Twenty-five years of international exchanges of plant genetic resources facilitated by the CGIAR genebanks: a case study on global interdependence

This article analyses 25 years of data about international movements of plant genetic resources for food and agriculture (PGRFA), facilitated by the gene banks hosted by seven centres of the Consultative Group on International Agricultural Research. It identifies trends in the movements of PGRFA for use in research and development, and describes the diversity of those resources transferred over time. The paper also presents data on the number of countries involved in the global exchanges, analyses their development status and describes their role as providers and/or recipients, providing a picture of the breadth of these global exchanges. We highlight that it is primarily developing and transition economies that have participated in the flows, and that the transferred germplasm has been largely used within their public agricultural research and development programmes. We conclude that, when provided the opportunity of facilitated access, countries will use a wide diversity of germplasm from many other countries, sub-regions and continents as inputs into their agricultural research and development programmes. We highlight the importance of enabling the continuation of the non-monetary benefits from international access to germplasm. We discuss the implications for the process of development and reform of the multilateral system of access and benefit sharing under International Treaty on Plant Genetic Resources for Food and Agriculture.     

High levels of cyclic‐di‐GMP in plant‐associated Pseudomonas correlate with evasion of plant immunity

The plant innate immune system employs plasma membrane‐localized receptors that specifically perceive pathogen/microbe‐associated molecular patterns (PAMPs/MAMPs). This induces a defence response called pattern‐triggered immunity (PTI) to fend off pathogen attack. Commensal bacteria are also exposed to potential immune recognition and must employ strategies to evade and/or suppress PTI to successfully colonize the plant. During plant infection, the flagellum has an ambiguous role, acting as both a virulence factor and also as a potent immunogen as a result of the recognition of its main building block, flagellin, by the plant pattern recognition receptors (PRRs), including FLAGELLIN SENSING2 (FLS2). Therefore, strict control of flagella synthesis is especially important for plant‐associated bacteria. Here, we show that cyclic‐di‐GMP [bis‐(3′‐5′)‐cyclic di‐guanosine monophosphate], a central regulator of bacterial lifestyle, is involved in the evasion of PTI. Elevated cyclic‐di‐GMP levels in the pathogen Pseudomonas syringae pv. tomato (Pto) DC3000, the opportunist P. aeruginosa PAO1 and the commensal P. protegens Pf‐5 inhibit flagellin synthesis and help the bacteria to evade FLS2‐mediated signalling in Nicotiana benthamiana and Arabidopsis thaliana. Despite this, high cellular cyclic‐di‐GMP concentrations were shown to drastically reduce the virulence of Pto DC3000 during plant infection. We propose that this is a result of reduced flagellar motility and/or additional pleiotropic effects of cyclic‐di‐GMP signalling on bacterial behaviour.     

Acellularization-Induced Changes in Tensile Properties Are Organ Specific - An In-Vitro Mechanical and Structural Analysis of Porcine Soft Tissues

Introduction

Though xenogeneic acellular scaffolds are frequently used for surgical reconstruction, knowledge of their mechanical properties is lacking. This study compared the mechanical, histological and ultrastructural properties of various native and acellular specimens.

Materials and Methods

Porcine esophagi, ureters and skin were tested mechanically in a native or acellular condition, focusing on the elastic modulus, ultimate tensile stress and maximum strain. The testing protocol for soft tissues was standardized, including the adaption of the tissue’s water content and partial plastination to minimize material slippage as well as templates for normed sample dimensions and precise cross-section measurements. The native and acellular tissues were compared at the microscopic and ultrastructural level with a focus on type I collagens.

Results

Increased elastic modulus and ultimate tensile stress values were quantified in acellular esophagi and ureters compared to the native condition. In contrast, these values were strongly decreased in the skin after acellularization. Acellularization-related decreases in maximum strain were found in all tissues. Type I collagens were well-preserved in these samples; however, clotting and a loss of cross-linking type I collagens was observed ultrastructurally. Elastins and fibronectins were preserved in the esophagi and ureters. A loss of the epidermal layer and decreased fibronectin content was present in the skin.

Discussion

Acellularization induces changes in the tensile properties of soft tissues. Some of these changes appear to be organ specific. Loss of cross-linking type I collagen may indicate increased mechanical strength due to decreasing transverse forces acting upon the scaffolds, whereas fibronectin loss may be related to decreased load-bearing capacity. Potentially, the alterations in tissue mechanics are linked to organ function and to the interplay of cells and the extracellular matrix, which is different in hollow organs when compared to skin.     

A novel mutation in the mitochondrial tRNAAla gene (m.5636T>C) in a patient with progressive external ophthalmoplegia

We report a heteroplasmic novel mutation m.5636T>C in the mt-tRNA^Ala in a patient with bilateral ptosis and ophthalmoparesis in whom a muscle biopsy showed cytochrome c oxdidase (COX) negative and ragged red fibers. Using laser capture microdissection we have isolated COX negative fibers and COX positive fibers from the muscle of the patient and determined that the mutation load was clearly increased in COX negative muscle fibers. Additionally, the mutated m.5636T nucleotide is conserved in all the mammal and non-mammal species analyzed and might be structurally relevant as it is located in a position involved in the formation of tertiary structure of canonical mitochondrial tRNAs.     

Chromosomal defects in 34 male homosexuals,half of them with HIV antibodies

Most of the research about viral interactions with human chromosomes was done during the sixties and early seventies and very few was performed after the human immunodeficiency virus (HIV) appearance as an epidemic in the eighties. The objective of this work was to estimate if particular chromosomal changes follow the infection of homosexual males by HIV and to determine if the lifestyle, habits, sexual practices, of our sample of male homosexuals predisposes them to chromosomal abnormalities at a higher rate than the background level of cytogenetic damage the general population has. This was a double blinded case-control study, 17 individuals positive for HIV antibodies (HIV+) detected by enzyme-linked immunosorbent assay (ELISA) and confirmed by western blot (WB) were the cases, and 17 individuals negative for HIV antibodies (HIV-) the controls. These men were a very homogeneous population in terms of age, social status, lifestyles, drug abuse, sexual practices and education. Blood was collected between September 1988 and October 1989. Fresh whole blood was cultured in duplicate for 72 hr. Cell harvest followed conventional methods. Once all cell cultures were gathered, the tubes were picked up at random and air dried chromosome preparations were trypsin-giemsa banded (GTG) after overnight incubation at 60 C degrees. The percentage of gaps and breaks these men had was not different from the reported for the general population, nor were there significant difference among both groups (O.R. = 1.8) in items of amount of chromosomal fragility. The distinction among them was at the level of the specific chromosomal sites where the gaps and breaks located, being sites at 2p21 and at 3p21 four times more frequent among HIV+. These probably represent viral modification sites on chromosomes which are known to look like non-staining gaps which are caused by the virus or viral products. This presumption is supported by an earlier report of repeated breaks at 3p21.1, in fact this was the most common lesion site in this study of chromosomal aberrations of male homosexuals and the authors even considered the probability of "a new type of chromosome marker". Furthermore, years later the CKR5 structural gene was mapped to human chromosome 3p21. This gene codes for the chemokine receptor 5 (CKR5) protein which serves as a secondary receptor on CD4+ T lymphocytes for certain strains of HIV-1. It is possible that this gene was being transcribed in HIV+ men and the consequent "staggering" of DNA contributed to the production of gaps and breaks at 3p21.     

Genotoxicity assessment in aquatic environments under the influence of heavy metals and organic contaminants

The genotoxicity of river water and sediment including interstitial water was evaluated by microscreen phage-induction and Salmonella/microsome assays. Different processes used to fractionate the sediment sample were compared using solvents with different polarities. The results obtained for mutagenic activity using the Salmonella/microsome test were negative in the water and interstitial water samples analysed using the direct concentration method. The responses in the microscreen phage-induction assay showed the presence of genotoxic or indicative genotoxic activity for at least one water sample of each site analysed using the same concentration method. Similar results were obtained for interstitial water samples, i.e. absence of mutagenic activity in the Salmonella/microsome test and presence of genotoxic activity in the microscreen phage-induction assay. Metal contamination, as evidenced by the concentrations in stream sediments, may also help explain some of these genotoxic results. Stream sediment organic extracts showed frameshift mutagenic activity in the ether extract detected by Salmonella/microsome assay. The concentrates evaluated by microscreen phage-induction assay identified the action of organic compounds in the non-polar, medium polar and polar fractions. Thus, the microscreen phage-induction assay has proven to be a more appropriate methodology than the Salmonella/microsome test to analyse multiple pollutants in this ecosystem where both organic compounds and heavy metals are present.     

Characterization of enhanced monovalent and bivalent thrombin DNA aptamer binding using single molecule force spectroscopy

Thrombin aptamer binding strength and stability is dependent on sterical parameters when used for atomic force microscopy sensing applications. Sterical improvements on the linker chemistry were developed for high-affinity binding. For this we applied single molecule force spectroscopy using two enhanced biotinylated thrombin aptamers, BFF and BFA immobilized on the atomic force microscopy tip via streptavidin. BFF is a dimer composed of two single-stranded aptamers (aptabody) connected to each other by a complementary sequence close to the biotinylated end. In contrast, BFA consists of a single DNA strand and a complementary strand in the supporting biotinylated part. By varying the pulling velocity in force-distance cycles the formed thrombin-aptamer complexes were ruptured at different force loadings allowing determination of the energy landscape. As a result, BFA aptamer showed a higher binding force at the investigated loading rates and a significantly lower dissociation rate constant, k_off, compared to BFF. Moreover, the potential of the aptabody BFF to form a bivalent complex could clearly be demonstrated.     

Identification of a Bifunctional Maize C- and O-Glucosyltransferase

Flavonoids accumulate in plant vacuoles usually as O-glycosylated derivatives, but several species can also synthesize flavonoid C-glycosides. Recently, we demonstrated that a flavanone 2-hydroxylase (ZmF2H1, CYP93G5) converts flavanones to the corresponding 2-hydroxy derivatives, which are expected to serve as substrates for C-glycosylation. Here, we isolated a cDNA encoding a UDP-dependent glycosyltransferase (UGT708A6), and its activity was characterized by in vitro and in vivo bioconversion assays. In vitro assays using 2-hydroxyflavanones as substrates and in vivo activity assays in yeast co-expressing ZmF2H1 and UGT708A6 show the formation of the flavones C-glycosides. UGT708A6 can also O-glycosylate flavanones in bioconversion assays in Escherichia coli as well as by in vitro assays with the purified recombinant protein. Thus, UGT708A6 is a bifunctional glycosyltransferase that can produce both C- and O-glycosidated flavonoids, a property not previously described for any other glycosyltransferase.     

Metallothionein responses in the Asiatic clam (Corbicula fluminea) after exposure to trivalent arsenic

The main objective of this work was to evaluate arsenic effects on metallothionein (MT) induction by exposing a freshwater Asiatic clam (Corbicula fluminea) to different concentrations of this metalloid. The presence of MT-like proteins was detected by sodium dodecyl sulfate polyacrylamide gel electrophoresis and compared with a standard rabbit MT. In addition, the polarographic response showed good correspondence between standard MT and MT-like curves from C. fluminea, allowing MT quantification. The results show that clams exposed to different concentrations of arsenic are able to induce significant levels of MTs. Although variability was found in MT induction, significant differences in MT levels were found after 28 days of exposure in all treatments in comparison with the controls, suggesting that exposure to arsenic induced MT-like proteins in C. fluminea.