Encyonopsis linensis Marquardt, Wengrat & C.E. Wetzel, 2015

Wengrat, Simone, Marquardt, Gisele C., Bicudo, Denise C., Bicudo, Carlos E. De M., Wetzel, Carlos E. & Ector, Luc, 2015, Type analysis of Cymbella schubartii and two new Encyonopsis species (Bacillariophyceae) from southeastern Brazil, Phytotaxa 221 (3), pp. 247-264 : 256-261

Encyonopsis linensis Marquardt, Wengrat & C.E. Wetzel , sp. nov. ( Figs 55–69 )

Light microscopy ( Figs 55–65 ): valves slightly asymmetric, slightly dorsiventral, elliptic-lanceolate to linear-lanceolate, dorsal margin convex, ventral margin slightly convex; rounded apices, subcapitate to capitate; length 15.2–22.6 μm, width 3.7–4.7 μm, l/w ratio 3.7–4.8. Axial area narrow, linear to slightly lanceolate; central area only faintly dorsally marked or absent. Raphe branches straight or faintly undulate, linear, about in the median valve region, proximal ends of raphe slightly rounded, dorsally bent; terminal fissures indistinct or ventrally bent. Striae almost parallel to slightly radiating at the center, becoming weakly radiate towards the apices. Dorsal striae 10–12 in 10 μm, ventral striae (9) 10–12 in 10 μm, at the ends 11–13 striae in 10 μm; areolae not discernible in LM; stigmoid absent. Scanning electron microscopy ( Figs 66–69 ): striae uniseriate composed by elongate foramina appearing as narrow slits or varied slits, as Y-shaped or X-shaped ( Figs 66–68 ), turning shorter towards the apices. Stigmoid absent. Raphe slightly sinuous externally, curved to the dorsal side at the center ( Figs 66–67 ); terminal fissures hooked towards the ventral margin ( Fig. 66 ). Inner raphe fissure straight, with an intermissio near the central nodule ( Fig. 69 ). Internal areolae: struts provide structural support to the foramen, whose opening bears three spines at each side ( Fig. 69 ). 42–44 areolae in 10 μm.

Type: — BRAZIL. São Paulo: Lins, marsh, 21º 43’ 53.2” S, 49º 42’ 31.9” W, C. E. M. Bicudo, L. R. Godinho & C. I. Santos, 14 August 2001 (holotype SP!, preparation 355776, Herbário Científico do Estado Maria Eneyda P. Kauffmann Fidalgo, São Paulo, Brazil, depicted in Figs 68–82 . Isotype BR!, slide 4423, Botanic Garden, Meise, Belgium).

Etymology: —Specific epithet after Lins, the municipality from which the type material was collected.

Morphological examination: — Encyonopsis linensis observed in periphyton samples differs from E. sanctipaulensis by being more markedly dorsiventral, the margins more linear, narrower axial area and smaller valves. Encyonopsis linensis resembles E. hustedtii Bahls (2013: 19) in its linear-lanceolate valves, the capitate extremities and the circular central area. However, E. hustedtii has noticeably smaller dimensions (length 14.4–19.6 μm, width 3.3–4.0 μm) and greater striae density in 10 μm (23–26) ( Bahls 2013).

Ecology: —Representative specimens of E. linensis were collected from a marsh with pH water of 6.3 (Lins city), associated with Encyonopsis difficilis , Encyonema silesiacum (Bleisch in Rabenhorst 1861 –1882: no. 1802) D.G. Mann in Round et al. (1990: 667), Placoneis witkowskii Metzeltin, Lange-Bertalot & García-Rodríguez (2005: 200) and Gomphonema lagenula Kützing (1844: 85) . Besides pH, no other limnological information is available about the collecting site.

Encyonopsis difficilis (Krasske) Krammer, Bibliotheca Diatomologica 36: 121, fig. 163: 9–19. 1997 b ( Figs 70–86 ). Basionym: Cymbella difficilis Krasske, Archiv für Hydrobiologie 35: 403, pl. 12, figs 19–21. 1939.

Type: — CHILE. Pond in Dachstein-Westplatte. Syntype: preparation D III 163 KA.

Light microscopy:—( Figs 70–80 ) Valves slightly asymmetric, slightly dorsiventral, elliptic-lanceolate; dorsal margin convex, ventral margin slightly convex; rounded, rostrate to subcapitate apices; length 21.1–34.5 μm, width 5.5–6.5 μm, l/w ratio 3.5–5.0; axial area lanceolate, wider at the ventral side; central area absent; raphe branches straight or slightly undulate, linear, approximately at the median region of valve, proximal ends of raphe somewhat rounded, dorsally bent; terminal fissures indistinct or ventrally bent; striae almost parallel to slightly radiate towards the apices. Dorsal and ventral striae 8–10 in 10 μm, at the ends 9–10 striae in 10 μm; areolae not discernible in LM; stigmoid absent.

Scanning electron microscopy: —( Figs 81–86 ) Striae uniseriate composed by round striae to rounded lineolae ( Figs 81–86 ). Stigmoid absent. Raphe sinuous externally, bent to the dorsal side at the center of valve, ending in very slightly bent, expanded pores ( Fig. 81 ); terminal fissures hooked to the ventral margin ( Fig. 81 ). Inner raphe fissure straight, intermissio near the central nodule ( Fig. 85 ). Distal raphe endings terminating in helictoglossae near the areolae of the last long transapical striae ( Fig. 83–84, 86 ). Shortened striae continuing around the apex ( Fig. 82 ). Internally, the areola opening lacks occlusion. 36–38 areolae in 10 μm.

Morphological examination: —Dimensions of the studied specimens are in agreement with those in the original description by Krasske (1939), based on material collected in southern Chile. Later on, the species was recorded by Krasske (1948) for the State of São Paulo, this population showing more asymmetrical forms. Besides the latter ones, other specimens with larger sizes and more symmetrical valves, similar to those in Krammer (1997b, fig. 163: 14–16), were also currently observed in the São Paulo samples. Encyonopsis difficilis differs from E. difficiliformis , which has a larger valve size and higher striae density in 10 μm.

Ecology: —Representative specimens of E. difficilis were collected from the periphyton of a pond with Eichhornia , Nymphaea and Cyperaceae (São Carlos city) and from a marsh with water pH of 6.3 (Lins city), associated with Encyonopsis linensis , Encyonema silesiacum , Placoneis witkowskii and Gomphonema lagenula . There is no information available about these environments’ trophic state.

Geometric morphometry

Morphological differences between the four studied groups were evidenced by using their valve shape analysis ( Fig. 87 ). Similarity tests (ANOSIM) performed on the Cartesian coordinates of resulting groups in the NMDS, revealed statistically significant differences (p = <0.0001) between the four studied taxa ( Table 2), except for E. schubartii (type and São Paulo population specimens) and for E. sanctipaulensis (Billings and Ribeirão do Campo Reservoir samples), which belong to the same group.

Final remarks and conclusion

Comparison and examination of E. schubartii type material and species complex provided complementary information on some ultrastructural details such as raphe characteristics, striation pattern and shape of areolae, hence revealing some slight differences in these characters among the four species studied.

However, the present species complex showed significant overlapping of both metric and morphological characteristics ( Table 3). Consequently, despite the valve outlines being statistically different, the four species were not clearly separated in the graph ( Fig. 87 ). Besides, there is a considerable overlapping of the small specimens of E. sanctipaulensis (Rio Pequeno branch, Billings Reservoir), which are close to those of E. difficilis , perhaps due to the non-capitate apices and the valve length and width measures. Nevertheless, when LM and SEM photomicrographs are considered, they are clearly different species as discussed before.

Geometric morphological approach was also important to confirm that both populations of the E. schubartii (from São Paulo State, which showed the greatest size variation and consequently in its valve morphology, and that of the nomenclatural type) represent the same species. Moreover, population of E. sanctipaulensis from Ribeirão do Campo Reservoir, that was constant in their morphological characteristics, and that from the Rio Pequeno branch (Billings Reservoir) also represents the same species.

Furthermore, E. sanctipaulensis populations are somewhat similar to E. subcapitata ( Krammer 1997b, fig. 180: 1–4), i.e. the longer specimens showed more elongate and capitate apices, whereas the shorter specimens have shorter, subcapitate apices, thus leading to misidentification. The latter was especially true in the Rio Pequeno branch for E. sanctipaulensis . Such a morphological variability could be associated to the trophic state of the system, since Rio Pequeno branch is the only oligo-mesotrophic water body where the species occurred (higher nutrient availability), whereas the population from Ribeirão do Campo Reservoir (oligotrophic) showed more stable morphological characteristics. Therefore, population analysis was essential for the characterization of the species in the complex.

The application of the geometric morphological approach introduced an important tool for morphological investigations and gave statistical evidence to the identification of the four taxa within the E. schubartii complex. This approach has become a standard tool of taxonomic studies and proved to be a useful tool in clarifying difficult species complexes, such as Achnanthidium Kützing (1844: 75) ( Potapova & Hamilton 2007), Sellaphora Mereschkowsky (1902: 186) ( Falasco et al. 2009), Fragilaria construens var. subsalina Hustedt (1925: 106) ( Cejudo-Figueiras et al. 2011), Hippodonta Lange-Bertalot, Metzeltin & Witkowski (1996: 254) ( Peng et al. 2014), and many others. Furthermore, our results are in line with studies that consider geometric valve shape analysis as a complementary tool and a validation of the chosen criteria for the identification of these species ( Novais et al. 2009, Peng et al. 2014), where the traditional taxonomic studies with LM and SEM are the most important analysis ( Falasco et al. 2009).

Finally, the species of the Encyonopsis schubartii complex can be differentiated by a combination of characters including valve outline, stria density, length/width ratio and, mainly, by the shape of the valve ends: except for E. linensis , which mostly differs by its smaller valves, for the remaining three species the valve ends varied from rounded to subrostrate, not detached from the valve ( E. difficilis ), to rostrate, elongate and prostrate ( E. sanctipaulensis ) or subcapitate and capitate ( E. schubartii ). Regarding ecological preferences, all species of the E. schubartii complex collected in the State of São Paulo occurred in oligotrophic systems (except for the E. sanctipaulensis population from Rio Pequeno branch, Billings Reservoir, an oligo-mesotrophic system). Consequently, up to now, there are not recognizable differences in the ecological preferences for all the taxa within this complex.

Formal taxonomic transfer

Cymbella perpusilla var. moreirae has all the features of the genus Encyonopsis and is particularly very similar to E. schubartii , therefore, the proposal of its transfer to this genus is justified as follows:

Figures

FIGURE 87. Nonparametric multidimensional scaling plot of normalized coordinates for the morphological landmarks digitized on LM images of selected populations of Encyonospsis. Confidence ellipses (α = 0.95) shown.

FIGURES 55–69. Type material of Encyonopsis linensis. Light and scanning electron micrographs.All pictures taken from the holotype population (SP355776). 55–65. LM views showing variation in size and valve outline. 66–67. SEM external view of entire valve showing the raphe and striae structure. 68. SEM external view of central area. Note the elongate foramina appearing as narrow slits and varied slits. 69. SEM internal view detail of areolae and central area with intermissio. LM scale bar = 10 μm (in Figs 55–65).

FIGURES 70–86: Encyonopsis difficilis, São Paulo populations. Light and scanning electron micrographs. 70–80. LM views showing variation in size and valve outline. 81. SEM external view of entire valve showing the raphe and striae structure. 82. SEM internal view of entire valve. Note the shortened striae continuing around the apex. 83–84, 86. SEM internal detail of valve apex with helictoglossa. 85. SEM internal view of central area. Note the intermission. LM scale bar = 10 μm (in Figs 70–80).