Insight into a natural novel histidine decarboxylase gene deletion in Enterobacter hormaechei RH3 from traditional Sichuan-style sausage.

Abstract

Histamine (HIS) is primarily formed from decarboxylated histidine by certain bacteria with histidine decarboxylase (hdc) activity and is the most toxic biogenic amine. Hdc, which is encoded by the hdc gene, serves as a key enzyme that controls HIS production in bacteria. In this paper, we characterized the changes in microbial and biogenic amines content of traditional Sichuan-style sausage before and after storage and demonstrated that Enterobacteriaceae play an important role in the formation of HIS. To screen for Enterobacteriaceae with high levels of HIS production, we isolated strain RH3 which has a HIS production of 2.27mg/mL from sausages stored at 37°C for 180 days, using selective media and high-performance liquid chromatography. The strain RH3 can produce a high level of HIS after 28h of fermentation with a significant hysteresis. Analysis of the physicochemical factors revealed that RH3 still retained its ability to partially produce HIS in extreme environments with pH 3.5 and 10.0. In addition, RH3 exhibited excellent salt tolerance (6.0% NaCl and 1.0% NaNO2 ). Subsequently, RH3 was confirmed as Enterobacter hormaechei with hdc gene deletion by PCR, western blot, and whole-genome sequencing analysis. Furthermore, RH3 exhibited pathogenicity rate of 75.60% toward the organism, indicating that it was not a food-grade safe strain, and demonstrated a high level of conservation in intraspecific evolution. The results of this experiment provide a new reference for studying the mechanism of HIS formation in microorganisms. PRACTICAL APPLICATION: This study provides a new direction for investigating the mechanism of histamine (HIS) formation by microorganisms and provides new insights for further controlling HIS levels in meat products. Further research can control the key enzymes that form HIS to control HIS levels in food.