AbstractMass releases of sterilized male insects, in the frame of sterile insect technique programs, have helped suppress insect pest populations since the 1950s. In the major horticultural pests Bactrocera dorsalis, Ceratitis capitata, and Zeugodacus cucurbitae, a key phenotype white pupae (wp) has been used for decades to selectively remove females before releases, yet the gene responsible remained unknown. Here we use classical and modern genetic approaches to identify and functionally characterize causal wp− mutations in these distantly related fruit fly species. We find that the wp phenotype is produced by parallel mutations in a single, conserved gene. CRISPR/Cas9-mediated knockout of the wp gene leads to the rapid generation of novel white pupae strains in C. capitata and B. tryoni. The conserved phenotype and independent nature of the wp− mutations suggest that this technique can provide a generic approach to produce sexing strains in other major medical and agricultural insect pests.

BackgroundRecent advances in genomics have addressed the challenge that divergent haplotypes pose to the reconstruction of haploid genomes. However for many organisms, the sequencing of either field-caught individuals or a pool of heterogeneous individuals is still the only practical option. Here we present methodological approaches to achieve three outcomes from pooled long read sequencing: the generation of a contiguous haploid reference sequence, the sequences of heterozygous haplotypes; and reconstructed genomic sequences of individuals related to the pooled material. Results PacBio long read sequencing, DovetailHi-C scaffolding and linkage map integration yielded a haploid chromosome-level assembly for the diamondback moth (Plutella xylostella), a global pest of Brassica crops, from a pool of related individuals. The final assembly consisted of 573 scaffolds, with a total assembly size of 343.6Mbp a scaffold N50 value of 11.3Mbp (limited by chromosome size) and a maximum scaffold size of 14.4Mbp. This assembly was then integrated with an existing RAD-seq linkage map, anchoring 95% of the assembled sequence to defined chromosomal positions. ConclusionsWe describe an approach to resolve divergent haplotype sequences and describe multiple validation approaches. We also reconstruct individual genomes from pooled long-reads, by applying a recently developed k-mer binning method.

18Although there are many honey bee pathogens/parasites, it is still not understood how 19 they change their gene expression to adapt to the host environment or how the host 20 simultaneously responds to pathogen/parasite infection by modifying its own gene 21 expression. Such interactions must lead to changes in the physiological states of both 22 host and parasite. To address this question, we studied a trypanosomatid, 23 Lotmaria passim, which can be cultured in medium and inhabit the honey bee hindgut. 24 We found that L. passim dynamically modifies the expression of mRNAs associated 25 with protein translation and the electron transport chain to adapt to the anaerobic and 26 nutritionally poor honey bee hindgut at early stages of infection, and to become dormant 27 at late stages of infection. Meanwhile, several genes are continuously up-or 28 down-regulated during infection, including GP63 as well as genes coding for host cell 29 signaling pathway modulators (up-regulated), and those involved in detoxification of 30 radical oxygen species as well as flagellar formation (down-regulated). L. passim 31 infection only slightly increases honey bee mortality and does not affect the number of 32 microorganisms in the gut microbiota; but it induces honey bee innate immune response. 33 Upon infection, the host appears to be in poor nutritional status, indicated by the 34 increase in the levels of mRNAs for take-out and facilitated trehalose transporter and 35 the decrease of vitellogenin mRNA level. Simultaneous gene expression profiling of L. 36 passim and honey bee during infection provided insight into how both parasite and host 37 modify their gene expressions. This study presents one of the best models to understand 38 host-parasite interactions at the molecular and cellular levels in honey bee. 39 40 41 42