In this study, we report the purification, cDNA cloning, and characterization of the novel growth hormone-releasing peptide, ghrelin, in the chicken (Gallus gallus). Chicken ghrelin is composed of 26 amino acids (GSSFLSPTYKNIQQQKDTRKPTARLH) and possesses 54% sequence identity with human ghrelin. The serine residue at position 3 (Ser(3)) is conserved between the chicken and mammalian species, as its acylation by either n-octanoic or n-decanoic acid. Chicken ghrelin mRNA is predominantly expressed in the stomach, where it is present in the proventriculus but absent in the gizzard. Using RT-PCR analysis, low levels of expression were also detectable in brain, lung, and intestine. Administration of chicken ghrelin increases plasma GH levels in both rats and chicks, with a potency similar to that of rat or human ghrelin. In addition, chicken ghrelin also increases plasma corticosterone levels in growing chicks at a lower dose than in mammals. The present results indicate that the stimulatory effect of ghrelin on GH secretion is evolutionarily conserved, whereas its effect on adrenal function seems to be unique in the chicken.

CHICKEN GHRELIN : PURIFICATION, cDNA CLONING AND BIOLOGICAL ACTIVITY

A ‘meta-analysis’ was performed to determine effects of post-hatch food and water deprivation (PHFWD) on chicken development, performance and welfare (including health). Two types of meta-analysis were performed on peer-reviewed scientific publications: a quantitative ‘meta-analysis’ (MA) and a qualitative analysis (QA). Previously reported effects of PHFWD were quantified in the MA, for variables related to performance, mortality and relative yolk sac weight. The QA counted the number of studies reporting (non-)significant effects when five or more records were available in the data set (i.e. relative heart, liver and pancreas weight; plasma T3, T4 and glucose concentrations; relative duodenum, jejunum and ileum weight; duodenum, jejunum and ileum length; and villus height and crypt depth in duodenum, jejunum and ileum). MA results indicated that 24 hours of PHFWD (i.e. ≥12–36 hours) or more resulted in significantly lower body weights compared to early-fed chickens up to six weeks of age. Body weights and food intake were more reduced as durations of PHFWD (24, 48, 72, ≥84 hours) increased. Feed conversion rate increased in chickens up to 21 and 42 days of age after ≥84 hours PHFWD in comparison with chickens fed earlier. Total mortality at day 42 was higher in chickens after 48 hours PHFWD compared to early fed chickens or chickens after 24 hours PHFWD. First week mortality was higher in chickens after ≥84 hours PHFWD than in early fed chickens. The MA for relative yolk sac weight was inconclusive for PHFWD. The QA for plasma T3, T4 and glucose concentrations indicated mainly short-term decreases in T3 and glucose in PHFWD chickens compared to early fed chickens, and no effects of PHFWD on T4 concentrations. Relative weights of liver, pancreas and heart were lower after PHFWD, but only in the first week of life. A retarded development of gut segments (duodenum, jejunum and ileum) was found in the first week of life, measured as shorter, lower relative weight, and lower villus height and crypt depth. It is concluded that 48 hours (≥36–60 hours) PHFWD leads to lower body weights and higher total mortality in chickens up to six weeks of age, the latter suggesting compromised chicken welfare, but effects of PHFWD on organ development and physiological status appear to be mainly short-term.

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A 'meta-analysis' of effects of post-hatch food and water deprivation on development, performance and welfare of chickens.

In this study, gestational sows were fed control or betaine-supplemented diets (3 g/kg) throughout the pregnancy, and the newborn piglets were used to elucidate whether maternal dietary betaine affected offspring hepatic gluconeogenic genes through epigenetic mechanisms. Neonatal piglets born to betaine-supplemented sows had significantly higher serum and hepatic betaine contents, together with significantly greater expression of methionine metabolic enzymes in the liver. Interestingly, significantly higher serum concentrations of lactic acid and glucogenic amino acids, including serine, glutamate, methionine and histidine, were detected in the piglets born to betaine-supplemented sows, which were coincident with higher hepatic glycogen content and PEPCK1 enzyme activity, as well as greater protein expression of gluconeogenic enzymes, pyruvate carboxylase (PC), cytoplasmic phosphoenolpyruvate carboxykinase (PEPCK1), mitochondrional phosphoenolpyruvate carboxykinase (PEPCK2) and fructose-1, 6-bisphosphatase (FBP1). Moreover, maternal betaine significantly changed the methylation status of both CpGs and histones on the promoter of gluconeogenic genes. The lower PEPCK1 mRNA was associated with DNA hypermethylation and more enriched repression histone mark H3K27me3, while the up-regulated PEPCK2 and FBP1 mRNA was associated with DNA hypomethylation and more enriched activation histone mark H3K4me3. Furthermore, the expression of two miRNAs predicted to target PC and 6 miRNAs predicted to target PEPCK1 was dramatically suppressed in the liver of piglets born to betaine-supplemented sows. Our results provide the first evidence that maternal betaine supplementation affects hepatic gluconeogenic genes expression in newborn piglets through enhanced hepatic methionine metabolism and epigenetic regulations, which involve DNA and histone methylations, and possibly miRNAs-mediated post-transcriptional mechanism.

Betaine supplementation in maternal diet modulates the epigenetic regulation of hepatic gluconeogenic genes in neonatal piglets

Infectious bursal disease virus (IBDV) of chickens is a birnavirus with a bi-segmented double-stranded RNA genome, the segments designated as A and B. We performed phylogenetic analysis using a 366-bp fragment of segment A (nt 785-1150) and a 508-bp fragment of segment B (nt 328-835) of IBDV. A total of 463 segment A and 434 segment B sequences from GenBank, including the sequences of eight recent Bangladeshi isolates, were used in the analysis. The analysis revealed eight genogroups of segment A under serotype 1, designated as A1 (classical), A2 (US antigenic variant), A3 (very virulent), A4 (dIBDV), A5 (atypical Mexican), A6 (atypical Italian), A7 (early Australian) and A8 (Australian variant), and a single genogroup under serotype 2, designated as A0. On the other hand, segment B could be categorized into five genogroups irrespective of serotype, these being B1 (classical-like), B2 (very virulent-like), B3 (early Australian-like), B4 (Polish & Tanzanian) and B5 (Nigerian). Segment B of serotype 2 strains clustered within genogroup B1. With the bi-segmented genome of IBDV, these differences would allow for a total of 45 possible assortments. Based on the combinations of segment A and segment B genogroups observed in 463 IBDV strains, a total of 15 genotypes could be recognized. Recent Bangladeshi IBDV strains, isolated in 2016, appeared to be segment reassortants having segment A of genogroup A3 (very virulent) and segment B of genogroup B3 (early Australian-like). An extended system of nomenclature of IBDV strains is proposed.

A unified genotypic classification of infectious bursal disease virus based on both genome segments.

Infectious bursal disease virus (IBDV) belongs to the family Birnaviridae and is economically important to the poultry industry worldwide. IBDV infects B cells in the bursa of Fabricius (BF), causing immunosuppression and morbidity in young chickens. In addition to strains that cause classical Gumboro disease, the so-called 'very virulent' (vv) strain, also in circulation, causes more severe disease and increased mortality. IBDV has traditionally been controlled through the use of live attenuated vaccines, with attenuation resulting from serial passage in non-lymphoid cells. However, the factors that contribute to the vv or attenuated phenotypes are poorly understood. In order to address this, we aimed to investigate host cell-IBDV interactions using a recently described chicken primary B-cell model, where chicken B cells are harvested from the BF and cultured ex vivo in the presence of chicken CD40L. We demonstrated that these cells could support the replication of IBDV when infected ex vivo in the laboratory. Furthermore, we evaluated the gene expression profiles of B cells infected with an attenuated strain (D78) and a very virulent strain (UK661) by microarray. We found that key genes involved in B-cell activation and signalling (TNFSF13B, CD72 and GRAP) were down-regulated following infection relative to mock, which we speculate could contribute to IBDV-mediated immunosuppression. Moreover, cells responded to infection by expressing antiviral type I IFNs and IFN-stimulated genes, but the induction was far less pronounced upon infection with UK661, which we speculate could contribute to its virulence.

Differential gene expression in chicken primary B cells infected ex vivo with attenuated and very virulent strains of infectious bursal disease virus (IBDV).

Differential expression of pro-inflammatory and anti-inflammatory genes of layer chicken bursa after experimental infection with infectious bursal disease virus.

Infectious bursal disease virus (IBDV) infection causes immunosuppression in chickens and increases their susceptibility to secondary infections. To explore the interaction between host and IBDV, RNA-Seq was applied to analyse the transcriptional profiles of the responses of chickens’ bursas of Fabricius in the early stage of IBDV infection. The results displayed that a total of 15546 genes were identified in the chicken bursa libraries. Among the annotated genes, there were 2006 and 4668 differentially expressed genes in the infection group compared with the mock group on day 1 and day 3 post inoculation (1 and 3 dpi), respectively. Moreover, there were 676 common up-regulated and 83 common down-regulated genes in the bursae taken from the chickens infected with IBDV on both 1 and 3 dpi. Meanwhile, there were also some characteristic differentially expressed genes on 1 and 3 dpi. On day 1 after inoculation with IBDV, host responses mainly displayed immune response processes, while metabolic pathways played an important role on day three post infection. Six genes were confirmed by quantitative reverse transcription-PCR. In conclusion, the differential gene expression profile demonstrated with RNA-Seq might offer a better understanding of the molecular interactions between host and IBDV during the early stage of infection.

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Jinyou Ma

Papers

Differential expression of pro-inflammatory and anti-inflammatory genes of layer chicken bursa after experimental infection with infectious bursal disease virus.

Transcription profiles of the responses of chicken bursae of Fabricius to IBDV in different timing phases

Identification and assessment of pathogenicity of a naturally reassorted infectious bursal disease virus from Henan, China.

Effects of development and delayed feed access on ghrelin expression in neonatal broiler chickens.

M protein is sufficient for assembly and release of Peste des petits ruminants virus-like particles.