Building the first SSU database of phagotrophic euglenidsusing single-cell approaches

SSU PhylogenyRAxML GTR gamma1000 bootstraps957 sites

Simpson Lab website:

protists.biology.dal.ca

Taxon sampling of phagotrophic euglenids is extremely limited for all molecular markers including the SSU gene, mainly because there are very few cultures available (see Table 1).

The culture-independent approach demonstrated here enables collection of micrographic data (for identification) and SSU genes from a single cell, using either multiple-displacement amplification (MDA) and PCR, partial shotgun-genomics, or single-cell transcriptomics.

The wide range of taxa successfully sampled with these methods shows that they can rapidly produce much-improved (and much-needed) coverage of phagotrophic euglenid diversity.

Fig. 1: Examples of SSU-typed phagotrophic euglenid cells. Top row, left to right: Anisonema acinus (SDB4), Chasmostoma nieuportense (UB23), Urceolus sp. (UB25), Petalomonas planus (UB22). Bottom row, left to right: Ploeotia cf. vitrea (U4), Neometanema sp. (SDB8), Dinema sp. (UB26), Heteronema globuliferum (BP3). Colors and symbols correspond to Table 1. Scale bars are 20µm.

MethodsPHOTOTROPHS

OSMOTROPHS

SYMBIONTIDS

OUTGROUP

Table 1: Genera of phagotrophic euglenids ordered by availability of SSU data (outside this study). 2 = sequences from 2 species only, 1 = 1 species only, 0 = no sequences available. New data obtained in this study shown in colored symbols (correspond to Fig. 1).

sedimentsample

extraction /analysisof SSU

pick single cells

photo-documentation

transcriptome

shotgungenome

seq.

SSU-PCR(via MDA)

Gordon Lax & Alastair Simpson Dalhousie University

PetalomonasNotosolenusAnisonemaPeranemaPloeotiaHeteronemaNeometanemaDinemaKeelungiaEntosiphonUrceolusJenningsiaSphenomonasLentomonasChasmostomaCalycimonasScytomonasDoliumDylakosoma

2

1

0

# SSU via PCR

# shotgun genome data

# transcriptomes

28 new SSU sequences+1 Entosiphon sulcatum(not shown)

0.1

Anisonema sp. W1

Lepocinclis ovum

Ploeotia costata (brackish)

Neobodo saliens

Anisonema sp. SDB1

Bihospites bacati isolate 2

Parmidium circulare

Trypanoplasma borreli

Ploeotia corrugata UB21

Dinema platysomum U5

Notosolenus ostium U1

Neometanema sp. BP5Neometanema exaratum U2

Anisonema sp. SMS2

Dinema litorale UBJ2

Peranema trichophorum CCAP1260/1B

Petalomonas sphagnophila GU477296

Peranema trichophorum PtR

Cryptobia helicis

Ploeotia cf. vitrea U4

Cryptoglena pigraMonomorphina pyrum

Keelungia pulex

Heteronema scaphurum

Peranema trichophorum

Ploeotia costata AF525486

Trachelomonas abrupta

Diplonema papillatum

Petalomonas cantuscygni

Colacium vesiculosum

Anisonema acinus B1

Heteronema globuliferum BP3

Environmental sequence AY821956

Petalomonas cantuscygni CCAP 1259/1

Eutreptia viridis

Urceolus sp. UB25

Bodo saltans

Rapaza viridis

Anisonema sp. SDB4

Distigma gracilis

Euglena viridis

Gyropaigne lefevrei

Dimastigella trypaniformis

Notosolenus urceolatus M49

Petalomonas planus UB22

Rhynchobodo sp.

Phacus parvulus

Anisonema sp. G1

Rhynchomonas nasuta

Dinema sulcatum

Calkinsia aureus

Notosolenus ostium

Peranema sp. AY048919

Environmental sequence GQ330643

Environmental sequence EF100248

Environmental sequence HM749952

Urceolus sp. UB24

Anisonema sp. PP1

Environmental sequence AY821958

Anisonema/Dinema sp. U3

Dinema/Heteronema sp. CD5

Neometanema sp. BP6

Neometanema sp. SDB8

Distigma curvatum

Petalomonas sphagnophila GU477297

Ichthyobodo necator

Neometanema larsenii SDB3

Neometanema parovale M51

Dinema sp. UB26

Environmental sequence AY821957

Petalomonas sphagnophila GU477295

Environmental sequence AB505539

Anisonema acinus

Anisonema sp. SAL5

Dinema litorale UB10

Menoidium pellucidum

Environmental sequence DQ310255

Heteronema sp. PP2

Rhabdomonas incurva

Ploeotia costata (marine)

Bihospites bacati isolate 1

97

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100

100

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100

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57

100

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95

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100

78

91

67

100

100

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60

100

100

100

50

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100

80

100

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100

100

100

89

52

93

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97

59

86

100

92

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10095

98