Genetic diversity and patterns of population structure in Creole goats from the Americas.

Biodiversity studies are more efficient when large numbers of breeds belonging to several countries are involved, as they allow for an in-depth analysis of the within- and between-breed components of genetic diversity. A set of 21 microsatellites was used to investigate the genetic composition of 24 Creole goat breeds (910 animals) from 10 countries to estimate levels of genetic variability, infer population structure and understand genetic relationships among populations across the American continent. Three commercial transboundary breeds were included in the analyses to investigate admixture with Creole goats. Overall, the genetic diversity of Creole populations (mean number of alleles = 5.82 ± 1.14, observed heterozygosity = 0.585 ± 0.074) was moderate and slightly lower than what was detected in other studies with breeds from other regions. The Bayesian clustering analysis without prior information on source populations identified 22 breed clusters. Three groups comprised more than one population, namely from Brazil (Azul and Graúna; Moxotó and Repartida) and Argentina (Long and shorthair Chilluda, Pampeana Colorada and Angora-type goat). Substructure was found in Criolla Paraguaya. When prior information on sample origin was considered, 92% of the individuals were assigned to the source population (threshold q ≥ 0.700). Creole breeds are well-differentiated entities (mean coefficient of genetic differentiation = 0.111 ± 0.048, with the exception of isolated island populations). Dilution from admixture with commercial transboundary breeds appears to be negligible. Significant levels of inbreeding were detected (inbreeding coefficient > 0 in most Creole goat populations, P < 0.05). Our results provide a broad perspective on the extant genetic diversity of Creole goats, however further studies are needed to understand whether the observed geographical patterns of population structure may reflect the mode of goat colonization in the Americas.

Genetic relationships among American donkey populations: insights into the process of colonization.

This study presents the first insights into the genetic diversity and structure of the American donkey metapopulation. The primary objectives were to detect the main structural features underlying variability among American donkey populations, identify boundaries between differentiated gene pools, and draw the main colonization pathways since the introduction of donkeys into America in the 15th century. A panel of 14 microsatellite markers was applied for genotyping 350 American donkeys from 13 countries. The genetic structure of this metapopulation was analysed using descriptive statistics and Bayesian model-based methods. These populations were then compared to a database containing information on 476 individuals from 11 European breeds to identify the most likely ancestral donor populations. Results showed the presence of two distinct genetic pools, with confluence of the two in Colombia. The southern pool showed a unique genetic signature subsequent to an older founder event, but lacked any significant influence of modern gene flow from Europe. The northern pool, conversely, may have retained more ancestral polymorphisms and/or have experienced modern gene flow from Spanish breeds. The Andalusian and, to a lesser extent, the Catalan breeds have left a more pronounced footprint in some of the American donkey populations analysed.

Iterative Denoising Accelerated 3D FLAIR Sequence for Hydrops MR Imaging at 3T.

BACKGROUND AND PURPOSE: 3D FLAIR sequences have become the criterion standard for identifying endolymphatic hydrops, but scan time remains an important limitation to their widespread use. Our purpose was to evaluate the diagnostic performance and image quality of an accelerated 3D FLAIR sequence combined with an iterative denoising algorithm. MATERIALS AND METHODS: This was a retrospective study performed on 30 patients with clinical suspicion of endolymphatic hydrops who underwent 3T MR imaging 4 hours after gadolinium injection using two 3D FLAIR sequences. The first (conventional FLAIR) was accelerated with a conventional turbo factor of 187. The second was accelerated with an increased turbo factor of 263, resulting in a 33% scan time reduction (5 minutes 36 seconds versus 8 minutes 15 seconds, respectively). A sequence was reconstructed in-line immediately after the accelerated 3D FLAIR acquisition from the same raw data with iterative denoising (accelerated-FLAIR iterative denoising). The signal intensity ratio image quality score and endolymphatic hydrops diagnosis were evaluated. RESULTS: The mean signal intensity ratio for symptomatic and asymptomatic ears of accelerated-FLAIR iterative denoising was significantly higher than the mean SNR of conventional FLAIR (29.5 versus 19 and 25.9 versus 16.3, P < .001). Compared with the conventional FLAIR sequence, the image-quality score was higher with accelerated-FLAIR iterative denoising (mean image-quality score, 3.8 [SD, 0.4] versus 3.3 [SD, 0.6] for accelerated-FLAIR iterative denoising and conventional FLAIR, respectively, P = .003). There was no significant difference in the diagnosis of endolymphatic hydrops between the 2 sequences. Interreader agreement was good-to-excellent. CONCLUSIONS: The iterative denoising algorithm applied to an accelerated 3D FLAIR sequence for exploration of endolymphatic hydrops enabled significantly reducing the scan time without compromising image quality and diagnostic performance.

Combination of chemical suppression techniques for dual suppression of fat and silicone at diffusion-weighted MR imaging in women with breast implants.

OBJECTIVES: Silicone breast prostheses prove technically challenging when performing diffusion-weighted MR imaging in the breasts. We describe a combined fat and chemical suppression scheme to achieve dual suppression of fat and silicone, thereby improving the quality of diffusion-weighted images in women with breast implants. METHODS: MR imaging was performed at 3.0 and 1.5 T in women with silicone breast implants using short-tau inversion recovery (STIR) fat-suppressed echo-planar (EPI) diffusion-weighted MR imaging (DWI) on its own and combined with the slice-select gradient-reversal (SSGR) technique. Imaging was performed using dedicated breast imaging coils. RESULTS: Complete suppression of the fat and silicone signal was possible at 3.0 T using EPI DWI with STIR and SSGR, evaluated with dedicated breast coils. However, a residual silicone signal was still perceptible at 1.5 T using this combined approach. Nevertheless, a further reduction in silicone signal at 1.5 T could be achieved by employing thinner slice partitions and the addition of the chemical-selective fat-suppression (CHESS) technique. CONCLUSIONS: DWI using combined STIR and SSGR chemical suppression techniques is feasible to eliminate or reduce silicone signal from prosthetic breast implants. KEY POINTS: Breast magnetic resonance imaging (MRI) is frequently needed following breast implants. Unsuppressed signal from silicone creates artefacts on diffusion-weighted MR sequences. Dual fat/chemical suppression can eliminate signal from fat and silicone. STIR with slice selective gradient reversal can suppress fat and silicone signal.

Mechanical response of individual collagen fibrils in loaded tendon as measured by atomic force microscopy.

A precise analysis of the mechanical response of collagen fibrils in tendon tissue is critical to understanding the ultrastructural mechanisms that underlie collagen fibril interactions (load transfer), and ultimately tendon structure-function. This study reports a novel experimental approach combining macroscopic mechanical loading of tendon with a morphometric ultrascale assessment of longitudinal and cross-sectional collagen fibril deformations. An atomic force microscope was used to characterize diameters and periodic banding (D-period) of individual type-I collagen fibrils within murine Achilles tendons that were loaded to 0%, 5%, or 10% macroscopic nominal strain, respectively. D-period banding of the collagen fibrils increased with increasing tendon strain (2.1% increase at 10% applied tendon strain, p<0.05), while fibril diameter decreased (8% reduction, p<0.05). No statistically significant differences between 0% and 5% applied strain were observed, indicating that the onset of fibril (D-period) straining lagged macroscopically applied tendon strains by at least 5%. This confirms previous reports of delayed onset of collagen fibril stretching and the role of collagen fibril kinematics in supporting physiological tendon loads. Fibril strains within the tissue were relatively tightly distributed in unloaded and highly strained tendons, but were more broadly distributed at 5% applied strain, indicating progressive recruitment of collagen fibrils. Using these techniques we also confirmed that collagen fibrils thin appreciably at higher levels of macroscopic tendon strain. Finally, in contrast to prevalent tendon structure-function concepts data revealed that loading of the collagen network is fairly homogenous, with no apparent predisposition for loading of collagen fibrils according to their diameter.

Comparison of a new whole-body continuous-table-movement protocol versus a standard whole-body MR protocol for the assessment of multiple myeloma.

OBJECTIVES: To evaluate a whole body (WB) continuous-table-movement (CTM) MR protocol for the assessment of multiple myeloma (MM) in comparison to a step-by-step WB protocol. METHODS: Eighteen patients with MM were examined at 1.5T using a WB CTM protocol (axial T2-w fs BLADE, T1-w GRE sequence) and a step-by-step WB protocol including coronal/sagittal T1-w SE and STIR sequences as reference. Protocol time was assessed. Image quality, artefacts, liver/spleen assessability, and the ability to depict bone marrow lesions less than or greater than 1 cm as well as diffuse infiltration and soft tissue lesions were rated. Potential changes in the Durie and Salmon Plus stage and the detectability of complications were assessed. RESULTS: Mean protocol time was 6:38 min (CTM) compared to 24:32 min (standard). Image quality was comparable. Artefacts were more prominent using the CTM protocol (P = 0.0039). Organ assessability was better using the CTM protocol (P < 0.001). Depiction of bone marrow and soft tissue lesions was identical without a staging shift. Vertebral fractures were not detected using the CTM protocol. CONCLUSIONS: The new protocol allows a higher patient throughput and facilitates the depiction of extramedullary lesions. However, as long as vertebral fractures are not detectable, the protocol cannot be safely used for clinical routine without the acquisition of an additional sagittal sequence.

Even illumination in total internal reflection fluorescence microscopy using laser light.

In modern fluorescence microscopy, lasers are a widely used source of light, both for imaging in total internal reflection and epi-illumination modes. In wide-field imaging, scattering of highly coherent laser light due to imperfections in the light path typically leads to nonuniform illumination of the specimen, compromising image analysis. We report the design and construction of an objective-launch total internal reflection fluorescence microscopy system with excellent evenness of specimen illumination achieved by azimuthal rotation of the incoming illuminating laser beam. The system allows quick and precise changes of the incidence angle of the laser beam and thus can also be used in an epifluorescence mode.

Sub-100-nanometre resolution in total internal reflection fluorescence microscopy.

Combining total internal reflection fluorescence microscopy with structured illumination allows optical wide-field imaging with sub-100-nanometre resolution. We present a novel objective-launch set-up for standing wave illumination that takes advantage of a tunable transmission diffraction grating and transparent phase shifters actuated by electro-active polymers to control the excitation pattern in three dimensions. Image acquisition is completed in less than 1 s. To reconstruct the extended image spectrum, we apply a new apodization function that results in a lateral resolution of 89 nm for green emission wavelength.

Writing subwavelength-sized structures into aluminium films by thermo-chemical aperture-less near-field optical microscopy.

An optical near-field at the tip of an atomic force microscope probe is utilised to pattern aluminium thin films on glass substrates by photo-thermally induced corrosion in water. Aluminium forms a thin passivating oxide layer when immersed into neutral water at room temperature. Owing to the high energy density of the near-field, the metal below the probe tip can be heated to 100 degrees C due to absorption of the light, which then provokes breakdown of the passivation and metal corrosion. The localised near-field is generated by tip-induced enhancement of an evanescent field originating from a laser beam, that is totally internally reflected at the glass-aluminium-water interface. The process is governed by surface plasmons excited in the aluminium film by the evanescent waves and the field enhancement of the probe tip. Holes of 40 nm diameter and lines below 100 nm width have been written into a 20-nm-thick aluminium film. Applications of the scanning probe lithography process may include the one-step fabrication of point contacts or contact masks for near-field optical lithography and reactive ion etching.

Imaging biological macromolecules by STM: quantitative interpretation of topographs.

Methods are discussed which permit the calibration of x-, y-, z-sensitivities, non-linearities and frequency responses of the scanning device of a scanning tunneling microscope (STM) either by interferometry or directly from STM topographs. A technique is presented to measure the frequency response of the complete STM feedback unit and to derive a maximum speed in z direction which allows one to estimate the maximum scanning speed still permitting one to track surface corrugations. The signal transfer characteristics of a STM are evaluated in a direct comparison with high resolution transmission electron microscopy on an identical specimen area. The various effects of contaminants between tip and specimen and the finite tip radius receive special attention.

Biological structures imaged in a hybrid scanning transmission electron microscope and scanning tunneling microscope.

A hybrid scanning transmission electron microscope (STEM) and scanning tunneling microscope (STM) is described which allows simultaneous imaging of biological structures adsorbed to electron-transparent specimen supports in both modes of scanning microscopy, as demonstrated on uncoated phage T4 polyheads. We further discuss the reproducibility and validity of height data obtained from STM topographs of biomacromolecules and present raw data from topographs of freeze-dried, metal-coated nuclear envelopes from Xenopus laevis oocytes.

Fast contact-mode atomic force microscopy on biological specimen by model-based control.

The dynamic behavior of the piezoelectric tube scanner limits the imaging rate in atomic force microscopy (AFM). In order to compensate for the lateral dynamics of the scanning piezo a model based open-loop controller is implemented into a commercial AFM system. Additionally, our new control strategy employing a model-based two-degrees-of-freedom controller improves the performance in the vertical direction, which is important for high-speed topographical imaging. The combination of both controllers in lateral and vertical direction compensates the three-dimensional dynamics of the AFM system and reduces artifacts that are induced by the systems dynamic behavior at high scan rates. We demonstrate this improvement by comparing the performance of the model-based controlled AFM to the uncompensated and standard PI-controlled system when imaging pUC 18 plasmid DNA in air as well as in a liquid environment.

Scanning tunneling and transmission electron microscopy on identical areas of biological specimens.

A methodology for tip and specimen-support manufacturing is described which allows for scanning tunneling microscopy (STM) and transmission electron microscopy (TEM) on identical areas of biological specimens, at comparable resolution. Topographs and dI/ds-maps were used to investigate tip-specimen interaction on native air-dried phage T4 polyheads where individual capsomeres and structural alterations upon repetitive scans have been observed at a tunnel current of 0.5 pA.

"Planned" management of squamous cell carcinoma of the mandibular oropharynx.

The term "planned," where management of squamous cell carcinoma of the mandibular oropharynx is concerned, has been put into proper perspective with several case examples in order to clarify its relationship to the pathophysiology of the disease and the anticipated morbidity involved in its cure.

Tendon glycosaminoglycan proteoglycan sidechains promote collagen fibril sliding-AFM observations at the nanoscale.

The extracellular matrix of tendon is mainly composed of discontinuous Type-I collagen fibrils and small leucine rich proteoglycans (PG). Macroscopic tendon behaviors like stiffness and strength are determined by the ultrastructural arrangement of these components. When a tendon is submitted to load, the collagen fibrils both elongate and slide relative to their neighboring fibrils. The role of PG glycosaminoglycan (GAG) sidechains in mediating inter-fibril load sharing remains controversial, with competing structure-function theories suggesting that PGs may mechanically couple neighboring collagen fibrils (cross-linking them to facilitate fibril stretch) or alternatively isolating them (promoting fibril gliding). In this study, we sought to clarify the functional role of GAGs in tensile tendon mechanics by directly investigating the mechanical response of individual collagen fibrils within their collagen network in both native and GAG depleted tendons. A control group of Achilles tendons from adult mice was compared with tendons in which GAGs were enzymatically depleted using chondroitinase ABC. Tendons were loaded to specific target strains, chemically fixed under constant load, and later sectioned for morphological analysis by an atomic force microscope (AFM). Increases in periodic banding of the collagen fibrils (D-period) or decreases in fibril diameter was considered to be representative of collagen fibril elongation and the mechanical contribution of GAGs at the ultrascale was quantified on this basis. At high levels of applied tendon strain (10%), GAG depleted tendons showed increased collagen stretch (less fibril sliding). We conclude that the hydrophilic GAGs seem thus not to act as mechanical crosslinks but rather act to promote collagen fibril sliding under tension.

A hybrid breath hold and continued respiration-triggered technique for time-resolved 3D MRI perfusion studies in lung cancer.

PURPOSE: Assessment of lung cancer perfusion is impaired by respiratory motion. Imaging times for contrast agent wash-out studies often exceed breath hold capabilities, and respiration triggering reduces temporal resolution. Temporally resolved volume acquisition of entire tumors is required to assess heterogeneity. Therefore, we developed and evaluated an MR measurement technique that exceeds a single breath hold, and provides a variable temporal resolution during acquisition while suspending breath-dependent motion. MATERIALS AND METHODS: 20 patients with suspected lung cancer were subjected to perfusion studies using a spoiled 3D gradient echo sequence after bolus injection of 0.07 mmol/kg body weight of Gd-DTPA. 10 acquisitions in expiratory breath hold were followed by 50 navigator-triggered acquisitions under free breathing. Post-processing allowed for co-registration of the 3D data sets. An ROI-based visualization of the signal-time curves was performed. RESULTS: In all cases motion-suspended, time-resolved volume data sets (40 x 33 x 10 cm(3), voxel size: 2.1 x 2.1 x 5.0 mm(3)) were generated with a variable, initially high temporal resolution (2.25 sec) that was synchronized with the breath pattern and covered up to 8 1/2 min. In 7 / 20 cases a remaining offset could be reduced by rigid co-registration. The tumors showed fast wash-in, followed by rapid signal decay (8 / 20) or a plateau. CONCLUSION: The feasibility of a perfusion study with hybrid breath hold and navigator-triggered time-resolved 3D MRI which combines high initial temporal resolution during breath hold with a long wash-out period under free breathing was demonstrated.

Design of a village breeding programme for a llama population in the High Andes of Bolivia.

No national breeding programme for llamas is in place in Bolivia. Initiatives for genetic improvement are rarely found and are usually carried out by NGOs working in rural development or improvement of livestock production or research stations. Farmers in the Province of Ayopaya in the District of Cochabamba have formed a breeders' organization with the aim of improving fibre production. In this study, a detailed outline of a breeding programme with a focus on organizational and technical details is described. Facing constraints like illiteracy of farmers, bad infrastructure and lack of finances, a simple breeding programme is set up. The breeding goal is a higher fleece weight while keeping the fleece quality at the current high level. Greasy fleece weight and fibre diameter are identified as main selection criteria. Mass selection of males is carried out. Selected males are either exchanged between farmers and used in the herds or are kept during the mating season in a central mating station owned by the breeders' organization. Model calculations were carried out with the program zplan, which is based on a deterministic approach. zplan evaluates the genetic and economic efficiency of breeding strategies considering one cycle of selection. Scenarios with only intra-herd use, using only the central mating station or combinations of those were compared in terms of expected genetic gain and expected increase of inbreeding. Fastest genetic progress is achieved when the males are kept in a central mating station as the selection intensity is on a high level. Rates of inbreeding vary between 0.08 and 0.32% per generation.

A hybrid scanning force and light microscope for surface imaging and three-dimensional optical sectioning in differential interference contrast.

The design of a scanned-cantilever-type force microscope is presented which is fully integrated into an inverted high-resolution video-enhanced light microscope. This set-up allows us to acquire thin optical sections in differential interference contrast (DIC) or polarization while the force microscope is in place. Such a hybrid microscope provides a unique platform to study how cell surface properties determine, or are affected by, the three-dimensional dynamic organization inside the living cell. The hybrid microscope presented in this paper has proven reliable and versatile for biological applications. It is the only instrument that can image a specimen by force microscopy and high-power DIC without having either to translate the specimen or to remove the force microscope. Adaptation of the design features could greatly enhance the suitability of other force microscopes for biological work.

Fabrication of near-field optical apertures in aluminium by a highly selective corrosion process in the evanescent field.

A simple, one-step process to fabricate high-quality apertures for scanning near-field optical microscope probes based on aluminium-coated silicon nitride cantilevers is presented. A thin evanescent optical field at a glass-water interface was used to heat the aluminium at the tip apex due to light absorption. The heat induced a breakdown of the passivating oxide layer and local corrosion of the metal, which selectively exposed the front-most part of the probe tip from the aluminium. Apertures with a protruding silicon nitride tip up to 72 nm in height were fabricated. The height of the protrusion was controlled by the extent of the evanescent field, whereas the diameter depended on the geometry of the probe substrate. The corrosion process proved to be self-terminating, yielding highly reproducible tip heights. Near-field optical resolution in a transmission mode of 85 nm was demonstrated.