A new diatom genus Spargeria gen.nov.(Bacillariophyceae)from Xizang, China, and the description of two new species*

Yan LIU, John Patrick KOCIOLEK, Li WANG, Xinxin LU, Huan ZHU,Guoxiang LIU, Fengyang SUI, Yawen FAN,**

1 College of Life Science and Technology, Harbin Normal University, Harbin 150025, China

2 Museum of Natural History and Department of Ecology and Evolutionary Biology, University of Colorado, Boulder CO 80309, USA

3 Key Laboratory of Algal Biology, Institute of Hydrobiology, Chinese Academy of Sciences, Wuhan 430072, China

Abstract During our investigation of diatom biodiversity in Xizang, two species exhibited unique morphological features discriminative from all previously known genera.Herein we describe these two species and describe as new the genus, Spargeria gen.nov.The new genus features narrow to wide rectangular valves, narrow valve mantles, filiform raphe branches that occur on the valve face only,terminal raphe fissures straight or slightly deflected to same side, bow-tie shaped central areas, chambered striae present on the valve face only, being absent from the mantle, wider striae near the axial area and very narrow near the margin, multiseriate striae with small and round areolae that are occluded externally.Comparatively, Spargeria zhuii sp.nov.has larger and robust valves, radiate striae, with one divergent stria near the apices, while Spargeria chenia sp.nov.is smaller, with narrow valves, striae slightly radiate in the middle, becoming convergent or parallel near apices.This new genus belongs to the family Pinnulariaceae, and it was compared and contrasted with other genera of this family.Our work suggests the need for continued studies to document the biodiversity of diatoms in Xizang.

Keyword: Spargeria; new genus; Pinnulariaceae; morphology; Xizang

1 INTRODUCTION

The family Pinnulariaceae was established by Mann in Round et al.(1990) and originally included 4 genera, namelyPinnulariaEhrenberg andDiatomellaGreville, both of which are living freshwater diatoms, as well as?strupiaHeiden &Hustedt andDimidiataHajos that are genera of living marine and fossil diatoms, respectively.Later Cox (2015) published a new classification scheme and, in addition to the former 4 genera, three freshwater genera, includingAlveovallumLange-Bertalot & Krammer,CaloneisCleve, andHygropetraKrammer & Lange-Bertalot, and one marine genus,PinnunavisOkuno, were added to the family.Alveovallumwas subsequently combined intoPinnulariaby Liu et al.(2018b).The family Pinnulariaceae now is comprised of 7 genera.Freshwater genera in this family usually have biraphid valves, chambered striae, and areolae arranged into multiseriate rows.

Pinnulariais one of the most species-rich raphid diatom genera (Poulí?ková et al., 2018), with more than 3 600 recorded names worldwide (Kociolek et al., 2022).Compared with other “catch-all” genera,likeCymbellaAgardh andAchnanthidiumKützing,the genusPinnulariahas seemed very “stable” and has not been split into several genera.Pinnulariaspecies are usually small to large sized, and most have naviculoid symmetry and organization.Striae are alveolate, the raphe can range from filiform to lateral, with variously-shaped central and terminal raphe endings.SomePinnulariaspecies have an axial plate and form images of longitudinal lines.Pinnulariaspecies are usually abundant in low slightly acid waters of low conductivity (Spaulding and Edlund,2009).As the most species-rich genus in this family,Pinnulariahas received considerable attention from diatomists.Krammer (2000) considered (mostly)European species ofPinnulariaand made a relatively comprehensive revision for the genus.Some new species were described recently by Moreno et al.(2020), Das et al.(2020), Van De Vijver et al.(2020),Poulí?ková et al.(2018), Kochman-K?dziora et al.(2018), among others.

Caloneisis morphologically similar toPinnularia,thus Round et al.(1990) did not accept it; they combined it with the genusPinnularia.However, in most of the research,Caloneishas been identified as a separate genus (Bruder et al., 2008) having chambered striae, obvious longitudinal lines, and a fascia in the central area or irregularly-shaped structures in central area.Nearly 1 000 names have been recorded (Kociolek et al., 2022).Studies by Levkov and Williams (2014), Kulikovskiy et al.(2012), and Levkov et al.(2007) reported new species of this genus.

Diatomellahas a striae structure similar toPinnulariaandCaloneis, but it can be distinguished from them by having a scalariform valvocopula(mostly known as a septum-like structure formed by the valvocopula) (Van De Vijver et al., 2020).This genus is rare in samples, making the illustration of the fine structure of their valves more difficult.Van De Vijver et al.(2020) presented detailed observations on frustule morphology of the type species,Diatomella balfourianaGreville; and the formation of the scalariform valvocopula and its differences with septa were discussed specifically.Diatomellais a relatively “small” genus with only 13 taxa were recorded (Kociolek et al., 2022).No new species have been described in the genus since 1996.

Hygropetrais a recently described genus within the family, since Krammer (2000) established it based onHygropetra balfouriana(Grunow & Cleve)Krammer & Lange-Bertalot, and described a new speciesHygropetra elongataKrammer & Lange-Bertalot.In addition, two new species and one new combination have been reported subsequently by Mayama and Idei (2009), Kociolek et al.(2014),and Li and Qi (2014).Hygropetrahas an overall structure of the striae that is similar to other members of the family.It differs, however, by having areolar occlusions positioned on the interior of the valve, while species in most other genera of the family lack areolar occlusions (Round et al.,1990; Mayama and Idei, 2009) or they are positioned on the valve exterior.Hygropetraalso has the feature of a relatively simple raphe, with the terminal raphe fissures not extending onto the valve mantle (Spaulding, 2008).

In China,PinnulariaandCaloneisare well documented, of which 535 taxa and 66 taxa,respectively, have been recorded at present.DiatomellaandHygropetrahave 1 and 2 records only,respectively.NewPinnulariaspecies were published by Liu et al.(2018b) and Deng et al.(2021).New variety and forms ofCaloneiswere reported by Zhu and Chen (2000).But no new genus or new species from other genera within Pinnulariaceae have been reported.

Xizang is a biodiversity hotspot (Xu et al., 2017;Yu et al., 2020), which also is seeing threats to that diversity (Liu et al., 2021).Diatom research in Xizang began in the early 20thcentury (Mereschkowsky,1906; Skvortzow, 1935).In the 1960s, research expeditions to Xizang culminated in several works,including those by Jao (1964) and Jao et al.(1974)on living diatoms and by Huang (1979), (including a new taxon,Pinnularia molarisvar.brevistriataHuang 1979: 191) and Li (1983) on fossil diatoms.Four taxa were described by Zhu and Chen (1996) and a large work on the diatoms of Xizang was published by Zhu and Chen (2000).In that large treatment,Zhu and Chen (2000) reported 104 taxa belonging to three genera (Pinnularia,Caloneis, andDiatomella)in the Pinnulariaceae, but no new species were described.During the re-investigation of biodiversity of Qinghai-Xizang Plateau, diatom samples were collected and one new genus,Spargeriagen.nov.,with two new species are described as new to science.

2 MATERIAL AND METHOD

Samples were collected from Xizang, China,during a biodiversity investigation from 2018 to 2021.Benthic diatoms were taken from several counties located in north-eastern Xizang.Samples containing the new genus were benthic and collected from hot springs in Nima County (92°09′64.74E, 31°74′37.31N),which is located in northern Xizang, belonging to the plateau subfrigid zone and semiarid climate zone, at an elevation 4 639 m above sea level.

Samples were fixed in 4% formaldehyde in the field.The samples were cleaned with HNO3, then washed and settled using distilled water until the pH was neutral.For light microscope (LM) observations,cleaned diatoms were mounted to make permanent slides with Naphrax.These permanent slides were examined with a Zeiss Imager A2 microscope equipped with a digital camera (AxioCam MRc 5)and observed with differential interference contrast(DIC) optics (Zeiss, Jena, Germany).For scanning electron microscope (SEM) observations, cleaned material was air-dried and coated with gold palladium, and observations made with a Hitachi S-4800 field emission SEM (Hitachi, Tokyo, Japan) at an operating voltage of 15 kV.Diatom images were treated technically with Photoshop 7.0.Cleaned material and permanent slides are archived at the Key Laboratory of Biodiversity of Aquatic Organisms,Harbin Normal University.

3 RESULT

SpargeriaLiu & Kociolek gen.nov.

Description: Valves narrow to wide rectangular,with obtuse to round apices, linear and narrow axial area, and nearly bow-tie shaped central area.Raphe only present on the valve face, barely expanded on to the mantle.Central raphe endings slight expanded, positioned close to each other; terminal raphe fissures short and deflected to the same side of the valve.Striae chambered, wider near the axial area and become narrower near margins, nearly long triangle-shaped, only present on the valve face,multiseriate with 3-4 rows of small and round areolae.Areolae are occluded externally by hymens.Internally, interstriae are simply thickened and the striae are sunken as alveoli.Central sternum is flat,central raphe endings are slightly deflected to the same side, raphe terminates as an almost flat helictoglossa near the apex.

Type species:Spargeria zhuiiLiu & Kociolek

Etymology: Named for its habitat, this new genus was collected from a hot spring.

Remarks: Distinguished byPinnulariaandHygropetraby its simple raphe system, normal size of the distance between two proximal raphe ends,and striae are much wider near the axial area than near the margin.

Spargeria zhuiiLiu & Kociolek sp.nov.Figs.1-4

LM observations: Valves nearly rectangular,with obtuse round ends, length 16.0-27.5 μm, breadth 7.0-9.5 μm (n=20).Raphe filiform, straight in the middle of the valve slightly deflected at the apices.Axial area linear and narrow, central area bow-tie shaped, with one to two very short striae on one or both sides.Striae strongly radiate in valve center,becoming parallel to convergent near the apices.A very short and divergent stria evident near the apices(Fig.1a).Striae 10-12 per 10 μm.

Fig.1 LM views of short and divergent striae (arrows) of Spargeria zhuii sp.nov.

SEM observations: Externally, the raphe is straight (Fig.2a), with short terminal raphe fissures only present on the valve face and deflected to the same side of the valve (Fig.3c & d), central raphe endings dilated (Fig.3e & f).Striae relatively short only present on the valve face, not extending onto the mantle (Fig.2a).Striae multiseriate with 3-4 rows of small and round areolae.Finely structured hymenes are observed on the exterior of each areola(Fig.3a, b, e, & f).Girdle view narrow, band relatively wide, with very 1 or 2 rows of small poroids (Fig.2d).Internally, central raphe endings are straight (Fig.4d) or slightly deflected to the same side (Fig.4e), positioned on a small, rounded central nodule.Terminal raphe fissures are straight and there are no obvious helictoglossae (Fig.4b & c).Interstriae are internally thickened between the striae (Fig.4a).

Holotype: HANU, Slide no.THXZNR17008,illustrated as Fig.1b.Isotype: Key laboratory of Algal Biology, Institute of Hydrobiology, Chinese Academy of Sciences (Wuhan, China), slide No.NR17008-1#.

Type locality: Hot spring in Nima County from Xizang, China.

Etymology: This species was named after the diatomist Huizhong ZHU, who conducted investigations of diatoms from Xizang during 1960s.

Remarks: Distinguished withSpargeria chenisby its larger and robust valve, radiate striae, and one divergent stria near apices.

Spargeria chenisLiu & Kociolek sp.nov.Figs.5-7

LM observations: Valves narrow rectangular,with parallel sides and round apices, length 12.0-16.0 μm, breadth 3.0-4.0 μm (n=30).Raphe filiform,straight in the middle of the valve, appearing deflected at the terminal.Axial area linear, very narrow.Central area rectangular to “bow-tie” shaped.Striae parallel to radiate in the center, becoming parallel to convergent towards the apices, 10-13 per 10 μm.

SEM observations: Externally, raphe straight(Fig.6a), central raphe endings slightly expanded(Fig.6d), terminal raphe fissures deflected to the same side of the valve (Fig.6a-c).Striae narrower near the margin, expanded towards the axial area,relatively short, only present on the valve face, not extending to the mantle (Fig.6a).Striae multiseriate,composed of 3-4 rows of areolae (Fig.6a-d).Areolae are small and round, occluded by hymenes which occur at the exterior of each areola (Fig.6e).Girdle view, band narrow, and poroids absent(Fig.6f).Internally, central raphe endings are straight(Fig.7d), positioned on small and rounded central nodule, terminal raphe fissures are straight and helictoglossae are not evident to very weakly developed (Fig.7b & c).Interstriae are slightly thickened between the striae (Fig.7e).

Holotype: HANU, slide No.THXZNR17010,illustrated as Fig.5c.Isotype: Key laboratory of Algal Biology, Institute of Hydrobiology, Chinese Academy of Sciences (Wuhan, China), slide No.NR17010-1#.

Type locality: Hot spring in Nima County from Xizang, China.

Etymology: This species was named after diatomist Jiayou CHEN, who conducted the investigations of diatoms from Xizang during 1960s.

Remarks: Distinguished by its narrow valves,striae slight radiate in the middle, become parallel or convergent near apices.

4 DISCUSSION

Fig.4 SEM internal views of Spargeria zhuii sp.nov.

During our investigation of the diatoms from the family Pinnulariaceae from Xizang, two species were identified from a hot spring in Nima County.The morphological features of these two species are similar to the genusPinnulariain having multiseriate striae in alveoli.But the features of the raphe (filiform, with slightly curved external terminal raphe fissures that are restricted to the valve face), very narrow valve mantle, nearly long triangle-shaped fields of areolae and presence of hymenate occlusions on the valve exterior showed these two species may not belong to any known members of the Pinnulariaceae, hence we established a new genus,Spargeriagen.nov.

4.1 Comparison with similar genera

Based on valve morphology, the new genus resembles bothPinnulariaandHygropetra.Compared withPinnularia,the most distinctive differences between the new species andPinnulariais the feature of raphe.InPinnularia, raphe may be described as filiform or complex, with long hooked terminal raphe fissures (Round et al., 1990) and various central raphe endings externally, well developed helictoglossae and an intermissio(Krammer, 2000).For the striae and areolae, both genera have alveolate stria, but inPinnularia,longitudinal lines are observed in some species,formed by the smaller opening internally or axial plate covering the striae; these structures are lacking in our specimens.Each stria inPinnulariais nearly equal in size transapically and extend onto the mantle, but the striae are wider near axial area and become narrower near the margins in the new genus from axial area to the margin, and they are only present on the valve face.

Hygropetrais a “small” genus, both in terms of valve size and number of species, which is similar toSpargeria.Both genera are small-sized and have straight raphe branches, as well as chambered and multiseriate striae.However, differences between the two genera are obvious.Hygropetrahas a relatively long gap between two central raphe endings, and the striae usually extend to the mantle(Mayama and Idei, 2009), but in the new genus, for both species, the central raphe endings resemblePinnulariaspecies, and the striae only are present on the valve face, never extending to the mantle.ForHygropetra, the striae are narrower near the axial area and become wider near the margins, composed of 2-4 rows of areolae, while in our new genus the striae are narrow near the margin, composed by 1-2 rows of areolae and become robust toward the axial area, composed by 3-4 rows of areolae.

Position of alveolar occlusions (external versus internal) is considered a feature that distinguishes several lineages from one another amongst naviculoid diatoms sensu lato (Kulikovskiy et al.,2020), especially for small-sized taxa, without much more “conspicuous” taxonomical characters, such as raphe, striae, stigma, etc.Although the position of areolar occlusions was considered an important difference betweenPinnulariaandHygropetra(Mayama and Idei, 2009), the position of the occlusions yet requires more observational data from more species to clarify the character in both genera.The occlusions were barely reported in the genusPinnularia.Hygropetrawas described as“The large areolae in each alveolus are closed between interior and exterior of the valves by vela”in Krammer (2000, p.206) and documented as“occluded internally by a scarcely domed hymenes”in Mayama and Idei (2009, p.291) for their new speciesHygropetra gelasina, and “Hymenes with regularly arranged perforations are located near the inner openings of the areolae” forHygropetra balfouriana.The cleaning and coating process may disturb the morphology of hymenes.Based on observation for the new genus, hymenes are located in or near the valve exterior similar toPinnulariaspecies and differing from the internally-placed hymenate occlusions reported inHygropetra.

Fig.7 SEM internal view of Spargeria chenia sp.nov.

Other small-sized genera with multiseriate stria areDiatomellaandFrankophilaLange-Bertalot(Lange-Bertalot, 1997) as well asSichuaniella(Li et al., 2013).Diatomellaspecies have scalariform valvocopula, which make it very distinctive compared with most biraphid genera.Sichuaniella,an endemic to China (Li et al., 2013; Wu et al.,2020) with representatives in Xizang (Georgiev et al., 2015), has multiseriate striae and two prominent internal pillars of silica, one on either side of the central nodule.It has been recommended to be a member of the Pinnulariacae (Li et al., 2013),although the presence of apical pore fields reported by Georgiev et al.(2015) suggested that this genus may be better placed amongst the Cymbellales, a group with many genera endemic to Asia (Kociolek,2018).Frankophilais a genus with most representatives reported from the southern hemisphere (e.g., Rumrich et al., 2000; Lowe et al.,2006; Furey et al., 2012; Rivera and Cruces, 2015;Van De Vijver et al., 2020).This genus differs fromHygropetraby having very short raphe branches near the apices; it may also have spines around the valve margin (Lange-Bertalot, 1997).The relationship betweenHygropetraandFrankophilaremains unclear, as Mayama and Idei (2009) suggested that they cannot be separated from one another.More work is needed to explore the relationships between them.Comparison between the new genus and similar genera are showed in Table 1.

4.2 Comparison with species of Hygropetra

In China, previously, the genusHygropetrawas observed from Harbin, Heilongjiang Province, NE China only (Li and Qi, 2014).Two taxa were recorded only by Skvortzow (1936, 1938) in the genusPinnularia.Skvortzow reportedPinnularia balfouriana(=Hygropetra balfouriana(Grunow &Cleve) Krammer & Lange-Bertalot) andPinnularia balfourianavar.stauropteraSkvortzow (1936, p.43,16/15), which was later changed into a new combination asHygropetra balfourianavar.staurophora(Skvortzow) Li and Qi (2014).H.balfourianavar.staurophorais similar toSpargeriachenia, but the latter is larger and has coarser striae.Comparison betweenH.balfourianavar.

Table 1 Comparison between Spargeria and similar genera

staurophoraandSpargeria cheniais difficult due to a single line-drawing offered by Skvortzow (1936)and Li and Qi (2014).We went back to the locality as per Skvortzow (1938) to try to findH.balfourianavar.staurophora.Unfortunately, in the over 80 years since Skvortzow made his collection,the region has changed dramatically and all the habitats no longer existed.We were not able to recognize similarities in the flora generally, and could not locate specimens ofH.balfourianavar.staurophoraspecifically.Based on the only linedrawing, it is hard to tell whether the species which Skvortzow described even belongs to the genusHygropetra.

Valve structure and morphological characters of our two new species were compared with previously describedHygropetrataxa (Table 2).The striae in the two new species are very unique, as both of them have striae that are narrower from central to mantle, being “comet shaped”, and all the striae are present in the valve face only, never reach the mantle.Spargeria zhuiihas relatively large and robust valves and can be easily separated from other known species.Spargeria cheniais most similar toH.elongata, the differences between them are related to the outline and the central area, asH.elongatahaving linear-elliptical outlines with convex sides and broadly rounded apices.On the other hand,S.cheniahas narrow rectangular valves with parallel sides, and round apices.The central area inH.elongatais an asymmetric fascia,sometimes with 1-2 shortened striae, whileS.cheniahas a rectangular to bow-tie shaped central area.

Historical research, the research of others more recently (e.g., Li et al., 2010; Georgiev et al., 2015;Liu et al., 2018b, 2019, 2020; Yang et al., 2019) and our recent explorations on diatom biodiversity in Xizang (Ge et al., 2022) have obtained interesting data.Further work is necessary for more complete understanding of freshwater diatom biodiversity in Xizang.

Table 2 Comparison between species in Pinnularifomris and Hygropetra

5 DATA AVAILABILITY STATEMENT

Data sharing not applicable to this article as no datasets were generated or analyzed during the current study.All data generated and /or analyzed during this study are included in this published article.

6 ACKNOWLEDGMENT

The authors thank Ying ZHANG (Harbin Normal University) for her help on observation and identification of the diatoms.Special thanks are given to the anonymous reviewers for their help.