Short-range phenotypic divergence among genetically distinct parapatric populations of an Australian funnel-web spider.

Speciation involves divergence at genetic and phenotypic levels. Where substantial genetic differentiation exists among populations, examining variation in multiple phenotypic characters may elucidate the mechanisms by which divergence and speciation unfold. Previous work on the Australian funnel-web spider Atrax sutherlandi Gray (2010; Records of the Australian Museum62, 285-392; Mygalomorphae: Hexathelidae: Atracinae) has revealed a marked genetic structure along a 110-kilometer transect, with six genetically distinct, parapatric populations attributable to past glacial cycles. In the present study, we explore variation in three classes of phenotypic characters (metabolic rate, water loss, and morphological traits) within the context of this phylogeographic structuring. Variation in metabolic and water loss rates shows no detectable association with genetic structure; the little variation observed in these rates may be due to the spiders' behavioral adaptations (i.e., burrowing), which buffer the effects of climatic gradients across the landscape. However, of 17 morphological traits measured, 10 show significant variation among genetic populations, in a disjunct manner that is clearly not latitudinal. Moreover, patterns of variation observed for morphological traits serving different organismic functions (e.g., prey capture, burrowing, and locomotion) are dissimilar. In contrast, a previous study of an ecologically similar sympatric spider with little genetic structure indicated a strong latitudinal response in 10 traits over the same range. The congruence of morphological variation with deep phylogeographic structure in Tallaganda's A. sutherlandi populations, as well as the inconsistent patterns of variation across separate functional traits, suggest that the spiders are likely in early stages of speciation, with parapatric populations independently responding to local selective forces.

Effects of substrate salinity on oviposition, embryonic development and survival in the Australian plague locust, Chortoicetes terminifera (Walker).

The Australian plague locust, Chortoicetes terminifera (Walker), is an important agricultural pest that oviposits into soil across vast semi-arid and arid regions. This study aimed to determine whether gravid female locusts can discriminate among substrates of increasing salinity (0, 4, 8, 12, 16, 20, 24, and 28ppt NaCl) when attempting oviposition, and quantify the effects of saline substrate on direct developing egg viability, and subsequent hatchling nymph body weight and survival. Gravid female locusts increasingly excavated and withdrew prior to completing oviposition in substrates of increasing salinity, but similar numbers of completed egg pods were observed across treatments. Egg weight at 50% total development time and successful egg development to nymph emergence decreased with increasing substrate salinity. Water balance equilibrium between the egg and the substrate occurred at approximately 12ppt NaCl corresponding to a water activity of ∼0.995. Eggs oviposited into sand containing ⩽12ppt NaCl weighed ⩾6.26±0.91mg and had ⩾76.8% successful development to nymph emergence. Eggs oviposited into sand containing >12ppt NaCl weighed ⩽5.16±1.27mg and had ⩽45.6% successful development to nymph emergence. Hatchling nymph body weight and survival to second instar also decreased with increasing substrate salinity. Nymphs that hatched and emerged from sand containing ⩽12ppt NaCl weighed ⩾5.55±0.43mg at emergence and had ⩾68.9% survival. Nymphs that hatched and emerged from sand containing >12ppt NaCl weighed ⩽5.28±0.67mg at emergence and had ⩽52.0% survival. These results indicate that C. terminifera is sufficiently resilient to develop and survive in saline substrates over most of their range.

Embryonic diapause in the Australian plague locust relative to parental experience of cumulative photophase decline.

The Australian plague locust Chortoicetes terminifera (Walker) exhibits facultative embryonic diapause during autumn. To approximate natural photoperiod changes during late summer and autumn, locust nymphs were reared under different total declines in laboratory photophase (-0.5, -0.75, -1.0, -1.25, -1.5, -1.75, -2 h each lowered in 15 min steps) in a 24 h photoperiod to quantify any effect on the subsequent production of diapause eggs. Induction of diapause eggs was significantly affected by accumulated photoperiod decline experienced by the parental generation throughout all development stages from mid-instar nymph to fledgling adult. The incidence of embryonic diapause ranged from nil at -0.5 h to 86.6% diapause at -2 h. Continued declines in photoperiod for post-teneral locusts (transitioned from -1h until fledging to -1.75 h) produced a further increase in the proportion of diapause eggs. The results were unaffected by time spent at any given photoperiod, despite a previously indicated maximal inductive photoperiod of 13.5h being used as the mid-point of all treatments. Implications for the seasonal timing processes of photoperiodism in C. terminifera, which has a high migratory capacity and a latitudinal cline in the timing of diapause egg production across a broad geographic range, are discussed.

Cold tolerance of the Australian spur-throated locust, Austracris guttulosa.

The cold tolerance of overwintering adult Spur-throated locusts, Austracris guttulosa, was examined using measures of supercooling point relative to gender, environmental acclimation and feeding state as well as mortality for a range of sub-zero temperature exposure treatments. Freezing was lethal and supercooling points ranged from -6 to -12.8°C, but were statistically independent of fresh mass, body water content, acclimation, and/or gut content in fed and starved individuals. A significant interaction effect of gender and feeding status showed that the larger bodied females had decreased supercooling capacity with increased food material in the digestive tract. Post-freezing dissections revealed differences in the amount of freshly consumed and retained food material in the digestive tract between fed and starved individuals of each gender, which could explain this effect based on inoculation of ice crystallisation by food particles. Above supercooling temperatures, neither gender nor the rate of cooling had a significant effect on mortality. When cooled from 25°C at 0.1 or 0.5°Cmin(-1) to a range of experimental minimum temperatures held for 3h, survival was ~74% to -7°C, but declined sharply to ~37% when cooled to -8°C or lower. Although the laboratory experiments reported here suggest that A. guttulosa is not freeze tolerant and unable to rapidly cold harden, exposure to typical cold and frosty nights that very rarely reach below -8°C as a night minimum in the field would be unlikely to cause mortality in the vast majority of overwintering aggregations.

Cold tolerance of first-instar nymphs of the Australian plague locust, Chortoicetes terminifera.

The cold tolerance of first-instar nymphs of the Australian plague locust, Chortoicetes terminifera, was examined using measures of total body water content, supercooling point and mortality for a range of sub-zero temperature exposure regimes. The supercooling points for starved and fed nymphs were -13.1+/-0.9 and -12.6+/-1.6 degrees C, and freezing caused complete mortality. Above these temperatures, nymphs were cold tolerant to different degrees based on whether they were starved or given access to food and water for 24h prior to exposure. The rate of cooling also had a significant effect on mortality. Very rapid cooling to -7 degrees C caused 84 and 87% mortality for starved and fed nymphs respectively, but this significantly decreased for starved nymphs if temperature declined by more ecologically realistic rates of 0.5 and 0.1 degrees C min(-1). These results are indicative of a rapid cold hardening response and are discussed in terms of the likely effects of cold nights and frost on first-instar nymphal survival in the field.

Field evaluation of vaporised ethyl formate and carbon dioxide for fumigation of stored wheat.

BACKGROUND: Vapormate is a cylinderised non-flammable mixture of ethyl formate (16.7% by weight) and carbon dioxide (CO(2)) that has been developed as a rapid fumigant of stored grain. Four field trials were undertaken on wheat in 50 t farm silos to demonstrate the feasibility of dynamic application. To assess treatment efficacy, each trial tested mixed stages of Rhyzopertha dominica F. (>11,000), Tribolium castaneum (Herbst.) (>1500) and Sitophilus oryzae (L.) (>13,000) in mesh cages positioned through the centre of the grain bulk and on the grain surface. Ethyl formate and CO(2) concentrations were measured in the silo during fumigation and in ambient air outside the 6 m fumigation zone. Application rates of 420, 660 and 940 g m(-3) of ethyl formate/CO(2) formulation and exposure times of 24, 3 and 72 h, respectively, were examined using wheat of 10.4-11.7% moisture content and grain temperatures between 2 and 32 degrees C. RESULTS: All life stages of R. dominica and T. castaneum were fully controlled under all conditions tested, and mortality of all life stages of S. oryzae was greater than 98%. CONCLUSION: Dynamic application of vaporised ethyl formate and CO(2) to 50 t silos proved safe to operators and rapidly effective against stored-grain insects on cold to warm grain.

Efficacy of vaporised ethyl formate/carbon dioxide formulation against stored-grain insects: effect of fumigant concentration, exposure time and two grain temperatures.

BACKGROUND: The ethyl formate/carbon dioxide (CO(2)) formulation Vapormate is a rapid-acting fumigant being developed for the control of stored-grain insects. The effects have been investigated of concentration, exposure times of 1, 3, 24 and 72 h and two grain temperatures, 15 and 25 degrees C, on its efficacy against mixed-stage cultures of Sitophilus oryzae (L.) Tribolium castaneum (Herbst) and strongly phosphine-resistant Rhyzopertha dominica (F.) strain QRD569. RESULTS: High mortalities (> or = 92%) of mixed-stage cultures of all three species were obtained when grain was fumigated with the formulation (193 g m(-3) ethyl formate) for 1 h. Complete control of R. dominica QRD569 and T. castaneum was achieved with 63 and 76 g m(-3) ethyl formate respectively, with exposure for 24 h, whereas mean mortality of S. oryzae was 86% under the same conditions. Mortalities of S. oryzae juvenile stages were significantly lower than adults under the conditions tested, which was due to pronounced tolerance of mid-stage pupae to the fumigant. Reducing grain temperature from 25 to 15 degrees C had no effect on insect mortality. CONCLUSION: Ethyl formate/CO(2) formulation is highly effective against stored-grain insects over a range of concentrations and exposure times. Efficacious fumigations were conducted in as little as 1 h, and a strongly phosphine-resistant R. dominica strain was readily controlled with the fumigant.

Effect of opioid compounds on feeding and activity of the cockroach, Periplaneta americana.

Opioid peptides have been implicated in regulation of feeding in invertebrates. Studies have suggested that receptors for opioids are present in cockroaches and that these receptors play roles in affecting both behaviour and feeding. We examined the effect of micro, delta, and kappa opioid receptor agonists and antagonists on feeding, mass changes and activity in the cockroach, Periplaneta americana. The kappa antagonist, nor-binaltorphimine, significantly increased food intake, while naltrexone (general antagonist) and naloxonazine (micro antagonist) both reduced feeding. A large mass loss was observed in cockroaches treated with nor-binaltorphimine, despite the increased food intake. Males did not lose as much mass during the 3h as females, although drug treatment did have some effect on the loss. Time of activity (%) was not influenced by any drug. Water loss experiments suggested that nor-binaltorphimine increased water loss, accounting for the mass loss despite the increased feeding. We suggest that two populations of opioid receptors are present as previously reported, with one affecting feeding and the other involved with evaporative water loss.

Neural regulation of discontinuous gas exchange in Periplaneta americana.

Patterns of gas exchange among terrestrial arthropods are highly variable from continuous to discontinuous with discretely partitioned phases. The underlying initiation and co-ordination of these patterns is relatively poorly understood. Here we present a novel method for the simultaneous measurement of central nervous system (CNS) activity of the metathoracic ganglion and VCO(2) in medium to large sized live terrestrial arthropods. Using Periplaneta americana at four oxygen levels (40%, 21%, 10% and 2% at 25 degrees C; n=6 per treatment), we present minimally invasive visualization of nervous output relative to typical resting discontinuous gas exchange (DGE) data for the first time. DGE was maintained when cockroaches were exposed to hyperoxia or moderate hypoxia, but was lost in severe hypoxia. CNS activity was manifested in three signal types: large CNS output coinciding with peak CO(2) production during a burst, moderate CNS output coinciding with CO(2) sawtoothing and fluttering, and minimal CNS activity during the closed phase of DGE in normoxia. Large and moderate CNS outputs were associated with observed abdominal pumping and congruent CO(2) peaks. At 10% oxygen, VCO(2) was significantly elevated during the inter-burst period in association with almost constant moderate CNS output between the periodic large CNS output. At 2% oxygen, DGE and large CNS output are lost to continuous CO(2) release and largely continuous moderate CNS output. As previously reported for this species, a central pattern generator for ventilation in the metathoracic ganglion is supported and we infer the presence of localized oxygen chemoreceptors based on clear CNS response to a change in oxygen tension.

Effects of phosphine on the neural regulation of gas exchange in Periplaneta americana.

Phosphine is used for fumigating stored commodities, however an understanding of the physiological response to phosphine in insects is limited. Here we show how the central pattern generator for ventilation in the central nervous system (CNS) responds to phosphine and influences normal resting gas exchange. Using the American cockroach, Periplaneta americana, that perform discontinuous gas exchange (DGE) at rest, we simultaneously measure ventilatory nervous output from the intact CNS, VCO(2) and water loss from live specimens. Exposure to 800 ppm phosphine at 25 degrees C for 2 h (n=13) during recording did not cause any mortality or obvious sub-lethal effects. Within 60 s of introducing phosphine into the air flow, all animals showed a distinct CNS response accompanied by a burst release of CO(2). The initial ventilatory response to phosphine displaced DGE and was typically followed by low, stable and continuous CO(2) output. CNS output was highest and most orderly under normoxic conditions during DGE. Phosphine caused a series of ventilatory CNS spikes preceding almost complete cessation of CNS output. Minimal CNS output was maintained during the 2 h normoxic recovery period and DGE was not reinstated. VCO(2) was slightly reduced and water loss significantly lower during the recovery period compared with those rates prior to phosphine exposure. A phosphine narcosis effect is rejected based on animals remaining alert at all times during exposure.

Physiology complements population structure of two endemic log-dwelling beetles.

Given rapid, global land modification and the likelihood of major global climate changes, it is becoming increasingly important to understand the physiological limits and capabilities of species to allow more accurate prediction of species' distributions under different scenarios of climate and landscape management. We studied whether the different habitat requirements of two species of tenebrionid beetles in temperate eucalypt forest could explain their patterns of dispersal and gene flow by applying flow-through respirometry to analysis of their physiological responses to different, ecologically relevant temperatures. Both Adelium calosomoides and Apasis puncticeps showed sensitivity to increasing temperatures (in terms of water loss), but Ap. puncticeps lost more water per unit of CO2 produced than did Ad. calosomoides. Recovery time from chill coma was also significantly longer for Ap. puncticeps than Ad. calosomoides. This supported prior qualitative assessment that Ap. puncticeps is more of a habitat specialist than Ad. calosomoides, at least concerning the critical factor of moisture requirements, and is consistent with stronger population genetic patterning and inferred low mobility of Ap. puncticeps. Despite its relatively lower mobility as deduced from population genetic structure, Ap. puncticeps walked four times faster than Ad. calosomoides in a laboratory assay, indicating that, for these species, mobility and gene flow are influenced more by physiological limitations than by speed.

Effects of temperature and oxygen availability on water loss and carbon dioxide release in two sympatric saproxylic invertebrates.

Water loss and VCO(2) relative to temperature and oxygen tension was investigated in a log-dwelling onychophoran (Euperipatoides rowelli) and a sympatric, un-described millipede species using flow-through respirometry. Onychophorans possess a tracheal system featuring permanently open spiracles. Total body water loss was consistently very high in E. rowelli and there was a positive correlation with increasing temperature. CO(2) output was continuous, increasing with higher temperatures and decreasing under lower oxygen concentrations. The millipede which has occludible spiracles also showed continuous gas exchange; however water loss was up to an order of magnitude lower than E. rowelli. An ability to survive under hypoxia is apparent for both species and corresponds with reports of hypoxic conditions within rotting logs. The rotting log habitat common to both taxa is characterized by high relative humidity and typically cool temperatures that approach 0 degrees C at night in winter. Consequently, dispersal through the higher temperatures and lower humidity of the exposed and dry understorey between suitable habitat may be hazardous for E. rowelli due to high desiccation susceptibility.

Cyclic gas exchange in the giant burrowing cockroach, Macropanesthia rhinoceros: effect of oxygen tension and temperature.

The giant burrowing cockroach, Macropanesthia rhinoceros, is endemic to north-eastern Australia and excavates a permanent burrow up to 1m deep into soil. Using flow-through respirometry, we investigated gas exchange and water loss at three different oxygen tensions (21%, 10% and 2% at 20 degrees C) and temperatures (10, 20 and 30 degrees C at 21% oxygen). M. rhinoceros employ cyclic gas exchange (CGE) making the species by far the largest insect known to engage in discontinuous ventilation. CGE featured rhythmic bursts of CO(2) dispersed among inter-burst periods of reduced output. CGE was most commonly observed at 20 degrees C and degraded at <10% oxygen. Mild hypoxia (10% oxygen) resulted in a lengthening of the burst period by approximately two-fold; this result is complementary to oxygen consumption data that suggests that the burst period is important in oxygen uptake. When exposed to severe hypoxia (2% oxygen), CGE was degraded to a more erratic continuous pattern. Also, during severe hypoxia, total water loss increased significantly, although CO(2) release was maintained at the same level as in 21% oxygen. During CGE, an increase in temperature from 10 to 20 degrees C caused both water loss and CO(2) output to double; from 20 to 30 degrees C, CO(2) output again doubled but water loss increased by only 31%.