SummaryIn animals and yeast, voltage-dependent chloride channels of the CLC family play a role in basic cellular functions such as epithelial transport, plasma membrane excitability, and control of pH and membrane potential in intracellular compartments. To assess the function of CLCs in plants, we searched for CLC insertion mutants in a library of Arabidopsis lines transformed by Agrobacterium tumefaciens transferred DNA (T-DNA). Using a polymerase chain reaction-based screening procedure, an Arabidopsis line that carries a T-DNA insertion within the C-terminus of the AtCLC-a coding sequence was identi®ed. Progeny from this plant line, clca-1, showed dramatically altered transcription of the AtCLC-a gene. Plants homozygous for the clca-1 mutation exhibited normal development and a morphology indistinguishable from the wild-type. However, their capacity to accumulate nitrate under conditions of nitrate excess was reduced in roots and shoots, by approximately 50%, while chloride, sulphate and phosphate levels were similar to the wild-type. In addition, the herbicide chlorate, an analogue of nitrate, induced a faster and more pronounced chlorosis in mutant plants. Hypersensitivity to chlorate as well as decreased nitrate levels co-segregated with the T-DNA insertion. They were found at various time points of the clca-1 life cycle, supporting the idea that AtCLC-a has a general role in the control of the nitrate status in Arabidopsis. Concordant with such a function, AtCLC-a mRNA was found in roots and shoots, and its levels rapidly increased in both tissues upon addition of nitrate but not ammonium to the culture medium. The speci®city of AtCLC-a function with respect to nitrate is further supported by a similar free amino acid content in wild-type and clca-1 plants. Although the cellular localization of AtCLC-a remains unclear, our results suggest that AtCLC-a plays a role in controlling the intracellular nitrate status.

Quarantine measures to prevent insect invasions tend to focus on well-known pests but a large proportion of the recent invaders were not known to cause significant damage in their native range, or were not even known to science before their introduction. A novel method is proposed to detect new potential pests of woody plants in their region of origin before they are introduced to a new continent. Since Asia is currently considered to be the main supplier of insect invaders to Europe, sentinel trees were planted in China during 2007-2011 as an early warning tool to identify the potential for additional Asian insect species to colonize European trees. Seedlings (1-1.5 m tall) of five broadleaved (Quercus petraea, Q. suber, Q. ilex, Fagus sylvatica, and Carpinus betulus) and two conifer species (Abies alba and Cupressus sempervirens) were planted in blocks of 100 seedlings at two widely separated sites (one in a nursery near Beijing and the other in a forest environment near Fuyang in eastern China), and then regularly surveyed for colonization by insects. A total of 104 insect species, mostly defoliators, were observed on these new hosts, and at least six species were capable of larval development. Although a number of the insects observed were probably incidental feeders, 38 species had more than five colonization events, mostly infesting Q. petraea, and could be considered as being capable of switching to European trees if introduced to Europe. Three years was shown to be an appropriate duration for the experiment, since the rate of colonization then tended to plateau. A majority of the identified species appeared to have switched from agricultural crops and fruit trees rather than from forest trees. Although these results are promising, the method is not appropriate for xylophagous pests and other groups developing on larger trees. Apart from the logistical problems, the identification to species level of the specimens collected was a major difficulty. This situation could be improved by the development of molecular databases.

Tissue culture has been shown to induce the transposition of plant transposable elements; their insertion at novel sites results in somaclonal variation. Introduction of the tobacco retrotransposon Tnt1 into Arabidopsis thaliana by co-cultivation of root explants with Agrobacterium tumefaciens induces its transposition at a high frequency, but no transposed copies are found in plants transformed by the in planta procedure. Transposition occurs in the transformed root cells or in the calli derived from them, allowing the regeneration of transformed plants with up to 26 transposed copies of Tnt1. Analysis of Tnt1 integration sites in Arabidopsis shows that the Tnt1 endonuclease does not show any cleavage-site specificity at the sequence level. The insertion sites are unlinked and distributed on all five Arabidopsis chromosomes. The fact that the majority of the integration sites are located in coding regions, and none in repeated sequences, demonstrates the potential of Tnt1 as a tool for gene tagging.

Retracing the routes of invasions and determining the origins of invading species is often critical in understanding biological invasions. The Western conifer seed bug, Leptoglossus occidentalis, an insect native of western North America, was first accidentally introduced to eastern North America and then to Europe. The colonization of the entire European continent occurred in ca. 10-15 years, probably promoted by independent introductions in different parts of Europe. A multi-marker approach (mtDNA and microsatellites) combined with approximate Bayesian computation analyses was used to track the origin of European populations and to determine whether this rapid invasion was caused by multiple introductions. Our results show that at least two independent introductions of L. occidentalis have occurred in Europe. Moreover, the analyses showed a stronger genetic similarity of European invasive populations with the eastern North American populations than with those of the native range, suggesting that invasive North American population acted as a bridgehead for European invasion. The results also revealed that natural dispersal as well as human-mediated transportations as hitchhikers probably enhanced the rapid spread of this invasive pest across Europe. This study illustrates the complexity of a rapid invasion and confirms that bridgehead and multiple introductions have serious implications for the success of invasion.