Scielo RSS <![CDATA[Journal of Venomous Animals and Toxins including Tropical Diseases]]> http://www.scielo.br/rss.php?pid=1678-919920140001&lang=es vol. 20 num. lang. es <![CDATA[SciELO Logo]]> http://www.scielo.br/img/en/fbpelogp.gif http://www.scielo.br <![CDATA[Envenomation by caterpillars (erucism): proposal for simple pain relief treatment]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S1678-91992014000100102&lng=es&nrm=iso&tlng=es <![CDATA[Current challenges for confronting the public health problem of snakebite envenoming in Central America]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S1678-91992014000100203&lng=es&nrm=iso&tlng=es Snakebite envenoming is a serious public health problem in Central America, where approximately 5,500 cases occur every year. Panama has the highest incidence and El Salvador the lowest. The majority, and most severe, cases are inflicted by the pit viper Bothrops asper (family Viperidae), locally known as ‘terciopelo’, ‘barba amarilla’ or ‘equis’. About 1% of the bites are caused by coral snakes of the genus Micrurus (family Elapidae). Despite significant and successful efforts in Central America regarding snakebite envenomings in the areas of research, antivenom manufacture and quality control, training of health professionals in the diagnosis and clinical management of bites, and prevention of snakebites, much remains to be done in order to further reduce the impact of this medical condition. This essay presents seven challenges for improving the confrontation of snakebite envenoming in Central America. Overcoming these challenges demands a coordinated partnership of highly diverse stakeholders though inter-sectorial and inter-programmatic interventions. <![CDATA[A historical approach to scorpion studies with special reference to the 20 <sup>th</sup> and 21st centuries]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S1678-91992014000100204&lng=es&nrm=iso&tlng=es This work provides historical context about scorpion studies from the end of the 19 th century to the present day. The content is mainly addressed to non-zoologists, working in research fields that embrace scorpion biology, notably to those working with venoms and toxins. The historical aspects described include academic professional scholars who worked on scorpion classification and general distribution patterns; and to a lesser extent, on studies of ecology and natural history. The aim is not to provide an exhaustive description of all scholars who in one way or another became involved with scorpions, but rather of those who greatly contributed during a given period to the research of these organisms. No critical analysis of the work of previous researchers is undertaken, but some comments are proposed to bring clarification on ‘who’s who’. Since a global consensus in relation to classification and/or distribution patterns has not been reached among modern experts, these different approaches are also presented without judgment. Consequently, distinct approaches remain open for discussion. <![CDATA[Septic arthritis: immunopathogenesis, experimental models and therapy]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S1678-91992014000100205&lng=es&nrm=iso&tlng=es Septic arthritis is an inflammatory disease of the joints that is started by an infection whose most common agent is Staphylococcus aureus. In this review we discuss some of the most arthritogenic bacterial factors and the contribution of innate and specific immune mechanisms to joint destruction. Special emphasis is given to the induction of experimental arthritis by S. aureus in mice. The improvement of therapy by association of antibiotics with down-modulation of immunity is also included. <![CDATA[Snake venom L-amino acid oxidases: an overview on their antitumor effects]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S1678-91992014000100206&lng=es&nrm=iso&tlng=es The L-amino acid oxidases (LAAOs) constitute a major component of snake venoms and have been widely studied due to their widespread presence and various effects, such as apoptosis induction, cytotoxicity, induction and/or inhibition of platelet aggregation, hemorrhage, hemolysis, edema, as well as antimicrobial, antiparasitic and anti-HIV activities. The isolated and characterized snake venom LAAOs have become important research targets due to their potential biotechnological applications in pursuit for new drugs of interest in the scientific and medical fields. The current study discusses the antitumor effects of snake venom LAAOs described in the literature to date, highlighting the mechanisms of apoptosis induction proposed for this class of proteins. <![CDATA[Hump-nosed viper bite: an important but under-recognized cause of systemic envenoming]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S1678-91992014000100207&lng=es&nrm=iso&tlng=es Hump-nosed viper bites are common in the Indian subcontinent. In the past, hump-nosed vipers (Hypnale species) were considered moderately venomous snakes whose bites result mainly in local envenoming. However, a variety of severe local effects, hemostatic dysfunction, microangiopathic hemolysis, kidney injury and death have been reported following envenoming byHypnale species. We systematically reviewed the medical literature on the epidemiology, toxin profile, diagnosis, and clinical, laboratory and postmortem features of hump-nosed viper envenoming, and highlight the need for development of an effective antivenom. <![CDATA[Systemic effects induced by intralesional injection of ?-conotoxin MVIIC after spinal cord injury in rats]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S1678-91992014000100311&lng=es&nrm=iso&tlng=es Background : Calcium channel blockers such as conotoxins have shown a great potential to reduce brain and spinal cord injury. MVIIC neuroprotective effects analyzed in in vitro models of brain and spinal cord ischemia suggest a potential role of this toxin in preventing injury after spinal cord trauma. However, previous clinical studies with MVIIC demonstrated that clinical side effects might limit the usefulness of this drug and there is no research on its systemic effects. Therefore, the present study aimed to investigate the potential toxic effects of MVIIC on organs and to evaluate clinical and blood profiles of rats submitted to spinal cord injury and treated with this marine toxin. Rats were treated with placebo or MVIIC (at doses of 15, 30, 60 or 120 pmol) intralesionally following spinal cord injury. Seven days after the toxin administration, kidney, brain, lung, heart, liver, adrenal, muscles, pancreas, spleen, stomach, and intestine were histopathologically investigated. In addition, blood samples collected from the rats were tested for any hematologic or biochemical changes. Results : The clinical, hematologic and biochemical evaluation revealed no significant abnormalities in all groups, even in high doses. There was no significant alteration in organs, except for degenerative changes in kidneys at a dose of 120 pmol. Conclusions : These findings suggest that MVIIC at 15, 30 and 60 pmol are safe for intralesional administration after spinal cord injury and could be further investigated in relation to its neuroprotective effects. However, 120 pmol doses of MVIIC may provoke adverse effects on kidney tissue. <![CDATA[Purification and characterization of a hyaluronidase from venom of the spider <em>Vitalius dubius </em> (Araneae, Theraphosidae)]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S1678-91992014000100312&lng=es&nrm=iso&tlng=es Background Venom hyaluronidase (Hyase) contributes to the diffusion of venom from the inoculation site. In this work, we purified and characterized Hyase from the venom of Vitalius dubius (Araneae, Theraphosidae), a large theraphosid found in southeastern Brazil. Venom obtained by electrical stimulation of adult male and female V. dubius was initially fractionated by gel filtration on a Superdex® 75 column. Active fractions were pooled and applied to a heparin-sepharose affinity column. The proteins were eluted with a linear NaCl gradient. Results Active fractions were pooled and assessed for purity by SDS-PAGE and RP-HPLC. The physicochemical tests included optimum pH, heat stability, presence of isoforms, neutralization by flavonoids and assessment of commercial antivenoms. Hyase was purified and presented a specific activity of 148 turbidity-reducing units (TRU)/mg (venom: 36 TRU/mg; purification factor of ~4). Hyase displayed a molecular mass of 43 kDa by SDS-PAGE. Zymography in hyaluronic-acid-containing gels indicated an absence of enzyme isoforms. The optimum pH was 4-5, with highest activity at 37°C. Hyase was stable up to 60°C; but its activity was lost at higher temperatures and maintained after several freeze-thaw cycles. The NaCl concentration (up to 1 M) did not influence activity. Hyase had greater action towards hyaluronic acid compared to chondroitin sulfate, and was completely neutralized by polyvalent antiarachnid sera, but not by caterpillar, scorpion or snakes antivenoms. Conclusion The neutralization by arachnid but not scorpion antivenom indicates that this enzyme shares antigenic epitopes with similar enzymes in other spider venoms. The biochemical properties of this Hyase are comparable to others described. <![CDATA[Identification of two novel cytolysins from the hydrozoan <em>Olindias sambaquiensis</em> (Cnidaria)]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S1678-91992014000100316&lng=es&nrm=iso&tlng=es Background : Although the hydrozoan Olindias sambaquiensis is the most common jellyfish associated with human envenomation in southeastern and southern Brazil, information about the composition of its venom is rare. Thus, the present study aimed to analyze pharmacological aspects of O. sambaquiensis venom as well as clinical manifestations observed in affected patients. Crude protein extracts were prepared from the tentacles of animals; peptides and proteins were sequenced and submitted to circular dichroism spectroscopy. Creatine kinase, cytotoxicity and hemolytic activity were evaluated by specific methods. Results : We identified two novel cytolysins denominated oshem 1 and oshem 2 from the tentacles of this jellyfish. The cytolysins presented the amino acid sequences NEGKAKCGNTAGSKLTFKSADECTKTGQK (oshem 1) and NNSKAKCGDLAGWSKLTFKSADECTKTGQKS (oshem 2) with respective molecular masses of 3.013 kDa and 3.375 kDa. Circular dichroism revealed that oshem 1 has random coils and small α-helix conformation as main secondary structure whereas oshem 2 presents mainly random coils as its main secondary structure probably due to the presence of W (13) in oshem 2. The hemolysis levels induced by oshem 1 and oshem 2 using a peptide concentration of 0.2 mg/mL were, respectively, 51.7 ± 6.5% and 32.9 ± 8.7% (n = 12 and p ≤ 0.05). Oshem 1 and oshem 2 showed significant myonecrotic activity, evaluated by respective CK level measurements of 1890.4 ± 89 and 1212.5 ± 103 (n = 4 and p ≤ 0.05). In addition, myonecrosis was also evaluated by cell survival, which was measured at 72.4 ± 8.6% and 83.5 ± 6.7% (n = 12 and p ≤ 0.05), respectively. The structural analysis showed that both oshem 1 and oshem 2 should be classified as a small basic hemolytic peptide. Conclusion : The amino acid sequences of two peptides were highly similar while the primary amino acid sequence analysis revealed W (22th) as the most important mutation. Finally oshem 1 and oshem 2 are the first cytolytic peptides isolated from the Olindias sambaquiensis and should probably represent a novel class of cytolytic peptides. <![CDATA[Comparative analysis of the venom proteome of four important Malaysian snake species]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S1678-91992014000100317&lng=es&nrm=iso&tlng=es Background Naja kaouthia, Ophiophagus hannah, Bungarus fasciatus and Calloselasma rhodostoma are four venomous snakes indigenous to Malaysia. In the present study, their proteomic profile by two-dimensional gel electrophoresis (2-DE) have been separated and compared. Results The 2-DE of venoms of the four species snake demonstrated complexity and obvious interspecies differences in proteome profiles. A total of 63 proteins were identified in the four species: C. rhodostoma - 26, N. kaouthia - 16, O. hannah - 15 and B. fasciatus - 6. Conclusions Despite the identifications of major proteins in the four snake species, a large number of protein spots from the 2-DE were unidentified even though the spots displayed high-quality MALDI-TOF-MS spectra. Those identified included phospholipase A2 proteins in all four venoms, long neurotoxins in both cobra species and the C. rhodostoma venom found with the most varied types of peptidases, i.e. metalloproteinase kistomin, halystase and L-amino acid oxidase. <![CDATA[<em>In vitro </em>screening and evaluation of antivenom phytochemicals from <em>Azima tetracantha </em>Lam. leaves against <em>Bungarus caeruleus </em>and <em>Vipera russelli</em>]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S1678-91992014000100318&lng=es&nrm=iso&tlng=es Background : Snakebites are considered a neglected tropical disease that affects thousands of people worldwide. Although antivenom is the only treatment available, it is associated with several side effects. As an alternative, plants have been extensively studied in order to obtain an alternative treatment. In folk medicine, Azima tetracantha Lam. is usually used to treat snakebites. The present study aims to provide a scientific explanation for the use of this plant against snakebite. The extracts of shade dried leaves of A. tetracantha were tested for in vitro inhibitory activity on toxic venom enzymes like phosphomonoesterase, phosphodiesterase, acetylcholinesterase, hyaluronidase etc. from Bungarus caeruleus and Vipera russelli venoms. Results : The ethylacetate extract rendered a significant inhibitory effect on the phosphomonoesterase, phosphodiesterase, phospholipase A2 and acetylcholinesterase enzymes. Conclusions : The present study suggests that ethylacetate extract of A. tetracantha leaves possesses compounds that inhibit the activity of toxic enzymes from Bungarus caeruleus and Vipera russelli venom. Further pharmacological and in vivo studies would provide evidence that this substance may lead to a potential treatment against these venoms. <![CDATA[Association among <em>H. pylori </em> virulence markers <em>dupA, cagA</em> and <em>vacA</em> in Brazilian patients]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S1678-91992014000100319&lng=es&nrm=iso&tlng=es Background: Only a few Helicobacter pylori-infected individuals develop severe gastric diseases and virulence factors of H. pylori appear to be involved in such clinical outcomes. Duodenal ulcer promoting gene A (dupA) is a novel virulence factor of Helicobacter pylori that is associated with duodenal ulcer development and reduced risk for gastric carcinoma in some populations. The aims of the present study were to determine the presence of dupA gene and evaluate the association among dupA and other virulence factors including cagA and vacA in Brazilian patients. Gastric biopsies were obtained from 205 dyspeptic patients (100 children and 105 adults). DNA was extracted and analyzed for the presence of H. pylori and its virulence factors using the polymerase chain reaction method. Results: Patients with gastritis tested positive for H. pylori more frequently. The dupA gene was detected in 41.5% of them (85/205); cagA gene was found in 98 isolates (47.8%) and vacA genotype s1/m1 in 50.2%, s1/m2 in 8.3%, s2/m2 in 36.6%, s2/m1 in 0.5% and s1/s2/m1/m2 in 4.4%. We also verified a significant association between cagA and dupA genes [p = 0.0003, relative risk (RR) 1.73 and confidence interval [CI] = 1.3-2.3]. The genotypes s1/m1 were also associated with dupA gene (p = 0.0001, RR: 1.72 and CI: 1.3-2.2). The same associations were found when analyzing pediatric and adult groups of patients individually. Conclusion: Ours results suggest that dupA is highly frequent in Brazilian patients and is associated with cagA gene and vacA s1/m1 genotype, and it may be considered an important virulence factor in the development of gastric diseases in adults or children. <![CDATA[Hematology and plasma biochemistry of wild-caught Indian cobra <em>Naja naja </em>(Linnaeus, 1758)]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S1678-91992014000100320&lng=es&nrm=iso&tlng=es Background : Hematology and plasma biochemistry parameters are useful in the assessment and management of snake physiological status. Although reference ranges are readily available for many snake species, they are lacking for most venomous ophidians. We determined hematology and plasma biochemistry reference ranges for the wild-caught Indian cobra, Naja naja. Results : Blood samples, taken from the ventral tail vein, were assessed for erythrocyte count, total leukocyte count, hemoglobin concentration, hematocrit, mean corpuscular volume, mean corpuscular hemoglobin, and mean corpuscular hemoglobin concentration, considering the sex of snakes. Results revealed the erythrocyte numbers (male, 390000 ± 12503.33/mm3 and female, 347500 ± 7505.55/mm3), shapes and the centrally located oval nuclei. Leukocytes were round, circular or disk-shaped, and the mean size was larger in male than female snakes. The maximum number of leukocytes was found to be 11700 ± 100/mm3 in male and 12100 ±200/mm3 in female snakes, and mean values of differential leukocyte count differed statistically between male and female snakes. The total leukocyte levels were found to be higher in female snakes, but the levels of hemoglobin, hematocrit, and MCV values were higher in male snakes. However, the MCH and MCHC values remained higher in female snakes throughout the study period. Mean protein and cholesterol contents differed significantly between male (45.32 ± 1.76 and 3.76 ± 0.06 mg/mL) and female (12.47 ± 0.82 and 4.72 ± 0.2 mg/mL) snakes. Conclusions : In conclusion, monitoring snake hematological and biochemical parameters can serve as a means to evaluate the physiological and health status of N. naja populations, which may be a useful indicator of their environmental status. <![CDATA[Clinical aspects of envenomation caused by <em>Tityus obscurus </em> (Gervais, 1843) in two distinct regions of Pará state, Brazilian Amazon basin: a prospective case series]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S1678-91992014000100321&lng=es&nrm=iso&tlng=es Background : Scorpion envenomations are a major public health problem in Brazil, whose most dangerous cases are attributable to the genus Tityus. This study was designed to compare the clinical and demographic features of envenomations by 77tyus obscurus in two areas of the state of Pará located in the Amazon basin. Were compared demographic findings, local and systemic signs and symptoms of human envenomations caused by T. obscurus that occurred in western and eastern areas of the state. Results : Forty-eight patients with confirmed envenomation by T. obscurus were evaluated from January 2008 to July 2011. Most of them came from the eastern region, where male and female patients were present in similar numbers, while males predominated in the west. Median age groups were also similar in both areas. Most scorpion stings took place during the day and occurred significantly more frequently on the upper limbs. The time between the sting and admission to the health center was less than three hours in both areas. Most eastern patients had local manifestations while in the west, systemic manifestations predominated. Local symptoms were similar in both areas, but systemic signs and symptoms were more common in the west. Symptoms frequently observed at the sting site were local and radiating pain, paresthesia, edema, erythema, sweating, piloerection and burning. The systemic manifestations were significantly higher in patients from the west. Futhermore, neurological symptoms such as general paresthesia, ataxia, dysarthria, myoclonus, dysmetria, and electric shock-like sensations throughout the body were reported only by patients from the west. Conclusion : The present study shows that two regions of Para state differ in the clinical manifestations and severity of confirmed envenomation by T. obscurus which suggests a toxicity variation resulting from the diversity of T. obscurus venom in different areas of the Brazilian Amazon basin, and that T. serrulatus antivenom can be successfully used against T. obscurus. <![CDATA[Mild reproductive effects of the <em>Tityus bahiensis </em> scorpion venom in rats]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S1678-91992014000100322&lng=es&nrm=iso&tlng=es Background Scorpion envenoming is a public health problem in Brazil, where Tityus serrulatus and T. bahiensis are considered the most dangerous scorpions. They are well adapted to urbanized environments, and there is an increasing probability of human exposure to these venoms, including during pregnancy. Not much is known about the effects of prenatal exposure to the venom, and no information is available to aid in the rational treatment of victims stung during pregnancy. Thus, this study aimed to investigate whether venom from the scorpion T. bahiensis administered once to pregnant female rats at a dose that causes a moderate envenomation may lead to deleterious effects on the reproductive performance of the dams and on the development of their offspring. This is the first work demonstrating that T. bahiensis venom, when administered experimentally to rats, alters maternal reproductive performance and the morphological development of fetuses. The venom was given to dams on the 5th (GD5) or on the 10th (GD10) gestational day. After laparotomy, on GD21, fetuses and placentas were counted, weighed and externally analyzed. The corpora lutea were counted. The sex and vitality of fetuses were evaluated, and each litter was then randomly divided for visceral or skeletal analyses. Data were analyzed by ANOVA followed by the Tukey-Kramer test and Fisher's exact test. The significance level for all tests was set at p &lt; 0.05. Results : GD5 group presented an increased number of pre-implantation losses. Weight gains in fetuses and placentas were observed in the GD5 and GD10 groups. Weights of the heart and lungs were elevated in GD5 and GD10 and liver weight in GD10. Conclusions : Moderate envenomation by T. bahiensis scorpion venom alters maternal reproductive performance and fetal development. However, these are preliminary results whose causes should be investigated more carefully in future studies. <![CDATA[Evaluation of canine and feline leishmaniasis by the association of blood culture, immunofluorescent antibody test and polymerase chain reaction]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S1678-91992014000100323&lng=es&nrm=iso&tlng=es Background : This study aimed to evaluate the occurrence of Leishmania spp. in dogs and cats from Botucatu, São Paulo state, and Campo Grande, Mato Grosso do Sul state, Brazil, by the association of three diagnostic tests: blood culture in liver infusion tryptose medium, immunofluorescent antibody test and polymerase chain reaction. Fifty blood samples of dogs and cats from the Center for Zoonosis Control in Campo Grande, an area endemic for canine visceral leishmaniasis, were collected randomly, as well as canine and feline blood samples from the Municipal Kennel and Animal Protection Association in Botucatu, currently considered a transmission-free, non-endemic area. Results : Of the 50 dog blood cultures from Botucatu, three (6%) were positive and of the 50 cats, two (4%) were positive. In Campo Grande, 29 dog blood cultures (58%) were positive and all (100%) cats negative by this test. Polymerase chain reaction detected Leishmania spp. in 100% of dog and cat samples from Botucatu but found all the cats from Campo Grande to be negative. On the other hand, 36 dogs from Campo Grande were positive (72%) by the same technique. Immunofluorescent antibody test in Botucatu found 100% of dogs and cats non-reactive, while in Campo Grande, it detected positivity in 32 dogs (64%) and 15 cats (30%). Conclusions : The results show the importance of not only continuous epidemiological surveillance in areas not endemic for leishmaniasis, but also research for accurate diagnosis of this zoonosis. <![CDATA[Epidemiology of bee stings in Campina Grande, Para√≠ba state, Northeastern Brazil]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S1678-91992014000100324&lng=es&nrm=iso&tlng=es Background : The present study aims to investigate the clinical-epidemiological characteristics of bee sting cases recorded between 2007 and 2012 in the city of Campina Grande, Paraiba state, Brazil. Data were collected from the database of the Injury Notification Information System of the Brazilian Ministry of Health.  Results : A total of 459 bee sting cases were retrospectively analyzed. The average annual incidence was 19 cases per 100,000 inhabitants. Cases were distributed in all months of the year, with higher prevalence in September and February. Most victims were men aged between 20 and 29 years. The highest incidence of cases was recorded in urban areas. Victims were stung mainly on the head and torso and received medical assistance predominantly 1 to 3 hours after being stung. The most frequent clinical manifestations were pain, edema and itching. Most cases were classified as mild, and three deaths were reported.  Conclusions : The high incidence of envenomations provoked by bees in Campina Grande suggests that it may be an important risk area for accidents. Since several medical records lacked information, clinical-epidemiological profile of bee sting cases in the studied region could not be accurately determined. The current study provides relevant data for the development of strategies to promote control and prevention of bee stings in this area. Further training for health professionals seems to be necessary to improve their skills in recording clinical-epidemiological information as well as in treating bee sting victims. <![CDATA[Leptospira¬†spp. infection in sheep herds in southeast Brazil]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S1678-91992014000100325&lng=es&nrm=iso&tlng=es Background With the aim of studying Leptospira spp. infection in sheep herds, blood samples and respective kidney and liver fragments were collected from 100 animals from twenty different properties during slaughter at a meat company in the Sorocaba region, São Paulo state, southeast Brazil. The microscopic agglutination test (MAT) was performed with 29 strains of Leptospira spp. To identify the agent in the liver and kidney, 100 samples of each tissue were submitted to culture in Fletcher medium and analyzed by the polymerase chain reaction (PCR) for Leptospira spp. Results MAT detected 23 samples serologically positive for one or more Leptospira spp. serovars and significantly more for Autumnalis. Eight (4%) samples were positive in culture (four kidneys and four livers), corresponding to five animals with positive serology (one animal simultaneously positive for both kidney and liver) and two negatives. PCR detected Leptospira spp. in 14 samples (seven kidneys and seven livers) corresponding to 12 positive animals (two animals simultaneously positive for kidney and liver), of which ten were serologically positive and two negative. Conclusions PCR was faster, more practical and more sensitive than culture for detecting leptospires. The results reinforce the importance of sheep in the epidemiological context of leptospirosis. <![CDATA[Tracking false-negative results in molecular diagnosis: proposal of a triplex-PCR based method for leishmaniasis diagnosis]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S1678-91992014000100326&lng=es&nrm=iso&tlng=es Background : Molecular biological methods have become increasingly relevant to the diagnosis and control of infectious diseases, such as leishmaniasis. Since various factors may affect the sensitivity of PCR assays, including DNA yield and purity, an optimal extraction method is pivotal. Losses of a parasite’s DNA during extraction may significantly impair its detection by PCR and lead to false-negative results. This study proposes a triplex PCR assay targeting the parasite’s DNA, an external control (pUC18) and an internal control (G3PD) for accurate diagnosis of leishmaniasis. Results : Two primer pairs were designed to detect the plasmid pUC18 and a triplex PCR assay targeting theLeishmania braziliensis kinetoplast DNA, the external control and the internal control was standardized. The triplex PCR assay was assessed for its ability to detect the three target DNA fragments simultaneously. PCR products from pUC18 DNA resulted in bands of 368 (P1) and 316 (P2) base pairs (bp). The triplex PCR optimized with the chosen external control system (P1) allowed the simultaneous detection of the internal control (G3PD – 567 bp) as well as of small quantities (10 pg) of the target parasite’s DNA, detected by amplification of a 138 bp product. Conclusions : The new tool standardized herein enables a more reliable interpretation of PCR results, mainly by contributing to quality assurance of leishmaniasis diagnosis. Furthermore, after simple standardization steps, this protocol could be applied to the diagnosis of other infectious diseases in reference laboratories. This triplex PCR enables the assessment of small losses during the DNA extraction process, problems concerning DNA degradation (sample quality) and the detection of L. braziliensis kDNA. <![CDATA[Gene polymorphism of interleukin 1 and 8 in chronic gastritis patients infected with <em>Helicobacter pylori</em>]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S1678-91992014000100327&lng=es&nrm=iso&tlng=es Background : Epidemiological investigations have indicated that Helicobacter pylori induces inflammation in the gastric mucosa regulated by several interleukins. The genes IL1B and IL8 are suggested as key factors in determining the risk of gastritis. The aim of this paper was to evaluate the association of gene polymorphism of interleukin-1 and interleukin-8 with chronic gastrits in H. pylori infected patients. A total of 60 patients underwent endoscopic procedure. Biopsy samples were collected for urease test, histopathological and molecular exams. The DNA of theses samples was extracted for detection of H. pylori and analysis of the genes mentioned above. Patients with gastritis had a higher frequency of H. pylori-positive samples. Results : H. pylori was detected in 30/60 patients (50%) by PCR. As for polymorphism of interleukin 8 (-251) gene we observed a statistical difference when analyzed TA (p = 0.039) and TT (p = 0.047) genotypes. In the IL1B31 there was a statistical difference in TT (p = 0.01) genotype and in theIL1B-511 there wasn’t any statistical difference. Conclusion : Our results suggest a strong correlation between the presence of chronic gastritis and infection byH. pylori and that IL1B-31TT and IL8-251TT genotypes appear to act as protective factors againstH. pylori infection while IL8-251TA genotype may comprise a risk factor for infection with this bacterium. <![CDATA[Detection and identification of huwentoxin-IV interacting proteins by biotin-avidin chemistry combined with mass spectrometry]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S1678-91992014000100328&lng=es&nrm=iso&tlng=es Background : Numerous spider toxins are of interest as tools for neurophysiological research or as lead molecules for the development of pharmaceuticals and insecticides. Direct detection and identification of the interacting proteins of a spider toxin are helpful for its action-mechanism analysis and practical application. The present study employed a combinative strategy for the analysis of interacting proteins of huwentoxin-IV (HWTX-IV), a peptidic neurotoxin from the venom of the spider Selenocosmia huwena. Results : HWTX-IV was first lightly labeled with biotin under the optimized mild experimental conditions and the toxin labeled with a single biotin group (monobiotinylated HWTX-IV) was demonstrated by electrophysiological experiments to retain its original bioactivity and was used in combination with far-western blotting to detect its interacting proteins. Comparative experiments indicated that some membrane proteins from rat neuromuscular junction preparations bind to monobiotinylated HWTX-IV after being transferred onto a PVDF membrane from the SDS-gel. With capillary high performance liquid chromatography-tandem mass spectrometry, several membrane proteins with which HWTX-IV potentially interacted were identified from the preparations and then bioinformatically analyzed. Conclusions : This work has provided not only a new insight into the action mechanism of HWTX-IV but also a reference technology for the relevant researches. <![CDATA[<em>In vitro</em> antibacterial effect of wasp (<em>Vespa orientalis</em>) venom]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S1678-91992014000100329&lng=es&nrm=iso&tlng=es Background The emergence of antibacterial resistance against several classes of antibiotics is an inevitable consequence of drug overuse. As antimicrobial resistance spreads throughout the globe, new substances will always be necessary to fight against multidrug-resistant microorganisms. Venoms of many animals have recently gained attention in the search for new antimicrobials to treat infectious diseases. Thefore, the present study aimed to study the antibacterial effects of wasp (Vespa orientalis) crude venom. Two gram-positive bacteria (Staphylococcus aureus and Bacillus subtilis) and two gram-negative ones (Escherichia coli and Klesiella pneumonia) were compared for their sensitivity to the venom by determining the inhibition zone (Kirby-Bauer method) and minimum inhibitory concentration (MIC). A microbroth kinetic system based on continuous monitoring of changes in the optical density of bacterial growth was also used for determination of antimicrobial activity. Results The venom exhibited a well-recognized antimicrobial property against the tested bacterial strains. The inhibition zones were determined to be 12.6, 22.7, 22.4 and 10.2 mm for S. aureus, B. subtilis,E. coli and K. pneumonia, respectively. The corresponding MIC values were determined to be 64, 8, 64 and 128 μg/mL, respectively. The MIC50 and MIC90 values of the venom were respectively determined to be 63.6 and 107 μg/mL for S. aureus, 4.3 and 7.0 μg/mL for B. subtilis, 45.3 and 65.7 μg/mL for E. coli and 74.4 and 119.2 μg/mL for K. pneumonia. Gram-positive bacteria were generally more sensitive to the venom than gram-negative ones. Conclusions Results revealed that the venom markedly inhibits the growth of both gram-positive and gram-negative bacteria and could be considered a potential source for developing new antibacterial drugs. <![CDATA[Molecular cloning of a hyaluronidase from <em>Bothrops pauloensis</em> venom gland]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S1678-91992014000100330&lng=es&nrm=iso&tlng=es Background Hyaluronate is one of the major components of extracellular matrix from vertebrates whose breakdown is catalyzed by the enzyme hyaluronidase. These enzymes are widely described in snake venoms, in which they facilitate the spreading of the main toxins in the victim’s body during the envenoming. Snake venoms also present some variants (hyaluronidases-like substances) that are probably originated by alternative splicing, even though their relevance in envenomation is still under investigation. Hyaluronidases-like proteins have not yet been purified from any snake venom, but the cDNA that encodes these toxins was already identified in snake venom glands by transcriptomic analysis. Herein, we report the cloning and in silico analysis of the first hyaluronidase-like proteins from a Brazilian snake venom. Methods The cDNA sequence of hyaluronidase was cloned from the transcriptome of Bothrops pauloensisvenom glands. This sequence was submitted to multiple alignment with other related sequences by ClustalW. A phylogenetic analysis was performed using MEGA 4 software by the neighbor joining (NJ) method. Results The cDNA from Bothrops pauloensis venom gland that corresponds to hyaluronidase comprises 1175 bp and codifies a protein containing 194 amino acid residues. The sequence, denominated BpHyase, was identified as hyaluronidase-like since it shows high sequence identities (above 83%) with other described snake venom hyaluronidase-like sequences. Hyaluronidases-like proteins are thought to be products of alternative splicing implicated in deletions of central amino acids, including the catalytic residues. Structure-based sequence alignment of BpHyase to human hyaluronidase hHyal-1 demonstrates a loss of some key secondary structures. The phylogenetic analysis indicates an independent evolution of BpHyal when compared to other hyaluronidases. However, these toxins might share a common ancestor, thus suggesting a broad hyaluronidase-like distribution among venomous snakes. Conclusions This work is the first report of a cDNA sequence of hyaluronidase from Brazilian snake venoms. Moreover, the in silico analysis of its deduced amino acid sequence opens new perspectives about the biological function of hyaluronidases-like proteins and may direct further studies comprising their isolation and/or recombinant production, as well as their structural and functional characterization. <![CDATA[Molecular detection of <em>Leishmania</em> spp. in road-killed wild mammals in the Central Western area of the State of São Paulo, Brazil]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S1678-91992014000100331&lng=es&nrm=iso&tlng=es Background Road-killed wild animals have been classified as sentinels for detecting such zoonotic pathogens asLeishmania spp., offering new opportunities for epidemiological studies of this infection. Methods This study aimed to evaluate the presence of Leishmania spp. and Leishmania chagasi DNA by PCR in tissue samples (lung, liver, spleen, kidney, heart, mesenteric lymph node and adrenal gland) from 70 road-killed wild animals. Results DNA was detected in tissues of one Cavia aperea (Brazilian guinea pig), five Cerdocyon thous(crab-eating fox), one Dasypus septemcinctus (seven-banded armadillo), two Didelphis albiventris(white-eared opossum), one Hydrochoerus hydrochoeris (capybara), two Myrmecophaga tridactyla(giant anteater), one Procyon cancrivorus (crab-eating raccoon), two Sphiggurus spinosus(porcupine) and one Tamandua tetradactyla (lesser anteater) from different locations in the Central Western part of São Paulo state. The Leishmania chagasi DNA were confirmed in mesenteric lymph node of one Cerdocyon thous. Results indicated common infection in wild animals. Conclusions The approach employed herein proved useful for detecting the environmental occurrence ofLeishmania spp. and L. chagasi, as well as determining natural wild reservoirs and contributing to understand the host-parasite interaction. <![CDATA[A case of stings in humans caused by <em>Sclerodermus </em> sp. in Italy]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S1678-91992014000100503&lng=es&nrm=iso&tlng=es In the last years, stings of Sclerodermus species in humans have been sporadically reported in Italy. In order to draw attention to these bethylid wasps of medical importance, we report the case of documented Sclerodermus sp. stings on the dorsum, abdomen, arms, and thighs of a 40-year-old man and his wife. The sting sites developed raised red itchy rash. The source of environmental contamination was identified in a worm-eaten sofa purchased from a used furniture dealer and placed in the living room about a month and half earlier. The lesions on the man and his wife rapidly healed within 3 to 4 days once they left the house and treatment for the lesions was instituted. Physicians, dermatologists, medical and public health entomologists, as well as specific categories of workers should be aware of the risk of exposure to Sclerodermus stings. <![CDATA[Presumptive thrombotic thrombocytopenic purpura following a hump-nosed viper (<em>Hypnale hypnale</em>) bite: a case report]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S1678-91992014000100504&lng=es&nrm=iso&tlng=es Hump-nosed viper bites are frequent in southern India and Sri Lanka. However, the published literature on this snakebite is limited and its venom composition is not well characterized. In this case, we report a patient with thrombotic thrombocytopenic purpura-like syndrome following envenoming which, to the best of our knowledge, has not been reported in the literature before. A 55-year-old woman from southern Sri Lanka presented to the local hospital 12 hours after a hump-nosed viper (Hypnale hypnale) bite. Five days later, she developed a syndrome that was characteristic of thrombotic thrombocytopenic purpura with fever, thrombocytopenia, microangiopathic hemolysis, renal impairment and neurological dysfunction in the form of confusion and coma. Her clinical syndrome and relevant laboratory parameters improved after she was treated with therapeutic plasma exchange. We compared our observations on this patient with the current literature and concluded that thrombotic thrombocytopenic purpura is a theoretically plausible yet unreported manifestation of hump-nosed viper bite up to this moment. This study also provides an important message for clinicians to look out for this complication in hump-nosed viper bites since timely treatment can be lifesaving. <![CDATA[Seroprevalence of flaviviruses antibodies in water buffaloes <em>(Bubalus bubalis) </em>in Brazilian Amazon]]> http://www.scielo.br/scielo.php?script=sci_arttext&pid=S1678-91992014000100600&lng=es&nrm=iso&tlng=es Background The state of Pará encompasses 26% of Brazilian Amazon where an enormous diversity of arboviruses has been found. This study aimed to assess the prevalence and distribution of hemagglutination-inhibition antibodies against antigens of six Flavivirus (yellow fever virus, Ilheus virus, Saint Louis encephalitis virus, Cacipacore virus, Bussuquara virus and Rocio virus) in water buffaloes in Pará state, Brazil. The prevalence of antibodies in these farm animals is important to determine the circulating arboviruses. Findings All investigated arboviruses were detected in the species studied and our results indicate that water buffaloes are susceptible to Flavivirus infection. Furthermore, there is solid evidence of active circulation of these viruses in the Brazilian Amazon. Conclusions Water buffaloes showed higher prevalence of heterotypic antibody reactions and we hypothesized that they can serve as sentinels to detect the movement of such arboviruses in the Brazilian Amazon.