Phylogenetic Relationships Among Basal-Most Arthrodontous Mosses with Special Emphasis on the Evolutionary Signiﬁcance of the Funariineae

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The Bryologist 103(2), pp. 212–223 Copyright ᭧ 2000 by the American Bryological and Lichenological Society, Inc. View metadata, citation and similar papers at core.ac.uk brought to you by CORE

provided by Sapientia Phylogenetic Relationships Among Basal-most Arthrodontous Mosses with Special Emphasis on the Evolutionary Signiﬁcance of the Funariineae

BERNARD GOFFINET Department of Botany, Box 90339, Duke University, Durham, NC 27708-0339, U.S.A. Current address: Department of Ecology and Evolutionary Biology U-43, University of Connecticut, Storrs, CT 06269-3043, U.S.A.

CYMON J. COX Department of Botany, The Natural History Museum, Cromwell Road, London, England, SW7 5BD, U.K. Current address: Department of Botany, Box 90339, Duke University, Durham, NC 27708-0339, U.S.A.

Abstract. The classification of the Bryopsida (mosses) has been based primarily on the variation of sporophytic characters i.e., architectural features of the peristome teeth that line the capsule mouth. Five arthrodontous peristome types have been recognized. Whether peristome types define natural groups and how they are evolutionary related has, however, remained unclear. Nucleotide sequence data from one nuclear and two chloroplast loci are generated and compiled to test two contrasting hypotheses regarding the ancestral peristome type in the Arthrodonteae. The genomic data partitions are incongruent with regard to the phylogenetic signal they carry. All phylogenetic analyses converge toward the polyphyly of the Funariineae and the Funariaceae. The Funariaceae are defined by the loss of a codon in the rps4 gene. Goniomitrium acuminatum, the type of the genus, lacks this deletion, and is always resolved within the Haplolepideae. Con- sequently Goniomitrium is transferred to the Pottiaceae. The Ephemeraceae and Splachnobry- aceae are tentatively retained as distinct, but with strong affinities to the Pottiineae. Neither the combined nor the independent data sets yield well supported topologies under the parsimony optimality criterion. Hence, the relationships among major lineages remain ambiguous. Inferences from chloroplast data alone yield a basal dichotomy between taxa with alternate peristomes (Orthotrichales and Bryales sensu lato) and those with opposite peristomes (Encalyptineae, Dis- celiaceae, Funariaceae, Timmiaceae, and the Haplolepideae). In contrast, analyses of the combined data resolve the Timmiaceae as sister to the split between the two peristomial lineages. It is hypothesized that the symmetric divisions of the IPL cells, is a synapomorphy for at least the Encalyptineae-Funariineae clade. The endostomial appendages of the timmiaceous peristome could, under either phylogenetic hypothesis, be regarded as homologous to the cilia in the bryoid peristome. Although the relationships among major lineages of arthrodontous mosses remain ambiguous, this study suggests that taxa with reduced or no peristomes, such as the Disceliaceae and the Gigaspermaceae, may be crucial in resolving the early evolutionary history of the Ar- throdonteae when using DNA sequences.

Arthrodontous mosses are deﬁned by the archi- remnants of two columns of cells (hence the name tecture of their peristome teeth. In contrast to ne- diplolepideous). In the Funaria-type peristome, the matodontous mosses (the Polytrichales and Tetra- exostome teeth lie opposite the endostome seg- phidales) whose teeth are solid, and built from ments. By contrast, in the Orthotrichum- and whole cells, the teeth that line the mouth of arthro- Bryum-types, the outer teeth alternate with the in- dontous mosses, are composed of cell wall rem- ner segments. The latter type is further unique by nants, and in particular of periclinal cell plates. The the presence of endostomial cilia between the seg- architecture of the peristome teeth varies among ments, a character correlated to the presence of ad- major lineages of arthrodontous mosses. Vitt (1981) ditional cells in the inner most peristomial layer recognized four main peristome types that differ in (IPL). These architectural differences observed at their basic architecture at maturity. The haplolepi- maturity are complemented by ontogenetic varia- deous peristome or Dicranum-type peristome is tions in the type of cell divisions in the IPL. In the typically reduced to an endostome of 16 teeth, Funaria-type peristome, the divisions that lead to a whose inner surface is built from one and a half doubling of the number of IPL cells from eight to columns of cells. The remaining three types are 16 are all symmetric, and the new anticlinal walls diplolepideous, that is they are typically composed are perfectly aligned, with the anticlinal walls of of two rows of teeth, with the outer teeth bearing the adjacent primary peristomial layer (PPL – Shaw

0007-2745/00/212–223$1.35/0 2000] GOFFINET & COX: ARTHRODONTOUS MOSSES 213 et al. 1989a), whereas in the remaining three types, dating the relationships among major lineages of these divisions are asymmetric, and hence the new the Bryopsida (sensu Vitt 1984). wall, is not aligned with the adjacent PPL wall The Funariineae sensu Vitt (1984) comprise five (Goffinet et al. 1999; Shaw et al. 1989a,b). Vitt families, namely the Funariaceae, Pseudoditricha- (1984) later also considered the peristome of the ceae, Disceliaceae, Gigaspermaceae, and Ephem- Encalyptineae to be distinct enough to warrant its eraceae. Shaw (1984) excluded the monospecific recognition as a distinct architectural type. In the Pseudoditrichaceae and transferred it near the Bry- Encalyptineae the teeth and segments appear op- aceae (Bryineae) based on peristomial characters. posite as in Funaria and in the Haplolepideae (see The Gigaspermaceae and the Ephemeraceae are Vitt et al. 1998), but the exostome can also bear composed of gymnostomous taxa only. The peri- intermediate teeth alternating with the endostomial stome of the Disceliaceae is reduced; the endosto- segments (Edwards 1984; Horton 1982). me consists of a thin transparent membrane, adher- The polarity of peristome-type evolution has re- ent to the exostome, whereas the exostome is well mained controversial, due to the uncertainty regard- developed and composed of sixteen teeth (Shaw & ing the sister-group to the Arthrodonteae, to the Allen 1985). The longitudinal anticlinal walls of the lack of a robust phylogeny for mosses, and to the IPL appear aligned with those of the adjacent PPL, difficulty of assessing homology among peristomial a feature present only in the Funariineae and the features (Goffinet et al. 1999; Vitt et al. 1998). Vitt Encalyptineae (Shaw & Allen 1985). In the Funar- (1981, 1984) proposed that the Funaria-type peri- iaceae, the peristome is either well developed, re- stome represents the most basic peristome architec- duced and simple, or lacking (Fife 1985). When ture from which other types are derived. By con- well developed, the peristome of the Funariaceae is trast, Shaw and Rohrer (1984) considered the peri- diplolepideous, with the endostome segments op- stome of Funaria to be the product of reduction posite or co-radially aligned to the exostome teeth. from a ciliate bryalean type peristome. The oppo- Cilia are always lacking and the peristome formula site arrangement of peristomes would consequently is typically 4:2:4, with the IPL lacking additional appear as a derived feature. Recent reconstructions cellular divisions, which are characteristic of typi- of the phylogeny of mosses from mitochondrial cal Bryales with cilia. The cells of the IPL are all (Beckert et al. 1999), chloroplast (Goffinet et al., similar, if not identical, in size as a consequence of unpubl.), and nuclear (Newton et al. 2000) se- the symmetric divisions (Shaw et al. 1989a). In the quence variations converge toward the Diphysci- absence of sporophytic features for most taxa, the aceae (i.e., Diphyscium Mohr, Muscoflorschutzia Funariineae have traditionally been defined by the Crosby, and Theriotia Card.) forming the sister- similarities in vegetative characters, such as smooth group to the Arthrodonteae. Because the IPL divi- cells, costate leaves, undifferentiated alar cells, and sion is asymmetric in Diphyscium, Goffinet et al. large calyptrae (Vitt 1982). Except for the Gigas- (1999) argued that this character should at present permaceae that are cladocarpous (LaFarge-England be considered plesiomorphic in the Arthrodontae, 1996), the Funariineae sensu Vitt are typical acro- rather than derived from an ancestral symmetric di- carpous taxa (Fife 1980). Koponen (1981) erected vision as suggested by Vitt (1981, 1984). This hy- the Splachnobryaceae, a small family restricted to pothesis does, however, not preclude an early origin the genus Splachnobryum C. Mu¨ll., and transferred of the Funariineae in the evolutionary history of the it from the Pottiineae to the Funariineae, based on Arthrodonteae. Analyses of rbcL sequences re- the diplolepideous rather than haplolepideous ar- solved the Funariaceae sister to a clade comprising chitecture of the peristome. Koponen (1981) further the Orthotrichineae, Dicranineae, Bryineae, and argued for Splachnobryum having a rather basal po- Splachnineae (Goffinet et al. 1998), with the En- sition among arthrodontous moss phylogeny, on the calyptineae sister to either this large clade or only basis of the lateral solitary gametangia. The peri- to the Dicranineae. Cox and Hedderson (1999), an- stome is, however, reduced to a single row of 16 alyzing a more extensive taxon sample using chlo- rather short teeth, whose architecture Allen and roplast and nuclear data, examined the relationships Pursell (2000) interpret as of the haplolepideous within the Arthrodonteae rooted to Funaria based type. The circumscription of the Funariidae (sensu on evidence from a broader analysis of 18S rDNA Vitt 1984), and the relationships among the families sequences (Hedderson et al. 1996, 1998). Their currently accepted within the order, have not been analyses resolve the Orthotrichineae and the critically examined using a phylogenetic approach. Splachnineae nested within the Bryineae, and the Assuming that Diphyscium and its close relatives Encalyptineae and Haplolepideae composing a bas- compose the closest outgroup to the Arthrodonteae, al grade. Furthermore, they indicated that the Tim- this study will address the circumscription of the miaceae, that were resolved in a nested position Funariineae, the relationships among its families, within this basal grade, may be crucial for eluci- and examine the significance of the Funaria-type 214 THE BRYOLOGIST [VOL. 103 peristome in the evolution of peristome types in the Fitch 1971) was chosen as the optimality criterion for the Arthrodonteae. Specifically, the following questions phylogenetic analyses. Parsimony analyses were performed with equal weighting of characters and transfor- will be addressed 1) are the Funariineae sensu Vitt mations. For each analysis searches were replicated 100 (1984) monophyletic? 2) what is the significance of times, with random sequence addition. The steepest de- the Funariineae in the early diversification of the scent option was selected. A partition homogeneity test Arthrodonteae? and 3) what are the trends in the was performed using the same set of options to examine whether phylogenetic signal carried by the nuclear and evolution of peristomial characters in the Arthro- chloroplast sequences are congruent or whether they re- donteae? cover distinct histories. Consistency indices (CI) and rescaled consistency indices (RC) were calculated with PAUP. Fifty thousand random trees were generated and MATERIAL AND METHODS the g1 (Huelsenbeck 1991) statistic describing the tree length frequency distribution was compute with Paup. Taxon sampling.—Exemplars of arthrodontous taxa po- Relative strength of support for particular branches was tentially representing early cladogenic events, with a main estimated using bootstrap analysis (Felsenstein 1985; Hil- emphasis on the Funariineae, sensu Vitt (1984) were sam- lis & Bull 1993) based on 100 bootstrap replicates of the pled (Table 1). Duplicate taxonomic sampling was per- heuristic search with the same set of options in effect as formed for selected taxa as primarily analyses progressed, above (except for 10 search replicates). Support for the to confirm sequences; duplicate taxa were retained even branches was evaluated by Bremer support analysis, using when the additional sequences diverged by a single nu- the program Autodecay (Eriksson 1998). cleotide. The matrix was completed by retrieving sequenc- Finally, constraint trees were employed to assess the es from GenBank, primarily for diplolepideous mosses effect of alternative phylogenetic hypotheses on the tree (Cox & Hedderson 1999). Vouchers are deposited in DUKE length. Tree scores of the constrained trees were compared unless otherwise indicated in Table 1. to that of unconstrained trees by non-parametric (Temple- DNA extraction, PCR amplification, and sequencing.— ton 1983) and parametric (Kishino & Hasegawa 1989) Apical portions of stems or branches, or in the case of testing. ephemeral taxa, operculate capsule(s), were removed from dried herbarium collections. DNA was extracted from plant tissues following the protocol outlined in Goffinet RESULTS et al. (1998). Plant material was ground using a small glass test tube, in 250 ␮L of 2X CTAB (hexadecyltrime- Sequence variation.—Sequences for the 18S thylamonium bromide)-0.2% beta-mercaptoethanol, heat- rRNA and rps4 genes and for the trnL-trnF region ed to 60ЊC, and incubated at this temperature for at least 30 min. An equal volume of chloroform-isoamyl (24:1) were assembled for 39 taxa. 18S rDNA sequences was added. The emulsified solution was centrifuged for were not obtained for Discelium nudum (Disceli- one min. at 6,500 rpm, and the aqueous phase was trans- aceae) and the taxon is therefore not included in ferred to a new tube to which an equal volume of ice cold the combined analysis. Eighty-two sequences were Њ isopropanol was added. DNA was precipitated at 4 C for generated in the course of this study (Table 1). 30–60 min. Tubes were centrifuged first for 10 min. at 13,000 rpm. The pellet was washed with 70% ethanol, and Alignment of sequences resulted in a matrix of the tubes centrifuged for three min at 13,000 rpm. The 3,430 characters of which 807 were excluded, ei- pellet was dried in a vacuum centrifuge and suspended in ther because these sites corresponded to the ampli- 100 ␮L TE (Tris-EDTA, pH 8.0). Protocols (of) for the fication primer sequences, or the homology as- amplification(s) of the trnL-trnF and rps4 regions as well as the sequencing of the fragments followed those pre- sumptions required for these sites were considered sented in Buck et al. (2000). Amplification and sequenc- ambiguous (the matrix is available from the senior ing of the 18S rRNA gene followed the protocol described author). The 2,623 characters included in the anal- in Cox et al. (2000). Labeled fragments yielded by the yses consisted of 1,740 nt from the18S rRNA gene, sequencing reactions were separated on polyacrylamide 570 nt of the rps4 gene and 313 nt of the trnL-trnF gels (Long Range Singel; FMC Bioproducts), using an ABI Prism௢ 373 or 377 automated DNA sequencer (Per- region. Approximately 13% (i.e., 331) of the sites kin Elmer). Nucleotide sequences were edited using Se- used, were parsimony informative, with the 18S quencher 3.0 (Gene Codes Corporation), entered in Paup rDNA accounting for 34% (i.e., 111 sites) of these version 4.0b2a (Swofford 1999) and manually aligned. sites, whereas the rps4 gene and the trnL-trnF re- The intron, exon, and spacer composing the trnL-trnF product were delimited by comparing the sequences with gion carried 149 (i.e., 45% of all informative sites) available GenBank accessions. Aligned trnL-trnF se- and 71 (i.e., 21%) informative sites, respectively. quences were trimmed of the 5’exon of the trnL, and the Although indels per se were not examined for their trnF exon (leaving the trnL intron, trnL-3Ј exon, and the phylogenetic signal, all members of the Funariaceae trnL-trnF spacer), while the first 27 sites (including the except Goniomitrium acuminatum shared the ab- annealing site of the primer) and the intergenic spacer following the stop codon were excluded from the rps4 sence of a codon in the rps4 gene. The distribution sequences. Sequences of the 18S rRNA gene were of the tree length of 50,000 randomly generated trimmed at both ends at sites corresponding to the ampli- trees from the data used here, was significantly left- fication primer annealing sites. All sequences obtained in skewed (p Ͻ 0.01), suggesting that the data set ap- this study were submitted to GenBank (Table 1). Phylogenetic analysis.—All analyses and tests were pears more structured than what would be expected performed using PAUP version 4.0b2a (Swofford 1999) from a random set of data, and thus that the data on a Macintosh G3 400 MHz. Maximum parsimony (MP, may carry the signal necessary for resolving the 2000] GOFFINET & COX: ARTHRODONTOUS MOSSES 215 phylogenetic history of the taxa (Hillis & Huelsen- clade, neither the Encalyptaceae, the Gigasperma- beck 1992). The partition homogeneity test indi- ceae, nor the Haplolepideae was defined as a mono- cates that the chloroplast and nuclear data partitions phyletic taxon, and the Orthotrichaceae were nested are incongruent (p Ͻ 0.01). That is, the chloroplast within the Haplolepideae. Similarly, the genera and nuclear sequences appear to possess conflicting Aphanorrhegma (Funariaceae) and Lorentziella historical signals based on the assumptions implicit (Gigaspermaceae) were resolved as para- or poly- in the use of the parsimony criterion and the op- phyletic. tions invoked during the tree search procedure. The analysis of the chloroplast data yielded two Phylogenetic analyses.—All analyses consistent- most parsimonious trees (Fig. 2), of length 889, ly resolve the Funariaceae and Funariineae as poly- characterized by a consistency index (CI) of 0.56 phyletic lineages. Goniomitrium acuminatum, (0.46 when autapomorphic characters are removed) which lacks the codon loss in the rps4 shared by and a rescaled consistency index (RC) of 0.39. Here all other Funariaceae, appears nested within the the Haplolepideae, Funariaceae (excl. Goniomi- Haplolepideae. The Ephemeraceae and Splachnob- trium acuminatum), and the Gigaspermaceae are ryaceae are also resolved within the Haplolepideae. monophyletic. Furthermore, these taxa compose a The phylogenetic position of the Splachnobryaceae natural group that is sister to the Timmiaceae. This is determined primarily by chloroplast data, as anal- inclusive clade shares a common ancestor with the yses of 18S rDNA sequence data result in it being remaining diplolepideous mosses. This topology is nested within the Funariaceae. However, Splach- retained upon the exclusion of the third codon po- nobryum does possess the codon that is absent from sition of the rps4 gene, or the inclusion of Disce- the other members of the Funariaceae. Enforcing lium in the analysis. In the resulting single MPT in the monophyly of either the Funariaceae or the Fu- the latter analysis (tree length ϭ 909, CI ϭ 0.56, nariineae sensu Vitt (1984) using the 18S sequences RC ϭ 0.38) Discelium is nested between the Gi- results in topologies whose length is significantly gaspermaceae and the Funariaceae-Encalyptaceae worse (p Ͻ 0.001). clade (Fig. 2). Most basal internodes remain de- Phylogenetic inferences from a combined data fined by low BT and DI values. set resulted in two most parsimonious hypotheses ϭ (tree length 1,363, CI with autapomorphic char- DISCUSSION acters excluded ϭ 0.441, RC ϭ 0.384; Fig. 1). In both trees the Timmiaceae are resolved as the sis- The Funariineae are primarily defined by vege- ter-group to the remaining ingroup taxa. The Ar- tative characters, because many taxa (genera or chidiaceae are consistently nested within the Hap- even families) are gymnostomous or characterized lolepideae. The Orthotrichales, Splachnales (includ- by otherwise reduced peristomes. Such a systematic ing Meesiaceae), and Bartramiales form a mono- concept of the Funariineae does not withstand phy- phyletic group characterized by a bootstrap logenetic testing. Neither nuclear nor chloroplast percentage (BP) of 86 and a decay index (DI) of data, whether analyzed in combination or separate- three. In the sister clade to this diplolepideous ally, support a monophyletic Funariineae as tradition- ternate clade, the Haplolepideae share a common ally defined (e.g., Vitt 1984). Moreover, the mono- ancestor with a lineage comprising the Gigasper- phyly of these taxa is even strongly rejected by the maceae, Encalyptineae, and the Funariineae. Within data presented here. The Ephemeraceae, a small, the latter clade the Funariaceae appear more closely but widespread family of minute ephemeral mosses, related to the Encalyptaceae than to the Gigasper- are consistently resolved within the Haplolepideae. maceae. None of the branches defining the relation- The lack of a peristome and the lack of strong ga- ships among these major lineages is defined by BP metophytic differentiation of the Ephemeraceae higher than 50, and the DI vary between one and preclude reciprocal testing of this hypothesis using two. Considering the possible heterogeneity of the morphological data at present. A placement within signal carried by both partitions (nuclear and chlo- the Pottiineae, and maybe even within the Potti- roplast data), separate analyses were performed. aceae, is certainly a viable hypothesis, considering When using the 18S rDNA sequences alone, at that the Pottiaceae include many taxa having un- least 5,000 equally most parsimonious trees (tree dergone severe morphological reduction (see Zan- length ϭ 414, CI without autapomorphies ϭ 0.496, der 1993). The affinities of the Ephemeraceae with- and RC ϭ 0.502) were obtained (result not shown). in the Haplolepideae are currently being further ex- The Funariaceae (excl. Goniomitrium acuminatum) amined and preliminary results, based on rps4 data were resolved sister to clade composed of all other for species of Micromitrium corroborate a relation- Arthrodonteae, wherein the Timmiaceae were re- ship of the Ephemeraceae with the Pottiaceae (Gof- solved as the basal-most lineage (see also Cox et finet et al., unpubl.). Goniomitrium acuminatum, al. 2000; Newton et al. 2000). Within the large traditionally included in the Funariaceae (Fife 216 THE BRYOLOGIST [VOL. 103 F) trn L- trn 4/ rps (18S/ GenBank accession number /AF223063/AF229920 AF223019/AF223048/AF229905 AF223013/AF223042/AF229899 AF223014/AF223043/AF229900 AF223012/AF223041/AF229898 X74114/AF023776/AF03716 AF230415/AF223034/AF229891 AF223007/AF223036/AF229892 — AF223018/AF223047/AF229904 AF223028/AF223057/AF229914 AF223015/AF223044/AF229901 AF223017/AF223046/AF229903 AF223016/AF223045/AF229902 AF223026/AF223055/AF229912 AF023679/AF023778/AF023718 AF223010/AF223039/AF229896 AF223011/AF223040/AF229897 AF023680/AF023777/AF023717 AF223009/AF223038/AF229895 AF025291/AF023814/AF023727 AF223032/AF223061/AF229918 AF223022/AF223051/AF229908 AF223020/AF223049/AF229906 AF223021/AF223050/AF229907 AF223023/AF223052/AF229909 AF223024/AF223053/AF229910 ) ) (H) NY ( NYSM ( lken & Venter 575 ¬ (voucher 822)* * eld 90527 eld 98872 net 5601 net 6326 net 4524 net 4580 net 4492 net s.n. net s.n. net 5586 net 5348 net 5613 fi fi fi fi fi fi fi fi fi fi fi fi F region have been included in the analyses (all vouchers for which sequences were generated Vitt 27234 Cox & Hedderson (1999) Gof Gof Mitzutani 13257 Gof Cox & Hedderson (1999) Gof Gof Gof Gof Oliver, To Scho McGill Schinini 24785 Scho Cox & Hedderson (1999) Gof Smith 47503 Streimann 48855 Whitehouse s.n. Gof Cox & Hedderson (1999) Gof Rushing trn L- trn 4 genes and the unless otherwise noted; * refers to collections represented by axenic cultures maintained by Michael Christianson, University rps DUKE ll. ¨ r. & Dix. ´ Hook. & Wils. (Hook. f. & Wils.) Sull. 2 (Hook. f. & Wils.) Sull. 1 (Hedw.) Hampe The Mitt. (Hook.) Mitt. Schimp. C. Mu n (Williams) Horton (Mitt.) Broth. 1 (Mitt.) Broth. 2 Hedw. (Hedw.) Bruch & Schimp. ´ (Hook.) Lindb. 1 (Hook.) Lindb. 2 Schwaegr. Bruch & Schimp. (Hedw.) Mohr Taxon Voucher or reference (Taylor) Broth. Hook. & Tayl. (Mitt.) Fife Card. Duse (Dicks.) Brid. Hedw. 1. Taxa for which the 18S rRNA and ABLE Physcomitrella patens Physcomitrium lorentzii Physcomitrium pyriforme Goniomitrium acuminatum Aphanorrhegma serratum Entosthodon bonplandii Entosthodon laevus Funaria apophysata Encalypta rhaocarpa Funaria hygrometrica Lorentziella imbricata Gigaspermum repens Lorentziella imbricata Orthotrichum lyellii Zygodon intermedius Diphyscium foliosum Discelium nudum Aphanorrhegma serratum Ephemerum spinulosum Chamaebryum pottioides Gigaspermum repens Bryobrittonia longipes Encalypta armata Drummondia obtusifolia Theriotia lorifolia Encalypta ciliata IPHYSCIACEAE ISCELIACEAE IGASPERMACEAE RTHOTRICHACEAE PHEMERACEAE NCALYPTACEAE UNARIACEAE T RTHOTRICHINEAE E D D E O G F UXBAUMIINEAE NCALYPTINEAE UNARIINEAE F O E of Kansas). B during the course of this study are deposited in 2000] GOFFINET & COX: ARTHRODONTOUS MOSSES 217 F) trn L- trn 4/ rps (18S/ GenBank accession number AF223027/AF223056/AF229913 AF223008/AF223037/AF229894 AF023698/AF023799/AF023756 X80980/AF023702/AF023736 AF223033/AF223062/AF229919 AF223006/AF223035/AF229892 AF023678/AF023775/AF023715 AF223025/AF223054/AF229911 AF023682/AF023831/AF023722 AF023689/AF023831/AF023722 AF023684/AF023780/AF023723 AF223030/AF223059/AF229916 AF223031/AF223060/AF229917 AF223029/AF223058/AF229915 ) UBC ( ) NY ( 1470 Ð eld 102331 eld 98363 eld 89289 net 5263 fi fi fi fi Scho Cox & Hedderson (1999) Cox & Hedderson (1999) Scho Cox & Hedderson (1999) Gof Scho Risk 1536 Cox & Hedderson (1999) Cox & Hedderson (1999) Cox & Hedderson (1999) Churchill 15606 Buck 29822 Goward 95 ll. ¨ (Brid.) Mitt. (Brid.) C. Mu Lesq. Hedw. (Hedw.) Wils. Hedw. (Hedw.) Bruch & Schimp. Taxon Voucher or reference Aust. (Hook.) Mitt. Hook. Hedw. Hedw. Lind. & Arnell (Hedw.) G.M.S. 1. Continued. EESIACEAE ABLE Tayloria scabriseta Splachnum sphaericum Splachnobryum obtusum Bartramia stricta Leptobryum pyriforme Amblyodon dealbatus Timmia austriaca Timmia siberica Fissidens subbasilaris Bryoxiphium norvegicum Archidium donnellii Tortula ruralis Ptychomitrium gardneri Scouleria aquatica RCHIDIACEAE RIMMIACEAE ARTRAMIACEAE RYACEAE RYOXIPHIACEAE IMMIACEAE OTTIACEAE PLACHNACEAE PLACHNOBRYACEAE ISSIDENTACEAE T RIMMIINEAE M T S B F B A P G B S ICRANINEAE RCHIDIINEAE RYINEAE ISSIDENTINEAE OTTIINEAE PLACHNINEAE A F G D S B P 218 THE BRYOLOGIST [VOL. 103

FIGURE 1. Strict consensus of two equally most parsimonious trees obtained from analyzing the nuclear and chloroplast data combined. Bootstrap percentages (Ͼ 50%) are presented below the branches and decay indices above. Boxed taxa belonged to the Funariineae sensu Vitt, and the shaded boxes further more composed the Funariaceae sensu Vitt (1984). 2000] GOFFINET & COX: ARTHRODONTOUS MOSSES 219

FIGURE 2. Phylogram of single most parsimonious trees obtained from analyzing chloroplast data (rps4 and trnL- trnF region) independently for a taxon sample that includes Discelium. Bootstrap percentages (Ͼ 50%) are presented below the branches and decay indices above. Boxed taxa belonged to the Funariineae sensu Vitt, and the shaded boxes further more composed the Funariaceae sensu Vitt (1984). 220 THE BRYOLOGIST [VOL. 103

1985) and considered with strong affinities to Fu- two conspecific samples) whereas the chloroplast naria (Stone 1981), is also shown to be more close- data consistently resolved at least the peristomate ly related to the Haplolepideae and to Tortula (Pot- taxa in major groups in accordance with traditional tiaceae) in particular (BT ϭ 96 and DI ϭ 8). The peristome-based systematic concepts (e.g., Funari- genus Goniomitrium comprises five species, and is ineae, Dicranineae). We will therefore discuss pri- defined by an overall reduce morphology (Fife marily the topologies inferred from chloroplast 1985). The phylogenetic affinities of Goniomitrium data. within the Pottiaceae is beyond the scope of this In the analyses of the combined data or the chlo- study and will be examined critically elsewhere. roplast data alone the Encalyptaceae form a sister The phylogenetic relationships of Splachnob- group to the Funariaceae. The peristome of Funaria ryum, a pantropical genus also composed of minute hygrometrica is composed of two rows, with the mosses, have remained controversial. Robinson endostome segments positioned opposite the exos- (1971) and Crum and Anderson (1981) among oth- tome teeth. The segments are raised on a high basal ers, placed the genus within the Pottiaceae, whereas membrane formed from an IPL that is composed of Vitt (1982, 1984) retained the genus within the 32 identical cells as a result of perfectly symmetric Splachnaceae as proposed earlier by Brotherus divisions (Shaw et al. 1989a). The peristome of the (1924). Koponen (1981) accommodated the genus Encalyptaceae varies in architecture (Edwards in its own family, and argued against affinities with 1984; Horton 1982, 1983), with regard to the num- the Pottiaceae, on the basis of diplolepideous fea- ber and structure of the teeth. Edwards (1984) tures of the peristome. Affinities to the Splachna- found no evidence of sesquilepideae (i.e., a 2:3 cell ceae were rejected due to the lack of differentiated pattern between the PPL and IPL characteristic of hypophysis. Instead, Koponen proposed to transfer the Dicranum-type peristome). The endostome seg- the Splachnobryaceae to the Funariineae. As men- ments never bear a vertical line on their outer sur- tioned above, the Funariineae have been defined face, and are always facing the exostome teeth, as mainly by vegetative characters. Many of these in the Funaria-type peristome. Some of the species characters are, however, considered plesiomorphic have narrow intermediate teeth that alternate with in mosses (Crosby 1980; Vitt 1982), and could be the segments. This situation is not homologous to reacquired following reduction (e.g., loss of papil- that found in the Bryum-type peristome, since, in lae results in smooth cells). The most parsimonious the latter, the segments bear remnants of the lon- inferences from nuclear sequences are congruent gitudinal anticlinal wall of the PPL on the outer with Koponen’s hypothesis, even though the genus surface (Edwards 1984). This arrangement is there- is resolved in a nested position within the Funari- fore only analogous to that of the Bryales sensu aceae. Such paraphyly of the Funariaceae seems, lato. Developmental studies further indicate that the however, incompatible with Splachnobryum lacking divisions in the IPL are also aligned with the plane the loss of a codon within its rps4 sequence, a char- of the adjacent PPL wall, although the timing may acter otherwise shared by all other Funariaceae be different from that observed in Funaria (Shaw, sampled (except Goniomitrium acuminatum, see pers. comm.). The sister relationship between the above). In contrast, phylogenetic inferences from Encalyptineae and the Funariineae, as suggested by chloroplast data suggest that Splachnobryum is al- chloroplast data, is consequently not incongruent lied to the Haplolepideae. Allen and Pursell (2000) with the sporophytic features. This phylogenetic recently re-examined the peristome of Splachnob- hypothesis is, although the most parsimonious, ryum and argued for a haplolepideous rather than poorly supported and in conflict with analyses diplolepideous architecture, an interpretation con- based on broader character sampling (see Cox et al. gruent with the chloroplast-based phylogenetic hy- 2000; Newton et al. 2000). The ancestor to this pothesis presented here. group may be mostclosely related to the Disceli- Phylogenetic inferences regarding the early di- aceae (Fig. 2). The peristome of Discelium, albeit versification of Arthrodontous mosses based on nu- reduced (Shaw & Allen 1985), is consistent with clear (18S rDNA) and chloroplast (rps4 and trnL- defining this broad clade by an opposite peristome. trnF) sequences separately yielded incongruent re- Furthermore, anatomical studies of immature cap- sults. However, neither the nuclear nor the chloro- sules reveal that the anticlinal walls of the IPL are plast data yielded topologies with internal branches all perfectly aligned with those of the PPL, sug- well supported by bootstrap values or decay indi- gesting that the divisions are symmetric (Shaw & ces. The nuclear data resolved many well estab- Allen 1985). lished taxa as polyphyletic (e.g., the Haplolepideae The Gigaspermaceae are resolved as the sister- were polyphyletic and included the Orthotricha- group to this broad diplolepideous-opposite clade. ceae; the Encalyptaceae were paraphyletic; and so Its members are, however, consistently gymnosto- were Aphanorrhegma and Lorentziella based on mous, and therefore offer no information for recon- 2000] GOFFINET & COX: ARTHRODONTOUS MOSSES 221 structing ancestral peristome types. The develop- sent an ancestral type of division from which more ment of the sporophyte in the Gigaspermaceae has severe asymmetries have evolved (i.e., those typical been studied by Rushing and Snider (1980). The of the Haplolepideae and the Bryales sensu lato; ontogenetic patterns of cell divisions within the am- Shaw et al. 1989a,b). Weak asymmetries occur also phithecium of Lorentziella imbricata, a species in- in Tetraphis (Goffinet et al. 1999; Shaw & Ander- cluded here, does not follow that of the fundamen- son 1988), whereas in Diphyscium, they are more tal square as in other mosses, and the divisions pronounced (Shaw et al. 1987). If indeed the divi- within the layers are not synchronized. Rushing and sions in Timmia are asymmetric, then the symmet- Snider (1980) do not specifically address each set ric division represents a synapomorphy for the En- of divisions, but from their figure 7, within which calyptineae and Funariineae. three of the eight IPL have undergone an anticlinal The peristome of the Timmiaceae is unique division, it appears that the new walls are laid down among diplolepideous mosses in the architecture of in the same plane as the adjacent PPL walls. The the endostome. This inner row is composed of a divisions can thus be characterized as symmetric. high basal membrane supporting 64 filamentous ap- This mode of IPL cell division may serve as a syn- pendages, arranged into groups of four that are op- apomorphy for the clade comprising the Gigasper- posite the exostome teeth at maturity. As in other maceae, Encalyptineae, and Funariineae. opposite peristomes, these appendages lack any an- Inferences from chloroplast data suggest that the ticlinal wall remnants of the PPL on their outer sur- sister-group to this Funarialean-Encalyptalean clade face (Murphy 1988; Shaw & Rohrer 1984). Wheth- is the Haplolepideae; a hypothesis retained upon er the opposite arrangement of the peristomes is an analysis of the combined data set, but in conflict apomorphy or a plesiomorphy for the clade com- with the analyses by Cox et al. (2000) and Newton prising the Funariineae, Encalyptineae, Haplolepi- et al. (2000). The peristome of the Haplolepideae deae, and Timmiaceae (as presented in Fig. 1) re- is typically single, and composed of the endostome mains unclear since the endostome of Diphyscium only. When the exostome is present, the teeth are (the Theriotia peristome was not examined) con- always poorly developed and adherent to the en- sists of a pleated membrane that lacks individual dostomial segments, and clearly positioned oppo- segments. The exostome of Diphyscium is com- site to the latter. That this observation is not an posed of 16 teeth separated by 16 intermediate artifact can be established by the position of the teeth. The latter are characterized by a median ver- teeth with regard to the anticlinal walls of the PPL. tical wall on their inner surface, and are fused to The teeth are indeed always formed between two the outer pleats of the endostome (Edwards 1984). consecutive anticlinal walls of the PPL (Shaw et al. The true teeth thus alternate with the pleats. A basal 1989b), as in the Funaria-type peristome and can membrane of the endostome is particularly pleated thus only lie opposite the exostomial teeth (Vitt et in taxa with alternate peristomes (see Shaw et al. al. 1998). The clade composed of the Funariineae, 1989a), but also albeit less so, in lineages with op- Encalyptineae, and the Haplolepideae can therefore posite peristomes (Edwards 1984; Schwartz 1994). be characterized by its opposite peristomes. Unlike The pleats themselves are thus not an indication of in the Funariineae, the development of the haplo- the arrangement of the peristomes. Consequently, lepideous peristome proceeds through a stage char- the lack of differentiated segments in Diphyscium acterized by an asymmetric division. Whether the precludes from determining its ‘‘putative’’ arrange- asymmetric division in the ILP should be consid- ment. Inclusion of the nuclear data in the analysis ered plesiotypic compared to the symmetric one of (or analyzing the nuclear data alone) resolves Tim- the Funariineae and Encalyptineae is not clear. mia sister to a dichotomy between lineages with The Timmiaceae and a clade composed of the either opposite or alternate peristome, suggesting Orthotrichineae and Bryineae sensu lato form a that the latter peristome configuration is derived. grade at the base of the Arthrodonteae (Figs. 1–2). The typical Bryum-type peristome bears cilia be- The development of the timmiaceous peristome has tween the segments. The occurrence of cilia is not been studied, except for the latest stages (Mur- seemingly correlated to additional divisions in the phy 1988). Prior to the deposition of the wall ma- IPL, leading to at least 48 cells composing the IPL. terial, anticlinal walls of IPL adjacent to those of In the Orthotrichaceae, cilia are always lacking and the PPL, appear in some cases well aligned, where- the IPL is composed of 32 cells at most. The re- as in others, they are not (Murphy 1988). The pat- lationships of the Orthotrichaceae remain ambigu- tern deviates from the perfect alignment of all walls ous (Cox & Hedderson 1999; Cox et al. 2000; Gof- in well-developed peristomes of the Funariineae finet et al. 1998), but a sister-group relationship to (Schwartz 1994, see also Goffinet et al. 1999), but the Bryales sensu lato as resolved here, rather than is similar to that observed in the Splachnineae and a nested position within the latter, cannot be ruled the Orthotrichineae. Partial asymmetry may repre- out. Depending on the homology assumption made 222 THE BRYOLOGIST [VOL. 103 for the appendages of the Timmiaceae, cilia could, shared some of his inherited axenic cultures. Assistance based on the chloroplast based phylogeny have by William Buck (NY) and Norton Miller (N.Y. State Mu- seum) in locating material of Lorentziella imbricata and arisen in the ancestor to either the Arthrodonteae Discelium nudum is very much appreciated. Sandra Boles (excluding the ‘‘Diphysciaceae’’) or to the Bryales (DUKE) provided valuable assistance for which we are only. The appendages present in the endostome of grateful. Support from the Green Plant Phylogeny Re- the Timmiaceae are monomorphic, whereas in the search Coordination Group through USDA grant 94- 37105-0713 from DOE/NSF/USDA for attendance of the Bryales sensu lato, these are dimorphic; cilia and XVI International Botanical Congress and various work- segments have distinct architectures (Shaw & Roh- shops is gratefully acknowledged. Finally, we would like rer 1984). The appendages present in the Timmi- to thank the curators of the herbaria (H, NY, UBC) for their aceae are similar to cilia in that they lack remnants permission to sample the collection for DNA extractions. of the anticlinal PPL walls on the outer surface, and in their position opposite the teeth. The endostome LITERATURE CITED of the Timmiaceae is also characterized by additional divisions leading to more than 32 cells in the ALLEN, B. & R. A. PURSELL. 2000. A reconsideration of the systematic position of Splachnobryum. Journal of IPL (Murphy 1988; Shaw & Rohrer 1984), as is the Hattori Botanical Laboratory 88: (in press). also typical of ciliate bryoid peristomes. It is thus BECKERT, S., S. STEINHAUSER,H.MUHLE &V.KNOOP. possible that at least some appendages of the tim- 1999. A molecular phylogeny of bryophytes based on miaceous endostome are homologous to the cilia of nucleotide sequences of the mitochondrial nad5 gene. Plant Systematics and Evolution 218: 179–192. the typical Bryum-type peristome, as suggested by BROTHERUS, V. F. 1924. Splachnaceae, pp. 333–334. In A. Shaw and Rohrer (1984). Support for this hypoth- Engler (ed.), Die natu¨rlichen Pflanzenfamilien 10, ed. esis will depend on the phylogenetic affinities of 2. Leipzig. the diplolepideous alternate mosses lacking cilia BUCK, W. R., B. GOFFINET &A.J.SHAW. 2000. Testing (e.g., Orthotrichales, and Splachnales; see Goffinet morphological concepts of orders of pleurocarpous mosses (Bryophyta) using phylogenetic reconstruc- et al. 1999) and the interpretation of their peristome tions based on trnL-trnF and rps4 sequences. Molec- (see Cox et al. 2000). ular Phylogenetics and Evolution 10: (in press). The phylogenetic relationships among basal- COX,C.J.&T.A.HEDDERSON. 1999. Phylogenetic rela- most arthrodontous mosses hypothesized here tionships among the ciliate arthrodontous mosses: evidence from chloroplast and nuclear DNA sequences. based on chloroplast data are congruent with infer- Plant Systematics and Evolution 215: 119–139. ences made from mitochondrial sequences (Beckert ,B.GOFFINET,A.E.NEWTON,J.SHAW &T.A.J. et al. 1999), but are incongruent with those derived HEDDERSON. 2000. Phylogenetic relationships among from analyses of the nuclear 18S rRNA gene (this the diplolepideous-alternate mosses (Bryidae) inferred study, and see also Cox et al. 2000; Hedderson et from nuclear and chloroplast DNA sequences. THE BRYOLOGIST 103: 224–241. al. 1996, 1998; Newton et al. 2000). Whether this CROSBY M. R. 1980. The diversity and relationships of conflict results from ancestral hybridization or re- mosses, pp. 115–129. In R. J. Taylor & A. E. Leviton flects the inadequacy of either set of characters for (eds.), The Mosses of North America. California deep-level reconstructions needs to be addressed Academy of Science, San Francisco. CRUM,H.A.&L.E.ANDERSON. 1981. Mosses of Eastern further. However, both the nuclear and the chloro- North America. 1. Columbia University Press, NY. plast do concur with regard to the polyphyly of the EDWARDS, S. R. 1984. Homologies and inter-relations of Funariineae and the Funariaceae. These results moss peristomes, pp. 658–695. In R. M. Schuster highlight again the usefulness of the molecular (ed.), New Manual of Bryology, Vol. 2. Hattori Bo- characters in addressing the circumscription of taxa tanical Laboratory, Nichinan, Japan. ERIKSSON, T. 1998. AutoDecay ver. 4.0 (program distrib- characterized by morphological reduction (i.e., loss uted by the author). Department of Botany, Stockholm or reversal of characters). Our analyses reveal fur- University. Stockholm. ther that taxa lacking critical morphological fea- FELSENSTEIN, J. 1985. Confidence limits on phylogenies: tures should not be omitted from phylogenetic anal- An approach using the bootstrap. Evolution 39: 783– 791. yses because of their limited contribution in resolv- FIFE, A. J. 1980. The affinities of Costesia and Neoshar- ing trends of morphological evolution, as these piella and notes on the Gigaspermaceae. THE BRYOL- taxa, such as the Disceliaceae or the Gigasperma- OGIST 83: 466–476. ceae, may be highly relevant for resolving the re- . 1985. A generic revision of the Funariaceae (Bry- lationships among lineages of arthrodontous moss- ophyta: Musci). Part I. Journal of the Hattori Botanical Laboratory 58: 149–196. es. FITCH, W. M. 1971. Toward defining the course of evolution: Minimum change for a specific tree topology. ACKNOWLEDGEMENTS Systematic Zoology 20: 406–416. GOFFINET, B., R. J. BAYER &D.H.VITT. 1998. Circum- This study was made possible through the financial sup- scription and phylogeny of the Orthotrichales (Bryop- port provided by Duke-University, NSF-grant DEB- sida) inferred from rbcL sequence analysis. American 9806955, and the Natural History Museum (London) Re- Journal of Botany 85: 1324–1337. search Fund. Michael Christiansen (U. of Kansas) kindly ,A.J.SHAW,L.E.ANDERSON &B.D.MISHLER. 2000] GOFFINET & COX: ARTHRODONTOUS MOSSES 223

1999. Peristome development in mosses in relation to stome-forming layers in the Funariaceae. International systematics and evolution. V. Diplolepideae: Orthotri- Journal of Plant Sciences 155: 640–657. chaceae. THE BRYOLOGIST 102: 581–594. SHAW, J. 1984. A reinterpretation of peristome structure HEDDERSON, T. A., R. L. CHAPMAN &W.L.ROOTES. 1996. in Pseudoditrichum mirabile Steere & Iwats. (Pseu- Phylogenetic relationships of bryophytes inferred from doditrichaceae). THE BRYOLOGIST 87: 314–318. nuclear-encoded rRNA gene sequences. Plant System- &B.H.ALLEN. 1985. Anatomy and morphology atics and Evolution 200: 213–224. of the peristome in Discelium nudum (Musci: Disce- ,R.CHAPMAN &C.J.COX. 1998. Bryophytes and liaceae). THE BRYOLOGIST 88: 263–267. the origins and diverification of land plants: new evi- &L.E.ANDERSON. 1988. Peristome development dence from molecules, pp. 65–77. In J. W. Bates, N. in mosses in relation to systematics and evolution. II. W. Ashton & J. G. Duckett (eds.), Bryology for the Tetraphis pellucida (Tetraphidaceae). American Jour- nal of Botany 75: 1019–1032. Twenty-First Century. Maney Publishing and British , &B.D.MISHLER. 1987. Peristome de- Bryological Society, Leeds, U. K. velopment in mosses in relation to systematics and HILLIS,D.M.&J.P.HUELSENBECK. 1992. Signal, noise, evolution. I. Diphyscium foliosum (Buxbaumiaceae). and reliability in molecular phylogenetic analyses. Memoirs of the New York Botanical Garden 45: 55– Journal of Heredity 83: 189–195. 70. &J.J.BULL. 1993. An empirical test for boot- , & . 1989a. Peristome develop- strapping as a method for assessing confidence in phy- ment in mosses in relation to systematics and evolu- logenetic analysis. Systematic Biology 42: 182–192. tion. III. Funaria hygrometrica, Bryum pseudocapil- HORTON, D. G. 1982. A revision of the Encalyptaceae lare, and B. bicolor. Systematic Botany 14: 24–36. (Musci) with particular reference to the North Amer- ,B.D.MISHLER &L.E.ANDERSON. 1989b. Peri- ican taxa. Part I. Journal of the Hattori Botanical Lab- stome development in mosses in relation to system- oratory 53: 365–418. atics and evolution. IV. Haplolepideae: Ditrichaceae . 1983. A revision of the Encalyptaceae (Musci) and Dicranaceae. THE BRYOLOGIST 92: 314–325. with particular reference to the North American taxa. &J.R.ROHRER. 1984. Endostomial architecture Part II. Journal of the Hattori Botanical Laboratory 54: in diplolepideous mosses. Journal of the Hattori Bo- 353–532. tanical Laboratory 57: 41–61. STONE, I. G. 1981. Spore morphology and some features HUELSENBECK, J. P. 1991. Tree length distribution skew- of Goniomitrium Hook. & Wils. (Funariaceae). Journal ness: an indicator of phylogenetic information. Sys- of Bryology 11: 491–500. tematic Zoology 41: 17–48. SWOFFORD, D. L. 1999. PAUP.* Phylogenetic Analysis us- KISHINO,H.&HASEGAWA, M. 1989. Evaluation off the ing Parsimony (*and other methods). Version 4. Sin- maximum likelihood estimate of the evolutionary tree auer Associates, Sunderland, MA. topologies from DNA sequence data, and the branch- TEMPLETON, A. R. 1983. Phylogenetic inference from re- ing order in Homonoidea. Journal of Molecular Evo- striction endonuclease cleavage site maps with partic- lution 29: 170–179. ular reference to the humans and apes. Evolution 37: KOPONEN, A. 1981. Splachnobryaceae, a new moss family. 221–244. Annales Botanici Fennici 18: 123–132. VITT, D. H. 1981. Adaptive modes of the moss sporo- LA FARGE-ENGLAND, C. 1996. Growth form, branching phyte. The Bryologist 84: 166–186. pattern, and perichaetial position in mosses: Cladocar- . 1982. Sphagnopsida and Bryopsida, pp. 305, py and pleurocarpy revisited. The Bryologist 99: 170– 307–336. In S. P. Parker (ed.), Synopsis and Classifi- 186. cation of living organisms, Vol. 1. McGraw-Hill, NY. MURPHY, S. A. 1988. Development of the Peristome of . 1984. Classification of the Bryopsida, pp. 696– Timmia megapolitana Hedw. M.Sc. thesis, University 759. In R. M. Schuster (ed.), New Manual of Bryol- of Iowa, Iowa City, Iowa. ogy, Vol. 2. Hattori Botanical Laboratory, Nichinen, Japan. NEWTON, A. E., C. COX,J.G.DUCKETT,J.WHEELER,B. ,B.GOFFINET &T.A.HEDDERSON. 1998. The or- GOFFINET,T.A.J.HEDDERSON &B.D.MISHLER. 2000. dinal classification of the mosses: Questions and an- Evolution of the major moss lineages: phylogenetic swers for the 1990’s, pp. 143–159. In J. W. Bates, N. analyses based on multiple gene sequences and mor- W. Ashton & J. G. Duckett (eds.), Bryology for the phology. THE BRYOLOGIST 103: 187–211. Twenty-first Century.Maney Publishing and British ROBINSON, H. 1971. A revised classification for the orders Bryological Society, Leeds, U.K. and families of mosses. Phytologia 21: 289–293. ZANDER, R. H. 1993. Genera of the Pottiaceae: mosses of RUSHING, A. E. & J. A. SNIDER. 1980. Observations of harsh environments. Bulletin of the Buffalo Society of sporophyte development in Lorentziella imbricata Natural Sciences 32: 1–378. (Mitt.) Broth. Journal of the Hattori Botanical Labo- ratory 47: 35–44. SCHWARTZ, O. M. 1994. The development of the peri- ms. received Nov. 15, 1999; accepted Feb. 8, 2000.