Mapping the malaria parasite: Several species of the parasite Plasmodium cause human malarial diseases, and, despite determined control efforts, a huge global disease burden remains. Howick et al. present a single-cell analysis of transcription across the malaria parasite life cycle (see the Perspective by Winzeler). Single-cell transcriptomes generated from 10 different life-cycle stages of the rodent-model malaria parasite P. berghei identified 20 “modules” among 5156 core transcriptome genes. These clusters enabled functional assignment of hypothetical and conserved genes, and they hint at further substructure of established life-cycle stages. The atlas also allowed for P. falciparum and P. malariae transcriptomes from patient isolates to be deconvoluted and for classification of parasitemia according to developmental stage. Science , this issue p. eaaw2619 ; see also p. 753

A possible malaria control approach involves the dissemination in mosquitoes of inherited symbiotic microbes to block Plasmodium transmission. However, in the Anopheles gambiae complex, the primary African vectors of malaria, there are limited reports of inherited symbionts that impair transmission. We show that a vertically transmitted microsporidian symbiont (Microsporidia MB) in the An. gambiae complex can impair Plasmodium transmission. Microsporidia MB is present at moderate prevalence in geographically dispersed populations of An. arabiensis in Kenya, localized to the mosquito midgut and ovaries, and is not associated with significant reductions in adult host fecundity or survival. Field-collected Microsporidia MB infected An. arabiensis tested negative for P. falciparum gametocytes and, on experimental infection with P. falciparum, sporozoites aren’t detected in Microsporidia MB infected mosquitoes. As a microbe that impairs Plasmodium transmission that is non-virulent and vertically transmitted, Microsporidia MB could be investigated as a strategy to limit malaria transmission.

Spiroplasma is a diverse bacterial clade that includes many vertically transmitted insect endosymbionts, including Spiroplasma poulsonii , a natural endosymbiont of Drosophila melanogaster . These bacteria persist in the hemolymph of their adult host and exhibit efficient vertical transmission from mother to offspring. In this study, we analyzed the mechanism that underlies their vertical transmission, and here we provide strong evidence that these bacteria use the yolk uptake machinery to colonize the germ line. We show that Spiroplasma reaches the oocyte by passing through the intercellular space surrounding the ovarian follicle cells and is then endocytosed into oocytes within yolk granules during the vitellogenic stages of oogenesis. Mutations that disrupt yolk uptake by oocytes inhibit vertical Spiroplasma transmission and lead to an accumulation of these bacteria outside the oocyte. Impairment of yolk secretion by the fat body results in Spiroplasma not reaching the oocyte and a severe reduction of vertical transmission. We propose a model in which Spiroplasma first interacts with yolk in the hemolymph to gain access to the oocyte and then uses the yolk receptor, Yolkless, to be endocytosed into the oocyte. Cooption of the yolk uptake machinery is a powerful strategy for endosymbionts to target the germ line and achieve vertical transmission. This mechanism may apply to other endosymbionts and provides a possible explanation for endosymbiont host specificity. IMPORTANCE Most insect species, including important disease vectors and crop pests, harbor vertically transmitted endosymbiotic bacteria. Studies have shown that many facultative endosymbionts, including Spiroplasma , confer protection against different classes of parasites on their hosts and therefore are attractive tools for the control of vector-borne diseases. The ability to be efficiently transmitted from females to their offspring is the key feature shaping associations between insects and their inherited endosymbionts, but to date, little is known about the mechanisms involved. In oviparous animals, yolk accumulates in developing eggs and serves to meet the nutritional demands of embryonic development. Here we show that Spiroplasma coopts the yolk transport and uptake machinery to colonize the germ line and ensure efficient vertical transmission. The uptake of yolk is a female germ line-specific feature and therefore an attractive target for cooption by endosymbionts that need to maintain high-fidelity maternal transmission.

Spiroplasma poulsonii is a maternally transmitted bacterial endosymbiont that is naturally associated with Drosophila melanogaster. S. poulsonii resides extracellularly in the hemolymph, where it must acquire metabolites to sustain proliferation. In this study, we find that Spiroplasma proliferation specifically depletes host hemolymph diacylglyceride, the major lipid class transported by the lipoprotein, Lpp. RNAi-mediated knockdown of Lpp expression, which reduces the amount of circulating lipids, inhibits Spiroplasma proliferation demonstrating that bacterial proliferation requires hemolymph-lipids. Altogether, our study shows that an insect endosymbiont acquires specific lipidic metabolites from the transport lipoproteins in the hemolymph of its host. In addition, we show that the proliferation of this endosymbiont is limited by the availability of hemolymph lipids. This feature could limit endosymbiont over-proliferation under conditions of host nutrient limitation as lipid availability is strongly influenced by the nutritional state.

Insects commonly harbor facultative bacterial endosymbionts, such as Wolbachia and Spiroplasma species, that are vertically transmitted from mothers to their offspring. These endosymbiontic bacteria increase their propagation by manipulating host reproduction or by protecting their hosts against natural enemies. While an increasing number of studies have reported endosymbiont-mediated protection, little is known about the mechanisms underlying this protection. Here, we analyze the mechanisms underlying protection from parasitoid wasps in Drosophila melanogaster mediated by its facultative endosymbiont Spiroplasma poulsonii . Our results indicate that S. poulsonii exerts protection against two distantly related wasp species, Leptopilina boulardi and Asobara tabida . S. poulsonii -mediated protection against parasitoid wasps takes place at the pupal stage and is not associated with an increased cellular immune response. In this work, we provide three important observations that support the notion that S. poulsonii bacteria and wasp larvae compete for host lipids and that this competition underlies symbiont-mediated protection. First, lipid quantification shows that both S. poulsonii and parasitoid wasps deplete D. melanogaster hemolymph lipids. Second, the depletion of hemolymphatic lipids using the Lpp RNA interference ( Lpp RNAi ) construct reduces wasp success in larvae that are not infected with S. poulsonii and blocks S. poulsonii growth. Third, we show that the growth of S. poulsonii bacteria is not affected by the presence of the wasps, indicating that when S. poulsonii is present, larval wasps will develop in a lipid-depleted environment. We propose that competition for host lipids may be relevant to endosymbiont-mediated protection in other systems and could explain the broad spectrum of protection provided. IMPORTANCE Virtually all insects, including crop pests and disease vectors, harbor facultative bacterial endosymbionts. They are vertically transmitted from mothers to their offspring, and some protect their host against pathogens. Here, we studied the mechanism of protection against parasitoid wasps mediated by the Drosophila melanogaster endosymbiont Spiroplasma poulsonii . Using genetic manipulation of the host, we provide strong evidence supporting the hypothesis that competition for host lipids underlies S. poulsonii -mediated protection against parasitoid wasps. We propose that lipid competition-based protection may not be restricted to Spiroplasma bacteria but could also apply other endosymbionts, notably Wolbachia bacteria, which can suppress human disease-causing viruses in insect hosts.

The population-level dynamics of maternally transmitted endosymbionts, including reproductive parasites, depends primarily on the fitness effects and transmission fidelity of these infections. Although experimental laboratory studies have shown that within-host endosymbiont density can affect both of these factors, the existence of such effects in natural populations has not yet been documented. Using quantitative PCR, we survey the density of male-killingWolbachiain natural populations ofDrosophila innubilafemales from the Chiricahua Mountains of Arizona. We find that there is substantial (20 000-fold) variation inWolbachiadensity among wild flies and that within-hostWolbachiadensity is positively correlated with both the efficacy of male killing and maternal transmission fidelity. MeanWolbachiadensity increases three- to five-fold from early to late in the season. This pattern suggests thatWolbachiadensity declines with fly age, a conclusion corroborated by a laboratory study ofWolbachiadensity as a function of age. Finally, we suggest three alternative hypotheses to account for the approximately lognormal distribution ofWolbachiadensity among wild flies.