Abstract in English:In parasitology, routine laboratory diagnosis involves conventional methods, such as optical microscopy, used for the morphological identification of parasites. Currently, molecular biology techniques are increasingly used to diagnose parasite structures in order to enhance the identification and characterization of parasites. The objective of the present study was to review the main current and new diagnostic techniques for confirmation of parasite infections, namely: polymerase chain reaction (PCR), real-time polymerase chain reaction (RT-PCR), loop-mediated isothermal amplification (LAMP), Luminex xMAP, random amplified polymorphic DNA (RAPD), amplified fragment length polymorphism (AFLP), and restriction fragment length polymorphism (RFLP), in addition to microsatellites. Molecular assays have comprehensively assisted in the diagnosis, treatment and epidemiological studies of parasitic diseases that affect people worldwide, helping to control parasitic disease mortality.
Abstract in English:Duffy gene (FY) codifies the transmembrane glycoprotein Duffy (gp-Fy) of 35 to 43 kDa which is moderately immunogenic. This glycoprotein is polymorphic, and constitutes the antigens of the Duffy histo-blood system which were designated receptors for chemokines and denominated DARC (Duffy antigen/receptor for chemokine). This receptor has an important role in the regulation of chemokine levels in the circulation, as it binds and adsorbs them on the surface of red cells as a reservoir. It plays a "sink" role, which can contribute to homeostasis by removing inflammatory chemokines from circulation as well as maintaining them in plasmatic levels. Chronic Chagas' cardiopathy (CCC) is the most frequent form of the disease. It is an inflammatory disease, in which infiltrated inflammatory cells play an important role in the development and progress of the infection. High chemokine levels in the plasma have been associated with the disease severity in patients with heart failure. In this context, the profile of DARC expression could play an important function as a receptor for chemokines in Chagas' disease, in patients with CCC, as it can modulate damage from this inflammatory disease.
Abstract in English:Fasciolosis is a disease that affects the liver parenchyma and bile ducts of numerous animals, including humans, which causes economic losses and threatens public health. The present work aimed at reporting the distribution and factors connected with Fasciola hepatica infection in cattle in ten municipalities in southern Espírito Santo, Brazil, between 2008 and 2009. During this period, 50 rural properties in ten cities were analyzed and fecal samples from 10% of the cattle on each farm were collected. Of the 717 samples analyzed, 154 (21.33%) presented Fasciola hepatica eggs. As for the 50 rural properties studied, 32 (64%) had animals positive for fasciolosis. Throughout the evaluation, it was observed that 42 (84%) rural properties had flooded areas, 33 (66%) were located at altitudes less than 226 meters, in 36 (72%) mollusks were found, 47 (94%) had other definitive hosts (horses, sheep, goats) and 25 (50%) had previous cases of bovine fasciolosis. The present results showed a high rate of bovine fasciolosis in the analyzed region whereas infected animals are present in all the municipalities. Based on statistical analysis, it was concluded that there was an association between the presence of flooded areas and other definitive hosts in rural properties where bovine fasciolosis was detected.
Abstract in English:Schistosomiasis is a major public health problem with 207 million people infected and more than 779 million at risk. The drug of choice for treating schistosomiasis is praziquantel (PZQ); however, it is inefficient against immature forms of schistosomes. The aim of this study was to test new imidazolidine derivatives LPSF/PT09 and LPSF/PT10 against adult Schistosoma mansoni worms. IC50, cytotoxicity, immune response and cell viability assays were also available for these imidazolidines. Different concentrations of imidazolidine, from 32 to 320 ¼M, promoted motor abnormalities in breeding and unpaired worms, and death in 24 hours at higher concentrations. Although LPSF/PT09 and LPSF/PT10 did not affect IFN-³ and IL-10 production, they induced nitric oxide production and showed a similar behavior to praziquantel on cell death test. Thus, these new imidazolidine derivatives should undergo further study to develop schistosomiasis drugs.
Abstract in English:Snakes that can ingest prey that are proportionally large have high metabolic rates during digestion. This great increase in metabolic rate (specific dynamic action - SDA) may create a significant augment in the animal's body temperature. The present study investigated postprandial thermogenesis in Bothrops moojeni. Briefly, two groups of snakes were fed meals equivalent to 17 ± 3% and 32 ± 5% of their body weight and were observed for 72 hours, in which thermal images of each snake were taken with an infrared camera in a thermostable environment with a constant air temperature of 30°C. The results showed a significant increase in snake surface temperature, with a thermal peak between 33 and 36 hours after feeding. The meal size had a great impact on the intensity and duration of the thermogenic response. Such increase in temperature appears to be connected with the huge increase in metabolic rates during digestion of relatively large prey by snakes that feed infrequently. The ecologic implication of the thermogenic response is still not well understood; however, it is possible that its presence could affect behaviors associated with the snake digestion, such as postprandial thermophily.
Abstract in English:There is an interest in the use of IL-12 as a possible anti-cancer drug to induce immune responses and anti-IL-13 formulations to treat the undesirable effects of IL-13. Thus, the present study aimed at analyzing IL-12 and IL-13 profiles, viral hepatitis serology and blood cultures in acute myeloid leukemia (AML) patients. Forty individuals (20 without septicemia - Group A, and 20 with septicemia - Group B) and 20 healthy controls were evaluated. Hepatitis B virus antigens (HBsAg) and hepatitis C virus antibodies (HCV Ab) were quantified using commercial ELISA kits. IL-12 and IL-13 levels were estimated in culture supernatant of mitogen-stimulated peripheral blood mononuclear cells by ELISA. Significantly low IL-12 values were observed among AML patients compared to controls whereas the opposite was observed regarding IL-13. IL-12 levels were found to be elevated in the follow-up cases. M4 and M5 subtypes of AML presented higher IL-12 levels than M1 and M2 subtypes. The isolated organisms from AML with septicemia were Staphylococcus aureus (35%), Esherichia coli (25%), coagulase-negative staphylococci (25%), and Candida (15%). Fungemia cases showed higher IL-12 values than bacteremia cases. In conclusion, IL-12 and IL-13 should be further tested in large-scale studies to provide future immunotherapy against AML.
Abstract in English:Cutaneous leishmaniasis is an infectious disease caused by protozoans of the genus Leishmania, which is transmitted through the bite of hematophagous insects of the genus Lutzomyia. This study aimed at testing in vitro the phototoxic effect of aluminum phthalocyanine tetrasulfonate (AlPcS4) on the viability of Leishmania major and Leishmania braziliensis. Stationary phase promastigote forms were treated with AlPcS4 at 1.0 µM and 10.0 µM and incubated for one hour. Then 659 nm laser was applied at 5 and 10 J/cm². Parasite viability was determined by differential count using the trypan blue dye exclusion method and by monitoring growth curves for nine days. Trypan blue exclusion assay showed a significant reduction of viable parasites compared to controls, L. major seemed more sensitive to the toxic effects of AlPcS4 in the dark. The most effective photodynamic therapy (PDT) was obtained with AlPcS4 at 10.0 µM and 10 J/cm² whereas L. braziliensis showed the highest mortality rate after treatment.
Abstract in English:The skin has an important role in the transmission of visceral leishmaniasis (VL) as the infection pathway in dogs. To better characterize the inflammatory response of intact skin in VL, sixty infected dogs (30 symptomatic and 30 asymptomatic) and six non-infected controls were studied. Diagnosis of visceral leishmaniasis was confirmed by RIFI and ELISA; direct visualization of the parasite in bone marrow aspirate; imprints of popliteal lymph nodes, spleen, liver and skin; culture in NNN-phase liquid Schneider's medium; and PCR (performed only in the ear skin). Amastigote forms of the parasite in intact skin were found only in symptomatic dogs. Inflammatory infiltrates were observed in all groups, varying from intense and/or moderate in symptomatic to discrete and/or negligible in asymptomatic and control animals. Parasite load was associated with the intensity of the inflammatory response and with clinical manifestations in canine visceral leishmaniasis. AgNOr as active transcription markers were expressed in inflammatory cells and within apoptotic bodies in all groups, including controls, with no statistical difference. Therefore, cell activation and transcription do occur in both symptomatic and asymptomatic canine visceral leishmaniasis and may result in more necrosis and inflammation or in apoptosis and less symptoms, depending on the parasite load.
Abstract in English:Cell-free antigens (CFAg) derived from Paracoccidioides brasiliensis have typically been used in immunodiffusion reactions for serodiagnosis or therapeutic follow-up of paracoccidioidomycosis patients. Thus, we investigated the usefulness of CFAg obtained from cultures at different ages, to evaluate cellular immunity by the footpad test, in experimental murine paracoccidioidomycosis. Male mice infected with P. brasiliensis 265 strain were challenged in the footpad with CFAg obtained from four- (4d CFAg) or 11-day-old cultures (11d CFAg). The increase in footpad swelling provoked by 4d CFAg and 11d CFAg was similar and showed significant difference in relation to control groups. However, the infiltrate pattern was strikingly different: 4d CFAg induced a predominant mononuclear infiltrate whereas 11d CFAg provoked a predominant polymophonuclear infiltrate. These different inflammatory patterns were associated with distinct electrophoretic characteristics. By comparison with 11d CFAg, 4d CFAg showed more numerous and intense bands, including a strong one of 43 kDa (gp43). These results suggest that CFAg derived from Pb 265 isolate can be used as a reagent to evaluate cellular immunity; however, the culture's age is critical because only young cultures are able to induce a typical mononuclear infiltrate. The efficacy of this new paracoccidioidin to assay the cellular immunity in infections caused by other P. brasiliensis isolates is under investigation.
Abstract in English:Among the various methods for evaluating animal venom toxicity, the calculation of the median lethal dose (LD50) is the most widely used. Although different protocols can be used to calculate the LD50, the source of the venom and the method of extraction, as well as the strain, age, and sex of the animal model employed, should be taken into consideration. The objective of the present study was to evaluate the influence of sex and age on the toxicity of Tityus serrulatus scorpion venom in Swiss mice. Although the symptoms of envenomation were similar in male and female animals, female mice proved to be more resistant to the venom. In females, age had no impact on the susceptibility to scorpion envenomation. Male mice were more sensitive to T. serrulatus venom. Moreover, in males, age was an important parameter since sensitivity to the venom increased with age.
Abstract in English:In this study, we evaluated the actions of Crotalus durissus cumanensis venom (CDCmV), and its crotoxin (Crtx) fraction, on renal and vascular functions in Wistar rats. In isolated perfused kidneys, CDCmV (10 µg/mL) significantly increased the perfusion pressure (PP) from 110.7 ± 2.4 to 125.3 ± 2.8 mmHg after 30 minutes. This effect was accompanied by an increased renal vascular resistance (RVR) from 5.4 ± 0.1 to 6.2 ± 0.2 mmHg/mL.g-1.min-1. We observed decreases in urinary flow (UF) from 0.13 ± 0.01 to 0.05 ± 001 mL.g-1.min-1 and glomerular filtration rate (GFR) from 0.66 ± 0.06 to 0.18 ± 0.02 mL.g-1.min-1. Crtx did not change PP or RVR, but diminished GFR (from 0.65 ± 0.05 to 0.26 ± 003 mL.g-1.min-1) and UF (from 0.11 ± 0.008 to 0.09 ± 0.008 mL.g-1.min-1). Both CDCmV and Crtx reduced the percentage of tubular transport of sodium, chloride and potassium. The cytotoxicity of these substances against MDCK cells was tested by the MTT method: only CDCmV caused a decrease in the cell viability with an IC50 of 5.4 µg/mL. In endothelium-intact isolated aortic rings, CDCmV (0.1 to 30 µg/mL) increased the sustained phenylephrine-induced contraction to a value of 130.0 ± 6.6% of its corresponding control, but showed a relaxant effect in endothelium-denuded preparations. Similar results were observed in aortic rings contracted with potassium (40 mM). Crtx was ineffective in aortic ring assays. Thus, it is reasonable to suggest that the renal effects induced by the CDCmV may be due to its influence on the endothelium's ability to release factors that can alter the contractile behavior of vascular smooth muscle. In conclusion, CDCmV is toxic to kidney cells. It changes parameters of the renal function including the glomerular filtration rate, renal vascular resistance and tubular transport. The actions induced by CDCmV also involve endothelium-dependent vasoactive properties. Their effects may be only partially attributed to Crtx.
Abstract in English:Uninfected dogs (n = 10) and those naturally infected with leishmaniasis (n = 10) were subjected to several diagnostic tests, namely: hemoculture, polymerase chain reaction (PCR) of hemoculture, indirect immunofluorescence (RIFI), cytological examination of lymph node aspirate, culture of lymph node aspirate and PCR of lymph node aspirate. RIFI - followed by PCR of lymph node aspirate culture - presented more positive results in infected dogs than in uninfected ones. In infected animals, RIFI was more effective than PCR of lymph node aspirate culture. There was no statistical difference in positivity between RIFI and hemoculture; lymph node aspirate culture/cytological examination of lymph node aspirate and PCR of hemoculture; and between PCR of lymph node aspirate culture and PCR of hemoculture. All infected and uninfected animals had positive and negative results in at least one test. In conclusion, the association of several tests improves the efficacy of canine visceral leishmaniasis diagnosis.
Abstract in English:Giardia duodenalis is a complex species that comprises at least seven distinct genetic groups (A to G), but only genotypes A and B are known to infect humans and a wide variety of other mammals. Regardless of biological, biochemical and antigenic analysis, several isolates maintained in vitro were not genetically typed yet. So, in the present study, five Brazilian axenic isolates obtained from asymptomatic and symptomatic patients were typed in order to determine the major genetic groups to which the isolates belonged. DNA was extracted from axenic trophozoites, fragments of glutamate dehydrogenase (gdh) and triosephosphate isomerase (tpi) genes were amplified by PCR and the isolate genotyping was carried out using restriction fragment length polymorphism (RFLP) and DNA sequencing for both genes. The results revealed that all isolates were assigned to genotype A at both analyzed loci. Indeed, DNA sequence analysis classified the four isolates obtained from asymptomatic individuals into subtype AII, while the isolate obtained from the symptomatic patient was typed as subtype AI. Despite of the limited number of isolates assessed, the findings presented herein provide interesting insights on the occurrence of Giardia genotypes in Brazil and hold the perspective for future molecular and epidemiological investigations.
Abstract in English:Cerebral infarction after a viper bite is relatively uncommon. A combination of factors has been implicated in the pathophysiology of infarct following snakebite. In this case report, the clinical outcome after a posterior circulation infarct and various possibilities that could lead to such a catastrophic event are discussed. The present study stresses the need to keep hydration, blood pressure and central venous pressure optimal in all snakebite patients. Cerebral infarction should be considered a differential diagnosis, in any patient with neurological deterioration following snakebite. Prognosis of such patients with posterior circulation stroke remains poor and decompressive craniectomy has not been found to be helpful.