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Wolbachia are maternally inherited endosymbiotic bacteria, widespread among arthropods thanks to host reproductive manipulations that increase their prevalence into host populations. The most commonly observed manipulation is cytoplasmic incompatibility (CI). CI leads to embryonic death in crosses between i) infected males and uninfected females and ii) individuals infected with incompatible Wolbachia strains. CI can be conceptualized as a toxin-antidote system where a toxin deposited by Wolbachia in the sperm would induce embryonic death unless countered by an antidote produced by Wolbachia present in the eggs. In Drosophila melanogaster, transgenic expression of Wolbachia effector cidB revealed its function of CI-inducing toxin. Moreover in Culex pipiens, the diversity of cidB variants present in wPip strains accounts for the diversity in crossing-types. We conducted cytological analyses to determine the CI mechanisms that lead to embryonic death in C. pipiens, and assess whether diversity in crossing-types could be based on variations in these mechanisms. We revealed that paternal chromatin condensation and segregation defects during the first embryonic division are always responsible for embryonic death. The strongest observed defects lead to an exclusion of the paternal chromatin from the first zygotic division, resulting in haploid embryos unable to hatch. The proportion of unhatched haploid embryos, developing with only maternal chromatin, which reflects the frequency of strong defects can be considered as a proxy of CI intensity at the cellular level. We thus studied the putative effect of variations in crossing types and cidB diversification on CI defects intensity. Incompatible crosses involving distinct wPip strains revealed that CI defects intensity depends on the Wolbachia strains hosted by the males and is linked to the diversity of cidB genes harbored in their genomes. These results support that, additionally to its implication in C. pipiens crossing type variability, cidB diversification also influences the strength of CI embryonic defects. [ABSTRACT FROM AUTHOR]

Background: Aedes albopictus is one of the most invasive human disease vectors. Its control has been largely based on insecticides, such as the larvicide temephos. Temephos resistance has been associated with the up-regulation, through gene amplification, of two carboxylesterase (CCE) genes closely linked on the genome, capable of sequestering and metabolizing temephos oxon, the activated form of temephos. Principal findings: Here, we investigated the occurrence, geographical distribution and origin of the CCE amplicon in Ae. albopictus populations from several geographical regions worldwide. The haplotypic diversity at the CCEae3a locus revealed high polymorphism, while phylogenetic analysis showed an absence of correlation between haplotype similarity and geographic origin. Two types of esterase amplifications were found, in two locations only (Athens and Florida): one, previously described, results in the amplification of both CCEae3a and CCEae6a; the second is being described for the first time and results in the amplification of CCEae3a only. The two amplification events are independent, as confirmed by sequence analysis. All individuals from Athens and Florida carrying the CCEae3a-CCEae6a co-amplicon share a common haplotype, indicating a single amplification event, which spread between the two countries. Significance: The importance of passive transportation of disease vectors, including individuals carrying resistance mechanisms, is discussed in the light of efficient and sustainable vector control strategies. [ABSTRACT FROM AUTHOR]

Gene copy-number variations are widespread in natural populations, but investigating their phenotypic consequences requires contemporary duplications under selection. Such duplications have been found at the ace-1 locus (encoding the organophosphate and carbamate insecticides’ target) in the mosquito Anopheles gambiae (the major malaria vector); recent studies have revealed their intriguing complexity, consistent with the involvement of various numbers and types (susceptible or resistant to insecticide) of copies. We used an integrative approach, from genome to phenotype level, to investigate the influence of duplication architecture and gene-dosage on mosquito fitness. We found that both heterogeneous (i.e., one susceptible and one resistant ace-1 copy) and homogeneous (i.e., identical resistant copies) duplications segregated in field populations. The number of copies in homogeneous duplications was variable and positively correlated with acetylcholinesterase activity and resistance level. Determining the genomic structure of the duplicated region revealed that, in both types of duplication, ace-1 and 11 other genes formed tandem 203kb amplicons. We developed a diagnostic test for duplications, which showed that ace-1 was amplified in all 173 resistant mosquitoes analyzed (field-collected in several African countries), in heterogeneous or homogeneous duplications. Each type was associated with different fitness trade-offs: heterogeneous duplications conferred an intermediate phenotype (lower resistance and fitness costs), whereas homogeneous duplications tended to increase both resistance and fitness cost, in a complex manner. The type of duplication selected seemed thus to depend on the intensity and distribution of selection pressures. This versatility of trade-offs available through gene duplication highlights the importance of large mutation events in adaptation to environmental variation. This impressive adaptability could have a major impact on vector control in Africa. [ABSTRACT FROM AUTHOR]

The global expansion of Aedes albopictus together with the absence of vaccines for most of the arboviruses transmitted by this mosquito has stimulated the development of sterile-male strategies aiming at controlling disease transmission through the suppression of natural vector populations. In this context, two environmentally friendly control strategies, namely the terile nsect echnique (SIT) and the Wolbachia-based ncompatible nsect echnique (IIT) are currently being developed in several laboratories worldwide. So far however, there is a lack of comparative assessment of these strategies under the same controlled conditions. Here, we compared the mating capacities, i.e. insemination capacity, sterilization capacity and mating competitiveness of irradiated (35 Gy) and incompatible Ae. albopictus males at different ages and ratios under laboratory controlled conditions. Our data show that there was no significant difference in insemination capacity of irradiated and incompatible males, both male types showing lower capacities than untreated males at 1 day but recovering full capacity within 5 days following emergence. Regarding mating competitiveness trials, a global observed trend is that incompatible males tend to induce a lower hatching rate than irradiated males in cage controlled confrontations. More specifically, incompatible males were found more competitive than irradiated males in 5:1 ratio regardless of age, while irradiated males were only found more competitive than incompatible males in the 1:1 ratio at 10 days old. Overall, under the tested conditions, IIT seemed to be slightly more effective than SIT. However, considering that a single strategy will likely not be adapted to all environments, our data stimulates the need for comparative assessments of distinct strategies in up-scaled conditions in order to identify the most suitable and safe sterilizing technology to be implemented in a specific environmental setting and to identify the parameters requiring fine tuning in order to reach optimal release conditions. [ABSTRACT FROM AUTHOR]

Background: The control of Aedes albopictus, a major vector for viral diseases, such as dengue fever and chikungunya, has been largely reliant on the use of the larvicide temephos for many decades. This insecticide remains a primary control tool for several countries and it is a potential reliable reserve, for emergency epidemics or new invasion cases, in regions such as Europe which have banned its use. Resistance to temephos has been detected in some regions, but the mechanism responsible for the trait has not been investigated. Principal findings: Temephos resistance was identified in an Aedes albopictus population isolated from Greece, and subsequently selected in the laboratory for a few generations. Biochemical assays suggested the association of elevated carboxylesterases (CCE), but not target site resistance (altered AChE), with this phenotype. Illumina transcriptomic analysis revealed the up-regulation of three transcripts encoding CCE genes in the temephos resistant strain. CCEae3a and CCEae6a showed the most striking up-regulation (27- and 12-folds respectively, compared to the reference susceptible strain); these genes have been previously shown to be involved in temephos resistance also in Ae. aegypti. Gene amplification was associated with elevated transcription levels of both CCEae6a and CCEae3a genes. Genetic crosses confirmed the genetic link between CCEae6a and CCEae3a amplification and temephos resistance, by demonstrating a strong association between survival to temephos exposure and gene copy numbers in the F2 generation. Other transcripts, encoding cytochrome P450s, UDP-glycosyltransferases (UGTs), cuticle and lipid biosynthesis proteins, were upregulated in resistant mosquitoes, indicating that the co-evolution of multiple mechanisms might contribute to resistance. Significance: The identification of specific genes associated with insecticide resistance in Ae. albopictus for the first time is an important pre-requirement for insecticide resistance management. The genomic resources that were produced will be useful to the community, to study relevant aspects of Ae. albopictus biology. [ABSTRACT FROM AUTHOR]

In response to the widespread use of control strategies such as Insecticide Treated Nets (ITN), Anopheles mosquitoes have evolved various resistance mechanisms. Kdr is a mutation that provides physiological resistance to the pyrethroid insecticides family (PYR). In the present study, we investigated the effect of the Kdr mutation on the ability of female An. gambiae to locate and penetrate a 1cm-diameter hole in a piece of netting, either treated with insecticide or untreated, to reach a bait in a wind tunnel. Kdr homozygous, PYR-resistant mosquitoes were the least efficient at penetrating an untreated damaged net, with about 51% [39-63] success rate compared to 80% [70-90] and 78% [65-91] for homozygous susceptible and heterozygous respectively. This reduced efficiency, likely due to reduced host-seeking activity, as revealed by mosquito video-tracking, is evidence of a recessive behavioral cost of the mutation. Kdr heterozygous mosquitoes were the most efficient at penetrating nets treated with PYR insecticide, thus providing evidence for overdominance, the rarely-described case of heterozygote advantage conveyed by a single locus. The study also highlights the remarkable capacity of female mosquitoes, whether PYR-resistant or not, to locate holes in bed-nets. [ABSTRACT FROM AUTHOR]

Many insect species harbor Wolbachia bacteria that induce cytoplasmic incompatibility (CI), i.e. embryonic lethality in crosses between infected males and uninfected females, or between males and females carrying incompatible Wolbachia strains. The molecular mechanism of CI remains unknown, but the available data are best interpreted under a modification–rescue model, where a mod function disables the reproductive success of infected males’ sperm, unless the eggs are infected and express a compatible resc function. Here we examine the evolution of CI in the mosquito Culex pipiens, harbouring a large number of closely related Wolbachia strains structured in five distinct phylogenetic groups. Specifically, we used a worldwide sample of mosquito lines to assess the hypothesis that genetic divergence should correlate with the divergence of CI properties on a low evolutionary scale. We observed a significant association of Wolbachia genetic divergence with CI patterns. Most Wolbachia strains from the same group were compatible whereas those from different groups were often incompatible. Consistently, we found a strong association between Wolbachia groups and their mod-resc properties. Finally, lines from the same geographical area were rarely incompatible, confirming the conjecture that the spatial distribution of Wolbachia compatibility types should be constrained by selection. This study indicates a clear correlation between Wolbachia genotypes and CI properties, paving the way toward the identification of the molecular basis of CI through comparative genomics. [ABSTRACT FROM AUTHOR]

Many insect species harbor Wolbachia bacteria that induce cytoplasmic incompatibility (CI), i.e. embryonic lethality in crosses between infected males and uninfected females, or between males and females carrying incompatible Wolbachia strains. The molecular mechanism of CI remains unknown, but the available data are best interpreted under a modification–rescue model, where a mod function disables the reproductive success of infected males’ sperm, unless the eggs are infected and express a compatible resc function. Here we examine the evolution of CI in the mosquito Culex pipiens, harbouring a large number of closely related Wolbachia strains structured in five distinct phylogenetic groups. Specifically, we used a worldwide sample of mosquito lines to assess the hypothesis that genetic divergence should correlate with the divergence of CI properties on a low evolutionary scale. We observed a significant association of Wolbachia genetic divergence with CI patterns. Most Wolbachia strains from the same group were compatible whereas those from different groups were often incompatible. Consistently, we found a strong association between Wolbachia groups and their mod-resc properties. Finally, lines from the same geographical area were rarely incompatible, confirming the conjecture that the spatial distribution of Wolbachia compatibility types should be constrained by selection. This study indicates a clear correlation between Wolbachia genotypes and CI properties, paving the way toward the identification of the molecular basis of CI through comparative genomics. [ABSTRACT FROM AUTHOR]

Several mosquito-borne diseases affect the Western Indian Ocean islands. Culex pipiens quinquefasciatus is one of these vectors and transmits filariasis, Rift Valley and West Nile viruses and the Japanese encephalitis. To limit the impact of these diseases on public health, considerable vector control efforts have been implemented since the 50s, mainly through the use of neurotoxic insecticides belonging to Organochlorines (OC), Organophosphates (OP) and pyrethroids (PYR) families. However, mosquito control failures have been reported on site, and they were probably due to the selection of resistant individuals in response to insecticide exposure. In this study, we used different approaches to establish a first regional assessment of the levels and mechanisms of resistance to various insecticides. Bioassays were used to evaluate resistance to various insecticides, enzyme activity was measured to assess the presence of metabolic resistances through elevated detoxification, and molecular identification of known resistance alleles was investigated to determine the frequency of target-site mutations. These complementary approaches showed that resistance to the most used insecticides families (OC, OP and PYR) is widespread at a regional scale. However, the distribution of the different resistance genes is quite heterogeneous among the islands, some being found at high frequencies everywhere, others being frequent in some islands and absent in others. Moreover, two resistance alleles displayed clinal distributions in Mayotte and La Réunion, probably as a result of a heterogeneous selection due to local treatment practices. These widespread and diverse resistance mechanisms reduce the capacity of resistance management through classical strategies (e.g. insecticide rotation). In case of a disease outbreak, it could undermine the efforts of the vector control services, as only few compounds could be used. It thus becomes urgent to find alternatives to control populations of Cx. p. quinquefasciatus in the Indian Ocean. [ABSTRACT FROM AUTHOR]