William F. Humphreys

Evolution of subterranean diving beetles (Coleoptera: Dytiscidae: Hydroporini, Bidessini) in the arid zone of Australia.

Abstract Calcrete aquifers in arid inland Australia have recently been found to contain the worlds most diverse assemblage of subterranean diving beetles (Coleoptera: Dytiscidae). In this study we test whether the adaptive shift hypothesis (ASH) or the climatic relict hypothesis (CRH) is the most likely mode of evolution for the Australian subterranean diving beetles by using a phylogeny based on two sequenced fragments of mitochondrial genes (CO1 and 16S‐tRNA‐ND1) and linearized using a relaxed molecular clock method. Most individual calcrete aquifers contain an assemblage of diving beetle species of distantly related lineages and/or a single pair of sister species that significantly differ in size and morphology. Evolutionary transitions from surface to subterranean life took place in a relatively small time frame between nine and four million years ago. Most of the variation in divergence times of the sympatric sister species is explained by the variation in latitude of the localities, which correlates with the onset of aridity from the north to the south and with an aridity maximum in the Early Pliocene (five mya). We conclude that individual calcrete aquifers were colonized by several distantly related diving beetle lineages. Several lines of evidence from molecular clock analyses support the CRH, indicating that all evolutionary transitions took place during the Late Miocene and Early Pliocene as a result of aridification.

Cryptic speciation in two widespread subterranean amphipod genera reflects historical drainage patterns in an ancient landscape

The landscape of the Pilbara region of Western Australia has been relatively unchanged for 100 million years. The ancient river systems of this region might be expected to be sources of isolation and divergence for aquatic species. Hence, the occurrence of widespread groundwater taxa in this landscape offers the opportunity to examine associations between genetic diversity and drainage patterns. Pilbarus and Chydaekata are two widespread genera of subterranean amphipods endemic to the Pilbara, each occupying multiple tributaries. We used molecular data to examine the roles of drainage patterns in structuring genetic diversity. Gene flow within a tributary may be facilitated by the occasional occurrence of these amphipods in springs, which results in their downstream dispersal during episodic flooding. However, tributary boundaries may form hydrological barriers to gene flow, resulting in localised isolation of populations and divergence. Samples of both genera, collected throughout three river basins, were examined for sequence divergence in the cytochrome c oxidase I mitochondrial gene. There was no evidence of contemporary gene flow among populations of either genus, and each tributary contained highly divergent lineages, which were not associated with similar morphological differentiation. This suggests cryptic speciation has occurred, and similar phylogenetic signals in both taxa imply similar evolutionary histories. Surface populations may have been driven into subterranean refugia by the cessation of flow in the rivers, associated with Tertiary climate change, while morphological evolution may have been constrained by stabilising selection. The lack of congruence between molecular diversity and morphology raises important practical issues for conservation.

Subterranean archipelago in the Australian arid zone: mitochondrial DNA phylogeography of amphipods from central Western Australia

In 1998, a unique subterranean ecosystem was discovered in numerous isolated calcrete (carbonate) aquifers in the arid Yilgarn region of Western Australia. Previous morphological and genetic analyses of a subterranean water beetle fauna suggest that calcrete aquifers are equivalent to closed island habitats that have been isolated for millions of years. We tested this hypothesis further by phylogeographic analyses of subterranean amphipods (Crangonyctoidea: Paramelitidae and Hyalidae) using mitochondrial DNA sequence data derived from the cytochrome oxidase I gene. Phylogenetic analyses and population genetic analyses (samova) provided strong evidence for the existence of at least 16 crangonyctoid and six hyalid divergent mitochondrial lineages, each restricted in their distribution to a single calcrete aquifer, in support of the ‘subterranean island (archipelago) hypothesis’ and extending its scope to include entirely water respiring invertebrates. Sequence divergence estimates between proximate calcrete populations suggest that calcretes have been isolated at least since the Pliocene, coinciding with a major aridity phase that led to the intermittent drying of surface water. The distribution of calcretes along palaeodrainage channels and on either side of drainage divides, have had less influence on the overall phylogeographic structure of populations, with evidence that ancestral crangonyctoid and hyalid species moved between catchments multiple times prior to their isolation within calcretes. At least two potential modes of evolution may account for the diversity of subterranean amphipod populations: dispersal/vicariance of stygobitic species or colonization of calcretes by surface species and independent evolution of stygobitic characteristics.

Rising from Down Under: developments in subterranean biodiversity in Australia from a groundwater fauna perspective

Over the last two decades, Australia has undergone a renaissance in studies of subterranean biology. This paper sets these recent developments into context from the perspective of groundwater fauna. Owing to its obligate subterranean life, typical local endemicity and the geological persistence of subterranean habitats, stygofauna is an excellent subject for biogeographic study. Groundwater containing diverse faunas range from freshwater to marine salinities in both coastal and continental locations. They occur in typical karst, alluvial, and fractured rock aquifers, but also in novel matrices formed during the hydrogeochemical evolution of groundwater (goethite pisolites and groundwater calcretes) in the Tertiary. This range of habitats, water quality and the diverse origins of the fauna (Gondwanan, Pangaean and Tethys) support a phylogenetically highly diverse fauna. Several taxa, notably among the Podocopida, Bathynellacea, Amphipoda, and Dytiscidae show remarkable species diversity. Typically there is fine spatial scale endemicity of species associated with local aquifers, but there are inexplicable regional differences, such as the change of fauna between the Yilgarn and Pilbara, contiguous areas on the long emergent Western Shield. The anchialine taxa representing higher taxa are highly disjunct from their congeners in the North Atlantic. The emerging species richness, the fine scale patchwork of endemicity, and the distinct regional differences, respectively, contribute to a substantial increase in α, β and γ diversity of the aquatic fauna, especially in arid Australia. This diversity is posing challenging issues for proponents and regulators of mineral development because much of this diversity has emerged in the two most mineraliferous provinces of Australia. The scientific capacity to respond is challenged by the sheer scale of the emerging issues.

Protecting the innocent: studying short-range endemic taxa enhances conservation outcomes

A major challenge confronting many contemporary systematists is how to integrate standard taxonomic research with conservation outcomes. With a biodiversity crisis looming and ongoing impediments to taxonomy, how can systematic research continue to document species and infer the ‘Tree of Life’, and still maintain its significance to conservation science and to protecting the very species it strives to understand? Here we advocate a systematic research program dedicated to documenting short-range endemic taxa, which are species with naturally small distributions and, by their very nature, most likely to be threatened by habitat loss, habitat degradation and climate change. This research can dovetail with the needs of industry and government to obtain high-quality data to inform the assessment of impacts of major development projects that affect landscapes and their biological heritage. We highlight how these projects are assessed using criteria mandated by Western Australian legislation and informed by guidance statements issued by the Environmental Protection Authority (Western Australia). To illustrate slightly different biological scenarios, we also provide three case studies from the Pilbara region of Western Australia, which include examples demonstrating a rapid rise in the collection and documentation of diverse and previously unknown subterranean and surface faunas, as well as how biological surveys can clarify the status of species thought to be rare or potentially threatened. We argue that ‘whole of biota’ surveys (that include all invertebrates) are rarely fundable and are logistically impossible, and that concentrated research on some of the most vulnerable elements in the landscape – short-range endemics, including troglofauna and stygofauna – can help to enhance conservation and research outcomes.

Subterranean archipelago: mitochondrial DNA phylogeography of stygobitic isopods (Oniscidea:Haloniscus ) from the Yilgarn region of Western Australia

The arid Yilgarn region of Western Australia contains numerous isolated calcrete aquifers, within which a diverse subterranean fauna has been discovered. Genetic and morphological studies of subterranean dytiscid beetles and amphipods have suggested that individual calcretes are equivalent to closed island habitats, which have been isolated for millions of years. Here we test this ‘subterranean island’ hypothesis further by phylogeographic analyses of subterranean oniscidean isopods (Haloniscus), using mitochondrial DNA (mtDNA) sequence data derived from the cytochrome c oxidase subunit I gene. Phylogenetic and population genetic analyses provided evidence for significant phylogeographic structuring of isopod populations, with evidence for at least 24 divergent mtDNA lineages, each restricted in their distribution to a single calcrete aquifer. The high level of divergence among calcrete populations (generally >25%) and several mtDNA lineages within calcretes (>16%) suggests that each lineage is likely to represent a distinct species. These analyses, together with comparative phylogeographic data from dytiscid beetles and amphipods, provide strong support for the ‘subterranean island’ hypothesis, applying to both air-breathing and fully aquatic arthropod groups. The finding of several epigean lineages that grouped with stygobitic Haloniscus populations, and the overall phylogeographic structure of populations, suggests that the majority of stygobitic species evolved within individual calcretes following independent colonisation by epigean ancestors.

Shrimps Down Under: Evolutionary Relationships of Subterranean Crustaceans from Western Australia (Decapoda: Atyidae: Stygiocaris )

Background We investigated the large and small scale evolutionary relationships of the endemic Western Australian subterranean shrimp genus Stygiocaris (Atyidae) using nuclear and mitochondrial genes. Stygiocaris is part of the unique cave biota of the coastal, anchialine, limestones of the Cape Range and Barrow Island, most of whose nearest evolutionary relations are found in coastal caves of the distant North Atlantic. The dominance of atyids in tropical waters and their food resources suggest they are pivotal in understanding these groundwater ecosystems. Methodology/Principle Findings Our nuclear and mitochondrial analyses all recovered the Mexican cave genus Typhlatya as the sister taxon of Stygiocaris, rather than any of the numerous surface and cave atyids from Australia or the Indo-Pacific region. The two described Stygiocaris species were recovered as monophyletic, and a third, cryptic, species was discovered at a single site, which has very different physiochemical properties from the sites hosting the two described species. Conclusions/Significance Our findings suggest that Stygiocaris and Typhlatya may descend from a common ancestor that lived in the coastal marine habitat of the ancient Tethys Sea, and were subsequently separated by plate tectonic movements. This vicariant process is commonly thought to explain the many disjunct anchialine faunas, but has rarely been demonstrated using phylogenetic techniques. The Cape Ranges geological dynamism, which is probably responsible for the speciation of the various Stygiocaris species, has also led to geographic population structure within species. In particular, Stygiocaris lancifera is split into northern and southern groups, which correspond to population splits within other sympatric subterranean taxa.

Phylogeography of the ancient Parabathynellidae (Crustacea : Bathynellacea) from the Yilgarn region of Western Australia

The crustacean order Bathynellacea is a primitive group of subterranean aquatic (stygobitic) invertebrates that typically inhabits freshwater interstitial spaces in alluvia. A striking diversity of species from the bathynellacean family Parabathynellidae have been found in the calcretes of the Yilgarn palaeodrainage system in Western Australia. Taxonomic studies show that most species are restricted in their distribution to a single calcrete, which is consistent with the findings of other phylogeographic studies of stygofauna. In this, the first molecular phylogenetic and phylogeographic study of interspecific relationships among parabathynellids, we aimed to explore the hypothesis that species are short-range endemics and restricted to single calcretes, and to investigate whether there were previously unidentified cryptic species. Analyses of sequence data based on a region of the mitochondrial (mt) DNA cytochrome c oxidase 1 gene showed the existence of divergent mtDNA lineages and species restricted in their distribution to a single calcrete, in support of the broader hypothesis that these calcretes are equivalent to closed island habitats comprising endemic taxa. Divergent mtDNA lineages were also observed to comprise four new and 12 recognised morphospecies. These results reflect the findings of previous studies of stygobitic arthropods (beetles, amphipods and isopods) from the Yilgarn region and reinforce the usefulness of using DNA-sequence data to investigate species boundaries and the presence of cryptic species.

DNA barcoding of stygofauna uncovers cryptic amphipod diversity in a calcrete aquifer in Western Australia's arid zone.

The arid Yilgarn region of Western Australia contains numerous subterranean calcrete aquifers with unique assemblages of obligate groundwater invertebrates (stygofauna). We aimed to establish a DNA barcoding framework for the macro‐invertebrates present in a single calcrete, as a basis for future assessment of biodiversity of the Yilgarn calcretes and for investigating food webs. Intense sampling of a bore field grid in the Sturt Meadows calcrete was undertaken to obtain representatives of the entire macro‐invertebrate ecosystem. A 623‐bp fragment of the mitochondrial cytochrome c oxidase 1 (COI) gene was used to provide DNA barcodes for stygobiont macro‐invertebrates plus terrestrial organisms that are found in the calcrete. Phylogenetic analyses revealed the existence of 12 divergent monophyletic groups of haplotypes. Subterranean amphipods (Chiltoniidae) showed three groups of COI haplotypes with sequence divergences between them of >11%. Allozyme analyses found a large number of fixed allelic differences between these three amphipod groups, indicating that there are three morphologically cryptic species within the Sturt Meadows calcrete. Unlike the sister triplet of dytiscid beetles present, the amphipods are not sister clades and are more closely related to other Yilgarn and non‐Yilgarn amphipods than to each other. Our results show that the aquifer contains at least 12 macro‐invertebrate species and DNA barcoding provides a useful means for discriminating species in this system.

Twenty-six New Dytiscidae (Coleoptera) of the Genera Limbodessus Guignot And Nirripirti Watts Humphreys, from Underground Waters in Australia

Summary Twenty-six new species of stygobitic Dytiscidae from inland Western Australia and Central Australia are described: Limbodessus atypicalis, L. barwidgeeensis, L. cooperi, L. exilis, L. leysi, L. gumwellensis, L. harleyi, L. macrohinkleri, L. millbilliensis, L micrommatoion, L. mirandaae, L. melitaensis, L. narryerensis, L. nambiensis, L. palmulaoides, L. phoebeae, L. raeae, L. surreptitius, L. usitatus, L. yandalensis, Nirripirti macrosturtensis, N. megamacrocephalus, N. mesosturtensis, N. microsturtensis, N. septum and N. tetrameres. This brings the total of stygobitic Dytiscidae described from Australia to 80 species in four genera. A key to the known species of Australian stygobitic Dytiscidae is given as well as notes on the physico/chemical properties of selected collecting sites. Geographically the study reinforced our working hypothesis that stygal Dytiscidae in Australia are confined to two discrete areas; the Ngalia Basin northwest of Alice Springs in Central Australia and the Yilgarn Craton in Central Western Australia. As in previous years (see Watts & Humphreys 2004) the stygofauna was found, together with a rich stygobitic fauna largely comprising crustaceans, in those portions of shallow aquifers that ran through areas of calcrete formation. The following new combinations are proposed: Limbodessus bialveus (Watts & Humphreys, 2003), (Tjirtudessus); Limbodessus cunyuensis (Watts & Humphreys, 2003), (Tjirtudessus); Limbodessus hillviewensis (Watts & Humphreys, 2004), (Tjirtudessus); Limbodessus jundeeensis (Watts & Humphreys, 2003), (Tjirtudessus); Limbodessus karalundiensis (Watts & Humphreys, 2003), (Tjirtudessus); Limbodessus macrotarsus (Watts & Humphreys, 2003), (Tjirtudessus); Limbodessus microocular (Watts & Humphreys, 2004), (Tjirtudessus); Limbodessus occidentalis (Watts & Humphreys, 2004), (Boongurrus); Limbodessus padburyensis (Watts & Humphreys, 2004), (Tjirtudessus); Limbodessus silus (Watts & Humphreys, 2003), (Tjirtudessus); Limbodessus sweetwatersensis (Watts & Humphreys, 2003), (Tjirtudessus); Limbodessus wilunaensis (Watts & Humphreys, 2003), (Tjirtudessus); Limbodessus wogarthaensis (Watts & Humphreys, 2004), (Tjirtudessus); and Limbodessus yuinmeryensis (Watts & Humphreys, 2003), (Tjirtudessus). This is the seventh paper in our series describing the stygobitic Dytiscidae of Australia (Watts & Humphreys 1999, 2000, 2001, 2003, 2004; Balke et al. 2004). In it we describe the new species found during fieldwork in Western Australia in the winters of 2003 and 2004.