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Penelope Mavragani-tsipidou - One of the best experts on this subject based on the ideXlab platform.
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Genomic structure, organization and localization of the acetylcholinesterase locus of the olive fruit fly, Bactrocera oleae.
Bulletin of entomological research, 2012Co-Authors: Evdoxia G. Kakani, Elena Drosopoulou, Penelope Mavragani-tsipidou, M. Trakala, Kostas D. MathiopoulosAbstract:Acetylcholinesterase (AChE), encoded by the ace gene, is a key enzyme of cholinergic neurotransmission. Insensitive acetylcholinesterase (AChE) has been shown to be responsible for resistance to OPs and CBs in a number of arthropod species, including the most important pest of olives trees, the olive fruit fly Bactrocera oleae. In this paper, the organization of the B. oleae ace locus, as well as the structural and functional features of the enzyme, are determined. The organization of the gene was deduced by comparison to the ace cDNA sequence of B. oleae and the organization of the locus in Drosophila melanogaster. A similar structure between insect ace gene has been found, with conserved exon-intron positions and junction sequences. The B. oleae ace locus extends for at least 75 kb, consists of ten exons with nine introns and is mapped to division 34 of the chromosome arm IIL. Moreover, according to bioinformatic analysis, the Bo AChE exhibits all the common features of the insect AChE. Such structural and functional similarity among closely related AChE enzymes may implicate similarities in insecticide resistance mechanisms.
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Acetobacter tropicalis is a major symbiont of the olive fruit fly (Bactrocera oleae).
Applied and environmental microbiology, 2009Co-Authors: Ilias Kounatidis, Elena Crotti, Panagiotis Sapountzis, Luciano Sacchi, Aurora Rizzi, Bessem Chouaia, Claudio Bandi, Alberto Alma, Daniele Daffonchio, Penelope Mavragani-tsipidouAbstract:Following cultivation-dependent and -independent techniques, we investigated the microbiota associated with Bactrocera oleae, one of the major agricultural pests in olive-producing countries. Bacterial 16S rRNA gene libraries and ultrastructural analyses revealed the presence of several bacterial taxa associated with this insect, among which Acetobacter tropicalis was predominant. The recent increased detection of acetic acid bacteria as symbionts of other insect model organisms, such as Anopheles stephensi (G. Favia et al., Proc. Natl. Acad. Sci. USA 104:9047-9051, 2007) or Drosophila melanogaster (C. R. Cox and M. S. Gilmore, Infect. Immun. 75:1565-1576, 2007), prompted us to investigate the association established between A. tropicalis and B. oleae. Using an A. tropicalis-specific PCR assay, the symbiont was detected in all insects tested originating from laboratory stocks or field-collected from different locations in Greece. This acetic acid bacterium was successfully established in cell-free medium, and typing analyses, carried out on a collection of isolates, revealed that different A. tropicalis strains are present in fly populations. The capability to colonize and lodge in the digestive system of both larvae and adults and in Malpighian tubules of adults was demonstrated by using a strain labeled with a green fluorescent protein.
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Isolation and characterization of microsatellite markers from the olive fly, Bactrocera oleae, and their cross-species amplification in the Tephritidae family
BMC genomics, 2008Co-Authors: Antonios A. Augustinos, E. E. Stratikopoulos, Antigone Zacharopoulou, Penelope Mavragani-tsipidou, Eleni Drosopoulou, Evdoxia G. Kakani, Kostas D. MathiopoulosAbstract: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.
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Genetic and cytogenetic analysis of the olive fruit fly Bactrocera oleae (Diptera: Tephritidae).
Genetica, 2002Co-Authors: Penelope Mavragani-tsipidouAbstract:The genetic and cytogenetic characteristics of one of the major agricultural pests, the olive fruit fly Bactrocera oleae, are presented here. The mitotic metaphase complement of this insect consists of six pairs of chromosomes including one pair of heteromorphic sex chromosomes, with the male being the heterogametic sex. The analysis of the polytene complements of three larval tissues, the fat body, the salivary glands and the Malpighian tubules of this pest has shown (a) a total number of five long chromosomes (10 polytene arms) that correspond to the five autosomes of the mitotic nuclei and a heterochromatic mass corresponding to the sex chromosomes, (b) the constancy of the banding pattern of the three somatic tissues, (c) the absence of a typical chromocenter as an accumulation of heterochromatin, (d) the existence of reverse tandem duplications, and (e) the presence of toroid tips of the chromosome arms. The in situhybridization of genes or DNA sequences to the salivary gland polytene chromosomes of B. oleaeprovided molecular markers for all five autosomes and permitted the establishment of chromosomal homologies among B. olea, B. tryoniand Ceratitis capitata. The heat shock response of B. oleae, as revealed by heat-inducible puffing and protein pattern, shows a higher thermotolerance than Drosophila melanogaster.
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Organophosphate Resistance and Allelic Frequencies of Esterases in the Olive Fruit Fly Bactrocera oleae (Diptera: Tephritidae) 1
2001Co-Authors: Panagiotis Stasinakis, Vassilis Katsares, Penelope Mavragani-tsipidouAbstract:The polymorphic esterase loci, Est A and Est B, of a major agricultural pest, the olive fruit fly, Bactrocera oleae (Dacus oleae), were stud- ied using a standard laboratory population (Ls) that had never been subjected to insecticides, two "resistant" populations (Rs and RRs) constructed after exposure of larvae to sublethal doses of the organophosphate insecticide dimethoate, and a field population (Fs) subjected to long-term pesticide expo- sure. Data showed that dimethoate, as a selective agent, was responsible for the induction of the changes in the frequencies of alleles of the Est A locus, whereas it did not affect the Est B locus. Our results also showed that the changes in the frequencies of some active alleles of Est A locus, observed in resistant populations, were similar to those found in the field population.
Kostas D. Mathiopoulos - One of the best experts on this subject based on the ideXlab platform.
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Targeted somatic mutagenesis through CRISPR/Cas9 ribonucleoprotein complexes in the olive fruit fly, Bactrocera oleae.
Archives of insect biochemistry and physiology, 2020Co-Authors: Angela Meccariello, Kostas D. Mathiopoulos, Konstantina T. Tsoumani, Andrea Gravina, Pasquale Primo, Martina Buonanno, Giuseppe SacconeAbstract:The olive fruit fly, Bactrocera oleae (Diptera: Tephritidae), is the most destructive insect pest of olive cultivation, causing significant economic and production losses. Here, we present the establishment of the clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 methodology for gene disruption in this species. We performed targeted mutagenesis of the autosomal gene white (Bo-we), by injecting into early embryos in vitro preassembled and solubilized Cas9 ribonucleoprotein complexes loaded with two gene-specific single-guide RNAs. Gene disruption of Bo-we led to somatic mosaicism of the adult eye color. Large eye patches or even an entire eye lost the iridescent reddish color, indicating the successful biallelic mutagenesis in somatic cells. Cas9 induced either indels in each of the two simultaneously targeted Bo-we sites or a large deletion of the intervening region. This study demonstrates the first efficient implementation of the CRISPR/Cas9 technology in the olive fly, providing new opportunities towards the development of novel genetic tools for its control.
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targeted somatic mutagenesis through crispr cas9 ribonucleoprotein complexes in the olive fruit fly Bactrocera oleae
Archives of Insect Biochemistry and Physiology, 2020Co-Authors: Kostas D. Mathiopoulos, Angela Meccariello, Konstantina T. Tsoumani, Andrea Gravina, Pasquale Primo, Martina Buonanno, Giuseppe SacconeAbstract:The olive fruit fly, Bactrocera oleae (Diptera: Tephritidae), is the most destructive insect pest of olive cultivation, causing significant economic and production losses. Here, we present the establishment of the clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 methodology for gene disruption in this species. We performed targeted mutagenesis of the autosomal gene white (Bo-we), by injecting into early embryos in vitro preassembled and solubilized Cas9 ribonucleoprotein complexes loaded with two gene-specific single-guide RNAs. Gene disruption of Bo-we led to somatic mosaicism of the adult eye color. Large eye patches or even an entire eye lost the iridescent reddish color, indicating the successful biallelic mutagenesis in somatic cells. Cas9 induced either indels in each of the two simultaneously targeted Bo-we sites or a large deletion of the intervening region. This study demonstrates the first efficient implementation of the CRISPR/Cas9 technology in the olive fly, providing new opportunities towards the development of novel genetic tools for its control.
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Genomic structure, organization and localization of the acetylcholinesterase locus of the olive fruit fly, Bactrocera oleae.
Bulletin of entomological research, 2012Co-Authors: Evdoxia G. Kakani, Elena Drosopoulou, Penelope Mavragani-tsipidou, M. Trakala, Kostas D. MathiopoulosAbstract:Acetylcholinesterase (AChE), encoded by the ace gene, is a key enzyme of cholinergic neurotransmission. Insensitive acetylcholinesterase (AChE) has been shown to be responsible for resistance to OPs and CBs in a number of arthropod species, including the most important pest of olives trees, the olive fruit fly Bactrocera oleae. In this paper, the organization of the B. oleae ace locus, as well as the structural and functional features of the enzyme, are determined. The organization of the gene was deduced by comparison to the ace cDNA sequence of B. oleae and the organization of the locus in Drosophila melanogaster. A similar structure between insect ace gene has been found, with conserved exon-intron positions and junction sequences. The B. oleae ace locus extends for at least 75 kb, consists of ten exons with nine introns and is mapped to division 34 of the chromosome arm IIL. Moreover, according to bioinformatic analysis, the Bo AChE exhibits all the common features of the insect AChE. Such structural and functional similarity among closely related AChE enzymes may implicate similarities in insecticide resistance mechanisms.
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Isolation and characterization of microsatellite markers from the olive fly, Bactrocera oleae, and their cross-species amplification in the Tephritidae family
BMC genomics, 2008Co-Authors: Antonios A. Augustinos, E. E. Stratikopoulos, Antigone Zacharopoulou, Penelope Mavragani-tsipidou, Eleni Drosopoulou, Evdoxia G. Kakani, Kostas D. MathiopoulosAbstract: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.
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Organophosphate resistance in olive fruit fly, Bactrocera oleae, populations in Greece and Cyprus.
Pest management science, 2006Co-Authors: Panagiotis J. Skouras, Kostas D. Mathiopoulos, John T. Margaritopoulos, Nicos Seraphides, Ioannis M. Ioannides, Evi G Kakani, J.a. TsitsipisAbstract:The olive fruit fly Bactrocera oleae (Gmelin) (Diptera: Tephritidae) is the most important pest of olives in countries around the Mediterranean basin. Its control has been based mostly on bait sprays with organophosphate insecticides (usually dimethoate or fenthion) for about 40 years. In the present study, the resistance status of olive fruit fly populations to dimethoate was examined in Greece and Cyprus over 2 years. Thirty-one populations from various regions of Greece, nine from Cyprus and one laboratory susceptible strain, which served as a control, were assayed by topical application of dimethoate. Considerable variation in the resistance levels to dimethoate was recorded in the populations of B. oleae, with resistance ratios ranging from 6.3 to 64.4 (ED50 values 12.5–128.7 ng dimethoate per insect). The highest resistance ratios were found in populations from Crete, and the lowest in those from Cyprus. This variation could be attributed to different selection pressures from insecticidal applications among populations from the various regions. Migration of resistant genotypes, either autonomous or via commerce, may also be involved. Copyright © 2006 Society of Chemical Industry
Charalambos Savakis - One of the best experts on this subject based on the ideXlab platform.
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transinfection of the olive fruit fly Bactrocera oleae with wolbachia towards a symbiont based population control strategy
Journal of Applied Entomology, 2011Co-Authors: A. Apostolaki, Charalambos Savakis, Ioannis Livadaras, Aggeliki Saridaki, A. Chrysargyris, Kostas BourtzisAbstract:The olive fruit fly Bactrocera oleae is responsible for worldwide economic damage. In this report, we describe the first B. oleae lines transinfected with the Wolbachia strain wCer2, an endosymbiont of the cherry fruit fly Rhagoletis cerasi. Immunostaining followed by confocal microscopy, detects high numbers of Wolbachia in embryos as well as in ovarioles and sperm from individuals of both transinfected lines. wCer2 was uniformly distributed in B. oleae egg chambers and the cortex of preblastoderm embryos. Wolbachia is known to manipulate host reproduction with several strategies, one of which is cytoplasmic incompatibility (CI), resulting in embryonic mortality in incompatible crosses. Wolbachia was found to induce complete CI in the novel host, suggesting that symbiont-based approaches can be used as novel environmentally friendly tools for the control of natural olive fruit fly populations.
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Transinfection of the olive fruit fly Bactrocera oleae with Wolbachia: towards a symbiont‐based population control strategy
Journal of Applied Entomology, 2011Co-Authors: A. Apostolaki, Charalambos Savakis, Ioannis Livadaras, Aggeliki Saridaki, A. Chrysargyris, Kostas BourtzisAbstract:The olive fruit fly Bactrocera oleae is responsible for worldwide economic damage. In this report, we describe the first B. oleae lines transinfected with the Wolbachia strain wCer2, an endosymbiont of the cherry fruit fly Rhagoletis cerasi. Immunostaining followed by confocal microscopy, detects high numbers of Wolbachia in embryos as well as in ovarioles and sperm from individuals of both transinfected lines. wCer2 was uniformly distributed in B. oleae egg chambers and the cortex of preblastoderm embryos. Wolbachia is known to manipulate host reproduction with several strategies, one of which is cytoplasmic incompatibility (CI), resulting in embryonic mortality in incompatible crosses. Wolbachia was found to induce complete CI in the novel host, suggesting that symbiont-based approaches can be used as novel environmentally friendly tools for the control of natural olive fruit fly populations.
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The transformer gene in Bactrocera oleae: the genetic switch that determines its sex fate
Insect molecular biology, 2007Co-Authors: D. Lagos, M. Koukidou, Charalambos Savakis, Katia KomitopoulouAbstract:Transformer (tra) is the second gene of a regulatory cascade based on RNA splicing that determines sex in Drosophila melanogaster. Splicing of tra transcripts is regulated by the master gene Sex lethal and tra itself regulates splicing of the transcriptional regulator doublesex (dsx). We present the isolation and characterization of Botra, the olive fruit fly Bactrocera oleae orthologue to the Drosophila gene transformer. As in Drosophila, Botra transcripts are spliced in a sex-specific manner so that only females encode a functional polypeptide of 422 amino acids, whereas males encode presumably nonfunctional peptide(s). The identification of multiple TRA/TRA-2 binding sites within the Botra male-specific exons, suggests an autoregulation mechanism of tra, through TRA/TRA2 activities. The fundamental role of the TRA protein in sex determination of Bactrocera was investigated by RNA interference, where the introduction of Botra dsRNA into embryos resulted in complete transformation of XX flies into fertile males.
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Germ line transformation of the olive fly Bactrocera oleae using a versatile transgenesis marker.
Insect molecular biology, 2006Co-Authors: M. Koukidou, Apostolos Klinakis, C. Reboulakis, Laskaro Zagoraiou, Nektarios Tavernarakis, I. Livadaras, A. P. Economopoulos, Charalambos SavakisAbstract:The olive fruit fly (olive fly) Bactrocera oleae (Dacus), recently introduced in North America, is the most destructive pest of olives worldwide. The lack of an efficient gene transfer technology for olive fly has hampered molecular analysis, as well as development of genetic techniques for its control. We have developed a Minos-based transposon vector carrying a self-activating cassette which overexpresses the enhanced green fluorescent protein (EGFP). Efficient transposase-mediated integration of one to multiple copies of this vector was achieved in the germ line of B. oleae by coinjecting the vector along with in vitro synthesized Minos transposase mRNA into preblastoderm embryos. The self-activating gene construct combined with transposase mRNA present a system with potential for transgenesis of very diverse species.
Boaz Yuval - One of the best experts on this subject based on the ideXlab platform.
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symbiotic bacteria affect oviposition behavior in the olive fruit fly Bactrocera oleae
Journal of Insect Physiology, 2019Co-Authors: Polpass Arul Jose, Edouard Jurkevitch, Michael Benyosef, Boaz YuvalAbstract:Microbial associations are widespread across the insects. In the olive fruit fly Bactrocera oleae (Diptera: Tephritidae), vertically transmitted gut symbionts contribute to larval development inside the olive host, and to adult nutrition. Nevertheless, their effect on behavioural decisions of adults is unknown. In this study, we show that symbiotic bacteria affect oviposition behaviour in B. oleae. We studied the effect of different fruits as hosts and different gut-bacteria as gut-symbionts on oviposition attempts and fly development in B. oleae. Untreated flies that had native gut-symbionts attempted oviposition significantly more times than axenic flies as well as flies treated with medfly-associated Pantoea or Klebsiella bacteria. Axenic flies provided with a diet containing the homogenized gut of symbiotic flies recovered the same number of oviposition attempts as their symbiotic counterparts. As for as the different hosts, green olives (unripe) and grapes were preferred while black olives (ripe) elicited the least number of oviposition attempts, with an interactive effect of host and bacterial treatments. It appears that both the host attributes and the native gut-symbionts drive oviposition preference towards green olives in B. oleae. Moreover, both bacterial treatments and hosts significantly affected the development of B. oleae larvae. Though grapes elicited as many oviposition attempts as green olives, they yielded no pupae. Taken together, our results suggest that the intimate association between B. oleae and their gut-microbes, extends beyond nutritional support to behaviour.
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Symbiotic bacteria enable olive flies (Bactrocera oleae) to exploit intractable sources of nitrogen
Journal of evolutionary biology, 2014Co-Authors: Michael Ben-yosef, Zohar Pasternak, Edouard Jurkevitch, Boaz YuvalAbstract:Insects are often associated with symbiotic micro-organisms, which allow them to utilize nutritionally marginal diets. Adult fruit flies (Diptera: Tephritidae) associate with extracellular bacteria (Enterobacteriaceae) that inhabit their digestive tract. These flies obtain nutrients by foraging for plant exudates, honeydew and bird droppings scattered on leaves and fruit – a nutritional niche which offers ample amounts of carbohydrates, but low quantities of available nitrogen. We identified the bacteria resident in the gut of the olive fly (Bactrocera oleae) – a worldwide pest of olives and examined their contribution to nitrogen metabolism in the adult insect. By suppressing bacteria in the gut and monitoring female fecundity, we demonstrate that bacteria contribute essential amino acids and metabolize urea into an available nitrogen source for the fly, thus significantly elevating egg production. In an ecological context, bacteria were found to be beneficial to females subsisting on bird droppings, but not on honeydew – two natural food sources. We suggest that a main gut bacterium (Candidatus Erwinia dacicola) forms an inseparable, essential part of this fly’s nutritional ecology. The evolution of this symbiosis has allowed adult flies to utilize food substrates which are low or imbalanced in assimilable nitrogen and thereby to overcome the nitrogen limitations of their natural diet.
John Vontas - One of the best experts on this subject based on the ideXlab platform.
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Efficient genome editing in the olive fruit fly, Bactrocera oleae.
Insect molecular biology, 2020Co-Authors: Venetia Koidou, Shane Denecke, Panagiotis Ioannidis, I. Vlatakis, Ioannis Livadaras, John VontasAbstract:The olive fruit fly, Bactrocera oleae, causes great damage to the quality and quantity of olive production worldwide. Pest management approaches have proved difficult for a variety of reasons, a fact that has brought about a need for alternative tools and approaches. Here we report for the first time in B. oleae the development of the clustered regularly interspaced palindromic repeats (CRISPR)/CRISPR associated protein 9 (Cas9) gene editing tool, using the well-known eye colour marker gene scarlet. Two synthetic guide RNAs targeting the coding region of the scarlet gene were synthesized and shown to work efficiently in vitro. These reagents were then microinjected along with purified Cas9 protein into early-stage embryos. Successful CRISPR-induced mutations of both copies of the scarlet gene showed a striking yellow eye phenotype, indicative of gene disruption. Multiple successful CRISPR events were confirmed by PCR and sequencing. The establishment of an efficient CRISPR-based gene editing tool in B. oleae will enable the study of critical molecular mechanisms in olive fruit fly biology and physiology, including the analysis of insecticide resistance mechanisms and the discovery of novel insecticide targets, as well as facilitate the development of novel biotechnology-based pest control strategies.
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Molecular characterization of pyrethroid resistance in the olive fruit fly Bactrocera oleae
Pesticide biochemistry and physiology, 2018Co-Authors: Nena Pavlidi, Anastasia Kampouraki, Vasilis Tseliou, Nicky Wybouw, Wannes Dermauw, Emmanouil Roditakis, Ralf Nauen, Thomas Van Leeuwen, John VontasAbstract:Α reduction of pyrethroid efficacy has been recently recorded in Bactrocera oleae, the most destructive insect of olives. The resistance levels of field populations collected from Crete-Greece scaled up to 22-folds, compared to reference laboratory strains. Sequence analysis of the IIS4-IIS6 region of para sodium channel gene in a large number of resistant flies indicated that resistance may not be associated with target site mutations, in line with previous studies in other Tephritidae species. We analyzed the transcriptomic differences between two resistant populations versus an almost susceptible field population and two laboratory strains. A large number of genes was found to be significantly differentially transcribed across the pairwise comparisons. Interestingly, gene set analysis revealed that genes of the 'electron carrier activity' GO group were enriched in one specific comparison, which might suggest a P450-mediated resistance mechanism. The up-regulation of several transcripts encoding detoxification enzymes was qPCR validated, focusing on transcripts coding for P450s. Of note, the expression of contig00436 and contig02103, encoding CYP6 P450s, was significantly higher in all resistant populations, compared to susceptible ones. These results suggest that an increase in the amount of the CYP6 P450s might be an important mechanism of pyrethroid resistance in B. oleae.
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Draft Genome Sequence of Chryseobacterium Strain CBo1 Isolated from Bactrocera oleae.
Genome announcements, 2017Co-Authors: Frances Blow, John Vontas, Alistair C. DarbyAbstract:Bacteria of the genus Chryseobacterium have previously been identified as mutualists of plants and insects. Chryseobacterium strain CBo1 was cultured from the gut of the agricultural pest Bactrocera oleae and its whole genome sequenced. This genomic resource will aid investigations into the transition of microbes between plant and invertebrate hosts.
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Draft Genome Sequence of Stenotrophomonas maltophilia SBo1 Isolated from Bactrocera oleae.
Genome announcements, 2016Co-Authors: Frances Blow, John Vontas, Alistair C. DarbyAbstract:ABSTRACT Bacteria of the genus Stenotrophomonas are ubiquitous in the environment and are increasingly associated with insects. Stenotrophomonas maltophilia SBo1 was cultured from the gut of Bactrocera oleae. The draft genome sequence presented here will inform future investigations into the nature of the interaction between insects and their microbiota.
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Draft Genome Sequence of the Bactrocera oleae Symbiont “Candidatus Erwinia dacicola”
Genome announcements, 2016Co-Authors: Frances Blow, Anastasia Gioti, David Starns, Michael Ben-yosef, Zohar Pasternak, Edouard Jurkevitch, John Vontas, Alistair C. DarbyAbstract:"Candidatus Erwinia dacicola" is a Gammaproteobacterium that forms a symbiotic association with the agricultural pest Bactrocera oleae Here, we present a 2.1-Mb draft hybrid genome assembly for "Ca. Erwinia dacicola" generated from single-cell and metagenomic data.
