A proteinase inhibitor from Caesalpinia echinata (pau-brasil) seeds for plasma kallikrein, plasmin and factor XIIa

Caesalpinia echinata is a tree belonging to the Leguminosae family. The red color of the trunk, looking like burning wood ('brasa' in Portuguese), is the origin of the name Brazil. Seeds of leguminous plants contain high amounts of serine proteinase inhibitors that can affect different biological processes. Here we show that a protein isolated from seeds of C. echinata is able to inhibit enzymes that participate in blood coagulation and fibrinolysis. This inhibitor (CeKI) was purified to homogeneity by ion exchange and reversed-phase chromatography. SDS-PAGE indicated a single polypeptide chain with a molecular mass of 20 kDa. CeKI inhibits human plasma kallikrein ( K i =3.1 nM), plasmin ( K i =0.18 nM), factor XIIa ( K i =0.18 nM), trypsin ( K i =21.5 nM) and factor Xa ( K i =0.49 mM). CeKI inhibited kinin release from highmolecular- mass kininogen by kallikrein in vitro . The N-terminal sequence, determined by automatic Edman degradation, identified the inhibitor as a member of the Kunitz family. The secondary structure, determined by circular dichroism, is mainly a random coil followed by beta-sheet structure. The action of CeKI on enzymes of the blood-clotting intrinsic pathway was confirmed by prolongation of the activated partial thromboplastin time.     

TNF-alpha, H2O2 and NO response of peritoneal macrophages to Yersinia enterocolitica O:3 derivatives

In this study, the effect of Yersinia derivatives on nitric oxide (NO), hydrogen peroxide (H2O2) and tumor necrosis factor-alpha (TNF-alpha) production by murine peritoneal macrophages was investigated. Addition of lipopolysaccharide (LPS) to the macrophage culture resulted in NO production that was dose dependent. On the other hand, bacterial cellular extract (CE) and Yersinia outer proteins (Yops) had no effect on NO production. The possible inhibitory effect of Yops on macrophage cultures stimulated with LPS was investigated. Yops partially inhibited NO production (67.4%) when compared with aminoguanidine. The effects of Yersinia derivatives on H2O2 production by macrophages were similar to those on NO production. LPS was the only derivative that stimulated H2O2 release in a dose-dependent manner. All Yersinia derivatives provoked the production of TNF-alpha, but LPS had the strongest effect, as observed for NO production. CE and Yops stimulated TNF-alpha production to a lesser extent than LPS. The results indicate the possibility that in vivo Yops may aid the evasion of the bacteria from the host defense mechanism by impairing the secretion of NO by macrophages.     

Assessment of the serodiagnostic potential of nine novel proteins from Mycobacterium tuberculosis

To identify antigens that would improve the accuracy of serological diagnosis of active tuberculosis, we cloned the genes encoding nine potentially immunogenic secreted or surface-associated proteins of Mycobacterium tuberculosis. Recombinant proteins were reacted with sera from HIV-negative individuals with extrapulmonary tuberculosis (EP-TB) or HIV-positive individuals with pulmonary tuberculosis (TBH). Specific and high level antibody responses were obtained for four recombinant proteins, of which antigen GST-822 was recognized by 60% of EP-TB and 42% of TBH and antigen MBP-506 was recognized by 45% of EP-TB and 61% of TBH. These results suggest that these proteins are strong candidates as subunits in a polyvalent serodiagnostic test.     

Primary sequence determination of a Kunitz inhibitor isolated from Delonix regia seeds

A serine proteinase inhibitor was purified from Delonix regia seeds a Leguminosae tree of the Caesalpinioideae subfamily. The inhibitor named DrTI, inactivated trypsin and human plasma kallikrein with K(i )values 2.19x10(-8) M and 5.25 nM, respectively. Its analysis by SDS-PAGE 10-20% showed that the inhibitor is a protein with a single polypeptide chain of M(r) 22 h Da. The primary sequence of the inhibitor was determined by Edman degradation, thus indicating that it contained 185 amino acids and showed that it belongs to the Kunitz type family; however, its reactive site did not contain Arg or Lys at the putative reactive site (position 63, SbTI numbering) or it was displaced when compared to other Kunitz-type inhibitors.     

Can molecular data place each neotropical monkey in its own branch?

Four different DNA datasets, representative of all extant neotropical primate genera, were tandemly aligned, comprising some 6,763 base pairs (bp) with 2,086 variable characters and 674 informative sites. Maximum Parsimony, Maximum Likelihood and Neighbor-Joining analyses suggested three monophyletic families (Atelidae, Pitheciidae and Cebidae) that emerged almost at the same time during primate radiation. Combined molecular data showed congruent branching inside the atelid clade, placing Alouatta as the most basal lineage followed by Ateles and a more derived branch including Brachyteles and Lagothrix as sister groups. In the Pitheciidae, Callicebus was the most basal lineage with respect to Pithecia and to the more derived sister groups (Cacajao and Chiropotes). Conjoint analysis strongly supported the monophyly of the Cebidae, grouping Aotus, Cebus and Saimiri with the small callitrichines. Within callitrichines, Cebuella merged with Callithrix, Callimico appeared as a sister group of Callithrix/Cebuella, Leontopitecus as a sister group of the previous clade, and Saguinus was the earliest callitrichine offshoot. Two major points remained to be clarified in platyrrhine phylogeny: (i) the exact branching pattern of Aotus, Cebus, Saimiri and the callitrichines, and (ii), which two of these three families (Atelidae, Pitheciidae and Cebidae) are more closely related to one another.     

Parasitism rate of Myzus persicae (Sulzer) by Diaeretiella rapae (McIntosh) in the presence of an alternative,resistant host

The aphids Lipaphis pseudobrassicae (Davis) and Myzus persicae (Sulzer) (Hemiptera: Aphididae) are important Brassicaceae pests, occurring worldwide and causing significant damage to crops. Interspecific variations in the resistance to natural enemies can potentially impact the interaction among aphid populations. Here we evaluated the hypothesis of associational resistance by determining if the presence of resistant aphids (L. pseudobrassicae) reduces the rate of parasitism by Diaeretiella rapae (McIntosh) on non-resistant aphids (M. persicae). The experiment was conducted using collard green plants infested with M. persicae and L. pseudobrassicae either resistant or susceptible to D. rapae. The percentage of parasitism by D. rapae was greater on L. pseudobrassicae in the susceptible than in the resistant treatment, but parasitism rates on M. persicae did not differ between the treatments. There was no difference in average growth rate between M. persicae and susceptible L. pseudobrassicae populations, but resistant L. pseudobrassicae had greater growth rate than M. persicae. These results suggest that over a short period of time the presence of resistant L. pseudobrassicae does not affect the rate of parasitism by D. rapae on M. persicae.     

Abundance of invasive peacock bass increases with water residence time of reservoirs in southeastern Brazil

Neotropical freshwater ecosystems are experiencing a great expansion in the number of invasive species, which is especially alarming since this region harbours 30% of the world’s fish biodiversity with high levels of endemism. We aimed to evaluate the main predictors of peacock basses (Cichla spp.) abundance outside their native range, which are the Amazon and Tocantins-Araguaia river basins. We used multivariate ordination techniques and multimodel inference to analyse peacock basses abundance in twelve reservoirs of the Paraíba do Sul river basin, southeastern Brazil. Interestingly, reservoirs at higher (southernmost) latitudes, located in more populated areas, had higher water temperature and lower turbidity, due to increased water residence time, and these three variables were also positively correlated with abundance of this warm-water invasive fish. Habitat structure was less important in explaining peacock basses abundance, which was not significantly related to biotic factors (fish species richness and time since peacock basses introduction). We hypothesize that the observed effects of reservoir management on limnological features and peacock bass abundance, particularly water residence time (as a mediator of temperature and turbidity), may apply to other Neotropical basins and could influence the impact of this invader.     

Lethal Effect and Behavioral Responses of Leaf-Cutting Ants to Essential Oil of <Emphasis Type="Italic">Pogostemon cablin</Emphasis> (Lamiaceae) and Its Nanoformulation

Leaf-cutting ants belonging to the genus Atta (Formicidae: Myrmicinae) are important pests in agricultural and forest environments. In the present study, we evaluated the formicidal activity of the essential oil of Pogostemon cablin and its nanoformulation on the leaf-cutting ants: Atta opaciceps (Borgmeier, 1939), Atta sexdens (Linnaeus, 1758), and Atta sexdens rubropilosa Forel, 1908. The nanoformulation was developed by magnetic stirring using polyoxyethylene (36%), pure ethanol (36%), essential oil of P. cablin (18%), and water (10%). Bioassays of acute toxicity by fumigation and behavioral bioassays in treated arenas, with and without choice, were performed. The essential oil of P. cablin and its nanoformulation demonstrated efficient insecticidal activity and irritability to ant species. The concentration required to kill 50% of workers varied from 1.06 to 2.10 μL L^?1, with a mean time to death of less than or equal to 42 h. The essential oil of P. cablin and its nanoformulation reduced the displacement and velocity speed of the workers of A. opaciceps and A. sexdens rubropilosa in totally treated arenas. In the bioassays with choices, the three species of ants walked less and at a greater speed on the treated side of arena. This work demonstrates the potential of the essential oil of P. cablin and its nanoformulation to the generation of new formicidal products.     

Absence of cellular damage in tropical newly hatched sharks (<Emphasis Type="Italic">Chiloscyllium plagiosum</Emphasis>) under ocean acidification conditions

Sharks have maintained a key role in marine food webs for 400 million years and across varying physicochemical contexts, suggesting plasticity to environmental change. In this study, we investigated the biochemical effects of ocean acidification (OA) levels predicted for 2100 (pCO₂ ~?900 μatm) on newly hatched tropical whitespotted bamboo sharks (Chiloscyllium plagiosum). Specifically, we measured lipid, protein, and DNA damage levels, as well as changes in the activity of antioxidant enzymes and non-enzymatic ROS scavengers in juvenile sharks exposed to elevated CO₂ for 50 days following hatching. Moreover, we also assessed the secondary oxidative stress response, i.e., heat shock response and ubiquitin levels. Newly hatched sharks appear to cope with OA-related stress through a range of tissue-specific biochemical strategies, specifically through the action of antioxidant enzymatic compounds. Our findings suggest that ROS-scavenging molecules, rather than complex enzymatic proteins, provide an effective defense mechanism in dealing with OA-elicited ROS formation. We argue that sharks’ ancient antioxidant system, strongly based on non-enzymatic antioxidants (e.g., urea), may provide them with resilience towards OA, potentially beyond the tolerance of more recently evolved species, i.e., teleosts. Nevertheless, previous research has provided evidence of detrimental effects of OA (interacting with other climate-related stressors) on some aspects of shark biology. Moreover, given that long-term acclimation and adaptive potential to rapid environmental changes are yet experimentally unaccounted for, future research is warranted to accurately predict shark physiological performance under future ocean conditions.     

Genomic selection prediction models comparing sequence capture and SNP array genotyping methods

The successful application of genomic selection (GS) approaches is dependent on genetic makers derived from high-throughput and low-cost genotyping methods. Recent GS studies in trees have predominantly relied on SNP arrays as the source of genotyping, though this technology has a high entry cost. The recent development of alternative genotyping platforms, tailored to specific species and with low entry cost, has become possible due to advances in next-generation sequencing and genome complexity reduction methods such as sequence capture. However, the performance of these new platforms in GS models has not yet been evaluated, or compared to models developed from SNP arrays. Here, we evaluate the impact of these genotyping technologies on the development of GS prediction models for a Eucalyptus breeding population composed of 739 trees phenotyped for 13 wood quality and growth traits. Genotyping data obtained with both methods were compared for linkage disequilibrium, minor allele frequency, and missing data. Phenotypic prediction methods RR-BLUP and BayesB were employed, while predictive ability using cross validation was used to evaluate the performance of GS models derived from the different genotyping platforms. Differences in linkage disequilibrium patterns, minor allele frequency, missing data, and marker distribution were detected between sequence capture and SNP arrays. However, RR-BLUP and BayesB GS models resulted in similar predictive abilities. These results demonstrate that both genotyping methods are equivalent for genomic prediction of the traits evaluated. Sequence capture offers an alternative for species where SNP arrays are not available, or for when the initial development cost is too high.