Description of Brooksia lacromae sp. nov. (Tunicata, Thaliacea) from the Adriatic Sea Author Garić, Rade Institute for Marine and Coastal Research, University of Dubrovnik, Kneza Damjana Jude 12, 20000 Dubrovnik, Croatia. & Email: rade. garic @ unidu. hr rade.garic@unidu.hr Author Batistić, Mirna Institute for Marine and Coastal Research, University of Dubrovnik, Kneza Damjana Jude 12, 20000 Dubrovnik, Croatia. & Corresponding author: mirna. batistic @ unidu. hr mirna.batistic@unidu.hr text European Journal of Taxonomy 2016 2016-05-12 196 1 13 journal article 21967 10.5852/ejt.2016.196 0d3456bf-150b-4cd2-ae80-e050a4852458 2118-9773 3837952 38CC4797-768A-44C5-8B85-EA3DCB6678CB Brooksia lacromae sp. nov. urn:lsid:zoobank.org:act: 2B9ACCBD-8B20-437D-B013-468F8E956864 Figs 1–4 Diagnosis Oozooid (solitary form; Fig. 2 ) Muscles II and III fuse dorsally before fusing with muscle I mid-dorsally. Muscles III and IV fuse briefly dorsolaterally. Muscles IV and V fuse before approaching muscles VI and VII mid-dorsally. There is one longitudinal ventral muscle which extends anteriorly into the proboscis. In addition, the proboscis contains two lateral longitudinal muscles. The ventral longitudinal muscle branches posteriorly into two branches. There are two slit-like openings in the ventral longitudinal muscle, one below the anterior end of the endostyle, and the other slightly anteriorly from the first one. Muscle I fuses with branches of the ventral longitudinal muscle at their anterior part, while muscles VI and VII fuse with them posteriorly. Muscles II, III, IV and V converge ventrally towards the junction between muscle I and the branches of the ventral longitudinal muscle. Muscle II can be slightly fused with muscle I just before its connection with branches of the ventral longitudinal muscle. Endostyle straight. Stolon emerges from the test ventrally, slightly posteriorly from the nucleus. There is one test processus just posteriorly from the stolon ( Fig. 4D ). Muscles I–VII with 52–58 muscle fibers in total, measured laterally in two examined individuals. Fig. 2. Oozooid of Brooksia lacromae sp. nov. A . Dorsal view, drawing. B . Ventral view, drawing. C . Dorsal view, photograph of an individual dyed with Janus Green, with an arrow indicating breakage in the proboscis during collection. Abbreviations: as = atrial siphon; bl = branching of the ventral longitudinal muscle; br = branchial bar; dt = dorsal tubercle; en = endostyle; g = ganglion; im = intermediate muscle; lm = longitudinal muscle; lo = openings in the longitudinal muscle; nu = nucleus; os = oral siphon; p = proboscis; pb = peripharyngeal band; st = stolon; tp = test processus; I–VII = body muscles. Blastozooid (aggregate form; Figs 3 , 4 A–C) Sinistral individual – Four muscles on the left side, three muscles on the right side. Dorsal IM1 continues ventrally as the ventral muscle IM1, without extending to the anterior attachment processus (ap1). A branch of dorsal muscle IM1 (IM1-a) extends to the anterior attachment processus (ap1). Dorsal muscles IR, IIR and IIIR branch dichotomously. Dorsal muscle IR-a continues ventrally as ventral muscle IR. Dorsal muscle IR-b continues ventrally as muscle IIR-c. A small branch from muscle IR-b/IIR-c enters the lateral attachment processus (ap2). Dorsal muscle IIR-a continues ventrally as ventral muscle IIR-a. Muscle IIR-b ends blindly on the ventral side of the animal without entering the posterior attachment processus (ap3). Dorsal and ventral muscles IIIR enter the posterior attachment processus (ap3) without joining. Muscles IVL-a and IIIR-a extend from muscles IVL and IIIR, respectively, towards the nucleus. Endostyle curved anteriorly. Muscles IL-IIIL, IR-a,b and IIR-a,b,c with 3 muscle fibers each in all examined individuals. Muscle IVL with from 3 to 9 muscle fibers, muscle IIIR with from 5 to 7 muscle fibers. Fig. 3. Drawing of the blastozooid of Brooksia lacromae sp. nov. (sinistral individual). A . Dorsal view. B . Ventral view. Abbreviations: ap1–3 = attachment processes; as = atrial siphon; br = branchial bar; dt = dorsal tubercle; e = embryo; en = endostyle; g = ganglion; h = heart; nu = nucleus; os = oral siphon; pb = peripharyngeal band; I L –IV L = left side muscles; I R –III R = right side muscles; IM 1 and IM 2 = intermediate muscles. Dextral individual Mirror image of sinistral individual. Etymology Brooksia lacromae sp. nov. is named after the island of Lokrum near which it has been found. The Latin name of the island of Lokrum is Lacroma. Material examined Two oozooids and 19 blastozooids. Holotype One oozooid ( 14.8 mm body length, 23.9 mm total length including proboscis), collected from a 0–50 m depth layer on 3 Oct. 2014 in a 53 μm mesh plankton net. It is deposited in the Tunicata collection of the Croatian Natural History Museum under inventory number CNHM Inv. br. 44/1. Fig. 4. Brooksia lacromae sp. nov. A . Photograph of the ventral side of the dextral blastozooid. B . Photograph of the dorsal side of the sinistral blastozooid. C . Photograph of the lateral side of a blastozooid. D . Detail of the ventral posterior end of an oozooid. All indivuduals were dyed with Janus Green dye. Abbreviations: st = stolon; tp = test processus. Allotype One sinistral blastozooid ( 4.2 mm body length, without attachment processes), collected from a 0–50 m depth layer on 3 Oct. 2014 in a 53 μm mesh plankton net. It is deposited in the Tunicata collection of the Croatian Natural History Museum under inventory number CNHM Inv. br. 44/2 Paratypes One oozooid ( 11.5 mm body length, 18.2 mm total length including proboscis) and one dextral blastozooid ( 4.8 mm body length, without attachment processes), collected from a 50–100 m depth layer on 3 Oct. 2014 using a 53 μm mesh plankton net, deposited in the Institute for Coastal and Marine Research (University of Dubrovnik) under inventory number IMP-002; one dextral blastozooid ( 4.1 mm body length, without attachment processes) collected from a 0–50 m depth layer on 3 Oct. 2014 with a 200 μm mesh plankton net, deposited in the Dubrovnik Natural History Museum under inventory number PMD 2106. Other material 16 blastozooids deposited in the Institute for Coastal and Marine research (University of Dubrovnik) under inventory number IMP-003. Type locality 42°37'21"N , 018°06'05"E , off Dubrovnik, South Adriatic (Mediterranean Sea; Fig. 1 ). The temperature average in the 0–50 m layer was 19.0°C, with a range between 16.2°C and 22.2°C, while the salinity average was 38.36, with a range between 37.79 and 38.68. In the 50–90 m layer the temperature average was 15.8°C, with a range between 15.5°C and 16.1°C, while the salinity average was 38.71, with a range between 38.67 and 38.77. Remarks Oozooid. Brooksia rostrata and Brooksia berneri oozooids are very similar, differing only in the fact that in B. berneri oozooid muscle I, joined with the intermediate muscle (im), is separated from muscle II and discontinuous mid-dorsally. Due to this fact we will only compare the B. lacromae sp. nov. oozooid to the B. rostrata oozooid. The Brooksia lacromae sp. nov. oozooid has one ventral longitudinal muscle which extends into the proboscis, while B. rostrata has two ( Fig. 2 ). The proboscides in both collected oozooids broke off during collection, but were found in the sample ( Fig. 2c ). The proboscis of Brooksia lacromae sp. nov. seems to be thinner at the base than in Brooksia rostrata and in both species it contains two lateral longitudinal muscles. The ventral longitudinal muscle in B. lacromae sp. nov. has two small slit-like openings anteriorly, one below the endostyle and the other just anteriorly to it, partly overlapping with the anterior end of the endostyle. In Brooksia lacromae sp. nov. , in contrast to B. rostrata , body muscles are not arranged in a barrel-like structure where muscles are perpendicular to the body axis dorsally, as well as ventrally, but they converge to the posterior third of the body. Only muscles I, VI and VII fuse with the branches of the ventral longitudinal muscle, while muscles II, III, IV and V converge to the connection between muscle I and branches of the ventral longitudinal muscle, without fusing with either. Only muscle II can sometimes be slightly connected with muscle I ventrally. The stolon in Brooksia lacromae sp. nov. seems to emerge from the test, slightly posteriorly from the nucleus ( Fig. 4D ), while in B. rostrata it emerges from the anterior part of the nucleus ( Thompson 1948 ). Blastozooid (sinistral individual). The Brooksia lacromae sp. nov. and Brooksia rostrata blastozooids are very similar. In Brooksia lacromae sp. nov. , muscle IIR-b ends blindly on the ventral side of the animal without entering the posterior attachment processus (ap3), while in Brooksia rostrata it enters the posterior attachment processus (ap3). The left intermediate muscle (IM1) in Brooksia lacromae sp. nov. is continuous dorsally and ventrally. Its branch IM1-a enters the anterior attachment processus (ap1). In Brooksia rostrata the left intermediate muscle is discontinuous. Its dorsal and ventral counterparts both enter the anterior attachment processus. Because of this, the anterior attachment processus in B. lacromae sp. nov. possesses two muscles (IM1-a and IM2), while in B. rostrata it possesses three muscles (dorsal and ventral left intermediate muscle and right intermediate muscle). According to Thompson (1948) , the right intermediate muscle of the sinistral individual of B. rostrata ends blindly, while after Tokioka (1954) and Godeaux (1998) it connects to muscle I. In B. lacromae sp. nov. the right intermediate muscle (IM2) ends blindly. Genetic analysis There were no differences between blastozooid and oozooid 18S sequences. Between two cox1 sequences there were 7 substitutions out of 837 nucleotides, which results in 0.84% uncorrected pairwise distance. Cox1 sequences were translated using ascidian mitochondrial code and there were no differences in amino acid sequence between them. The uncorrected pairwise distance between the Brooksia rostrata 18S sequence ( HQ015403 ) and the Brooksia lacromae sp. nov. 18S sequence ( KR057223 ) was 1.5% (including gaps). Out of 26 differences in 1740 nucleotides between these two sequences, there were 21 transitions, 4 transversions and one single nucleotide gap.