Hemocytes limit the capacity of mosquitoes to transmit human pathogens. Here we profile the transcriptomes of 8506 hemocytes of Anopheles gambiae and Aedes aegypti mosquito vectors. Our data reveal the functional diversity of hemocytes, with different subtypes of granulocytes expressing distinct and evolutionarily conserved subsets of effector genes. A previously unidentified cell type in An. gambiae, which we term “megacyte,” is defined by a specific transmembrane protein marker (TM7318) and high expression of lipopolysaccharide-induced tumor necrosis factor–α transcription factor 3 (LL3). Knockdown experiments indicate that LL3 mediates hemocyte differentiation during immune priming. We identify and validate two main hemocyte lineages and find evidence of proliferating granulocyte populations. This atlas of medically relevant invertebrate immune cells at single-cell resolution identifies cellular events that underpin mosquito immunity to malaria infection.

e Leptospira interrogans is the primary causative agent of the most widespread zoonotic disease, leptospirosis. An in-depth structural characterization of L. interrogans is needed to understand its biology and pathogenesis. In this study, cryo-electron tomography (cryo-ET) was used to compare pathogenic and saprophytic species and examine the unique morphological features of this group of bacteria. Specifically, our study revealed a structural difference between the cell envelopes of L. interrogans and Leptospira biflexa involving variations in the lipopolysaccharide (LPS) layer. Through cryo-ET and subvolume averaging, we determined the first three-dimensional (3-D) structure of the flagellar motor of leptospira, with novel features in the flagellar C ring, export apparatus, and stator. Together with direct visualization of chemoreceptor arrays, DNA packing, periplasmic filaments, spherical cytoplasmic bodies, and a unique "cap" at the cell end, this report provides structural insights into these fascinating Leptospira species.

Green fluorescent protein (GFP) fusions, immunofluorescence microscopy, and cryo-electron tomography revealed that the chemoreceptors of the Lyme disease spirochete Borrelia burgdorferi form long, thin arrays near both cell poles. These arrays are in close proximity to the flagellar motors. This information provides a basis for further understanding motility, chemotaxis, and protein localization in spirochetes.Bacterial chemotaxis is a complex sensory transduction pathway that enables cells to sense and respond to environmental stimuli. Although chemotaxis has been well studied in the paradigm models of Escherichia coli, Salmonella enterica serovar Typhimurium, and Rhodobacter sphaeroides (17,33,35), the understanding of this mechanism in spirochetes is at an early stage (4,7,10,16,24,28,30). Spirochetes have two bundles of periplasmic flagella (PF) that are subterminally attached at each cell pole (7,8,15,18). Due to this unique geometry, the PF necessarily rotate asymmetrically during translational motility, with one bundle rotating counterclockwise (CCW) and the other rotating clockwise (CW) (7,24,30). One enigmatic question is how spirochetes coordinate the directional rotation (CCW or CW) of the two bundles of PF during chemotaxis (7,24). Although several models have been proposed, the mechanism involved still remains unknown (7,16,24,30). Flagellar rotation is modulated by chemotaxis (3, 35). Methylaccepting chemotaxis proteins (MCPs) form clusters that reside at or near the cell poles in most motile bacteria, and the spatial organization and polar positioning of chemotaxis arrays are extremely important for the process of tactic responses (2,5,6,19,32,33). However, the location of chemotaxis arrays in spirochetes is still obscure (7, 24). Here we asked whether the MCPs are located at only one cell pole or both cell poles. Although Gestwicki et al. found using fluorescent antibodies that the MCPs in Spirochaeta aurantia were localized at either one or both cell poles, no quantitative data were presented with respect to the percentage of cells that have MCPs at one or both cell ends (14). We also asked whether the MCPs are in close proximity to the subterminally located flagellar motors, as Briegel et al., using cryo-electron tomography (cryo-ET), found that the MCPs were subpolarly localized in Borrelia burgdorferi (6). To address these two questions, we first used green fluorescent protein (GFP) and immunofluorescence assays (IFA) to determine the approximate cellular locations of two different MCPs and then applied cryo-ET to reveal the precise cellular position of the chemoreceptor arrays and their spatial orientation to the flagellar motors.The strategy to localize the chemotaxis arrays of B. burgdorferi. MCPs typically interact with each other and other chemotaxis proteins to form arrays at cell poles (5,17,29,33,34). There are five putative MCPs in B. burgdorferi, including MCP1 (BB0578), MCP2 (BB0596), MCP3 (BB0597), MCP4 (BB0680), and MCP5 (BB0681) (7,12). Among these proteins, MCP3 and MCP5 are mo...