Research strategies that combine molecular data from multiple levels of genome expression (i.e., multi-omics data), often referred to as a systems biology strategy, has been advocated as a route to discovering gene functions. In this study we conducted an evaluation of this strategy by combining lipidomics, metabolite mass-spectral imaging and transcriptomics data from leaves and roots in response to mutations in two AuTophaGy-related (ATG) genes of Arabidopsis. Autophagy is an essential cellular process that degrades and recycles macromolecules and organelles, and this process is blocked in the atg7 and atg9 mutants that were the focus of this study. Specifically, we quantified abundances of ~100 lipids and imaged the cellular locations of ~15 lipid molecular species and the relative abundance of ~26,000 transcripts from leaf and root tissues of WT, atg7 and atg9 mutant plants, grown either in normal (nitrogen-replete) and autophagy-inducing conditions (nitrogen-deficient). The multi-omics data enabled detailed molecular depiction of the effect of each mutation, and a comprehensive physiological model to explain the consequence of these genetic and environmental changes in autophagy is greatly facilitated by the a priori knowledge of the exact biochemical function of the ATG7 and ATG9 proteins.

Floral nectar is a rich secretion produced by the nectary gland and is offered as reward to attract pollinators leading to improved seed set. Nectars are composed of a complex mixture of sugars, amino acids, proteins, vitamins, lipids, organic and inorganic acids. This composition is influenced by several factors, including floral morphology, mechanism of nectar secretion, time of flowering, and visitation by pollinators. The objective of this study was to determine the contributions of flowering time, plant phylogeny, and pollinator selection on nectar composition in Nicotiana. The main classes of nectar metabolites (sugars and amino acids) were quantified using gas chromatography/mass spectrometric analytical platforms to identify differences among fifteen Nicotiana species representing day- and night-flowering plants from ten sections of the genus that are visited by five different primary pollinators. The nectar metabolomes of different Nicotiana species can predict the feeding preferences of the target pollinator(s) of each species, and the nectar sugars (i.e., glucose, fructose, and sucrose) are a distinguishing feature of Nicotiana species phylogeny. Moreover, comparative statistical analysis indicate that pollinators are a stronger determinant of nectar composition than plant phylogeny.

Host cell proteins (HCP) are a problematic set of impurities in downstream processing (DSP) as they behave most similarly to the target protein during separation. Approaching DSP with the knowledge of HCP separation behavior would be beneficial for the production of high purity recombinant biologics. Therefore, this work was aimed at characterizing the separation behavior of complex mixtures of HCP during a commonly used method: anion-exchange chromatography (AEX). An additional goal was to evaluate the performance of a statistical methodology, based on the characterization data, as a tool for predicting protein separation behavior. Aqueous two-phase partitioning followed by two-dimensional electrophoresis provided data on the three physicochemical properties most commonly exploited during DSP for each HCP: pI (isoelectric point), molecular weight, and surface hydrophobicity. The protein separation behaviors of two alternative expression host extracts (corn germ and E. coli) were characterized. A multivariate random forest (MVRF) statistical methodology was then applied to the database of characterized proteins creating a tool for predicting the AEX behavior of a mixture of proteins. The accuracy of the MVRF method was determined by calculating a root mean squared error value for each database. This measure never exceeded a value of 0.045 (fraction of protein populating each of the multiple separation fractions) for AEX. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 32:1453-1463, 2016.

Protein biomarkers can be used to predict biological traits or diseases. There is a need for robust methods to identify protein biomarkers that are linked to livestock production traits. Serum from blood is often used for detecting and screening for biomarkers. Albumin is a protein that comprises 50% or more of the total content of serum, and has historically been removed from serum before biomarker development and testing. We have shown that it is possible to successfully identify potential biomarkers without albumin removal, thus reducing steps and potentially cost in both development and routine testing phases.

The goal of this study was to find potential gene expression biomarkers in blood of piglets that can be used to predict pigs’ future feed efficiency. Using RNA-seq technology, we found 453 genes were differentially expressed (false discovery rate (FDR) ≤ 0.05) in the blood of two Yorkshire lines of pigs divergently selected for feed efficiency (FE) based on residual feed intake (RFI). Genes involved in several biosynthetic processes were overrepresented among genes more highly expressed in the low RFI line compared to the high RFI line. Weighted gene co-expression network analysis (WGCNA) also revealed genes involved in some of these biosynthesis processes and having similar patterns of expression formed clusters. The average expression in the clusters was highly associated with lines (p < 3.9E-07, R2 > 0.59). Current findings implied these biosynthesis pathways might be more active in the high RFI line. After further stringent validation, some of the differentially expressed genes (DEGs) will be selected for validation as biomarkers for feed efficiency.

and Implications The objective of this study was to determine the effect of selection for reduced feed intake (RFI) on scale activity for Yorkshire gilts. A total of 192 Yorkshire gilts were used, 96 were from a line that had been selected for low residual feed intake over 5 generations (LRFI) and 96 from a randomly bred control line (CRFI). Gilts were housed in 12 pens (16 gilts/pen; 0.82 m 2 /gilt) containing 8 gilts from each line in a conventional grow-finish unit. Gilts were weighed every 2-wks for a maximum of eight scores per gilt. Gilts were scored while on the weigh scale for activity using a whole number scale of one to five (1 = calm, minimal movement; 5 = continuous rapid movement and an escape attempt). Analyses were done using Proc Mixed of SAS. The LRFI line began with a lower scale activity score, but did not experience as great of a drop in their score as the CRFI gilts. The CRFI gilts scored lower by the end of the rounds compared to the LRFI gilts. In conclusion, selection for lower residual feed intake in purebred Yorkshires has a related effect on scale activity score but this relationship is complicated and thus warrants further research. Therefore, scale activity may not be an easy measure to be added to the list of already described traits in pigs as a factor which relates to selection for lower RFI.

We evaluated the preventative effects of Lactobacillus acidophilus strain NP51 fed to Balb/c mice infected with a virulent strain of Mycobacterial avium subspecies paratuberculosis (MAP) isolated from a clinical cow at the National Animal Disease Center in Ames, Iowa. Mice were randomized to treatment groups that were fed either viableor heat-killed NP51 and inoculated with either viable- or heat-killed MAP or sterile phosphate-buffered saline. Feeding the NP51 elevated numbers of T lymphocytes in the spleen and increased the concentration of interferongamma (IFN-γ) in the supernatant of splenocytes stimulated in vitro with MAP antigen. Most importantly, feeding the NP51 lowered the number of viable MAP CFU in the livers of infected mice on day 180 of the study. These results suggest that feeding the NP51 to BALB/c mice has potential to prevent the advance of MAP infection in mice.

The objective of this study was to determine if Yorkshire gilts become habituated to the process of weighing. A total of 192 pure bred Yorkshire gilts were used. The gilts were housed in 12 pens, 16 gilts/pen providing a space allowance of 0.82 m2/gilt. Gilts were weighed every 2-wks for a maximum of eight scores/ gilt. Gilts were scored while on the weigh scale for activity using a whole number scale of one to five (1 = calm, minimal movement; 5 = continuous rapid movement and an escape attempt) at two different time points, T = 0 (as soon as the back gate was closed and T = 15, 15 sec after the back gate was closed. Analyses were done using Proc Mixed of SAS. The differences during round one between T = 0 and T = 15 was 1 point on the 5 point scale. By round eight this had dropped down to a difference of 0.2. During the first round over both time points the gilts scored on average 2.5, by round 5 this had dropped down to around 1.5 at which point it stabilized. In conclusion, Yorkshire gilts appear to become habituated to the process of weighing over the course of a trial, and as such do not seem to find the process highly aversive.

Growth performance and carcass composition of 40 Yorkshire pigs (74.8±9.9 kg or 164.9±21.8 lbs), 20 pigs from a line selected for low residual feed intake for 5 generations and 20 pigs from a control line, was observed while fed on either an ad libitum or NRC maintenance (weight-stasis) basis over a 6 week period. The aim of the latter diet treatment was to keep pigs at a constant weight for six weeks. In the ad libitum treatment, there was no difference in initial (p < 0.49) or final body weights (p < 0.65) but the low residual feed intake line consumed 9% less feed compared to the control (p < 0.08). Similarly, there was no difference in LEA (p < 0.57) but the low residual feed intake line had slightly less backfat compared to the control (p < 0.21). These same results were found from chemical analysis of the carcass, as there was no difference in protein percentage (p < 0.60), but the ad libitum low residual feed intake pigs had a slightly lower fat percentage (p < 0.21). For the weight stasis treatment, the low residual feed intake pigs weighed 3.5% more than the control (p < 0.08), despite attempts to maintain a static body weight, and consumed 7.6% less feed overall (p < 0.09). Both lines had a decrease in backfat; however, the low residual feed intake line had an increase in loin eye area while the control line had a decrease. No differences were observed in chemical carcass composition between the two lines on the weight stasis treatment. These data show that the low residual feed intake line is more efficient, with only slight differences in carcass composition.

The objective of this study was to examine effects of feeding Lactobacillus acidophilus strain NP51 to mice challenged with Mycobacterium avium subspecies paratuberculosis ( MAP ). Mice were randomized to ten treatment groups; sentinels, control, heat‐killed MAP, viable MAP, heat‐killed NP51, viable NP51, heat‐killed NP51 plus heat‐killed MAP, heat‐killed NP51 plus viable‐MAP, viable‐NP51 plus heat‐killed MAP, viable‐NP51 plus viable‐MAP. Mice were fed 1 × 10 6 CFU of NP51· mice −1 · day −1 . On day 45, mice were challenged with 1 × 10 8 CFU of MAP intraperitonealy. We hypothesized that feeding NP51would increase Th‐1 responses and decrease progression of Johne's disease in mice. Ten mice from each group were euthanized on days 45, 90, 135 and 180. Supernatante from in vitro splenocyte culture was examined for IFN‐γ production. Feeding Heat‐killed NP51 to heat‐killed MAP‐ or viable MAP‐infected mice increased interferon ( IFN )‐γ by 1.4‐ and 4‐fold (44.32 and 127.14 vs. 31.69 pg/mL, respectively) over that of heat‐killed MAP‐infected control. Similarly, feeding viable‐NP51 to heat‐killed MAP‐ or viable MAP‐infected mice increased IFN‐γ by 3‐ and 4‐ fold (98.6 and 130.68 vs. 31.69, respectively) over that of heat‐killed MAP‐infected control. These data suggest that feeding NP51 can simulate IFN‐γ production and prevent progression of Johne's disease in mice. Funding source: Nutrition Physiology Company, Guymon, OK. Grant Funding Source : Nutrition Physiology Company

and Implications The objectives of this study were to determine the effect of selection for reduced residual feed intake (RFI) on lesion scores in gilts in their home pen. A total of 192 gilts were used; 96 were from a line that had been selected for low residual feed intake over 5 generations (LRFI) and 96 from a randomly bred control line (CRFI). Gilts were housed in 12 pens (16 gilts/pen; 0.82 m/gilt) containing 8 gilts from each line in a conventional grow-finish unit. Lesion scores were collected the day after placement and every 4 weeks for 3 subsequent periods. The gilt’s body was divided into 4 regions, with each region receiving a score of 0 (0 lesions) to 3 (5+ lesions). All analyses were done using Proc Mixed of SAS. The data were analyzed separately for the day after placement and the subsequent three rounds. Lesion scores for each region of the body were analyzed as repeated measures. Gilts from the LRFI line had lower (P 0.05) differences between the genetic lines. In conclusion, gilts from the line selected for low RFI had lower lesions scores on the day after placement into the grow-finish environment and this may be a useful tool to use in a selection program for more efficient gilts.

Residual feed intake (RFI) is a measure of feed efficiency defined as the difference between observed and predicted feed intake based on average requirements for maintenance and production. At Iowa State University, two lines of Yorkshire pigs were developed to study the effects of selection for RFI during the grow/finish phase of production (RFIG/F). One line was selected over 7 generations for decreased RFIG/F (LRFI) to improve feed efficiency and the other line (HRFI) was selected randomly for 4 generations and then for increased RFIG/F. The main objectives of this dissertation were to evaluate feeding behavior traits and sow reproductive performance and lactation efficiency. Pigs from the LRFI line had significantly lower feed intake (FI) per day than did HRFI pigs. After adjusting for FI per day, number of visits (NV) per day and per hour did not differ significantly between the two lines but the trend was for LRFI pigs to have fewer visits, particularly during peak eating times. Furthermore, pigs from the LRFI line ate faster and spent less time in the feeder per day, per visit, and per hour than HRFI pigs. Feeding behavior traits were moderately to highly heritable, with heritabilities ranging from 0.36 for FI per visit to 0.71 for occupation time (OT) per day. Feed intake rate was also highly heritable at 0.59. Heritabilities of NV per day, OT per visit, and FI per day were similar (0.44, 0.42, and 0.42, respectively). FI per day was strongly correlated, both phenotypically and genetically, with RFIG/F, average daily gain (ADG), and backfat depth (BF). FI per visit was moderately correlated, both phenotypically and genetically, with ADG and BF. OT per day was moderately correlated, both phenotypically and genetically, with RFIG/F and BF. Other correlations between feeding behavior traits and performance traits were low. For each feeding behavior trait, one or two genomic regions were identified as being important in a whole genome association study. SNPs located adjacent to MC4R (a gene already shown to be associated with FI, fatness, and growth) were significant for FI per day. Other genes with nearby SNPs found to be associated with feeding behavior traits included several related to different transcription regulators. After 7 generations, selection for decreased RFIG/F has improved piglet performance and increased sow weight loss during lactation. LRFI sows had more piglets farrowed, born alive, and weaned than did HRFI sows. LRFI piglets were heavier at birth and had better litter growth than did HRFI piglets. However, this increased piglet performance came at a cost to the sow During lactation, LRFI sows consumed less feed and lost more body weight, fat mass, and BF than did HRFI sows. LRFI sows had a greater negative energy balance but more favorable lactation efficiency and RFI during lactation than HRFI sows. Heritabilities were high (h2 > 0.4) for sow weights, body composition, and maintenance requirements and piglet birth weights. Piglet growth during lactation, mobilization of the sow's body tissue, sow feed intake and total born were moderately heritable (0.2 < h2 < 0.4). Correlations with RFIG/F were not significant for most traits. However, strong, positive genetic correlations with RFIG/F were found for sow weight at farrowing and weaning, sow maintenance requirements, and sow RFI and strong, negative genetic correlations with RFIG/F were found for sow protein mass loss and lactation efficiency. In conclusion, feed efficiency may be affected by feed intake behavior because selection for decreased RFIG/F has resulted in pigs which spend less time eating and eat faster. A large genetic component to feeding behavior is evident and measuring and selecting for these traits may allow for other opportunities to improve traits of economic importance. Selection for RFIG/F has positively affected piglet performance and lactation efficiency but has negatively affected sow body condition change and energy balance during lactation.

The genetic mechanisms controlling appetite and feeding behaviors are not well understood. In this study, transcriptional profiling was used to identify porcine genes and pathways that respond to a fasting treatment or to a missense mutation (D298N) in the melanocortin‐4 receptor (MC4R) gene, which has been associated with increased growth and feed efficiency. Prepubertal gilts (n=24) homozygous for D298N MC4R or wildtype were either fed ad lib. or fasted for 3 days in a completely randomized block design with 2×2 factorial treatment structure. The Affymetrix Porcine Genome was used to profile gene expression of liver and subcutaneous fat. Due to fasting, 7,029 genes in adipose tissue and 1,831 genes in liver were declared differentially expressed (q<0.05), but an effect of MC4R was not observed on expression in both tissues under same criterion. Pathway analyses indicated that 23 of the tested set of 298 genes that were down‐regulated (p<0.05, fold change>2) in the adipose tissue due to fasting were directly regulated by sterol regulatory element binding transcription factor‐1 (SREBF1). We confirmed the expression levels of SREBF1 and of its targets such as fatty acid synthase, aconitase‐1, acetyl CoA carboxylase alpha, and acetyl CoA synthase in the adipose tissue by qPCR. Results indicate that SREBF1 may play a key role in determining the pathways that respond to fasting in pigs. (USDA‐NRI‐2005‐3560415618).