Extension of the known range of the snapping shrimp Alpheus christofferseni Anker, Hurt and Knowlton, 2007 (Caridea: Alpheidae)

Abstract Alpheus christofferseni Anker, Hurt and Knowlton, 2007 was described based on four specimens from Atol das Rocas, northeastern Brazil, and one specimen from Bocas del Toro, the Caribbean coast of Panama. Here, we report the collection of two specimens from Pernambuco, Brazil (~ 8°41’S), thus increasing the knowledge on species distribution along the Brazilian coast by approximately five latitudinal degrees south of its type locality (Atol das Rocas, Rio Grande do Norte, ~ 3°51'S). We also report a substantial increase in the known bathymetric distribution from shallow intertidal to approximately 51 m. Notes on morphological variation are provided based on the new material, and the first sequence of the 16S ribosomal subunit gene for the Brazilian coast is provided and compared with congeners.


INTRODUCTION
Alpheus christofferseni Anker, Hurt and Knowlton, 2007 has been reported from Atol das Rocas, Brazil (type locality) and from the Atlantic coast of Panama, within burrows in very shallow waters (up to 1.5 m), occasionally associated with echiuran worms (Anker et al., 2007;De Grave and Anker, 2017).Herein, we report two specimens of A. christofferseni collected on the coast of Pernambuco State, northeastern Brazil, extending the southernmost known distribution for this shrimp in the western Atlantic by approximately five latitudinal degrees.Additionally, we report a substantial increase in the known bathymetric distribution of the species and the first sequence of the 16S ribosomal subunit gene for one specimen from the Brazilian coast, which is compared with sequences of other Alpheus Fabricius, 1798 species and with the sequence of A. christofferseni from Panama.We provide drawings illustrating the diagnostic characters and report some morphological variation observed between newly collected material and the description of A. christofferseni.

MATERIAL AND METHODS
We collected the first specimen (female) in one of several dredgings carried out off Recife (08°13'25.4"S34°37'43.2"W,Fig. 1A) in 2018, at a depth of 51 m, with the aim to survey crustacean fauna associated with substrate of biogenic origin (e.g., calcareous algae, and dead coral).Calcareous algae and dead corals were brought to the laboratory, and the endolithic fauna was carefully extracted with a hammer and chisel, via substrate fragmentation.The single specimen obtained was anesthetized on ice and fixed in 70% ethanol for later identification.
We collected the second specimen (male) in 2019, at Praia dos Carneiros, Tamandaré (08°41'32"S 35°04' 25"W, Fig. 1A), on the reef near the mouth of the Formoso River.The reefs of Tamandaré are a set of beach rocks approximately 1 km offshore from the beach.The Tamandaré reef complex is part of the Environmental Protection Area "Costa dos Corais", the largest Federal Marine conservation area of Brazil (Maida and Ferreira, 2004).The specimen was collected by using artificial refuge structures (ARS) as described by Bochini et al. (2020).
The female specimen was deposited in the Museum of Oceanography "Prof.Petrônio Alves Coelho" of the Federal University of Pernambuco, Brazil (MOUFPE) and the male was deposited in the Coleção de Crustáceos do Departamento de Zoologia da Universidade Federal do Rio Grande do Sul, Brazil (DZ/UFRGS).The male specimen was used for drawings made under a dissecting stereomicroscope equipped with a camera lucida and for genetic data.Drawings of the specimen were made by using Adobe Illustrator CS6, according to the methods described by Coleman (2003;2009).The carapace length (CL, in mm -from the postorbital angle to the posterior margin of the carapace) was measured using a stereomicroscope equipped with a camera lucida and image capture system.
DNA extraction from the male specimen, PCR amplification with specific primers for a region of the 16S ribosomal subunit gene (~550 base pairs), amplicon cleanup, and sequencing were conducted following the laboratory protocols described by Soledade et al. (2019).The obtained sequence and some sequences from the GenBank were used for analysis: one sequence from A. christofferseni from Panama, six sequences from species morphologically close to A. christofferseni belonging to the brevirostris group [Alpheus barbatus Coutière, 1897, Alpheus bellulus Miya and Miyake, 1969, Alpheus floridanus Kingsley, 1878, Alpheus glaber (Olivi, 1792) and Alpheus aff.macroskeles Alcock and Anderson, 1899] and two sequences used as the outgroup (Fig. 1B).Other 16S gene sequences from closely related species were not available in GenBank.Sequences were aligned by using the MUSCLE software (Edgar, 2004) with default parameters, and Maximum Likelihood performed using RAxML (Stamatakis, 2014) on the platform Cyberinfrastructure for Phylogenetic Research (Miller et al., 2010).The topology consistency was measured by the bootstrap method, and the phylogenic trees were viewed and edited in MEGA X (Kumar et al., 2018).Only support values above 75% are shown.A genetic distance (p distance) was calculated in MEGA X (Kumar et al., 2018).The sequence obtained in this study was deposited in GenBank (Fig. 1B).
Habitat.The species is mainly found in burrows on intertidal or shallow seagrass flats (0-1.5 m), associated with echiurans (Anker et al., 2007); reef areas (beach rock) 3-6 m; and substrate of biogenic origin such as calcareous algae, and dead coral, 51 m (this study).
Genetic data.The topology obtained (Fig. 1B) showed a clade formed by the two sequences of A. christofferseni and a clear separation of these from the other species.The intraspecific genetic distance of A. christofferseni was 0.9%, much smaller than the divergence of this species from other species of the brevirostris group (9.9% -14.4%).
Remarks.The distribution range of A. christofferseni is extended by approximately five degrees south from the type locality (Atol das Rocas, ~3°51"S, Fig. 1A).The current southernmost record of the species is Praia dos Carneiros, Tamandaré, Pernambuco (~8°41'S).Furthermore, the bathymetric distribution increased significantly considering the collection of a specimen at 51 m depth.The specimens from Pernambuco (female and male) resemble the holotype.However, some morphological variations between the two specimens from Pernambuco and the holotype were observed, such as the exopod of the third maxilliped (Fig. 2B, C) reaches the penultimate segment of the endopod (vs.not reaching the penultimate segment of the third maxilliped in the holotype); telson (Fig. 2H, I) with mesial spiniform setae 4.5 times longer than the lateral spiniform setae (vs.mesial spiniform setae twice the size of the lateral spiniform setae in the holotype); third and fourth pereiopods (Fig. 2F, G) armed with four spiniform setae along the ventral margin and a pair of distal spiniform setae (vs.five spiniform setae and a pair of distal spiniform setae in the holotype).