title: Soft X-ray microscopy analysis of cell volume and hemoglobin content in erythrocytes infected with asexual and sexual stages of Plasmodium falciparum.
abstract: Plasmodium falciparum, the most virulent agent of human malaria, undergoes both asexual cycling and sexual differentiation inside erythrocytes. As the intraerythrocytic parasite develops it increases in size and alters the permeability of the host cell plasma membrane. An intriguing question is: how is the integrity of the host erythrocyte maintained during the intraerythrocytic cycle? We have used water window cryo X-ray tomography to determine cell morphology and hemoglobin content at different stages of asexual and sexual differentiation. The cryo stabilization preserves native structure permitting accurate analyses of parasite and host cell volumes. Absorption of soft X-rays by protein adheres to Beer-Lambert's law permitting quantitation of the concentration of hemoglobin in the host cell compartment. During asexual development the volume of the parasite reaches about 50% of the uninfected erythrocyte volume but the infected erythrocyte volume remains relatively constant. The total hemoglobin content gradually decreases during the 48h cycle but its concentration remains constant until early trophozoite stage, decreases by 25%, then remains constant again until just prior to rupture. During early sexual development the gametocyte has a similar morphology to a trophozoite but then undergoes a dramatic shape change. Our cryo X-ray tomography analysis reveals that about 70% of the host cell hemoglobin is taken up and digested during gametocyte development and the parasite eventually occupies about 50% of the uninfected erythrocyte volume. The total volume of the infected erythrocyte remains constant, apart from some reversible shrinkage at stage IV, while the concentration of hemoglobin decreases to about 70% of that in an uninfected erythrocyte.
authors.affiliation: Electron Microscopy Unit, Bio21 Molecular Science and Biotechnology Institute, The University of Melbourne, Melbourne, VIC 3010, Australia. ehanssen@unimelb.edu.au
authors.forename: Eric
authors.initials: E
authors.lastname: Hanssen
authors.forename: Christian
authors.initials: C
authors.lastname: Knoechel
authors.forename: Megan
authors.initials: M
authors.lastname: Dearnley
authors.forename: Matthew W A
authors.initials: MW
authors.lastname: Dixon
authors.forename: Mark
authors.initials: M
authors.lastname: Le Gros
authors.forename: Carolyn
authors.initials: C
authors.lastname: Larabell
authors.forename: Leann
authors.initials: L
authors.lastname: Tilley
mesh_terms: D002138:Calibration; D048430:Cell Shape; D048429:Cell Size; D004912:Erythrocytes / Q000378:metabolism / Q000469:parasitology* / Q000648:ultrastructure; D005854:Germ Cells / Q000502:physiology / Q000648:ultrastructure; D006454:Hemoglobins / Q000378:metabolism*; D006801:Humans; D008853:Microscopy; D046529:Microscopy, Electron, Transmission; D010963:Plasmodium falciparum / Q000502:physiology* / Q000648:ultrastructure; D012100:Reproduction, Asexual; D012710:Serum Albumin, Bovine / Q000737:chemistry; D059010:Single-Cell Analysis; D033761:Spores, Protozoan / Q000648:ultrastructure; D014056:Tomography, X-Ray; D056928:X-Ray Absorption Spectroscopy
chemical_list: D006454:Hemoglobins; D012710:Serum Albumin, Bovine