Ephemeroptera (Insecta) from the metropolitan region of Santarém, Pará, Brazil

Abstract With the objective of improving the knowledge about Ephemeroptera in the North region of Brazil, the group’s first survey was carried out for the metropolitan region of Santarém, Pará, Brazil. The specimens analyzed were collected from 2019 to 2020 in 27 aquatic environments distributed in the region. The nymphs were captured in the streams with aquatic entomological nets, in addition to manual collection on the substrates. Adults were collected using light traps and flight intercept traps. To complement the inventory, a bibliographic survey of species previously recorded in the area was carried out. For this, the Taxonomic Catalog of Fauna of Brazil, the website Ephemeroptera in South America and scientific articles were consulted. As a result, eight families, 23 genera and 31 species/morphospecies were recorded for the region, with 13 new records for Pará and 19 for the metropolis of Santarém. This study demonstrates the relevance of surveys for the knowledge of biodiversity and for understanding the geographic distribution of taxa.


Introduction
Ephemeroptera (Insecta) is one of the most important groups of aquatic insects.Representatives of this order are found in lotic and lentic freshwater ecosystems, associated with various substrate types, and participate actively in nutrient cycling and energy flow.In general, they need clean, cold, and well-oxygenated waters for their survival and permanence in the place (Alba-Tercedor 2015), being considered excellent indicators of good water quality because they are sensitive to environmental disturbances (Barbola et al. 2011).
The order is widely distributed on the terrestrial globe, and there are about 3,500 valid species in 450 genera and 42 families (Sartori & Briittain 2015).According to the Brazilian Fauna Taxonomic Catalog (in Portuguese: Catálogo Taxonômico da Fauna do Brasil), about 432 species distributed in 84 genera and 10 families are registered from Brazil.For Pará state, there were 41 species distributed in 33 genera and nine families (Salles et al. 2022).
The knowledge concerning mayflies (Ephemeroptera: Insecta) in Brazil has increased considerably in the last years.With the increase of taxonomic papers and published geographical distribution, the number of species reported from the country almost doubled in two decades (Da-Silva & Salles 2012, Salles et al. 2022).However, this knowledge is concentrated in some regions (Salles et al. 2022).Most of the studies on the order conducted with material collected in Pará were directed to descriptions of species of some families (e.g., Dias et al. 2007, Gonçalves 2010, Souto et al. 2016, Boldrini et al. 2017).This situation not only hampers any attempts to uncover the diversity of the aquatic biota in the region, but also prevents directly applicable studies, such as biomonitoring for example.
The metropolitan region of Santarém in Para state, Brazil, has been suffering anthropic impacts for years, including deforestation for the planting of soybeans, rice and corn, pisciculture, the release of untreated sewage in water bodies and channeling rivers (Da Trindade & Cordeiro 2011, Soares et al. 2016, Gomes et al. 2017).Therefore, understanding of Ephemeroptera biodiversity in this region is priority and can contribute to the realization of future actions in the face of these impacts, especially in decision making and public policies for conservation of natural habitats.

Study area
Santarém metropolitan region is in the west of Pará state, comprising the cities of Santarém, Mojuí dos Campos and Belterra, covering a territory of 27,285.426km² (Figure 1).It has a tropical monsoon climate and an average temperature of 25.6 °C, with an average relative humidity of 80% and annual rainfall of 2,000 mm (Fapespa 2015, De Andrade 2017).The sampled sites are distributed between the Tapajós and Mojuí rivers basins (Lima 2012), including streams, waterfalls, rivers and lakes ecosystems.The region also has a conservation unit -the Floresta Nacional do Tapajós (Tapajós National Forest), an area of approximately 527,319 hectares, covering the following cities: Aveiro, Belterra, Placas and Rurópolis (ICMBIO 2019).

Collection, Identification and illustrations
The collections were carried out at 27 points distributed in the metropolitan region of Santarém-PA, being 18 in Santarém, seven in Belterra and five in Mojuí dos Campos.The mayflies were collected between June/2019 and October/2020.Nymphs were collected with aquatic entomological nets in addition to manual collection with forceps directly from de substrates.At each collection point, the largest number of possible substrates (sand, gravel, bottom litter, stones, marginal vegetation) was sampled.Imagos and subimagos were collected with light traps (white sheet illuminated with a 15W emergency lamp, Pennsylvania trap and Malaise trap).The subimagos collected alive were kept for about twelve hours in empty microtubules, until they moulted to the imago stage, and then they were fixed in 80% ethanol.
The identifications were based on taxonomic keys (e.g., Domínguez et al. 2006, Salles 2006, Salles et al. 2018) and updated articles for each group (e.g., Boldrini et al. 2018, Araújo & Dias 2020, Oliveira et al. 2020).When needed, structures of the bodies were dissected and mounted between slide and coverslip, using Euparal ® as the mounting medium.The specimens are deposited in the Laboratory of Ecology and Taxonomy of Aquatic Invertebrates (UFOPA).
Multilayer photographs were obtained using a Leica M165C stereomicroscope in conjunction with Leica DFC 420 image capturing equipment and LED dome lighting for uniform reflection of light on the specimens (Kawada and Buffington 2016) and an Olympus BX51 compound microscope in conjunction with an Olympus digital image acquisition system (DP 72 model using the Cell D program).The final images were generated using Digital Leica Application Suite v.3.7 and Helicon Focus (6.7.1 Pro) software.

Geographical distribution and material examined
The geographic distribution of the taxa collected in the present study was compiled from the Brazilian Fauna Taxonomic Catalog (Salles et al. 2022), from the Ephemeroptera in South America website and from several articles using data of collections carried out in the region.The new records for Pará state were marked with a positive symbol (+) and the new records only for the metropolis of Santarém were marked with a black asterisk (*).

Results
Based on the literature and new samples, a total of eight families, 23 genera and 31 species/morphospecies have been compiled for the metropolitan region of Santarém, with 13 new records for Pará and 19 new records for the metropolitan region of Santarém (Table 1).
Comment.Species described by Lugo-Ortiz & McCafferty (1996a) based on material collected in Paraná do Tapará, near Santarém, but not collected during the sampling of the present study.
Geographic distribution.North: Amazonas, Pará + .Comment.Until now, this species was registered only for its typelocality (Amazonas) (Salles et al. 2011).According to the description presented in Salles et al. (2011), the costal area of the anterior wing of A. maculata has four veins.However, based on the material collected in this study, it was observed that this characteristic is variable, with specimens also showing five veins (Figure 2E).This is the first record of this species for Pará state.
Comment.This is the first record of the genus and species for Pará state.
Comment.This is the first record of this species for Pará state.Comment.This is the first record of this species for the Pará state.
Comment.This is the first record of this species for the state.
Comment.This species was registered for Pará for the first time by Molineri (2002) as Tricorythodes australis, for Altamira municipality.Later, Dias et al. (2005) proposed a new genus to allocate this species and recorded its occurrence for Oriximiná (PA).This is the first record for the metropolitan region of Santarém.
Comment.This is the first record of this species for Pará state.Geographic distribution.North: Pará*.Northeast: Maranhão.Midwest: Goiás.
Comment.Species described by Domínguez et al. (1996) based on material collected in Altamira (PA) municipality.This is the first record for the metropolitan region of Santarém.
Comment.This is the first record of the genus and species for Pará state.
Comment.This is the first record of the genus and species for Pará state.
Comment.This species was previously registered for Pará state by Nascimento et al. (2017), occurring in Santarém.Comment.This species was registered for Pará state by Molineri & Salles (2017), for Tucuruí municipality.This is the first record for the metropolitan region of Santarém.
Comment.This species was registered for the municipality of Santarém by Lestage (1923); however, it was not found during the sampling of the present study.Comment.This is the first record of this species for Pará state.

Discussion
Ephemeroptera is a group of aquatic insects well studied in some locations due to the permanence of researchers in local institutions and because this is a key group in studies of environmental quality (Alba-Tercedor 2015).Despite the great ecological importance of the group, most of applied studies deal with Ephemeroptera only at the level of genus or family (Chen et al. 2017, Nicacio et al. 2020).
The metropolitan area of Santarém presented a representative diversity of Ephemeroptera for the state, which may be related to the variety of habitats among the sampled sites, mainly in places far from the urban areas of the city.Environmental heterogeneity is considered one of the main factors to explain the high richness of taxa in an area (Chisholm et al. 2011), as they provide more resources and niches for species (Bazzaz 1975).However, it is important to emphasize that the metropolitan region of Santarém is experiencing great anthropic pressures (Sousa et al. 2020) and that a large part of the natural environments is at risk of destruction.
Until the completion of this study, a total of 41 species, distributed in 33 genera and nine families were registered for Pará, of these, only 10 species, 16 genera and six families were registered for the metropolis of Santarém.After this research, Pará state is now represented by 54 species/morphospecies, 36 genera and nine families; the metropolis of Santarém is now represented by 31 species/morphospecies, 23 genera and eight families.Thus, with the present study, there was an increase of 23% in the number of new records of Ephemeroptera registered for Pará state and an increase of 65% in what there was registered for the metropolitan area of Santarém (Table 1).And the metropolitan region of Santarém becomes the area with the largest number of Ephemeroptera  Baetidae and Leptophlebidae were the most representative families in the metropolitan region of Santarém, with 11 and eight species/morphospecies, respectively, followed by Caenidae (n = 3), Polymitarcidae (n = 3), Leptohyphidae (n = 3), Euthyplocidae (n = 1), Ephemeridae (n = 1) and Coryphoridae (n = 1).This pattern is in line with general studies on the order Ephemeroptera in Brazil, where Baetidae is always identified as the most representative family, followed by Leptophlebiidae and Leptohyphidae (e.g., Francischetti 2007, Salles et al. 2010, Shimano et al. 2011, Lima et al. 2012).
Cloeodes, Hermanellopsis and Tupiara stand out as the first records for the state of Pará.Even if recorded for some Brazilian states (Salles & Boldrinni 2022), on a more precise geographic scale, we still see gaps in their distributions, probably because of sampling being concentrated in certain locations.
Although the three cities are seen as a single region, Hydrosmilodon gilliesae and Simothraulopsis inaequalis were the only species sampled in the three cities.Callibaetis gelidus, Caenis cuniana and Campsurus lucidus (and others), for example, were sampled in only one of the cities.Likewise, some taxa registered in previous (ecological) studies carried out in the region were not collected in the present study.Some points were sampled several times to obtain nymphs or adults for the complete identification of the species.Even so, despite the efforts, it was not possible to obtain the necessary stage.These observations underscore the importance of constant surveys with increased sampling efforts to obtain complete knowledge about the diversity of Ephemeroptera in the region.
The results presented in this study increased, in general, the knowledge about the diversity and distribution of Ephemeroptera.This knowledge is fundamental for the understanding of aquatic diversity, both in terms of naming the taxa, given future descriptions of new species collected, and for understanding the distribution of these taxa.In addition, the naming of taxa allows greater accuracy in ecological studies, including studies of aquatic biomonitoring in the region.

Figure 1 .
Figure 1.(A-C) Maps.(A) Brazil.(B) Pará state in detail.(C) Metropolitan Region of Santarém with the location of the sampled sites.

Table 1 .
Ephemeroptera (Insecta) recorded for the Metropolitan Region of Santarém.New records: (+) new record for the state; (*) new record for the metropolitan region of Santarém; (?) locality not specified.