Background: The olive fruit fly, Bactrocera oleae, is the most important pest in the olive fruit agribusiness industry. This is because female flies lay their eggs in the unripe fruits and upon hatching the larvae feed on the fruits thus destroying them. The lack of a high-quality genome and other genomic and transcriptomic data has hindered progress in understanding the fly’s biology and proposing alternative control methods to pesticide use. Results: Genomic DNA was sequenced from male and female Demokritos strain flies, maintained in the laboratory for over 45 years. We used short-, mate-pair-, and long-read sequencing technologies to generate a combined male-female genome assembly (GenBank accession GCA_001188975.2). Genomic DNA sequencing from male insects using 10x Genomics linked-reads technology followed by mate-pair and long-read scaffolding and gap-closing generated a highly contiguous 489 Mb genome with a scaffold N50 of 4.69 Mb and L50 of 30 scaffolds (GenBank accession GCA_001188975.4). RNA-seq data generated from 12 tissues and/or developmental stages allowed for genome annotation. Short reads from both males and females and the chromosome quotient method enabled identification of Y-chromosome scaffolds which were extensively validated by PCR. Conclusions: The high-quality genome generated represents a critical tool in olive fruit fly research. We provide an extensive RNA-seq data set, and genome annotation, critical towards gaining an insight into the biology of the olive fruit fly. In addition, elucidation of Y-chromosome sequences will advance our understanding of the Y-chromosome’s organization, function and evolution and is poised to provide avenues for sterile insect technique approaches.

The Bactrocera dorsalis species complex currently harbors approximately 90 different members. The species complex has undergone many revisions in the past decades, and there is still an ongoing debate about the species limits. The availability of a variety of tools and approaches, such as molecular-genomic and cytogenetic analyses, are expected to shed light on the rather complicated issues of species complexes and incipient speciation. The clarification of genetic relationships among the different members of this complex is a prerequisite for the rational application of sterile insect technique (SIT) approaches for population control.

Background: The Tephritidae family of insects includes the most important agricultural pests of fruits and vegetables, belonging mainly to four genera (Bactrocera, Ceratitis, Anastrepha and Rhagoletis). The olive fruit fly, Bactrocera oleae, is the major pest of the olive fruit. Currently, its control is based on chemical insecticides. Environmentally friendlier methods have been attempted in the past (Sterile Insect Technique), albeit with limited success. This was mainly attributed to the lack of knowledge on the insect's behaviour, ecology and genetic structure of natural populations. The development of molecular markers could facilitate the access in the genome and contribute to the solution of the aforementioned problems. We chose to focus on microsatellite markers due to their abundance in the genome, high degree of polymorphism and easiness of isolation. Results: Fifty-eight microsatellite-containing clones were isolated from the olive fly, Bactrocera oleae, bearing a total of sixty-two discrete microsatellite motifs. Forty-two primer pairs were designed on the unique sequences flanking the microsatellite motif and thirty-one of them amplified a PCR product of the expected size. The level of polymorphism was evaluated against wild and laboratory flies and the majority of the markers (93.5%) proved highly polymorphic. Thirteen of them presented a unique position on the olive fly polytene chromosomes by in situ hybridization, which can serve as anchors to correlate future genetic and cytological maps of the species, as well as entry points to the genome. Cross-species amplification of these markers to eleven Tephritidae species and sequencing of thirty-one of the amplified products revealed a varying degree of conservation that declines outside the Bactrocera genus. Conclusion: Microsatellite markers are very powerful tools for genetic and population analyses, particularly in species deprived of any other means of genetic analysis. The presented set of microsatellite markers possesses all features that would render them useful in such analyses. This could also prove helpful for species where SIT is a desired outcome, since the development of effective SIT can be aided by detailed knowledge at the genetic and molecular level. Furthermore, their presented efficacy in several other species of the Tephritidae family not only makes them useful for their analysis but also provides tools for phylogenetic comparisons among them.

Ceratitis fasciventrisis a serious agricultural pest of the Tephritidae family that belongs to the AfricanCeratitisFAR species complex. Species limits within the FAR complex are obscure and multidisciplinary approaches have attempted to resolve phylogenetic relationships among its members. These studies support the existence of at least three additional species in the complex,C.anonnae,C.rosaandC.quilicii, while they indicate the presence of two structured populations (F1 and F2) within theC.fasciventrisspecies. In the present study we present the mitotic karyotype, polytene chromosome maps,in situhybridization data and the complete mitochondrial genome sequence of an F2 population ofC.fasciventris. This is the first polytene chromosome map and complete mitogenome of a member of the FAR complex and only the second reported for theCeratitisgenus. Both polytene chromosomes and mitochondrial sequence could provide valuable information and be used as reference for comparative analysis among the members of the complex towards the clarification of their phylogenetic relationships.

The marine turtle Caretta caretta is a widely distributed species that is facing critical population decline, especially in the Mediterranean rookeries. Molecular markers, such as mitochondrial DNA (mtDNA) sequences, are of great importance for the description and monitoring of turtle migratory populations. The complete sequence of the C. caretta mitochondrial genome is presented here. The genome comprises 16,440 base pairs, containing 37 genes (13 protein-coding genes, 22 tRNA genes, and 2 rRNA genes), and a control region, all organized similar to the majority of vertebrate mitogenomes. MtDNA length polymorphism and heteroplasmy were observed among, and within, individuals due to the variable size of a microsatellite repeat residing at the 3′ end of the control region. The use of the above repeat as a marker for individual fingerprinting is discussed. Furthermore, phylogenetic analyses among Testudines based on complete mitogenomes, as well as among marine turtles based on partial mtDNA sequences, are considered.

Bactrocera carambolae is one of the approximately 100 sibling species of the Bactrocera dorsalis complex and considered to be very closely related to B. dorsalis. Due to their high morphological similarity and overlapping distribution, as well as to their economic impact and quarantine status, the development of reliable markers for species delimitation between the two taxa is of great importance. Here we present the complete mitochondrial genome of B. carambolae sourced from its native range in Malaysia and its invaded territory in Suriname. The mitogenome of B. carambolae presents the typical organization of an insect mitochondrion. Comparisons of the analyzed B. carambolae sequences to all available complete mitochondrial sequences of B. dorsalis revealed several species-specific polymorphic sites. Phylogenetic analysis based on Bactrocera mitogenomes supports that B. carambolae is a differentiated taxon though closely related to B. dorsalis. The present complete mitochondrial sequences of B. carambolae could be used, in the frame of Integrative Taxonomy, for species discrimination and resolution of the phylogenetic relationships within this taxonomically challenging complex, which would facilitate the application of species-specific population suppression strategies, such as the sterile insect technique.