Virginia Research Day 2022

Graduate Student Research Biomedical

11 Combatting Streptococcus Suis: A Combined Sequencing Approach

S. Tristan Stoyanof 1 , Kevin K. Lahmers 1 , Kevin S. McIver 2 , Clayton C. Caswell 1 Corresponding author: tstoy@vt.edu

1 Virginia-Maryland College of Veterinary Medicine 2 University of Maryland

The Tn-seq experiments established a list of genes essential for survival by first isolating total genomic DNA from an unstressed library of mutants, then sequencing the pool to map transposon insertional sites. This same Tn- seq library was then grown in either mucin or was allowed to form a biofilm, which led to the characterization identification of genes that contributed to the overall fitness of S. suis in those conditions. Combined, these experimental findings begin to lay the groundwork for better defining virulence mechanisms in S. suis.

The RNA-seq experiments demonstrated that specific genes are differentially expressed in the presence of either pig mucin, cerebral spinal fluid, serum, or within a biofilm. We hypothesize that an operon encoding four YhgE/Pip domain-containing proteins, which is upregulated when the bacteria are exposed to mucin, plays a critical role in the overall virulence of S. suis. In addition to measuring total gene expression, multiple novel small RNAs (sRNAs) were predicted and confirmed via northern blot analyses. Among these sRNAs, three appear to be more abundant when isolated from a biofilm.

Streptococcus suis poses as a looming economic threat to the pork industry and a growing health concern for those working in close proximity to the infected pigs. Although outbreaks have been recorded across the world, with many instances displaying high lethality in both pigs and humans, S. suis remains a relatively understudied bacterium. In response, this project utilized multiple high-throughput sequencing methods to better understand the conditional expression and importance of certain genes, and to locate potential therapeutic targets within S. suis. This was done via transposon sequencing (Tn-seq) and RNA sequencing (RNA-seq) approaches.

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