Comparative transcriptional analysis of homologous pathogenic and non-pathogenic Lawsonia intracellularis isolates in infected porcine cells.
Lawsonia intracellularis is the causative agent of proliferative enteropathy. This disease affects various animal species, including nonhuman primates, has been endemic in pigs, and is an emerging concern in horses. Non-pathogenic variants obtained through multiple passages in vitro do not induce disease, but bacterial isolates at low passage induce clinical and pathological changes. We hypothesize that genes differentially expressed between pathogenic (passage 10) and non-pathogenic (passage 60) L. intracellularis isolates encode potential bacterial virulence factors. The present study used high-throughput sequencing technology to characterize the transcriptional profiling of a pathogenic and a non-pathogenic homologous L. intracellularis variant during in vitro infection. A total of 401 genes were exclusively expressed by the pathogenic variant. Plasmid-encoded genes and those involved in membrane transporter (e.g. ATP-binding cassette), adaptation and stress response (e.g. transcriptional regulators) were the categories mostly responsible for this wider transcriptional landscape. The entire gene repertoire of plasmid A was repressed in the non-pathogenic variant suggesting its relevant role in the virulence phenotype of the pathogenic variant. Of the 319 genes which were commonly expressed in both pathogenic and non-pathogenic variants, no significant difference was observed by comparing their normalized transcription levels (fold change±2; p<0.05). Unexpectedly, these genes demonstrated a positive correlation (r(2) = 0.81; p<0.05), indicating the involvement of gene silencing (switching off) mechanisms to attenuate virulence properties of the pathogenic variant during multiple cell passages. Following the validation of these results by reverse transcriptase-quantitative PCR using ten selected genes, the present study represents the first report characterizing the transcriptional profile of L. intracellularis. The complexity of the virulence phenotype was demonstrated by the diversity of genes exclusively expressed in the pathogenic isolate. The results support our hypothesis and provide the basis for prospective mechanistic studies regarding specific roles of target genes involved in the pathogenesis, diagnosis and control of proliferative enteropathy.
Department of Veterinary and Biomedical Science, College of Veterinary Medicine, University of Minnesota, St. Paul, Minnesota, United States of America. vannu008@umn.edu
Fabio A
FA
Vannucci
Douglas N
DN
Foster
Connie J
CJ
Gebhart
D000818:Animals; D002460:Cell Line; D020639:Lawsonia Bacteria / Q000235:genetics / Q000472:pathogenicity* / Q000502:physiology*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017422:Sequence Analysis, DNA / Q000379:methods*; D013552:Swine; D013553:Swine Diseases / Q000235:genetics / Q000382:microbiology; D014158:Transcription, Genetic / Q000235:genetics