The Life Cycle of Isospora felis Wenyon, 1923, a Coccidium of the Cat*

SYNOPSIS. A pure strain of Isospora felis derived from a single oocyst was used to study the endogenous cycle. One and a half to two-month-old laboratory-reared, coccidia-free kittens were used thruout the study. The endogenous stages occurred in the epithelial cells of the distal parts of the villi in the ileum and occasionally duodenum and jejunum. All stages lay above the host cell nucleus. There were 3 asexual generations. The 1st generation schizonts were 11–30 by 10–23 μ when mature and contained 16–17 banana-shaped merozoites 11–15 by 3–5 μ. They became mature in 96 or sometimes in 120 hours. The 1st generation merozoites entered new host cells, rounded up and formed 2nd generation schizonts. These formed within themselves 2–10 or more spindle-shaped bodies resembling 1st generation merozoites in shape and size. These were 2nd generation merozoites. They were uninucleate 120 hours after inoculation, but by 144 hours they became larger, multinucleate and some lost their elongate shape and became ovoid. They were then 3rd generation schizonts. They were 12–16 by 4–5 μ. Each formed up to 6 or more banana-shaped merozoites 6–8 by 1–2 μ. The 3rd generation schizonts and merozoites developed within the same host cell and parasitophorous vacuole as the 2nd generation schizonts and merozoites. Mature schizonts containing only 3rd generation merozoites appeared 144 hours after inoculation, were most abundant 168 hours after inoculation, and might be present as late as 216 hours after inoculation. They were 14–36 by 13–22 μ and contained 36 to more than 70 merozoites. The 3rd generation merozoites entered the sexual cycle. The mature microgametocytes were 24–72 by 18–32 μ and contained a central residuum and a large number of microgametes 5–7 by 0.8 μ with 2 posteriorly-directed flagella. The mature macrogametes were 16–22 by 8–13 μ. Gametogony occurred 144–216 hours after inoculation. The prepatent period was 168–192 hours and the patent period 10–11 days. Peak oocyst production occurred on the 6th day of the patent period.     

Experimental Isospora canis and Isospora felis Infection in Mice,Cats, and Dogs*

SYNOPSIS. Two 6-month-old gnotobiotic dogs and 4 five-day-old dogs became infected but did not shed oocysts within 15 days after ingesting the feline coccidian, Isospora felis. Infection of the dogs was evidenced by the shedding of I. felis oocysts by cats consuming extra-intestinal organs of dogs fed I. felis. Likewise, cats became infected with the canine coccidian, Isospora canis, without producing oocysts. Dogs also became infected after ingesting mice previously fed I. canis oocysts. The prcpatent period for I. canis was slightly shorter in dogs fed infected mice (8–9 days) than in those fed oocysts (9–11 days).     

Feline Toxoplasmosis from Acutely Infected Mice and the Development of Toxoplasma Cysts*

SYNOPSIS. The development of Toxoplasma cysts was studied in mice inoculated with tachyzoites by several routes. After 1–30 days of infection, murine tissues were examined microscopically, and portions or whole carcasses were fed to mice and cats. The feces of the cats were examined for oocyst shedding. Cyst-like structures containing distinct PAS-positive granules were first seen after 3 days of infection with tachyzoites, and became numerous by 6 days. Argyrophilic walls were first seen after 6 days, and became numerous by 16 days of infection with tachyzoites. Prepatent periods to oocyst shedding (PPO) were either “short” (3–10 days) or “long” (19–48 days). The “short” PPO was found only in cats that had ingested mice infected for 3 days or longer, and was related to the development of PAS-positive granules in T. gondii, and to high, 60–100%, oral infectivity rates for cats. The “long” PPO followed the ingestion of mice infected for only 1–2 days, and was related to tachyzoites without distinct PAS-positive granules and low, 32% or less, infectivity for cats. The “long” PPO followed also the ingestion of oocysts and the parenteral inoculation of tachyzoites, bradyzoites, or sporozoites. Using the “short” PPO as a criterion for detecting cysts in tissues, it was shown that (a) numerous cysts developed in mice 5 days after inoculation with tachyzoites, 7–9 days after inoculation with cysts, and 9–10 days after inoculation with oocysts, and (b) cysts developed faster and more frequently in the brain and muscle than in lungs, liver, spleen, and kidneys of mice inoculated with tachyzoites.     

Sarcocystis Idahoensis Sp. N. In Deer Mice Peromyscus Maniculatus (Wagner) and Geopher Snakes Pituophis Melanoleucus (Daudin)*,†

SYNOPSIS Deer mice Peromyscus maniculatus (Wagner) were trapped near Hammett, Idaho, as a possible source of Besnoitia jellisoni Frenkel and species of Sarcocystis to be used for life cycle studies. Forty-nine deer mice were necropsied; 20 (40.8%) were positive for sarcocysts structurally identical with those of Sarcocystis idahoensis sp. N. the source of S. idahoensis used for life cycle studies was a Great Basin gopher snake Pituophis melanoleucus deserticola Stejneger killed near Hammett, Idaho; 20 sporulated sporocysts measured 11.1 × 13.4 (11-12 × 13-14) μm. Structurally identical sporocysts were found in 7 of 14 Pacific gopher snakes P. m. catenifer (Blainville), and in 6 of 10 San Diego gopher snakes, P. m. annectens Baird & Girard. Totals of 148 deer mice and 17 gopher snakes were necropsied in the course of life cycle studies. Development of the first generation meronts took place within the hepatocytes of deer mice 2-10 days post-inoculation (PI) with sporulated sporocysts. Rosette-shaped meronts (6-8 days PI) contained tachyzoites attached by their posterior poles to a residual body. After release from the residual body, tachyzoites were initially retained in a meront wall and later released from the hot cells Within muscle cells a single tachyzoite-shaped structure was found 11 days PI and PAS-negative metrocyte-containing sarcocysts (2nd generation meronts) 13-34 days PI. PAS-positive material was first seen in sarcocysts 34 days II at which time bradyzoite formation became apparent. At 160 days PI, 10 sarcocysts measured 0.4 × 5.8 (0.2-0.9 × 1.8-9.9) μ and appeared to be mature and structurally identical with those from naturally infected deer mice. After ingestion of S. idahoensis-infected deer mice by gopher snakes, bradyzoites developed directly into microgamonts and macrogametes. These stages were first seen 5 days PI. Microgamonts were generally located above and macrogametes below the epithelial host cell nucleus. Seven to 11 days PI microgamonts were seen with mature microgametes, and oocysts which had not yet begun sporogony were found with oocyst walls. Clinical signs of illness were generally not observed in infected gopher snakes; however, one snake developed anorexia and cachexia, and became moribund after repeated ingestion of heavily infected deer mice. Acute hepatitis associated with developing meronts often was noted in deer mice given over 15,000 sporocysts each. Five to 6 days PI anorexia, weakness, ataxia, and dyspnea were observed: these clinical signs increased in severity until 6-8 days PI, when mice became recumbent and died, or were killed while moribund. Hepatosplenomegaly, petechial hemorrhage o the serosal and cut surfaces of the liver, and icterus were common. Diffuse coagulative necrosis with cellular infiltration (primarily neutrophils) was noted on microscopic examination.     

Life Cycle of Isospora rivolta (Grassi, 1879) in Cats and Mice*

SYNOPSIS. The endogenous development of Isospora rivolta (Grassi) was studied in cats fed oocysts, and was compared with the endogenous cycle after feeding them mice infected with I. rivolta. For the mouse-induced cycle, 14 newborn cats were killed 12 to 240 h after having been fed mesenteric lymph nodes and spleens of mice. Asexual and sexual development occurred throughout the small intestine, in epithelial cells of the villi and glands of Lieberkuhn. The number of asexual generations was not determined with certainty, but there were at least 3 structurally different meronts. Type I meronts appeared at 12–48 h postinoculation (HPI). They were 8.5(6–13) × 5.1(3–6) μm, contained 2–8 merozoites, and divide by binary division or endodyogeny. Type II meronts were multinucleate merozoite-shaped meronts within a single parasitophorous vacuole. They were found at 48–172 HPI and measured 12.6(9–18) × 9.8(9–13) μm. Individual multinucleate merozoite-shaped meronts were 7–13 × 3–5 μm in sections and contained 2–30 slender (5.5 × 1.0 μm) merozoites. Type III meronts occurred at 72–192 HPI and gamonts at 72–96 HPI. Mature microgamonts measured 11.3(9–15) × 8.0(6–9) μm in sections and up to 21.5 × 14 μm in smears, and contained up to 70 microgametes. Macrogamonts measured 13.3(11–18) × 9.0(5–13) μm in sections and 18 × 16 μm in smears. Oocysts were 10–15 × 9–15 μm in sections and 19.8(17–24) × 18.0(17–23) μm in fixed and stained smears. Unsporulated oocysts in feces were 22.3(18–25) × 19.7(16–23) μm and spomlated oocysts 25.4(23–29) × 23.4(20–26) μm. Sporulation was completed within 24 h at 22–26 C. For the study of the oocyst-induced cycle in cats, 18 newborn cats were killed between 6 and 192 HPI. The endogenous development was essentially similar to the mouse-induced cycle, but merogony and gametogony occurred 12–48 h later than in the latter cycle. Isospora rivolta was pathogenic for newborn but not for weaned cats. Newborn cats fed 10⁵ sporocysts or infected mice usually developed diarrhea 3–4 days after inoculation. Microscopically, desquamation of the tips of the villi and cryptitis were seen in the ilium and cecum in association with meronts and gamonts. For the study of the development of I. rivolta in mice, mice were killed from day 1 to 23 months after having been fed 10⁵–10⁵ sporocysts, and their tissues were examined for the parasites microscopically, and by feeding to cats. The following conclusions were drawn. (A) Isospora rivolta most frequently invaded the mesenteric lymph nodes of mice and remained there for 23 months at least. It also invaded the spleen, liver, and skeletal muscles of mice. This species could not be passed from mouse to mouse. Sporozoites increased in size from ?6.8 × 4.9 μm on day 1 to ?13.4 × 6.9 μm on day 31 postinoculation. Division was not seen. Prepatent period was 4–7 days and patent periods ranged from 2 to several weeks.     

Life Cycles of Two Isospora Species in the Canary,Serinus canarius Linnaeus*

SYNOPSIS. The endogenous stages of Isospora serini Aragão and Isospora canaria Box are described from experimentally infected canaries, Serinus canarius Linnaeus. Unlike other Coccidia, the first part of the I. serini life cycle takes place in mono-nuclear phagocytes. Five asexual generations are described from this cell type; 2 additional asexual generations and the sexual stages take place in the intestinal epithelium. Isospora canaria, on the other hand, has a conventional coccidian life cycle in that all of the endogenous stages are in the epithelium of the small intestine, with 3 asexual generations and the sexual generation described in the duodenal epithelium. The 2 species differ in their position relative to the nucleus of the intestinal epithelial cell. Isospora serini is usually on the lumenal side of the nucleus while I. canaria is below the nucleus, toward the basement membrane. The prepatent period is 4–5 days for I. canaria and 9–10 days for I. serini. Patency lasts for 11–13 days in I. canaria infections, but duration of oocyst output is more chronic in I. serini infections, persisting for as long as 231 days. Both species have a diurnal periodicity of oocyst discharge which occurs in late afternoon and evening.     

Development of patent infection in immunosuppressed C57Bl/6 mice with a single Cryptosporidium meleagridis oocyst

The ability of Cryptosporidium meleagridis to produce patent infection was studied in adult C57BL/6 mice that were immunosuppressed with dexamethasone phosphate provided in the drinking water at a dosage of 16 microg/ml. Four days after the onset of immunosuppression, mice were orally challenged with 1, 3, 10, or 1,000 C. meleagridis TU1867 oocysts per mouse. The mice were monitored daily for 18 days postinoculation for oocyst shedding. Five of 10 mice given a single oocyst, 4 of 5 mice given 3 oocysts, and all 9 mice given either 10 or 1,000 oocysts became infected and began shedding oocysts 5-7 days after challenge and continued to shed oocysts until the end of the experiment on day 18 postchallenge. Approximately 10(7) oocysts per mouse per day were excreted, regardless of the challenge dose. Neither the noninfected, immunosuppressed nor the inoculated, nonimmunosuppressed control mice shed oocysts. The excreted oocysts were confirmed to be those of C. meleagridis by polymerase chain reaction-restriction fragment length polymorphism analysis. We show that C. meleagridis, originally classified as an avian pathogen but recently found in humans with cryptosporidiosis, can produce patent infection in mice infected with a single oocyst. Moreover, we demonstrate that the immunosuppressed C57BL/6 adult mouse is an ideal host for the propagation of clonal populations of C. meleagridis isolates for laboratory studies.     

Characteristics of a line of Eimeria necatrix after 16 successive passages of oocysts collected after peak oocyst production

A line of Eimeria necatrix was selected by repeated passages of oocysts that were collected after peak oocyst production from feces or cecal contents of previously infected chickens. When compared with the parent strain, the new line of E. necatrix after 16 successive passages had the following characteristics: (1) the peak of oocyst production was delayed by 2 days; (2) the sizes of 9 endogenous development stages became larger; and (3) the reproductive capacity and the immunogenicity were both enhanced. This new line of E. necatrix may be used for the development of new coccidiosis vaccines.     

Ultrastructural Studies of the Zygote and Oocyst Wall Formation of Eimeria truncata of the Lesser Snow Goose1

ABSTRACT. Zygote development and oocyst wall formation of Eimeria truncata occurred in epithelial cells in renal tubules and ducts of experimentally infected lesser snow geese (Anser c. caerulescens). Post-fertilization stages were present throughout the kidneys beginning nine days post-inoculation. Initially, a single plasmalemma enclosed the zygote, and type 1 wall-forming bodies (WF1) became labyrinthine and moved toward the surface. There, WF1 degranulated and formed the outer layer of the oocyst wall between the plasmalemma and a newly formed second subpellicular membrane. Several WF2 fused and formed the inner layer, of the oocyst wall between the third and fourth subpellicular membranes. Six subpellicular membranes were observed during wall formation. Other features of oocyst development were similar to those of other eimerian species.     

Life cycle of Isospora rivolta (Grassi, 1879) in cats and mice

The endogenous development of Isospora rivolta (Grassi) was studied in cats fed oocysts, and was compared with the endogenous cycle after feeding them mice infected with I. rivolta. For the mouse-induced cycle, 14 newborn cats were killed 12 to 240 h after having been fed mesenteric lymph nodes and spleens ofmice. Asexual and sexual development occurred throughout the small intestine, in epithelial cells of the villi and glands of Lieberkühn. The number of asexual generations was not determined with certainty, but there were at least 3 structurally different meronts. Type I meronts appeared at 12-48 h postinoculation (HPT). They were 8.5(6-13) x 5.1(3-6) micrometer, contained 2-8 merozoites, and divide by binary division or endodyogeny. Type II meronts were multinucleate merozoite-shaped meronts within a single parasitophorous vacuole. They were found at 48-172 HPI and measured 12.6(9-18) x 9.8(9-13) micrometer. Individual multinucleate merozoite-shaped meronts were 7-13 x 3-5 micrometer in sections and contained 2-30 slender (5.5 x 1.0 micrometer) merozoites. Type III meronts occurred at 72-192 HPI and gamonts at 72-96 HPI. Mature microgamonts measured 11.3(9-15) x 8.0(6-9) micrometer in sections and up to 21.5 x 14 micrometer in smears, and contained up to 70 microgametes. Macrogamonts measured 13.3(11-18) x 9.0(5-13) micrometer in sections and 18 x 16 micrometer in smears, and contained up to 70 microgametes. Macrogamonts measured 13.3(11-18) x 9.0(5-13) micrometer. Sporulation was completed within 24 h at 22-26 C. For the study of the oocyst-induced cycle in cats, 18 newborn cats were killed between 6 and 192 HPI. The endogenous development was essentially similar to the mouse-induced cycle, but merogony and gametogony occurred 12-48 h later than in the latter cycle. Isospora rivolta was pathogenic for newborn but not for weaned cats. Newborn cats fed 10(6) sporocysts or infected mice usually developed diarrhea 3-4 days after inoculation. Microscopically, desquamation of the tips of the villi and cryptitis were seen in the ilium and cecum in association with meronts and gamonts. For the study of the development of I. rivolta in mice, mice were killed from day 1 to 23 months after having been fed 10(5)-10(6) sporocysts, and their tissues were examined for the parasites microscopically, and by feeding to cats. The following conclusions were drawn. (A) Isospora rivolta most freqeuntly invaded the mesenteric lymph nodes ofmice and remained there for 23 months at least. Ii also invaded the spleen, liver, and skeletal muscles of mice. This species could not be passed from mouse to mouse. Sporozoites increased in size from approximately 6.8 x 4.9 micrometer on day 1 to approximately 13.4 x 6.9 micrometer on day 31 postinoculation. Division was not seen. Prepatent period was 4-7 days and patent periods ranged from 2 to several weeks.     

Shedding of Oocysts in Piglets Experimentally Infected with Isospora suis

Forty-seven piglets were inoculated with doses of 100 to 50,000 sporulated oocysts of Isospora suis. After 5-7 days oocysts were found in faeces. The patent period extended from 8 to 16 days. The shedding of oocysts showed a cyclic pattern with 2-3 peaks separated by intervals of approximately 5 days. Subpatent periods were often seen between the peaks. The level of oocyst shedding during the initial days of the patent period reflected, to some extent, the inoculation dose. However, a maximum of OPG at the 100,000 level was observed among one or more piglets from all groups, regardless of the inoculation dose. Among the majority of piglets inoculated with more than 100 oocysts, the highest OPG-figures were observed in the first peak of the cyclic pattern. Unlike this, the maximum of OPG was observed in the second peak of the cycle among 6 of the 7 piglets inoculated with 100 oocysts only. The triphasic pattern was most pronounced in the low dosed group. The marked upscaling of oocyst production, as particularly registered in the low dosed groups, seams to explain at least part of the problems met under practical conditions, when trying to eliminate the transmission of oocysts between successive litters in the farrowing boxes. The cyclic excretion pattern and an apparent absence of autoinfections may indicate that the development of I. suis in the host includes several oocyst producing generations descending from the same initial infection. The presence of subpatent periods can probably explain the marked variation in OPG, as they are often recorded when examining faecal samples from piglets, even when the samples are originating from the same litter.     

Improved production of Neospora caninum oocysts,cyclical oral transmission between dogs and cattle,and in vitro isolation from oocysts

Scarce information is available about Neospora caninum oocysts and sporozoites, in part because only small numbers of oocysts have typically been produced by experimentally infected dogs. We hypothesized that I reason for low experimental production of oocysts is that dogs have been fed tissues from experimentally infected mice instead of tissues from cattle (which are natural intermediate hosts of N. caninum). In this study, 9 dogs were fed tissues from N. caninum-infected calves, and oocyst production was compared with 6 dogs that were fed infected mouse carcasses. The number of oocysts produced by dogs that ingested infected calf tissues (mean = 160,700) was significantly greater (P = 0.03) than the number of oocysts shed by dogs that ingested infected mice (mean = 5,400). The second goal of our experiment was to demonstrate cyclical oral transmission of N. caninum between dogs and cattle. As few as 300 oocysts were used to successfully infect calves, and tissues from these calves induced patent infections in 2 of 3 dogs; oocysts from I of these dogs were administered to another calf, and tissues from this calf subsequently induced a third dog to shed oocysts. Oocysts were confirmed to be N. caninum using a species-specific polymerase chain reaction technique. In addition, sporulated oocysts were used to recover N. caninum in vitro after digestion in an acid-pepsin solution and inoculation of cell monolayers.     

Experimental infection of dogs with Sarcocystis from wapiti.

Ten domestic dogs became infected with Sarcocystis when fed simple portions of heart, esophagus and diaphragm from a two-year-old female wapiti (Cervus canadensis). The prepatent period was 14 days in all exposed dogs; the patent period ranged from 8 to 20 days. Neither the 10 control dogs, nor two dogs fed sporocysts collected from the infected dogs passed sporocysts within the study period. Sporocysts averaged 16.5 by 11.1 micron in size.     

Methods to produce and safely work with large numbers of Toxoplasma gondii oocysts and bradyzoite cysts

Two major obstacles to conducting studies with Toxoplasma gondii oocysts are the difficulty in reliably producing large numbers of this life stage and safety concerns because the oocyst is the most environmentally resistant stage of this zoonotic organism. Oocyst production requires oral infection of the definitive feline host with adequate numbers of T. gondii organisms to obtain unsporulated oocysts that are shed in the feces for 3-10 days after infection. Since the most successful and common mode of experimental infection of kittens with T. gondii is by ingestion of bradyzoite tissue cysts, the first step in successful oocyst production is to ensure a high bradyzoite tissue cyst burden in the brains of mice that can be used for the oral inoculum. We compared two methods for producing bradyzoite brain cysts in mice, by infecting them either orally or subcutaneously with oocysts. In both cases, oocysts derived from a low passage T. gondii Type II strain (M4) were used to infect eight-ten week-old Swiss Webster mice. First the number of bradyzoite cysts that were purified from infected mouse brains was compared. Then to evaluate the effect of the route of oocyst inoculation on tissue cyst distribution in mice, a second group of mice was infected with oocysts by one of each route and tissues were examined by histology. In separate experiments, brains from infected mice were used to infect kittens for oocyst production. Greater than 1.3 billion oocysts were isolated from the feces of two infected kittens in the first production and greater than 1.8 billion oocysts from three kittens in the second production. Our results demonstrate that oral delivery of oocysts to mice results in both higher cyst loads in the brain and greater cyst burdens in other tissues examined as compared to those of mice that received the same number of oocysts subcutaneously. The ultimate goal in producing large numbers of oocysts in kittens is to generate adequate amounts of starting material for oocyst studies. Given the potential risks of working with live oocysts in the laboratory, we also tested a method of oocyst inactivation by freeze-thaw treatment. This procedure proved to completely inactivate oocysts without evidence of significant alteration of the oocyst molecular integrity.     

True Intermediate Hosts for Eimeria funduli (Apicomplexa) from Estuarine Fishes1

Grass shrimp (Palaemonetes pugio) fed liver containing sporulated oocysts of Eimeria funduli permitted development of sporozoites that became infective to a variety of killifishes. The shrimp's gastric mill mechanically ruptured the oocysts. Sporozoites then excysted through an opening in the sporocyst, and by 12 and 13 h postinfection (p.i.) numerous empty sporocysts and free sporozoites occurred extracellularly in the intestine of the grass shrimp. Even at 5, 7, 8, 11, 46, 79, and 83 days p.i., and presumably for many months, numerous sporozoites still occurred free in the alimentary tract or between intestinal cells. The coccidium did not infect killifish at either 2 or 4 days p.i., but did at 5 days; after release from the sporocyst, it became more elongate with a distinct nucleus and two relatively large refractile bodies. Infections of E. funduli resulted in about one half of the fish that were fed either entire hepatopancreas or tips of hepatopancreas from experimentally infected shrimp. Feeding either the entire alimentary tract proximal to the first abdominal segment or any portion of that section from experimentally infected shrimp produced infections in nearly all tested fish. Feeding portions of the cephalothorax without any attached hepatopancreas or alimentary tract failed to produce an infection. Feeding killifish with wild grass shrimp from an enzootic area produced infections in only a fourth of the fish sample; however, feeding experimentally infected wild, laboratory-reared, and juvenile grass shrimp produced infections in nearly all fish. Palaemonid shrimps other than P. pugio also can serve as intermediate hosts for E. funduli, and these shrimps include Palaemonetes vulgaris, P. paludosus, P. kadiakensis, and Macrobrachium ohione. In contrast, a penaeid shrimp, mysidacean, amphipod, and crab fed liver with sporulated oocysts did not produce infections when fed to killifish.     

Experimental Studies of the Life Cycle of Leucocytozoon simondi in Ducks in Norway*

SYNOPSIS. Pekin ducks were infected naturally and experimentally. The life cycle of Leucocytozoon simondi was followed in the ducks and the vector, a new species of Simulium, close to Simulium dogieli. The prepatent period in the ducks was 4–5 days and developing megaloschizonts appeared in 6–7 days. Schizonts were found in liver, spleen, brain, and kidney. In the kidneys they were located in the glomeruli. Sporogony in some individuals was completed in 7 days at 13–14 C, other individuals developed more slowly, as ookinetes were found in flies 8 days after feeding. The rapid asexual cycle combined with a sporogonic cycle, in which some ookinetes develop rapidly and others more slowly, favors the maintenance of the parasite in an environment with relatively low daily temperatures. This and the size of the megaloschizonts indicate differences between this strain of the parasite and the one occurring in North America.     

Extra-Intestinal Stages of Isospora felis and I. rivolta (Protozoa: Eimeriidae) in Cats*

SYNOPSIS. Evidence is presented that Isospora felis and I. rivolta invade the extra-intestinal tissues of cats. Kittens were fed sporocysts of I. felis and I. rivolta. At specific intervals the kittens were killed and suspensions of extra-intestinal tissues were fed to indicator kittens less than a day old. Oocyst production by the indicator kittens within the regular prepatent period was taken as evidence that coccidian stages were present in the inoculum consisting of extra-intestinal tissues of cats. Tissues of kittens infected with I. felis for 5–104 days were infectious to newborn kittens as follows: liver and spleen mixture 3 out of 5 times, mesenteric lymph nodes 4 out of 4 times, brain and muscle mixture 1 out of 5 times, lungs 1 out of 5 times. The prepatent period in kittens consuming oocysts of I. felis was 7-11 days; after consuming extra-intestinal tissues of kittens it was 4–8 days. Distinct coccidian stages unlike those present in the gut were found singly and in groups of 2–15 in lymphoreticular cells of mesenteric lymph nodes of 2 kittens infected for 2–4 days. Tissues of kittens infected with I. rivolta for 5–21 days were infectious to newborn kittens as follows: liver and spleen mixture 3 out of 5 times, mesenteric lymph nodes 1 out of 5 times, brain, muscle and lung mixture none of 5 times. The prepatent period in kittens consuming oocysts of I. rivolta or extra-intestinal tissues of cats was 5–7 days. Coccidian stages occurred singly or in pairs, intracellularly or free in the mesenteric lymph nodes of 3 out of 10 kittens infected for 1–8 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.     

Role of intraepithelial lymphocytes in mucosal immune responses of mice experimentally infected with Cryptosporidium parvum

In order to investigate the role of intestinal intraepithelial lymphocytes (IELs) in host defense against Cryptosporidium parvum infection, conventionally bred immunocompetent (ImCT) ICR mice and immunosuppressed (ImSP) littermates were infected orally with 10(6) C. parvum oocysts. Then fecal oocyst excretion, the number and location of IELs, and their T lymphocyte subsets were observed on days 4, 7, 10, 13, 16, and 20 postinfection (PI). Uninfected ImCT and ImSP mice were used as controls. The starting point of oocyst excretion was day 4 PI in both ImCT- and ImSP-infected mice. The highest oocyst excretion occurred on day 7 PI in both groups, though the number of oocysts excreted was 3 times greater in ImSP than in ImCT mice. In ImCT mice, IELs greatly increased in number on days 16 and 20 PI (P < 0.05), but the increase was minimal in ImSP mice. IELs changed their location from the basal area to intermediate and apical areas of villous epithelial cells during the early stage of infection. In ImCT-infected mice, IEL phenotypes also changed; whereas CD4+ cells increased temporarily on day 7 PI (P < 0.05), CD8+ cells increased significantly on days 16 and 20 PI (P < 0.05). The results strongly suggest that IELs play a significant role in host defense against C. parvum infection, with helper T cells initiating control of the infection and cytotoxic T cells eliminating the parasites.     

The opossum (Didelphis virginiana) as a host for Sarcocystis debonei from cowbirds (Molothrus ater) and grackles (Cassidix mexicanus, Quiscalus quiscula).

Sarcocystis-infected muscles from ducks, cowbirds, and grackles were fed to cats, opossums, rats, and a dog. Only the opossum (Didelphis virginiana) was a suitable definitive host. All opossums that were fed Sarcocystis-infected cowbirds (Molothrus ater) and grackles (Cassidix mexicanus and Quiscalus quiscula) passed sporocysts in their feces. Opossums that ate the cowbirds had prepatent periods of 5 and 10 days and remained patent for at least 105 days. Opossums that ate the grackles became patent on day 10 after the infective meal and remained patent for over 90 (Quiscalus) and 105 (Cassidix) days. A single opossum fed infected muscle from a pintail duck (Anas acuta) passed sporocysts in the feces from days 13 through 18 after infection. No sporocysts were passed by opossums fed infected muscle from the green-winged teal (Anas carolinensis) and shoveller (Spatula clypeata). Sporocysts of duck, cowbird, and grackle origin were structurally similar. Mean dimensions of sporocysts were: duck-origin, 11.2 by 8.2 micron; cowbird-origin, 11.4 by 7.8 micron; Cassidix-origin, 11.2 by 7.8 micron; and Quiscalus-origin, 11.6 by 7.7 micron. We designate the sporocysts of cowbird and grackle origin as Sarcoycstis debonei Vogelsang, 1929 (Syn. Isospora boughtoni Volk, 1938).