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Peatmoss (Sphagnum) diversification associated with Miocene Northern Hemisphere climatic cooling?
Molecular Phylogenetics and Evolution 55 (2010) 1139–1145
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Peatmoss (Sphagnum) diversification associated with Miocene Northern Hemisphere climatic cooling? A. Jonathan Shaw a,*, Nicolas Devos b, Cymon J. Cox c, Sandra B. Boles a, Blanka Shaw a, Alex M. Buchanan d, Lynette Cave d, Rodney Seppelt e a
Department of Biology, Duke University, Durham, NC 27708, USA Institute of Botany, University of Liège, B22 Sart Tilman, B-4000 Liège, Belgium Centro de Ciencias do Mar, Universidade do Algarve, Campus de Gambelas, 8005-139 Faro, Portugal d Tasmanian Herbarium, Private Bag 4 Hobart, Tasmania 7001, Australia e Australian Antarctic Division, Channel Highway, Kingston, Tasmania 7050, Australia b c
a r t i c l e
i n f o
Article history: Received 24 September 2009 Revised 13 January 2010 Accepted 19 January 2010 Available online 25 January 2010 Keywords: Bryophyte evolution Miocene Peatlands Peatmosses Sphagnum
a b s t r a c t Global climate changes sometimes spark biological radiations that can feed back to effect significant ecological impacts. Northern Hemisphere peatlands dominated by living and dead peatmosses (Sphagnum) harbor almost 30% of the global soil carbon pool and have functioned as a net carbon sink throughout the Holocene, and probably since the late Tertiary. Before that time, northern latitudes were dominated by tropical and temperate plant groups and ecosystems. Phylogenetic analyses of mosses (phylum Bryophyta) based on nucleotide sequences from the plastid, mitochondrial, and nuclear genomes indicate that most species of Sphagnum are of recent origin (ca. <20 Ma). Sphagnum species are not only well-adapted to boreal peatlands, they create the conditions that promote development of peatlands. The recent radiation that gave rise to extant diversity of peatmosses is temporally associated with Miocene climatic cooling in the Northern Hemisphere. The evolution of Sphagnum has had profound influences on global biogeochemistry because of the unique biochemical, physiological, and morphological features of these plants, both while alive and after death. Ó 2010 Elsevier Inc. All rights reserved.
1. Introduction Changes in earth’s geology and climate have had major impacts on biological evolution. The separation of Australia and Antarctica during the Tertiary, for example, led to increasing aridity on the Australian continent, which in turn set the stage for spectacular adaptive radiations of certain plant groups suited to dry conditions (e.g., Eucalyptus, Acacia) (Crisp et al., 2004) Similarly, Mediterranean-type ecosystems, with their numerous endemic organisms, developed during the last 5–10 million years as a result of global climate changes (cooling, drying) and modification of atmospheric circulation patterns (Ackerly, 2009). Some such biological radiations can feedback to effect significant impacts on biogeochemistry and ecosystem function. Peatlands form where primary plant production exceeds organic matter decomposition and today they occupy some 3.5 million km2 in boreal and subarctic regions (Vasander and Kettunen, 2006). Boreal peatlands have functioned as a sink for atmospheric carbon throughout the Holocene (ca. 10,000 years ago until present) and currently store between 250 and 450 pg of carbon, a large * Corresponding author. Fax: +1 919 660 7293. E-mail address: [email protected] (A.J. Shaw). 1055-7903/$ - see front matter Ó 2010 Elsevier Inc. All rights reserved. doi:10.1016/j.ympev.2010.01.020
percentage of the total terrestrial carbon in the boreal zone and up to 30% of world’s soil carbon pool (Gorham, 1991). Consequently, peatlands play an important role in fluxes of the ‘‘greenhouse” gases carbon dioxide and methane, and require careful consideration in predictions about future (and past) climate change. Although peat can form from the accumulation of various types of partially decomposed plant material, most northern peatlands are formed predominantly from Sphagnum moss (Rydin et al., 2006). Indeed, it has been suggested that more biomass is bound up in the moss genus Sphagnum, living and dead, than in any other single genus of plants on earth (Clymo and Hayward, 1982). Sphagnum-dominated peatlands also harbor substantial biodiversity including a broad range of endemic invertebrates (Desrochers and van Duinen, 2006), and are determinants of regional hydrology (Siegel and Glaser, 2006). The moss genus Sphagnum, which forms the basis for most northern peatlands, includes about 300–500 species worldwide (Eddy, 1979; Shaw, 2000; Shaw et al., 2003a). Sphagnum is one of two genera in the class Sphagnopsida of the phylum Bryophyta (the mosses). The other, Ambuchanania, includes only a single species, Ambuchanania leucobryoides, first discovered in 1987 in Tasmania (Yamaguchi et al., 1990). The Sphagnopsida is one of four or five deep clades within the phylum Bryophyta (Cox et al.,
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2004). Phylogenetic analyses support inferences from comparative morphology and fossils that the Bryophyta (mosses) diverged from other land plants before the vascular plants diversified during the mid- to late-Paleozoic (Shaw and Renzaglia, 2004). This implies that the peatmosses are an ancient group. Nevertheless, indirect observations suggest that many or most extant species of peatmosses may not be ancient and relictual. Cryptic species, genetic differentiation among disjunct populations, interspecific hybridization, and complex patterns of allopolyploid speciation, give the impression that extant peatmoss species are actively evolving and do not appear to be ‘‘living fossils”. Furthermore, extant Sphagnum species appear to be genetically closely related rather than isolated and relictual. For example, DNA sequences for the nuclear ribosomal internal transcribed spacer region (nrITS) can be readily aligned across almost all species (e.g., Shaw et al., 2003b). Furthermore, microsatellite primers that were developed for one group of closely related Sphagnum species amplify homologous microsatellite-containing loci across the whole genus with no evidence of ascertainment bias (Shaw et al., 2008a,b). This is in sharp contrast to most organisms, where microsatellite primers developed for one species typically do not amplify even in closely related species (e.g., Crawford et al., 1998). Although the genus Sphagnum occurs throughout the world, it is exceptionally diverse and abundant in boreal and subarctic zones of the Northern Hemisphere. These cold-climate, northern latitude habitats where Sphagnum reaches its greatest development did not exist in that region until relatively late in the Tertiary when global climate underwent significant cooling. During the late Tertiary (i.e., Oligocene to Pliocene) climatic cooling led to the loss of tropical and then temperate floristic elements in what are today the boreal and subarctic zones (Elias et al., 2006; Grimsson and Denk, 2007; Mosbrugger et al., 2005; Williams et al., 2008). This chronology, combined with the observation that extant species of Sphagnum appear to be evolutionarily young and closely related, leads to the hypothesis that extant peatmosses represent a recent radiation linked to the expansion of boreal and subarctic environments. Testing our hypothesis of a recent diversification for extant peatmosses requires that the phylogeny be calibrated with absolute dates. Unfortunately, mosses have a very poor fossil record (Oostendorp, 1987). We estimated absolute divergence times by reconstructing phylogenetic relationships and using three approaches to calibrating ages on the tree. First, within the context of a Bayesian Markov chain Monte Carlo phylogenetic analysis, a prior probability of 405 Ma was used for the most recent common ancestor (MRCA) of mosses (Bryophyta) and liverworts (Marchantiophyta) based on fossil estimates (Shaw and Renzaglia, 2004; Oostendorp, 1987). Second, average substitution rates for plastid, mitochondrial, and nuclear genes were gleaned from the literature and the overall mean rate was used as a prior in a separate analysis. Finally, both priors were used. There is considerable uncertainty associated with these estimates but consistent results from the three analyses (Table 1) make our results plausible. Furthermore, divergence dates we obtained for major lineages within the mosses agree well with previous estimates using different calibration approaches (Newton et al., 2007).
2. Materials and methods 2.1. Taxon sampling We tested the hypothesis that peatmosses diversified relatively recently by reconstructing phylogenetic relationships among extant mosses based on nucleotide sequences from eight plastid, mitochondrial, and nuclear loci. Sequences were assembled from a total of 56 mosses in the phylum Bryophyta, plus four liverworts
(phylum Marchantiophyta) included as outgroups (Table 1). Sampling from within the mosses included all major lineages resolved by previous phylogenetic analyses (Cox et al., 2004). Sequences used in the analyses published by Cox et al. (2004) were supplemented with 108 new sequences generated for this study. Twenty-five species currently classified in Sphagnum and representing all four major sections of the genus were included, as were two additional Sphagnum species that previous analyses indicated are sister to the rest of the genus (Shaw et al., 2003a). A sample of A. leucobryoides, recently collected in Tasmania, was also included. 2.2. Molecular protocols and genomic sampling Extractions, amplification, and sequencing followed protocols as described in Shaw et al. (2003a) using primers cited in that paper. The following genes were sequenced: photosystem II (PSII) reaction center protein D1 (psbA), ribulose-bisphosphate carboxylase gene (rbcL), ribosomal small protein 4 (rps4), tRNA(Gly) (UCC) (trnG), and the trnL (UAA) 59 exon—trnF (GAA) region (trnL) from the plastid genome, NADH protein-coding subunit 7 (nad7) from the mitochondrial genome, 18S ribosomal RNA (18S) and 26S ribosomal RNA (26S) from the nuclear genome. Accession information and GenBank accession numbers are provided in Table 1. 2.3. Phylogenetic reconstruction and divergence time estimation A relaxed-clock employing an uncorrelated lognormal model of rate variation among branches in the tree was used to estimate divergence times in our phylogenetic tree. The relaxed-clock model is implemented in BEAST version 1.4.8 (Drummond et al., 2006; Drummond and Rambaut, 2007). In contrast to other dating algorithms, BEAST uses a Bayesian Markov Chain Monte Carlo method to simultaneously estimate topology along with divergence times (Drummond et al., 2002). All BEAST analyses were run in the absence of topological constraints on the combined data set. The data set was, however, divided into plastid, mitochondrial and nuclear partitions. Substitution patterns in each partition followed a general time reversible model with rate variation among sites modeled using a gamma shape parameter with four rate categories. Model parameters were unlinked between the partitions so that each partition has its own model parameters estimated. Molecular evolution model parameters used flat priors, while tree priors were modeled according to a Yule speciation process. Divergence dates estimated were integrated over tree topologies sampled throughout the MCMC analysis, and weighted in proportion to their posterior probabilities. The MCMC chains were each run for 40 million generations, and parameters were sampled every 1000th generation. Inspection of the results using Tracer version 1.4 (Rambaut and Drummond, 2007) confirmed that stationarity and acceptable mixing of the sampled parameters were achieved. Trees were summarized as maximum clade credibility trees after removing, as a burnin, the first 20% of samples. In the absence of fossil evidence, two techniques were used to calibrate the phylogenetic tree in order to obtain absolute estimates of divergence times in millions of years. (1) We first constrained the age of the root of the tree using prior knowledge about the origin of bryophytes (Shaw and Renzaglia, 2004; Newton et al., 2007). Accordingly, a normal probability distribution prior was fitted to the parameter that corresponds to the age of the most recent common ancestor of mosses and liverworts. The normal distribution was centered at 405 millions years and had a standard deviation of 43 millions years. In this context, the root of the tree can only take a value that is within the prior distribution, which calibrates the tree and allows for the estimation of divergence times for other nodes in the tree. (2) In a second analysis, we placed a prior on the overall absolute rate of nucleotide substitution (meanRate parameter in
Table 1 Voucher information and GenBank accession numbers for samples included in phylogenetic analyses. Taxon
DNA_code
Collector
Coll. #
Herbarium
Rps4
Trnl
Psba
Nuc18s
Nuc26s-1
Nuc26s-2
Rbcl
Nad5
Nad7
Trng
Alophosia azorica Ambuchanania leucobryoides Andreaea wilsonii Andreaeobryum macrosporum Aulacomnium turgidum Bartramia stricta Brachythecium salebrosum Buxbaumia aphylla Dendroligotrichum dendroides Diphyscium foliosum Encalypta ciliata Entosthodson laevus Fissiden subbasilaris Funaria hygrometrica Haplomitrium hookeri Hedwigia ciliata Hookeria lucens Leucobryum sp. Mielichhoferia elongata Mnium hornum Oedipodium griffithianum Orthodontium lineare Orthotrichum lyelli Pellia ephiphylla Polytrichadelphus purpureus Polytrichum pallidisetum Porella pinnata Preissia quadrata Pyrrhobryum vallis-gratiae Rhodobryum giganteum Scouleria aquatica Sphagnum palustre Takakia lepidozioides Tetraphis pellucida Tetraplodon mniodes Timmia megapolitana Sphagnum perichaetiale Sphagnum tenerum Sphagnum lescurii Sphagnum cyclophyllum Sphagnum strictum Sphagnum compactum Sphagnum recurvum Sphagnum portoricense Sphagnum cymbifolioides Sphagnum teres Sphagnum steerii Sphagnum aongstroemii
MDP332 SB3269 B75 SB472
Rumsey Buchanan Cox Schofield
s.n. s.n 00–668 78094
DUKE
GQ368649 AY312939
AY312891 GQ368606 AY312892 AY312893
GQ375079 AY330416 AJ275005
AY330424 GQ375083 AY330425 AY330426
GQ368591 GQ368584
AY312924 GQ368609 AY312925 AF231059
AY312867 GQ368600 AY312868
AY330453 GQ368603 AY330454
GQ368647 GQ368615
DUKE DUKE
AY330476 GQ368612 AY330477 AF306953
A37 E3 BB86 B759 BG977
Hedderson Longton Goffinet Belland Goffinet
6385 4871 4723 16889 5425
RNG RNG NY DUKE DUKE
AF023809 AF023799 AF143027 AF306959 AF208420
AF023728 AF023756 AF161120 AF231909 AY312940
AY312894 AY312895 AY312896 AY312897 AY312898
AF023687 AF023698 AY330417 Y17603 AY330418
AY330427 AY330428 AY330429 AY330430 AY330431
AJ275180 AY312926 AY312927 GQ368610 AF208411
AY312869 AY312870 AY312871 AY312872 AY312873
AY330455 AY330456 AY330457
BG712 BG713 BG980 BG984 C148 SB475 A31 A80 AV_G7 SH5 C115 BG656 A79 L16 B97 MDP353 BG768 BG1072 BG1080 11755 5073 5811 SB1209 SB473 MDP474 L3 BG1003 SB74 SB75 SB77 SB78 SB79 SB80 SB83 SB106 SB127 SB366 SB367 SB412
4492 98872 5601 5263 148 95224 11771 118 s.n. s.n. 115 98670 s.n. 5745 12,231 84/01 4581 4744 105579 11,775 5073 5811 28,667 86,563 105,858 s.n. 97,957 9213 9335 s.n. 8560 9406 9332 9196 26,770 47,192 7928 8574 7531
DUKE DUKE DUKE DUKE RNG DUKE RNG RNG
AF229891 AF229897 AY312941 AF229913 AF231175 AY312942 AF233587 AF215906 GQ368650 AF023766 AF182360 AF246290 AF023768 AF023727
AY330432 AY330433 AY330434 AY330435 AY330436 AY330437 AY330438 AY330439 GQ375084 AY330440 AY330441 AY330442 AY330443 AY330444 AF226030
AY312928 AY312929 AY312930 AF231304 AF005513 U87072 AF231073 AY312931
AY312874 AY312875 AY312876 AY312877 Z98959 AY608284 Z98966 Z98969 GQ368601 AY312878 AY312879 AY312880 AY312881 AY312882 AY608305 AY312883 AY312884 AY608308 AY608309 AY312885 AY312886 AY312887 AY312888 AY312889 AY908812
AY330459 AY330460 AY330461 AY330462 AY330463
AY312943 AY312944 AY312945 AY312946 AF023754 AF023737 AF023723 AF192634 AY312947 AF231908 AF023730 AY312948 AF192575 AF192588 AF192565 AF192562 AF192585 AF192578 AF192569 AF192577 AF192584 AF192596 AF192574 AF192619
AY312899 AY312900 AY312901 AY312902 AY312903 AY312904 AY312905 AY312906 GQ368607 AY312907 AY312908 AY312909 AY312910 AY312911 AY312912 AY312913 AY312914 AY312915 AY312916 AY312917 AY312918 AY312919 AY312920 AY312921
Y17765 AF223011 AY330419 AF223027 X74114 Y19006 AJ275010 AJ243168
RNG RNG DUKE RNG RNG DUKE DUKE DUKE DUKE DUKE RNG RNG RNG DUKE DUKE DUKE RNG DUKE DUKE DUKE DUKE DUKE DUKE DUKE DUKE DUKE NY DUKE DUKE DUKE
AJ251065 AF223040 AY330478 AF223056 AF023776 AJ251064 AJ251309 AJ251316 GQ368613 AF023793 AF023796 AF306968 AF023800 AF023814 AY330479 AY330480 AF306956 AY330481 AY330482 AF023825 AF023789 AF023780 AF231892 AF306950 AY908021 AF023804 AF222902 AY309724 AY309734 AY309723 AY309719 AY309732 AY309716 AY309728 AY309725 AY309720 AY309735 AY309731 AY309715
Sphagnum Sphagnum Sphagnum Sphagnum
SB538 SB542 SB543 SB571
Goffinet Schofield Goffinet Goffinet Cox Schofield Hedderson Cox Vanderpoorten Shaw Cox Schofield Hedderson Hedderson Risk & Gross Cox Goffinet Goffinet Schofield Hedderson Longton Hedderson Long Schofield Risk & Gross Soderstrom Schofield Shaw Shaw Shaw Shaw Shaw Shaw Shaw Anderson Streimann Hedderson Andrus Andrus & Flatberg Shaw Shaw Shaw Shaw
9723 9639 9796 9678
DUKE DUKE DUKE DUKE
AY309733 AY309714 AY309726 AY309721
AY298303 AY298005 AY298224 AY347095
AY309623 AY309604 AY309616 AY309611
GQ375059 GQ375058 GQ375067 GQ375070
AF197065 AF197066 AF197067 AF197068 AF197069 AF197072 AF197077 AF197076 AF197090 AF197081 AF197083
GQ368587 GQ368579 GQ368592 GQ368589 GQ368593 GQ368590 GQ368599 GQ368594 GQ368598 GQ368596 GQ368595 GQ368597 AF197999 AF198000 AF198001 AF198002 AF198003 AF198004 AF198007 AF198012 AF198011 AF198025 AF198016 AF198018
AF232693 AF226820 AY312932 AJ275174 AF005536 U87085 AY312933 AY312934 U87088 AY312935 AJ275179 AJ275176 AF226822 AF231887 AY312936 U87091 AY312937 AY312938 AY309700 AY309710 AY309699 AY309695 AY309708 AY309692 AY309704 AY309701 AY309696 AY309711 AY309707 AY309691
AY312890 AY309566 AY309576 AY309565 AY309561 AY309574 AY309558 AY309570 AY309567 AY309562 AY309577 AY309573 AY309557
AY330474 AY330475 AY309590 AY309600 AY309589 AY309585 AY309598 AY309582 AY309594 AY309591 AY309586 AY309601 AY309597 AY309581
GQ368642 GQ368641 GQ368639 GQ368640 GQ368636 GQ368637 GQ368638 GQ368635 GQ368617 GQ368634 GQ368632 GQ368633 GQ368629 GQ368630 GQ368631 GQ368626 GQ368625 GQ368627 GQ368628 GQ368620 GQ368621 GQ368622 GQ368623 GQ368624 GQ368619 GQ368618 GQ368616 AY309766 AY309776 AY309765 AY309761 AY309774 AY309758 AY309770 AY309767 AY309762 AY309777 AY309773 AY309757
AF197092 AF197086 AF197087 AF197091
AF198027 AF198021 AF198022 AF198026
AY309709 AY309690 AY309702 AY309697
AY309575 AY309556 AY309568 AY309563
AY309599 AY309580 AY309592 AY309587
AY309775 AY309756 AY309768 AY309763
AY330445 AY330446 AY330447 AY330448 AY330449 AY330450 AY330451 AF197061 AF226033 AY330452
GQ368574 GQ368575 GQ368580 GQ368581 GQ368582 GQ368583
AY330458
GQ368645 GQ368646 GQ368643 GQ368644
GQ368588 GQ368585 GQ368586 GQ368576 GQ368577 GQ368578
AY330464 AY330465 GQ368604
AY330466 AY330467 AY330468 AY330469 AY330470
AY330471 AY330472 AY330473 AJ309978
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subnitens angustifolium pulchrum fuscum
AY312922 AY312923 AY309614 AY309624 AY309613 AY309609 AY309622 AY309606 AY309618 AY309615 AY309610 AY309625 AY309621 AY309605
AF023708 X80985 AF228668 AF023697 AF025291 X80210 AY330420 AY330421 AY330422 X80211 AF023695 AF023699 AF023684 Y11370 AJ269686 U18527 AF023691 AY330423 GQ375078 GQ375060 GQ375069 GQ375063 GQ375080 GQ375073 GQ375065 GQ375062 GQ375064 GQ375076 GQ375077 GQ375074
GQ368570 GQ368571 GQ368572 GQ368573
AY309769 AY309760 AY309759 AY309764 AY309755 AY309771 AY309772 AY309778 GQ368648 AY313205 AY313207 AY313208 AY313209
Nad7
AY309593 AY309584 AY309583 AY309588 AY309579 AY309595 AY309596 AY309602 GQ368605 AY309569 AY309560 AY309559 AY309564 AY309555 AY309571 AY309572 AY309578 GQ368602
Nad5 Rbcl
AY309703 AY309694 AY309693 AY309698 AY309689 AY309705 AY309706 AY309712 GQ368611 AF198024 AY313206
Nuc26s-2
GQ375071 GQ375075 GQ375068 GQ375082
GQ375061 GQ375066 GQ375072 GQ375081
Nuc26s-1 Nuc18s Psba
AY309617 AY309608 AY309607 AY309612 AY309603 AY309619 AY309620 AY309626 GQ368608 AF192608 AF192633 AY298061 AY298153 AY297996 AY298280 AY298287 AY298357 GQ368651
Trnl Rps4
AY309727 AY309718 AY309717 AY309722 AY309713 AY309729 AY309730 AY309736 GQ368614 DUKE DUKE DUKE MICH DUKE DUKE DUKE DUKE DUKE 9682 9327 s.n. 1236 28,884 18,926 40,888 9855 11,835 Shaw Shaw Tan & Harrison Price et al. Long Yamaguchi Higuchi Shaw Andrus SB576 SB642 SB858 SB1052 SB1211 SB1239 SB1248 SB1288 SB3274
Herbarium Coll. # Collector DNA_code
quinquefarium cuspidatum sericeum lapazense affine sericeum squarrosum wulfianum palenae Sphagnum Sphagnum Sphagnum Sphagnum Sphagnum Sphagnum Sphagnum Sphagnum Sphagnum
Taxon
Table 1 (continued)
A.J. Shaw et al. / Molecular Phylogenetics and Evolution 55 (2010) 1139–1145
AF197089 AY309506 AY309505 AY309507 AY309504 AY309508 AY309509 AY309510
Trng
1142
BEAST). Plastid, nuclear, and mitochondrial genomes have different rates of substitution. Within each one of those genomes, different genes will harbor different rates of substitution. Within a gene, coding regions and non-coding regions also probably have different rates of nucleotide substitution. Estimating an overall absolute rate of substitution is thus difficult and has to be undertaken with caution. It is worth noting that although we do calibrate our tree with an overall absolute rate, BEAST estimates a separate rate for each partition. Substitution rates used in the analyses were as follows: Palmer (1991) and Sanderson (2002) provided overall mean substitution rates for the plastid genome: 5.6 10 4 and 5 10 4 substitutions/site/million years, respectively. Gaut (1998) provided an estimate of 1.09 10 4 substitutions/site/million years for the mitochondrial genome in land plants, and Sanderson (2002) estimated 1.09 10 4 substitutions/site/million years for 26S nrDNA. An overall average across the three genomes was 3 10 4 substitutions/site/million years; this is the value used for calibrating our phylogenetic tree. (3) Finally, we combined in the same analysis the two types of calibration (i.e., prior on the age of the root and prior on the overall absolute rate of nucleotide evolution). Both calibration methods and the combined approach yielded very similar time estimates for most nodes (Table 2). The consistency between inferences using a divergence date for mosses and liverworts, and substitution rates, suggests that our selected substitution rate is reasonable given the time since divergence between mosses and liverworts as estimated from fossils and previous analyses. 3. Results and discussion Our results indicate that two major lineages within the Bryophyta diverged during the Paleozoic at least 196 Ma, one daughter lineage comprising the classes Andreaeopsida and Bryopsida, and the other Takakiopsida and Sphagnopsida (Fig. 1). Important nodes were well-supported with the exception of the sister group relationship between the Takakiopsida and Sphagnopsida (and this was not critical for testing our hypothesis). Our results suggest that the Sphagnopsida diverged relatively early from an ancestor shared with Takakiopsida, between 128 and 319 Ma, but Sphagnum s.s. diversified very recently, with the most recent common ancestor of almost all extant peatmoss estimated at 7–20 Ma (Fig. 1; Table 1). Molecular data clearly suggest that the radiation of extant Sphagnum species was relatively recent and very sudden, irrespective of the absolute dating for that radiation. Two Sphagnum species, Sphagnum sericeum and Sphagnum lapazense, fall outside the
Table 2 Estimated ages for critical nodes in the phylogenetic tree for mosses using Bayesian molecular dating approach and three different types of calibration: (1) a prior on the root node for the most recent common ancestor of mosses and liverworts was set to 405 mya (root) and a standard deviation of 43 was used to account for uncertainty around that age, (2) a prior on the overall average substitution rate in the tree was set to 3 104substitutions/site/millions of years (rate) and a standard deviation of 1 10 4 was used to take into account uncertainty around that rate, and (3) both previously described priors were combined in a single analysis.
1 2 3 4 5 6 7 8 9 10
Root
Rate
Root + rate
391.6 [308.6–474.3] 285.9 [196.6–386.9] 225.6 [126–326.3] 67.2 [34.1–108.3] 14.2 [8.3–21.4] 248.8 [169.9–348.2] 218.5 [146.8–306.2] 205.4 [139.5–290.2] 194.1 [127.4–270] 299.1 [202–409.8]
304.6 [135.8–531.9] 228.0 [107.7–399.8] 185.9 [77.4–329.6] 52.4 [21.0–96.4] 11.0 [4.7–19.8] 197.6 [92.7–343.5] 173.4 [81.7–305.6] 163.5 [78.8–285.8] 154.0 [74.0–269.4] 230.4 [91.3–409.1]
383.7 [296.3 – 464.3] 279.9 [196.8–377.8] 223.8 [128.2–319.2 65.5 [34.1–102.1] 13.6 [7.6–20.6] 240.9 [167–324.9] 209.4 [144.4–283.2] 197.4 [134–264] 185.9 [123.7–247.8] 296.6 [185.6–397.1]
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296.6 My (185 - 397)
Liverworts
10 240.9 My (167 - 324)
Andreopsida
6 7
1
209.4 My (144 - 283)
383.7 My (296 - 464)
8 2
Bryopsida
197.4 My (134 - 264)
279.9 My (196 - 377)
9
185.9 My (123 - 247)
1 2 3
3 223.8 My (128 - 319)
Takakiopsida
4 65.5 My (34 - 102)
5
Sphagnopsida
13.6 (7 - 20)
Paleozoic 400
350
Mesozoic 300
250
200
150
Cenozoic 100
50
14.4
Age (My)
Fig. 1. Phylogenetic tree showing relationships among major moss lineages based on nucleotide sequences from eight mitochondrial, plastid, and nuclear genes. Dates (± standard errors) at nodes are based on the analysis using two priors (‘‘root + rate” in Table 1): estimated age for the common ancestor of mosses and liverworts (405 Ma) and average estimated substitution rates for the three data partitions (mtDNA, cpDNA, nDNA). Nodal dates estimated from three analytical approaches are compared in Table 1. Branches supported at < 95% posterior probability are thickened. Early diverging taxa of Sphagnopida, Ambuchanania leucobryoides, S. lapazense, and S. sericum, are indicated on the figure as ‘‘1”, ‘‘2”, and ‘‘3”, respectively. Two populations of S. sericeum were included in the analyses.
recent Sphagnum radiation, and are in fact more closely related to Ambuchanania than to Sphagnum s.s. (Fig. 1; upper clade within Sphagnopsida). A sister group relationship between S. lapazense and Ambuchanania is strongly supported whereas the relationship of S. sericeum to those two is more ambiguous. Nevertheless, it is clear that these three taxa are not within the monophyletic group including most peatmosses (Sphagnum s.s.). The divergence between Ambuchanania, S. lapazense, and S. sericeum from Sphagnum s.s. is dated at 34–102 Ma (Fig. 1, Table 1; node (4). The long branch in the tree leading to extant species of Sphagnopsida (Sphagnum s.s. plus Ambuchanania, S. lapazense, and S. sericeum) suggests that there has been extensive extinction of early-diverging lineages within the Sphagnopsida during the Mesozoic and early Tertiary. The diversification of Sphagnum s.s. was clearly sudden and recent relative to the divergence and diversification of other moss lineages. The estimated age for the diversification of Sphagnum agrees closely with the date, based on oxygen isotope data, for rapid late Tertiary cooling in the Northern Hemisphere (Zachos et al., 2001). Prior to that turning point, dated at 17 Ma and known as the Miocene Climatic Optimum, northern latitudes of the northern Hemisphere were dominated by tropical floras (Mosbrugger et al., 2005). At about that time, these tropical flora were replaced by more temperate elements (Tiffney and Manchester, 2001;
Mosbrugger et al., 2005; Grimsson and Denk, 2007; Utescher and Mosbruger, 2007). During the late Miocene and Pliocene, true boreal and arctic (i.e., treeless) ecosystems expanded at very high latitudes (Elias et al., 2006). It appears that the diversification of Sphagnum was contemporaneous with these environmental and floristic changes in the late Tertiary. The three species that are in the Sphagnopsida but outside Sphagnum s.s. are largely restricted to the Southern Hemisphere; S. lapazense is known from one site in Bolivia (Crum, 2001), S. sericeum occurs from New Caledonia northward to Taiwan (Eddy, 1977), and Ambuchanania occurs only in Tasmania (Seppelt, 2000). Our results suggest that these Southern Hemisphere taxa represent relictual, early-diverging lineages of Sphagnopsida. Our results do not permit a determination of where the Sphagnopsida originated because extinction of other early lineages may have occurred. In fact, the long stem branch leading to extant species of Sphagnopsida in our reconstruction, as well as the relatively long branch leading to extant species of Sphagnum s.s., suggests that extinction of earlier lineages has occurred. Paleozoic and Mesozoic fossils attributed to the Sphagnopsida (see below) may represent some such lineages. The genus Sphagnum, in contrast to the three relictual taxa, occurs on all continents except Antarctica. Sphagnum s.s. is very di-
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Sphagnum Rigida Cuspidata Subsecunda
Squarrosa Acutifolia
Fig. 2. Summary of phylogenetic inferences from analyses of two major sections within the genus Sphagnum (Shaw et al., 2005, 2008). In both sections, taxa distributed in the boreal zone appear to be basal and tropical and Southern Hemisphere taxa are derived. Left is a phylogenetic tree showing relationships among the sections of Sphagnum (redrawn from 12).
verse at high latitudes of the Northern Hemisphere, but the genus also occurs at tropical latitudes and in the Southern Hemisphere so it could be that boreal and subarctic peatmosses are derived within the genus, contradicting the hypotheses that extant peatmosses first diversified in connection with Miocene cooling in the Northern Hemisphere. However, several observations suggest that tropical and Southern Hemisphere species of Sphagnum s.s. are derived rather than basal within the genus. Of the four large sections within the genus Sphagnum (which together include over 90% of Sphagnum diversity), two have been subjected to phylogenetic analyses with sufficient sampling at the species and population levels to resolve polarity of geographic patterns, and in both cases (sections Acutifola Shaw et al., 2005, and Subsecunda Shaw et al., 2008a,b), boreal taxa appear to be basal and tropical / Southern Hemisphere taxa are derived (Fig. 2). In addition, patterns of phylogenetic diversity across the whole genus suggest that tropical and Southern Hemisphere taxa are very closely related genetically, consistent with the hypothesis that they represent recent radiations from boreal-subarctic ancestors (Shaw et al., 2003b). Although the fossil record for peatmosses is limited, those records that do exist are consistent with the chronology we present here based on the molecular data. The earliest fossils attributed to the Sphagnopsida belong to the Permian Protosphagnum (Neuberg, 1960), which has the unique dimorphic leaf cell structure of extant peatmosses but appears to have a leaf midrib, unlike any extant
species of Sphagnopsida. Other Paleozoic plants that may be related to Sphagnum include the genera Junjagia and Vorcutannularia the (Lacey, 1969; Oostendorp, 1987). Like Protosphagnum, these appear to have dimorphic leaf cells, a hallmark of Sphagnopsida (including Ambuchanania), but have a midrib. Sphagnum leaves and spores have also been described from the Mesozoic and Cenozoic (Lacey, 1969; Jie and Xiuyi, 1986), consistent with the phylogenetic inference that the Sphagnopsida, and the genus Sphagnum, originated early, even if the diversity of extant species reflects a recent radiation. It is noteworthy that although Ambuchanania differs from typical peatmosses in numerous morphological features, the close phylogenetic relationship of Ambuchanania to S. sericeum and S. lapazense, both of which are mainstream sphagna in morphology, suggests that the characteristic Sphagnum morphology and plant architecture originated early in the evolution of Sphagnopsida. In fact, phylogenetic results indicate that the distinctive morphology of Ambuchanania is derived rather than primitive within the Sphagnopsida. Sphagnum is not only well-adapted to cold-temperature, acidic, low nutrient, anoxic environments that lead to the formation of peatlands, they play an active role in creating those conditions as potent ecosystem engineers (Clymo and Hayward, 1982; van Breemen, 1995). Peatmosses thrive in water-logged boreal peatlands and these habitats form in large part because of the chemical and growth characteristics of peatmosses. It appears that rapid
A.J. Shaw et al. / Molecular Phylogenetics and Evolution 55 (2010) 1139–1145
speciation of peatmosses in the late Tertiary was facilitated by global climatic cooling, and that the diversification of Sphagnum itself promoted the expansion of peatlands and their role in control of global climates. Indeed, boreal peatlands, made up largely of partially decomposed Sphagnum remains, can be considered an extended phenotype of the Sphagnum plants themselves (van Breemen, 1995). Natural selection on living peatmosses could have favored those chemical and physiological characteristics that promoted the accumulation of peat because peatlands have biotic and abiotic characteristics that favor peatmosses over other plants (van Breemen, 1995). Previous studies have documented correlations between global climate changes and major evolutionary events in the history of life (Ackerly, 2009; Koch et al., 1992; Crisp et al., 2004); the recent and rapid diversification of Sphagnum appears to exemplify a case where climate change promoted the diversification of a major land plant lineage, which in turn promoted the expansion of a novel ecosystem with important feedbacks to global climate. Acknowledgments B. Goffinet, D. Quandt, D. Soltis, H. Stenoien, A. Vanderpoorten provided comments on an earlier draft of the manuscript. This research was supported by NSF Grant nos. DEB-0515749, DEB0918998 and National Geographic Committee for Research and Exploration Grant No. 7855-05. References Ackerly, D.D., 2009. Evolution, origin and age of lineages in the Californian and Mediterranean floras. J. Biogeogr. 36, 1221–1233. Clymo, R.S., Hayward, P.M., 1982. The ecology of Sphagnum. In: Smith, A.J.E. (Ed.), Bryophyte Ecology. Chapman and Hall Ltd., London, New York, pp. 229–289. Cox, C.J., Goffinet, B., Shaw, A.J., Boles, S.B., 2004. Phylogenetic relationships among the mosses based on heterogeneous Bayesian analysis of multiple genes from multiple genomic compartments. Syst. Bot. 29, 234–250. Crawford, A.M., Kappes, S.M., Peterson, K.A., degotari, M.J., Dodds, K.G., Freking, B.A., Stone, R.T., Beattie, C.W., 1998. Microsatellite evolution: testing the ascertainment bias hypothesis. J. Mol. Evol. 46, 256–260. Crisp, M., Cook, L., Steane, D., 2004. Radiation of the Australian flora: what can comparisons of molecular phylogenies across multiple taxa tell us about the evolution of diversity in present-day communities. Phil. Trans. R. Soc. Lond. B. 359, 1551–1571. Crum, H.A., 2001. Miscellaneous notes on the genus Sphagnum. 11. Contrib. Univ. Mich. Herb 23, 107–114. Desrochers, S., van Duinen, G.-J., 2006. Peatland fauna. In: Wieder, R.K., Vitt, D.H. (Eds.), Boreal Peatland Ecosystems. Springer-Verlag, Heidelberg, pp. 67–100. Drummond, A.J., Rambaut, A., 2007. BEAST: Bayesian evolutionary analysis by sampling trees. BMC Evol. Biol. 7, 214. Drummond, A.J., Ho, S.Y.W., Phillips, M.J., Rambaut, A., 2006. Relaxed phylogenetics and dating with confidence. PLoS Biol 4, e88. Drummond, A.J., Nicholls, G.K., Rodrigo, A.G., Solomon, W., 2002. Estimating mutation parameters, population history and genealogy simultaneously from temporally spaced sequence data. Genetics 161, 1307–1320. Eddy, A., 1977. Sphagnales of tropical Asia. Bull. British Museum (Nat. Hist.) Bot. 5, 359–445. Eddy, A., 1979. Taxonomy and evolution of Sphagnum. In: Clarke, G.C.S., Duckett, J.G. (Eds.), Bryophyte Systematics. Academic Press, London and New York, pp. 109–121. Elias, S.A., Kuzmina, S., Kiselyov, S., 2006. Later Tertiary origins of the arctic beetle fauna. Palaeogeogr. Palaeoclim. Palaeoecol. 241, 373–392. Gaut, B.S., 1998. Molecular clocks and nucleotide substitution rates in higher plants. Evol. Biol. 30, 93–120.
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Gorham, E., 1991. Northern peatlands: role in the carbon cycle and probable responses to climatic warming. Ecol. App. 1, 182–195. Grimsson, F., Denk, T., 2007. Floristic turn-over in Iceland from 15 to 6 Ma– extracting signals from fossil flora assemblages. J. Biogeo. 34, 1490–1504. Jie, L., Xiuyi, Z., 1986. Discovery of Tertiary Sphagnum coal in Jinsuo Basin, Yunnan Province and its significance. Kexue Tongbao 31, 1556–1559. Koch, P.L., Zachos, J.C., Gingerich, P.D., 1992. Correlation between isotope records in marine and continental carbon reservoirs near the Palaeocene/Eocene boundary. Nature 358, 319–322. Lacey, W.S., 1969. Fossil bryophytes. Biol. Rev. 44, 189–205. Mosbrugger, V., Utescher, T., Dilcher, D.L., 2005. Cenozoic continental climatic evolution of Central Europe. Proc. Natl. Acad. Sci. USA 102, 14964–14969. Neuberg, M.F., 1960. Leafy mosses from the Permian deposits of Angarida. Trudy Geol. Inst. Leningr. 19, 1–104 (in Russian). Newton, A.E., Wikstrom, N., Bell, N., Forrest, L.L., Ignatov, M.S., 2007. Dating the diversification of the pleurocarpous mosses. In: Newton, A.E., Tangney, R.S. (Eds.), Pleurocarpous Mosses: Systematics and Evolution. CRC Press, Boca Raton, pp. 337–366. The bryophytes of the Palaeozoic and the Mesozoic. Bryoph. Biblio. 34. J. Cramer, Berlin. Plastid chromosome: structure and evolution. In: Bogorad, L., Vasil, I.K. (Eds.). Cell Culture and Somatic Genetics of Plants, Vol. 7A Molecular Biology of Plastids. Academic Press, San Diego, pp. 5-53. Rambaut, A., Drummond, A.J., 2007. Tracer version 1.4 (computer program). Available from:
Contents lists available at ScienceDirect
Molecular Phylogenetics and Evolution journal homepage: www.elsevier.com/locate/ympev
Peatmoss (Sphagnum) diversification associated with Miocene Northern Hemisphere climatic cooling? A. Jonathan Shaw a,*, Nicolas Devos b, Cymon J. Cox c, Sandra B. Boles a, Blanka Shaw a, Alex M. Buchanan d, Lynette Cave d, Rodney Seppelt e a
Department of Biology, Duke University, Durham, NC 27708, USA Institute of Botany, University of Liège, B22 Sart Tilman, B-4000 Liège, Belgium Centro de Ciencias do Mar, Universidade do Algarve, Campus de Gambelas, 8005-139 Faro, Portugal d Tasmanian Herbarium, Private Bag 4 Hobart, Tasmania 7001, Australia e Australian Antarctic Division, Channel Highway, Kingston, Tasmania 7050, Australia b c
a r t i c l e
i n f o
Article history: Received 24 September 2009 Revised 13 January 2010 Accepted 19 January 2010 Available online 25 January 2010 Keywords: Bryophyte evolution Miocene Peatlands Peatmosses Sphagnum
a b s t r a c t Global climate changes sometimes spark biological radiations that can feed back to effect significant ecological impacts. Northern Hemisphere peatlands dominated by living and dead peatmosses (Sphagnum) harbor almost 30% of the global soil carbon pool and have functioned as a net carbon sink throughout the Holocene, and probably since the late Tertiary. Before that time, northern latitudes were dominated by tropical and temperate plant groups and ecosystems. Phylogenetic analyses of mosses (phylum Bryophyta) based on nucleotide sequences from the plastid, mitochondrial, and nuclear genomes indicate that most species of Sphagnum are of recent origin (ca. <20 Ma). Sphagnum species are not only well-adapted to boreal peatlands, they create the conditions that promote development of peatlands. The recent radiation that gave rise to extant diversity of peatmosses is temporally associated with Miocene climatic cooling in the Northern Hemisphere. The evolution of Sphagnum has had profound influences on global biogeochemistry because of the unique biochemical, physiological, and morphological features of these plants, both while alive and after death. Ó 2010 Elsevier Inc. All rights reserved.
1. Introduction Changes in earth’s geology and climate have had major impacts on biological evolution. The separation of Australia and Antarctica during the Tertiary, for example, led to increasing aridity on the Australian continent, which in turn set the stage for spectacular adaptive radiations of certain plant groups suited to dry conditions (e.g., Eucalyptus, Acacia) (Crisp et al., 2004) Similarly, Mediterranean-type ecosystems, with their numerous endemic organisms, developed during the last 5–10 million years as a result of global climate changes (cooling, drying) and modification of atmospheric circulation patterns (Ackerly, 2009). Some such biological radiations can feedback to effect significant impacts on biogeochemistry and ecosystem function. Peatlands form where primary plant production exceeds organic matter decomposition and today they occupy some 3.5 million km2 in boreal and subarctic regions (Vasander and Kettunen, 2006). Boreal peatlands have functioned as a sink for atmospheric carbon throughout the Holocene (ca. 10,000 years ago until present) and currently store between 250 and 450 pg of carbon, a large * Corresponding author. Fax: +1 919 660 7293. E-mail address: [email protected] (A.J. Shaw). 1055-7903/$ - see front matter Ó 2010 Elsevier Inc. All rights reserved. doi:10.1016/j.ympev.2010.01.020
percentage of the total terrestrial carbon in the boreal zone and up to 30% of world’s soil carbon pool (Gorham, 1991). Consequently, peatlands play an important role in fluxes of the ‘‘greenhouse” gases carbon dioxide and methane, and require careful consideration in predictions about future (and past) climate change. Although peat can form from the accumulation of various types of partially decomposed plant material, most northern peatlands are formed predominantly from Sphagnum moss (Rydin et al., 2006). Indeed, it has been suggested that more biomass is bound up in the moss genus Sphagnum, living and dead, than in any other single genus of plants on earth (Clymo and Hayward, 1982). Sphagnum-dominated peatlands also harbor substantial biodiversity including a broad range of endemic invertebrates (Desrochers and van Duinen, 2006), and are determinants of regional hydrology (Siegel and Glaser, 2006). The moss genus Sphagnum, which forms the basis for most northern peatlands, includes about 300–500 species worldwide (Eddy, 1979; Shaw, 2000; Shaw et al., 2003a). Sphagnum is one of two genera in the class Sphagnopsida of the phylum Bryophyta (the mosses). The other, Ambuchanania, includes only a single species, Ambuchanania leucobryoides, first discovered in 1987 in Tasmania (Yamaguchi et al., 1990). The Sphagnopsida is one of four or five deep clades within the phylum Bryophyta (Cox et al.,
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2004). Phylogenetic analyses support inferences from comparative morphology and fossils that the Bryophyta (mosses) diverged from other land plants before the vascular plants diversified during the mid- to late-Paleozoic (Shaw and Renzaglia, 2004). This implies that the peatmosses are an ancient group. Nevertheless, indirect observations suggest that many or most extant species of peatmosses may not be ancient and relictual. Cryptic species, genetic differentiation among disjunct populations, interspecific hybridization, and complex patterns of allopolyploid speciation, give the impression that extant peatmoss species are actively evolving and do not appear to be ‘‘living fossils”. Furthermore, extant Sphagnum species appear to be genetically closely related rather than isolated and relictual. For example, DNA sequences for the nuclear ribosomal internal transcribed spacer region (nrITS) can be readily aligned across almost all species (e.g., Shaw et al., 2003b). Furthermore, microsatellite primers that were developed for one group of closely related Sphagnum species amplify homologous microsatellite-containing loci across the whole genus with no evidence of ascertainment bias (Shaw et al., 2008a,b). This is in sharp contrast to most organisms, where microsatellite primers developed for one species typically do not amplify even in closely related species (e.g., Crawford et al., 1998). Although the genus Sphagnum occurs throughout the world, it is exceptionally diverse and abundant in boreal and subarctic zones of the Northern Hemisphere. These cold-climate, northern latitude habitats where Sphagnum reaches its greatest development did not exist in that region until relatively late in the Tertiary when global climate underwent significant cooling. During the late Tertiary (i.e., Oligocene to Pliocene) climatic cooling led to the loss of tropical and then temperate floristic elements in what are today the boreal and subarctic zones (Elias et al., 2006; Grimsson and Denk, 2007; Mosbrugger et al., 2005; Williams et al., 2008). This chronology, combined with the observation that extant species of Sphagnum appear to be evolutionarily young and closely related, leads to the hypothesis that extant peatmosses represent a recent radiation linked to the expansion of boreal and subarctic environments. Testing our hypothesis of a recent diversification for extant peatmosses requires that the phylogeny be calibrated with absolute dates. Unfortunately, mosses have a very poor fossil record (Oostendorp, 1987). We estimated absolute divergence times by reconstructing phylogenetic relationships and using three approaches to calibrating ages on the tree. First, within the context of a Bayesian Markov chain Monte Carlo phylogenetic analysis, a prior probability of 405 Ma was used for the most recent common ancestor (MRCA) of mosses (Bryophyta) and liverworts (Marchantiophyta) based on fossil estimates (Shaw and Renzaglia, 2004; Oostendorp, 1987). Second, average substitution rates for plastid, mitochondrial, and nuclear genes were gleaned from the literature and the overall mean rate was used as a prior in a separate analysis. Finally, both priors were used. There is considerable uncertainty associated with these estimates but consistent results from the three analyses (Table 1) make our results plausible. Furthermore, divergence dates we obtained for major lineages within the mosses agree well with previous estimates using different calibration approaches (Newton et al., 2007).
2. Materials and methods 2.1. Taxon sampling We tested the hypothesis that peatmosses diversified relatively recently by reconstructing phylogenetic relationships among extant mosses based on nucleotide sequences from eight plastid, mitochondrial, and nuclear loci. Sequences were assembled from a total of 56 mosses in the phylum Bryophyta, plus four liverworts
(phylum Marchantiophyta) included as outgroups (Table 1). Sampling from within the mosses included all major lineages resolved by previous phylogenetic analyses (Cox et al., 2004). Sequences used in the analyses published by Cox et al. (2004) were supplemented with 108 new sequences generated for this study. Twenty-five species currently classified in Sphagnum and representing all four major sections of the genus were included, as were two additional Sphagnum species that previous analyses indicated are sister to the rest of the genus (Shaw et al., 2003a). A sample of A. leucobryoides, recently collected in Tasmania, was also included. 2.2. Molecular protocols and genomic sampling Extractions, amplification, and sequencing followed protocols as described in Shaw et al. (2003a) using primers cited in that paper. The following genes were sequenced: photosystem II (PSII) reaction center protein D1 (psbA), ribulose-bisphosphate carboxylase gene (rbcL), ribosomal small protein 4 (rps4), tRNA(Gly) (UCC) (trnG), and the trnL (UAA) 59 exon—trnF (GAA) region (trnL) from the plastid genome, NADH protein-coding subunit 7 (nad7) from the mitochondrial genome, 18S ribosomal RNA (18S) and 26S ribosomal RNA (26S) from the nuclear genome. Accession information and GenBank accession numbers are provided in Table 1. 2.3. Phylogenetic reconstruction and divergence time estimation A relaxed-clock employing an uncorrelated lognormal model of rate variation among branches in the tree was used to estimate divergence times in our phylogenetic tree. The relaxed-clock model is implemented in BEAST version 1.4.8 (Drummond et al., 2006; Drummond and Rambaut, 2007). In contrast to other dating algorithms, BEAST uses a Bayesian Markov Chain Monte Carlo method to simultaneously estimate topology along with divergence times (Drummond et al., 2002). All BEAST analyses were run in the absence of topological constraints on the combined data set. The data set was, however, divided into plastid, mitochondrial and nuclear partitions. Substitution patterns in each partition followed a general time reversible model with rate variation among sites modeled using a gamma shape parameter with four rate categories. Model parameters were unlinked between the partitions so that each partition has its own model parameters estimated. Molecular evolution model parameters used flat priors, while tree priors were modeled according to a Yule speciation process. Divergence dates estimated were integrated over tree topologies sampled throughout the MCMC analysis, and weighted in proportion to their posterior probabilities. The MCMC chains were each run for 40 million generations, and parameters were sampled every 1000th generation. Inspection of the results using Tracer version 1.4 (Rambaut and Drummond, 2007) confirmed that stationarity and acceptable mixing of the sampled parameters were achieved. Trees were summarized as maximum clade credibility trees after removing, as a burnin, the first 20% of samples. In the absence of fossil evidence, two techniques were used to calibrate the phylogenetic tree in order to obtain absolute estimates of divergence times in millions of years. (1) We first constrained the age of the root of the tree using prior knowledge about the origin of bryophytes (Shaw and Renzaglia, 2004; Newton et al., 2007). Accordingly, a normal probability distribution prior was fitted to the parameter that corresponds to the age of the most recent common ancestor of mosses and liverworts. The normal distribution was centered at 405 millions years and had a standard deviation of 43 millions years. In this context, the root of the tree can only take a value that is within the prior distribution, which calibrates the tree and allows for the estimation of divergence times for other nodes in the tree. (2) In a second analysis, we placed a prior on the overall absolute rate of nucleotide substitution (meanRate parameter in
Table 1 Voucher information and GenBank accession numbers for samples included in phylogenetic analyses. Taxon
DNA_code
Collector
Coll. #
Herbarium
Rps4
Trnl
Psba
Nuc18s
Nuc26s-1
Nuc26s-2
Rbcl
Nad5
Nad7
Trng
Alophosia azorica Ambuchanania leucobryoides Andreaea wilsonii Andreaeobryum macrosporum Aulacomnium turgidum Bartramia stricta Brachythecium salebrosum Buxbaumia aphylla Dendroligotrichum dendroides Diphyscium foliosum Encalypta ciliata Entosthodson laevus Fissiden subbasilaris Funaria hygrometrica Haplomitrium hookeri Hedwigia ciliata Hookeria lucens Leucobryum sp. Mielichhoferia elongata Mnium hornum Oedipodium griffithianum Orthodontium lineare Orthotrichum lyelli Pellia ephiphylla Polytrichadelphus purpureus Polytrichum pallidisetum Porella pinnata Preissia quadrata Pyrrhobryum vallis-gratiae Rhodobryum giganteum Scouleria aquatica Sphagnum palustre Takakia lepidozioides Tetraphis pellucida Tetraplodon mniodes Timmia megapolitana Sphagnum perichaetiale Sphagnum tenerum Sphagnum lescurii Sphagnum cyclophyllum Sphagnum strictum Sphagnum compactum Sphagnum recurvum Sphagnum portoricense Sphagnum cymbifolioides Sphagnum teres Sphagnum steerii Sphagnum aongstroemii
MDP332 SB3269 B75 SB472
Rumsey Buchanan Cox Schofield
s.n. s.n 00–668 78094
DUKE
GQ368649 AY312939
AY312891 GQ368606 AY312892 AY312893
GQ375079 AY330416 AJ275005
AY330424 GQ375083 AY330425 AY330426
GQ368591 GQ368584
AY312924 GQ368609 AY312925 AF231059
AY312867 GQ368600 AY312868
AY330453 GQ368603 AY330454
GQ368647 GQ368615
DUKE DUKE
AY330476 GQ368612 AY330477 AF306953
A37 E3 BB86 B759 BG977
Hedderson Longton Goffinet Belland Goffinet
6385 4871 4723 16889 5425
RNG RNG NY DUKE DUKE
AF023809 AF023799 AF143027 AF306959 AF208420
AF023728 AF023756 AF161120 AF231909 AY312940
AY312894 AY312895 AY312896 AY312897 AY312898
AF023687 AF023698 AY330417 Y17603 AY330418
AY330427 AY330428 AY330429 AY330430 AY330431
AJ275180 AY312926 AY312927 GQ368610 AF208411
AY312869 AY312870 AY312871 AY312872 AY312873
AY330455 AY330456 AY330457
BG712 BG713 BG980 BG984 C148 SB475 A31 A80 AV_G7 SH5 C115 BG656 A79 L16 B97 MDP353 BG768 BG1072 BG1080 11755 5073 5811 SB1209 SB473 MDP474 L3 BG1003 SB74 SB75 SB77 SB78 SB79 SB80 SB83 SB106 SB127 SB366 SB367 SB412
4492 98872 5601 5263 148 95224 11771 118 s.n. s.n. 115 98670 s.n. 5745 12,231 84/01 4581 4744 105579 11,775 5073 5811 28,667 86,563 105,858 s.n. 97,957 9213 9335 s.n. 8560 9406 9332 9196 26,770 47,192 7928 8574 7531
DUKE DUKE DUKE DUKE RNG DUKE RNG RNG
AF229891 AF229897 AY312941 AF229913 AF231175 AY312942 AF233587 AF215906 GQ368650 AF023766 AF182360 AF246290 AF023768 AF023727
AY330432 AY330433 AY330434 AY330435 AY330436 AY330437 AY330438 AY330439 GQ375084 AY330440 AY330441 AY330442 AY330443 AY330444 AF226030
AY312928 AY312929 AY312930 AF231304 AF005513 U87072 AF231073 AY312931
AY312874 AY312875 AY312876 AY312877 Z98959 AY608284 Z98966 Z98969 GQ368601 AY312878 AY312879 AY312880 AY312881 AY312882 AY608305 AY312883 AY312884 AY608308 AY608309 AY312885 AY312886 AY312887 AY312888 AY312889 AY908812
AY330459 AY330460 AY330461 AY330462 AY330463
AY312943 AY312944 AY312945 AY312946 AF023754 AF023737 AF023723 AF192634 AY312947 AF231908 AF023730 AY312948 AF192575 AF192588 AF192565 AF192562 AF192585 AF192578 AF192569 AF192577 AF192584 AF192596 AF192574 AF192619
AY312899 AY312900 AY312901 AY312902 AY312903 AY312904 AY312905 AY312906 GQ368607 AY312907 AY312908 AY312909 AY312910 AY312911 AY312912 AY312913 AY312914 AY312915 AY312916 AY312917 AY312918 AY312919 AY312920 AY312921
Y17765 AF223011 AY330419 AF223027 X74114 Y19006 AJ275010 AJ243168
RNG RNG DUKE RNG RNG DUKE DUKE DUKE DUKE DUKE RNG RNG RNG DUKE DUKE DUKE RNG DUKE DUKE DUKE DUKE DUKE DUKE DUKE DUKE DUKE NY DUKE DUKE DUKE
AJ251065 AF223040 AY330478 AF223056 AF023776 AJ251064 AJ251309 AJ251316 GQ368613 AF023793 AF023796 AF306968 AF023800 AF023814 AY330479 AY330480 AF306956 AY330481 AY330482 AF023825 AF023789 AF023780 AF231892 AF306950 AY908021 AF023804 AF222902 AY309724 AY309734 AY309723 AY309719 AY309732 AY309716 AY309728 AY309725 AY309720 AY309735 AY309731 AY309715
Sphagnum Sphagnum Sphagnum Sphagnum
SB538 SB542 SB543 SB571
Goffinet Schofield Goffinet Goffinet Cox Schofield Hedderson Cox Vanderpoorten Shaw Cox Schofield Hedderson Hedderson Risk & Gross Cox Goffinet Goffinet Schofield Hedderson Longton Hedderson Long Schofield Risk & Gross Soderstrom Schofield Shaw Shaw Shaw Shaw Shaw Shaw Shaw Anderson Streimann Hedderson Andrus Andrus & Flatberg Shaw Shaw Shaw Shaw
9723 9639 9796 9678
DUKE DUKE DUKE DUKE
AY309733 AY309714 AY309726 AY309721
AY298303 AY298005 AY298224 AY347095
AY309623 AY309604 AY309616 AY309611
GQ375059 GQ375058 GQ375067 GQ375070
AF197065 AF197066 AF197067 AF197068 AF197069 AF197072 AF197077 AF197076 AF197090 AF197081 AF197083
GQ368587 GQ368579 GQ368592 GQ368589 GQ368593 GQ368590 GQ368599 GQ368594 GQ368598 GQ368596 GQ368595 GQ368597 AF197999 AF198000 AF198001 AF198002 AF198003 AF198004 AF198007 AF198012 AF198011 AF198025 AF198016 AF198018
AF232693 AF226820 AY312932 AJ275174 AF005536 U87085 AY312933 AY312934 U87088 AY312935 AJ275179 AJ275176 AF226822 AF231887 AY312936 U87091 AY312937 AY312938 AY309700 AY309710 AY309699 AY309695 AY309708 AY309692 AY309704 AY309701 AY309696 AY309711 AY309707 AY309691
AY312890 AY309566 AY309576 AY309565 AY309561 AY309574 AY309558 AY309570 AY309567 AY309562 AY309577 AY309573 AY309557
AY330474 AY330475 AY309590 AY309600 AY309589 AY309585 AY309598 AY309582 AY309594 AY309591 AY309586 AY309601 AY309597 AY309581
GQ368642 GQ368641 GQ368639 GQ368640 GQ368636 GQ368637 GQ368638 GQ368635 GQ368617 GQ368634 GQ368632 GQ368633 GQ368629 GQ368630 GQ368631 GQ368626 GQ368625 GQ368627 GQ368628 GQ368620 GQ368621 GQ368622 GQ368623 GQ368624 GQ368619 GQ368618 GQ368616 AY309766 AY309776 AY309765 AY309761 AY309774 AY309758 AY309770 AY309767 AY309762 AY309777 AY309773 AY309757
AF197092 AF197086 AF197087 AF197091
AF198027 AF198021 AF198022 AF198026
AY309709 AY309690 AY309702 AY309697
AY309575 AY309556 AY309568 AY309563
AY309599 AY309580 AY309592 AY309587
AY309775 AY309756 AY309768 AY309763
AY330445 AY330446 AY330447 AY330448 AY330449 AY330450 AY330451 AF197061 AF226033 AY330452
GQ368574 GQ368575 GQ368580 GQ368581 GQ368582 GQ368583
AY330458
GQ368645 GQ368646 GQ368643 GQ368644
GQ368588 GQ368585 GQ368586 GQ368576 GQ368577 GQ368578
AY330464 AY330465 GQ368604
AY330466 AY330467 AY330468 AY330469 AY330470
AY330471 AY330472 AY330473 AJ309978
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A.J. Shaw et al. / Molecular Phylogenetics and Evolution 55 (2010) 1139–1145
subnitens angustifolium pulchrum fuscum
AY312922 AY312923 AY309614 AY309624 AY309613 AY309609 AY309622 AY309606 AY309618 AY309615 AY309610 AY309625 AY309621 AY309605
AF023708 X80985 AF228668 AF023697 AF025291 X80210 AY330420 AY330421 AY330422 X80211 AF023695 AF023699 AF023684 Y11370 AJ269686 U18527 AF023691 AY330423 GQ375078 GQ375060 GQ375069 GQ375063 GQ375080 GQ375073 GQ375065 GQ375062 GQ375064 GQ375076 GQ375077 GQ375074
GQ368570 GQ368571 GQ368572 GQ368573
AY309769 AY309760 AY309759 AY309764 AY309755 AY309771 AY309772 AY309778 GQ368648 AY313205 AY313207 AY313208 AY313209
Nad7
AY309593 AY309584 AY309583 AY309588 AY309579 AY309595 AY309596 AY309602 GQ368605 AY309569 AY309560 AY309559 AY309564 AY309555 AY309571 AY309572 AY309578 GQ368602
Nad5 Rbcl
AY309703 AY309694 AY309693 AY309698 AY309689 AY309705 AY309706 AY309712 GQ368611 AF198024 AY313206
Nuc26s-2
GQ375071 GQ375075 GQ375068 GQ375082
GQ375061 GQ375066 GQ375072 GQ375081
Nuc26s-1 Nuc18s Psba
AY309617 AY309608 AY309607 AY309612 AY309603 AY309619 AY309620 AY309626 GQ368608 AF192608 AF192633 AY298061 AY298153 AY297996 AY298280 AY298287 AY298357 GQ368651
Trnl Rps4
AY309727 AY309718 AY309717 AY309722 AY309713 AY309729 AY309730 AY309736 GQ368614 DUKE DUKE DUKE MICH DUKE DUKE DUKE DUKE DUKE 9682 9327 s.n. 1236 28,884 18,926 40,888 9855 11,835 Shaw Shaw Tan & Harrison Price et al. Long Yamaguchi Higuchi Shaw Andrus SB576 SB642 SB858 SB1052 SB1211 SB1239 SB1248 SB1288 SB3274
Herbarium Coll. # Collector DNA_code
quinquefarium cuspidatum sericeum lapazense affine sericeum squarrosum wulfianum palenae Sphagnum Sphagnum Sphagnum Sphagnum Sphagnum Sphagnum Sphagnum Sphagnum Sphagnum
Taxon
Table 1 (continued)
A.J. Shaw et al. / Molecular Phylogenetics and Evolution 55 (2010) 1139–1145
AF197089 AY309506 AY309505 AY309507 AY309504 AY309508 AY309509 AY309510
Trng
1142
BEAST). Plastid, nuclear, and mitochondrial genomes have different rates of substitution. Within each one of those genomes, different genes will harbor different rates of substitution. Within a gene, coding regions and non-coding regions also probably have different rates of nucleotide substitution. Estimating an overall absolute rate of substitution is thus difficult and has to be undertaken with caution. It is worth noting that although we do calibrate our tree with an overall absolute rate, BEAST estimates a separate rate for each partition. Substitution rates used in the analyses were as follows: Palmer (1991) and Sanderson (2002) provided overall mean substitution rates for the plastid genome: 5.6 10 4 and 5 10 4 substitutions/site/million years, respectively. Gaut (1998) provided an estimate of 1.09 10 4 substitutions/site/million years for the mitochondrial genome in land plants, and Sanderson (2002) estimated 1.09 10 4 substitutions/site/million years for 26S nrDNA. An overall average across the three genomes was 3 10 4 substitutions/site/million years; this is the value used for calibrating our phylogenetic tree. (3) Finally, we combined in the same analysis the two types of calibration (i.e., prior on the age of the root and prior on the overall absolute rate of nucleotide evolution). Both calibration methods and the combined approach yielded very similar time estimates for most nodes (Table 2). The consistency between inferences using a divergence date for mosses and liverworts, and substitution rates, suggests that our selected substitution rate is reasonable given the time since divergence between mosses and liverworts as estimated from fossils and previous analyses. 3. Results and discussion Our results indicate that two major lineages within the Bryophyta diverged during the Paleozoic at least 196 Ma, one daughter lineage comprising the classes Andreaeopsida and Bryopsida, and the other Takakiopsida and Sphagnopsida (Fig. 1). Important nodes were well-supported with the exception of the sister group relationship between the Takakiopsida and Sphagnopsida (and this was not critical for testing our hypothesis). Our results suggest that the Sphagnopsida diverged relatively early from an ancestor shared with Takakiopsida, between 128 and 319 Ma, but Sphagnum s.s. diversified very recently, with the most recent common ancestor of almost all extant peatmoss estimated at 7–20 Ma (Fig. 1; Table 1). Molecular data clearly suggest that the radiation of extant Sphagnum species was relatively recent and very sudden, irrespective of the absolute dating for that radiation. Two Sphagnum species, Sphagnum sericeum and Sphagnum lapazense, fall outside the
Table 2 Estimated ages for critical nodes in the phylogenetic tree for mosses using Bayesian molecular dating approach and three different types of calibration: (1) a prior on the root node for the most recent common ancestor of mosses and liverworts was set to 405 mya (root) and a standard deviation of 43 was used to account for uncertainty around that age, (2) a prior on the overall average substitution rate in the tree was set to 3 104substitutions/site/millions of years (rate) and a standard deviation of 1 10 4 was used to take into account uncertainty around that rate, and (3) both previously described priors were combined in a single analysis.
1 2 3 4 5 6 7 8 9 10
Root
Rate
Root + rate
391.6 [308.6–474.3] 285.9 [196.6–386.9] 225.6 [126–326.3] 67.2 [34.1–108.3] 14.2 [8.3–21.4] 248.8 [169.9–348.2] 218.5 [146.8–306.2] 205.4 [139.5–290.2] 194.1 [127.4–270] 299.1 [202–409.8]
304.6 [135.8–531.9] 228.0 [107.7–399.8] 185.9 [77.4–329.6] 52.4 [21.0–96.4] 11.0 [4.7–19.8] 197.6 [92.7–343.5] 173.4 [81.7–305.6] 163.5 [78.8–285.8] 154.0 [74.0–269.4] 230.4 [91.3–409.1]
383.7 [296.3 – 464.3] 279.9 [196.8–377.8] 223.8 [128.2–319.2 65.5 [34.1–102.1] 13.6 [7.6–20.6] 240.9 [167–324.9] 209.4 [144.4–283.2] 197.4 [134–264] 185.9 [123.7–247.8] 296.6 [185.6–397.1]
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296.6 My (185 - 397)
Liverworts
10 240.9 My (167 - 324)
Andreopsida
6 7
1
209.4 My (144 - 283)
383.7 My (296 - 464)
8 2
Bryopsida
197.4 My (134 - 264)
279.9 My (196 - 377)
9
185.9 My (123 - 247)
1 2 3
3 223.8 My (128 - 319)
Takakiopsida
4 65.5 My (34 - 102)
5
Sphagnopsida
13.6 (7 - 20)
Paleozoic 400
350
Mesozoic 300
250
200
150
Cenozoic 100
50
14.4
Age (My)
Fig. 1. Phylogenetic tree showing relationships among major moss lineages based on nucleotide sequences from eight mitochondrial, plastid, and nuclear genes. Dates (± standard errors) at nodes are based on the analysis using two priors (‘‘root + rate” in Table 1): estimated age for the common ancestor of mosses and liverworts (405 Ma) and average estimated substitution rates for the three data partitions (mtDNA, cpDNA, nDNA). Nodal dates estimated from three analytical approaches are compared in Table 1. Branches supported at < 95% posterior probability are thickened. Early diverging taxa of Sphagnopida, Ambuchanania leucobryoides, S. lapazense, and S. sericum, are indicated on the figure as ‘‘1”, ‘‘2”, and ‘‘3”, respectively. Two populations of S. sericeum were included in the analyses.
recent Sphagnum radiation, and are in fact more closely related to Ambuchanania than to Sphagnum s.s. (Fig. 1; upper clade within Sphagnopsida). A sister group relationship between S. lapazense and Ambuchanania is strongly supported whereas the relationship of S. sericeum to those two is more ambiguous. Nevertheless, it is clear that these three taxa are not within the monophyletic group including most peatmosses (Sphagnum s.s.). The divergence between Ambuchanania, S. lapazense, and S. sericeum from Sphagnum s.s. is dated at 34–102 Ma (Fig. 1, Table 1; node (4). The long branch in the tree leading to extant species of Sphagnopsida (Sphagnum s.s. plus Ambuchanania, S. lapazense, and S. sericeum) suggests that there has been extensive extinction of early-diverging lineages within the Sphagnopsida during the Mesozoic and early Tertiary. The diversification of Sphagnum s.s. was clearly sudden and recent relative to the divergence and diversification of other moss lineages. The estimated age for the diversification of Sphagnum agrees closely with the date, based on oxygen isotope data, for rapid late Tertiary cooling in the Northern Hemisphere (Zachos et al., 2001). Prior to that turning point, dated at 17 Ma and known as the Miocene Climatic Optimum, northern latitudes of the northern Hemisphere were dominated by tropical floras (Mosbrugger et al., 2005). At about that time, these tropical flora were replaced by more temperate elements (Tiffney and Manchester, 2001;
Mosbrugger et al., 2005; Grimsson and Denk, 2007; Utescher and Mosbruger, 2007). During the late Miocene and Pliocene, true boreal and arctic (i.e., treeless) ecosystems expanded at very high latitudes (Elias et al., 2006). It appears that the diversification of Sphagnum was contemporaneous with these environmental and floristic changes in the late Tertiary. The three species that are in the Sphagnopsida but outside Sphagnum s.s. are largely restricted to the Southern Hemisphere; S. lapazense is known from one site in Bolivia (Crum, 2001), S. sericeum occurs from New Caledonia northward to Taiwan (Eddy, 1977), and Ambuchanania occurs only in Tasmania (Seppelt, 2000). Our results suggest that these Southern Hemisphere taxa represent relictual, early-diverging lineages of Sphagnopsida. Our results do not permit a determination of where the Sphagnopsida originated because extinction of other early lineages may have occurred. In fact, the long stem branch leading to extant species of Sphagnopsida in our reconstruction, as well as the relatively long branch leading to extant species of Sphagnum s.s., suggests that extinction of earlier lineages has occurred. Paleozoic and Mesozoic fossils attributed to the Sphagnopsida (see below) may represent some such lineages. The genus Sphagnum, in contrast to the three relictual taxa, occurs on all continents except Antarctica. Sphagnum s.s. is very di-
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A.J. Shaw et al. / Molecular Phylogenetics and Evolution 55 (2010) 1139–1145
Sphagnum Rigida Cuspidata Subsecunda
Squarrosa Acutifolia
Fig. 2. Summary of phylogenetic inferences from analyses of two major sections within the genus Sphagnum (Shaw et al., 2005, 2008). In both sections, taxa distributed in the boreal zone appear to be basal and tropical and Southern Hemisphere taxa are derived. Left is a phylogenetic tree showing relationships among the sections of Sphagnum (redrawn from 12).
verse at high latitudes of the Northern Hemisphere, but the genus also occurs at tropical latitudes and in the Southern Hemisphere so it could be that boreal and subarctic peatmosses are derived within the genus, contradicting the hypotheses that extant peatmosses first diversified in connection with Miocene cooling in the Northern Hemisphere. However, several observations suggest that tropical and Southern Hemisphere species of Sphagnum s.s. are derived rather than basal within the genus. Of the four large sections within the genus Sphagnum (which together include over 90% of Sphagnum diversity), two have been subjected to phylogenetic analyses with sufficient sampling at the species and population levels to resolve polarity of geographic patterns, and in both cases (sections Acutifola Shaw et al., 2005, and Subsecunda Shaw et al., 2008a,b), boreal taxa appear to be basal and tropical / Southern Hemisphere taxa are derived (Fig. 2). In addition, patterns of phylogenetic diversity across the whole genus suggest that tropical and Southern Hemisphere taxa are very closely related genetically, consistent with the hypothesis that they represent recent radiations from boreal-subarctic ancestors (Shaw et al., 2003b). Although the fossil record for peatmosses is limited, those records that do exist are consistent with the chronology we present here based on the molecular data. The earliest fossils attributed to the Sphagnopsida belong to the Permian Protosphagnum (Neuberg, 1960), which has the unique dimorphic leaf cell structure of extant peatmosses but appears to have a leaf midrib, unlike any extant
species of Sphagnopsida. Other Paleozoic plants that may be related to Sphagnum include the genera Junjagia and Vorcutannularia the (Lacey, 1969; Oostendorp, 1987). Like Protosphagnum, these appear to have dimorphic leaf cells, a hallmark of Sphagnopsida (including Ambuchanania), but have a midrib. Sphagnum leaves and spores have also been described from the Mesozoic and Cenozoic (Lacey, 1969; Jie and Xiuyi, 1986), consistent with the phylogenetic inference that the Sphagnopsida, and the genus Sphagnum, originated early, even if the diversity of extant species reflects a recent radiation. It is noteworthy that although Ambuchanania differs from typical peatmosses in numerous morphological features, the close phylogenetic relationship of Ambuchanania to S. sericeum and S. lapazense, both of which are mainstream sphagna in morphology, suggests that the characteristic Sphagnum morphology and plant architecture originated early in the evolution of Sphagnopsida. In fact, phylogenetic results indicate that the distinctive morphology of Ambuchanania is derived rather than primitive within the Sphagnopsida. Sphagnum is not only well-adapted to cold-temperature, acidic, low nutrient, anoxic environments that lead to the formation of peatlands, they play an active role in creating those conditions as potent ecosystem engineers (Clymo and Hayward, 1982; van Breemen, 1995). Peatmosses thrive in water-logged boreal peatlands and these habitats form in large part because of the chemical and growth characteristics of peatmosses. It appears that rapid
A.J. Shaw et al. / Molecular Phylogenetics and Evolution 55 (2010) 1139–1145
speciation of peatmosses in the late Tertiary was facilitated by global climatic cooling, and that the diversification of Sphagnum itself promoted the expansion of peatlands and their role in control of global climates. Indeed, boreal peatlands, made up largely of partially decomposed Sphagnum remains, can be considered an extended phenotype of the Sphagnum plants themselves (van Breemen, 1995). Natural selection on living peatmosses could have favored those chemical and physiological characteristics that promoted the accumulation of peat because peatlands have biotic and abiotic characteristics that favor peatmosses over other plants (van Breemen, 1995). Previous studies have documented correlations between global climate changes and major evolutionary events in the history of life (Ackerly, 2009; Koch et al., 1992; Crisp et al., 2004); the recent and rapid diversification of Sphagnum appears to exemplify a case where climate change promoted the diversification of a major land plant lineage, which in turn promoted the expansion of a novel ecosystem with important feedbacks to global climate. Acknowledgments B. Goffinet, D. Quandt, D. Soltis, H. Stenoien, A. Vanderpoorten provided comments on an earlier draft of the manuscript. This research was supported by NSF Grant nos. DEB-0515749, DEB0918998 and National Geographic Committee for Research and Exploration Grant No. 7855-05. References Ackerly, D.D., 2009. Evolution, origin and age of lineages in the Californian and Mediterranean floras. J. Biogeogr. 36, 1221–1233. Clymo, R.S., Hayward, P.M., 1982. The ecology of Sphagnum. In: Smith, A.J.E. (Ed.), Bryophyte Ecology. Chapman and Hall Ltd., London, New York, pp. 229–289. Cox, C.J., Goffinet, B., Shaw, A.J., Boles, S.B., 2004. Phylogenetic relationships among the mosses based on heterogeneous Bayesian analysis of multiple genes from multiple genomic compartments. Syst. Bot. 29, 234–250. Crawford, A.M., Kappes, S.M., Peterson, K.A., degotari, M.J., Dodds, K.G., Freking, B.A., Stone, R.T., Beattie, C.W., 1998. Microsatellite evolution: testing the ascertainment bias hypothesis. J. Mol. Evol. 46, 256–260. Crisp, M., Cook, L., Steane, D., 2004. Radiation of the Australian flora: what can comparisons of molecular phylogenies across multiple taxa tell us about the evolution of diversity in present-day communities. Phil. Trans. R. Soc. Lond. B. 359, 1551–1571. Crum, H.A., 2001. Miscellaneous notes on the genus Sphagnum. 11. Contrib. Univ. Mich. Herb 23, 107–114. Desrochers, S., van Duinen, G.-J., 2006. Peatland fauna. In: Wieder, R.K., Vitt, D.H. (Eds.), Boreal Peatland Ecosystems. Springer-Verlag, Heidelberg, pp. 67–100. Drummond, A.J., Rambaut, A., 2007. BEAST: Bayesian evolutionary analysis by sampling trees. BMC Evol. Biol. 7, 214. Drummond, A.J., Ho, S.Y.W., Phillips, M.J., Rambaut, A., 2006. Relaxed phylogenetics and dating with confidence. PLoS Biol 4, e88. Drummond, A.J., Nicholls, G.K., Rodrigo, A.G., Solomon, W., 2002. Estimating mutation parameters, population history and genealogy simultaneously from temporally spaced sequence data. Genetics 161, 1307–1320. Eddy, A., 1977. Sphagnales of tropical Asia. Bull. British Museum (Nat. Hist.) Bot. 5, 359–445. Eddy, A., 1979. Taxonomy and evolution of Sphagnum. In: Clarke, G.C.S., Duckett, J.G. (Eds.), Bryophyte Systematics. Academic Press, London and New York, pp. 109–121. Elias, S.A., Kuzmina, S., Kiselyov, S., 2006. Later Tertiary origins of the arctic beetle fauna. Palaeogeogr. Palaeoclim. Palaeoecol. 241, 373–392. Gaut, B.S., 1998. Molecular clocks and nucleotide substitution rates in higher plants. Evol. Biol. 30, 93–120.
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Gorham, E., 1991. Northern peatlands: role in the carbon cycle and probable responses to climatic warming. Ecol. App. 1, 182–195. Grimsson, F., Denk, T., 2007. Floristic turn-over in Iceland from 15 to 6 Ma– extracting signals from fossil flora assemblages. J. Biogeo. 34, 1490–1504. Jie, L., Xiuyi, Z., 1986. Discovery of Tertiary Sphagnum coal in Jinsuo Basin, Yunnan Province and its significance. Kexue Tongbao 31, 1556–1559. Koch, P.L., Zachos, J.C., Gingerich, P.D., 1992. Correlation between isotope records in marine and continental carbon reservoirs near the Palaeocene/Eocene boundary. Nature 358, 319–322. Lacey, W.S., 1969. Fossil bryophytes. Biol. Rev. 44, 189–205. Mosbrugger, V., Utescher, T., Dilcher, D.L., 2005. Cenozoic continental climatic evolution of Central Europe. Proc. Natl. Acad. Sci. USA 102, 14964–14969. Neuberg, M.F., 1960. Leafy mosses from the Permian deposits of Angarida. Trudy Geol. Inst. Leningr. 19, 1–104 (in Russian). Newton, A.E., Wikstrom, N., Bell, N., Forrest, L.L., Ignatov, M.S., 2007. Dating the diversification of the pleurocarpous mosses. In: Newton, A.E., Tangney, R.S. (Eds.), Pleurocarpous Mosses: Systematics and Evolution. CRC Press, Boca Raton, pp. 337–366. The bryophytes of the Palaeozoic and the Mesozoic. Bryoph. Biblio. 34. J. Cramer, Berlin. Plastid chromosome: structure and evolution. In: Bogorad, L., Vasil, I.K. (Eds.). Cell Culture and Somatic Genetics of Plants, Vol. 7A Molecular Biology of Plastids. Academic Press, San Diego, pp. 5-53. Rambaut, A., Drummond, A.J., 2007. Tracer version 1.4 (computer program). Available from: