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Developmental morphology of Sarcodia montagneana and S. grandifoliafrom New Zealand and a phylogeny of Sarcodia (Sarcodiaceae,Rhodophyta) based on rbcL sequence analysisConxi Rodríguez-Prietoa; Showe-Mei Linb; Wendy A. Nelsonc; Max H. Hommersandd

a University of Girona, Faculty of Sciences, Department of Environmental Sciences, Campus deMontilivi, 17071 Girona, Spain b Institute of Marine Biology, National Taiwan Ocean University,Keelung 20224, Taiwan, Republic of China c National Institute of Water and Atmospheric Research(NIWA), Kilbirnie, Wellington 6241, New Zealand d Department of Biology, University of NorthCarolina at Chapel Hill, Chapel Hill, North Carolina 27599-3280, USA

First published on: 23 May 2011

To cite this Article Rodríguez-Prieto, Conxi , Lin, Showe-Mei , Nelson, Wendy A. and Hommersand, Max H.(2011)'Developmental morphology of Sarcodia montagneana and S. grandifolia from New Zealand and a phylogeny ofSarcodia (Sarcodiaceae, Rhodophyta) based on rbcL sequence analysis', European Journal of Phycology, 46: 2, 153 — 170,First published on: 23 May 2011 (iFirst)To link to this Article: DOI: 10.1080/09670262.2011.580198URL: http://dx.doi.org/10.1080/09670262.2011.580198

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Developmental morphology of Sarcodia montagneana and

S. grandifolia from New Zealand and a phylogeny of Sarcodia(Sarcodiaceae, Rhodophyta) based on rbcL sequence analysis

CONXI RODRIGUEZ-PRIETO1, SHOWE-MEI LIN2, WENDY A. NELSON3 AND

MAX H. HOMMERSAND4

1University of Girona, Faculty of Sciences, Department of Environmental Sciences, Campus de Montilivi, 17071 Girona, Spain2Institute of Marine Biology, National Taiwan Ocean University, Keelung 20224, Taiwan, Republic of China3National Institute of Water and Atmospheric Research (NIWA), Private Bag 14-901, Kilbirnie, Wellington 6241, New Zealand4Department of Biology, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina 27599-3280, USA

(Received 12 January 2011; revised 27 March 2011; accepted 6 April 2011)

Despite its widespread distribution in the Indo-West Pacific Ocean, Sarcodia is one of the least understood genera among the

red algae. Samples investigated from different parts of the Indo-West Pacific Ocean were well separated phylogenetically,

based on rbcL base pair distances, and probably represent separate species. In this report we investigate the developmental

morphology of two species from New Zealand: the type species, Sarcodia montagneana, and S. grandifolia (including S.

‘flabellata’). Molecular studies indicate that S. montagneana is restricted to the north-eastern part of the North Island and that

S. grandifolia occurs from Wellington southwards as far as the Snares Islands. Thalli are multiaxial and consist of three

primary layers: a surface layer of uninucleate cells, a cortex of five or six layers of polygonal multinucleate cells, and a medulla

of multinucleate stellate cells. Secondary rhizoidal filaments are frequent to abundant in the medulla. Spermatangia occur in

chains and release spermatia through a pore. The female reproductive system is procarpic and consists of a supporting cell that

bears two side branches and a one-celled terminal carpogonium. The auxiliary cell is the basal cell of one of the side branches

and is undifferentiated prior to fertilization. After presumed fertilization, the carpogonium separates into a terminal cell and a

subterminal hypogynous cell that fuses with the auxiliary cell and deposits in it a single nucleus. The auxiliary cell cuts off a

gonimoblast initial obliquely that first forms a short linear chain of gonimoblast cells. Cystocarp formation proceeds rapidly

with the outward development of linear files derived from surface cells that differentiate into a terminal ostiole and inner and

outer pericarp layers separated by a central cavity. The mature gonimoblasts consist of an inner gonimoblast reticulum of

laterally fused cells that attach secondarily to outer cells of the inner pericarp and outer gonimoblast filaments that terminate

in short chains of carposporangia. Tetrasporangia are zonately divided and are scattered over the thallus surface with new

initials formed continuously. The morphological and molecular observations reported here contrast sharply with those of

previous studies and are largely new to the Sarcodia literature.

Key words: molecular systematics, morphology, New Zealand, phylogeny, Rhodophyta, Sarcodia, Sarcodiaceae, taxonomy

Introduction

J.G. Agardh (1852, p. 622) established Sarcodiain the tribe Sphaerococcoideae based on a singlespecies, Rhodomenia (Rhodymenia) montagneana,which J.D. Hooker & Harvey (1845) describedfrom the Bay of Islands, New Zealand. Agardhcharacterized thalli of the genus as flat, fleshy–membranous, 3–5 times dichotomously branched,and consisting of three layers: a surface layer ofsmall vertically rounded cells, an intermediatelayer of angular rounded cells, and an interiorlayer of laxly intertwining hyaline filiform cells.

The cystocarps were described as being hemispher-ical, raised along the margins and surfaces of thedichotomous branches, and bearing obovate-elon-gate carpospores on dichotomously branched fila-ments. Gonimoblast filaments were said tosurmount a raised central placenta inside an ostio-late pericarp of concentric layers of radiating cells.Tetrasporangia were described as being oblong,zonately quadripartite and not aggregated in soribut distributed individually over the frond. Thekey distinguishing characters given by Agardhthat separated Sarcodia from other members ofthe Sphaerococcoideae were the tripartite layeringof the vegetative thallus and the scattered, zonatelydivided tetrasporangia.

Correspondence to: Conxi Rodrıguez-Prieto. E-mail: conxi.

[email protected]

ISSN 0967-0262 print/ISSN 1469-4433 online/11/020153–170 � 2011 British Phycological Society

DOI: 10.1080/09670262.2011.580198

Eur. J. Phycol. (2011) 46(2): 153–170

Downloaded By: [Rodriguez-Prieto, Conxi] At: 05:13 24 May 2011

Kylin (1932, 1956) established the familySarcodiaceae to contain Sarcodia J. Agardh,Chondrymenia Zanardini, Dicurella Harvey,Trematocarpus Kutzing, and provisionallyNizymenia Sonder. Dicurella has been merged withTrematocarpus (see Searles, 1969) and Nizymenia isplaced in its own family (see Chiovitti et al., 1995).In establishing the Sarcodiaceae, Kylin emphasizedthe outward orientation of the gonimoblasts, itsdevelopment from an irregular fusion cell, and thezonate division of the tetrasporangia. Rasmussen(1964) reported that the carpogonial branchsystem of S. montagneana (J.D. Hooker &Harvey) J. Agardh consists of a supporting cell,one or two carpogonial branches of three cellseach, and two or three subsidiary branches, andalso that the hypogynous cell links with a nearbyundifferentiated auxiliary cell after presumed fertil-ization by means of a connecting filament. Hereferred to the gonimoblast fusion cell as a reticu-lum. Similarly, Norris (1987) described a connect-ing filament in S. dentata (Suhr) R.E. Norris thatlinked the carpogonial branch to a nearby auxiliarycell, and also referred to the inner gonimoblastregion as a reticulate fusion cell. Norris suggestedthat vegetative cells might play a role in the forma-tion of the fusion cell. More recently, Kraft (in Liaoet al., 1993) illustrated a three-celled carpogonialbranch in S. montagneana that was similar to theone described earlier by Searles (1968, 1969) in spe-cies of Trematocarpus.Kylin (1932, 1956) placed his family Sarcodiaceae

in the order Gigartinales; however, Saunders et al.(2004) specifically excluded the family from theGigartinales and instead inferred a weak relation-ship between the Sarcodiaceae and the Plocamialesbased on a phylogenetic analysis of nuclear small-subunit ribosomal DNA sequences. Guiry & Guiry(2011) list 20 species (and infraspecific) names intheir current database under Sarcodia, of which 12are flagged as currently accepted taxonomically.Even so, S. montagneana is the name commonlychosen by the majority of phycologists when report-ing a Sarcodia species from localities in the Indo-West Pacific Ocean (Silva et al., 1996; Guiry &Guiry, 2011) and from Antarctica and subantarcticwaters (Wiencke & Clayton, 2002, but seeHommersand et al., 2009). This is not surprising,since morphological variation overlaps amongmany collections and the habit of most falls withinthe range of forms reported for S. montagneanafrom New Zealand.In the present study we investigate the structure

and distribution of the two species of Sarcodiafound in New Zealand, S. montagneana andS. grandifolia (including S. ‘flabellata’) basedon a detailed analysis of their vegetative andreproductive development. A phylogeny of

representative species of Sarcodia from NewZealand, Australia and regions of the Indo-WestPacific Ocean is presented and evaluated based onanalyses of rbcL sequences.

Materials and methods

Morphological observations

Specimens of Sarcodia montagneana and S. grandifolia

were collected from the New Zealand coast subtidallyfrom –0.5m to c. –25m depth, or from the drift.Material was observed in surface view or sectionedwith a freezing microtome, and preparations werestained either with 1% acidified aqueous aniline blueor with a solution of haematoxylin according to themethod of Rodrıguez-Prieto & Hommersand (2009).Habit pictures were taken with a Canon EOS 350D(Canon, Tokyo, Japan) and photomicrographs weremade with an AxioCam MRc attached to an Axioskop2 plus microscope (Zeiss, Berlin, Germany). Habit scansof historically important collections were provided bythe Natural History Museum, London, UK (BM) andTrinity College, Dublin, Ireland (TCD). Voucher speci-mens were deposited in the herbaria of the Museum ofNew Zealand Te Papa Tongarewa, Wellington, NZ(WELT); Auckland Institute and Museum, Auckland,NZ (AK); Herbarium of the University of Girona,Spain (HGI); and herbarium of the University ofNorth Carolina, Chapel Hill, USA (NCU). Herbariumabbreviations follow Thiers (2011, continuouslyupdated) Index Herbariorum: http://sweetgum.nybg.org/ih/.

Molecular observations

DNA from silica gel dried specimens was extracted usingDNeasy Plant Mini Kits (Qiagen, Valencia, CA, USA)following the manufacturer’s instructions. DNAsequencing procedures were as described in Lin et al.(2001). New sequence data and those available fromGenBank were compiled and aligned with Sequencher(Gene Codes, Ann Arbor, MI, USA). Thirteen rbcLsequences were newly generated. Locality data for eachsample analysed are provided in Table 1.

Phylogenetic analyses were performed using maxi-mum parsimony (MP), maximum likelihood (ML)and Bayesian analyses (BA) using PAUP� v4.0(Swofford, 2002), GARLI 1.0 (Zwickl, 2006) andMrBayes 3.1.2 (Ronquist & Huelsenbeck, 2003), respec-tively, as described in Lin et al. (2001, 2004, 2011).Support values for MP and ML trees were assessed bycalculating 1000 and 100 bootstrap resamplings usingPAUP� v4.0 and GARLI 1.0, respectively. Thegeneral-time-reversible model for ML was used as thedefault in GARLI 1.0. A Bayesian analysis was per-formed in MrBayes 3.1.2 (Ronquist & Huelsenbeck,2003) using a GTRþ Iþ� model, which allowed forrate variation among different codon positions.The analysis used four chains, one cold and three incre-mentally heated. Each run started with a random treeand consisted of 106 generations with sampling every

C. Rodrıguez-Prieto et al. 154


http://sweetgum.nybg.org/ih/


Table

1.Listofspeciesusedin

rbcL

analysiswithGenBankaccessionnumbers.

Species

Locationandcollectingdata

Accessionno.

Source

Plocamium

‘cartilagineum’

(Linnaeus)

P.S.Dixon

New

Zealand,Stewart

Island,HalfMoonBay,Lonneker’sNugget,Coll.D.W

.Freshwater

NZ04-159,29October

2004(N

CU

589564)

HQ224543

Thisstudy

Trematocarpusantarcticus(H

ariot)

Fredericq&

Moe

Antarctica,South

ShetlandIslands,KingGeorgeIsland,Bahıa

Fildes,Punta

Collins,coll.

S.Fredericq&

J.Rhodriguez,15February1994(SF

2-15-94-1-8)

U21698

Hommersandet

al.,2009

Sarcodia

grandifoliaLevring

New

Zealand,Stewart

Island,0m,theneck,opposite

Cow

Island,Coll.J.F.Herrick,

07October

1994(A

K220918)

U26822(asS.‘flabellata’)

S.Fredericq,unpublished

Sarcodia

grandifoliaLevring

New

Zealand,South

Island,ShagPoint,Coll.D.W

.Freshwater&

M.H

.Hommersand

NZ04-056,24October

2004(asS.‘flabellata’)(N

CU

591798)

HQ224542

Thisstudy

Sarcodia

grandifoliaLevring

New

Zealand,NorthIsland,Wellington,Princess

Bay,low

intertidal,Coll.D.W

.

Freshwater&

M.H

.HommersandNZ04-599,17Novem

ber

2004(N

CU

591799-802)

HQ224540

Thisstudy

Sarcodia

grandifoliaLevring

New

Zealand,NorthIsland,Wellington,low

intertidal,westLyallBay,Coll.W.Nelson,

7Decem

ber

2009(W

ELT

A030183).

HQ224541

Thisstudy

Sarcodia

grandifoliaLevring

New

Zealand,StewartIsland,Lee

Bay,Coll.D.W

.FreshwaterNZ04-186,30October

2004

(asS.‘flabellata’)(N

CU

591797)

HQ224539

Thisstudy

Sarcodia

sp.

Australia,New

South

Wales,JervisBay,�20m,coll.A.J.K

.Millar&

DHardin,

15October

1995(N

CU

531772-773)

AF212194

Hommersand&

Fredericq,2003

Sarcodia

marginata

J.Agardh

Australia,Victoria,Warrnambool,coll.M.H

.Hommersand,13July1995(N

CU

591796)

AF212193

Hommersand&

Fredericq,2003

Sarcodia

montagneana

(J.D

.Hooker

&Harvey)

J.Agardh

New

Zealand,NorthIsland,Taipa,coll.WA

Nelson,2Novem

ber

1993

U21705AY294374

Hommersand&

Fredericq,2003

Sarcodia

montagneana

(J.D

.Hooker

&Harvey)

J.Agardh

New

Zealand,NorthIsland,BayofIslands,Russell,OneroaBay,�5to�8m,Coll.

R.D’A

rchino,2Decem

ber

2009(W

ELT

A030191)

HQ224538

Thisstudy

Sarcodia

sp.2

Indonesia,Bali,Lagoon,Coll.S.-M.Lin,23July

2008(SML23vii2008-ss2)

HQ224531

Thisstudy

Sarcodia

ceylanicaHarvey

ex

Kutzing

SriLanka,Hikkaduwa,Coll.E.Coppejans,1February

2007(H

EC16114)

HQ224534

Thisstudy

Sarcodia

ceylanicaHarvey

ex

Kutzing

SriLanka,Tangalla,Coll.E.Coppejans,7April2005(H

EC15578)

HQ224532

Thisstudy

Sarcodia

ceylanicaHarvey

ex

Kutzing

SriLanka,Weligama,Coll.E.Coppejans,5January

1996(H

EC1155a)

HQ224533

Thisstudy

Sarcodia

sp.1

Indonesia,Bali,Lagoon,Coll.S.-M.Lin,23July

2008(SML23vii2008-ss1)

HQ224535

Thisstudy

Sarcodia

dentata

(Suhr)

R.E.Norris

South

Africa,CapeProvince,Port

Alfred,TheKowie,collM.H

.Hommersand,10July

1993[silicagel

sample¼voucher]

U26816

Hommersand&

Fredericq,2003

Sarcodia

sp.

Taiwan,Orchid

Island,FiveCaves,Coll.S.-M.Lin,4April2003(SML4iv2003_FC-O

I)HQ224536

Thisstudy

Sarcodia

sp.

Taiwan,Keeling,Coll.L.-J.

Liu

&X.-R.Hsieh,27May2010(SML23vii2008-ss1)

HQ224537

Thisstudy

Morphology and phylogeny of Sarcodia 155


100 generations. Stationarity was reached at

generation 5600. Thus, trees saved until generation

5600 were the ‘burn-in’ of the chain, and inferences

about the phylogeny were based on those trees sampled

after generation 5600. A 50% consensus tree

(majority rule as implemented by PAUP� v4.0) was

computed from the 9944þ 1 trees saved after the

burn-in point.

Results

Morphological observations

Sarcodia montagneana (J.D. Hooker & Harvey)

J. Agardh 1852, p. 623

(Figs 2–44)BASIONYM: Rhodymenia montagneana J.D. Hooker& Harvey 1845, p. 544–545 (‘Rhodomenia’).LECTOTYPE (designated here): Cystocarpicspecimen in Herb. BM, BM001032008 labelled‘Bay of Islands, New Zealand, September L/27’collected by David Lyall, during the AntarcticExpedition of the Erebus and Terror 1839–1843(Fig. 2).TYPE LOCALITY: New Zealand, North Island, Bay ofIslands (Silva et al., 1996).SYNTYPES: Samples of S. montagneana collectedby either David Lyall or J.D. Hooker in theBay of Islands, North Island, New Zealand,between August and November 1841, during theAntarctic Expedition of the Erebus and Terror1839–1843. TCD0012148 labelled ‘N. Zeal., R.montagneana’ (this is the reverse of the image pub-lished without comment by Harvey in NereisAustralis 1849, pl. 48) (Fig. 3); TCD0012150,labelled ‘New Zealand J.D.H., R. MontagneanaH & H’; BM000044273 labelled ‘Rhod.Montagnei, H & H, New Zealand, Antarct.Exped. 1839–1843 J.D.H.’; BM000044274 labelled‘Bay of Islands, New Zealand, September JDH’(cystocarpic); BM001032007 labelled ‘R. ornata?Mont., Bay of Islands, New Zealand, September1841’; BM000044272 with added label ‘Rhod.Montagnei, H & H, New Zealand, Antarct.Exped. 1839–1843 J.D.H’; BM001032001 labelled‘R. Montagnei H & H, Bay of Islands, NewZealand, September 1841’; BM 00103200 labelled‘Rhod. Montagnena H & Harv, New Zealand,Herb. Dickie’; BM00004271 labelled‘Rhodomenia Montagnei H & H, N. Zealand’(cystocarpic); BM001032006 labelled ‘Bay ofIslands, New Zealand, September 1841’;BM001032005 labelled ‘Bay of Islands, NewZealand, September 1841’. (Note: not all the speci-mens in the type folder at BM are confirmed to beSarcodia montagneana.)DISTRIBUTION: Northern North Island, east coast tothe Bay of Plenty, New Zealand (Fig. 1).

HABITAT: From the intertidal region to �11mdepth, collected throughout the year.SPECIMENS EXAMINED: North Island, New Zealand:

Doubtless Bay, Mangonui, 34�590S, 173�320E(V.W. Lindauer, 05 December 1942, WELTA020586). Bay of Islands: Poraenui Point,35�11.40S 174�04.20E (R. D’Archino, 30 November2009, WELT A030196, tetrasporangial); Moturoa(S point), 35�130S, 174�050E, –5m (K.A. Johnson,15 November 1974, WELT A008920, tetrasporan-gial); Te Miko Reef (between Moturua andMotuarohia), 35�13.40S, 174�10.50E, �10.4m(R. D’Archino, 21 September 2010, WELTA030869, female); East of Motuarohia, 35�13.90S174�10.90E, �11m, on rhodolith bed(R. D’Archino, 02 December 2009, HGI-A 9545,female); Long Beach, 35�14.80S 174�07.30E, drift(R. D’Archino, 18 September 2010, WELTA030870, female); Oneroa Bay, 35�150S 174�070E,�5 to �8m, on forest of Carpophyllum flexuosum(Esper) Greville, sheltered site (R. D’Archino, 02December 2009, WELT A030191, female); Paihia,35�170S, 174�060E, drift (W.A. Nelson, 07November 1991, WELT A020101, female);Paihia, 35�170S, 174�060E, drift (W.A. Nelson, 27April 1990, WELT A019414, male); KahuwheraPoint, 35�260S, 174�180E, �6.3m (R. D’Archino,19 September 2010, WELT A030867, female;WELT A030868, tetrasporangial); Whitianga,Buffalo Beach, 36�490S, 175�420E, drift (U.V.Dellow, 10 December 1949, WELT A002706,female).

Fig. 1. Map of the distribution of the species of Sarcodia inNew Zealand. Open circles: collection localities of S. mon-

tagneana. Solid circles: collection localities of S. grandifolia.



Figs 2–13. Sarcodia montagneana, habit and vegetative development. Figs 6–8, 13: aniline blue; Figs 9–12: haematoxylin. 2.Lectotype: Rhodomenia (Rhodymenia) montagneana J.D. Hooker & Harvey 1845, Herb. BM001032008. Label reads: Bay ofIslands, New Zealand, September 28L/27. This specimen was collected by David Lyall and corresponds to the description anddiscussion of a female plant in Hooker & Harvey, 1845, pp. 544–545. 3. Rhodomenia (Rhodymenia) montagneana J.D. Hooker

& Harvey 1845, Herb. TCD0012148. The label reads: N. Zeal. R. Montagneana. The figure published by Harvey in NereisAustralis, 1849, pl. XLVIII is the reverse image of this plant. 4. Sarcodia montagneana (Hook. fil. & Harvey) J. Agardh.Female plant, WELT A020101, drift, Northland, Bay of Islands, Paihia, coll. W.A. Nelson, 07 November 1991. 5. Part of a

young branch of plant in Fig. 4 showing early development of cystocarps along thallus margins and surface. 6. Transversesection near apex of thallus (HGI-A 9545). 7. Transverse section showing stellate cells in young medulla (arrow) (WELTA030191). 8. Transverse section of surface layer and outer cortex (WELT A030191). 9. Detail of cortex and surface layer in

cross section, showing nucleus and plastid in surface cells (WELT A030870). 10. Surface view showing nucleus and plastid insurface cells (HGI-A 9545). 11. Inner multinucleate subcortical cell showing the small nuclei and dissected ribbon-like plastid(HGI-A 9545). 12. Multinucleate stellate cell in medulla compared to cortical cell in Fig. 11 (HGI-A 9545).

13. Transverse section of cortex and medulla showing distribution of cortical and stellate medullary cells and secondaryrhizoidal filaments (HGI-A 9545). Abbreviations: n: nucleus; p: plastid. Scale bars: 5 cm (Figs 2–5), 100mm (Figs 6, 13)or 20mm (Figs 7–12).



Figs 14–24. Sarcodia montagneana. Spermatangial and early stages of female reproductive development. Haematoxylin. 14.Transverse section of a spermatangial sorus derived from surface layer showing elongated spermatangial parent cells (WELT

A019414). 15. Spermatangia aligned in rows (WELT A019414). 16. Transverse section of female plant showing a potentiallyfertile surface area (WELT A030191). 17. Supporting cell undergoing nuclear division (WELT A030870). 18. Supporting cellbearing paired unicellular, uninucleate sterile branches (WELT A030869). 19. Supporting cell bearing two 2-celled sterile

branches (WELT A030191). 20. Carpogonium with short trichogyne partly covered by a 2-celled sterile branch(Continued)



Habit and vegetative structure: Thallus erect,flattened, up to 25 (–40) cm in length,attached to the substratum by a discoidholdfast, shortly stipitate, complanate, subdichoto-mously branched with rounded axils and apices(Figs 2–5). Axis up to 2 (–6) cm wide. Marginssmooth or occasionally proliferous (Figs 2–4),slightly thicker than the lamina. Thallus moderatelyfirm and rosy to dark red in colour.Structure multiaxial with a lax medulla

(Figs 6, 7). Outer cortex bilayered, composed ofanticlinal once-dichotomously branched filamentsthat are usually two cells long, without secondarypit connections (Figs 8, 9). Outer cortical cells uni-nucleate with a parietal plastid that is ovoid toelongate in transverse section and rounded topolygonal in surface view, up to 9 (–14) mm highand 6 (–12) mm in diameter (Figs 8–10). Subcortexcomposed of a multilayered network of cells con-nected to neighbouring cells by secondary pit con-nections, the outermost ovoid and small, becomingstellate and increasing in size inwardly, with thelargest up to 95mm in diameter (Figs 7–9, 11).Subcortical cells multinucleate (Figs 9, 11), theinner ones with dissected ribbon-like plastids(Fig. 11). Medulla composed of stellate cells withthe rounded central portion up to 55 mm in diam-eter and entangled slender rhizoidal filaments,some of them longitudinally orientated and up to12mm in diameter (Figs 6, 12, 13). Stellate cellsmultinucleate with dissected ribbon-like plastids(Fig. 12). Medullary rhizoidal filaments multi-nucleate, originating secondarily from inner corti-cal and stellate medullary cells.

Life cycle: Triphasic, with isomorphic gametophytesand tetrasporophytes. Gametophytes dioecious.

Male reproductive structures: Spermatangial parentcells initiated in sori from surface cortical cells thatelongate toward the thallus surface forming a tube(Fig. 14). Spermatangia ultimately formed in rowswith the terminal cells differentiating spermatia(Fig. 15). Upper part of the tube fusing with thethallus surface and releasing spermatia.

Female reproductive structures: Fertile areas clearlydistinguishable in cross section by their darklystaining cells, which are slightly larger than neigh-bouring cells (Fig. 16). Such cells undergo division(Fig. 17) to produce a basal cell (¼the supporting

cell) that bears two sterile uninucleate one-celledbranches (Fig. 18) which later divide to formtwo-celled branches (Fig. 19). The uninucleate sup-porting cell cuts off a one-celled carpogoniumtransversely with a straight trichogyne that elon-gates toward the thallus surface, flanked by thesterile branches (Figs 20–23). A pit connectionlinks the sterile branches to the supporting cell(Fig. 21, arrow). After presumed fertilization, thefirst cell of one of the sterile branches enlarges anddifferentiates into an auxiliary cell (Fig. 23) and thecarpogonium divides into a two-celled filament,with the basal cell becoming the hypogynous celladjacent to the auxiliary cell (Fig. 24). The carpo-gonial remnant degenerates and the auxiliary cellcuts off a gonimoblast initial (Fig. 25), which inturn gives rise to a row of cells that comprise theearliest gonimoblasts flanked by the sterilebranches (Figs 26–28).Cystocarp development begins when the outer

cortex resumes growth and forms files of cells adja-cent to and above the developing gonimoblasts(Fig. 29). The cells adjacent to the gonimoblastsbecome darkly staining and will form the innerpericarp (Fig. 29, circle), while those above arelighter staining and will form the outer pericarp(Fig. 30). The auxiliary cell sits basally betweenthe inner pericarp filaments and the originalouter cortex (Figs 29, 30). The outer pericarp sep-arates from the inner pericarp, forming the centralcavity, with some cells elongating and remaininginterconnected between the outer and inner peri-carps (Fig. 30, arrow). During cystocarp develop-ment, the gonimoblast filaments branch andprogressively fill the cavity between the inner andouter pericarp layers (Figs 31, 32). Gonimoblastcells adjacent to the inner pericarp and those inthe inner part of the central cavity are initially uni-nucleate, containing large nuclei (5–9 mm diame-ter). Inner gonimoblast cells fuse with oneanother to form a gonimoblast reticulum of multi-nucleate filaments with nuclei 5–9 mm in diameter(Figs 32, 33), and those adjacent to and above theinner pericarp link to inner pericarp cells (Fig. 32,arrow). The region above the gonimoblast reticu-lum is composed of elongated cells which give riseto the gonimoblast filaments that differentiate intoterminal chains of carposporangia (Figs 31, 32).Mature gonimoblasts reach up to 650mm in diam-eter (Fig. 32).

(Continued)

(WELT A030191). 21. Supporting cell bearing a carpogonium and two sterile branches. The arrow points to the pit connectionbetween the supporting cell and a sterile branch (WELT A030870). 22. Supporting cell bearing an elongating carpogoniumand two sterile branches (WELT A030870). 23. Supporting cell bearing a carpogonium and sterile branch in which the basal

cell has differentiated into an auxiliary cell (WELT A030870). 24. Supporting cell with two side branches in which the basalcell of one has differentiated into an auxiliary cell. The carpogonium has divided into a hypogynous cell and a terminal cell(WELT A030870). Abbreviations: ac: auxiliary cell; cp: carpogonium; hy: hypogynous cell; n: nucleus; sb: sterile branches; sc:

supporting cell; sg: spermatangium; spc: spermatangial parent cell; t: trichogyne. Scale bars: 20mm.



Figs 25–33. Sarcodia montagneana. Postfertilization stages. Figs 25–29, 31–33: haematoxylin; Fig. 30: aniline blue. 25.Supporting cell, sterile side branch and carpogonial remnant. The enlarged auxiliary cell has cut off the first gonimoblastcell (WELT A030191). 26–28. Two-celled stages in gonimoblast development from the auxiliary cell (WELT A030870). 29.

Early stage in cystocarp development showing differentiation of the darkly staining inner pericarp (inside circle) from outerpericarp (WELT A030191). 30. Initiation of the central cavity above the developing gonimoblasts, and separation of the innerand outer pericarp regions with some cells elongating (arrow) between the two (WELT A030191). 31. Radiation of gonimo-

blast filaments at the centre of inner pericarp and gonimoblast expansion into central cavity (HGI-A 9545). 32. Cystocarp withostiole and differentiation of gonimoblasts into an inner gonimoblast reticulum and outer covering of carposporangia-bearingfilaments. Cells adjacent to and above the inner pericarp link to inner pericarp cells without exchanging nuclei (arrow) (HGI-A

9545). 33. Organization of gonimoblast reticulum composed of fused uninucleate cells and transition to outer carposporangia-bearing layers (HGI-A 9545). Abbreviations: ac: auxiliary cell; c: carposporangia; cp: carpogonium; g1: gonimoblast initial;g2: gonimoblast cell developed from g1; gf: gonimoblast filaments; gr: gonimoblast reticulum; ip: inner pericarp; n: nucleus; o:

ostiole; op: outer pericarp; sb: sterile branches; sc: supporting cell. Scale bars: 20mm.



Cystocarps are sessile and hemispherical. Theyinitially form along the margins but progressively

become scattered over both surfaces of the blade

(Figs 2, 4, 5, 34). An ostiole forms apically in the

outer pericarp (Figs 32, 35, 36) and is a translucent

rose colour, through which the enclosed deep-red

gonimoblasts can be seen in both fresh and dried

specimens (Fig. 34). The outer pericarp consists of

a compact surface layer of small ovoid or slightly

elongated uninucleate cells that are not linked by

secondary pit connections (Fig. 37) and an inner

cortex of larger multinucleate cells that are second-

arily pit-connected (Fig. 37, arrow). The subcortex

of the outer pericarp is lax, with rounded or ovoid

cells that are more elongated at the base of

the cystocarp, whereas the cells surrounding the

gonimoblasts are stellate and more compact(Fig. 38). Mature carposporangia are uninucleateand 18–30 mm in diameter (Fig. 39). Theyoccasionally germinate in situ (Fig. 40, arrow).

Tetrasporangia: Tetrasporangia are scattered overthe thallus surface. They are not produced simulta-neously but differentiate continuously and tetra-sporangial initials can form adjacent to maturetetrasporangia. Tetrasporangial initials originatelaterally from a subcortical cell (Fig. 41).Tetrasporangia are ovoid when young and elon-gated and enlarged with age (Figs 41–44), with thepit connection between the tetrasporangium andthe subcortical cell situated close to the base of thetetrasporangium (Figs 41, 42, arrows), flanked byelongated cortical filaments (Fig. 44, arrowheads).

Figs 34–44. Sarcodia montagneana. Postfertilization stages and tetrasporangia development. Figs 35–37, 39, 41–44: haema-

toxylin; Figs 38, 40: aniline blue. 34. Portion of a fresh specimen with cystocarps borne along margins and over thallus surface(HGI-A 9545). 35–36. Detail of the ostiole in surface view and in cross section (WELT A030191). 37. Surface and inner cells ofthe outer pericarp (HGI-A 9545). 38. Mature outer pericarp showing the inner layer of slightly stellate cells (arrow) surround-

ing cystocarp cavity (HGI-A 9545). 39. Uninucleate mature carposporangia (HGI-A 9545). 40. Group of carposporangia inwhich one of them is germinating in situ (arrow) (WELT A030191). 41–44. Development of the tetrasporangial initials andzonately dividing tetrasporangia. Note that each tetrasporangium is pit-connected basally with a subcortical cell (arrows), and

that the mature tetrasporangia are surrounded by elongated cortical filaments (arrowheads) (WELT A030196). Abbreviation:n: nucleus. Scale bars: 20mm (Figs 35–37, 39–44), 1 cm (Fig. 34) or 100 mm (Fig. 38).



Mature tetrasporangia are zonately divided and upto 48mm broad by 78mm long (Fig. 44).

Sarcodia grandifolia Levring 1949, p. 395, pl. 42,

fig. 3

(Figs 45–84)HETEROTYPIC SYNONYM: Sarcodia flabellataLevring, p. 396, pl. 42, fig. 4.

LECTOTYPE: Specimen collected in Stewart Island(New Zealand), 24 June 1946, WELT A022966(VWL No. 7585) (Nelson & Phillips, 2001).Collector unnamed but probably Mrs EileenWilla (Fig. 45).ISOTYPE: WELT A022967.TYPE LOCALITY: Stewart Island, New Zealand.DISTRIBUTION: New Zealand: North Island (shoresof Cook Strait), South Island (Marlborough,

Figs 45–54. Sarcodia grandifolia, habit and vegetative development. Figs 48, 51–53: aniline blue; Figs 49, 50, 54: haematoxylin.45. Lectotype, Levring No. 7585, WELT A022926, Stewart Island, New Zealand, V.W. Lindauer No. 7585, 24 June 1946. 46.A broad, twice-forked female plant, The Neck, beach facing Cow Island, Stewart Island, J.F. Herrick, 07 October 1994

(AK220918). 47. Female plant. Attached, lower littoral, Princess Bay, Wellington, Auckland, New Zealand, coll. D.W.Freshwater (NCU 591801, upper left). 48. Longitudinal section through tip (NCU 591799-802). 49. Surface layer and outercortex in cross section (NCU 591803-811). 50. External view of uninucleate surface cells with parietal plastids (NCU 591799-

802). 51. Multinucleate inner cortical cell with dissected ribbon-shaped plastid (NCU 591803-811). 52. Stellate medullary cellwith a dissected ribbon-shaped plastid (NCU 591799-802). 53. Cross section of a thick thallus showing denselypacked secondary medullary rhizoidal filaments filling the medulla (NCU 591803-811). 54. Longitudinal section showing avacuolated multinucleate stellate cell on right and multinucleate secondary medullary rhizoidal filaments on left (NCU

591799-802). Abbreviations: mc: medullary cell; n: nucleus; p: plastid. Scale bars: 5 cm (Figs 45–47), 200mm (Fig. 48) or20mm (Figs 49–54).



Figs 55–67. Sarcodia grandifolia. Male and young female reproductive stages. Figs 55, 56, 58, 59: aniline blue; Figs 57, 60–67:

haematoxylin. 55. Transverse section of a mature spermatangial sorus (WELT A029965). 56. Developing elongated sperma-tangial parent cells forming tubes (WELT A029965). 57. As above, with differentiating spermatia inside spermatangial parentcells which function as spermatangia (WELT A029965). 58. Seriate spermatangia (WELT A029965). 59. Mature spermatan-

gial sorus showing elongated spermatangial parent cells, seriate spermatangia, terminal spermatia and released spermatia(WELT A029965). 60. Uninucleate supporting cell bearing a pair of one-celled, uninucleate sterile branches and a youngcarpogonium (NCU 591803-811). 61. Supporting cell bearing a carpogonium flanked by a pair of two-celled sterile branches(NCU 591803-811). 62. Supporting cell bearing an elongated carpogonium and trichogyne that has cut off a hypogynous cell

(NCU 591803-811). 63. Supporting cell bearing a carpogonium and hypogynous cell in which the basal cell of the adjacentsterile branch has differentiated into an auxiliary cell (NCU 591803-811). 64. Supporting cell bearing a carpogonium andhypogynous cell, in which the hypogynous cell has fused with the adjacent auxiliary cell (NCU 591803-811). 65. As above, but

with the carpogonium bearing a remnant trichogyne (NCU 591803-811). 66. Auxiliary cell situated at the base of a side branchhas cut off the first gonimoblast cell laterally which, in turn, bears two additional gonimoblast cells in a row (NCU 591803-811). 67. As above, with the auxiliary cell and gonimoblast cells seen in face view (NCU 591803-811). Abbreviations: ac:

auxiliary cell; cp: carpogonium; g: gonimoblast; g1: gonimoblast initial; g2 and g3: second and third gonimoblast cells; hy:hypogynous cell; n: nucleus; s: spermatium; sb: sterile branches; sc: supporting cell; sg: spermatangium; spc: spermatangialparent cell; t: trichogyne. Scale bars: 20mm.



Otago, Fiordland), Stewart Island and SnaresIslands (Fig. 1).HABITAT: Intertidal up to �25m depth collected allyear round.SPECIMENS EXAMINED: North Island, New Zealand:

Wellington, Lyall Bay, 40�200S, 174�480E (M.H.Hommersand, 19 September 1974, NCU 591803-811, female, tetrasporangial); Wellington, WestLyall Bay, 41�210S, 174�480E, low intertidal onrock, partially buried in gravel (W.A. Nelson, 07December 2009, WELT A030183, tetrasporangial);Wellington, Princess Bay, 41�200S, 174�470E,attached at low tide (D.W. Freshwater NZ04-559,17 November 2005, NCU 591799-802, female);Wellington, Island Bay, Taputerenga Island,41�210S, 174�460E, low water (N.M. Adams, 29September 1976, WELT A009499, female, male).South Island, New Zealand: Shag Point, 45�290S,170�490E, attached (D.W. Freshwater, NZ04-056,24 October 2004, NCU 591798, female);Fiordland, Chalky Inlet, Passage I. (SE point),46�200 S, 166�320E, �12 to �15m (C.H. Hay, 23April 1991, WELT A022576, female). Stewart

Island, New Zealand: Lee Bay, 46�520S, 168�070E,�1.5 to �3.5m (D.W. Freshwater NZ04-186, 30October 2004, NCU591797, female); HalfmoonBay, Lonneker’s Nugget, 46�540S, 168�80E, lowintertidal (W.A. Nelson, 07 September 1990,WELT A029965, male); Port Pegasus, BlindPassage, 47�130S, 167�400E (20 August 1946,WELT A001089 (¼ANZE 289), female); TheNeck, facing Cow Island, 46�570S, 168�100E, lowintertidal (J.F. Herrick, 07 October 1994,AK220918, female); Halfmoon Bay, HarroldsBay, 46�530S, 168�900E (A.L. Stewart & C.D.Roberts, 04 March 1992, WELT A029918, tetra-sporangial). Snares Islands, New Zealand: NWcorner of HoHo Bay, 48�10S, 166�370E, �22m(G.D. Fenwick, 22 February 1977, WELTA009701, female).

Habit and vegetative structure: Thallus erect,flattened, up to 19 (–31) cm in length, attachedto the substratum by a discoid holdfast, shortlystipitate, complanate, subdichotomously branchedand with rounded axils and apices (Figs 45–47).Axis up to 4 (–7.5) cm wide. The lectotype(Fig. 45) is a cystocarpic plant that is erodedtowards the base and has regenerated some 20 orso new branches. Plants that are sparingly andshallowly (Fig. 46) or deeply dichotomouslybranched (Fig. 47) have commonly been referredto S. montagneana or S. flabellata in variousherbaria and publications (Levring, 1949;Adams, 1994; Miller, 2003). Margins smooth orsometimes proliferous in older individuals.Texture moderately firm, the colour variable butoften deep red.

Structure multiaxial with abundant intercellularmedullary rhizoidal filaments in mature plants(Fig. 48). Outer cortex composed of anticlinal,dichotomously branched filaments, usually twocells long and lacking secondary pit connections(Fig. 49). Surface cells up to 12 (–19) mm highand 8 (–12)mm in diameter, rectangular in trans-verse section and rounded to polygonal in surfaceview, uninucleate, and with a parietal plastid(Figs 49, 50). Inner cortex multilayered, the cellsconnected to neighbouring cells by secondary pitconnections, the outermost ovoid and small,increasing in size inwardly and becoming stellate,with the largest reaching 65 mm in diameter (Figs51, 52). Subcortical cells multinucleate with theinnermost containing dissected ribbon-like plastids(Fig. 51). Medulla consisting of multinucleate stel-late cells with dissected ribbon-like plastids inwhich the rounded central portion is up to 40mmin diameter (Figs 52, 54). An abundance ofentangled slender rhizoidal filaments is usually pre-sent, some of which are clearly longitudinally ori-entated (Figs 48, 53, 54). The medullary rhizoidalfilaments originate near the apex but are of second-ary origin, arising from inner cortical and possiblystellate medullary cells.

Life cycle: Triphasic, with isomorphic gameto-phytes and tetrasporophytes. Gametophytesdioecious.

Male reproductive structures: Spermatangiadevelop in large, pale sori on both sides of the thal-lus (Fig. 55), the earliest stages consisting of sper-matangial parent cells which form elongate tubesthat extend to the thallus surface (Figs 56, 57).Such cells may function initially as spermatangiathat form a single spermatium inside the tube. Theupper part of the tube fuses with the thallus surfaceand releases the uninucleate spermatium. Inmature sori a row of rectangular spermatangiaappears to differentiate serially inside the tubularspermatangial parent cells (Fig. 58). The walls ofmature terminal spermatangia dissolve and the lib-erated spermatia appear rounded and vacuolate atthe ends of the rows, each with a terminal nucleusand proximal vesicle (Fig. 59).

Female reproductive structures: Immature femalereproductive structures develop from darkly stain-ing cortical cells that undergo divisions to producea basal intercalary cell (¼the supporting cell) andtwo uninucleate sterile branches. Subsequently,the supporting cell cuts off a carpogonium to theoutside, flanked by the sterile branches (Figs 60,61). These remain pit-connected to the supportingcell and are initially one-celled but later becometwo-celled. Both the supporting cell and the car-pogonium are uninucleate at this stage (Fig. 61).



The carpogonium expands, becomes conical, andinitiates a straight trichogyne that elongates to thethallus surface. The carpogonium divides after pre-sumed fertilization and gives rise to a hypogynouscell (Fig. 62). At the same time, the basal cell of oneof the sterile branches borne on the supportingcell differentiates into an auxiliary cell (Fig. 63).

The hypogynous cell fuses with the auxiliary cell(Figs 64, 65) and deposits a conspicuous nucleusinside it (Fig. 65). At this stage the remnant carpo-gonium and trichogyne can occasionally still beseen (Fig. 65). Finally, the carpogonial remnantdegenerates and the auxiliary cell that has fusedwith the hypogynous cell divides to form a

Figs 68–76. Sarcodia grandifolia. Development of cystocarp. Haematoxylin. 68. Young cystocarp showing inner and outerpericarp composed of longitudinal files of cells. The auxiliary cell sits at the base of the gonimoblasts flanked by darkly

staining inner pericarp filaments (NCU 591799-802). 69. Enlarged view of the inner pericarp. A uninucleate gonimoblast cell(arrow) is prominent inside the inner pericarp (NCU 591803-811). 70. Developing cystocarp showing gonimoblasts filling thecavity between the inner and outer pericarps. Some inner cells of the gonimoblast reticulum have linked up with cells of the

inner pericarp (arrows) (NCU 591799-802). 71. As above, but a later stage in which the outer gonimoblasts have producedterminal chains of carposporangia (NCU 591803-811). 72. Enlarged view of inner pericarp and inner gonimoblast reticulumshowing the linkage between the two (arrows) (NCU 591803-811). 73. Inner gonimoblast reticulum at higher magnification

(NCU 591799-802). 74. Transition between the inner gonimoblast reticulum and outer gonimoblast files that will bear thecarposporangia (NCU 591799-802). 75. Segment of a fresh specimen showing cystocarps scattered over thallus surface andalong margins (NCU 591803-811). 76. Surface view of an ostiole (NCU 591799-802). Abbreviations: ac: auxiliary cell; c:carposporangia; gf: gonimoblast filaments; ip: inner pericarp; n: nucleus; op: outer pericarp; gr: gonimoblast reticulum. Scale

bars: 20mm (Figs 67–69, 72–74, 76) or 200mm (Figs 70, 71).



gonimoblast initial, which in turn gives rise to a fileof gonimoblast cells (Figs 66, 67).Cystocarp development is initiated when cells of

the outer cortex divide to form files of aligned cellsthat differentiate into inner and outer pericarps(Figs 68). Occasionally, a gonimoblast cell maybe distinguished from inner pericarp cells by thepresence of a single large nucleus (Fig. 69,arrow). A central cavity is initiated by the separa-tion of outer and inner pericarps that soonbecomes filled with highly branched gonimoblastfilaments (Figs 70, 71). The inner gonimoblastscells unite laterally to form a gonimoblast reticu-lum (Fig. 72), which expands into a conspicuousnetwork inside the central cavity (Figs 72, 73),whereas the outer gonimoblasts consist ofbranched files of gonimoblast filaments (Fig. 74)that terminate in short chains of carposporangia(Fig. 70). The innermost cells of the gonimoblastreticulum may fuse with the outermost cells of theinner pericarp (Figs 70, 71, arrows). Mature goni-moblasts reach 600 mm in diameter (Figs 70, 71).Cystocarps are hemispherical and sessile, and are

borne along the margins and scattered over bothsides of the thallus surface (Figs 46, 47, 75); they

possess a small terminal ostiole (Fig. 76). The outer

pericarp and gonimoblasts are dark red in both

fresh and dried specimens (Fig. 75, seen as

black). The outer pericarp consists of a compact

outer cortex composed of small ovoid or slightly

elongated uninucleate cells that are not connected

by secondary pit connections, and a subcortex of

larger multinucleate cells connected by secondary

pit connections (Fig. 77). Subcortical cells are

rounded or ovoid (Figs 77, 78) becoming elongate

at the base of the cystocarp (Fig. 79). The cells

adjacent to the gonimoblasts are slightly stellate

and more compact (Fig. 78). Mature carposporan-

gia are uninucleate and 28–50 mm in diameter

(Fig. 80). A few were occasionally seen germinating

in situ (Figs 81, 82).

Tetrasporangia: Tetrasporangia are scattered overthe thallus surface and are produced more or lesscontinuously with sporangia of different ages adja-cent to one another. Tetrasporangial initialsdevelop laterally from subcortical cells and arebasally pit-connected to them (Figs 83, 84,arrows). Tetrasporangia are ovoid when youngand later become broadly elongated and zonately

Figs 77–84. Sarcodia grandifolia. Development of cystocarp and tetrasporangia. Haematoxylin. 77. Surface layer and innercells of the outer pericarp (NCU 591799-802). 78. Mature cystocarp showing the slightly stellate cells (arrow) in the outer

pericarp surrounding the gonimoblasts (NCU 591799-802). 79. Basal part of a cystocarp showing the relationships betweeninner and outer pericarp and the gonimoblast filaments (arrow) (NCU 591803-811). 80. Mature uninucleate carposporangia(NCU 591799-802). 81. Carposporangia and in situ germinating carposporangia (arrows) (NCU 591803-810). 82. In situ

germinating carposporangia (NCU 591803-811). 83. Tetrasporangium linked by a basal pit connection (arrow) to a subcorticalcell (WELT A030183). 84. Mature zonately divided tetrasporangium pit-connected basally (arrow) to a subcortical cell andflanked by elongated cortical filaments (arrowheads) (WELT A030183). Abbreviations: c: carposporangia; ip: inner pericarp;

n: nucleus; gr: gonimoblast reticulum. Scale bars: 20mm (Figs 77, 78, 81–84), 50 mm (Fig. 79) or 10 mm (Fig. 80).



divided, flanked by sterile branches (Fig. 84,arrowheads). Mature tetrasporangia are up to55mm broad and 75 mm long (Fig. 84).

Molecular analyses

Thirteen rbcL sequences were newly generated forspecies of Sarcodia, along with five sequences avail-able from GenBank (Table 1). A species ofPlocamium and one of Trematocarpus were selectedas the outgroup. The data matrix analysed included1388 base pairs with 120 parsimony-informativesites. The topologies of the ML, MP and BA treeswere largely congruent and only theML rbcL tree isshown (Fig. 85). The species of Sarcodia that wereanalysed formed a well-supported clade. Two col-lections of Sarcodia (sp. 1 and sp. 2) from Bali,Indonesia, two of Sarcodia sp. from Taiwan, S. den-tata (Suhr) R.E. Norris from SouthAfrica and threecollections of S. ceylanica Harvey ex Kutzing from

Sri Lanka clustered together with 100% support.On the other hand, the two samples of S. montag-neana, the generitype fromNewZealand, are closelyrelated to S. marginata J. Agardh and Sarcodia sp.from Australia and are only distantly related to thefive samples of S. grandifolia. The rbcL sequences ofS. ‘flabellata’ (see Table 1) were nearly identical tothose of S. grandifolia and the former is treated as asynonym of the latter in the rbcL phylogenetictree (Fig. 85).

Discussion

Sarcodia montagneana is currently treated as awidespread, morphologically variable species thatis distributed throughout the Indo-West PacificOcean (Silva et al., 1996; Guiry & Guiry, 2011).In reality, the type species of Sarcodia,S. montagneana, appears to be limited in its distri-bution to the northern part of the North Island ofNew Zealand (Fig. 1). Our molecular data (Fig. 85)

Fig. 85. RbcL phylogeny: ML tree (–lnL¼ 9560.8708) of the genus Sarcodia from the Indian and Pacific Oceans. Numbersabove branches are ML and MP bootstrap values, respectively, whereas numbers below branches are Bayesian posteriorprobabilities in %. Asterisks indicate support values >99%.



place S. montagneana closest to S. marginata fromsouthern Australia, which is a subdichotomouslybranched species, and to a small, undescribed spe-cies from New South Wales, Australia. Perhaps itmay also be related to the little known species, S.marginalis (Kutzing) Millar, in Millar &Prud’homme van Reine (2005) from NewCaledonia. Thus, the forms that superficiallyresemble S. montagneana are not the ones mostclosely related to it. A second species, S. grandifoliaLevring (1949) is widely distributed, occurring inthe Wellington area around Cook Strait in theNorth Island and throughout the South Island,Stewart Island and the Snares Islands. A third spe-cies, S. flabellata, was also described by Levring(1949) from New Zealand, but, in our opinion,this species is a smaller, variably branched flabel-late form of S. grandifolia, as suggested previouslyby Adams (1994). Indeed, Levring (1949) alreadystated that S. grandifolia is variable in shape, withyounger specimens clearly flabellate and larger spe-cimens with flabellately divided branches and anattenuated base. The New Zealand species ofSarcodia have long been confused due to their sim-ilarity in habit and gross morphology. Vegetativeplants can be readily distinguished in cross sectionby the form and height of their outer cortical cells,which are rounded or ovoid and up to 9 (–14) mmin length in S. montagneana (Fig. 9), and rectangu-lar up to 12 (–19) mm in length in S. grandifolia(Fig. 49). The most conspicuous character distin-guishing the two is the colour of the gonimoblastsand outer pericarp as seen in surface view in bothfresh and dried specimens. The gonimoblasts areconspicuously red inside a transparent rosy outerpericarp in S. montagneana, as reported in the orig-inal description by Hooker & Harvey (1845),whereas in S. grandifolia both the gonimoblastsand the outer pericarp are dark red and indistin-guishable in surface view. Other taxonomic char-acters have been suggested to differentiate the taxa,such as the length of adult specimens (Levring,1949; Adams, 1994) and the presence of stalkedcystocarps (Adams, 1994), but these were foundto be unreliable in the present study.Spermatangia have been described as developing

in surface patches in S. montagneana (Adams,1994; Liao et al., 1993) and S. dentata (Norris,1987), whereas those of S. grandifolia wereunknown. We observed that the development ofthe spermatangial sori is initiated by the differen-tiation of the outer cortical cells into spermatangialparent cells with tubular extensions that reachthe thallus surface. Paraphyses are absent.Spermatangial parent cells are initially non-septateand function as spermatangia, cutting off a singlespermatium. A mature spermatium contains aterminal nucleus and a proximal vesicle and is

released through a pore formed by the interactionof the spermatangial tube with the outer wall andcuticle. Subsequently the spermatangial parentcells produce spermatangia in rows, the terminalones differentiating into spermatia that are laterreleased. Similarly, Searles reported the presenceof spermatangia cut off in two-celled chains fromelongated spermatangial parent cells inTrematocarpus flabellatus (J. Agardh) De Toni[Searles, 1968, as Dicurella scutellata (Hering)Papenfuss], and Womersley (1994) recorded sper-matia in chains in T. concinnus (R. Brown exTurner) De Toni. On the other hand, Liao et al.(1993) reported that the spermatangia are formedsingly from surface cortical cells inS. montagneana.The development and structure of the female

apparatus is quite similar in S. montagneana andS. grandifolia and differs significantly from thedescriptions by Rasmussen (1964) and Liao et al.(1993), the only authors to have investigatedfemale development in detail. Rasmussen (1964)reported the presence of polycarpogonial systemscomposed of three-celled carpogonial branches andflattened trichogynes in S. montagneana, whereasLiao et al. (1993) reported monocarpogonial sys-tems with three-celled carpogonial branches in thesame species. Norris (1987) described a monocar-pogonial system in S. dentata in which the carpo-gonial branch was two or possibly three cells long.Three-celled carpogonial branches were illustratedby Searles (1968) in Trematocarpus fragilis (asDicurella fragilis) and T. flabellatus (as Dicurellascutellata) and in T. dichotomus (Searles, 1969)and were confirmed by Barrientos & Alveal(2005) and by one of us (Rodrıguez-Prieto, pers.observ.) for the Chilean T. dichotomus. The pub-lished evidence thus suggests that a three-celledrecurved carpogonial branch is the usual conditionin most members of the Sarcodiaceae.Our observations of a one-celled carpogonium

cut off terminally from the supporting cell beforefertilization in two New Zealand species is in sharpcontrast to the expected behaviour. We never sawthree-celled carpogonial branches in our materialand the few extended trichogynes we saw weretubular. If we interpreted our observations cor-rectly, then the Sarcodiaceae have evolved asecond pathway for the development of thefemale reproductive system, something that isvirtually unheard-of in the Rhodophyta. All ofour evidence hinges on relating the one-celled ter-minal carpogonium to postfertilization events. Weobserved that the carpogonium cuts off a hypogy-nous cell basally after fertilization at the same timethat the basal cell of one of the side branches differ-entiates into an auxiliary cell, and that this cell cutsoff a gonimoblast initial obliquely or laterally that



gives rise to an initially filamentous gonimoblastfollowed by the elaboration of a massive cystocarp.Fusion between the hypogynous cell and the auxil-iary cell took place directly indicating that thefemale reproductive apparatus is procarpic. Wedid not see a connecting filament linking the hypog-ynous cell with a nearby cortical cell functioning asan auxiliary cell, as reported by Rasmussen (1964)in S. montagneana or by Norris (1987) working onSouth African S. dentata. Norris concluded thatS. dentata was non-procarpic with the auxiliarycell being a neighbouring cell identical in size andposition to the supporting cell. The validity of ourcontrasting interpretation of postfertilizationevents in New Zealand species of Sarcodia hingeson the correctness of our observations of thesequence of nuclear events leading to gonimoblastformation. These should be examined again in NewZealand material and in other species of Sarcodia.Later stages in the development of the cystocarp

are not so controversial. Soon after gonimoblastinitiation, the surrounding cortical cells initiateparallel files of cells that form the pericarp. Cellsimmediately below the gonimoblast filaments aredensely filled with cytoplasm and darkly staining.We referred to these darkly staining inner files ofcells as the inner pericarp. Files of cells above thisarea were lightly staining and produced the outerpericarp and a central ostiole. Some of the fila-ments between the two regions elongated or sepa-rated to initiate the central cavity, which in turnwas filled with branched gonimoblast filaments.Gonimoblast cells were initially uninucleate andcontained enlarged nuclei making them readily dis-tinguishable from both inner and outer pericarpcells. The gonimoblast filaments differentiatedinto inner and outer gonimoblast regions, withthe inner gonimoblast filaments fusing laterally toform a gonimoblast reticulum in which the fusedcells were multinucleate, whereas the outer fila-ments grew linearly in branched files that ulti-mately bore short chains of uninucleatecarposporangia. As Norris (1987) noted, gonimo-blast cells and cells of the inner pericarp interact.Our observations confirm that cells of the gonimo-blast reticulum do attach to the outermost cells ofthe inner pericarp filaments. The fusion that takesplace does not seem to involve an exchange ofnuclei between the two regions, inasmuch asnuclei of different sizes were never seen in thesame cell. Where we have called the basal filaments‘inner pericarp’, Norris (1987) referred to them as‘a ring or partial platform of large, deeply stainingcells originated from the vegetative cells.’ Kylin(1956, in Sarcodia), Norris (1987, in S. dentata)and Womersley (1994, in S. marginata) alsoreported the presence of a reticulate fusion cell atthe base of the gonimoblasts. Outer gonimoblast

filaments do not differentiate cells or filaments thatconnect to the outer pericarp, as occurs inGracilaria and some other genera in theGracilariaceae. The similarity between the out-wardly developed cystocarp with a central ostioleand the structure seen in most Gracilariaceae is aparallel development and the two groups show nophylogenetic relationship.Molecular studies suggest a weak relationship

among the families Plocamiaceae, Pseudoanem-oniaceae and Sarcodiaceae (Saunders et al.,2004). All three are procarpic, with the carpogo-nium situated adjacent to the auxiliary cell, and allthree produce outwardly developed gonimoblastssurrounded by an external pericarp with a centralostiole. The supporting cell functions as the auxil-iary cell in Plocamium (Plocamiaceae) (Kylin, 1923;Hommersand & Fredericq, 1990) and Hummbrella(Pseudoanemoniaceae) (Hawkes & Johnson, 1981)and is thought to serve the same function in thesarcodiacean genus Trematocarpus (Searles, 1968,1969). These similarities need to be reinvestigatedin Trematocarpus and clarified more fully in theother two families. Sarcodia is multiaxial with anelaborate, highly differentiated cystocarp, whereasPlocamium and Hummbrella are uniaxial withsimple filamentous organization and a much sim-pler cystocarp. All three are ancient groups withfew genera that are widely separated phylogeneti-cally. The basis for the ancestral relationshipamong them remains to be determined. The studyby Norris (1987) suggests that important charac-ters may be discovered that distinguish the speciesof Sarcodia. The present work may be sufficientlydetailed to permit new comparisons to be madebetween species and genera. Likewise, further stud-ies of the Sarcodiaceae, the Plocamiaceae and thePseudoanemoniaceae may illuminate a basis forevaluating the developmental and phylogeneticsimilarities and differences among these threeseemingly related families.

Acknowledgements

We thank Roberta D’Archino for generously collect-ing specimens of Sarcodia montagneana. Scans of spe-cimens in the type collection at BM were kindlyprovided by J. Wilbraham and from TCD byJ. Parnell. We thank Sarah Pene Eftonga for addi-tional photographs of W.H. Harvey’s collection. Wethank Dr D.W. Freshwater and Dr G. Kraft for theirreviews and positive criticisms, which have improvedthe quality of the article. This project was supportedby two grants from the Spanish Ministry of Scienceand Technology (CGL2004-05556-C02-01 andCGL2008-00932); by grants from Taiwan’s NationalScience Council (NSC 99-2621-B-019-003-MY3)and NTOU’s Center of Excellence for Marine



Bioenvironment and Biotechnology (99529001G) toS.-M. Lin; by Foundation for Research Science &Technology contract C01X0502 (New Zealand)grant to W.A. Nelson, and by NSF grantDEB0937978 to J.M. Lopez Bautista, M.H.Hommersand and S. Fredericq.

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