Impact of sample preservation and manipulation on insect gut microbiome profiling : a test case with fruit flies (Diptera, Tephritidae)

High-throughput sequencing (HTS) techniques are of great value for the investigation of microbial communities, and have been extensively used to study the gut microbiome. While most studies focus on the human gut, many others have investigated insects. However, because of the rapid spread of HTS techniques, a lot of variation exists in the protocols for sample preparation. In the present study, we investigated the impact of two widely adopted sample-processing procedures preceding library preparation, i.e., preservation of insect tissue in 70% ethanol (EtOH) and sample dissection. We used the fruit fly Ceratitis capitata (Diptera: Tephritidae) as a model organism and set up two experiments, one comparing the effects of sample manipulation and preservation across life stages and the other across fruit samples from different sources. The results of this study showed no major effects of dissection on the outcome of HTS. However, EtOH preservation did have effects on the recovered gut microbiome, the main effect being a significant reduction of the dominant genus, Providencia, in EtOH-preserved samples. Less abundant bacterial groups were also affected resulting in altered microbial profiles obtained from samples preserved in 70% EtOH. These results have important implications for the planning of future studies and when comparing studies that used different sample preparation protocols

Assessing diversity among traditional Greek and foreign eggplant cultivars using molecular markers and morphometrical descriptors

Eggplant is a widely cultivated vegetable crop of great economic importance. Its long lasting history of domestication, selection and breeding has led to the development of numerous cultivars with variable traits. In the present study, we assessed the diversity levels within and among eleven Greek and foreign cultivars, using 22 morphological descriptors and two different classes of molecular markers (retrotransposon microsatellite amplified polymorphism-REMAP markers and nuclear microsatellites). Our results, in accordance with other studies in the field showed: a) the limited levels of genetic polymorphism within the cultivars; b) the high morphological and genetic divergence existing among them as indicated by the genetic distance values calculated, which could be attributed to selection, inbreeding and bottleneck effects; and c) the lack of concordance among morphological descriptors and molecular markers. Despite these, our analysis showed that the utilization of combinations of markers is an effective method for the characterization of plant material providing also useful diagnostic tools for the identification and authentication of the selected Greek cultivars

The effects of geographic origin and antibiotic treatment on the gut symbiotic communities of <i>Bactrocera oleae</i> populations

The olive fruit fly, Bactrocera oleae (Rossi) (Diptera: Tephritidae), is the major insect pest of olive orchards (Olea europaea L.), causing extensive damages on cultivated olive crops worldwide. Due to its economic importance, it has been the target species for a variety of population control approaches including the sterile insect technique (SIT). However, the inefficiency of the current mass-rearing techniques impedes the successful application of area-wide integrated pest management programs with an SIT component. It has been shown that insect mass rearing and quality of sterile insects can be improved by the manipulation of the insect gut microbiota and probiotic applications. In order to exploit the gut bacteria, it is important to investigate the structure of the gut microbial community. In the current study, we characterized the gut bacterial profile of two wild olive fruit fly populations introduced in laboratory conditions using next generation sequencing of two regions of the 16S rRNA gene. We compared the microbiota profiles regarding the geographic origin of the samples. Additionally, we investigated potential changes in the gut bacteria community before and after the first exposure of the wild adult flies to artificial adult diet with and without antibiotics. Various genera - such as Erwinia, Providencia, Enterobacter, and Klebsiella - were detected for the first time in B. oleae. The most dominant species was Candidatus Erwinia dacicola Capuzzo et al. and it was not affected by the antibiotics in the artificial adult diet used in the first generation of laboratory rearing. Geographic origin affected the overall structure of the gut community of the olive fruit fly, but antibiotic treatment in the first generation did not significantly alter the gut microbiota community

Manipulation of insect gut microbiota towards the improvement of <i>Bactrocera oleae</i> artificial rearing

Bactrocera oleae (Rossi) (Diptera: Tephritidae) is the main pest of olive trees (Olea europaeaL.), causing major damages in olive crops. Improvement of mass rearing is a prerequisite for the successful development of large-scale sterile insect technique (SIT) applications. This can be achieved through the enrichment of artificial diets with gut bacteria isolates. We assessed the efficiency of three gut bacteria previously isolated fromCeratitis capitata(Wiedemann), and four isolated fromB. oleae, as larval diet additives in both live and inactivated/dead forms. Our results showed that deadEnterobactersp. AA26 increased pupal weight, whereas both live and dead cells increased pupal and adult production and reduced immature developmental time, indicating that its bacterial cells serve as a direct nutrient source. LiveProvidenciasp. AA31 improved pupal and adult production, enhanced male survival under stress conditions, and delayed immature development. DeadProvidenciasp. AA31, however, did not affect production rates, indicating that live bacteria can colonize the insect gut and biosynthesize nutrients essential for larval development. Live and deadBacillussp. 139 increased pupal weight, accelerated immature development, and increased adult survival under stress. Moreover, liveBacillussp. 139 improved adult production, indicating thatBacilluscells are a direct source of nutrients. DeadSerratiasp. 49 increased pupal and adult production and decreased male survival under stress conditions whereas live cells decreased insect production, indicating that the live strain is entomopathogenic, but its dead cells can be utilized as nutrient source.Klebsiella oxytoca,Enterobactersp. 23, andProvidenciasp. 22 decreased pupal and subsequent adult production and were harmful forB. oleae. Our findings indicate that deadEnterobactersp. AA26 is the most promising bacterial isolate for the improvement ofB. oleaemass rearing in support of future SIT or related population suppression programs

The impact of fruit fly gut bacteria on the rearing of the parasitic wasp <i>Diachasmimorpha longicaudata</i>

Area-wide integrated pest management strategies against tephritid fruit flies include the release of fruit fly parasitic wasps in the target area. Mass rearing of parasitic wasps is essential for the efficient application of biological control strategies. Enhancement of fruit fly host fitness through manipulation of their gut-associated symbionts might also enhance the fitness of the produced parasitic wasps and improve the parasitoid rearing system. In the current study, we added three gut bacterial isolates originating from Ceratitis capitata (Wiedemann) and four originating from Bactrocera oleae (Rossi) (both Diptera: Tephritidae) to the larval diet of C. capitata and used the bacteria-fed larvae as hosts for the development of the parasitic wasp Diachasmimorpha longicaudata (Ashmead) (Hymenoptera: Braconidae). We evaluated the effect of the bacteria on wasp life-history traits and assessed their potential use for the improvement of D. longicaudata rearing. Enterobactersp. AA26 increased fecundity and parasitism rate and accelerated parasitoid emergence. Providencia sp. AA31 led to faster emergence of both male and female parasitoids, whereas Providencia sp. 22 increased the production of female progeny. Bacillus sp. 139 increased parasitoid fecundity, parasitism rate, and production of female progeny. Serratia sp. 49 accelerated parasitoid emergence for both males and females and increased production of female progeny. Klebsiella oxytoca delayed parasitoid emergence and Enterobacter sp. 23 decreased parasitoid fecundity and parasitism rate. Our findings demonstrate a wide range of effects of fruit fly gut symbionts on parasitoid production and reveal a great potential of bacteria use towards enhancement of parasitic wasp rearing

Isolation and characterization of microsatellite markers from the olive fly, Bactrocera oleae, and their cross-species amplification in the Tephritidae family

<p>Abstract</p> <p>Background</p> <p>The Tephritidae family of insects includes the most important agricultural pests of fruits and vegetables, belonging mainly to four genera (<it>Bactrocera, Ceratitis, Anastrepha </it>and <it>Rhagoletis</it>). The olive fruit fly, <it>Bactrocera oleae</it>, 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.</p> <p>Results</p> <p>Fifty-eight microsatellite-containing clones were isolated from the olive fly, <it>Bactrocera oleae</it>, 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 <it>in situ </it>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 <it>Bactrocera </it>genus.</p> <p>Conclusion</p> <p>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.</p

Symbionts do not affect the mating incompatibility between the Brazilian-1 and Peruvian morphotypes of the Anastrepha fraterculus cryptic species complex

The South American fruit fly, Anastrepha fraterculus, is clearly undergoing a speciation process. Among others, two of their morphotypes, the Brazilian-1 and Peruvian, have accumulated differences in pre- and post-zygotic mechanisms resulting in a degree of reproductive isolation. Both harbor a different strain of Wolbachia, which is a widespread endosymbiotic bacterium among many invertebrates producing a range of reproductive effects. In this paper, we studied the role of this bacterium as one of the factors involved in such isolation process. Infected and cured laboratory colonies were used to test pre- and post-zygotic effects, with special emphasis in uni- and bi-directional cytoplasmic incompatibility (CI). We showed that Wolbachia is the only known reproductive symbiont present in these morphotypes. Wolbachia reduced the ability for embryonic development in crosses involving cured females and infected males within each morphotype (uni-directional CI). This inhibition showed to be more effective in the Peruvian morphotype. Bi-directional CI was not evidenced, suggesting the presence of compatible Wolbachia strains. We conclude that Wolbachia is not directly involved in the speciation process of these morphotypes. Other mechanisms rather than CI should be explored in order to explain the reduced mating compatibility between the Brazilian-1 and Peruvian morphotypes.Instituto de BiotecnologíaFil: Devescovi, Francisco. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Agrobiotecnología y Biología Molecular; Argentina. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Genética. Laboratorio de Genética de Insectos de Importancia Económica; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Conte, Claudia Alejandra. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Genética. Laboratorio de Genética de Insectos de Importancia Económica; ArgentinaFil: Augustinos, Antonios A. Vienna International Centre. Joint FAO/IAEA Division of Nuclear Techniques in Food and Agriculture. Insect Pest Control Laboratory; AustriaFil: Cancio Martinez, Elena I. Vienna International Centre. Joint FAO/IAEA Division of Nuclear Techniques in Food and Agriculture. Insect Pest Control Laboratory; AustriaFil: Segura, Diego Fernando. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Agrobiotecnología y Biología Molecular; Argentina. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Genética. Laboratorio de Genética de Insectos de Importancia Económica; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Caceres, Carlos. Vienna International Centre. Joint FAO/IAEA Division of Nuclear Techniques in Food and Agriculture. Insect Pest Control Laboratory; AustriaFil: Lanzavecchia, Silvia Beatriz. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Genética "Ewald A. Favret"; ArgentinaFil: Bourtzis, Kostas. Vienna International Centre. Joint FAO/IAEA Division of Nuclear Techniques in Food and Agriculture. Insect Pest Control Laboratory; Austri

Wolbachia pipientis associated with tephritid fruit fly pests: from basic research to applications

Members of the true fruit flies (family Tephritidae) are among the most serious agricultural pests worldwide, whose control and management demands large and costly international efforts. The need for cost-effective and environmentally friendly integrated pest management (IPM) has led to the development and implementation of autocidal control strategies. These approaches include the widely used sterile insect technique and the incompatible insect technique (IIT). IIT relies on maternally transmitted bacteria (namely Wolbachia) to cause a conditional sterility in crosses between released mass-reared Wolbachia-infected males and wild females, which are either uninfected or infected with a different Wolbachia strain (i.e., cytoplasmic incompatibility; CI). Herein, we review the current state of knowledge on Wolbachia-tephritid interactions including infection prevalence in wild populations, phenotypic consequences, and their impact on life history traits. Numerous pest tephritid species are reported to harbor Wolbachia infections, with a subset exhibiting high prevalence. The phenotypic effects of Wolbachia have been assessed in very few tephritid species, due in part to the difficulty of manipulating Wolbachia infection (removal or transinfection). Based on recent methodological advances (high-throughput DNA sequencing) and breakthroughs concerning the mechanistic basis of CI, we suggest research avenues that could accelerate generation of necessary knowledge for the potential use of Wolbachia-based IIT in area-wide integrated pest management (AW-IPM) strategies for the population control of tephritid pests.Instituto de GenéticaFil: Mateos, Mariana. Texas A&M University. Departments of Ecology and Conservation Biology, and Wildlife and Fisheries Sciences; Estados UnidosFil: Martinez Montoya, Humberto. Universidad Autónoma de Tamaulipas. Unidad Académica Multidisciplinaria Reynosa Aztlan. Laboratorio de Genética y Genómica Comparativa; MéxicoFil: Lanzavecchia, Silvia Beatriz. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Genética; ArgentinaFil: Conte, Claudia Alejandra. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Genética; ArgentinaFil: Guillén, Karina. El Colegio de la Frontera Sur; MéxicoFil: Morán-Aceves, Brenda M. El Colegio de la Frontera Sur; MéxicoFil: Toledo, Jorge. El Colegio de la Frontera Sur; MéxicoFil: Liedo, Pablo. El Colegio de la Frontera Sur; MéxicoFil: Asimakis, Elias D. University of Patras. Department of Environmental Engineering; GreciaFil: Doudoumis, Vangelis. University of Patras. Department of Environmental Engineering; GreciaFil: Kyritsis, Georgios A. University of Thessaly. Department of Agriculture Crop Production and Rural Environment. Laboratory of Entomology and Agricultural Zoology; GreciaFil: Papadopoulos, Nikos T. University of Thessaly. Department of Agriculture Crop Production and Rural Environment. Laboratory of Entomology and Agricultural Zoology; GreciaFil: Augustinos, Antonios A. Hellenic Agricultural Organization. Institute of Industrial and Forage Crops. Department of Plant Protection; GreciaFil: Segura, Diego Fernando. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Agrobiotecnología y Biología Molecular; Argentina. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Genética. Laboratorio de Genética de Insectos de Importancia Económica; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Tsiamis, George. University of Patras. Department of Environmental Engineering; Greci

Isolation of microsatellite markers in the olive fruit fly, bactrocera oleae, and their use for the analyses of natural populations of the species

The olive fruit fly, Bactrocera oleae, is the main pest of the olive fruit. Because of its great economic importance, especially for the Mediterranean countries, there is a need for a more effective control method. The application of an integrated, environmental friendly, management of this pest requires a better knowledge of its biology and of the genetic structure of its natural populations. The aim of the present study was the development of DNA microsatellite markers for the analysis of the natural populations of the olive fruit fly. These markers are abundant in the genome of any species studied so far and highly polymorphic. Three different strategies were used for the isolation of microsatellite markers. The first was the construction and screening of genomic libraries of the insect, the second was the construction of genomic libraries, enriched for microsatellites and the third was the use of primer pairs that were designed for the amplification of microsatellite markers in the closely related species Bactrocera oleae and Ceratitis capitata(cross-species amplification). A total of 69 microsatellite containing clones were isolated from libraries. The next step was the design of primer pairs in the microsatellite flanking sequences. A total of 42 primer pairs was designed and tested for their abillty to amplify the expected product. Test was performed through PCR and analysis of the PCR products through electrophoresis on agarose gel. Twenty primer pairs designed for the amplification of Bactrocera tryoni’ s microsatellites and 42 primer pairs designed for the amplification of Ceratitis capitata microsatellites were also tested. All three strategies gave 67 primer pairs that amplified the expected product. The degree of polymorphism of these primer pairs and their ability to amplify easily resolvable alleles was tested through PCR with DNA template of nine individuals. PCR products were analysed through polyacrylamide gel electrophoresis. Twenty eight out of the 49 primer pairs tested produced clear bands and twenty five of them were polymorphic. A small scale population analysis was then performed, using tventy four of the markers available. The main purpose of this analysis was to demonstate the quality of the markers and lead to the exclusion of six markers: one of them was monomorphic, another didn’t show reproducible results and four more showed deviations from H-W equilibrium, probably because of the presence of null alleles. Twelve of the remaining loci were used in a large scale analysis of B. oleae’s populations in the European part of the Mediterranean basin. Nineteen samples, varying from nine to fifty individuals, were analysed. These samples were collected from six different countries (Greece, Cyprus, Turkey, Italy, Spain and Portugal). The analysis revealed relatively low genetic distances, which, however, demonsrated a statistically important differentiation of the samples in three subpopulations. The first consisted of the samples from Cyprus, the second of the samples from Greece, Turkey and Italy and the third of the samples from the Iberian Peninsula. The statistical analyses performed showed the statistically important contribution of geographic distance to the generation of genetic distance. These three groups of samples also demonstrate a clear loss of polymorphism towards West. This loss is statistically important and, if we take into account the hypothesis that the colonization process of a species is followed by a loss in polymorphism, it suggests a colonization process of the olive fruit fly towards West in the European part of the Mediterranean basin, with a first expansion area in the Eastern part of the Mediterranean basin. Cross-species amplification experiments indicate close phylogenetic relationships among the species studied, mainly between B. tryoni-B. oleae and B. oleae-C. capitata. These results support the usefulness of the markers isolated in phylogenetic studies in these species, as well as in other, closely related species. The identification of a high percentage if conserved microsatellites in species that have been well separated for millions of years is in agreement with the hypothesis that microsatellites are not useless genomic regions (junj DNA) but they perform specific functions in the genome. Polymorphism analysis in the crosses of individuals from laboratory strains was very encouraging. The high degree of polymorpism showes that they can be used in genetic mapping of the species.Ο δάκος της ελιάς, Bactrocera oleae, αποτελεί το κυριότερο παράσιτο του καρπού της ελιάς. Λόγω της μεγάλης οικονομικής σημασίας της ελιάς, ιδιαίτερα για τις Μεσογειακές χώρες, ο αποτελεσματικότερος έλεγχος του εντόμου αυτού είναι απαραίτητος. Η εφαρμογή μεθόδων ολοκληρωμένης διαχείρισης του παρασίτου αυτού, μεθόδων δηλαδή φιλικών προς το περιβάλλον, επιβάλλει την καλύτερη γνώση της βιολογίας του και ιδιαίτερα της γενετικής και της γενετικής δομής των φυσικών πληθυσμών του. Στα πλαίσια αυτά, στόχος της παρούσας μελέτης ήταν η ανάπτυξη στο δάκο DNA δεικτών, των μικροδορυφόρων, οι οποίοι είναι άφθονοι στα γονιδιώματα και επιπλέον υψηλά πολυμορφικοί, για την ανάλυση φυσικών πληθυσμών του. Για την απομόνωση των μικροδορυφορικών δεικτών ακολουθήθηκαν τρεις διαφορετικές στρατηγικές: η κατασκευή και διαλογή γονιδιωματικών βιβλιοθηκών, η κατασκευή εμπλουτισμένων σε μικροδορυφόρους βιβλιοθηκών και ο έλεγχος ζευγών εκκινητών που είχαν σχεδιαστεί για την ενίσχυση μικροδορυφόρων στα συγγενικά είδη B. tyroni και C. capitata για το κατά πόσο ενισχύουν τις αντίστοιχες περιοχές και στο γονιδίωμα του εντόμου B. oleae (cross-species amplification). Από τις βιβλιοθήκες απομονώθηκαν συνολικά 69 κλώνοι που περιείχαν μικροδορυφόρους. Ακολούθησε ο σχεδιασμός ζευγών εκκινητών στις μοναδικές περιοχές που περιβάλλουν τους μικροδορυφόρους. Συνολικά σχεδιάστηκαν 42 ζεύγη εκκινητών. Οι εκκινητες αυτοί ελέγχθηκαν για το αν ενισχύουν το αναμενόμενο προϊόν. Ο έλεγχος έγινε με PCR και ηλεκτροφόρηση των προϊόντων σε πήκτωμα αγαρόζης. Παράλληλα ελέγχθηκαν είκοσι ζεύγη εκκινητών που είχαν σχεδιαστεί για μικροδορυφόρους του εντόμου B. tryoni και 42 ζεύγη εκκινητών που είχαν σχεδιαστεί για μικροδορυφόρους του εντόμου C. capitata. Οι τρεις διαδικασίες απομόνωσης έδωσαν συνολικά 67 λειτουργικά ζεύγη εκκινητών. Στη συνέχεια ελέγχθηκε μέσω PCR ο πολυμορφισμός των εκκινητών αυτών και το αν ενισχύουν διακριτά αλληλόμορφα, με μήτρα DNA γενετικό υλικό εννέα ατόμων. Τα προϊόντα της PCR αναλύθηκαν σε πήκτωμα ακρυλαμιδίου. Από τα 49 ζεύγη εκκινητών που ελέχθησαν, τα 28 έδωσαν πολύ καθαρό σήμα, ενώ τα 25 από αυτά βρέθηκαν πολυμορφικά. Μια πρώτη πληθυσμιακή ανάλυση έγινε με τη χρησιμοποίηση 24 από αυτούς τους δείκτες για την ανάλυση μικρού δείγματος ατόμων. Βασικός σκοπός της ανάλυσης αυτής ήταν να ελεγχθεί η ποιότητα των δεικτών και οδήγησε στον αποκλεισμό έξι δεικτών: ο ένας ήταν μονομορφικός, ένας δεύτερος έδειξε ασθενή ενίσχυση και μικρή επαναληψιμότητα και άλλοι τέσσερις εμφάνισαν απόκλιση από την ισορροπία κατά H-W, πιθανότητα λόγω της παρουσίας null αλληλομόρφων. Δώδεκα από τους υπόλοιπους δεκαοκτώ δείκτες χρησιμοποιήθηκαν για μια εκτεταμένη ανάλυση των πληθυσμών του δάκου στην Ευρωπαϊκή πλευρά της λεκάνης της Μεσογείου. Αναλύθηκαν δεκαεννέα δείγματα, μεγέθους εννέα ως πενήντα ατόμων, από έξι διαφορετικές χώρες (Ελλάδα, Κύπρος, Τουρκία, Ιταλία, Ισπανία και Πορτογαλία). Η ανάλυση αποκάλυψε σχετικά μικρές γενετικές αποστάσεις, που έδειχναν όμως μια στατιστικά σημαντική διαφοροποίηση σε τρεις υποπληθυσμούς. Ο πρώτος αποτελείτο από τα δείγματα της Κύπρου, ο δεύτερος από τα δείγματα Ελλάδας, Τουρκίας και Ισπανίας και ο τρίτος από τα δείγματα της Ιβηρικής χερσονήσου. Οι στατιστικές αναλύσεις που έγιναν έδειξαν τη σημαντική επίδραση της γεωγραφικής απόστασης στη δημιουργία αυτών των ομαδοποιήσεων. Οι τρεις αυτές ομάδες χαρακτηρίζονται από διαφορά και στο επίπεδο του πολυμορφισμού, εμφανίζοντας μια καθαρή μείωσή του από την Ανατολή προς τη Δύση. Η μείωση αυτή είναι στατιστικά σημαντική και με βάση την υπόθεση ότι η πορεία εποικισμού ενός είδους συνοδεύεται από μείωση του πολυμορφισμού, δίνει σημαντικές ενδείξεις για μία προς Δυσμάς πορεία εποίκισης του είδους στον Ευρωπαϊκό χώρο, με πρώτο κέντρο εξάπλωσης την Ανατολική λεκάνη της Μεσογείου. Τα πειράματα δια-ειδικής ενίσχυσης έδειξαν στενές φυλογενετικές σχέσεις των ειδών που εξετάστηκαν και κυρίως μεταξύ B. tryoni - B. oleae και B.oleae - C. capitata. Τα πειράματα αυτά υποστηρίζουν την χρησιμότητα των δεικτών που απομονώθηκαν σε ενδεχόμενες φυλογενετικές μελέτες στα είδη αυτά, καθώς και σε άλλα συγγενικά είδη. Η εύρεση ενός μεγάλου ποσοστού συντηρημένων μικροδορυφόρων σε είδη που έχουν διαχωριστεί εδώ και πολλά εκατομμύρια χρόνια ενισχύει την υπόθεση ότι οι μικροδορυφόροι δεν είναι γενετικό υλικό χωρίς ρόλο (junk DNA), αλλά επιτελούν συγκεκριμένες λειτουργίες στο γονιδίωμα. Η ανάλυση πολυμορφισμού που έγινε στις ατομικές διασταυρώσεις ατόμων από εργαστηριακούς ήταν ιδιαίτερα ενθαρρυντική. Ο υψηλός πολυμορφισμός που βρέθηκε δείχνει την δυνατότητα χρησιμοποίησης τους και στη γενετική χαρτογράφηση του είδους

Medfly Gut Microbiota and Enhancement of the Sterile Insect Technique: Similarities and Differences of Klebsiella oxytoca and Enterobacter sp. AA26 Probiotics during the Larval and Adult Stages of the VIENNA 8D53+ Genetic Sexing Strain

The Mediterranean fruit fly, Ceratitis capitata, is a major agricultural pest worldwide. The development of genetic sexing strains (GSSs) for this species that allows male-only sterile insects releases has boosted the effectiveness of the environmental friendly pest control method known as the sterile insect technique. The last generation of these strains, the VIENNA 7 and VIENNA 8, are currently used in all mass rearing facilities worldwide and are considered as models for such pest control applications. The sterile insect technique depends on the rearing of sufficient numbers of adequate “biological quality” laboratory flies to be released in the field. Currently, there is an increasing amount of studies focusing on the characterization of the symbiotic communities and development of probiotic diets. In our study, two bacterial isolates, an Enterobacter sp. (strain AA26) and a Klebsiella oxytoca strain, were used as probiotics in larval and adult diet. These strains have been shown to be beneficial, affecting several aspects related to the rearing efficiency and biological quality of the medfly VIENNA 8D53+ GSS. Our results demonstrate the effect of K. oxytoca on the developmental duration of the immature stages and, to some extent, on flight ability. On the other hand, our study does not support the presence of any beneficial effect of (a) K. oxytoca on pupal and adult recovery and adults’ survival under stress conditions when provided as a larval diet supplement and (b) K. oxytoca and Enterobacter sp. AA26 on mating competitiveness when provided as adult diet supplements. Possible explanations for inconsistencies with previous studies and the need for universalizing protocols are discussed. Our findings, combined with previous studies can support the sterile insect technique, through the improvement of different aspects of mass rearing and biological properties of laboratory reared insect pests