**ASC-Mediated Inflammation and Pyroptosis Attenuates** *Brucella abortus* **Pathogenesis Following the Recognition of gDNA**

**Juselyn D. Tupik 1, Sheryl L. Coutermarsh-Ott 1, Angela H. Benton 1, Kellie A. King 1, Hanna D. Kiryluk 1, Clayton C. Caswell 1 and Irving C. Allen 1,2,\***


Department Virginia **\***Correspondence:icallen@vt.edu; Tel.: +1-540-231-7551;Fax:1-540-231-6033

Received: 31 October 2020; Accepted: 25 November 2020; Published: 30 November 2020

**Abstract:** *Brucella abortus* is a zoonotic pathogen that causes brucellosis. Because of *Brucella's* unique LPS layer and intracellular localization predominately within macrophages, it can often evade immune detection. However, pattern recognition receptors are capable of sensing *Brucella* pathogen-associated molecular patterns (PAMPS). For example, NOD-like receptors (NLRs) can form a multi-protein inflammasome complex to attenuate *Brucella* pathogenesis. The inflammasome activates IL-1β and IL-18 to drive immune cell recruitment. Alternatively, inflammasome activation also initiates inflammatory cell death, termed pyroptosis, which augments bacteria clearance. In this report, we assess canonical and non-canonical inflammasome activation following *B. abortus*infection. We conducted in vivo studies using *Asc*−/−mice and observed decreased mouse survival, immune cell recruitment, and increased bacteria load. We also conducted studies with *Caspase-11*−/−mice and did not observe any significant impact on *B. abortus* pathogenesis. Through mechanistic studies using *Asc*−/<sup>−</sup>macrophages, our data suggests that the protective role of ASC may result from the induction of pyroptosis through a gasdermin D-dependent mechanism in macrophages. Additionally, we show that the recognition of *Brucella* is facilitated by sensing the PAMP gDNA rather than the less immunogenic LPS. Together, these results refine our understanding of the role that inflammasome activation and pyroptosis plays during brucellosis.

**Keywords:** brucellosis; canonical inflammasome; non-canonical inflammasome; NLR; pyroptosis; ASC; caspase-11; caspase-1; IL-1β; gDNA
