Description of Andinoacara stalsbergi sp . n . ( Teleostei : Cichlidae : Cichlasomatini ) from Pacifi c coastal rivers in Peru , and annotations on the phylogeny of the genus

The South American cichlid genus Andinoacara MUSILOVÁ, ŘIČAN & NOVÁK, 2009 belonging to the tribe Cichlasomatini is one of the genera which were recently described as the result of analyses of morphological and molecular data and ensuing nomenclatural revisions. This genus involves six species previously placed in the genus Aequidens EIGENMANN & BRAY, 1894, viz. Cichlosoma biseriatum REGAN, 1913, Acara coeruleopunctata KNER, 1863, Acara latifrons STEINDACHNER, 1878, Cychlasoma pulchrum GILL, 1858, Chromis rivulata GÜNTHER, 1860 and Acara sapayensis REGAN, 1903. KULLANDER (1983) demonstrated that Aequidens was an unnatural catch-all group. He listed the six species mentioned above among the Aequidens species which ought to have been placed into a separate Description of Andinoacara stalsbergi sp. n. (Teleostei: Cichlidae: Cichlasomatini) from Pacifi c coastal rivers in Peru, and annotations on the phylogeny of the genus

genus and introduced the term 'Aequidens' pulcher group for them because of lack of an alternative generic allocation.In addition, he later (KULLANDER 1991) used the term 'Aequidens' rivulatus group for 'Aequidens' rivulatus and undescribed forms (cf.STA-WIKOWSKI & WERNER 1998) closely related to this species, and eventually showed that the 'Aequidens' pulcher-rivulatus group could have a generic status (KULLANDER 1998).MUSILOVÁ et al. (2008) confi rmed that the 'Aequidens' pulcher-rivulatus group represents a well supported yet unnamed genus.In their study of the phylogenetic relationships among cichlasomatine cichlids MUSILOVÁ et al. (2009) tentatively listed 'Aequidens' sp."Silbersaum" as an undescribed species among the members of the 'Aequidens' pulcher-rivulatus group, for which they established the new genus Andinoacara.
The species provisionally referred to as 'Aequidens' sp."Silbersaum" in Europe or as "Green Terror" in the USA has been known both in the aquarium trade and the popular literature for about forty years (LÜLING 1972;STAECK & LINKE 1985).In the older ichthyological and aquaristic literature it was treated as a form of A. rivulatus (e. g.REGAN 1905, EVERMAN & RADCLIFFE 1917, EIGENMANN 1922, LÜLING 1972, WERNER 1983).The formal description of this cichlid is one subject of this paper.In addition the phylogeny within the genus Andinoacara is discussed on the basis of a set of molecular data.This is the fi rst molecular phylogenetic analysis including all valid nominal species of Andinoacara.

Material and Methods
Type specimens were fi xed in formalin and later transferred into 75% ethanol.The holotype and paratypes are deposited in the fi sh collection of the Senckenberg Naturhistorische Sammlungen Dresden, Museum für Tierkunde, (MTD F).
The techniques for taking measurements and meristic data follow those described in KULLANDER (1986) and KULLANDER & NIJSSEN (1989).Measurements were made with an electronic digital caliper reading to the nearest 0.1 mm.Figures in brackets after counts indicate the number of specimens examined with that condition.Terminology and methods of measurements of jaws and teeth follow CASCIOTTA & ARRATIA (1993).Scale rows are numbered as described by KULLANDER (1990).Nomenclature of colour patterns follows KUL-LANDER (1983,1991).Vertical bars are numbered from the caudal fi n to the snout as described by KULLANDER & SILFVERGRIP (1991).According to this approach the caudal spot is counted as bar 1 (homolog to bar 1p in ŘÍČAN et al. 2005).The description follows the general format used by KULLANDER (1991).
In the phylogenetic analysis 11 species or forms of Andinoacara were studied.We analyzed specimens of seven nominal species, viz.Andinoacara biseriatus, A. coeruleopunctatus, A. latifrons, A. pulcher, A. rivulatus, A. sapayensis, A. stalsbergi and four apparently undescribed forms.All specimens used in the molecular analyses were obtained from direct imports to specialized aquarium trade companies in Europe.We studied 2-4 specimens of each species or form, except for A. coeruleopunctatus and A. sapayensis, as only one individual of these species was available.
In the molecular study eight genetic markers were sequenced, and a data set with the total length of 5627 bp was obtained.Both mitochondrial (16S rRNA, 12S rRNA, cytochrome b, ND 4) and nuclear coding (RAG1, rhodopsin, Tmo-4C4) and non-coding (intron 1 in S7 gene) markers were used.We analyzed a data set of eight genes with the exception of A. sapayensis because sequences of only two genes of this species were available.The set of primers is listed in Tab. 1.
PCR condition consisted of an initial denaturation step at 94 °C followed by the extension of DNA at Andinoacara stalsbergi sp.n.
On fi rst gill arch 2 or 3 small gill rakers on epibranchial, 1 in the angle and 9-10 (n=5) externally on ceratobranchial.Fourth ceratobranchial with 5 tooth plates and 3 to 7 teeth on each plate.Lower pharyngeal tooth plate (Fig. 5) robust, moderately long (width of bone 83-85% of its length; n=2), with well-ordered teeth; length of dentigerous area 85% of its width; 14-17 teeth in posterior row (n=2), 6-9 teeth in median row (n=2); teeth obviously lost were also counted.Oral jaw teeth conical with recurved tips.In upper jaw hemiseries 15-22 outer row teeth and in lower jaw hemiseries 17-23.Length of dentigerous arm of premaxilla shorter than length of ascending arm (premaxillary ascending arm length/dentigerous arm length ≈ 1.6); width of the ascending arm about 15% of its length.Lower jaw comparatively high (anguloarticular depth about 72% of length, coulter area depth about 43% of anguloarticular length).Coulter area deeper than its length (coulter area depth/coulter area width ≈ 1.3).Dorsal margin of hyoid more or less straight, without a deep notch.

Colouration in life.
Based on observations on specimens kept in aquarium and photos taken immediate-ly after capture.Forehead, nape and pre-dorsal part of dorsum uniformly greyish or light brown.On the body sides each scale with iridescent or metallic green centre and contrasting dark brown marginal line.The dark scale margins form a fi ne reticulate pattern which is particularly prominent in adult specimens.Cheeks with two to four narrow oblique opalescent green lines and several small buccal dots of the same colour.Dark cheek spot in the corner of the preopercle usually vis-  ible only during brood care.Lips, lower region of preopercle and gill cover iridescent green.Iris golden.
No horizontal lateral band.Midlateral spot black, squarish or rectangular, extending dorso-ventrally from 3 / 4 of El scales 8-10, 9-11 or 9-12 to all of these scales in E2 and E3 row above.Anterior and posterior to midlateral spot with contrasting narrow vertical white stripe which fades in dorsal and ventral region.Sometimes with three ill-defi ned wide dark vertical bars and narrow light interspaces behind the midlateral spot.Usually no caudal spot (if visible: small, vertically extended, positioned on level of lower lateral line).
Dorsal fi n grey, with narrow dark submarginal band and white lappets forming a conspicuous white margin; soft part with iridescent streaks on the membranes.Anal fi n grey with blackish margin and small iridescent green dots and short lines.Caudal fi n grey, with darker distal region, a conspicuous white posterior margin and a pattern of tiny greenish dots.Pelvic fi ns grey, darker along anterior margin, with green fi rst interradial membrane and greenish dots or short streaks inwardly.Pectoral fi ns hyaline and colourless.After spawning and during parental care both sexes develop a very dark, almost blackish colouration with two contrasting white vertical stripes anterior and posterior to midlateral spot.

Colouration in alcohol.
Based on holotype with notes on paratype specimens.Body sides grey, darker on nape and back.Scales on sides with narrow black margin.Snout and cheeks grey.Lips dark grey.Gill cover, preopercle and branchiostegal region dark grey.Two or three dark preorbital stripes and several irregular short streaks or dots.Supraorbital markings indistinct and faint.Suborbital stripe reduced to a short dark marking (often masked by dark head sides) in the corner of the preopercle.
No continious lateral band.Bar 1 (caudal spot) narrow, prominent and blackish, in the centre of the caudal-fi n base, not reaching ventral or dorsal edges of it; bar 2 on caudal peduncle; bar 3 on anterior part of caudal peduncle between posterior rays of dorsal fi n and anal fi n, in smaller specimens (< 70 mm SL) often split into two parallel vertical bars; bar 4 and 5 fused in adult specimens, but separate in smaller ones; bar 6 darker than the other bars, straight, not vertically split, on both sides with contrasting narrow light margin; bar 7 and 8 indistinct, usually fused.Midlateral spot black, on upper half of bar 6. Dorsal fi n dark grey with light marginal band and light grey streaks in its soft part.Anal fi n grey with darker marginal band and light grey streaks in posterior portion.Caudal fi n grey, with light grey streaks in posterior portion and contrasting light distal margin.Pelvic fi ns dark grey.Pectoral fi ns hyaline.
Sexual dimorphism.There are no obvious external sex differences in fi n length or intensity of colour pattern.However, observations under aquarium conditions revealed that there is a distinct size difference between males and females and that in addition dominant males develop a prominent nuchal hump.

Geographical distribution.
Andinoacara stalsbergi is distributed in trans-andean rivers and lakes at the Peruvian Pacifi c coast.The distribution of this species in the Pacifi c slope of western Peru between Río Chira (Depto.Piura) in the north and the Río Pisco (Depto.Ica) in the south is well documented (cf.STAWIKOWSKI  tween 1994 and 2008 in different years at many localities, collected additional ecological data at several collecting sites (from the south to north): (1) Depto.Ica: Río Pisco near Independencia (13º43'25" S, 75º58'59" W).At the collecting site the rather clear river was approx.10 m wide.Its bottom was covered with sand and rocks.There was no submerged vegetation.Water data: water temperature 25.5 ºC, pH 8.3, total hardness > 40 °dH, temporary hardness 7 dH.
The available ecological data reveal that Andinoacara stalsbergi is well adapted to very alkaline and hard water rich in dissolved minerals and even tolerates brackish water (LÜLING 1973).
Reproductive behaviour.Observations under aquarium conditions showed that Andinoacara stalsbergi is a monogamous substratum spawner and that both sexes share in all the duties of brood care.The female, however, is usually the more active partner as long as the pair cares for eggs or larvae, while the male defends the spawning territory against intruders.Like most other open brooders these cichlids deposit their eggs on a horizontal surface.At 27 °C hatching occurs about two days postspawning, and the fry attempt swimming seven days thereafter.The male and female fi sh practice long-term biparental defense of their mobile fry.A detailed description of the reproductive behaviour was published by WERNER (1983).
Etymology.Named in honour of ALF STALSBERG (Tjodalyng, Norway), the collector of the holotype, in recognition of his longstanding commitment to increase the knowledge about cichlid fi shes.

Discussion
Andinoacara stalsbergi is most closely related to A. rivulatus (Fig. 6), its sister species.Both differ from other species of Andinoacara by the lack of conspicuous dark nape markings (versus distinct dark supraorbital marks in species of both the A. pulcher group and of the related genera Bujurquina and Tahuantinsuyoa; cf.KULLANDER 1986KULLANDER , 1991)), larger size (adult males > 150 mm TL versus < 150 mm TL in the previously mentioned taxa; cf.EIGENMANN 1922) and a trend towards more gill rakers (usually 9 or 10 rakers on ceratobrachial of outer gill arch versus usually < 9 in the remaining species of Andinoacara; cf.REGAN 1905, 1913, EIGENMANN 1922).
Although A. stalsbergi is very similar to A. rivulatus in morphometric and meristic data, general appearance and colouration of its head, these two allopatric species can readily be distinguished for distinctive specifi c colour patterns: Andinoacara stalsbergi has light scale centres and dark scale edges forming a reticulate pattern on the fl anks (versus light scale edges and dark scale centres forming a pattern of horizontal lines in A. rivulatus) and a prominent white margin of the dorsal and caudal fi n (versus a broad orange margin in A. rivulatus).The genetic differences between A. stalsbergi and its congeners (Tab.3) provide additional arguments for its taxonomical separateness according to the evolutionary species concept (WILEY 1978).
Andinoacara biseriatus listed as a member of the pulcher group by KULLANDER (1998) and included in the rivulatus group by STAWIKOWSKI & WERNER (1998) clustered as the basal sister taxon of the rivulatusstals bergi clade (Fig. 7).This species differs from A. stalsbergi by the possession of dark rimmed scales on its nape (versus scales without dark posterior rim), a dark dot in the centre of each scale on body and opercle (versus dark edges, but no dark dots in A. stalsbergi), a caudal fi n with a narrow reddish margin (versus caudal fi n with broad whitish margin), a lateral spot positioned more dorsally than in A.stalsbergi, an additional spot (more prominent in females) in the dorsal fi n above the bar with lateral spot (versus no such spot in A. stalsbergi) and, according to REGAN (1913), frequently 2 rows of scales on its cheek (versus 3 rows of scales in A. stalsbergi).
In the result of the phylogenetic analysis Andinoacara sapayensis clustered as a sister taxon of the rivulatus-stalsbergi clade.Adult specimens of Andinoacara stalsbergi differ signifi cantly from A. sapayensis by the possession of only 3 bars between midlateral spot and caudal spot (versus four bars; compare  . stalsbergi versus <8 in A. sapayensis;cf. REGAN 1905, EIGENMANN 1922) and the lack of distinct dark supraorbital marks (versus possession of prominent supraorbital marks in A. sapayensis).
The molecular studies confi rm the status of Andinoacara stalsbergi and reveal its phylogenetic relationships to its congeners (Fig. 7).The reconstruction of the phylogeny within the genus Andinoacara results in the existence of two clades.One consists of three valid species (i.e. A. latifrons, the type species Vertebrate Zoology ■ 59 (2) 2009 distribution.However, in the results of our analyses we found only minor genetic difference between this form and A. latifrons living in eastern Colombia.Morphological comparisons of both are still missing because of the lack of material.
The specimens of A. pulcher used in our analyses were collected in Trinidad.They are genetically identical with A. pulcher obtained from Czech and German aquarium trade.Eventually A. sp."Venezuela" appears to represent another form of "blue acara" with uncertain taxonomic position, for conclusive evidence is missing as only few specimens were available.
of Andinoacara, A. coeruleopunctatus and A. pulcher) and several possibly undescribed forms: Andinoacara sp."Rio Chirgua" and A. sp."Orinoco" representing separate lineages of fi shes living in Venezuela and differing from A. pulcher and A. latifrons in their colouration.This group of "blue acaras" was referred to as "Aequidens" sp."Orinoco" in STAWIKOWSKI & WERNER (1998).But despite considerable differences, further morphological studies are required to confi rm their status as new species.Andinoacara sp."Maracaibo" (STAWIKOWSKI & WERNER 1998) is endemic to the drainages of Lago de Maracaibo in western Venezuela and represents another undescribed form with limited Tab. 3. Uncorrected p-distance between Andinoacara stalsbergi sp.n. and the two closely related species A. rivulatus (sister species) and A. biseriatus (1st & 2nd column).Average distance between nominal species of Andinoacara, including A. stalsbergi sp.n. in 3rd column.

A. stalsbergi vs.
A

Fig. 7 .
Fig. 7. Phylogenetic relationships within the genus Andinoacara based on a data set of eight genes (16S rRNA, 12S rRNA, cytochrome b, ND 4, S7 intron 1, RAG1, Rhodopsin, Tmo-4C4).Analysis was performed by using MrBayes software with two parallel runs of 20 million generations and sampling every 1000 trees.The model parameters suggested by jModeltest were applied.The phylogenetic position of Andinoacara stalsbergi sp.n. is marked by the indicator.
Tab. 1. Primers used for molecular analyses in this study.
Diagnosis.A species of the Andinoacara rivulatus group.It is most similar to A. rivulatus with which it shares the comparatively large size (TL > 200 mm in males), the colour pattern of the cheeks and a light vertical stripe anterior and posterior to the rectangular midlateral spot.It is readily distinguished from this species by specifi c colour characteristics, viz.having (1) a conspicuous white margin in both the dorsal and caudal fi n and (2) on the body sides scales with light centres and contrasting dark marginal lines forming a fi ne reticulate pattern.Description.Refer toFigs.1-4for general appearance and colour pattern.Morphometric data of eight specimens (89.6-113.0mmSL) are summarised in Tab. 2. Counts from 12 specimens (65.8-113.0mmSL), osteological characters from a dissected specimen (73.3 mm SL).Body moderately deep (body depth 42-49% of SL) and laterally compressed.