Detecting Sex-Related Changes to the Metabolome of a Critically Endangered Freshwater Crayfish During the Mating Season.

Captive breeding is a vital tool in the conservation of highly endangered species, as it is for the Margaret River hairy marron, Cherax tenuimanus, from the south west of Australia. A close relative, Cherax cainii, has almost completely displaced C. tenuimanus in the wild and is a successful aquaculture species, whereas C. tenuimanus has performed poorly in captivity. We used untargeted liquid chromatography-mass spectrometry to obtain metabolomic profiles of female and male C. tenuimanus held in controlled aquarium conditions during their reproductive period. Using repeated haemolymph sampling we tracked the metabolomic profiles of animals just prior to and for a period of up to 34 days after pairing with a similar sized potential mate. We identified 54 reproducible annotated metabolites including amino acids, fatty acids, biogenic amines, purine and pyrimidine metabolites and excretion metabolites. Hierarchical clustering analysis distinguished five metabolite clusters. Principal component-canonical variate analysis clearly distinguished females from males, both unpaired and paired; similar trends in profile changes in both sexes after pairing; and a striking shift in males upon pairing. We discuss three main patterns of metabolomic responses: differentiation between sexes; reactive responses to the disturbance of pairing; and convergent response to the disturbance of pairing for males. Females generally had higher concentrations of metabolites involved in metabolic rate, mobilisation of energy stores and stress. Responses to the disturbance of pairing were also related to elevated stress. Females were mobilising lipid stores to deposit yolk, whereas males had a rapid and strong response to pairing, with shifts in metabolites associated with gonad development and communication, indicating males could complete reproductive readiness only once paired with a female. The metabolomic profiles support a previously proposed potential mechanism for displacement of C. tenuimanus by C. cainii in the wild and identify several biomarkers for testing hypotheses regarding reproductive success using targeted metabolomics.

Mus musculus populations in Western Australia lack VKORC1 mutations conferring resistance to first generation anticoagulant rodenticides: Implications for conservation and biosecurity.

BACKGROUND: Humans routinely attempt to manage pest rodent populations with anticoagulant rodenticides (ARs). We require information on resistance to ARs within rodent populations to have effective eradication programs that minimise exposure in non-target species. Mutations to the VKORC1 gene have been shown to confer resistance in rodents with high proportions of resistance in mice found in all European populations tested. We screened mutations in Mus musculus within Western Australia, by sampling populations from the capital city (Perth) and a remote island (Browse Island). These are the first Australian mouse populations screened for resistance using this method. Additionally, the mitochondrial D-loop of house mice was sequenced to explore population genetic structure, identify the origin of Western Australian mice, and to elucidate whether resistance was linked to certain haplotypes. RESULTS: No resistance-related VKORC1 mutations were detected in either house mouse population. A genetic introgression in the intronic sequence of the VKORC1 gene of Browse Island house mouse was detected which is thought to have originated through hybridisation with the Algerian mouse (Mus spretus). Analysis of the mitochondrial D-loop reported two haplotypes in the house mouse population of Perth, and two haplotypes in the population of Browse Island. CONCLUSIONS: Both house mouse populations exhibited no genetic resistance to ARs, in spite of free use of ARs in Western Australia. Therefore weaker anticoagulant rodenticides can be employed in pest control and eradication attempts, which will result in reduced negative impacts on non-target species. Biosecurity measures must be in place to avoid introduction of resistant house mice, and new house mouse subspecies to Western Australia.

More evolution underground: Accelerated mitochondrial substitution rate in Australian burrowing freshwater crayfishes (Decapoda: Parastacidae).

To further understand the evolutionary history and mitogenomic features of Australia's highly distinctive freshwater crayfish fauna, we utilized a recently described rapid mitogenome sequencing pipeline to generate 24 new crayfish mitogenomes including a diversity of burrowing crayfish species and the first for Astacopsis gouldi, the world's largest freshwater invertebrate. Whole mitogenome-based phylogeny estimates using both Bayesian and Maximum Likelihood methods substantially strengthen existing hypotheses for systematic relationships among Australian freshwater crayfish with evidence of pervasive diversifying selection and accelerated mitochondrial substitution rate among the members of the clade representing strongly burrowing crayfish that may reflect selection pressures for increased energy requirement for adaptation to terrestrial environment and a burrowing lifestyle. Further, gene rearrangements are prevalent in the burrowing crayfish mitogenomes involving both tRNA and protein coding genes. In addition, duplicated control regions were observed in two closely related Engaeus species, together with evidence for concerted evolution. This study significantly adds to the understanding of Australian freshwater crayfish evolutionary relationships and suggests a link between mitogenome evolution and adaptation to terrestrial environments and a burrowing lifestyle in freshwater crayfish.

Multiple drivers of decline in the global status of freshwater crayfish (Decapoda: Astacidea).

Rates of biodiversity loss are higher in freshwater ecosystems than in most terrestrial or marine ecosystems, making freshwater conservation a priority. However, prioritization methods are impeded by insufficient knowledge on the distribution and conservation status of freshwater taxa, particularly invertebrates. We evaluated the extinction risk of the world's 590 freshwater crayfish species using the IUCN Categories and Criteria and found 32% of all species are threatened with extinction. The level of extinction risk differed between families, with proportionally more threatened species in the Parastacidae and Astacidae than in the Cambaridae. Four described species were Extinct and 21% were assessed as Data Deficient. There was geographical variation in the dominant threats affecting the main centres of crayfish diversity. The majority of threatened US and Mexican species face threats associated with urban development, pollution, damming and water management. Conversely, the majority of Australian threatened species are affected by climate change, harvesting, agriculture and invasive species. Only a small proportion of crayfish are found within the boundaries of protected areas, suggesting that alternative means of long-term protection will be required. Our study highlights many of the significant challenges yet to come for freshwater biodiversity unless conservation planning shifts from a reactive to proactive approach.