The algal flora of subaerial habitats in the tropics remains largely unexplored, despite the fact that it potentially encompasses a wealth of new evolutionary diversity. Here we present a detailed morphological and molecular characterization of an autosporic coccoid green alga isolated from decaying wood in a natural forest in Singapore. Depending on culture conditions, this alga formed globular to irregularly oval solitary cells. Autosporulation was the only mode of reproduction observed. The cell periphery was filled with numerous vacuoles, and a single parietal chloroplast contained a conspicuous pyrenoid surrounded by a bipartite starch envelope. The cell wall was composed of a thick inner layer and a thin trilaminar outer layer, and the cell surface was ornamented with a few delicate ribs. Phylogenetic analyses of 18S rRNA gene sequences placed our strain in the family Scenedesmaceae (Sphaeropleales, Chlorophyceae) as a strongly supported sister branch of the genus Desmodesmus. Analyses of an alternative phylogenetic marker widely used for the Scenedesmaceae, the ITS2 region, confirmed that the strain is distinct from any scenedesmacean alga sequenced to date, but is related to the genus Desmodesmus, despite lacking the defining phenotypic features of Desmodesmus (cell wall with four sporopolleninic layers ornamented with peculiar submicroscopic structures). Collectively, our results establish that we identified a novel, previously undocumented, evolutionary lineage of scenedesmacean algae necessitating its description as a new species in a new genus. We propose it be named Hylodesmus singaporensis gen. et sp. nov. A cryopreserved holotype specimen has been deposited into the Culture Collection of Algae of Charles University in Prague, Czech Republic (CAUP) as CAUP C-H8001.
AnS. S.,
FriedlT.,
HegewaldE.1999; Phylogenetic relationships of Scenedesmus and Scenedesmus -like coccoid green algae as inferred from IST-2 rDNA sequence comparisons. Plant Biol 1:418–428[CrossRef]
EliášM.,
NeustupaJ.,
ŠkaloudP.2008; Elliptochloris bilobata var. corticola var. nov. (Trebouxiophyceae, Chlorophyta), a novel subaerial coccal green alga. Biologia 63:791–798
FawleyM. W.,
FawleyK. P.,
OwenH. A.2005; Diversity and ecology of small coccoid green algae from Lake Itasca, Minnesota, USA, including Meyerella planktonica , gen. et sp. nov. Phycologia 44:35–48[CrossRef]
FelsensteinJ.2004phylip (Phylogeny Inference Package) version 3.6. Distributed by the author. Department of Genome Sciences University of Washington; Seattle: 2004http://evolution.genetics.washington.edu/phylip.html
FottB.,
NovákováM.1969; A monograph of the genus Chlorella . The fresh water species. In Studies in Phycology pp 10–74 Edited by
FottB.
Praha: Academia;
FriedlT.,
BesendahlA.,
PfeifferP.,
BhattacharyaD.2000; The distribution of group I introns in lichen algae suggests that lichenization facilitates intron lateral transfer. Mol Phylogenet Evol 14:342–352[CrossRef]
FritzL.,
TriemerR. E.1985; A rapid simple technique utilizing calcofluor white M2R for the visualization of dinoflagellate thecal plates. J Phycol 21:662–664
HanagataN.1998; Phylogeny of the subfamily Scotiellocystoideae (Chlorophyceae, Chlorophyta) and related taxa inferred from 18S ribosomal RNA gene sequence data. J Phycol 34:1049–1054[CrossRef]
HegewaldE.,
WolfM.2003; Phylogenetic relationships of Scenedesmus and Acutodesmus (Chlorophyta, Chlorophyceae) as inferred from 18S rDNA and IST-2 sequence comparisons. Plant Syst Evol 241:185–191[CrossRef]
HegewaldE.,
CoeselP. F. M.,
HegewaldP.2002; A phytoplankton collection from Bali, with the description of a new Desmodesmus species (Chlorophyta, Scenedesmaceae). Arch Hydrobiol 143:51–78
HegewaldE.,
SchmidtA.,
BrabandA.,
TsareknoP.2005; Revision of the Desmodesmus (Sphaeropleales, Scenedesmaceae) species with lateral spines. 2. The multi-spined to spineless taxa. Algol Stud 116:1–38[CrossRef]
HelmsG.,
FriedlT.,
RamboldG.,
MayrhoferH.2001; Identification of photobionts from the lichen family Physciaceae using algal-specific ITS rDNA sequencing. Lichenologist 33:73–86[CrossRef]
HussV. A.,
SoginM. L.1990; Phylogenetic position of some Chlorella species within the chlorococcales based upon complete small-subunit ribosomal RNA sequences. J Mol Evol 31:432–442[CrossRef]
HussV. A. R.,
FrankC.,
HartmannE. C.,
HirmerM.,
KloboucekA.,
SeidelB. M.,
WenzelerP.,
KesslerE.1999; Biochemical taxonomy and molecular phylogeny of the genus Chlorella sensu lato (Chlorophyta). J Phycol 35:587–598[CrossRef]
JeonS. L.,
HegewaldE.2006; A revision of the species Desmodesmus perforatus and D. tropicus (Scenedesmaceae, Chlorophyceae, Chlorophyta. Phycologia 45:567–584[CrossRef]
JohnsonJ. L.,
FawleyM. W.,
FawleyK. P.2007; The diversity of Scenedesmus and Desmodesmus (Chlorophyceae) in Itasca State Park. Minnesota, USA: J Phycol 46:214–229[CrossRef]
KatanaA.,
KwiatowskiJ.,
SpalikK.,
ZakrysB.,
SzalachaE.,
SzymanskaH.2001; Phylogenetic position of Koliella (Chlorophyta) as inferred from nuclear and chloroplast small subunit rDNA. J Phycol 37:443–451[CrossRef]
KomárekJ.,
FottB.1983; Familie Scenedesmaceae. In Das Phytoplankton des Süßwassers7 – Chlorococcales pp 747–945 Edited by
Huber-PestalozziG.
Stuttgart: Schweizerbart;
KrienitzL.,
HegewaldE.,
HepperleD.,
WolfM.2003; The systematics of coccoid green algae: 18S rRNA gene sequence data versus morphology. Biologia (Bratisl 58:437–446
LewisL. A.,
FlechtnerV. R.2004; Cryptic species of Scenedesmus (Chlorophyta) from desert soil communities of western North America. J Phycol 40:1127–1137[CrossRef]
MathewsD. H.,
DisneyM. D.,
ChildsJ. L.,
SchroederS. J.,
ZukerM.,
TurnerD. H.2004; Incorporating chemical modification constraints into a dynamic programming algorithm for prediction of RNA secondary structure. Proc Natl Acad Sci U S A 101:7287–7292[CrossRef]
MattoxK. R.,
StewardK. D.1984; Classification of the green algae: a concept based on comparative cytology. In Systematics of the Green Algae pp 265–292 Edited by
IrvineD. E. G.,
JohnD. M.
London: Academic Press;
NeustupaJ.,
EliášM.,
ŠejnohováL.2007; A taxonomic study of two Stichococcus species (Trebouxiophyceae, Chlorophyta) with a starch-enveloped pyrenoid. Nova Hedwigia 84:51–63[CrossRef]
NeustupaJ.,
NěmcováY.,
EliášM.,
ŠkaloudP.2009; Kalinella bambusicola gen. et sp. nov. (Trebouxiophyceae, Chlorophyta), a novel coccoid Chlorella -like subaerial alga from Southeast Asia. Phycol Res 57:159–169[CrossRef]
PoulíčkováA.,
ŽižkaZ.,
HašlerP.,
BenadaO.2007; Zygnematalean zygospores: morphological features and use in species identification. Folia Microbiol (Praha 52:135–145[CrossRef]
RindiF.,
López-BautistaJ. M.,
SherwoodA. R.,
GuiryM. D.2006; Morphology and phylogenetic position of Spongiochrysis hawaiiensis gen. et sp. nov., the first known terrestrial member of the order Cladophorales (Ulvophyceae, Chlorophyta). Int J Syst Evol Microbiol 56:913–922[CrossRef]
StamatakisA.2006; RAxML-VI-HPC: maximum likelihood-based phylogenetic analyses with thousands of taxa and mixed models. Bioinformatics 22:2688–2690[CrossRef]
ThompsonJ. D.,
GibsonT. J.,
PlewniakF.,
JeanmouginF.,
HigginsD. G.1997; The clustal_x windows interface: flexible strategies for multiple sequence alignment aided by quality analysis tools. Nucleic Acids Res 25:4876–4882[CrossRef]
TsarenkoP. M.,
HegewaldE.,
BrabandA.2005; Scenedesmus -like algae of Ukraine. 1. Diversity of taxa from water bodies in Volyn Polissia. Algol Stud 118:1–45[CrossRef]
van HannenE. J.,
FinkP.,
LürlingM.2002; A revised secondary structure model for the internal transcribed spacer 2 of the green algae Scenedesmus and Desmodesmus and its implication for the phylogeny of these algae. Eur J Phycol 37:203–208[CrossRef]
VanormelingenP.,
HegewaldE.,
BrabandA.,
KitschkeM.,
FriedlT.,
SabbeK.,
VyvermanW.2007; The systematics of small spineless Desmodesmus species, D. costato-granulatus (Sphaeropleales, Chlorophyceae), based on ITS2 rDNA sequence analyses and cell wall morphology. J Phycol 43:378–396[CrossRef]
WhiteT. J.,
BrunsT.,
LeeS.,
TaylorJ. W.1990; Amplification and direct sequencing of fungal ribosomal RNA genes for phylogenetics. In PCR Protocols: A Guide to Methods and Applications pp 315–322 Edited by
InnisM. A.,
GelfandD. H.,
SninskyJ. J.,
WhiteT. J.
New York: Academic Press;
WolfM.,
HegewaldE.,
HepperleD.,
KrienitzL.2003a; Phylogenetic position of the Golenkiniaceae (Chlorophyta) as inferred from 18S rDNA sequence data. Biologia (Bratisl 58:433–436
WuytsJ.,
De RijkP.,
Van de PeerY.,
PisonG.,
RousseeuwP.,
De WachterR.2000; Comparative analysis of more than 3000 sequences reveals the existence of two pseudoknots in area V4 of eukaryotic small subunit ribosomal RNA. Nucleic Acids Res 28:4698–4708[CrossRef]
ZhangJ. M.,
HussV. A. R.,
SunX. P.,
ChangK. J.,
PangD. B.2008; Morphology and phylogenetic position of a trebouxiophycean green alga (Chlorophyta) growing on the rubber tree, Hevea brasiliensis , with the description of a new genus and species. Eur J Phycol 43:185–193[CrossRef]