A mitochondrial species identification assay for Australian blacktip sharks (Carcharhinus tilstoni, C. limbatus and C. amblyrhynchoides) using real-time PCR and high-resolution melt analysis

Tropical Australian shark fisheries target two morphologically indistinguishable blacktip sharks, the Australian blacktip (Carcharhinus tilstoni) and the common blacktip (C. limbatus). Their relative contributions to northern and eastern Australian coastal fisheries are unclear because of species identification difficulties. The two species differ in their number of precaudal vertebrae, which is difficult and time consuming to obtain in the field. But, the two species can be distinguished genetically with diagnostic mutations in their mitochondrial DNA ND4 gene. A third closely related sister species, the graceful shark C. amblyrhynchoides, can also be distinguished by species‐specific mutations in this gene. DNA sequencing is an effective diagnostic tool, but is relatively expensive and time consuming. In contrast, real‐time high‐resolution melt (HRM) PCR assays are rapid and relatively inexpensive. These assays amplify regions of DNA with species‐specific genetic mutations that result in PCR products with unique melt profiles. A real‐time HRM PCR species‐diagnostic assay (RT‐HRM‐PCR) has been developed based on the mtDNA ND4 gene for rapid typing of C. tilstoni, C. limbatus and C. amblyrhynchoides. The assay was developed using ND4 sequences from 66 C. tilstoni, 33. C. limbatus and five C. amblyrhynchoides collected from Indonesia and Australian states and territories; Western Australia, the Northern Territory, Queensland and New South Wales. The assay was shown to be 100% accurate on 160 unknown blacktip shark tissue samples by full mtDNA ND4 sequencing.     

A novel field method to distinguish between cryptic carcharhinid sharks,Australian blacktip shark Carcharhinus tilstoni and common blacktip shark C. limbatus,despite the presence of hybrids

Multivariate and machine‐learning methods were used to develop field identification techniques for two species of cryptic blacktip shark. From 112 specimens, precaudal vertebrae (PCV) counts and molecular analysis identified 95 Australian blacktip sharks Carcharhinus tilstoni and 17 common blacktip sharks Carcharhinus limbatus. Molecular analysis also revealed 27 of the 112 were C. tilstoni × C. limbatus hybrids, of which 23 had C. tilstoni PCV counts and four had C. limbatus PCV counts. In the absence of further information about hybrid phenotypes, hybrids were assigned as either C. limbatus or C. tilstoni based on PCV counts. Discriminant analysis achieved 80% successful identification, but machine‐learning models were better, achieving 100% successful identification, using six key measurements (fork length, caudal‐fin peduncle height, interdorsal space, second dorsal‐fin height, pelvic‐fin length and pelvic‐fin midpoint to first dorsal‐fin insertion). Furthermore, pelvic‐fin markings could be used for identification: C. limbatus has a distinct black mark >3% of the total pelvic‐fin area, while C. tilstoni has markings with diffuse edges, or has smaller or no markings. Machine learning and pelvic‐fin marking identification methods were field tested achieving 87 and 90% successful identification, respectively. With further refinement, the techniques developed here will form an important part of a multi‐faceted approach to identification of C. tilstoni and C. limbatus and have a clear management and conservation application to these commercially important sharks. The methods developed here are broadly applicable and can be used to resolve species identities in many fisheries where cryptic species exist.     

New microsatellite loci for Carcharhinid sharks (Carcharhinus tilstoni and C. sorrah) and their cross‐amplification in other shark species

Twelve microsatellite DNA markers were isolated in the spot‐tail shark (Carcharhinus sorrah) and nine were isolated in Australian black‐tip shark (Carcharhinus tilstoni). These loci plus 18 others developed for sharks from the genera Negaprion, Ginglymostoma, Carcharodon and Isurus were tested for amplification success on four species of Carcharhinus (including C. sorrah and C. tilstoni) and four other species representing three diverse families. Cross‐amplification was most common within families. Five loci were subsequently tested for polymorphism on 50 C. sorrah and 60 C. tilstoni. The number of alleles per locus was two to 24 and the average heterozygosity was 0.54 (range 0.16–0.87) for C. sorrah and 0.64 (range 0.44–0.78) for C. tilstoni. These loci may be useful tools for genetic analyses of the Carcharhinidae.     

Size,sex and seasonal patterns in the assemblage of Carcharhiniformes in a sub‐tropical bay

Size, sex and seasonal patterns among Carcharhiniformes were examined in shallow regions of Moreton Bay, Queensland, Australia. A total of 1259 sharks were caught, comprising 13 species. The Australian sharpnose shark Rhizoprionodon taylori and the blacktip complex Carcharhinus limbatus–Carcharhinus tilstoni comprised 55% of all shark individuals. Neonates were observed for five species including the dusky shark Carcharhinus obscurus, which contrary to previous reports was relatively abundant in shallow, predominantly estuarine waters. Three contrasting patterns of occurrence were observed: smaller species were abundant and present throughout much of their ontogeny, larger species were mainly caught as neonates or juveniles and vagrant species were only caught during the warmer months. The shark assemblage differed significantly among seasons. While many species were observed during the warmer months, species diversity was lower in winter when C. obscurus comprised 43% of the catch. Overall, the results indicated that spatial and temporal distribution patterns were not synchronous for all species. The capture of small numbers of neonate C. obscurus in late autumn and winter demonstrates that parturition among Carcharhiniformes is not confined to spring and summer in sub‐tropical waters.     

Shark attacks in Dakar and the Cap Vert Peninsula, Senegal: low incidence despite high occurrence of potentially dangerous species

Background

The International Shark Attack File mentions only four unprovoked shark attacks on the coast of West Africa during the period 1828–2004, an area where high concentrations of sharks and 17 species potentially dangerous to man have been observed. To investigate if the frequency of shark attacks could be really low and not just under-reported and whether there are potentially sharks that might attack in the area, a study was carried out in Dakar and the Cap Vert peninsula, Senegal.

Methodology/Principal Findings

Personnel of health facilities, administrative services, traditional authorities and groups of fishermen from the region of Dakar were interviewed about the occurrence of shark attacks, and visual censuses were conducted along the coastline to investigate shark communities associated with the coasts of Dakar and the Cap Vert peninsula. Six attacks were documented for the period 1947–2005, including two fatal ones attributed to the tiger shark Galeocerdo cuvieri. All attacks concerned fishermen and only one occurred after 1970. Sharks were observed year round along the coastline in waters 3–15 m depth. Two species potentially dangerous for man, the nurse shark Ginglymostoma cirratum and the blacktip shark Carcharhinus limbatus, represented together 94% of 1,071 sharks enumerated during 1,459 hours of observations. Threatening behaviour from sharks was noted in 12 encounters (1.1%), including 8 encounters with C. limbatus, one with Galeocerdo cuvieri and 3 with unidentified sharks.

Conclusions/Significance

These findings suggest that the frequency of shark attacks on the coast of West Africa is underestimated. However, they also indicate that the risk is very low despite the abundance of sharks. In Dakar area, most encounters along the coastline with potentially dangerous species do not result in an attack. Compared to other causes of water related deaths, the incidence of shark attack appears negligible, at least one thousand fold lower.     

Distribution and phylogenetic relationships of Australian glow-worms Arachnocampa (Diptera, Keroplatidae)

Glow-worms are bioluminescent fly larvae (Order Diptera, genus Arachnocampa) found only in Australia and New Zealand. Their core habitat is rainforest gullies and wet caves. Eight species are present in Australia; five of them have been recently described. The geographic distribution of species in Australia encompasses the montane regions of the eastern Australian coastline from the Wet Tropics region of northern Queensland to the cool temperate and montane rainforests of southern Australia and Tasmania. Phylogenetic trees based upon partial sequences of the mitochondrial genes cytochrome oxidase II and 16S mtDNA show that populations tend to be clustered into allopatric geographic groups showing overall concordance with the known species distributions. The deepest division is between the cool-adapted southern subgenus, Lucifera, and the more widespread subgenus, Campara. Lucifera comprises the sister groups, A. tasmaniensis, from Tasmania and the newly described species, A. buffaloensis, found in a high-altitude cave at Mt Buffalo in the Australian Alps in Victoria. The remaining Australian glow-worms in subgenus Campara are distributed in a swathe of geographic clusters that extend from the Wet Tropics in northern Queensland to the temperate forests of southern Victoria. Samples from caves and rainforests within any one geographic location tended to cluster together within a clade. We suggest that the morphological differences between hypogean (cave) and epigean (surface) glow-worm larvae are facultative adaptations to local microclimatic conditions rather than due to the presence of cryptic species in caves.     

Adult blacktip sharks (Carcharhinus limbatus) use shallow water as a refuge from great hammerheads (Sphyrna mokarran)

A refuge can be any space that keeps an organism safe from danger. Prey usually seek protection in the closest refuge available to minimize cost while maximizing survival. Aerial drone footage of blacktip sharks, Carcharhinus limbatus, along the coast of southeast Florida, USA, shows adult blacktips fleeing to the shallow water adjacent to the beach when confronted with or chased by a predatory great hammerhead shark, Sphyrna mokarran. To authors’ knowledge, this is the first evidence of adult C. limbatus using shallow waters as a refuge.     

Feeding ecology and trophic comparisons of six shark species in a coastal ecosystem off southern Brazil

The diets of six shark species, Sphyrna lewini, Sphyrna zygaena, Carcharhinus obscurus, Carcharhinus limbatus, Rhizoprionodon lalandii and Galeocerdo cuvier, were investigated in a subtropical coastal ecosystem of southern Brazil. Stomach content data were obtained to assess foraging niche segregation and ontogenetic shifts in the diets of these sharks. Five of the shark species off the Paraná coast were ichthyophagous, with the exception of S. zygaena, which was teutophagous. With the exception of G. cuvier, which had a generalist diet, the other five species displayed specialization in their feeding. Ontogenetic shifts were observed in C. obscurus and S. lewini with large individuals consuming elasmobranchs. Owing to the diet overlap between C. obscurus and S. lewini, C. obscurus and C. limbatus and R. lalandii and C. limbatus, future studies on the spatial and temporal distributions of these species are needed to understand the extent of competitive interactions.     

Quantification of Massive Seasonal Aggregations of Blacktip Sharks (Carcharhinus limbatus) in Southeast Florida

Southeast Florida witnesses an enormous seasonal influx of upper trophic level marine predators each year as massive aggregations of migrating blacktip sharks (Carcharhinus limbatus) overwinter in nearshore waters. The narrow shelf and close proximity of the Gulf Stream current to the Palm Beach County shoreline drive tens of thousands of sharks to the shallow, coastal environment. This natural bottleneck provides a unique opportunity to estimate relative abundance. Over a four year period from 2011–2014, an aerial survey was flown approximately biweekly along the length of Palm Beach County. A high definition video camera and digital still camera mounted out of the airplane window provided a continuous record of the belt transect which extended 200 m seaward from the shoreline between Boca Raton Inlet and Jupiter Inlet. The number of sharks within the survey transect was directly counted from the video. Shark abundance peaked in the winter (January-March) with a maximum in 2011 of 12,128 individuals counted within the 75.6 km^-2 belt transect. This resulted in a maximum density of 803.2 sharks km^-2. By the late spring (April-May), shark abundance had sharply declined to 1.1% of its peak, where it remained until spiking again in January of the following year. Shark abundance was inversely correlated with water temperature and large numbers of sharks were found only when water temperatures were less than 25°C. Shark abundance was also correlated with day of the year but not with barometric pressure. Although shark abundance was not correlated with photoperiod, the departure of the sharks from southeast Florida occurred around the vernal equinox. The shark migration along the United States eastern seaboard corresponds spatially and temporally with the spawning aggregations of various baitfish species. These baseline abundance data can be compared to future studies to determine if shark population size is changing and if sharks are restricting their southward migration as global water temperatures increase.     

Utilisation of a tropical bay as a nursery area by sharks of the families Carcharhinidae and Sphyrnidae

Synopsis At least eight species of sharks of the families Carcharhinidae and Sphyrnidae use Cleveland Bay in northern Australia as a communal nursery area.Carcharhinus dussumieri, C. fitzroyensis, C. limbatus andC. tilstoni use the bay as a seasonal primary nursery, with juveniles occurring in it for only a few months each year immediately after birth. Alternatively,Carcharhinus sorrah, Rhizoprionodon acutus andR. taylori use the bay as a year-round primary and secondary nursery, with juveniles remaining in it up to the size at maturity. AdultR. taylori also persist in the bay, a behavioural pattern possibly explained by their small maximum size. While present immediately after birth the type of utilisation pattern displayed bySphyrna lewini could not be clarified in this study. Although diets of these species in the bay are similar, there is probably little direct competition for food due to the highly productive habitats in the bay supporting an abundance of food resources. The highest numbers of juveniles occur when prey species are the most abundant, and when temporal separation of some seasonally-occurring species of sharks in effect.     

Ecological Drivers of Shark Distributions along a Tropical Coastline

As coastal species experience increasing anthropogenic pressures there is a growing need to characterise the ecological drivers of their abundance and habitat use, and understand how they may respond to changes in their environment. Accordingly, fishery-independent surveys were undertaken to investigate shark abundance along approximately 400 km of the tropical east coast of Australia. Generalised linear models were used to identify ecological drivers of the abundance of immature blacktip Carcharhinus tilstoni/Carcharhinus limbatus, pigeye Carcharhinus amboinensis, and scalloped hammerhead Sphyrna lewini sharks. Results indicated general and species-specific patterns in abundance that were characterised by a range of abiotic and biotic variables. Relationships with turbidity and salinity were similar across multiple species, highlighting the importance of these variables in the functioning of communal shark nurseries. In particular, turbid environments were especially important for all species at typical oceanic salinities. Mangrove proximity, depth, and water temperature were also important; however, their influence varied between species. Ecological drivers may promote spatial diversity in habitat use along environmentally heterogeneous coastlines and may therefore have important implications for population resilience.     

Resource-use dynamics of co-occurring chondrichthyans from the First Coast,North Florida,USA

Recent studies on shark assemblages on the northeast Florida and southeast Georgia coast (hereafter referred to collectively as the “First Coast”) have demonstrated differences in species and age-class composition of catch from previously characterized estuaries and newly surveyed area beaches, demonstrating that these regions may provide a critical habitat to different segments (i.e., life stages) of local shark populations. In this study, carbon and nitrogen stable isotopes (δ¹³C and δ¹⁵N) from muscle tissue and blood plasma were used to examine trophic dynamics (and temporal variability thereof) of the three dominant co-occurring species found along First Coast beaches (the Atlantic Sharpnose shark Rhizoprionodon terraenovae, Blacknose shark Carcharhinus acronotus and Blacktip shark Carcharhinus limbatus) to determine if they exhibit overlap in resource use along with spatial and temporal habitat use. Although considered spatially segregated from the beach species, a dominant, age-class species found in First Coast estuaries (juvenile Sandbar sharks Carcharhinus plumbeus) was also included in this analysis for comparison. Temporal variability of resource-use characteristics was detected at the species level. Resource-use overlap among species varied by tissue type and was generally higher for blood plasma, suggesting greater resource sharing over more recent time periods. Over longer time periods Atlantic Sharpnose and Blacktip sharks exhibited resource-use expansion, whereas Blacknose sharks exhibited a narrowing in resource use, suggesting a more specialized foraging strategy compared to the other species. The resource-use breadth of Sandbar sharks also expanded between blood plasma and muscle tissue. Significant size relationships were detected in Blacktip and Sandbar sharks, indicating ontogenetic resource shifts for both species. A diversity of highly productive resource pools likely support shark populations along the First Coast such that resource-use differentiation is not required to facilitate species co-occurrence. This work may shed light on understanding patterns of species co-occurrence as well as aid in future conservation efforts.     

Rapid and Simultaneous Identification of Body Parts from the Morphologically Similar Sharks Carcharhinus obscurus and Carcharhinus plumbeus (Carcharhinidae) Using Multiplex PCR

Many commercially exploited carcharhinid sharks are difficult to identify to species owing to extensive morphological similarities. This problem is severely exacerbated when it comes to identifying detached shark fins, and the finless and headless shark carasses typically sold in markets. To assist in the acquisition of urgently needed conservation and management data on shark catch and trade, we have developed a highly streamlined approach based on multiplex polymerase chain reaction (PCR) that uses species-specific primers derived from nuclear ribosomal ITS2 sequences to achieve rapid species identification of shark body parts. Here we demonstrate the utility of this approach for identifying fins and flesh from two globally distributed, morphologically very similar carcharhinid sharks (Carcharhinus obscurus and Carcharhinus plumbeus) intensively targeted in fisheries worldwide, and often confused for each other even as whole animals. The assay is conducted in a 4-primer multiplex format that is structured to simultaneously achieve the following efficiency and cost-reduction objectives: it requires only a single-tube amplification reaction for species diagnosis, it incorporates an internal positive control to allow detection of false-negative results, and it is novel in that it allows species identification even when DNAs from two species are combined in the same tube during the PCR reaction. The latter innovation reduces the required effort for screening a set of unknown samples by 50%. The streamlined approach illustrated here should be amenable for use in a shark conservation and management context where large numbers of samples typically need to be screened; the approach shown may also provide a model for a rapid diagnostic method applicable to species identification in general. Received September 15, 2000; accepted December 15, 2000     

New insights on the trophic ecology of blue (Prionace glauca) and shortfin mako sharks (Isurus oxyrinchus) from the oceanic eastern South Pacific

The blue shark (Prionace glauca) and the shortfin mako shark (Isurus oxyrinchus) are two large and highly migratory sharks distributed in most oceans. Although they are often caught in the south Pacific Ocean long-line fisheries, their trophic ecology is poorly understood. Stable isotopes with Bayesian mixing and dependence concentration models were performed to determine the diet and trophic differences between the two species in the South-eastern Pacific Ocean. According to the mixing models, fishes are the most important prey of these sharks. Dolphin calves and remains were found in the stomachs of both species, which represents a novel finding in trophic ecology of South Pacific sharks. Intra-specific differences were found in P. glauca, but not in specimens of I. oxyrinchus. The two sharks showed a high degree of diet overlap (73%), primarily over mackerel and dolphin carcasses. Our results indicate that blue and shortfin mako sharks have a generalist feeding strategy in the eastern Pacific Ocean, with a strong preference for teleost fishes and also for dolphin carcasses. Therefore, trophic studies are useful to understand energy flow through the food web, and the trophic position of key species.     

Evoked Potential Audiograms of the Nurse Shark (Ginglymostoma cirratum) and the Yellow Stingray (Urobatis jamaicensis)

The hearing thresholds of the nurse shark, Ginglymostoma cirratum, and the yellow stingray, Urobatis jamaicensis, were measured using auditory evoked potentials (AEP). Stimuli were calibrated using a pressure-velocity probe so that the acoustic field could be completely characterized. The results show similar hearing thresholds for both species and similar hearing thresholds to previously measured audiograms for the lemon shark, Negaprion brevirostris, and the horn shark, Heterodontis francisi. All of these audiograms suggest poor hearing abilities, raising questions about field studies showing attraction of sharks to acoustic signals. By extrapolating the particle acceleration thresholds into estimates of their equivalent far-field sound pressure levels, it appears that these sharks cannot likely detect most of the sounds that have attracted sharks in the field.     

Potential expansion in the spatial distribution of subtropical and temperate west Australian sharks

Fishery-dependent and -independent data collected since 1975 were examined to explore the spatial distribution of 30 shark and ray species in the west coast of Australia. Bigeye sixgill (Hexanchus nakamurai), tiger (Galeocerdo cuvier) and spinner (Carcarhinus brevipinna) sharks, and scalloped hammerhead (Sphyrna lewini) were observed >1000 and 300 km to the east of the edge of their reported distributions. Broadnose sevengill sharks (Notorhyncus cepedianus) and southern sawsharks (Pristiophorus nudipinnis) were observed >1000 km to the west of the edge of their reported distributions. Our study highlights the value of collecting and examining long time-series of data for understanding the spatial distribution of large marine predators.     

Relationships of morphological groups in the northern flicker superspecies complex (Colaptes auratus & C. chrysoides)

Many species complexes have diversified rapidly and recently, resulting in morphologically diverse populations; however, the rapid pace of diversification often prevents identification of clear phylogeographic structure. Recently, the use of large genomic and reduced-representation genomic datasets has improved resolution of the evolutionary histories in such species and allowed identification of lineages on distinct evolutionary trajectories. The northern flicker (Colaptes auratus) and gilded flicker (Colaptes chrysoides) form a polytypic superspecies group with a complex taxonomic history. The superspecies group includes up to 13 described subspecies, which represent slight geographic variation among five main morphological groups: red-shafted flickers of western North America (cafer group), yellow-shafted flickers of eastern North America (auratus group), Cuban flickers of the Caribbean (chrysocaulosus group), gilded flickers of the U.S. south-west and Mexican north-west (chrysoides group), and Guatemalan flickers of Central America (mexicanoides group). These groups are largely differentiable by variation in feather shaft colour, malar colour, throat colour, crown colour, and back barring. Here, using mitochondrial DNA (mtDNA) and hundreds of single nucleotide polymorphisms (SNPs), we characterized the genetic relationships and genomic distinctiveness of the five morphological groups. We found the mexicanoides group to be the most genetically distinct in both mtDNA (～1.4% sequence divergence) and large SNP panels. The chrysocaulosus group is differentiated by a single basepair mutation in a small mtDNA fragment. In both mtDNA and SNP panels, there is little genetic distinctiveness between auratus, cafer, and chrysoides morphological groups, with evidence of admixture and a lack of fixed differences.     

Recreational fishing impacts on threatened river sharks: A potential conservation issue

The Adelaide River in Australia's Northern Territory is a popular recreational fishing area, as well as habitat for threatened and protected river sharks (Glyphis species). Both the Critically Endangered Speartooth Shark (Glyphis glyphis) and Endangered Northern River Shark (Glyphis garricki) are identified here in illegal catches from recreational angling. The identification of a decayed shark specimen using a DNA barcoding‐like approach is the first such application to the identification of protected sharks in a recreational fishery. While the extent of catches by recreational anglers is unknown, the threatened status of these sharks, their suspected low population sizes, restricted distributions and importance of the Adelaide River as a nursery area call for the consideration of this as a potential conservation issue. As such, appropriate measures should be taken to reduce interactions with recreational anglers. The primary target species in the river is the iconic sportfish, Barramundi, which is predominantly caught by unbaited lure. Sharks are rarely caught on lure, allowing an opportunity for mitigation to focus on a fishing activity (baited hooks) which would limit any regulatory impact on popular lure fishing. Potential mitigation measures range from increased angler education and compliance checks, to the implementation of a spatial closure to baited hook fishing (a lure‐only zone). Such measures may assist in meeting a stated objective of the Australian Government's river shark Recovery Plan to ‘reduce and, where possible, eliminate adverse impacts of recreational fishing'.     

Oceanic dispersal in a sedentary reef shark (Triaenodon obesus): genetic evidence for extensive connectivity without a pelagic larval stage

Aim Most reef fishes are site‐attached, but can maintain a broad distribution through their highly dispersive larval stage. The whitetip reef shark (Triaenodon obesus) is site‐attached, yet maintains the largest Indo‐Pacific distribution of any reef shark while lacking the larval stage of bony (teleost) fishes. Here we use mitochondrial DNA (mtDNA) sequence data to evaluate the enigma of the sedentary reef shark that maintains a distribution across two‐thirds of the planet. Location Tropical Pacific and Indian Oceans. Methods We analysed 1025 base pairs of the mtDNA control region in 310 individuals from 25 locations across the Indian and Pacific Oceans. Phylogeographic and population genetic analyses were used to reveal the dispersal and recent evolutionary history of the species. Results We resolved 15 mtDNA control region haplotypes, but two comprised 87% of the specimens and were detected at nearly every location. Similar to other sharks, genetic diversity was low (h = 0.550 ± 0.0254 and π = 0.00213 ± 0.00131). Spatial analyses of genetic variation demonstrated strong isolation across the Indo‐Pacific Barrier and between western and central Pacific locations. Pairwise Φ_ST comparisons indicated high connectivity among archipelagos of the central Pacific but isolation across short distances of contiguous habitat (Great Barrier Reef) and intermittent habitat (Hawaiian Archipelago). In the eastern Pacific only a single haplotype (the most common one in the central Pacific) was observed, indicating recent dispersal (or colonization) across the East Pacific Barrier. Main conclusions The shallow haplotype network indicates recent expansion of modern populations within the last half million years from a common ancestor. Based on the distribution of mtDNA diversity, this began with an Indo‐West Pacific centre of origin, with subsequent dispersal to the Central Pacific and East Pacific. Genetic differences between Indian and Pacific Ocean populations are consistent with Pleistocene closures of the Indo‐Pacific Barrier associated with glacial cycles. Pairwise population comparisons reveal weak but significant isolation by distance, and notably do not indicate the high coastal connectivity observed in other shark species. The finding of population structure among semi‐contiguous habitats, but population connectivity among archipelagos, may indicate a previously unsuspected oceanic dispersal behaviour in whitetip reef sharks.     

Navigating the mtDNA road map out of the morphological maze: interpreting morphological variation in the diverse Monomorium rothsteini (Forel) complex (Hymenoptera: Formicidae)

Monomorium is a large and diverse ant genus with speciose radiations in both the Afrotropical and Australian regions. According to the most recent taxonomic revision, many Australian species are characterised by very broad distributions and variable morphology, which suggests that some species may be unrecognised species complexes. With a continent‐wide distribution and diverse yet overlapping morphology, M. rothsteini (Forel) is representative of the greater challenge that exists in Australian Monomorium systematics. Here we investigate species boundaries in M. rothsteini using a molecular phylogenetic framework to interpret the complex overlap of nine morphological characters (with 31 states) and examine biogeographic relationships among the lineages. Bayesian inference resolved 38 mtDNA lineages that were morphologically separable, at least from their sister lineage. Although the morphological characters were intermixed across the phylogeny, instances of inseparable morphology among sister clades was rare. Seventeen lineages exhibited complete morphological overlap with one or more other lineages and could not be separated by Principal Component Analysis based on 12 morphometric variables. Two‐thirds of all lineages occurred sympatrically with one or more both genetically and morphologically divergent lineages. The two nuclear markers EF1αF2 and wingless were used to generate haplotype networks which were characterised by a star‐like pattern indicative of a rapid and recent radiation. Several haplotypes for both nuclear gene regions were shared among individuals occurring in separate mtDNA clades which we were also unable to distinguish morphologically or that were occurring in sympatry, indicating possible introgression in both the mtDNA and nuclear genomes. Clear biogeographic affinities among samples within a lineage were detected but there was no overall pattern in the biogeographic relationships among the lineages. We conclude that M. rothsteini is a large species complex that has undergone a complex evolutionary history following aridification of the Australian continent, and discuss the implications of this conclusion for the systematics of Australian Monomorium more generally.