Genetic diversity and maternal origin of Northeast African and South American donkey populations.

To investigate the mtDNA variation and origin of maternal lineages in South American donkeys and to reassess the domestication of donkeys in northeast Africa, we analyzed sequences (489 bp of the D-loop) from 323 domestic donkeys sampled from Peru, Brazil, Ethiopia and Egypt. Altogether, the 323 sequences displayed 53 different haplotypes (45 in Ethiopia, 14 in Egypt, eight in Peru and six in Brazil). Among the four populations, Egyptian donkeys possessed the highest haplotype diversity (0.910 ± 0.032), followed by Brazilian donkeys (0.879 ± 0.060). The Clade I haplotypes dominated in Peruvian donkeys (65%), whereas Clade II haplotypes dominated in Brazilian donkeys (67%). Estimates of FST values showed a high genetic differentiation between Peruvian and Brazilian donkey populations (FST  = 0.4066), which could be explained by the complex introduction history of South American donkeys. Phylogeographic analysis indicates that northeast Africa could be the most probable domestication center for Clade I donkeys. Analysis of molecular variance confirmed a weak genetic structure in domestic donkey populations among four continents (Europe, Asia, Africa and South America).

Genetic diversity and paternal origin of domestic donkeys.

Numerous studies have been conducted to investigate genetic diversity, origins and domestication of donkey using autosomal microsatellites and the mitochondrial genome, whereas the male-specific region of the Y chromosome of modern donkeys is largely uncharacterized. In the current study, 14 published equine Y chromosome-specific microsatellites (Y-STR) were investigated in 395 male donkey samples from China, Egypt, Spain and Peru using fluorescent labeled microsatellite markers. The results showed that seven Y-STRs-EcaYP9, EcaYM2, EcaYE2, EcaYE3, EcaYNO1, EcaYNO2 and EcaYNO4-were male specific and polymorphic, showing two to eight alleles in the donkeys studied. A total of 21 haplotypes corresponding to three haplogroups were identified, indicating three independent patrilines in domestic donkey. These markers are useful for the study the Y-chromosome diversity and population genetics of donkeys in Africa, Europe, South America and China.

Expression of the stem cell factor in fibroblasts, endothelial cells, and macrophages in periapical tissues in human chronic periapical diseases.

Stem cell factor (SCF), an important stem cell cytokine, has multiple functions. Fibroblasts (FBs), mature mast cells, endothelial cells (ECs), and eosinophil granulocytes can produce SCF in the inflammatory process. Therefore, we aimed to observe SCF expression in FBs, ECs, and macrophages (MPs) in periapical tissues in human chronic periapical disease and investigate the effects of cells expressing SCF in pathogenesis of the disease. Healthy (N = 20), periapical cyst (N = 15), and periapical granuloma (N = 15) tissues were fixed in 10% formalin for 48 h, embedded in paraffin, and stained with hematoxylin and eosin to observe histological changes. SCF expression was observed in FBs, ECs, and MPs in periapical tissues by double immunofluorescence. CD334, CD31, and CD14 are specific markers of FBs, ECs, and MPs, respectively. Results showed that densities of CD334-SCF double-positive FBs, CD31-SCF double-positive ECs, and CD14-SCF double-positive MPs were significantly increased in periapical tissue groups (P < 0.01). There were no significant differences in CD334-SCF double-positive FB and CD31-SCF double-positive EC levels between the two periapical tissue groups (P > 0.05). CD14-SCF double-positive MP density was considerably higher in periapical granulomas than in cysts (P < 0.01). FB, EC, and MP levels were significantly high and densities of CD334-SCF double-positive FBs, CD31-SCF double-positive ECs, and CD14-SCF double-positive MPs improved considerably in chronic periapical tissues, suggesting that the cells might be related to occurrence, development, and pathogenesis of chronic periapical disease.

LncRNA SNHG12 promotes cell growth and inhibits cell apoptosis in colorectal cancer cells.

Several long non-coding RNA (lncRNA) might be correlated with the prognosis of colorectal cancer (CRC) and serve as a diagnostic and prognostic biomarker. However, the exact expression pattern of small nucleolar RNA host gene 12 (SNHG12) in colorectal cancer and its clinical significance remains unclear. The level of SNHG12 was detected by qRT-PCR in CRC tissues and CRC cells. MTT assay and colony formation assay were performed to examine the cell proliferation of CRC cells transfected with pcDNA-SNHG12 or si-SNHG12. Flow cytometry technology was used to detect cell cycle and cell apoptosis of CRC cells transfected with pcDNA-SNHG12 or si-SNHG12. The protein level of cell cycle progression-related molecules, including cyclin-dependent kinases (CDK4, CDK6), cyclin D1 (CCND1) and cell apoptosis-related molecule caspase 3 was detected by western blot. The effect of SNHG12 knockdown was examined in vivo. Increased levels of SNHG12 were observed in CRC tissues and in CRC cells. SNHG12 promoted the cell proliferation of CRC cells. In addition, SNHG12 overexpression boosted the cell cycle progression of SW480 cells transfected with pcDNA-SNHG12 and SNHG12 knockdown inhibited the cell cycle progression of HT29 cells transfected with si-SNHG12. SNHG12 also inhibited the cell apoptosis of CRC cells. We also found that SNHG12 increased the expression of cell cycle-related proteins and suppressed the expression of caspase 3. Our results suggest that SNHG12 promoted cell growth and inhibited cell apoptosis in CRC cells, indicating that SNHG12 might be a useful biomarker for colorectal cancer.

MicroRNA-25 expression level is an independent prognostic factor in epithelial ovarian cancer.

OBJECTIVES: The present study aimed to investigate the expression level of MicroRNA-25 (miR-25) in epithelial ovarian cancer (EOC) tissue, and examine its relationship with clinicopathological factors and prognosis of patients with EOC. METHODS: Expression levels of miR-25 in 86 pairs of EOC tissue and adjacent normal tissue were measured by quantitative real-time reverse-transcription polymerase chain reaction (qRT-PCR). The comparison of the expression level of miR-25 between EOC tissue and adjacent normal tissue was performed using the two-sample Student's t test. The correlation between the expression of miR-25 and clinicopathological characters was assessed with the two-sample Student's t test. The overall survival was analyzed by log-rank test, and survival curves were plotted according to Kaplan-Meier. RESULTS: The expression level of miR-25 in EOC tissue was significantly higher than in adjacent normal tissue. The miR-25 expression level was significantly positively correlated with tumor stage, histology, and regional lymph node involvement (P < 0.05). Kaplan-Meier analysis showed that patients with higher levels of miR-25 had significantly poorer survival than those with lower expression of this miRNA in patients, with a 6-year overall survival of 15.96 and 45.89 %, respectively, (P = 0.001). In the multivariate Cox proportional hazards analysis, high miR-25 expression was independently associated with poor survival (P = 0.002; HR = 2.119; 95 % CI = 1.568-3.221). CONCLUSION: The increased expression of miR-25 is closely related to poor prognosis of EOC, indicating that miR-25 may serve as a predictive biomarker for the prognosis of EOC.

Effects of common polymorphisms rs2910164 in miR-146a and rs11614913 in miR-196a2 on susceptibility to colorectal cancer: a systematic review meta-analysis.

PURPOSE: Emerging evidence has shown that single nucleotide polymorphisms occurred in microRNAs may contribute to the development of colorectal cancer (CRC). rs2910164 in miR-146a and rs11614913 in miR-196a2 are suggested to be associated with the susceptibility to CRC, but individually published studies revealed inconclusive results. To systematically summarize the possible correlationship between these polymorphisms and CRC risk, we performed this meta-analysis. METHODS: We retrieved the relevant articles of the associations between these two microRNA polymorphisms and susceptibility to CRC for the period up to July 1, 2013. A total of seven articles were identified with 2,143 cases and 2,457 controls for miR-146a rs2910164, 1,594 cases and 2,252 controls for miR-196a2 rs11614913. Odds ratio and 95 % confidence interval were calculated to investigate the strength of the association. RESULTS: The pooled analysis showed that miR-146a rs2910164 did not reveal any correlation with CRC susceptibility. However, a decreased risk was observed between miR-196a2 rs11614913 and CRC in all genetic models. CONCLUSION: Our current meta-analysis demonstrates that miR-196a2 rs11614913 most likely contributes to decreased risk of CRC, whereas miR-146a rs2910164 may not be associated with the susceptibility to CRC.

Changes of gene expression profiles across different phases of vascular calcification in rats.

This study investigated the alteration of gene expression profiles in order to gain a deeper understanding into the molecular mechanism involved in different processes of vascular calcification (VC). Sprague Dawley (SD) rats were injected with 300,000 µg/kg vitamin D3 and gavaged with 25 mg/kg nicotine for 8 or 16 weeks to create 8- and 16-week VC calcification groups. Histological analysis and quantification of aortic calcium content were used to determine the severity of vascular calcification. The suppression subtractive hybridization (SSH) method was employed to screen for up and downregulated genes in early and later phases of vascular calcification. Changes in calcium and phosphorus levels in tissue were used as markers of vascular calcification. Quantification of aortic calcium content revealed that vascular calcification might regress over time. In the early phase of vascular calcification, many calcification-promoting genes were upregulated, including ossification, oxidation, and inflammatory genes. In contrast, in later phase of vascular calcification, various calcification-inhibitor genes were highly expressed, including pyrophosphoric acid synthesis genes, glutamate signal peptide-related, reduction activity, and apoptosis regulation genes. The relatively higher expression of calcification-inhibitor genes compared to that of calcification-promoting genes might explain the genetic mechanism leading to the regression of vascular calcification. Therefore, this study provides a genomic basis to facilitate understanding of the molecular mechanism underlying vascular calcification regression.

Altered expression of piRNAs and their relation with clinicopathologic features of breast cancer.

INTRODUCTION: PIWI interacting RNAs (piRNAs) are endogenous regulatory non-coding RNAs which are generally expressed in the germline across animal species. The expressions of piRNAs in tumor tissue were rarely reported until now. The aim of this study was to investigate the expression of piRNAs in breast cancer. MATERIALS AND METHODS: Deep sequencing was carried out in four tumor and four matched non-tumor tissues from resected breast cancer cases to screen out differentially expressed piRNAs. Furthermore, the result was evaluated by real-time reverse transcription-polymerase chain reaction (RT-PCR) technology in 50 patients with breast cancer and the piRNA expression levels were correlated with clinicopathologic features. RESULTS: Through deep sequencing, six differentially expressed piRNAs were screened out (5 up- and 1 down-regulated) in the four tumor tissues compared with matched non-tumor tissues. Among them, 4 piRNAs (piR-4987, piR-20365, piR-20485 and piR-20582) were confirmed to be up-regulated by realtime RT-PCR in 50 breast cancer (P < 0.001). Moreover, up-regulated piR-4987 expression was associated with lymph node positivity (P = 0.001). CONCLUSIONS: PiRNAs might play a role in breast cancer and act as tumor markers.

Morphological variation of mutant sunflowers (Helianthus annuus) induced by space flight and their genetic background detection by SSR primers.

After sunflower seeds were exposed to space conditions, various mutant plants were screened from the descendent plants. The morphological characters of plants changed in flower color from golden to yellow, light yellow, or even to yellowish green. The ligulate petals of the unisexual floret broadened, or became thin, while the short tubular petals of bisexual floret elongated to some extent, or even turned into semi-ligulate petals or ligulate petals, making the phenotype of the whole inflorescence like a chrysanthemum. The shape and thickness of leaves varied in some of these plants. Absolute sterile plants in mutant plants were found to possess neither normal bisexual florets nor unisexual florets, but the "pseudo-floret" only consisted of pieces of shield-like bracts on protuberant floral disc. Thirty-five pairs of simple sequence of repeat primers were used to detect the genetic variation of the plants, and the results showed that only a variation was tested in the mutant plants from 4 primers. The different PCR products obtained were extracted for sequencing and alignment analysis, and the aligned results showed that the DNA sequence changed by deletion, insertion and replacement that occurred at some sites. The results proved the high mutagenic efficacy of space flight, and ways of DNA transformation due to space conditions.

Longitudinal development of antisocial behaviors in young and early adolescent Puerto Rican children at two sites.

OBJECTIVE: This report provides descriptive longitudinal findings over three waves of a study designed to assess the development of antisocial behaviors in young and early adolescent Puerto Rican children at two sites. METHOD: Through the use of standard assessment measures, representative samples of Puerto Rican children of both genders 5 to 13 years of age and their adult caretakers were interviewed at two sites: the South Bronx in New York City (n = 1,138) and the Standard Metropolitan Areas in Puerto Rico (n = 1,353; N = 2,491). RESULTS: Although no differences in prevalence between the two sites were apparent at baseline, analyses of the longitudinal data show that site differences emerge over time, with a decrease in risk of antisocial behavior over time in the Standard Metropolitan Areas relative to the Bronx. CONCLUSIONS: The decreased risk of these disorders in the Standard Metropolitan Areas corroborates the low rates in Puerto Rico reported in previous research. Future analyses of these data are needed to identify the risk and protective factors associated with this difference.

A study of disruptive behavior disorders in Puerto Rican youth: II. Baseline prevalence, comorbidity, and correlates in two sites.

OBJECTIVE: This is the second of two associated articles. The prevalence, correlates, and comorbidities of disruptive behavior disorders (DBDs) in two populations are reported. METHOD: Probability community samples of Puerto Rican boys and girls ages 5-13 years in San Juan, and the south Bronx in New York City are included (n = 2,491). The Diagnostic Interview Schedule for Children-IV and measures of correlates were employed to look at the association between DBDs and potential correlates, taking comorbidity into account. Data presented in this report were collected primarily between 2002 and 2003 but spanned a 3-year period from August 2000 to August 2003. RESULTS: There were no significant age or site differences among males in rates of DBDs, but rates among females increased with age in the south Bronx and decreased with age in Puerto Rico. The salient comorbidity of DBDs was with attention-deficit/hyperactivity disorder. Multiple regression showed lack of parental warmth and approval, poor peer relationships, and parental report of aggressive behavior during the toddler years to be the most significant correlates of DBDs in this population. CONCLUSIONS: Cultural factors, such as level of acculturation, were not associated with DBDs. The results suggest that clinical and preventive efforts need to emphasize interpersonal factors such as parent-child relationships and peer interactions.

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.

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.

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.

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.