Tiny pollen grains: first evidence of Saururaceae from the Late Cretaceous of western North America

Introduction

Smith & Stockey (2007a) described inflorescences and flowers with in-situ pollen from the Eocene of North America that they assigned then to the modern genus Saururus (S. tuckerae S.Y.Sm. & Stockey). Saururaceae are a very small magnoliid family included in the Piperales (APG III, 2009), with six currently accepted species in four genera. In addition to Saururus cernuus L. and S. chinensis (Lour.) Baill., these are: Anemopsis californica Hook. & Arn., Gymnotheca chinensis Decne., Gymnotheca involucrata Pei, and Houttuynia cordata Thunb. An interesting pattern is the modern disjunct distribution of both of the two mutually monophyletic lineages in the Saururaceae (Anemopsis + Houttuynia vs. Gymnotheca + Saururus; Massoni, Forest & Sauquet, 2014) in North America and South/East Asia, suggesting that the family probably had a much wider distribution in the past (Table 1). The fossil record of Saururaceae is scanty (Table 2). Most of the fossils are fruits/seeds from the Eocene to Pliocene of western Eurasia and have been assigned to Saururus (S. bilobatus [Nikitin] Mai). In addition, Mai (1999) described fruits/seeds from the lower Miocene of Germany as Houttuynia bavarica Mai. The oldest fossil record so far is fossil wood from the Upper Cretaceous (no detailed stratigraphic information available) of Hokkaido described as Saururopsis niponensis Stopes & Fujii (1910, p. 58ff); the authors discuss carefully the affinity of the fossil and suggest that it could represent an ancestral member of the Saururaceae combining wood features typical for either Saururus or Houttuynia. The Eocene Saururus tuckerae (Smith & Stockey, 2007a) is so far the only fossil reported from North America. Though scanty, the fossil record confirms that Saururaceae were widespread by the Paleogene. The fossil record is also in line with the latest molecular dating estimates of a magnoliid dataset. According to the dating analyses of Massoni, Couvreur & Sauquet (2015b), the divergence between the two clades of the Saururaceae (Anemopsis + Houttuynia vs. Gymnotheca + Saururus) was established at the latest by the Eocene (>45 Ma), and the modern genera (and disjunctions) by the late Miocene (>10 Ma; Table 3). Two nodes in the phylogenetic neighbourhood of the Saururaceae were constrained using fossil age priors: the Saururus (≥44.3 Ma; ‘safe’ minimum constraint with reference to S. tuckerae) and Winteraceae root ages (=Canellales crown age; ≥ 126 Ma; Massoni, Doyle & Sauquet, 2015). Here, we document fossil pollen from the middle Upper Cretaceous Eagle Formation (Fm) of Wyoming, western North America, that is very similar to those of extant Saururus and nearly identical to that of pollen recovered in situ from Saururus tuckerae from the Eocene of British Columbia. Our findings are discussed in the context of newly documented dispersed Saururus pollen from the Eocene of British Columbia and Miocene of Central Europe (Austria), and the recent dating estimates for the family.

Material & Methods

Systematic Palaeobotany

Nomenclatural note. We believe that a fossil name should reflect the biological affinity indicated by the morphology of the fossil. Taking together all evidence, our Cretaceous pollen grains either represent an ancestral lineage within the Saururaceae that shared the primitive pollen morphology of extant and Cenozoic Saururus (hypothesis 1 below) or an early member of the Saururus-lineage (hypothesis 2). A name best reflecting hypothesis 1 would be to erect a new genus named, e.g. “Protosaururus” (Fig. 1B). However, this is impractical. The genus diagnosis could only be based on the Cretaceous pollen grains, and would be non-exclusive regarding pollen of the actual Saururus-lineage. If the currently prevalent cladistic-phylogenetic nomenclature should be followed (Fig. 1A) that only accepts taxa that have a (putative) inclusive common origin, i.e., are ‘monophyletic’ in a strict sense (Hennig, 1950; Hennig & Schlee, 1978), termed also ‘holophyletic’ by Ashlock (1971), the Cretaceous fossils would need to be addressed as “Saururaceae gen. et sp. indet.” (hypothesis 1) or Saururus (hypothesis 2). For consistency, our and future Saururus-type pollen grains would need to be named based on the currently accepted divergence ages for the Saururaceae (Fig. 1). An alternative solution that serves the requirements of the Botanical Code for unambiguous diagnoses is to follow the concept of “evolutionary classification” (e.g., Mayr & Bock, 2002; Hörandl, 2006; Hörandl, 2007), which allows naming also ‘paraphyletic’ groups to avoid that groups of directly related organisms with a non-inclusive common origin and similar or identical morphology are addressed by different names (Fig. 1B). In this case, one does not need to decide which hypothesis (paraphyletic Saururus pollen vs. holophyletic Saururus) applies when naming the pollen; and all Saururus-type pollen can be addressed as Saururus spp.

Saururus aquilae sp. nov. (Figs. 2–4)

Stratigraphy and age. Lettered Sands Member, Upper Eagle beds, Eagle Fm, Upper Cretaceous (Campanian); 82–81 Ma (Hicks, 1993; Van Boskirk, 1998).

Species diagnosis. Sculpture perforate, psilate to granulate; proximal face with about five perforations per μm²; perforations can have lobate outlines and up to six free-standing and/or protruding columellae; exine ≤400 nm and nexine <200 nm thick. All other pollen features that can be observed under LM and SEM as in the two modern species of the genus.

Description. Pollen, monad, shape oblate, form boat-like to globose, outline elliptic in equatorial and polar view; size very small, polar axis 3–5 μm long in SEM, equatorial diameter 6–11 μm in SEM; sulcate, sulcus with rounded ends (SEM); tectate; exine c. 400 nm thick, nexine c. 140 nm thick, nexine thinner than sexine (SEM); sculpture psilate in LM, perforate, psilate to granulate in SEM, 20–25 perforations per 4 μm², perforations tiny to small, circular, elliptic, irregular, irregular elongated to lobate in outline, perforations fewer and smaller on distal polar face (SEM), perforations are characterized by 1–6 free-standing and/or protruding columellae, free-standing columellae at periphery of perforations or sometimes filling them completely (SEM); sulcus membrane microechinate, microechini mostly with blunt apex, microechini densely packed to segregated (SEM).

Remarks. The description is based on c. 50 individual dispersed pollen grains studied both in LM and SEM. The Cretaceous S. aquilae sp. nov. pollen grains are very similar to or indistinguishable from the Eocene pollen of S. tuckerae that has been found both in situ in inflorescences/flowers (Smith & Stockey, 2007a) and dispersed in the same sediments (Zetter, 2006; this study). The only differences are found in the sculpture of the sulcus membrane: in some grains of S. aquilae sp. nov., the microechini can be densely packed (Figs. 2A, 2C; Figs. 4C, 4E, 4F), whereas they are widely spaced in S. tuckerae and the two modern species of Saururus (Table 4; File S1). The pollen grains of both taxa are even smaller than pollen of Miocene (S. stoobensis sp. nov., below) and extant Saururaceae except for Gymnotheca. They show the same basic SEM sculpture ranging from perforate, psilate to granulate; a variation also seen in the Miocene pollen but not to the same degree in extant members of the Saururaceae. The main diagnostic feature distinguishing S. aquilae sp. nov. from the Cretaceous and S. tuckerae from the Miocene and modern species of the genus is their high density of perforations (≥20 per 4 μm² on the proximal pollen face compared to ≤10 per 4 μm² in S. stoobensis sp. nov., S. cernuus, and S. chinensis). Furthermore, they both show up to six free-standing/protruding columellae at the periphery of perforations compared to a maximum of four in extant species of the Saururaceae. Occasionally lobate perforations in addition to the more common circular, elliptical and irregular perforations represent a feature seen only in the fossil Saururus pollen and the extant S. chinensis. Exine and nexine in both taxa are slightly but consistently thinner than in extant Sauruaceae. The S. aquilae sp. nov. pollen grains differ from those of Gymnotheca, Houttuynia, and Anemopsis. Pollen grains of Gymnotheca differ from S. aquilae sp. nov. and fossil and extant Saururus by their prominently striate and nanoechinate SEM sculpture; their perforations are without free-standing/protruding columellae. Houttuynia pollen grains are considerably larger than pollen of S. aquilae sp. nov., and are unique within Saururaceae in having a microverrucate sulcus membrane; their exine is much thicker than in S. aquilae sp. nov. Anemopsis pollen grains have sulcus membranes that are echinate to rugulate, a feature not seen in any other fossil or extant Saururaceae.

Derivation of name. The species is named after the Eagle (lat. aquila) Fm.

Saururus tuckerae S.Y.Sm. & Stockey (Figs. 4H,5,6,7A–7D)

Age. Middle Eocene, c. 48 Ma (Moss, Greenwood & Archibald, 2005).

Description. Pollen, monad, shape oblate, form boat-like, outline elliptic in equatorial and polar view; size very small, polar axis 3–5 μm long in SEM, equatorial diameter 6–11 μm in SEM; sulcate, sulci with rounded ends (SEM); tectate; exine c. 370 nm thick, nexine c. 150 nm thick, nexine thinner than sexine (TEM); sculpture psilate in LM, perforate, psilate to granulate in SEM, 23–26 perforations per 4 μm², perforations tiny to small, circular, elliptic, irregular, irregular elongated or lobate in outline, perforations fewer and smaller on distal polar face (SEM), perforations are characterized by 2–6 freestanding and/or protruding columellae, freestanding-columellae at periphery of perforations or sometimes filling it completely (SEM); sulcus membrane microechinate, microechini mostly with blunt apex, microechini segregated (SEM).

Remarks. The description is based on c. 200 individual dispersed pollen grains studied under LM and SEM, and compared with the in-situ grains figured in Smith & Stockey (2007a, 2007b). For additional remarks see remarks for Saururus aquilae.

Saururus stoobensis sp. nov. (Figs. 7E–7G)

Holotype. IPUW 7513/124 (Figs. 7E–7G).

Type locality. Opencast clay pit, Stoob-Warasdorf-Forest, Burgenland, Austria.

Age. Miocene (Pannonian ?; = Tortonian to Messinian, c. 12–6 Ma; Klaus, 1982)

Species diagnosis. Sculpture perforate, psilate to granulate; perforations occasionally with lobate outlines. All other pollen features (size, form, sculpture of sulcus membrane, number of perforations per μm²) that can be observed under LM and SEM as in the two modern species of the genus.

Description. Pollen, monad, shape oblate, form boat-like, outline elliptic in equatorial view; size very small, polar axis 4–5 μm long in SEM, equatorial diameter 10–11 μm in SEM; sulcate, sulci with rounded ends (SEM); tectate; sculpture psilate in LM, perforate, psilate to granulate in SEM, 7–10 perforations per 4 μm², perforations tiny to small, circular, elliptic or irregular in outline, perforations fewer and smaller on distal polar face (SEM), perforations are characterized by 2–4 freestanding and/or protruding columellae, freestanding columellae at periphery of perforations (SEM).

Remarks. The Miocene Saururus stoobensis sp. nov. pollen is more similar to the pollen of extant Saururus (Table 4; File S1) than the Cretaceous and Eocene Saururus pollen. It is of similar size, has the same density of perforations and the same number of free-standing columellae. It slightly differs from both modern species in the variation of the sculpture seen in SEM, and the occasional occurrence of perforations with lobate outline, which can be found in S. chinensis and the older fossil taxa, but has so far not been observed in S. cernuus or other Saururaceae genera.

Discussion

Fossil records of Saururaceae on the backdrop of latest molecular age estimates (Massoni, Couvreur & Sauquet, 2015b)

Until now the fossil record of Saururaceae has been confined to the Cenozoic except for wood remains from the Late Cretaceous of Japan described a century ago (Stopes & Fujii, 1910). Most specimens have been linked to the extant genus Saururus (Table 2). Figure 8 shows the fossil record in comparison to the magnoliid subtree that includes the Saururaceae, extracted from the dated trees provided by Massoni, Couvreur & Sauquet (2015a). The fossil pollen S. aquilae sp. nov. described here from the middle Late Cretaceous (Campanian) of Wyoming, conflicts with the youngest dating estimates, which infer a Late Cretaceous to Paleocene root age for the Saururaceae. Under the oldest age scenario (Massoni, Couvreur & Sauquet, 2015a, 2015b), the Wyoming pollen falls (time-wise) in the (arithmetic) middle between the Saururaceae root and crown divergence ages. On the backdrop of the dating estimates, S. aquilae could be the pollen produced by a potential precursor of all extant Saururaceae genera (hypothesis 1). Hypothesis 1 would fit also with the interpretation of the Cretaceous fossil wood described as Saururopsis nipponensis from Japan (Stopes & Fujii, 1910). Although being more similar to wood of Saururus, Stopes and Fujii state that some features are reminiscent of Houttuynia, which belongs to the second lineage of extant Saururaceae (e.g., Massoni, Forest & Sauquet, 2014), and discussed the possibility that the wood comes from an ancestral member of the family. On the other hand, the tip ages are poorly constrained (likely too young) using Massoni, Couvreur & Sauquet’s (2015b) dataset, who focussed on (much) deeper nodes. Hence, S. aquilae sp. nov. could represent an early member of the Saururus-lineage (hypothesis 2). Notably, the age of S. aquilae sp. nov. is close to the lower boundary of the highest posterior density (HPD) intervals for the older age scenarios (Fig. 8; Massoni, Couvreur & Sauquet, 2015a). Being interested in large-scale magnoliid processes, Massoni, Couvreur & Sauquet (2015b) did not use any age priors from within the Piperales subtree (Massoni, Doyle & Sauquet, 2015) and relied on relatively slow-evolving gene regions. It is a common observation that divergence ages towards the leaves of a tree tend to be (severely) underestimated in studies using large datasets when compared to focused studies that rely on ingroup constraints. Typically, the latter are in better agreement with the fossil record, as e.g., in the case of the Fagaceae (Hubert et al., 2014; Grímsson et al., 2015; Grímsson et al., 2016a; Renner et al., 2016). A similar observation can be made in the sister group of the Piperales, the Canellales. Figure 8 also shows the oldest records of the Winteraceae, which are much older than the age estimates by Massoni, Couvreur & Sauquet (2015a, 2015b). Studies focusing on either Canellaceae (Müller et al., 2015) or Winteraceae (Marquínez et al., 2009; Thomas et al., 2014), using different sets of age priors including Canellaceae and Winteraceae crown group fossils, obtained (much) older ages than the oldest age scenario (angiosperm root fixed to max. 200 Ma) in the set of analyses performed by Massoni, Couvreur & Sauquet (2015b). Differences range from at least 13 Ma for the Canellaceae and Winteraceae roots to more than 40 Ma for close-to-tips nodes.

Based on pollen morphology, neither hypotheses can be rejected. Saururus aquilae sp. nov. is essentially indistinguishable from pollen linked to the c. 30 Ma younger S. tuckerae, and all modern Saururaceae pollen types (≥35 Ma younger) differ only in a few characters (Table 4; File S1). The differences are often expressed as a range of variability (such as outlines of perforations or sculpture of the pollen surface). This demonstrates that pollen morphology is a very conservative trait in the lineage, and provides an argument for hypothesis 2 that S. aquilae sp. nov. was produced by an early member of the genus Saururus. On the other hand, the extant six species are clearly only the last survivors of a once more widespread group (Table 1), and may be unrepresentative regarding the actual variation in each generic lineage and the family over time. The pollen of Saururus may simply be primitive within the family. Ancestral members of the Saururaceae including precursors of all modern genera may have produced essentially the same pollen (hypothesis 1), whereas pollen morphologies of the extant species of the other genera are more derived (Fig. 9).

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