Transport stress induces paradoxical increases in airway inflammatory responses in beef stocker cattle

The development of Bovine Respiratory Disease (BRD ) in beef
cattle is associated with stressful events, including auction and
transport. To characterize immunity at the primary site of pathogen
colonization in BRD, we evaluated stress responses in lung cells from beef
stocker cattle to understand mucosal immune changes. Stocker calves were
sampled via bronchoalveolar lavage fluid collection within 24 hours of
auction and transport to a new facility (Stressed, n=12), or allowed to
acclimate for 2 months at the new facility before sampling (Acclimated,
n=7). Lavage cellular RNA was extracted and sequenced for bulk RNA-seq
gene expression. The resulting fastq files are included in this repository
and named based on the calf's unique identifier at sampling. The
metadata file provides the key to which experimental group each calf
belongs, as well as pertinent sequencing metadata values and MD5 sums to
ensure file quality. Raw data counts are included in the counts.txt file,
and are organized with genes as the row names and calf unique identifiers
as the column names. Code files for DESeq2 and CIBERSORTx analysis are
included as final analysis code.R and Cibersortx analysis.R. Differential
gene expression analysis of RNA sequencing data demonstrated a profound
upregulation of inflammatory genes in Stressed calves compared to
Acclimated calves, including expression of CXCL8, CSF3R, IL1B, and CCL22.
The top pathway upregulation in Stressed calves involved neutrophil
migration and chemotaxis, and cytokine signaling. To predict cellular
population proportions from the data, CIBERSORTx was used to deconvolute
bulk RNA-seq gene counts. This analysis showed that Stressed calves had
significantly increased BALF neutrophils compared to Acclimated calves (p
=0.003). Neutrophilic infiltration occurred in the absence of pathogen
colonization of the lungs in most calves, as demonstrated by a
multi-pathogen respiratory qPCR screen. As such, the stress induced an
inflammatory response in the lungs not explained by pathogen exposure.
This study provides strong evidence that shipping stress in beef stocker
calves can paradoxically trigger increased rather than suppressed
inflammatory pulmonary mucosal immune responses, which have important
implications for the pathogenesis of BRD. All animals were sampled under
institutional IACUC approval.