Coscinodiscophyceae and Fragilariophyceae ( Diatomeae ) in the Iguaçu River , Paraná , Brazil

A taxonomic survey was carried out on Coscinodiscophyceae and Fragilariophyceae found in the Iguaçu River catchment area within Iguaçu National Park, in the state of Paraná, Brazil. Between September 2007 and August 2008, we collected 24 samples from two stations on the Iguaçu River, upstream and downstream of the falls. We identified 37 taxa, including 22 specific and infraspecific taxa of Coscinodiscophyceae, together with 15 specific and infraspecific taxa of Fragilariophyceae. Melosira ruttneri Hustedt and Fragilaria alpestris Krasske ex Hustedt represent new


Introduction
Diatoms are algae with siliceous cell walls that are fairly well-represented in aquatic systems, in terms of richness as well as abundance (Hoek et al. 1995).The identification of these organisms is complex (Stoermer & Smol 1999) because of variations in the form and frustule ornamentation.In water quality studies, diatoms are excellent bioindicators (Round 1991;Descy & Ector 1999), mainly because of their short life cycle and their selective sensitivity to certain limnological conditions, making them respond promptly to environmental changes (Lobo et al. 2002;Stevenson & Smol 2003).
Taxonomic studies are of extreme importance for expanding knowledge on the biodiversity of diatoms (Lobo et al. 2002).They enable the proposal of indices that might express the ecological significance of the complex relationships among species in the environments studied (Senna & Magrin 1999;Dinnerstein et al. 1995).Floristic surveys of diatoms in the watershed of the Iguaçu River started with Moreira-Filho et al. (1973), Contin (1990) and Lozovei & Shirata (1990), in a stretch of the river near the municipality of Curitiba.Subsequently, other studies of diatoms were conducted along the watershed.Ludwig & Flôres (1995;1997) conducted studies in rivers within the catchment area of the Segredo hydroelectric power plant, and Brassac et al. (1999) studied the area around the Salto Caxias hydroelectric power plant, both identifying centric diatoms.Similar studies were conducted by Tremarin et al. (2009a), in the Maurício River, and by Santos et al. (2011), in the Salto Amazonas River and in an artificial lake in the municipality of General Carneiro.Other studies of diatoms in the Iguaçu River were performed by Brassac & Ludwig (2003;2005;2006), Ludwig et al. (2008) and by Bartozek et al. (2013).At Iguaçu Falls, Metzeltin & Lange-Bertalot (1998;2007) identified 45 taxa, of which only six were centric and none were araphid.
The aim of this study was to inventory the species and taxonomic varieties of the classes Coscinodiscophyceae and Fragilariophyceae in the Iguaçu River, within the catchment area of Iguaçu National Park, thereby expanding the knowledge on the flora of the diatoms of the state of Paraná and providing support for future studies on this community of rheophilic species in freshwater environments.

Material and methods
Iguaçu National Park, which comprises approximately 169765.00ha, is the largest protected area within the Atlantic Forest biome of Brazil.The area has a humid temperate climate (Ibama 1999); the average annual rainfall in the Iguaçu River watershed is approximately 1500 mm/year; and the rainfall distribution is fairly irregular in time and space, without a well-defined wet period (Lactec 2005).The mean annual temperature is 26 °C, monthly averages ranging from 3 °C to 40 °C (Salamuni et al. 2002).
The Iguaçu River watershed, located at the southern portion of the state of Paraná, is the largest drainage basin in the state and extends into the state of Santa Catarina as well as into some areas of Argentina (Maack 2002).The river is 1275 km in length, and its drainage area covers 70800 km 2 and flowing in an east-west direction from the source at the Serra do Mar mountain range to the mouth in the Paraná River (Paraná 2010).
Two sampling stations were selected, in the backwater, on the right bank of the Iguaçu River, in the municipality of Foz do Iguaçu (Fig. 1): -Station 1 (25°35'72"S, 54°23'63"W), located upstream of Iguaçu Falls, with a width of 869.89-1200.00m, a flow rate of 0.424 m.s -1 and a depth ranging from 0.90 m to 4.62 m, depending on the season (Paraná 2010).
A total of 24 samples were monthly collected between September 2007 and August 2008 from subsurface water at both stations (Tab.1).The samples were fixed with Transeau's solution (Bicudo & Menezes 2006) and oxidized by the technique proposed by Simonsen (1974) modified by Moreira-Filho & Valente-Moreira (1981).Microscope slides were prepared using Naphrax ® as mountant (refractive index = 1.73;Brunel Microscopes Ltd., Chippenham, UK).The analysis of the permanent slides was performed with an Olympus BX60 optical microscope coupled to an Olympus DP71 microscope digital camera (Olympus, Tokyo, Japan).The systematic classification and the terminology followed Round et al. (1990) and Houk & Klee (2004).
Frequencies of the taxa were based on the observation of samples in permanent slides and calculated with the following equation: where F is the frequency, p is the number of samples containing the species and P is the total number of samples analyzed.The diatom taxa identified in the 24 samples were classified into the following categories (Dajoz 2005): constant (F ≥ 70% in the samples), frequent (30% ≥ F ≤ 69%), sporadic (10% ≥ F ≤ 29%) and occasional (F ≤ 9%).
The material was deposited in the Herbarium of the Universidade Estadual do Oeste do Paraná (UNOP, Western Paraná State University), Cascavel campus.

Results and discussion
In the taxonomic evaluation, 37 taxa were identified.Of those, 22 belonged to the class Coscinodiscophyceae, within seven families and nine genera, with one taxon identified down to the genus level, 17 identified down to the species level, four identified down to the level of variety and two identified as non-typical taxonomic forms.In the class Fragilariophyceae, we identified 15 taxa, within one family and four genera, with one taxon identified down to the genus level, 13 taxa identified down to the species level and two taxa identified as non-typical taxonomic forms.

Discostella stelligera (Cleve
The morphometry of the specimens studied was coincident with the description of Thalassiosira rudis by Ludwig et al. (2008) for freshwater environments (Iguaçu River and its tributaries).Tuji et al. (2012)  of the rimoportula (Tuji et al. 2012).Spicaticribra differs from the genus Thalassiosira Cleve by the presence of a network of channels with an anastomosed (bifurcated) pattern and without central fultoportulae (Karthick & Kociolek 2011).

Melosira sp.
Fig. 6a-b Valve cylindrical in girdle view, forming non-rectilinear chains connected by the valve faces.Mantle straight to slightly concave with extremities facing outward in the region of the junction with the complementary valve.Frustule with one concave valve face and one convex valve face in order to dock with the adjacent valve, which has the opposing curvature.Valve surface with inconspicuous ornamentations under optical microscopy.MH: 27-32.5 μm; D: 25.3-33.2μm.Occasional taxon, occurring in only 4.2% of the samples.
This taxon was found in only one collection during the study period, and no similar specimens were found in the literature consulted.We opted to identify the taxon only down to the genus level and suggest studies involving a larger number of specimens, examined under electron microscopy to confirm a new species of Melosira Agardh.Occurrence in samples: UNOP 2800.
The morphological characters and the cellular dimensions of Melosira undulata var.normanii are in agreement with the descriptions of Brassac et al. (1999) for the state of Paraná and of Garcia (2009) for the state of Rio Grande do Sul.However, the cellular dimensions differ from those of the species found by Rosa et al. (1994, D: 45.4 μm) for lakes in Rio Grande do Sul.
As reported by Houk (1993), and using the studies of Krammer (1991) and Brassac et al. (1999) as examples, we found that the taxon studied here has been erroneously identified by several taxonomists as Orthoseira roeseana var.roeseana and Orthoseira roeseana morphotype spiralis, because of its complicated interpretation.The taxa have therefore been gathered in two morphological groups, considering the pattern of striae on the mantle and the presence or absence of undulating spirals in the cingulum.Occurrence in samples: UNOP 2708, 2729, 2800.
The specimen found is similar to the material examined by Metzeltin & Lange-Bertalot (2007, pl. 10, fig. 5) and identified as Orthoseira cf.dendrophila.However, Orthoseira epidendron differs from Orthoseira dendrophila (Ehrenberg) Round, Crawford & Mann by displaying dichotomous striae near the cingulum, the spines forming a continuous ring at the valve margin, with ribs that extend into the mantle.Conversely, Orthoseira dendroteres is characterized by uniseriate striae on the mantle and spines separated by areas with small pores near the valve margin (Houk 2003) Valve face approximately circular; circular areolae in radial lines; prominent ocelli present at an opposite position at the valve margin.Two to three rimoportulae at a right angle, with a more central location, transversal to the ocelli.D: short axis 70-93.7 μm, long axis 75.5-108.8μm; S: 11-12 in 10 μm; A: 10-11 in 10 μm.Frequent taxon, occurring in 54.1% of the samples.

Terpsinoë musica
The genera Terpsinoë Ehrenberg and Hydrosera Wallich were both found in the same collection months.As described by Round et al. (1990), the structure and ecology of the genus Terpsinoë are frequently related to those of Hydrosera.Occurrence in samples: UNOP 2705, 2706, 2707, 2708, 2709, 2710, 2711, 2712, 2714, 2716, 2726, 2728, 2731. Fragilariophyceae Round in Round et al. 1990 Valve linear, narrow, with quite round capitate to subcapitate extremities; axial area linear to linear-lanceolate; central area occasionally extending to the margins, with slight bilateral swelling; nearly parallel striae intercalated with those of the opposite margin.AA: 38 μm; TA: 4 μm; S: 13 in 10 μm.Occasional taxon, occurring in only one of the samples.
For this study, we considered the morphological characters of the valve outline and the central area, as described by Patrick & Reimer (1966), who distinguished Fragilaria capucina by its linear shape and thin striae.The study population was identified as F. capucina because the material did not show round attenuated apices typical of F. fragilarioides or unilateral swelling such as that observed for F. vaucheriae (Patrick & Reimer 1966).
This taxon differs from the other named varieties in that it exhibits greater swelling in the median region and thicker striae.The study specimen was coincident with the description provided by Contin (1990) for material also collected from the Iguaçu River (AA: 117.6 μm; TA: 4.75 μm; S: 12 in 10 μm), and with that provided by Patrick & Reimer (1966) for the taxon (AA: 90-120 μm; TA: 3.4 μm; S: 11-14 in 10 μm), except for the difference in the size of the transapical axis, which was larger in our specimen.Occurrence in samples: UNOP 2716.
Identifications were based on the constriction in the central area, which coincided with the illustrations of Patrick & Reimer (1966), Krammer & Lange-Bertalot (1991) and Ludwig & Flôres (1997).Occurrence in samples: UNOP 2705, 2706, 2707, 2708, 2712.Fragilaria socia (Wallace) Lange-Bertalot. Nova Hedwigia 33: 749. 1980. Fig. 50 Valves linear-lanceolate; extremities rostrate, attenuate to subcapitate; axial area linear to linear-lanceolate; central area occasionally extending to the margins, with a slight constriction preceding bilateral swelling in the median portion of the region; striae nearly parallel, intercalated with those of the opposite margin.AA: 25.6-26.5 μm; TA: 3.3-3.5 μm; S: 12-13 in 10 μm.Occasional taxon, occurring in only 8.3% of the samples.Patrick & Reimer (1966) commented that this taxon is similar to Fragilaria rumpens (Kützing) Carlson except for Acta bot.bras.28(1): 127-140.2014.the fact that its length/width ratio is typically smaller than that found in F. rumpens and that it is more lanceolate.Identifications were based on the swelling in the central area, which was more prominent than in similar species.The study material was coincident with that described by Patrick & Reimer (1966) and Contin (1990), which was identified as Synedra socia, except in that the number of striae was higher than that reported by those authors (17 and 15-17 in 10 μm, respectively).Ludwig & Flôres (1997) identified similar material as Fragilaria capucina var.fragilarioides.The species commonly displays valves of larger dimensions, without such a prominent constriction causing bilateral swelling in the median region, as is typical of Fragilaria socia.Occurrence in samples: UNOP 2730, 2800.
The specimen found is similar to the type specimen of Fragilaria fonticola Hustedt illustrated by Simonsen (1987, pl. 320: 21-26, AA: 20-35 μm; TA: 4-5 μm; S: 13-15 in 10 μm), but differs from it by the lanceolate valve outline and by the lanceolate and larger axial area.As shown by Krammer & Lange-Bertalot (1991), F. fonticola also possesses linear to lanceolate valve margins, similar to those shown in the illustrations of Patrick & Reimer [1966, pl. 5: 16 -as Synedra fasciculata var.truncata (Greville) Patrick], as well as in those of Metzeltin et al. (2005, pl. 14: 17-23), who also reported a higher number of striae.Patrick & Reimer (1966, pl: 5, fig. 6) documented a specimen very similar to the one found in this study as Synedra minuscula Grun.The authors described the specimen as having a linear valve, abruptly attenuate, with rounded extremities, and characterize the taxon by the shape, the absence of a central area and the size of the valve (AA: 15-39 μm; TA: 2-3.5 μm; S: 15-18 in 10 μm).In addition, the specimen described by Rumrich et al. (2000, pl. 7, fig. 7) as Fragilaria sp. is very similar to the specimen studied here, although the authors provided no morphological descriptions and it was therefore impossible to confirm other data.
We opted to identify the specimen only down to the genus level, because we found no another specimens with the same morphometric characters in the literature consulted.The specimen found in the present study was unique; whether it represents a variation in the population is uncertain.Occurrence in samples: UNOP 2799.
Smaller specimens of Staurosirella subcapitata (Frenguelli) Morales tend to lose their typical well-lanceolate outline, and become very similar to S. dubia, as seen by Morales & Manoylov (2006), and the two species can be confused without a detailed analysis.Occurrence in samples: UNOP 2730.(Ehrenberg) Willians & Round. Diatom Research 2: 274. 1987. Fig. 58 Valves oval to elliptic; extremities rounded; axial area lanceolate and narrow; striae robust, radial in the apices and almost parallel in the median portion of the valve, alternated with those of the opposite margin; absence of central area.AA: 8.8 μm; TA: 4 μm; S: 10 in 10 μm.Occasional taxon, occurring in only 4.2% of the samples.

Staurosirella pinnata
The specimen found was identified as Staurosirella pinnata because it displayed a valve structure that was more oval, similar to the specimens examined by Ehrenberg (1843).The axial area is pronounced, with coarser striae (Morales 2001).Our specimen was coincident with the descriptions of Fragilaria pinnata var.pinnata of Patrick & Reimer (1966), Krammer & Lange-Bertalot (1991) and Flôres et al. (1999).Morales & Manoylov (2006) stated that the valves of Staurosirella pinnata are smaller and more oval in shape than are those of Staurosirella martyi (Héribaud) E.A. Morales & Manoylov, which are large, elliptic and rarely oval, with rounded extremities.Paull et al. (2008) concluded that it is very difficult to distinguish Staurosira venter (Ehrenberg) Cleve & Müller from S. pinnata and that reliable characters are the width of the valve (mean: S. venter: 4.7 μm, S. pinnata: 3.7 μm) and the length of the areolae of the margin (mean: S. venter: 0.2 μm, S. pinnata: 0.5 μm), which can be visualized with a high-quality optical microscope.
The genus Melosira Agardh includes few freshwater species, represented by only ten taxa, six of which have been recorded for Brazil (Ludwig & Bigunas, 2006).At our study site, we identified three species of Melosira (M.varians, M. undulata var.normanii and M. ruttneri), as well as one population identified only down to the genus level.Melosira varians is one of the most common species of this genus (Round et al. 1990) and is commonly found in eutrophic rivers and lakes (Reynolds et al. 2002).Melosira undulata var.normanii, a taxon whose distribution in Brazilian waters is much more restricted than is that of M. varians (Rosa et al. 1994;Garcia 2009), has been recorded for France (Germain 1981) and for other tropical areas (Krammer & Lange-Bertalot 1991), as well as in large oligotrophic lakes and dystrophic rivers in North America (Manguin 1949).The geographic distribution of M. ruttneri remains largely unknown, and the present study provided its first record for Brazil.Melosira sp.displayed some structures that differ from those found in the available literature.There is a need for further studies, involving electron microscopy and a larger number of specimens, in order to discuss its identity in greater detail.
Questions remain regarding the true identity of the taxon Fragilaria sp., in comparison with the descriptions in the literature (Simonsen 1987;Krammer & Lange Bertalot 1991), because of the absence of illustrations or descriptions of morphologically similar individuals.Further detailed studies are required in order to determine whether this is a new species.

Table 1 .
transferred the species to the genus Spicaticribra Johansen, Kociolek & Lowe.Those authors differentiated Spicaticribra rudis from Spicaticribra kingstonii Johansen, Kociolek & Lowe by the pattern of the areolae in the central region of the valve and by the position Collection dates, UNOP Herbarium accession number of the herbarium samples from stations 1 and 2, upstream and downstream of Iguaçu Falls, respectively and samples collectors.Iguaçu National Park, state ofParaná, Brazil.