Artificial diet for aphids - thirty years' experience

Myzus persicae (Sulzer) was reared continuously for over thirty years (until it died out in December 2008) on a totally defined synthetic artificial diet, the procedure for which is described. Development time was extended on diet compared with rearing on Brussels sprout plants (Brassica oleracea L. var. gemmifera L.), and generation time was further increased by an added pre-reproductive period of 4 days. Fecundity was reduced by about two-thirds, and mean relative growth rate in weight (MRGR) was only 60% in comparison with plant-reared aphids. Applying 2 kg/cm(2) pressure to a 10% sucrose solution extended the adult longevity of Aphis fabae Scopoli by less than I day. In contrast, a short experience of half-strength diet Caused a sharp rise in honeydew excretion by A. fabae for several hours, and alternating full-strength diet with diluted diets (including water) Caused a greater weight increase. The poor performance of aphids on diet thus seems to have a behavioural rather than a mechanical explanation. The diet, designed to give optimal performance of the aphids, has proved not to be useful for nutritional studies, as any change is deleterious. Areas of aphid research where the diet has been useful, however, are studies on repellents/attractants/toxins, role of symbionts, maintenance of genotype collections, work on parasitoid behaviour in relation to plant chemistry, and collection of aphid saliva.

Plastic and evolutionary responses to heat stress in a temperate dung fly: negative correlation between basal and induced heat tolerance?

Extreme weather events such as heat waves are becoming more frequent and intense. Populations can cope with elevated heat stress by evolving higher basal heat tolerance (evolutionary response) and/or stronger induced heat tolerance (plastic response). However, there is ongoing debate about whether basal and induced heat tolerance are negatively correlated and whether adaptive potential in heat tolerance is sufficient under ongoing climate warming. To evaluate the evolutionary potential of basal and induced heat tolerance, we performed experimental evolution on a temperate source 4 population of the dung fly Sepsis punctum. Offspring of flies adapted to three thermal selection regimes (Hot, Cold and Reference) were subjected to acute heat stress after having been exposed to either a hot-acclimation or non-acclimation pretreatment. As different traits may respond differently to temperature stress, several physiological and life history traits were assessed. Condition dependence of the response was evaluated by exposing juveniles to different levels of developmental (food restriction/rearing density) stress. Heat knockdown times were highest, whereas acclimation effects were lowest in the Hot selection regime, indicating a negative association between basal and induced heat tolerance. However, survival, adult longevity, fecundity and fertility did not show such a pattern. Acclimation had positive effects in heat-shocked flies, but in the absence of heat stress hot-acclimated flies had reduced life spans relative to nonacclimated ones, thereby revealing a potential cost of acclimation. Moreover, body size positively affected heat tolerance and unstressed individuals were less prone to heat stress than stressed flies, offering support for energetic costs associated with heat tolerance. Overall, our results indicate that heat tolerance of temperate insects can evolve under rising temperatures, but this response could be limited by a negative relationship between basal and induced thermotolerance, and may involve some but not other fitness-related traits.

The influence of larval density, food availability and habitat longevity on the life history and population growth rate of the midge Chironomus riparius

Despite long-standing interest in the forms and mechanisms of density dependence, these are still imperfectly understood. However, in a constant environment an increase in density must reduce per capita resource availability, which in turn leads to reduced survival, fecundity and somatic growth rate. Here we report two population experiments examining the density dependent responses under controlled conditions of an important indicator species, Chironomus riparius. The first experiment was run for 35 weeks and was started at low density with replicate populations being fed three different rations. Increased ration reduced generation time and increased population growth rate (pgr) but had no effect on survival, fecundity and female body weight in the first generation. In the second generation there was a six-fold increase in generation time, presumably due to the greatly reduced per capita resource availability as the estimated initial densities of the second generation were 300 times greater than the first. Juvenile survival to emergence, fecundity, adult body weight and pgr declined by 90%, 75%, 35% and 99%, respectively. These large between-generation effects may have obscured the effects of the threefold variation in ration, as only survival to emergence significantly increased with ration in the second generation. These results suggest that some chironomid larvae survive a reduction in resource availability by growing more slowly. In the ephemeral habitats sometimes occupied by C. riparius, the effects of population density may depend crucially on the longevity of the environment. A second experiment was therefore performed to measure pgr from six different starting densities over an eight-week period. The relationship between pgr and density was concave, viewed from above. At densities above 16 larvae per cm(2), less than 1% of the population emerged and no offspring were produced. Under the conditions of experiment 2 - an 8-week habitat lifespan carrying capacity was estimated as 8 larvae per cm(2).

Cholinergic modulation of intrinsic fibre-evoked excitatory transmission contains a nicotinic component in immature but not adult rat piriform cortex, in vitro

The piriform cortex (PC) is highly prone to epileptogenesis, particularly in immature animals, where decreased muscarinic modulation of PC intrinsic fibre excitatory neurotransmission is implicated as a likely cause. However, whether higher levels of acetylcholine (ACh) release occur in immature vs. adult PC remains unclear. We investigated this using in vitro extracellular electrophysiological recording techniques. Intrinsic fibre-evoked extracellular field potentials (EFPs) were recorded from layers II to III in PC brain slices prepared from immature (P14-18) and adult (P>40) rats. Adult and immature PC EFPs were suppressed by eserine (1muM) or neostigmine (1muM) application, with a greater suppression in immature ( approximately 40%) than adult ( approximately 30%) slices. Subsequent application of atropine (1muM) reversed EFP suppression, producing supranormal ( approximately 12%) recovery in adult slices, suggesting that suppression was solely muscarinic ACh receptor-mediated and that some 'basal' cholinergic 'tone' was present. Conversely, atropine only partially reversed anticholinesterase effects in immature slices, suggesting the presence of additional non-muscarinic modulation. Accordingly, nicotine (50muM) caused immature field suppression ( approximately 30%) that was further enhanced by neostigmine, whereas it had no effect on adult EFPs. Unlike atropine, nicotinic antagonists, mecamylamine and methyllycaconitine, induced immature supranormal field recovery ( approximately 20%) following anticholinesterase-induced suppression (with no effect on adult slices), confirming that basal cholinergic 'tone' was also present. We suggest that nicotinic inhibitory cholinergic modulation occurs in the immature rat PC intrinsic excitatory fibre system, possibly to complement the existing, weak muscarinic modulation, and could be another important developmentally regulated system governing immature PC susceptibility towards epileptogenesis.

Fate of prions in soil: Longevity and migration of recPrP in soil columns

Research into transmissible spongiform encephalopathy (TSE) diseases has become a high priority worldwide in recent years yet remarkably little is known about the behaviour of TSE infectivity in the environment. The resilience and stability of prion proteins could lead to soils becoming a potential reservoir of TSE infectivity as a result of contamination from activities such as infected carcass burial or the dispersion of effluents from slaughter houses, or by contamination of pastures by infected animals, (e.g. scrapie in sheep). Knowledge of the fate of prion proteins in soils, and associated physico-chemical conditions which favour migration, can be used to help prevent re-infection of animals through grazing, to protect watercourses and develop good management practices. In two consecutive experiments of 9 and 6 months, the migration of recombinant ovine PrP (recPrP) in soil columns was followed under contrasting levels of microbial activity (normal versus reduced), under varying regimes of soil water content and redox potential, and in two different soil types (loamy sand and clay loam). At each analysis time (1, 3, 6 or 9 months), in both soil types, full-length recPrP was detected in the original contaminated layer, indicating the resilience and stability of recPrP under varied soil conditions, even in the presence of active soil microbial populations. Evidence of protein migration was found in every soil column at the earliest analysis time (1 or 3 months), but was restricted to a maximum distance of 1 cm, indicative of limited initial mobility in soils followed by strong adsorption over the following days to weeks. The survival of recPrP in the soil over a period of at least 9 months was demonstrated. In this study, recPrP was used as an indicator for potential TSE infectivity, although infectivity tests should be carried out before conclusions can be drawn regarding the infection risk posed by prions in soil. However, it has been demonstrated that soil is likely to act as a significant barrier to the dispersion of contaminated material at storage or burial sites. (c) 2007 Elsevier Ltd. All rights reserved.

Errors associated with preparation of adult and paediatric parenteral nutrition in UK hospital pharmacies

Rationale: In UK hospitals, the preparation of all total parenteral nutrition (TPN) products must be made in the pharmacy as TPNs are categorised as high-risk injectables (NPSA/2007/20). The National Aseptic Error Reporting Scheme has been collecting data on pharmacy compounding errors in the UK since August 2003. This study reports on types of error associated with the preparation of TPNs, including the stage at which these were identified and potential and actual patient outcomes. Methods: Reports of compounding errors for the period 1/2004 - 3/2007 were analysed on an Excel spreadsheet. Results: Of a total of 3691 compounding error reports, 674 (18%) related to TPN products; 548 adult vs. 126 paediatric. A significantly higher proportion of adult TPNs (28% vs. 13% paediatric) were associated with labelling errors and a significantly higher proportion of paediatric TPNs (25% vs. 15% adult) were associated with incorrect transcriptions (Chi-Square Test; p<0.005). Labelling errors were identified equally by pharmacists (42%) and technicians (48%) with technicians detecting mainly at first check and pharmacists at final check. Transcription errors were identified mainly by technicians (65% vs. 27% pharmacist) at first check. Incorrect drug selection (13%) and calculation errors (9%) were associated with adult and paediatric TPN preparations in the same ratio. One paediatric TPN error detected at first check was considered potentially catastrophic; 31 (5%) errors were considered of major and 38 (6%) of moderate potential consequence. Five errors (2 moderate, 1 minor) were identified during or after administration. Conclusions: While recent UK patient safety initiatives are aimed at improving the safety of injectable medicines in clinical areas, the current study highlights safety problems that exist within pharmacy production units. This could be used in the creation of an error management tool for TPN compounding processes within hospital pharmacies.

Developmental changes in the germinability, desiccation tolerance, hardseededness, and longevity of individual seeds of Trifolium ambiguum

Background and Aims: Using two parental clones of outcrossing Trifolium ambiguum as a potential model system, we examined how during seed development the maternal parent, number of seeds per pod, seed position within the pod, and pod position within the inflorescence influenced individual seed fresh weight, dry weight, water content, germinability, desiccation tolerance, hardseededness, and subsequent longevity of individual seeds. Methods: Near simultaneous, manual reciprocal crosses were carried out between clonal lines for two experiments. Infructescences were harvested at intervals during seed development. Each individual seed was weighed and then used to determine dry weight or one of the physiological behaviour traits. Key Results: Whilst population mass maturity was reached at 33–36 days after pollination (DAP), seed-to-seed variation in maximum seed dry weight, when it was achieved, and when maturation drying commenced, was considerable. Individual seeds acquired germinability between 14 and 44 DAP, desiccation tolerance between 30 and 40 DAP, and the capability to become hardseeded between 30 and 47 DAP. The time for viability to fall to 50 % (p50) at 60 % relative humidity and 45 °C increased between 36 and 56 DAP, when the seed coats of most individuals had become dark orange, but declined thereafter. Individual seed f. wt at harvest did not correlate with air-dry storage survival period. Analysing survival data for cohorts of seeds reduced the standard deviation of the normal distribution of seed deaths in time, but no sub-population showed complete uniformity of survival period. Conclusions: Variation in individual seed behaviours within a developing population is inherent and inevitable. In this outbreeder, there is significant variation in seed longevity which appears dependent on embryo genotype with little effect of maternal genotype or architectural factors.

Δ9-tetrahydrocannabivarin suppresses in vitro epileptiform and in vivo seizure activity in adult rat

Purpose: We assessed the anticonvulsant potential of the phytocannabinoid Δ9-tetrahydrocannabivarin (Δ9-THCV) by investigating its effects in an in vitro piriform cortex (PC) brain slice model of epileptiform activity, on cannabinoid CB1 receptor radioligand-binding assays and in a generalized seizure model in rats. Methods: Δ9-THCV was applied before (10 μmΔ9-THCV) or during (10–50 μmΔ9-THCV) epileptiform activity induced by Mg2+-free extracellular media in adult rat PC slices and measured using multielectrode array (MEA) extracellular electrophysiologic techniques. The actions of Δ9-THCV on CB1 receptors were examined using [3H]SR141716A competition binding and [35S]GTPS assays in rat cortical membranes. Effects of Δ9-THCV (0.025–2.5 mg/kg) on pentylenetetrazole (PTZ)–induced seizures in adult rats were also assessed. Results: After induction of stable spontaneous epileptiform activity, acute Δ9-THCV application (≥20 μm) significantly reduced burst complex incidence and the amplitude and frequency of paroxysmal depolarizing shifts (PDSs). Furthermore, slices pretreated with 10 μmΔ9-THCV prior to induction of epileptiform activity exhibited significantly reduced burst complex incidence and PDS peak amplitude. In radioligand-binding experiments, Δ9-THCV acted as a CB1 receptor ligand, displacing 0.5 nm [3H]SR141716A with a Ki∼290 nm, but exerted no agonist stimulation of [35S]GTPS binding. In PTZ-induced seizures in vivo, 0.25 mg/kg Δ9-THCV significantly reduced seizure incidence. Discussion: These data demonstrate that Δ9-THCV exerts antiepileptiform and anticonvulsant properties, actions that are consistent with a CB1 receptor–mediated mechanism and suggest possible therapeutic application in the treatment of pathophysiologic hyperexcitability states.

Temperature sensitivity of the low-moisture-content limit to negative seed longevity-moisture content relationships in hermetic storage

Background and Aims The negative logarithmic relationship between orthodox seed longevity and moisture content in hermetic storage is subject to a low-moisture-content limit (m(c)), but is m(c) affected by temperature? Methods Red clover (Trifolium pratense) and alfalfa (Medicago sativa) seeds were stored hermetically at 12 moisture contents (2-15 %) and five temperatures (-20, 30, 40, 50 and 65 degrees C) for up to 14.5 years, and loss in viability was estimated. Key Results Viability did not change during 14.5 years hermetic storage at -20 degrees C with moisture contents from 2.2 to 14.9 % for red clover, or 2.0 to 12.0 % for alfalfa. Negative logarithmic relationships between longevity and moisture contents > m(c) were detected at 30-65 degrees C, with discontinuities at low moisture contents; m(c) varied between 4.0 and 5.4 % (red clover) or 4.2 and 5.5 % (alfalfa), depending upon storage temperature. Within the ranges investigated, a reduction in moisture content below m(c) at any one temperature had no effect on longevity. Estimates of m(c) were greater the cooler the temperature, the relationship (P < 0.01) being curvilinear. Above m(c), the estimates of C-H and C-Q (i.e. the temperature term of the seed viability equation) did not differ (P > 0.10) between species, whereas those of K-E and C-W did (P < 0.001). Conclusions The low-moisture-content limit to negative logarithmic relationships between seed longevity and moisture content in hermetic storage increased the cooler the storage temperature, by approx. 1.5 % over 35 degrees C (4.0-4.2 % at 65 degrees C to 5.4-5.5 % at 30-40 degrees C) in these species. Further reduction in moisture content was not damaging. The variation in m(c) implies greater sensitivity of longevity to temperature above, compared with below, m(c). This was confirmed (P < 0.005).

Seed longevity - moisture content relationships in hermetic and open storage

In seed storage research, moisture content can be maintained by providing a stable relative humidity (e.g. over saturated salt solutions) or by hermetic storage, but the two approaches provide different gaseous environments which might affect longevity. Seeds of timothy (Phleum pratense L.) and sesame (Sesamum indicum L.) were stored at 45 degrees C or 50 degrees C, respectively, with different moisture contents maintained by hermetic storage in laminated-aluminium-foil packets, or by desiccators above either saturated salt solutions or moistened silica gel. Seeds were withdrawn from storage at intervals from 1 to 28 d for up to 480 d and viability estimated. Within a species, the negative logarithmic relation between seed longevity and moisture content did not differ (P> 0.25, timothy; >0.05, sesame) between storage in desiccators over either moistened silica gel or saturated salt solutions, whereas the relation was much steeper (P< 0.005) in hermetic storage: longevity was similar at high moisture contents, but at low values much greater with hermetic storage. This effect of storage method on seed longevity's sensitivity to moisture content implies that oxygen is relatively more deleterious to seeds at lower than at greater moisture contents and confirms that hermetic storage is preferable for long-term seed storage at low moisture contents.

Quantitative response of the longevity of seed of twelve crops to temperature and moisture in hermetic storage

Seeds of 39 seed lots of a total of twelve different crops were stored hermetically in a wide range of air-dry environments (2-25% moisture content at 0-50 degrees C), viability assessed periodically, and the seed viability equation constants estimated. Within a species, estimates of the constants which quantify absolute longevity (K-E) and the relative effects on longevity of moisture content (C-W) and temperature (C-H and C-Q) did not differ (P >0.05 to P >0.25) among lots. Comparison among the 12 crops provided variant estimates of K-E and C-W (P< 0.01), but common values of C-H and C-Q (0.0322 and 0.000454, respectively, P >0.25). Maize (Zea mays) provided the greatest estimate of K-E (9.993, s.e.= 0.456), followed by sorghum (Sorghum bicolor) (9.381, s.e. 0.428), pearl millet (Pennisetum typhoides) (9.336, s.e.= 0.408), sugar beet (Beta vulgaris) (8.988, s.e.= 0.387), African rice (Oryza glaberrima) (8.786, s.e.= 0.484), wheat (Triticum aestivum) (8.498, s.e.= 0.431), foxtail millet (Setaria italica) (8.478, s.e.= 0.396), sugarcane (Saccharum sp.) (8.454, s.e.= 0.545), finger millet (Eleusine coracana) (8.288, s.e.= 0.392), kodo millet (Paspalum scrobiculatum) (8.138, s.e.= 0.418), rice (Oryza sativa) (8.096, s.e.= 0.416) and potato (Solanum tuberosum) (8.037, s.e.= 0.397). Similarly, estimates of C-W were ranked maize (5.993, s.e.= 0.392), pearl millet (5.540, s.e.= 0.348), sorghum (5.379, s.e.=0.365), potato (5.152, s.e.= 0.347), sugar beet (4.969, s.e.= 0.328), sugar cane (4.964, s.e.= 0.518), foxtail millet (4.829, s.e.= 0.339), wheat (4.836, s.e.= 0.366), African rice (4.727, s.e.= 0.416), kodo millet (4.435, s.e.= 0.360), finger millet (4.345, s.e.= 0.336) and rice (4.246, s.e.= 0.355). The application of these constants to long-term seed storage is discussed.

Drying method influences the development of germinability, dessication tolerance and subsequent longevity of immature seeds of sumauma (Ceiba pentandra (L.) Gaertn. [Bombacaceae])

The ability to germinate, tolerate desiccation and survive in air-dry storage was investigated during early seed development in planta and subsequent ex planta maturation of sumauma (Ceiba pentandra). Immature fruits were collected on three different dates (i.e. from about 5 days before until 7 days after mass maturity). Immature fresh seeds were not able to germinate. Fruits or seeds were subjected immediately after each collection to three different drying treatments with progressively slower rates of dessication: (i) seeds were extracted from the fruits and dried immediately; (ii) fruits were dried in a thin layer; (iii) fruits were dried in a tied polyethylene bag (with 10 holes of 1cm diameter). Drying was in a room maintained at 25 degrees C +/- 3 degrees C and 65%+/- 5% r.h. For treatment (i) the seeds were dried for 6 days in order to reduce moisture content to around 13% ( +/- 2%) moisture content. For treatments (ii) and (iii) the fruits were subjected to different periods of drying depending upon collection date. The results of these post-collection treatments showed generally that the more immature the seeds the slower the rate of drying that is required to improve ability to germinate, ability to tolerate desiccation and potential longevity, but at the third harvest, 7 days after mass maturity, the intermediate drying rate treatment was the most beneficial. Thus post fruit collection treatments can be modified depending upon the stage of seed development in order to provide good to high quality seeds of sumauma when collection has to be made at a site with difficult access at less than ideal times. The results are relevant to seed collection practices for both forestry and ex situ plant biodiversity conservation.

Mutant alleles at the rugosus loci in pea affect seed moisture sorption isotherms and the relations between seed longevity and moisture content

Pea (Pisum sativum L.) mutant near-isogenic lines (RRrbrb, rrRbRb, rrrbrb) with lower starch but higher lipid contents, brought about by lesions in the starch biosynthetic pathway, had seed moisture sorption isotherms displaced below that of the wild type (RRRbRb). The negative logarithmic relationship between seed longevity and seed storage moisture content (%, f.wt basis), determined in hermetic storage at 65 degreesC, also differed: longevity in the mutant near-isogenic lines was poorer and less sensitive to moisture content than in the wild type (i.e. C-w was lower). The low-moisture-content limit (m(c)) to this relation also differed, being lower in the mutant near-isogenic lines (5.4-5.9%) than in the wild type (6.1%). In contrast, all four near-isogenic lines showed no difference (P >0.25) in the negative semilogarithmic relationship between equilibrium relative humidity (ERH) and seed longevity. It is concluded that the effect of these alleles at the r and rb loci on seed longevity. was largely indirect; a consequence of their effect on seed composition and hence on moisture sorption isotherms. However, this explanation could not be invoked at moisture contents below mc where differences in longevity remained substantial (RRRbRb double that of rrrbrb). Hence, these mutant alleles affected seed longevity directly at very low moisture contents.