Serological and parasitological prevalence of Toxoplasma gondii in wild birds from Colorado

Ground-feeding birds are considered important in the epidemiology of Toxoplasma gondii because they serve as indicators of soil contamination by oocysts, and birds of prey are indicators of T. gondii prevalence in rodents and other small mammals. Cats excrete environmentally resistant oocysts after consuming tissues of T. gondii -infected birds. In the present study, sera and tissues from 382 wild birds from Colorado were tested for T. gondii infection. Antibodies to T. gondii were found in 38 birds with the use of the modified agglutination test (MAT, 1∶25 titer). Tissues (brains, hearts) of 84 birds were bioassayed in mice. Viable T. gondii was isolated from 1 of 1 barn owl (Tyto alba), 1 of 5 American kestrels (Falco sparverius), 1 of 7 ferruginous hawks (Buteo regalis), 1 of 4 rough-legged hawks (Buteo lagopus), 2 of 13 Swainson's hawks (Buteo swainsoni), and 1 of 25 red-tailed hawks (Buteo jamaicensis). This is the first time T. gondii has been isolated from the barn owl, ferruginous hawk, rough-legged hawk, and Swainson's hawk.     

The red-tailed hawk, Buteo jamaicensis, a native definitive host of Frenkelia microti (Apicomplexa) in North America.

Oral inoculation of prairie voles, Microtus ochrogaster, with coccidian sporocysts isolated from the feces of a red-tailed hawk, Buteo jamaicensis, in Kansas, USA, resulted in formation of Frenkelia microti (Apicomplexa: Sarcocystidae) tissue cysts in the brains of the voles. Five additional isolates of morphologically similar sporocysts collected from red-tailed hawks or other Buteo spp. in Kansas failed to result in detectable infections in rodents. These results are the first to verify that red-tailed hawks are natural definitive host in North America for F. microti.     

Prevalence and isolation of Toxoplasma gondii from white-tailed deer in Alabama

Nineteen white-tailed deer (Odocoileus virginianus) from 5 counties in Alabama were examined for infection with Toxoplasma gondii. Twenty-gram samples of heart tissue were bioassayed in mice, serum was examined for T. gondii antibodies using the direct agglutination test, and sections of heart muscle were examined histologically for tissue cysts. Toxoplasma gondii was isolated from 4 of 19 (21%) white-tailed deer hearts. Antibody titers of greater than or equal to 1:50 were found in sera from 7 of 16 (44%) white-tailed deer. Histological examinations of tissue sections from white-tailed deer hearts were negative for T. gondii.     

Prevalence of Toxoplasma gondii in rats (Rattus norvegicus) in Grenada, West Indies

Cats are important in the natural epidemiology of Toxoplasma gondii, because they are the only hosts that can excrete environmentally resistant oocysts. Cats are infected with T. gondii via predation on infected birds and rodents. During 2005, 238 rats (Rattus norvegicus) were trapped in Grenada, West Indies, and their sera along with tissue samples from their hearts and brains were examined for T. gondii infection. Antibodies to T. gondii were assayed by the modified agglutination test (MAT, titer 1:40 or higher); only 2 (0.8%) of 238 rats were found to be infected. Brains and hearts of all rats were bioassayed in mice. Toxoplasma gondii was isolated from the brain and the heart of only 1 rat, which had a MAT titer of 1:320. All of 5 mice inoculated with the heart tissue, and the 5 mice inoculated with the brain tissue of the infected rat remained asymptomatic, even though tissue cysts were found in their brains. Genetically, the isolates of T. gondii from the heart and the brain were identical and had genotype III by using the SAG1, SAG2, SAG3, BTUB, and GRA6 gene markers. These data indicate that rats are not important in the natural history of T. gondii in Grenada.     

Toxoplasma gondii antibodies in naturally exposed wild coyotes, red foxes, and gray foxes and serologic diagnosis of Toxoplasmosis in red foxes fed T. gondii oocysts and tissue cysts

Antibodies to Toxoplasma gondii were determined in sera from 222 coyotes (Canis latrans), 283 red foxes (Vulpes vulpes), and 97 gray foxes (Urocyon cinereoargenteus) from Indiana, Kentucky, Michigan, and Ohio during 1990-1993. Sera were examined in 1:25, 1:100, and 1:500 dilutions by the modified direct agglutination test (MAT) with formalinized whole tachyzoites plus mercaptoethanol. Antibodies were found in 131 (59.0%) of 222 coyotes, 243 (85.9%) of 283 red foxes, and 73 (75.3%) of 97 gray foxes. Antibodies were also measured by different serologic tests in 4 littermate T. gondii-free red foxes fed T. gondii tissue cysts or oocysts; the fifth littermate fox was not fed T. gondii. Antibodies were measured in fox sera obtained 0, 14, and 36-55 days after infection with T. gondii. All 4 foxes fed T. gondii developed MAT and dye test antibody titers of 1:200 or more 14 days later. The latex agglutination test (LAT) and indirect hemagglutination test (IHAT) were less sensitive than MAT for the diagnosis of T. gondii infection in foxes. Antibodies were not detected by LAT (titer 1:64) in the 2 foxes fed tissue cysts nor by IHAT in 1 of the foxes fed tissue cysts. Toxoplasma gondii was isolated by bioassay in mice from tissues of all 4 foxes fed T. gondii. The control fox had no T. gondii antibodies detectable by any of the serologic tests.     

Isolation, tissue distribution, and molecular characterization of Toxoplasma gondii from free-range chickens from Guatemala

The prevalence of Toxoplasma gondii in free-ranging chickens is a good indicator of the prevalence of T. gondii oocysts in the soil because chickens feed from the ground. The prevalence of T. gondii in 50 free-range chickens (Gallus domesticus) from Guatemala was determined. Antibodies to T. gondii were assayed by the modified agglutination test (MAT). Antibodies were found in 37 (74%) chickens with titers of 1:5 (11), 1:10 (7), 1:20 (11), 1:40 (1), 1:80 (1), 1:160 (3), 1:1,280 (2), and 1:2,560 (1). Hearts, pectoral muscles, and brains of 19 chickens with MAT titers of 1:20 or more were bioassayed individually in mice. Tissues from the remaining 31 chickens with titers of 1:10 or lower were pooled and fed to 4 T. gondii-free cats (13 chickens with titers of less than 1:5 to 1 cat, 11 chickens with titers of 1:5 to 2 cats, and 7 chickens with titers of 1:10 to 1 cat). Feces of cats were examined for oocysts; they did not shed oocysts. Toxoplasma gondii was isolated from 8 chickens with MAT titers of 1:20 or more (from 1 of 11 chickens with a titer of 1:20 and all 7 chickens with a titer of 1:80 or more) from the heart, brain, and pectoral muscle (3); heart and pectoral muscle (1); and heart alone (4). Genotyping of these 8 isolates with the SAG2 locus indicated that 5 were type III and 3 were type 1. This is the first report of isolation of T. gondii from chickens from Guatemala.     

Infectious diseases in wild animals in Utah. VI. Experimental infection of birds with Rickettsia rickettsii

Lundgren, D. L. (University of Utah, Salt Lake City), B. D. Thorpe, and C. D. Haskell. Infectious diseases in wild animals in Utah. VI. Experimental infection of birds with Rickettsia rickettsii. J. Bacteriol. 91:963-966. 1966.-Chickens, pigeons, pheasants, sparrow hawks, red-tailed hawks, ravens, magpies, and a marsh hawk were inoculated with Rickettsia rickettsii, the etiological agent of Rocky Mountain spotted fever. The development and persistence of complement-fixing (CF) antibodies and rickettsemias were tested for in these birds. Rickettsiae were recovered from the blood of a number of birds up to the 16th day after inoculation, whereas only the pigeon was found to develop high CF antibody titers. It was concluded that certain species of birds have the potential of contributing to the dissemination of R. rickettsii in nature, and that the CF test is generally unsuitable for serological diagnosis of this organism in birds.     

Experimental tissue cyst induced Toxoplasma gondii infections in dogs

Tissue cyst induced Toxoplasma gondii infections were examined in 2 beagle dogs orally inoculated with tissue cysts. Neither dog developed clinical signs of toxoplasmosis. Both dogs developed low antibody titers to T. gondii. The MAT and IFAT were superior to the LAT and IHT tests for detecting antibodies to T. gondii.     

Fatal pox infection in a rough-legged hawk.

Natural pox infection occurred in a free-living rough-legged hawk (Buteo lagopus) in northeastern North Dakota. Gross, histological and electron microscopic findings were typical of pox infection, and characteristic lesions developed in red-tailed hawks (Buteo jamaicensis) but not in great horned owls (Bubo virginianus) following inoculation with case material. Death of the rough-legged hawk was attributed to starvation rsulting from inability to capture prey and to blood loss from foot lesions.     

Effects of high-pressure processing on Toxoplasma gondii tissue cysts in ground pork

Ingestion of Toxoplasma gondii tissue cysts can result in severe disease in immunocompromised individuals and pregnant women. Treatment of meat and meat products to eliminate viable T. gondii tissue cysts would provide a means to protect consumers. In this study, we examined the effects of high-pressure processing (HPP) on ground pork containing viable tissue cysts of the VEG strain of T. gondii. Ground pork containing tissue cysts was exposed to 400, 300, 200, 100, or 0 MPa treatment for 30, 60, or 90 sec in a commercial HPP unit. The HPP-treated ground pork was subjected to acid-pepsin digestion and bioassayed in mice. The results of the mouse bioassay revealed that none of the mice inoculated with tissue cysts exposed to 400 or 300 MPa became infected, whereas all mice inoculated with tissue cysts exposed to 200, 100, or 0 MPa became infected with T. gondii regardless of exposure time. Results indicate that HPP treatment of ground pork with 300 MPa of pressure will render tissue cysts of T. gondii nonviable and make pork safe for human consumption.     

Virulence for BALB/c mice and antigenic diversity of eight Toxoplasma gondii strains isolated from animals and humans in Brazil

With the purpose of establishing alternative parameters to determine the virulence of Toxoplasma gondii strains, the antigenic diversity of eight strains of the parasite isolated in Brazil was evaluated. BALB/c mice were inoculated i.p. with 10(0), 10(1), 10(2) and 10(3) tachyzoites from each strain. The mortality and time to death of the animals showed that T. gondii strains may be divided in three groups: three strains resulted in 100% of mortality, 5-10 days post inoculation (DPI); three strains resulted in 100% of mortality, 7-19 DPI and brain cysts were observed in the mice which were inoculated; two strains resulted in 0% of mortality, 30 DPI. The analysis of the antigenic profile of different T. gondii strains through Western blotting, using rabbit antiserum to T. gondii, revealed that most antigens are similar to all strains. The mAb 4C3H4 recognized antigens only in the RH, N, AS28 and ME49 strains.     

Experimental congenital toxoplasmosis in Wistar and Holtzman rats

Congenital toxoplasmosis was evaluated in Wistar and Holtzman rats using two strains of Toxoplasma gondii isolated in Brazil. Pregnant rats were inoculated by subcutaneous or intraperitoneal routes with 10(6) or 8 x 10(6) tachyzoites of N strain (virulent for mice) and by subcutaneous or oral routes with 10(2) or 1.2 x 10(3) cysts of P strain (avirulent for mice). The tissues of rat pups born from these rats were bioassayed for T. gondii infection. T. gondii was not observed in the pups born from rats inoculated with N strain. In the animals inoculated with P strain, congenital toxoplasmosis occurred in 22.8% (Wistar rats inoculated with 10(2) cysts by the subcutaneous route), 11.4% (Wistar rats inoculated with 10(2) cysts by the oral route), 21.2% (Wistar rats inoculated with 1.2 x 10(3) cysts by the oral route) and 2.9% of fetal infection (Holtzman rats inoculated with 10(2) cysts by the oral route). None of the pups born from chronically infected mother were infected with T. gondii.     

Use of recombinant antigens for detection of Toxoplasma gondii infection in swine

Five recombinant Toxoplasma gondii antigens, designated B427, C51, C55, V22, and MBP30 were assessed for their potential use in an enzyme-linked immunoassay (EIA) for detection of T. gondii infection in swine. The antigens were evaluated with sera from young pigs that had been fed 1-10,000 T. gondii oocysts of the VEG or GT-1 strains. Results were compared with an EIA using a native T. gondii antigen extract. All 5 recombinant antigens, as well as native antigen, detected antibody responses as soon as 3 wk after infection in pigs inoculated with 1 or 10 oocysts of the VEG strain. This antibody response persisted, at varying levels, for 14 wk when the experiment was terminated. All antigens also detected antibody responses in pigs 4 wk after inoculation with 10,000 oocysts of the GT-1 strain. The antibody response recognized by native antigen remained high through 51 wk after inoculation. However, there was considerable animal-to-animal variation in responses to the individual recombinant antigens. Only antigens C51 and MBP30 consistently detected a positive antibody response over the entire 51-wk course of the experiment. These results suggest that these antigens might be useful for the serological detection of T. gondii infection in pigs.     

Prevalence of Toxoplasma gondii in chickens from an area in southern Brazil highly endemic to humans

The prevalence of Toxoplasma gondii in free-range chickens from Campos dos Goytacazes, Rio de Janeiro State, Brazil, was examined to evaluate environmental contamination by oocysts. Antibodies against T. gondii were assayed by the modified agglutination test (MAT) in sera of chickens. Antibodies against the parasite were found in 129 of 198 chickens with MAT titers > or = 1:25. Brains and hearts of 86 of the 198 chickens were bioassayed in mice for the presence of T. gondii. Viable parasites were isolated from 61 (70.9%) of the 86 chickens. Importantly, viable T. gondii were recovered even from seronegative chickens (MAT titer < or = 1:10). The distribution of parasite-positive chickens by MAT titer was 4 of 17 (titer < or = 1:10), 3 of 4 (titer of 1:20), 2 of 6 (titer of 1:40), and 52 of 59 (titer > or = 1:80). Thus, the high recovery rate of T. gondii observed in mice is indicative of high levels of environmental contamination of free-range chickens by T. gondii oocysts in this area that is endemic to humans.     

Direct evidence for Toxoplasma gondii infection in a wild serow (Capricornis crispus) from mainland Japan

Toxoplasma gondii infection was studied in 41 Japanese serows ( Capricornis crispus ), a goat-antelope in mainland Japan. Blood and muscle specimens were collected from 41 subjects between 2006 and 2010. Presence of antibodies to T. gondii in the sera was examined by using the latex agglutination test (cutoff titer 1:32); 10 of 41 (24.4%) were seropositive. Toxoplasma gondii DNA was detected in muscle tissue of 1 seropositive serow using a semi-nested PCR assay for the B1 gene. A partial nucleotide sequence (220 bp) corresponding to the B1 gene of T. gondii was obtained by direct sequencing; the sequence was 99.1% identical to that of the RH strain. This study is the first report to show direct evidence for the T. gondii infection in Japanese serows.     

Effect of Age and Sex on the Acquisition of Immunity to Toxoplasmosis in Cats*

SYNOPSIS. The effects of age and sex of the cat on oocyst shedding, multiplication of Toxoplasma gondii in tissues of cats, and acquisition of immunity were investigated after oral inoculation of cats with Toxoplasma cysts. Twenty-five cats varying in age from 1 week to 39 months were killed 7-97 days after inoculation with T. gondii. Homogenates of brain, heart, mesenteric lymph nodes, retina, and blood from these cats were inoculated into mice to test for Toxoplasma infectivity. Toxoplasma was isolated more frequently and in higher titers in mice receiving inocula from cats of the youngest age group (1 week old). Toxoplasma gondii was isolated from tissues of only 2 of 21 cats older than 2 months (at the time of inoculation), although all of the animals shed oocysts within 1 week after ingesting the parasites. The number of oocysts shed varied among littermates of the same sex and between sexes. Generally, cats younger than 12 months shed more oocysts than older cats. The number of oocysts shed by older cats varied considerably; males generally shed more oocysts than the females. However, the numbers of cats examined were too small for statistical comparison. Nevertheless, the observations suggest that cats older than 12 months should not be used in experiments where numbers of oocysts shed is critical.     

First biologic and genetic characterization of Toxoplasma gondii isolates from chickens from Africa (Democratic Republic of Congo, Mali, Burkina Faso, and Kenya)

The prevalence of Toxoplasma gondii in free-ranging chickens (Gallus domesticus) is a good indicator of the prevalence of T. gondii oocysts in the soil because chickens feed from the ground. In the present study, prevalence of T. gondii in chickens from Democratic Republic of Congo, Mali, Burkina Faso, and Kenya is reported. The prevalence of T. gondii antibodies in sera of 50 free-range chickens from Congo was 50% based on the modified agglutination test (MAT); antibody titers were 1:5 in 7, 1:10 in 7, 1:20 in 6, 1:40 in 1, and 1:160 or more in 4 chickens. Hearts, pectoral muscles, and brains of 11 chickens with titers of 1:20 or more were bioassayed individually in mice; T. gondii was isolated from 9, from the hearts of 9, brains of 3, and muscles of 3 chickens. Tissues of each of the 14 chickens with titers of 1:5 or 1:10 were pooled and bioassayed in mice; T. gondii was isolated from 1 chicken with a titer of 1:10. Tissues from the remaining 25 seronegative chickens were pooled and fed to 1 T. gondii-free cat. Feces of the cat were examined for oocysts, but none was seen. The results indicate that T. gondii localizes in the hearts more often than in other tissues of naturally infected chickens. Genotyping of these 10 isolates using the SAG2 locus indicated that 8 were isolates were type III, 1 was type II, and 1 was type I. Two isolates (1 type I and 1 type III) were virulent for mice. Toxoplasma gondii was isolated by mouse bioassay from a pool of brains and hearts of 5 of 48 chickens from Mali and 1 of 40 chickens from Burkina Faso; all 6 isolates were avirulent for mice. Genetically, 4 isolates were type III and 2 were type II. Sera were not available from chickens from Mali and Burkina Faso. Toxoplasma gondii antibodies (MAT 100 or more) were found in 4 of 30 chickens from Kenya, and T. gondii was isolated from the brain of 1 of 4 seropositive chickens; this strain was avirulent for mice and was type II. This is the first report on isolation and genotyping of T. gondii from any source from these 4 countries in Africa.     

Toxoplasma gondii does not persist in goldfish (Carassius auratus)

Recent reports of toxoplasmosis in marine mammals raise concern that cold-blooded marine animals are a potential source of Toxoplasma gondii infection. To examine the transmissibility of T. gondii to fish, we observed the development of T. gondii tachyzoites inoculated into oviduct epithelial cells of goldfish (Carassius auratus) microscopically in vitro. Further, the survival period of tachyzoites inoculated into goldfish muscle was bioassayed in mice and through PCR analysis. In cell cultures at 37 C, both RH and Beverley strains of T. gondii tachyzoites had penetrated into cells at 6 hr post inoculation, and were multiplying. In cell cultures at 33 C, many tachyzoites of both strains attached to the host cells, but no intracellular tachyzoites were observed at 24 hr post inoculation. In the T. gondii inoculated goldfish kept at 33 C, tachyzoite DNA was detected in the inoculated region on day 3, but not on day 7. When inoculated goldfish were kept at 37 C, live tachyzoites were seen at the inoculation site on day 3, but not on day 7. These results suggest that T. gondii does not persist in fish.     

Caryospora uptoni and Frenkelia sp.-like coccidial infections in red-tailed hawks (Buteo borealis)

The feces from 16 red-tailed hawks (Buteo borealis) were examined by fecal flotation for the presence of coccidial oocysts or sporocysts. Oocysts of Caryospora uptoni were found in five (31%), sporocysts of a Frenkelia sp.-like coccidium were found in eight (50%), and mixed infections with both species of coccidia were found in three (19%) red-tailed hawks. Neither oocysts nor sporocysts were found in six (38%) red-tailed hawks. Sexual stages of C. uptoni were found in the duodenum and jejunum of a naturally infected red-tailed hawk. Sexual stages were located in enterocytes on the distal three-fourths of the villi. This study shows that over 60% of red-tailed hawks may be passing coccidial oocysts/sporocysts in their feces and provides morphological information for diagnosing C. uptoni infections in histological sections.     

Biological and molecular characterizations of Toxoplasma gondii strains obtained from southern sea otters (Enhydra lutris nereis)

Toxoplasma gondii was isolated from brain or heart tissue from 15 southern sea otters (Enhydra lutris nereis) in cell cultures. These strains were used to infect mice that developed antibodies to T. gondii as detected in the modified direct agglutination test and had T. gondii tissue cysts in their brains at necropsy. Mouse brains containing tissue cysts from 4 of the strains were fed to 4 cats. Two of the cats excreted T. gondii oocysts in their feces that were infectious for mice. Molecular analyses of 13 strains indicated that they were all type II strains, but that they were genetically distinct from one another.