List of angiosperm species in an Atlantic Forest fragment reveals collection gaps in Espírito Santo state , Brazil

This study presents a list of angiosperm species in an Atlantic Forest fragment in the southern portion of Espírito Santo state, Brazil, a region that represents a collection gap within the Atlantic Forest. The studied site is a relatively small fragment of 144 ha located within a conservation unit, the Mata das Flores State Park. The site belongs to a conservation priority area for the Atlantic Forest in Espírito Santo, and is under strong anthropic pressure. Of the 239 species listed here, 21 are new records for the state, eight are endemic, and 20 figure either in the country’s or the state’s Red Lists of endangered species. Rubiaceae and Piperaceae were the families with the highest number of species. We show that small fragments that were never inventoried before can reveal a relatively large number of threatened species and that collection gaps need to be filled in order to refine our understanding about conservation priorities within the Atlantic Forest Biome.


INTRODUCTION
Collection gaps affect the knowledge about species conservation statuses and endemism levels, and hamper effective management planning for conservation areas (Grand et al. 2007;Joppa et al. 2011).This is of great concern considering the high endemism levels of Southeast Brazil and that most known species have small distribution ranges (Pimm et al. 2014).Fragments with less than 100 ha correspond to 30-40% of all remnants in the biome (Ribeiro et al. 2009), which has high levels of floristic richness, endangered species, and endemism (e.g., Mori et al. 1981;Peixoto 1992).Most of this knowledge is based on floristic studies and local floras, which can also lead to the description of new species and new occurrences (Baitello 2001;Deble 2005;Deble et al. 2006;Oliveira and Deble 2006;Coelho 2010;Brotto and Baitello 2012).The description and identification of new species are important topics for biodiversity research agendas (see Joly et al. 2014).Moreover, this biome has the highest number of endangered species in Brazil and the southeast portion of the biome houses the majority of these species (Martinelli et al. 2013).
The number of endemic species of an area is directly affected by collection effort (see Murray-Smith et al. 2009;Werneck et al. 2011).It is noteworthy that from the 8,630 endemic species of the Atlantic Forest, 1,104 (12.8 %) are indigenous from Rio de Janeiro state, whereas the neighboring state of Espírito Santo, with nearly the same total area of Rio de Janeiro, have almost half (551) of this value (List of Species of the Brazilian Flora 2014).Despite the increased knowledge about

Lists of species
the Atlantic Forest flora (Stehmann et al. 2009;Forzza et al. 2010), some regions of this biome remain poorly known.Studies about centers of endemism have shown a geographical bias of sampling effort in the central and northern portions of the Espírito Santo state (Murray-Smith et al. 2009;Werneck et al. 2011), while the southern portion have insufficient data of species occurrences.Nevertheless, the floristic composition of the remnants will reflect their history, making each remnant unique (Santos and Kinoshita 2003).
Floristic studies are crucial in order to fill these collection gaps.Field surveys are often viewed as the obvious, though expensive, way to do this.In the absence of information about the species richness, alternative approaches, such as mapping of vegetation cover (Ribeiro et al. 2009), landscape structure (Metzger 2000), or predictive models (Bini et al. 2006) can be used as surrogates for the expensive and time consuming field surveys, in order to subsidize conservation strategies.Such approaches have the advantages of covering larger spatial extents and providing faster solutions.However, systematic field surveys are the obvious and best available solution to increase the knowledge about biodiversity (Margules and Pressey 2000).This is because most of these models are based on local inventories, and they suffer with collection gaps and lack of knowledge about the number of new, endemic or endangered species in a given site.Therefore, even though systematic field surveys have several operational shortcomings, they are the definitive solution to reveal how many species a site holds.Here we report the results of an intensive field survey on a 144 ha of an Atlantic Forest remnant within a collection gap in the southern portion of the Espírito Santo state, Brazil.

Study site
The study was conducted in a 144 ha forest fragment (Figure 1) located in in the Mata das Flores State Park (MFSP; 20°35ʹ54ʺ S, 041°10ʹ53ʺ W), Municipality of Castelo, Espírito Santo state (ES).The area comprises a Lowland Ombrophilous Forest, with mean annual precipitation of 1,200 mm (Oliveira-Filho et al. 2005), and elevation ranging between 100 m and 800 m.The park is adjacent to the urban area of Castelo and is surrounded by cattle farms and coffee crops (IEMA 2012).There are evidences of past selective logging within the Park.Nonetheless, the MFSP is an important corridor connected to two other state parks located at higher altitudes, Forno Grande and Pedra Azul.

Floristic study
Plant samples were collected on a weekly basis from August 2012 to June 2014 through expeditious walks on the studied fragment.Fertile specimens were collected and dried following usual procedures in plant collection (Peixoto and Maia 2013), and identified using specialized literature, consults to specialists (see Acknowledgements) and to the collections of RB, VIES (acronyms follow Thiers 2015), as well as the virtual herbarium REFLORA (http://reflora.jbrj.gov.br/jabot/PrincipalUC/PrincipalUC.do).The samples were deposited at VIES, with duplicates sent to RB.
Family and genera were listed in alphabetical order following APG III (2009), including only indigenous species.Endemic species were marked with a circle (•) and new records for ES with a diamond (◊).Identifications made by specialists based on non-fertile specimens were included in the list and marked with an asterisk (*), without designation of a collector number.The currently accepted names were verified in the List of Species of the Brazilian Flora (LSBF; http://floradobrasil.jbrj.gov.br/).Conservation status was attributed following the Red Book of the Brazilian Flora (RBBF; Martinelli et al. 2013) and the Endangered Species Flora of the Espírito Santo State (ESFES; Fraga et al. 2007).
The LSBF and SL (CRIA, http://splink.cria.org.br/)websites were used as sources to assign a species as a new occurrence to ES.We considered that a species has a new occurrence in ES when it was not listed in LSBF, and a voucher specimen identified by a specialist was not located in herbaria collections using SL.The LSBF was also used to establish a comparison between the species richness of the families in MFSP and ES.The SL was also used to check how many specimens for a particular species are included in herbaria collections.Both databases were consulted after the last updating of the LSBF (between 14 August and 17 August 2015).

RESULTS
The inventory resulted in 61 families, 159 genera and 239 species (Table 1).The families with the highest number of species were Rubiaceae (25 spp.), Piperaceae (19 spp.), Myrtaceae (13 spp.), Moraceae (10 spp.) and Fabaceae (9 spp.).These families comprised about 31% of all identified  (Martinelli et al. 2013) or in the ESFES (Fraga et al. 2007).Amongst Rubiaceae, Rudgea coronata subsp.sainthilaire (Standl.)Zappi (Figure 2a, b) is endemic of the ES, and is indicated as critically endangered (CR) in both RBBF and ESFES lists.This taxon is relatively common in the park, occurring strictly within the forest, and their individuals aggregate in relatively small populations.The occurrence of Rudgea coronata subsp.saint-hilaire in the ES was attested by only two collections, both from the Linhares municipality, northern Espírito Santo.Thus, the collections of MFSP expanded the geographic distribution of this species to the southwards of the state.Another species of the same genus, R. reflexa Zappi (Figure 2c) is considered endangered (EN) in RBBF, but is not endemic to the Espírito Santo, also occurring in Bahia state.Rudgea reflexa and R. coronata subsp.sainthilaire are very common understory species in the MFSP, being exclusively found in this layer of the forest.
None of the Piperaceae species that occur in the MFSP is endemic to Espírito Santo, and Piper piliovarium Yunck is a new occurrence for this state.Individuals of this species occur strictly within the forest and show an aggregated distribution.Piper vicosanum Yunck.(Figure 2d) is the only species categorized under some threat criteria.The species is considered endangered (EN) in the ESFES list.In the MFSP, isolated individuals occur both inside the forest and in roadsides.
Amongst the three species of Myrtaceae that are endemic of ES and occur in the MFSP, two are categorized in some of the IUCN (2013) threat category.Myrcia limae Barroso and Peixoto (Figure 2e) is also an endemic species of the ES, categorized as endangered (EN) by RBBF and vulnerable (VU) by the ESFES lists.This species was previously known by few collections from four municipalities (Conceição da Barra, Nova Venécia, Linhares and Santa Teresa).The distribution of M. limae is therefore expanded in Espírito Santo.Moreover, the specimens collected in the MFSP also add to the knowledge about the morphology of the fruits of this species, which were not described at the species' protologue.Another species of note in the same genus is Myrcia follii Barroso & Peixoto.This species is indicated as critically endangered (CR) in the RBBF and vulnerable (VU) in the ESFES, and was previously known for 20 samples gathered in three municipalities of Espírito Santo.
The genus Dorstenia (Moraceae) comprises four species in MFSP, which apparently occur in different habitats.Topography and light seem to explain the diff erences in the spatial organization of the different species in the park.Populations of D. arifolia (Figure 3a, b) are always ombrophilous and occur in aggregated patches, preferentially in hill slopes.D. bonijesu (Figure 3c) does not show an aggregate pattern of organization.Its isolated individuals occur in both shaded areas of hilltops, and open areas of roadsides.This species was classified as vulnerable (VU) in the ESFES.However, in the MFSP this species is quite common, occurring even in disturbed areas at roadsides.Individuals show a low size variation.Finally, D. elata (Figure 3d, e) occurs exclusively in shaded forest interiors instead.At last, populations of D. hirta (Figure 3f, g) were found in shaded areas and below canopy gaps.In these sites, observed individuals varied from 20 cm to 1 m height, respectively.
Even though Rutaceae presented low species richness in the studied site, Almeidea rubra A.St.-Hil. is the most commonly found understory species, as well as Clavija caloneura Mart.ex Miq.(Primulaceae).The individuals of A. rubra in MFSP show an aggregated distribution in space, occurring from valleys to hilltops.Individuals of C. caloneura, on the contrary, were commonly observed at the valleys, and apparently are not distributed in aggregate populations.

DISCUSSION
Our results show that the conservation value of small sized fragments (ca.150 ha) can be quite high, considering their overall species richness and number of endangered species they can hold.It is noteworthy that such small remnants can comprise about 12% of what remains in forest cover from this biome (Ribeiro et al. 2009).Our results also highlight the need for time consuming, intensive, floristic inventories in order to encompass the flowering time of most plant species occurring in a site.Because snapshot inventories may not detect several plant species with short flowering periods, we concentrated the sampling effort within this 144 ha remnant for a relatively long period (two years), with a high visiting frequency (weekly).
The overall pattern of species richness, especially within the families Rubiaceae, Piperaceae, and Myrtaceae, is that expected for Lowland Dense Ombrophilous Forests (Carvalho 2006) and other physiognomies of the Atlantic Forest (Amorim et al. 2009;Carvalho et al. 1996;Carvalho et al. 2000).Floristic surveys in different vegetation types of Espírito Santo confirm the high richness of Myrtaceae and Rubiaceae in this state (Assis et al. 2004;Jesus and Rolim 2005;Rolim et al. 2006;Saiter et al. 2011).However, the high species richness of Piperaceae was an unexpected result, considering that Myrtaceae is the richest angiosperm family of the Brazilian flora (Forzza et al 2010), and is highly representative in the Atlantic Forest (Mori et al. 1983;Oliveira-Filho and Fontes 2000).A possible explanation for this result is the fact that the increased heterogeneity created by trails and roads, and their related edge effects, seems to favor species of Piperaceae more than species of Myrtaceae.The open areas within the MFSP are represented by roads that cross the fragment and by trails inside the forest reportedly created by motocross practice in the past.Piper species usually occurs near roadsides and in trails inside the forest.The fruit dispersal made by bats is one possible reason to explain this spatial occupation (Thiers and Kalko 2004).These animals predominantly forage in open areas (Coffin 2007), leading seeds to fall during the flight.
The results found in MFSP reveal the importance of small forest fragments in conserving endemic species known for a small number of populations.Many other collection gaps in southeast Brazil deserve future floristic inventories, especially in the southern portion of Espírito Santo state.Field studies on small sized remnants within areas with collection gaps are essential to improve herbaria collections and biodiversity databases of the Atlantic Forest.

Figure 1 .
Figure 1.Location of the studied fragment.Top left, location of Espírito Santo state in Brazil.Top right, location of Castelo Municipality within Espírito Santo.Bottom, location of the fragment within Mata das Flores State Park (800 ha. of total area) in Castelo.

Table 1 .
List of angiosperms from Mata das Flores State Park, ES, Brazil.◊ New records for ES • Endemic species of ES; * Species identified based on non-fertile specimens.Red List (IUCN 2013) Status in Brazil: LC = least concern, VU = vulnerable, EN = endangered, CR = critically endangered.RBBF -Red Book of the Brazilian Flora; ESFES -Endangered Species of the Espírito Santo state Flora.

Table 1 .
Continued.Luber et al. | Angiosperms from Mata das Flores, Brazil species.Twenty-two families were represented by only one species each, totalizing about 9% of all identified species.Considering the criteria previously explained, 21 species are new records for ES, eight are endemic to this state, and 20 are listed as vulnerable, endangered or critically endangered in the RBBF