Determination of Body Parts in Holstein Friesian Cows Comparing Neural Networks and k Nearest Neighbour Classification.

Machine learning methods have become increasingly important in animal science, and the success of an automated application using machine learning often depends on the right choice of method for the respective problem and data set. The recognition of objects in 3D data is still a widely studied topic and especially challenging when it comes to the partition of objects into predefined segments. In this study, two machine learning approaches were utilized for the recognition of body parts of dairy cows from 3D point clouds, i.e., sets of data points in space. The low cost off-the-shelf depth sensor Microsoft Kinect V1 has been used in various studies related to dairy cows. The 3D data were gathered from a multi-Kinect recording unit which was designed to record Holstein Friesian cows from both sides in free walking from three different camera positions. For the determination of the body parts head, rump, back, legs and udder, five properties of the pixels in the depth maps (row index, column index, depth value, variance, mean curvature) were used as features in the training data set. For each camera positions, a k nearest neighbour classifier and a neural network were trained and compared afterwards. Both methods showed small Hamming losses (between 0.007 and 0.027 for k nearest neighbour (kNN) classification and between 0.045 and 0.079 for neural networks) and could be considered successful regarding the classification of pixel to body parts. However, the kNN classifier was superior, reaching overall accuracies 0.888 to 0.976 varying with the camera position. Precision and recall values associated with individual body parts ranged from 0.84 to 1 and from 0.83 to 1, respectively. Once trained, kNN classification is at runtime prone to higher costs in terms of computational time and memory compared to the neural networks. The cost vs. accuracy ratio for each methodology needs to be taken into account in the decision of which method should be implemented in the application.

Structural Asymmetry and Kinetic Limping of Single Rotary F-ATP Synthases.

F-ATP synthases use proton flow through the FO domain to synthesize ATP in the F1 domain. In Escherichia coli, the enzyme consists of rotor subunits γεc10 and stator subunits (αß)3δab2. Subunits c10 or (αß)3 alone are rotationally symmetric. However, symmetry is broken by the b2 homodimer, which together with subunit δa, forms a single eccentric stalk connecting the membrane embedded FO domain with the soluble F1 domain, and the central rotating and curved stalk composed of subunit γε. Although each of the three catalytic binding sites in (αß)3 catalyzes the same set of partial reactions in the time average, they might not be fully equivalent at any moment, because the structural symmetry is broken by contact with b2δ in F1 and with b2a in FO. We monitored the enzyme's rotary progression during ATP hydrolysis by three single-molecule techniques: fluorescence video-microscopy with attached actin filaments, Förster resonance energy transfer between pairs of fluorescence probes, and a polarization assay using gold nanorods. We found that one dwell in the three-stepped rotary progression lasting longer than the other two by a factor of up to 1.6. This effect of the structural asymmetry is small due to the internal elastic coupling.

Oxygenic photosynthesis: history, status and perspective.

Cyanobacteria and plants carry out oxygenic photosynthesis. They use water to generate the atmospheric oxygen we breathe and carbon dioxide to produce the biomass serving as food, feed, fibre and fuel. This paper scans the emergence of structural and mechanistic understanding of oxygen evolution over the past 50 years. It reviews speculative concepts and the stepped insight provided by novel experimental and theoretical techniques. Driven by sunlight photosystem II oxidizes the catalyst of water oxidation, a hetero-metallic Mn4CaO5(H2O)4 cluster. Mn3Ca are arranged in cubanoid and one Mn dangles out. By accumulation of four oxidizing equivalents before initiating dioxygen formation it matches the four-electron chemistry from water to dioxygen to the one-electron chemistry of the photo-sensitizer. Potentially harmful intermediates are thereby occluded in space and time. Kinetic signatures of the catalytic cluster and its partners in the photo-reaction centre have been resolved, in the frequency domain ranging from acoustic waves via infra-red to X-ray radiation, and in the time domain from nano- to milli-seconds. X-ray structures to a resolution of 1.9 Å are available. Even time resolved X-ray structures have been obtained by clocking the reaction cycle by flashes of light and diffraction with femtosecond X-ray pulses. The terminal reaction cascade from two molecules of water to dioxygen involves the transfer of four electrons, two protons, one dioxygen and one water. A rigorous mechanistic analysis is challenging because of the kinetic enslaving at millisecond duration of six partial reactions (4e-, 1H+, 1O2). For the time being a peroxide-intermediate in the reaction cascade to dioxygen has been in focus, both experimentally and by quantum chemistry. Homo sapiens has relied on burning the products of oxygenic photosynthesis, recent and fossil. Mankind's total energy consumption amounts to almost one-fourth of the global photosynthetic productivity. If the average power consumption equalled one of those nations with the highest consumption per capita it was four times greater and matched the total productivity. It is obvious that biomass should be harvested for food, feed, fibre and platform chemicals rather than for fuel.

The Phylogenetic Signature Underlying ATP Synthase c-Ring Compliance.

The proton-driven ATP synthase (FOF1) is comprised of two rotary, stepping motors (FO and F1) coupled by an elastic power transmission. The elastic compliance resides in the rotor module that includes the membrane-embedded FO c-ring. Proton transport by FO is firmly coupled to the rotation of the c-ring relative to other FO subunits (ab2). It drives ATP synthesis. We used a computational method to investigate the contribution of the c-ring to the total elastic compliance. We performed principal component analysis of conformational ensembles built using distance constraints from the bovine mitochondrial c-ring x-ray structure. Angular rotary twist, the dominant ring motion, was estimated to show that the c-ring accounted in part for the measured compliance. Ring rotation was entrained to rotation of the external helix within each hairpin-shaped c-subunit in the ring. Ensembles of monomer and dimers extracted from complete c-rings showed that the coupling between collective ring and the individual subunit motions was independent of the size of the c-ring, which varies between organisms. Molecular determinants were identified by covariance analysis of residue coevolution and structural-alphabet-based local dynamics correlations. The residue coevolution gave a readout of subunit architecture. The dynamic couplings revealed that the hinge for both ring and subunit helix rotations was constructed from the proton-binding site and the adjacent glycine motif (IB-GGGG) in the midmembrane plane. IB-GGGG motifs were linked by long-range couplings across the ring, while intrasubunit couplings connected the motif to the conserved cytoplasmic loop and adjacent segments. The correlation with principal collective motions shows that the couplings underlie both ring rotary and bending motions. Noncontact couplings between IB-GGGG motifs matched the coevolution signal as well as contact couplings. The residue coevolution reflects the physiological importance of the dynamics that may link proton transfer to ring compliance.

Spatio-temporal resolution of primary processes of photosynthesis.

Technical progress in laser-sources and detectors has allowed the temporal and spatial resolution of chemical reactions down to femtoseconds and Å-units. In photon-excitable systems the key to chemical kinetics, trajectories across the vibrational saddle landscape, are experimentally accessible. Simple and thus well-defined chemical compounds are preferred objects for calibrating new methodologies and carving out paradigms of chemical dynamics, as shown in several contributions to this Faraday Discussion. Aerobic life on earth is powered by solar energy, which is captured by microorganisms and plants. Oxygenic photosynthesis relies on a three billion year old molecular machinery which is as well defined as simpler chemical constructs. It has been analysed to a very high precision. The transfer of excitation between pigments in antennae proteins, of electrons between redox-cofactors in reaction centres, and the oxidation of water by a Mn4Ca-cluster are solid state reactions. ATP, the general energy currency of the cell, is synthesized by a most agile, rotary molecular machine. While the efficiency of photosynthesis competes well with photovoltaics at the time scale of nanoseconds, it is lower by an order of magnitude for crops and again lower for bio-fuels. The enormous energy demand of mankind calls for engineered (bio-mimetic or bio-inspired) solar-electric and solar-fuel devices.

ATP synthase.

Oxygenic photosynthesis is the principal converter of sunlight into chemical energy. Cyanobacteria and plants provide aerobic life with oxygen, food, fuel, fibers, and platform chemicals. Four multisubunit membrane proteins are involved: photosystem I (PSI), photosystem II (PSII), cytochrome b6f (cyt b6f), and ATP synthase (FOF1). ATP synthase is likewise a key enzyme of cell respiration. Over three billion years, the basic machinery of oxygenic photosynthesis and respiration has been perfected to minimize wasteful reactions. The proton-driven ATP synthase is embedded in a proton tight-coupling membrane. It is composed of two rotary motors/generators, FO and F1, which do not slip against each other. The proton-driven FO and the ATP-synthesizing F1 are coupled via elastic torque transmission. Elastic transmission decouples the two motors in kinetic detail but keeps them perfectly coupled in thermodynamic equilibrium and (time-averaged) under steady turnover. Elastic transmission enables operation with different gear ratios in different organisms.

Quantification of the effects of fur, fur color, and velocity on Time-Of-Flight technology in dairy production.

With increasing herd sizes, camera based monitoring solutions rise in importance. 3D cameras, for example Time-Of-Flight (TOF) cameras, measure depth information. These additional information (3D data) could be beneficial for monitoring in dairy production. In previous studies regarding TOF technology, only standing cows were recorded to avoid motion artifacts. Therefore, necessary conditions for a TOF camera application in dairy cows are examined in this study. For this purpose, two cow models with plaster and fur surface, respectively, were recorded at four controlled velocities to quantify the effects of movement, fur color, and fur. Comparison criteria concerning image usability, pixel-wise deviation, and precision in coordinate determination were defined. Fur and fur color showed large effects (η (2)=0.235 and η (2)=0.472, respectively), which became even more considerable when the models were moving. The velocity of recorded animals must therefore be controlled when using TOF cameras. As another main result, body parts which lie in the middle of the cow model's back can be determined neglecting the effect of velocity or fur. With this in mind, further studies may obtain sound results using TOF technology in dairy production.

Evaluation of different lactation curve models fitted for milk viscosity recorded by an automated on-line California Mastitis Test.

Laboratory somatic cell count (LSCC) records are usually recorded monthly and provide an important information source for breeding and herd management. Daily milk viscosity detection in composite milking (expressed as drain time) with an automated on-line California Mastitis Test (CMT) could serve immediately as an early predictor of udder diseases and might be used as a selection criterion to improve udder health. The aim of the present study was to clarify the relationship between the well-established LSCS and the new trait,'drain time', and to estimate their correlations to important production traits. Data were recorded on the dairy research farm Karkendamm in Germany. Viscosity sensors were installed on every fourth milking stall in the rotary parlour to measure daily drain time records. Weekly LSCC and milk composition data were available. Two data sets were created containing records of 187,692 milkings from 320 cows (D1) and 25,887 drain time records from 311 cows (D2). Different fixed effect models, describing the log-transformed drain time (logDT), were fitted to achieve applicable models for further analysis. Lactation curves were modelled with standard parametric functions (Ali and Schaeffer, Legendre polynomials of second and third degree) of days in milk (DIM). Random regression models were further applied to estimate the correlations between cow effects between logDT and LSCS with further important production traits. LogDT and LSCS were strongest correlated in mid-lactation (r = 0.78). Correlations between logDT and production traits were low to medium. Highest correlations were reached in late lactation between logDT and milk yield (r = -0.31), between logDT and protein content (r = 0.30) and in early as well as in late lactation between logDT and lactose content (r = -0.28). The results of the present study show that the drain time could be used as a new trait for daily mastitis control.

Structure and energy transfer in photosystems of oxygenic photosynthesis.

Oxygenic photosynthesis is the principal converter of sunlight into chemical energy on Earth. Cyanobacteria and plants provide the oxygen, food, fuel, fibers, and platform chemicals for life on Earth. The conversion of solar energy into chemical energy is catalyzed by two multisubunit membrane protein complexes, photosystem I (PSI) and photosystem II (PSII). Light is absorbed by the pigment cofactors, and excitation energy is transferred among the antennae pigments and converted into chemical energy at very high efficiency. Oxygenic photosynthesis has existed for more than three billion years, during which its molecular machinery was perfected to minimize wasteful reactions. Light excitation transfer and singlet trapping won over fluorescence, radiation-less decay, and triplet formation. Photosynthetic reaction centers operate in organisms ranging from bacteria to higher plants. They are all evolutionarily linked. The crystal structure determination of photosynthetic protein complexes sheds light on the various partial reactions and explains how they are protected against wasteful pathways and why their function is robust. This review discusses the efficiency of photosynthetic solar energy conversion.

Polymorphisms within the APOBR gene are highly associated with milk levels of prognostic ketosis biomarkers in dairy cows.

Essentially all high-yielding dairy cows experience a negative energy balance during early lactation leading to increased lipomobilization, which is a normal physiological response. However, a severe energy deficit may lead to high levels of ketone bodies and, subsequently, to subclinical or clinical ketosis. It has previously been reported that the ratio of glycerophosphocholine to phosphocholine in milk is a prognostic biomarker for the risk of ketosis in dairy cattle. It was hypothesized that this ratio reflects the ability to break down blood phosphatidylcholine as a fatty acid resource. In the current study, 248 animals from a previous study were genotyped with Illumina BovineSNP50 BeadChip, and genome-wide association studies were carried out for the milk levels of phosphocholine, glycerophosphocholine, and the ratio of both metabolites. It was demonstrated that the latter two traits are heritable with h2 = 0.43 and h2 = 0.34, respectively. A major quantitative trait locus was identified on cattle chromosome 25. The APOBR gene, coding for the apolipoprotein B receptor, is located within this region and was analyzed as a candidate gene. The analysis revealed highly significant associations of polymorphisms within the gene with glycerophosphocholine as well as the metabolite ratio. These findings support the hypothesis that differences in the ability to take up blood phosphatidylcholine from low-density lipoproteins play an important role in early lactation metabolic stability of dairy cows and indicate APOBR to contain a causative variant.

A refined reaction-diffusion model of tau-microtubule dynamics and its application in FDAP analysis.

Fluorescence decay after photoactivation (FDAP) and fluorescence recovery after photobleaching (FRAP) are well established approaches for studying the interaction of the microtubule (MT)-associated protein tau with MTs in neuronal cells. Previous interpretations of FDAP/FRAP data have revealed dwell times of tau on MTs in the range of several seconds. However, this is difficult to reconcile with a dwell time recently measured by single-molecule analysis in neuronal processes that was shorter by two orders of magnitude. Questioning the validity of previously used phenomenological interpretations of FDAP/FRAP data, we have generalized the standard two-state reaction-diffusion equations by 1), accounting for the parallel and discrete arrangement of MTs in cell processes (i.e., homogeneous versus heterogeneous distribution of tau-binding sites); and 2), explicitly considering both active (diffusion upon MTs) and passive (piggybacking upon MTs at rates of slow axonal transport) motion of bound tau. For some idealized cases, analytical solutions were derived. By comparing them with the full numerical solution and Monte Carlo simulations, the respective validity domains were mapped. Interpretation of our FDAP data (from processes of neuronally differentiated PC12 cells) in light of the heterogeneous formalism yielded independent estimates for the association (∼2 ms) and dwell (∼100 ms) times of tau to/on a single MT rather than in an MT array. The dwell time was shorter by orders of magnitude than that in a previous report where a homogeneous topology of MTs was assumed. We found that the diffusion of bound tau was negligible in vivo, in contrast to an earlier report that tau diffuses along the MT lattice in vitro. Methodologically, our results demonstrate that the heterogeneity of binding sites cannot be ignored when dealing with reaction-diffusion of cytoskeleton-associated proteins. Physiologically, the results reveal the behavior of tau in cellular processes, which is noticeably different from that in vitro.

Interplay between phosphorylation and palmitoylation mediates plasma membrane targeting and sorting of GAP43.

Phosphorylation and lipidation provide posttranslational mechanisms that contribute to the distribution of cytosolic proteins in growing nerve cells. The growth-associated protein GAP43 is susceptible to both phosphorylation and S-palmitoylation and is enriched in the tips of extending neurites. However, how phosphorylation and lipidation interplay to mediate sorting of GAP43 is unclear. Using a combination of biochemical, genetic, and imaging approaches, we show that palmitoylation is required for membrane association and that phosphorylation at Ser-41 directs palmitoylated GAP43 to the plasma membrane. Plasma membrane association decreased the diffusion constant fourfold in neuritic shafts. Sorting to the neuritic tip required palmitoylation and active transport and was increased by phosphorylation-mediated plasma membrane interaction. Vesicle tracking revealed transient association of a fraction of GAP43 with exocytic vesicles and motion at a fast axonal transport rate. Simulations confirmed that a combination of diffusion, dynamic plasma membrane interaction and active transport of a small fraction of GAP43 suffices for efficient sorting to growth cones. Our data demonstrate a complex interplay between phosphorylation and lipidation in mediating the localization of GAP43 in neuronal cells. Palmitoylation tags GAP43 for global sorting by piggybacking on exocytic vesicles, whereas phosphorylation locally regulates protein mobility and plasma membrane targeting of palmitoylated GAP43.

Single-molecule tracking of tau reveals fast kiss-and-hop interaction with microtubules in living neurons.

The microtubule-associated phosphoprotein tau regulates microtubule dynamics and is involved in neurodegenerative diseases collectively called tauopathies. It is generally believed that the vast majority of tau molecules decorate axonal microtubules, thereby stabilizing them. However, it is an open question how tau can regulate microtubule dynamics without impeding microtubule-dependent transport and how tau is also available for interactions other than those with microtubules. Here we address this apparent paradox by fast single-molecule tracking of tau in living neurons and Monte Carlo simulations of tau dynamics. We find that tau dwells on a single microtubule for an unexpectedly short time of ∼40 ms before it hops to the next. This dwell time is 100-fold shorter than previously reported by ensemble measurements. Furthermore, we observed by quantitative imaging using fluorescence decay after photoactivation recordings of photoactivatable GFP-tagged tubulin that, despite this rapid dynamics, tau is capable of regulating the tubulin-microtubule balance. This indicates that tau's dwell time on microtubules is sufficiently long to influence the lifetime of a tubulin subunit in a GTP cap. Our data imply a novel kiss-and-hop mechanism by which tau promotes neuronal microtubule assembly. The rapid kiss-and-hop interaction explains why tau, although binding to microtubules, does not interfere with axonal transport.

Lateral pH gradient between OXPHOS complex IV and F(0)F(1) ATP-synthase in folded mitochondrial membranes.

Ion-driven ATP synthesis by rotary F0F1 ATP-synthase powers aerobic life. Since Mitchell's seminal hypothesis, this synthesis has been discussed in terms of the proton-motive force between two bulk phases, each in equilibrium. In active mitochondria, a steady proton flow cycles between pumps and the distant ATP synthase. Here we determine the lateral pH profile along the p-side of cristae in situ by attaching a ratiometric fluorescent pH-sensitive GFP variant to OXPHOS complex IV, a proton pump, and the dimeric F0F1 ATP-synthase, a proton consumer. In respiring HeLa cells, we observe that the local pH at F0F1 dimers is 0.3 units less acidic than that at complex IV. This finding is consistent with the calculated pH profile for steady proton diffusion from CIV to F0F1. The observed lateral variation in the proton-motive force necessitates a modification to Peter Mitchell's chemiosmotic proposal. The experimental technique can be extended to other pH-dependent reactions in membrane microcompartments.

Calibration of an automated California mastitis test with focus on the device-dependent variation.

The aim of the paper was to estimate the accuracy of the metrology of an installed indirect on-line sensor system based on the automated California Mastitis Test (CMT) with focus on the prior established device-dependent variation. A sensor calibration was implemented. Therefore, seven sensors were tested with similar trials on the dairy research farm Karkendamm (Germany) on two days in July 2011 and January 2012. Thereby, 18 mixed milk samples from serial dilutions were fourfold recorded at every sensor. For the validation, independent sensor records with corresponding lab somatic cell score records (LSCS) in the period between both trials were used (n = 1,357). From these records for each sensor a polynomial regression function was calculated. The predicted SCS (PSCS) was obtained for each sensor with the previously determined regression coefficients. Pearson correlation coefficients between PSCS and LSCS were established for each sensor and ranged between r = 0.57 and r = 0.67. Comparing the results with the correlation coefficients between the on-line SCS (OSCS) and the LSCS (r = 0.20 to 0.57) for every sensor, the calibration showed the tendency to improve the installed sensor system.

Feasibility of automated body trait determination using the SR4K time-of-flight camera in cow barns.

As herd sizes have increased in the last decades, computerized monitoring solutions, which provide fast, objective and accurate evaluations of the herd status, gain more and more importance. This study analyzes the feasibility of a Time-of-Flight-camera-based system for gathering body traits in dairy cows for use under cow barn conditions. Recording, determination of body condition score on a 5 point scale by visual and manual inspection, and measuring the backfat thickness with ultrasound took place from July 2011 to May 2012 at the dairy research farm Karkendamm of the Institute of Animal Breeding and Husbandry, Christian-Albrechts-University Kiel (Germany) and between August 2010 and July 2012 at the Institute for Agricultural Engineering and Animal Husbandry of Bavarian State Research Center for Agriculture in Grub (Germany). The two breeds Holstein Friesian cows (Karkendamm) and Fleckvieh (Grub) were considered in this study. Software for recording, image sorting and evaluation, determining the body parts needed, and extracting traits from the images was written and assembled to an automated system. Sorting the images and finding ischeal tuberosities, base of the tail, and dishes of the rump, backbone, and hips had error rates of 0.2%, 1.5%, 0.1%, and 2.6%, respectively. 13 traits were extracted and compared to backfat thickness and body condition score as well as between breeds. All traits depend significantly on the animal and showed very large effect sizes. Coefficients of determination restricted to individual animals were reaching up to 0.89. The precision in measuring the traits and gathering backfat thickness was comparable. Results indicated that the application of Time-Of-Flight in determination of body traits is feasible.

Investigation of the hemostatic effect of a transdermal patch containing 0.55 mg ethinyl estradiol and 2.1 mg gestodene compared with a monophasic oral contraceptive containing 0.03 mg ethinyl estradiol and 0.15 mg levonorgestrel: an open-label, randomized, crossover study.

BACKGROUND: Transdermal delivery of contraceptives offers several advantages over combined oral contraceptives (COCs), including effective absorption and the provision of relatively constant serum concentrations. Ethinyl estradiol (EE) and the progestin gestodene are well-absorbed through the skin and, therefore, well-suited for use in a transdermal contraceptive patch. OBJECTIVE: The objective of this study was to investigate the impact of a once-weekly transparent, transdermal patch delivering low doses of EE and gestodene equivalent to a COC containing 0.02 mg EE and 0.06 mg gestodene on hemostasis parameters compared with a monophasic COC containing 0.03 mg EE and 0.15 mg levonorgestrel. METHODS: In this single-center, open-label, randomized, crossover study, 30 women (aged 18-35 years) received three cycles of each treatment, separated by a two-cycle washout period. The primary outcome measure was the absolute change from baseline in prothrombin fragments 1 + 2 and D-dimer. RESULTS: For both treatments, prothrombin fragments 1 + 2 remained stable during the first treatment period, and increased only slightly in the second period (mean absolute change 0.025 and 0.028 nmol/L in the novel Bayer patch and COC groups, respectively). Increases in D-dimer were observed in both periods (mean absolute change 107.0 ± 147.2 ng/L for the novel Bayer patch and 113.7 ± 159.0 ng/L for the COC). There were no statistically significant treatment differences in prothrombin 1 + 2 or D-dimer (p = 0.667 and p = 0.884, respectively) and no statistically significant treatment sequence or period effects. CONCLUSION: A COC containing 0.03 mg EE and 0.15 mg levonorgestrel and the novel Bayer patch have comparable influence on hemostatic endpoints. Both treatments were well-tolerated by subjects.

Half a century of molecular bioenergetics.

Molecular bioenergetics deals with the construction, function and regulation of the powerhouses of life. The present overview sketches scenes and actors, farsighted goals and daring hypotheses, meticulous tool-making, painstaking benchwork, lucky discovery, serious scepticism, emphatic believing and strong characters with weak and others with hard arguments, told from a personal, admittedly limited, perspective. Bioenergetics will blossom further with the search focused on both where there is bright light for ever-finer detail and the obvious dark spots for surprise and discovery.