Identification of an operon and its regulator required for autoaggregation in Tetragenococcus halophilus

Abstract

ABSTRACT Bacterial autoaggregation is associated with self- and non-self-recognition, surface colonization, and biofilm formation. Numerous aggregation factors have been cloned for basic and applied purposes. Snowflake Forming Collagen Binding Aggregation Factor (SFCBAF) is one such aggregation factor isolated from a few lactic acid bacteria species; however, the regulatory mechanisms of SFCBAF remain poorly understood. Here, we report the aggR and aggABC genes associated with autoaggregation in the halophilic lactic acid bacterium Tetragenococcus halophilus . A draft genome sequence analysis of five aggregating strains and subsequent comparative genomic analysis revealed that all of them have aggR and aggABC genes, which are possibly involved in autoaggregation. SFCBAF, a sortase, and a hypothetical protein are encoded by aggABC , which constitutes the agg operon. To analyze the function of these genes, phenotype- and genotype-based mutant screening was carried out. Mutants of either aggR or aggA lack autoaggregation and collagen binding abilities, indicating that they are both essential for these phenotypes. aggR was considered a positive regulator for the agg operon because aggR encodes a helix-turn-helix protein and aggR mutation resulted in diminished agg operon expression. Our results provide genetic bases underlying the mechanisms of aggregation and its regulation in T. halophilus . IMPORTANCE Tetragenococcus halophilus is a halophilic lactic acid bacterium generally used as a starter culture in fermenting soy and fish sauces. Aggregating strains can be useful in fermenting and obtaining clear soy sauce because cell clumps are trapped by the filter cake when the soy sauce mash is pressed. However, the genetic mechanisms of aggregation in T. halophilus are unknown. In this study, we identified genes encoding aggregation factor and its regulator. These findings may provide a foundation for developing improved T. halophilus starter cultures for soy sauce fermentation, leading to more efficient and consistent clear soy sauce production.