Light and electron microscopy on the sporulation of the oocysts of Eimeria brunetti. II. Development into the sporocyst and formation of the sporozoite

The later stages of sporulation in oocysts of Eimeria brunetti were examined in samples which had been allowed to sporulate at 27 degrees C for 24, 36 and 48 hours. It was observed that the sporoblasts became ellipsoidal and the nucleus underwent the final division. A nucleus with associated Golgi bodies was not observed at either end of the organism. The cytoplasm was limited by two unit membranes and contained rough endoplasmic reticulum, dense bodies, electron translucent vacuoles and mitochondria. The first evidence of sporozoite formation was the appearance of a dense plaque at either end of the organism. This appeared in the vicinity of the nuclei, and adjacent to the limiting membrane of the soroblast. At this stage the sporocyst wall was still unformed. Then the two sporozoites were formed from opposite ends of the organism by growth of the dense plaques and invaginations of the plasmalemma which thus formed the pellicles of the developing sporozoites. A conoid and subpellicular microtubules were observed at this stage as development continued, a number of vacuoles were found between the nucleus and the conoid. These vacuoles constituted the precursors of the rhoptries and micronemes. At the same stage a large dense body had appeared within the forming sporozoite. As the sporozoite developed, this body, anterior refractile body, is followed by the nucleus and another dense body which formed the posterior refractile body. During this period, the thin sporocyst wall was formed and Stieda and sub-Stieda bodies were now present at one end of the sporocyst. Each mature sporocyst contained two sporozoites.     

Electron microscopy of stages of Isospora felis of the cat in the mesenteric lymph node of the mouse.

Stages of Isospora felis of the cat in the mesenteric lymph node of the mouse 25 days after oral inoculation with oocysts, have been described at the ultrastructural level. The organisms occurred singly within parasitophorous vacuoles in host cell cytoplasm and were sporozoite-like, having a large crystalloid body up to 5.5 mum in length posterior to the nucleus. The size and appearance of the parasitophorous vacuole varied. Some vacuoles contained numerous, small, electron dense granules about 30 nm in diameter. Because of the aggregation of granules and their arrangement within the parasitophorous vacuole, the impression was sometimes gained by light microscopy that parasites were surrounded by a sheath or cyst wall. However, a cyst wall was not present. In host cells, spherical, membrane-bound bodies with a homogeneous, electron dense core and a maximum diameter of 0.25 mum were filed along the limiting membrane of the parasitophorous vacuole. These extra-intestinal parasites were considered to be waiting stages, with a biological function similar to that of the tissue cyst stage of other general of isosporan coccidia.     

Gametogenesis and Sporogony of Hepatozoon Mocassini (Apicomplexa: Adeleina: Hepatozoidae) In an Experimental Mosquito Host, Aedes Aegypti

ABSTRACT. The sexual life cycle of the hemogregarine Hepatozoon mocassini was studied in Aedes aegypti , an experimental mosquito host, using transmission electron microscopy. Gamonts were observed leaving the host snake erythrocyte as early as 30 min after mosquitoes ingested infected blood, and some gamonts had penetrated the gut epithelial cells by this time. Six hours post-feeding, gamonts were identified within cells of the abdominal fat body. Twenty-four hours post-feeding, gamonts were often entrapped within the peritrophic membrane, but were no longer observed within the gut wall. Parasites pairing up in syzygy and undergoing sexual differentiatioe were observed within fat cells at this time, and by 48 hours post-feeding, well-developed macro- and microgametocytes as well as microgametes were discernible. Developing zygotes observed 3 days post-feeding were enclosed within a panoitophorous vacuole. By day 6, multinucleate oocysts with crystalloid bodies in the cytoplasm were seen. Sporazoites developing within sporocysts appeared by day 12. Seventeen days post-feeding, mature oocysts with sporocysts containing approximately 16 sporozoites were observed upon dissection of mosquitoes. Large crystalloid bodies no longer bound by rough endopbsmic reticulum were located anterior and posterior to the sporozoite nucleus. Free sporozoites were not observed.     

The fine structure of the developing oocyst of Pterospora floridiensis (Apicomplexa,Urosporidae)

Developing oocysts of the gregarine Pterospora floridiensis Landers 2001 were examined by transmission electron microscopy. Each oocyst had an outer capsule and an inner capsule that contained 8 sporozoites. In early stages of development the inner capsular wall was separated from the developing sporozoites and residual mass, and was not appressed to the sporozoites. Early stage sporozoites were connected to a residual mass and were filled with endoplasmic reticulum, golgi and numerous developing secretory vesicles. In late stages of oocyst and sporozoite development, the inner capsular wall was closely appressed to the sporozoite surface. The inner capsular wall was ～60-100 nm thick and the outer capsular wall was ～160-320 nm thick. There were no extensions on the outer wall for which the genus was named. Late stage sporozoites had no residual mass connection, were more electron dense, and contained three distinct types of dense secretory structures: 1) small oval/spherical dense vesicles, 2) large (350-400 nm) vesicles near the anterior end, and 3) elongated dense tubular bodies that converged at the apex. Few ultrastructural reports exist of developing gregarine oocysts and sporozoites, and as more studies are completed these morphological characteristics may be important in interpreting molecular phylogenetic analyses.     

THE TRANSFORMATION OF THE PLASMODIUM GALLINACEUM OOCYST IN AEDES AEGYPTI MOSQUITOES

Sporoblast and sporozoite formation from oocysts of the avian malarial parasite, Plasmodium gallinaceum, after the seventh day of infection in Aedes aegypti mosquitoes offers an interesting example of differentiation involving the appearance and modification of several cellular components. Sporoblast formation is preceded by (a) invaginations of the oocyst capsule into the oocyst cytoplasm, (b) subcapsular vacuolization and cleft formation, (c) the appearance of small tufts of capsule material on the previously noted invaginations, and (d) linear dense areas located just below the oocyst plasma membrane which predetermine the site of emerging sporozoites from the sporoblast. The subcapsular clefts subdivide the once-solid oocyst into sporoblast peninsulae. Within the sporoblast, nuclei migrate from the random distribution seen in the solid oocyst and come to lie at the periphery of the sporoblast just below the linear dense areas noted in the earlier stage. A typical nuclear fiber apparatus occurs in most of the nuclei seen in random sections at this stage although such a fiber apparatus may occasionally be seen in the solid oocyst stage. The nucleus, its associated fiber apparatus, and the overlying dense area appear to induce the onset of sporozoite budding from the sporoblast as well as the formation of the sporozoite pellicular complex and the paired organelle precursor. Several mitochondria are present in each sporozoite, in contrast to the single mitochondrion seen in the merozoites of the erythrocytic and exoerythrocytic stages of avian malaria infection. The paired organelles and associated dense inclusion bodies are formed by condensation of an irregular meshwork of membrane-bound, coarse, dense material. The nature of small, particulate cytoplasmic inclusions is described.     

Ultrastructural studies on oocysts, sporulation and sporozoites of Schellackia cf. agamae from the intestine of the starred lizard Agama stellio.

Young intracellular oocysts of Schellackia cf. agamae in the gut epithelium of agama stellio were bound by several fine membranes. Later-stage oocysts and sporoblasts in the lamina propria were intercellular and were bound by a thin but firm tri-layered wall. Oocysts had a large central refractile body which, during sporulation, sent extensions into the developing sporozoites. Sporozoites escaped into the gut tissue, leaving a large oocyst residuum with the remains of a refractile body. These sporozoites invaded a variety of connective tissue cells, endothelial cells and circulatory leucocytes in the lamina propria. Sporozoites caused lysis of the host cell cytoplasm at their perimeter and multiple sporozoite infections led to complete degradation of the host cell.     

The Fine Structure of the Sporozoite of Lankesteria culicis

SYNOPSIS. The sporozoite of Lankesteria culicis was studied by light and electron microscopy, after excystation in the intestine of Aedes aegypti 1st stage larvae. The sporozoite was 9.5–10.0 μ long with a blunt anterior end and a tapered posterior region. The organism was enclosed by a typical pellicle consisting of an outer and an inner membrane with underlying subpellicular microtubules. The anterior end had a conoid with 2 associated rings, a polar ring which served as a termination of the subpellicular microtubules and a flask-shaped structure situated internal and posterior to the conoid. A micropyle consisting of a collar formed from the inner membrane and lacking an invagination of the outer membrane was present near the anterior end of the parasite. The nucleus was centrally located and had a peripheral concentration of chromatin and a central nucleolus. One or more mitochondria were observed in the vicinity of the nucleus.     

Ultrastructure of sporozoites and zoites of Hammondia heydorni

The ultrastructure of sporozoites and zoites of Hammondia heydorni was studied in cultured bovine cells. In addition to ultrastructural features typical of coccidian parasites, H. heydorni sporozoites and zoites contain rhoptries that are located posteriorly as well as anteriorly. Also, sporozoites contain a posteriorly located crystalloid body (1.2 micron in diameter); a small crystalloid body (0.5 micron in diameter) was occasionally seen in the anterior end. Zoites resulting from the 1st division of endodyogeny contain a posteriorly located crystalloid body, which is absent in zoites formed by subsequent divisions. Zoites contain posteriorly located amylopectin granules and a relatively large anterior vacuole which is not present in sporozoites. During penetration, the host cell plasmalemma ballooned laterally around the sporozoite creating a large cavity, which later disappeared. Sporozoites and zoites undergoing cell penetration usually exhibit partially empty anterior rhoptries; no changes occur in posterior rhoptries. Lysosomes fuse with the parasitophorous vacuole surrounding killed sporozoites but not live sporozoites.     

Ultrastructure of Sporozoites and Zoites of Hammondia heydorni

The ultrastructure of sporozoites and zoites of Hammondia heydorni was studied in cultured bovine cells. In addition to ultrastructural features typical of coccidian parasites, H. heydorni sporozoites and zoites contain rhoptries that are located posteriorly as well as anteriorly. Also, sporozoites contain a posteriorly located crystalloid body (1.2 μm in diameter); a small crystalloid body (0.5 μm in diameter) was occasionally seen in the anterior end. Zoites resulting from the 1st division of endodyogeny contain a posteriorly located crystalloid body, which is absent in zoites formed by subsequent divisions. Zoites contain posteriorly located amylopectin granules and a relatively large anterior vacuole which is not present in sporozoites. During penetration, the host cell plasmalemma ballooned laterally around the sporozoite creating a large cavity, which later disappeared. Sporozoites and zoites undergoing cell penetration usually exhibit partially empty anterior rhoptries; no changes occur in posterior rhoptries. Lysosomes fuse with the par-asitophorous vacuole surrounding killed sporozoites but not live sporozoites.     

Ultrastructural observations on the developmental stages of Lankesterella minima (Apicomplexa) in experimentally infected Rana catesbeiana tadpoles

During May to August 1988, the prevalence of Lankesterella minima in bullfrog tadpoles and adults in the vicinity of Lake Sasajewun, Algonquin Park, Ontario, was 54.8% and 29.4%, respectively. The parasite was transmitted to laboratory-reared tadpoles through the bite of experimentally infected, laboratory-reared leeches (Batracobdella picta). Intraerythrocytic sporozoites were observed in the experimentally infected tadpoles 36 days postexposure (PE) to leech bites. Sporogonic stages in these tadpoles were examined by light and electron microscopy. Oocysts developed in vascular endothelial cells of several organs. Multinucleate oocysts at 29 days PE lay in a parasitophorous vacuole and contained several large inclusions. Mature oocysts, observed 32 days PE, contained about 70 sporozoites that exhibited typical apicomplexan features. In the posterior region of the sporozoites, 12 helically arranged chains of electron-lucent intrapellicular extended anteriorly from a dense terminal ring.     

Nematopsis gigas n. sp. (Apicomplexa), a parasite of Nerita ascencionis (Gastropoda, Neritidae) from Brazil

A new species of Nematopsis (Apicomplexa, Porosporidae) is described from the mantle tissues of the seawater gastropod, Nerita ascencionis (Neritidae), collected in the Atlantic North off the coast of "Fernando de Noronha" Island (3 degrees 47' 57' S, 32 degrees 25' 12' W) situated about 350 km from the northeast coast of Brazil. Numerous oocysts, each contained in a parasitophorous vacuole, were found in the cytoplasm of phagocytes in the mantle tissue of the host. The phagocytes were surrounded by a thin wall composed of lucent material. The phagocyte cytoplasm contained a nucleus surrounded by numerous vesicles and some dense masses. The oocysts were 21.9 +/- 0.5 microm long, and 11.5 +/- 0.6 microm wide. The oocyst wall was 0.18-0.25 microm thick, and the apical zone contained a micropyle, 1.0-1.2 microm in diameter, covered by a canopy-like operculum about 0.25 microm thick. Externally, the oocyst wall was surrounded by numerous anastomosing microfibrils attached to the wall and extending towards the periphery of the parasitophorous vacuole. Some microfibrils formed a dense complex network that surrounded the oocyst in the middle of the parasitophorous vacuole, which opened only at the apical zone near the external region of the opercular system. On the basis of the data obtained by light and transmission electron microscopy and host specificity, the gregarine Nematopsis gigas is distinguished from the nearest species as a new species. The taxonomic affinities and morphological comparisons with other similar species of the same genus are discussed.     

A simple method of DNA extraction for Eimeria species

A new, simple method is described for extracting DNA from coccidia (Eimeriidae) oocysts. In our hands this method works well for all Eimeria oocysts and, presumably, will work equally well for oocysts of other coccidia genera. This method combines the two steps of breaking oocyst and sporocyst walls, and dissolving the sporozoite membrane in one step. This greatly simplifies the currently used DNA extraction procedures for Eimeria species and overcomes the disadvantages of existing DNA extraction methods based on glass-bead grinding and sporozoite excystation procedures. Because all the procedures are done in a 1.5-ml microfuge tube, which minimizes the loss of DNA in the extraction procedures, this method is especially suitable for samples with small number of oocysts. In addition, this method directly lyses the oocyst and sporocyst walls as well as the sporozoite membrane in a continuous incubation; therefore, it does not require the sporozoites to be alive. The results of PCR experiments indicate that this method generates better quality of DNA than what the existing glass-bead grinding method does for molecular analysis, and is suitable for both large or small number (<10(2) oocysts) of living or dead oocyst samples.     

Ultrastructural Features of Cystic and Merogonic Stages of Hepatozoon sipedon (Apicomplexa: Adeleorina) in Northern Leopard Frogs (Rana pipiens) and Northern Water Snakes (Nerodia sipedon) from Ontario, Canada

The cystic and merogonic stages of the haemogregarine Hepatozoon sipedon , infecting Northern water snakes ( Nerodia sipedon sipedon ) and Northern leopard frogs ( Rana pipiens ), respectively, in Ontario, Canada, were investigated by transmission electron microscopy. Cysts, which were observed in the liver of Northern leopard frogs ( Rana pipiens ) after these anurans ingested mosquitoes ( Culex pipiens ) containing oocysts of the parasite, harboured two cystozoites, each of which contained a large crystalloid inclusion anterior to the nucleus. Two types of meronts were observed in snakes that were fed the liver of infected frogs. Macromeronts, which matured in endothelial cells of the liver approximately 16 d after snakes ingested infected frogs, contained about 50 large macromerozoites. Macromerozoites emerged from macromeronts, entered the bloodstream of the snake, and reinfected endothelial cells. Micromeronts, which matured about 34 d post-inoculation, contained about 150 micromerozoites that infected erythrocytes and transformed into gamonts. The ultrastructural features of micromeronts and macromeronts differed only slightly: immature macromeronts and macromerozoites contained numerous amylopectin and lipid inclusions, whereas immature micromeronts and micromerozoites did not contain amylopectin inclusions and featured fewer, smaller lipid inclusions. A comparison of cystic stages among Hepatozoon species in different groups of vertebrates is presented with respect to their structure and evolutionary significance.     

Description of the oocysts of Eimeria paludosa (Apicomplexa: Eimeriidae) from Fulica americana (Aves: Gruiformes), with comments on synonyms of eimerian species from related birds

Between November and December 1988, fecal and intestinal contents were collected from 25 northern American coots, Fulica americana americana, in Arkansas and Texas, and examined for coccidial parasites. Seventeen (68%) of the coots were infected with Eimeria paludosa, herein described; for the first time, photomicrographs of the species are presented. Sporulated oocysts are ovoid, 16.5 x 12.6 (15-23 x 11-14) microns, with a lightly to heavily pitted single-layered wall; an oocyst residuum is absent, but a prominent micropyle is present. A large, or several smaller, polar granule(s) is present, usually located beneath the micropyle. Sporocysts are elongate-ovoid, 10.8 x 6.2 (10-12 x 5-7) microns, with Stieda and substieda bodies. A sporocyst residuum is present, normally composed of very fine faint granules scattered among the sporozoites or, rarely, as a spherical mass. Sporozoites are elongate, 8.7 x 2.7 (7-11 x 2-3) microns, in situ. Each sporozoite contains a spherical-ellipsoid posterior refractile body and occasionally a spherical anterior refractile body. A nucleus is located immediately anterior to the posterior refractile body. The occurrence of E. paludosa in F. a. americana is a new host and geographic record for the parasite. In addition, several of the previously described eimerian species from gruiform birds are proposed to be synonyms of E. paludosa.     

Fine Structure of Differentiating Oocysts and Mature Sporozoites of Haemoproteus metchnikovi in its Intermediate Host Chrysops callidus*

The fine structure of the sporogonic stages of Haemoproteus metchnikovi has been investigated by electron microscopy. Young oocysts are found beneath the basement membrane of midgut epithelial cells. These eventually protrude outward into the haemocoel space and are surrounded by a distinct oocyst capsule. Sporozoite formation begins with a subcapsular vacuolation. Evagination of the oocyst cytoplasm occurs in regions of membrane thickenings and 100–200 sporozoites are formed about a single sporoblastoid body. Remnants of the ookinete pellicle can be observed in maturing oocysts and always are found in the residual body. The fine structure of the mature sporozoite is essentially similar to that which has been described for other haemosporidia and a spherical body is described in association with the mitochondrion of the sporozoite. The sporogonic stages of H. metchnikovi have features common to the sporogonic stages of Plasmodium and Leucocytozoon that are not held in common by the latter 2 genera, including pattern of sporozoite formation and number of sporozoites formed, the presence of a cytostome and of “crystalloid” in the sporozoite.     

Sporogonic development of Leucocytozoon smithi.

The sporogonic development of Leucocytozoon smithi in its black fly vector was studied by light and electron microscopy and was compared with that of other haemosporidians. Within 18 to 24 h after ingestion of gametocytes by black flies, ookinetes passing through the midgut epithelium were observed. Intracellular migration of ookinetes resulted in the apparent disruption and degeneration of host cells. Intercellular migration also occurred as was evidenced by the presence of ookinetes between midgut cells. Transformation of ookinete to spherical oocyst occurred extracellularly in three different sites. Although most oocysts were found between the host cell basal membrane and the basal lamina, large numbers also were found attached to the external surface of the basal lamina, projecting into the hemocoel. Ectopic development of oocysts in the midgut epithelium between cells was observed much less frequently than development on the basal side of the midgut. The oocyst wall of dense granules, believed to be of parasite origin, was distinguishable from the basal lamina of the host's midgut epithelium. As in other Leucocytozoidae, the cytoplasm of the oocyst differentiated into a single sporoblastoid from which 30-50 sporozoites were formed. Beginning on the third day post infection, elongation of segregated dense sporoblastoid material associated with pellicle thickening led to the formation of the finger-like sporozoite buds which projected into the oocyst cavity. Sporozoites within mature oocysts and salivary glands were structurally similar to sporozoites as described for other haemosporidians.     

Sporogonic Development of Leucocytozoon smithi

The sporogonic development of Leucocytozoon smithi in its black fly vector was studied by light and electron microscopy and was compared with that of other haemosporidians. Within 18 to 24 h after ingestion of gametocytes by black flies, ookinetes passing through the midgut epithelium were observed. Intracellular migration of ookinetes resulted in the apparent disruption and degeneration of host cells. Intercellular migration also occurred as was evidenced by the presence of ookinetes between midgut cells. Transformation of ookinete to spherical oocyst occurred extracellularly in three different sites. Although most oocysts were found between the host cell basal membrane and the basal lamina, large numbers also were found attached to the external surface of the basal lamina, projecting into the hemocoel. Ectopic development of oocysts in the midgut epithelium between cells was observed much less frequently than development on the basal side of the midgut. The oocyst wall of dense granules, believed to be of parasite origin, was distinguishable from the basal lamina of the host's midgut epithelium. As in other Leucocytozoidae, the cytoplasm of the oocyst differentiated into a single sporoblastoid from which 30–50 sporozoites were formed. Beginning on the third day post infection, elongation of segregated dense sporoblastoid material associated with pellicle thickening led to the formation of the finger-like sporozoite buds which projected into the oocyst cavity. Sporozoites within mature oocysts and salivary glands were structurally similar to sporozoites as described for other haemosporidians.     

Storage Polysaccharide in Coccidial Sporozites after Excystation and Penetration of Cells

SYNOPSIS. Eimeria acervulina, E. necatrix , and E. meleagrimitis sporozoites were examined for carbohydrates by cytochemical methods during dormancy, after excystation, and after penetration of cells. The only carbohydrate found was amylopectin, a homogeneous polymer of glucose. It was distributed in 3 regions: (a) in front of the anterior refractile globule, (b) around the nucleus, and (c) behind the posterior refractile globule. The relative amounts decreased after excystation and penetration of cells until only small amounts remained around the nucleus. The quantity of amylopectin decreased following excystation from 30.0-36.7 to 9.4-13.3 μg glucose/10⁶ oocysts. Over a 6 yr period of storage at 4 C, there was a decrease in the quantity of amylopectin in dormant sporozoites of E. acervulina from 33.3 μg glucose/10⁶ oocysts at 3 mos to 1.5 μg at 6 years. Coincidentally, 3 month- and 1 year-old oocysts of E. acervulina produced patent infections in chicks with a dosage of 5 × 10⁴ oocysts, but only a few of the oocysts that had been stored for 2 years were infective; a dosage of 2 × 10⁶ oocysts was necessary to produce a patent infection. Oocysts which had been stored 6 years did not produce a patent infection.
It was concluded that amylopectin is the energy source for excystation and subsequent penetration of cells. Small amounts of amylopectin are used during dormancy and, when the content in the sporozoite falls below a certain level, the sporozoites lack sufficient energy to infect cells.     

Ultrastructure of shizonts and merozoites of Sarcocystis falcatula in the lungs of budgerigars (Melopsittacus undulatus).

Transmission electron microscopy was used to study the ultrastructure of schizogony of Sarcocystis falcatula in the lungs of budgerigars (Melopsittacus undulatus). Schizogony occurred exclusively by endopolygeny within endothelial cells of pulmonary capillaries, venules, and small veins. Early schizonts were elongate with a large nucleus and nucleolus, surrounded by a pellicle consisting of a plasmalemma and an inner single membrane, and contained most of the organelles and inclusion bodies found in merozoites of Sarcocystis species. As development proceeded, schizonts increased in size and conformed to the shapes of the pulmonary blood vessels. As micronemes, dense granules, the conoid, and subpellicular microtubules disappeared, there was an increase in the size and number of mitochondria, Golgi complexes, and Golgi adjuncts (apicoplasts). As the nucleus elongated, there was a progressive increase in the number of spindles located at various intervals along the nuclear envelope. Eventually, 2 merozoites formed internally immediately above each spindle. During endopolygeny, a portion of the nucleus was incorporated into each merozoite bud along with 1 or 2 Golgi adjuncts, a Golgi complex, mitochondria, endoplasmic reticulum, and ribosomes. During merozoite formation, micronemes appeared in close association with the Golgi complex and gradually increased in number. The pellicle invaginated around the merozoites so they budded at the schizont surface leaving behind a small, central residual body. Dense granules appeared after merozoites were completely formed. Schizonts were 24 x 6.8 microm and contained 24-96 merozoites. Merozoites were 5.1 x 1.8 microm and were found free in the pulmonary air passages and pulmonary capillaries and within nearly all cells of the lung except red blood cells.     

Plasmodium vivax:A Monoclonal Antibody Recognizes a Circumsporozoite Protein Precursor on the Sporozoite Surface

Gonzalez-Ceron, L., Rodriguez, M. H., Wirtz, R. A., Sina, B. J., Palomeque, O. L., Nettel, J. A., and Tsutsumi, V. 1998.Plasmodium vivax:A monoclonal antibody recognizes a circumsporozoite protein precursor on the sporozoite surface.Experimental Parasitology90, 203–211. The major surface circumsporozoite (CS) proteins are known to play a role in malaria sporozoite development and invasion of invertebrate and vertebrate host cells.Plasmodium vivaxCS protein processing during mosquito midgut oocyst and salivary gland sporozoite development was studied using monoclonal antibodies which recognize different CS protein epitopes. Monoclonal antibodies which react with the CS amino acid repeat sequences by ELISA recognized a 50-kDa precursor protein in immature oocyst and additional 47- and 42-kDa proteins in older oocysts. A 42-kDa CS protein was detected after initial sporozoite invasion of mosquito salivary glands and an additional 50-kDa precursor CS protein observed later in infected salivary glands. These data confirm previous results with otherPlasmodiumspecies, in which more CS protein precursors were detected in oocysts than in salivary gland sporozoites. A monoclonal antibody (PvPCS) was characterized which reacts with an epitope found only in the 50-kDa precursor CS protein. PvPCS reacted with allP. vivaxsporozoite strains tested by indirect immunofluorescent assay, homogeneously staining the sporozoite periphery with much lower intensity than that produced by anti-CS repeat antibodies. Immunoelectron microscopy using PvPCS showed that the CS protein precursor was associated with peripheral cytoplasmic vacuoles and membranes of sporoblast and budding sporozoites in development oocysts. In salivary gland sporozoites, the CS protein precursor was primarily associated with micronemes and sporozoite membranes. Our results suggest that the 50-kDa CS protein precursor is synthesized intracellularly and secreted on the membrane surface, where it is proteolytically processed to form the 42-kDa mature CS protein. These data indicate that differences in CS protein processing in oocyst and salivary gland sporozoites development may occur.