RNA-Seq Analysis Reveals Spatial and Sex Differences in Pectoralis Major Muscle of Broiler Chickens Contributing to Difference in Susceptibility to Wooden Breast Disease.

Brilynn Brothers,Michael B Papah,Behnam Abasht,Zhu Zhuo

doi:10.3389/fphys.2019.00764

Abstract

Wooden Breast Disease (WBD) is a novel myopathy affecting the pectoralis major muscle of modern broiler chickens. The etiology of WBD is not currently known, but has been linked to increased feed efficiency, growth rate, and muscle yield in broiler chickens. Differential effect of WBD has been detected between regions of the P. major and between sexes of broilers—male birds and the cranial aspect of the muscle tend to be more severely affected by the disease than females and the caudal aspect. This study aimed to characterize biological differences in the P. major between regions of the muscle and sexes of birds. Samples were taken from the cranial and caudal aspects of P. major muscles of 3-week-old, unaffected male and female birds for RNA sequencing. RNA was extracted and used for preparation of cDNA libraries, which were sequenced by the Delaware Biotechnology Institute (DBI) using HiSeq2500. Sequence reads were aligned to the chicken reference genome with HISAT, and genes were analyzed for differential expression between regions of the breast muscle and sexes of birds using CuffDiff. Functional analysis was performed on differentially expressed genes (DEGs) between sex groups using DAVID and Ingenuity Pathway Analysis (IPA). There were 12 DEGs between cranial and caudal samples, and 260 between male and female birds. Out of the 260 genes differentially expressed between sexes, 189 were upregulated in males. Of this subset, 103 genes (55%) were located on the Z-chromosome. There was increased expression of genes involved in fat metabolism and oxidative stress responses in the cranial region of the P. major muscle, as well as increased expression of fat metabolism, oxidative stress response, antiangiogenesis, and connective tissue proliferation genes in male broilers. These results support the hypothesis that there are biological characteristics in male broilers and the cranial region of the breast muscle that may make them more susceptible to WBD, as well as raising the possibility of a metabolic switch in modern broiler chickens that may be more prominent in males.

Highlights

Demand for poultry meat has greatly increased over the last 50 years
This study aims to provide answers for a few questions: Why is the cranial aspect of the pectoralis major more affected by Wooden Breast Disease (WBD) than the caudal, and why are males more affected than females? why does the breast muscle grow larger and faster in male birds than females, and why does the cranial aspect grow faster and become thicker than the caudal aspect? This study utilizes RNA sequencing techniques to look at differential gene expression between cranial and caudal sections of the pectoralis major, as well as between male and female birds in search of differing biological pathways that may lead to differing susceptibility to WBD
We explored the functions of these genes individually: Perilipin 1 (PLIN1), G0/G1 switch gene 2 (G0S2), and fatty acid-binding protein 4 (FABP4) are all involved in lipid metabolism; neuritin 1 (NRN1) is involved in nervous system development; kyphoscoliosis peptidase (KY) is involved in muscle growth, and homeobox A11 (HOXA11) and pituitary homeobox 2 (PITX2) are transcription factors

Summary

Introduction

To meet the still-growing demand for poultry meat, the poultry industry has implemented methods to increase growth rate, feed efficiency, and muscle yield, especially that of the breast muscle, in turkeys and chickens These improvements in the efficiency of the poultry industry may have inadvertently induced a higher incidence of muscle disorders and abnormalities, such as deep pectoral muscle disease and white striping (Petracci et al, 2013). Later stages of WBD demonstrate diffuse myodegeneration, inflammatory cell infiltration, fibrosis, necrosis, and lipid infiltration (Papah et al, 2017) Though they may occur independently of each other, WBD is often coexistent with another muscle condition, namely, white striping on the muscle (Sihvo et al, 2014; Mudalal et al, 2015; Dalle Zotte et al, 2017; Griffin et al, 2018)

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