Two new endemic species in the Midas cichlid species complex from ...

AQUA19(4)-LAYOUT:AQUA 30/10/13 15:22 Pagina 207

aqua, International Journal of Ichthyology

Two new endemic species in the Midas cichlid species complex from Nicaraguan crater lakes: Amphilophus tolteca and Amphilophus viridis (Perciformes, Cichlidae) Hans Recknagel1, 3, Henrik Kusche1, 2, Kathryn R. Elmer1, 3 and Axel Meyer1, 2*

1) Lehrstuhl für Zoologie und Evolutionsbiologie, Department of Biology, University of Konstanz, Universitätsstrasse 10, 78457 Konstanz, Germany. 2) International Max Planck Research School for Organismal Biology, University of Konstanz, Germany. 3) Institute of Biodiversity, Animal Health and Comparative Medicine, College of Medical, Veterinary and Life Sciences, University of Glasgow, Glasgow, UK. *Corresponding author: E-mail [email protected] Received: 04 June 2013 – Accepted: 10 October 2013

Abstract The Neotropical Midas cichlid species complex (Amphilophus citrinellus Günther 1864) has become a model system for investigating the mechanisms of speciation and adaptive radiation. In several instances ancestral Midas cichlids from the great Nicaraguan lakes have colonized nearby crater lakes where they continued to evolve in isolation. Each crater lake can be seen as a “natural experiment” of sympatric and allopatric divergence. Several ecologically and genetically well-differentiated crater lake species have already been identified, but the species complex is not fully taxonomically resolved. Here, two new endemic Nicaraguan crater lake cichlids species are described: Amphilophus tolteca n. sp., a slender-bodied species which is endemic to the ca. 1,245 year old Lake Asososca Managua and Amphilophus viridis n. sp., an endemic benthic species from Lake Xiloá. Amphilophus tolteca morphologically resembles previously described limnetic species from the crater lakes Apoyo and Xiloá with a depressed, elongated body. However, A. tolteca is geographically isolated and genetically distinct from those species and from the putative generalist ancestral species. Amphilophus viridis resembles the Xiloá species A. amarillo in terms of body shape, but is distinct in coloration and ecology, and is genetically differentiated from all other syntopic species.

Resumen El complejo de especies de la mojarra es un sistema modelo para investigar los mecanismos de especiación y la formación de radiaciones adaptativas. Diversas mojarras, que son distintas en su ecología, morfología y genética; han sido descritas tanto en escenarios alopátricos como simpátricos en las cadenas de lagunas cratéricas nicaragüenses, las cuales forman un “experimento natural”. En este estudio, describimos dos nuevas especies de cíclidos endémicos en las lagunas cratéricas de Nicaragua: Amphilophus tolteca 207

n. sp., una especie de cuerpo estrecho endémica en la laguna de Asososca Managua y Amphilophus viridis n. sp., una especie endémica que ocupa el espacio bentónico de la laguna de Jiloá. Con su cuerpo alargado, la morfología de A. tolteca es semejante a especies anteriormente descritas en las lagunas cratéricas de Apoyo y Jiloá. Sin embargo, A. tolteca es genéticamente distinta a estas especies limnéticas y las supuestas especies ancestrales. En su morfología, A. viridis se parece a la especie sintópica A. amarillo en Jiloá, pero es distinta en su coloración, ecología y genética.

Zusammenfassung Die neotropische Gruppe der Midas-Cichliden (Amphilophus citrinellus Günther 1864) gilt inzwischen als Modellsystem, um die Mechanismen der Artbildung und adaptiven Radiation zu studieren. Mehrfach haben die Vorfahren der Midas-Cichliden von den großen Seen Nicaraguas aus nahe gelegene Kraterseen besiedelt, wo sie sich isoliert weiterentwickelt haben. Jeder der Kraterseen lässt sich damit als „Naturexperiment“ sympatrischer und allopatrischer Divergenz auffassen. Einige ökologisch und genetisch gut unterscheidbare Kratersee-Arten konnten bereits bestimmt werden, aber die Artengruppe ist damit noch nicht vollständig taxonomisch durchgearbeitet. Hier werden zwei neue endemische nicaraguanische KraterseeCichlidenarten beschrieben: Amphilophus tolteca n. sp., eine eher limnische Art, die endemisch im ca. 1245 Jahre alten Lake Asososca Managua lebt, sowie Amphilophus viridis n. sp., eine benthische Art vom Lake Xiloá. Die Exemplare von Amphilophus tolteca ähneln zwar vorher beschriebenen limnischen Arten von den Kraterseen Apoyo und Xiloá mit ihren länglichen Körpern sind aber von diesen und von der mutmaßlichen Vorläufer-Art geografisch isoliert und auch genetisch unterschiedlich. Amphilophus viridis ähnelt der Xiloá-Art A. amarillo nach dem Maßstab der Körperform, unterscheidet sich aber aqua vol. 19 no. 4 - 25 October 2013


Two new endemic species in the Midas cichlid species complex from Nicaraguan crater lakes: Amphilophus tolteca and Amphilophus viridis

nach Farbgebung und Ökologie und lässt sich genetisch von allen anderen syntopischen Arten unterscheiden.

Résumé Le complexe néotropical du Cichlide Amphilophus citrinellus Günther 1864 est devenu un système modèle dans la recherche des mécanismes de la spéciation et de la radiation adaptative. A plusieurs égards, les ancêtres de ces cichlides des grands lacs du Nicaragua ont colonisé des lacs de cratère proches où ils ont continué à évoluer en isolés. Chaque lac de cratère peut être vu comme un « laboratoire naturel » de divergence sympatrique et allopratique. Plusieurs espèces de lacs de cratère bien distincts écologiquement et génétiquement ont déjà été identifiées, mais le complexe d’espèces n’a pas encore été exhaustivement décrit. Ici sont décrites deux nouvelles espèces du Nicaragua de lacs de cratère: Amphilophus tolteca n. sp., une espèce au corps allongé, endémique du lac Asososca Managua, vieux de 1.245 ans et Amphilophus viridis n. sp., une espèce endémique benthique du lac Xiloa. Amphilophus tolteca ressemble morphologiquement à des espèces limniques déjà décrites des lacs de cratère Apoyo et Xiloá au corps déprimé et allongé. Toutefois, A. tolteca est géographiquement isolé et génétiquement distinct de ces espèces et des espèces ancestrales supposées. Amphilophus viridis ressemble à l’espèce du Xiloá, A. amarillo, en ce qui concerne la forme du corps, mais s’en distingue par la colorationet l’écologie, et diffère génétiquement de toutes les autres espèces synoptiques.

Sommario Il complesso di specie di ciclidi neotropicali midas (Amphilophus citrinellus Günther 1864) è diventato un sistema modello per lo studio dei meccanismi di speciazione e di radiazione adattativa. In diversi casi i midas ancestrali dei grandi laghi del Nicaragua hanno colonizzato vicini laghi vulcanici dove continuarono a evolversi in isolamento. Ogni lago del cratere può essere visto come un "esperimento naturale" della divergenza simpatrica e allopatrica. In questi laghi sono già state individuate diverse specie ben differenziate ecologicamente e geneticamente, ma il complesso di specie non è stato tassonomicamente risolto. Qui sono descritte due nuove specie endemiche nicaraguensi di ciclidi di laghi craterici: Amphilophus tolteca n. sp., una specie dal corpo snello, che è endemica del lago Asososca Managua vecchio di circa 1245 anni e Amphilophus viridis n. sp., una specie endemica bentonica dal Lago Xiloá. Amphilophus tolteca assomiglia morfologicamente alle specie limnetiche con un corpo allungato e depresso, precedentemente descritte dai laghi cratere Apoyo e Xiloá. Tuttavia, A. tolteca è geograficamente isolata e geneticamente distinta da queste specie e dalle generalisti putative specie ancestrali. Amphilophus viridis assomiglia alla specie dello Xiloá, A. amarillo, in termini di forma del corpo, ma si distingue per la colorazione e l'ecologia ed è geneticamente differenziata da tutte le altre specie sintopiche.

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INTRODUCTION Midas cichlid fishes form a young species complex in the freshwaters of Central America, with their highest abundance and species richness in Nicaragua (Barluenga & Meyer, 2010). They occur in a geographical setting of two great lakes (Lake Nicaragua and Lake Managua, which are occasionally connected) and a chain of nearby isolated, young crater lakes (Fig. 1). This spatial arrangement of a “natural experiment” of a large source population and small, unconnected peripheral populations makes Midas cichlids ideal study objects for speciation research (Barluenga et al., 2006; Elmer et al., 2010a; Wilson et al., 2000). Midas cichlids within each crater lake studied to date have been found to be genetically more similar to each other than to their relatives in any other lake (Barluenga & Meyer, 2010; McKaye et al., 2002). Each lake population can hence be investigated as its own, closed and independent system and sympatric speciation is likely to contribute to the formation of new Midas cichlid species (Barluenga et al., 2006). Parallel evolution has probably occurred (Elmer et al., 2010a; Elmer & Meyer, 2011) and some crater lake populations constitute early evolutionary stages of diversification in sympatry, opening the possibility to test speciation hypotheses (Elmer et al., 2010a). Parallel evolution and parallel speciation processes are considered as strong evidence for the role of natural selection in speciation (Rundle et al., 2000; Schluter & Nagel, 1995). A valid taxonomy is a prerequisite for a thorough investigation of how new species arise. Taxonomic status in the Midas cichlid species complex Midas cichlids have long been known for their astonishing degree of phenotypic variability and rank among the most diverse fishes in Central America (Elmer et al., 2010a; Meyer, 1990a, 1990b). The fleshy lipped A. labiatus Günther 1864 and the high-bodied generalist A. citrinellus Günther 1864, which occur most abundantly in the great lakes, were described first. The taxonomy has long been in flux and a number of synonyms for these species existed in the past (reviewed in Barlow & Munsey 1976). Even long ago, the astonishing variability of A. citrinellus (sensu lato) was a point of interest for researchers. In 1907, the ichthyologist S. E. Meek noted, upon sampling A. citrinellus from great and crater lakes: Of all the species in these [Nicaraguan] lakes, this one is by far the most variable. I made many repeated 208


Hans Recknagel, Henrik Kusche, Kathryn R. Elmer and Axel Meyer

efforts to divide this material listed below in from two to a half-dozen or more species, but in all cases I was unable to find any tangible constant characters to define them. To regard them as more than one species meant only to limit the number by the material at hand, and so I have lumped them all in one. In an attempt to distill the taxonomy, in the 1970s, a series of studies addressed the biology of Nicaraguan cichlids, including Midas cichlids (Thorson, 1976). This research described the first crater lake endemic: in Lake Apoyo the open-water species with an elongated body shape named A.

zaliosus (Barlow & Munsey, 1976). Since the 1990s, five additional endemic species from Apoyo (A. astorquii, A. chancho, A. flaveolus, A. globosus and A. supercilius) (Geiger et al., 2010b; Stauffer Jr et al., 2008) and three new endemic species in crater lake Xiloá (A.amarillo, A. sagittae and A. xiloaensis) (Stauffer & McKaye, 2002) have been described. This brings the Midas cichlid complex to eleven described species to date. However, it has been suggested by a number of researchers (Elmer et al., 2010a; Geiger et al., 2010a; Waid et al., 1999) that more Midas cichlid species await scien-

Fig. 1. Map of Nicaraguan bodies of water containing Midas cichlids that were examined in this study. L. = Lake; As. = Asososca. 209




tific description in allopatry and in sympatry with currently described species. In particular, isolated crater lakes (e.g. Asososca León, Tiscapa, Monte Galán, Masaya), more well-studied crater lakes (e.g. Xiloá and Apoyo), the great lakes, and the associated rivers are habitats that may be home to new species upon further study (Barluenga & Meyer, 2010; Elmer et al., 2010a; Geiger et al., 2010a). The integration of ecological, morphological and genetic data (integrative taxonomy) (Padial et al., 2010) is necessary to describe the actual biological richness of the Midas cichlid complex and seek to identify the biological relevance of the long recognized abundant diversity. We see Midas cichlid species as genetically distinct and therefore largely reproductively isolated units with certain phenotypic features that allow distinguishing them from other such units. The Midas cichlid communities in two isolated crater lakes – Asososca Managua and Xiloá – are the focus of the current study. Notes on crater lake Asososca Managua Asososca Managua is maximally only ca. 1,245 years old (Pardo et al., 2008) and therefore ranks among the youngest crater lakes in Nicaragua (Elmer et al., 2010a). This crater lake is very small (0.73 km2; 40 000 m3) compared to the other Nicaraguan crater lakes that house multiple described Midas cichlid species. The slope of the littoral zone in Asososca Managua is extremely steep in comparison to other Nicaraguan crater lakes (Recknagel et al., in review), therefore this crater lake probably exhibits among the lowest proportion of littoral zone (Waid et al., 1999). The presence of a narrow littoral zone may limit Midas cichlid diversification since this zone provides the main breeding and feeding grounds (Elmer et al., 2009; Lehtonen et al., 2011; Recknagel et al., in review). Asososca Managua is a natural reservoir and well-protected because it serves as a drinking water supply for the city of Managua. Access to the lake is strictly controlled by the Nicaraguan government. Thus, its fauna is relatively pristine and aquatic and terrestrial vertebrates that are rare in other crater lakes can be observed easily, such as crocodiles (Crocodilus acutus) and the green iguana (Iguana iguana). It has been previously reported that Midas cichlids from Asososca Managua have distinct body shapes and elongated individuals were reported occasionally (Elmer et al., 2010a). To date, Midas cichlids that have an elongated body shape, relative to A. citrinellus, were believed to be restricted to lakes Apoyo (A. zaliosus) aqua vol. 19 no. 4 - 25 October 2013

Table I. Holotypes’ and paratypes’ catalogue numbers deposited at the Natural History Museum of London. Species

Type

Sex

BMNH No.

A. tolteca A. tolteca A. tolteca A. tolteca A. viridis A. viridis A. viridis

Holotype Paratype Paratype Paratype Holotype Paratype Paratype

male male male female female male female

BMNH 2012.9.2.1 BMNH 2012.9.2.2 BMNH 2012.9.2.3 BMNH 2012.9.2.4 BMNH 2012.9.2.5 BMNH 2012.9.2.6 BMNH 2012.9.2.7

and Xiloá (A. sagittae). The current study examines the phenotypic and genetic diversity of the Asososca Managua Midas cichlids and proposes a new species. Notes on crater lake Xiloá Lake Xiloá is estimated to be ca. 6,100 years old (Kutterolf et al., 2007), making it the second oldest of the Nicaraguan crater lakes (Elmer et al., 2010a). Although lakes Apoyo and Masaya are larger than Xiloá, the highest number of fish species of all crater lakes is found in Lake Xiloá (reviewed in Elmer et al., 2010a), probably due to its close proximity and possible occasional connection to Lake Managua (Villa, 1976). The lake has three main habitats: i) sandy, muddy and gently sloping bottom, ii) hot sulfurous springs with weed beds, and iii) a steep, rocky shore with large jumbled boulders (McKaye et al., 2002). Amphilophus amarillo Stauffer & McKaye 2002 has mostly been observed close to the shore in the weed habitat and A. xiloaensis Stauffer & McKaye 2002 mostly on rocky habitat, whereas A. sagittae Stauffer & McKaye 2002 seems to have a preference for both sandy and a mixed habitat structure of weed and rocks and breeds deeper than A. amarillo, but not as deep as A. xiloaensis (Elmer et al., 2009; McKaye et al., 2002). In recent sampling trips, variants with greenish ground coloration were occasionally collected and attributed to A. amarillo. Further sampling and analyses of morphological, meristic, genetic and ecological differentiation from the syntopic Midas cichlid species warranted a species description. MATERIALS AND METHODS Sampling Fishes from Nicaraguan crater lakes Asososca Managua (N 12°08.273’W 86°18.792’) and Xiloá (N 12°12.888’W 86°18.862’) were caught with gill nets, hook and line, or by harpooning in November-December 2010 (NXiloá = 60, NAsososca 210



Table II. Accession numbers of 79 mtDNA control region sequences that were downloaded from Genbank. ID

Species

Lake

AMCK00278 AMCK00327 AMCK00328 AMCK00331 AMCK00345 AMCK00346 cAsM5553 cAsM5562 cAsM5565 cAsM5573 cAsM5574 cAsM5586 cAsM5587 cAsM5591 AMCK00534 AMCK01257 cXil5456 cXil5457 cXil5458 cXil5459 cXil5461 cXil5475 cXil5550 cXil5552 AMCK00911 AMCK01251 AMCK01253 AMCK01256 cXil5449 cXil5468 cXil5471 cXil5477 cXil5481 cXil5484 cXil5500 cXil5038 cXil5055 cXil5269 cXil5271 cXil5283

A. tolteca A. tolteca A. tolteca A. tolteca A. tolteca A. tolteca A. tolteca A. tolteca A. tolteca A. tolteca A. tolteca A. tolteca A. tolteca A. tolteca A. amarillo A. amarillo A. amarillo A. amarillo A. amarillo A. amarillo A. amarillo A. amarillo A. amarillo A. amarillo A. viridis A. viridis A. viridis A. viridis A. viridis A. viridis A. viridis A. viridis A. viridis A. viridis A. viridis A. sagittae A. sagittae A. sagittae A. sagittae A. sagittae

As. Managua As. Managua As. Managua As. Managua As. Managua As. Managua As. Managua As. Managua As. Managua As. Managua As. Managua As. Managua As. Managua As. Managua Xiloá Xiloá Xiloá Xiloá Xiloá Xiloá Xiloá Xiloá Xiloá Xiloá Xiloá Xiloá Xiloá Xiloá Xiloá Xiloá Xiloá Xiloá Xiloá Xiloá Xiloá Xiloá Xiloá Xiloá Xiloá Xiloá

Accession number GU017018 GU017036 GU017037 GU017040 GU017047 GU017048 HM183710 HM183689 HM184591 HM184585 HM183711 HM183703 HM183704 HM184588 GU017062 GU017077 HM184649 HM184650 HM184651 HM184652 HM184653 HM184015 HM184007 HM183946 GU017071 GU017073 GU017074 GU017076 HM184892 HM183893 HM183895 HM183834 HM184775 HM184062 HM183897 HM184046 HM184065 HM184498 HM184700 HM184049

Managua = 37). Other specimens were collected from lakes Managua, Nicaragua, and Apoyo in 2010 for comparison (total material analyzed for all lakes: N = 144). Standardized photographs, tissue samples (fin, muscle), and stomach contents were taken in the field. All samples were preserved in ethanol or 10% formalin and vouchered in the

211

ID

Species

Lake

cXil5543 AMCK00891 AMCK00892 AMCK00893 AMCK00894 AMCK00898 AMCK00902 cManMir3002 cManMir3009 cManMir3019 cManMir3022 cManSAn3934 cManSAn3939 cManSAn3947 cManSAn3959 cNicOme5759 cNicOme5761 cNicOme5764 cNicOme5765 cNicOme5767 cNicOme5773 cNicOme5776 cNicOme5785 cNicOme5786 lManMir5002 lManMom5606 lManMom5823 lManMom5828 lManMom5829 lNicIsl5609 lNicIsl5610 lNicIsl5612 lNicIsl5614 lNicIsl5615 lNicIsl5617 lNicIsl5621 lNicIsl5623 lNicIsl5626 lNicIsl5635

A. sagittae A. xiloaensis A. xiloaensis A. xiloaensis A. xiloaensis A. xiloaensis A. xiloaensis A. citrinellus A. citrinellus A. citrinellus A. citrinellus A. citrinellus A. citrinellus A. citrinellus A. citrinellus A. citrinellus A. citrinellus A. citrinellus A. citrinellus A. citrinellus A. citrinellus A. citrinellus A. citrinellus A. citrinellus A. labiatus A. labiatus A. labiatus A. labiatus A. labiatus A. labiatus A. labiatus A. labiatus A. labiatus A. labiatus A. labiatus A. labiatus A. labiatus A. labiatus A. labiatus

Xiloá Xiloá Xiloá Xiloá Xiloá Xiloá Xiloá Managua Managua Managua Managua Managua Managua Managua Managua Nicaragua Nicaragua Nicaragua Nicaragua Nicaragua Nicaragua Nicaragua Nicaragua Nicaragua Managua Managua Managua Managua Managua Nicaragua Nicaragua Nicaragua Nicaragua Nicaragua Nicaragua Nicaragua Nicaragua Nicaragua Nicaragua

Accession number HM184072 GU017080 GU017081 GU017082 GU017083 GU017086 GU017079 HM184119 HM183729 HM183949 HM183730 HM184145 HM183741 HM183741 HM184160 HM183758 HM184866 HM183759 HM184867 HM183760 HM184260 HM184264 HM184272 HM184275 HM184783 HM184103 HM184777 HM184779 HM184780 HM183863 HM183749 HM183864 HM184209 HM183750 HM184213 HM184210 HM184220 HM184211 HM183754

Axel Meyer Collection Konstanz (AMCK). Holotypes and some of the paratypes (N = 7) are deposited at the Natural History Museum in London (Table I), where Günther’s first specimens of this species complex are also located. All other paratypes of the type series (N = 43) are deposited at the University of Konstanz. aqua vol. 19 no. 4 - 25 October 2013



Analysis of morphological characters A set of 24 morphometric characters was measured to the nearest 0.1 mm with digital calipers. Also nine meristic counts were taken. Snout acuteness was determined with a goniometer following Meyer 1990a. Measurements were performed on the left side of adult voucher specimens. Comparisons were based on absolute numbers (meristic traits, snout acuteness) or ratios of morphometric characters to standard length (SL) or head length (HL). All variables were z-transformed and the geometric mean was calculated for all individuals following Mosimann (1970). A regression of all morphometric variables on the geometric mean was performed to correct for significant allometric effects (p < 0.001) and regression residuals were used in downstream analyses (Klingenberg, 1996). Multivariate analysis of variance (MANOVA), principal component analysis (PCA), and linear discriminant analysis (LDA) were calculated. Meristic measurements were compared in a separate analysis. The newly described species from Lake Asososca Managua was compared to the two putative ancestral species that seeded Asososca Managua (Barluenga & Meyer 2010), A. citrinellus and A. labiatus from lakes Managua and Nicaragua, and the two other described elongated Midas cichlid species, A. zaliosus from Lake Apoyo and A. sagittae from Lake Xiloá. The new species from Lake Xiloá was contrasted to the three other sympatric Midas cichlid species from Xiloá: A. amarillo, A. sagittae, A. xiloaensis. Genetic analyses Mitochondrial control region DNA sequences from 79 individuals relevant to our study were downloaded from Genbank (Table II). Photographs of individuals with published mtDNA control region DNA sequences were individually checked and species status was identified based on coloration, body shape, pharyngeal jaw shape (following Barlow & Munsey, 1976; Geiger et al., 2010b; Stauffer et al., 2008; Stauffer & McKaye, 2002) and by cross-checking field note books. A Maximum likelihood tree was computed using RAxML performing a bootstrap search with 1,000 pseudo-replicates and the best tree was used for constructing a haplotype network of taxonomic grouping relevant to this study. DNA was amplified for a total 13 microsatellite loci for all Asososca Managua individuals used in the morphometric analyses using previously published conditions (M1M, M2, M7, M12 (Noack et al., aqua vol. 19 no. 4 - 25 October 2013

2000), UNH002 (Kellogg et al., 1995), UNH011, UNH012, UNH013 (McKaye et al., 2002), Abur45, Abur82, Abur151 (Sanetra et al., 2009), Burtkit F 474/R672 (Salzburger et al., 2007), TmoM7 (Zardoya et al., 1996)). Genotypic data for individuals from lakes Nicaragua, Managua and Xiloá were already available (Elmer et al., 2010b; Elmer et al., in prep.). Analyses were run using STRUCTURE vers. 2.3.3 (Pritchard et al., 2000) with a burn-in period of 100,000 and 500,000 Markov Chain Monte Carlo (MCMC) repetitions under an admixture model and correlated allele frequencies (Falush et al., 2003). Analyses using all species (k = 1-9) and a second intralacustrine analysis restricted to Lake Xiloá individuals (k = 1-8) were performed and analyzed using methodologies published in Evanno et al. 2005. F-statistics were computed using ARLEQUIN 3.1 performing 10,000 permutations (Excoffier et al., 2005). Comparative material Amphilophus amarillo (Xiloá): AMCK 13037, AMCK 13039, AMCK 13041-44, AMCK 13046, AMCK 13048, AMCK 13052, AMCK 13141-42, AMCK 13268, AMCK 13636-37; A. citrinellus: AMCK 13456, AMCK 13780-82, AMCK 13790, AMCK 13793-94, AMCK 13796, AMCK 13798802, AMCK 13804; A. labiatus: AMCK 12598, AMCK 12600, AMCK 12602, AMCK 12605-09, AMCK 13768-69, AMCK 13771, AMCK 13773, AMCK 13785-86, AMCK 13995, AMCK 13998, AMCK 14000, AMCK 14001, AMCK 14004; A. sagittae (Xiloá): AMCK 13114, AMCK 13122, AMCK 13124, AMCK 13126-27, AMCK 13129, AMCK 13166-72, AMCK 13174-78, AMCK 13280, AMCK 13450; A. xiloaensis (Xiloá): AMCK 13033-35, AMCK 13272, AMCK 1327576, AMCK 13281, AMCK 13284, AMCK 13440, AMCK 13446, AMCK 13653; A. zaliosus (Apoyo): AMCK 12460, AMCK 12523-25, AMCK 12528-30, AMCK 12533-34, AMCK 12536-37, AMCK 12539, AMCK 12573. RESULTS

Amphilophus tolteca n. sp. (Figs 2-3, Table III) Holotype: BMNH 2012.9.2.1, adult male, 131.1 mm standard length (SL), 14. Dec. 2010, Managua, Lake Asososca Managua, N 12°08.390’W 086°18.792’. 212



Paratypes: All paratypes with same collection data as holotype. BMNH 2012.9.2.2-4, adult male, 143.6 mm SL, adult male, 129.6 mm SL, adult female, 126.3 mm SL, AMCK 13965, 115.0 mm SL, AMCK 13966, 124.0 mm SL, AMCK 13968, 149.6 mm SL, AMCK 13969, 135.9 mm SL, AMCK 13970, 103.6 mm SL, AMCK 13972,

130.6 mm SL, AMCK 14041, 107.2 mm SL, AMCK 14061, 130.8 mm SL, AMCK 14067, 116.9 mm SL,

AMCK 13978, 107.3 mm SL, AMCK 14059, 113.5 mm SL, AMCK 14066, 123.2 mm SL, AMCK 14070,

105.3 mm SL, AMCK 14058, 100.0 mm SL, AMCK 14063, 128.8 mm SL, AMCK 14069, 129.3 mm SL,

Fig. 2. Lateral view of A. tolteca, holotype, BMNH 2012.9.2.1, Lake Asososca Managua, Nicaragua. Photo by H. Kusche.

Fig. 3. Amphilophus tolteca, non-type, live coloration shortly after capture. Photo by G. Machado-Schiaffino. 213




Table III. Morphometric (in mm), snout acuteness and meristic values of type material. All type series specimens for species A. tolteca and A. viridis are included. Abbreviations correspond to the following distance-based measurements: ADAA = anterior insertion dorsal fin to anterior insertion anal fin; ADPA = anterior insertion dorsal fin to posterior insertion anal fin; PDAA = posterior insertion dorsal fin to anterior insertion anal fin; PDPA = posterior insertion dorsal fin to posterior insertion anal fin; PDP = posterior insertion of dorsal fin to insertion of pelvic fin; PAA: insertion of pelvic fin to anterior insertion of anal fin; PPA = pelvic fin to posterior insertion of anal fin; APP = anterior insertion of pectoral fin to pelvic fin. Species holotype standard length head length

A. tolteca (N=37) x range

sd

holotype

A. viridis (N=12) x range

sd

131.1 46.2

118.6 42.4

100.0-149.6 36.0-52.5

13 4.6

150 49.5

164.1 55.1

126.1-191.7 46.3-62.9

21.5 6.4

%SL head length body depth ADAA ADPA PDAA PDPA PDP PAA PPA APP caudal peduncle length caudal peduncle depth pectoral fin length caudal fin length front length throat length dorsal fin base length

35.3 37.9 47.7 59.5 31.5 13.7 55.0 28.6 49.5 14.2 13.2 13.0 26.0 28.2 40.4 39.7 54.8

35.7 39.9 49.3 61.3 33.7 14.5 53.3 24.3 47.8 15.1 13.9 13.5 26.6 27.8 41.2 41.0 56.3

34.6-37.1 35.4-43.7 46.1-53.0 57.9-64.5 31.3-35.9 13.0-16.3 49.6-55.9 18.2-28.6 43.4-50.8 12.9-17.3 12.3-15.2 12.8-14.4 23.1-28.7 24.0-31.5 39.3-43.1 38.9-43.0 51.3-59.4

0.7 1.8 2.0 1.6 1.1 0.7 1.6 2.2 1.7 1.2 0.7 0.5 1.2 1.9 3.8 1.0 2.5

33.0 44.4 55.0 67.9 36.2 16.7 56.2 24.2 49.6 16.1 14.0 15.3 25.6 24.1 41.1 39.4 60.7

3.7 44.0 54.1 64.8 35.4 15.5 55.8 24.6 49.3 16.7 14.1 14.3 26.2 25.2 40.5 40.2 58.4

32.1-37.0 41.3-47.0 51.8-55.9 62.7-67.9 34.0-37.5 14.4-16.7 52.7-59.8 21.5-27.0 45.7-52.2 15.7-18.1 13.1-15.6 13.2-15.5 24.5-29.1 23.1-28.7 37.8-44.3 37.8-42.8 56.1-61.9

1.2 1.7 1.3 1.8 0.9 0.7 1.8 1.5 1.6 0.8 0.7 0.7 1.1 1.5 1.8 1.3 1.8

%HL snout length cheek depth interorbital width eye length eye depth lower jaw length

35.2 29.8 34.4 28.4 25.4 41.4

36.9 29.3 35.6 26.5 25.1 41.8

33.0-43.9 26.7-32.1 32.1-40.3 23.6-29.7 21.2-28.2 39.2-44.5

2.3 1.2 1.9 1.6 1.6 1.2

41.0 37.8 44.3 25.1 24.1 40.5

40.0 34.3 39.7 23.2 22.5 38.8

33.3-43.7 31.5-37.8 33.6-44.3 20.7-25.4 20.6-25.0 35.6-40.7

3.0 1.7 3.4 1.7 1.6 1.8

Angle snout angle

66.9

66.4

57.9-75.0

4.6

83.2

76.0

72.1-83.2

3.2

holotype

mode

range

%

holotype

mode

range

%

33 22 11 12 17 9 6 14 5

34 21 12 12 17 9 7 14 5

29-35 20-25 6-14 11-13 16-17 8-10 6-8 13-15 5-5

33 50 50 58 83 67 83 83 100

34 21 13 11 17 8 7 14 5

33 21 11 12 17 9 7 15 5

30-37 20-24 10-13 11-12 16-17 8-9 6-7 14-15 5-5

32 41 35 73 78 70 78 68 100

meristics lateral line scales upper lateral line scales lower lateral line scales dorsal fin rays dorsal fin spines anal fin rays anal fin spines pectoral fin rays pelvic fin rays

AMCK 14071, 106.6 mm SL, AMCK 14074, 137.5 mm SL, AMCK 14077, 121.4 mm SL,

112.2 mm SL, AMCK 14073, 102.4 mm SL, AMCK 14076, 118.5 mm SL, AMCK 14079,


AMCK 14072, 104.7 mm SL, AMCK 14075, 132.1 mm SL, AMCK 14078, 111.7 mm SL,

AMCK 14082, 117.0 mm SL, AMCK 14085, 110.5 mm SL, AMCK 14104, 111.7 mm SL.

119.8 mm SL, AMCK 14084, 113.1 mm SL, AMCK 14094, 104.6 mm SL,

AMCK 14083, 107.3 mm SL, AMCK 14089, 107.8 mm SL, AMCK 14108,

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Table IV. F-statistics for all species included in this study. Fst pairwise distances are listed for mtDNA haplotypes below diagonal and for microsatellite loci above diagonal. Probability values: NS P > 0.05, * P < 0.05, ** P < 0.01, *** P < 0.001. A. xiloaensis A. xiloaensis A. sagittae A. viridis A. amarillo A. tolteca A. labiatus A. citrinellus

0.308* 0.125* 0.176** 0.463*** 0.078NS 0.141**

A. sagittae

A. viridis

A. amarillo

A. tolteca

A. labiatus

A. citrinellus

0.020***

0.103*** 0.097***

0.061*** 0.058*** 0.062***

0.224*** 0.274*** 0.236*** 0.227***

0.089*** 0.132*** 0.104*** 0.093*** 0.115***

0.121*** 0.159*** 0.108*** 0.123*** 0.139*** 0.019***

0.163* 0.216* 0.503*** 0.175** 0.22***

0.069* 0.395*** 0.054NS 0.085**

0.421*** 0.048NS 0.06*

0.290** 0.334**

0.041*

Fig. 4. Linear discriminant analysis of the two putative ancestral species, limnetic species and A. tolteca. Amphilophus citrinellus and A. labiatus represent the two ancestral species, the two elongated limnetic species from lake Apoyo and lake Xiloá are A. zaliosus and A. sagittae, respectively. Each symbol represents a single individual in morphometric space, except the ones close to each Midas cichlid that indicate correspondence between symbol and species. The first two axes explain 90.8% of the total variance. 215




Fig. 5. A haplotype network showing the relationship between the putatively ancestral Midas cichlids from the great Nicaraguan lakes (A. citrinellus and A. labiatus) and those of derived crater lakes Xiloá (A. amarillo, A. viridis, A. xiloaensis and A. sagittae) and lake Asososca Managua (A. tolteca). The haplotype network is based on 767 bp of mitochondrial DNA control region, estimated by ML analysis. Each line constitutes a single mutational step between connected haplotypes. Open circles illustrate mutational steps that were not recovered in this study. Size of each circle reflects the number of individuals that share a particular haplotype (see legend). Genbank numbers are listed in Table II.

Fig. 6. View of crater lake Asososca Managua, type locality of A. tolteca. Lake Asososca Managua is characterized by steep slopes and a small littoral zone. Photo by H. Kusche. aqua vol. 19 no. 4 - 25 October 2013

216



Diagnosis and comparisons: Amphilophus tolteca is clearly distinguished from its putative ancestral species A. citrinellus by a reduced body depth (mean = 39.9% SL, range = 35.4-43.7% SL vs. A. citrinellus, mean = 45.9% SL, range = 43.8-48.3% SL; see Fig. 4). Amphilophus labiatus is less elongated and is clearly distinctive by the presence of hypertrophied lips. Description: Morphometric and meristic measurements of the holotype and paratypes are given in Table III. Slender-bodied Midas cichlid with laterally and dorso-ventrally compressed body. Head moderately wide (interorbital width: 32.1-40.3% SL) and notably large (head length: 34.6-37.1% SL) with pointed mouth (snout angle: 57.9-75.0°) and slightly prolated lower jaw (lower jaw length: 39.244.5% SL). Jaws isognathous. Eyes large (eye length: 23.6-29.7% SL; eye depth: 21.2-28.3% SL). Caudal peduncle slightly longer than deep (mean caudal peduncle length: 13.9% SL, mean caudal peduncle depth: 13.5% SL). Flank scales ctenoid. Lateral line scales in holotype and on average 33, ranging from 29-35. Scales on upper lateral line 20-25 and lower lateral line 6-14. Scales between upper lateral line and dorsal fin anteriorly 6-7 and posteriorly 3-5; 2 scale rows between lateral lines. Dorsal and anal fins with squamation between fin rays extending to about ¼ of fin; caudal fin scales extending to about ⅓ of the fin. First to 4th dorsal fin spines increasing in length, with first spine of variable length, but rarely more than ⅓ of 4th spine. 5th to 17th spines of similar length. First three anal fin spines increasing in length, following 5 spines of subequal length. Pelvic fin inserted slightly posterior to vertical through pectoral-fin insertion, first ray longest, reaching up to insertion of anal fin rays. C o l o r a t i o n i n l i f e : Male and female coloration similar. Head and ground coloration from light greenish/grayish to black. 5-7 dark vertical bars along lateral body axis. Dark spot above the lateral line in transition from caudal peduncle to caudal fin. Throat and belly pale and in some cases pinkish. Gold color morphs of this species exist (e.g. AMCK 14041, AMCK 14058, AMCK 14059) at a low frequency (ca.