of this species under this nomen from outside its type locality in Japan. It is essential to note, however, that the
earlier report of L. arachnoideus (as Aseroë arachnoidea)
from Karnataka by Narasimhan (1932) needs a critical evaluation. Description of anomalous specimen of L.
arachnoideus by Narasimhan (1932) with red
colored arms has a close resemblance with the description of A. coccinea
from our study. In fact, while
describing the species A. coccinea, Kasuya
(2007) pointed out that the description of Indian specimens provided by
Narasimhan (1932) shares some characters with A. coccinea. We believe that the species collected by
Narasimhan (1932) is same as the species in our collection, indicating that A.
coccinea is distributed in both Maharashtra and Karnataka part of the
Western Ghats. One notable difference
among the specimens studied by Narasimhan (1932), type studied by Kasuya (2007) and our observation is the size of the
fruiting bodies. Type studied by Kasuya (2007) is a small specimen as compared to the
specimens we studied from northern Western Ghats, while the specimen dimensions
provided by Narasimhan (1932) are very large.
For instance, Kasuya (2007) reports the length
of arm as 4 to 10 mm, we report it as 25–50 mm, while Narasimhan (1932) reports
it as 38 to 40 mm. Because the number of
specimens studied in all three studies are limited the variation in the size
cannot be explained at the moment.
Genetically,
A. coccinea from our study is sister taxon to L. arachnoideus (Figure 1). Nevertheless, the two species are separated
by a raw genetic distance of 5.4%. This
result is not very surprising, because even in the original description of A.
coccinea, Kasuya (2007) pointed out close
resemblance between the two species. The fact is, however, that A. coccinea
is nested within the well supported Lysurus
clade, suggests that the species needs to be transferred to the genus. In the current communication, we refrain from
transferring A. coccinea to Lysurus
because of two reasons, (1) there is need for detailed taxonomic study of the
group, preferably including the material from the type locality, for delimiting
the generic boundary for Aseroë by studying
its type species A. rubra from throughout its
distributional range and reassessment of A. coccinea, and (2) taxonomic
sampling for the genetic analysis in our study is not adequate because of
limited information available on 18S and ITS markers of family Phallaceae.
Nevertheless, we make the data available for two genetic markers, 18S
and ITS region, of specimens from our study, which can facilitate further
comparative genetic studies on this group.
Our
report of A. coccinea, along with its genetic information,
suggests that there is a need for more exploratory surveys for understanding
diversity, distribution and taxonomy of Phallales of
India.
For
figure & image - - click here
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