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Phytotaxa 561 (1): 053–064
https://www.mapress.com/pt/
Copyright © 2022 Magnolia Press Article PHYTOTAXA
ISSN 1179-3155 (print edition)
ISSN 1179-3163 (online edition)
Accepted by Jian-Kui Liu: 13 Aug. 2022; published: 6 Sept. 2022
https://doi.org/10.11646/phytotaxa.561.1.5
53
Madagascaromyces cannae sp. nov. (Mycosphaerellaceae) on Canna indica
(Cannaceae) from Guangdong, China
JINGWEN CHEN1,2,4, ISHARA S. MANAWASINGHE1,2,5, JIEYING LIN1,2,6, YANHONG LIN1,2,7, DHANUSHKA N.
WANASINGHE3,8 & YUNXIA ZHANG1,2,9*
1 Innovative Institute for Plant Health, Zhongkai University of Agriculture and Engineering, Guangzhou 510225, P.R. China
2 Key Laboratory of Green Prevention and Control on Fruits and Vegetables in South China, Ministry of Agriculture and Rural Affairs,
Guangzhou 510225, P.R. China
3 Center for Mountain Futures, Kunming Institute of Botany, Chinese Academy of Sciences, Honghe 654400, P.R. China
4
�
Jingwen075@163.com; https://orcid.org/0000-0003-3048-1653
5
�
ishara9017@gmail.com; https://orcid.org/0000-0001-5730-3596
6
�
linjieying178@163.com; https://orcid.org/0000-0003-0328-4523
7
�
yanhong1624@163.com; https://orcid.org/0000-0001-6415-6467
8
�
dnadeeshan@gmail.com; https://orcid.org/0000-0003-1759-3933
9
�
yx_zhang08@163.com; https://orcid.org/0000-0002-2815-2554
*Corresponding author:
�
yx_zhang08@163.com
Abstract
Microfungi associated with Cannaceae are distributed in tropical and subtropical regions. Even though many fungal taxa
have been reported from Cannaceae hosts, Mycospharellaceae on Canna indica are less studied. In this study, a saprobic
Mycosphaerellaceae species was isolated on C. indica, from Guangzhou, Guangdong province, China. To identify the spe-
cies, morphological characteristics and phylogenetic analysis of LSU, ITS and rpb2 were employed. Based on multi-gene
phylogenetic analyses and morphological comparisons, we introduce our collection as a new species, Madagascaromyces
cannae. The novel species is characterized by the septate, pale brown conidiophores, pale brown, terminal and lateral conid-
iogenous cells and pale brown, clavate, guttulate conidia with 0–6 septa. In addition, a detailed comparison of morphologi-
cal characters of Madagascaromyces species is provided. To our knowledge, this is the first report of a Madagascaromyces
species associated with C. indica in China. Our collection will be an addition to the knowledge of fungi associated with
Cannaceae.
Keywords: 1 new species, asexual morph, hyaline conidia, multi-gene
Introduction
Cannaceae comprises a single genus, which includes 55 Canna species (Kaul 1988). Most Cannaceae species are native
to tropical areas in Central and South America, West Indian Islands (Jesús et al. 2018) and China (Sun et al. 2020). In
China, these species are grown as ornamental plants in tropical and subtropical regions including Guangdong Province.
So far, 13 Canna species have been identified from Guangdong Province, China (http://www.nsii.org.cn, accessed on
14 March 2022). There are 93 fungal species described from Canna hosts around the world of which 27 were reported
from China (Farr & Rossman 2022). Among these, nine species viz. Alternaria alternata, A. bulbotrichum, Botrytis
cinerea, Colletotrichum gloeosporioides, Curvularia microspore, Fusarium fujikuroi, Puccinia thaliae, Pyricularia
cannicola and Dactylaria cannae were identified as pathogens (Farr & Rossman 2022).
Mycosphaerellaceae (Capnodiales, Dothideomycetes) species are well-known plant pathogens, saprobes,
endophytes and epiphytes, which are able to colonize in diverse niches with various life modes (Videira et al. 2017).
Mycosphaerellaceae was initially considered polyphyletic (Crous et al. 2009a, b), and later separated into several
families viz. Cladosporiaceae (Schubert et al. 2007, Dugan et al. 2008, Bensch et al. 2012, 2015), Dissoconiaceae,
Schizothyriaceae (Batzer et al. 2008) and Teratosphaeriaceae (Crous et al. 2009c, Li et al. 2012, Quaedvlieg et al.
2014 , Hongsanan et al. 2020a). Based on these facts, it is evident that the mycosphaerella-like morphology has
evolved multiple times, thus a new circumscription of Mycosphaerella is required. Hyde et al. (2013) re-circumscribed
CHEN ET AL.
54 • Phytotaxa 561 (1) © 2022 Magnolia Press
Mycosphaerellaceae and accepted 46 genera in this family. Videira et al. (2017) resurrected several old generic names
and described 32 additional genera and accepted 120 genera in Mycosphaerellaceae based on phylogenetic analyses.
Hongsanan et al. (2020b) accepted that 112 genera are based on molecular while considering the remaining 107 genera
mentioned to be reevaluated. Wijayawardene et al. (2022) followed Abdollahzadeh et al. (2020) and Hongsanan et al.
(2020b) and treated Mycosphaerellaceae under Mycosphaerellales and accepted 119 genera with molecular data.
Madagascaromyces is one of the pathogenic genera in Mycosphaerellaceae, which can cause severe defoliation
and die-black of Eucalyptus spp. (Crous et al. 2009d). Madagascaromyces was described by Videira et al. (2017)
to accommodate Madagascaromyces intermedius (= Passalora intermedia) which was isolated as an endophyte on
Eucalyptus calmadulensis from Madagascar.
In the present study, a saprobic Madagascaromyces species was isolated from Canna indica from Guangdong
Province China. The isolated taxon was identified as a novel taxon; Madagascaromyces cannae based on multi-gene
phylogeny and morphology. Herein complete species description and illustration are given for the newly identified
species.
Materials & methods
Isolation and morphological characterization
A diseased Canna indica leaf was collected from the Haizhu National Wetland Park in Guangzhou City, Guangdong
Province, China on October 4, 2021. The sample was collected and taken into the laboratory. Macro morphological
characters of the fungi were observed using a Cnoptec SZ650 (Chongqing Optec Instrument Co., Ltd., Chongqing,
China) series stereo microscope, and photographs were taken using the Nikon Eclipse 80i and the industrial DigitaL
Sight DS-Fi1 (Panasonic, Osaka, Japan) microscope imaging system. Digital images of micromorphological structures
were recorded with a Nikon Eclipse 80i microscope (Nikon, Tokyo, Japan).
Single spore isolation was done to obtain pure cultures (Senanayake et al. 2020). Germinated conidia were each
transferred to potato dextrose agar (PDA) plates using a sterile needle. The pure cultures were incubated at 25 °C with
natural light/dark. Colonies were sub-cultured in 2 % potato dextrose agar (PDA), malt extract agar (MEA) and oatmeal
agar (OA), and incubated under continuous 12h light/dark at 25 °C to promote sporulation (Crous et al. 2009d). All
pure cultures obtained in this study were deposited in the culture collection of Zhongkai University of Agriculture and
Engineering (ZHKUCC). Herbarium material (as dry cultures) is deposited in the herbaria of Zhongkai University of
Agriculture and Engineering (ZHKU).
DNA extraction, PCR amplification and sequencing
Genomic DNA was extracted from fresh mycelium grown on PDA for five days using the DNA rapid Extraction Kit
(Aidlab Biotechnologies Co., Ltd, Beijing, China) following the manufacturers’ protocols. The internal transcribed
spacer regions and intervening 5.8S nrRNA gene (ITS) of the nrDNA operon was amplified using ITS4 (White et al.
1990) and ITS-V9G (de Hoog & Gerrits van den Ende 1998) primer pair. The 28S nrRNA gene (LSU) region was
amplified with LSU1Fd (Crous et al. 2009e) and LR5 (Vilgalys & Hester 1990) and the RNA polymerase II second
largest subunit (rpb2) region was amplified using 5F2 (Sung et al. 2007) and 7CR (White et al. 1990). The PCR
reaction mixture contained 25 μL of total volume, which consisted of 12.5 μL 2 × FastTaq Premix (mixture of FastTaq
TM DNA Polymerase, buffer, dNTP Mixture, and stabilizer) (Vazyme Biotech Co., Ltd., Nanjing, China), 1 μL of each
forward and reverse primers, 9.5 μL ddH2O and 1 μL DNA. PCR amplification was performed in BIO-RAD (Hangzhou
Bio-Gener Technology Co., Ltd., Hangzhou, China) thermal cycler. Thermal cycler conditions for each primer pair are
given in TABLE 1. PCR amplification conditions for LSU and ITS followed the methods of Videira et al. (2017) and
PCR amplification conditions for rpb2 in this study was showed in TABLE 1. The PCR products were checked on 1%
agarose electrophoresis gels stained with ethidium bromide under UV light using a Gel DocTM XR Molecular Imager
(Bio-Rad, USA). The unpurified PCR products were sequenced by Sangon Biotech Co., Ltd, China.
MADAGASCAROMYCES CANNAE SP. NOV. Phytotaxa 561 (1) © 2022 Magnolia Press • 55
TABLE 1. Gene regions, primer pairs and respective thermal cycler conditions used in this study.
Locus Primer Amplification Reference
LSU 1Fd Initial denaturation at 94 ˚C for 3 min, followed by 35 cycles consisting of
denaturation at 94 ˚C for 30 s, annealing at 52 ˚C for 30 s and extension at 72
˚C for 45 s, and a final extension at 72 ˚C for 5 min
Crous et al. (2009e)
LR5 Vilgalys & Hester (1990)
ITS V9G Initial denaturation at 94 ˚C for 3 min, followed by 35 cycles consisting of
denaturation at 94 ˚C for 30 s, annealing at 52 ˚C for 30 s and extension at 72
˚C for 45 s, and a final extension at 72 ˚C for 5 min
de Hoog & Gerrits van den
Ende (1998)
ITS4 White et al. (1990)
rpb25F2 Initial denaturation at 95 ˚C for 5 min, followed by 35 cycles consisting of
denaturation at 94 ˚C for 1 min, annealing at 53 ˚C for 30 s and extension at 72
˚C for 1:30 min, and a final extension at 72 ˚C for 10 min
Sung et al. (2007)
7CR Liu et al. (1999)
Phylogenetic analysis
The sequence quality was assured by checking chromatograms of resulted sequences using BioEdit v.7.0.9.1 (Hall
1999). Geneious v.9.1.2 (Biomatters 2005) was used to combine the sequences generated by the forward and reverse
primers. Sequences obtained in this study were analyzed using the National Center for Biotechnology Information
(NCBI) search engine BLASTn (https://blast.ncbi.nlm.nih.gov/Blast.cgi) for initial species confirmation. Based on
blast results, reference sequences (TABLE 2) were obtained from GenBank following Videira et al. (2017) and Crous
et al. (2021). Sequences obtained in this study were aligned with sequences downloaded from GenBank using MAFFT
v. 7 (Katoh et al. 2019). The sequences of LSU, ITS, and rpb2 were combined by BioEdit v.7.0.9.1 (Hall 1999).
Using Alignment Transformation Environment online (https://sing.ei.uvigo.es/ALTER/) files were converted to run
phylogenetic trees. Phylogenetic analyses were conducted using maximum likelihood (ML) inferred in RAxML v.
8.2.12 (Stamatakis 2014), maximum parsimony (MP) implied on PAUP v. 4.0b10 (Swofford 2003), and Bayesian
analysis on MrBayes v. 3.2.7a (Huelsenbeck & Ronqvist 2001).
Maximum parsimony analysis was performed in PAUP (phylogenetic analysis using parsimony) v.4.0b10
(Swofford 2003) using the heuristic search option with tree bisection-reconnection (TBR) branch swapping and 1000
random sequence additions. Ambiguous regions in the alignment were excluded, and gaps were treated as missing
data. The stability of the trees was evaluated by 1000 bootstrap replications. Branches of zero length were collapsed
and all multiple parsimonious trees were saved. Descriptive statistics including tree length (TL), consistency index
(CI), retention index (RI), relative consistency index (RC), and homoplasy index (HI) were calculated.
The best evolution model was determined by MrModeltest v. 2.2 for each gene. Maximum likelihood analyses
were accomplished using RAxML-HPC v.8 on XSEDE v. 8.2.12 (Stamatakis et al. 2008 & Stamatakis 2014) in
the CIPRES Science Gateway platform (Miller 2010) using the GTRGAMMA model of evolution with 1000 non-
parametric bootstrapping iterations. MrBayes v. 3.2.7a was (Huelsenbeck & Ronqvist 2001) used for the Bayesian
analyses. The Markov Chain Monte Carlo sampling (BMCMC) analysis was conducted with four simultaneous Markov
chains. They were run for 2,000,000 generations: sampling the trees at every 100th generation. From the 20,000 trees
obtained, the first 5,000 representing the burn-in phase were discarded. The remaining 15,000 trees were used for
calculating posterior probabilities in the majority rule consensus tree. Species delineations were based on Chethana
et al. (2021), Manawasinghe et al. (2021) and Pem et al. (2021). Taxonomic novelties were submitted to the Faces
of Fungi database (Jayasiri et al. 2015) and Index Fungorum (http://www.indexfungorum.org). All sequences derived
from this study are deposited in GenBank (TABLE 2).
CHEN ET AL.
56 • Phytotaxa 561 (1) © 2022 Magnolia Press
TABLE 2. GenBank accession numbers of the sequences used in phylogenetic analyses. The newly generated sequences are
indicated in bold.
Species Culture collection
number1,2
GenBank accession number
LSU ITS rpb2
Annellosympodiella juniperi CBS 137992TKJ869204 KJ869147 MF951436
Australosphaerella nootherensis CBS 130522TKF901835 MF951293 MF951440
Brunneosphaerella jonkershoekensis CPC 13902ET JN712503 JN712439 MF951441
B. nitidae CBS 130595TGU214396 GU214625 MF951442
B. protearum CBS 130597ET GU214397 GU214626 MF951443
Chuppomyces handelii CBS 113302 GU214437 EU167581 MF951475
Epicoleosporium ramularioides CBS 141103TGU214688 GU214688 KX288433
CPC 10673 MF951160 KX287289 KX288434
Exosporium livistonae CBS 131313TJQ044446 JQ044427 MF951494
E. livistonicola MUCC 190 MF951161 MF951315 MF951495
Hyalinozasmidium aerohyalinosporum CBS 125011TKF901930 GQ852839 MF951504
Hyalinozasmidium sideroxyli CBS 142191TMF951169 MF951323 MF951505
Madagascaromyces intermedius CBS 124154TFJ790297 FJ790267 MF951511
M. intermedius CPC 15719 MF951170 FJ790251 MF951512
M. cannae ZHKUCC 220038TON188677 ON188673 ON204605
M. cannae ZHKUCC 220039 ON188675 ON188676 ON204606
M. cannae ZHKUCC 220040 ON188678 ON188675 ON204607
Microcyclosporella mali CBS 126136TGU570547 GU570535 KX288436
M. mali CBS 126132 MF951171 MF951324 MF951513
Mycodiella sumatrensis CBS 118501 JX901872 DQ303049 MF951525
Mycosphaerelloides madeirae CBS 112895TKF902017 AY725553 KX348057
M. madeirae CBS 116066 KX286989 AY853188 KX288444
Neoceratosperma cyatheae CPC 18580 KT037580 KT037539 MF951530
N. eucalypti CBS 137998TKJ869210 KJ869153 MF951531
N. haldinae CBS 142190TMF951184 MF951328 MF951533
N. legnophoricola CBS 142189TMF951183 MF951327 MF951532
N. yunnanensis CBS 119975TKF901962 KF901628 MF951534
Neomycosphaerella pseudopentameridis CBS 136407TKF777226 KF777173 MF951545
Neomycosphaerella guibourtiae CPC 39348TMW883821 MW883429 N/A3
Neopenidiella nectandrae CBS 734.87TKF901982 MF951335 MF951546
Nothopericoniella persea-macranthae CBS 122097 GU452682 MF951354 MF951583
N. persea-macranthae CBS 122282 GU452681 MF951355 MF951584
Pachyramichloridium pini CBS 461.82TEU041859 EU041802 MF951552
Paramycosphaerella brachystegiae CBS 136436TKF777230 KF777178 MF951567
P. intermedia CBS 114356TKF902026 KF901681 MF951569
P. marksii CBS 110750 KF902056 DQ267596 MF951573
Paramycosphaerella sp. A CBS 118825 MF951204 MF951347 MF951574
Paramycosphaerella sp. A CBS 118849 MF951205 MF951348 MF951575
Paramycosphaerella sp. B CBS 118968 MF951206 MF951349 MF951576
Paramycosphaerella sp. B CBS 125300 MF951207 MF951350 MF951577
Paramycosphaerella wachendorfiae CBS 129579TJF951163 JF951143 MF951578
Polyphialoseptoria tabebuiae-serratifolia CBS 112650TKF251716 KF251213 MF951613
P. terminaliae CBS 135106TKF251717 KF251214 MF951615
CBS 135475 KF251718 KF251215 MF951614
Pseudopericoniella levispora CBS 873.73TEU041837 EU041780 MF951633
Pseudopericoniella sp. CBS 330.51 GU214413 GU214632 MF951634
Pseudozasmidium eucalypti CBS 121101TKF901931 KF901606 MF951637
P. vietnamense CBS 119974TJF700944 DQ632675 MF951639
Ruptoseptoria unedonis CBS 755.70 KF251732 KF251229 MF951659
Virosphaerella irregularis CBS 123242TKF902126 KF901769 MF951685
V. pseudomarksii CBS 123241TKF902127 KF901770 MF951686
Xenomycosphaerella elongata CBS 120735TJF700942 EF394833 MF951687
Xenosonderhenia eucalypti CBS 138858TKP004485 KP004457 MF951688
Xenosonderhenioides indonesiana CBS 142239TMF951261 MF951396 MF951689
Zasmidium arcuatum CBS 113477TEU041836 EU041779 MF951692
...continued on the next page
MADAGASCAROMYCES CANNAE SP. NOV. Phytotaxa 561 (1) © 2022 Magnolia Press • 57
TABLE 2. (Continued)
Species Culture collection
number1,2
GenBank accession number
LSU ITS rpb2
Z. cellare CBS 892.85 MF951262 MF951397 KT356875
Z. citri-griseum CPC 15291 KF902152 KF901793 MF951696
Z. daviesiae CBS 116002 FJ839669 FJ839633 MF951698
Z. eucalyptorum CBS 118500TMF951266 KF901652 MF951702
Z. grevilleae CBS 124107TFJ839670 FJ839634 MF951705
Z. gupoyu CBS 122099 MF951267 MF951401 MF951706
Z. iteae CBS 113094TMF951271 MF951405 MF951711
Z. musae CBS 121384 MF951272 EU514292 MF951713
Z. musae-banksii CBS 121710TEU041852 EU041795 MF951716
Z. musigenum CBS 190.63 EU041857 EU041800 MF951718
Z. nocoxi CBS 125009TKF251788 KF251284 MF951719
Z. proteacearµm CBS 116003 FJ839671 FJ839635 MF951721
Z. pseudoparkii CBS 110999TJF700965 DQ303023 MF951723
Z. pseudotsugae rapssd EF114704 EF114687 N/A3
Z. pseudovespa CBS 121159TKF901836 MF951407 MF951724
Z. scaevolicola CBS 127009TKF251789 KF251285 MF951726
Z. tsugae ratstk EF114705 EF114688 N/A3
Z. velutinum CBS 101948ET EU041838 EU041781 MF951731
Dothiora ceratoniae CBS 477.69TKF251655 KF251151 MF951419
1CBS: Culture collection of the Westerdijk Fungal Biodiversity Institute, Utrecht, The Netherlands; CPC: Culture collection of Pedro
Crous, housed at the Westerdijk Institute; MUCC (in TSU): Culture Collection, Laboratory of Plant Pathology, Mie University, Tsu, Mie
Prefecture, Japan; ZHKUCC: Zhongkai University of Agriculture and Engineering Culture Collection. 2Status of the strains: (T) ex-type,
(ET) ex-epitype. 3N/A: information is not available.
Morphological characterization
To observe macro and micro morphologies, pure cultures were cultivated on malt extract agar (MEA) and oatmeal agar
(OA) at 25 °C in natural light/dark for up to 30 days (Crous et al. 2009d). Conidiogenous structures were observed at
25 °C in natural light/dark after 2 weeks on MEA and spermatogonia formed on OA after about 4–5 weeks. Conidial
length and width were measured for at least 30 spores and processed by TaroSoft® Image Frame Work program v0.9.7
(Panasonic, Osaka, Japan). The mean values were calculated along with standard deviations (SDs). Adobe Photoshop
CC 2019 and Adobe Illustrator CC 2019 software (Adobe Systems Inc., San Jose, America) were used to develop
images.
Results
DNA phylogeny
Phylogenetic trees were generated using combined LSU (725 bp), ITS (456 bp), and rpb2 (779 bp) sequence data.
The tree topologies generated by Bayesian, ML and MP were similar and the best scoring Bayesian tree is shown in
FIGURE 1. The sequence alignment comprised 74 taxa of representative strains of Mycosphaerellaceae, including
three different isolates obtained in this study. Dothiora ceratoniae (CBS 477.69) was used as the outgroup taxon.
Maximum parsimony analysis consisted of 1,034 constant characters and 786 informative characters resulting in
parsimonious trees (CI = 0.239, RI = 0.606, RC = 0.145, HI = 0.761). The best scoring ML tree had an optimization
likelihood value of -32914.445388. The matrix had 940 distinct alignment patterns with a 3.40% proportion of gaps
and completely undetermined characters. Estimated base frequencies were as follows: A = 0.242936, C = 0.255753,
G = 0.289728, T = 0.211583; substitution rates: AC = 1.673757, AG = 3.549537, AT = 1.350349, CG = 1.065921,
CT = 7.177086, GT = 1.0; gamma distribution shape parameter α = 0.265345. Incomplete portions at the ends of the
sequences were excluded from the analysis. Three isolates from this study clustered within the Madagascaromyces
clade which forms a sister clade to Neomycosphaerella in Mycosphaerellaceae. Our isolates constituted a sister
relationship to Madagascaromyces intermedius with 100 % ML, 100 % MP, and 1.00 Bayesian posterior probabilities
(BYPP) supports (FIGURE 1).
CHEN ET AL.
58 • Phytotaxa 561 (1) © 2022 Magnolia Press
FIGURE 1. Bayesian posterior probabilities tree derived from analysis of a combined LSU, ITS, and rpb2 sequence dataset. Bootstrap
support values for Bayesian posterior probabilities (BYPP) greater than 0.95 and maximum likelihood (ML) greater than 75 % and
maximum parsimony (MP) greater than 75 % are given at the nodes. Dothiora ceratoniae (CBS 477.69) taxa was used as the outgroup.
Strain numbers are given after the species names. The new species identified are highlighted in red. Ex-type strains are in bold.
Taxonomy
Madagascaromyces cannae Y.X. Zhang, J.W. Chen & Manawas. sp. nov. (FIGURE 2)
Index Fungorµm number: IF 559623
Facesoffungi number: FoF 10814
Etymology: Epithet refers to the host genus from which the fungus was isolated
Holotype: ZHKU 220029
MADAGASCAROMYCES CANNAE SP. NOV. Phytotaxa 561 (1) © 2022 Magnolia Press • 59
FIGURE 2. Madagascaromyces cannae (ZHKU 220029 Holotype): a, b. Specimen observed. c, d. Colonies on MEA at 25 °C (b from
the bottom). e. Spermatogonium forming on OA. f. Spermatia. g, h. Conidiophores. i, j. Conidiogenous cells. k–p. Conidia. Scale bars =
10 µm.
CHEN ET AL.
60 • Phytotaxa 561 (1) © 2022 Magnolia Press
Associated with leaf spot of Canna indica. Sexual morph: not observed. Asexual morph: Conidiophores (18–)20–
40(–78) × 2–4(–5) µm (
x
= 31 × 3 µm, n = 50), 0–4 septate, pale brown. Conidiogenous cells (4–)7–15(–20) × (2–
)4–5 µm (
x
= 10 × 4 µm, n = 50), pale brown, terminal and lateral. Conidia (17–)30–50(–70) × 3–5 µm (n = 50), pale
brown, clavate, guttulate, apex subobtuse, base long obconically subtruncate, 0–3(–6) septa (5 septa not observed),
the septa are slightly constricted, some are slightly curved. Spermatogonia forming on OA, 510–770 × 540–770 µm
(
x
= 648 × 653 µm, n = 20), black dots, semi-immersed, globose. Spermatia 2.5–4.5 × 1.5–2 µm (
x
= 3 × 2 µm, n =
50), smooth, nearly cylindrical.
Cultural characteristics: Colonies on MEA reaching 27 mm diam after one month, flourishes aerial mycelium
and smooth, surface white-grey; reverse dark green. Colonies on OA reaching 25 mm diam after one month at 25 ˚C,
spreading with moderate aerial mycelium and smooth, surface Brown, reverse olivaceous-grey.
Material examined: CHINA, Guangdong Province, Guangzhou, Haizhu National Wetland Park, on fresh leaves
of Canna indica, 4 October 2021, Jieying Lin, (dried culture ZHKU 220029 holotype; ZHKU 220030, ZHKU 220031
paratype), living cultures ZHKUCC 220038, ex-type; ZHKUCC 220039, ZHKUCC 220040 ex-paratype.
Notes: Three isolates obtained in the present study clustered with Madagascaromyces species in the phylogenetic
analysis (FIGURE 1). Our isolate differs from Madagascaromyces intermedius by the size of the conidia and the
number of septa (TABLE 3). Madagascaromyces intermedius develop conidia with 1–8 septa, while our isolate
Madagascaromyces cannae has 0–6 septa (5 septa not observed), mostly 0–3 septa. Conidia sizes of our isolate (22–45
× 4 µm) are shorter and wider than Madagascaromyces intermedius [(35–)50–75(–100) × (2.5–)3 µm]. Based on a
polyphasic approach, M. intermedius is identified as a new species.
TABLE 3. Comparison of Asexual morph characters of Madagascaromyces species.
Species name Madagascaromyces cannae ZHKUCC 220038 Madagascaromyces intermedius CBS 124154
Conidiophores (18–)20–43(–78) × 2–4(–5) µm, 0–4 septa, pale
brown.
70 × 4 µm, 0–3 septa, medium brown, smooth.
Conidiogenous cell (4–)7–16(–20) × (2–)4–5 µm, pale brown, terminal
and lateral.
15–20 × 3–3.5 µm, pale to medium brown,
smooth.
Conidia (17–)30–50(–70) × 3–5 µm, 0–3(–6) septa (5 septa
not observed).
(35–)50–75(–100) × (2.5–)3 µm, 1–8 septa.
Spermatia 2.5–4.5 × 1.5–2 µm 3–5 × 1 µm
Cultural characteristic surface white grey, reverse dark green. surface isabelline, margin sepia, reverse sepia
to brown-vinaceous.
Growth rate MEA growing, 27 mm diam and OA growing, 25 mm
after one month, at 25°C.
MEA growing, 25 mm diam and OA growing,
20 mm after one month, at 25°C.
References This study Videira et al. (2017)
Discussion
In the present study, a novel fungus was isolated and described from Canna indica from the subtropical climate
of Guangdong Province China (Guangdong Map Publishing House 2021). High temperature, annual rainfall and
humidity facilitate fungal disease on economical important hosts such as ornamental plants (Adikaram 1998). Thus,
the identification and characterization of fungal species associated with these plants have ecological and economic
impacts. Several fungal species have been reported to cause severe diseases in Canna plants. Among these, Canna
rust is a devastating disease in many Canna species worldwide (Jeeva et al. 2010, Neo et al. 2010, Padamsee et
al. 2012, Talhinhas et al. 2016, Jesús et al. 2018, Reddy et al. 2020). Three Puccinia species; P. cannae, P. thaliae
and P. cannacearum have been recorded to cause Canna rust (Thurston et al. 1940, Jesús et al. 2018, Reddy et al.
2020). Three Cercospora (Mycosphaerellaceae) species; C. apii, C. cannae and Cercospora sp. have been reported on
Cannaceae (Farr & Rossaman 2022). However, there are no records of Madagascaromyces species on Canna indica
(Farr & Rossaman 2022). Therefore, the novel taxon described in this study is the first report of Madagascaromyces
species reported on Canna indica.
Mycosphaerellaceae is a family rich in opportunistic fungal pathogens (Crous et al. 2009). The asexual morph of
Madagascaromyces is characterized by conidia that are initially solitary, smooth, guttulate, subcylindrical that become
1- or multi-septate upon maturity, apex subobtuse, base long obconically subtruncate and straight to slightly curved.
Spermatogonia on OA are cylindrical with obtuse ends, smooth and hyaline (Crous et al. 2009 & Videira et al. 2017).
Based on these morphological characteristics and combined with phylogenetic analysis, only Madagascaromyces
MADAGASCAROMYCES CANNAE SP. NOV. Phytotaxa 561 (1) © 2022 Magnolia Press • 61
intermedius is accepted in Madagascaromyces (Crous et al. 2009, Videira et al. 2017, Wijayawardene et al. 2020,
Wijayawardene et al. 2022). Thus, our novel collection will be an addition to this genus.
Our isolate Madagascaromyces cannae is distinguished from Madagascaromyces intermedius by its shorter
and wider conidia and the number of septa. While our samples were associated with disease leaves of Canna
indica, Madagascaromyces intermedius was introduced as an endophytic species of Eucalyptus camaldulensis
leaf from Madagascar (Crous et al. 2009). These two species represent different ecological niches and lifestyles of
Mycosphaerellaceae. However, the pathogenicity of Madagascaromyces cannae is unknown, thus further studies are
necessary to understand the pathogenicity of this novel taxa.
Acknowledgements
We would like to thank Dr Shaun Pennycook, Nomenclature Editor, Mycotaxon, for his guidance on the species
name. This study was gratefully acknowledged the financial support received from the Key Realm R&D Program
of Guangdong Province (grant no. 2018B020205003), the Modern Agricultural Industry Technology System Flower
Innovation Team of Guangdong Province (grant no: 2021KJ121 & 2022KJ121) and the Project of Educational
Commission of Guangdong Province of China (grant no: 2021KTSCX045). Dhanushka Wanasinghe would like
to thank CAS President’s International Fellowship Initiative (grant number 2021FYB0005), the National Science
Foundation of China (NSFC) under the project code 32150410362 and the Postdoctoral Fund from Human Resources
and Social Security Bureau of Yunnan Province.
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