Vector Biological Studies in Leishmaniasis

我们以前提供了确凿的证据，证明利什曼原虫的无脊椎阶段在它们在沙蝇媒介的生长和发育过程中完全能够有性循环。我们2012年研究的重点是将性能力的发现扩展到利什曼原虫的其他物种和毒株以及其他沙蝇媒介物种。我们已经能够从长掌Lutzomyia longipalpis中恢复L. major杂交种，这表明在新大陆发现的主要沙蝇属能够支持它们传播的利什曼原虫的有性生殖周期。杂种在感染后3天就恢复了，这表明早期发育阶段，如原环和/或nectomonad promastigotes，能够交配。我们还成功地将来自西非、中东和俄罗斯南部的L. major子菌株进行了杂交，证明了在其整个地理分布范围内的L. major分离株都具有性能力。此外，我们还首次成功地培育出了大乳杆菌和小乳杆菌的种间杂交品种。通过对300多个菌株的多位点基因分型，证实了自然种群中存在种内和种间等位基因杂交。

We have previously provided conclusive evidence that the invertebrate stages of Leishmania are fully capable of a sexual cycle during their growth and development in the sand fly vector. The priority of our studies in 2012 has been to extend the findings of sexual competence to other species and strains of Leishmania and to additional sand fly vector species. We have been able to recover L. major hybrids from Lutzomyia longipalpis, demonstrating that the main genus of sand flies found in the New World are capable of supporting the sexual reproductive cycle of the Leishmania parasites that they transmit. Hybrids were recovered as early as three days post infection, indicating that early developmental stages, e.g. procyclic and/or nectomonad promastigotes, are able to mate. We have also been able to successfully cross L. major sub-strains from West Africa, the Middle East, and Southern Russia, demonstrating that L. major isolates from across the full range of their geographic distribution are sexually competent. Furthermore, we have been able for the first time to generate interspecies hybrids between L. major and L. infantum. The occurrence of intra- and inter-species allelic hybrids amongst natural populations has been substantiated by multi-locus genotyping of over 300 strains. The differentiation of midgut promastigotes into the infectious, metacyclic stage is essential for the development of transmissible infections. We have been able to predict transmission success based on the efficiency of this differentiation process in the anterior midgut. Meta-analysis of multiple transmission experiments allowed us to establish a percent-metacyclic cutoff value that strongly predicted transmission competence. We have also identified the normal midgut microbiome as an essential factor controlling metacyclogenesis. L. major infected P. duboscqi sand flies treated with penicillin/streptomycin were completely inhibited in their ability to support the normal differentiation of promastigotes into the metacyclic stage. The bacterial diversity in normal and antibiotic treated flies was characterized by both culture dependent and independent methods by sequencing of the 16s rDNA. The possible roles of the normal bacterial communities in providing nutritional supplements essential to parasite differentiation, or displacement of the normal midgut microbiota by communities that are harmful to the parasite, will be investigated in future studies.