Golden bananas in the field: elevated fruit pro‐vitamin A from the expression of a single banana transgene

Vitamin A deficiency remains one of the world's major public health problems despite food fortification and supplements strategies. Biofortification of staple crops with enhanced levels of pro‐vitamin A (PVA) offers a sustainable alternative strategy to both food fortification and supplementation. As a proof of concept, PVA‐biofortified transgenic Cavendish bananas were generated and field trialed in Australia with the aim of achieving a target level of 20 μg/g of dry weight (dw) β‐carotene equivalent (β‐CE) in the fruit. Expression of a Fe'i banana‐derived phytoene synthase 2a (MtPsy2a) gene resulted in the generation of lines with PVA levels exceeding the target level with one line reaching 55 μg/g dw β‐CE . Expression of the maize phytoene synthase 1 (ZmPsy1) gene, used to develop ‘Golden Rice 2’, also resulted in increased fruit PVA levels although many lines displayed undesirable phenotypes. Constitutive expression of either transgene with the maize polyubiquitin promoter increased PVA accumulation from the earliest stage of fruit development. In contrast, PVA accumulation was restricted to the late stages of fruit development when either the banana 1‐aminocyclopropane‐1‐carboxylate oxidase or the expansin 1 promoters were used to drive the same transgenes. Wild‐type plants with the longest fruit development time had also the highest fruit PVA concentrations. The results from this study suggest that early activation of the rate‐limiting enzyme in the carotenoid biosynthetic pathway and extended fruit maturation time are essential factors to achieve optimal PVA concentrations in banana fruit.     

“If You Tell People That You Had Sex with a Fellow Man,It Is Hard to Be Helped and Treated”: Barriers and Opportunities for Increasing Access to HIV Services among Men Who Have Sex with Men in Uganda

Background

Despite the high HIV prevalence among men who have sex with men (MSM) in sub-Saharan Africa, little is known about their access to HIV services. This study assessed barriers and opportunities for expanding access to HIV services among MSM in Uganda.

Methods

In October-December 2013, a cross-sectional qualitative study was conducted in 12 districts of Uganda. Semi-structured in-depth interviews were conducted with 85 self-identified MSM by snowball sampling and 61 key informants including HIV service providers and policy makers. Data were analysed using manifest content analysis and Atlas.ti software.

Results

Three quarters of the MSM (n = 62, 72.9%) were not comfortable disclosing their sexual orientation to providers and 69 (81.1%) felt providers did not respect MSM. Half (n = 44, 51.8%) experienced difficulties in accessing health services. Nine major barriers to access were identified, including: (i) unwelcoming provider behaviours; (ii) limited provider skills and knowledge; (iii) negative community perceptions towards MSM; (iv) fear of being exposed as MSM; (v) limited access to MSM-specific services; (vi) high mobility of MSM, (vii) lack of guidelines on MSM health services; viii) a harsh legal environment; and ix) HIV related stigma. Two-thirds (n = 56, 66%) participated in MSM social networks and 86% of these (48) received support from the networks to overcome barriers to accessing services.

Conclusions

Negative perceptions among providers and the community present barriers to service access among MSM. Guidelines, provider skills building and use of social networks for mobilization and service delivery could expand access to HIV services among MSM in Uganda.     

Phytochemical differences between Calia secundiflora (Leguminosae) growing at two sites in Mexico

The ecology and quinolizidine alkaloid chemistry of Calia secundiflora (Ortega) Yakovlev growing at two sites in Mexico were compared. At one site (Hidalgo) the vegetation was dominated by Flourensia resinosa and C. secundiflora, at the other site (Queretaro) C. secundiflora and Dodanaea viscosa were dominant. The Hidalgo site had shallower soils with less organic matter, N, P, and CaCO3. Seeds of C. secundiflora from each site accumulated a similar range of quinolizidine alkaloids, but the profile of alkaloids in the leaves and roots were different. The leaves and roots of plants at Hidalgo accumulated a similar range of alkaloids to the seeds with cytisine and/or N-methylcytisine being most abundant, whereas at Queretaro the leaves and roots accumulated lupinine, with other alkaloids being relatively minor constituents. The latter profile has not been reported previously for C. secundiflora.     

Sexual Behavior among Persons Living with HIV in Uganda: Implications for Policy and Practice

Introduction

HIV epidemics are sustained and propagated by new cases of infection which result from transmission from infected persons to uninfected susceptible individuals. People living with HIV (PLHIV) play a critical role in prevention if they adopt safer sexual behaviors. This study estimated the prevalence of and factors associated with safer sexual behaviors among PLHIV seeking care from civil society organizations (CSOs).

Methods

In a cross sectional study PLHIV were interviewed about their sexual practices, use of alcohol, HIV status of their regular sexual partners, desire for more children and about their socio-demographic characteristics. We calculated the proportion of PLHIV who abstained and consistently used condoms in the previous twelve months. Independent associations between safer sex and other variables were estimated using adjusted prevalence ratios (aPR) and their 95% confidence intervals (CI).

Results

Of the 939 PLHIV, 54% (508) were either abstaining or using condoms consistently and 291 (31%) desired more children. The prevalence of consistent condom use among the sexually active was 41.3% (300/731). Consistent condom use was higher among PLHIV who: didn''t use alcohol (aPR 1.30, CI 1.03–1.63); were educated about re-infection with a new strain of HIV (aPR 1.84, CI 1.08–3.12) and had regular sexual partner who was HIV negative (aPR 1.29, CI 1.05–1.57). Prevalence of abstinence was 22.2% (208/939). Abstinence increased with age from 9.4% among PLHIV <25 years to 40.5% among those >50 years. Abstinence was extremely low (2.5%) among PLHIV who were married.

Conclusions

Effective interventions that reduce alcohol consumption among PLHIV are needed to avert HIV transmission, prevent acquisition of new HIV strains and STIs. In addition, strategies are needed to address needs of PLHIV who desire more children.     

Prevalence,Awareness and Control of Hypertension in Uganda

Background

Prevention and control of hypertension are critical in reducing morbidity and mortality attributable to cardiovascular diseases. Awareness of hypertension is a pre-condition for control and prevention. This study estimated the proportion of adults who were hypertensive, were aware of their hypertension and those that achieved adequate control.

Methods

We conducted a community based cross sectional survey among people≥15 years in Buikwe and Mukono districts of Uganda. People had their blood pressure measured and were interviewed about their social-demographic characteristics. Hypertension was defined as systolic blood pressure ≥140 mmHg, or diastolic blood pressure ≥90 mmHg, or previous diagnosis of hypertension. Participants were classified as hypertensive aware if they reported that they had previously been informed by a health professional that they had hypertension. Control of hypertension among those aware was if systolic blood pressure was <140 mmHg and diastolic blood pressure was <90 mmHg.

Results

The age standardized prevalence of hypertension was 27.2% (95% CI 25.9–28.5) similar among females (27.7%) and males (26.4%). Prevalence increased linearly with age, and age effect was more marked among females. Among the hypertensive participants, awareness was 28.2% (95% CI 25.4–31.0) higher among females (37.0%) compared to males (12.4%). Only 9.4% (95% CI 7.5–11.1) of all hypertensive participants were controlled. Control was higher among females (13.2%) compared to males (2.5%).

Conclusion

More than a quarter of the adult population had hypertension but awareness and control was very low. Measures are needed to enhance control, awareness and prevention of hypertension.     

Gene expression study of monocytes/macrophages during early foreign body reaction and identification of potential precursors of myofibroblasts

Foreign body reaction (FBR), initiated by adherence of macrophages to biomaterials, is associated with several complications. Searching for mechanisms potentially useful to overcome these complications, we have established the signaling role of monocytes/macrophages in the development of FBR and the presence of CD34(+) cells that potentially differentiate into myofibroblasts. Therefore, CD68(+) cells were in vitro activated with fibrinogen and also purified from the FBR after 3 days of implantation in rats. Gene expression profiles showed a switch from monocytes and macrophages attracted by fibrinogen to activated macrophages and eventually wound-healing macrophages. The immature FBR also contained a subpopulation of CD34(+) cells, which could be differentiated into myofibroblasts. This study showed that macrophages are the clear driving force of FBR, dependent on milieu, and myofibroblast deposition and differentiation.     

Functional connectivity fMRI of the rodent brain: comparison of functional connectivity networks in rat and mouse

At present, resting state functional MRI (rsfMRI) is increasingly used in human neuropathological research. The present study aims at implementing rsfMRI in mice, a species that holds the widest variety of neurological disease models. Moreover, by acquiring rsfMRI data with a comparable protocol for anesthesia, scanning and analysis, in both rats and mice we were able to compare findings obtained in both species. The outcome of rsfMRI is different for rats and mice and depends strongly on the applied number of components in the Independent Component Analysis (ICA). The most important difference was the appearance of unilateral cortical components for the mouse resting state data compared to bilateral rat cortical networks. Furthermore, a higher number of components was needed for the ICA analysis to separate different cortical regions in mice as compared to rats.     

DNA Barcoding for Identification of ‘Candidatus Phytoplasmas’ Using a Fragment of the Elongation Factor Tu Gene

Background

Phytoplasmas are bacterial phytopathogens responsible for significant losses in agricultural production worldwide. Several molecular markers are available for identification of groups or strains of phytoplasmas. However, they often cannot be used for identification of phytoplasmas from different groups simultaneously or are too long for routine diagnostics. DNA barcoding recently emerged as a convenient tool for species identification. Here, the development of a universal DNA barcode based on the elongation factor Tu (tuf) gene for phytoplasma identification is reported.

Methodology/Principal Findings

We designed a new set of primers and amplified a 420–444 bp fragment of tuf from all 91 phytoplasmas strains tested (16S rRNA groups -I through -VII, -IX through -XII, -XV, and -XX). Comparison of NJ trees constructed from the tuf barcode and a 1.2 kbp fragment of the 16S ribosomal gene revealed that the tuf tree is highly congruent with the 16S rRNA tree and had higher inter- and intra- group sequence divergence. Mean K2P inter−/intra- group divergences of the tuf barcode did not overlap and had approximately one order of magnitude difference for most groups, suggesting the presence of a DNA barcoding gap. The use of the tuf barcode allowed separation of main ribosomal groups and most of their subgroups. Phytoplasma tuf barcodes were deposited in the NCBI GenBank and Q-bank databases.

Conclusions/Significance

This study demonstrates that DNA barcoding principles can be applied for identification of phytoplasmas. Our findings suggest that the tuf barcode performs as well or better than a 1.2 kbp fragment of the 16S rRNA gene and thus provides an easy procedure for phytoplasma identification. The obtained sequences were used to create a publicly available reference database that can be used by plant health services and researchers for online phytoplasma identification.     

Cerebral blood flow assessment with indocyanine green bolus transit detection by near-infrared spectroscopy in the rat

In this study we evaluated the feasibility of measuring cerebral blood flow in rats by monitoring the transit of an indocyanine green bolus through the brain with multiwavelength near-infrared spectroscopy. Different volumes of a 1 mg/ml indocyanine green solution (5, 15, 25, 50 microl) were injected intravenously in the search for an optimal dose. Clear transit curves were obtained with all doses and a blood flow index could easily be determined. The indocyanine green signal obtained with the bolus of 5 microl rapidly returned to baseline and interfered minimally with the haemoglobin and cytochrome oxidase signals. This dose was used in a second study to evaluate the reproducibility of the signal and the effect of hypercapnia. Two groups of rats received 7 repetitive boli of indocyanine green. In one group, 7% CO(2) was added to the gas mixture before the second, fourth and sixth indocyanine green injection. Hypercapnia consistently caused a significant increase in blood flow index, cerebral haemoglobin concentration and O(2)-saturation. In the control group these variables remained stable in time. We conclude that monitoring of the transit of an indocyanine green bolus with multiwavelength near-infrared spectroscopy can be used to assess cerebral blood flow qualitatively in rats in combination with continuous monitoring of brain oxygenation.