CSIRO PUBLISHING

© CSIRO 10 November 2003 10.1071/SB03012 1030-1887/03/050581

www.publish.csiro.au/journals/asb Australian Systematic Botany 16, 581–594

A molecular estimate of the phylogeny of Styphelieae (Ericaceae)

Christopher J. QuinnA,C, Darren M. CraynA, Margaret M. HeslewoodA, Elizabeth A. BrownA and Paul A. GadekB

ANational Herbarium of New South Wales, Royal Botanic Gardens and Domain Trust, Mrs Macquaries Road, Sydney, NSW 2000, Australia.

BSchool of Tropical Biology, James Cook University, PO Box 6811, Cairns, Qld 4870, Australia.C Corresponding author; email: chris.quinn@rbgsyd.nsw.gov.au

Abstract. Parsimony analyses of sequence data derived from two regions of the chloroplast genome, matK and theatpΒ−rbcL intergenic spacer, separately and together, are reported for Styphelieae. Taxonomic sampling includesrepresentatives of all currently accepted genera apart from the two non-Australian monotypics, Cyathopsis Brongn.& Gris and Decatoca F.Muell., and of all subgenera or informal infrageneric groups except for Trochocarpasubgenus Pseudocyathodes Sleumer. A well resolved estimate of the phylogeny of the tribe is obtained, with highlevels of jackknife support for terminal groupings. The results provide support for the current concepts ofAcrotriche R.Br., Androstoma Hook.f., Croninia J.M.Powell, Cyathodes Labill. sensu Weiller, LeptecophyllaC.M.Weiller, Leucopogon R.Br. sensu Taaffe et al., Pentachondra R.Br., Planocarpa C.M.Weiller and TrochocarpaR.Br., but the following genera appear paraphyletic: Astroloma R.Br., Brachyloma Sond., Cyathodes sensu Labill.,Leucopogon R.Br. sensu lato, Lissanthe R.Br., Monotoca R.Br. and Styphelia Sm. (sensu Bentham). Several robustclades are identified as potential new genera but formalisation of nomenclatural changes is left, pendingmorphological analyses of the various clades to identify synapomorphies with which to characterise genericgroupings. The general approach to be adopted in this endeavour is discussed, with narrower generic concepts beingfavoured as more informative and less disruptive of the existing nomenclature.SB03012Phyl ogeny of Stypheli eaeC. J. Qui nn et al .

IntroductionThe Styphelieae is the largest of the seven currentlyrecognised tribes of epacrids (subfamily Styphelioideae,Ericaceae; Kron et al. 2002). It comprises species withindehiscent fruits, with the exception of the WesternAustralian monotypic endemics Needhamiella L.Watson andOligarrhena R.Br., which constitute tribe Oligarrheneae.Styphelieae are woody plants that range from prostrate,mat-forming plants of montane herbfields [e.g.Pentachondra pumila (J.R.Forst. & G.Forst. ) R.Br.] to smallrainforest trees (e.g. Trochocarpa laurina R.Br.). Althoughpredominantly Australian in distribution, it is the mostwidely dispersed of the epacrid tribes, with representatives inNew Zealand, the Pacific east to Hawaii, Malesia andSouth-East Asia as far north as Myanmar. The Styphelieaecomprises ~350 species in 19 genera, the largest beingLeucopogon R.Br. sensu lato (sens. lat.; approximately 230spp.), Astroloma R.Br. (18 spp.), Styphelia Sm. (14 spp.) andAcrotriche R.Br. (14 spp.) (Powell 1992; Kron et al. 2002).

The first critical treatment of the Australian epacrids(Order Epacrideae) was provided by Brown (1810). He

recognised 134 species in 24 genera and establishedinfrafamilial groups based on fruit type. Section Onecomprised 96 species in 14 genera with fleshy fruits; SectionTwo comprised 38 species in 9 genera with capsular fruits.

Bentham (1869) recognised 15 genera of Styphelieae(which corresponds to Brown’s Section One) in Australia.Broader generic concepts have been proposed from time totime (e.g. Sprengel 1825; Drude 1889). Most noteworthy arethose of Mueller (1867, 1889), who expanded Styphelia toinclude Acrotriche, Astroloma, Cyathodes Labill.,Cyathopsis Brongn. & Gris, Leucopogon R.Br., LissantheR.Br., Melichrus R.Br., Monotoca R.Br. and PentachondraR.Br. However, few subsequent authors [Sleumer (1964)notwithstanding] took up his broad concepts, with Bentham(1869) citing reasons of floral diversity and generalacceptance for retaining the narrower ones established byBrown (1810).

Bentham’s treatment remained essentially unmodified,apart from the addition of species, until recently, whenPowell (1993) described the monotypic genus Croninia forLeucopogon kingianus (F.Muell.) C.A.Gardner, and Weiller

582 Australian Systematic Botany C. J. Quinn et al.

(1996a) revised Cyathodes, describing two new segregategenera (Leptecophylla, Weiller 1999; Planocarpa, Weiller1996b) and reinstating another (Androstoma Hook.f.)(Weiller 1996c). This has brought the total number ofaccepted genera to 19 (Kron et al. 2002).

Powell and co-workers compiled a morphologicaldatabase for Styphelieae and used cladistic methods to testthe generic concepts (Powell et al. 1987, 1997). Theyconcluded that Leucopogon, inter alia, was polyphyletic, andproposed the new generic names ‘Axonanthus’ and‘Gynoconus’ for clades of Leucopogon sens. lat. Recentwork based on sequence data from the atpB–rbcL intergenicspacer (Crayn and Quinn 2000; Taaffe et al. 2001) does notsupport either of these segregates as circumscribed by Powell(1992). Furthermore, in-depth studies on Astroloma(Streiber 1999; A. Wilson unpublished data) and Monotoca(Owens 1996) have highlighted the degree to which currenttaxonomic concepts in Styphelieae fail to accurately reflectphylogenetic relationships.

As part of ongoing work (Crayn et al. 1996a, 1996b,1998; Crayn and Quinn 2000; Cherry et al. 2001; Taaffe etal. 2001) to resolve meaningful genera ahead of a Flora ofAustralia treatment of the epacrids, we present here acladistic analysis of DNA sequence data from two plastidregions: the matK gene and the atpΒ–rbcL intergenic spacer(hereafter referred to as the spacer). The database containsnear-complete generic representation of Styphelieae, lackingonly the monotypic genera Cyathopsis and DecatocaF.Muell., which are endemic to New Caledonia and PapuaNew Guinea, respectively, and for which suitable material formolecular work has yet to be obtained.

Materials and methods

The database comprised 121 taxa, 115 of which were derived fromStyphelieae, including 30 of the 33 terminal taxa employed for the tribeby Powell et al. (1997). No suitable material was available for molecularwork on either species of Trochocarpa subgenus PseudocyathodesSleumer. Outgroups comprised two members of Epacrideae, bothmembers of Oligarrheneae, and one member each from Archerieae andPrionoteae. Since the latter tribe has been demonstrated to represent thefirst diverging lineage within the Styphelioideae (Kron et al. 2002), itwas used to root the trees. Conostephium Benth. is represented bydifferent species in the spacer and matK databases. Details of vouchers,GenBank accessions for sequences and authorities for binomials aregiven in Appendix 1. Non-standard abbreviations are used for genericnames where necessary to avoid confusion; these are defined in thefigure captions.

Total DNA was extracted either from fresh or silica-dried leafmaterial, or from herbarium specimens. Leaf material was ground inliquid nitrogen and the DNA extracted either by the hot CTAB(hexadecyltrimethyl ammonium bromide) method of Doyle and Doyle(1990) or with the Plant DNeasy Minikit (Qiagen, Clifton Hill, Vic.).Double-stranded templates were amplified in a Thermocycler (PerkinElmer, Norwalk, CT) and purified with Wizard Preps PCR columns(Promega, Annandale, NSW). Sequences were obtained either bymanual sequencing using radionucleotide labelling or automatedsequencing on an ABI Prism 377 DNA Sequencer (Perkin Elmer) usingfluorescent-dye-labelled terminators. Details of primers are given in

Cherry et al. (2001). Electropherograms were processed usingSequence Navigator software (ABI, v1.0.1). Sequences were aligned byeye in MacClade (version 4.05; Maddison and Maddison 2002).Insertions/deletions (indels) were positioned so as to best conform tothe indel types of Golenberg et al. (1993). In aligning the spacer data,overlapping indels of different lengths, and insertions of the samelength but bearing different relationships to surrounding sequence,were treated as having independent origins. Potentially informativeindels were scored as additional present or absent characters and addedto the database. Sequences of the matK gene were translated inMacClade to check for internal stop codons.

The matK and spacer datasets were first analysed separately, thensimultaneously after testing for data combinability using theincongruence-length difference test (Mickevich and Farris 1981; Farriset al. 1994: implemented as the ‘partition-homogeneity test’ in PAUP*).Heuristic searches were conducted in PAUP* Version 4.0b10 (Swofford2002) using tree bisection reconnection branch-swapping to recover themost-parsimonious trees. Simple heuristic searches of the separatedatasets exhausted available memory before completion. In order tosample the tree space adequately, multiple replicates of random taxonaddition were conducted as above, saving only 100 trees per replicate,with subsequent use of the ‘condense’ option to delete duplicate trees.The analysis of the combined data went to completion, and heuristicsearching was done saving all trees and with 100 replicates of randomtaxon addition to detect multiple islands of trees. The combined datasetwas reanalysed following deletion of the indel characters in order togauge their effect on the results. Relative support for the cladesidentified by parsimony analysis was estimated using the jackknifeoption in PAUP* (10000 replicates with 33% character deletion, ‘fast’step-wise addition and emulate ‘Jac’ resampling). Constraint trees wereconstructed in MacClade and imported into PAUP, and the analysesconducted as above.

Results

Analysis of spacer data

The spacer dataset comprised 123 sequences (Appendix 1)ranging from 611 base pairs (bp) in Monotoca tamariscina to808 bp in Leucopogon lanceolatus. Missing data totalled6.2% of the sequence data. Sequences for the following taxawere substantially incomplete: Lissanthe brevistyla,Astroloma tectum, As. pallidum and Leucopogon juniperinushad 49.8%, 33.9%, 30.9% and 26.2% missing data,respectively. Alignment necessitated many indels rangingfrom 1–144 bp. The aligned database comprised 1250characters, with gaps comprising 38.2% of the data. Seventypotentially informative indels were scored and added to thedatabase. There were 568 (43%) variable characters in thetotal database, 389 (31.8%) of which were informative,indicating potential to resolve relationships within the studygroup.

Limited heuristic searching with 500 replicates of randomtaxon addition yielded 48397 trees of 1280 steps[consistency index excluding uninformative characters (CI)= 0.54; retention index (RI) = 0.81; rescaled consistencyindex (RC) = 0.49]. The strict consensus of these trees isshown in Fig. 1.

Archeria comberi is placed in a polytomy with threeprimary clades, respectively comprising Epacrideae, allStyphelieae except Monotoca tamariscina, and

Phylogeny of Styphelieae Australian Systematic Botany 583

Oligarrheneae plus Monotoca tamariscina. The first twoclades are highly robust (96% and 99% jackknife support,respectively), but the last is only weakly supported (62%).

Within the primary clade of Styphelieae numerous robustgroups are recognised but relationships between these are,for the most part, unresolved. Representatives of thefollowing genera are grouped, mostly with strong support:Acrotriche (99%), Cyathodes sensu Weiller (93%, Cysubclade), Leptecophylla (80%, Lp subclade), Leucopogonsensu stricto (sens. str.; 99%, L1), Pentachondra (100%),Planocarpa (97%) and Trochocarpa (70%). The monophylyof the remaining genera is not supported. Only one speciesof a genus currently assigned to Styphelieae (Kron et al.2002) is not included in this main Styphelieae clade:Monotoca tamariscina (group B of Powell et al. 1997) isplaced within the Oligarrheneae as sister to Oligarrhenamicrantha with 100% support.

In addition to the monophyletic genera listed above,several clades within the Styphelieae primary clade receivedsome jackknife support (see Fig. 1); these are referred to asthe Androstoma clade (81%, An), the Astroloma clade (65%,As), the Conostephium clade (66%, Co), the Cyathodesclade (53%, Cs), the Leucopogon crassifolius clade (59%,L2), the Melichrus clade (100%, Me) and the Monotocaclade (82%, Mo). Forty of the representatives of Leucopogonsens. lat. that were excluded from the genus by Taaffe et al.(2001) are placed within four of these clades, one in An, 17in As, 10 in Cs and 12 in L2. Leucopogon blepharolepis is ona separate lineage that is grouped without support withCroninia kingiana and the As and L2 clades.

Astroloma xerophyllum (group B of Powell et al. 1997) isrobustly grouped (100%) with As. stomarrhena (group C) ona lineage that is the first to diverge within the As clade. Threerepresentatives of Astroloma group A (As. epacridis, As.humifusum and As. macrocalyx), As. ciliatum (group E) andthe unassigned species, As. tectum, As. ‘recurvum’ andAstroloma sp. ‘FNQ’, are all placed within the wellsupported (84%) core of the As clade. The otherrepresentatives of the genus are placed at a distance from theAs clade: Astroloma pinifolium (group D) is stronglygrouped (86%) with As. conostephioides (group F) withinthe Co clade; As. pallidum (group A) is grouped with As.baxteri (group G), but without support. Neither is theresupport for the relationships of these two clades to any otherclade within the tribe.

The Tasmanian Cyathodes dealbata and the New ZealandCy. pumila, neither of which was treated in the recentrevision of that genus (Weiller 1996a, 1996b, 1996c, 1999),are grouped with 96% support and placed as thewell-supported (82%) sister group of Monotoca (excludingMon. tamariscina) in the Mo clade.

Three Lissanthe species (Li. brevistyla, Li. sapida and Li.strigosa) form a strongly supported group (97%) within theAc subclade of the Cs clade. The strongly supported (93%)

sister to Lissanthe is a robust clade (99%) that includes twoof the four species of Leucopogon Group E (Powell et al.1997), namely Leucopogon pluriloculatus andLe. rubicundus, plus an undescribed species from WesternAustralia (Lissanthe ‘powelliae’).

Brachyloma preissii (group A of Powell et al. 1997) issister to B. daphnoides (group B), but there is no support forthis relationship, and nor is there any evidence of a closerelationship between these and B. scortechinii (group C).

All Australian representatives of Styphelia sensu Benth.are placed within the As clade, but the genus appearsparaphyletic, since S. tenuifolia is strongly grouped (89%)with Le. cuneifolius, S. exserta is placed in a polytomy(without support), and the remaining representatives aregrouped with Le. esquamatus with only weak support (59%).The New Guinea representative of Styphelia sensu Sleumer,Styph. suaveolens, is placed with 97% support among agroup of species of Leucopogon sens. lat. in the L4 subcladeof the Cs clade.

Analysis of matK data

Sequences were obtained for 93 accessions (Appendix 1),including at least one species from each of the genera listedabove, and from each of the subclades in Fig. 1. Positions1–27 were excluded before analysis since these weredetermined in only a small proportion of taxa. Severalsequences were substantially incomplete: Leucopogonappressus (30% determined), Le. spathaceus (34%),Astroloma xerophyllum (49%), Le. pluriloculatus (63%),Cyathodes platysoma (63%), Le. cicatricatus (66%),Monotoca rotundifolia (68%), Mon. tamariscina (71%),Mon. ‘aristata’ (73%). However, missing data constitutedonly 7.7% of the total sequence data. Alignment requiredseveral indels, all of which involved from 1–8 completecodons (3–24 characters). Six of these were potentiallyinformative and were scored and added to the database. Gapsmade up 5.2% of the aligned data. The total databasecomprised 1512 characters, of which 660 (43.6%) werevariable and 363 (24%) potentially informative.

Five hundred replicates of limited heuristic searchingyielded 49983 trees of 1328 steps (CI = 0.55, RI = 0.77,RC = 0.52), the strict consensus of which is shown in Fig. 2.Relationships between the outgroups are resolved, with 75%support for Archeria being the first lineage to diverge. AStyphelieae clade excluding Monotoca tamariscina is againrobustly supported (100%), as are the other two primaryclades (Epacrideae 100%, Oligarrhena clade 95%). Withinthe Styphelieae clade, there is a high degree of congruencewith the clades recognised in Fig. 1, with the notableexception of the Cs clade. The subclades of Cs are allretrieved as separate lineages within the large basalpolytomy: Cyathodes sens. str. (Cy), Leptecophylla (Lp), andthe Acrotriche (Ac), L3 and L4 subclades. Basal relationshipswithin the Styphelieae clade, however, are unresolved.

584 Australian Systematic Botany C. J. Quinn et al.

Prionotes cerinthoides OUTGROUPArcheria comberi

Epacris impressaLysinema ciliatum Epacrideae

As. conostephioides FAs. pinifolium DConostephium preissii

Co

B. daphnoides BB. preissii A

Mel. procumbensMel. urceolatus'Pseudactinia'

Me

Pl. nitidaPl. petiolaris

T. laurinaTrochocarpa sp. A

T. gunnii

Cy. pumilaCy. dealbata

Mon. 'aristata' AMon. rotundifolia AMon. ledifolia A

Mon. elliptica CMon. scoparia A

Mo

As. baxteri GAs. pallidum A

Androstoma empetrifoliaLe. milliganii An

Le. australisLe. bossiaeaLe. alternifoliusLe. amplexicaulisLe. lanceolatusLe. microphyllus

Le. assimilisLe. glabellusLe. cordatusLe. gibbosusLe. plumuliflorus

L1

B. scortechinii CLe. blepharolepisCroninia kingianaLe. crassiflorus

Le. muticusLe. ericoidesLe. leptospermoidesLe. margarodes

Le. crassifoliusLe. corynocarpus

Le. cymbiformisLe. tamminensisLe. pogonocalyxLe. appressusLe. blakei

As. xerophyllum BAs. stomarrhena C

Col. myrtoides

As. macrocalyx A

As. tectumAs. 'recurvum'As. ciliatum E

As. epacridis AAs. humifusum A

S. tenuifoliaLe. cuneifolius

Le. cordifoliusLe. allittiiLe. propinquus

Le. strictus

Le. dielsianusLe. pendulus

Le. nutansLe. ovalifoliusLe oxycedrus

S. exsertaLe. esquamatusS. exarrhena

S. tubifloraS. viridis

Le. fletcheriLe. setiger

Le. neoanglicusLe. fraseri NSWAstroloma sp. 'FNQ'

Le. fraseri NZLe. juniperinus

As

L2

Mon. tamariscina BOligarrhena micranthaNeedhamiella pumilo Oligarrheneae

621(1) 2

4(1)

Pe. ericifoliaPe. involucrataPe. pumila

70

(1)

1

70 51

1 671

6193 1(2)

82

(1)

6667

59

91

2

58 (2)

54

651

84

88

89(1)

63(1) 90

1

6972

69

67

59

5558

76

64(1)

Sty

ph

elie

ae

Ac. divaricataAc. affinisAc. cordataAc. patula

Li. sapidaLi. strigosaLi. brevistyla

Le. pluriloculatus

Le. rubicundusLi. 'powelliae'

Ac

Le. colensoiStyph. suaveolensLe. hookeriLe. maccraeiLe. montanusLe. spathaceus

L4

Cy. glaucaCy. platysomaCy. straminea

CyLe. cicatricatus

Le. pedicellatusLe. pleiospermus L3

Lept. juniperina var. parvifolia

Lept. robustaLept. tamaeimaeie

Lept. pendulosaLept. juniperina var. junipernia NZLept. juniperina var. juniperina TASLept. divaricataLept. juniperina var. oxycedrusLept. abietina

Lp

21

67

93

2(1)2

22

1(2)

84

94

67

1(2)

8094

641

1

89

931(1)

91 7611(4)

Cs

1(1)

1

8666

81

53

Phylogeny of Styphelieae Australian Systematic Botany 585

Astroloma pallidum (group A) is grouped with othermembers of Astroloma A and E (with 64% support) withinthe robust (97%) core of the As clade, and As. xerophyllum(group B) is still sister to this, although now without support.There is 73% support for a larger clade, the Styphelia clade(Sy in Fig. 2), comprising the As and L2 clades plus Croniniaand Leucopogon blepharolepis. Conostephium pendulum isplaced in the same position of sister to Astrolomaconostephioides + As. pinifolium, within the Co clade (98%),as was Con. preissii in Fig. 1. There is moderate support(77%) for placing Brachyloma preissii (group A) withB. scortechinii (group C) in a clade labelled B1 in Fig. 2,whereas Brachyloma daphnoides (group B) is now groupedwith Astroloma baxteri with 94% support in the B2 clade,which is the unsupported sister of the Co clade.

When the distributions of indel characters were mappedon the figures, 39 of the 70 spacer indels (Fig. 1) and threeof the six indels in matK (Fig. 2) were consistent with asingle origin (though three of the former required one ormore reversals). Another four would require a single originif the topology was resolved, and three could be madeconsistent with a single origin by changing weak elements ofthe topology. Among the remaining 24 obviously homoplasicindels, two types are common: duplications of adjacentsequence (e.g. an 18-bp insertion that occurs in Leucopogongibbosus and Le. microphyllus; a 5-bp duplication that arisesin Astroloma conostephioides + As. pinifolium as well as inBrachyloma scortechinii, Le. cicatricatus and Stypheliatubiflora) account for 14 indels; another seven involve shortdeletions. Both types were recognised as common forms oflength mutation in the study by Golenberg et al. (1993).

Analysis of combined data

In view of the close similarity between the topologiesobtained from these two analyses, and the fact that acombinability test showed no significant conflict betweenthe phylogenetic signals in the two datasets (P = 0.12), acombined analysis was performed on a subset of 93 taxa forwhich both sequences had been determined. Heuristicsearching with 100 replicates of random taxon additionyielded a single island of 3266 trees of 2479 steps (CI = 0.54;RI = 0.77; RC = 0.50). The strict consensus of these is shownin Fig. 3. There is a close resemblance to the topologiesrevealed by the separate analyses, with the same major cladesbeing present (including both Cs and Sy), and an increase in

resolution of, and support for, relationships. There is, forexample, resolution of the relationships between most of theclades recognised within the Styphelieae clade, and supportfor both the As and Sy clades is now very strong (98% and100%, respectively). Additionally, Astroloma, Brachyloma,Lissanthe, Monotoca and Styphelia again appearparaphyletic, and species of Leucopogon sens. lat. are placedwithin the same set of clades (An, Cs and Sy). One of themost parsimonious trees is shown in Fig. 4 with branchlengths proportional to the amount of change.

Analysis after deletion of the indel characters yielded5048 equally parsimonious trees, but the strict consensus ofthese was identical with Fig. 3 in all major aspects. Therewas a small change to the internal topology of both the L1clade and the Lp subclade, and some minor changes (mostlyreductions) in the support values.

Discussion

Our results (Fig. 1) provide strong support for the monophylyof several long-accepted genera (viz. Acrotriche,Pentachondra, Trochocarpa), and the recent segregatesrecognised from Cyathodes and Leucopogon (viz.Androstoma, Croninia, Cyathodes sens. str., Leptecophylla,Leucopogon sens. str. and Planocarpa) are also indicated tobe well founded. The remaining generic concepts are in needof modification if they are to reflect more accurately thephylogenetic relationships indicated here.

The representatives of Monotoca groups A and Cconstitute a clade that received support in all three analyses,although Mon. elliptica, the sole member assigned to groupC by Powell et al. (1997), is robustly embedded within groupA as sister to Mon. scoparia (99% support). Hence, group Csimply represents a specialisation from within group A. Onthe other hand, Mon. tamariscina, the sole representative inthese analyses of the exclusively Western Australian groupB, is placed outside the Styphelieae clade as the robustlysupported (100%) sister taxon to Oligarrhena micrantha.This placement is also supported by three unique spacerindels: a 144-bp deletion characterises all members of theOligarrhena clade, and both O. micrantha and Mon.tamariscina share two smaller deletions of 1 and 10 bp,respectively. Constraint analysis revealed that an additional118 steps are required on the combined data to renderMonotoca monophyletic. This suggests that members ofgroup B must be removed from the genus and the tribe, a

Fig. 1. Strict consensus of 48397 trees of 1280 steps found from heuristic searching of the spacer data; consistency index excluding uninformativecharacters (CI) = 0.54; retention index (RI) = 0.81; rescaled consistency index (RC) = 0.49. Thick branches received at least 95% jackknife support;other values > 50% shown above branches. Square brackets indicate tribes according to Kron et al. (2002). Abbreviations: A–G, subgeneric groupsof Powell et al. (1997); Ac., Acrotriche; As., Astroloma; B., Brachyloma; Col., Coleanthera; Cy., Cyathodes; Le., Leucopogon; Lept., Leptecophylla;Li., Lissanthe; Mel., Melichrus; Mon., Monotoca; Pe., Pentachondra; Pl., Planocarpa; S., Styphelia sensu Benth.; Styph., Styphelia sensu Sleumer;T., Trochocarpa. Labels: Ac, Acrotriche subclade; An, Androstoma clade; As, Astroloma clade; Co, Conostephium clade; Cs, Cyathodes clade; Cy,Cyathodes sens. str. subclade; L1, Leucopogon sens. str. clade; L2–4, clades including mainly Leucopogon sens. lat.; Lp, Leptecophylla subclade;Me, Melichrus clade; Mo, Monotoca clade. Numbers below the branches indicate the number of unique indels arising on the branch, with thenumber of homoplasic indels within parentheses. Indels arising on terminal branches not shown.

586 Australian Systematic Botany C. J. Quinn et al.

conclusion supported by the morphological analysesconducted by Owens (1996) in which three members of thegroup were represented. We conclude that this group (viz.Mon. leucantha, Mon. oligarrhenoides, Mon. tamarascina)should be moved from the Styphelieae to the Oligarrheneae,and included within an expanded concept of Oligarrhena.

Cyathodes pumila, from the South Island of New Zealand,bears a close morphological resemblance to Cy. dealbatafrom western Tasmania, and the two are robustly grouped(96%) in the spacer analysis (Fig. 1) and placed as sister toMonotoca in the Mo clade with 82% support. This positionis supported by the placement of Cy. dealbata in the same

Prionotes cerinthoides OUTGROUPArcheria comberi

Epacris impressaLysinema ciliatum Epacrideae

Androstoma empetrifoliaLe. milliganii An

As. conostephioides FAs. pinifolium DConostephium pendulum

Co

As. baxteri GB. daphnoides B B 2

Cy. dealbataMon. 'aristata' A

Mon. elliptica CMon. ledifolia AMon. scoparia AMon. rotundifolia A Mo

Le. lanceolatusLe. microphyllus

Le. alternifoliusLe. amplexicaulis

Le. cordatusLe. glabellusLe. plumuliflorus

T. laurina

Pl. nitidaPl. petiolaris

Melichrus procumbens'Pseudactinia' MePe. ericifoliaPe. involucrataPe. pumila

Croninia kingiana

Astroloma sp. 'FNQ'Le. fraseri NSWLe. neoanglicus

As. 'recurvum'As. epacridis AAs. humifusum AAs. ciliatum EAs. macrocalyx AAs. pallidum A

As. tectum

S. exarrhenaS. tubiflora S. viridisLe. esquamatus

Le. setigerLe. fraseri NZ

S. exserta

Col. myrtoidesS. tenuifoliaLe. cuneifolius

Le. allittii

Le. oxycedrusLe. propinquus

Le. nutansLe. pendulus

As. xerophyllum B

Le. blepharolepis

Le. crassifoliusLe. pogonocalyxLe. cymbiformisLe. appressus

Le. muticusLe. ericoidesLe. leptospermoides

L 2

Cy. glaucaCy. platysomaCy. straminea

Cy

Le. cicatricatusLe. pedicellatus L 3

Mon. tamariscina BOligarrhena micranthaNeedhamiella pumilo Oligarrhena clade

As

Sy

L 1

Le. hookeriLe. maccraeiStyph. suaveolens Le. spathaceus

L470

Lept. juniperina var. parvifolia

Lept. juniperina var. juniperina TASLept. pendulosaLept. divaricataLept. tameiameiaeLept. abietinaLept. juniperina var. juniperina NZLept. juniperina var. oxycedrus

Lp

88

(1)

Ac. divaricataLe. pluriloculatus

Li. sapidaLi. strigosa

Ac1

94

51

8885

67

6563

73

1(1)

(1)

75

5964

89

67

1

69

60(1)

77

B. scortechinii CB. priessii A B 1

77

94

Sty

ph

elie

ae c

lad

e

Fig. 2. Strict consensus of 49983 equally parsimonious trees of 1328 steps found from heuristic searching of the matK database; CI = 0.55;RI = 0.77; RC = 0.52. Thick branches received at least 95% jackknife support; other values > 50% shown above the branches. B1, B2, Brachylomaclades; Sy, Styphelia clade; other abbreviations and labels for clades as in Fig. 1. Numbers below the branches indicate the number of unique indelsarising on the branch, with the number of homoplasic indels within parentheses.

Phylogeny of Styphelieae Australian Systematic Botany 587

position on the matK data (Fig. 2, 77% support). Constraintanalyses revealed that an additional 14 steps are required onthe spacer data to include both taxa within Cyathodes sens.str., and 28 extra steps on the combined data to incorporateCy. dealbata in that subclade. Both taxa were excluded fromCyathodes sens. str. by Weiller (1996a) but were notincluded within any of the segregate genera (Weiller 1996b,1996c, 1999), pending molecular study to determine their

relationships. The spacer sequences obtained from the twotaxa differ by only a single base-pair; the same divergencewas observed between two sequences obtained fromdifferent accessions of Cy. dealbata. This suggests the twopopulations may well be conspecific, although detailedmorphological study is still required to establish the statusthat should be accorded them. Nevertheless, theirmorphological distinctiveness from Monotoca is readily

Prionotes cerinthoides OUTGROUPArcheria comberi

Mon. tamariscina BOligarrhena micranthaNeedhamiella pumilo

Oligarrhena clade

Ac. divaricataLe. pluriloculatus

Li. sapidaLi. strigosa Ac

Cy. straminea

Cy. glaucaCy. platysoma Cy

Lept. juniperina var. parvifolia

Lept. abietinaLept. juniperina var. oxycedrusLept. juniperina var. juniperina TASLept. pendulosaLept. divaricataLept. tameiameiaeLept. juniperina var. juniperina NZ

Lp

Le. hookeriLe. maccraeiStyph. suaveolensLe. spathaceus

L4

Le. cicatricatusLe. pedicellatus L3

Pe. pumila

Pe. ericifoliaPe. involucrata

Pl. nitidaPl. petiolaris

As. xerophyllum B

Le. nutansLe. oxycedrusLe. pendulusLe. allittiiLe. propinquusS. tenuifoliaLe. cuneifoliusCol. myrtoidesS. exserta

As. tectum

As. 'recurvum'As. epacridis AAs. humifusum A

As. ciliatum E

As. macrocalyx AAs. pallidum A

Le. esquamatusS. exarrhena

S. tubifloraS. viridis

Astroloma sp. 'FNQ'Le fraseri NSWLe neoanglicusLe fraseri NZLe. setiger

Le. muticusLe. ericoidesLe. leptospermoides

Le. crassifoliusLe. appressus

Le. pogonocalyxLe. cymbiformis

Le. blepharolepisCroninia kingiana

L2

As

Sy

Cy. dealbataMon. 'aristata' A

Mon. elliptica CMon. scoparia A

Mon. rotundifolia AMon. ledifolia A Mo

Melichrus procumbens'Pseudactinia' Me

T. laurina

B. scortechinii CB. priessii A B1

As. baxteri GB. daphnoides B B2

As. conostephioides FAs. pinifolium DConostephium

Co

Le. lanceolatusLe. microphyllus

Le. alternifoliusLe. amplexicaulis

Le. cordatusLe. glabellusLe. plumuliflorus

L1

Androstoma empetrifoliaLe. milliganii An

Epacris impressaLysinema Epacrideae

Sty

ph

elie

aecl

ade

Cs

89

69

80

52

89

93

74

9385

8977

88

83

73

70

89

6488

61

9174

54

9073

89

9090

60

84

Fig. 3. Strict consensus of 3266 equally parsimonious trees of 2479 steps (CI = 0.54, RI = 0.77, RC = 0.50) found from heuristic searching ofthe combined database with 100 replicates of random taxon addition. Abbreviations and labels as in Figs 1 and 2.

588 Australian Systematic Botany C. J. Quinn et al.

apparent: they display none of the features that characterisethat genus (e.g. glabrous corolla, 1–2 locular ovary, A-typepseudomonad pollen). We conclude that it would beinappropriate to include them within Monotoca.

All three analyses group Melichrus and ‘Pseudactinia’with robust support, but in the spacer analysis (Fig. 1), wheretwo species of the former were included, relationshipsbetween the three taxa were not resolved. Hence there is avery close relationship between the three, but our analysesprovide no support for separate generic status for the

undescribed taxon. Collections of a second undescribedtaxon that has been referred to ‘Pseudactinia’ are in ourpossession. More intensive study of a larger representationof the clade is required to elucidate the appropriatetaxonomic treatment.

Only a single representative of each of the three groupsrecognised by Powell et al. (1997) within Brachyloma wasincluded in these analyses. The spacer data provide nosupport for any relationship between them (Fig. 1). ThematK data, however, provide moderate support (77%) for a

Archeria comberi

Pe. ericifoliaPe. involucrata

Pe. pumila

Lept. juniperina var. juniperina TAS

Cy. glaucaCy. platysoma

Cy. straminea

Le. pluriloculatus

Li. sapidaLi. strigosa

Ac. divaricata

10 changes

Prionotes cerinthoides OUTGROUP

Androstoma empetrifoliaLe. milliganii An

Le. cordatusLe. glabellus

Le. plumuliflorusLe. amplexicaulis

Le. alternifoliusLe. lanceolatus

L1

Le. microphyllus

As. conostephioides FAs. pinifolium D

ConostephiumMelichrus procumbens

'Pseudactinia'

Co

Me

As. baxteriB. daphnoides B

B. preissii AB. scortechinii C

T. laurina

B2

B1

Mon. rotundifolia

Mon. scoparia

Cy. dealbataMon. 'aristata'

Mon. elliptica

Mon. ledifolia Mo

Croninia kingianaLe. crassifolius

Le. cymbiformisLe. pogonocalyx

Le. appressus

Pl. nitidaPl. petiolaris

Astroloma sp. 'FNQ'Le. fraseri NSWLe. neoanglicus

Le. fraseri NZLe. setiger

S. exarrhena

S. tubifloraS. viridis

Le. esquamatus

As.' recurvum'Ast epacridis A

As. humifusum A

As. ciliatum E

As. macrocalyx AAs. pallidum A

As. tectum

Col. myrtoides

Le. allittiiLe. propinquus

Le. nutansLe. oxycedrus

Le. pendulusS. tenuifolia

Le. cuneifolius

S. exsertaAs. xerophyllumLe. blepharolepis

Le. ericoidesLe. muticus

Le. leptospermoidesL2

As

Sy

Le. cicatricatusLe. pedicellatus

Le. hookeriLe. maccraeiStyph. suaveolens

Le. spathaceus

L3

L4

Ac

Cy

Lept. abietinaLept. juniperina var. oxycedrus

Lept. juniperina var. juniperina NZLept. divaricata

Lept. pendulosaLept tamaeimaeieLept. juniperina var. parvifolia

Lp

Cs

Epacris impressaLysinema ciliatum Epacrideae

Mon. tamariscinaOligarrhena micrantha

Needhamiella pumiloOligarrhena clade

Sty

phel

ieae

clad

e

Fig. 4. One of the equally parsimonious trees obtained from the combined database, with branch lengths proportional to the amount of change.Abbreviations and labels as in Figs 1 and 2.

Phylogeny of Styphelieae Australian Systematic Botany 589

sister relationship between B. preissii (group A) andB. scortechinii (group C), whereas B. daphnoides (groupB) is strongly grouped (94%) with As. baxteri (Fig. 2).This appears to be at least partly a reflection of the moreconserved nature of the matK data, where pairwisedivergences are markedly lower. Even in the combineddataset, all four taxa are on extremely long terminalbranches (Fig. 4). The current concept of Brachyloma isclearly paraphyletic, but further sampling of groups A andC is required to establish the appropriate taxonomictreatment.

Constraint analysis revealed that an additional 30 stepsare required on the spacer data, and 67 extra steps on thecombined data, to render Astroloma monophyletic,indicating that there is considerable strength in the moleculardata against the current concept of the genus. Astrolomaconostephioides and As. pinifolium are strongly grouped in aclade that is sister to Conostephium in all analyses, and thereis no support for a relationship between this clade and anyother clade containing species of Astroloma (see Fig. 3).Similarly, As. xerophyllum and As. stomarrhena constitute ahighly robust clade, which is the first to diverge within theAs clade in Fig. 1, and this sister relationship with the coremembers of the clade receives robust support in thecombined analysis (98% in Fig. 3). There is strong support(97% in Fig. 3) for the inclusion of all other representativesof the genus within the As clade, and all but one constitute asingle subclade. The placement of As. pallidum outside theAs clade in Fig. 1 appears to be a reflection of theincompleteness of its spacer sequence (30.9% of positionsundetermined). The presence of several long polyAsequences in this species, as well as in As. macrocalyx andAs. tectum, prevented us from generating completesequences, even with the use of additional internalsequencing primers. This appears to be a primary cause ofthe lack of resolution of relationships between them in Fig. 1,and the absence of support for the topology within thesubclade in Fig. 3. Astroloma sp. ‘FNQ’ is grouped with 88%jackknife support with Leucopogon fraseri andLe. neoanglicus (Fig. 3), indicating that this undescribedtaxon is only distantly related to the species of Astrolomagroups A and E.

All representatives of Styphelia sensu Benth. are placedwithin the As clade, but three of them constitute a separateclade that is strongly grouped (91% in Fig. 3) withLe. esquamatus, whereas the other two are placed within theColeanthera subclade with 90% support, S. tenuifolia beingthe strongly supported sister of Le. cuneifolius. Constraintanalyses revealed an extra 16 steps are required on thecombined data to render Styphelia sensu Benth.monophyletic, and an extra 12 steps if Leucopogonesquamatus is included within the clade. Hence, the currentconcept of Styphelia is paraphyletic and requiresmodification.

The distinctiveness and monophyly of Leucopogon sens.str. (clade L1), as defined by Taaffe et al. (2001), is stronglysupported by all three analyses. Clade L2 receives only weaksupport in Figs 1 and 2 (59% and 51%, respectively), butsupport increases to 89% in the combined analysis. In theanalysis of the spacer data, the clade comprises two highlyrobust subclades and Le. crassiflorus, which is not groupedwith either subclade; one of the subclades, however, isunresolved on the matK data (Fig. 2). Both subclades includespecies from each of the postulated segregate genera,‘Axonanthus’ and ‘Gynoconus’ (Powell et al. 1997), whichaccords with the conclusions of Taaffe et al. (2001) thatneither of these concepts is monophyletic.

All representatives of Leucopogon placed within the Asclade were assigned to ‘Axonanthus’. Althoughrelationships within the clade are only poorly resolved ineither Figs 1 or 2, the combined analysis (Fig. 3) providesmuch greater resolution. Four subclades are present: theLe. setiger and Le. esquamatus + Styphelia subclades asunsupported sister groups, the Astroloma sens. str. subclade(which receives no support), and the strongly supportedColeanthera subclade. The placement of Coleantheramyrtoides within a strongly supported (90%) subcladecontaining Styphelia tenuifolia, S. exserta and a group ofLeucopogon species means that recognition of Coleantheraat the generic level would require the elevation of at leasttwo (and probably three) other groups in the subclade togeneric status. Given the placement of the type species ofStyphelia (S. tubiflora) in the Le. esquamatus subclade, thename Coleanthera is available for application to this robustsubclade of the As clade.

The variable nature of the support values for the internaltopology of the As clade is at least partly due to insufficientdivergence between the sequences; this is shown by the shortinternodes in Fig. 4, most notably within the Le. setigersubclade. This indicates the need for additional moleculardata before a reliable estimate of relationships within muchof the As clade can be made. Other loci are beinginvestigated for their potential in this endeavour.

Leucopogon blepharolepis and Croninia kingiana eachoccupy isolated positions terminating early-diverginglineages of the Sy clade (Fig. 3). This supports the decisionof Powell (1993) to erect the latter genus and suggests thatsimilar treatment must also be accorded the former.

The L4 subclade of the Cs clade receives good support,especially from the spacer data (97%, Fig. 1), but therelationships of it to the Cy, Lp and L3 subclades receive littleor no support, even in the combined analysis (Fig. 3). Allmembers of L4 occupy montane habitats in eastern Australia(Le. hookeri, Le. maccraei, Le. montanus), New Zealand(Le. colensoi) or New Guinea (Styphelia suaveolens), and itseems likely that some New Caledonian taxa referred toStyphelia sensu Sleumer also belong in this clade (unpubl.data).

590 Australian Systematic Botany C. J. Quinn et al.

Three representatives of Lissanthe are placed within theone subclade of the Ac clade, and this is strongly groupedwith Li. ‘powelliae’, Leucopogon pluriloculatus andLe. rubicundus (93% in Fig. 1). The last two species weremembers of Leucopogon group E of Powell et al. (1997), andthose authors concluded the entire group should betransferred to Lissanthe. Whereas the molecular analysissupports this conclusion in the case of Le. pluriloculatus andLe. rubicundus, the placement of the two other species ofgroup E (Le. pedicellatus, Le. pleiospermus) indicates themto be strongly grouped with Le. cicatricatus (91% in Fig. 1)and closer to Leptecophylla, Cyathodes sens. str. and the L4subclade (Fig. 3). They are only distantly related toLissanthe. Some of the taxonomic changes necessitatedwithin this group are addressed in an accompanying paper(Crayn et al. 2003).

Leucopogon milliganii is robustly grouped in all threeanalyses with the New Zealand monotypic, Androstoma, andthey share a unique 4-bp deletion in the spacer region.However, even in the combined analysis (Fig. 3), there is nosupport for a relationship between this clade and any otherlineage in the Styphelieae clade. This suggests that the mostappropriate taxonomic treatment would be to expand theconcept of Androstoma to accommodate the former species.

The distributions of many of the indels support elementsin the topologies obtained, and some indicate that thephylogeny may actually be different in certain respects.Hence, three unique indels, two in matK and one in thespacer, reinforce monophyly of the Styphelieae clade, andone spacer indel, an 8-bp deletion at position 621, supportsthe sister relationship between the Styphelieae andOligarrhena clades. The distributions of three spacer indels(a 3-bp insertion at position 431, a 10-bp insertion at 450, a6-bp insertion at 511) support the Le. cicatricatus (L3)subclade being the first lineage to diverged within the Csclade, rather than the topology in Fig. 3, where there is 80%jackknife support for the Acrotriche (Ac) subclade being thefirst to diverge. The distributions of some indels suggest aresolution of polytomies. A 32-bp deletion from the spacersequence is shared by the B1, B2 and Me clades, suggestingthat all three lineages share a common root. Two 6-bpinsertions in the spacer region are shared by Melichrusurceolatus and 'Pseudactinia', but neither is present in Mel.procumbens. This suggests that the first two are sister taxa,which would render Melichrus paraphyletic if the putativenew genus 'Pseudactinia' (Powell et al. 1997) is erected.

The lack of resolution or support for the basalrelationships within the Styphelieae clade, even with themore conservative matK data, contrasts with the strongsupport values obtained for more inclusive clades ofEricaceae by Kron et al. (2002). Hence this is not a result ofsaturation of the database obscuring the phylogenetic signal.Examination of branch lengths in Fig. 4 reveals that basalbranches within the tribe are extremely short, indicating that

there has been a relatively rapid diversification in the earlystages of the evolution of this lineage. Nevertheless, there isstrong support in both databases for sizeable clusters of taxa,which may be interpreted as groups sharing strongphylogenetic relationship, and which can be considered aspotential generic or infrageneric taxa.

Conclusion

Given the breadth of the taxonomic sampling in the aboveanalyses and the strength of the support obtained, we believewe have presented a reliable estimate of the phylogeneticframework of the tribe. The revision of generic conceptsmade necessary by this estimate will be made in a series ofpapers addressing each of the main clades in turn, the first ofwhich is to be found in this volume (Crayn et al. 2003).Detailed morphological analyses of a broadenedrepresentation of the constituent clades will be required toestablish morphological synapomorphies on which to foundthe new taxonomic concepts. However, some considerationof the approach we plan to adopt in selecting appropriategeneric concepts is in order at this stage. With the soleexception of the Sy clade, no large aggregations of taxawithin the Styphelieae clade receive robust support. Neitherdoes there appear to be a unique morphologicalsynapomorphy to define the Sy clade. While this clade is lessinclusive than Styphelia sensu Mueller, it still displayspronounced morphological diversity, including memberscurrently assigned to Astroloma, Coleanthera, Styphelia andLeucopogon. Recognition of the Sy clade as a single genuswould cause no less nomenclatural upheaval than theerection of further segregate genera, and the resultant largegenus would have much lower information content, andhence lower predictive value to the user of the taxonomy,than the remaining genera in the tribe. We consider such ataxonomic treatment of the tribe would be unbalanced, andwould be less likely to prove acceptable to the broaderbotanical community. Hence, we concur with Bentham(1869, p. 146), who concluded that Brown’s narrower genera‘have now been so long and so generally recognised bybotanists and horticulturists, … [that adoption of a broaderconcept of Styphelia] would rather lead to practicalconfusion … [and add] to the already overloaded synonymyof the tribe.’

Acknowledgments

We thank Dr Peter G Wilson for much helpful advice, Dr JMPowell for useful discussion, and the following for plantmaterial: Drs MW Chase, GP Findlay, KA Kron,DJ Mabberley, Y Menadue, B Molloy, PG Wilson, Mr RJohns, Mr PJ de Lange, Mr IR Telford, Mr AR Bean, Mrs JDe Nardi, Mrs GM Russell and Ms N Streiber.

Some of the sequences are from the Honours/Grad. Dip.projects of Mr Wayne Cherry, Mrs Corinne Hawley (neeOwens), Ms Nikola Streiber and Mrs Gabrielle Russell (nee

Phylogeny of Styphelieae Australian Systematic Botany 591

Taaffe); Ms Marcelle O’Brien and Dr Deryn Alpers assistedwith other sequencing work. This project was supported byAustralian Research Council grant no. A19700885.

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592 Australian Systematic Botany C. J. Quinn et al.

Appendix 1. Details of GenBank numbers and voucher specimens for sequence data

Specimen numbers listed using Index Herbariorum abbreviations; RBG indicates living collection voucher. Bold abbreviations distinguish different provenances of a taxon: NSW, New South Wales; NZ, New Zealand; Tas., Tasmania

Plant name Source/Voucher GenBank accession numbermatK spacer

Acrotriche affinis DC. NSW422938 – AY372541Ac. cordata (Labill.) R.Br. NSW416321 – AY372545Ac. divaricata R.Br. NSW242116 AY005094 AF155855Ac. patula R.Br. NSW416315 – AY372542Androstoma empetrifolia Hook.f. UNSW22891 AY372597 AY372670Archeria comberi Hook.f. UNSW22567 AF015632 AF155840Astroloma baxteri A.Cunn. ex DC. NSW413773 AY372598 AY372543As. ciliatum (Lindl.) Druce UNSW23214 AY372599 AF208748As. conostephioides Benth. RBG(NSW)951859 AY372600 AY372544As. epacridis (DC.) Druce NSW414299 AY372601 AY372546As. humifusum (Cav.) R.Br. UNSW22502 AY372602 AF155866As. macrocalyx Behr & F.Muell. ex Sond. NSW415689 AY372603 AY372547As. pallidum R.Br. NSW415929 AY372604 AY372548As. pinifolium (R.Br.) Benth. RBG(NSW)903072 AY372605 AY372549As. stomarrhena Behr & F.Muell. ex Sond. NSW415519 – AY372550As. tectum R.Br. NSW416352 AY372606 AY372551As. xerophyllum (DC.) Sond. UNSW23035 AY372607 AY372554Astroloma sp. nov. ‘FNQ’ NSW409420 AY372608 AY372552Astroloma sp. nov. ‘recurvum’ NSW415757 AY372609 AY372553Brachyloma daphnoides (Sm.) Benth. UNSW22543 AF015633 AF155859B. preissii Sond. NSW415942 AY372610 AY372555B. scortechinii F.Muell. UNSW23180 AY372611 AF208749Coleanthera myrtoides Stschegl. NSW416334 AY372612 AY372556Conostephium preissii Sond. NSW415589 – AY372557C. pendulum Benth. UNSW23032 AY372613 –Croninia kingiana J.M.Powell UNSW23031b AY372614 AF208750Cyathodes dealbata R.Br. UNSW23786 AY372615 AY372559Cy. glauca Labill. UNSW22521 AY005095 AF155858Cy. platysoma C.M.Weiller UNSW23903 AY372616 AY372560Cy. pumila Hook.f. P.J. de Lange 5419 – AY372558Cy. straminea R.Br. UNSW22564 AY372617 AF208752Epacris impressa Labill. RBG(E)680556 AF015636 AF155849Leptecophylla abietina (Labill.) C.M.Weiller UNSW23964 AY372618 AY372561Lept. divaricata (Hook.f.) C.M.Weiller UNSW23748 AY372619 AY372562Lept. juniperina (J.R.Forst. & G.Forst.) C.M.Weiller

subsp. juniperina NZ UNSW23899 AY372621 AY372563Tas. UNSW23901 AY372622 AY372564

Lept. juniperina subsp. oxycedrus (Labill.) C.M.Weiller UNSW17190 AY372623 AY372565Lept. juniperina subsp. parvifolia (R Br.) C.M.Weiller UNSW22558 AY372624 AY372566Lept. pendulosa (Jarman) C.M.Weiller UNSW17195 AY372625 AY372567Lept. robusta (Hook.f.) C.M.Weiller P.J. de Lange CH262 – AY372568Lept. tameiameiae (Cham. & Schltdl.) C.M.Weiller NSW456149 AY372626 AY372569Leucopogon allittii F.Muell. NSW419082 AY372627 AF208753Le. alternifolius R.Br. NSW416062 AY372628 AF208754Le. amplexicaulis (Rudge) R.Br. NSW411658 AY372629 AF208755Le. appressus R.Br. RBG(NSW)910642 AY372630 AF208756Le. assimilis R.Br. NSW416186 – AF208757Le. australis R.Br. NSW416061 – AF208758Le. blakei Pedley NSW492715 – AY372570Le. blepharolepis (F.Muell.) Benth. NSW413775 AY372631 AY372571Le. bossiaea (F.Muell.) Benth. NSW416197 – AF208759Le. cicatricatus J.M.Powell NSW500119 AY372632 AY372572Le. colensoi Hook.f. P.J. de Lange 5415 – AY372573

Phylogeny of Styphelieae Australian Systematic Botany 593

Appendix 1. (continued)

Plant name Source/Voucher GenBank accession numbermatK spacer

Le. cordatus Behr & F.Muell. ex Sond. RBG(NSW)854043 AY372633 AF208760Le. cordifolius Lindl. NSW419081 – AF208761Le. corynocarpus Sond. NSW414780 – AF208762Le. crassiflorus (F.Muell.) Benth. NSW415523 – AF208763Le. crassifolius Sond. NSW416319 AY372634 AF208764Le. cuneifolius Stschegl. NSW415764 AY372635 AF208765Le. cymbiformis DC. NSW415754 AY372636 AF208766Le. dielsianus E.Pritzel NSW415695 – AF208767Le. ericoides (Sm.) R.Br. UNSW22538 AY372637 AF208768Le. esquamatus R.Br. UNSW23811 AY372638 AF208769Le. fletcheri Maiden & E.Betche UNSW23855b – AF208770Le. fraseri DC. NSW NSW414293 AY372639 AF208771

NZ UNSW22892 AY372640 AF208772Le. gibbosus Stschegl. UNSW23000 – AF155863Le. glabellus R.Br. NSW415932 AY372620 AF208773Le. hookeri Behr & F.Muell. ex Sond. UNSW22503 AY372641 AF208774Le. juniperinus R.Br. UNSW23527 – AF208775Le. lanceolatus (Sm.) R.Br. UNSW21174 AY372642 AF208776Le. leptospermoides R.Br. UNSW23155 AY372643 AF208777Le. maccraei F.Muell. RBG(NSW)860529 AY372644 AF208778Le. margarodes R.Br. NSW434477 – AY372574Le. microphyllus (Cav.) R.Br. UNSW21169 AY005097 AF155862Le. milliganii Rodway NSW459519 AY372645 AY372575Le. montanus (R.Br.) J.H.Willis UNSW23799 – AY372576Le. muticus R.Br. NSW392764 AF015638 AF155864Le. neoanglicus Benth. UNSW23596 AY372646 AF208779Le. nutans E.Pritzel NSW415613 AY372647 AF208780Le. ovalifolius Sond. NSW415517 – AF208781Le. oxycedrus Behr & F.Muell. ex Sond. NSW415665 AY372648 AF208782Le. pedicellatus C.T.White NSW504182 AY372649 AY372577Le. pleiospermus (F.Muell.) Benth. NSW492873 – AY372578Le. pendulus R.Br. NSW415769 AY372650 AF208784Le. plumuliflorus (F.Muell.) Benth. NSW415532 AY372651 AF208785Le. pluriloculatus F.Muell. UNSW23597 AY372652 AF208786Le. pogonocalyx Benth. NSW415696 AY372653 AF208787Le. propinquus R.Br. NSW415772 AY372654 AF208788Le. rubicundus F.Muell. NSW416370 – AY372579Le. setiger R.Br. NSW412089 AY372655 AF208790Le. spathaceus Pedley NSW363698 AY372656 AY372580Le. strictus Benth. NSW413777 – AF208791Le. tamminensis E.Pritzel NSW416195 – AF208792Lissanthe brevistyla A.R.Bean AQ666659 – AY372581Li. sapida R.Br. UNSW23213 AY372657 AF208793Li. strigosa (Sm.) R.Br. RBG(NSW)931330 AY372658 AF208794Lissanthe sp. nov. ‘powelliae’ NSW415501 – AY372582Lysinema ciliatum R.Br. UNSW22193 AF015639 AF155848Melichrus procumbens (Cav.) Druce NSW392758 AY005098 AF155856Mel. urceolatus R.Br. UNSW23870 – AY372595Monotoca elliptica (Sm.) R.Br. UNSW22856 AY005099 AY005085Mon. ledifolia A.Cunn exDC. CBG 8704096 AY372660 AY372584Mon. rotundifolia J.H.Willis UNSW22868 AY372661 AY372585Mon. scoparia (Sm.) R.Br. NSW392761 AF015640 AF155857Mon. tamariscina F.Muell. UNSW22968 AY372662 AY372586Monotoca sp. nov. ‘aristata’ RBG(NSW)893165 AY372659 AY372583Oligarrhena micrantha R.Br. UNSW22974 AF539985 AF155854Needhamiella pumilio (R.Br.) L.Watson UNSW22976 AF539984 AF155853Pentachondra ericifolia Hook.f. CBG 8900276 AY005100 AY005086

594 Australian Systematic Botany C. J. Quinn et al.

http://www.publish.csiro.au/journals/asb

Appendix 1. (continued)

Plant name Source/Voucher GenBank accession numbermatK spacer

Pe. involucrata R.Br. NSW268191 AY005101 AY005087Pe. pumila (J.R.Forst. and G.Forst.) R.Br RBG(E)681038 AF015641 AF155860Planocarpa nitida (Jarman) C.M.Weiller UNSW23798 AY372663 AY372593Pl. petiolaris (DC.) C.M.Weiller UNSW22563 AY372664 AY372594Prionotes cerinthoides (Labill.) R.Br. M.W.Chase 890 (K) AF015642 AF155838‘Pseudactinia’ gen. nov. NSW416093 AY372665 AY372596Styphelia exarrhena (F.Muell.) F.Muell. NSW422929 AY372666 AY372587S. exserta (F.Muell.) Sleumer NSW416385 AY372667 AY372588S. suaveolens (Hook.f.) Warb. R.Johns 9397 (K) AY372668 AY372589S. tenuifolia Lindl. NSW413776 AY372669 AY372590S. tubiflora Sm. UNSW24020 AY372670 AY372591S. viridis Andrews UNSW23062 AY005105 AF155865Trochocarpa gunnii (Hook.f.) Benth. UNSW22574 – AF155861T. laurina (Rudge) R.Br. RBG(NSW)822985 AY005106 AY005092Trochocarpa sp. nov. ‘A’ sensu Powell, 1992 NSW504425 – AY372592