Analysis of the Ontogenetic Variation in the Venom Proteome/Peptidome of Bothrops jararaca Reveals Different Strategies to Deal with Prey

Previous studies have demonstrated that the pharmacological activities displayed by Bothrops jararaca venom undergo a significant ontogenetic shift. Variation in the venom proteome is a well-documented phenomenon; however, variation in the venom peptidome is poorly understood. We report a comparative proteomic and peptidomic analysis of venoms from newborn and adult specimens of B. jararaca and correlate it with the evaluation of important venom features. We demonstrate that newborn and adult venoms have similar hemorrhagic activities, while the adult venom has a slightly higher lethal activity in mice; however, the newborn venom is extremely more potent to kill chicks. The coagulant activity of newborn venom upon human plasma is 10 times higher than that of adult venom. These differences were clearly reflected in their different profiles of SDS-PAGE, gelatin zimography, immunostaining using specific antibodies, glycosylation pattern, and concanavalin A-binding proteins. Furthermore, we report for the first time the analysis of the peptide fraction of newborn and adult venoms by MALDI-TOF mass spectrometry and LC-MS/MS, which revealed different contents of peptides, while the bradykinin potentiating peptides (BPPs) showed rather similar profiles and were detected in the venoms showing their canonical sequences and also novel sequences corresponding to BPPs processed from their precursor protein at sites so far not described. As a result of these studies, we demonstrated that the ontogenetic shift in diet, from ectothermic prey in early life to endothermic prey in adulthood, and in animal size are associated with changes in the venom proteome in B. jararaca species.

A quantitative analysis of the nerve fibres of the acetabular periosteum of dogs

There are many techniques for the treatment of hip dysplasia, and novel research is currently being undertaken in the hope of obtaining more efficient and less traumatic techniques. The denervation of the hip joint capsule is a simple and effective technique that allows recovery of the functional activity of the affected limbs in significantly less time than other techniques. This surgical procedure consists of removing the acetabular periosteum, thus eliminating the nerve fibres with consequent analgesia. The aim of this investigation was to quantify the number of nerve fibres present in different regions of the acetabular periosteum. The knowledge of regional differences is potentially valuable for the refining of the denervation technique of the hip joint capsule. Thirty canine acetabular fragments were used to compare the nerve fibre density of the periosteum. The results showed a significant difference between the mean density of nerve fibres at the cranial and dorsal-lateral portion (approximately 75 fibres/mm(2)) and caudal lateral portion (approximately 60 fibres/mm(2)) of the acetabulum. Those fibres at the pedosteum are almost positioned in a sagittal plane, pointing towards the joint capsule, suggesting the some density in the latter region. These results indicate a new approach to the articular denervation technique, thus obtanining even better results for the treatment of hip dysplasia in dogs.

Cost-effectiveness analysis of universal childhood vaccination against varicella in Brazil

This study conducts a cost-effectiveness analysis of a childhood universal varicella vaccination program in Brazil. An age and time-dependent dynamic model was developed to estimate the incidence of varicella for 30 years. Assuming a single-dose schedule; vaccine efficacy of 85% and coverage of 80%, the program could prevent 74,422,058 cases and 2905 deaths. It would cost R$ 3,178,396,110 and save R$ 660,076,410 to the society and R$ 365,602,305 to the healthcare system. The program is cost-effective (R$ 14,749 and R$ 16,582 per life-year saved under the societal and the healthcare system`s perspective, respectively). The program`s cost-effectiveness is highly sensitive to the vaccine price and number of doses. (C) 2008 Elsevier Ltd. All rights reserved.

Differential diagnosis of oocysts of Hammondia-like organisms of dogs and cats by PCR-RFLP analysis of 70-kilodalton heat shock protein (HSP70) gene

Organisms of the genera Toxoplasma, Hammondia and Neospora, the Hammondia-like organisms, are closely related coccidian with similarly sized oocysts. Therefore, a diagnosis based on microscopy of oocysts in feces is not a method of choice for species identification of these important parasites. In this paper, we present a polymerase chain reaction coupled with restriction fragment length polymorphism (PCR-RFLP) method to differentially diagnose oocysts of Toxoplasma gondii from oocyst of Hammondia hammondi. Another PCR-RFLP was designed to differentiate oocysts of Hammondia heydorni from oocysts of Neospora spp. Both PCR-RFLP are based on nucleotide sequences of the Hsp70 coding gene. In conclusion, we presented two alternative molecular diagnostic assays that can be successfully applied for the differentiation of oocysts of Hammondia-like organisms shed by felids and canids.

Genetic analysis of ticks belonging to the Rhipicephalus sanguineus group in Latin America

Phylogenetic analyses based on mitochondrial 16S rDNA sequences were generated from Rhipicephalus sanguineus group specimens collected in 29 localities among 9 Latin-American countries, plus ticks collected in South Africa, Spain, and Italy. Sequences from Latin America generated six different haplotypes (A, B, C, D, E, and F). Phylogenetic analyses generated trees that segregated our tick sequences into two distinct clades: one is represented by haplotypes A-C, and South African R. sanguineus and Rhipicephalus turanicus ticks; the second clade is represented by haplotypes D-F, and European R. sanguineus and R. turanicus ticks. When haplotypes A-Fare plotted in the Latin America map according to their geographical coordinates, it is clearly seen that haplotypes D-F are restricted to the southern portion of this continent, whereas haplotypes A-C are distributed in areas between northern Mexico and Brazil (except for the extreme south of this last country, where haplotype E was present). Hence, our phylogenetic analyses separated New World specimens of R. sanguineus into two distinct clades, one represented by tropical and subtropical populations (haplotypes A-C), here designated as the `tropical` species. On the other hand, haplotypes D-F are here designated as the `temperate` species because of their distribution in the southern portion of South America. Until recently, it was assumed that the R. sanguineus group was represented by a single species in the New World, namely R. sanguineus. While the present results coupled with recent studies support the presence of at least two species under the taxon R. sanguineus in the New World, they also show that even in the Old World, the taxon R. sanguineus might be represented by more than one species, since our phylogenetic analysis segregated European and South African R. sanguineus ticks into two distinct clades. The same can be applied for Spanish and South African R. turanicus. (C) 2010 Elsevier B.V. All rights reserved.

Comparative analysis of rabies virus isolates from Brazilian canids and bats based on the G gene and G-L intergenic region

Rabies virus (RABV) isolates from two species of canids and three species of bats were analyzed by comparing the C-terminal region of the G gene and the G-L intergenic region of the virus genome. Intercluster identities for the genetic sequences of the isolates showed both regions to be poorly conserved. Phylogenetic trees were generated by the neighbor-joining and maximum parsimony methods, and the results were found to agree between the two methods for both regions. Putative amino acid sequences obtained from the G gene were also analyzed, and genetic markers were identified. Our results suggest that different genetic lineages of RABV are adapted to different animal species in Brazil.

Analysis of marine bivalve shellfish from the fish market in Santos city, Sao Paulo state, Brazil, for Toxoplasma gondii

The aim of this study was to determine if Toxoplasma gondii are present in oysters (Crassostrea rhizophorae) and mussels (Mytella guyanensis) under natural conditions using a bioassay in mice and molecular detection methods. We first compared two standard protocols for DNA extraction, phenol-chloroform (PC) and guanidine-thiocyanate (GT), for both molluscs. A total of 300 oysters and 300 mussels were then acquired from the fish market in Santos city, Sao Paulo state, Brazil, between March and August of 2008 and divided into 60 groups of 5 oysters and 20 groups of 15 mussels. To isolate the parasite, five mice were orally inoculated with sieved tissue homogenates from each group of oysters or mussels. For molecular detection of T. gondii, DNA from mussels was extracted using the PC method and DNA from oysters was extracted using the GT method. A nested-PCR (Polymerase Chain Reaction) based on the amplification of a 155 bp fragment from the B1 gene of T. gondii was then performed. Eleven PCR-RFLP (Restriction Fragment Length Polymorphism) markers, SAG1, SAG2, SAG3, BTUB, GRA6, c22-8, c29-2, L358, PK1, CS3 and Apico, were used to genotype positive samples. There was no isolation of the parasite by bioassay in mice. T. gondii was not detected in any of the groups of mussels by nested-PCR. DNA of T. gondii was apparently detected by nested-PCR in 2 groups of oysters (3.3%). Genotyping of these two positive samples was not successful. The results suggest that oysters of the species C. rhizophorae, the most common species from the coast of Sao Paulo, can filter and retain T. gondii oocysts from the marine environment. Ingestion of raw oysters as a potential transmission source of T. gondii to humans and marine mammals should be further investigated. (C) 2010 Elsevier B.V. All rights reserved.

Complete genome analysis of a rabies virus isolate from Brazilian wild fox

The complete genome sequence of wild-type rabies virus (RABV) isolated from a wild Brazilian hoary fox (Dusicyon sp.), the BR-Pfx1 isolate, was determined and compared with fixed RABV strains. The genome structure and organization of the BR-Pfx1 isolate were composed of 11,924 nt and included the five standard genes of rhabdoviruses. Sequences of mRNA start and stop signals for transcription were highly conserved among all structural protein genes of the BR-Pfx1 isolate. All amino acid residues in the glycoprotein (G) gene associated with pathogenicity were retained in the BR-Pfx1 isolate, while unique amino acid substitutions were found in antigenic region I of the nucleoprotein gene and III of G. These results suggest that although the standard genome structure and organization of the RABV isolate are common between the BR-Pfx1 isolate and fixed RABV strains, the unique amino acid substitutions in functional sites of the BR-Pfx1 isolate may result in different biological characteristics from fixed RABV strains.

An Economic Analysis of the Legal Concept of Trustees' Power of Investment

Authorised trustee status is a legal concept which has economic implications; one of the major implications is that it assists in the direction of investment funds into particular securities and areas of the economy. The concept of authorised trustee status, while attempting to achieve specific outcomes for the beneficiaries of trusts cannot be relied upon to secure these results. Economic analysis of the role of the trustee maintains that this role is one of portfolio manager; a role which is complex but which is explicable in terms of definable procedures and practices. The role of trustee as portfolio manager is one which requires greater financial knowledge than can be assumed is possessed by all trustees. The trustee as portfolio manager is required to maintain a review of decisions make under powers to invest trust assets. A solution to the problem of authorised trustee status is proposed. The solution takes two parts: the first is the adoption of the prudent person approach but with the codification of duties of the trustee and the explicit listing of the factors that a trustee should consider in using the investment powers. The second part of the proposed solution is to link the investment powers of trustees to the best practice features of securities advisers who are now licensed by a regulatory body, the Australian Securities and Investment Commission.