[The analysis of related factors on the unilateral aldosterone secretion in the functional diagnosis of primary aldosteronism and establishment of a predictive model].

Objective: To explore related factors associated with unilateral aldosterone secretion of patients with primary aldosteronism (PA) and construct a predictive model. Methods: The clinical data of patients who were diagnosed as PA in West China Hospital from April 2016 to September 2020 was analyzed retrospectively. According to the results of the adrenal enhancement CT, patients were divided into 3 groups, namely non-nodular group with no bilateral adrenal glands lesions, only with unilateral adrenal hyperplasia or bilateral adrenal hyperplasia, unilateral nodule group with unilateral adrenal nodules and the contralateral adrenal glands with hyperplasia or not, and bilateral nodules group with nodules in both adrenal glands. Regarding the related factors of dominant side of aldosterone secretion, univariate analysis and binary logistic regression were used. Receiver operating characteristic curve and nomogram were used to evaluate the diagnostic performance of regression models. Results: A total of 237 patients with PA were included, of which, 118 males and 119 females, the median age was 39 years, and the body mass index (BMI) was (25.2±3.5) kg/m2. There were 157 (66.2%) of 237 patients with typical imaging findings. There were 32 cases in no-nodular group, 183 cases in unilateral nodule group, and 22 cases in bilateral nodules group. Multivariate analysis showed that age (OR=0.876, P<0.001), blood potassium concentration (OR=0.430, P=0.004), and typical imaging findings (OR=2.202, P=0.035) were associated with unilateral aldosterone secretion. As for unilateral nodule group, multivariate analysis showed that age (OR=0.900, P<0.001), plasma aldosteronism concentration (PAC) (OR=1.050, P=0.018), and typical imaging findings (OR=2.637, P=0.025) were associated with unilateral aldosterone secretion. The agreement rate between the dominant side of the adrenal CT and AVS was only 50.2%. Multivariate analysis showed that age (OR=0.954, P=0.001), BMI (OR=0.893, P=0.024) and PAC (OR=1.043, P=0.011) were independently associated with concordance between AVS and CT. The cut-off value of the ROC curve was 0.43; the model sensitivity was 56.3%; the specificity was 86.7% and the area under the ROC curve was 0.742. Conclusions: Age is an important predictor in the diagnosis of PA subtypes. It is recommended to refer to subgroup based on imaging results for clinical decision. For patients with no obvious lesions or bilateral lesions on CT, AVS should be performed as far as possible to confirm the subtypes in diagnosis of PA.

Value of preoperative computed tomography in predicting the local staging of cancer of the tongue at primary surgery.

Preoperative staging is essential for the planning of treatment of cancer. This study was designed to evaluate the accuracy of computed tomography (CT) in predicting the local stage of tongue cancer by comparing it with the gold standard of histopathology. A total of 233 patients with newly-diagnosed tongue cancer was retrospectively reviewed, and the size of the tumour and the status of the cervical lymph node were compared between CT images and histopathological results. Patients with stage II cancer were followed up to assess the influence of inaccurate preoperative staging on prognosis. The accuracy of local staging by CT was 47.6% (111/233), with 59.7% (139/233) for tumour stage, and 70.4% (164/233) for nodal stage. The greatest dimension of the tumour on the CT image was about 2mm less than that measured by histopathology. The estimated volume of tumour was a quarter smaller. The accuracy of predicting malignant lymph nodes by CT was 68.9% (n=161). Among patients with stage II disease, simultaneous neck dissection was less likely in the understaged group than in the accurately staged one. The reoperation rate was a little higher but not significantly so. We conclude that the accuracy of CT in predicting local staging for tongue cancer was only moderate, because it underestimated the size of the tumour and needed to improve the criteria for detecting malignant lymph nodes. Understaging on CT images may influence the prognosis of patients with early stage tongue cancer.

[Meta analysis of clinical safety and efficacy evaluation of peripheral arterial disease patients with critical limb ischemia who accepted bypass surgery or endovascular therapy].

Objective: To evaluate the safety and efficacy of peripheral arterial disease patients with critical limb ischemia who accepted BSX or EVT. Methods: According to the requirements of systematic review, we searched MEDLINE (1980-2014), Emabse (1980-2014), Journals @ Ovid Full Text (1980-2014) databases, selected literature and extracted data, meta-analysis was performed through STATA11.2. Results: A total of 17 studies (3 RCTs, 14 non-randomized studies) and 5 515 patients (BSX group: 2 454, EVT group: 2 769) were deemed eligible. Meta-analysis showed there were no differences between the two groups in 30-day mortality (OR=1.110, P=0.523). However, BSX was significantly associated with increasing overall complications (OR=2.456, P=0.003), infections (OR=3.163, P<0.001) and thrombosis (OR=3.069, P=0.002), but showed a reduction in dissection and pseudoaneurysm (OR=0.537, P=0.012). During the follow-up, BSX had significant advantages. The 1-year and 3-year primary patency of BSX patients were higher than EVT group (1 year, OR=1.415, P=0.008; 3 years, OR=1.619, P<0.001); so were in secondary patency( 1 year, OR=2.156, P<0.001; 3 years, OR=2.547, P<0.001). Meanwhile, the 5-year overall survival rate was also higher in BSX group (OR=1.243, P=0.007). Conclusions: EVT has potential advantages in reducing surgical trauma and early postoperative complications, shortening hospital stay and so on. Concerning long-term results, BSX is better in reducing long-term mortality and improving long-term patencies than EVT group.

[Evaluation between bypass surgery and endovascular therapy to peripheral arterial disease patients with critical limb ischemia].

Bypass surgery(BSX) and endovascular therapy(EVT) are the most important therapeutic method to critical limb ischemia.EVT has potential advantages in reducing surgical trauma and early postoperative complications, shortening hospital stay and so on. Concerning long-term results, BSX is better in reducing long-term mortality and improving long-term patency than EVT group. Therefore, control indications reasonably and select individualized methods, avoid the abuse of EVT are more meaningful for patients.

Prevalence of Toxoplasma gondii in cats from Colombia, South America and genetic characterization of T. gondii isolates.

Cats are important in the epidemiology of Toxoplasma gondii infection because they are the only hosts that can excrete the environmentally-resistant oocysts. In the present study, prevalence of T. gondii was determined in serum, feces, and tissues of 170 unwanted cats from Colombia, South America. Antibodies to T. gondii were assayed by the modified agglutination test and found in 77 of 170 (45.2%) cats with titers of <1:5 in 93, 1:5 in eight, 1:10 in 17, 1:20 in 10, 1:40 in seven, 1:80 in four, 1:160 in eight, 1:320 in six, and 1:640 or higher in 17 cats. T. gondii oocysts were not found in feces of any cat as ascertained by bioassay in mice. Tissues (brain, heart, tongue) of 116 cats were bioassayed in mice or cats. T. gondii was isolated from tissues of 15 of the 42 cats with titers of 1:40 or higher and not from any of the 90 cats titers of 1:20 or lower. Of the 29 cats whose tissues were bioassayed individually, T. gondii was isolated from the tongues of nine, hearts of eight, and brains of five. Mice inoculated with tissues of 12 of 15 infected cats died of toxoplasmosis; with nine T. gondii isolates all infected mice died. Overall, 65 of 92 (70%) of T. gondii-infected mice died of toxoplasmosis. Genotyping of these 15 isolates using polymorphisms at the SAG1, SAG2, SAG3, BTUB, and GRA6 loci revealed that three isolates (TgCtCo1, 2, and 7) had Type I alleles and one isolate (TgCtCo8) had Type II allele at all five loci. Eleven isolates contained the combination of Type I and III alleles and were divided into three genotypes, with TgCtCo3,5,6,9,12,13 and 15 had alleles I, I, III, I and III, TgCtCo4,10,11 had alleles I, III, III, I and I, and TgCtCo14 had alleles I, III, III, III, and III, at loci SAG1, SAG2, SAG3, BTUB and GRA6, respectively. All infected mice from each group had identical genotype except one mouse infected with TgCtCo5 had a Type III allele at locus BTUB and a unique allele (u-1) at locus SAG1 indicating mixed infection for TgCtCo5, whereas the rest seven mice had a Type I alleles at both loci.

Characterization of Toxoplasma gondii isolates in free-range chickens from Chile, South America.

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 85 free-range chickens (Gallus domesticus) from Chile was determined. Antibodies to T. gondii were assayed by the modified agglutination test (MAT), and found in 47 of 85 (55.3.9%) chickens with titers of 1:5 in six, 1:10 in four, 1:20 in four 1: 40 in three, 1: 80 in nine, 1: 160 in four 1:320 in nine, and 1: 640 or higher in eight. Hearts and brains of 47 chickens with titers of 1:5 or higher were pooled for each chicken and bioassayed in mice. Tissues from 16 seronegative (MAT<1:5) chickens were pooled and fed to one T. gondii-free cat. Feces of the cat were examined for oocysts but none was found based on bioassay of fecal floats in mice. Hearts and brains from seven seronegative (<1:5) were pooled and bioassayed in mice; T. gondii was not isolated. T. gondii was isolated by bioassay in mice from 22 chickens with MAT titers of 1:20 or higher. Genotyping of these 22 isolates using polymorphisms at the loci SAG1, SAG2, SAG3, BTUB and GRA6 revealed three genotypes. Seventeen isolates had type II alleles and four isolates had type III alleles at all loci. One isolate contained the combination of type I and III alleles. This is the first report of genetic characterization of T. gondii isolates from Chile, South America.

Biologic and genetic characteristics of Toxoplasma gondii isolates in free-range chickens from Costa Rica, Central America.

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 144 free-range chickens (Gallus domesticus) from Costa Rica was determined. Antibodies to T. gondii were assayed by the modified agglutination test (MAT), and found in 60 (40.1%) of 144 chickens with titers of 1:5 in 16, 1:10 in 5, 1:20 in 2, 1:40 in 3, 1:80 in 5, and 1:160 or higher in 29. Tissues of all chickens were bioassayed for T. gondii in mice or cats. Hearts and brains of 52 chickens with titers of 1:5 or higher and 16 chickens with doubtful titers were pooled and bioassayed in mice. Tissues from 76 chickens with MAT titers of 1:10 or less were pooled and fed to three T. gondii-free cats. Fecal floats of cats were bioassayed orally in mice but were negative for T. gondii oocysts. T. gondii was isolated by bioassay in mice from 32 chickens with MAT titers of 1:10 or higher. All infected mice from 4 of the 32 isolates died of toxoplasmosis. Genotyping of these 32 isolates using polymorphisms at the loci SAG1, SAG2, SAG3, BTUB and GRA6 revealed five genotypes. Five isolates had type I alleles and one isolate had type III alleles at all loci. The rest 26 isolates contained the combination of type I and II or I and III alleles and were divided into three genotypes. None was found to have genotype II alleles at all five loci. This is the first report of genetic characterization of T. gondii isolates from Costa Rica, Central America.

Genetic and biologic characteristics of Toxoplasma gondii isolates in free-range chickens from Colombia, South America.

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 77 free-range chickens (Gallus domesticus) from Colombia, South America was determined. Antibodies to T. gondii were assayed by the modified agglutination test (MAT), and found in 32 (44.4%) of 72 chickens with titers of 1:5 in 4, 1:10 in 3, 1:20 in 1, 1:40 in 1, 1:80 in 8, 1:160 in 8, 1:320 in 3, and 1:640 or higher in 4. Hearts and brains of 31 seropositive chickens were pooled and bioassayed in mice. Tissues from 32 (16+16) seronegative chickens were pooled and fed to two, T. gondii-free cats, and tissues from nine chickens without matching sera were fed to one T. gondii-free cat. Feces of cats were examined for oocysts. T. gondii oocysts were excreted by a cat that was fed tissues of 16 seronegative chickens. T. gondii was isolated by bioassay in mice from 23 chickens with MAT titers of 1:20 or higher. All infected mice from 16 of the 23 isolates died of toxoplasmosis. Overall, 82 (81.1%) of 101 mice that became infected after inoculation with chicken tissues died of toxoplasmosis. Genotyping of these 24 isolates using polymorphisms at the SAG2 locus indicated that seven T. gondii isolates were Type I, 17 were Type III, and none was Type II. Phenotypically, T. gondii isolates from chickens from Colombia were similar to isolates from Brazil but different from the isolates from North America; most isolates from chickens from Brazil and Colombia were lethal for mice whereas isolates from North America did not kill inoculated mice. Genetically, none of the T. gondii isolates from Colombia and Brazil was SAG2 Type II, whereas most isolates from chickens from North America were Type II. This is the first report of genetic characterization of T. gondii isolates from Colombia, South America.

Isolation, tissue distribution, and molecular characterization of Toxoplasma gondii from chickens in Grenada, West Indies.

The prevalence of Toxoplasma gondii in free-range 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 102 free-range chickens (Gallus domesticus) from Grenada was determined. Antibodies to T. gondii were assayed by the modified agglutination test (MAT). Antibodies were found in 53 (52%) chickens with titers of 1:5 in 6, 1:10 in 4, 1:20 in 4, 1:40 in 4, 1:80 in 15, 1:160 in 9, 1: 320 in 5, 1:640 in 4, and 1:1,280 or greater in 2. Hearts, pectoral muscles, and brains of 43 seropositive chickens with MAT titers of 1:20 or greater were bioassayed individually in mice. Tissues of each of 10 chickens with titers of 1:5 and 1:10 were pooled and bioassayed in mice. Tissues from the remaining 49 seronegative chickens were pooled and fed to 4 T. gondii-free cats. Feces of cats were examined for oocysts; they did not shed oocysts. T. gondii was isolated from 35 of 43 chickens with MAT titers of 1:20 or greater; from the hearts, brains, and pectoral muscles of 2, hearts and brains of 20, from the hearts alone of 11, and brains alone of 2. T. gondii was isolated from 1 of 10 chickens with titers of 1:5 or 1:10. All 36 T. gondii isolates were avirulent for mice. Genotyping of these 36 isolates using polymorphisms at the SAG2 locus indicated that 29 were Type III, 5 were Type I, 1 was Type II, and 1 had both Type I and Type III. Genetically, the isolates from Grenada were different from those from the United States; Type II was the predominant type from the United States. Phenotypically, all isolates from Grenada were avirulent for mice, whereas those from Brazil were mouse-virulent. This is the first report of isolation of T. gondii from chickens from Grenada, West Indies.

Prevalence of viable Toxoplasma gondii in beef, chicken, and pork from retail meat stores in the United States: risk assessment to consumers.

The prevalence of viable Toxoplasma gondii was determined in 6,282 samples (2,094 each of beef, chicken, and pork) obtained from 698 retail meat stores from 28 major geographic areas of the United States. Each sample consisted of a minimum of 1 kg of meat purchased from the retail meat case. To detect viable T. gondii, meat samples were fed to T. gondii-free cats and feces of cats were examined for oocyst shedding. Initially, 100 g of meat from 6 individual samples of a given species were pooled (total, 600 g), fed to a cat over a period of 3 days, and feces were examined for oocysts for 14 days; the remaining meat samples were stored at 4 C for 14 days (until results of the initial cat fecal examination were known). When a cat fed pooled samples had shed oocysts, 6 individual meat samples from each pool were bioassayed for T. gondii in cats and mice. Toxoplasma gondii isolates were then genetically characterized using the SAG2 locus and 5 hypervariable microsatellite loci. In all, 7 cats fed pooled pork samples shed oocysts. Toxoplasma gondii oocysts were detected microscopically in the feces of 2 of the cats; 1 isolate was Type II and the second was Type III. Analyzed individually, T. gondii was detected by bioassay in 3 of the 12 associated samples with genetic data indicating T. gondii isolates present in 2. The remaining 5 pooled pork samples had so few oocysts that they were not initially detected by microscopic examination, but rather by mouse bioassay of cat feces. Two were Type I, 1 was Type II, and 2 were Type III. None of the cats fed chicken or beef samples shed oocysts. Overall, the prevalence of viable T. gondii in retail meat was very low. Nevertheless, consumers, especially pregnant women, should be aware that they can acquire T. gondii infection from ingestion of undercooked meat, and in particular, pork. Cooking meat to an internal temperature of 66 C kills T. gondii.

Tissue distribution and molecular characterization of chicken isolates of Toxoplasma gondii from Peru.

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 antibodies in sera of 50 free-range chickens (Gallus domesticus) from Peru was 26% on the basis of the modified agglutination test (MAT). Hearts, pectoral muscles, and brains of seropositive (MAT > or =1:5) chickens were bioassayed individually in mice. Tissues from the remaining 37 seronegative chickens were pooled and fed to 2 T. gondii-free cats. Feces of cats were examined for oocysts; they did not shed oocysts. Toxoplasma gondii was isolated from the hearts of 10 seropositive chickens but not from their brains and pectoral muscles. Genotyping of these isolates using the SAG2 locus indicated that 7 isolates were type I and 3 were type III. Six of the 7 type-I isolates were avirulent for mice, which was unusual because type-I isolates are considered virulent for mice. The T. gondii isolates were from chickens from different properties that were at least 200 m apart. Thus, each isolate is likely to be different. This is the first report of isolation of T. gondii from chickens from Peru.

Toxoplasma gondii infections in cats from Paraná, Brazil: seroprevalence, tissue distribution, and biologic and genetic characterization of isolates.

Cats are important in the epidemiology of Toxoplasma gondii because they are the only hosts that can excrete environmentally resistant oocysts. The prevalence of T. gondii was determined in 58 domestic cats from 51 homes from Santa Isabel do Ivai, Parana State, Brazil where a water-associated outbreak of acute toxoplasmosis had occurred in humans. Antibodies to T. gondii were found with the modified agglutination test in 49 of 58 (84.4%) cats at a serum dilution of 1:20. Tissues (brain, heart, and skeletal muscle) of 54 of these cats were bioassayed in T. gondii-free, laboratory-reared cats; T. gondii oocysts were excreted by 33 cats that were fed feline tissues. Brains from these 54 cats were bioassayed in mice; T. gondii was isolated from 7. Skeletal muscles and hearts of 15 cats were also bioassayed in mice; T. gondii was isolated from skeletal muscles of 9 and hearts of 13. The results indicate that T. gondii localizes in muscle tissue more than the brains of cats. In total there were 37 T. gondii isolates from 54 cats. Most isolates of T. gondii were virulent for mice. Genotyping of the 37 isolates of T. gondii, using the SAG2 locus, revealed that 15 isolates were type I and 22 were type III. The absence of type II genotype in cats in this study is consistent with the previous studies on T. gondii isolates from Brazil and is noteworthy because most T. gondii isolates from the United States are type II. These findings support the view that Brazilian and North American T. gondii isolates are genetically distinct. This is the first report of genotyping of T. gondii isolates from the domestic cat.

High prevalence of Toxoplasma gondii in a commercial flock of chickens in Israel, and public health implications of free-range farming.

Little is known of the prevalence of Toxoplasma gondii in commercially raised chickens. In the present study, the prevalence of T. gondii in 96 free-range chickens (Gallus domesticus) from a commercial farm in Israel was assessed. Blood, heart, and brain from each chicken were examined for T. gondii infection. Antibodies to T. gondii, assayed with the modified agglutination test (MAT > or = 1:5), were found in 45 of the 96 chickens. Hearts and brains of seropositive (MAT > or = 1:5) chickens were bioassayed in mice. Additionally, hearts and brains of 51 seronegative (MAT < 1:5) chickens were bioassayed in two T. gondii-free cats. T. gondii was isolated from 19 of the 45 (42.2%) seropositive chickens by bioassay in mice. Both the cats fed tissues pooled from seronegative chickens shed T. gondii oocysts. Tachyzoites and tissue cysts of all 21 isolates of T. gondii from chickens were avirulent for mice. Seventeen of the 19 isolates genotyped were found to be type II, and 2 were type III. Understanding of the sources of infection on such farms could be the key to the development of better prevention strategies.

Molecular and biologic characteristics of Toxoplasma gondii isolates from wildlife in the United States.

Toxoplasma gondii isolates can be grouped into 3 genetic lineages. Type I isolates are considered more virulent in outbred mice and have been isolated predominantly from clinical cases of human toxoplasmosis, whereas types II and III isolates are considered less virulent for mice and are found in humans and food animals. Little is known of genotypes of T. gondii isolates from wild animals. In the present report, genotypes of isolates of T. gondii from wildlife in the United States are described. Sera from wildlife were tested for antibodies to T. gondii with the modified agglutination test, and tissues from animals with titers of 1:25 (seropositive) were bioassayed in mice. Toxoplasma gondii was isolated from the hearts of 21 of 34 seropositive white-tailed deer (Odocoileus virginianus) from Mississippi and from 7 of 29 raccoons (Procyon lotor); 5 of 6 bobcats (Lynx rufus); and the gray fox (Urocyon cinereoargenteus), red fox (Vulpes vulpes), and coyote (Canis latrans) from Georgia. Toxoplasma gondii was also isolated from 7 of 10 seropositive black bears (Ursus americanus) from Pennsylvania by bioassay in cats. All 3 genotypes of T. gondii based on the SAG2 locus were circulating among wildlife.

Isolation and molecular characterization of Toxoplasma gondii from chickens and ducks from Egypt.

The prevalence of Toxoplasma gondii in free range chickens is a good indicator of the prevalence of T. gondii oocysts in the environment because chickens feed from the ground. In the present study, prevalence of T. gondii in 121 free range chickens (Gallus domesticus) and 19 ducks (Anas sp.) from a rural area surrounding Giza, Egypt was assessed. Blood, heart, and brain from each animal were examined for T. gondii infection. Antibodies to T. gondii, assayed with the modified agglutination test (MAT), were found in 49 (40.4%) chickens in titers of 1:5 in 11, 1:10 in four, 1:20 in four, 1:40 in eight, 1:80 in 10, and 1:160 or more in 12 chickens. Antibodies were found in three ducks each with a titer of 1:80. Hearts and brains of seropositive (MAT > or = 1:5) chickens and ducks were bioassayed in mice. Additionally, hearts and brains of seronegative (MAT<1:5) animals were bioassayed in T. gondii-free cats. T. gondii was isolated from 19 of 49 seropositive chickens (one with a titer of 1:5, two with a titer of 1:20, one with a titer of 1:40, five with a titer of 1:80, three with a titer of 1:160, and seven with a titer of > or = 1:360). One cat fed tissues pooled from 15 seronegative chickens shed T. gondii oocysts, while two cats fed tissues of 34 seronegative chickens did not shed oocysts. T. gondii was isolated from one of the seropositive ducks by bioassay in mice. The two cats fed tissues from 16 seronegative ducks did not shed oocysts. Genotyping of 20 chicken isolates of T. gondii using the SAG 2 locus indicated that 17 isolates were type III and three were type II. The duck isolate of T. gondii was type III. The mice inoculated with tissue stages of all 21 isolates of T. gondii from chickens and ducks remained asymptomatic, indicating that phenotypically they were not type I because type I strains are lethal for mice. Infections with mixed genotypes were not found.

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.

Establishment of Besnoitia darlingi from opossums (Didelphis virginiana) in experimental intermediate and definitive hosts, propagation in cell culture, and description of ultrastructural and genetic characteristics.

Besnoitia darlingi from naturally infected opossums (Didelphis virginiana) from Mississippi, USA, was propagated experimentally in mice, cats, and cell culture and was characterised according to ultrastructural, genetic, and life-history characteristics. Cats fed tissue cysts from opossums shed oocysts with a prepatent period of nine or 11 days. Oocysts, bradyzoites, or tachyzoites were infective to outbred and interferon-gamma gene knockout mice. Tachyzoites were successfully cultivated and maintained in vitro in bovine monocytes and African green monkey cells and revived after an 18-month storage in liquid nitrogen. Schizonts were seen in the small intestinal lamina propria of cats fed experimentally-infected mouse tissues. These schizonts measured up to 45 x 25 microm and contained many merozoites. A few schizonts were present in mesenteric lymph nodes and livers of cats fed tissue cysts. Ultrastructurally, tachyzoites and bradyzoites of B. darlingi were similar to other species of Besnoitia. A close relationship to B. besnoiti and an even closer relationship to B. jellisoni was indicated for B. darlingi on the basis of the small subunit and ITS-1 portions of nuclear ribosomal DNA.

Biological and genetic characterisation of Toxoplasma gondii isolates from chickens (Gallus domesticus) from São Paulo, Brazil: unexpected findings.

In spite of a wide host range and a world wide distribution, Toxoplasma gondii has a low genetic diversity. Most isolates of T. gondii can be grouped into two to three lineages. Type I strains are considered highly virulent in outbred laboratory mice, and have been isolated predominantly from clinical cases of human toxoplasmosis whereas types II and III strains are considered avirulent for mice. In the present study, 17 of 25 of the T. gondii isolates obtained from asymptomatic chickens from rural areas surrounding São Paulo, Brazil were type I. Antibodies to T. gondii were measured in 82 chicken sera by the modified agglutination test using whole formalin-preserved tachyzoites and mercaptoethanol and titres of 1:10 or more were found in 32 chickens. Twenty-two isolates of T. gondii were obtained by bioassay in mice inoculated with brains and hearts of 29 seropositive (> or =1:40) chickens and three isolates were obtained from the faeces of cats fed tissues from 52 chickens with no or low levels (<1:40) of antibodies. In total, 25 isolates of T. gondii were obtained by bioassay of 82 chicken tissues into mice and cats. All type I isolates killed all infected mice within 4 weeks whereas type III isolates were less virulent to mice. There were no type II strains. Tissue cysts were found in mice infected with all 25 isolates and all nine type I isolates produced oocysts. Infected chickens were from localities that were 18-200 km apart, indicating no common source for T. gondii isolates. This is the first report of isolation of predominantly type I strains of T. gondii from a food animal. Epidemiological implications of these findings are discussed.

Effects of high temperature and disinfectants on the viability of Sarcocystis neurona sporocysts.

The effect of moist heat and several disinfectants on Sarcocystis neurona sporocysts was investigated. Sporocysts (4 million) were suspended in water and heated to 50, 55, 60, 65, and 70 C for various times and were then bioassayed in interferon gamma gene knockout (KO) mice. Sporocysts heated to 50 C for 60 min and 55 C for 5 min were infective to KO mice, whereas sporocysts heated to 55 C for 15 min and 60 C or more for 1 min were rendered noninfective to mice. Treatment with bleach (10, 20, and 100%), 2% chlorhexidine, 1% betadine, 5% o-benzyl-p-chlorophenol, 12.56% phenol, 6% benzyl ammonium chloride, and 10% formalin was not effective in killing sporocysts. Treatment with undiluted ammonium hydroxide (29.5% ammonia) for 1 hr killed sporocysts, but treatment with a 10-fold dilution (2.95% ammonia) for 6 hr did not kill sporocysts. These data indicate that heat treatment is the most effective means of killing S. neurona sporocysts in the horse feed or in the environment.

The gamma interferon knockout mouse model for sarcocystis neurona: comparison of infectivity of sporocysts and merozoites and routes of inoculation.

The dose-related infectivity of Sarcocystis neurona sporocysts and merozoites of 2 recent isolates of S. neurona was compared in gamma interferon knockout (KO) mice. Tenfold dilutions of sporocysts or merozoites were bioassayed in mice, cell culture, or both. All 8 mice, fed 1,000 sporocysts, developed neurological signs with demonstrable S. neurona in their tissues. Of 24 mice fed low numbers of sporocysts (100, 10, 1), 18 became ill by 4 wk postinoculation, and S. neurona was demonstrated in their brains; antibodies (S. neurona agglutination test) to S. neurona and S. neurona parasites were not found in tissues of the 6 mice that were fed sporocysts and survived for >39 days. One thousand culture-derived merozoites of these 2 isolates were pathogenic to all 8 mice inoculated subcutaneously (s.c.). Of the 24 mice inoculated s.c. with merozoites numbering 100, 10, or 1, only 3 mice had demonstrable S. neurona infection; antibodies to S. neurona were not found in the 21 mice that had no demonstrable organisms. As few as 10 merozoites were infective for cell cultures. These results demonstrate that at least 1,000 merozoites are needed to cause disease in KO mice. Sarcocystis neurona sporocysts were infective to mice by the s.c. route.