Development of Caryospora simplex (Apicomplexa: Eimeriidae) from sporozoites to oocysts in human embryonic lung cell culture

Leighton tubes containing monolayers of human embryonic lung cells were inoculated with 70,000 or 30,000 sporozoites of the viperid coccidium Caryospora simplex and examined at 1, 2, 4, 6, 8, 10, 12, 14, 16, and 18 days post-inoculation (PI). By day 1 PI, sporozoites had penetrated cells and were within parasitophorous vacuoles. Most sporozoites became spherical and then underwent karyokinesis several times between days 2 and 6 PI. Mature Type I meronts were found on days 6-16 PI and contained 8 to 22 short, stout merozoites. Mature Type II meronts were present on days 10-18 PI and contained 8 to 22 long, slender merozoites. Developing gamonts (undifferentiated sexual stages) were observed on days 14 and 16 PI. Mature micro- and macrogametes and thin-walled unsporulated oocysts were present on days 16 and 18 PI. Attempts to sporulate oocysts in tissue culture medium or in a 2.5% (w/v) aqueous solution of K2Cr2O7 at 25 degrees C and 37 degrees C were unsuccessful; only a few oocysts developed to the contracted sporont stage. Four Swiss-Webster mice injected intraperitoneally with merozoites obtained from Leighton tubes on day 10 PI did not acquire infections. This is the second coccidium reported to complete its entire development, from sporozoite to oocyst, in cell culture.     

Complete Development of Isospora suis of Swine in Chicken Embryos1

Development of the swine coccidium, Isospora suis, in embryonated chicken eggs is described. The allantoic cavities of eight-to-ten-day-old white Leghorn embryos were inoculated with either 100,000 or 200,000 sporozoites. Developmental stages morphologically similar to those found in the intestines of piglets were present in the endodermal layer of the chorioallantoic membrane (CAM), beginning three days post inoculation (PI). No stages were found in the mesodermal or ectodermal layers of the CAM and none were observed in heart, lung, liver, or spleen. Type I meronts and merozoites were found on days 3 through 10 PI. Type II meronts and merozoites were found days 4 through 10 PI. Mature microgamonts, macrogamonts, and oocysts were found on days 7 through 10 PI. Oocysts appeared to be retained in the endodermal cells and in ovo sporulation did not occur. Attempts to sporulate CAM-derived oocysts were not successful. Isospora suis was not pathogenic for embryos under the conditions of this study. This study represents the first fully documented report of complete development of a mammalian coccidium in chicken embryos.     

Complete development of Isopora suis of swine in chicken embryos

Development of the swine coccidium , Isospora suis, in embryonated chicken eggs is described. The allantoic cavities of eight-to-ten-day-old white Leghorn embryos were inoculated with either 100,000 or 200,000 sporozoites. Developmental stages morphologically similar to those found in the intestines of piglets were present in the endodermal layer of the chorioallantoic membrane (CAM), beginning three days post inoculation (PI). No stages were found in the mesodermal or ectodermal layers of the CAM and none were observed in heart, lung, liver, or spleen. Type I meronts and merozoites were found on days 3 through 10 PI. Type II meronts and merozoites were found days 4 through 10 PI. Mature microgamonts , macrogamonts, and oocysts were found on days 7 through 10 PI. Oocysts appeared to be retained in the endodermal cells and in ovo sporulation did not occur. Attempts to sporulate CAM-derived oocysts were not successful. Isospora suis was not pathogenic for embryos under the conditions of this study. This study represents the first fully documented report of complete development of a mammalian coccidium in chicken embryos.     

The Life Cycle of Cryptosporidium baileyi n. sp. (Apicomplexa, Cryptosporidiidae) Infecting Chickens

ABSTRACT. The life cycle and morphology of a previously undescribed species of Cryptosporidium isolated from commercial broiler chickens is described. The prepatent period for Cryptosporidium baileyi n. sp. was three days post oral inoculation (PI) of oocysts, and the patent period was days 4–24 PI for chickens inoculated at two days of age and days 4–14 for chickens inoculated at one and six months of age. During the first three days PI, most developmental stages of C. baileyi were found in the microvillous region of enterocytes of the ileum and large intestine. By day 4 PI, most parasites occurred in enterocytes of the cloaca and bursa of Fabricius (BF). Mature Type I meronts with eight merozoites first appeared 12 h PI and measured 5.0 × 4.9 μm. Mature Type II meronts with four merozoites and a large granular residuum first appeared 48 h PI and measured 5.1 × 5.1 μm. Type I meronts with eight short merozoites and a large homogeneous residuum first appeared 72 h PI and measured 5.2 × 5.1 μm. Microgamonts (4.0 × 4.0 μm) produced 16 micro-gametes that penetrated into macrogametes (4.7 × 4.7 μm). Macrogametes gave rise to two types of oocysts that sporulated within the host cells. Most were thick-walled oocysts (6.3 × 5.2 μm), the resistant forms that passed unaltered in the feces. Some were thin-walled oocysts whose wall (membrane) readily ruptured upon release from the host cell. Sporozoites from thin-walled oocysts were observed penetrating enterocytes in mucosal smears. The presence of thin-walled, autoinfective oocysts and the recycling of Type I meronts may explain why chickens develop heavy intestinal infections lasting up to 21 days. Oocysts of C. baileyi were inoculated orally into several animals to determine its host specificity. Cryptosporidium baileyi did not produce infections in suckling mice and goats or in two-dayold or two-week-old quail. One of six 10-day-old turkeys had small numbers of asexual stages only in the BF. Four of six one-day-old turkeys developed mild infections only in the BF, and sexual stages of the parasite were observed in only one of the four. All seven one-day-old ducks and seven two-day-old geese developed heavy infections only in the BF with all known developmental stages present.     

The life cycle of Cryptosporidium baileyi n. sp. (Apicomplexa, Cryptosporidiidae) infecting chickens

The life cycle and morphology of a previously undescribed species of Cryptosporidium isolated from commercial broiler chickens is described. The prepatent period for Cryptosporidium baileyi n. sp. was three days post oral inoculation (PI) of oocysts, and the patent period was days 4-24 PI for chickens inoculated at two days of age and days 4-14 for chickens inoculated at one and six months of age. During the first three days PI, most developmental stages of C. baileyi were found in the microvillous region of enterocytes of the ileum and large intestine. By day 4 PI, most parasites occurred in enterocytes of the cloaca and bursa of Fabricius (BF). Mature Type I meronts with eight merozoites first appeared 12 h PI and measured 5.0 x 4.9 micrometers. Mature Type II meronts with four merozoites and a large granular residuum first appeared 48 h PI and measured 5.1 x 5.1 micrometers. Type III meronts with eight short merozoites and a large homogeneous residuum first appeared 72 h PI and measured 5.2 x 5.1 micrometers. Microgamonts (4.0 x 4.0 micrometers) produced approximately 16 microgametes that penetrated into macrogametes (4.7 x 4.7 micrometers). Macrogametes gave rise to two types of oocysts that sporulated within the host cells. Most were thick-walled oocysts (6.3 x 5.2 micrometers), the resistant forms that passed unaltered in the feces. Some were thin-walled oocysts whose wall (membrane) readily ruptured upon release from the host cell. Sporozoites from thin-walled oocysts were observed penetrating enterocytes in mucosal smears. The presence of thin-walled, autoinfective oocysts and the recycling of Type I meronts may explain why chickens develop heavy intestinal infections lasting up to 21 days. Oocysts of C. baileyi were inoculated orally into several animals to determine its host specificity. Cryptosporidium baileyi did not produce infections in suckling mice and goats or in two-day-old or two-week-old quail. One of six 10-day-old turkeys had small numbers of asexual stages only in the BF. Four of six one-day-old turkeys developed mild infections only in the BF, and sexual stages of the parasite were observed in only one of the four. All seven one-day-old ducks and seven two-day-old geese developed heavy infections only in the BF with all known developmental stages present.     

Extraintestinal Development of Caryospora simplex (Apicomplexa: Eimeriidae) in Experimentally Infected Mice,Mus musculus1

Developmental stages of Caryospora simplex were found in connective tissue of the cheek, tongue, and nose of Swiss-Webster and C57 BL/6 mice (Mus musculus) from 8 through 70 days after oral inoculation with 50,000 or 250,000 oocysts, or 60,000 free sporocysts of the same species obtained from an Ottoman viper, Vipera xanthina xanthina. The earliest developmental stages were seen on day 8 post-inoculation (PI) and consisted of two types of meronts and gamonts (undifferentiated sexual stages). Gamonts, microgametocytes, macrogametes, and unsporulated oocysts were found on days 10 and 12 PI. Fully sporulated, thin-walled oocysts containing eight sporozoites surrounded by a thin sporocyst membrane were first seen 12 days PI. Monozoic cysts (caryocysts) were first seen 12 days PI and appeared fully viable throughout the duration of the study, 70 days PI. Four mice injected intra-peritoneally with 150,000 free sporozoites and killed 12 days PI contained unsporulated and sporulated oocysts in connective tissues of the cheek, tongue, and nose, suggesting that sporozoites may be carried to the site of infection via the lymphatic/circulatory system. Four cotton rats, Sigmodon hispidus, inoculated orally with 250,000 oocysts all had unsporulated and sporulated oocysts of C. simplex in connective tissue of the cheek, tongue, and nose when killed on day 12 PI, indicating extraintestinal development in the secondary host is not species specific. This is the first report of a heteroxenous coccidium with both asexual and sexual development in the primary (predator) and secondary (prey) hosts.     

Endogenous development of Eimeria minasensis in experimentally infected goats

The endogenous development of Eimeria minasensis was studied in 9 coccidia-free goat kids inoculated with 10(5) sporulated oocysts/kg body weight. Kids were killed 4, 7 (2 animals), 10, 13, 16, 18, 19, and 22 days after inoculation (DAI). In tissue sections of the intestines stained with hematoxylin and eosin and examined by light microscopy, 2 generations of meronts, gamonts, gametes, and oocysts were found. The first generation of meronts developed in cells deep in the lamina propria of the jejunum and ileum. Mature giant meronts (299.4x243.8 microm) found 16 DAI were visible to the naked eye and contained a large number of crescent-shaped merozoites. The second generation of meronts developed in the epithelial cells of crypts of the ileum and above the host cell nuclei. Mature meronts (11.5x10.1 microm) with 18-28 comma-shaped merozoites were first seen 16 DAI. Gametogenesis took place in epithelial cells of the crypts and villi of the terminal part of the ileum, cecum, and colon. Macrogametes (27.8x17.6 microm), mature microgamonts (21.3x17.0 microm), microgametes, and oocysts (30.5x19.4 microm) were found 19 DAI. Sexual stages were below the host cell nucleus.     

Extraintestinal development of Caryospora simplex (Apicomplexa: Eimeriidae) in experimentally infected mice, Mus musculus

Developmental stages of Caryospora simplex were found in connective tissue of the cheek, tongue, and nose of Swiss-Webster and C57 BL/6 mice (Mus musculus) from 8 through 70 days after oral inoculation with 50,000 or 250,000 oocysts, or 60,000 free sporocysts of the same species obtained from an Ottoman viper, Vipera xanthina xanthina. The earliest developmental stages were seen on day 8 post-inoculation (PI) and consisted of two types of meronts and gamonts (undifferentiated sexual stages). Gamonts, microgametocytes, macrogametes, and unsporulated oocysts were found on days 10 and 12 PI. Fully sporulated, thin-walled oocysts containing eight sporozoites surrounded by a thin sporocyst membrane were first seen 12 days PI. Monozoic cysts (caryocysts) were first seen 12 days PI and appeared fully viable throughout the duration of the study, 70 days PI. Four mice injected intra-peritoneally with 150,000 free sporozoites and killed 12 days PI contained unsporulated and sporulated oocysts in connective tissues of the cheek, tongue, and nose, suggesting that sporozoites may be carried to the site of infection via the lymphatic/circulatory system. Four cotton rats, Sigmodon hispidus, inoculated orally with 250,000 oocysts all had unsporulated and sporulated oocysts of C. simplex in connective tissue of the cheek, tongue, and nose when killed on day 12 PI, indicating extraintestinal development in the secondary host is not species specific. This is the first report of a heteroxenous coccidium with both asexual and sexual development in the primary (predator) and secondary (prey) hosts.     

A Comparison of Endogenous Development of Three Isolates of Cryptosporidium in Suckling Mice1

ABSTRACT. Suckling mice were used as a model host to compare the endogenous development of three different isolates of Cryptosporidium: one from a naturally infected calf, one from an immunocompetent human with a short-term diarrheal illness, and one from a patient with acquired immune deficiency syndrome (AIDS) and persistent, life-threatening, gastrointestinal cryptosporidiosis. After oral inoculation of mice with oocysts, no differences were noted among developmental stages of the three isolates in their sites of infection, times of appearance, and duration, morphology, and fine structure. Sporozoites excysted within the lumen of the duodenum and ileum, penetrated into the microvillous region of villous enterocytes, and developed into type I meronts with six or eight merozoites. Type I merozoites penetrated enterocytes and underwent cyclic development as type I meronts or they became type II meronts with four merozoites. Type II merozoites did not exhibit cyclic development but developed directly into sexual forms. Microgamonts produced £16 small, bullet-shaped microgametes, which were observed attaching to and penetrating macrogametes. Approximately 80% of the oocysts observed in enterocytes had a thick, two-layered wall. After sporulating within the parasitophorous vacuole, these thick-walled oocysts passed through the gut unaltered and were the resistant forms that transmitted the infection to a new host. Approximately 20% of the oocysts in enterocytes consisted of four sporozoites and a residuum surrounded only by a single oocyst membrane that ruptured soon after the parasite was released from the host cell. The presence of thin-walled, autoinfective oocysts and recycling of type I meronts may explain why a small oral inoculum can produce an overwhelming infection in a suitable host and why immune deficient persons can have persistent, life-threatening cryptosporidiosis in the absence of repeated oral exposure to thick-walled oocysts.     

Development of the Swine Coccidium Eimeria debliecki Douwes, 1921 in Mammalian Cell Cultures1

Sporozoites of Eimeria debliecki entered human fetal lung and porcine kidney cells grown in cultures and underwent one merogenous cycle, terminating in the production of second-generation trophozoites. Sporozoites were intracellular 1 h post-inoculation (PI) and developed into sporozoite-shaped meronts at 40 h PI. These meronts, one of which was motile, had from two to ten nuclei. Sporozoite-shaped meronts then developed into elongate or spheroidal meronts with 10 to 24 nuclei by two days PI. Ten to 26 first-generation merozoites were formed by budding from the meront surface. Mature first-generation merozoites were most numerous three days PI. Most meronts had ruptured and released nonmotile merozoites into the culture medium by four days PI. Merozoites that were not released became rounded and developed into second-generation trophozoites. Refractile bodies were present in all developmental stages. No further development was observed five through eight days PI.     

In vitro Development of First-Generation Schizonts of Eimeria canadensis (Bruce, 1921)*†

SYNOPSIS. In vitro development of Eimeria canadensis from cattle was studied in monolayer cultures of various bovine cell lines grown on coverslips in Leighton tubes. Excysted sporozoites were used for inoculation of the cell cultures. Sporozoites entered the host cells within a few minutes, but apart from a reduction in the number of refractile bodies, changed little in appearance during the first 9 days. Beginning at 91/2 days postinoculation, sporozoites developed into sporozoite-shaped schizonts or, less frequently, transformed into trophozoites. Sporozoite-shaped schizonts with as many as 8 nuclei were observed transforming into spheroid schizonts. At 111/2 days, intermediate schizonts had a characteristic single mass of refractile granules and 60–80 nuclei. Deep invaginations, which resulted in the formation of several blastophores, usually occurred when schizonts had about 100 nuclei. Merozoites were formed as a result of radial outgrowth from the surface of spheroid schizonts as well as of blastophores. Mature merozoites were seen 1st after 13 days.     

The life cycle of Eimeria falciformis var. pragensis (Sporozoa: Coccidia) in the mouse, Mus musculus

The life cycle of Eimeria falciformis var. pragensis, established from a single oocyst, is described in experimentally infected mice (Mus musculus). The coccidium had a prepatent period of 7 days and a patent period of 10--16 days. Oocysts were spherical to ellipsoidal in shape and measured 21.2 x 18.3 micron. Sporulation time was 3 to 3.5 days. Sporocysts measured 12.2 x 7.2 micron and contained a circular to avoid granular sporocyst residuum measuring 5.5 X 5.0 micron. One, 2 or 3 circular to rectangular polar granules were observed within each sporulated oocyst. The endogenous stages developed primarily in the cecum and colon and only occasionally in the lower ileum. Four generations of schizonts were found. Mature 1st-generation schizonts, first observed 48 hr postinfection (PI), measured 17.8 x 12.3 micron and had 12 merozoites that measured 13.3 x 2.0 micron. Mature 2nd-generation schizonts appeared 78 hr PI. They measured 10.2 x 9.3 micron and had 8 merozoites measuring 5.0 x 1.6 micron. Mature 3rd-generation schizonts appeared first at 114 hr PI and measured 17.5 x 10.2 micron and had 10 merozoites that measured 12.4 x 1.8 micron. Mature 4th-generation schizonts appeared first at 144 hr PI. They measured 18.2 x 15.3 micron and had 18 merozoites. The merozoites of the 4th-generation schizont were 4.5 x 1.2 micron. Mature macrogamonts and microgamonts developed simultaneously appearing at 156 hr PI. Macrogamonts measured 16 x 14.5 micron and microgamonts were 18.2 x 15.3 micron. In experimentally infected rats (Rattus norvegicus), development of E. falciformis var. pragensis progressed only as far as mature 1st-generation schizonts.     

Complete Development of Caryospora bigenetica (Apicomplexa: Eimeriidae) In Vitro1

The development of Caryospora bigenetica in vitro is described by light microscopy. Sporozoites from snake-derived oocysts were purified and inoculated onto cultures of primary testicle cells of the cotton rat, cotton rat kidney cells, and human fetal lung cells. Intracellular sporozoites were observed one and two days postinoculation (DPI). Motile, extracellular first-generation merozoites were present 3 DPI, and second-generation merozoites were present 5 DPI. Mature gamonts were observed 9 DPI and developed into unsporulated oocysts by 10 DPI. Oocysts sporulated in vitro, and excystation was observed. Cells that were penetrated by in vitro-produced sporozoites formed caryocysts by 16 DPI. To test infectivity of in vitro-derived stages, merozoites were removed from cultured cells 5 DPI and inoculated intraperitoneally into a mouse; infection resulted. Sporulated oocysts removed from cell cultures 12 DPI produced facial swelling in an orally inoculated cotton rat.     

Further studies on the development of gamonts and oocysts of Eimeria magna in cultured cells

Monolayer, cell-line cultures of embryonic bovine trachea, Madin-Darby bovine kidney (MDBK), and monolayers (RK-1) or aggregates of primary rabbit kidney cells were inoculated with merozoites obtained from rabbits that had been inoculated 3 to 5 1/2 days earlier with Eimeria magna. Merozoites obtained from from rabbits 3 days entered cells and underwent only merogony, whereas 3 1/2-5 1/2-day-old merozoites formed gamonts as well as meronts. Merozoites arising from the first or second meront generation in culture formed another meront generation or gamonts. Third-generation merozoites formed only gamonts. Most merozoites remained within the parasitophorous vacuole of the original host cell and transformed into macro- or microgamonts or meronts. Some such macro- and microgamonts then fused with each other to form larger multinucleated bodies. Such microgamonts formed microgametes, but multinucleate macrogamonts did not form oocysts. Mature microgamonts were 34 microns in diameter, and contained several hundred biflagellate microgametes. Mature macrogamonts measured 29.1 x 21.5 microns, unsporulated oocysts were 31.2 x 22 microns, and sporulated oocysts were 32 x 23.1 microns. Oocysts obtained from cell cultures were sporulated and then inoculated by gavage into rabbits, which passed E. magna oocysts 6--10 days later. Sporozoites, obtained from oocysts produced in culture or from rabbits that had been inoculated with the vitro-produced oocysts, developed to first- and second-generation meronts in MDBK or RK-1 cultures.     

Carbon Dioxide as the Initial Stimulus for Excystation of Eimeria tenella Oocysts*

SYNOPSIS. The stimulus necessary to initiate in vitro excystation of the chicken coccidium Eimeria tenella was provided by exposure of intact sporulated oocysts to an atmosphere of carbon dioxide. This stimulus produced a thinning and indentation at the micropylar region and oocysts became permeable to trypsin and bile. Sporozoites became active and began to escape from sporocysts into the oocyst cavity and then to the outside thru the altered micropyle after incubation in the enzyme-bile mixture. Activation of sporozoites when CO₂-pretreated oocysts were incubated in trypsin and bile, was used as the criterion to determine the number of oocysts responding to the initial stimulus. Thus, activation of sporozoites within intact oocysts was an indirect measurement of the number of oocysts stimulated during CO₂-pretreatment. Approximately 90% of the oocysts contained active sporozoites after 18 hr of pretreatment with carbon dioxide and 8 hr incubation in trypsin and bile at 38 or 41 C, respectively. Pretreatment of oocysts with air, N₂, O₂, or He resulted in 8% or less activation during incubation in trypsin and bile. Approximately 83% of the oocysts responded to the stimulus during 8 hr CO₂-pretreatment at 41 C, whereas at 38 C, 16 hr of pretreatment were required for a similar response. The stimulus did not elicit a response from oocysts held at 23 C during the pretreatment gasphase. No significant difference occurred in number of oocysts containing active sporozoites after sufficient CO₂-pretreatment for maximum stimulation of oocysts and incubation in trypsin and bile at 38 or 41 C.     

Observations on the endogenous stages of Eimeria crandallis in domestic lambs (Ovis aries)

Lambs reared coccidia-free were inoculated orally with various numbers of sporulated oocysts of E. crandallis and were killed between 1 and 22 days after inoculation; tissues were examined histologically. Sporozoites were seen 1, 2 and 3 days after inoculation (DAI) in crypt epithelial cells in the mid-jejunum. Infected cells migrated into the lamina propria where the parasite within them developed into a firstgeneration meront containing about 250,000 merozoites at 10 DAI. A second generation of meronts was seen at 10–12 DAI, each containing up to about 10 merozoites, situated mainly at the bases of crypts in the jejunum and ileum but also in the caecum. From 11 DAI pro-gamonts were seen which were enveloped by the host cell nucleus and which divided in synchrony with the host cell for an undetermined number of generations. Mature gamonts began to develop from them by 16 DAI. Oocyst output began at 16 DAI and rose to a peak at about 22 DAI. Up to 10⁸ oocysts were produced per oocyst inoculated. They showed wide variation in size and colour.     

A comparison of endogenous development of three isolates of Cryptosporidium in suckling mice

Suckling mice were used as a model host to compare the endogenous development of three different isolates of Cryptosporidium: one from a naturally infected calf, one from an immunocompetent human with a short-term diarrheal illness, and one from a patient with acquired immune deficiency syndrome (AIDS) and persistent, life-threatening, gastrointestinal cryptosporidiosis. After oral inoculation of mice with oocysts, no differences were noted among developmental stages of the three isolates in their sites of infection, times of appearance, and duration, morphology, and fine structure. Sporozoites excysted within the lumen of the duodenum and ileum, penetrated into the microvillous region of villous enterocytes, and developed into type I meronts with six or eight merozoites. Type I merozoites penetrated enterocytes and underwent cyclic development as type I meronts or they became type II meronts with four merozoites. Type II merozoites did not exhibit cyclic development but developed directly into sexual forms. Microgamonts produced approximately 16 small, bullet-shaped microgametes, which were observed attaching to and penetrating macrogametes. Approximately 80% of the oocysts observed in enterocytes had a thick, two-layered wall. After sporulating within the parasitophorous vacuole, these thick-walled oocysts passed through the gut unaltered and were the resistant forms that transmitted the infection to a new host. Approximately 20% of the oocysts in enterocytes consisted of four sporozoites and a residuum surrounded only by a single oocyst membrane that ruptured soon after the parasite was released from the host cell. The presence of thin-walled, autoinfective oocysts and recycling of type I meronts may explain why a small oral inoculum can produce an overwhelming infection in a suitable host and why immune deficient persons can have persistent, life-threatening cryptosporidiosis in the absence of repeated oral exposure to thick-walled oocysts.     

Developmental stages of Hepatozoon hemprichii sp. nov. infecting the skink Scincus hemprichii and the tick Hyalomma impeltatum from Saudi Arabia

The life cycle of Hepatozoon hemprichi n. sp. is described; the vertebrate host is Scincus hemprichii and it is vectored by Hyalomma impeltatum. Erythrocytic stages of 18 ± 1.8 × 4 ± 0.8 μm developed in the hemocoel of ticks to sporozoites within 16-18 days. Schizogony occurred in the liver parenchyma and the endothelial cells of blood capillaries in lung and spleen. Mature schizonts measuring 27 ± 3.11 × 20.13 ± 3.0 μm produced 28 merozoites (on average). The merozoites were 13 ± 1.21 × 1.21 ± 0.72 μm with nuclei 5 ± 0.65 × 2.1 ± 0.51 μm. Syzygy and differentiation of gamonts took place in tick's hemocoel up to the third day post-infection (PI). The microgamont (16 ± 0.31 × 18 ± 0.42 μm) produced 4, uniflagellated microgametes at 4-5 days PI. The microgamete measured 15.2 ± 0.31 μm while the flagellum was always at least 26 μm. The macrogamete was very large in size (31 ± 3.11 μm) with a central nucleus. After fertilization, (5-6 days PI) zygotes developed into oocysts (55 ± 3.41 × 52 ± 4.11 μm) in which repeated mitotic divisions with centripetal invaginations occurred; each contained 18 banana-shaped sporozoites, 13.61 ± 0.8 × 1.2 ± 0.31 μm in size. Experimental transmission was successfully carried out by oral administration or by intra-peritoneal inoculation of the infective stages (sporozoites) to uninfected skinks and led to the appearance of blood stages after 5 wk and 4 wk, respectively.     

New Observations on First-Generation Merogony of Eimeria tuskegeensis in Sigmodon hispidus1

First-generation development of Eimeria tuskegeensis was evaluated using light microscopy. Sporozoite-shaped meronts containing a prominent refractile body were observed in small intestinal cells of an experimentally infected cotton rat at 24 h post inoculation (PI). Mature spherical or subspherical meronts containing crescent-shaped merozoites were observed at 36 h PI. Refractile bodies were observed in some of these merozoites. Sporozoite-shaped meronts that were isolated from host intestinal cells and inoculated onto human fetal lung cell cultures penetrated the cultured cells by 2 h PI. A mature, subspherical, first-generation meront containing seven merozoites was observed at 9 h PI in cell culture, indicating that sporozoite-shaped meronts isolated from the host retained their infectivity.     

Development of the swine coccidium Eimeria debliecki Douwes, 1921 in mammalian cell cultures

Sporozoites of Eimeria debliecki entered human fetal lung and porcine kidney cells grown in cultures and underwent one merogenous cycle, terminating in the production of second-generation trophozoites. Sporozoites were intracellular 1 h post-inoculation (PI) and developed into sporozoite-shaped meronts at 40 h PI. These meronts, one of which was motile, had from two to ten nuclei. Sporozoite-shaped meronts then developed into elongate or spheroidal meronts with 10 to 24 nuclei by two days PI. Ten to 26 first-generation merozoites were formed by budding from the meront surface. Mature first-generation merozoites were most numerous three days PI. Most meronts had ruptured and released nonmotile merozoites into the culture medium by four days PI. Merozoites that were not released became rounded and developed into second-generation trophozoites. Refractile bodies were present in all developmental stages. No further development was observed five through eight days PI.