Wolbachia in the Culex pipiens group mosquitoes: introgression and superinfection.

Wolbachia bacteria in mosquitoes induce cytoplasmic incompatibility (CI), where sperm from Wolbachia-infected males can produce inviable progeny. The wPip strain in the Culex pipiens group of mosquitoes produces a complexity of CI crossing types. Several factors are thought to be capable of influencing the expression of CI including Wolbachia strain type and host genotype. In this study, the unidirectional CI that occurs between 2 C. pipiens complex laboratory strains, Col and Mol, was further investigated by nuclear genotype introgression. The unidirectional CI between Col and Mol was not found to be influenced by host genetic background, in contrast to a previous introgression study carried out using bidirectionally incompatible C. pipiens group strains. A line containing both wPip strain variants superinfection was also generated by embryonic cytoplasmic transfer. The same crossing type as the parental Col strain was observed in the superinfected line. Quantitative polymerase chain reaction demonstrated a low density of the injected wPipMol variant in the superinfected line after 18 generations, which was considered likely to be responsible for the crossing patterns observed. The Wolbachia density was also shown to be lower in the parental Mol strain males compared with Col strain males, and no inverse relationship between WO phage and Wolbachia density could be detected

Wolbachia strain wAu efficiently blocks arbovirus transmission in Aedes albopictus

The global incidence of arboviral diseases transmitted by Aedes mosquitoes, including dengue, chikungunya, yellow fever, and Zika, has increased dramatically in recent decades. The release of Aedes aegypti carrying the maternally inherited symbiont Wolbachia as an intervention to control arboviruses is being trialled in several countries. However, these efforts are compromised in many endemic regions due to the co-localization of the secondary vector Aedes albopictus, the Asian tiger mosquito. Ae. albopictus has an expanding global distribution following incursions into a number of new territories. To date, only the wMel and wPip strains of Wolbachia have been reported to be transferred into and characterized in this vector. A Wolbachia strain naturally infecting Drosophila simulans, wAu, was selected for transfer into a Malaysian Ae. albopictus line to create a novel triple-strain infection. The newly generated line showed self-compatibility, moderate fitness cost and complete resistance to Zika and dengue infections

The Wolbachia strain wAu provides highly efficient virus transmission blocking in Aedes aegypti

Introduced transinfections of the inherited bacteria Wolbachia can inhibit transmission of viruses by Aedes mosquitoes, and in Ae. aegypti are now being deployed for dengue control in a number of countries. Only three Wolbachia strains from the large number that exist in nature have to date been introduced and characterized in this species. Here novel Ae. aegypti transinfections were generated using the wAlbA and wAu strains. In its native Ae. albopictus, wAlbA is maintained at lower density than the co-infecting wAlbB, but following transfer to Ae. aegypti the relative strain density was reversed, illustrating the strain-specific nature of Wolbachia-host co-adaptation in determining density. The wAu strain also reached high densities in Ae. aegypti, and provided highly efficient transmission blocking of dengue and Zika viruses. Both wAu and wAlbA were less susceptible than wMel to density reduction/incomplete maternal transmission resulting from elevated larval rearing temperatures. Although wAu does not induce cytoplasmic incompatibility (CI), it was stably combined with a CI-inducing strain as a superinfection, and this would facilitate its spread into wild populations. Wolbachia wAu provides a very promising new option for arbovirus control, particularly for deployment in hot tropical climates

First Steps towards Underdominant Genetic Transformation of Insect Populations

The idea of introducing genetic modifications into wild populations of insects to stop them from spreading diseases is more than 40 years old. Synthetic disease refractory genes have been successfully generated for mosquito vectors of dengue fever and human malaria. Equally important is the development of population transformation systems to drive and maintain disease refractory genes at high frequency in populations. We demonstrate an underdominant population transformation system in Drosophila melanogaster that has the property of being both spatially self-limiting and reversible to the original genetic state. Both population transformation and its reversal can be largely achieved within as few as 5 generations. The described genetic construct {Ud} is composed of two genes; (1) a UAS-RpL14.dsRNA targeting RNAi to a haploinsufficient gene RpL14 and (2) an RNAi insensitive RpL14 rescue. In this proof-of-principle system the UAS-RpL14.dsRNA knock-down gene is placed under the control of an Actin5c-GAL4 driver located on a different chromosome to the {Ud} insert. This configuration would not be effective in wild populations without incorporating the Actin5c-GAL4 driver as part of the {Ud} construct (or replacing the UAS promoter with an appropriate direct promoter). It is however anticipated that the approach that underlies this underdominant system could potentially be applied to a number of species. Figure

Genome Evolution of Wolbachia Strain wPip from the Culex pipiens Group

The obligate intracellular bacterium Wolbachia pipientis strain wPip induces cytoplasmic incompatibility (CI), patterns of crossing sterility, in the Culex pipiens group of mosquitoes. The complete sequence is presented of the 1.48-Mbp genome of wPip which encodes 1386 coding sequences (CDSs), representing the first genome sequence of a B-supergroup Wolbachia. Comparisons were made with the smaller genomes of Wolbachia strains wMel of Drosophila melanogaster, an A-supergroup Wolbachia that is also a CI inducer, and wBm, a mutualist of Brugia malayi nematodes that belongs to the D-supergroup of Wolbachia. Despite extensive gene order rearrangement, a core set of Wolbachia genes shared between the 3 genomes can be identified and contrasts with a flexible gene pool where rapid evolution has taken place. There are much more extensive prophage and ankyrin repeat encoding (ANK) gene components of the wPip genome compared with wMel and wBm, and both are likely to be of considerable importance in wPip biology. Five WO-B–like prophage regions are present and contain some genes that are identical or highly similar in multiple prophage copies, whereas other genes are unique, and it is likely that extensive recombination, duplication, and insertion have occurred between copies. A much larger number of genes encode ankyrin repeat (ANK) proteins in wPip, with 60 present compared with 23 in wMel, many of which are within or close to the prophage regions. It is likely that this pattern is partly a result of expansions in the wPip lineage, due for example to gene duplication, but their presence is in some cases more ancient. The wPip genome underlines the considerable evolutionary flexibility of Wolbachia, providing clear evidence for the rapid evolution of ANK-encoding genes and of prophage regions. This host–Wolbachia system, with its complex patterns of sterility induced between populations, now provides an excellent model for unraveling the molecular systems underlying host reproductive manipulation

Wolbachia Stimulates Immune Gene Expression and Inhibits Plasmodium Development in Anopheles gambiae

The over-replicating wMelPop strain of the endosymbiont Wolbachia pipientis has recently been shown to be capable of inducing immune upregulation and inhibition of pathogen transmission in Aedes aegypti mosquitoes. In order to examine whether comparable effects would be seen in the malaria vector Anopheles gambiae, transient somatic infections of wMelPop were created by intrathoracic inoculation. Upregulation of six selected immune genes was observed compared to controls, at least two of which (LRIM1 and TEP1) influence the development of malaria parasites. A stably infected An. gambiae cell line also showed increased expression of malaria-related immune genes. Highly significant reductions in Plasmodium infection intensity were observed in the wMelPop-infected cohort, and using gene knockdown, evidence for the role of TEP1 in this phenotype was obtained. Comparing the levels of upregulation in somatic and stably inherited wMelPop infections in Ae. aegypti revealed that levels of upregulation were lower in the somatic infections than in the stably transinfected line; inhibition of development of Brugia filarial nematodes was nevertheless observed in the somatic wMelPop infected females. Thus we consider that the effects observed in An. gambiae are also likely to be more pronounced if stably inherited wMelPop transinfections can be created, and that somatic infections of Wolbachia provide a useful model for examining effects on pathogen development or dissemination. The data are discussed with respect to the comparative effects on malaria vectorial capacity of life shortening and direct inhibition of Plasmodium development that can be produced by Wolbachia

TRIM69 inhibits vesicular stomatitis Indiana virus (VSIV)

Vesicular stomatitis Indiana virus (VSIV), formerly known as vesicular stomatitis virus (VSV) Indiana (VSVIND), is a model virus that is exceptionally sensitive to the inhibitory action of interferons (IFNs). Interferons induce an antiviral state by stimulating the expression of hundreds of interferon-stimulated genes (ISGs). These ISGs can constrain viral replication, limit tissue tropism, reduce pathogenicity, and inhibit viral transmission. Since VSIV is used as a backbone for multiple oncolytic and vaccine strategies, understanding how ISGs restrict VSIV not only helps in understanding VSIV-induced pathogenesis but also helps us evaluate and understand the safety and efficacy of VSIV-based therapies. Thus, there is a need to identify and characterize the ISGs that possess anti-VSIV activity. Using arrayed ISG expression screening, we identified TRIM69 as an ISG that potently inhibits VSIV. This inhibition was highly specific as multiple viruses, including influenza A virus, HIV-1, Rift Valley fever virus, and dengue virus, were unaffected by TRIM69. Indeed, just one amino acid substitution in VSIV can govern sensitivity/resistance to TRIM69. Furthermore, TRIM69 is highly divergent in human populations and exhibits signatures of positive selection that are consistent with this gene playing a key role in antiviral immunity. We propose that TRIM69 is an IFN-induced inhibitor of VSIV and speculate that TRIM69 could be important in limiting VSIV pathogenesis and might influence the specificity and/or efficacy of vesiculovirus-based therapies

Simultaneous genome sequencing of symbionts and their hosts

Second-generation sequencing has made possible the sequencing of genomes of interest for even small research groups. However, obtaining separate clean cultures and clonal or inbred samples of metazoan hosts and their bacterial symbionts is often difficult. We present a computational pipeline for separating metazoan and bacterial DNA in silico rather than at the bench. The method relies on the generation of deep coverage of all the genomes in a mixed sample using Illumina short-read sequencing technology, and using aggregate properties of the different genomes to identify read sets belonging to each. This inexpensive and rapid approach has been used to sequence several nematode genomes and their bacterial endosymbionts in the last year in our laboratory and can also be used to visualize and identify unexpected contaminants (or possible symbionts) in genomic DNA samples. We hope that this method will enable researchers studying symbiotic systems to move from gene-centric to genome-centric approaches

Steady states in a structured epidemic model with Wentzell boundary condition

We introduce a nonlinear structured population model with diffusion in the state space. Individuals are structured with respect to a continuous variable which represents a pathogen load. The class of uninfected individuals constitutes a special compartment that carries mass, hence the model is equipped with generalized Wentzell (or dynamic) boundary conditions. Our model is intended to describe the spread of infection of a vertically transmitted disease, for example Wolbachia in a mosquito population. Therefore the (infinite dimensional) nonlinearity arises in the recruitment term. First we establish global existence of solutions and the Principle of Linearised Stability for our model. Then, in our main result, we formulate simple conditions, which guarantee the existence of non-trivial steady states of the model. Our method utilizes an operator theoretic framework combined with a fixed point approach. Finally, in the last section we establish a sufficient condition for the local asymptotic stability of the positive steady state

Invertebrate Post-Segregation Distorters: A New Embryo-Killing Gene

Cytoplasmic incompatibility induced by inherited intracellular bacteria of arthropods, and Medea elements found in flour beetles, are both forms of postsegregation distortion involving the killing of embryos in order to increase the ratio of progeny that inherit them. The recently described peel-zeel element of Caenorhabditis elegans also uses this mechanism; like Medea the genes responsible are in the nuclear genome but it shares a paternal mode of action with the bacteria. The peel-1 gene has now been shown to encode a potent toxin that is delivered by sperm, and rescued by zygotic transcription of the linked zeel-1. The predominance of self-fertilization in C. elegans has produced an unusual distribution pattern for a selfish genetic element; further population and functional studies will shed light on its evolution. The element might also have potential for use in disease control