Non-lethal method of DNA sampling in euglossine bees supported by mark–recapture experiments and microsatellite genotyping

Non-lethal sampling methods are of great interest for conservation genetic studies to prevent the death of individuals in populations that are threatened or in decline. With this aim, we tested a non-lethal method of partial antennae removal for DNA sampling in two euglossine bee species: Euglossa cordata and Eulaema nigrita. We validated the survival of the individuals through mark–recapture experiments during 16 months. The quality and quantity of the tissue for DNA analysis was verified through amplification and genotyping of nine and eleven microsatellite loci, respectively. Our results from the mark–recapture experiments showed equal recapture rates of individuals with intact and removed antennae (E. cordata χ² = 2.492, df = 1, p = 0.114; E. nigrita χ² = 1.683, df = 1, p = 0.194). Microsatellite loci were successfully genotyped in 97.1 and 97.6 % of the E. cordata and E. nigrita individuals, respectively. Our results validate the feasibility of using antennae tissue for DNA genetic analysis without compromising the survival of individual bees.     

Effect of Postweaning Handling Strategies on Welfare and Productive Traits in Lambs

Postweaning management strategies that include an element of social enrichment may reduce weaning stress and improve welfare and productive performance. We analyzed the effect of postweaning handling strategies on welfare and production traits in lambs. After weaning, 36 lambs were assigned to 3 experimental groups with 12 lambs each (control [C], fattening with gentle human female contact [H], and fattening with 2 adult ewes [E]). The average daily gain (ADG) was estimated. Blood samples were taken, and infrared thermography was used to estimate stress variables. There were significant differences among treatments (in favor of alternative strategies) regarding production and stress variables (cortisol, glucose, and creatine kinase). The results suggest that the lambs handled gently during the fattening were less reactive and better able to modulate their physiological stress. The E group adapted better to acute stress than the C group but was less efficient in modulating chronic stress. Both treatments showed higher slaughter live weights and better ADGs compared with the control. The use of social enrichment at weaning, especially to establish a positive human–nonhuman animal bond, alleviates lamb weaning stress and improves welfare and performance.     

Ribosomal protein S3 selectively affects colon cancer growth by modulating the levels of p53 and lactate dehydrogenase

Molecular Biology Reports - Ribosomal protein S3 (RPS3) is a component of the 40S ribosomal subunit. It is known to function in ribosome biogenesis and as an endonuclease. RPS3 has been shown to be...     

Prevalence of Entamoeba nuttalli infection in wild rhesus macaques in Nepal and characterization of the parasite isolates

We have recently resurrected the name Entamoeba nuttalli Castellani, 1908 for a potentially virulent ameba isolate, P19-061405, obtained from a rhesus macaque in Kathmandu, Nepal. The ameba was morphologically indistinguishable from Entamoeba histolytica/Entamoeba dispar/Entamoeba moshkovskii, but located phylogenetically between E. histolytica and E. dispar. To evaluate the prevalence of E. nuttalli infection in wild rhesus macaques, 112 fecal samples were collected in four locations of the Kathmandu Valley. PCR analysis of DNA extracted from the feces showed positive rates of E. nuttalli, E. dispar, E. histolytica and E. moshkovskii of 51%, 12%, 0% and 0%, respectively. A total of 14 E. nuttalli isolates were obtained from four locations, of which 6 were established as axenic cultures. The sequences of the serine-rich protein gene of E. nuttalli isolates differed among four locations although no differences were found in the composition of sequence motifs. Isoenzyme pattern was analyzed in 8 isolates obtained from three locations. In hexokinase, the mobility of the slower migrating band was located between E. histolytica and E. dispar regardless of the culture conditions. These results demonstrate that E. nuttalli is highly prevalent in wild rhesus macaques in Nepal. Rhesus macaques appear to be one of the natural hosts and heterogeneity of the serine-rich protein gene might be useful for geographical typing of isolates.     

Effect of long-term GHRH and somatostatin administration on GH release and body weight in prepubertal female rats

In order to find a chronic GHRH administration capable of stimulating growth rate without depleting pituitary GH content, prepubertal female rats were subcutaneously (sc) treated with GHRH (1-29)-NH2 and somatostatin (SS). In experiment 1, the rats received sc injections of GHRH and cyclic natural SS for 19 days. In the second study, female rats were continuously treated during 21 days with GHRH, using a slow release pellet, alone or combined with one daily injection of long acting SS (octreotide). In experiment 1, body weight was significantly increased when GHRH was administered at the highest daily dosage (1200 microg/day), accompanied by an slight increment in pituitary GH content. Hypothalamic SS concentrations decreased when GHRH or SS were administered alone whereas the combined treatment with both peptides did not modify this parameter, which suggests the existence of a balance between the chronic actions of both peptides on hypothalamus. In experiment 2, the continuous infusion of GHRH increased plasma GH levels and tended to enhance pituitary GH content. Nevertheless, GHRH effect was not effective enough to increase body weight. By adding one daily injection of SS both GHRH effects on the pituitary gland were abolished. Our study indicates that female rats retain responsiveness to chronic GHRH and SS treatments at both pituitary and hypothalamic levels.     

The Inner Foreskin of Healthy Males at Risk of HIV Infection Harbors Epithelial CD4+ CCR5+ Cells and Has Features of an Inflamed Epidermal Barrier

Male circumcision provides partial protection against multiple sexually transmitted infections (STIs), including HIV, but the mechanisms are not fully understood. To examine potential vulnerabilities in foreskin epithelial structure, we used Wilcoxon paired tests adjusted using the false discovery rate method to compare inner and outer foreskin samples from 20 healthy, sexually active Peruvian males who have sex with males or transgender females, ages 21–29, at elevated risk of HIV infection. No evidence of epithelial microtrauma was identified, as assessed by keratinocyte activation, fibronectin deposition, or parakeratosis. However, multiple suprabasal tight junction differences were identified: 1) inner foreskin stratum corneum was thinner than outer (p = 0.035); 2) claudin 1 had extended membrane-bound localization throughout inner epidermis stratum spinosum (p = 0.035); 3) membrane-bound claudin 4 was absent from inner foreskin stratum granulosum (p = 0.035); and 4) occludin had increased membrane deposition in inner foreskin stratum granulosum (p = 0.042) versus outer. Together, this suggests subclinical inflammation and paracellular transport modifications to the inner foreskin. A setting of inflammation was further supported by inner foreskin epithelial explant cultures secreting higher levels of GM-CSF (p = 0.029), IP-10 (p = 0.035) and RANTES (p = 0.022) than outer foreskin, and also containing an increased density of CCR5+ and CD4+ CCR5+ cells (p = 0.022). Inner foreskin dermis also secreted more RANTES than outer (p = 0.036), and had increased density of CCR5+ cells (p = 0.022). In conclusion, subclinical changes to the inner foreskin of sexually active males may support an inflammatory state, with availability of target cells for HIV infection and modifications to epidermal barriers, potentially explaining the benefits of circumcision for STI prevention.     

Assessing Leukocyte-endothelial Interactions Under Flow Conditions in an Ex Vivo Autoperfused Microflow Chamber Assay

Leukocyte-endothelial interactions are early and critical events in acute and chronic inflammation and can, when dysregulated, mediate tissue injury leading to permanent pathological damage. Existing conventional assays allow the analysis of leukocyte adhesion molecules only after the extraction of leukocytes from the blood. This requires the blood to undergo several steps before peripheral blood leukocytes (PBLs) can be ready for analysis, which in turn can stimulate PBLs influencing the research findings. The autoperfused micro flow chamber assay, however, allows scientists to study early leukocytes functional dysregulation using the systemic flow of a live mouse while having the freedom of manipulating a coated chamber. Through a disease model, the functional expression of leukocyte adhesion molecules can be assessed and quantified in a micro-glass chamber coated with immobilized endothelial adhesion molecules ex vivo. In this model, the blood flows between the right common carotid artery and left external jugular vein of a live mouse under anesthesia, allowing the interaction of native PBLs in the chamber. Real-time experimental analysis is achieved with the assistance of an intravital microscope as well as a Harvard Apparatus pressure device. The application of a flow regulator at the input point of the glass chamber allows comparable physiological flow conditions amongst the experiments. Leukocyte rolling velocity is the main outcome and is measured using the National Institutes of Health open-access software ImageJ. In summary, the autoperfused micro flow chamber assay provides an optimal physiological environment to study leukocytes endothelial interaction and allows researchers to draw accurate conclusions when studying inflammation.     

Indicators measuring the performance of malaria programs supported by the global fund in Asia, progress and the way forward

Introduction

In 2010, the Global Fund provided more than 75% of external international financing for malaria control. The Global Fund uses performance based funding in the grants it finances. This paper analyses the indicators used to measure the performance of Global Fund supported malaria grants in Asia.

Methods

Indicators used in the performance frameworks for all Global Fund supported malaria grants in Asia were retrieved from grant database and grouped into impact, outcome, output and input categories and categorized by service delivery areas. Indicators of each group were compared over rounds. Indicators used in performance frameworks were compared with internationally adopted indicators included in the Monitoring and Evaluation Toolkit developed by the Global Fund and international technical agencies.

Results

Between 2002 and 2010, 1,434 indicators were included in the performance frameworks of the 48 malaria grants awarded in Asia, including 229 impact and 227 outcome indicators, 437 output and 541 input indicators, with an average of 29.9 indicators per grant. The proportion of impact and outcome indicators increased over rounds, with that of input indicators declining from 44.1% in Round 1 to 22.7% in Round 9.

Conclusions

Input indicators, which have predominated the performance frameworks of the Global Fund supported malaria programs in Asia have declined between Rounds 1 and 9. However, increased alignment with internationally adopted indicators included in the Monitoring and Evaluation Toolkit is needed to improve the validity of reported results.     

Identification of selective tubulin inhibitors as potential anti-trypanosomal agents

The potency of a series of sulfonamide tubulin inhibitors against the growth of Trypanosoma brucei (T. brucei), as well as human cancer and primary fibroblast cells were evaluated with the aim of determining whether compounds that selectively inhibit parasite proliferation could be identified. Several compounds showed excellent selectivity against T. brucei growth, and have the potential to be used for the treatment of Human African trypanosomiasis. A T. brucei tubulin protein homology model was built based on the crystal structure of the bovine tubulin. The colchicine-binding domain, which is also the binding site of the tested sulfonamide tubulin inhibitors, showed clear differences between the tubulin structures and presumably explained the selectivity of the compounds.     

Gene variation and genetic differentiation among populations of the solitary mud dauber wasp Trypoxylon (Trypargilum) albitarse Fabricius 1804 (Hymenoptera, Crabronidae)

Trypoxylon is a genus of solitary crabronid wasps whose populationgenetics is poorly known. The purpose of the present study was to investigate thegenetic variation and differentiation among five populations of Trypoxylonalbitarse, a species widely distributed throughout the Neotropics, withrecords from Panama to northern Argentina. Eight species-specific microsatellite lociwere used for genotyping 96 adult wasps (one female per nest) sampled at five sitesin Brazil. The analysis of allelic richness and private alleles indicated highgenetic diversity in the populations sampled. Pairwise comparisons using theF_st and D_est indices revealed significant differentiation for all, but one pair ofpopulations. F_st, D_est, AMOVA and assignment test values pointed to inter-population differentiation.Additionally, the analysis of population structure using Bayesian and PCA methodscharacterized two alternative genetic groups. The Mantel test indicated nocorrelation between genetic and geographic distances. Despite evidence ofconsiderable dispersal capacity for T. albitarse, the data indicatelow to moderate population structuring in this species.