Characterization and Isolation of Segmented Filamentous Bacteria from Commercial Turkey

Abstract

This work aimed to characterize and isolate segmented filamentous bacteria from turkey hosts in order to determine the role these organisms play in improving the health of commercial turkeys. Segmented filamentous bacteria (SFB) have been associated with healthy growth outcomes of turkey flocks and are assumed to play an immunostimulatory role in the development of the innate and adaptive immune systems of the host and have potential as novel direct-fed microbials as the demand for alternatives to antibiotic use rises. The approach involved several methods for the in vitro and in silico analysis of turkey-associated SFB, including whole genome sequencing, predictive genomics and metabolomics assays, extraction and enrichment of SFB from ileal homogenates, development of a genetic detection technique, and comparative growth studies in various media formulations. Additionally, attempts to isolate SFB-turkey from other members of the extracted intestinal co-cultures were attempted using dilution-to-extinction, growth in “spent media”, separation via filtration and targeted use of species-specific bacteriophage lysates. Turkey-associated SFB are very distinct among the SFB, possessing several distinguishing metabolic characteristics including an increased capacity to degrade amino acids, metabolize a broader array of mono/disaccharide substrates, break down complex glycans, hydrolyze and recycle primary bile acids, and synthesize biotin. In examining the ability of SFB-turkey to grow and reproduce in vitro, it was initially determined that turkey- associated SFB were able to grow to high proportions independently from intestinal cell lines in liquid cultures grown under microaerobic conditions after 48 hours of incubation. Following several failed attempts to separate and purify SFB-turkey from other microbial members contained within the obtained co-cultures it was determined that cultures presumed to contain SFB-turkey were found to consist of high proportions of Bacillus marisflavi, an organism observed to display filamentous growth without the formation of transverse septa. The presented work underscores the highly auxotrophic nature of SFB and overall dependence on the host intestinal environment for consistent growth and development.