Population structure and the co-evolution between social parasites and their hosts

Co-evolutionary trajectories of host-parasite interactions are strongly affected by the antagonists' evolutionary potential, which in turn depends on population sizes as well as levels of recombination, mutation, and gene flow. Under similar selection pressures, the opponent with the higher evolutionary rate is expected to lead the co-evolutionary arms race and to develop local adaptations. Here, we use mitochondrial DNA sequence data and microsatellite markers to assess the amount of genetic variability and levels of gene flow in two host-parasite systems, each consisting of an ant social parasite--the European slavemaker Harpagoxenus sublaevis and the North American slavemaker Protomognathus americanus--and its two main host species. Our population genetic analyses revealed limited gene flow between individual populations of both host and parasite species, allowing for a geographic mosaic of co-evolution. In a between-system comparison, we found less genetic variability and more pronounced structure in Europe, where previous behavioural studies demonstrated strong local adaptation. Within the European host-parasite system, the larger host species Leptothorax acervorum exhibited higher levels of both genetic variability and gene flow, and previous field data showed that it is less affected by the social parasite H. sublaevis than the smaller host Leptothorax muscorum, which has genetically depleted and isolated populations. In North America, the parasite P. americanus showed higher levels of gene flow between sites, but overall less genetic diversity than its hyper-variable main host species, Temnothorax longispinosus. Interestingly, recent ecological and chemical studies demonstrated adaptation of P. americanus to local host populations, indicating the importance of migration in co-evolutionary interactions.     

Population structuring of the ubiquitous stingless bee Tetragonisca angustula in southern Brazil as revealed by microsatellite and mitochondrial markers

Tetragonisca angustula is one of the most widespread stingless bees in the Neotropics. This species swarms frequently and is extremely successful in urban environments. In addition, it is one of the most popular stingless bee species for beekeeping in Latin America, so nest transportation and trading is common. Nest transportation can change the genetic structure of the host population, reducing inbreeding and increasing homogenization. Here, we evaluate the genetic structure of 17 geographic populations of T. angustula in southern Brazil to quantify the level of genetic differentiation between populations. Analyses were conducted on partially sequenced mitochondrial genes and 11 microsatellite loci of 1002 workers from 457 sites distributed on the mainland and on 3 islands. Our results show that T. angustula populations are highly differentiated as demonstrated by mitochondrial DNA (mtDNA) and microsatellite markers. Of 73 haplotypes, 67 were population‐specific. MtDNA diversity was low in 9 populations but microsatellite diversity was moderate to high in all populations. Microsatellite data suggest 10 genetic clusters and low level of gene flow throughout the studied area. However, physical barriers, such as rivers and mountain ranges, or the presence or absence of forest appear to be unrelated to population clusters. Factors such as low dispersal, different ecological conditions, and isolation by distance are most likely shaping the population structure of this species. Thus far, nest transportation has not influenced the general population structure in the studied area. However, due to the genetic structure we found, we recommend that nest transportation should only occur within and between populations that are genetically similar.     

Multilocus genetic analyses and spatial modeling reveal complex population structure and history in a widespread resident North American passerine (Perisoreus canadensis)

An increasing body of studies of widely distributed, high latitude species shows a variety of refugial locations and population genetic patterns. We examined the effects of glaciations and dispersal barriers on the population genetic patterns of a widely distributed, high latitude, resident corvid, the gray jay (Perisoreus canadensis), using the highly variable mitochondrial DNA (mtDNA) control region and microsatellite markers combined with species distribution modeling. We sequenced 914 bp of mtDNA control region for 375 individuals from 37 populations and screened seven loci for 402 individuals from 27 populations across the gray jay range. We used species distribution modeling and a range of phylogeographic analyses (haplotype diversity, Φ_ST, SAMOVA, F_ST, Bayesian clustering analyses) to examine evolutionary history and population genetic structure. MtDNA and microsatellite markers revealed significant genetic differentiation among populations with high concordance between markers. Paleodistribution models supported at least five potential areas of suitable gray jay habitat during the last glacial maximum and revealed distributions similar to the gray jay's contemporary during the last interglacial. Colonization from and prolonged isolation in multiple refugia is evident. Historical climatic fluctuations, the presence of multiple dispersal barriers, and highly restricted gene flow appear to be responsible for strong genetic diversification and differentiation in gray jays.     

Loss of genetic connectivity and diversity in urban microreserves in a southern California endemic Jerusalem cricket (Orthoptera: Stenopelmatidae: Stenopelmatus n. sp. “santa monica”)

Microreserves may be useful in protecting native arthropod diversity in urbanized landscapes. However, species that do not disperse through the urban matrix may eventually be lost from these fragments. Population extinctions may be precipitated by an increase in genetic differentiation among fragments and loss of genetic diversity within fragments, and these effects should become stronger with time. We analyzed population genetic structure in the dispersal limited Jerusalem cricket Stenopelmatus n. sp. “santa monica” in the Santa Monica Mountains and Simi Hills north of Los Angeles, California (CA), to determine the impacts of fragmentation over the past 70 years. MtDNA divergence was greater among urban fragments than within contiguous habitat and was positively correlated with fragment age. MtDNA genetic diversity within fragments increased with fragment size and decreased with fragment age. Genetic divergence across 38 anonymous nuclear Inter-Simple Sequence Repeat (ISSR) loci was influenced by the presence of major highways and highway age, but there was no effect of additional urban fragmentation. ISSR diversity was not correlated with fragment size or age. Differing results between markers may be due to male-biased dispersal, or different effective population sizes, sorting rates, or mutation rates among sampled genes. Results suggest that genetic connectivity among populations has been disrupted by highways and urban development, prior to declines in local population sizes. We emphasize that genetic connectivity can rapidly erode in fragmented landscapes and that flightless arthropods can serve as sensitive indicators for these effects.     

Genetic costructure in a meta‐community under threat of habitat fragmentation

Habitat fragmentation increasingly threatens the services provided by natural communities and ecosystem worldwide. An understanding of the eco‐evolutionary processes underlying fragmentation‐compromised communities in natural settings is lacking, yet critical to realistic and sustainable conservation. Through integrating the multivariate genetic, biotic and abiotic facets of a natural community module experiencing various degrees of habitat fragmentation, we provide unique insights into the processes underlying community functioning in real, natural conditions. The focal community module comprises a parasitic butterfly of conservation concern and its two obligatory host species, a plant and an ant. We show that both historical dispersal and ongoing habitat fragmentation shape population genetic diversity of the butterfly Phengaris alcon and its most limited host species (the plant Gentiana pneumonanthe). Genetic structure of each species was strongly driven by geographical structure, altitude and landscape connectivity. Strikingly, however, was the strong degree of genetic costructure among the three species that could not be explained by the spatial variables under study. This finding suggests that factors other than spatial configuration, including co‐evolutionary dynamics and shared dispersal pathways, cause parallel genetic structure among interacting species. While the exact contribution of co‐evolution and shared dispersal routes on the genetic variation within and among communities deserves further attention, our findings demonstrate a considerable degree of genetic parallelism in natural meta‐communities. The significant effect of landscape connectivity on the genetic diversity and structure of the butterfly also suggests that habitat fragmentation may threaten the functioning of the community module on the long run.     

Host dispersal as the driver of parasite genetic structure: a paradigm lost?

Understanding traits influencing the distribution of genetic diversity has major ecological and evolutionary implications for host–parasite interactions. The genetic structure of parasites is expected to conform to that of their hosts, because host dispersal is generally assumed to drive parasite dispersal. Here, we used a meta‐analysis to test this paradigm and determine whether traits related to host dispersal correctly predict the spatial co‐distribution of host and parasite genetic variation. We compiled data from empirical work on local adaptation and host–parasite population genetic structure from a wide range of taxonomic groups. We found that genetic differentiation was significantly lower in parasites than in hosts, suggesting that dispersal may often be higher for parasites. A significant correlation in the pairwise genetic differentiation of hosts and parasites was evident, but surprisingly weak. These results were largely explained by parasite reproductive mode, the proportion of free‐living stages in the parasite life cycle and the geographical extent of the study; variables related to host dispersal were poor predictors of genetic patterns. Our results do not dispel the paradigm that parasite population genetic structure depends on host dispersal. Rather, we highlight that alternative factors are also important in driving the co‐distribution of host and parasite genetic variation.     

Evolution of Pathogen Specialisation in a Host Metapopulation: Joint Effects of Host and Pathogen Dispersal

Metapopulation processes are important determinants of epidemiological and evolutionary dynamics in host-pathogen systems, and are therefore central to explaining observed patterns of disease or genetic diversity. In particular, the spatial scale of interactions between pathogens and their hosts is of primary importance because migration rates of one species can affect both spatial and temporal heterogeneity of selection on the other. In this study we developed a stochastic and discrete time simulation model to specifically examine the joint effects of host and pathogen dispersal on the evolution of pathogen specialisation in a spatially explicit metapopulation. We consider a plant-pathogen system in which the host metapopulation is composed of two plant genotypes. The pathogen is dispersed by air-borne spores on the host metapopulation. The pathogen population is characterised by a single life-history trait under selection, the infection efficacy. We found that restricted host dispersal can lead to high amount of pathogen diversity and that the extent of pathogen specialisation varied according to the spatial scale of host-pathogen dispersal. We also discuss the role of population asynchrony in determining pathogen evolutionary outcomes.     

The socially parasitic ant Polyergus mexicanus has host‐associated genetic population structure and related neighbouring colonies

The genetic structure of populations can be both a cause and a consequence of ecological interactions. For parasites, genetic structure may be a consequence of preferences for host species or of mating behaviour. Conversely, genetic structure can influence where conspecific interactions among parasites lay on a spectrum from cooperation to conflict. We used microsatellite loci to characterize the genetic structure of a population of the socially parasitic dulotic (aka “slave‐making”) ant (Polyergus mexicanus), which is known for its host‐specificity and conspecific aggression. First, we assessed whether the pattern of host species use by the parasite has influenced parasite population structure. We found that host species use was correlated with subpopulation structure, but this correlation was imperfect: some subpopulations used one host species nearly exclusively, while others used several. Second, we examined the viscosity of the parasite population by measuring the relatedness of pairs of neighbouring parasitic ant colonies at varying distances from each other. Although natural history observations of local dispersal by queens suggested the potential for viscosity, there was no strong correlation between relatedness and distance between colonies. However, 35% of colonies had a closely related neighbouring colony, indicating that kinship could potentially affect the nature of some interactions between colonies of this social parasite. Our findings confirm that ecological forces like host species selection can shape the genetic structure of parasite populations, and that such genetic structure has the potential to influence parasite‐parasite interactions in social parasites via inclusive fitness.     

Phoretic dispersal influences parasite population genetic structure

Dispersal is a fundamental component of the life history of most species. Dispersal influences fitness, population dynamics, gene flow, genetic drift and population genetic structure. Even small differences in dispersal can alter ecological interactions and trigger an evolutionary cascade. Linking such ecological processes with evolutionary patterns is difficult, but can be carried out in the proper comparative context. Here, we investigate how differences in phoretic dispersal influence the population genetic structure of two different parasites of the same host species. We focus on two species of host‐specific feather lice (Phthiraptera: Ischnocera) that co‐occur on feral rock pigeons (Columba livia). Although these lice are ecologically very similar, “wing lice” (Columbicola columbae) disperse phoretically by “hitchhiking” on pigeon flies (Diptera: Hippoboscidae), while “body lice” (Campanulotes compar) do not. Differences in the phoretic dispersal of these species are thought to underlie observed differences in host specificity, as well as the degree of host–parasite cospeciation. These ecological and macroevolutionary patterns suggest that body lice should exhibit more genetic differentiation than wing lice. We tested this prediction among lice on individual birds and among lice on birds from three pigeon flocks. We found higher levels of genetic differentiation in body lice compared to wing lice at two spatial scales. Our results indicate that differences in phoretic dispersal can explain microevolutionary differences in population genetic structure and are consistent with macroevolutionary differences in the degree of host–parasite cospeciation.     

Genetic diversity,population structure and sex‐biased dispersal in three co‐evolving species

Genetic diversity and spatial structure of populations are important for antagonistic coevolution. We investigated genetic variation and population structure of three closely related European ant species: the social parasite Harpagoxenus sublaevis and its two host species Leptothorax acervorum and Leptothorax muscorum. We sampled populations in 12 countries and analysed eight microsatellite loci and an mtDNA sequence. We found high levels of genetic variation in all three species, only slightly less variation in the host L. muscorum. Using a newly introduced measure of differentiation (Jost’s D_est ), we detected strong population structuring in all species and less male‐biased dispersal than previously thought. We found no phylogeographic patterns that could give information on post‐glacial colonization routes – northern populations are as variable as more southern populations. We conclude that conditions for Thompson’s geographic mosaic of coevolution are ideal in this system: all three species show ample genetic variation and strong population structure.     

Life history determines genetic structure and evolutionary potential of host-parasite interactions

Measures of population genetic structure and diversity of disease-causing organisms are commonly used to draw inferences regarding their evolutionary history and potential to generate new variation in traits that determine interactions with their hosts. Parasite species exhibit a range of population structures and life-history strategies, including different transmission modes, life-cycle complexity, off-host survival mechanisms and dispersal ability. These are important determinants of the frequency and predictability of interactions with host species. Yet the complex causal relationships between spatial structure, life history and the evolutionary dynamics of parasite populations are not well understood. We demonstrate that a clear picture of the evolutionary potential of parasitic organisms and their demographic and evolutionary histories can only come from understanding the role of life history and spatial structure in influencing population dynamics and epidemiological patterns.     

Population genetic structure and male-biased dispersal in the queenless ant Diacamma cyaneiventre

In this study we investigated the population genetic structure of the queenless ant Diacamma cyaneiventre. This species, lacking winged queens, is likely to have a restricted female dispersal. We used both mitochondrial and microsatellite markers to assess the consequence of such restricted female dispersal at three geographical scales: within a given locality (< 1 km), between localities within a given region (< 10 km) and between regions (> 36 km). Within a locality, a strong population structure was observed for mitochondrial DNA (mtDNA) whereas weak or nonexistent population genetic structure was observed for the microsatellites (around 5% of the value for mtDNA). Male gene flow was estimated to be about 20-30 times higher than female gene flow at this scale. At a larger spatial scale, very strong genetic differentiation for both markers was observed between localities - even within a single region. Female dispersal is nonexistent at these scales and male dispersal is very restricted, especially between regions. The phylogeographical structure of the mtDNA haplotypes as well as the very low genetic diversity of mtDNA within localities indicate that new sites are colonized by a single migration event from adjacent localities, followed by successive colony fissions. These patterns of genetic variability and differentiation agree with what is theoretically expected when colonization events are kin-structured and when, following colonization, dispersion is mainly performed by males.     

Very low genetic variability in the Indian queenless ant Diacamma indicum

Abstract We developed microsatellite markers and combined them with mitochondrial markers to analyse the population genetic structure of the queenless ant Diacamma indicum. This species, lacking winged queens, is likely to have a restricted female dispersal but exhibits various life history traits suggesting higher dispersal abilities than the other Diacamma species. Only 4 of 11 microsatellites were polymorphic and only 1 had more than 4 alleles over 166 individuals originating from 7 populations from the south of India. Only one mitochondrial DNA (mtDNA) haplotype was detected throughout India (including one population in the north) and Sri Lanka. Such a level of polymorphism is particularly low compared with other Diacamma species having much smaller ranges in the south of India. A strong genetic differentiation was observed between populations separated by more than a few kilometres. We also analysed the genetic differentiation between the Indian populations and two populations from the Japanese island of Okinawa, which are morphologically similar and might belong to the same species. The genetic differentiation was high for both markers, suggesting an absence of ongoing gene flow between these populations.     

Male‐biased dispersal in a fungus‐gardening ant symbiosis

For nearly all organisms, dispersal is a fundamental life‐history trait that can shape their ecology and evolution. Variation in dispersal capabilities within a species exists and can influence population genetic structure and ecological interactions. In fungus‐gardening (attine) ants, co‐dispersal of ants and mutualistic fungi is crucial to the success of this obligate symbiosis. Female‐biased dispersal (and gene flow) may be favored in attines because virgin queens carry the responsibility of dispersing the fungi, but a paucity of research has made this conclusion difficult. Here, we investigate dispersal of the fungus‐gardening ant Trachymyrmex septentrionalis using a combination of maternally (mitochondrial DNA) and biparentally inherited (microsatellites) markers. We found three distinct, spatially isolated mitochondrial DNA haplotypes; two were found in the Florida panhandle and the other in the Florida peninsula. In contrast, biparental markers illustrated significant gene flow across this region and minimal spatial structure. The differential patterns uncovered from mitochondrial DNA and microsatellite markers suggest that most long‐distance ant dispersal is male‐biased and that females (and concomitantly the fungus) have more limited dispersal capabilities. Consequently, the limited female dispersal is likely an important bottleneck for the fungal symbiont. This bottleneck could slow fungal genetic diversification, which has significant implications for both ant hosts and fungal symbionts regarding population genetics, species distributions, adaptive responses to environmental change, and coevolutionary patterns.     

Development of microsatellite markers for Diaphanosoma dubium (Crustacea,Cladocera) and application to seasonal population dynamics

Diaphanosoma, the “tropical Daphnia”, is common and ubiquitous in South China. Like other ctenopods, Diaphanosoma has a reproductive mode similar to Daphnia’s, but its resting eggs are rarely observed and lack an ephippium. With limited dispersal and reduced buffer effect from resting egg banks, Diaphanosoma is expected to have a population genetic structure different from that of temperate Daphnia. To facilitate genetic comparison, we developed microsatellite markers using next-generation sequencing for the most common species in tropical and subtropical East Asia, Diaphanosoma dubium. Thirty-one polymorphic microsatellite markers were obtained, and 29 of them were efficient for the congeneric species D. excisum, D. orghidani, D. mongolianum and D. chankensis. The markers allowed intra- and interspecific genetic analysis, including population structure, hybridization and introgression. We used 11 selected microsatellite markers to analyze spatial and temporal heterogeneity of genetic diversity in four (sub)tropical D. dubium populations from two large reservoirs and two temporary ponds. In contrast to temperate Cladocera, higher genetic diversity in summer rather than in spring suggested weak contribution from resting eggs in spring. Clustering of DAPC and STRUCTURE analyses indicated a clear-cut genetic structure in the four populations. Variation partitioning revealed that water storage and depth were key factors in genetic differentiation. Within large reservoirs, we detected backward (reversing time) gene flow from resting egg banks. We conclude that resting eggs have an effective contribution to the genetic diversity in large water bodies during growing seasons and that large water bodies can host higher genetic diversity in summer due to environmental heterogeneity and high carrying capacity. Spatial and temporal heterogeneity of genetic diversity detected by our microsatellite markers showed the newly developed markers can be applied for further study of populations of D. dubium and other species of Diaphanosoma at a contemporary scale.

     

Lousy grouse: Comparing evolutionary patterns in Alaska galliform lice to understand host evolution and host–parasite interactions

Understanding both sides of host–parasite relationships can provide more complete insights into host and parasite biology in natural systems. For example, phylogenetic and population genetic comparisons between a group of hosts and their closely associated parasites can reveal patterns of host dispersal, interspecies interactions, and population structure that might not be evident from host data alone. These comparisons are also useful for understanding factors that drive host–parasite coevolutionary patterns (e.g., codivergence or host switching) over different periods of time. However, few studies have compared the evolutionary histories between multiple groups of parasites from the same group of hosts at a regional geographic scale. Here, we used genomic data to compare phylogenomic and population genomic patterns of Alaska ptarmigan and grouse species (Aves: Tetraoninae) and two genera of their associated feather lice: Lagopoecus and Goniodes. We used whole‐genome sequencing to obtain hundreds of genes and thousands of single‐nucleotide polymorphisms (SNPs) for the lice and double‐digest restriction‐associated DNA sequences to obtain SNPs from Alaska populations of two species of ptarmigan. We found that both genera of lice have some codivergence with their galliform hosts, but these relationships are primarily characterized by host switching and phylogenetic incongruence. Population structure was also uncorrelated between the hosts and lice. These patterns suggest that grouse, and ptarmigan in particular, share habitats and have likely had historical and ongoing dispersal within Alaska. However, the two genera of lice also have sufficient dissimilarities in the relationships with their hosts to suggest there are other factors, such as differences in louse dispersal ability, that shape the evolutionary patterns with their hosts.     

Strong dispersal in a parasitoid wasp overwhelms habitat fragmentation and host population dynamics

The population dynamics of a parasite depend on species traits, host dynamics and the environment. Those dynamics are reflected in the genetic structure of the population. Habitat fragmentation has a greater impact on parasites than on their hosts because resource distribution is increasingly fragmented for species at higher trophic levels. This could lead to either more or less genetic structure than the host, depending on the relative dispersal rates of species. We examined the spatial genetic structure of the parasitoid wasp Hyposoter horticola, and how it was influenced by dispersal, host population dynamics and habitat fragmentation. The host, the Glanville fritillary butterfly, lives as a metapopulation in a fragmented landscape in the Åland Islands, Finland. We collected wasps throughout the 50 by 70 km archipelago and determined the genetic diversity, spatial population structure and genetic differentiation using 14 neutral DNA microsatellite loci. We compared the genetic structure of the wasp with that of the host butterfly using published genetic data collected over the shared landscape. Using maternity assignment, we also identified full‐siblings among the sampled parasitoids to estimate the dispersal range of individual females. We found that because the parasitoid is dispersive, it has low genetic structure, is not very sensitive to habitat fragmentation and has less spatial genetic structure than its butterfly host. The wasp is sensitive to regional rather than local host dynamics, and there is a geographic mosaic landscape for antagonistic co‐evolution of host resistance and parasite virulence.     

Colony structure of a slavemaking ant. II. Frequency of slave raids and impact on the host population

The parasite pressure exerted by the slavemaker ant Protomognathus americanus on its host species Leptothorax longispinosus was analyzed demographically and genetically. The origin of slaves found in colonies of the obligate slavemaker was examined with nuclear and mitochondrial DNA markers to make inferences about the frequency and severity of slave raids. Relatedness of enslaved L. longispinosus workers in the same nest was very low, and our data suggest that, on average, each slavemaker nest raids six host colonies per season. Therefore, the influence of slavemaker species on their hosts is much stronger than simple numerical ratios suggest. We also found that slave relatedness was higher in small than in large slavemaker nests; thus, larger nests wield a much stronger influence on the host. We estimated that in the study population, on average, a host nest has a 50% chance of being attacked by a slavemaker colony per year. Free-living Leptothorax colonies in the vicinity of slavemaker nests did not represent the source of slaves working in P. americanus colonies, which suggests that raided nests either do not survive or migrate after being raided. Colony composition and intranest relatedness of free-living L. longispinosus colonies differed markedly between areas with slavemakers and those that are parasite-free. In the presence of slavemakers, host colonies were less likely to be polygynous and had fewer workers and a higher relatedness among worker brood. Host nests with slavemaker neighbors allocated more resources into sexuals, possibly caused by these shifts in nest demography. Finally, enslaved Leptothorax workers in P. americanus nests appeared to be less efficient than their counterparts in free-living colonies. Thus, slavemakers exert a much stronger impact on their hosts than had previously been suspected and represent an unique system to study parasite-host coevolution.     

Absence of isolation by distance patterns at the regional scale in the fungal plant pathogen Leptosphaeria maculans

Outcomes of host-pathogen coevolution are influenced by migration rates of the interacting species. Reduced gene flow with increasing spatial distance between populations leads to spatial genetic structure, as predicted by the isolation by distance (IBD) model. In wind-dispersed plant-pathogenic fungi, a significant spatial genetic structure is theoretically expected if local spore dispersal is more frequent than long-distance dispersal, but this remains to be documented by empirical data. For 29 populations of the oilseed rape fungus Leptosphaeria maculans sampled from two French regions, genetic structure was determined using eight minisatellite markers. Gene diversity (H = 0.62-0.70) and haplotypic richness (R = 0.96-1) were high in all populations. No linkage disequilibrium was detected between loci, suggesting the prevalence of panmictic sexual reproduction. Analysis of molecular variance showed that > 97% of genetic diversity was observed within populations. Genetic differentiation was low among populations (F(st) < 0.05). Although direct methods previously revealed short-distance dispersal for L. maculans, our findings of no correlation between genetic and geographic distances among populations illustrate that the IBD model does not account for dispersal of the fungus at the spatial scale we examined. These results indicate high gene flow among French populations of L. maculans, suggesting high dispersal rates and/or large effective population sizes, two characteristics giving the pathogen high evolutionary potential against the deployment of resistant oilseed rape cultivars.     

Historic hybridization and introgression between two iconic Australian anemonefish and contemporary patterns of population connectivity

Endemic species on islands are considered at risk of extinction for several reasons, including limited dispersal abilities, small population sizes, and low genetic diversity. We used mitochondrial DNA (D-Loop) and 17 microsatellite loci to explore the evolutionary relationship between an endemic anemonefish, Amphiprion mccullochi (restricted to isolated locations in subtropical eastern Australia) and its more widespread sister species, A. akindynos. A mitochondrial DNA (mtDNA) phylogram showed reciprocal monophyly was lacking for the two species, with two supported groups, each containing representatives of both species, but no shared haplotypes and up to 12 species, but not location-specific management units (MUs). Population genetic analyses suggested evolutionary connectivity among samples of each species (mtDNA), while ecological connectivity was only evident among populations of the endemic, A. mccullochi. This suggests higher dispersal between endemic anemonefish populations at both evolutionary and ecological timeframes, despite separation by hundreds of kilometers. The complex mtDNA structure results from historical hybridization and introgression in the evolutionary past of these species, validated by msat analyses (NEWHYBRIDS, STRUCTURE, and DAPC). Both species had high genetic diversities (mtDNA h > 0.90, π = 4.0%; msat genetic diversity, gd > 0.670). While high gd and connectivity reduce extinction risk, identifying and protecting populations implicated in generating reticulate structure among these species should be a conservation priority.