First record of Eptesicus furinalis (Chiroptera, Vespertilionidae) from the state of Maranhão, Brazil Primer registro de Eptesicus furinalis (Chiroptera, Vespertilionidae) en el estado de Maranhão, Brasil

A specimen of Eptesicus furinalis was collected in the municipality of Cândido Mendes, in the state of Maranhão, Brazil. It was a non-lactating adult female, with dark chestnut dorsal coloration, yellow venter, hairless membranes, short and rounded ears, and a pointed tragus. The analysis of the DNA barcode of the COI mitochondrial gene revealed a 99.80 % similarity with the sequence of E. furinalis deposited in the BOLDSystems platform. The combined analysis of the morphological and molecular data confirmed the occurrence of E. furinalis in the state of Maranhão. This extends the known distribution of the species 676.1 km from the nearest recorded locality in the Ceará

Species of the genus Eptesicus exhibit close morphological similarity relationships with Myotis (Tirira 2017).These species are distinguished by the absence of hair at the base of the uropathagio and by the smaller distance between the canine and first molar and reduced number of premolars (pm 1/2) (Reis et al. 2017).These bats are insectivorous, and present sexual dimorphism, with males being larger than the females (Laval and Fitch 1977).
The present study applies molecular tools sequencing mitochondrial Cytochrome Oxidase I (COI) gene, together with morphological and morphometric data, to confirm the occurrence of E. furinalis in the state of Maranhão, which extends the known distribution of this species in Brazil.

MATERIALS AND METHODS
The E. furinalis specimen was collected in an area of dense forest in the municipality of Cândido Mendes (1°27'21" South, 45°43'32" West) on April 26th 2017 by Costa, CLS, Ventura, MCS and Mendes, SB.This municipality is within the Amazon domain, and its vegetation is dominated by dense rainforest (Brasil c2015).The specimen was collected with a mist-net, 3x 12 m, with a 25 mm mesh.The specimen was weighed using a Pesola balance and the basic external and cranial measurements were using a manual calliper Carbografite (0-150 mm, precision 0,02mm).The specimen was classified as a young adult, based on the ossification of the phalangeal epiphyses (Anthony 1988).The identification and morphological and craniometric measurements were obtained followed by Taddei et al. (1998), Barquez et al. (1999), Van Cakenberghe et al. (2002) Díaz et al. (2011) and Reis et al. (2011Reis et al. ( , 2013Reis et al. ( , 2017)).
The skull was extracted through the buccal aperture.Once clean, the skull was clarified with 10 % peroxide and dried in a stove at 30 °C, after which, it was labeled and stored
Paraíba, in João Pessoa, Brazil where it is deposited and received the voucher n°12155.
Total DNA was extracted from muscle tissue using a Wizard Genomic DNA Purification kit from Promega, following the company instructions.The mitochondrial Cytochrome Oxidase subunit I (COI) gene was amplified by Polymerase Chain Reaction (PCR) using the primers LCO-1490 and HCO-2198 described by Folmer et al. (1994).The samples were sequenced by the dideoxyterminal method (Sanger et al. 1977) with a Big Dye kit in an automatic ABI Prism™ 3500 DNA sequencer.The COI gene sequences were edited and aligned using CLUSTAL W (Thompson et al. 1994) using default program parameters.The MEGA X program (Kumar et al. 2018) was used to obtain the Maximum Likelihood (MV) phylogenetic tree and the evolutionary model General Time Reversible (GTR+G+I).The estimated significance of the clusters was assessed by bootstrap analysis (Felsenstein 1985).The database consisted of twelve sequences with 581 base pairs, with species of Eptesicus occurring in Brazil available on online platforms.The species and Genbank access codes are: E. furinalis (JF454656, EF080339, JF448033, EU096738 and KU295475), E. brasiliensis (JF444299 and MG191855), E. chiriquinus (EU096715 and JF448026), E.
fuscus (MG423513 and JF498646).The species Lasiurus ega Gervais, 1856 (KP734216) and L. blossevillii Lesson & Garnot, 1826 (JF446799 and JF448048) were used as an outgroup.The similarity of the COI sequence with those of the Eptesicus was verified on the BOLD Systems V3 (Barcode of Life Data) platform (Ratnasingham and Hebert 2007).The sequence was deposited at Genbank with the acess code OL504970.
A COI sequence of 687 base pairs was obtained from the E. furinalis specimen, which was 99.80% similar to the E. fu- rinalis sequences with which it was compared.This analysis of the COI marker thus confirmed the morphological identification of E. furinalis.The clusters observed in the Maximum Likelihood analysis showed a strongly supported clade with 99% bootstrap clustering E. furinalis specimens from Maranhão (present study), Guyana, Suriname and French Guiana; the other species grouped in separate and well-supported clades (Fig. 3).This is the first record of E. furinalis specimen in the Brazilian state of Maranhão, in the municipality of Cândido Mendes (Fig. 4) extends the known distribution of the species by 676.1 km to the west and north from the nearest known locality at the RPPN Serra das Almas, in the Brazilian state of Ceará (05°05' South, 40°50' West) (Silva et al. 2015)

DISCUSSION
In the present study, E. furinalis was recorded in the Amazon biome of Maranhão, which may be linked to the enormous diversity of insects and the abundance of other feeding resources, which would support the occurrence of the species in this biome (Bernard et al. 2011).The paucity of data on this species in most studies, is likely related to the difficulty of capturing this bat in mist-nets, given that it typically flies at much high levels, and may thus be under-represented in most scientific collections (Reis et al. 2013).
Although E. furinalis appears to have wide distribution (Reis et al. 2007), its occurrence in Brazil is still poorly understood, and many areas have yet to be inventoried ad- in a clean recipient.The study was authorized by the Chico Mendes Institute for Biodiversity Conservation (ICMBio) through licenses IBAMA/SISBIO 42670-3 and 54384-1.The specimen with field number (MRR 128) was fixed in 10 % formaldehyde and preserved in 70 % ethanol at the Laboratory of Genetics and Molecular Biology, Caxias/MA, and then transferred to the mammal collection of the Federal University of
equately.When new areas are surveyed, as in the present study, the occurrence of E. furinalis in a new region is almost invariably confirmed.The E. furinalis specimen collected in the present study in the Amazon forests of western Maranhão extend the known distribution of the species in Brazil 791 km to the north, from Colinas, in Tocantins state, 719.5 km in a

Figure 2 .
Figure 2. Skull of Eptesicus furinalis.a. Dorsal and side view showing the small braincase, b.Presence of poorly developed sagittal crest, c. side and front view of small molars with a series of maxillary teeth.Source: Olímpio, A.P.M.

Figure 3 .
Figure 3. Maximum Likelihood phylogenetic tree generated using the General Time Reversible model based on COI gene data, for species of the Eptesicus genus.The numbers on the branches correspond to bootstrap values.CM-BRA = Cândido Mendes-Brazil, GUY= Guyana, GT= Guatemala, GUF= French Guyana, SUR= Suriname, PAN= Panama, CAN= Canada, and CR= Costa Rica.

Figure 4 .
Figure 4. Geographic distribution of E. furinalis in Brazil.The black dots indicate the known occurrence localities of the species in Brazil, the red dot, the new record, in the state of Maranhão.

Table 1 .
Craniometric measurements (mm) of the Eptesicus furinalis specimen from Cândido Mendes, Maranhão, Brazil (present study), with values recorded in specimens of the same species from the Yucatán Peninsula (Mexico) and Chaco Borealis of Argentina, Bolivia and Paraguay.