scholarly journals A transgenic female killing system for the genetic control of Drosophila suzukii

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Marc F. Schetelig ◽  
Jonas Schwirz ◽  
Ying Yan
Keyword(s):  
Genetic Control ◽  
Control Strategies ◽  
Fruit Fly ◽  
Female Offspring ◽  
Early Gene ◽  
Invasive Pest ◽  
Drosophila Suzukii ◽  
Female Specific

AbstractThe spotted wing Drosophila (Drosophila suzukii) is an invasive pest of soft-skinned fruit crops. It is rapidly transmitted in Europe and North America, causing widespread agricultural losses. Genetic control strategies such as the sterile insect technique (SIT) have been proposed as environment-friendly and species-restricted approaches for this pest. However, females are inefficient agents in SIT programs. Here we report a conditional female-killing (FK) strategy based on the tetracycline-off system. We assembled sixteen genetic constructs for testing in vitro and in vivo. Twenty-four independent transgenic strains of D. suzukii were generated and tested for female-specific lethality. The strongest FK effect in the absence of tetracycline was achieved by the construct containing D. suzukii nullo promoter for early gene expression, D. suzukii pro-apoptotic gene hidAla4 for lethality, and the transformer gene intron from the Mediterranean fruit fly Ceratitis capitata for female-specific splicing. One strain carrying this construct eliminated 100% of the female offspring during embryogenesis and produced only males. However, homozygous females from these FK strains were not viable on a tetracycline-supplemented diet, possibly due to the basal expression of hidAla4. Potential improvements to the gene constructs and the use of such FK strains in an SIT program are discussed.

scholarly journals Immune Response of Drosophila suzukii Larvae to Infection with the Nematobacterial Complex Steinernema carpocapsae–Xenorhabdus nematophila

Insects ◽  
2020 ◽  
Vol 11 (4) ◽  
pp. 210 ◽  
Author(s):  
Anna Garriga ◽  
Maristella Mastore ◽  
Ana Morton ◽  
Fernando Garcia del Pino ◽  
Maurizio Francesco Brivio
Keyword(s):  
Immune Response ◽  
Entomopathogenic Nematodes ◽  
Humoral Response ◽  
Inhibitory Effect ◽  
Steinernema Carpocapsae ◽  
Invasive Pest ◽  
Xenorhabdus Nematophila ◽  
Drosophila Suzukii

Entomopathogenic nematodes have been proposed as biological agents for the control of Drosophila suzukii, an invasive pest of small-stone and soft-skinned fruits. Larvae of the fly are susceptible to Steinernema carpocapsae infection but the reaction of immune defenses of the host are unknown. To determine the immune response, larvae were infected with S. carpocapsae and Xenorhabdus nematophila to evaluate the effector mechanisms of both humoral and cellular processes. The symbiont bacteria presented an inhibitory effect on the phenoloxidase cascade with a low level of melanization. Besides, X. nematophila activated the synthesis of putative antimicrobial peptides on the hemolymph of infected larvae. However, those peptides presented a lower antimicrobial activity compared to hemolymph from larvae infected with non-symbiont bacteria. Xenorhabdus nematophila avoided also the phagocytosis response of hemocytes. During in vitro and in vivo assays, S. carpocapsae was not encapsulated by cells, unless the cuticle was damaged with a lipase-treatment. Hemocyte counts confirmed differentiation of lamellocytes in the early phase of infection despite the unrecognition of the nematodes. Both X. nematophila and S. carpocapsae avoided the cellular defenses of D. suzukii larvae and depressed the humoral response. These results confirmed the potential of entomopathogenic nematodes to control D. suzukii.

scholarly journals The use of light spectrum blocking films to reduce populations of Drosophila suzukii Matsumura in fruit crops

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Michelle T. Fountain ◽  
Amir Badiee ◽  
Sebastian Hemer ◽  
Alvaro Delgado ◽  
Michael Mangan ◽  
...  
Keyword(s):  
Control Strategies ◽  
Field Trials ◽  
Biological Data ◽  
Insect Vision ◽  
Invasive Pest ◽  
Effective Control ◽  
Fruit Crops ◽  
Light Spectrum ◽  
Drosophila Suzukii ◽  
The Impact

Abstract Spotted wing drosophila, Drosophila suzukii, is a serious invasive pest impacting the production of multiple fruit crops, including soft and stone fruits such as strawberries, raspberries and cherries. Effective control is challenging and reliant on integrated pest management which includes the use of an ever decreasing number of approved insecticides. New means to reduce the impact of this pest that can be integrated into control strategies are urgently required. In many production regions, including the UK, soft fruit are typically grown inside tunnels clad with polyethylene based materials. These can be modified to filter specific wavebands of light. We investigated whether targeted spectral modifications to cladding materials that disrupt insect vision could reduce the incidence of D. suzukii. We present a novel approach that starts from a neuroscientific investigation of insect sensory systems and ends with infield testing of new cladding materials inspired by the biological data. We show D. suzukii are predominantly sensitive to wavelengths below 405 nm (ultraviolet) and above 565 nm (orange & red) and that targeted blocking of lower wavebands (up to 430 nm) using light restricting materials reduces pest populations up to 73% in field trials.

Transcriptional activation by herpes simplex virus type 1 VP16 in vitro and its inhibition by oligopeptides

1994 ◽  
Vol 14 (5) ◽  
pp. 3484-3493
Author(s):  
T J Wu ◽  
G Monokian ◽  
D F Mark ◽  
C R Wobbe
Keyword(s):  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Transcriptional Activation ◽  
Deletion Mutant ◽  
Full Length ◽  
Early Gene ◽  
Immediate Early ◽  
Simplex Virus

VP16 is a herpes simplex virus (HSV)-encoded transcriptional activator protein that is essential for efficient viral replication and as such may be a target for novel therapeutic agents directed against viral gene expression. We have reconstituted transcriptional activation by VP16 in an in vitro system that is dependent on DNA sequences from HSV immediate-early gene promoters and on protein-protein interactions between VP16 and Oct-1 that are required for VP16 activation in vivo. Activation increased synergistically with the number of TAATGARAT elements (the cis-acting element for VP16 activation in vivo) upstream of the core promoter, and mutations of this element that reduce Oct-1 or VP16 DNA binding reduced transactivation in vitro. A VP16 insertion mutant unable to interact with Oct-1 was inactive, but, surprisingly, a deletion mutant lacking the activation domain was approximately 65% as active as the full-length protein. The activation domains of Oct-1 were necessary for activation in reactions containing the VP16 deletion mutant, and they contributed significantly to activation by full-length VP16. Addition of a GA-rich element present in many HSV immediate-early gene enhancers synergistically stimulated VP16-activated transcription. Finally, oligopeptides that are derived from a region of VP16 thought to contact a cellular factor known as HCF (host cell factor) and that inhibit efficient VP16 binding to the TAATGARAT element also specifically inhibited VP16-activated, but not basal, transcription. Amino acid substitutions in one of these peptides identified three residues that are absolutely required for inhibition and presumably for interaction of VP16 with HCF.

DNA sequence requirements for replication of polyomavirus DNA in vivo and in vitro

1987 ◽  
Vol 7 (10) ◽  
pp. 3694-3704
Author(s):  
C Prives ◽  
Y Murakami ◽  
F G Kern ◽  
W Folk ◽  
C Basilico ◽  
...  
Keyword(s):  
Dna Replication ◽  
Simian Virus 40 ◽  
T Antigen ◽  
Early Gene ◽  
Wild Type ◽  
The Core ◽  
Cell Extracts ◽  
Sequence Requirements

Cell extracts of FM3A mouse cells replicate polyomavirus (Py) DNA in the presence of immunoaffinity-purified Py large T antigen, deoxynucleoside triphosphates, ATP, and an ATP-generating system. This system was used to examine the effects of mutations within or adjacent to the Py core origin (ori) region in vitro. The analysis of plasmid DNAs containing deletions within the early-gene side of the Py core ori indicated that sequences between nucleotides 41 and 57 define the early boundary of Py DNA replication in vitro. This is consistent with previously published studies on the early-region sequence requirements for Py replication in vivo. Deleting portions of the T-antigen high-affinity binding sites A and B (between nucleotides 57 and 146) on the early-gene side of the core ori led to increased levels of replication in vitro and to normal levels of replication in vivo. Point mutations within the core ori region that abolish Py DNA replication in vivo also reduced replication in vitro. A mutant with a reversed orientation of the Py core ori region replicated in vitro, but to a lesser extent that wild-type Py DNA. Plasmids with deletions on the late-gene side of the core ori, within the enhancer region, that either greatly reduced or virtually abolished Py DNA replication in vivo replicated to levels similar to those of wild-type Py DNA plasmids in vitro. Thus, as has been observed with simian virus 40, DNA sequences needed for Py replication in vivo are different from and more stringent than those required in vitro.

scholarly journals PENGARUH IRADIASI SINAR GAMMA [60Co] TERHADAP BACTROCERA CARAMBOLAE DREW & HANCOCK IN VITRO DAN IN VIVO

2015 ◽  
Vol 15 (1) ◽  
pp. 17
Author(s):  
Endang Sri Ratna ◽  
Kemas Usman ◽  
Indah Arastuti ◽  
Dadan Hindayana
Keyword(s):  
Gamma Ray ◽  
Lethal Dose ◽  
Fruit Fly ◽  
Probit Analysis ◽  
Adult Survival ◽  
Significance Level ◽  
Bactrocera Carambolae ◽  
Gamma Ray Irradiation

Effect of gamma irradiation [60Co] against Bactrocera carambolae Drew & Hancock in vitro and in vivo. Bactrocera carambolae Drew & Hancock is one of the most important pests on guava fruit. According to a quarantine regulation in export-import commodities, irradiation treatment is a suitable methods for eradicating infested organism, which is relatively safe for the environment. The aim of this research was to determine mortality doses and an effective dose of [60Co] gamma ray irradiation for the eradication purpose, and its implication on the survival of fruit fly B. carambolae. Two irradiation methods of in vitro dan in vivo were carried out, by exposing egg and 3rd instar larvae of B. carambolae obtained from the laboratory reared insect. Eleven doses of gamma ray irradiation of 0, 30, 50, 75, 100, 125, 150, 175, 200, 300, 450, and 600 Gy were applied, respectively. The level of 99% fruit fly mortality was estimated by the value of LD99 using probit analysis and the number of larvae, pupae and adult survival were evaluated by analysis of variance (ANOVA), and the means compared by Tukey’s test, at 5% of significance level. These result showed that the effective lethal dose (LD99) of irradiation that could be successful to eradicate eggs and 3rd instar larvae in vitro were 2225 and 2343 Gy and in vivo were 3165 dan 3177 Gy, respectively. Almost all of the treated larvae survived and developed to pupae, therefore only the minimum irradiation dose of 30 Gy allowed the pupae to develop into adults.

scholarly journals Efficient and Stable Delivery of Multiple Genes to Fish Cells by a Modified Recombinant Baculovirus System

2018 ◽  
Vol 19 (12) ◽  
pp. 3767 ◽  
Author(s):  
Qian Wang ◽  
Jian Fang ◽  
Qihua Pan ◽  
Yizhou Wang ◽  
Ting Xue ◽  
...  
Keyword(s):  
Mammalian Cells ◽  
Fluorescent Protein ◽  
Recombinant Baculovirus ◽  
Transgenic Fish ◽  
Early Gene ◽  
Drug Selection ◽  
Baculovirus System ◽  
Fish Cells

The recombinant baculovirus has been widely used as an efficient tool to mediate gene delivery into mammalian cells but has barely been used in fish cells. In the present study, we constructed a recombinant baculovirus containing the dual-promoter cytomegalovirus (CMV) and white spot syndrome virus (WSSV) immediate-early gene 1 (ie1) (WSSV ie1), followed by a puromycin–green fluorescent protein (Puro-GFP, pf) or puromycin–red fluorescent protein (Puro-RFP, pr) cassette, which simultaneously allowed for easy observation, rapid titer determination, drug selection, and exogenous gene expression. This recombinant baculovirus was successfully transduced into fish cells, including Mylopharyngodon piceus bladder (MPB), fin (MPF), and kidney (MPK); Oryzias latipes spermatogonia (SG3); and Danio rerio embryonic fibroblast (ZF4) cells. Stable transgenic cell lines were generated after drug selection, which was further verified by Western blot. A cell monoclonal formation assay proved the stable heredity of transgenic MPB cells. In addition, a recombinant baculovirus containing a pr cassette and four transcription factors for induced pluripotent stem cells (iPSC) was constructed and transduced into ZF4 cells, and these exogenous genes were simultaneously delivered and transcribed efficiently in drug-selected ZF4 cells, proving the practicability of this modified recombinant baculovirus system. We also proved that the WSSV ie1 promoter had robust activity in fish cells in vitro and in vivo. Taken together, this modified recombinant baculovirus can be a favorable transgenic tool to obtain transient or stable transgenic fish cells.

Exogenous L-lactate promotes astrocyte plasticity but is not sufficient for enhancing striatal synaptogenesis or motor behavior in mice

2020 ◽  
Author(s):  
Adam J. Lundquist ◽  
Tyler J. Gallagher ◽  
Giselle M. Petzinger ◽  
Michael W. Jakowec
Keyword(s):  
Motor Behavior ◽  
Early Gene ◽  
Specific Gene ◽  
Motor Circuits ◽  
Specific Manner ◽  
Primary Astrocytes ◽  
Exercise Induced ◽  
The Brain

AbstractL-lactate is an energetic and signaling molecule that is key to the metabolic and neuroplastic connection between astrocytes and neurons and may be involved in exercise-induced neuroplasticity. This study sought to explore the role of L-lactate in astrocyte reactivity and neuroplasticity. Using in vitro cultures of primary astrocytes, we show L-lactate increased expression of plasticity-related genes, including neurotrophic factors, Bdnf, Gdnf, Cntf and the immediate early gene cFos. L-lactate’s promotion of neurotrophic factor expression may be mediated in part by the lactate receptor HCAR1 since application of the HCAR1 agonist 3,5-DHBA also increased expression of Bdnf in primary astrocytes. In vivo L-lactate administration to healthy mice caused a similar increase in the expression of plasticity-related genes as well as increased astrocyte morphological complexity in a region-specific manner, with increased astrocytic response found in the striatum but not the ectorhinal cortex, regions of the brain where increases in regional cerebral blood flow are increased or unaltered, respectively, with motor behavior. Additionally, L-lactate administration did not cause synaptogenesis or improve motor behavior based on the latency to fall on the accelerating rotarod, suggesting that L-lactate administration can initiate astrocyte-specific gene expression, but the activation of motor circuits is necessary to initiate striatal neuroplasticity. These results suggest that peripheral L-lactate is likely an important molecular component of exercise-induced neuroplasticity by acting in an astrocyte-specific manner to prime the brain for neuroplasticity.

scholarly journals In vivo and in vitro analysis of transcriptional activation mediated by the human cytomegalovirus major immediate-early proteins.

1993 ◽  
Vol 13 (2) ◽  
pp. 1238-1250 ◽  
Author(s):  
K M Klucher ◽  
M Sommer ◽  
J T Kadonaga ◽  
D H Spector
Keyword(s):  
Human Cytomegalovirus ◽  
Hela Cells ◽  
Transcriptional Activation ◽  
Histone H1 ◽  
Early Gene ◽  
Immediate Early ◽  
Early Promoter ◽  
Drosophila Embryos

To define mechanistically how the human cytomegalovirus (HCMV) major immediate-early (IE) proteins induce early-gene transcription, the IE1 72-kDa protein, the IE2 55-kDa protein, and the IE2 86-kDa protein were analyzed for their ability to activate transcription from an HCMV early promoter in vivo and in vitro. In transient-expression assays in U373MG astrocytoma/glioblastoma and HeLa cells, only the IE2 86-kDa protein was able to activate the HCMV early promoter to high levels. In HeLa cells, the IE1 72-kDa protein was able to activate the promoter to a low but detectable level, and the level of promoter activity observed in response to the IE2 86-kDa protein was increased synergistically following cotransfection of the constructs expressing both IE proteins. To examine the interaction of the HCMV IE proteins with the RNA polymerase II transcription machinery, we assayed the ability of Escherichia coli-synthesized proteins to activate the HCMV early promoter in nuclear extracts prepared from U373MG cells, HeLa cells, and Drosophila embryos. The results of the in vitro experiments correlated well with those obtained in vivo. The basal activity of the promoter was minimal in both the HeLa and U373MG extracts but was stimulated 6- to 10-fold by the IE2 86-kDa protein. With a histone H1-deficient extract from Drosophila embryos, the HCMV early promoter was quite active and was stimulated two- to fourfold by the IE2 86-kDa protein. Addition of histone H1 at 1 molecule per 40 to 50 bp of DNA template significantly repressed basal transcription from this promoter. However, the IE2 86-kDa protein, but none of the other IE proteins, was able to counteract the H1-mediated repression and stimulate transcription at least 10- to 20-fold. The promoter specificity of the activation was demonstrated by the inability of the IE2 86-kDa protein to activate the Drosophila Krüppel promoter in either the presence or absence of histone H1. These results suggest that one mechanism of transcription activation by the IE2 86-kDa protein involves antirepression.

scholarly journals Location of sequences in polyomavirus DNA that are required for early gene expression in vivo and in vitro.

1984 ◽  
Vol 4 (12) ◽  
pp. 2594-2609 ◽  
Author(s):  
C R Mueller ◽  
A M Mes-Masson ◽  
M Bouvier ◽  
J A Hassell
Keyword(s):  
Gene Expression ◽  
Thymidine Kinase ◽  
Dna Sequences ◽  
Viral Genome ◽  
Simian Virus 40 ◽  
Early Gene ◽  
Wild Type ◽  
Transcription In Vitro

To define the DNA sequences required for the expression of the polyomavirus early transcription unit, we cloned part of the viral genome in a plasmid vector, isolated mutants bearing lesions introduced in vitro within DNA sequences upstream of the transcriptional start site, and measured the capacity of these various mutant genomes to transform cells and to function as templates for transcription in vitro by comparison with wild-type DNA. One set of mutants bore 5' unidirectional deletions beginning at position -810 and extending downstream to position +4. Another set of mutants bore 3' undirectional deletions starting at position +4 and progressing upstream to position -311. The last set of mutants bore internal deletions between positions -810 and +4. Analyses of the properties of these mutant DNAs led us to conclude that the region between positions -403 and -311 includes an enhancer of gene expression. Deletion of this area from the viral genome reduced gene expression in vivo to 1 to 2% of wild-type levels, as measured by transformation assays. Moreover, this region increased the frequency of transformation of thymidine kinase-negative Rat-2 cells by the herpes simplex virus thymidine kinase (tk) gene from 5- to 20-fold. This occurred only if the polyomavirus sequences were covalently linked to the tk gene and then occurred independently of their orientation or position relative to the tk gene. A second transcriptional element is located downstream of the enhancer between positions -311 and -213. This element together with the enhancer was sufficient to bring about transformation of Rat-1 cells at nearly wild-type frequencies, and together these elements constitute the minimal sequences required for gene expression in vivo. The sequences making up the second element may be functionally duplicated downstream of position -165 (between positions -165 and -60). This was revealed by the characterization of mutant genomes with deletions between positions -349 and -60. The role of these redundant elements is not known; however, they may be analogous to the 21-base-pair repeats of simian virus 40. Finally, sequences between positions -57 and -1 were required for accurate and efficient transcription in vitro. However, this DNA stretch, which includes the TATA box and major transcriptional start sites, was not absolutely required for gene expression in vivo. We conclude that the polyomavirus promoter comprises multiple functional elements which are distributed across a DNA stretch of about 400 base pairs.

scholarly journals Retinoic acid increases zif268 early gene expression in rat preosteoblastic cells.

1991 ◽  
Vol 11 (5) ◽  
pp. 2503-2510 ◽  
Author(s):  
L J Suva ◽  
M Ernst ◽  
G A Rodan
Keyword(s):  
Retinoic Acid ◽  
Primary Cultures ◽  
Simian Virus 40 ◽  
Simian Virus ◽  
Early Gene ◽  
Regulatory Sequences ◽  
Fetal Rat ◽  
In Vitro Data

In this study we demonstrate that retinoic acid (RA) increases the expression of transcription factor zif268 mRNA in primary cultures of fetal rat calvarial cells and in simian virus 40-immortalized clonal rat calvarial preosteoblastic cells (RCT-1), which differentiate in response to RA, but not in the more differentiated RCT-3 and ROS 17/2.8 cells. The increased expression of zif268 mRNA is rapid (maximal within 1 h), transient (returns to basal levels by 3 h), detectable at RA doses of 10(-12)M, and independent of protein synthesis. The relative stimulation of zif268 mRNA by RA was much larger than that of other early genes, including c-fos, c-jun, and junB. The rate of transcription of RA-stimulated RCT-1 cells, estimated by nuclear run-on assays, was elevated, suggesting that RA regulation of zif268 gene transcription was at least in part transcriptional. Moreover, RA stimulated the transcriptional activity of a Zif268CAT (chloramphenicol acetyltransferase) plasmid containing 632 bp of zif268 5' regulatory sequences in RCT-1 cells but not in the more differentiated RCT-3 cells. These in vitro data support the in vivo observations which localize zif268 and RA receptor-gamma transcripts to bone and cartilage during development, suggesting that both RA and zif268 may play a role in osteoblast differentiation.

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