Phylogenetics of the Old World screwworm fly and its significance for planning control and monitoring invasions in Asia.

Phylogenetic, genealogical and population relationships of Chrysomya bezziana, the Old World screwworm fly (OWSF), were inferred from DNA sequences of mitochondrial cytochrome b (cyt b), nuclear elongation factor-1α (EF-1α) and nuclear white eye colour (white), using sequences of Chrysomya megacephala and Chrysomya rufifacies as outgroups. Cyt b (717bp, 754 specimens), EF-1α (361bp, 256 specimens) and white (577bp, 242 specimens) were analysed from up to two African and nine Asian countries, including 10 Indonesian islands. We show that OWSF occurs as distinctive African and Asian lineages based on cyt b and white, and that there is a marked differentiation between Sumatran and Javan populations in Indonesia, supported by the genealogy and analysis of molecular variance of cyt b alone. Four cyt b sub-lineages are recognised in Asia: only 2.1 occurs on the Asian mainland, from Yemen to Peninsular Malaysia; only 2.2, 2.3 and 2.4 occur in central Indonesia; 2.4 predominates on New Guinea; and 2.1 co-occurs with others only on Sumatra in western Indonesia. This phylogeography and the genetic distances between cyt b haplotypes indicate pre-historic, natural dispersal of OWSF eastwards into Indonesia and other Malesian islands, followed by vicariant evolution in New Guinea and central Indonesia. OWSF is absent from Australia, where there is surveillance for importation or natural invasion. Judged by cyt b haplotype markers, there is currently little spread of OWSF across sea barriers, despite frequent shipments of Australian livestock through Indonesian seas to the Middle East Gulf region. These findings will inform plans for integrated pest management, which could be applied progressively, for example starting in East Nusa Tenggara (central Indonesia) where OWSF has regional cyt b markers, and progressing westwards to Java where any invasion from Sumatra is unlikely. Cyt b markers would help identify the source of any re-emergence in treated areas.

Multiple genetic divergences and population expansions of a Mediterranean sandfly, Phlebotomus ariasi, in Europe during the Pleistocene glacial cycles.

Phlebotomus ariasi is one of the two sandflies transmitting the causative agent of zoonotic leishmaniasis, Leishmania infantum, in France and Iberia, and provides a rare case study of the postglacial re-colonization of France by a Mediterranean species. Four DNA sequences were analysed-mitochondrial cytochrome b (cyt b), nuclear elongation factor-1α (EF-1α) and two anonymous nuclear loci-for 14-15 French populations and single populations from northeast Spain, northwest Spain, Portugal and Morocco. The presence of cryptic sibling species was not revealed by phylogenetic analyses and testing for reproductive isolation between sympatric populations defined by the two most divergent cyt b haplogroups. No locus was shown to be under positive directional or balancing selection and, therefore, molecular variation was explained demographically. Each nuclear locus showed shallow isolation by distance from Portugal to the French Pyrenees, but for both cyt b and EF-1α there was then a step change to the upland Massif Central, where leading-edge populations showed low diversity at all loci. Multiple genetic divergences and population expansions were detected by analyses of cyt b and dated to the Pleistocene. Endemicity of one cyt b sub-lineage suggested the presence of a refuge north of the Pyrenees during the last glacial period. Monopolization of the Massif Central by genetically differentiated populations of P. ariasi might possibly hinder the northwards spread of leishmaniasis.

Phylogeography of Androctonus species (Scorpiones: Buthidae) in Tunisia: Diagnostic characters for linking species to scorpionism.

A fragment of the mitochondrial (mt) 16S ribosomal RNA gene was amplified by PCR and sequenced from individual adult scorpions of the genus Androctonus, which were sampled from central and southern Tunisia and identified using an explicit set of morphological characters. Phylogenetic analyses placed the mtDNA haplotypes in three well-supported monophyletic lineages, corresponding to the morphospecies Androctonusaeneas, Androctonusamoreuxi and Androctonusaustralis. The latter species was the most abundant and widespread, and it was characterized by two mtDNA sub-lineages each of which predominated only north or south of the Chott el Jerid, a seasonally flooded saline depression that divides non-Mediterranean Tunisia. The divergence of the two mtDNA lineages was dated by mtDNA molecular clocks, indicating that the formation of the Chott el Jerid is unlikely to have been the barrier generating the vicariant evolution of the two lineages of A. australis, although it may have impeded their mixing following secondary contact. Both regional mtDNA lineages were found in A. australis hector and A. australisgarzonii, indicating that these two morphological forms are neither monophyletic nor geographically isolated and, therefore, should not be treated as species or subspecies. It is recommended that no subspecies of A. australis should be recognized in North Africa and toxicologists should cease the taxonomic error of referring to a species "Androctonus australis Hector". The morphological form "hector" has no proven association with an increased risk of scorpionism compared with "garzonii". However, it might be prudent to produce anti-venom in Tunisia by using both morphological forms of A. australis collected each side of the Chott el Jerid, because of the evidence for regional variation in toxins. The highest risk for scorpion stings occurs in the central region, where the new diagnostic markers should be used to discover any association between Androctonus species and scorpionism.

Postglacial dispersal of Phlebotomus perniciosus into France.

Phlebotomus perniciosus was identified morphologically in samples from France and northeast Spain, and individuals were then characterized at three polymorphic isoenzyme loci (by isoelectrofocusing) and at the mitochondrial DNA locus (by comparative DNA sequence analysis of a fragment of the Cytochrome b gene). The four polymorphic loci gave conflicting patterns of population relationships, which can be explained by hypothesizing different amounts of gene introgression at each locus when two distinctive lineages met in southern France or northeast Spain after isolation in southern Italy and Spain during the Pleistocene Ice Ages. P. perniciosus is an important vector of leishmania infantum and so these population differentiation studies are relevant for predicting the emergence and spread of leishmaniasis in relation to environmental changes, including climate.