High richness, new occurrences, and threatened species in a savanna grassland remnant in the largest Brazilian metropolis

Parque Estadual do Juquery is the largest protected remnant of savanna grassland in the metropolitan region of São Paulo (MRSP). We conducted a floristic inventory and created a photographic record of the angiosperm flora of the savanna formations in this area and compared it with those of other Cerrado sites using Jaccard’s similarity index. We present a list with 366 species, distributed in 58 botanical families. Asteraceae and Poaceae were the families with the highest species richness. Eighty-six new occurrences were recorded in the park. According to the São Paulo state Red List, 14 species are threatened by extinction, of which four are presumably extinct. Most species are herbaceous (36.0%) and sub-shrub plants (26.5%). No floristic similarity was observed with other savanna formations in the countryside of the state of São Paulo. Our results emphasize the importance of the Parque Estadual do Juquery for the conservation of savanna grassland formations in the MRSP and the protection of locally endangered species.


Introduction
In the Neotropical region, savannas cover more than two million square kilometers (Mistry 2000) and represent the second-largest vegetation formation in tropical America (Ratter et al. 2006). Among Neotropical savannas, the Cerrado is the most extensive and biodiverse (Ratter et al. 2006), with more than 12,000 plant species (Mendonça et al. 2008) distributed in a mosaic of physiognomies according to the forest-ecotone-grassland concept (Coutinho 1978). This mosaic includes the savanna forest, termed "cerradão", and a gradient of savanna-grassland formations consisting of a mixed flora with forest and grassland elements, termed "cerrado stricto sensu", "campo cerrado", "campo sujo", and "campo limpo" (Coutinho 1978).
Despite the importance of the plant diversity of Cerrado, two decades ago, less than 20% of its area was estimated to be undisturbed (Myers et al. 2000) and approximately 53% had been converted into pasture and monocultures (Klink and Machado 2005). These land cover changes reveal that the Cerrado biome has been highly influenced by anthropogenic pressure, especially agriculture, which could lead to the loss of up to 34% of the current 112 million hectares of remaining natural areas of Cerrado by 2050 (Strassburg et al. 2017). This scenario may be optimistic because it does not consider the current urban expansion rates of cities located in the Cerrado biome (Rumble et al. 2019). Based on the history of degradation of natural landscapes and the threat to species richness and endemism in the Cerrado, this biome has long been considered one of the priority areas for the conservation of global biodiversity (Myers et al. 2000).
Savanna-grassland formations (open habitats) comprise 64.4% of the Cerrado plant taxa, and 39.2% of its richness is represented by herbaceous plants (Filgueiras 2002;Mendonça et al. 2008), which are vital for the maintenance of ecological processes (Hoffmann et al. 2012;Parr et al. 2014). Also, the floral and faunal diversity in these formations (Tubelis and Cavalcanti 2001;Vynne et al. 2011) are essential for the maintenance of provisioning and regulating services in the ecosystem Veldmann et al. 2015;Murphy et al. 2016), mainly hydrological processes (Scherer-Lorenzen et al. 2003;Honda and Durigan 2016). Despite their environmental role, grassland formations have fewer legally protected areas than tropical forest formations and, therefore, are more susceptible to land-use conversion (Veldman et al. 2015). This global reality is also observed in the Cerrado, especially in the state of São Paulo.
The prominent remnants of Cerrado in São Paulo are concentrated in the central region of the state (Durigan et al. 2003), represented mainly by more dense habitats, such as cerrado stricto sensu and cerradão, associated with deep, sandy soils (Ishara and Maimoni-Rodella 2012). Despite this biogeographical concentration, grassland fragments in the metropolitan region of São Paulo (MRSP) have also been reported (e.g., Usteri 1911;Joly 1950;Baitello et al. 2013;Beraldo et al. 2018). In this region is observed a typical flora of open Cerrado habitats, which are very distinct from those found in the central areas of the state (Durigan et al. 2003). The few sites that harbor this flora are relicts of the original vegetation of this biome (Raimundo 2006), and these are necessary for the conservation of rare and threatened species (Baitello et al. 2013;Beraldo et al. 2018). Still, few studies have been carried out in these fragments in the MRSP (e.g., Baitello et al. 2013;Beraldo et al. 2018;Silva et al. 2018). The lack of floristic surveys of the grasslands in the MRSP reveals a gap in knowledge on species diversity, especially for herbaceous plants, needed to understand the physiognomic and ecological characteristics and the conservation status of these areas.
New surveys and botanical lists of the Cerrado grassland vegetation complement the current knowledge of the flora. Moreover, along with the systematization of the existing information, we report data on potentially threatened plant species, which are of great importance for the conservation of these formations' biodiversity. Surveys can be an essential step toward creating identification keys and photographic reference guides for other studies. Information on the floristic composition of grasslands of Cerrado formations in MRSP becomes even more important when considering the surrounding anthropogenic pressure of constant urban sprawl, which potentially leads to loss of habitats and, consequently, of species.
In this study we aim to provide an updated floristic inventory and create a photographic record of the angiosperm flora in the last and largest protected remnant of Brazilian grassland savanna in the MRSP. We also compared this vegetation's floristic similarity with those from other savannas (Cerrado) remnants, especially from São Paulo state.

Study Area
We carried out this study in the Parque Estadual do Juquery in Portuguese (hereafter acronym PEJY), Franco da Rocha, São Paulo state, Brazil (23°20′52″S, 046°41′50″W at its center; Fig. 1A). This park comprises remnants of the Atlantic Forest and Cerrado, with most of the vegetation cover known as campo sujo; this park represents the largest remnant of this Cerrado phytophysiognomy in the MRSP. PEJY is a protected area created in 1993 and with an area of 2,058 ha. The land originally belonged to the State Psychiatric Hospital, which was managed by Dr. Franco da Rocha at the former Juquery Farm (Baitello et al. 2013). Because of the history of low-intensity use (Fraletti 1986), the natural cover of grassland and forest formations of Juquery Farm was maintained, despite the growing urbanization in its surroundings.
In PEJY, the altitude ranges from 730 to 950 m, the mean annual rainfall varies between 1200 and 2000 mm, and the average temperature is between 20 and 21 °C (Baitello et al. 2013). The climate in the region is influenced by the ocean, and there is no defined dry season (Cfb in the Köppen-Geiger classification system; Alvares et al. 2013). The park's geology is within the São Roque Group, with a lithological composition of phyllites and metasiltites and to a lesser extent, with eluvium-colluvium deposits and quartzites (Amorim et al. 2017). Haplyc acrisol and dystrophic cambisol are dominant (Rossi 2017), with a relief characterized by the presence of low hills with flat tops and steep valleys in mamelonized slopes covered mainly by campo sujo vegetation, in addition to campo cerrado in a few areas, and a drainage basin covered by ombrophilous forest (Baitello et al. 2013). These characteristics create a heterogeneous landscape of Cerrado campo sujo interspersed with ombrophilous Atlantic Forest and other savanna formations (Fig. 1B). PEJY also includes springs that are tributaries of the Juquery River and the Paiva Castro reservoir of the Cantareira system, one of the primary water sources that supply the MRSP.

Methods
Sampling and recording the flora. We recorded the flora of PEJY by systematically surveying woody and herbaceous plants as part of a larger project on the area's grassland community structure. We surveyed in plots and sub-plots distributed in areas with a predominance of campo sujo (Fig. 1A). Woody plants with a circumference at ground height (CGH) >3 cm were sampled in 12 plots of 10 × 25 m. For herbaceous and woody plants with CGH <3 cm, in each of these plots we delimited five 1 × 1 m sub-plots 4 m equidistant from each other, totaling 60 sub-plots. These systematic surveys were carried out in the dry (August 2017) and wet (February 2018) seasons.
During these periods, we also made additional monthly surveys of specimens with flowers and fruits during walks, covering roads and areas close to the plots, including campo cerrado. We made photographic records of species in loco during surveys. The images were not treated by any imagen software or any post-production, except for a photo of Polygala pumila Norlind taken in the laboratory under a stereomicroscope. The photographs were all taken at PEJY by the first author and comprise a personal image library spanning 2014 to 2020.
Creating a species list. We identified the specimens collected consulting specialized literature for the plants of the state of São Paulo (e.g., Wanderley et al. 2001Wanderley et al. , 2002Wanderley et al. , 2005Wanderley et al. , 2007Wanderley et al. , 2016Martins et al. 2009) and for specific families and groups (e.g., Caruzo and Cordeiro 2007;Borges and Forzza 2008;Nakajima 2008, 2011;Heiden et al. 2009;Silva et al. 2018). We consulted specialists in the local flora (see Acknowledgments) to identify or confirm species. We compared our samples with those of the Forestry Institute Herbarium (Instituto Florestal; SPSF) and the Reflora Virtual Herbarium (2020). We prepared vouchers of species with flowers or fruits, especially for those considered rare, threatened, or endangered, or that had not been reported at the study area by Baitello et al. (2013). All vouchers are deposited at the Universidade Federal do ABC Herbarium (HUFABC), and additional duplicates are available at the Universidade Federal de São Paulo Herbarium (HUFSP) and the Forestry Institute Herbarium (SPSF).
Species nomenclature and synonyms were verified and updated using the Reflora Project (Flora do Brasil 2020). The conservation status of species followed the São Paulo state's Red List of threatened plants (Mamede et al. 2007) and its updated version (São Paulo 2016).
After creating our species list, we added the species presented by Baitello et al. (2013) for the open formations (campo limpo, campo sujo, campo cerrado, and cerrado strico sensu) to produce the final PEJY flora. We checked the names of species on Baitello et al.'s (2013) list to avoid synonyms.
Floristic similarity. We compared the flora composition with those from other Cerrado remnants in the states of São Paulo, Paraná, and Goiás, and in the Federal District (Table 1). Prior to analysis, we updated the names in the species lists in these areas according to the Flora do Brasil 2020 Project (Flora do Brasil 2020) to avoid synonyms. We carried out a comparison among areas using PAST v. 3.13 software (Hammer et al. 2001) based on the Jaccard similarity index applied to a hierarchical cluster analysis using the UPGMA algorithm (Legendre and Legendre 1998). We also tested our dataset using the Sørensen similarity index. As the structure of our cluster remained the same using both indexes, we only show here the one using Jaccard's similarity index. We did not include in the analysis species occurring in only one of the areas. recently burned areas and roadsides of PEJY. The survey by Baitello et al. (2013) in PEJY found 280 species in open habitats, 79 more than those identified by us. Despite this, only 115 species are common to both surveys. Our research found 86 new species occurring at PEJY that were not listed by Baitello et al. (2013).
The new records represent an addition of almost a third of the species richness previously recorded, totaling 366 species found in open habitats of PEJY. The families Poaceae (26 spp.), Asteraceae (24 spp.), Fabaceae (7 spp.), and Rubiaceae (6 spp.) had the most significant increase in numbers of species compared to the list by Baitello et al. (2013). Our survey doubled the list of threatened species of the park, from seven (Baitello et al. 2013) to 14, and found one more potentially invasive species in PEJY.
Of the total species presented here, including those of Baitello et al. (2013) We provide short descriptions, vernacular names (when present), comments, and geographic distribution in Brazil for 42 species; these are all threatened species and Poaceae species that we collected in our study.
The cluster analysis of the floristic composition of PEJY revealed three large groups (Fig. 2): (1) with the areas in the countryside of the state of São Paulo (Assis, Itirapina, Mogi Guaçu, Pedregulho, Pirassununga, Pratânia, and Santa Rita do Passa Quatro), where sandy and deep soils predominate as well as one seasonal subtropical climate (Table 1); (2) areas in MRSP and Paraná (Butantã, Jaraguá, Juquery, and Ponta Grossa), characterized by shallow clayey soils and a humid subtropical climate (Table 1); and (3) the areas in Alto Paraíso and Brasília, which are present in the core region of the Cerrado biome, with deep soils and a typical savanna climate (Table 1). The floristic similarity was observed (Jaccard >25%, Mueller-Dombois and Ellenberg 1974) between PEJY and the grasslands of Butantã (Fig. 2), currently nearly nonexistent in the city of São Paulo. Despite the PEJY and the Jaraguá State Park both being located in the MRSP, our analysis indicated a dissimilarity (Jaccard < 25%) between their flora. The flora of the Cerrado sites in the countryside of São Paulo state was similar to each other but dissimilar to grassland formations of the MRSP, Ponta Grossa, Alto Paraíso, and Brasília (Fig. 2). The cluster analysis by floristic similarity calculated with the Sørensen index did not change the cluster structure substantially.  Figure 20F Identification. Small prostrate herb, cylindrical stem very branched. Differentiated by its alternate to sub-opposite, sessile or sub-sessile, elliptic-ovate leaves with mucronulate apex, 3.5-7 mm length and 1.5-5 mm wide.

Polygala pumila Norlind
Green-yellowish flowers in a spike.

Discussion
The high species richness observed in PEJY gives this savanna grassland high conservation priority, especially when the park's surroundings have intense anthropogenic pressures. The species richness obtained in our survey was similar to what has been reported for other areas of Cerrado in São Paulo (e.g., Batalha et al. 1997;Tannus and Assis 2004;Almeida et al. 2005;Ishari and Maimoni-Rodella 2012). When our list is combined with the survey by Baitello et al. (2013), we have a glimpse of the megadiversity present on the open formation of the Cerrado of PEJY (Mendonça et al. 2008). We observed a predominance of species from the families Asteraceae and Poaceae, as expected for the open habitats of the Cerrado (e.g., Carvalho et al. 2010;Ishara and Maimoni-Rodella 2012). The species richness by family in descending order was consistent with that observed by Mendonça et al. (2008) for the Cerrado biome: Asteraceae > Poaceae > Fabaceae > Melastomataceae > Rubiaceae > Myrtaceae > Lamiaceae; these data reinforce the importance of PEJY as the last preserved Cerrado grassland remnant in the MRSP.
We found U. brizantha, an invasive exotic species that can cause declines in savanna grassland diversity. This species is one of the main threats to native plant populations due to its fast recover and spread after fires (Durigan et al. 2007;Pereira-Silva et al. 2019), being an especially severe threat to PEJY's rare and threatened species.
The variation in species richness observed between our survey and Baitello et al.'s (2013) study might be  due to differences in sampling methods. Baitello et al. (2013) made the continuous floristic survey in a non-systematic and long-term way for six years.. In our study, plots were systematically sampled for a set amount of time, and these data were complemented by collections made during walks. The use of plots facilitates the finding of small, inconspicuous, rare species (e.g., Richardia schumannii and Polygala pumila) which occur sparingly in the vegetation (Garrard et al. 2008;Chen et al. 2009). This method revealed seven locally threatened species that had not yet been reported from the study area.
The species richness assessment by plant habit indicates that 62.5% of the species sampled are herbaceous or sub-shrub plants. Percentages close to that were observed in other remnants of savanna grassland in São Paulo, such as Pirassununga (72.3%; Batalha et al. 1997) and Itirapina (78.5%; Tannus and Assis 2004). Despite the predominance of non-woody plants, some portions of the rugged and mountainous terrain of PEJY (Amorim et al. 2017) seem to favor the shrubtree vegetation, resembling a campo cerrado/cerrado stricto sensu. Large speciesindividuals dominate in these areas; examples include Dalbergia miscolobium Benth, Kielmeyera coriacea Mart. & Zucc., Roupala montana Aubl., and Plenckia populnea Reissek, which are typical woody plant species of the Cerrado biome (Eiten 1972). As the diversity of soils, reliefs, and physiognomies promotes environmental heterogeneity and consequently higher species richness in plants (Kumar et al. 2006), the richness observed in PEJY might be associated with local habitat diversity, despite its small and isolated area.
The floristic dissimilarity observed between PEJY and the remnants of savanna forest (cerradão) and savanna (cerrado stricto sensu) in the countryside of the state was expected (e.g., Durigan et al. 2003). The cluster analysis by floristic similarity suggests that geographical proximity, soil, and climate may influence the remnants flora composition in this analysis. The temperature, both in Ponta Grossa and the MRSP (Juquery, Jaraguá, and Butantã areas), is mild, and humidity is high due to the greater latitude and proximity to the coast (Alvares et al. 2013). On the other hand, the countryside remnants (Assis, Itirapina, Mogi-Guaçú, Pedregulho, Pirassununga, Pratânia, and Santa Rita do Passa Quatro) have a hotter, seasonal climate, more similar to those observed on the core areas of the Cerrado (Brasília and Alto Paraíso; Alvares et al. 2013). Despite this climatic similarity, we observed a strong floristic dissimilarity between the core sites and the southern sites (São Paulo state).
Other studies also reported floristic dissimilarities (Jaccard <25 %) among grassland formations of Cerrado (e.g., Durigan et al. 2003;Garcia et al. 2009). On the other hand, when comparing only woody formations in the Cerrado of São Paulo, they are similar to each other than when comparing only grassland formations (e.g., Durigan et al. 2003;Pereira-Silva et al. 2006;Carvalho et al. 2010; Ishara and Maimoni-Rodella, 2012). The floristic dissimilarities might be related to the concentration of woody physiognomies in the central-western portion of the state (cerradão and cerrado stricto sensu) on sandy and deep soils (Durigan et al. 2003). The lower similarity of the grassland formations of São Paulo may be associated with the geographic location of fragments, with a disjunction between the eastern (MRSP and Vale do Paraíba) urban matrix and the central-western agrosilvopastoral matrix. The floristic dissimilarity observed between these two areas may be reinforced by the low natural dispersion capacity of small herbaceous species (Levin et al. 2003), which hinders the flow of propagules among communities and reduces the ability to recolonize disturbed grassland areas (Sheth et al. 2020).
The proximity of the grasslands of Butantã (Joly 1950) and PEJY (24 km apart) and their floristic similarity suggest that in the last seven decades, these areas could have been at least functionally connected. Historical evidence indicates that a diverse landscape of grasslands was present on hilltops and Atlantic Forest extended throughout the depressions and valley bottoms (Usteri 1911;Raimundo 2006). A similar scenario currently exists in PEJY (Baitello et al. 2013). Among the species in common between PEJY and the grasslands of Butantã are many species typical of the Cerrado biome, such as Aegiphila verticillata Vell., Baccharis aphylla (Vell.) DC., Byrsonima intermedia A.Juss., Cambessedesia espora (A.St.-Hil. ex Bonpl.) DC., Erythroxylum campestre A.St.-Hil., Echinolaena inflexa (Poir.) Chase, and Loudetiopsis chrysotrix (Nees) Conert (Eiten 1972;Ratter et al. 2006;Ribeiro and Walter 2006). Additonally, some locally threatened species were also observed in both areas, such as Mesosetum ferrugineum (Trin.) Chase, Paspalum erianthum Nees ex Trin., and Polygala pumila. With the growth of São Paulo city and its surroundings, the grasslands of Butantã were quickly replaced by densely urbanized areas, leaving only small fragments of grassland (Raimundo 2006). Therefore, PEJY is of critical importance as the last preserved grassland remnant of Cerrado within Brazil's largest metropolis. PEJY also has historical relevance as an example of the pristine landscape in the MRSP.
The species diversity found in this area by us and a previous study (Baitello et al. 2013) is in contrast to the site's surroundings within the MRSP. The park is surrounded by growing dense urban areas, and threats such as increased fires and the spread of invasive plant species. The park is also relatively small (ca. 2,000 ha) and physically and possibly functionally isolated from other grassland savannas. These urbanized landscape surrounding PEJY reinforce the need for conservation of this grassland savanna and its species, as recommended in other parts of the world (Bond and Parr 2010;Parr et al. 2014;Veldman et al. 2015;Bond 2016). We suggest a combination of in situ conservation strategies, involving the protection and rehabilitation of threatened species, some of which were identified herein. However, ensuring the protection of species genetic resources of this area in its urban context requires local actions for inspection, management, and control of adverse human impacts, such as the indiscriminate use of fire and other recurrent threats in protected areas of the MRSP (Durigan et al. 2007;Arce et al. 2014).
Despite its small size and the human disturbance of its urban matrix, this protected area is vital for the biological conservation of grassland species, including those of restricted occurrence threatened by extinction. The low similarity of PEJY with other areas analyzed demonstrates its importance as one of the last remnants for the conservation of savanna grassland in the MRSP. Our results contribute to the understanding of the current conservation status of the savanna grassland of PEJY at a local scale and demonstrate the need for additional information on this vegetation type in the state of São Paulo. Additional surveys are needed to provide a framework for developing environmental protection and conservation policies for the Brazilian savanna grassland. To ensure the protection of PEJY, we also recommend additional inventories and studies to assess the current conservation status at the regional scale.