skip to main content
US FlagAn official website of the United States government
dot gov icon
Official websites use .gov
A .gov website belongs to an official government organization in the United States.
https lock icon
Secure .gov websites use HTTPS
A lock ( lock ) or https:// means you've safely connected to the .gov website. Share sensitive information only on official, secure websites.


Title: The midgut epithelium of mosquitoes adjusts cell proliferation and endoreplication to respond to physiological challenges
Abstract BackgroundHematophagous mosquitoes transmit many pathogens that cause human diseases. Pathogen acquisition and transmission occur when female mosquitoes blood feed to acquire nutrients for reproduction. The midgut epithelium of mosquitoes serves as the point of entry for transmissible viruses and parasites. ResultsWe studied midgut epithelial dynamics in five major mosquito vector species by quantifying PH3-positive cells (indicative of mitotic proliferation), the incorporation of nucleotide analogs (indicative of DNA synthesis accompanying proliferation and/or endoreplication), and the ploidy (by flow cytometry) of cell populations in the posterior midgut epithelium of adult females. Our results show that the epithelial dynamics of post-emergence maturation and of mature sugar-fed guts were similar in members of theAedes,Culex, andAnophelesgenera. In the first three days post-emergence, ~ 20% of cells in the posterior midgut region of interest incorporated nucleotide analogs, concurrent with both proliferative activity and a broad shift toward higher ploidy. In mature mosquitoes maintained on sugar, an average of 3.5% of cells in the posterior midgut region of interest incorporated nucleotide analogs from five to eight days post-emergence, with a consistent presence of mitotic cells indicating constant cell turnover. Oral bacterial infection triggered a sharp increase in mitosis and nucleotide analog incorporation, suggesting that the mosquito midgut undergoes accelerated cellular turnover in response to damage. Finally, blood feeding resulted in an increase in cell proliferation, but the nature and intensity of the response varied by mosquito species and by blood source (human, bovine, avian or artificial). InAn. gambiae, enterocytes appeared to reenter the cell cycle to increase ploidy after consuming blood from all sources except avian. ConclusionsWe saw that epithelial proliferation, differentiation, and endoreplication reshape the blood-fed gut to increase ploidy, possibly to facilitate increased metabolic activity. Our results highlight the plasticity of the midgut epithelium in mosquitoes’ physiological responses to distinct challenges. Graphical Abstract  more » « less
Award ID(s):
2024252
PAR ID:
10488033
Author(s) / Creator(s):
; ; ; ; ; ; ;
Publisher / Repository:
Springer Science + Business Media
Date Published:
Journal Name:
BMC Biology
Volume:
22
Issue:
1
ISSN:
1741-7007
Format(s):
Medium: X
Sponsoring Org:
National Science Foundation
More Like this
  1. Kittayapong, Pattamaporn (Ed.)
    BackgroundSugar feeding is an important behavior which may determine vector potential of female mosquitoes. Sugar meals can reduce blood feeding frequency, enhance survival, and decrease fecundity, as well as provide energetic reserves to fuel energy intensive behaviors such as mating and host seeking. Sugar feeding behavior can be harnessed for vector control (e.g. attractive toxic sugar baits). Few studies have addressed sugar feeding ofAedes albopictus, a vector of arboviruses of public health importance, including dengue and Zika viruses. To address this knowledge gap, we assessed sugar feeding patterns ofAe.albopictusfor the first time in its invasive northeastern USA range. Methodology/Principal findingsUsing the cold anthrone fructose assay with robust sample sizes, we demonstrated that a large percentage of both male (49.6%) and female (41.8%)Ae.albopictusfed on plant or homopteran derived sugar sources within 24 hrs prior to capture. Our results suggest that sugar feeding behavior increases when environmental conditions are dry (high saturation deficit) and may vary by behavioral status (host seeking vs. resting). Furthermore, mosquitoes collected on properties with flowers (>3 blooms) had higher fructose concentrations compared to those collected from properties with few to no flowers (0–3). Conclusions/SignificanceOur results provide the first evidence ofAe.albopictussugar feeding behavior in the Northeastern US and reveal relatively high rates of sugar feeding. These results suggest the potential success for regional deployment of toxic sugar baits. In addition, we demonstrate the impact of several environmental and mosquito parameters (saturation deficit, presence of flowers, host seeking status, and sex) on sugar feeding. Placing sugar feeding behavior in the context of these environmental and mosquito parameters provides further insight into spatiotemporal dynamics of feeding behavior forAe.albopictus, and in turn, provides information for evidence-based control decisions. 
    more » « less
  2. Coffey, Lark L (Ed.)
    The mosquito midgut functions as a key interface between pathogen and vector. However, studies of midgut physiology and virus infection dynamics are scarce, and inCulex tarsalis—an extremely efficient vector of West Nile virus (WNV)—nonexistent. We performed single-cell RNA sequencing onCx. tarsalismidguts, defined multiple cell types, and determined whether specific cell types are more permissive to WNV infection. We identified 20 cell states comprising 8 distinct cell types, consistent with existing descriptions ofDrosophilaandAedes aegyptimidgut physiology. Most midgut cell populations were permissive to WNV infection. However, there were higher levels of WNV RNA (vRNA) in enteroendocrine cells (EE), suggesting enhanced replication in this population. In contrast, proliferating intestinal stem cells (ISC) had the lowest levels of vRNA, a finding consistent with studies suggesting ISC proliferation in the midgut is involved in infection control. ISCs were also found to have a strong transcriptional response to WNV infection; genes involved in ribosome structure and biogenesis, and translation were significantly downregulated in WNV-infected ISC populations. Notably, we did not detect significant WNV-infection induced upregulation of canonical mosquito antiviral immune genes (e.g.,AGO2,R2D2, etc.) at the whole-midgut level. Rather, we observed a significant positive correlation between immune gene expression levels and vRNA load in individual cells, suggesting that within midgut cells, high levels of vRNA may trigger antiviral responses. Our findings establish aCx. tarsalismidgut cell atlas, and provide insight into midgut infection dynamics of WNV by characterizing cell-type specific enhancement/restriction of, and immune response to, infection at the single-cell level. 
    more » « less
  3. Abstract IntroductionMechanical forces provide critical biological signals to cells. Within the distal lung, tensile forces act across the basement membrane and epithelial cells atop. Stretching devices have supported studies of mechanical forces in distal lung epithelium to gain mechanistic insights into pulmonary diseases. However, the integration of curvature into devices applying mechanical forces onto lung epithelial cell monolayers has remained challenging. To address this, we developed a hammock-shaped platform that offers desired curvature and mechanical forces to lung epithelial monolayers. MethodsWe developed hammocks using polyethylene terephthalate (PET)-based membranes and magnetic-particle modified silicone elastomer films within a 48-well plate that mimic the alveolar curvature and tensile forces during breathing. These hammocks were engineered and characterized for mechanical and cell-adhesive properties to facilitate cell culture. Using human small airway epithelial cells (SAECs), we measured monolayer formation and mechanosensing using F-Actin staining and immunofluorescence for cytokeratin to visualize intermediate filaments. ResultsWe demonstrate a multi-functional design that facilitates a range of curvatures along with the incorporation of magnetic elements for dynamic actuation to induce mechanical forces. Using this system, we then showed that SAECs remain viable, proliferate, and form an epithelial cell monolayer across the entire hammock. By further applying mechanical stimulation via magnetic actuation, we observed an increase in proliferation and strengthening of the cytoskeleton, suggesting an increase in mechanosensing. ConclusionThis hammock strategy provides an easily accessible and tunable cell culture platform for mimicking distal lung mechanical forces in vitro. We anticipate the promise of this culture platform for mechanistic studies, multi-modal stimulation, and drug or small molecule testing, extendable to other cell types and organ systems. 
    more » « less
  4. Abstract BackgroundCulexmosquitoes are the primary vectors of West Nile virus (WNV) across the USA. Understanding when these vectors are active indicates times when WNV transmission can occur. This study determined the proportion of femaleCulexmosquitoes that were in diapause during the fall and winter and when they terminated diapause and began blood feeding in the spring. MethodsMosquitoes were collected from parks using various traps and/or aspirated from culverts in Franklin County, Ohio, from October to mid-May from 2019 to 2022.Culexmosquitoes were morphologically identified to species, and the ovaries of females were dissected to determine their diapause and parity statuses. ResultsBy early October 2021, roughly 95% ofCulex pipienscollected in culverts were in diapause and 98% ofCx. erraticuswere in diapause. Furthermore, gravid and blood-fedCulex salinarius,Cx. pipiens, andCx. restuanswere collected in late November in 2019 and 2021 in standard mosquito traps. In the winter of 2021, the proportions of non-diapausingCulexdecreased within culverts. The last non-diapausingCx. erraticuswas collected in late December 2021 while the final non-diapausingCx. pipienswas collected in mid-January 2022, both in culverts. Roughly 50% ofCx. pipiensterminated diapause by mid-March 2022, further supported by our collections of gravid females in late March in all 3 years of mosquito collection. In fact, male mosquitoes ofCx. pipiens,Cx. restuans, andCx. territanswere collected by the 1st week of May in 2022, indicating that multiple species ofCulexproduced a second generation that reached adulthood by this time. ConclusionsWe collected blood-fed and gravidCulexfemales into late November in 2 of the 3 years of our collections, indicating that it might be possible for WNV transmission to occur in late fall in temperate climates like Ohio. The persistence of non-diapausingCx. pipiensandCx. erraticusthroughout December has important implications for the winter survival of WNV vectors and our overall understanding of diapause. Finally, determining whenCulexterminate diapause in the spring may allow us to optimize mosquito management programs and reduce the spread of WNV before it is transmitted to humans. Graphical Abstract 
    more » « less
  5. One of the first organizing processes during animal development is the assembly of embryonic cells into epithelia. Common features unite epithelialization across select bilaterians, however, we know less about the molecular and cellular mechanisms that drive epithelial emergence in early branching nonbilaterians. In sea anemones, epithelia emerge both during embryonic development and after cell aggregation of dissociated tissues. Although adhesion is required to keep cells together, it is not clear whether cell polarization plays a role as epithelia emerge from disordered aggregates. Here, we use the embryos of the sea anemoneNematostella vectensisto investigate the evolutionary origins of epithelial development. We demonstrate that lateral cell polarization is essential for epithelial organization in both embryos and aggregates. With disrupted lateral polarization, cell contact in the aggregate is not sufficient to trigger epithelialization and further tissue development. Specifically, knockdown of the conserved lateral polarity and tumor suppressor protein Lethal giant larvae (Lgl) disrupts epithelia in developing embryos and impairs the capacity of dissociated cells to epithelialize from aggregates. In contrast to other systems, cells inNematostella lglknockdown embryos do not undergo excessive proliferation. Cells with reduced Lgl levels lose their columnar shape and proper positioning of their mitotic spindles and basal bodies. Due to misoriented divisions and aberrant shapes, cells arrange nonuniformly without forming a monolayer. Together our data show that, inNematostella,Lgl drives epithelialization in embryos and cell aggregates through its effect on cell shape and organelle localization. 
    more » « less