Figure 1.
Early development of Gregarina cuneata observed using a light microscope.
A. Earliest stages of trophozoites with developed epimerites (asterisks) under transmission light (left) and in phase contrast (right). B. Early trophozoites with polymorphous epimerites (asterisks). Transmission light. C. Detached maturing trophozoite with an epimerite surrounded by the host cell (arrowhead). Phase contrast. D. Three maturing trophozoites exhibiting obvious injury to their epimerites (asterisks) after forced separation from the host tissue by specimen processing. Transmission light. E. Maturing two-segmented individual exhibiting a well-developed protomerite and a cylindrical deutomerite. Note the rounded top of the protomerite lacking an epimerite. Phase contrast. F. Detailed view of the rounded protomerite top of a living gamont (left) and of the lance-shaped protomerite top of a chemically fixed gamont (right). Transmission light. G. Living gamonts associated in syzygy; primite (p), satellite (s). Note the rounded top of the primite protomerite with some remnants of the host tissue (arrowhead). Phase contrast. H. Chemically fixed gamonts associated in syzygy; lance-shaped top of the primite protomerite (asterisk), primite (p), satellite (s). I. Localisation of F-actin in early trophozoites (left) and maturing gamont (right); epimerite (asterisk), ruptured epimerite (arrowhead), septum (arrows) separating the protomerite from the deutomerite. Note that the septum (arrow) in the gamont is bulging into the protomerite. Direct fluorescence. J. Localisation of actin in maturing gamont. Note the patchy accumulation of actin with a very intense signal (green) in the protomerite cytoplasm. Immunofluorescence, counterstained with Evans blue. K. Localisation of myosin in trophozoites; epimerite (asterisk), ruptured epimerite (arrowhead). Immunofluorescence. L. Localisation of myosin in maturing individuals. The top of the protomerite exhibits more (left) or less (right) intense labelling, suggesting the presence of host tissue fragments. The inset shows the protomerite of more advanced stage of maturing gamont. Immunofluorescence, counterstained with Evans blue. M. Localisation of myosin in single maturing gamonts after detachment from host epithelium. The protracted (left) and retracted (right) protomerite tops exhibit strong labelling, suggesting the presence of host tissue fragments. Immunofluorescence; fluorescence and combination of fluorescence with transmission light. N. Localisation of myosin in mature gamonts associated in syzygies. Note the primite (left) with a lance-shaped top of the protomerite (asterisk) exhibiting distinct labelling in the peripheral area at its base (arrow) as well as the primite (right) with fragments of the host tissue covering its protomerite top (arrowhead). Immunofluorescence; combination of fluorescence with transmission light.
Figure 2.
Early development of Gregarina cuneata observed using a transmission electron microscope.
A. Invading sporozoite; host cell microvilli (mv), sporozoite nucleus (n). B–G. Sporozoite transforming into the trophozoite stage; conoid (arrow), developing epimeritic bud (asterisks), host cell (hc), host cell microvilli (mv), micronemes (arrowheads), microtubule (white arrow), nucleus (n), rhoptry-like organelle (r), subpellicular microtubules (double arrowheads). H. Early trophozoite stage. Note the anterior part of the gregarine, covered by a developing cortical vesicle (asterisks), causing an invagination of the host cell (hc) plasma membrane; host cell microvilli (mv), membrane fusion site (in circle), mitochondria (arrowheads), nucleus (n), pellicle (double arrow). Insets show details of the membrane fusion sites. I. Early trophozoite. Note the folded plasma membrane covering the cortical vesicle (asterisks) and forming numerous digitations; host cell (hc), host cell microvilli (mv), membrane fusion site (in circle), nucleus (n). J. Developing trophozoite; cortical vesicle (asterisks), endoplasmic reticulum (er), host cell (hc), host cell microvilli (mv), membrane fusion site (in circle), nucleus (n), pellicle with raising epicytic folds (double arrow). K. The apical end of another maturing trophozoite; amylopectin granules (am), cortical vesicle (asterisks), endoplasmic reticulum (er), host cell (hc), membrane fusion site (in circle), nucleus (n), unknown structure (st).
Figure 3.
Trophozoites of Gregarina cuneata observed using a transmission electron microscope.
A. Trophozoite with a well-developed epimerite (asterisk); deutomerite (d) with a nucleus, host cell microvilli (mv), protomerite (p). B. A more detailed view of the epimerite (asterisk) shown in Fig. 3A; host cell (hc), protomerite (p) cytoplasm packed with numerous inclusions. C–E. Decreasing epimerite (asterisks) forming numerous rhizoids and digitations (arrowheads) in more advanced stages of trophozoites as observed in different planes of sectioning; host cell (hc), host cell microvilli (mv), membrane fusion site (in circle), protomerite (p). The inset in Fig. 3E shows the membrane fusion site in detail. F–H. Host cell-epimerite interactions visualised by a freeze-etching technique. Fig. 3G shows a more detailed view of the membrane fusion site (in circle) from Fig. 3F. Note the border (arrowheads) between the epimerite and host cell (hc); cortical vesicle (asterisks), epicytic folds (ef) of the protomerite region (p), host cell microvilli (mv), host cell plasma membrane (white arrow), membrane-like structure limiting the cortical vesicle on its cytoplasmic face (white arrowheads), parasite plasma membrane (arrow). I. Trophozoite exhibiting a quite completely decreased epimerite (asterisks) with numerous mitochondria and gradual detachment (arrowhead) from host cell (hc), membrane fusion sites (in circles), protomerite (p).
Figure 4.
Gamonts of Gregarina cuneata observed using an electron microscope.
A. Individual exhibiting a lance-shaped top (arrowhead) of the protomerite (p) in contact with a host cell (hc); deutomerite (d), microvilli (mv), septum (arrow). B. Higher magnification of the protomerite top shown in Fig. 4A. Note the close contact of protomerite (p) with the host cell (hc) in some areas. C. Longitudinal section of the protomerite (p) separated from the deutomerite (d) by a septum (arrow). Note that the tapered protomerite top, which is in contact (arrowheads) with host cell microvilli (mv), lacks the epicytic folds (ef) covering the rest of the gregarine body. D. A view of the protomerite (p) top (arrowheads) in close contact with the microvillous surface (mv) of host epithelial cells (hc); amorphous material (×), deutomerite (d). E. Scanning electron micrograph showing the protomerite top covered by a wrinkled plasma membrane; protomerite epicytic folds (ef). The apical end of the protomerite is obviously damaged (arrowheads), probably due to mechanical separation of the gregarine from the host tissue during specimen processing. F. A more detailed view of the protomerite top exhibiting small remnants of host tissue still attached to its plasma membrane. G. A general view of the protomerite (p) separated from the deutomerite (d) by a distinct septum (arrow); epicytic folds (ef). Arrowheads indicate the rounded protomerite top in contact with host cell microvilli. H. Scanning electron micrograph showing the rounded protomerite top (arrowhead) with a scrap of host tissue (t) attached; epicytic folds (ef) covering the rest of protomerite. I. A more detailed view of the plasma membrane covering the protomerite top shown in Fig. 4H; scrap of host tissue (t).
Figure 5.
Host-parasite interactions in Gregarina cuneata as observed by transmission electron microscopy.
A, B. A more detailed view of the protomerite top (p) in close contact with host microvilli (mv) or epithelial cells (hc); amorphous material (×). C–E. A detail of the protomerite (p) apical region covered by a three-layered pellicle underlined by a dense layer; Golgi apparatus (g), host cells (hc), microvilli (mv). Note the ducts (arrows) passing to the exterior, numerous dense bodies (asterisks), semi-empty (Fig. 5C) and filled (Fig. 5D) dense structures (in circle) directly linked to the pore-like structures (arrowheads) interrupting the inner membrane complex. F. Higher magnification of the Golgi apparatus frequently observed in the protomerite cytoplasm. G. A view of the protomerite top (p) exhibiting a more undulating pattern (arrowheads) in the area adjacent to the host epithelium (hc) with microvilli (mv); epicytic folds (ef), numerous dense bodies (asterisks). H. A higher magnification showing the protomerite (p) apical region with unusual duct-like structures (arrows). This region is obviously covered by a typical three-layered pellicle consisting of a plasma membrane and inner membrane complex underlined by a dense layer, but it lacks epicytic folds. Note that the inner membrane complex is discontinuous in a periodic pattern (interrupted by pore-like structures); amorphous material (×), dense bodies (asterisks), host cells (hc).