Plant-mediated rifampicin treatment of Bemisia tabaci disrupts but does not eliminate endosymbionts

Abstract

Whiteflies are among the most important global insect pests in agriculture; their sustainable control has proven challenging and new methods are needed. Bacterial symbionts of whiteflies are poorly understood potential target of novel whitefly control methods. Whiteflies harbour an obligatory bacterium, Candidatus Portiera aleyrodidarum, and a diverse set of facultative bacterial endosymbionts. Function of facultative microbial community is poorly understood largely due to the difficulty in their selective elimination without removal of the primary endosymbiont. Since the discovery of secondary endosymbionts, antibiotic rifampicin has emerged as the most used tool for their manipulation. Its effectiveness is however much less clear, with contrasting reports on its effects on the endosymbiont community. The present study builds upon most recent method of rifampicin application in whiteflies and evaluates its ability to eliminate obligatory Portiera and two facultative endosymbionts (Rickettsia and Arsenophnus). Our results show that rifampicin reduces but does not eliminate any of the three endosymbionts. Additionally, rifampicin causes direct negative effect on whiteflies, likely by disrupting mitochondria. Taken together, results signify the end of a rifampicin era in whitefly endosymbiont studies. Finally, we propose refinement of current quantification and data analysis methods which yields additional insights in cellular metabolic scaling.

Geographic distribution of Bemisia tabaci species in Sicily and patterns in facultative endosymbiont community composition

“…The issue was not resolved by varying primer annealing temperature. As a part of the troubleshooting process, we attempted to use strain‐specific primers, designed for novel Arsenophonus strain described recently by Milenovic, Gouttepifre, et al (2022), on a subset of 30 samples of the localities 3, 4, and 5 from the Table 1, which included nine Q1_1 individuals, one Q1_4, and 20 Q2 individuals. The results of this test reveal Arsenophonus presence only in Q2 samples, with the presence confirmed in 19/20 samples.…”

“…Bemisia tabaci identity was determined by alignment to the mtCOI reference dataset (Boykin et al, 2017) followed by phylogenetic tree construction (Figure S1) as previously described in Milenovic, Gouttepifre, et al (2022).…”

“…Whiteflies were individually removed from ethanol, briefly dried and DNA was extracted according to the protocol described in Milenovic, Gouttepifre, et al (2022). Ten randomly selected, unsexed individuals were selected from each location (or less if <10 individuals were available), and DNA was extracted.…”

“…After cooling on ice, samples were diluted by adding 180 μL of PCR-grade H 2 O to each well. Separate master mixes were prepared using Q5® Hot Start High-Fidelity 2× Master Mix (New England Biolabs) and a primer pair for whitefly COX1 gene, and each of the endosymbionts as described in Milenovic, Gouttepifre, et al (2022). To avoid human error in setting up over 1000 PCR reactions, Eppendorf epMotion 5073C (Eppendorf) automated pipetting system was programmed to distribute master mix and add DNA sample to each well.…”

Life history parameters of Bemisia tabaci MED (Hemiptera: Aleyrodidae) in the present and future climate of central Europe, predicted by physically realistic climatic chamber simulation

“…The previously characterized laboratory whitefly colony of B. tabaci MED was used as a source of insect material (Milenovic et al 2022). The population harbors primary endosymbiont Portiera (group P1), and two secondary endosymbionts, Rickettsia (group R1, with scattered phenotype), and Arsenophonus (group similar to A2c, with bacteriocyte confined phenotype) (Milenovic et al 2022).…”