Virginia Research Day 2021

Faculty Research Biomedical

02 Removal of a Small, Hypothetical Protein Attenuates B . Abortus Survivalin Mice and Macrophages

Angela H Benton; Kellie A King; Clayton C Caswell Corresponding author: ahbenton@vt.edu

Virginia-Maryland College of Veterinary Medicine Department of Biomedical Sciences and Pathobiology

Brucella abortus is a causative agent of brucellosis in both humans and cattle. This Gram-negative bacterium causes spontaneous abortion and sterility in the natural host (cattle), and a debilitating flu- like illness in humans. B. abortus is an intracellular bacterium that not only survives but replicates inside the macrophage. This is of particular interest given the harsh environment inside this immune cell, including a lack of nutrients. Inside the macrophage, the brucellae utilize glucose; however, bacterial metabolism in this environment is not well understood. Other potential carbon sources exist inside macrophages, including L-fucose. L-fucose is found in the digestive tract of

mammals, the rectum of cows, and the lysosomes of macrophages. This 6-carbon sugar was previously discovered to be important in B. abortus for growth in vitro. Wildtype brucellae grow to a significantly higher cell density in minimal media supplemented with L-fucose than without supplementation. Also, a 12-gene operon in B. abortus 2308 was found to be up-regulated in the presence of L-fucose. Within this operon is a gene encoding a small, 33 amino acid, hypothetical protein (BAB1_0245). The function of this protein is unknown; however, a mutant strain lacking bab1_0245 had diminished growth in minimal media even when supplemented with additional

carbon sources. Quantitative PCR also revealed a significant increase in gene expression of bab1_0245 when exposed to fucose, a carbon source that the brucellae may encounter while in the macrophage. A Δbab1_0245 mutant showed significant decreases in survival and replication in a macrophage model. Also, significantly less bacteria was recovered from mice infected with a Δbab1_0245 mutant in a chronic infection model. Together these data indicate BAB1_0245 is necessary for full virulence of B. abortus both in vitro and in vivo .

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