A comparative analysis of the chromosomes of three FARQ species complex members, Ceratitis rosa, C. quilicii, and C. fasciventris F2 (Diptera: Tephritidae).

The Ceratitis FARQ species complex consists of four highly destructive agricultural pests of Africa, namely C. fasciventris, C. anonae, C. rosa, and C. quilicii. The members of the complex are considered very closely related and the species limits among them are rather obscure. Their economic significance and the need for developing biological methods for their control makes species identification within the complex an important issue, which has become clear that can only be addressed by multidisciplinary approaches. Chromosomes, both mitotic and polytene, can provide a useful tool for species characterization and phylogenetic inference among closely related dipteran species. In the current study, we present the mitotic karyotype and the polytene chromosomes of C. rosa and C. quilicii together with in situ hybridization data. We performed a comparative cytogenetic analysis among the above two species and C. fasciventris, the only other cytogenetically studied member of the FARQ complex, by comparing the mitotic complement and the banding pattern of the polytene chromosomes of each species to the others, as well as by studying the polytene chromosomes of hybrids between them. Our analysis revealed no detectable chromosomal rearrangements discriminating the three FARQ members studied, confirming their close phylogenetic relationships.

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

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.

The Effect of an Irradiation-Induced Recombination Suppressing Inversion on the Genetic Stability and Biological Quality of a White Eye-Based Aedes aegypti Genetic Sexing Strain.

Aedes aegypti is the primary vector of diseases such as dengue, chikungunya, Zika fever, and yellow fever. The sterile insect technique (SIT) has been proposed as a species-specific and environment-friendly tool for the suppression of mosquito vector populations as a major component of integrated vector management strategies. As female mosquitoes are blood-feeders and may transmit pathogenic microorganisms, mosquito SIT depends on the release of sterile males. Genetic sexing strains (GSS) can be used for the efficient and robust separation of males from females. Two Ae. aegypti GSS were recently developed by exploiting eye colour mutations, resulting in the Red-eye GSS (RGSS) and the White-eye GSS (WGSS). In this study, we compared two WGSS, with and without the chromosomal inversion 35 (Inv35), and evaluated their biological quality, including genetic stability. Our results suggest that the WGSS/Inv35 presents a low recombination rate and long-term genetic stability when recombinants are removed from the colony (filtering) and a slow accumulation of recombinants when they are not removed from the colony (non-filtering). The two strains were similar with respect to fecundity, pupal and adult recovery rates, pupation curve, and pupal weight. However, differences were detected in fertility, survival rate of females, and flight ability of males. The WGSS/Inv35 presented lower fertility, higher survival rate of females, and better flight ability of males compared to the WGSS.

Marker-assisted mapping enables forward genetic analysis in Aedes aegypti, an arboviral vector with vast recombination deserts.

Aedes aegypti is a major vector of arboviruses that cause dengue, chikungunya, yellow fever, and Zika. Although recent success in reverse genetics has facilitated rapid progress in basic and applied research, integration of forward genetics with modern technologies remains challenging in this important species, as up to 47% of its chromosome is refractory to genetic mapping due to extremely low rate of recombination. Here, we report the development of a marker-assisted mapping strategy to readily screen for and genotype only the rare but informative recombinants, drastically increasing both the resolution and signal-to-noise ratio. Using marker-assisted mapping, we mapped a transgene that was inserted in a >100-Mb recombination desert and a sex-linked spontaneous red-eye (re) mutation just outside the region. We subsequently determined, by CRISPR/Cas9-mediated knockout, that cardinal is the causal gene of re, which is the first forward genetic identification of a causal gene in Ae. aegypti. The identification of the causal gene of the sex-linked re mutation provides the molecular foundation for using gene editing to develop versatile and stable genetic sexing methods. To facilitate genome-wide forward genetics in Ae. aegypti, we generated and compiled a number of lines with markers throughout the genome. Thus, by overcoming the challenges presented by the vast recombination deserts and the scarcity of markers, we have shown that effective forward genetic analysis is increasingly feasible in this important arboviral vector species.

Genetic Stability and Fitness of Aedes aegypti Red-Eye Genetic Sexing Strains With Pakistani Genomic Background for Sterile Insect Technique Applications.

The mosquito species Aedes aegypti is the primary transmitter of viruses that cause endemic diseases like dengue in Pakistan. It is also a cause of other vector-borne diseases like yellow fever, Zika fever, and chikungunya, which significantly impact human health worldwide. In the absence of efficient vaccines (except for yellow fever) or drugs, vector control methods, such as the sterile insect technique (SIT), have been proposed as additional tools for the management of these diseases. Mosquito SIT programs are based on the release of sterile males and it is important female releases to be ideally zero or to be kept at a minimum, since females are the ones that bite, blood-feed and transmit pathogens. Recently, an Ae. aegypti genetic sexing strain (GSS), with and without a recombination-suppressing inversion (Inv35), was developed using the eye color as a selectable marker, with males having black eyes and females red eyes. In the present study, we introgressed the sexing features and the Inv35 of the Ae. aegypti red-eye GSS into the Pakistani genomic background aiming to their future use for SIT applications in the country. Both introgressed strains, the Red-eye GSS-PAK and the Red-eye GSS/Inv35-PAK, were evaluated in respect to their genetic stability and biological quality by assessing parameters like recombination rate, fecundity, fertility, pupal and adult recovery, time of development, pupal weight, survival, and flight ability in comparison with a wild Pakistani population (PAK). The results suggest that the sexing features and the recombination suppression properties of Inv35 were not affected after their introgression into the local genomic background; however, some biological traits of the two newly constructed strains were affected, positively or negatively, suggesting that a thorough quality control analysis should be performed after the introgression of a GSS into a new genomic background prior to its use in SIT field trials or applications. The importance of using GSS with local genomic background for SIT applications against Aedes aegypti is also discussed.

Introgression of the Aedes aegypti Red-Eye Genetic Sexing Strains Into Different Genomic Backgrounds for Sterile Insect Technique Applications.

Aedes aegypti is an invasive mosquito species and major vector of human arboviruses. A wide variety of control methods have been employed to combat mosquito populations. One of them is the sterile insect technique (SIT) that has recently attracted considerable research efforts due to its proven record of success and the absence of harmful environmental footprints. The efficiency and cost-effectiveness of SIT is significantly enhanced by male-only releases. For mosquito SIT, male-only releases are ideally needed since females bite, blood-feed and transmit the pathogens. Ae. aegypti genetic sexing strains (GSS) have recently become available and are based on eye colour mutations that were chosen as selectable markers. These genetic sexing strains were developed through classical genetics and it was shown to be subjected to genetic recombination, a phenomenon that is not suppressed in males as is the case in many Diptera. The genetic stability of these GSS was strengthened by the induction and isolation of radiation-induced inversions. In this study, we used the red eye mutation and the inversion Inv35 line of the Ae. aegypti red-eye GSS s and introgressed them in six different genomic backgrounds to develop GSS with the respective local genomic backgrounds. Our goal was to assess whether the recombination frequencies in the strains with and without the inversion are affected by the different genomic backgrounds. In all cases the recombination events were suppressed in all Inv35 GSS strains, thus indicating that the genomic background does not negatively affect the inversion result. Absence of any effect that could be ascribed to genetic differences, enables the introgression of the key elements of the GSS into the local genomic background prior to release to the target areas. Maintaining the local background increases the chances for successful matings between released males and wild females and addresses potential regulatory concerns regarding biosafety and biosecurity.

Interactions Between Tsetse Endosymbionts and Glossina pallidipes Salivary Gland Hypertrophy Virus in Glossina Hosts.

Tsetse flies are the sole cyclic vector for trypanosomosis, the causative agent for human African trypanosomosis or sleeping sickness and African animal trypanosomosis or nagana. Tsetse population control is the most efficient strategy for animal trypanosomosis control. Among all tsetse control methods, the Sterile Insect Technique (SIT) is one of the most powerful control tactics to suppress or eradicate tsetse flies. However, one of the challenges for the implementation of SIT is the mass production of target species. Tsetse flies have a highly regulated and defined microbial fauna composed of three bacterial symbionts (Wigglesworthia, Sodalis and Wolbachia) and a pathogenic Glossina pallidipes Salivary Gland Hypertrophy Virus (GpSGHV) which causes reproduction alterations such as testicular degeneration and ovarian abnormalities with reduced fertility and fecundity. Interactions between symbionts and GpSGHV might affect the performance of the insect host. In the present study, we assessed the possible impact of GpSGHV on the prevalence of tsetse endosymbionts under laboratory conditions to decipher the bidirectional interactions on six Glossina laboratory species. The results indicate that tsetse symbiont densities increased over time in tsetse colonies with no clear impact of the GpSGHV infection on symbionts density. However, a positive correlation between the GpSGHV and Sodalis density was observed in Glossina fuscipes species. In contrast, a negative correlation between the GpSGHV density and symbionts density was observed in the other taxa. It is worth noting that the lowest Wigglesworthia density was observed in G. pallidipes, the species which suffers most from GpSGHV infection. In conclusion, the interactions between GpSGHV infection and tsetse symbiont infections seems complicated and affected by the host and the infection density of the GpSGHV and tsetse symbionts.

Τhe complete mitochondrial genomes of Ceratitis rosa and Ceratitis quilicii, members of the Ceratitis FAR species complex (Diptera: Tephritidae).

Ceratitis FAR is an African species complex comprising insect pests of great economic interest and obscure species limits. Here, we report the mitochondrial genomes of two members of the FAR complex, namely Ceratitis rosa and the recently characterized Ceratitis quilicii. A phylogenetic analysis based on PCGs of available Tephritidae mitogenomes is presented. The current mitochondrial sequences from the FAR complex could contribute toward the resolution of phylogenetic relationships and species limits within this taxonomically challenging group, which is also an important issue for the development of environment-friendly and species-specific control methods, such as the sterile insect technique (SIT).

Genetic sexing strains for the population suppression of the mosquito vector Aedes aegypti.

Aedes aegypti is the primary vector of arthropod-borne viruses including dengue, chikungunya and Zika. Vector population control methods are reviving to impede disease transmission. An efficient sex separation for male-only releases is crucial for area-wide mosquito population suppression strategies. Here, we report on the construction of two genetic sexing strains using red- and white-eye colour mutations as selectable markers. Quality control analysis showed that the Red-eye genetic sexing strains (GSS) is better and more genetically stable than the White-eye GSS. The introduction of an irradiation-induced inversion (Inv35) increases genetic stability and reduces the probability of female contamination of the male release batches. Bi-weekly releases of irradiated males of both the Red-eye GSS and the Red-eye GSS/Inv35 fully suppressed target laboratory cage populations within six and nine weeks, respectively. An image analysis algorithm allowing sex determination based on eye colour identification at the pupal stage was developed. The next step is to automate the Red-eye-based genetic sexing and validate it in pilot trials prior to its integration in large-scale population suppression programmes. This article is part of the theme issue 'Novel control strategies for mosquito-borne diseases'.

Irradiation induced inversions suppress recombination between the M locus and morphological markers in Aedes aegypti.

BACKGROUND: Aedes aegypti is the primary vector of arthropod-borne viruses and one of the most widespread and invasive mosquito species. Due to the lack of efficient specific drugs or vaccination strategies, vector population control methods, such as the sterile insect technique, are receiving renewed interest. However, availability of a reliable genetic sexing strategy is crucial, since there is almost zero tolerance for accidentally released females. Development of genetic sexing strains through classical genetics is hindered by genetic recombination that is not suppressed in males as is the case in many Diptera. Isolation of naturally-occurring or irradiation-induced inversions can enhance the genetic stability of genetic sexing strains developed through genetically linking desirable phenotypes with the male determining region. RESULTS: For the induction and isolation of inversions through irradiation, 200 male pupae of the 'BRA' wild type strain were irradiated at 30 Gy and 100 isomale lines were set up by crossing with homozygous 'red-eye' (re) mutant females. Recombination between re and the M locus and the white (w) gene (causing a recessive white eye phenotype when mutated) and the M locus was tested in 45 and 32 lines, respectively. One inversion (Inv35) reduced recombination between both re and the M locus, and wand the M locus, consistent with the presence of a rather extended inversion between the two morphological mutations, that includes the M locus. Another inversion (Inv5) reduced recombination only between w and the M locus. In search of naturally-occurring, recombination-suppressing inversions, homozygous females from the red eye and the white eye strains were crossed with seventeen and fourteen wild type strains collected worldwide, representing either recently colonized or long-established laboratory populations. Despite evidence of varying frequencies of recombination, no combination led to the elimination or substantial reduction of recombination. CONCLUSION: Inducing inversions through irradiation is a feasible strategy to isolate recombination suppressors either on the M or the m chromosome for Aedes aegypti. Such inversions can be incorporated in genetic sexing strains developed through classical genetics to enhance their genetic stability and support SIT or other approaches that aim to population suppression through male-delivered sterility.

Genetic structure and symbiotic profile of worldwide natural populations of the Mediterranean fruit fly, Ceratitis capitata.

BACKGROUND: The Mediterranean fruit fly, Ceratitis capitata, is a cosmopolitan agricultural pest of worldwide economic importance and a model for the development of the Sterile Insect Technique (SIT) for fruit flies of the Tephritidae family (Diptera). SIT relies on the effective mating of laboratory-reared strains and natural populations, and therefore requires an efficient mass-rearing system that will allow for the production of high-quality males. Adaptation of wild flies to an artificial laboratory environment can be accompanied by negative effects on several life history traits through changes in their genetic diversity and symbiotic communities. Such changes may lead to reduced biological quality and mating competitiveness in respect to the wild populations. Profiling wild populations can help understand, and maybe reverse, deleterious effects accompanying laboratory domestication thus providing insects that can efficiently and effectively support SIT application. RESULTS: In the present study, we analyzed both the genetic structure and gut symbiotic communities of natural medfly populations of worldwide distribution, including Europe, Africa, Australia, and the Americas. The genetic structure of 408 individuals from 15 distinct populations was analyzed with a set of commonly used microsatellite markers. The symbiotic communities of a subset of 265 individuals from 11 populations were analyzed using the 16S rRNA gene-based amplicon sequencing of single individuals (adults). Genetic differentiation was detected among geographically distant populations while adults originated from neighboring areas were genetically closer. Alpha and beta diversity of bacterial communities pointed to an overall reduced symbiotic diversity and the influence of the geographic location on the bacterial profile. CONCLUSIONS: Our analysis revealed differences both in the genetic profile and the structure of gut symbiotic communities of medfly natural populations. The genetic analysis expanded our knowledge to populations not analyzed before and our results were in accordance with the existing scenarios regarding this species expansion and colonization pathways. At the same time, the bacterial communities from different natural medfly populations have been characterized, thus broadening our knowledge on the microbiota of the species across its range. Genetic and symbiotic differences between natural and laboratory populations must be considered when designing AW-IPM approaches with a SIT component, since they may impact mating compatibility and mating competitiveness of the laboratory-reared males. In parallel, enrichment from wild populations and/or symbiotic supplementation could increase rearing productivity, biological quality, and mating competitiveness of SIT-important laboratory strains.

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.

Multiple Factors Determine the Structure of Bacterial Communities Associated With Aedes albopictus Under Artificial Rearing Conditions.

Insect symbionts are major manipulators of host's behavior. Their effect on parameters such as fecundity, male mating competitiveness, and biological quality in general, can have a major influence on the effectiveness of the sterile insect technique (SIT). SIT is currently being developed and applied against human disease vectors, including Ae. albopictus, as an environment-friendly method of population suppression, therefore there is a renewed interest on both the characterization of gut microbiota and their exploitation in artificial rearing. In the present study, bacterial communities of eggs, larvae, and adults (both males and females) of artificially reared Ae. albopictus, were characterized using both culture-dependent and culture-independent approaches. Mosquito-associated bacteria corresponding to thirteen and five bacteria genera were isolated from the larval food and the sugar solution (adult food), respectively. The symbiont community of the females was affected by the provision of a blood meal. Pseudomonas and Enterobacter were either introduced or enhanced with the blood meal, whereas Serratia were relatively stable during the adult stage of females. Maintenance of these taxa in female guts is probably related with blood digestion. Gut-associated microbiota of males and females were different, starting early after emergence and continuing in older stages. Our results indicate that eggs contained bacteria from more than fifteen genera including Bacillus, Chryseobacterium, and Escherichia-Shigella, which were also main components of gut microbiota of female adults before and after blood feeding, indicating potential transmission among generations. Our results provided a thorough study of the egg- and gut-associated bacteria of artificially reared Ae. albopictus, which can be important for further studies using probiotic bacteria to improve the effectiveness of mosquito artificial rearing and SIT applications.

Wolbachia infection in Argentinean populations of Anastrepha fraterculus sp1: preliminary evidence of sex ratio distortion by one of two strains.

BACKGROUND: Wolbachia, one of the most abundant taxa of intracellular Alphaproteobacteria, is widespread among arthropods and filarial nematodes. The presence of these maternally inherited bacteria is associated with modifications of host fitness, including a variety of reproductive abnormalities, such as cytoplasmic incompatibility, thelytokous parthenogenesis, host feminization and male-killing. Wolbachia has attracted much interest for its role in biological, ecological and evolutionary processes as well as for its potential use in novel and environmentally-friendly strategies for the control of insect pests and disease vectors including a major agricultural pest, the South American fruit fly, Anastrepha fraterculus Wiedemann (Diptera: Tephritidae). RESULTS: We used wsp, 16S rRNA and a multilocus sequence typing (MLST) scheme including gatB, coxA, hcpA, fbpA, and ftsZ genes to detect and characterize the Wolbachia infection in laboratory strains and wild populations of A. fraterculus from Argentina. Wolbachia was found in all A. fraterculus individuals studied. Nucleotide sequences analysis of wsp gene allowed the identification of two Wolbachia nucleotide variants (named wAfraCast1_A and wAfraCast2_A). After the analysis of 76 individuals, a high prevalence of the wAfraCast2_A variant was found both, in laboratory (82%) and wild populations (95%). MLST analysis identified both Wolbachia genetic variants as sequence type 13. Phylogenetic analysis of concatenated MLST datasets clustered wAfraCast1/2_A in the supergroup A. Paired-crossing experiments among single infected laboratory strains showed a phenotype specifically associated to wAfraCast1_A that includes slight detrimental effects on larval survival, a female-biased sex ratio; suggesting the induction of male-killing phenomena, and a decreased proportion of females producing descendants that appears attributable to the lack of sperm in their spermathecae. CONCLUSIONS: We detected and characterized at the molecular level two wsp gene sequence variants of Wolbachia both in laboratory and wild populations of A. fraterculus sp.1 from Argentina. Crossing experiments on singly-infected A. fraterculus strains showed evidence of a male killing-like mechanism potentially associated to the wAfraCast1_A - A. fraterculus interactions. Further mating experiments including antibiotic treatments and the analysis of early and late immature stages of descendants will contribute to our understanding of the phenotypes elicited by the Wolbachia variant wAfraCast1_A in A. fraterculus sp.1.

Enterobacter sp. AA26 gut symbiont as a protein source for Mediterranean fruit fly mass-rearing and sterile insect technique applications.

BACKGROUND: Insect species have established sophisticated symbiotic associations with diverse groups of microorganisms including bacteria which have been shown to affect several aspects of their biology, physiology, ecology and evolution. In addition, recent studies have shown that insect symbionts, including those localized in the gastrointestinal tract, can be exploited for the enhancement of sterile insect technique (SIT) applications against major insect pests such as the Mediterranean fruit fly (medfly) Ceratitis capitata. We previously showed that Enterobacter sp. AA26 can be used as probiotic supplement in medfly larval diet improving the productivity and accelerating the development of the VIENNA 8 genetic sexing strain (GSS), which is currently used in large scale operational SIT programs worldwide. RESULTS: Enterobacter sp. AA26 was an adequate nutritional source for C. capitata larvae, comprising an effective substitute for brewer's yeast. Incorporating inactive bacterial cells in the larval diet conferred a number of substantial beneficial effects on medfly biology. The consumption of bacteria-based diet (either as full or partial yeast replacement) resulted in decreased immature stages mortality, accelerated immature development, increased pupal weight, and elongated the survival under stress conditions. Moreover, neither the partial nor the complete replacement of yeast with Enterobacter sp. AA26 had significant impact on adult sex ratio, females' fecundity, adults' flight ability and males' mating competitiveness. The absence of both yeast and Enterobacter sp. AA26 (deprivation of protein source and possible other important nutrients) from the larval diet detrimentally affected the larval development, survival and elongated the immature developmental duration. CONCLUSIONS: Enterobacter sp. AA26 dry biomass can fully replace the brewer's yeast as a protein source in medfly larval diet without any effect on the productivity and the biological quality of reared medfly of VIENNA 8 GSS as assessed by the FAO/IAEA/USDA standard quality control tests. We discuss this finding in the context of mass-rearing and SIT applications.

Τhe Complete Mitochondrial Genome of Bactrocera carambolae (Diptera: Tephritidae): Genome Description and Phylogenetic Implications.

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

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.

Medfly-Wolbachia symbiosis: genotype x genotype interactions determine host's life history traits under mass rearing conditions.

BACKGROUND: Wolbachia pipientis is a widespread, obligatory intracellular and maternally inherited bacterium, that induces a wide range of reproductive alterations to its hosts. Cytoplasmic Incompatibility (CI) is causing embryonic lethality, the most common of them. Despite that Wolbachia-borne sterility has been proposed as an environmental friendly pest control method (Incompatible Insect Technique, IIT) since 1970s, the fact that Wolbachia modifies important fitness components of its hosts sets severe barriers to IIT implementation. Mass rearing of Mediterranean fruit fly, Ceratitis capitata (medfly), is highly optimized given that this pest is a model species regarding the implementation of another sterility based pest control method, the Sterile Insect Technique (SIT). We used the medfly-Wolbachia symbiotic association, as a model system, to study the effect of two different Wolbachia strains, on the life history traits of 2 C. capitata lines with different genomic background. RESULTS: Wolbachia effects are regulated by both C. capitata genetic background and the Wolbachia strain. Wolbachia infection reduces fertility rates in both C. capitata genetic backgrounds and shortens the pre-pupa developmental duration in the GSS strain. On the other hand, regardless of the strain of Wolbachia (wCer2, wCer4) infection does not affect either the sex ratio or the longevity of adults. wCer4 infection imposed a reduction in females' fecundity but wCer2 did not. Male mating competitiveness, adults flight ability and longevity under water and food deprivation were affected by both the genetic background of medfly and the strain of Wolbachia (genotype by genotype interaction). CONCLUSION: Wolbachia infection could alter important life history traits of mass-reared C. capitata lines and therefore the response of each genotype on the Wolbachia infection should be considered toward ensuring the productivity of the Wolbachia-infected insects under mass-rearing conditions.

Taxonomy, Diet, and Developmental Stage Contribute to the Structuring of Gut-Associated Bacterial Communities in Tephritid Pest Species.

Insect-symbiont interactions are receiving much attention in the last years. Symbiotic communities have been found to influence a variety of parameters regarding their host physiology and fitness. Gut symbiotic communities can be dynamic, changing through time and developmental stage. Whether these changes represent real differential needs and preferential relationships has not been addressed yet. In this study, we characterized the structure of symbiotic communities of five laboratory populations that represent five Tephritidae species that are targets for pest control management through the sterile insect technique (SIT), namely Bactrocera oleae, Anastrepha grandis, Anastrepha ludens, and two morphotypes of Anastrepha fraterculus (sp.1 and the Andean lineage). These populations are under artificial or semi artificial rearing conditions and their characterization was performed for different developmental stages and age. Our results demonstrate the presence of a symbiotic community comprising mainly from different Enterobacteriaceae genera. These communities are dynamic across developmental stages, although not highly variable, and appear to have a species-specific profile. Additional factors may contribute to the observed structuring, including diet, rearing practices, and the degree of domestication. Comparison of these results with those derived from natural populations could shed light to changes occurring in the symbiotic level during domestication of Tephritidae populations. Further studies will elucidate whether the changes are associated with modification of the behavior in laboratory strains and assess their effects in the quality of the mass rearing insects. This could be beneficial for improving environmentally friendly, species-specific, pest control methods, such as the SIT.

The chromosomes of Drosophila suzukii (Diptera: Drosophilidae): detailed photographic polytene chromosomal maps and in situ hybridization data.

The spotted wing drosophila, D. suzukii, is a serious agricultural pest attacking a variety of soft fruits and vegetables. Although originating from East Asia it has recently invaded America and Europe raising major concern about its expansion potential and the consequent economic losses. Since cytogenetic information on the species is scarce, we report here the mitotic karyotype and detailed photographic maps of the salivary gland polytene chromosomes of D. suzukii. The mitotic metaphase complement contains three pairs of autosomes, one of which is dot-like, and one pair of heteromorphic (XX/XY) sex chromosomes. The salivary gland polytene complement consists of five long polytene arms, representing the two metacentric autosomes and the acrocentric X chromosome, and one very short polytene element, which corresponds to the dot-like autosome. Banding pattern as well as the most characteristic features and prominent landmarks of each polytene chromosome arm are presented and discussed. Furthermore, twelve gene markers have been mapped on the polytene chromosomes of D. suzukii by in situ hybridization. Their distribution pattern was found quite similar to that of D. melanogaster revealing conservation of synteny although the relative position within each chromosome arm for most of the genes differed significantly between D. suzukii and D. melanogaster. The chromosome information presented here is suitable for comparative cytogenetic studies and phylogenetic exploration, while it could also facilitate the assembly of the genome sequence and support the development of genetic tools for species-specific and environment-friendly biological control applications such as the sterile insect technique.