Characterization of tetranucleotide microsatellite loci in a Lake Victorian,haplochromine cichlid fish: a Pundamilia pundamilia × Pundamilia nyererei hybrid

The haplochromine cichlid fishes inhabiting Lake Victoria in East Africa are of great interest to evolutionary biologists. We have isolated and optimized six tetranucleotide and a single dinucleotide locus from a hybrid of the haplochromine cichlid fishes, Pundamilia pundamilia and P. nyererei. Characterization in 18 individuals of P. nyererei from a single wild population revealed between six and 18 alleles per locus, with expected heterozygosity ranging from 0.68 to 0.94. These loci will prove useful for investigations of population structure, and elucidating relationships between closely related species. An additional 26 unoptimized loci have been deposited with GenBank and the MEN database.     

Fitness correlates of male coloration in a Lake Victoria cichlid fish

Sexual selection by female choice has contributed to the rapidevolution of phenotypic diversity in the cichlid fish speciesflocks of East Africa. Yet, very little is known about the ecologicalmechanisms that drive the evolution of female mating preferences.We studied fitness correlates of male nuptial coloration ina member of a diverse Lake Victoria cichlid lineage, Pundamilianyererei. In this species, male red coloration is subject tointraspecific sexual selection by female mate choice. Male nuptialcoloration plays a critical role also in reproductive isolationbetween this species and the closely related sympatric speciesP. pundamilia. Here, we show that P. nyererei male colorationis carotenoid based, illustrating the potential for honest signalingof individual quality. In a wild population, we found that variationin male coloration was not associated with variation in a setof strongly intercorrelated indicators of male dominance: malesize, territory size, and territory location. Instead, the 2male characters that predominantly determine female choice,territory size and red coloration, may be independent predictorsof male quality: males with bright red coloration and largeterritories had lower parasite infestation rates. As a result,female preferences tended to select against heavily parasitizedmales. Consistent with parasite-mediated sexual selection, maleshad higher and more variable parasite loads than females.     

Construction of a BAC library for Haplochromis chilotes,a cichlid fish from Lake Victoria

Cichlid fishes in Lake Victoria are model organisms for studying rapid radiation and speciation. On the way to examine the molecular basis of how these cichlid fishes achieved such a remarkable morphological diversification, we constructed a bacterial artificial chromosome (BAC) library derived from a cichlid species, Haplochromis chilotes, from Lake Victoria. The library includes 157,056 clones with the average insert size of 128 kb, corresponding to a 10-fold coverage of the H. chilotes genome. Given that the cichlid fishes endemic to Lake Victoria are closely related to one another phylogenetically and their genomes are nearly identical, this BAC library can be utilized to isolate genes from the more than 200 Haplochromine cichlid species in Lake Victoria.     

PERMANENT GENETIC RESOURCES: Isolation and characterization of microsatellite loci from the Arctic cisco (Coregonus autumnalis)

Eight polymorphic microsatellite loci were isolated and characterized for the Arctic cisco, Coregonus autumnalis. Loci were evaluated in 21 samples from the Colville River subsistence fishery. The number of alleles per locus ranged from two to 18. Observed heterozygosity of loci varied from 0.10 to 1.00, and expected heterozygosity ranged from 0.09 to 0.92. All eight microsatellite markers were in Hardy-Weinberg equilibrium. The loci presented here will be useful in describing population structure and exploring populations of origin for Arctic cisco.     

Male-male competition and speciation: aggression bias towards differently coloured rivals varies between stages of speciation in a Lake Victoria cichlid species complex

Sympatric speciation driven by sexual selection by female mate choice on a male trait is a much debated topic. The process is problematic because of the lack of negative frequency-dependent selection that can facilitate the invasion of a novel colour phenotype and stabilize trait polymorphism. It has recently been proposed that male-male competition for mating territories can generate frequency-dependent selection on male colouration. Rare male cichlid fish would enjoy a fitness advantage if territorial defenders bias aggression towards male cichlid fish of their own colour. We used blue (ancestral type) and red phenotypes of the Lake Victoria cichlid species complex Pundamilia. We tested the aggression bias of wild-caught territorial blue male cichlid fish from five separate populations for blue vs. red rival male cichlid fish using simulated intruder choice tests. The different populations vary in the frequency of red male cichlid fish, and in the degree of reproductive isolation between red and blue, reflecting different stages of speciation. Blue male cichlid fish from a population that lack red phenotypes biased aggression towards blue stimulus male cichlid fish. The same was found in two populations where blue and red are reproductively isolated sister species. This aggression bias may facilitate the invasion of a novel colour phenotype and species coexistence. Blue male cichlid fish from two populations where red and blue are hybridizing incipient species biased aggression towards red stimulus male cichlid fish. Thus, after a successful invasion of red, aggression bias alone is not likely to generate frequency dependence required to stabilize the coexistence of phenotypes. The findings show that aggression bias varies between stages of speciation, but is not enough to stabilize the process of speciation.     

PERMANENT GENETIC RESOURCES: Isolation and characterization of polymorphic microsatellite loci in common evening primrose (Oenothera biennis)

We developed nine polymorphic microsatellite loci for evening primrose (Oenothera biennis). These loci have two to 18 alleles per locus and observed heterozygosities ranging from 0 to 0.879 in a sample of 34 individuals. In a pattern consistent with the functionally asexual reproductive system of this species, 17/36 pairs of loci revealed significant linkage disequilibrium and three loci showed significant deviations from Hardy-Weinberg equilibrium. The loci will be informative in identifying genotypes in multigenerational field studies to assess changes in genotype frequencies.     

Shaping of the lower jaw bone during growth of Nile tilapia Oreochromis niloticus and a Lake Victoria cichlid Haplochromis chilotes: a geometric morphometric approach

East African cichlids have evolved feeding apparatus morphologies to adapt to diverse feeding environments. However, little is known about how the morphologies are formed during development. Here, we assessed the shape changes of the lower jaw bone during growth of the Nile tilapia Oreochromis niloticus and a Lake Victoria cichlid Haplochromis chilotes using geometric morphometric methods. 'Early Juvenile to Late Juvenile' and 'Late Juvenile to Adult' transitions of the shape change during growth of both O. niloticus and H. chilotes were detected. The 'Early Juvenile to Late Juvenile' transition of the shape change in H. chilotes occurred slightly earlier than in O. niloticus. We also compared the shape changes during growth of the two cichlids. Principal component analysis showed both commonalities and differences in the morphological changes between the cichlids. Our data suggest that most shape change may have a similar pattern during the growth of O. niloticus and H. chilotes, and that the differences in adult shapes may be due to differences arising early in development, not to the difference of shape change during growth. These data provide a basis for understanding the developmental mechanisms underlying this adaptive trait of East African cichlids.     

PERMANENT GENETIC RESOURCES: Isolation and characterization of microsatellite loci in the Mediterranean shore crab Carcinus aestuarii (Decapoda, Portunidae)

We characterized nine polymorphic microsatellites in the Mediterranean shore crab Carcinus aestuarii (Decapoda: Portunidae). Microsatellites were isolated from a partial genomic library enriched for multiple motifs. All loci were polymorphic, with number of alleles ranging from two to 16 and a mean observed heterozygosity of 0.75. Seven loci were in Hardy-Weinberg equilibrium, and two showed weak heterozygote deficiency. No linkage disequilibrium was found between loci. In addition, we tested Hardy-Weinberg and linkage equilibrium of three Carcinus maenas loci, already reported to cross-amplify in C. aestuarii. These molecular markers will be potentially useful to investigate genetic structure of this species.     

PERMANENT GENETIC RESOURCES: Isolation and characterization of polymorphic trinucleotide and dinucleotide microsatellite loci for the ice goby, Leucopsarison petersii

Eleven microsatellite markers were isolated and characterized for the ice goby, Leucopsarion petersii, from genomic libraries enriched for (ATG)(9) and (CA)(16) . Twenty individuals from a single population were used to screen polymorphism in these loci. The number of alleles per locus and observed heterozygosity ranged from four to 22 and from 0.35 to 1.00, respectively. All loci did not significantly deviated from Hardy-Weinberg equilibrium, and there was no evidence of linkage disequilibrium between all loci-pairs. These loci showed Mendelian inheritance in a full-sib family. The high level of polymorphism of these loci will be useful for studies of population genetics.     

PERMANENT GENETIC RESOURCES: Isolation of microsatellite loci from the endangered plant Sibara filifolia (Brassicaceae)

Sibara filifolia (Brassicaceae) is a federally endangered annual herb found on two of the California Channel Islands. Previous studies based on allozymes revealed little genetic variability on San Clemente Island. Nine polymorphic microsatellite loci were isolated from individuals on San Clemente Island. We found low levels of allelic variation (mean N_A = 2.3), with seven loci exhibiting significant deviations from Hardy–Weinberg equilibrium (P < 0.01) and 10 pairs of loci exhibiting significant linkage disequilibrium (P < 0.01). Most of the observed variability (mean H_O = 0.003) occurred among populations or in rare homozygous individuals.     

PERMANENT GENETIC RESOURCES: Development of polymorphic microsatellite loci for the endangered Topeka shiner, Notropis topeka

We have developed a set of eight polymorphic microsatellite markers for the endangered Topeka shiner, Notropis topeka. Allelic diversity at each of these loci was assessed in a single isolated population from eastern South Dakota, USA. The allelic diversity ranged from four to 15 alleles. These are the first microsatellite markers to be reported for this species. These markers are being used in a more thorough study of the population structure throughout the remaining range of this species.     

Population-structure and genetic diversity in a haplochromine cichlid fish [corrected] of a satellite lake of Lake Victoria

The approximately 500 species of the cichlid fish species flock of Lake Victoria, East Africa, have evolved in a record-setting 100,000 years and represent one of the largest adaptive radiations. We examined the population structure of the endangered cichlid species Xystichromis phytophagus from Lake Kanyaboli, a satellite lake to Lake Victoria in the Kenyan Yala wetlands. Two sets of molecular markers were analysed--sequences of the mitochondrial control region as well as six microsatellite loci--and revealed surprisingly high levels of genetic variability in this species. Mitochondrial DNA sequences failed to detect population structuring among the three sample populations. A model-based population assignment test based on microsatellite data revealed that the three populations most probably aggregate into a larger panmictic population. However, values of population pairwise FST indicated moderate levels of genetic differentiation for one population. Eleven distinct mitochondrial haplotypes were found among 205 specimens of X. phytophagus, a relatively high number compared to the total number of 54 haplotypes that were recovered from hundreds of specimens of the entire cichlid species flock of Lake Victoria. Most of the X. phytophagus mitochondrial DNA haplotypes were absent from the main Lake Victoria, corroborating the putative importance of satellite lakes as refugia for haplochromine cichlids that went extinct from the main lake in the last decades and possibly during the Late Pleistocene desiccation of Lake Victoria.     

PERMANENT GENETIC RESOURCES: Isolation and characterization of microsatellite DNA loci from the southern flounder, Paralichthys lethostigma

Paucity of polymorphic molecular markers in southern flounder (Paralichthys lethostigma) has been a major limitation in genetic improvement of this important economic fish. Hence, we constructed a repeat-enriched genomic library from P. lethostigma. A total of 39 new microsatellites were identified, for which 33 primer pairs were designed. After validating and scoring, 10 of these loci were polymorphic in a test population with the range of alleles from two to nine per locus. The observed and expected heterozygosities ranged from 0.2500 to 0.9000 and from 0.4469 to 0.8514, respectively. These polymorphic microsatellites will be useful for genetic diversity analysis and linkage map construction for P. lethostigma.     

PERMANENT GENETIC RESOURCES: Isolation and characterization of microsatellite loci in the North Island brown kiwi, Apteryx mantelli

We report the isolation and characterization of eight polymorphic and five monomorphic microsatellites in North Island brown kiwi (NIBK, Apteryx mantelli), using two polymerase chain reaction (PCR) techniques employing either short-tandem repeat primers (STR method) or random PCR-based isolation of microsatellite arrays (PIMA method). Microsatellite polymorphism was subsequently determined using 65 individuals. There were two to seven alleles for each polymorphic locus with heterozygozity ranging between 0.04 and 0.86. These primers will be used in future studies to determine the level of extra-pair copulation, dispersal patterns, and genetic diversity within and between wild populations of NIBK.     

PERMANENT GENETIC RESOURCES: Development and characterization of 14 polymorphic microsatellite loci in the endangered tree Euptelea pleiospermum (Eupteleaceae)

Fourteen polymorphic microsatellite loci were developed and characterized for the endangered and tertiary relict tree, Euptelea pleiospermum. A genomic DNA enrichment protocol was used to isolate microsatellite loci and polymorphism was explored using 32 individuals from one natural population. The observed number of alleles ranged from two to nine. The ranges of observed and expected heterozygosities were 0.25-1.00 and 0.22-0.85, respectively. These microsatellite markers provide powerful tools for the ongoing conservation genetic studies of E. pleiospermum.     

PERMANENT GENETIC RESOURCES: Isolation, characterization and cross-species amplifications of microsatellite loci from Conradina (Lamiaceae)

We report the isolation of microsatellite loci from three species in the genus Conradina (Lamiaceae). To ensure their utility for multiple species, loci were screened for amplification and variability in all six Conradina species; 11 loci demonstrated high levels of amplification and polymorphism in most species. These 11 loci were characterized in 20 individuals from one population of Conradina brevifolia; alleles per locus ranged from five to 15, and observed heterozygosity ranged from 0.30 to 0.90. These microsatellites will be used to clarify species limits, detect interspecific hybridization, and understand the partitioning of genetic variation in each species of Conradina.     

PERMANENT GENETIC RESOURCES: Isolation and characterization of 12 microsatellite markers in the middle-spotted woodpecker (Dendrocopos medius)

Twelve polymorphic microsatellite loci were developed for Dendrocopos medius. Polymorphism was assessed for 27 individuals from the southwesternmost population of this woodpecker species. The number of alleles per locus ranged from three to seven, with observed heterozygosity values from 0.444 to 0.852. Genotypic frequencies conformed to Hardy-Weinberg equilibrium, and no evidence for linkage disequilibrium was observed. Multilocus genotypes resulting from this set of markers will be useful to determine genetic diversity and differentiation within and among habitat patches inhabited by D. medius. Three of the loci were polymorphic for Picoides articus.     

A test of fitness consequences of hybridization in sibling species of Lake Victoria cichlid fish

Several hundred species of haplochromine cichlid fish have evolved rapidly in Lake Victoria. Divergent sexual and ecological selection probably played an important role in this radiation, generating divergent mating preferences and preference-trait covariance. However, the segregation of hybrid inviability or infertility genes could also potentially generate preference-trait covariance, and the mechanisms that cause the evolution of divergent mating preferences have not been investigated in detail in any cichlid species pair. We investigated intrinsic fitness of hybrids between two sister species in the genus Pundamilia, one of the most species-rich genera of Lake Victoria cichlids. Fitness-related traits were measured in nonhybrid offspring of both species, and in the first and second hybrid generations. There were no differences in fecundity, fertility, sex ratio or growth rates either between the sister species or between these and their hybrids. By contrast, there was a difference in offspring survival between the two sister species. Offspring survival was dependent only on the species of the mother, regardless of whether the cross was conspecific or heterospecific. Further, eggs tended to be larger in the hybrids than in one of the parental species. Hence, hybrids suffered no intrinsic fitness reduction relative to nonhybrids. Our data suggest that intrinsic hybrid incompatibilities are unlikely to have caused speciation in Pundamilia, nor to maintain species boundaries in this system.