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SUMMARYTwenty-four gilts were used at each of two centres in an experiment designed to examine the effects of dietary energy intake during lactation on yield and composition of sows' milk and the growth of their litters. Gestation feed intake was standardized at 2 kg per day supplying 6·28 Meal digestible energy (DE) and 280 g crude protein. The energy levels fed during lactation ranged from 12·2 to 18·25 Meal DE per day for gilts, 12·9 to 19·6 Meal DE per day for second parity and from 13·2 to 20·25 Meal DE per day for third parity sows. Daily crude protein intake and amino acid balance ofthe protein were kept constant.In the first lactation milk yield and composition and growth of litters were not influenced by dietary energy intake. In the second lactation milk yield was depressed by lower energy intakes and this was reflected in litter weights at 21 though not at 42 days. In the third lactation lower energy intakes seriously depressed milk yield and daily output of milk nutrients. On the lowest level of energy, milk yield was 32% lower at day 24 and 20% lower at day 41 of lactation than on the highest energy level. The corresponding depressions in litter weights were 17·2 and 13·5% at 21 and 42 days respectively.

BackgroundTalitrus saltator is an amphipod crustacean that inhabits the supralittoral zone on sandy beaches in the Northeast Atlantic and Mediterranean. T. saltator exhibits endogenous locomotor activity rhythms and time-compensated sun and moon orientation, both of which necessitate at least one chronometric mechanism. Whilst their behaviour is well studied, currently there are no descriptions of the underlying molecular components of a biological clock in this animal, and very few in other crustacean species.MethodsWe harvested brain tissue from animals expressing robust circadian activity rhythms and used homology cloning and Illumina RNAseq approaches to sequence and identify the core circadian clock and clock-related genes in these samples. We assessed the temporal expression of these genes in time-course samples from rhythmic animals using RNAseq.ResultsWe identified a comprehensive suite of circadian clock gene homologues in T. saltator including the ‘core’ clock genes period (Talper), cryptochrome 2 (Talcry2), timeless (Taltim), clock (Talclk), and bmal1 (Talbmal1). In addition we describe the sequence and putative structures of 23 clock-associated genes including two unusual, extended isoforms of pigment dispersing hormone (Talpdh). We examined time-course RNAseq expression data, derived from tissues harvested from behaviourally rhythmic animals, to reveal rhythmic expression of these genes with approximately circadian period in Talper and Talbmal1. Of the clock-related genes, casein kinase IIβ (TalckIIβ), ebony (Talebony), jetlag (Taljetlag), pigment dispensing hormone (Talpdh), protein phosphatase 1 (Talpp1), shaggy (Talshaggy), sirt1 (Talsirt1), sirt7 (Talsirt7) and supernumerary limbs (Talslimb) show temporal changes in expression.DiscussionWe report the sequences of principle genes that comprise the circadian clock of T. saltator and highlight the conserved structural and functional domains of their deduced cognate proteins. Our sequencing data contribute to the growing inventory of described comparative clocks. Expression profiling of the identified clock genes illuminates tantalising targets for experimental manipulation to elucidate the molecular and cellular control of clock-driven phenotypes in this crustacean.

The live-weight changes of 48 sows were recorded over three reproductive cycles. All sows were given standard amounts of feed in gestation and one of four energy allowances during lactation. The daily intakes of digestible energy during lactation ranged from 12-2 to 18-2 Meal in the first, 12-9 to 19-6 Meal in the second and 13-2 to 20-2 Meal in the third lactation. Weight change in lactation responded to energy intake; sows on the lowest energy intake lost weight whilst those on the highest intake gained. At weaning of their third litters, sows on the highest energy allowance weighed 44 kg more than those on the lowest level and this was equivalent to a carcass difference of 37-3 kg at slaughter. All the sows were slaughtered after the third litters had been weaned and the carcasses were dissected into lean, subcutaneous fat+skin, and bone. Sows from all treatments had less subcutaneous fat+ skin and more muscle and bone than non-pregnant gilts killed at a mean weight of 129 kg, a weight equivalent to that of the experimental animals at their first mating. The reduction in fat reserves was linearly correlated with dietary energy intake of the sows. Sows receiving the lowest energy intake in lactation possessed fat reserves which were estimated to be only 25% as great as in gilts comparable to those used to initiate the experiment. This reduction in body reserves has implications with regard to the long-term effects of feeding lactating sows low intakes of energy during 6-week lactations.

Crustacean hyperglycemic hormone (CHH) has been extensively studied in decapod crustaceans where it is known to exert pleiotropic effects, including regulation of blood glucose levels. Hyperglycemia in decapods seems to be temporally gated to coincide with periods of activity, under circadian clock control. Here, we used gene cloning, in situ hybridization and immunohistochemistry to describe the characterization and localization of CHH in two peracarid crustaceans, Eurydice pulchra and Talitrus saltator. We also exploited the robust behavioral rhythmicity of these species to test the hypothesis that CHH mRNA expression would resonate with their circatidal (12.4h) and circadian (24h) behavioral phenotypes. We show that both species express a single CHH transcript in the cerebral ganglia, encoding peptides featuring all expected, conserved characteristics of other CHHs. E. pulchra preproCHH is an amidated 73 amino acid peptide N-terminally flanked by a short, 18 amino acid precursor related peptide (CPRP) whilst the T. saltator prohormone is also amidated but 72 amino acids in length and has a 56 residue CPRP. The localization of both was mapped by immunohistochemistry to the protocerebrum with axon tracts leading to the sinus gland and into the tritocerebrum, with striking similarities to terrestrial isopod species. We substantiated the cellular position of CHH immunoreactive cells by in situ hybridization. Although both species showed robust activity rhythms, neither exhibited rhythmic transcriptional activity indicating that CHH transcription is not likely to be under clock control. These data make a contribution to the inventory of CHHs that is currently lacking for non-decapod species.

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ABSTRACT

1. Five hundred and fifty-two pigs were used in an experiment having six treatments. High and low protein rations were given (a) unsupplemented, (b) supplemented with 0·1% copper sulphate (CuSO4·5H2O) and (c) supplemented with 0·1% copper sulphate and 0·025% zinc carbonate (ZnCO3).2. Performance was significantly reduced and 18 pigs died from copper toxicosis, when the low protein ration supplemented with copper sulphate was fed.3. There were no indications of toxicity and performance was equal to the controls when the low protein diet supplemented with copper and zinc was fed.4. The addition of copper to the high protein diet slightly reduced performance and six pigs died from copper toxicosis.5. The addition of copper and zinc to the high protein diet improved growth rate and eliminated the toxicity associated with copper supplementation.6. No significant treatment effects on haemoglobin or packed cell volume were observed, other than a small effect of protein level on PCV.7. Copper supplementation of the diets significantly increased liver and kidney copper levels and liver zinc levels. It reduced liver and kidney iron contents. The addition of copper and zinc to the diet reduced the increase in liver copper associated with copper supplementation, and increased the level of zinc in both livers and kidneys.