Shrimps of the genus Thor Kingsley, 1878 (Caridea, Thoridae): description of a new species using integrative data, remarks on Thor manningi Chace, 1972, and a world identification key

( versus absent in the new species), and the pterygostomial margin rounded and unarmed in T. manningi ( versus angular, with tooth in the new species). Thor manningi is morphologically closer to the western Atlantic Thor dobkini Chace, 1972 and Thor floridanus Kingsley, 1878


intrOductiOn
The small-sized (< 1 cm in total length) shrimps of the genus Thor Kingsley, 1878 have a wide diversity of lifestyles and sexual systems, including some species being symbiotic with anemones or corals and others living in association with seaweed or seagrass (Bauer, 1986;Chace, 1997;Khan et al., 2004). Thor includes 16 valid species (De Grave and Fransen, 2011;Xu and Li, 2014;Anker and Baeza, 2021;WoRMS, 2022): four in the western Atlantic, three in the eastern Pacific, eight in the northwestern Pacific and Indo-West Pacific, and one with circumtropical distribution. Some species are known only from the type locality, such as Thor spinipes Bruce, 1983 (Australia, Northern Territory) and Thor singularis Xu and Li, 2015 (China, Hainan Island). In opposition, Thor amboinensis (de Man, 1888[in de Man, 1887-1888) and Thor manningi Chace, 1972, are widely distributed. Titus et al. (2018) performed a molecular analysis of the circumtropical T. amboinensis and revealed the existence of at least five cryptic lineages: four in the Indo-West Pacific and one in the Western Atlantic. The latter lineage was recently described as Thor dicaprio Anker and Baeza, 2021; and the undescribed Indo-West Pacific species within that complex are treated herein as T. amboinensis.
Thor manningi was initially described from Antigua and new records have extended its distribution to other Caribbean localities, Bermuda, the eastern coast of the USA, Brazil (seamounts of the North Brazilian Chain, Fernando de Noronha, Ceará to São Paulo), and Ascension Island (Central Atlantic) (Chace, 1972;Manning and Chace, 1990;Christoffersen, 1998;Coelho Filho, 2006). The occurrence of T. manningi in Brazil was probably based on the identification key proposed by Chace (1972), which was based on the analysis of specimens from the Caribbean Sea. In fact, a few morphological accounts (and figures) from Brazilian material of T. manningi have been provided by Almeida et al. (2008) based on material from Pernambuco, and the DNA sequences from São Paulo were made available by Terossi et al. (2017;. The taxonomy of this genus has not been completely clarified due to the morphological similarity among species, which hinders species identification, especially when they occur in the same region (Chace, 1972). This author reported difficulty in finding distinctive morphological characters among some species due to the occurrence of protandry. The sexual system of the four species reported from the western Atlantic has already been studied. Two species have been confirmed as gonochoric by Bauer and VanHoy (1996), i.e., Thor dobkini Chace, 1972 and Thor floridanus Kingsley, 1878. Simple protandry has been reported in T. dicaprio (see Baeza and Piantoni, 2010 as T. amboinensis) and protandry with primary males (half the population does not change to female) in T. manningi (see Bauer, 1986).
This taxonomic context led us to analyze specimens previously identified as T. manningi from Panama and Brazil, using a combination of morphological and molecular data, as well as the morphology of the holotype from Antigua. Our findings result in the discovery of a new species of Thor from the Atlantic coast of Panama. In this study, we provide the description of the new species as well as additional morphological observations on T. manningi, including Shrimps of the genus Thor, including new species Nauplius, 30: e2022028 illustration of their mouthparts. Both species were morphologically compared with congeners, and a world identification key for the genus is also provided.

Materials and MethOds
The specimens analyzed are deposited in the following collections: Coleção

Morphological data
Males were identified by the presence of the appendix masculina on the endopod of the second pair of pleopods and/or presence of the prehensile third pereopod, whereas females were identified by the presence of embryos or associated breeding material on their pleon, or by the absence of male diagnostic characters (Bauer, 1986). Mouthparts were dissected from specimens from samples CCDB 2593 (T. manningi), UF 44393, and UF 44518 (new species). Measurements and observations were performed with a Nikon SMZ800 stereomicroscope. The following measurements were taken: carapace length, length of the fifth and sixth pleonal segment, length, and proximal width of the telson, length of the endopod and exopod of the telson, length and greatest width of the scaphocerite, total length of the three articles of the third maxilliped, length from the articles of the first to the fifth pereopod (basis, ischium, merus, carpus, propodus, and dactylus, including the carpus articles of the second pereopod). Spiniform setae on the distal region of the flexor margin of the fifth pereopod were not counted since some specimens presented tufts of simple setae, making it impossible to see the structures. The illustrations of the morphology and the mouthparts were made, respectively, with the aid of a Nikon SMZ800 stereomicroscope and Olympus CX31 microscope equipped with a camera lucida, later vectorized into the Adobe Illustrator ® CS6. Abbreviations are: CL, carapace length; coll. (s), collector (s).
The identification key was based on, and modified from, a previous proposal: Chace (1997), a worldwide key with 10 species; Komai et al. (2015), an Indo-West Pacific key with nine species; and on all original descriptions from the 17 valid Thor species (present study).

Genetic data
Total genomic DNA was extracted from the muscle tissue of the pleon of the specimens, using the Qiagen Extraction Kit, following the manufacturer's recommendation. A region of ~600 base pairs (bp) of the 16S mtDNA (16S) and ~700 bp of the Cytochrome Oxidase subunit I (COI) genes were amplified from the extracted DNA through a polymerase chain reaction (PCR), using specific primers: 1472 (5´-AGATAGAAACCAACCTGG-3´) / 16SL2 (5´-GCCTGTTTATCAAAAACAT-3´) (Schubart et al., 2000;2002) for the 16S; and COL6b (5´-ACAAATCATAAAGATATYGG-3´) / COH6 (5´-TADACTTCDGGRTGDCCAAARAAYCA-3) (Schubart and Huber, 2006) for the COI. PCR products were purified with ExoSAP-IT Kit, following the manufacturer's recommendation, and sequenced by ACTGENE (Análises Moleculares LTDA). All the sequences were confirmed by sequencing both strands, and the consensus sequence was obtained in the software Bioedit 7.0.5 (Hall, 2005). The non-specific readings were corrected manually, when necessary.
Sequences of five specimens of T. manningi were obtained from three locations: one from Panama, two from Brazil (São Paulo and Pernambuco), and three specimens of the new species from Panama. We also included T. manningi sequences obtained from GenBank from São Paulo and Puerto Rico.
Available sequences from other species of Thor (T. amboinensis complex and Thor hainanensis Xu and Li, 2014) and the outgroup composed of Thinora maldivensis Borradaile, 1915 were retrieved from Genbank. This latter species was chosen for its morphological and phylogenetic proximity to Thor Shrimps of the genus Thor, including new species Nauplius, 30: e2022028 (see De Grave et al., 2014 andKomai et al., 2015), although there was only one sequence available for the 16S; and Eualus cranchii (Leach, 1817[in Leach, 1815-1875) that was selected because it belongs to the same family (De Grave et al., 2014, as Thoralus cranchii).
The final sequences were aligned by the MUSCLE method (Edgar, 2004), implemented in the Cyberinfrastructure for Phylogenetic Research (CIPRES) platform (Miller et al., 2010). For each gene, genetic distance matrices and phylogenetic trees were generated separately by the methods of Maximum Likelihood (ML) and Bayesian Inference (BI). The best substitution model was defined by the software JModelTest 2 (Darriba et al., 2012) implemented on the CIPRES platform, using the Bayesian Information Criterion.
ML was performed with RAxML (Stamatakis, 2014), implemented on the CIPRES platform with the standard GTR model. The topology consistency was measured by the bootstrap method and the ML trees were viewed and edited in Mega 7 (Kumar et al., 2016). Only support values above 90 % are shown.
BI was carried out in the MrBayes 3.2.6 (Ronquist et al., 2012) implemented on the CIPRES platform. The data from the obtained substitution model were analyzed in Tracer v1.6. (Rambaut et al., 2018). The first 15 % of the parameters and trees was discarded as burn-in. The final tree was generated, using the software Tree Annotator 1.8.4 (Drummond et al., 2012). The tree was viewed and edited in the software Fig Tree 1.4.4 (http://tree.bio.ed.ac.uk/software/ figtree/). Only support values of posterior probabilities above 0.90 are shown.
The genetic distance matrices were calculated for each gene by pairwise comparisons between sequences, using the p-distance with the software MEGA (Kumar et al., 2016). Etymology. The new species is named in honor of our great friend, Dra. Paula Beatriz de Araujo (Federal University of Rio Grande do Sul, Brazil), who has devoted her career to the study of biology, ecology, and systematics of crustaceans.
Description. Rostrum (Fig. 1A, B) slightly descending, falling short of distal margin of first article of antennular peduncle to reaching distal margin of second article; dorsal margin armed with 4-5 teeth, posteriormost tooth located at about rostral base; ventral margin with minute subapical tooth, rarely 2 teeth or toothless. Supraorbital tooth vestigial or absent. Antennal tooth present, acute. Pterygostomial margin angular, with 1 tooth.

Shrimps of the genus Thor, including new species
Nauplius, 30: e2022028  Cornea generally wider and slightly shorter than eyestalk. First article of antennular peduncle with tooth on median region of ventral margin (Fig. 1D); acute stylocerite of variable length from reaching distal margin of first article of antennular peduncle to distal margin of second article of antennular peduncle (Fig. 1C); stylocerite with tooth on proximal dorsolateral margin; second article of antennular Shrimps of the genus Thor, including new species Nauplius, 30: e2022028 peduncle with tooth on lateral margin with variable length from reaching half to slightly exceeding distal margin of third article of antennular peduncle; third article of antennular peduncle with triangular plate articulated on dorsodistal margin; inner flagellum overreaching outer flagellum; inner flagellum slender and elongate; outer f lagellum stout, with dense row of aesthetascs (Fig. 1A). Antennal basicerite with 2 ventral teeth; carpocerite variable in length, not reaching or slightly exceeding half-length of scaphocerite; scaphocerite 2.5 times longer than median width, overreaching distal margin of third article of antennular peduncle, with tooth on lateral distal margin (Fig. 1E); antennal flagellum long, filiform, articulated slightly broader at base than tip.
First maxilliped ( Fig. 2D) with coxal and basal endites similar in length, covered by numerous plumose setae; both endites with distal margin slightly rounded; undivided endopod, with plumose setae; exopod well developed and undivided, with some plumose setae on proximal region and numerous plumose setae on distal region; proximal and distal lobes of large epipod similar in width. Second maxilliped ( Fig. 2E) with protopodite with plumose setae; endopod with 4 articles, merus 3.1 times as long as dactylus, carpus 3 times as long as dactylus, propodus 10 times as long as dactylus, propodus and dactylus with numerous plumose setae; long, slender, undivided exopod, some plumose setae on proximal region and numerous plumose setae on distal region. Third maxilliped (Fig. 2G) overreaching antennular peduncle; antepenultimate article 0.9 times as long as ultimate article, with some plumose setae, distal region unarmed or with 1 papillose seta ( Fig. 2F) and 0 or 1 spiniform seta, with variable positions; penultimate article 0.3 times as long as ultimate article, with some simple and plumose setae; ultimate article with many simple and plumose setae, tip with 7 or 8 spiniform setae; exopod not reaching distal margin of antepenultimate article, with tuft of plumose setae on tip.
First maxilliped (Fig. 3D) with coxal and basal endites similar in length, covered by numerous plumose setae; distal margin of coxal endite slightly rounded; distal margin of basal endite slightly concave on median region and rounded on tips; undivided endopod, slightly more slender distally, with some plumose setae on proximal region; exopod well developed and undivided, with some plumose setae on proximal region and numerous plumose setae on distal region; proximal and distal lobes of large epipod similar in width. Second maxilliped (Fig. 3E) with protopodite with plumose setae; endopod with 4 articles, merus 5 times as long as dactylus, carpus 3 times as long as dactylus, propodus 12 times as long as dactylus, propodus and dactylus with numerous plumose setae; long, slender, undivided exopod, some plumose setae on proximal region and numerous plumose setae on distal region; podobranch on epipod. Third maxilliped (Fig. 3F) overreaching antennular peduncle; antepenultimate article 0.9 times as long as ultimate article, proximal region with some plumose setae, distal region unarmed or with 1 papillose seta and 0, 1, 2, or 3 spiniform setae, with variable positions; penultimate article 0.3 times as long as ultimate article, with some simple and plumose setae; ultimate article with many simple and plumose setae, tip with 7 or 8 spiniform setae; exopod not reaching distal margin of antepenultimate article, with tuft of plumose setae on tip.
Variations. The variations in the length of structures or number of spiniform setae of the pereopods did not show any relation with geographic distribution. The number of spiniform setae on the distal region of the merus of the third and fourth pereopods and on the telson dorsolateral surface may vary on each side of the body of the same specimen.

Genetic data
Final matrices with 520 bp (16S) and 570 bp (COI) were used in the analyses. The best-fitting substitution model for the 16S was TPM3uf + G, with nucleotide frequencies of A = 0.3623, C = 0.1030, G = 0.1797, and T = 0.3550, and gamma distribution = 0.3070. For COI, the best-fitting substitution model was TPM1uf + G, with nucleotide frequencies of A = 0.2894, C = 0.1585, G = 0.1535, and T = 0.3986, and gamma distribution = 0.2240.
We obtained almost the same topology for the phylogeny using the ML and BI analyses. Thus, we selected the ML tree as the basis for discussions, showing bootstrap supporting values, but also including posterior probabilities (expressed as a percentage) for BI analyses (Fig. 4). Both trees recovered T. manningi and T. paulae sp. nov. as two separate lineages with high support values. The T. manningi clade included specimens from Brazil (Pernambuco and São Paulo), Panama, and Puerto Rico; and T. paulae sp. nov. included only specimens from Panama. For the 16S gene, T. manningi and T. paulae sp. nov. were recovered as sister group (Fig. 4A), whereas for COI gene, T. paulae sp. nov. was included in a clade with specimens from T. amboinensis complex and T. hainanensis, being T. manningi sister group of this huge clade (Fig. 4B).
The genetic distance analysis corroborated the phylogenetic reconstruction results. The intraspecific variation of both genes was null for T. manningi and T. paulae sp. nov. In contrast, the values of interspecific variation between these two species were high (16S: 20 %, COI: 23 %). Such values are similar or even higher when these species are compared to different lineages of the T. amboinensis complex (T.

discussiOn
This study describes a new species from Panama Atlantic coast (previously identified as T. manningi) and provides remarks on T. manningi, including material from Brazil. The phylogenetic analysis (ML and BI) separating two distinct clades with high support values for both analyzed genes, the presence of a supraorbital prominence in T. manningi (versus absent or vestigial in the new species), and the pterygostomial margin rounded and unarmed in T. manningi (versus angular with one tooth in the new species) showed that these species are clearly separated, based on molecular and morphological data. Moreover, the proximal tooth on the dorsolateral margin of the stylocerite is more developed in the new species than in T. manningi.
Thor paulae sp. nov. is the seventeenth species described for the genus (Tab. 1). The new species can be separated from the other four Atlantic species. The new species differs from T. dobkini, T. floridanus, and T. manningi by the absence or presence of a vestigial Shrimps of the genus Thor, including new species Nauplius, 30: e2022028 supraorbital tooth (versus presence of a prominence in the other three species) and by the pterygostomial margin angular with one tooth (versus rounded and unarmed in the other three species) (see Fig. 1A, and Chace, 1972, figs. 57a, 58a, 59a, 60a). Otherwise, T. paulae sp. nov. shares these characters (supraorbital tooth absent and pterygostomial margin angular with one tooth) with T. dicaprio (see Anker and Baeza, 2021). However, T. paulae sp. nov. differs from the latter species in morphology (3 dorsolateral setae on telson versus 4 in T. dicaprio; Fig. 1I; Anker and Baeza, 2021, fig. 1D), by genetics ( Fig. 4; 22 % for COI), and by coloration since the new species does not have the "sexy shrimp pattern" as T. dicaprio and other lineages of the T. amboinensis complex (see Anker and Baeza, 2021; photos of T. paulae sp. nov. are available at http://specifyportal.flmnh.ufl.edu/ iz/, UF 44518 and UF 44393).
The new species is currently restricted to Bocas del Toro, Panama. One of us (FLM) has collected some individuals previously identified as Thor sp. in the STRI vicinities during a taxonomic field activity in August 2011. The material was deposited in the Zoological Collection of the University of Louisiana, Lafayette (ULLZ) and recently moved to USNM. The identity of this material should be checked in the future since we were not able to do so at the moment due to pandemic restrictions. With the new species, four species of Thor have been reported from the Panama Atlantic coast: T. dicaprio, T. floridanus, T. manningi, and T. paulae sp. nov. (De Grave and Anker, 2017;Anker and Baeza, 2021;this study). However, the records of T. floridanus by Heck (1977; could actually refer to T. manningi (see De Grave and Anker, 2017). Our molecular analysis has confirmed the occurrence of at least three species on the Panama Atlantic coast: T. dicaprio, T. manningi, and T. paulae sp. nov. (Fig. 4).
As the new species material has been previously identified as T. manningi, we analyzed specimens of T. manningi from Antigua and Barbuda (holotype), Panama, and Brazil, and added more detail to the original description provided by Chace (1972), mainly regarding mouthparts and pereopods. We provide below some comparative observations on this species: the Atlantic T. dobkini, T. manningi, and T. floridanus differ from all other species of the genus by having a small supraorbital prominence; four species have a strong supraorbital tooth (see Tab. 1) and a supraorbital prominence or tooth is absent in the remaining ten species (Tab. 1).
Thor manningi can be separated from T. dobkini by the distal half of flexor margin of merus of first pereopod unarmed (versus armed with 1 or 2 setae in T. dobkini) (Chace, 1972). Chace (1972) mentioned spines (= spiniform setae) on the proximal half of the flexor margin of the merus of the first pereopod in some specimens of T. manningi and T. floridanus. These setae were also observed in T. manningi. Chace (1972) mentioned that T. manningi could be separated from T. floridanus based on the number of spinules (= spiniform setae) on the dactylus of the fourth and fifth pereopods (3, sometimes 2 or 4, in T. manningi versus and 4 or 5, rarely 3 or 6, in T. floridanus). However, such characters must be analyzed with caution, considering the possibility of overlap. In this study, only a large female of T. manningi was found with 4 spiniform setae on the dactylus of the fourth and fifth pereopods, which agrees with Chace's (1972) report that this condition was rare in the species. Specimens from Brazil have been included in the analysis of T. manningi, expanding the knowledge on the variation of some characters. Analyzing more specimens of T. floridanus is recommended in order to find characters for more precise separation between these closely related species.
Finally, this study included all species with available sequences in a phylogenetic analysis (i.e., only five of the 17 described species), which is not enough to recover and discuss the phylogeny of the genus. Despite this unfavorable scenario, two considerations can be made: T. amboinensis complex is not monophyletic and the hypothesis proposed by Anker and Baeza (2021) that T. hainanensis "possibly representing one of the clades in Titus et al. (2018)" is refuted because here we included the five clades of Titus et al. (2018) and Shrimps of the genus Thor, including new species Nauplius, 30: e2022028 T. hainanensis is clearly separate from them. More studies on the phylogenetic relationships of this genus need to be conducted.

Identification key to Thor species worldwide
Remarks on the proposed key: Xu and Li (2015) mentioned an indistinct tooth on the proximal dorsolateral margin of the stylocerite of T. leptochelus (as Thinora leptochelus Xu and Li, 2015) while Komai et al. (2015) reported no trace of this tooth; Anker and Baeza (2021), when describing T. dicaprio, did not report morphological differences between the species and other T. amboinensis lineages, instead only genetics, geography and coloration.