Mitochondrial Superoxide Dismutase Overexpression and Low Oxygen Conditioning Hormesis Improve the Performance of Irradiated Sterile Males

The Sterile Insect Technique (SIT) is a successful autocidal control method that uses ionizing radiation to sterilize insects. However, irradiation in normal atmospheric conditions can be damaging for males, because irradiation generates substantial biological oxidative stress that, combined with domestication and mass-rearing conditions, may reduce sterile male sexual competitiveness and quality. In this study, biological oxidative stress and antioxidant capacity were experimentally manipulated in Anastrepha suspensa using a combination of low-oxygen conditions and transgenic overexpression of mitochondrial superoxide dismutase (SOD2) to evaluate their role in the sexual behavior and quality of irradiated males. Our results showed that SOD2 overexpression enhances irradiated insect quality and improves male competitiveness in leks. However, the improvements in mating performance were modest, as normoxia-irradiated SOD2 males exhibited only a 22% improvement in mating success compared to normoxia-irradiated wild type males. Additionally, SOD2 overexpression did not synergistically improve the mating success of males irradiated in either hypoxia or severe hypoxia. Short-term hypoxic and severe-hypoxic conditioning hormesis, per se, increased antioxidant capacity and enhanced sexual competitiveness of irradiated males relative to non-irradiated males in leks. Our study provides valuable new information that antioxidant enzymes, particularly SOD2, have potential to improve the quality and lekking performance of sterile males used in SIT programs

Prevalence of trypanosomes, salivary gland hypertrophy virus and Wolbachia in wild populations of tsetse flies from West Africa

Background: Tsetse flies are vectors of African trypanosomes, protozoan parasites that cause sleeping sickness (or human African trypanosomosis) in humans and nagana (or animal African trypanosomosis) in livestock. In addition to trypanosomes, four symbiotic bacteria Wigglesworthia glossinidia, Sodalis glossinidius, Wolbachia, Spiroplasma and one pathogen, the salivary gland hypertrophy virus (SGHV), have been reported in different tsetse species. We evaluated the prevalence and coinfection dynamics between Wolbachia, trypanosomes, and SGHV in four tsetse species (Glossina palpalis gambiensis, G. tachinoides, G. morsitans submorsitans, and G. medicorum) that were collected between 2008 and 2015 from 46 geographical locations in West Africa, i.e. Burkina Faso, Mali, Ghana, Guinea, and Senegal. Results: The results indicated an overall low prevalence of SGHV and Wolbachia and a high prevalence of trypanosomes in the sampled wild tsetse populations. The prevalence of all three infections varied among tsetse species and sample origin. The highest trypanosome prevalence was found in Glossina tachinoides (61.1%) from Ghana and in Glossina palpalis gambiensis (43.7%) from Senegal. The trypanosome prevalence in the four species from Burkina Faso was lower, i.e. 39.6% in Glossina medicorum, 18.08%; in Glossina morsitans submorsitans, 16.8%; in Glossina tachinoides and 10.5% in Glossina palpalis gambiensis. The trypanosome prevalence in Glossina palpalis gambiensis was lowest in Mali (6.9%) and Guinea (2.2%). The prevalence of SGHV and Wolbachia was very low irrespective of location or tsetse species with an average of 1.7% for SGHV and 1.0% for Wolbachia. In some cases, mixed infections with different trypanosome species were detected. The highest prevalence of coinfection was Trypanosoma vivax and other Trypanosoma species (9.5%) followed by coinfection of T. congolense with other trypanosomes (7.5%). The prevalence of coinfection of T. vivax and T. congolense was (1.0%) and no mixed infection of trypanosomes, SGHV and Wolbachia was detected. Conclusion: The results indicated a high rate of trypanosome infection in tsetse wild populations in West African countries but lower infection rate of both Wolbachia and SGHV. Double or triple mixed trypanosome infections were found. In addition, mixed trypanosome and SGHV infections existed however no mixed infections of trypanosome and/or SGHV with Wolbachia were found

Verbesserung der Sterilinsekten-Technik für Tsetse-Fliegen durch Erforschung ihrer Symbionten und Pathogene

Arbeit an der Bibliothek noch nicht eingelangt - Daten nicht geprueftAbweichender Titel nach Übersetzung der Verfasserin/des VerfassersAfrikanische Trypanosomiasen sind bedeutende, jedoch wenig beachtete Tropenkrankheiten der Menschen (Afrikanische Schlafkrankheit oder Human African Trypanosomiasis (HAT)), und Tieren (Nagana, oder Animal African Trypanosomiasis (AAT)), die in den subsaharischen Regionen Afrikas vorkommen und durch Tsetsefliegen übertragen werden (Dipteria: Glossinidae). Aufgrund fehlender effektiver Impfungen, der zunehmenden Resistenz gegen anti-trypanosomale Medikamente, und des Mangels an verfügbaren und leistbaren Therapien gegen diese Erkrankungen ist die Kontrolle der Tsetsefliegen Populationen momentan die effektivste Lösung um diese Krankheitsvorkommen zu reduzieren. Die Sterile Insekten Technik (SIT) ist eine erwiesene, effektive und nachhaltige Methode, wenn sie im Rahmen einer flächendeckenden, integrierten Schädlingsbekämpfungsstrategie ("Area-Wide Integrated Pest Management (AW-IPM)”) für die Dezimierung oder Ausrottung der Tsetsefliegen Populationen angewendet wird. Die SIT benötigt die künstliche Erzeugung einer großen Anzahl des Schadeninsekts, die Sterilisierung der Männchen durch ionisierende Strahlung, und deren Aussetzung in das Zielgebiet. Diese sterilen Männchen konkurrieren mit den in der Natur lebenden fertilen Männchen um die Weibchen, und erfolgreiche Paarungen der sterilen Männchen erzeugen kein Nachkommen. Bei einer verhältnismäßig höheren Anzahl der freigelassenen sterilen Männchen im Vergleich zu den natürlich vorhandenen fertilen Männchen steigen die Chancen der gewollten Paarungen und dadurch kann die Populationen des Zielinsekts stark verringert und sogar komplett ausgerottet werden. Trotz erfolgreiches Einsetzen der SIT/AW-IPM gegen Tsetsefliegen sind einige Beschränkungen vorhanden, die überwunden werden können um die Effizienz der Technik zu verbessern. Eine dieser Beschränkungen ist die niedrige Produktivität dieser Insekten in der Zucht, und deren Infektion mit einem schädlichen Virus. Eine zusätzliche Problematik ist die Fähigkeit der männlichen Tsetsefliegen, wegen deren haematophagen Ernährungsverhalten die erwähnten Krankheiten zu übertragen. Dadurch entsteht das Risiko einer erhöhten Übertragungsrate in der Region. Diese Probleme könnten gelöst werden, indem man die Interaktionen zwischen der Tsetsefliege und deren Symbionten und Pathogenen besser zu verstehen lernt. Sowohl Laborkolonien, als auch in der Natur existierenden Tsetsefliegenpopulationen sind zu hohem Anteil mit der Glossina pallidipes salivary gland hypertrophy virus (GpSGHV; Familie hytrosaviridae) infiziert und sind gleichzeitig Träger der endosymbiotischen Bakterien Wigglesworthia glossinidius, Sodalis glossinidia, Wolbachia, und Spiroplasma. GpSGHV infizierte Glossina pallidipes Fliegen zeigen Abnormitäten in den Ovarien und Testikel und Speicheldrüsen-Hypertrophie (salivary gland hypertrophy (SGH)). Ohne einer Strategie zur Virenkontrolle kann eine SGH Virus-Infektion in der Tsetsefliegenkolonie deren Produktivität stark verringern und sogar zum völligen Kollaps bringen. Es ist deswegen wichtig, die Auswirkungen dieser Viren auf die verschiedenen Tsetsefliegenspezien zu eruieren. Zusätzlich ist sowohl die Auswertung der Wirkung radioaktiver Strahlen auf Tsetse Symbiontenals auch auf die Krankheitsübertragungskapazität der Fliegen entscheidend. Das Ziel dieser wissenschaftlicher Arbeit war: (i) die Identifizierung der Tsetsefliegen Spezies durch molekulare Methoden, (ii) die Prävalenz von Viren, Wolbachia, und Trypanosomen in natürlichen Populationen der Tsetsefliegen, (iii) die Suszeptibilität der verschiedenen Tsetse Spezies zu Viren, (iv) die Eruierung der Auswirkungen von GpSGHV Infektionen auf deren Wirten, (v) die Eruierung der Wirkung ionisierender Strahlen auf die Tsetse Symbionten Sodalis und die Möglichkeiten einer kombinierten Paratransgenese und SIT Methode, und (vi) die Eruierung der Auswirkung einer Depletion der Symbionten und radioaktiver Strahlung auf das kutikuläre Hydrokarbonprofil (KHD Profil) der Tsetsefliegen G. m. morsitans. Die Ergebnisse weisen darauf hin, dass verschiedene Tsetse Spezies anfällig auf GpSGHV sind, jedoch nur G. pallidipes die Fähigkeit einer transgenerationalen Übertragung der Viren aufweist. Ionisierende Strahlen haben weder Auswirkung auf das KHD Profil, noch auf die Krankheitsübertragungskapazität der Fliegen, jedoch konnte eine Auswirkung auf die Dichte der Tsetse Symbionten nachgewiesen werden. Die Bestrahlung von 22 Tage alten Puppen ermöglicht die Wiederherstellung der Sordalis Dichte und somit die Kombination von Paratransgenese und SIT, um das Risiko der Krankheitsübertragung in SIT Anwendungsgebieten zu verringern. Letztlich ermöglichen verschiedene molekulare Methoden eine kosteneffektive, einfache und präzise Speziesidentifikation.African trypanosomoses are major neglected tropical diseases in both humans (sleeping sickness or Human African Trypanosomosis (HAT)) and animals (nagana or Animal African Trypanosomosis (AAT)) in sub-Saharan Africa and are transmitted by cyclical vector tsetse flies (Dipteria: Glossinidae). Due to the lack of efficient vaccines, an increased resistance against anti-trypanosomal drugs, and lack of inexpensive drugs against the diseases, tsetse fly control is currently considered the most powerful, innovative and efficient pest control tactic. The Sterile Insect Technique (SIT) has been proven to be one of the most effective and sustainable methods when it is applied as a part of Area-Wide Integrated Pest Management (AW-IPM) programmes for the suppression and/or eradication of tsetse fly populations. SIT requires the production of a large-number of males, sterilized by exposure to ionizing radiation and their release into the target area where they compete with wild males for wild females. Matings between sterile males and virgin wild females result in no offspring. A repeated release of a higher ratio of sterile males to wild males increases the chances of successful sterile matings and thus results in the suppression and/or eradication of the target insect population over time. Despite the fact that successful implementation of SIT/AW-IPM programmes for eradication tsetse flies in several infested areas has been shown, several constraints need to be addressed to enhance its efficiency. One of the important challenges of the SIT for tsetse is the mass production of sufficient sterile males due to their low productivity and infection with a pathogenic virus. Another challenge facing tsetse SIT programs is the ability of sterile males to transmit the disease due to their hematophagous feeding nature, which represents a potential risk of increasing the disease incidence in a SIT treatment site. Some of the constraints could be solved by understanding the interactions of tsetse flies with their symbionts and pathogens. Laboratory colonies, as well as field populations of many tsetse species, are infected by Glossina pallidipes salivary gland hypertrophy virus (GpSGHV; family hytrosaviridae) and harbor endosymbiotic bacteria Wigglesworthia glossinidius, Sodalis glossinidia, Wolbachia, and Spiroplasma. GpSGHV infected flies of Glossina pallidipes show ovarian abnormalities and testicular degeneration and salivary gland hypertrophy (SGH) symptoms. In the absence of an effective virus management strategy, the SGHV can reduce the colony productivity and may even result in the collapse of the infected colony. Therefore, it is important to assess the virus’ impact on several tsetse species. In addition, it is important to assess the impact of irradiation on tsetse symbionts as well as on its vectorial capacity. The aim of this research was to investigate (i) tsetse species identification using molecular approaches, (ii) prevalence of virus, Wolbachia and trypanosome in natural populations, (iii) the susceptibility of virus to different laboratory tsetse species, (iv) the impact of GpSGHV infection on heterogenous tsetse hosts, (v) the impact of irradiation on the tsetse symbiont Sodalis and the potential of using a combined approach of paratransgenesis and SIT, and lastly (vi) the impact of symbiont depletion and radiation treatment on G. m. morsitans cuticular hydrocarbon profiles and mate choice. The results indicate the susceptibility of different tsetse species to GpSGHV infection, however only G. pallidipes permit virus trans-generation transmission. Irradiation did not affect the CHC profile or male vectorial capacity, however, it reduced the density of symbionts. Exposing treatment on 22-day old puparia to radiation allows a significant recovery of the Sodalis density which enables a combination of paratransgenesis and SIT to eliminate the risk of increasing the disease incidence in areas using SIT. Finally, novel molecular tools provide easy, cheap and precise species identification methods where morphological identification methods formerly lacked accuracy.25

Susceptibility of Tsetse Species to Glossina pallidipes Salivary Gland Hypertrophy Virus (GpSGHV)

Salivary gland hytrosaviruses (SGHVs, family Hytrosaviridae) are non-occluded dsDNA viruses that are pathogenic to some dipterans. SGHVs primarily replicate in salivary glands (SG), thereby inducing overt salivary gland hypertrophy (SGH) symptoms in their adult hosts. SGHV infection of non-SG tissues results in distinct pathobiologies, including reproductive dysfunctions in tsetse fly, Glossina pallidipes (Diptera: Glossinidae) and house fly. Infection with the G. pallidipes virus (GpSGHV) resulted in the collapse of several laboratory colonies, which hindered the implementation of area wide integrated pest management (AW-IPM) programs that had a sterile insect technique (SIT) component. Although the impact of GpSGHV infection has been studied in some detail in G. pallidipes, the impact of the virus infection on other tsetse species remains largely unknown. In the current study, we assessed the susceptibility of six Glossina species (G. pallidipes, G. brevipalpis, G. m. morsitans, G. m. centralis, G. f. fuscipes, and G. p. gambiensis) to GpSGHV infections, and the impact of the viral infection on the fly pupation rate, adult emergence, and virus replication and transmission from the larval to adult stages. We also evaluated the ability of the virus to infect conspecific Glossina species through serial passages. The results indicate that the susceptibility of Glossina to GpSGHV varied widely amongst the tested species, with G. pallidipes and G. brevipalpis being the most susceptible and most refractory to the virus, respectively. Further, virus injection into the hemocoel of teneral flies led to increased viral copy number over time, while virus injection into the third instar larvae delayed adult eclosion. Except in G. pallidipes, virus injection either into the larvae or teneral adults did not induce any detectable SGH symptoms, although virus infections were PCR-detectable in the fly carcasses. Taken together, our results indicate that although GpSGHV may only cause minor damage in the mass-rearing of tsetse species other than G. pallidipes, preventive control measures are required to avoid viral contamination and transmission in the fly colonies, particularly in the facilities where multiple tsetse species are reared

Impact of Glossina pallidipes salivary gland hypertrophy virus (GpSGHV) on a heterologous tsetse fly host, Glossina fuscipes fuscipes

Abstract Background Tsetse flies (Diptera: Glossinidae) are the vectors of African trypanosomosis, the causal agent of sleeping sickness in humans and nagana in animals. Glossina fuscipes fuscipes is one of the most important tsetse vectors of sleeping sickness, particularly in Central Africa. Due to the development of resistance of the trypanosomes to the commonly used trypanocidal drugs and the lack of effective vaccines, vector control approaches remain the most effective strategies for sustainable management of those diseases. The Sterile Insect Technique (SIT) is an effective, environment-friendly method for the management of tsetse flies in the context of area-wide integrated pest management programs (AW-IPM). This technique relies on the mass-production of the target insect, its sterilization with ionizing radiation and the release of sterile males in the target area where they will mate with wild females and induce sterility in the native population. It has been shown that Glossina pallidipes salivary gland hypertrophy virus (GpSGHV) infection causes a decrease in fecundity and fertility hampering the maintenance of colonies of the tsetse fly G. pallidipes. This virus has also been detected in different species of tsetse files. In this study, we evaluated the impact of GpSGHV on the performance of a colony of the heterologous host G. f. fuscipes, including the flies’ productivity, mortality, survival, flight propensity and mating ability and insemination rates. Results Even though GpSGHV infection did not induce SGH symptoms, it significantly reduced all examined parameters, except adult flight propensity and insemination rate. Conclusion These results emphasize the important role of GpSGHV management strategy in the maintenance of G. f. fuscipes colonies and the urgent need to implement measures to avoid virus infection, to ensure the optimal mass production of this tsetse species for use in AW-IPM programs with an SIT component

Combining paratransgenesis with SIT: impact of ionizing radiation on the DNA copy number of Sodalis glossinidius in tsetse flies

Abstract Background Tsetse flies (Diptera: Glossinidae) are the cyclical vectors of the causative agents of African Trypanosomosis, which has been identified as a neglected tropical disease in both humans and animals in many regions of sub-Saharan Africa. The sterile insect technique (SIT) has shown to be a powerful method to manage tsetse fly populations when used in the frame of an area-wide integrated pest management (AW-IPM) program. To date, the release of sterile males to manage tsetse fly populations has only been implemented in areas to reduce transmission of animal African Trypanosomosis (AAT). The implementation of the SIT in areas with Human African Trypanosomosis (HAT) would require additional measures to eliminate the potential risk associated with the release of sterile males that require blood meals to survive and hence, might contribute to disease transmission. Paratransgenesis offers the potential to develop tsetse flies that are refractory to trypanosome infection by modifying their associated bacteria (Sodalis glossinidius) here after referred to as Sodalis. Here we assessed the feasibility of combining the paratransgenesis approach with SIT by analyzing the impact of ionizing radiation on the copy number of Sodalis and the vectorial capacity of sterilized tsetse males. Results Adult Glossina morsitans morsitans that emerged from puparia irradiated on day 22 post larviposition did not show a significant decline in Sodalis copy number as compared with non-irradiated flies. Conversely, the Sodalis copy number was significantly reduced in adults that emerged from puparia irradiated on day 29 post larviposition and in adults irradiated on day 7 post emergence. Moreover, irradiating 22-day old puparia reduced the copy number of Wolbachia and Wigglesworthia in emerged adults as compared with non-irradiated controls, but the radiation treatment had no significant impact on the vectorial competence of the flies. Conclusion Although the radiation treatment significantly reduced the copy number of some tsetse fly symbionts, the copy number of Sodalis recovered with time in flies irradiated as 22-day old puparia. This recovery offers the opportunity to combine a paratransgenesis approach – using modified Sodalis to produce males refractory to trypanosome infection – with the release of sterile males to minimize the risk of disease transmission, especially in HAT endemic areas. Moreover, irradiation did not increase the vector competence of the flies for trypanosomes

Interactions between Glossina pallidipes salivary gland hypertrophy virus and tsetse endosymbionts in wild tsetse populations

Abstract Background Tsetse control is considered an effective and sustainable tactic for the control of cyclically transmitted trypanosomosis in the absence of effective vaccines and inexpensive, effective drugs. The sterile insect technique (SIT) is currently used to eliminate tsetse fly populations in an area-wide integrated pest management (AW-IPM) context in Senegal. For SIT, tsetse mass rearing is a major milestone that associated microbes can influence. Tsetse flies can be infected with microorganisms, including the primary and obligate Wigglesworthia glossinidia, the commensal Sodalis glossinidius, and Wolbachia pipientis. In addition, tsetse populations often carry a pathogenic DNA virus, the Glossina pallidipes salivary gland hypertrophy virus (GpSGHV) that hinders tsetse fertility and fecundity. Interactions between symbionts and pathogens might affect the performance of the insect host. Methods In the present study, we assessed associations of GpSGHV and tsetse endosymbionts under field conditions to decipher the possible bidirectional interactions in different Glossina species. We determined the co-infection pattern of GpSGHV and Wolbachia in natural tsetse populations. We further analyzed the interaction of both Wolbachia and GpSGHV infections with Sodalis and Wigglesworthia density using qPCR. Results The results indicated that the co-infection of GpSGHV and Wolbachia was most prevalent in Glossina austeni and Glossina morsitans morsitans, with an explicit significant negative correlation between GpSGHV and Wigglesworthia density. GpSGHV infection levels > 103.31 seem to be absent when Wolbachia infection is present at high density (> 107.36), suggesting a potential protective role of Wolbachia against GpSGHV. Conclusion The result indicates that Wolbachia infection might interact (with an undefined mechanism) antagonistically with SGHV infection protecting tsetse fly against GpSGHV, and the interactions between the tsetse host and its associated microbes are dynamic and likely species specific; significant differences may exist between laboratory and field conditions. Graphical Abstrac

Data_Sheet_1.PDF

<p>Salivary gland hytrosaviruses (SGHVs, family Hytrosaviridae) are non-occluded dsDNA viruses that are pathogenic to some dipterans. SGHVs primarily replicate in salivary glands (SG), thereby inducing overt salivary gland hypertrophy (SGH) symptoms in their adult hosts. SGHV infection of non-SG tissues results in distinct pathobiologies, including reproductive dysfunctions in tsetse fly, Glossina pallidipes (Diptera: Glossinidae) and house fly. Infection with the G. pallidipes virus (GpSGHV) resulted in the collapse of several laboratory colonies, which hindered the implementation of area wide integrated pest management (AW-IPM) programs that had a sterile insect technique (SIT) component. Although the impact of GpSGHV infection has been studied in some detail in G. pallidipes, the impact of the virus infection on other tsetse species remains largely unknown. In the current study, we assessed the susceptibility of six Glossina species (G. pallidipes, G. brevipalpis, G. m. morsitans, G. m. centralis, G. f. fuscipes, and G. p. gambiensis) to GpSGHV infections, and the impact of the viral infection on the fly pupation rate, adult emergence, and virus replication and transmission from the larval to adult stages. We also evaluated the ability of the virus to infect conspecific Glossina species through serial passages. The results indicate that the susceptibility of Glossina to GpSGHV varied widely amongst the tested species, with G. pallidipes and G. brevipalpis being the most susceptible and most refractory to the virus, respectively. Further, virus injection into the hemocoel of teneral flies led to increased viral copy number over time, while virus injection into the third instar larvae delayed adult eclosion. Except in G. pallidipes, virus injection either into the larvae or teneral adults did not induce any detectable SGH symptoms, although virus infections were PCR-detectable in the fly carcasses. Taken together, our results indicate that although GpSGHV may only cause minor damage in the mass-rearing of tsetse species other than G. pallidipes, preventive control measures are required to avoid viral contamination and transmission in the fly colonies, particularly in the facilities where multiple tsetse species are reared.</p