Liver Transcriptome Responses to Heat Stress and Newcastle Disease Virus Infection in Genetically Distinct Chicken Inbred Lines

Abstract

Heat stress results in reduced productivity, anorexia, and mortality in chickens. The objective of the study was to identify genes and signal pathways associated with heat stress and Newcastle disease virus (NDV) infection in the liver of chickens through RNA-seq analysis, using two highly inbred chicken lines (Leghorn and Fayoumi). All birds were held in the same environment until 14 days of age. On day 14, half the birds were exposed to 38 °C with 50% relative humidity for 4 h, then 35 °C until the end of the experiment. The remaining birds were kept at 25 °C throughout the experiment. The heat-treated birds were inoculated at 21 days of age with 107 EID50 (One EID50 unit is the amount of virus that will infect 50 percent of inoculated embryos) NDV La Sota strain to investigate the effects of both heat stress and NDV infection. Physiological parameters were recorded as blood phenotypes at three stages: acute heat (AH), chronic heat (CH1), and chronic heat combined with NDV infection (CH&NDV), at 4 h, 7 days, and 10 days post-initiation of heat treatment, respectively. Our previous work revealed that the heat-resilient Fayoumi line maintained a more stable acid-base balance in their blood compared to the Leghorn line. Liver samples were harvested on both AH and CH&NDV to characterize the transcriptome profiles of these two inbred lines. Both genetic lines and treatments had large impact on the liver transcriptome. Fayoumi birds had more differentially expressed genes (DEGs) than Leghorn birds for both treatments. Metabolic and immune-related genes were on the DEG list, with Fayoumi having more immune-related DEGs than Leghorns, which was confirmed by gene functional enrichment analysis. Weighted correlation network analysis (WGCNA) indicated that the driver genes such as Solute Carrier Family genes could be very important for stabilizing the acid-base balance in Fayoumi birds during heat stress. Therefore, candidate genes such solute carrier family genes could be potential genetic targets that are regulated by Fayoumis to maintain physical hemostasis under heat stress. Differential gene expression showed that Leghorns mainly performed metabolic regulation in response to heat stress and NDV infection, while Fayoumis regulated both immune and metabolic functions. This study provides novel insights and enhances our understandings of liver response to heat stress of heat resilient and susceptible inbred chicken lines.

Transcriptome Response of Liver and Muscle in Heat-Stressed Laying Hens

“…Of the liver DEGs that were found in multiple heat-control contrasts, HSPA5 and protein disulfide isomerase family A member 2 ( PDIA2 ), which both codes for proteins involved with regulation of protein folding, have been identified as differentially expressed in other heat stress experiments [ 48 , 49 ]. Basic helix-loop-helix family member A15 ( BHLHA15 ), which has a regulatory role in transcription, had also been identified as a liver DEG in a chicken heat stress experiment [ 25 ]. Lastly, ovochymase 2 ( OVCH2 ), of which the cellular function is not well characterized beyond having endopeptidase activity, was identified as one of the genes responsible for impaired lipid utilization in liver tissue of riboflavin-deficient chicken embryos [ 50 ].…”

“…There were many more differentially expressed genes shared among the time-contrasts in the heat-treated tissues. Identified in multiple previous chicken heat stress experiments [ 25 , 41 , 48 , 62 ], DNAJA4 , a HSP coding gene, was identified as a DEG in response to heat treatment for both the 2-week and 4-week compared to 3 h time contrasts in muscle. We found PDK4 , a mitochondrial gene that codes for a metabolic protein, to be differentially expressed at the 2-week to 3 h time contrast for both muscle and liver, and PDK4 has also been previously identified as a DEG in multiple chicken heat stress experiments [ 63 , 64 ].…”

“…Birds were housed in individual battery cages with ad libitum access to mash layer feed and water in climate-and light-controlled rooms at VT. Heat-treated groups were exposed to cyclic heat for 4 weeks (from 24 weeks to 28 weeks of age), with daily heat cycles consisting of 7 h of elevated temperature at 35 • C and 17 h of ambient temperature at 30 • C. Control birds remained at 23 • C throughout the entire experiment. Birds were euthanized and the tissues were collected at 3 time points in 2-week intervals that evenly divided the 4-week experiment: acute heat stress (exposure to heat for 3 h), and chronic heat stress at 2 and 4 weeks of cyclic heat because previous studies have demonstrated that the transcriptomic response is quite different upon first and later exposures to heat [11,24,25]. At each time point, 1 cm cubes were dissected from the pectoralis major muscle and liver for 8 birds from the heat treatment group and 8 birds from the control group, diced and immediately placed in RNAlater (Sigma-Aldrich, St. Louis, MO, USA), transferred to Iowa State University, and stored at −80 • C until RNA isolation.…”

Liver Transcriptome Response to Heat Stress in Beijing You Chickens and Guang Ming Broilers

“…In poultry, the optimum temperature for handling most birds is between 18 and 20 degrees Celsius, and an increase in this temperature can negatively affect growth rate, productivity, and other biochemical characteristics [ 2 , 3 , 4 ]. This rise of temperature combined with humidity and other factors during prolonged hot weather results in heat stress [ 5 , 6 , 7 ]. Stress is a biological response to the animal’s adaptation to events that cause it to lose its normal physiological state [ 5 ].…”

“…Furthermore, the liver plays an important role in maintaining homeostasis, general metabolism, and protein synthesis by regulating fat and glucose metabolism [ 36 ]. Thus, additional molecular and cellular mechanistic investigations using liver transcriptome profiling may be beneficial for characterizing gene expression changes and elucidating the complex genomic response to heat stress, thereby providing additional insight into the genetic regulation of heat tolerance in chickens [ 7 ].…”