Biocomplexity and Selective modulation of whitefly symbiotic composition

Whiteflies are sap-sucking insects that harbor obligatory symbiotic bacteria to fulfill their dietary needs, as well as a facultative microbial community with diverse bacterial species. The sweetpotato whitefly Bemisia tabaci (Gennadius) is a severe agricultural pest in many parts of the world. This speciesconsists of several biotypes that have been distinguished largely on the basis of biochemical or molecular diagnostics, but whose biological significance is still unclear. The original objectives of the project were (i) to identify the specific complement of prokaryotic endosymbionts associated with select, well-studied, biologically and phylogeographically representative biotypes of B. tabaci, and (ii) to attempt to 'cure’ select biotypes of certain symbionts to permit assessment of the affect of curing on whitefly fitness, gene flow, host plant preference, and virus transmission competency.To identify the diversity of bacterial community associated with a suite of phylogeographically-diverseB. tabaci, a total of 107 populations were screened using general Bacteria primers for the 16S rRNA encoding gene in a PCR. Sequence comparisons with the available databases revealed the presence of bacteria classified in the: Proteobacteria (66%), Firmicutes (25.70%), Actinobacteria (3.7%), Chlamydiae (2.75%) and Bacteroidetes (<1%). Among previously identified bacteria, such as the primary symbiont Portiera aleyrodidarum, and the secondary symbionts Hamiltonella, Cardinium and Wolbachia, a Rickettsia sp. was detected for the first time in this insect family. The distribution, transmission, and localization of the Rickettsia were studied using PCR and fluorescence in situ hybridization (FISH). Rickettsia was found in all 20 Israeli B. tabaci populations screened as well as some populations screened in the Arizona laboratory, but not in all individuals within each population. FISH analysis of B. tabaci eggs, nymphs and adults, revealed a unique concentration of Rickettsia around the gut and follicle cells as well as its random distribution in the haemolymph, but absence from the primary symbiont housing cells, the bacteriocytes. Rickettsia vertical transmission on the one hand and its partial within-population infection on the other suggest a phenotype that is advantageous under certain conditions but may be deleterious enough to prevent fixation under others.To test for the possible involvement of Wolbachia and Cardiniumin the reproductive isolation of different B. tabacibiotypes, reciprocal crosses were preformed among populations of the Cardinium-infected, Wolbachia-infected and uninfected populations. The crosses results demonstrated that phylogeographically divergent B. tabaci are reproductively competent and that cytoplasmic incompatibility inducer-bacteria (Wolbachia and Cardinium) both interfered with, and/or rescued CI induced by one another, effectively facilitating bidirectional female offspring production in the latter scenario.This knowledge has implications to multitrophic interactions, gene flow, speciation, fitness, natural enemy interactions, and possibly, host preference and virus transmission. Although extensive and creative attempts undertaken in both laboratories to cure whiteflies of non-primary symbionts have failed, our finding of naturally uninfected individuals have permitted the establishment of Rickettsia-, Wolbachia- and Cardinium-freeB. tabaci lines, which are been employed to address various biological questions, including determining the role of these bacteria in whitefly host biology.