ELISA absorbance cut-off method affects malaria sporozoite rate determination in wild Afrotropical Anopheles

ABSTRACT. Malaria sporozoite infection rates in a mixed species group of 244 Anopheles gambiae Giles sensu lato and 115 An. funestus Giles wild female mosquitoes were compared using three methods to determine cutoff absorbance values for positivity of a Plasmodium falciparum Welch enzyme-linked immunosorbent assay (ELISA). Positive controls were based on P. falciparum circumsporozoite protein. As negative controls, four wild male Anopheles were included on each microtitre plate; tests were repeated on four consecutive days for each plate.
Infection rates were estimated at 13.1–22.8% using the mean absorbance value of negative controls plus three standard deviations, 11.7–12.8% using double the mean and 12.5–13.6% using the fixed cut-off value of 0.20 (allowing for 20% variation in negative control absorbance values).
Observed agreement for positivity or negativity among samples tested four times was 98.6% for the 2× mean method, 97.2% for the fixed cut-off 0.20 value, but only 82.7% for the mean +3 SD method. It was concluded that the 2× mean cut-off method is most reliable for field studies. P. falciparum sporozoite rates of 12.2% in An. funestus and 11.9% in An. gambiae s. l . were thus determined on the basis of the 2× mean cut-off method.
This comparative evaluation demonstrates that vector infectivity rates can be seriously over-estimated from sporozoite ELISA tests, by as much as 87% in one case considered here, depending on the absorbance cut-off method applied for negative controls.     

Quantitation of malaria sporozoites transmitted in vitro during salivation by wild Afrotropical Anopheles

Abstract. The malaria transmission potential of wild, infective Anopheles from western Kenya was evaluated by determining the number of sporozoites transmitted in vitro by salivation when their mouthparts were inserted into capillary tubes containing either sucrose or blood. With sucrose, 86.6% of 102 infective Anopheles transmitted a geometric mean (GM) of 3.84 sporozoites (range 1–34). With blood, 23.1% of 104 infective Anopheles , tested on the day of collection, transmitted a GM of 2.30 sporozoites (range 1–117). For Anopheles held 5 days postcapture before testing with blood, 53.6% of 56 transmitted a GM of 6.04 sporozoites (range 1–420). Transmitting Anopheles contained significantly more salivary gland sporozoites than non-transmitters. No significant differences were detected between Anopheles gambiae Giles sensu lato and Anopheles funestus Giles in sporozoite transmission by individuals with sporozoites in their salivary glands.
Sporozoites were detected microscopically in the salivary duct from heads in 80.3% of 117 infective Anopheles (GM=11.2, range 1–71). Sporozoite detection in mosquito heads by ELISA was 25% less efficient than microscopic detection.
Over 98% of the infective Anopheles transmitted less than twenty-five sporozoites. Transmitted sporozoites represented only about 3% of the total sporozoites in the salivary glands suggesting that sporozoite transmission may be restricted to sporozoites in the salivary duct at the time of feeding. Results are discussed in relation to anti-sporozoite vaccine development.     

Herbicides which inhibit photosystem II or produce chlorosis and their effect on production and transformation of pigments in etiolated radish seedlings (Raphanus sativus)

Abstract The herbicides DCMU, bentazon, amitrole, and SAN 6706 were tested for their ability to influence the carotenoid and pro-tochlorophyll(ide) composition as well as the protochloro-phyll(ide) phototransformation and the Shibata shift in dark-grown radish seedlings (Raphanus sativus L. cv. Saxa Treib). Bentazon enhanced the formation of lutein and carotenes, while SAN 6706 suppressed the biosynthesis of carotenoids. Amitrole led to a reduced accumulation of phototransformable pro-tochlorophyll(ide). The phototransformation of pro-tochlorophyll(ide) and the Shibata shift were not affected by any of the tested herbicides, irrespective of the presence or absence of activated phytochrome. From this we conclude that herbicides inhibiting photosystem II or producing chlorosis partly affect, but do not block, carotenoid and chlorophyll biosynthesis in dark-grown plants. The main herbicide effect becomes visible only after prolonged illumination.     

Coordinated approaches to quantify long‐term ecosystem dynamics in response to global change

Many serious ecosystem consequences of climate change will take decades or even centuries to emerge. Long‐term ecological responses to global change are strongly regulated by slow processes, such as changes in species composition, carbon dynamics in soil and by long‐lived plants, and accumulation of nutrient capitals. Understanding and predicting these processes require experiments on decadal time scales. But decadal experiments by themselves may not be adequate because many of the slow processes have characteristic time scales much longer than experiments can be maintained. This article promotes a coordinated approach that combines long‐term, large‐scale global change experiments with process studies and modeling. Long‐term global change manipulative experiments, especially in high‐priority ecosystems such as tropical forests and high‐latitude regions, are essential to maximize information gain concerning future states of the earth system. The long‐term experiments should be conducted in tandem with complementary process studies, such as those using model ecosystems, species replacements, laboratory incubations, isotope tracers, and greenhouse facilities. Models are essential to assimilate data from long‐term experiments and process studies together with information from long‐term observations, surveys, and space‐for‐time studies along environmental and biological gradients. Future research programs with coordinated long‐term experiments, process studies, and modeling have the potential to be the most effective strategy to gain the best information on long‐term ecosystem dynamics in response to global change.     

Rapid oxygen exchange across the leaves of Littorella uniflora provides tolerance to sediment anoxia

1. Littorella uniflora and Lobelia dortmanna are prominent small rosette species in nutrient‐poor, soft‐water lakes because of efficient root exchange of CO₂ and O₂. We hypothesise that higher gas exchange across the leaves of L. uniflora than of L. dortmanna ensures O₂ uptake from water and underlies its greater tolerance to sediment anoxia following organic enrichment. 2. We studied plant response to varying sediment O₂ demand and biogeochemistry by measuring photosynthesis, gas exchange across leaves and O₂ dynamics in plants during long‐term laboratory and field studies. Frequent non‐destructive sampling of sediment pore water was used to track changes in sediment biogeochemistry. 3. Addition of organic matter triggered O₂ depletion and accumulation of , Fe²⁺ and CO₂ in sediments. Gas exchange across leaf surfaces was 13–16 times higher for L. uniflora than for L. dortmanna. Oxygen in the leaf lacunae of L. uniflora remained above 10 kPa late at night on anoxic sediments despite organic enrichment. Leaf content of N and P of L. uniflora remained sufficient to keep up photosynthesis despite prolonged sediment anoxia, whereas nutrient content was too low for long‐term survival of L. dortmanna. 4. High gas exchange across L. uniflora leaves improves its performance and survival on anoxic sediments compared with L. dortmanna. Lobelia dortmanna uses the same gas‐tight leaves in air and water, which makes it highly susceptible to sediment anoxia but more cost‐effective in ultra‐oligotrophic environments because of slow leaf turnover.     

Contrasting effects of repeated summer drought on soil carbon efflux in hydric and mesic heathland soils

Current predictions of climate change include altered rainfall patterns throughout Europe, continental USA and areas such as the Amazon. The effect of this on soil carbon efflux remains unclear although several modelling studies have highlighted the potential importance of drought for carbon storage. To test the importance of drought, and more importantly repeated drought year-on-year, we used automated retractable curtains to exclude rain and produce repeated summer drought in three heathlands at varying moisture conditions. This included a hydric system limited by water-excess (in the UK) and two mesic systems with seasonal water limitation in Denmark (DK) and the Netherlands (NL). The experimental rainfall reductions were set to reflect single year droughts observed in the last decade with exclusion of rain for 2–3 months of the year resulting in a 20–26% reduction in annual rainfall and 23–38% reduction in mean soil moisture during the drought period. Unexpectedly, sustained reduction in soil moisture over winter (between drought periods) was also observed at all three sites, along with a reduction in the maximum water-holding capacity attained. Three hypotheses are discussed which may have contributed to this lack of recovery in soil moisture: hydrophobicity of soil organic matter, increased water use by plants and increased cracking of the soil. The responses of soil respiration to this change in soil moisture varied among the sites: decreased rates were observed at the water-limited NL and DK sites whilst they increased at the UK site. Reduced sensitivity of soil respiration to soil temperature was observed at soil moisture contents above 55% at the UK site and below 20% and 13% at the NL and DK sites, respectively. Soil respiration rates recovered to predrought levels in the NL and DK sites during the winter re-wetting period that indicates any change in soil C storage due to changes in soil C efflux may be short lived in these mesic systems. In contrast, in the hydric UK site after 2 years of drought treatment, the persistent reduction in soil moisture throughout the year resulted in a year-round increase in soil respiration flux, a response that accelerated over time to 40% above control levels. These findings suggest that carbon-rich soils with high organic matter content may act as a significant source of CO₂ to the atmosphere following repeated summer drought. Nonrecovery of soil moisture and a persistent increase in soil respiration may be the primary mechanism underlying the reported substantial losses of soil carbon from UK organic soils over the last 20 years. These findings indicate that the water status of an ecosystem will be a critical factor to consider in determining the impact of drought on the soil carbon fluxes and storage.     

Response of plant species richness and primary productivity in shrublands along a north–south gradient in Europe to seven years of experimental warming and drought: reductions in primary productivity in the heat and drought year of 2003

We used a nonintrusive field experiment carried out at six sites – Wales (UK), Denmark (DK), the Netherlands (NL), Hungary (HU), Sardinia (Italy – IT), and Catalonia (Spain – SP) – along a climatic and latitudinal gradient to examine the response of plant species richness and primary productivity to warming and drought in shrubland ecosystems. The warming treatment raised the plot daily temperature by ca. 1 °C, while the drought treatment led to a reduction in soil moisture at the peak of the growing season that ranged from 26% at the SP site to 82% in the NL site. During the 7 years the experiment lasted (1999–2005), we used the pin‐point method to measure the species composition of plant communities and plant biomass, litterfall, and shoot growth of the dominant plant species at each site. A significantly lower increase in the number of species pin‐pointed per transect was found in the drought plots at the SP site, where the plant community was still in a process of recovering from a forest fire in 1994. No changes in species richness were found at the other sites, which were at a more mature and stable state of succession and, thus less liable to recruitment of new species. The relationship between annual biomass accumulation and temperature of the growing season was positive at the coldest site and negative at the warmest site. The warming treatment tended to increase the aboveground net primary productivity (ANPP) at the northern sites. The relationship between annual biomass accumulation and soil moisture during the growing season was not significant at the wettest sites, but was positive at the driest sites. The drought treatment tended to reduce the ANPP in the NL, HU, IT, and SP sites. The responses to warming were very strongly related to the Gaussen aridity index (stronger responses the lower the aridity), whereas the responses to drought were not. Changes in the annual aboveground biomass accumulation, litterfall, and, thus, the ANPP, mirrored the interannual variation in climate conditions: the most outstanding change was a decrease in biomass accumulation and an increase in litterfall at most sites during the abnormally hot year of 2003. Species richness also tended to decrease in 2003 at all sites except the cold and wet UK site. Species‐specific responses to warming were found in shoot growth: at the SP site, Globularia alypum was not affected, while the other dominant species, Erica multiflora, grew 30% more; at the UK site, Calluna vulgaris tended to grow more in the warming plots, while Empetrum nigrum tended to grow less. Drought treatment decreased plant growth in several studied species, although there were some species such as Pinus halepensis at the SP site or C. vulgaris at the UK site that were not affected. The magnitude of responses to warming and drought thus depended greatly on the differences between sites, years, and species and these multiple plant responses may be expected to have consequences at ecosystem and community level. Decreases in biodiversity and the increase in E. multiflora growth at the SP site as a response to warming challenge the assumption that sensitivity to warming may be less well developed at more southerly latitudes; likewise, the fact that one of the studied shrublands presented negative ANPP as a response to the 2003 heat wave also challenges the hypothesis that future climate warming will lead to an enhancement of plant growth and carbon sequestration in temperate ecosystems. Extreme events may thus change the general trend of increased productivity in response to warming in the colder sites.     

Significance of cold-season respiration and photosynthesis in a subarctic heath ecosystem in Northern Sweden

While substantial cold-season respiration has been documented in most arctic and alpine ecosystems in recent years, the significance of cold-season photosynthesis in these biomes is still believed to be small. In a mesic, subartic heath during both the cold and warm season, we measured in situ ecosystem respiration and photosynthesis with a chamber technique at ambient conditions and at artificially increased frequency of freeze–thaw (FT) cycles during fall and spring. We fitted the measured ecosystem exchange rates to respiration and photosynthesis models with R²-values ranging from 0.81 to 0.85. As expected, estimated cold-season (October, November, April and May) respiration was significant and accounted for at least 22% of the annual respiratory CO₂ flux. More surprisingly, estimated photosynthesis during this period accounted for up to 19% of the annual gross CO₂ uptake, suggesting that cold-season photosynthesis partly balanced the cold-season respiratory carbon losses and can be significant for the annual cycle of carbon. Still, during the full year the ecosystem was a significant net source of 120 ± 12 g C m⁻² to the atmosphere. Neither respiration nor photosynthetic rates were much affected by the extra FT cycles, although the mean rate of net ecosystem loss decreased slightly, but significantly, in May. The results suggest only a small response of net carbon fluxes to increased frequency of FT cycles in this ecosystem.     

Haeckelia (= Euchlora) and Hydroctena and the phylogenetic interrelationships of Cnidaria and Ctenophora

A scrutiny of the literature shows that the ctenophore Haeckelia (= Euchlora) ruba has only kleptocnidae and that Hydroctena salenskii is a ctenophore without special cnidarian affinities. The “missing links” between cnidarians and ctenophores have thus turned out to be based on misinterpretations and must be excluded from future discussions on phylogeny.