Workshop report on the extraction of foetal DNA from maternal plasma

Objective Cell free foetal DNA (cff DNA) extracted from maternal plasma is now recognized as a potential source for prenatal diagnosis but the methodology is currently not well standardized. To evaluate different manual and automated DNA extraction methods with a view to developing standards, an International Workshop was performed. Methods Three plasma pools from RhD-negative pregnant women, a DNA standard, real-time-PCR protocol, primers and probes for RHD were sent to 12 laboratories and also to one company (Qiagen, Hilden, Germany). In pre-tests, pool 3 showed a low cff DNA concentration, pool 1 showed a higher concentration and pool 2 an intermediate concentration. Results The QIAamp DSP Virus Kit, the High Pure PCR Template Preparation Kit, an in-house protocol using the QIAamp DNA Blood Mini Kit, the CST genomic DNA purification kit, the Magna Pure LC, the MDx, the M48, the EZ1 and an in-house protocol using magnetic beads for manual and automated extraction were the methods that were able to reliably detect foetal RHD. The best results were obtained with the QIAamp DSP Virus Kit. The QIAamp DNA Blood Mini Kit showed very comparable results in laboratories that followed the manufacturer's protocol and started with ≥ 500 µL plasma. One participant using the QIAamp DNA Blood Midi Kit failed to detect reliably RHD in pool 3. Conclusions This workshop initiated a standardization process for extraction of cff DNA in maternal plasma. The highest yield was obtained by the QIAamp DSP Virus Kit, a result that will be evaluated in more detail in future studies. Copyright © 2007 John Wiley & Sons, Ltd.     

Economy-wide impacts of climate change: a joint analysis for sea level rise and tourism

While climate change impacts on human life have well defined and different origins, the interactions among the diverse impacts are not yet fully understood. Their final effects, however, especially those involving social-economic responses, are likely to play an important role. This paper is one of the first attempts to disentangle and highlight the role of these interactions. It focuses on the economic assessment of two specific climate change impacts: sea-level rise and changes in tourism flows. By using a Computable General Equilibrium (CGE) model the two impacts categories are first analysed separately and then jointly. Considered separately, in 2050, the forecasted 25 cm. of sea level rise imply a GDP loss ranging from (−) 0.1% in South East Asia to almost no loss in Canada, while redistribution of tourism flows – which in terms of arrivals favours Western Europe, Japan, Korea and Canada and penalises all the other world regions – triggers GDP losses ranging from (−) 0.5% in Small Island States to (−) 0.0004% in Canada. GDP gainers are Australia, New Zealand, Western Europe, Middle East and South Asia. The impact of sea level rise and tourism were simulated jointly and the results compared with those of the two disjoint simulations. From a qualitative point of view, the joint effects are similar to the outcomes of the disjoint exercises; from a quantitative perspective, however, impact interaction does play a significant role. In six cases out of 16 there is a detectable (higher than 2% and peaking to 70%) difference between the sum of the outcomes in the disjoint simulation and the outcomes of the joint simulations. Moreover, the relative contribution of each single impact category has been disentangled from the final result. In the case under scrutiny, demand shocks induced by changes in tourism flows outweigh the supply-side shock induced by the loss of coastal land.

Comparison the effects of bioaugmentation versus biostimulation on marine microbial community by PCR–DGGE: A mesocosm scale

In order to better understand the effects of biostimulation and bioaugmentation processes on a marine microbial community, three different mesocosm experiments were planned. Natural seawater(10.000 L) was artificially polluted with crude oil(1 L) and(1) inorganic nutrients(Biostimulating Mesocosm, BM),(2) inorganic nutrients and an inoculum of Alcanivorax borkumensis SK2(Single Bioaugmentation Mesocosm, SBM),(3) inorganic nutrients and inoculums of A. borkumensis SK2 and Thalassolituus oleivorans MIL-1(Consortium Bioaugmentation Mesocosm, CBM). During the experimental period(20 days), samples were taken from each mesocosm and the community structure was analyzed by PCR–DGGE. The 16 S r RNA gene DGGE banding patterns and sequence analysis demonstrated that biostimulation had the lowest effect on microbial biodiversity in the mesocosms; however, the biodiversity of the marine microbial community dramatically decreased in the CBM(Shannon index was 0.6 in T3). The community structures among the three mesocosms were also markedly different,and major bacteria derived from DGGE bands were related to uncultured Gamma Proteobacteria. The biodegradation results show that the Single Bioaugmentation Mesocosm(SBM) system had the highest percentage of degradation(95%) in comparison to the BM mesocosm(80%) and CBM(70%).     

Applying the dark diversity concept to nature conservation

Linking diversity to biological processes is central for developing informed and effective conservation decisions. Unfortunately, observable patterns provide only a proportion of the information necessary for fully understanding the mechanisms and processes acting on a particular population or community. We suggest conservation managers use the often overlooked information relative to species absences and pay particular attention to dark diversity (i.e., a set of species that are absent from a site but that could disperse to and establish there, in other words, the absent portion of a habitat‐specific species pool). Together with existing ecological metrics, concepts, and conservation tools, dark diversity can be used to complement and further develop conservation prioritization and management decisions through an understanding of biodiversity relativized by its potential (i.e., its species pool). Furthermore, through a detailed understanding of the population, community, and functional dark diversity, the restoration potential of degraded habitats can be more rigorously assessed and so to the likelihood of successful species invasions. We suggest the application of the dark diversity concept is currently an underappreciated source of information that is valuable for conservation applications ranging from macroscale conservation prioritization to more locally scaled restoration ecology and the management of invasive species.     

Relevance of wound-activated compounds produced by diatoms as toxins and infochemicals for benthic invertebrates

Plants evolve the production of wound-activated compounds (WACs) to reduce grazing pressure. In addition, several plant-produced WACs are recognized by various invertebrates, playing the role of infochemicals. Due to co-evolutionary processes, some invertebrates recognize plant infochemicals and use them to identify possible prey, detect the presence of predators or identify algae containing various classes of toxic metabolites. Different metabolites present in the same algae can play the role of toxins, infochemicals or both simultaneously. We investigated the infochemical activity of compounds extracted from three diatoms epiphytes of the seagrass Posidonia oceanica, by conducting choice experiments on invertebrates living in the same community or in close proximity. Furthermore, the specific toxicity of the extracts obtained from the same algae was tested on sea urchin embryos using a standard bioassay procedure, to detect the presence of toxins. The comparison of the two effects demonstrated that invertebrates are subjected to diatom wound-activated toxicants when these algae are not associated with their own habitat, but they are able to recognize volatile infochemicals derived from diatoms associated with their habitats. The specific toxicity of WACs was shown to be inversely correlated to the perceptive ability of invertebrates towards volatile compounds liberated by the same algae. Hence, when the recognition of specific algae by a given invertebrate species evolves, their detrimental effects on the receiving organism may be lost.     

Males are faster foragers than females: intersexual differences of foraging behaviour in the Apennine chamois

Availability of food resources and individual characteristics can influence foraging behaviour, which can differ between males and females, leading to different patterns of food/habitat selection. In dimorphic species, females are usually more selective in food choice, show greater bite rates and spend more time foraging than males. We evaluated sexual differences in foraging behaviour in Apennine chamois Rupicapra pyrenaica ornata, during the warm season, before the rut. Both sexes selected nutritious vegetation patches and spent a comparable amount of time feeding. However, males had a significantly greater feeding intensity (bite rate) and a lower search effort for feeding (step rate), as well as they spent more time lying down than females. Females selected foraging sites closer to refuge areas than males. In chamois, sexual size dimorphism is seasonal, being negligible in winter–spring, but increasing to 30–40 % in autumn. Our results suggest that males enhance their energy and mass gain by increasing their food intake rate during the warm season, to face the costs of the mating season (November). Conversely, females seem to prioritize a fine-scale selection of vegetation and the protection of offspring. A great food intake rate of males in the warm season could have developed as a behavioural adaptation leading herbivores to the evolutionary transition from year-round monomorphism to permanent dimorphism, through seasonal dimorphism.     

Differences in olfactory species recognition in the females of two Australian songbird species

Although birds have recently been shown to possess olfactory abilities and to use chemical cues in communication, limited effort has been made to demonstrate the use of odorants in social contexts. Even less is known regarding the use of odorants in species recognition. The ability to recognize conspecifics should be more pronounced in social species. This study investigated the importance of olfactory cues in species recognition in females of two estrildid finch species with different levels of sociality. Combining odor preference tests with chemical analyses, we surveyed whether female zebra finches and diamond firetails are able to distinguish between the species based on volatile traits and whether individuals exhibit species-specific differences in body odorants. Zebra finches are more social than diamond firetails; nevertheless, both species have an overlapping distribution area. Applying an experimental Y-maze paradigm, we showed that zebra finches can use differences in their species odor fingerprints and displayed a significant preference for the odor of conspecifics over that of heterospecifics, whereas diamond firetails did not reveal a preference. Using gas chromatography and mass spectrometry, we demonstrated that body odorants of the two species were significantly different in relative composition. This finding demonstrates the potential importance of olfactory cues in species recognition, at least in social bird species. Even these two closely related species displayed remarkable differences in their responsiveness to similar chemical cues, which might be caused by species-specific differences in ecology, physiology, or evolution.     

The effect of larval and early adult experience on behavioural plasticity of the aphid parasitoid <Emphasis Type="Italic">Aphidius ervi</Emphasis> (Hymenoptera,Braconidae, Aphidiinae)

The relevance of the integration of preimaginal and eclosion experiences on the subsequent habitat preferences and mate finding by the adult has been rarely tested in holometabolous insects. In this work, the effect of larval and early adult experiences on the behavioural responses of adult males of the aphid parasitoid, Aphidius ervi, towards volatiles from the host–plant complex (HPC) and from conspecific females were evaluated. Two experience factors were considered: host diet (normal diet=ND; artificial diet=AD), and eclosion, i.e. extraction or non-extraction of the parasitoid larva from the parasitised aphid (extracted=EX; non-extracted=NE). Thus, four treatments were set up: ND/NE, ND/EX, AD/NE and AD/EX. Glass Y-tube olfactometers were used to investigate the responses of adult A. ervi males to the odour sources used. Males from the ND/NE treatment showed a shorter latency to the first choice of olfactometer arms, displayed a marked preference towards the HPC olfactometer arm, and spent more time in the HPC arm than males from the other treatments. Only the interaction of host diet and eclosion experiences proved to be relevant in explaining the differences in latency to first choice, time spent in olfactometers arms, and behaviours displayed in the olfactometer arms. These results show the importance of the integration of larval and eclosion experiences in the development of stereotyped responses of the adults. This process may involve memory retention from the preimaginal and emergence period, but further research is needed to disentangle the contribution of each stage. The response to conspecific females was much less affected by the treatments in relation to first arm choice and times in olfactometer arms, suggesting a pheromone-mediated behaviour, even though a prompter and more intense wing fanning courtship behaviour was registered in the ND/NE males compared to males from the AD/NE treatment. These results show that sexual behaviours are less affected by early experiences than behaviours related with finding a HPC for foraging or oviposition. Taken together, our results demonstrate for first time that larval and eclosion experiences affect in a differential way the early responses of the adult towards environment-derived cues and mate related cues.     

Organochlorine pesticides and polychlorinated biphenyls in common buzzard (<Emphasis Type="Italic">Buteo buteo</Emphasis>) from Sicily (Italy)

In the present study, we investigated the concentrations and distribution of organochlorine pesticides (OCs) and polychlorinated biphenyls (PCBs) in intestine, liver, and muscle samples of 11 common buzzards (Buteo buteo) from Sicily used as bioindicator for monitoring pollution in environment. All samples of common buzzards were collected at the “Recovery Center of Wild Fauna” of Palermo, through the Zooprophilactic Institute. Quantitative determination of OCs and PCBs in the samples examined has been carried out using HRGC-ECD and GC-MS. The results obtained show the presence of concentrations of ∑DDT and ∑PCB in almost all samples. Regarding ∑DDT (4,4^′-DDE, 2,4^′-DDD, 4,4^′-DDD, 2,4^′-DDT, and 4,4^′-DDT), the highest concentrations were found in intestine (0.411 ± 0.050 μg/g) followed by muscle (0.130 ± 0.017 μg/g) and liver samples (0.109 ± 0.014 μg/g). As regards the ∑PCB congeners (PCB-28, PCB-52, PCB-95, PCB-99, PCB-101, PCB-110, PCB-138, PCB-146, PCB-149, PCB-151, PCB-153, PCB-170, PCB-177, PCB-180, PCB-183, and PCB-187), the highest concentrations were found in intestine (1.686 ± 0.144 μg/g) followed by liver (1.064 ± 0.162 μg/g) and muscle samples (0.797 ± 0.078 μg/g). Our data deserve particular attention not only for their significance but especially because they were recorded in Sicily, a region with a very low risk of environmental pollution due to the shortage of industries.