Direct PCR of indigenous and invasive mosquito species: a time- and cost-effective technique of mosquito barcoding.

Millions of people die each year as a result of pathogens transmitted by mosquitoes. However, the morphological identification of mosquito species can be difficult even for experts. The identification of morphologically indistinguishable species, such as members of the Anopheles maculipennis complex (Diptera: Culicidae), and possible hybrids, such as Culex pipiens pipiens/Culex pipiens molestus (Diptera: Culicidae), presents a major problem. In addition, the detection and discrimination of newly introduced species can be challenging, particularly to researchers without previous experience. Because of their medical importance, the clear identification of all relevant mosquito species is essential. Using the direct polymerase chain reaction (PCR) method described here, DNA amplification without prior DNA extraction is possible and thus species identification after sequencing can be achieved. Different amounts of tissue (leg, head; larvae or adult) as well as different storage conditions (dry, ethanol, -20 and -80 °C) and storage times were successfully applied and showed positive results after amplification and gel electrophoresis. Overall, 28 different indigenous and non-indigenous mosquito species were analysed using a gene fragment of the COX1 gene for species differentiation and identification by sequencing this 658-bp fragment. Compared with standard PCR, this method is time- and cost-effective and could thus improve existing surveillance and control programmes.

Genetic diversity and population structure of Mongolian domestic Bactrian camels (Camelus bactrianus).

The tradition of animal husbandry in the context of a nomadic lifestyle has been of great significance in the Mongolian society. Both Bactrian camels and horses have been invaluable for the survival and development of human activities in the harsh arid environment of the Mongolian steppe. As camels offer unique and sustainable opportunities for livestock production in marginal agro-ecological zones, we investigated the current genetic diversity of three local Mongolian camel breeds and compared their levels of variation with common native Mongolian camels distributed throughout the country. Based on mitochondrial and nuclear markers, we found levels of genetic diversity in Mongolian populations similar to that reported for Chinese Bactrian camels and for dromedaries. Little differentiation was detected between single breeds, except for a small group originating from the northwestern Mongolian Altai. We found neither high inbreeding levels in the different breeds nor evidence for a population decline. Although the Mongolian camel census size has severely declined over the past 20 years, our analyses suggest that there still exists a stable population with adequate genetic variation for continued sustainable utilization.

High mitochondrial differentiation levels between wild and domestic Bactrian camels: a basis for rapid detection of maternal hybridization.

Hybridization between wild species and their domestic congeners often threatens the gene pool of the wild species. The last wild Bactrian camel (Camelus ferus) populations in Mongolia and China are examples of populations facing such a hybridization threat. To address this key issue in the conservation of wild camels, we analysed wild, hybrid and domestic Bactrian camels (Camelus bactrianus) originating from Mongolia, China and Austria. Through screening of an 804-base-pair mitochondrial fragment, we identified eight mitochondrial haplotypes and found high sequence divergence (1.9%) between C. ferus and C. bactrianus. On the basis of a mitochondrial DNA sequence fixed difference, we developed a diagnostic PCR restriction fragment length polymorphism (PCR-RFLP) assay to differentiate between wild and domestic camel samples. We applied the assay to 81 individuals and confirmed the origin of all samples including five hybrids with known maternal ancestry. The PCR-RFLP system was effective for both traditional (blood, skin) and non-invasive samples (faeces, hair), as well as for museum specimens. Our results demonstrate high levels of mitochondrial differentiation between wild and domestic Bactrian camels and that maternal hybridization can be detected by a rapid and reliable PCR-RFLP system.