Polyparasitism and zoonotic parasites in dogs from a rural area of the Argentine Chaco.

Dogs play an important role as reservoirs and hosts of multiple pathogens shared with humans and wildlife, which contribute significantly to the global burden of disease. Here, we assessed the occurrence of a broad range of zoonotic and non-zoonotic parasites in dogs from a rural area in the humid Chaco; determined the occurrence of polyparasitism; and explored its association with selected risk factors. In total, 212 dogs were examined serologically to determine Trypanosoma cruzi infection and 152 of them also were examined for Ehrlichia canis, Borrelia bugderfori, Anaplasma phagocitophylum, Dirofilaria immitis and Toxoplasma gondii. Fecal samples from 85 dogs were examined for intestinal parasites. Seventeen parasite species were seen, 77% of which are zoonotic. The most prevalent parasites were Ancylostoma caninum (68.2%), T. gondii (55.3%, first report for dogs in Argentina), Giardia sp. (25.9%), Cryptosporidium sp. (20.0%), T. cruzi (16.5%), trematodes (15.3%) and Toxocara canis (14.1%). Polyparasitism was found in 96% of the dogs, with up to six parasite species in a single dog, and was significantly associated with age of dog but not with host body condition or sex. The most frequent pair of parasites found together were T. gondii-A. caninum (46%), A. caninum-T. cruzi (34%) and T. gondii-T. cruzi (27%). The prevalence of anemia and leukocytosis was significantly higher in dogs showing the worst body condition. Our findings likely reflect structural poverty, poor sanitation and lack of a safe water supply. Importantly, many of the prevalent parasites seen are threats to human health. 243 words.

Is the infectiousness of dogs naturally infected with Trypanosoma cruzi associated with poly-parasitism?

Interactions among different species of parasites co-infecting the same host could be synergistic or antagonistic. These interactions may modify both the frequency of infected hosts and their infectiousness, and therefore impact on transmission dynamics. This study determined the infectiousness of Trypanosoma cruzi-seropositive dogs (using xenodiagnosis) and their parasite load (quantified by qPCR), and tested the association between both variables and the presence of concomitant endoparasites. A cross-sectional serosurvey conducted in eight rural villages from Pampa del Indio and neighboring municipalities (northeastern Argentina) detected 32 T. cruzi-seropositive dogs out of 217 individuals examined for infection. Both the infectiousness to the vector Triatoma infestans and parasite load of T. cruzi-seropositive dogs examined were heterogeneous. A statistically significant, nine-fold higher mean infectiousness was registered in T. cruzi-seropositive dogs co-infected with Ancylostoma caninum and a trematode than in T. cruzi-seropositive dogs without these infections. The median parasite load of T. cruzi was also significantly higher in dogs co-infected with these helminths. An opposite trend was observed in T. cruzi-seropositive dogs that were serologically positive to Toxoplasma gondii or Neospora caninum relative to dogs seronegative for these parasites. Using multiple logistic regression analysis with random effects, we found a positive and significant association between the infectiousness of T. cruzi-seropositive dogs and co-infections with A. caninum and a trematode. Our results suggest that co-infections may be a modifier of host infectiousness in dogs naturally infected with T. cruzi.

The dynamics of erythrocyte infection in bovine anaplasmosis: a flow cytometry-based analysis.

Anaplasma marginale (A. marginale) is an obligate intracellular bacterium that infects bovine erythrocytes causing extravascular hemolysis and anemia. In the present work, we combine SYTO16 labeling of parasitized cells with the statistical power of flow cytometry to study the evolution of erythrocyte infection during bovine anaplasmosis.

Disturbance of cellular iron uptake and utilisation by aluminium.

Aluminium (Al) affects erythropoiesis but the real mechanism of action is still unknown. Transferrin receptors (TfR) in K562 cells are able to bind Tf, when carrying either iron (Fe) or Al, with similar affinity. Then, the aim of this work was to determine whether Al could interfere with the cellular Fe uptake and utilisation. K562 cells were induced to erythroid differentiation by either haemin (H) or sodium butyrate (B) and cultured with and without Al. The effect of Al on cellular Fe uptake, Fe incorporation to haem and cell differentiation was studied. H- and B-stimulated cells grown in the presence of 10 microM Al showed a reduction in the number of haemoglobinised cells (by 18% and 56%, respectively) and high amounts of Al content. Al(2)Tf inhibited both the (59)Fe cellular uptake and its utilisation for haem synthesis. The removal of Al during the (59)Fe pulse, after a previous incubation with the metal, allowed the cells to acquire Fe quantities in the normal range or even exceeding the amounts incorporated by the respective control cells. However, the Fe incorporated to haem could not reach control values in B-stimulated cells despite enough Fe acquisition was observed after removing Al. Present results suggest that Al might exert either reversible or irreversible effects on the haemoglobin synthesis depending on cellular conditions.

Dissimilar behavior of lymph cells in response to the action of aluminium. In vitro and in vivo studies.

In order to detect possible immunological effects of aluminum (Al) on lymph cells, mice were orally overloaded with pharmacological doses for 22 weeks. The in vitro response of lymph cells to mitogens (phytohemmaglutinin, PHA and concanavaline A, Con A) was examined at the end of the treatment. The chronic ingestion of Al affected the lymphatic nodes that were found to be 2- to 10-fold larger than those of the control mice. Concurrently, the in vitro proliferation of lymphatic node cells was found enhanced, while spleen cell cultures were unaffected. An acute direct action of Al on lymph cells from different sources was also examined. The blastogenic response to PHA of human peripheral lymphocytes was not disturbed by the presence of Al concentrations ranging from 0.09 to 900 microM. However, the response of mouse lymph cells was quite different, given that an Al dose-dependent inhibition was observed for lymphatic node cells, whereas for spleen cells the inhibition was only detected at Al concentrations higher than 90 microM. This work shows that Al might induce alterations in cell immune responses. The opposite results observed in mouse lymphatic node cells after in vitro and in vivo Al treatment, let us suggest that either the stimulating or suppressing effects of Al on the immune system might depend on the dose, route of administration and length of exposure, as well as on the cell population assayed.

Interference of aluminium on iron metabolism in erythroleukaemia K562 cells.

It has been suggested that aluminium (Al) has a deleterious effect on erythropoiesis. However, there is still uncertainty as to its action mechanism. The present work was designed to determine how Al could affect the iron (Fe) metabolism in the human erythroleukaemia cell line K562. These cells, that express surface transferrin receptors (TfRs), were induced to erythroid differentiation by either haemin or hydroxyurea in 72 h cultures in media containing apotransferrin (apoTf). In the presence of aluminium citrate, the number of benzidine-positive cells decreased 18% when the cultures were induced by haemin, and 30% when hydroxyurea was the inducer. Cell viability was always unaffected. From competition assays, surface binding of 125I-Tf-Fe2 was found to be inversely related (p < 0.05) to Tf-Al2 concentration (from 2.5 to 10 nM). The dissociation constants (Kd) of the binding reaction between TfRs and the ligands Tf-Fe2 and Tf-Al2 were calculated. Kd values of the same order of magnitude demonstrated that TfR has a similar affinity for Tf-Fe2 (Kd = 1.75 x 10(-9) M) and Tf-Al2 (Kd = 1.37 x 10(-9) M). The number of surface TfRs, measured by kinetic 125I-Tf-Fe2 binding assays, was higher in induced cells cultured in the presence of Al. Nevertheless, in spite of the inhibition of cell haemoglobinization observed, 59Fe incorporation values were not different from those measured in control cultures for 72 h. As a consequence, it can be suggested that cellular Fe utilisation, and not Fe uptake, might be the main metabolic pathway impaired by Al.

Morphologic and functional alterations of erythroid cells induced by long-term ingestion of aluminium.

Anaemia has been associated with aluminium (Al) accumulation in plasma and/or bone tissue in patients with chronic renal insufficiency. Nevertheless, in previous works, we have found shortened red-cell life span, increased osmotic resistance and inhibition of colony-forming units-erythroid (CFU-E) development in Al-overloaded rats with normal renal function. To elucidate further the action of Al on in vivo erythropoiesis, aluminium citrate was provided to Sprague Dawley rats (n = 18) in the drinking water for 8 months. Significant decreases in haematocrit (38.8 +/- 4.29 versus 43.1 +/- 3.58%, p < 0.05) and blood haemoglobin concentration (137 +/- 10.1 versus 148 +/- 8.5 g/l, p < 0.05), reticulocytosis (1.8/1.3-4.2 versus 1.2/0.4-3.7%, p < 0.05), and severe inhibition of CFU-E growth (670/120-950 versus 1530/810-2440 CFU-E/2 x 10(5) cells, p < 0.005) were found. Anysocytosis, poikilocytosis and schistocytosis were detected in peripheral blood stained films. Scanning electron microscopy revealed the presence of erythrocytes with abnormal shape, including crenated and target cells. Aluminium was localised specially inside the schistocytes by EDAX analysis. Decreased haptoglobin concentration (107/83-127 versus 139/89-169 mg/l, p < 0.05) supports the assumption of haemolytic nature of the anaemia. Rats were not iron depleted, as plasma iron concentration and total iron binding capacity were found in the range of control values, and sideroblasts and haemosiderin deposits were observed in bone marrow smears. Total 59Fe uptake and 59Fe incorporated to haem by the bone marrow cells were found decreased. In conclusion, the erythropoiesis impairment induced by Al may be a combined effect of direct action on circulating erythrocytes and interference with the cellular iron metabolism in erythroid progenitors.

Oral aluminum administration to rats wih normal renal function. 1. Impairment of erythropoiesis.

Aluminum (Al) toxicity has been mainly investigated in uremic patients although healthy subjects and patients without renal insufficiency are not exempt from its potential deleterious effects. This experimental study aims to elucidate the action of different doses of Al citrate on in vivo erythropoiesis and find out whether the metal exerts a local toxic effect upon the bone marrow late erythroid progenitor cells. The groups in the first experimental series were: C1 (n=5) controls and TAl-1 (n=5) rats receiving 1 micromol Al citrate/g body weight/day by gavage. Colony-forming units-erythroid (CFU-E) development was inhibited in the TAl-1 group, but the median osmotic fragility (MOF) and hematocrit (Ht) values were similar to those of the C1 group. The groups in the second series were C2 (n=5) controls and TAl-2 (n=5) rats receiving Al citrate in drinking water (100 mmol/l). The TAl-2 group showed decreased Ht, hemoglobin concentration, MOF and red blood-cell life-span values (P<0.05), and a marked inhibition of the CFU-E development (P<0.01). Serum and bone Al concentrations were increased in both Al-treated groups (P < 0.01). There was a dose-dependent increase in bone Al levels (P < 0.01) and a dose-dependent decrease of CFU-E development (P<0.05). The CFU-E development was inversely correlated with the bone Al content (r=-0.79; P<0.05). The results demonstrate that even very low doses of Al citrate impair erythropoiesis in vivo and higher doses exert a deleterious action on both CFU-E and mature erythrocytes. This might show a local effect of Al on CFU-E caused by the bone sensitivity to the metal accumulation.

Oral aluminum administration to rats with normal renal function. 2. Body distribution.

Aluminum (Al) has no known physiological function and is not considered an essential dietary compound. Nowadays, it is recognized as an element that can produce adverse effects on biological systems. The present study determined Al partitioning in the body compartments of rats that have been orally exposed for 15 weeks. Three experimental groups were studied: Controls (C, n=19), TAl-1 (n=10) rats receiving daily doses of Al citrate (1.0 micromol/g body weight) by gavage and TAl-2 (n=13), receiving Al citrate with the drinking water (100 mmol/ l). At the end of the experimental period, the Al contents of organs and sera were determined. Results are expressed as median and range values. Comparing the TAl-2 rats with the control ones, remarkable Al accumulation could be observed in serum (4.8/2.7-16.3 vs 0.4/0.2-1.2 micromol/l, P<0.001), bone (3.33/1.78-4.85 vs 1.00/0.48-1.59 micromol/ g, P<0.001), kidney (2.33/0.96-3.15 vs 0.52/0.22-2.07 micromol/g, P<0.001), spleen (2.22/0.70-4.19 vs 0.27/ 0.11 - 0.36 micromol/g, P< 0.001) and liver (0.60/0.42-0.91 vs 0.24/0.14-0.78 micromol/g, P<0.01) while brain Al content was not significantly increased. Aluminum levels were raised in the TAl-1 group only in serum (2.8/1.3 - 10.4 micromol/ g, P < 0.001), bone (1.85/1.00-3.41 micromol/g, P < 0.001) and kidney (1.74/0.96-2.07 micromol/g, P<0.01). Bone Al concentration increased in a dose-dependent manner (TAl-2 vs TAl-1, P<0.001). The results demonstrate different tissue Al accumulation in rats chronically exposed to Al citrate, irrespective of their intact renal function.

Depressed erythroid progenitor cell activity in aluminum-overloaded mice.

The aim of the present study was to evaluate the effect of aluminum (Al) on in vivo erythropoiesis in mice that had received short- and long-term Al overloading. At the end of each treatment period, clonal assays of late erythroid progenitor cells (CFU-E) stimulated in vitro with erythropoietin were carried out, hematological parameters determined, and histological aluminon staining performed on bone and liver. After 2 weeks of oral ingestion of either Al chloride or Al citrate (10 mumol per day), poor CFU-E growth was obtained but no differences in hematocrit (Ht) and hemoglobin (Hb) values were found when comparing the treated and control groups. CFU-E development, determined after loading mice with Al citrate during 22 weeks (10 mumol/day), proved to be heavily depressed, and significant reductions in Ht, Hb concentration and erythrocyte osmotic fragility were also observed. No Al accumulation could be demonstrated in tissue, using histological aluminon staining. The results suggest that, even in the absence of signs of anemia, ingested Al may depress hematopoiesis by affecting red blood cell production and cell destruction.

The inhibitory action of aluminum on mouse bone marrow cell growth: evidence for an erythropoietin- and transferrin-mediated mechanism.

Aluminum (Al) has been associated with anemia in chronic renal failure patients under hemodialysis as well as in Al-overloaded animals. In an attempt to elucidate further the mechanism of Al toxicity we have investigated the effect of this ion on erythropoiesis in vitro. Mouse bone marrow cells were stimulated in vitro with erythropoietin (Epo) in the presence of Al3+ ion and erythroid colony-forming units were then determined. Results of this study indicate that Al compounds (chloride and citrate) at concentrations as low as 0.37 mumol Al/l inhibit erythropoiesis in vitro through a mechanism dependent upon the availability of transferrin to bind to aluminum. This process cannot be reversed by increasing Epo doses. This inhibition only occurs in the presence of Epo at early stages during the interaction of the hormone with its target cell.

Glycosaminoglycans in urolithiasis.

To determine if there are differences in urinary glycosaminoglycan (GAG) concentrations, 43 stone-forming patients and 37 healthy control subjects of both sexes were studied. Urinary concentrations of calcium, magnesium, creatinine, uric acid and GAGs were determined. GAGs were measured by the Di Ferrante precipitation procedure followed by the Bitter and Muir reaction. Urinary GAG concentration and daily output were significantly lower in stone-forming patients. The present study clearly demonstrates the decreased urinary GAG concentration and excretion in stone-forming patients and suggests an interaction between GAGs and urate that could modify the inhibitory potency of GAGs.

The inhibitory action of chlorogenic acid on the intestinal iron absorption in rats.

The polyphenols are part of the composition of many foods, it is known the inhibitory effect of tea and coffee through the tannins on iron intestinal absorption; the "yerba mate" (Ilex Paraguarensis) is a beverage widely used in South America, that has a high content of a polyphenol named chlorogenic acid. The present work shows the effect of this substance in nonhem iron absorption. An intestinal loop, was made in rats, to form a closed cavity in a small section of intestine tieing it from the pilorous to a distance of six cm. In this closed cavity a solution of 59Fe was injected with different doses of chlorogenic acid; it was living 20, 40 and 120 minutes into the loop, and after this different times, the blood, spleen, liver, femur and intestine were removed to measure the 59Fe uptake to be compared with the control group. The results gave an intense inhibitory effect on the intestinal iron absorption with doses of 0.58 and 1.7 mM per rat of chlorogenic acid at the different times studied.

The inhibitory action of chlorogenic acid on the intestinal iron absorption in rats.

The polyphenols are part of the composition of many foods, it is known the inhibitory effect of tea and coffee through the tannins on iron intestinal absorption; the [quot ]yerba mate[quot ] (Ilex Paraguarensis) is a beverage widely used in South America, that has a high content of a polyphenol named chlorogenic acid. The present work shows the effect of this substance in nonhem iron absorption. An intestinal loop, was made in rats, to form a closed cavity in a small section of intestine tieing it from the pilorous to a distance of six cm. In this closed cavity a solution of 59Fe was injected with different doses of chlorogenic acid; it was living 20, 40 and 120 minutes into the loop, and after this different times, the blood, spleen, liver, femur and intestine were removed to measure the 59Fe uptake to be compared with the control group. The results gave an intense inhibitory effect on the intestinal iron absorption with doses of 0.58 and 1.7 mM per rat of chlorogenic acid at the different times studied.

The inhibitory action of chlorogenic acid on the intestinal iron absorption in rats.

The polyphenols are part of the composition of many foods, it is known the inhibitory effect of tea and coffee through the tannins on iron intestinal absorption; the [quot ]yerba mate[quot ] (Ilex Paraguarensis) is a beverage widely used in South America, that has a high content of a polyphenol named chlorogenic acid. The present work shows the effect of this substance in nonhem iron absorption. An intestinal loop, was made in rats, to form a closed cavity in a small section of intestine tieing it from the pilorous to a distance of six cm. In this closed cavity a solution of 59Fe was injected with different doses of chlorogenic acid; it was living 20, 40 and 120 minutes into the loop, and after this different times, the blood, spleen, liver, femur and intestine were removed to measure the 59Fe uptake to be compared with the control group. The results gave an intense inhibitory effect on the intestinal iron absorption with doses of 0.58 and 1.7 mM per rat of chlorogenic acid at the different times studied.