Large-scale analysis of sheep rumen metagenome profiles captured by reduced representation sequencing reveals individual profiles are influenced by the environment and genetics of the host.

BACKGROUND: Producing animal protein while reducing the animal's impact on the environment, e.g., through improved feed efficiency and lowered methane emissions, has gained interest in recent years. Genetic selection is one possible path to reduce the environmental impact of livestock production, but these traits are difficult and expensive to measure on many animals. The rumen microbiome may serve as a proxy for these traits due to its role in feed digestion. Restriction enzyme-reduced representation sequencing (RE-RRS) is a high-throughput and cost-effective approach to rumen metagenome profiling, but the systematic (e.g., sequencing) and biological factors influencing the resulting reference based (RB) and reference free (RF) profiles need to be explored before widespread industry adoption is possible. RESULTS: Metagenome profiles were generated by RE-RRS of 4,479 rumen samples collected from 1,708 sheep, and assigned to eight groups based on diet, age, time off feed, and country (New Zealand or Australia) at the time of sample collection. Systematic effects were found to have minimal influence on metagenome profiles. Diet was a major driver of differences between samples, followed by time off feed, then age of the sheep. The RF approach resulted in more reads being assigned per sample and afforded greater resolution when distinguishing between groups than the RB approach. Normalizing relative abundances within the sampling Cohort abolished structures related to age, diet, and time off feed, allowing a clear signal based on methane emissions to be elucidated. Genus-level abundances of rumen microbes showed low-to-moderate heritability and repeatability and were consistent between diets. CONCLUSIONS: Variation in rumen metagenomic profiles was influenced by diet, age, time off feed and genetics. Not accounting for environmental factors may limit the ability to associate the profile with traits of interest. However, these differences can be accounted for by adjusting for Cohort effects, revealing robust biological signals. The abundances of some genera were consistently heritable and repeatable across different environments, suggesting that metagenomic profiles could be used to predict an individual's future performance, or performance of its offspring, in a range of environments. These results highlight the potential of using rumen metagenomic profiles for selection purposes in a practical, agricultural setting.

Combining host and rumen metagenome profiling for selection in sheep: prediction of methane, feed efficiency, production, and health traits.

BACKGROUND: Rumen microbes break down complex dietary carbohydrates into energy sources for the host and are increasingly shown to be a key aspect of animal performance. Host genotypes can be combined with microbial DNA sequencing to predict performance traits or traits related to environmental impact, such as enteric methane emissions. Metagenome profiles were generated from 3139 rumen samples, collected from 1200 dual purpose ewes, using restriction enzyme-reduced representation sequencing (RE-RRS). Phenotypes were available for methane (CH4) and carbon dioxide (CO2) emissions, the ratio of CH4 to CH4 plus CO2 (CH4Ratio), feed efficiency (residual feed intake: RFI), liveweight at the time of methane collection (LW), liveweight at 8 months (LW8), fleece weight at 12 months (FW12) and parasite resistance measured by faecal egg count (FEC1). We estimated the proportion of phenotypic variance explained by host genetics and the rumen microbiome, as well as prediction accuracies for each of these traits. RESULTS: Incorporating metagenome profiles increased the variance explained and prediction accuracy compared to fitting only genomics for all traits except for CO2 emissions when animals were on a grass diet. Combining the metagenome profile with host genotype from lambs explained more than 70% of the variation in methane emissions and residual feed intake. Predictions were generally more accurate when incorporating metagenome profiles compared to genetics alone, even when considering profiles collected at different ages (lamb vs adult), or on different feeds (grass vs lucerne pellet). A reference-free approach to metagenome profiling performed better than metagenome profiles that were restricted to capturing genera from a reference database. We hypothesise that our reference-free approach is likely to outperform other reference-based approaches such as 16S rRNA gene sequencing for use in prediction of individual animal performance. CONCLUSIONS: This paper shows the potential of using RE-RRS as a low-cost, high-throughput approach for generating metagenome profiles on thousands of animals for improved prediction of economically and environmentally important traits. A reference-free approach using a microbial relationship matrix from log10 proportions of each tag normalized within cohort (i.e., the group of animals sampled at the same time) is recommended for future predictions using RE-RRS metagenome profiles.

Infant mortality and intra-household competition in the Northern Islands of Orkney, Scotland, 1855-2001.

This study applies principles from the theory of household life cycles to the study of early childhood mortality in the population of the Northern Orkney Islands, Scotland. The primary hypothesis is that unfavorable household economic conditions resulting from changes in household demographic composition increase the risk of death for children under the age of 5 years because of limited resources and intra-household competition. We apply Cox proportional hazards models to nearly 5,000 linked birth and death records from the Northern Orkney Islands, Scotland, from the period 1855 to 2001. The dependent variable is the child's risk of death before age 5. Findings suggest that children in households with unfavorable age compositions face higher risk of death. This elevated risk of death continues once heterogeneity among children, islands, and households is controlled. Results also show differential risk of death for male children, children of higher birth orders, and twin births. The analyses present evidence for intra-household competition in this historic setting. The most convincing evidence of competition is found in the effects of household consumer/producer ratios and twinning on child mortality risks.

Genetic parameter analysis of bareness and tail traits in New Zealand sheep.

Physical traits that improve welfare and disease outcomes for sheep are becoming increasingly important due to both increased climate challenges and societal expectations. Such traits include tail length, the amount of skin (vs. wool) on the underside of the tail, and the area of no-wool (hair) on the belly and breech areas (surrounding the anus) of the animal. An industry dataset consisting of records from individual stud breeders and industry progeny tests was available to estimate the genetic parameters associated with these traits and to investigate the potential for within-breed genetic selection. The heritability estimate for tail length was 0.68 ± 0.01 when breed was not fitted, and 0.63 ± 0.01 when breed was fitted. Similar trends were observed for breech and belly bareness which had heritability estimates around 0.50 (± 0.01). The estimates for these bareness traits are both higher than previous reports from animals of the same age. There was, however, between breed variation in the starting point for these traits, with some breeds having significantly longer tails and a wooly breech and belly, and limited variability. Overall, the results of this study show that flocks exhibiting some variation will be able to make rapid genetic progress in selecting for bareness and tail length traits, and therefore have the potential to make progress towards a sheep that is easier to look after and suffers fewer welfare insults. For those breeds that showed limited within-breed variation, outcrossing may be required to introduce genotypes that exhibit shorter tail length and bareness of belly and breech to increase the rate of genetic gain. Whatever approach is taken by the industry, these results support that genetic improvement can be used to breed "ethically improved sheep".

Physical traits that improve welfare and disease outcomes for sheep are becoming increasingly important as a result of both increased climate challenges and societal expectations. Such physical traits include genetically short tails and bare (hair vs. wool) in the breech and belly region. This study demonstrates that these traits are highly heritable, indicating that a lot of the variation observed between individuals is due to genetic variation. Overall, the results of this study show that flocks exhibiting some variation will be able to make rapid genetic progress in selecting for bareness and tail length traits, and therefore make progress towards a sheep that is easier to look after and suffers fewer welfare insults. Some breeds, however, showed very little within-breed variation, and outcrossing may be required to introduce genotypes that exhibit the traits. Whatever approach is taken by the industry, these results support that genetic improvement can be used to breed "ethically improved sheep".

Genetic parameters for residual feed intake, methane emissions, and body composition in New Zealand maternal sheep.

There is simultaneous interest in improving the feed efficiency of ruminant livestock and reducing methane (CH4) emissions. The relationship (genetic and phenotypic) between feed efficiency (characterized as residual feed intake: RFI) and greenhouse gases [methane (CH4) and carbon dioxide (CO2)] traits in New Zealand (NZ) maternal sheep has not previously been investigated, nor has their relationship with detailed estimates of body composition. To investigate these relationships in NZ maternal sheep, a feed intake facility was established at AgResearch Invermay, Mosgiel, NZ in 2015, comprising automated feeders that record individual feeding events. Individual measures of feed intake, feeding behavior (length and duration of eating events), and gas emissions (estimated using portable accumulation chambers) were generated on 986 growing maternal ewe lambs sourced from three pedigree recorded flocks registered in the Sheep Improvement Limited database (www.sil.co.nz). Additional data were generated from a subset of 591 animals for body composition (estimated using ultrasound and computed tomography scanning). The heritability estimates for RFI, CH4, and CH4/(CH4+CO2) were 0.42 ± 0.09, 0.32 ± 0.08, and 0.29 ± 0.06, respectively. The heritability estimates for the body composition traits were high for carcass lean and fat traits; for example, the heritability for visceral fat (adjusted for body weight) was 0.93 ± 0.19. The relationship between RFI and CH4 emissions was complex, and although less feed eaten will lead to a lowered absolute amount of CH4 emitted, there was a negative phenotypic and genetic correlation between RFI and CH4/(CH4+CO2) of -0.13 ± 0.03 and -0.41 ± 0.15, respectively. There were also genetic correlations, that were different from zero, between both RFI and CH4 traits with body composition including a negative correlation between the proportion of visceral fat in the body and RFI (-0.52 ± 0.16) and a positive correlation between the proportion of lean in the body and CH4 (0.54 ± 0.12). Together the results provide the first accurate estimates of the genetic correlations between RFI, CH4 emissions, and the body composition (lean and fat) in sheep. These correlations will need to be accounted for in genetic improvement programs.

Household-level predictors of the presence of servants in Northern Orkney, Scotland, 1851-1901.

Servants were an important part of the northwestern European household economy in the preindustrial past. This study examines household-level characteristics that are predictive of the presence of rural servants using data from Orkney, Scotland. The number of servants present in a household is related to household composition, landholding size, and the marital status of the household head. In addition, the sex of the particular servant hired reveals that the labor of male and female servants is not fungible. The sex of the servant hired is related to the ratio of male and female household members of working age, the occupation of the head, household composition, and the size of the household's landholding.

Relationships among Consumer Liking, Lipid and Volatile Compounds from New Zealand Commercial Lamb Loins.

Loin sections (m. Longissimus lumborum) were collected at slaughter from forty-eight lamb carcasses to evaluate consumer-liking scores of six types of typical New Zealand commercial lamb and to understand the possible underlying reasons for those ratings. A consumer panel (n = 160) evaluated tenderness, juiciness, flavor liking, and overall liking of the different types of lamb loins. Consumer scores differed among the types of lamb meat for all the evaluated attributes (p < 0.05). Further segmentation based on overall liking scores showed two consumer clusters with distinct ratings. Correlation and external preference map analyses indicated that one consumer cluster (n = 75) liked lamb types that had lower total lipid content, a lower proportion of branched-chain fatty acids, oleic and heptadecanoic acids; and a higher proportion of polyunsaturated fatty acids and volatile compounds (green and fruity descriptors). Consumer liking of the other segment (n = 85) was less influenced by fatty acids and volatiles, except hexanoic, heptanoic and octanoic acids (rancid, fatty, and sweaty descriptors). Thus, the fatty acid profile and the volatile compounds derived from their oxidation upon cooking seem to be a stronger driver of consumer liking of lamb for some consumers than others.

Genomic Tools for the Identification of Loci Associated with Facial Eczema in New Zealand Sheep.

Facial eczema (FE) is a significant metabolic disease that affects New Zealand ruminants. Ingestion of the mycotoxin sporidesmin leads to liver and bile duct damage, which can result in photosensitisation, reduced productivity and death. Strategies used to manage the incidence and severity of the disease include breeding. In sheep, there is considerable genetic variation in the response to FE. A commercial testing program is available for ram breeders who aim to increase tolerance, determined by the concentration of the serum enzyme, gamma-glutamyltransferase 21 days after a measured sporidesmin challenge (GGT21). Genome-wide association studies were carried out to determine regions of the genome associated with GGT21. Two regions on chromosomes 15 and 24 are reported, which explain 5% and 1% of the phenotypic variance in the response to FE, respectively. The region on chromosome 15 contains the ß-globin locus. Of the significant SNPs in the region, one is a missense variant within the haemoglobin subunit ß (HBB) gene. Mass spectrometry of haemoglobin from animals with differing genotypes at this locus indicated that genotypes are associated with different forms of adult ß-globin. Haemoglobin haplotypes have previously been associated with variation in several health-related traits in sheep and warrant further investigation regarding their role in tolerance to FE in sheep. We show a strategic approach to the identification of regions of importance for commercial breeding programs with a combination of discovery, statistical and biological validation. This study highlights the power of using increased density genotyping for the identification of influential genomic regions, combined with subsequent inclusion on lower density genotyping platforms.

Fatty Acid Composition and Volatile Profile of M. longissimus thoracis from Commercial Lambs Reared in Different Forage Systems.

Animal production factors can affect the fatty acid and volatile profile of lamb meat. The fatty acid and volatile composition of the M. longissimus thoracis was evaluated from 150 lambs from 10 groups of commercial lambs that differed in age, sex, diet and breed, from three farms, which represent typical forage lamb production systems in New Zealand. The meat from 4-month-old composite lambs slaughtered at weaning had a similar polyunsaturated to saturated fatty acid ratio compared to 6- to 8-month-old composite lambs, but a greater ratio than that of 12-month-old Merino lambs (p < 0.05), with all ratios being lower than the recommended ≥0.45. All lamb production systems produced meat with an omega-6 to omega-3 ratio below 1.5, well below the recommended ratio ≤ 4.0. Meat from 4-month-old lambs had higher C12:0, C14:0 and C16:0 and lower C18:0, reflecting the composition of the milk diet, resulting in higher atherogenic index than meat from other animal groups, while meat from 12-month-old Merino lambs, with lower content of polyunsaturated fatty acids, showed higher thrombogenic index. Meat from lambs processed at weaning contained the greatest concentration of eicosapentaenoic and docosahexaenoic acids, which would qualify as a 'source' or 'good source' of these target fatty acids based on the Commission of Regulation of the European Union or the Food Standards Australia New Zealand guidelines, respectively. Volatiles were extracted from the headspace of raw lean meat and 36 volatile compounds were identified. The abundance of carbon disulphide, isododecane, heptanal, 2,5-hexanediol and 3-octanone and pentanoic, octanoic, nonanoic and heptanoic acids was similar between all groups of lambs. Meat from 12-month-old Merino lambs had low abundance of acetic, propanoic, butanoic and hexanoic acids, and hexanal, octanal and dimethyl sulphide. For 6- to 8-month-old composite lambs, hexanal, octanal and nonanal were present at higher relative abundance in meat from lambs that grazed on chicory than perennial ryegrass. The significant differences in the fatty acid and volatile profiles in meat from 12-month-old Merino lambs compared with lambs slaughtered at weaning or further grazed on red clover, chicory or mixed pasture may result in distinctive nutritional value and lamb flavour.

Comprehensive Evaluation of Parameters Affecting One-Step Method for Quantitative Analysis of Fatty Acids in Meat.

Despite various direct transmethylation methods having been published and applied to analysis of meat fatty acid (FA) composition, there are still conflicting ideas about the best method for overcoming all the difficulties posed by analysis of complex mixtures of FA in meat. This study performed a systematic investigation of factors affecting a one-step method for quantitative analysis of fatty acids in freeze-dried animal tissue. Approximately 280 reactions, selected using factorial design, were performed to investigate the effect of temperature, reaction time, acid concentration, solvent volume, sample weight and sample moisture. The reaction yield for different types of fatty acids, including saturated, unsaturated (cis, trans and conjugated) and long-chain polyunsaturated fatty acids was determined. The optimised condition for one-step transmethylation was attained with four millilitres 5% sulfuric acid in methanol (as acid catalyst), four millilitres toluene (as co-solvent), 300 mg of freeze-dried meat and incubation at 70 °C for 2 h, with interim mixing by inversion at 30, 60 and 90 min for 15 s. The optimised condition was applied to meat samples from different species, covering a broad range of fat content and offers a simplified and reliable method for analysis of fatty acids from meat samples.

Self-reported impulsivity in Huntington's disease patients and relationship to executive dysfunction and reward responsiveness.

INTRODUCTION: Few studies have directly investigated impulsivity in Huntington's disease (HD) despite known changes in dopaminergic and frontal functioning, changes that have been associated with impulsivity in other disorders and in the normal population. This study sought to further categorize impulsivity in HD through examining differences in self-reported impulsivity between community controls and HD patients, the relationship between executive dysfunction and impulsivity, and the relationship of a reward/punishment behavioral inhibition task in relation to these self-report measures. It was expected that HD patients would report higher impulsivity and executive dysfunction and that these measures would relate to a reward/punishment behavioral inhibition task. METHOD: The Barratt Impulsivity Scale (BIS-11) and Behavioral Inhibition/Behavioral Activation Scale (BIS/BAS) were completed, and the Mini-Mental State Examination (MMSE) and a reward-based flanker task with punishing and rewarding conditions were administered to 22 HD patients and 14 control participants. RESULTS: HD patients reported higher trait impulsivity (BIS-11) and executive dysfunction (Frontal Systems Behavior Scale, FrSBE) but not increased impulsivity on the BIS/BAS relative to controls. Higher BIS-11 scores were related to increased self-reported executive dysfunction and the attention/working memory factor of the MMSE. On a reward/punishment behavioral inhibition task, BAS was uniquely related to increased accuracy on rewarding trials of the flanker task, but was not related to punishing trials in HD patients. CONCLUSIONS: The relationships found suggest that trait impulsivity is reported higher in HD and may not be driven by altered reward evaluation and the appetitive nature of stimuli but rather by increased executive dysfunction and lack of sensitivity to punishment. Impulsivity in HD may represent a combination of trait impulsivity, altered dopaminergic circuitry, and executive dysfunction. Understanding impulsivity in HD is important as it is related to increased risk to the patient and difficult behaviors for the caregiver, and sheds light on the disease process.

Pre-clinical medical students' perceptions of their patient safety skills in a simulated emergency department.

OBJECTIVES: Patient safety has emerged as an important topic for inclusion in medical curricula. However, there is limited literature describing how medical students are taught, learn and self-assess patient safety skills. The present study aimed to seek pre-clinical medical students' perceptions of (i) their individual performance at a range of safety skills; and (ii) how they define patient safety in a simulated ED. METHODS: Data were collected in the form of questionnaire responses at the end of the Bond University Simulated ED activity in October 2014. The simulated ED is a session for pre-clinical medical students prior to their clinical rotations, aimed at preparing them for the clinical environment. Likert scale and short answer responses were used to describe students' perceptions of their safety skills and to identify themes related to patient safety. RESULTS: Seventy-eight out of 92 students completed the questionnaires (85% response rate). The majority of students thought their patients were safe from adverse events during their shift. However, students' perception of patient safety was wide-ranging and demonstrated a number of misconceptions. The most frequent strategies employed by students to ensure patient safety were asking for help from nursing staff or senior doctors, ensuring good communication with the patient and checking allergies before administering medication. CONCLUSION: Students had a favourable opinion of their own safety skills. However, answers to free text question revealed misconceptions about the nature of patient safety despite significant teaching on this topic.

Pediatric misophonia with comorbid obsessive-compulsive spectrum disorders.

OBJECTIVE: Misophonia is a potentially debilitating condition characterized by increased sensitivity to specific sounds, which cause subsequent behavioral and emotional responses. The nature, clinical phenomenology and etiology of misophonia remain unclear, and misophonic clinical presentations are not currently accounted for by existing psychiatric or audiological disorders. METHOD: We present a case of pediatric misophonia in the context of comorbid obsessive-compulsive disorder and Tourette's syndrome. RESULTS: Given the interrelationships among obsessive-compulsive spectrum disorders and misophonia, these disorders may share underlying pathophysiology, particularly within the dopaminergic and serotonergic neural systems. Clinical (i.e., treatment) and theoretical implications are discussed.

Use of the oxford multilocus sequence typing protocol and sequencing of the flagellin short variable region to characterize isolates from a large outbreak of waterborne Campylobacter sp. strains in Walkerton, Ontario, Canada.

The Walkerton (Ontario, Canada) outbreak of waterborne Escherichia coli O157:H7 and Campylobacter jejuni was quite limited in both space and time, making it a good model for exploring the utility of different typing and subtyping methods for the characterization of relationships among isolates of these organisms. We have extended previous work with these organisms through analysis by the Oxford multilocus sequence typing (MLST) and the flagellin short variable region (fla-SVR) sequencing methods. Additional isolates not epidemiologically related to the Walkerton outbreak have also been included. Both sequencing methods identified and differentiated between Walkerton outbreak strains 1 and 2. When these strains were compared with isolates that were not part of the outbreak, the information produced by the fla-SVR method more often correlated with epidemiological findings than that produced by MLST, though both methods were required for optimal discrimination. The MLST data were more relevant in terms of the overall population structure of the organisms. Both mutation and recombination appeared to be responsible for generating diversity among the isolates tested.