Distribution, colour polymorphism and habitat use of the African killifish Nothobranchius furzeri, the vertebrate with the shortest life span

Intensive collection in southern Mozambique across and outside the potential range of Nothobranchius furzeri , the species with the shortest recorded life span among vertebrates used as a model in ageing research, revealed that, contrary to previous data, it is a widespread species. It occurs in small freshwater pools south of the Save River and north of the Incomati River, including basins of the Limpopo, Changane, Chefu, Mazimechopes and Vaneteze Rivers. During collection in February 2008 (the second part of the rainy season), populations were strongly female biased (mean, 28% of males across 19 populations), and there was a spatial pattern in female bias among metapopulations. Populations varied in the proportion of male colour morphs. Fourteen populations were composed exclusively of the red male phenotype, three populations of the yellow male phenotype and 12 populations were mixed. Overall, the red phenotype was more common, but there was strong geographical variation in morph proportion, with yellow males more abundant at the periphery and red male dominance in the centre of the range of N. furzeri in the Limpopo basin. Nothobranchius furzeri was sympatric with Nothobranchius orthonotus (35% of investigated pools) and Nothobranchius rachovii (27% of sites). Analysis of habitat use of N. furzeri is presented; N. furzeri was associated with pools containing a soft muddy substratum and turbid water.     

Nothobranchius furzeri

Nothobranchius furzeri The African killifish Nothobranchius furzeri lives in seasonal ponds. Through diapauses it can survive the often very extended dry season. After hatching N. furzeri grows extremely fast and becomes sexually mature after 4–5 weeks. Certain strains of N. furzeri show an extremely short lifespan of only 3–4 months, which is the shortest lifespan of a vertebrate that can be kept in captivity. Despite its short lifespan N. furzeri shows typical signs of aging on a morphological, behavioral and molecular level. Old animals display shortening of telomeres and a loss of the number and activity of mitochondria. First crossing experiments between the short‐ and the long‐living strains have revealed the existence of at least four genomic loci, which contribute to the differences in lifespan.     

Mapping of quantitative trait loci controlling lifespan in the short-lived fish Nothobranchius furzeri--a new vertebrate model for age research

The African annual fish Nothobranchius furzeri emerged as a new model for age research over recent years. Nothobranchius furzeri show an exceptionally short lifespan, age-dependent cognitive/behavioral decline, expression of age-related biomarkers, and susceptibility to lifespan manipulation. In addition, laboratory strains differ largely in lifespan. Here, we set out to study the genetics of lifespan determination. We crossed a short- to a long-lived strain, recorded lifespan, and established polymorphic markers. On the basis of genotypes of 411 marker loci in 404 F(2) progeny, we built a genetic map comprising 355 markers at an average spacing of 5.5 cM, 22 linkage groups (LGs) and 1965 cM. By combining marker data with lifespan values, we identified one genome-wide highly significant quantitative trait locus (QTL) on LG 9 (P < 0.01), which explained 11.3% of the F(2) lifespan variance, and three suggestive QTLs on LG 11, 14, and 17. We characterized the highly significant QTL by synteny analysis, because a genome sequence of N. furzeri was not available. We located the syntenic region on medaka chromosome 5, identified candidate genes, and performed fine mapping, resulting in a c. 40% reduction of the initial 95% confidence interval. We show both that lifespan determination in N. furzeri is polygenic, and that candidate gene detection is easily feasible by cross-species analysis. Our work provides first results on the way to identify loci controlling lifespan in N. furzeri and illustrates the potential of this vertebrate species as a genetic model for age research.     

Extremely short lifespan in the annual fish Nothobranchius furzeri

Evolutionary theories of senescence postulate that lifespan is determined by the age-dependent decrease in the effects of natural selection. Factors that influence survival and reproduction at early life stages have a larger impact on fitness than factors that influence later life stages. According to these views, selection for rapid sexual maturation and a steep age-dependent decrease in fitness drive the evolution of short lifespans. Here, we report on the survival trajectory of Nothobranchius furzeri (Pisces: Ciprinodontidae): a member of a group of annual species found in temporary bodies of water whose life expectancy in the wild is limited to a few months. We find that maximum survival of N. furzeri in the laboratory is less than 12 weeks. The temporal trajectory of survival shows an age-dependent increase in the mortality rate that is typical of organisms with defined lifespans. The lifespan of N. furzeri is exceptionally short for a vertebrate: owing to its small size and the possibility of propagation in captivity, N. furzeri could be used as a convenient model for ageing research.     

Annual fishes of the genus Nothobranchius as a model system for aging research

Aging research in vertebrates is hampered by the lack of short-lived models. Annual fishes of the genus Nothobranchius live in East African seasonal ponds. Their life expectancy in the wild is limited by the duration of the wet season and their lifespan in captivity is also short. Nothobranchius are popular aquarium fishes and many different species are kept as captive strains, providing rich material for comparative studies. The present paper aims at reviving the interest in these fishes by reporting that: (1) Nothobranchius can be cultured, and their eggs stored dry at room temperature for months or years, offering inexpensive methods of embryo storage; (2) Nothobranchius show accelerated growth and expression of aging biomarkers at the level of histology and behaviour; (3) the species Nothobranchius furzeri has a maximum lifespan of only 3 months and offers the possibility to perform investigations thus far unthinkable in a vertebrate, such as drug screening with life-long pharmacological treatments and experimental evolution; (4) when the lifespan of different species is compared, a general correlation is found between wet season duration in their natural habitat and longevity in captivity; and (5) vertebrate aging-related genes, such as p66Shc and MTP, can be easily isolated in Nothobranchius by homology cloning. These fishes can become excellent models for aging studies. They can be employed to test the effects of experimental manipulation on aging at a pace comparable with that of Drosophila and to probe the effects of natural selection on the evolution of aging-related genes.     

Phylogeography of colour polymorphism in the coral reef fish Pseudochromis fuscus, from Papua New Guinea and the Great Barrier Reef

Body colour has played a significant role in the evolution of coral reef fishes, but the phylogenetic level at which colour variation is expressed and the evolutionary processes driving the development and persistence of different colour patterns are often poorly understood. The aim of this study was to examine the genetic relationships between multiple colour morphs of Pseudochromis fuscus (family Pseudochromidae), both within and among geographic locations. Pseudochromis fuscus is currently described as a single species, but exhibits at least six discrete colour morphs throughout its range. In this study, P. fuscus from Papua New Guinea (PNG) and the Great Barrier Reef (GBR), Australia, formed three genetically distinct clades based on mitochondrial DNA (control region) sequence data: (1) yellow and brown morphs from the GBR and southern PNG, as well as an orange morph from southern PNG; (2) a pink morph from southern PNG; and (3) all three morphs (pink, orange and grey) found in Kimbe Bay, northern PNG. The three groups showed deep levels of divergence (d=14.6–25.4%), suggesting that P. fuscus is a complex of polychromatic species, rather than a single widespread species with many different colour morphs. Population genetic analyses indicate that the three clades have experienced unique evolutionary histories, possibly from differential effects of sea level fluctuations, barriers to gene flow and historical biogeography.     

Unexpected patterns of genetic structuring among locations but not colour morphs in Acropora nasuta (Cnidaria; Scleractinia)

Symbiotic relationships have contributed greatly to the evolution and maintenance of biological diversity. On the Great Barrier Reef, species of obligate coral-dwelling fishes (genus Gobiodon) coexist by selectively recruiting to colonies of Acropora nasuta that differ in branch-tip colour. In this study, we investigate genetic variability among sympatric populations of two colour morphs of A. nasuta ('blue-tip' and 'brown-tip') living in symbiosis with two fish species, Gobiodon histrio and G. quinquestrigatus, respectively, to determine whether gobies are selecting between intraspecific colour polymorphisms or cryptic coral species. We also examine genetic differentiation among coral populations containing both these colour morphs that are separated by metres between local sites, tens of kilometres across the continental shelf and hundreds of kilometres along the Great Barrier Reef. We use three nuclear DNA loci, two of which we present here for the first time for Acropora. No significant genetic differentiation was detected between sympatric colour morphs at these three loci. Hence, symbiotic gobies are selecting among colour morphs of A. nasuta, rather than cryptic species. Significant genetic geographical structuring was observed among populations, independent of colour, at regional (i.e. latitudinal separation by < 500 km) and cross-shelf (< 50 km) scales, alongside relative homogeneity between local populations on within reef scales (< 5 km). This contrasts with the reported absence of large-scale genetic structuring in A. valida, which is a member of the same species group as A. nasuta. Apparent differences in biogeographical structuring between species within the A. nasuta group emphasize the need for comparative sampling across both spatial (i.e. within reefs, between reefs and between regions) and taxonomic scales (i.e. within and between closely related species).     

A comprehensive phylogeny of the bumble bees (Bombus)

Bumble bees ( Bombus Latreille) occupy a wide diversity of habitats, from alpine meadows to lowland tropical forest, yet they appear to be similar in morphology throughout their range, suggesting that behavioural adaptations play a more important role in colonizing diverse habitats. Notwithstanding their structural homogeneity, bumble bees exhibit striking inter- and intraspecific variation in colour pattern, purportedly the outcome of mimetic evolution. A robust phylogeny of Bombus would provide the framework for elucidating the history of their wide biogeographical distribution and the evolution of behavioural and morphological adaptations, including colour pattern. However, morphological studies of bumble bees have discovered too few phylogenetically informative characters to reconstruct a robust phylogeny. Using DNA sequence data, we report the first nearly complete species phylogeny of bumble bees, including most of the 250 known species from the 38 currently recognized subgenera. Bayesian analysis of nuclear (opsin, EF-1α, arginine kinase, PEPCK) and mitochondrial (16S) sequences results in a highly resolved and strongly supported phylogeny from base to tips, with clear-cut support for monophyly of most of the conventional morphology-based subgenera. Most subgenera fall into two distinct clades ( short-faced and long-faced ) associated broadly with differences in head morphology. Within the short-faced clade is a diverse New World clade, which includes nearly one-quarter of the currently recognized subgenera, many of which are restricted to higher elevations of Central and South America. The comprehensive phylogeny provides a firm foundation for reclassification and for evaluating character evolution in the bumble bees.  © 2007 The Linnean Society of London, Biological Journal of the Linnean Society , 2007, 91 , 161–188.     

New World biogeography and the evolution of polychromatism: evidence from the bee assassin genus Apiomerus (Heteroptera: Reduviidae: Harpactorinae)

The crassipes and pictipes species groups of Apiomerus Hahn together contain 12 species, many with high intraspecific chromatic variability, which represent the majority of Nearctic species in this New World assassin bug genus. Because of their geographical distribution and their varying degrees of polychromatism, these two species groups provide a unique opportunity to study the evolution of polychromatism and analyse relationships among areas of endemism in the Nearctic, as well as determine the boundary between the Nearctic and Neotropical regions. The results of a morphology based phylogenetic analysis allowed investigation of these questions while also determining relationships among the 12 species in the two species groups. The crassipes and pictipes species groups were each supported as monophyletic and as sister taxa. Apiomerus rufipennis (Fallou) was not included in the original concept of the crassipes species group, but is shown here to be a member of the group. Apiomerus barrocoloradoi Forero, Berniker & Szerlip, which had been hypothesized previously to belong to the pictipes species group, is excluded from this group. Intraspecific polychromatism for each species was identified as being present in one of three states: no polychromatism; limited polychromatism; or polychromatism as discrete colour morphs. Limited polychromatism was here found to be the ancestral state for Apiomerus, and species with discrete colour morphs are restricted to the crassipes and pictipes species groups. Polychromatism appears to be a greatly homoplastic character within the genus. A Brooks parsimony analysis recovered distinct Nearctic and Neotropical clades. The Nearctic clade is divided between areas in the central and eastern U.S.A. and areas in the Western U.S.A. and Mexico. The Nearctic–Neotropical boundary for the taxa included in the analysis is along the Isthmus of Tehuantepec in southern Mexico.     

Female fecundity traits in wild populations of African annual fish: the role of the aridity gradient

The evolution of life history is shaped by life expectancy. Life‐history traits coevolve, and optimal states for particular traits are constrained by trade‐offs with other life‐history traits. Life histories contrast among species, but may also diverge intraspecifically, at the level of populations. We studied the evolution of female reproductive allocation strategy, using natural populations of two sympatric species of African annual fishes, Nothobranchius furzeri and Nothobranchius orthonotus. These species inhabit pools in the Mozambican savanna that are formed in the rainy season and persist for only 2–10 months. Using 207 female N. furzeri from 11 populations and 243 female N. orthonotus from 14 populations, we tested the effects of genetic background (intraspecific lineage) and life expectancy (position on the aridity gradient determining maximum duration of their temporary habitat) on female fecundity traits. First, we found that variation in female body mass was small within populations, but varied considerably among populations. Second, we found that fecundity was largely defined by female body mass and that females spawned most of their eggs in the morning. Third, we found that the trade‐off between egg size and egg number varied among lineages of N. furzeri and this outcome has been confirmed by data from two separate years. Overall, we demonstrate that local conditions were important determinants for Nothobranchius growth and fecundity and that eggs size in arid region was less limited by female fecundity than in humid region.     

Reproductive isolation of sympatric morphs in a population of Darwin's finches

Recent research on speciation has identified a central role for ecological divergence, which can initiate speciation when (i) subsets of a species or population evolve to specialize on different ecological resources and (ii) the resulting phenotypic modes become reproductively isolated. Empirical evidence for these two processes working in conjunction, particularly during the early stages of divergence, has been limited. We recently described a population of the medium ground finch, Geospiza fortis, that features large and small beak morphs with relatively few intermediates. As in other Darwin's finches of the Galápagos Islands, these morphs presumably diverged in response to variation in local food availability and inter- or intraspecific competition. We here demonstrate that the two morphs show strong positive assortative pairing, a pattern that holds over three breeding seasons and during both dry and wet conditions. We also document restrictions on gene flow between the morphs, as revealed by genetic variation at 10 microsatellite loci. Our results provide strong support for the central role of ecology during the early stages of adaptive radiation.     

A conservative test of genetic drift in the endosymbiotic bacterium Buchnera: slightly deleterious mutations in the chaperonin groEL

The obligate endosymbiotic bacterium Buchnera aphidicola shows elevated rates of sequence evolution compared to free-living relatives, particularly at nonsynonymous sites. Because Buchnera experiences population bottlenecks during transmission to the offspring of its aphid host, it is hypothesized that genetic drift and the accumulation of slightly deleterious mutations can explain this rate increase. Recent studies of intraspecific variation in Buchnera reveal patterns consistent with this hypothesis. In this study, we examine inter- and intraspecific nucleotide variation in groEL, a highly conserved chaperonin gene that is constitutively overexpressed in Buchnera. Maximum-likelihood estimates of nonsynonymous substitution rates across Buchnera species are strikingly low at groEL compared to other loci. Despite this evidence for strong purifying selection on groEL, our intraspecific analysis of this gene documents reduced synonymous polymorphism, elevated nonsynonymous polymorphism, and an excess of rare alleles relative to the neutral expectation, as found in recent studies of other Buchnera loci. Comparisons with Escherichia coli generally show patterns predicted by their differences in N(e). The sum of these observations is not expected under relaxed or balancing selection, selective sweeps, or increased mutation rate. Rather, they further support the hypothesis that drift is an important force driving accelerated protein evolution in this obligate mutualist.     

Community assembly in Nothobranchius annual fishes: Nested patterns,environmental niche and biogeographic history

The assembly of local communities from regional species pools is shaped by historical aspects of distribution, environmental conditions, and biotic interactions. We studied local community assembly patterns in African annual killifishes of the genus Nothobranchius (Cyprinodontiformes), investigating data from 168 communities across the entire range of regionally co‐existing species. Nothobranchius are small fishes associated with annually desiccating pools. We detected a nested pattern of local communities in one region (Southern Mozambique, with Nothobranchius furzeri as the core and dominant species), but no nestedness was found in the second region (Central Mozambique, with Nothobranchius orthonotus being the dominant species). A checkerboard pattern of local Nothobranchius community assembly was demonstrated in both regions. Multivariate environmental niche modeling revealed moderate differences in environmental niche occupancy between three monophyletic clades that largely co‐occurred geographically and greater differences between strictly allopatric species within the clades. Most variation among species was observed along an altitudinal gradient; N. furzeri and Nothobranchius kadleci were absent from coastal plains, Nothobranchius pienaari, Nothobranchius rachovii, and Nothobranchius krysanovi were associated with lower altitude and N. orthonotus was intermediate and geographically most widespread species. We discuss implications for ecological and evolutionary research in this taxon.     

Phylogenetic analysis of nuclear and mitochondrial genes reveals evolutionary relationships and mitochondrial introgression in the sertifer species group of the genus Neodiprion (Hymenoptera: Diprionidae)

Neodiprion Rohwer (Hymenoptera: Diprionidae) is a Holarctic genus of conifer-feeding sawflies with a remarkable amount of inter- and intraspecific diversity in host use, behavior, and development. This variation is thought to play a central role in Neodiprion diversification, but speciation hypotheses remain untested due to a lack of a robust phylogenetic estimate. Here, we utilize sequence data from three nuclear genes (CAD, ANL43, EF1alpha) to obtain a phylogenetic estimate for the genus. These analyses suggest that: (1) North American and Eurasian Neodiprion are monophyletic sister clades, (2) the sertifer group is paraphyletic with respect to the monophyletic lecontei group, and (3) on at least two occasions, dispersal from eastern to western North America proceeded via southern host bridges. Based on these results and host biogeography, we revise a previous scenario for the evolution of Neodiprion and suggest maximum ages for the genus and for the lecontei group (25 My and 14 My, respectively). In addition, because a previous study reported rampant mitochondrial introgression in the lecontei group, we assess its prevalence in the sertifer group. Analysis of three mitochondrial genes (COI, tRNA-leucine, and COII) reveals that mito-nuclear discordance is prevalent in the sertifer group, and patterns of species monophyly are consistent with those expected under frequent mitochondrial introgression. As was the case for lecontei group species, we find that introgression appears to be most pronounced between species that occasionally share hosts, suggesting that divergent host use is an important barrier to gene flow in Neodiprion. Finally, we suggest that the lack of phylogenetic resolution and prevalence of species non-monophyly in the non-Pinus feeding Neodiprion may result from the rapid divergence (possibly with gene flow) of these species following their entry into a novel adaptive zone.     

Intraspecific variation of flower colour and its distribution within a sea lavender, Limonium wrightii (Plumbaginaceae), in the northwestern Pacific Islands

Differentiation of flower colour is thought to be one of the most important factors promoting plant speciation. We describe the intraspecific variation of flower colour and its distribution in Limonium wrightii. We conducted a survey on 36 islands in the northwestern Pacific and discriminated six morphs of flower colour variation. Two flower colour morphs, pink and yellow, were most frequently observed, and their geographical distributions were basically allopatric. These two morphs were in contact in a narrow zone on Okinoerabu Island, located in the middle region of the Ryukyu Archipelago. In addition, orange, white, and ivory flower morphs were also found in this zone. The geographical distribution of pink and yellow morphs showed a “leapfrog” pattern; the distribution of pink flowers was divided into two areas, intercalated by the distribution of the yellow flower morph. The orange morph may have resulted from hybridization between the pink and yellow flower morphs.     

The evolution of male nuptial colour in a sexually dimorphic group of fishes (Percidae: Etheostomatinae)

To test hypotheses explaining variation in elaborate male colouration across closely related species groups, ancestral‐state reconstructions and tests of phylogenetic signal and correlated evolution were used to examine the evolution of male body and fin colouration in a group of sexually dichromatic stream fishes known as darters (Percidae: Etheostomatinae). The presence or absence of red–orange and blue–green male colour traits were scored across six body regions in 99 darter species using a recently estimated amplified fragment length polymorphism (AFLP) phylogeny for comparative analyses. Ancestral‐state reconstructions infer the most recent common ancestor of darters to lack red–orange colour and possess blue–green colour on different body regions, suggesting variation between species is due to independent gains of red–orange and losses of blue–green. Colour traits exhibit substantial phylogenetic signal and are highly correlated across body regions. Comparative analyses were repeated using an alternative phylogenetic hypothesis based on one mitochondrial and two nuclear genes, yielding similar results to analyses based on the AFLP phylogeny. Red–orange colouration in darters appears to be derived; whereas, blue–green appears to be ancestral, which suggests that different selection mechanisms may be acting on these two colour classes in darters.     

Linkage mapping of a polymorphic plumage locus associated with intermorph incompatibility in the Gouldian finch (Erythrura gouldiae)

Colour polymorphism is known to facilitate speciation but the genetic basis of animal pigmentation and how colour polymorphisms contribute to speciation is poorly understood. Restricted recombination may promote linkage disequilibrium between the colour locus and incompatibility genes. Genomic rearrangement and the position of relevant loci within a chromosome are important factors that influence the frequency of recombination. Therefore, it is important to know the position of the colour locus, gene order and recombination landscape of the chromosome to understand the mechanism that generates incompatibilities between morphs. Recent studies showed remarkable pre- and postzygotic incompatibilities between sympatric colour morphs of the Gouldian finch (Erythrura gouldiae), in which head feather colour is genetically determined by a single sex-linked locus, Red. We constructed a genetic map for the Z chromosome of the Gouldian finch (male-specific map distance=131 cM), using 618 captive-bred birds and 34 microsatellite markers, to investigate the extent of inter- and intraspecific genomic rearrangements and variation in recombination rate within the Z chromosome. We refined the location of the Red locus to a ~7.2-cM interval in a region with a moderate recombination rate but outside the least-recombining, putative centromeric region. There was no evidence of chromosome-wide genomic rearrangements between the chromosomes carrying the red or black alleles with the current marker resolution. This work will contribute to identifying the causal gene, which will in turn enable alternative explanations for the association between incompatibility and colouration, such as fine-scale linkage disequilibrium, genomic rearrangements and pleiotropy, to be tested.     

Evolution of flower morphology, pollen dimorphism, and nectar composition in Arcytophyllum, a distylous genus of Rubiaceae

A phylogenetic study of Arcytophyllum based on ITS was conducted and compared with an earlier study based on cpDNA. The position of the widespread A. thymifolium as sister to all other species was confirmed and several well-supported clades could be retrieved. The Central American A. lavarum is well embedded between exclusively or predominantly South American species. To understand the expression of heterostyly in the genus, we analyzed inter- and intraspecific variation in floral morphology, nectar, pollen–ovule (P/O) ratio and seed set of ten species in 11 populations. Stigma and anther levels differed significantly between the morphs in the species/populations investigated, except for A. filiforme, in which anther levels did not differ significantly between the two morphs. Different expressions of heterostyly in Arcytophyllum seem independent of phylogenetic relationships. Nectar sugar composition was similar between the morphs. Nectar of most species presented a larger proportion of hexoses than of sucrose; only the most derived species, A. macbridei and A. vernicosum, have higher sucrose proportions. There is a significant positive correlation between corolla tube length and the proportion of sucrose. Pollen dimorphism, with regard to both number (long-styled>short-styled) and size (short-styled>long-styled), was observed in all taxa investigated except A. filiforme. According to the P/O ratios, the breeding systems range from facultative autogamy to facultative xenogamy. The lowest P/O ratios were found in A. filiforme, and the highest in A. rivetii. Hymenoptera, Diptera and Coleoptera were observed as flower visitors. Seed production did not differ significantly between the morphs in eight of the 11 species/populations investigated. There is, however, a tendency in all species/populations (except in A. macbridei Peru) for the short-styled morph to have a higher percentage of seeds per ovule, indicating that the short-styled morphs display higher female reproductive success.