bioRxiv preprint doi: https://doi.org/10.1101/358333; this version posted June 29, 2018. The copyright holder for this preprint (which was not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. 1 Wolbachia pipientis associated to tephritid fruit fly pests: from basic research to 2 applications 3 Mariana Mateos, Humberto Martinez, Silvia B. Lanzavecchia, Claudia Conte, Karina 4 Guillén, Brenda M. Morán-Aceves, Jorge Toledo, Pablo Liedo, Elias D. Asimakis, 5 Vangelis Doudoumis, Georgios A. Kyritsis, Nikos T. Papadopoulos, Antonios A. 6 Avgoustinos, Diego F. Segura, George Tsiamis, and Kostas Bourtzis 7 8 Affiliations 9 (MM) Department of Wildlife and Fisheries Sciences, Texas A&M University, College 10 Station, Texas, USA; corresponding author [email protected] 11 (PL JT BMM KG) El Colegio de la Frontera Sur, Tapachula, Chiapas 30700, Mexico. 12 (HM) Departamento de Tecnología de Alimentos, Unidad Académica Multidisciplinaria 13 Reynosa Aztlan, Universidad Autónoma de Tamaulipas, Mexico 14 (SBL DFS CC) Laboratorio de Genética de Insectos de Importancia Económica, Instituto de 15 Genética ‘Ewald A. Favret’, CICVyA, Instituto Nacional de Tecnología Agropecuaria 16 (INTA), Hurlingham, Buenos Aires, Argentina 17 (KB AAA) Insect Pest Control Laboratory, Joint FAO/IAEA Division of Nuclear Techniques in 18 Food and Agriculture, Vienna, Austria 19 (GT EDA VD) Department of Environmental and Natural Resources Management, University 20 of Patras, 2 Seferi St, 30100 Agrinio, Greece 21 (NTP, GAK) Laboratory of Entomology and Agricultural Zoology, Department of Agriculture 22 Crop Production and Rural Environment, University of Thessaly, Phytokou St., 38446 N. 23 Ionia Magnisia, Greece 1 bioRxiv preprint doi: https://doi.org/10.1101/358333; this version posted June 29, 2018. The copyright holder for this preprint (which was not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. 24 Abstract 25 Members of the true fruit flies (family Tephritidae) are among the most serious 26 agricultural pests worldwide, whose control and management demands large and costly 27 international efforts. The need for cost-effective and environmentally-friendly integrated 28 pest management (IPM) has led to the development and implementation of autocidal 29 control strategies. Autocidal approaches include the widely used sterile insect technique 30 (SIT) and the incompatible insect technique (IIT). IIT relies on maternally transmitted 31 bacteria (namely Wolbachia), to cause a conditional sterility in crosses between released 32 mass-reared Wolbachia-infected males and wild females, which are either uninfected or 33 infected with a different Wolbachia strain (i.e., cytoplasmic incompatibility; CI). Herein, 34 we review the current state of knowledge on Wolbachia-tephritid interactions including 35 infection prevalence in wild populations, phenotypic consequences, and their impact on 36 life history traits. Numerous pest tephritid species are reported to harbor Wolbachia 37 infections, with a subset exhibiting high prevalence. The phenotypic effects of 38 Wolbachia have been assessed in very few tephritid species, due in part to the difficulty 39 of manipulating Wolbachia infection (removal or transinfection). Based on recent 40 methodological advances (high-throughput DNA sequencing) and a breakthrough 41 concerning the mechanistic basis of CI, we suggest research avenues that could 42 accelerate generation of necessary knowledge for the potential use of Wolbachia-based 43 IIT in area-wide integrated pest management (AW-IPM) strategies for the population 44 control of tephritid pests. 2 bioRxiv preprint doi: https://doi.org/10.1101/358333; this version posted June 29, 2018. The copyright holder for this preprint (which was not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. 45 1 Background 46 1.1 The economic importance of tephritids as pests 47 Flies in the family Tephritidae (Diptera) include some of the world’s most important 48 agricultural pests. The family is comprised of ~4900 described species within 481 49 genera, of which seven (Anastrepha, Bactrocera, Ceratitis, Dacus, Rhagoletis, 50 Toxotrypana and Zeugodacus) contain ~70 major pest species [1-5]. In addition to 51 causing billions of dollars in direct losses to a wide variety of fruit, vegetable and flower 52 crops, pest tephritids limit the development of agriculture in many countries due to strict 53 quarantines implemented in fruit-buying countries, and to the costs associated with 54 efforts aimed at prevention, containment, suppression, and eradication. 55 56 To prevent or minimize the harmful effects of tephritid pests, growers of affected crops 57 must comply with health and safety standards required by the market, applying an area- 58 wide management approach involving chemical, biological, cultural, and autocidal 59 control practices [6, 7]. Autocidal refers to methods that use the insect to control itself, 60 by releasing insects that are sterile or induce sterility upon mating with wild insects in the 61 next or subsequent generations [8-10]. Autocidal strategies include the sterile insect 62 technique (SIT); one of the most widespread control methods used against fruit flies. 63 SIT relies on the mass-rearing production, sterilization and recurrent release of insects 64 (preferentially males) of the targeted species. Sterilization is attained by radiation, in a 65 way that does not impair male mating and insemination capabilities. Wild females that 66 mate with sterilized males lay unfertilized eggs. At the appropriate sterile:wild (S:W) 67 ratio, the reproductive potential of the target population can be reduced [11-13]. 68 Historically, at least 28 countries have used the SIT at a large-scale for the suppression 69 or eradication of pest insects [14-16]. SIT has been applied successfully for several 70 non-tephritid insect pests, for example: the New World screw worm (Cochliomyia 3 bioRxiv preprint doi: https://doi.org/10.1101/358333; this version posted June 29, 2018. The copyright holder for this preprint (which was not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. 71 hominivorax Coquerel); several species of tsetse fly (Glossina spp.); and the codling 72 moth (Cydia pomonella L.) [reviewed in 17, 18]. 73 74 Successful SIT programs as part of Area-wide Integrated Pest Management (IPM) 75 strategies have also been implemented for several tephritids: Ceratitis capitata 76 Wiedemann; Anastrepha ludens Loew; Anastrepha obliqua Macquart; Anastrepha 77 fraterculus Wiedemann; Zeugodacus cucurbitae Coquillett; Bactrocera dorsalis Hendel; 78 and Bactrocera tryoni Froggatt [6, 12, 13, 15]. SIT is currently being developed for two 79 additional tephritid species: Dacus ciliatus Loew and Bactrocera tau Walker [19, 20]. 80 The advantages of the SIT over other pest control approaches (e.g. use of pesticides) 81 are that it is the most environmentally friendly and resistance is unlikely to evolve [21, 82 22]. 83 84 Another autocidal strategy where mating between mass-reared and wild insects can be 85 used to suppress pest populations is the incompatible insect technique (IIT). IIT also 86 relies on the principle of reducing female fertility, but utilizes endosymbiotic bacteria 87 instead of radiation, to induce a context-dependent sterility in wild females. It is based 88 on the ability of certain maternally inherited bacteria (namely from the genus Wolbachia) 89 to induce a form of reproductive incompatibility known as cytoplasmic incompatibility (CI; 90 explained in the section below). Herein we review the current knowledge on taxonomic 91 diversity of Wolbachia-tephritid associations and their phenotypic consequences, and 92 identify gaps in knowledge and approaches in the context of potential application of IIT in 93 AW-IPM programs to control tephritid pests. We also discuss scenarios where these 94 two autocidal strategies, SIT and IIT, could be potentially combined for the population 95 suppression of tephritid pests. 96 4 bioRxiv preprint doi: https://doi.org/10.1101/358333; this version posted June 29, 2018. The copyright holder for this preprint (which was not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. 97 1.2 The influence of Wolbachia on host ecology 98 Insects and other arthropods are common hosts of maternally inherited bacteria 99 [reviewed in 23]. These heritable endosymbionts can have a strong influence on host 100 ecology. Such vertically transmitted bacteria are typically vastly (or fully) dependent on 101 the host for survival and transmission. Certain associations are obligate for both 102 partners, and generally involve a nutritional benefit to the host. Other heritable bacteria 103 are facultative, with such associations ranging from mutualistic to parasitic from the 104 host’s perspective. Among these, Wolbachia is the most common and widespread 105 facultative symbiont of insects and arthropods [24-27]. 106 107 Wolbachia is a diverse and old genus [possibly older than 200 million years; 28] of 108 intracellular gram-negative Alphaproteobacteria (within the order Rickettsiales) 109 associated with arthropods and filarial nematodes. Wolbachia cells resemble small 110 spheres 0.2–1.5 μm, occur in all tissue types, but tend to be more prevalent in ovaries 111 and testicles of infected hosts,