The micelle-bound structure of an antimicrobial peptide derived from the alpha-chain of bovine hemoglobin isolated from the tick Boophilus microplus

Hemoglobin is known to be a source of peptides involved in several functions. The peptide FLSFPTTKTYFPHFDLSHGSAQVKGHGAK (Hb33-61) is a proteolytic product of the bovine hemoglobin alpha-chain found in the gut content of the cattle tick, Boophilus microplus, and it possesses antimicrobial activity. Since in the past we showed that the amidated form of Hb33-61, Hb33-61a, is active against a few Gram-positive bacteria and fungi strains at micromolar concentration [Fogaca et al. (1999) J. Biol. Chem. 274, 25330-25334], we have been prompted to shed more light on its functional and structural features. Here we show that the peptide is able to disrupt the bacterial membrane ofMicrococcus luteus A270. As for its structure, it has a random conformation in water, and it does not interact with zwitterionic micelles. On the other hand, it binds to negatively charged micelles acquiring a finite structural organization. The 3D structure of Hb33-61a bound to SDS micelles exhibits a nonconventional conformation for an antimicrobial peptide. The backbone is characterized by the presence of a beta-turn in the N-terminus and by a beta-turn followed by a alpha-helical stretch in the C-terminus. A hinge, whose spatial organization is stabilized by side-chain-side-chain interactions, joins these two regions. Interestingly, it preserves structural features present in the corresponding segment of the bovine hemoglobin alpha-chain. Hb33-61a does not possess a well-defined amphipathic nature, and H/D exchange experiments show that while the C-terminal region is embedded in the SDS micelle, one face of the N-terminal half is partly exposed to the solvent.     

A tetravalent dengue nanoparticle stimulates antibody production in mice

Background

Dengue is a major public health problem worldwide, especially in the tropical and subtropical regions of the world. Infection with a single Dengue virus (DENV) serotype causes a mild, self-limiting febrile illness called dengue fever. However, a subset of patients experiencing secondary infection with a different serotype progresses to the severe form of the disease, dengue hemorrhagic fever/dengue shock syndrome. Currently, there are no licensed vaccines or antiviral drugs to prevent or treat dengue infections. Biodegradable nanoparticles coated with proteins represent a promising method for in vivo delivery of vaccines.

Findings

Here, we used a murine model to evaluate the IgG production after administration of inactivated DENV corresponding to all four serotypes adsorbed to bovine serum albumin nanoparticles. This formulation induced a production of anti-DENV IgG antibodies (p < 0.001). However, plaque reduction neutralization assays with the four DENV serotypes revealed that these antibodies have no neutralizing activity in the dilutions tested.

Conclusions

Our results show that while the nanoparticle system induces humoral responses against DENV, further investigation with different DENV antigens will be required to improve immunogenicity, epitope specicity, and functional activity to make this platform a viable option for DENV vaccines.     

Distribution and natural infection status of synantrophic triatomines (Hemiptera: Reduviidae), vectors of Trypanosoma cruzi,reveals new epidemiological scenarios for chagas disease in the Highlands of Colombia

IntroductionUpdating the distribution and natural infection status of triatomine bugs is critical for planning, prioritizing, and implementing strategies to control Chagas disease (CD), especially after vector reduction programs. After carrying out a control program, the Department of Boyaca contains the highest number of Colombian municipalities certified by PAHO to be free of intradomiciliary transmission of Trypanosoma cruzi by Rhodnius prolixus. The present work describes the spatial distribution, natural infection (NI), and molecular characterization of T. cruzi in synanthropic triatomines from the Department of Boyaca in 2017 and 2018.Materials and methodsAn entomological survey was conducted in 52 municipalities in Boyaca known to have had previous infestations of triatomine bugs. Insects were collected through active searches carried out by technical personnel from the Secretary of Health and community members using Triatomine Collection Stations (PITs-acronym in Spanish). For evaluation of natural infection, triatomines were identified morphologically and grouped in pools of one to five individuals of the same species collected in the same household. DNA derived from the feces of each pool of insects was analyzed by PCR for the presence of T. cruzi using primers flanking the satellite DNA of the parasite. SL-IR primers were used to differentiate TCI from the other DTUs and to identify different genotypes. The distribution of the collected triatomines was analyzed to determine any vector hotspots using spatial recreation.ResultsA total of 670 triatomine bugs was collected, belonging to five species: Triatoma dimidiata (73.2%), Triatoma venosa (16.7%), Panstrongylus geniculatus (5.7%), Rhodnius prolixus (4.4%), and Panstrongylus rufotuberculatus (0.4%), from 29 of the 52 municipalities. In total, 71.6% of the bugs were collected within houses (intradomiciliary) and the rest around the houses (peridomiciliary). Triatoma dimidiata was the most widely distributed species and had the highest natural infection index (37.8%), followed by T. venosa and P. geniculatus. TcI was the only DTU found, with the TcI Dom genotype identified in 80% of positive samples and TcI sylvatic in the other insects. Spatial analysis showed clusters of T. dimidiata and T. venosa in the northeast and southwest regions of Boyaca.ConclusionsAfter some municipalities were certified free of natural transmission within houses (intradomiciliary transmission) of T. cruzi by R. prolixus, T. dimidiata has become the most prevalent vector present, and represents a significant risk of resurgent CD transmission. However, T. venosa, P. geniculatus, and P. rufotuberculatus also contribute to the increased risk of transmission. The presence of residual R. prolixus may undo the successes achieved through vector elimination programs. The molecular and spatial analysis used here allows us to identify areas with an ongoing threat of parasite transmission and improve entomological surveillance strategies.     

Transovarial passage of Leishmania infantum kDNA in artificially infected Rhipicephalus sanguineus

Phlebotomine sand flies are the only proven biological vectors of Leishmania parasites. However, Rhipicephalus sanguineus ticks have long been suspected to transmit Leishmania infantum in studies carried out in laboratory and natural conditions. In the present study, 5 μl of L. infantum promastigotes (1 × 10⁶ cells per ml) was injected into the hemocel through the coxa I of four engorged females (F1, F2, F3 and F4). Control ticks (F5 and F6) were injected with sterile phosphate-buffered saline (PBS) using the same procedure. Then, these females, their eggs, and the originated larvae were tested by real time polymerase chain reaction (real-time PCR) for the presence of L. infantum kinetoplast DNA (kDNA). Females and eggs were tested after the end of the oviposition period (about 5 weeks post-inoculation) whereas larvae were tested about 4 months after the inoculation of females. All artificially infected females were positive for L. infantum kDNA. In addition, two pools of eggs (one from F2 and other from F4) and four pools of larvae (one from each F1 and F4 and two from F2) were positive for L. infantum kDNA. These results showed, for the first time, the transovarial passage of L. infantum kDNA in R. sanguineus ticks, thus suggesting that the transovarial transmission of L. infantum protozoa in ticks is worth to be investigated.     

Genbank accession #: AF 135190

Plant Molecular Biology -     

Particle image velocimetry (PIV) study of rotating cylindrical filters for animal cell perfusion processes

In the present work, the main fluid flow features inside a rotating cylindrical filtration (RCF) system used as external cell retention device for animal cell perfusion processes were investigated using particle image velocimetry (PIV). The motivation behind this work was to provide experimental fluid dynamic data for such turbulent flow using a high‐permeability filter, given the lack of information about this system in the literature. The results shown herein gave evidence that, at the boundary between the filter mesh and the fluid, a slip velocity condition in the tangential direction does exist, which had not been reported in the literature so far. In the RCF system tested, this accounted for a fluid velocity 10% lower than that of the filter tip, which could be important for the cake formation kinetics during filtration. Evidence confirming the existence of Taylor vortices under conditions of turbulent flow and high permeability, typical of animal cell perfusion RCF systems, was obtained. Second‐order turbulence statistics were successfully calculated. The radial behavior of the second‐order turbulent moments revealed that turbulence in this system is highly anisotropic, which is relevant for performing numerical simulations of this system. © 2012 American Institute of Chemical Engineers Biotechnol. Prog., 2012     

Reduction of the response to sex pheromone in the oriental fruit moth,Grapholita molesta (Lepidoptera: Tortricidae) following successive pheromonal exposures

The effects of prior pheromonal experience upon the pheromone- mediated upwind flight response was examined in the oriental fruit moth, Grapholita molesta(Busck) (Lepidoptera: Tortricidae). Adult male G. molestawere subjected to a parallel series of staggered and repeated pheromonal exposures in a sustained-flight wind tunnel. Levels of response to pheromone in male G. molestasignificantly decreased in a (a) rectilinear function with increased ages of individuals, (b) logarithmic function of successive trials, and (c) steeper logarithmic function of successive trials with increased dosage of sex pheromone. The baseline levels of responding were not affected by either the (a) dosage of sex pheromone, (b) posteclosion ages of individuals for their initial exposures once the main effect of age itself was estimated, (c) elapsed time in hours between trials, or (d) discrete days of testing as integral intervals, disregarding hours within days.     

Comparison of two introduction methods for African elephants (Loxodonta africana)

Managers must consider an animal's potential for aggression when they decide to change or form a captive social group formation. In this study we compared two introduction methods (termed “sequential” and “nonsequential” introductions) in African elephants to assess their effectiveness in managing aggression and minimizing stress. Both introduction methods included four phases: baseline, visual contact, limited tactile contact, and physical introduction. In the sequential introduction, these steps were followed sequentially, and empirical data were considered during decision‐making. In the nonsequential introduction, these steps were not followed sequentially, and decision‐making was based primarily on intuitive assessments by animal managers. Behavioral data and fecal corticoid concentrations were measured throughout both types of introduction. The behavior categories measured included active aggression, passive aggression, submissive behavior, undesirable/stress‐related behavior, and affiliative behavior. While the role of affiliative behavior was surprising, general behavior patterns were characterized by increases in behavior as animals progressed to the next phase of introduction regardless of introduction type. These increases then attenuated over time during each phase. Overall, less behavior was observed during the sequential introduction, as predicted. The data suggest that the sequential introduction managed aggression more effectively. Similar patterns were predicted for undesirable/stress‐related behavior and fecal corticoid concentration. Undesirable/stress‐related behavior was a poor predictor of observed behavior patterns. Although the patterns differed from those predicted, higher concentrations of fecal corticoids were measured during the nonsequential introduction and correlated significantly only with submissive behavior. While more investigation is warranted, the data indicate that the nonsequential introduction brought about an increased physiological response. Overall, the sequential introduction method appeared to manage aggression and stress better than the nonsequential technique. Every introduction is subject to factors that can influence success, such as staff experience, the design of the facility, and the animals' social histories. It is hoped that the rigorous sequential protocol will be a useful tool in the animal manager's “toolbox” for planning and implementing introductions. Applications of this introduction method are also discussed. Zoo Biol 23:109–126, 2004. © 2004 Wiley‐Liss, Inc.