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Wolbachia pipientis is an obligate intracellular bacterium within the family Anaplasmataceae that infects many terrestrial arthropods and arthropod-transmitted nematodes (filariae). Several filarial species are major human pathogens, and antibiotics with activity against Wolbachia offer a promising new therapeutic approach, since the adult worms are relatively refractory to conventional anthelmintics but depend on Wolbachia for reproduction and viability. In a natural filarial parasite of cattle, Onchocerca ochengi, intermittent chemotherapy is adulticidal whereas the equivalent dose administered as a continuous treatment is not. To investigate this further and to aid the design of efficacious regimens for human therapy, we used Wolbachia-infected Aedes albopictus mosquito cells in vitro. Here, we describe for the first time the accelerated depletion of bacteria after antibiotic withdrawal relative to the rate of elimination in the continuous presence of the drug. Mosquito cells were incubated with doxycycline while changes in 16S (bacterial) and 18S (host) rRNA and rRNA genes were determined by quantitative PCR assays. In cultures treated for 7 or 14 days followed by 7 days of drug withdrawal, the Wolbachia-to-Aedes rRNA ratio declined by approximately 6 log, whereas immediately after 14 or 21 days of continuous treatment, the reduction was only approximately 4 log (P &lt; 0.05). However, low levels of 16S rRNA remained after 21 days of treatment, irrespective of whether doxycycline was withdrawn. Application of similar methodology to related intracellular bacteria may reveal that this posttreatment effect is not restricted to Wolbachia and could have wider implications for the design of intermittent regimens for antibiotic chemotherapy.

Rate of Elimination of Wolbachia pipientis by Doxycycline In Vitro Increases following Drug Withdrawal