Ichthyofauna of the Nascentes do Rio Parnaíba National Park: watershed divide in the main agricultural encroachment area in the Brazilian Cerrado

Abstract The Nascentes do Rio Parnaíba National Park (NPNP), located in Brazilian Cerrado, protects portions of three river basins: headwaters of Parnaíba, a small portion in upper Tocantins-Araguaia, and tributaries of the middle section of the São Francisco. This protected area is in the boundaries of the states of Maranhão, Tocantins, Piauí, and Bahia, also known as MATOPIBA, a region that has established rapid agricultural expansion associated to land conflicts between corporate agriculture, family agriculture, protected areas, indigenous territories, and quilombola communities. The knowledge of the ichthyofauna in this region is scarce while the impacts on the aquatic environments are increasing. We made a survey of fishes in the NPNP and its surroundings areas, estimating richness, endemics, introduced and shared species between river basins. During 2014, 54 sampling sites were visited, 19 in the NPNP and 35 in the surrounding area. Ninety-five species, 51 genera, 22 families, and six orders were recorded, of which 38 (40%) are in the NPNP. One introduced species were recorded in the Parnaíba River basin and no endangered species were registered. Our data corroborate the importance of the network of protected areas for the preservation of Cerrado fishes, including some possibly undescribed and restricted-range species that may be threatened due to intense agricultural encroachment in the MATOPIBA region.


Introduction
The Cerrado is a tropical woodland-savannah in central Brazil under intense anthropogenic pressure, with land conflicts and high deforestation rates, which have been increasing since the 1990s.It possesses a high level of endemism and is considered a global hotspot for biodiversity conservation (Myers et al. 2000, Strassburg et al. 2017).Thenceforth, approximately 50% of its area was transformed into agricultural areas or pastures (Beuchle et al. 2015), with the loss of natural landscapes approximately 47% larger than in the Amazon rainforest in the same period (Carvalho et al. 2019).Among the most intensely impacted areas is the MATOPIBA, an acronym formed by the initials of the Brazilian states of Maranhão, Tocantins, Piauí and Bahia, one of the main areas of agricultural expansion in Brazil, based on high technology productivity of vast soybean and corn plantations (Miranda et al. 2014, Lopes et al. 2020).
Despite the biological importance and the historical deforestation, when compared with other Brazilian biomes, few conservation units are in the Cerrado, protecting less than 10% of its original area.Among the federal protected areas, are the Ecological Station Serra Geral do Tocantins (7,163 km²), Araguaia National Park (5,577 km²), Chapada dos Veadeiros National Park (2,406 km²), Serra das Confusões National Park (8,234 km²), and Nascentes do Rio Parnaíba National Park (NPNP) (7,498 km²).This latter is located on the boundaries of the MATOPIBA states, encompassing three hydrographic basins and freshwater ecoregions, the Parnaíba, Tocantins-Araguaia, and São Francisco rivers (sensu Abell et al. 2008, Brasil 2019) (Figure 1).The NPNP and surrounding areas are considered of great importance for the conservation of Cerrado's biodiversity, given the advance of deforestation due to cash crops and cattle raising (Carvalho et al. 2019).
The Cerrado contributes to 43% of Brazil's surface water, outside the Amazon region (Strassburg et al. 2017), comprising headwaters from at least five hydrographic basins: Amazon, Tocantins-Araguaia, São Francisco, Paraná-Paraguay, and Parnaíba rivers (Lima & Silva 2005, Abell et al. 2008).The knowledge of fish diversity in these drainages is heterogeneous, varying among them and along different stretches of the same basin, usually reflecting the distance from research institutions (Langeani et al. 2009, Dagosta & de Pinna 2019).The richness of fish species in the Cerrado drainages is estimated in approximately 1,200 species (Klink & Machado 2005).
Few ichthyological studies focused on the Tocantins and São Francisco River basins in the MATOPIBA region.The most important was conducted by Lima & Caires (2011) at the Serra Geral do Tocantins Ecological Station and surrounding areas, encompassing headwaters of both drainages, including a shared one (known in Portuguese as 'águas emendadas').As evidence of the exchange of fish species through this natural connection, Lima & Caires (2011) reported the presence of two species that have unequivocally traversed this common headwater, the characin Astyanax novae Eigenmann 1911 and the cichlid Cichlasoma sanctifranciscense Kullander 1983.
As many Cerrado drainages from the region, the MATOPIBA rivers are threatened by the deforestation of riparian vegetation, the input of fertilizers and pesticides from nearby agriculture areas, and the construction of dams (ICMBio 2018, Latrubesse et al. 2019, Chamon et al. 2022).A mosaic of federal and private Protected Areas (PAs) in MATOPIBA encompass several tributaries of one of the largest and least known PAs of the Cerrado (Nogueira et al. 2011, ISA 2015).Thus, this study aimed to inventory the ichthyofauna in the drainages that are protected by the NPNP and its surroundings areas.With this, we intend to expand the knowledge of the fish diversity of the headwaters of the upper Parnaíba and Tocantins drainages and middle São Francisco River basins in the MATOPIBA region that is a knowledge gap presenting high vulnerability due to the intense agricultural expansion.
Entirely inserted in the Cerrado biome, temperatures in the NPNP region vary between 20 °C and 32 °C, with rainfall concentrated from October to March (MMA 2019, Macena et al. 2008).The relief presents the Chapada das Mangabeiras as its main geological formation, which interconnects with the Urucuia Formation, known for its sandstone composition and high soil permeability.These characteristics enable the formation of headwaters in the foothills, in addition to waterfalls and rapids which make up watercourses that later consolidate the drainages in the region (Brasil 2019).
The Parnaíba River headwaters are situated at the foothills of the Chapada das Mangabeiras, which is formed from numerous springs of the Água Quente (between PI and MA), Curriola, and Lontra rivers (PI).To the south of the Chapada das Mangabeiras, at the northern limit of NPNP, are the headwaters of the Sono River, a tributary of the upper Tocantins river basin.The southeast portion of the NPNP (BA) drains the São Francisco river basin, forming the Sapão River, one of the headwaters of the Preto River.However, the headwaters of the Sapão River (São Francisco) and the Galheiros River (Novo River, tributary of the Tocantins River), the region of 'águas emendadas', is situated outside the NPNP according to the shapefiles of the Agência Nacional de Águas e Saneamento Básico (ANA 2021) and federal conservation units by the Instituto Chico Mendes de Conservação da Biodiversidade (ICMBio 2018).

Collection and identification
The collections were carried out in June 2014 at 54 sampling sites distributed in the São Francisco (2), Parnaíba (40), and Tocantins (12) river basins (Table 1, Figure 2), under permits n° 20088/2014 and 30532/2011 issued by the ICMBio/SISBIO.The effort for the collection followed the AquaRAP approach (Rapid Assessment
To record the natural live coloration, some specimens were photographed in aquarium during collections.The specimens were anesthetized in eugenol alcoholic solution (9:1 dilution of ethanol 92° GL and eugenol) (Lucena et al. 2013) and fixed in formalin aqueous solution (9:1 dilution of water and a solution of formalin containing a minimum of 37% formaldehyde).Tissue samples or entire specimens were fixed in ethanol P.A. (99.9%) for molecular studies.
In laboratory, they were transferred to an ethanol 70% solution, and separated into lots.
The specimens were sorted and identified using the ichthyological literature available for the Parnaíba, São Francisco, and Tocantins river basins (Fowler 1941, Menezes 1969, Nijssen & Isbrücker 1976, Kullander 1983, Britski et al. 1984, Ploeg 1991, Lima & Gerhard 2001, Staeck & Schindler 2006, Costa 2016, Melo & Espindola 2016, Ramos et al. 2017a, b, Ramirez et al. 2017, Rocha et al. 2018, Varella et al. 2018, Zanata et al. 2018, Lustosa-Costa et al. 2022).Whenever necessary, the taxonomic term 'aff.' was used for a taxon that has an affinity with another species but that can be recognized as distinct, 'cf.' when additional studies are necessary to confirm the identification, 'gr.' for taxa with affinities to, but which must be distinct from the nominal taxon to which it was assigned, and 'sp.' indicating possibly undescribed species or simply specimens that could not be identified at the specific level (Winston 1999).The collected material was deposited in the ichthyological collection of the Universidade Federal do Rio Grande do Norte (UFRN) (Table 2).
The systematic classification and nomenclature of the species followed Fricke et al. (2023).Endemism was attributed to species restricted to a single hydrographic ecoregion as proposed by Albert et al. (2011).The geographic distribution of species was verified in Reis et al. (2003), Buckup et al. (2007), Ramos et al. (2014), andLima et al. (2017).To ascertain the conservation status, the official national lists of endangered fish species from Brazil were used (MMA 2014(MMA , 2022)).
Of the 95 species recorded, 58 (61%) occurred in only one of the three hydrographic ecoregions that compose the NPNP and its surroundings area.Thirty-three species (34.7%) were identified at the generic level and may represent species belonging to a complex or represent potential undescribed species.Colossoma macropomum (Cuvier 1816) was recorded near the NPNP in the Parnaíba river basin and was the only introduced species recorded in the survey.No recorded species is currently classified in any threatened IUCN category.
In the Parnaíba River basin, 59 species (62%) were recorded in 40 sampling sites, while in the Tocantins River basin 34 species (36%) in 12 localities, and in the São Francisco River basin, 17 species (18%) in two sampling sites (Tables 1-2, Figure 3c).Nineteen localities were sampled in the NPNP, 17 in the Parnaíba, and two in the Tocantins river basins, with 38 species (40%) belonging to 28 genera, 14 families, and six orders (Tables 1-2).In the NPNP, the order Characiformes was the most representative, with 19 species representing 50% of those recorded in the PA, followed by Siluriformes and Cichliformes, with seven species (18%), each.The Characidae family was the richest in the NPNP, with nine species (24%), followed by Cichlidae, with seven (18%) (Table 2).

Discussion
The knowledge of fish diversity in the Parnaíba, São Francisco and Tocantins river basins has advanced considerably in recent decades (e.g., Lima & Caires 2011, Ramos et al. 2014, Lima et al. 2017, Chamon et al. 2022), although the headwaters of these drainages, part of which are located in the NPNP, remain relatively unexplored.As other Brazilian protected areas, the NPNP does not have a management plan and this fish survey can support future environmental policies, in addition to taxonomic, systematics, ecological, biogeographic, and conservation studies of the aquatic biota in the Cerrado.Although Brazil is responsible for the majority of new PAs in the world (approximately 75%) established since 2003 (Jenkins & Joppa 2009), the freshwater fauna has generally not been considered for demarcation strategies (Agostinho et al. 2005).For example, at the Tocantins River basin the number of threatened fish species is greater than of other vertebrates (ICMBio 2018).
No endangered species was recorded in the NPNP drainages and surroundings area.However, it is noteworthy that in the Tocantins basin there are at least 51 threatened species, 42 in the São Francisco, and one in the Parnaíba River basin (Chamon et al. 2022, Lima et al. 2017, MMA 2014, 2022).Additionally, the impacts resulting from agricultural expansion can adversely affect species that inhabit the 'veredas' (i.e., palm swamps dominated by the Mauritia flexuosa L.f ) (Figure 2h).In these habitats, three species of non-annual rivulids were found.This family includes the largest number of threatened freshwater fish species in Brazil (MMA 2014(MMA , 2022)), mainly due to the loss of their shallow and lentic habitats (Costa 2002).Melanorivulus sp. is possibly an undescribed species recorded in some 'veredas' of the NPNP in the Parnaíba and Tocantins drainages (Figure 3n, Table 2).Melanorivulus jalapensis has not been recorded since its original description and was only known from its type locality (Costa 2010).The new records expands its distribution by 55 km to the northeast (Figure 1).Anteriorly known only from two localities (Costa 2016), Melanorivulus parnaibensis were collected in several 'veredas' sampling sites, expanding the range of the species in about 240 km to the south.In the middle and lower Parnaíba, Tocantins and São Francisco River basins, previous studies recorded several introduced species (Ramos et al. 2014, Silva et al. 2015, Garavello et al. 2010, Bartolette et al. 2017, Barbosa et al. 2017).In the NPNP no exotic species was recorded, however, in the surrounding region, juveniles of 'tambaqui' (Colossoma macropomum) were collected.It is a well-known species that has a native range encompassing the western and central Amazon basins, and the Llanos portion of the Orinoco basin (Araújo- Lima & Goulding 1997, Jégu & Keith 1999).Its occurrence out of its native range, as in the rivers of northeastern Brazil, are the result of escapes from aquaculture facilities, since it is one of the most cultivated freshwater fishes in South America.
Thirty-three species were identified only at the generic level as they have diagnostic characters different from the known species in the sampled basins, requiring further taxonomic studies.Some of these were previously identified by Ramos et al. (2014), Silva et al. (2015), and Melo et al. (2016) in the Parnaíba River basin, and by Lima & Caires (2011) in the Tocantins River basin.Recent taxonomic revisions using material of the NPNP expedition from the Parnaíba River basin resulted in the description of four new species, the cascudinho Parotocinclus cabessadecuia (Ramos et al. 2017), the characid Characidium tapuia (Zanata et al. 2018), the small heptapterid Phenacorhamdia cabocla (Rocha et al. 2018), and the armored catfish Hypostomus velhomonge (Lustosa-Costa et al. 2022).
It is estimated that the Parnaíba River harbor approximately 155 species (Ramos et al. 2014, Lima et al. 2017).In the portion of the NPNP that protects the headwaters of this drainage, 20% (32 species) of the basin were recorded.According to Chamon et al. (2022), in the Tocantins-Araguaia River basin there are 751 species.Although the number of species recorded in the Tocantins River basin in the NPNP is relatively low (13 species), this area protects the headwaters, small environments characterized by low diversity and high endemism (Richardson 2019).Finally, the São Francisco River basin, with 304 fish species, and approximately 60% of these are endemic (Barbosa et al. 2017), does not have its headwaters protected by the NPNP.However, it presented a distinct and diverse ichthyofauna, with 17 species recorded in just two localities.
The headwaters and tributaries of the São Francisco River that cross Bahia State remain little studied (Langeani et al. 2009, Silva et al. 2020b).However, some stretches of the São Francisco River in northwestern Bahia harbor one of the largest and best-preserved areas of Cerrado, including streams and 'veredas' (Nogueira et al. 2011), which should be integrated in the mosaic of PAs, known as the Jalapão Region Ecological Corridor Project (ICMBio 2023), in the MATOPIBA region.During collections, the area apparently most impacted by agricultural expansion was in the São Francisco River basin, with vast landscape completely deforested for soybean and cotton plantations (Figure 2l), with some croplands close to the rivers.
While there is no standardization in our sampling effort to support more accurate comparisons across watersheds, the shared species may indicate some evidence.Four shared species were recorded between the Parnaíba and São Francisco River basins (Table 2), possibly evidencing a recent hydrological connection (about 400,000 years) as suggested by Costa et al. (2018) between these drainages (Coutinho-Abreu et al. 2008).Five species were shared between the São Francisco and Tocantins River basins (Table 2).Although some of these species are widely distributed and the taxonomy poorly understood (e.g.Gymnotus aff.carapo, Hoplias aff.malabaricus, and Rhamdia aff.quelen), the occurrence of some species may be due to the natural connection between these basins.Among the species that occur in the upper Tocantins and middle São Francisco River basins are Astyanax novae, Cichlasoma sanctifranciscense, Eigenmannia besouro, Hyphessobrycon diastatos Dagosta, Marinho & Camelier 2014 and Pristella crinogi Lima, Caires, Conde-Saldaña, Mirande & Carvalho 2021 (Garutti & Venere 2009, Lima & Caires 2011, Dagosta et al. 2014, Birindelli et al. 2015, Lima et al. 2021).Lima & Caires (2011) and Dagosta et al. (2014) discussed the connections between the headwaters of the three drainages and the possible routes for the exchange of freshwater organisms, including those between the Novo River (Tocantins basin) and the Sapão river (São Francisco River basin), and between the São Domingos River (Tocantins basin) and the Grande River (São Francisco River basin).Since this is a region with unique hydrographic aspects (preserved 'veredas', clear water rivers used for tourism, and shared headwaters between drainages) and inhabited by many possibly undescribed species, we suggest that additional studies be carried out in the region to assess the possibility of expanding the NPNP encompassing portions of the Sapão River, thereby also protecting part of the fish fauna from the headwaters of the middle São Francisco River drainage.
While some headwaters of the Parnaíba and Tocantins river basins are protected in the NPNP, the intense agricultural expansion, as well as the construction of hydroelectric dams and water diversion projects (Daga et al. 2020, Chamon et al. 2022) may compromise the downstream habitats in the tributaries that drain the MATOPIBA region.To ensure the conservation of the fish species of this watershed divide, the freshwater environments must be included in the creation or expansion of the protected areas (Azevedo-Santos et al. 2019).

Figure 1 .
Figure 1.Sampling sites in the Nascentes do Rio Parnaíba National Park (NPNP) and surroundings in MATOPIBA (black line) in the Brazilian Cerrado (orange).White dots, sampling sites in the NPNP; black dots, sampling sites outside of the NPNP.The sites in Maranhão (MA) e Piauí (PI) states flow to Parnaíba River basin, those in Tocantins (TO) to Tocantins River basin and, those in Bahia (BA) to São Francisco River basin.

Figure 4 .
Figure 4. Taxonomic composition of the fishes from the Nascentes do Rio Parnaíba National Park and surroundings.A. Sampled orders; B. Sampled families.