Cloning of the replication region on the bacteriocinogenic plasmid pRJ9 of Staphylococcus aureus

Abstract A 5.8-kb Cla I fragment of pRJ9, a bacteriocinogenic plasmid of Sphylococcus aureus , was cloned in the unique Cla I site of pRJ5. The recombinant plasmid obtained, pRJ23, failed to confer bacteriocin production and immunity to bacteriocin on host cells. The cloned fragment was shown to contain the complete replicon of pRJ9. Attempts to clone the 4.4-kb Cla I fragment of pRJ9 were unsuccessful, apparently due to the inactivation of the basic replicon of the cloning vector. Therefore, plasmid pRJ5 cut at its Cla I site appears to be a suitable vector for cloning replication regions of plasmids that cab replicate in S. aureus .     

Effect of vitamin D3 on behavioural and biochemical parameters in diabetes type 1‐induced rats

Diabetes is associated with long‐term complications in the brain and reduced cognitive ability. Vitamin D₃ (VD₃) appears to be involved in the amelioration of hyperglycaemia in streptozotocin (STZ)‐induced diabetic rats. Our aim was to analyse the potential of VD₃ in avoiding brain damage through evaluation of acetylcholinesterase (AChE), Na⁺K⁺‐adenosine triphosphatase (ATPase) and delta aminolevulinate dehydratase (δ‐ALA‐D) activities and thiobarbituric acid reactive substance (TBARS) levels from cerebral cortex, as well as memory in STZ‐induced diabetic rats. Animals were divided into eight groups (n = 5): control/saline, control/metformin (Metf), control/VD₃, control/Metf + VD₃, diabetic/saline, diabetic/Metf, diabetic/VD₃ and diabetic/Metf + VD₃. Thirty days after treatment, animals were submitted to contextual fear‐conditioning and open‐field behavioural tests, after which they were sacrificed and the cerebral cortex was dissected. Our results demonstrate a significant memory deficit, an increase in AChE activity and TBARS levels and a decrease in δ‐ALA‐D and Na⁺K⁺‐ATPase activities in diabetic rats when compared with the controls. Treatment of diabetic rats with Metf and VD₃ prevented the increase in AChE activity when compared with the diabetic/saline group. In treated diabetic rats, the decrease in Na⁺K⁺‐ATPase was reverted when compared with non‐treated rats, but the increase in δ‐ALA‐D activity was not. VD₃ prevented diabetes‐induced TBARS level and improved memory. Our results show that VD₃ can avoid cognitive deficit through prevention of changes in important enzymes such as Na⁺K⁺‐ATPase and AChE in cerebral cortex in type 1 diabetic rats. Copyright © 2014 John Wiley & Sons, Ltd.     

Moth pollination of woody species in the Cerrados of Central Brazil: a case of so much owed to so few?

We present a survey of moth pollination in woody species of the Cerrado of Central Brazil. Although with the exception of Roupala montana (which has simple polysepalous flowers arranged in dense cymes) all moth pollinated species in this community have tubular flowers, or a pseudo-tube formed from a single folded petal in Qualea grandiflora, settling moth flowers (tube less than 15 mm) vs. hawkmoth flowers (tube more than 50 mm) are markedly different in size. Moths visit some 20 woody Cerrado species, but they are probably effective pollinators of only 13 species or ca.14% of the woody taxa studied, and even in these latter species they are often very sparse visitors. Nevertheless, it is notable that moths are pollinators for 21% of the 38 most commonly distributed woody Cerrado species. Moreover, the reproductive efficacy of the studied moth pollinated species was similar to that of species pollinated by other kind of vectors.     

Framing the Salmonidae Family Phylogenetic Portrait: A More Complete Picture from Increased Taxon Sampling

Considerable research efforts have focused on elucidating the systematic relationships among salmonid fishes; an understanding of these patterns of relatedness will inform conservation- and fisheries-related issues, as well as provide a framework for investigating evolutionary mechanisms in the group. However, uncertainties persist in current Salmonidae phylogenies due to biological and methodological factors, and a comprehensive phylogeny including most representatives of the family could provide insight into the causes of these difficulties. Here we increase taxon sampling by including nearly all described salmonid species (n = 63) to present a time-calibrated and more complete portrait of Salmonidae using a combination of molecular markers and analytical techniques. This strategy improved resolution by increasing the signal-to-noise ratio and helped discriminate methodological and systematic errors from sources of difficulty associated with biological processes. Our results highlight novel aspects of salmonid evolution. First, we call into question the widely-accepted evolutionary relationships among sub-families and suggest that Thymallinae, rather than Coregoninae, is the sister group to the remainder of Salmonidae. Second, we find that some groups in Salmonidae are older than previously thought and that the mitochondrial rate of molecular divergence varies markedly among genes and clades. We estimate the age of the family to be 59.1 MY (CI: 63.2-58.1 MY) old, which likely corresponds to the timing of whole genome duplication in salmonids. The average, albeit highly variable, mitochondrial rate of molecular divergence was estimated as ∼0.31%/MY (CI: 0.27–0.36%/MY). Finally, we suggest that some species require taxonomic revision, including two monotypic genera, Stenodus and Salvethymus. In addition, we resolve some relationships that have been notoriously difficult to discern and present a clearer picture of the evolution of the group. Our findings represent an important contribution to the systematics of Salmonidae, and provide a useful tool for addressing questions related to fundamental and applied evolutionary issues.     

A Combined Approach for the Assessment of Cell Viability and Cell Functionality of Human Fibrochondrocytes for Use in Tissue Engineering

Temporo-mandibular joint disc disorders are highly prevalent in adult populations. Autologous chondrocyte implantation is a well-established method for the treatment of several chondral defects. However, very few studies have been carried out using human fibrous chondrocytes from the temporo-mandibular joint (TMJ). One of the main drawbacks associated to chondrocyte cell culture is the possibility that chondrocyte cells kept in culture tend to de-differentiate and to lose cell viability under in in-vitro conditions. In this work, we have isolated human temporo-mandibular joint fibrochondrocytes (TMJF) from human disc and we have used a highly-sensitive technique to determine cell viability, cell proliferation and gene expression of nine consecutive cell passages to determine the most appropriate cell passage for use in tissue engineering and future clinical use. Our results revealed that the most potentially viable and functional cell passages were P5–P6, in which an adequate equilibrium between cell viability and the capability to synthesize all major extracellular matrix components exists. The combined action of pro-apoptotic (TRAF5, PHLDA1) and anti-apoptotic genes (SON, HTT, FAIM2) may explain the differential cell viability levels that we found in this study. These results suggest that TMJF should be used at P5–P6 for cell therapy protocols.