Does toxoplasmosis cause DNA damage? An evaluation in isogenic mice under normal diet or dietary restriction

Toxoplasmosis is an anthropozoonotic widespread disease, caused by the coccidian protozoan parasite Toxoplasma gondii. Since there are no data regarding the genotoxicity of the parasite in vivo, this study was designed to evaluate the genotoxic potential of the toxoplasmosis on isogenic mice with normal diet or under dietary restriction and submitted to a treatment with sulfonamide (375 mug/kg per day). DNA damage was assessed in peripheral blood, liver and brain cells using the comet assay (tail moment). The results for leucocytes showed increases in the mean tail moment in mice under dietary restriction; in infected mice under normal diet; in infected, sulfonamide-treated mice under normal diet; in infected mice under dietary restriction and in infected sulfonamide-treated mice under dietary restriction. In liver and brain cells, no statistically significant difference was observed for the tail moment. These results indicated that dietary restriction and T. gondii were able to induce DNA damage in peripheral blood cells, as detected by the comet assay. (C) 2004 Elsevier B.V. All rights reserved.

Short- and long-term reproductive effects of prenatal and lactational growth restriction caused by maternal diabetes in male rats

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Genetic characterisation of Giardia duodenalis in dairy cattle in Brazil

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Taenia saginata: Differential diagnosis of human taeniasis by polymerase chain reaction-restriction fragment length polymorphism assay

Speciation of Taenia in human stool is important because of their different clinical and epidemiological features. DNA analysis has recently become possible which overcomes the problems of differentiating human taeniid cestodes morphologically. In the present study, we evaluated PCR coupled to restriction fragment length polymorphism to differentiate Taenia solium from Taenia saginata eggs present in fecal samples from naturally infected patients. A different Dral-RFLP pattern: a two-band pattern (421 and 100 bp) for T saginata and a three-band pattern (234, 188, and 99 bp) for T solium was observed allowing the two species to be separated.. The lower detection limit of the PCR-RFLP using a non-infected fecal sample prepared with a given number of T saginata eggs was 34 eggs in 2 g stool sediment. The 521 bp mtDNA fragment was detected in 8 out of 12 Taenia sp. carriers (66.6%). of these, three showed a T solium pattern and five a T saginata pattern. (c) 2005 Elsevier B.V. All rights reserved.

Comparative evaluation of enzyme-linked immunosorbent assay based on crude and purified antigen in the diagnosis of canine visceral leishmaniasis in symptomatic and oligosymptomatic dogs

This study evaluated the performance of crude total antigen (CTA) and fucose-mannose ligand antigen (FML) in an enzyme-linked immunosorbent assay for diagnosis of canine visceral leishmaniasis (CVL). The assays used sera from known negative controls (n = 30), clinically symptomatic (n = 30) and oligosymptomatic (n = 30) parasitologically proven infection (by microscopy). Aspirates of popliteal lymph node from infected canines were colleted to score parasitism and compared with the ELISA results. The study indicated that FML used in ELISA provided high sensitivity for detecting oligosymptomatic dogs (90%) and CTA showed greater sensitivity than FML for symptomatic canines (90%). In oligosymptomatic dogs, specificity was 100% for CTA-ELISA, but in symptomatic dogs, FML specificity was higher (96.7%) than CTA-ELISA (93.3%). A significant correlation was observed between the degree of parasitism and the results obtained in CTA-ELISA. Since no available antigen offers 100% specificity and sensitivity for CVL diagnosis, the choice of antigen used must depend on the aim of the investigation. (C) 2008 Elsevier B.V. All rights reserved.

Serological diagnosis of visceral leishmaniasis by an enzyme immunoassay using protein A in naturally infected dogs

Um ensaio de imunoadsorção enzimática para detecção de anticorpos contra Leishmania chagasi, utilizando antígeno total de formas promastigotas lisados foi desenvolvido. Cinqüenta cães com sintomas clínicos de leishmaniose visceral foram examinados. Esta técnica utilizou anti-IgG de cão conjugado a peroxidase ou proteína A conjugado a peroxidase. Foi verificado que nos animais positivos diagnosticados por exame parasitológico direto o ensaio ELISA utilizando proteína A conjugada a peroxidase (média da densidade óptica ± desvio padrão 2,078 ± 0,631) detecta mais anticorpos do que o sistema utilizando anti-IgG de cão conjugado a peroxidase (média da densidade óptica ± desvio padrão 1,008 ± 0,437), enquanto para os animais negativos o resultado obtido nos dois sistemas de detecção são similares. Esse resultado sugere que o sistema de ELISA utilizando proteína A conjugado a peroxidase pode ser útil na detecção de animais na fase aguda da infecção e desta forma auxiliar na identificação dos animais positivos e no controle desta importante zoonose.

Distribution and hydrolytic enzyme characteristics of Candida albicans strains isolated from diabetic patients and their non-diabetic consorts

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Disparity between Multilocus Enzyme Electrophoresis, Microsatellite Markers and Pulsed-Field Gel Electrophoresis in epidemiological tracking of Candida albicans

Various molecular systems are available for epidemiological, genetic, evolutionary, taxonomic and systematic studies of innumerable fungal infections, especially those caused by the opportunistic pathogen C. albicans. A total of 75 independent oral isolates were selected in order to compare Multilocus Enzyme Electrophoresis (MLEE), Electrophoretic Karyotyping (EK) and Microsatellite Markers (Simple Sequence Repeats - SSRs), in their abilities to differentiate and group C. albicans isolates (discriminatory power), and also, to evaluate the concordance and similarity of the groups of strains determined by cluster analysis for each fingerprinting method. Isoenzyme typing was performed using eleven enzyme systems: Adh, Sdh, M1p, Mdh, Idh, Gdh, G6pdh, Asd, Cat, Po, and Lap (data previously published). The EK method consisted of chromosomal DNA separation by pulsed-field gel electrophoresis using a CHEF system. The microsatellite markers were investigated by PCR using three polymorphic loci: EF3, CDC3, and HIS3. Dendrograms were generated by the SAHN method and UPGMA algorithm based on similarity matrices (S(SM)). The discriminatory power of the three methods was over 95%, however a paired analysis among them showed a parity of 19.7-22.4% in the identification of strains. Weak correlation was also observed among the genetic similarity matrices (S(SM)(MLEE) x S(SM)(EK) x S(SM)(SSRs)). Clustering analyses showed a mean of 9 +/- 12.4 isolates per cluster (3.8 +/- 8 isolates/taxon) for MLEE, 6.2 +/- 4.9 isolates per cluster (4 +/- 4.5 isolates/taxon) for SSRs, and 4.1 +/- 2.3 isolates per cluster (2.6 +/- 2.3 isolates/taxon) for EK. A total of 45 (13%), 39(11.2%), 5 (1.4%) and 3 (0.9%) clusters pairs from 347 showed similarity (Si) of 0.1-10%, 10.1-20%, 20.1-30% and 30.1-40%, respectively. Clinical and molecular epidemiological correlation involving the opportunistic pathogen C. albicans may be attributed dependently of each method of genotyping (i.e., MLEE, EK, and SSRs) supplemented with similarity and grouping analysis. Therefore, the use of genotyping systems that give results which offer minimum disparity, or the combination of the results of these systems, can provide greater security and consistency in the determination of strains and their genetic relationships. (C) 2010 Elsevier B.V. All rights reserved.

Myocardial dysfunction induced by food restriction is related to morphological damage in normotensive middle-aged rats

Previous works from our laboratory have revealed that food restriction (FR) promotes discrete myocardial dysfunction in young rats. We examined the effects of FR on cardiac function, in vivo and in vitro, and ultrastructural changes in the heart of middle-aged rats. Twelve-month-old Wistar- Kyoto rats were fed a control (C) or restricted diet (daily intake reduced to 50% of the control group) for 90 days. Cardiac performance was studied by echocardiogram and in isolated left ventricular (LV) papillary muscle by isometric contraction in basal condition, after calcium chloride (5.2 mM) and beta- adrenergic stimulation with isoproterenol (10(-6) M). FR did not change left ventricular function, but increased time to peak tension, and decreased maximum rate of papillary muscle tension development. Inotropic maneuvers promoted similar effects in both groups. Ultrastructural alterations were seen in most FR rat muscle fibers and included, absence and/or disorganization of myofilaments and Z line, hyper-contracted myofibrils, polymorphic and swollen mitochondria with disorganized cristae, and a great quantity of collagen fibrils. In conclusion, cardiac muscle sensitivity to isoproterenol and elevation of extracellular calcium concentration is preserved in middle-aged FR rats. The intrinsic muscle performance depression might be related to morphological damage.

Exercise training increases myocardial inotropic response in food restricted rats

This study evaluated the effects of exercise training on myocardial function and ultrastructure of rats submitted to different levels of food restriction (FR). Male Wistar-Kyoto rats, 60 days old, were submitted to free access to food, light FR (20%), severe FR (50%) and/or to swimming training (one hour per day with 5% of load, five days per week for 90 days). Myocardial function was evaluated by left ventricular papillary muscle under basal condition (calcium 1.25 mM), and after extracellular calcium elevation to 5.2 mM and isoproterenol (I PM) addition. The ultrastructure of the myocardium was examined in the papillary muscle. The training effectiveness was verified by improvement of myocardial metabolic enzyme activities. Both 20% and 50% food restriction protocols presented minor body and ventricular weights gain. The 20%-FR, in sedentary or trained rats, did not alter myocardial function or ultrastructure. The 50%-FR, in sedentary rats, caused myocardial dysfunction under basal condition, decreased response to inotropic stimulation, and promoted myocardial ultrastructural damage. The 50%-FR, in exercised rats, increased myocardial dysfunction under basal condition but increased response to inotropic stimulation although there was myocardial ultrastructural damage. In conclusion, the exercise training in severe restriction caused marked myocardial dysfunction at basal condition but increased myocardial response to inotropic stimulation. (c) 2005 Elsevier B.V.. All rights reserved.

Myocardial dysfunction induced by food restriction is related to calcium cycling and beta-adrenergic system changes

Food restriction (FR) has been shown to promote myocardial dysfunction in rats. The aim of this study was to verify the participation of calcium and beta-adrenergic system on myocardial mechanical alteration in rats submitted to FR. Myocardial performance was studied in isolated left ventricular papillar muscle from young Wistar-Kyoto rats (WKY) submitted to FR or to control diet. The groups subjected to FR were fed 50% less food than the control group for 90 days. Mechanical function was studied in isometric contraction at post-rest contraction of 30 seconds (PRC), calcium chloride concentration 5.20 mM, and beta-adrenergic stimulation with isoproterenol 10(-6) M. FR decreased the body weight, and left and right ventricular weight. In basal condition (1.25 MM of calcium) time to peak tension (TPT) and time from peak tension to 50% relaxation (RT50) were greater in the FR group. Muscle function was. The same in both PRC groups. TPT decrease in both high calcium groups, more in FR rats; RT50 dropped only in FR animals. TPT decreased in both Isoproterenol groups, more intensely in the FR group. This result suggests that food restriction impairs myocardial performance and these changes may be attributed to alterations in the intracellular calcium cycling and beta-adrenergic system. (C) 2003 Elsevier B.V. All rights reserved.

Food restriction impairs myocardial inotropic response to calcium and beta-adrenergic stimulation in spontaneously hypertensive rats

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Severe food restriction induces myocardial dysfunction related to SERCA2 activity

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)