Maria P. Ikonomopoulou

Spider venomics: implications for drug discovery

Over a period of more than 300 million years, spiders have evolved complex venoms containing an extraordinary array of toxins for prey capture and defense against predators. The major components of most spider venoms are small disulfide-bridged peptides that are highly stable and resistant to proteolytic degradation. Moreover, many of these peptides have high specificity and potency toward molecular targets of therapeutic importance. This unique combination of bioactivity and stability has made spider-venom peptides valuable both as pharmacological tools and as leads for drug development. This review describes recent advances in spider-venom-based drug discovery pipelines. We discuss spider-venom-derived peptides that are currently under investigation for treatment of a diverse range of pathologies including pain, stroke and cancer.

Trace element concentrations in nesting flatback turtles (Natator depressus) from Curtis Island, Queensland, Australia

The blood and eggs of the flatback turtle (Natator depressus) sampled when nesting at Curtis Island, Queensland, Australia. In the blood, zinc was present at the highest concentration of 151.15 ± 1.45 μg/L followed by copper (7.74 ± 0.09 μg/L). Lead was found only in some individuals. The measured trace elements in the blood were maternally transferred into the eggs. Other metals and metalloids detected in eggs were chromium, manganese, arsenic and selenium. Eggs showed a more complex trace element profile than blood, suggesting that they provided more representative tissues for determining maternal levels of trace element accumulation in N. depressus. Intra-clutch variation was over 15% for most of the studied trace elements suggesting one egg is not in sufficient to determine trace element accumulation within a clutch. Copper was the only element which was positively correlated with breeding age. Furthermore, no detectable levels of tin compound derivates were measured in N. depressus.

How the cobra got its flesh-eating venom: cytotoxicity as a defensive innovation and its co-evolution with hooding, aposematic marking, and spitting

The cytotoxicity of the venom of 25 species of Old World elapid snake was tested and compared with the morphological and behavioural adaptations of hooding and spitting. We determined that, contrary to previous assumptions, the venoms of spitting species are not consistently more cytotoxic than those of closely related non-spitting species. While this correlation between spitting and non-spitting was found among African cobras, it was not present among Asian cobras. On the other hand, a consistent positive correlation was observed between cytotoxicity and utilisation of the defensive hooding display that cobras are famous for. Hooding and spitting are widely regarded as defensive adaptations, but it has hitherto been uncertain whether cytotoxicity serves a defensive purpose or is somehow useful in prey subjugation. The results of this study suggest that cytotoxicity evolved primarily as a defensive innovation and that it has co-evolved twice alongside hooding behavior: once in the Hemachatus + Naja and again independently in the king cobras (Ophiophagus). There was a significant increase of cytotoxicity in the Asian Naja linked to the evolution of bold aposematic hood markings, reinforcing the link between hooding and the evolution of defensive cytotoxic venoms. In parallel, lineages with increased cytotoxicity but lacking bold hood patterns evolved aposematic markers in the form of high contrast body banding. The results also indicate that, secondary to the evolution of venom rich in cytotoxins, spitting has evolved three times independently: once within the African Naja, once within the Asian Naja, and once in the Hemachatus genus. The evolution of cytotoxic venom thus appears to facilitate the evolution of defensive spitting behaviour. In contrast, a secondary loss of cytotoxicity and reduction of the hood occurred in the water cobra Naja annulata, which possesses streamlined neurotoxic venom similar to that of other aquatic elapid snakes (e.g., hydrophiine sea snakes). The results of this study make an important contribution to our growing understanding of the selection pressures shaping the evolution of snake venom and its constituent toxins. The data also aid in elucidating the relationship between these selection pressures and the medical impact of human snakebite in the developing world, as cytotoxic cobras cause considerable morbidity including loss-of-function injuries that result in economic and social burdens in the tropics of Asia and sub-Saharan Africa.

Accumulation of trace metals in the embryos and hatchlings of Chelonia mydas from Peninsular Malaysia incubated at different temperatures

A variety of trace metals were measured in the egg contents of three clutches of Chelonia mydas collected from Kuala Terengganu state in Peninsular Malaysia. We quantified Mn, Cu, Zn, Se (essential trace metals) and As (anthropogenic pollutant) at several developmental stages obtained by incubating eggs at two different temperatures (27 °C and 31 °C). The incubation temperatures were chosen because they produce predominantly male or predominantly female hatchlings, respectively. The eggs were removed from the sand and washed before being placed in incubators, to ensure that the only possible source of the detected metals was maternal transfer. Other metals: Mo, Co, Ni, Cd, Sn, Sb, Hg, Tl and Pb (all non-essential metals) were detected at concentrations below the lower limit of quantitation (LLOQ). Trace metal concentrations, particularly [Zn], increased during development, other metals (Cu, As, Se and Cr) accumulated to a lesser degree than zinc but no significant differences were observed between the incubation temperatures at any stage of incubation. To date, only a few studies on trace metals in turtle embryos and hatchlings have been reported; this study will provide basic knowledge on the accumulation of trace metals during development at two different incubation temperatures.

Molecular basis of the remarkable species selectivity of an insecticidal sodium channel toxin from the African spider Augacephalus ezendami

The inexorable decline in the armament of registered chemical insecticides has stimulated research into environmentally-friendly alternatives. Insecticidal spider-venom peptides are promising candidates for bioinsecticide development but it is challenging to find peptides that are specific for targeted pests. In the present study, we isolated an insecticidal peptide (Ae1a) from venom of the African spider Augacephalus ezendami (family Theraphosidae). Injection of Ae1a into sheep blowflies (Lucilia cuprina) induced rapid but reversible paralysis. In striking contrast, Ae1a was lethal to closely related fruit flies (Drosophila melanogaster) but induced no adverse effects in the recalcitrant lepidopteran pest Helicoverpa armigera. Electrophysiological experiments revealed that Ae1a potently inhibits the voltage-gated sodium channel BgNaV1 from the German cockroach Blattella germanica by shifting the threshold for channel activation to more depolarized potentials. In contrast, Ae1a failed to significantly affect sodium currents in dorsal unpaired median neurons from the American cockroach Periplaneta americana. We show that Ae1a interacts with the domain II voltage sensor and that sensitivity to the toxin is conferred by natural sequence variations in the S1–S2 loop of domain II. The phyletic specificity of Ae1a provides crucial information for development of sodium channel insecticides that target key insect pests without harming beneficial species.

Changes in milk composition during lactation in the eastern barred bandicoot (Perameles gunnii) (Marsupialia: Peramelidae)

Perameles gunnii is a small to medium-sized omnivorous marsupial. We measured milk components from Week 4 until weaning at Week 8; these showed marked quantitative and qualitative changes. The milk produced in the early stages of lactation was dilute, ~28% solid (w/w). At four weeks carbohydrate, protein and lipids were also at low levels: 2.0 g (100 mL)–1, 4.5 g (100 mL)–1 and 3.1 g (100 mL)–1 respectively. At the mid-phase of lactation solids reached 36%, carbohydrate 5.5 g (100 mL)–1, protein 10.0 g (100 mL)–1 and lipids 8.5 g (100 mL)–1. The energy content of the milk at 4 weeks of lactation was 2.25 kJ mL–1 while at pouch vacancy it reached 8.86 kJ mL–1. At 8 weeks the pouch young was almost weaned and all the constituents but carbohydrates had increased significantly. We compared the milk components of the eastern barred bandicoot with those of other marsupial species and with the Wombaroo milk formula used by zookeepers and carers to hand-raise orphaned bandicoots. We found the milk formula to be deficient in both protein and lipid.

Hormone and Metabolite Profiles in Nesting Green and Flatback Turtles: Turtle Species with Different Life Histories

Herbivorous turtle, Chelonia mydas, inhabiting the south China Sea and breeding in Peninsular Malaysia, and Natator depressus, a carnivorous turtle inhabiting the Great Barrier Reef and breeding at Curtis Island in Queensland, Australia, differ both in diet and life history. Analysis of plasma metabolites levels and six sex steroid hormones during the peak of their nesting season in both species showed hormonal and metabolite variations. When compared with results from other studies progesterone levels were the highest whereas dihydrotestosterone was the plasma steroid hormone present at the lowest concentration in both C. mydas and N. depressus plasma. Interestingly, oestrone was observed at relatively high concentrations in comparison to oestradiol levels recorded in previous studies suggesting that it plays a significant role in nesting turtles. Also, hormonal correlations between the studied species indicate unique physiological interactions during nesting. Pearson correlation analysis showed that in N. depressus the time of oviposition was associated with elevations in both plasma corticosterone and oestrone levels. Therefore, we conclude that corticosterone and oestrone may influence nesting behaviour and physiology in N. depressus. To summarise, these two nesting turtle species can be distinguished based on the hormonal profile of oestrone, progesterone, and testosterone using discriminant analysis.

An Investigation of Organochlorine and Polychlorobiphenyl Concentrations in the Blood and Eggs of the Carnivorous Flatback Turtle, Natator depressus, from Queensland, Australia

Abstract Low concentrations of several pesticides including organochlorines and polychlorinated biphenyls were found in the blood and eggs of nesting female flatback turtles, Natator depressus, collected from Curtis Island in Queensland, Australia.

The Development of Endothermy during Pouch Life in the Eastern Barred Bandicoot (Perameles gunnii), a Marsupial

Marsupials are born ectothermic and gradually become endothermic during pouch occupancy. In order to study the timing of this transition, we measured the metabolic rates of eight pouch‐young eastern barred bandicoots (Perameles gunnii) at 25°C (the thermoneutral zone of adults), at 35°C (pouch temperature), and after injection with norepinephrine. From 5 to 7 wk of age, oxygen consumption (V̇o2) was higher at 35°C than at 25°C. At 8 wk, V̇o2 was significantly higher at 25°C than at 35°C. Norepinephrine administration at 5 and 6 wk had no effect on metabolic rate, but at 8 wk, near pouch vacation, it resulted in a significant increase in metabolic rate. Our results indicate that signs of thermoregulation for P. gunnii begin at 7 wk, and at 8 wk the species is endothermic, that is, able to increase heat production in response to both cold and norepinephrine. The appearance of fur, first seen at the age of 6 wk and fully developed at 8 wk, coincides with the onset of endothermy.

Gomesin peptides prevent proliferation and lead to the cell death of devil facial tumour disease cells

The Tasmanian devil faces extinction due to devil facial tumour disease (DFTD), a highly transmittable clonal form of cancer without available treatment. In this study, we report the cell-autonomous antiproliferative and cytotoxic activities exhibited by the spider peptide gomesin (AgGom) and gomesin-like homologue (HiGom) in DFTD cells. Mechanistically, both peptides caused a significant reduction at G0/G1 phase, in correlation with an augmented expression of the cell cycle inhibitory proteins p53, p27, p21, necrosis, exacerbated generation of reactive oxygen species and diminished mitochondrial membrane potential, all hallmarks of cellular stress. The screening of a novel panel of AgGom-analogues revealed that, unlike changes in the hydrophobicity and electrostatic surface, the cytotoxic potential of the gomesin analogues in DFTD cells lies on specific arginine substitutions in the eight and nine positions and alanine replacement in three, five and 12 positions. In conclusion, the evidence supports gomesin as a potential antiproliferative compound against DFTD disease.