Humoral immune response of pigs infected with Toxocara cati.

Toxocara cati is one of the causative agents of human toxocariasis. Serological methods are used for diagnosis in paratenic hosts like humans but the humoral immune response triggered by this parasite is unknown. We characterized the humoral immune response to T. cati excretory-secretory antigens (TES) in pigs as animal model during the acute and chronic stages of infection. ELISA and Western Blot techniques were used to determine antibody response. Pigs were experimentally inoculated with 100,000 infective Toxocara cati eggs. Blood was collected at 7, 14, 21 and 28 days post-inoculation (d.p.i.) to assess the acute stage of infection and 90, 120 and 180 d. p.i. for chronic stage analysis. ELISA showed values higher than the cut-off of specific IgM and IgG at 7 d. p.i. with significant differences at 0 and 7 d. p.i. for IgM and at 14, 21 and 28 d. p.i. for IgG in the acute stage. Higher and stable levels were detected in the chronic stage. Western Blot showed bands from 102 to 38 kDa detected by specific IgM and IgG. More immunogenic bands were identified by specific IgG. In the chronic stage of infection a band near 31 kDa was the only band detected by IgM until 150 d. p.i. Specific IgG recognized bands between 102 and 31 kDa. This study demonstrates how the humoral immune response evolves in the acute and chronic stages of infection and provides evidence on the role of the pig as a paratenic host of T. cati.

Infection of Rat Osteoblasts with Junin Virus Promotes the Expression of Bone Morphogenetic Protein 6, an Osteogenic Differentiation Inducer.

BACKGROUND: The arenavirus Junin virus (JUNV), causative agent of the argentine hemorrhagic fever, is able to modulate several signaling pathways involved in cell survival and multiplication. OBJECTIVES: We aimed to characterize the infection of rat osteoblasts (OBCs) with JUNV and its consequence on the modulation of osteogenic genes expression, thus studying the ability of this virus to induce cell differentiation. In addition, we evaluated the effect of purinergic agonists on viral replication. METHOD: Quantification of infectivity by plaque forming unit (PFU) assay, synthesis of viral proteins by western blot and immunofluorescence, and expression of osteogenic differentiation markers (ODM) by quantitative real-time polymerase chain reaction were employed. RESULTS: Infection of OBCs with JUNV (MOI 0.01 PFU/cell) showed a peak of infectivity, reaching 1.5 × 105 PFU/mL at the second day post-infection (p.i.). A marked restriction in multiplication was detected at day 7 p.i. that did not impair the establishment of a persistent stage of infection in OBCs. Analysis of mRNAs corresponding to ODM such as alkaline phosphatase, bone sialo-protein, and bone morphogenetic proteins (BMPs) 4 and 6 revealed that only the levels of BMP-6 were significantly higher in infected cells. Treatment with the purinergic agonists ATPγS, UTP, ADP, or UDP diminished viral titer and reduced the expression of the viral nucleoprotein. Also, treatment with 10 µM ATPγS reduced the stimulation of BMP-6 expression induced by the infection. CONCLUSIONS: These data demonstrate that JUNV is capable of infecting OBCs and point out BMP-6 as a key factor during this process.

First inter-laboratory comparison of Echinococcus granulosus sensu lato diagnosis in Latin America.

OBJECTIVE: To compare the performance of polymerase chain reaction (PCR) and enzyme-linked immunosorbent assay (ELISA) tests for diagnosing Echinococcus granulosus in dog feces among national reference laboratories in Argentina, Chile, Peru, and Uruguay. METHODS: National laboratories affiliated with the Ministry of Health/Agriculture of each country exchanged panels of 10 positive/negative samples obtained from their regular national surveillance programs in November 2015 - November 2016. All laboratories applied PCR; two also applied ELISA techniques. Sensitivity and specificity were determined for each laboratory and concordance of results among the laboratories was evaluated by Cohen Kappa coefficient. RESULTS: Poor concordance (3 of 10 paired comparisons had values of Kappa > 0.4), low sensitivity and specificity across all laboratories, and poor performance of both techniques in detecting E. granulosus in canine feces was demonstrated in this study. An ex-post comparison of the laboratories' test protocols showed substantial heterogeneity that could partially explain poor concordance of results. CONCLUSION: The results underscore the heterogeneity of canine echinococcosis diagnosis across the region and indicate possible sources of variability. Efforts to standardize canine echinococcosis testing must be included in the plan of action for the Regional Initiative for the Control of Cystic Echinococcosis. Future comparisons with fecal samples of known parasite load are needed.

Involvement of GSK3/ß-catenin in the action of extracellular ATP on differentiation of primary cultures from rat calvaria into osteoblasts.

Modulation of purinergic receptors play an important role in the regulation of osteoblasts differentiation and bone formation. In this study, we investigated the involvement of the GSK3/ßcatenin signaling in the action of ATPγ-S on osteogenic differentiation of primary cell cultures from rat calvaria. Our results indicate that the cell treatment with 10 or 100 µM ATPγ-S for 96 h increase the cytoplasmic levels of ß-catenin and its translocation to nucleus respect to control. A similar effect was observed after cell treatment with the GSK3 inhibitor LiCl (10 mM). Cell treatments with 4-10 mM LiCl significantly stimulated ALP activity respect to control at 4 and 7 days, suggesting that inhibition of GSK-3 mediates osteoblastic differentiation of rat calvarial cells. Effects comparison between ATP and LiCl shown that ALP activity was significantly increased by 10 µM ATPγ-S and decreased by 10 mM LiCl at 10 day of treatment, respect to control, suggesting that the effect of ATPγ-S was less potent but more persistent than of LiCl in stimulating this osteogenic marker in calvarial cells. Cell culture mineralization was significantly increased by treatment with 10 µM ATPγ-S and decreased by 10 mM LiCl, respect to control. In together, these results suggest that GSK3 inhibition is involved in ATPγ-S action on rat calvarial cell differentiation into osteoblasts at early steadies. In addition such inhibition by LiCl appear promote osteoblasts differentiation at beginning but has a deleterious effect on its function at later steadies as the extracellular matrix mineralization.

Effect of Combined Action of Extracellular ATP and Elevated Calcium on Osteogenic Differentiation of Primary Cultures From Rat Calvaria.

The in vitro osteogenic differentiation has been intensively studied. However, it is not yet clear precisely how osteogenesis can be optimized. Changes in extracellular Ca(2+) concentration ([Ca(2+) ]e ), as well as modulation of purinergic receptors play an important role in the regulation of osteoblasts differentiation and bone formation. In this study, we investigated the effects of a combined treatment of ATPγ-S and high [Ca(2+) ]e (5.35 mM) on osteogenic differentiation and function of primary cell cultures from rat calvaria. Our results indicate that ATPγ-S stimulates cell transition from the G0 to S phase of cell cycle, involving the PI3K signaling pathway. Treatment with 10 or 100 µM ATPγ-S and [Ca(2+) ]e (ATP-[Ca(2+) ]e ) for 48 h increases cell number significantly above the control. ATPγ-S treatment in osteogenic medium containing [Ca(2+) ]e stimulates the gene expression of BMP-4, BMP-5, and OPN at 16, 48, and 72 h, respectively, above control. In same conditions, treatment for 6 days with 10 µM UTP or 100 µM UDP significantly increased the ALP activity respect to control. Cells grown in osteogenic medium showed a statistically significant increase in calcium deposits at 15 and 18 days, for 10 µM ATPγ-S treatment, and at 18 and 22 days, for [Ca(2+) ]e treatment, respect to control but ATP-[Ca(2+) ]e treatment shown a significant greater mineralization at 15 days respect to ATPγ-S, and at 18 days respect to both agonists. In conclusion, we demonstrated that an osteogenic medium containing 10 µM ATPγ-S and 5.35 mM [Ca(2+) ]e enhance osteogenesis and mineralization by rat primary calvarial cells cultures. J. Cell. Biochem. 117: 2658-2668, 2016. © 2016 Wiley Periodicals, Inc.

[Study of infestation of dogs with Echinococcus granulosus in the province of La Rioja, Argentina]. / Estudio de infestación de caninos con Echinococcus granulosus en la provincia de La Rioja, Argentina.

This work was conducted in the province of La Rioja, located in northwestern Argentina. The aim of this study was to estimate the percentage of dog feces showing the presence of antigens of Echinococcus sp. in different regions of the province. A total of 269 samples of dried canine stool were taken, which were analyzed by the copro-ELISA technique. The most affected area was zone IV, which had 30.5% of positive samples. Zone I corresponding to the Capital Department of the province had 12% of positivity. In other areas, the percentages ranged between 11.4% and 14.8%. This is the first study in the province of La Rioja on the existence of this disease in dogs. The lack of control strategies has allowed the spread of echinococcosis.

Manipulating the bioactivity of hydroxyapatite nano-rods structured networks: effects on mineral coating morphology and growth kinetic.

BACKGROUND: Nano-hydroxyapatite particles have better bioactivity than the coarse crystals. So, they can be utilized for engineered tissue implants with improved efficiency over other materials. The development of materials with specific bioactive characteristics is still under investigation. METHODS: The surface properties of four hydroxyapatite materials templated by different micelle-polymer structured network are studied. The synergistic interaction of each block copolymer in contact with CTAB rod-like micelles results in crystalline HAp nano-rods of 25-50nm length organized in hierarchical structures with different micro-rough characteristics. RESULTS: It was observed that the material in vitro bioactivity strongly depends on the surface structure while in a minor extent on their Ca/P ratio. So, MIII and MIV materials with Skewness parameter Rsk>2.62 favored the formation on their surfaces of net-like phase with a high growth kinetic constant; while MI and MII (Rsk≤2.62) induced the appearance of spherulitic-like structures and a growth rate 1.75 times inferior. Material biocompatibility was confirmed by interaction with rat calvarial osteoblasts. CONCLUSIONS: The different structures growth is attributed to a dissimilar matching of crystal planes in the material and the apatite layer formed. In specific synthesis conditions, a biocompatible material with a Ca/P ratio close to that for the trabecular bone and a morphology that are considered essential for bone-bonding was obtained. GENERAL SIGNIFICANCE: The creation of implantable devices with a specific bioactive characteristic may be useful to manipulate the attachment of cells on mineral coating directly affecting the stability and life of the implant.

ATP and UTP stimulate bone morphogenetic protein-2,-4 and -5 gene expression and mineralization by rat primary osteoblasts involving PI3K/AKT pathway.

The modulation of purinergic receptors plays an important role in the regulation of bone formation by the osteoblast. On the other hand, bone morphogenetic proteins (BMPs), members of the transforming growth factor-ß superfamily, regulate the differentiation of osteoprogenitor bone cells and stimulate bone formation. In this study, we investigate the effects of several nucleotides on osteoblast differentiation and function, and their relation with the gene expression of osteogenic proteins BMP-2, BMP-4 and BMP-5 as well as of differentiation markers alkaline phosphatase (ALP) and bone sialoprotein (BSP). Our results indicate that 100µM ATP, ATPγS and UTP, but not ADP, ADPßS or UDP, promote ALP activity in rat primary osteoblasts, showing a peak about day 7 of the treatment. ATP, ATPγS and UTP also increase the mRNA levels of ALP, BMP-2, BMP-4, BMP-5 and BSP. Both the ALP activity and ALP and BMP-4 mRNA increments induced by ATP and UTP are inhibited by Ly294002, a PI3K inhibitor, suggesting the involvement of PI3K/AKT signaling pathway in purinergic modulation of osteoblast differentiation. Furthermore, bone mineralization enhance 1 and 1.5 fold after culturing osteoblasts in the presence of 100µM ATP or UTP, respectively, but not of ADP or UDP for 22 days. This information suggests that P2Y2 receptors (responsive to ATP, ATPγS and UTP) enhance osteoblast differentiation involving PI3K/AKT signaling pathway activation and gene expression induction of ALP, BMP-2, BMP-4, BMP-5 and BSP. Our findings state a novel molecular mechanism that involves specific gene expression activation of osteoblast function by the purinoreceptors, which would be of help in setting up new pharmacological strategies for the intervention in bone loss pathologies.

[Diagnosis of cystic echinococcosis in faeces scattered in areas of Puna and Quebrada. Province of Jujuy, Argentina]. / Diagnóstico de situación de la equinococosis quística en heces dispersas en las zonas de Quebrada y Puna, provincia de Jujuy, Argentina.

Echinococcosis is a parasitic disease common in livestock, caused by the cestode Echinococcus granulosus, the dog being the principal definitive host. The province of Jujuy is an endemic area located in the Northwest of Argentina. Due to the restricted ecological conditions of Quebrada and Puna, the most important activity of the population is formal cattle pastoralism and transhumance, especially of sheep and camelids. The dog acquires the double function of company and shepherd in these communities. The objective of the present study was to conduct a diagnosis of the situation in areas of La Quebrada and Puna where the circulation of E. granulosus is suspected. Five hundred and twenty three (523) samples of canine fecal material scattered in the environment were collected from 2002 to 2012. Prevalence varied from 2% in Susques to 27.7% in Humahuaca, the largest in the province. In Tumbaya, prevalence was 21% in the year 2007, reaching 0% in the year 2010 but increasing again to 10.5% in the year 2011. These results may be related to health education on preventive measures and mass deworming held prior to sample taking in the year 2010. A prevalence between 19.4% and 2% was observed in the rest of the regions studied, suggesting that a lack of strategies for echinococcosis control has allowed the spread of the disease.

Toxocariasis: seroprevalence in abandoned-institutionalized children and infants.

Toxocariasis is an infection that has worldwide distribution. Toxocara canis is the most relevant agent due to its frequent occurrence in humans. Soil contamination with embryonated eggs is the primary source of T. canis. This study aimed to determine the seroprevalence of toxocariasis in 10-month to 3 year-old abandoned infants, considered to be at high risk because of their orphanhood status and early age. Blood samples were collected from 120 children institutionalized in an orphanage in the city of La Plata. In this study, we observed 38.33% of seropositive cases for T. canis by ELISA and 45% by Western blot techniques; significant differences among groups A (<1 year), B (1-2 years) and C (>2 years) were also found. In research group A, children presented a seropositivity rate of 23.91%, in group B of 42.85% and in group C of 56%, which indicates an increase in frequency as age advances, probably because of greater chances of contact with infective forms of the parasite since canines and soil are frequently infected with T. canis eggs. Abandoned children come from poor households, under highly unsanitary conditions resulting from inadequate or lack of water supply and sewer networks, and frequent promiscuity with canines, which promotes the occurrence of parasitic diseases. These children are highly vulnerable due to their orphanhood status and age.

[Cystic echinococcosis in dogs and children in the province of Río Negro, Argentina]. / Vigilancia de la equinococosis quística en perros y niños en la provincia de Río Negro, Argentina.

Cystic echinococcosis (CE) is an endemic disease in the province of Río Negro, Argentina. The control program against CE has developed monitoring surveillance systems. Currently, the coproELISA/Western blot (WB) test is used to determine transmission in livestock farms (epidemiological units or EU) from collected dry field-dispersed dog feces. The objective was to evaluate the prevalence of CE on livestock farms and its relationship with CE cases in children aged 0-14 years. Canine fecal samples were obtained from randomly selected livestock farms and processed by the coproELISA/WB test. Furthermore, new cases in children in the same age group mentioned above were identified. In 278 EU, 571 samples of canine feces were obtained. There were 37 positive samples for coproELISA/WB (6.5%) and the presence of transmission was demonstrated in 37 EU (13.3%). There were no significant differences (p=0.9) with the survey conducted in the period 2003-2004 while there were significant differences (p=0.02) with the EU survey of native populations conducted in 2009-2010. With respect to animal density in the work area, the EU yielding negative results had an average of 2 dogs (SD 2.1) per EU while in the EU having positive results the average was 3 dogs (SD 4.2), showing statistically significant differences (p=0.02). In children under 15 years of age, 12 cases were diagnosed. This study has identified, on average, that the cases of hydatid disease are closer in the positive fields than in the negative fields (p=0.00307).The coproELISA/WB test allowed to identify the dispersion of CE on livestock farms and its relationship with the occurrence of cases in children in 2009-2010.

[Evaluation of a purified antigen for the diagnosis of toxocariosis]. / Evaluación de un antígeno purificado para el diagnóstico de toxocariosis.

Toxocarosis is a zoonotic disease caused by the ingestion of infective eggs of Toxocara spp. The diagnosis is based on the detection of antibodies in serum or other biological fluids. One of the current serological techniques for the diagnosis of toxocariasis is ELISA using excretory - secretory antigens of third stage larvae (ES/L3). These antigens are glycoproteins, which originate in the secretory organs of the parasite and are non species-specific. Sera from patients with other helminthiases and non- parasitic diseases were used to evaluate the specificity of ELISA using the excretory - secretory antigen (ES/L3). The reactivity of these sera was between 11 and 70%. Western blot using patients' sera revealed that the glycoprotein triplet having a molecular weight of 120 kDa was responsible for cross-reactivity. With these results, and for the purpose of purifying the antigen, ion exchange chromatography was performed. When the sera from patients with various parasitic and non-parasitic diseases were analyzed with the purified antigen ES/ L3, they were only reactive between 10 to 20%. The sensitivity of the ELISA test determined by program Epidat 3.0 for the two antigens was 100%, but the following differences in specificity were observed: 84% for the total antigen ES/L3 and 99% for purified ES/L3. Using the ES/L3 purified antigen, it can be considered that the reactive sera, with compatible symptoms correspond to patients who are or were parasitized with Toxocara canis.

[Coproantigens detection for the diagnosis of canine Echinococcosis in the border area of La Quiaca-Villazón]. / Detección de coproantígenos para el diagnóstico de echinococosis canina en la zona fronteriza de La Quiaca-Villazón.

Cystic Echinococcosis is a major public health issue. Immunodiagnostic techniques based on the identification of parasite antigens in dog dry faeces have been developed as alternatives for the surveillance of canine Echinococcosis. The environmental conditions favouring the parasite cycle were met in the border of La Quiaca-Villazón, given the presence of definitive (dog) and intermediate hosts (sheep and goats). The most important activity in La Puna is sheep and goat rearing; slaughtering is carried out almost exclusively in the field by sheep's owners, and preventive health measures do not apply. The aim of this study was to determine the presence of dogs parasitized by Echinococcus granulosus in this border region. A hundred and sixty eight (168) stool specimens were collected during 2006. La Quiaca samples were taken from the following selected areas: the semi-rural area of Barrios, the rural areas of Santa Catalina, Yavi Chico, El Portillo, Pumahuasi and Cara Cara and the urban area of La Quiaca; selected urban areas in Villazón and the semi-rural area of Ojo de Agua, Lampaya and Matancillas. The samples were analyzed by copro-ELISA -Western blot test. The cities of San Francisco and Barrios had a prevalence of 14.3% and 6.7%, respectively. A prevalence of 3.4% was observed in the urban area of Villazón, which indicates that dogs become infected in the rural areas and bring the risk into the city. Lampaya showed a prevalence of 30%. These findings suggest the need to implement strategies for the control of hydatidosis, both in urban and rural areas to avoid the increase and spread of Echinococcosis in the region.

Killing Bacteria by Faradaic Processes through Nano-Hydroxyapatite/MoOx Platforms.

Following the secular idea of ″restitutio ad integrum″, regeneration is the pursued option to restore bones lost after a disease; accordingly, complementing antibiotic and regeneration capacity to bone grafts represents a great scientific success. This study is a framework proposal for understanding the antimicrobial effect of biocompatible nano-hydroxyapatite/MoOx (nano-HA/MoOx) platforms on the basis of their electroactive behavior. Through cyclic voltammetry and chronoamperometry measurements, the electron transference capacity of nano-HA and nano-HA/MoOx electrodes was determined in the presence of pathogenic organisms: Pseudomonas aeruginosa and Staphylococcus aureus. Faradaic processes were confirmed and related to the switch of MoO42-/PO43- groups in the original hexagonal nano-HA crystal lattice and to the extent of OH vacancies that act as electron acceptors. Microscopic analysis of bacteria's ultrastructure showed a disruptive effect on the cytoplasmic membrane upon direct contact with the materials, which is not evident in the presence of eukaryotic cells. Experiments support the existence of a type of extracellular electron transfer (EET) process that alters the function of the bacterial cytoplasmic membrane, accelerating their death. Our findings provide strong quantitative support for a drug-independent biocidal physical approach based on EET processes between microorganisms and phosphate ceramics that can be used to combat local orthopedic infections associated with implants.

[Retroperitoneal hydatidosis secondary to hepatic hydatid cyst]. / Hidatidosis retroperitoneal secundaria a quiste hidatídico de localización hepática.

Hydatid disease is a worldwide zoonosis. It is caused by a parasitic platyhelminth of the genus Echinococcus. We present a patient with a fluctuating lumbar tumor in the retroperitoneal space, secondary to a hepatic cyst. the initial diagnosis was made by identification of rostellar hooks from protoscoleces in the fluid aspirated from the abscess. We herein describe the clinical manifestations, diagnosis and medical and surgical treatment of this unusual case and conclude that the development of an accurate diagnosis requires a proper analysis of the patient's epidemiological history, clinical manifestations, imaging studies and laboratory tests. a multidisciplinary approach and differential diagnosis is paramount to be able to establish a cause of the disease to deliver appropriate treatment.

Activation of the PI3K/Akt signaling pathway through P2Y2 receptors by extracellular ATP is involved in osteoblastic cell proliferation.

We studied the PI3K/Akt signaling pathway modulation and its involvement in the stimulation of ROS 17/2.8 osteoblast-like cell proliferation by extracellular ATP. A dose- and time-dependent increase in Akt-Ser 473 phosphorylation (p-Akt) was observed. p-Akt was increased by ATPγS and UTP, but not by ADPßS. Akt activation was abolished by PI3K inhibitors and reduced by inhibitors of PI-PLC, Src, calmodulin (CaM) but not of CaMK. p-Akt was diminished by cell incubation in a Ca²âº-free medium but not by the use of L-type calcium channel blockers. The rise in intracellular Ca²âº induced by ATP was potentiated in the presence of Ro318220, a PKC inhibitor, and attenuated by the TPA, a known activator of PKC. ATP-dependent p-Akt was diminished by TPA and augmented by Ro318220 treatment in a Ca²âº-containing but not in a Ca²âº-free medium. ATP stimulated the proliferation of both ROS 17/2.8 cells and rat osteoblasts through PI3K/Akt. In the primary osteoblasts, ATP induces alkaline phosphatase activity via PI3K, suggesting that the nucleotide promotes osteoblast differentiation. These results suggest that ATP stimulates osteoblast proliferation through PI-PLC linked-P2Y2 receptors and PI3K/Akt pathway activation involving Ca²âº, CaM and Src. PKC seems to regulate Akt activation through Src and the Ca²âº influx/CaM pathway.

Protein phosphatases: possible bisphosphonate binding sites mediating stimulation of osteoblast proliferation.

We investigated the existence of a bisphosphonate (BP) target site in osteoblasts. Binding assays using [³H]-olpadronate ([³H]OPD) in whole cells showed the presence of specific, saturable and high affinity binding for OPD (K(d)=1.39 ± 0.33 µM) in osteoblasts. [³H]OPD was displaced from its binding site by micromolar concentrations of lidadronate, alendronate and etidronate (K(d)=1.42 ± 0.15 µM, 2.00 ± 0.2 µM and 2.4 ± 0.4 µM, respectively), and by millimolar concentrations of the non-permeant protein phosphatase (PP) substrates p-nitrophenylphosphate and α-naphtylphosphate. PP inhibitors orthovanadate, NaF or vpb(bipy) did not displace [³H]OPD. As expected, specific OPD binding was detected in the plasma membrane of ROS 17/2.8 cells, although significant BP binding was also found intracellularly. Moreover, OPD increased DNA synthesis in these cells with a temporal profile similar to the protein tyrosine phosphatase (PTP) inhibitors, Na3VO4 and vpb(bipy); but different from a general PP inhibitor (NaF). The stimulatory effect of OPD and PTP inhibitors on osteoblast proliferation was inhibited by the protein tyrosine kinase inhibitors genistein and geldanamycin. These results provide new evidence on the existence of a BP target in osteoblastic cells, presumably a PTP, which may be involved in the stimulatory action of BPs on osteoblast proliferation.

[Circulation of Echinococcus granulosus in Los Antiguos, Santa Cruz]. / Circulación del parásito Echinococcus granulosus en Los Antiguos, Santa Cruz.

Hydatidosis - caused by the parasite Echinococcus granulosus - is a zoonosis endemic to the province of Santa Cruz, associated with areas of livestock production. Once infected, man may remain asymptomatic for a prolonged period but the disease has an important impact on public health owing to the complexity and costs of its treatment. This study aimed at assessing the prevalence of environmental E. granulosus infection in rural and peri-urban areas of the town of Los Antiguos, through the detection of antigens in scattered canine feces, and to identify risk factors for transmission. From May-2016 to April-2017, 38 peri-urban and rural farms, defined as "epidemiological units", were visited; 144 samples of canine feces were analyzed with the copro-ELISA technique. Rural settlers were enrolled in an epidemiological survey on cultural patterns related to hydatidosis. An environmental contamination index of 17.3% was found and 44.7% of the epidemiological units were positive. Risk practices were found, such as domiciliary slaughter (34.2%), canine feeding with raw viscera (52.6%), and lack of dog deworming (86.8%). In turn, about half of the surveyed population ignored the modes of transmission of the infection and the measures to prevent it. This work shows a high infection index in the area and establishes a baseline for future comparisons. It also reinforces the need to implement education, prevention, and control activities at the local level - according to national program guidelines - in order to reduce the prevalence of environmental contamination of the disease.

La hidatidosis, causada por el parásito Echinococcus granulosus, es una zoonosis endémica en la provincia de Santa Cruz asociada a áreas de producción ganadera. El hombre puede permanecer asintomático durante un largo período luego de la infección. Una vez desarrollada, la enfermedad representa un importante problema de salud pública debido a la complejidad y el costo de su tratamiento. Los objetivos del trabajo fueron determinar la prevalencia de contaminación ambiental por E. granulosus en zona rural y periurbana de la localidad de Los Antiguos, mediante la detección de antígenos específicos en heces caninas, e identificar factores de riesgo de transmisión. Entre mayo-2016 y abril-2017, se visitaron 38 chacras periurbanas y estancias rurales, definidas como "unidades epidemiológicas". Se analizaron 144 muestras de heces de caninos con técnica copro-ELISA. Se realizó una encuesta epidemiológica de los pobladores rurales sobre patrones culturales relacionados a la hidatidosis. Se encontró un índice de contaminación ambiental del 17.3% y el 44.7% de las unidades epidemiológicas fueron positivas. Se identificaron prácticas de riesgo como faena domiciliaria (34.2%), alimentación de caninos con vísceras crudas (52.6%), y ausencia de desparasitación de perros (86.8%). Alrededor de la mitad de la población encuestada desconocía las formas de contagio y las medidas de prevención de la enfermedad. Este trabajo muestra un índice alto de contaminación y establece una línea de base para realizar comparaciones a futuro. También refuerza la necesidad de implementar medidas de educación, prevención y control de hidatidosis a nivel local de acuerdo a los programas nacionales.

Development of a New LAMP Assay for the Detection of Ancylostoma caninum DNA (Copro-LAMPAc) in Dog Fecal Samples.

Ancylostoma caninum is a zoonotic nematode which is able to affect animals and humans. Diagnosis in the definitive host and environmental detection are key to prevent its dissemination and achieve control. Herein, a new coprological LAMP method for the detection of A. caninum (Copro-LAMPAc) DNA was developed. DNA extraction was performed using a low-cost method and a fragment of the cox-1 gene was used for primer design. The analytical sensitivity, evaluated with serial dilutions of genomic DNA from A. caninum adult worms, was 100 fg. A specificity of 100% was obtained using genomic DNA from the host and other pathogens. The Copro-LAMPAc was evaluated using environmental canine fecal samples. When compared with gold standard optical microscopy in epidemiological studies, it proved to be more sensitive. This new LAMP assay can provide an alternative protocol for screening and identification of A. caninum for epidemiological studies in endemic areas.

ATP stimulates the proliferation of MCF-7 cells through the PI3K/Akt signaling pathway.

We studied the modulation of the PI3K/Akt signaling pathway by ATP in MCF-7 cells. Western blot analysis showed that ATP stimulated the phosphorylation of Akt in a dose- and time-dependent manner. Akt phosphorylation in response to nucleotides followed the potency order ATP=UTP=ATPgammaS>>ADP=UDP>ADPbetaS=adenosine, suggesting participation of P2Y(2/4) receptors. Inhibitors of PI3K, PLC, PKC and Src or Src antisense oligonucleotides prevented ATP-induced phosphorylation of Akt. Incubation of cells with 2-APB or in a nominally Ca(2+)-free medium plus EGTA showed that Akt phosphorylation by ATP depends on intracellular calcium release but is independent of calcium influx. The PI3K inhibitor was not effective in reducing MAPKs phosphorylation by ATP. ATP and UTP stimulated MCF-7 cell proliferation, effect that was inhibited by PI3K, PLC, PKC, Src and MAPKs inhibitors. These findings suggest that ATP modulation of P2Y(2/4) receptors increases MCF-7 cell proliferation by activation of the PI3K/Akt signaling pathway through PLC/IP(3)/Ca(2+), PKC and Src.