Wolbachia pipientis Associated With Tephritid Fruit Fly Pests: From Basic Research to Applications

Abstract

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.

Keywords: cytoplasmic incompatibility; endosymbiont; incompatible insect technique; insect control; symbiosis.

FIGURE 1

(A,B) Qualitative illustration of uni- and bidirectional cytoplasmic incompatibility (CI) on the basis of the Wolbachia infection status of the parent generation. Empty male and female symbols signify absence of Wolbachia. Blue and yellow ovals represent distinct Wolbachia strains. Green tick marks = Successful offspring production. Red crosses = no offspring production. (C) A special case of unidirectional incompatibility in which one Wolbachia strain (see text) can rescue another strain (i.e., the yellow one), but not vice versa.

FIGURE 2

Recommended steps to screen for Wolbachia infections in tephritids and other arthropods. A PCR is performed with Wolbachia-specific primers on DNA isolated from whole, or parts of (e.g., abdomens), insect. Agarose gel electrophoresis of the PCR products is used to determine whether the amplicon is of the expected size. Amplicons of expected size are directly sequenced (e.g., Sanger method). High sequence identity to other Wolbachia suggests Wolbachia infection. Clean chromatograms are consistent with a single Wolbachia strain. Otherwise, a cloning step to identify different Wolbachia alleles is required. Other genes are then amplified and sequenced for further genetic characterization of the strain. As an optional step, localization of Wolbachia cells within host tissues can be achieved by Fluorescent In Situ Hybridization (FISH) with Wolbachia-specific rRNA probe or immunolabeling with antibody specific to Wolbachia protein. An amplicon of an unexpected size might indicate the occurrence of a horizontally transmitted Wolbachia genome fragment to the insect chromosome, rather than a current infection. Similarly, multiple nucleotide polymorphisms (NP) or insertions/deletions, compared to known strains, are suggestive of Wolbachia pseudogenes (e.g., horizontally transferred to host genome). This can be further tested by in situ hybridization of Wolbachia-specific probe to host chromosomes, and/or by Whole Genome Sequencing of host. Photo of fly (Anastrepha obliqua) by Fabiola Roque (ECOSUR-UT). Image from Fast et al. (2011) freely available at https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4030408/bin/NIHMS391830-supplement-Supporting_Online_Material.pdf (accessed April 01, 2019). Original sources of other photographs are available in open access journals (Doudoumis et al., 2012; Brelsfoard et al., 2014; Joubert et al., 2016).

FIGURE 3

Backcrossing procedure. Wolbachia infection is indicated by blue oval. Host nuclear backgrounds are indicated by colors: white represents the initial nuclear background of Wolbachia-infected host; red (darkest) indicates the host background of the Wolbachia-free line contributing males every generation. Different shades of red represent the increasing replacement of “white” nuclear background over backcrossing generations (F₁ to F₈) by “red” nuclear background.

FIGURE 4

Use of bi-directional CI in IIT-based population suppression programs. (A) Patterns of compatibility with two bi-directionally incompatible strains, when doubly infected hosts occur. Empty male and female symbols signify absence of Wolbachia. Blue and yellow ovals represent distinct (mutually incompatible) Wolbachia strains. Green tick marks = Successful offspring production. Red crosses = no offspring production. (B) Options for implementing IIT-based population suppression when the target wild population harbors a CI inducing strain (yellow = “native”), according to the patterns of compatibility depicted in panel a. Bi-directional CI is achieved when the released males only harbor a strain (blue = “foreign”) that is incompatible with the native strain. Additional options exist, including double infections of both target and released insects with different Wolbachia strains (not shown), such as in Aedes albopictus (Moretti et al., 2018b).